NATIONAL TOXICOLOGY PROGRAM 

Technical Report Series 

No. 383 

 

TOXICOLOGY AND CARCINOGENESIS 

STUDIES OF 

1-AMINO-2,4-DIBROMOANTHRAQUINONE 

(CAS NO. 81-49-2) 

IN F344/N RATS AND B6C3F1 MICE 

(FEED STUDIES) 

U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES 

Public Health Service 

National Institutes of Health 
FOREWORD

The National Toxicology Program (NTP) is made up of four charter agencies of the U.S. Department of

Health and Human Services (DHHS): the National Cancer Institute (NCI), National Institutes of Health; the

National Institute of Environmental Health Sciences (NIEHS), National Institutes of Health; the National

Center for Toxicological Research (NCTR), Food and Drug Administration; and the National Institute for

Occupational Safety and Health (NIOSH), Centers for Disease Control. In July 1981, the Carcinogenesis

Bioassay Testing Program, NCI, was transferred to the NIEHS. The NTP coordinates the relevant programs,

staff, and resources from these Public Health Service agencies relating to basic and applied research and to

biological assay development and validation.

The NTP develops, evaluates, and disseminates scientific information about potentially toxic and hazardous

chemicals. This knowledge is used for protecting the health of the American people and for the primary

prevention of disease.

The studies described in this Technical Report were performed under the direction of the NIEHS and were

conducted in compliance with NTP laboratory health and safety requirements and must meet or exceed all

applicable federal, state, and local health and safety regulations. Animal care and use were in accordance

with the Public Health Service Policy on Humane Care and Use of Animals. The prechronic and chronic

studies were conducted in compliance with Food and Drug Administration (FDA) Good Laboratory Practice

Regulations, and all aspects of the chronic studies were subjected to retrospective quality assurance audits

before being presented for public review.

These studies are designed and conducted to characterize and evaluate the toxicologic potential, including

carcinogenic activity, of selected chemicals in laboratory animals (usually two species, rats and mice).

Chemicals selected for NTP toxicology and carcinogenesis studies are chosen primarily on the bases of

human exposure, level of production, and chemical structure. The interpretive conclusions presented in this

Technical Report are based only on the results of these NTP studies. Extrapolation of these results to other

species and quantitative risk analyses for humans require wider analyses beyond the purview of these studies.

Selection per se is not an indicator of a chemical's carcinogenic potential. 

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NTP TECHNICAL REPORT

ON THE

TOXICOLOGY AND CARCINOGENESIS

STUDIES OF 

1-AMINO-2,4-DIBROMOANTHRAQUINONE

(CAS NO. 81-49-2)

IN F344/N RATS AND B6C3F1 MICE

(FEED STUDIES)

NATIONAL TOXICOLOGY PROGRAM

P.O. Box 12233

Research Triangle Park, NC 27709

August 1996

NTP TR 383

NIH Publication No. 96-2838

U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES

Public Health Service

National Institutes of Health
2 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

CONTRIBUTORS

National Toxicology Program

Evaluated and interpreted results and reported findings

J.E. Huff, Ph.D., Study Scientist

G.A. Boorman, D.V.M., Ph.D.

D.A. Bridge, B.S.

J.R. Bucher, Ph.D.

M.R. Elwell, D.V.M., Ph.D.

T.J. Goehl, Ph.D.

J.K. Haseman, Ph.D.

G.N. Rao, D.V.M., Ph.D.

G.S. Travlos, D.V.M.

D.B. Walters, Ph.D.

K.L. Witt, M.S., Oak Ridge Associated Universities

EG&G Mason Research Institute

Conducted studies, evaluated pathology findings

H.S. Lilja, Ph.D., Principal Investigator

A.J. Block, Ph.D.

H.J. Esber, Ph.D.

R.W. Fleischman, D.V.M.

M. Hagopian, Ph.D.

C.F. Moyer, D.V.M.

L.E. Sendelbach, Ph.D.

Experimental Pathology Laboratories, Inc.

Provided pathology quality assurance

J.F. Hardisty, D.V.M., Principal Investigator

S. Botts, D.V.M.

H.R. Brown, D.V.M., M.S.

Dynamac Corporation

Prepared quality assurance audits

S. Brecher, Ph.D., Principal Investigator

Analytical Sciences, Inc.

Provided statistical analyses

R.W. Morris, Ph.D., Principal Investigator

N.G. Mintz, B.S.

S. Rosenblum, M.S.

NTP Pathology Working Group

Evaluated slides, prepared pathology report on rats 

(25 January 1991)

R.M. Kovatch, D.V.M., Chair

Pathology Associates, Inc.

T. Anderson, D.V.M., Ph.D.

Hoffmann-LaRoche, Inc.

H.R. Brown, D.V.M., M.S.

Experimental Pathology Laboratories, Inc.

J. Eighmy, D.V.M.

USUHS, Bethesda, MD

J.R. Hailey, D.V.M.

National Toxicology Program

M.P. Jokinen, D.V.M.

National Toxicology Program

M.M. McDonald, D.V.M., Ph.D.

National Toxicology Program

Evaluated slides, prepared pathology report on mice 

(23 January 1991)

R.M. Sauer, V.M.D., Chair

PATHCO, Inc.

S. Botts, D.V.M.

Experimental Pathology Laboratories, Inc.

R. Cattley, V.M.D., Ph.D.

Chemical Industry Institute of Toxicology

C.K.S. Elangbam, Ph.D., Observer

Oklahoma State University

J.R. Hailey, D.V.M.

National Toxicology Program

M.P. Jokinen, D.V.M.

National Toxicology Program

M.M. McDonald, D.V.M., Ph.D.

National Toxicology Program

Biotechnical Services, Inc.

Prepared Technical Report

D.D. Lambright, Ph.D., Principal Investigator

S.R. Gunnels, M.A.

M.J. Nicholls, B.S.

K.L. Shaw, B.A.
3

CONTENTS

ABSTRACT ................. ................. ................. ................. ......... 5

EXPLANATION OF LEVELS OF EVIDENCE OF CARCINOGENIC ACTIVITY ................. . 13

TECHNICAL REPORTS REVIEW SUBCOMMITTEE ................. ................. ...... 14

SUMMARY OF TECHNICAL REPORTS REVIEW SUBCOMMITTEE COMMENTS .............. 15

INTRODUCTION ................. ................. ................. ................. .... 17

MATERIALS AND METHODS ................. ................. ................. .......... 21

RESULTS ................. ................. ................. ................. ........... 31

DISCUSSION AND CONCLUSIONS ................. ................. ................. ..... 81

REFERENCES ................. ................. ................. ................. ....... 85

APPE ND IX A Summary of Lesions in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone ................. ................. ...... 93

APPE ND IX B Summary of Lesions in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone ................. ................. ...... 151

APPE ND IX C Summary of Lesions in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone ................. ................. ...... 205

APPE ND IX D Summary of Lesions in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone ................. ................. ...... 249

APPE ND IX E Summary of Lesions in Male Rats in the Stop-Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone ................. ................. ...... 289

APPE ND IX F Summary of Lesions in Female Rats in the Stop-Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone ................. ................. ...... 307

APPE ND IX G Genetic Toxicology ................. ................. ................. ..... 325

APPE ND IX H Organ Weights and Organ-Weight-to-Body-Weight Ratios ................. ...... 335

APPE ND IX I Chemical Characterization and Dose Formulation Studies ................. ....... 343

APPENDIX J Feed and Compound Consumption in the 2-Year Feed Studies 

of 1-Amino-2,4-dibromoanthraquinone ................. ................. ...... 353

APPE ND IX K Ingredients, Nutrient Composition, and Contaminant Levels

in NIH-07 Rat and Mouse Ration ................. ................. ........... 361

APPE ND IX L Sentinel Animal Program ................. ................. ................. 367
4 1-Amino-2,4-dibromoanthraquinone, NTP TR 383
5

ABSTRACT

1-AMINO-2,4-DIBROMOANTHRAQUINONE

CAS No. 81-49-2

Chemical Formula: C14H7Br2NO2 Molecular Weight: 381.04

Synonym: ADBAQ 

1-Amino-2,4-dibromoanthraquinoneisananthraquinonederivedvatdye, a member of a class of insolubledyes

that are impregnated into textile fibers. Five

anthraquinone-derived dyes with representative and

diverse structures, as well as the parent chemical,

anthraquinone,wereselectedfortoxicology andcarcinogenesis evaluation. Similartothebenzidinedye initiative, the rationale for selecting these vat dyes was to

generatesufficienttoxicologicdatatopermitmorereliable

predictionsof carcinogenicitytobemadeonotherchemicalsinthisclass,therebyeliminatingorreducingtheneed

to study every anthraquinone dye. 1-Amino2,4-dibromoanthraquinone is the last anthraquinonederived dye in this group to be studied.

Groups of male and female F344/Nrats and B6C3F1

micewere exposed to 1-amino-2,4-dibromoanthraquinone (87% to 97% pure) for 13 weeks or for 9, 15,

or 24 months. Because 1-amino-2,4-dibromoanthraquinonewaspredictedtobe carcinogenic,thesestudies

were designed to evaluate the potential for tumor

progression and regression. Absorptionandexcretion

studieswere carried out in male F344/Nrats. Genetic

toxicity was determined in vitro using Salmonella

typhimurium and cultured Chines e

hamsterovarycells. Extensive chemical analyseswere

performedtoidentifyand characterize impuritiesofthe

1-amino-2,4-dibromoanthraquinoneusedinthesestudies.

13-W EEK S T U DY IN R A T S

Groupsof10maleand10female ratsweregiven0,2,500,

5,000, 10,000, 25,000, or 50,000 ppm 1-amino2,4-dibromoanthraquinone in feed for 13weeks. These

levels correspond to approximately 150 to 3,200 mg

1-amino-2,4-dibromoanthraquinone/kgbodyweightper

dayformales and to approximately170 to 3,200mg/kg

for females. Chemical-relatedmortalitywaslimitedto

onemaleandonefemaleinthe50,000ppmgroups. Final

meanbodyweights andbodyweightgainsof allexposed

groupsofratswere significantlylowerthanthoseofthe

controls. Feed consumptionbyall exposed groupswas

less than that by the controls throughout the study and

generallydecreasedwithincreasingexposureconcentration. Pink-redstainingofthe fur andtailwasobservedin

allexposedgroups. Absoluteandrelative liverweightsof

all exposedgroupsweregenerallysignificantlygreater

than those of the controls.
6 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

Chemical-related lesions were present in the liver,

kidney, and spleen of male and female rats. Nonneoplasticlesionsinthe liverincludedfociofhepatocellular

alteration, diffuse hepatocellular hypertrophy (cytomegaly),hepatocellular cytoplasmic vacuolation,bile

duct hyperplasia, inflammation, and pigmentation.

Thesedifferenceswereobservedprimarilyinthe25,000

and 50,000 ppm groups of males and females; the

spectrumofproliferative lesionsofthebileducts(hyperplasia,fibrosis,andnecrotizingcholangitis)inthe25,000

and50,000ppmgroupswasmorphologicallyconsistent

withthe lesiondescribedas cholangiofibrosis. Pigmentationwas present in the renal tubuleepitheliumof all

groups of exposed rats; nuclear enlargement

(karyomegaly) was also present in the renal tubule

epitheliuminsome ofthe exposedrats. Accumulationof

hyaline droplets in the cytoplasm of the renal tubule

epithelium and tubule lumina was present in 2,500,

5,000, 10,000, and 25,000 ppm males. Incidences of

hematopoiesisofthe spleeninexposedgroupsofmales

and females were increased compared to those in the

controls. 

13-WEEK STUDY IN MICE

Groups of 10 male and 10 female mice were given 0,

2,500,5,000,10,000,25,000,or 50,000ppm1-amino2,4-dibromoanthraquinone infeedfor13weeks. These

levelscorrespond to approximately 500 to 10,600mg

1-amino-2,4-dibromoanthraquinone/kgbodyweightper

dayformales and approximately660 to 11,700mg/kg

per day for females. There was no chemical-related

mortality. Feed consumption and final mean body

weights of exposedgroupswere similarto those ofthe

controls. Red staining of the fur was observed in all

exposedgroups. Absolute andrelative liverweightsof

the exposed groups were greater than those of the

controls except for the absolute liver weight of

2,500ppmmales. Absolute andrelativekidneyweights

of25,000and50,000ppmmaleswere lowerthanthose

of the controls. 

Chemical-related lesions were limitedto the livers of

malesandconsistedofpigmentationofhepatocytesatall

exposureconcentrationsandcentrilobularhepatocellular

hypertrophyat10,000,25,000, and50,000ppm. Minimalpigmentwaspresentinthe liverofonefemale inthe

25,000ppmgroupandinone female inthe50,000ppm

group.

2-YEAR STUDY IN RATS

Groupsof70maleand70femaleratsweregiven0,5,000,

or10,000ppm1-amino-2,4-dibromoanthraquinoneinfeed

for 103 weeks. In addition, groups of 50 male and 50

female rats were given 2,000 ppm 1-amino2,4-dibromoanthraquinoneinfeedfor104weeks. These

exposureconcentrationswereapproximatelyequalto90,

240,or490mg1-amino-2,4-dibromoanthraquinone/kg

body weight for males and 110, 285, or 600 mg/kg for

females. Tenanimalsfromeachgroupwereevaluatedfor

histopathology at 9 months. Additional groups of

10 animals from the 0 and 10,000 ppm groups were

evaluated for histopathology at 15 months.

Survival, Body Weights, Feed Consumption,

and Clinical Findings

Inthe2-yearstudy,survivalofthe10,000ppmmalesand

femaleswassignificantlylowerthanthatofthe controls.

Survivalofthe 2,000and5,000ppmgroupswassimilar

to that ofthe controls. During the last year ofthe study,

the mean body weights of exposed males were 80% to

91% those of controls, and the mean body weights of

exposed females were 67% to 84% those of controls.

Feedconsumptionamongexposedgroupswasgenerally

similar, but wasless than that by controls. The fur and

urineof all exposedmaleandfemalegroupswerediscolored. 

Pathology Findings

Inthe2-yearstudy, 1-amino-2,4-dibromoanthraquinone

wasassociatedwithsignificantchemical-relatedincreases

in the incidencesof benign andmalignant neoplasmsin

the liver, large intestine, kidney, and urinarybladder of

males and females. Chemical-related nonneoplastic

proliferative and degenerative lesions occurred in the

liver,kidney, urinarybladder, andforestomachofmales

and females.

The incidences offoci of hepatocellular alteration and

pigmentation in the liver of males and females were

increased at the 9-month interim evaluation, and a

hepatocellular adenoma was presentin one 5,000 ppm

male. At the 15-month interim evaluation,

hepatocellular adenoma or carcinoma (combined)

occurred in all males and nine females in the

10,000 ppm groups. By the end of the 2-yearstudy,

hepatocellular adenoma, carcinoma, cholangioma, or

cholangiocarcinoma were observed in males and

 
1-Amino-2,4-dibromoanthraquinone, NTP TR 383 7

females in the 5,000 and 10,000 ppm groups. In the

2,000ppmgroups,similarliverneoplasmswerepresent

in 63% of themales and in 83%ofthe females. Of the

hepatocellularcarcinomasinthe5,000and10,000ppm

groups ofmales and females,31% to 49% were metastatic to the lungs or other sites. Increases in the incidences offoci of hepatocellular alteration(basophilic,

eosinophilic,andclear cell)andpigmentationofthe liver

were also observed in exposed groups of males and

females. 

Adenomatouspolyps(adenoma)ofthe large intestine

werepresentin six10,000ppmmales at the 15-month

interimevaluation. Incidences of adenomatouspolyp

(adenoma) and carcinoma of the large intestine were

significantlyincreasedinexposedgroupsofmalesand

females after 2 years; multiple benign and malignant

intestinal neoplasms were observed in many of these

rats. 

In the kidney, incidencesofrenaltubule adenoma and

carcinoma were significantly increased in exposed

groupsofmalesandfemalesafter2years. Renaltubule

adenomas were present in two 10,000 ppm males at

15months. Therewerealsochemical-relatedincreases

inthe incidences andseveritiesofrenaltubuleepithelial

hyperplasia,pigmentation,andtransitionalcellhyperplasia in the kidneyofmalesandfemales. Hyalinedroplet

accumulation was present in all exposed male rats at

9 months.

Incidencesoftransitional cellpapilloma andcarcinoma

oftheurinarybladderwereincreasedat2yearsinmales

andfemalesinthe10,000ppmgroups. Transitional cell

hyperplasiawasobservedinexposedmalesandfemales

atthe15-monthinterimevaluation. Othernonneoplastic

lesions observed in the urinary bladder at 2 years included metaplasia of the transitional epithelium and

submucosal stromal tissue.

In the forestomach, the incidences and severities of

inflammation,ulceration,hyperkeratosis, andhyperplasia of the squamous mucosa were increased in all

exposedgroupsofmales andfemalesat2years,butnot

at the 9- or 15-month interim evaluations.

In exposed males and females, the incidences of

mononuclearcellleukemiaweresignificantlydecreased.

The incidencesof atrophyofthe seminal vesiclewere

increased in exposed male rats in the 2-year study.

Stop-Exposure Evaluation in Rats

Groups of 40 male and 40 female rats were given

20,000ppm1-amino-2,4-dibromoanthraquinone infeed

for 9 or 15 months. At 9 months, 10 males and 10 femaleswereevaluatedforhistopathology (9-monthinterim

evaluation groups). After 9 months of exposure, an

additional10males and10femaleswerefedcontroldiet

until the end of the 15-month evaluation (9-month

stop-exposure groups), and 20 males and 20 females

continuedtoreceive20,000ppm1-amino-2,4-dibromoanthraquinoneuntilthe endofthe evaluation(15-month

exposuregroups). The approximatedailyconsumptionof

1-amino-2,4-dibromoanthraquinonewas1,335mg/kgfor

malesand1,790mg/kgforfemalesin the 9-monthstopexposure groups and 1,115 mg/kg for males and

1,435 mg/kg for females in the 15-month exposure

groups.

Survivalwassimilaramonggroupsexceptforthe females

inthe15-monthexposuregroup;thesurvivalofthisgroup

was lower than that of the controls. Lower mean body

weightswererelatedtoincreasedexposureduration. The

meanbodyweightsof exposedmaleswere76%to 82%

that of controls, and themeanbody weights of exposed

females were 73% to 84% that of controls. 

Forthestop-exposureevaluation,similarchemical-related

neoplasmsandnonneoplasticlesionswereobservedinthe

same sites as in the 2-year study: liver, large intestine,

kidney, urinary bladder, and forestomach.

After 9monthsof dietaryexposure toa concentrationof

20,000 ppm 1-amino-2,4-dibromoanthraquinone,

hepatocellularadenoma andcarcinomaoccurredinmales

andfemales. Nonneoplastic chemical-relatedlesionsin

the liver of exposed rats included pigmentation, focal

hepatocellular alteration, and bile duct hyperplasia.

Neoplasms atothersitesinmalesincludedoneadenomatous polyp (adenoma) in the large intestine and one

transitionalcellpapilloma intheurinarybladder. Hyaline

droplet accumulation was present in the kidney of exposed males at 9 months. 

In the stop-exposure groups examined at 15 months,

hepatocellular adenoma and carcinoma werepresentin

most males and females. Adenomatous polyp

(adenoma)ofthe colon,renaltubulecell adenoma,and

urinary bladder transitional cell papilloma and
8 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

carcinoma also occurred in males and females.

Nonneoplasticchemical-relatedlesionsincludedfociof

hepatocellularalteration in the liver andhyperplasiaof

the renaltubuleepitheliumand urinarybladdertransitional epithelium. Hyperplasia,hyperkeratosis,inflammation, and ulceration were observed in the

forestomachsofsomemaleandfemale ratscontinuously

exposed for 15 months.

2-YEAR STUDY IN MICE

Groups of 60 male and 60 female mice were given 0,

10,000, or 20,000 ppm 1-amino -

2,4-dibromoanthraquinone infeedfor104weeks. The

dailycompoundconsumptionwasapproximately1,690

or3,470mg1-amino-2,4-dibromoanthraquinone/kgbody

weightformalesand1,950or4,350mg/kgforfemales.

Ten animals from each group were evaluated for

histopathology at 15 months.

Survival, Body Weights, Feed Consumption,

and Clinical Findings

In the 2-year study, survival of exposed males was

significantlylowerthanthatofthe controls. Survivalof

exposedfemaleswassimilartothatofthe controls. The

finalmeanbodyweightsof exposedmaleswere83%to

85%thatof controls,andthe finalmeanbodyweightsof

exposed females were 81% to 86% that of controls.

Feed consumption by exposed groups was generally

similar to that by controls. Discoloration of the fur,

urine, and feces was observed in all exposed groups. 

Pathology Findings

Inthe2-yearstudy, 1-amino-2,4-dibromoanthraquinone

was associated with significant chemical-related increases in the incidences of benign and malignant

neoplasmsinthe liver,forestomach,and lungofmales

and females.

Incidencesof hepatocellular adenoma and carcinoma

were increased in exposed groups at the 15-month

interimevaluationandat2years. At 2years,therewere

significant increases in the incidences of multiple

hepatocellular adenoma and carcinoma in males and

females and in the incidences of hepatoblastoma in

males. Centrilobular hypertrophy of hepatocytes in

malesandfociofhepatocellularalterationandpigmentation in the liver of males and females were also

chemical-related changes.

Squamous cell papilloma of the forestomach mucosa

occurred in 10,000ppmfemales and 20,000ppmmales

andfemales atthe 15-monthinterimevaluation, andthe

incidences of squamous cell papilloma and carcinoma

were significantlyincreasedinexposedgroupsofmales

andfemales at 2 years. Chemical-relatedhyperplasiaof

forestomachepitheliumwasalsopresentat15monthsand

at 2 years.

Alveolar/bronchiolaradenomaswerepresentonlyinthe

exposedgroups ofmales and females at 15months,and

the incidences of alveolar/bronchiolar adenoma were

significantlyincreased in exposedmalesandfemales at

2years. The incidencesofmultiplealveolar/bronchiolar

adenoma were also increased in exposed males. 

In the kidney, pigmentation was present in the renal

tubules of most mice after 2 years of exposure.

DISPOSITION

AND METABOLISM STUDIES

Adult male F344/N rats were given [14C]-labeled

1-amino-2,4-dibromoanthraquinone as a singleintravenous dose of 0.4 mg/kg body weight or as a single oral

dose of 2, 23, 118, 814, or 1,473 mg/kg. A 6-hour bile

cannulation study was also performed. From day 0

throughday3afterintravenousadministration,about50%

ofthe 14Cwas excretedinthe feces,15%intheurine, and

6% in expired air. Unmetabolized 1-amino2,4-dibromoanthraquinone accountedforlessthan3%of

the excreted14Cafterintravenousadministration. Fororal

doses administered, the amount of the dose that was

absorbed fit the equation: absorbed dose = 6.6 

log(dose). Afterintravenousadministration,themetabolitesof1-amino-2,4-dibromoanthraquinoneinbloodwere

primarilyin the plasma fraction (blood:plasma ratio of

approximately0.5:1). The highest concentrationsof 14C

in tissues 15 minutes after intravenous dosing were in

excretory organs, lung, kidney, small intestine, liver,

adipose tissue, and adrenal gland. 

GENETIC TOXICOLOGY

1-Amino-2,4-dibromoanthraquinone was mutagenic

inSalmonellatyphimuriumstrainsTA98andTA1537in

the absence of S9; with S9, an equivocal response was

observedinTA1537. 1-Amino-2,4-dibromoanthraquinone

resulted in an equivocal response in
1-Amino-2,4-dibromoanthraquinone, NTP TR 383 9

strain TA100 with and without S9, and no mutagenic

activitywas detected with strainTA1535. In cultured

Chinese hamster ovary cells, 1-amino2,4-dibromoanthraquinone induced sister chromatid

exchangeswithandwithoutS9; chromosomal aberrations were induced only in the absence of S9. 

CONCLUSIONS

Underthe conditionsofthese2-yearfeedstudies,there

was clear evidence of carcinogenic activity* of

1-amino-2,4-dibromoanthraquinoneinmaleandfemale

F344/Nratsbasedonincreasedincidencesofneoplasms

inthe liver,large intestine,kidney, andurinarybladder.

There was clear evidence of carcinogenic activity of

1-amino-2,4-dibromoanthraquinoneinmaleandfemale

B6C3F1 mice based on increased incidences of

neoplasms in the liver, forestomach, and lung.

Exposure of male and female rats to 1-amino2,4-dibromoanthraquinonefor2yearswasassociatedwith

basophilic focus (males only), clear cell focus,

eosinophilic focus, and pigmentationin the liver;renal

tubulehyperplasia,renaltubulepigmentation,andtransitional cell hyperplasia in the kidney; transitional cell

hyperplasia,squamousmetaplasia,andstromalmetaplasia

(femalesonly)intheurinarybladder;squamoushyperplasia, hyperkeratosis,ulceration, andinflammationofthe

forestomach mucosa; and seminal vesicle atrophy.

Exposure of male and female mice to 1-amino2,4-dibromoanthraquinonefor2yearswasassociatedwith

centrilobularhepatocellularhypertrophy(malesonly),

basophilic focus, clear cell focus (females only),

eosinophilicfocus,andpigmentationinthe liver;pigmentationinthekidney; andhyperplasia,basal cellhyperplasia,hyperkeratosis, andinflammationofthe forestomach

mucosa. 

__________

*

Explanation of Levels of Evidence of Carcinogenic Activity is on page 13. A summary of the Technical Reports Review

 Subcommittee comments and the public discussion on this Technical Report appears on page 15.
10 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

Font Size:11ptSummary of the 2-Year Carcinogenesis and Genetic Toxicology Studies 

of 1-Amino-2,4-dibromoanthraquinone

Male

F344/N Rats

Female

F344/N Rats

Male 

B6C3F1 Mice

Female

B6C3F1 Mice

Doses 0, 2,000, 5,000, or

10,000 ppm

[approximately 

90, 240, or

490 mg/kg/day]

0, 2,000, 5,000, or

10,000 ppm

[approximately 

110, 285, or

600 mg/kg/day]

0, 10,000, or 20,000

ppm

[approximately 

1,690 or

3,470 mg/kg/day]

0, 10,000, or

20,000 ppm

[approximately 

1,950 or

4,350 mg/kg/day]

Body weights Exposed groups lower

than controls

Exposed groups lower

than controls

Exposed groups lower

than controls

Exposed groups lower

than controls

2-Year survival

rates

26/50, 24/40, 21/60,

10/50

38/50, 32/40, 38/60,

12/49

40/50, 22/51, 23/50 39/50, 34/50, 33/50

Nonneoplastic

effects

Liver: basophilic

focus (9/50, 12/40,

24/59, 22/50); clear

cell focus (3/50,

26/40, 39/59, 27/50);

eosinophilic focus

(1/50, 13/40, 14/59,

6/50); pigmentation

(3/50, 19/40, 48/59,

39/50)

Kidney: renal tubule

hyperplasia (9/50,

30/40, 25/59, 19/50);

renal tubule

pigmentation (5/50,

40/40, 58/59, 49/50);

transitional cell

hyperplasia (30/50,

40/40, 51/59, 35/50)

Urinary bladder:

transitional cell

hyperplasia (1/50,

5/38, 17/58, 30/50);

squamous metaplasia

(0/50, 0/38, 0/58,

3/50)

Liver: clear cell focus

(3/50, 28/40, 39/60,

17/48); eosinophilic

focus (7/50, 23/40,

12/60, 1/48);

pigmentation (1/50,

19/40, 51/60, 45/48)

Kidney: renal tubule

hyperplasia (1/50,

12/40, 23/60, 27/48);

renal tubule

pigmentation (0/50,

40/40, 60/60, 48/48);

transitional cell

hyperplasia (10/50,

16/40, 44/60, 21/48)

Urinary bladder:

transitional cell

hyperplasia (1/50,

2/40, 41/60, 41/46);

squamous metaplasia

(0/50, 1/40, 4/60,

8/46); stromal

metaplasia (0/50,

0/40, 4/60, 2/46)

Liver: centrilobular

hepatocyte

hypertrophy (0/50,

17/51, 13/50);

basophilic focus

(0/50, 4/51, 3/50);

eosinophilic focus

(0/50, 6/51, 1/50);

pigmentation (1/50,

50/51, 47/50)

Kidney: renal tubule

pigmentation (0/50,

42/51, 43/50)

Forestomach:

hyperplasia (1/50,

9/50, 4/50); basal cell

hyperplasia (0/50,

0/50, 2/50);

hyperkeratosis (1/50,

7/50, 6/50);

inflammation (2/50,

6/50, 13/50)

Liver: basophilic

focus (0/50, 4/50,

5/50); clear cell focus

(0/50, 10/50, 9/50);

eosinophilic focus

(0/50, 4/50, 2/50);

pigmentation (0/50,

44/50, 49/50)

Kidney: renal tubule

pigmentation (0/50,

43/50, 43/50)

Forestomach:

hyperplasia (9/48,

15/50, 19/50); basal

cell hyperplasia (0/48,

7/50, 3/50);

hyperkeratosis (10/48,

14/50, 17/50);

inflammation (7/48,

10/50, 21/50)

(continued)
1-Amino-2,4-dibromoanthraquinone, NTP TR 383 11

Summary of the 2-Year Carcinogenesis and Genetic Toxicology Studies 

of 1-Amino-2,4-dibromoanthraquinone (continued)

Male

F344/N Rats

Female

F344/N Rats

Male 

B6C3F1 Mice

Female

B6C3F1 Mice

Nonneoplastic

effects

(continued)

Forestomach: 

squamous hyperplasia

(3/49, 19/39, 25/59,

26/49); hyperkeratosis

(5/49, 18/39, 21/59,

20/49); ulcer (3/49,

10/39, 15/59, 16/49);

inflammation (3/49,

12/39, 11/59, 11/49);

Seminal vesicle:

atrophy (1/49, 30/40,

35/59, 23/50)

Forestomach: 

squamous hyperplasia

(2/49, 7/40, 26/60,

33/47); hyperkeratosis

(2/49, 7/40, 23/60,

28/47); ulcer (1/49,

2/40, 7/60, 17/47);

inflammation (0/49,

1/40, 13/60, 10/47)

Neoplastic effects Liver: hepatocellular

adenoma (1/50,

20/40, 40/59, 34/50);

hepatocellular

carcinoma (1/50,

12/40, 55/59, 46/50);

hepatocholangiocarcinoma (0/50,

0/40, 6/59, 2/50)

Large intestine (all

sites): adenomatous

polyp (adenoma)

(0/50, 13/40, 51/59,

40/50); carcinoma

(0/50, 1/40, 11/59,

17/50)

Kidney (renal tubule):

adenoma (2/50,

10/40, 11/59, 14/50);

carcinoma (0/50,

0/40, 2/59, 1/50)

Urinary bladder:

transitional cell

papilloma (0/50, 1/38,

2/58, 8/50);

transitional cell

carcinoma (0/50,

0/38, 1/58, 4/50)

Liver: hepatocellular

adenoma (0/50,

28/40, 47/60, 29/48);

hepatocellular

carcinoma (0/50,

12/40, 57/60, 45/48);

hepatocholangiocarcinoma (0/50,

0/40, 11/60, 13/48)

Large intestine (all

sites): adenomatous

polyp (adenoma)

(0/50, 28/40, 53/60,

43/49); carcinoma

(0/50, 2/40, 21/60,

8/49)

Kidney (renal tubule):

adenoma (0/50, 3/40,

16/60, 16/48);

carcinoma (0/50,

0/40, 0/60, 2/48)

Urinary bladder:

transitional cell

papilloma (0/50, 2/40,

7/60, 9/46);

transitional cell

carcinoma (0/50,

0/40, 8/60, 16/46)

Liver: hepatocellular

adenoma (10/50,

38/51, 39/50);

hepatocellular

carcinoma (9/50,

18/51, 21/50);

hepatoblastoma (0/50,

3/51, 5/50)

Forestomach:

squamous cell

papilloma (0/50,

13/51, 16/50); 

squamous cell

carcinoma (0/50,

12/51, 13/50)

Lung: alveolar/

bronchiolar adenoma

(7/50, 26/51, 24/50)

Liver: 

hepatocellular

adenoma (6/50,

45/50, 49/50);

hepatocellular

carcinoma (0/50,

23/50, 27/50)

Forestomach:

squamous cell

papilloma (2/50,

16/50, 27/50);

squamous cell

carcinoma (0/50,

12/50, 11/50)

Lung: alveolar/

bronchiolar adenoma

(4/50, 17/50, 13/49)

(continued)
12 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

Summary of the 2-Year Carcinogenesis and Genetic Toxicology Studies 

of 1-Amino-2,4-dibromoanthraquinone (continued)

Male

F344/N Rats

Female

F344/N Rats

Male 

B6C3F1 Mice

Female

B6C3F1 Mice

Level of evidence

of carcinogenic

activity

Clear evidence Clear evidence Clear evidence Clear evidence

Genetic toxicology

Salmonella typhimurium gene mutation: Equivocal in strain TA100 with and without S9; negative in strain TA1535 with and without S9;

positive in strain TA1537 without S9, equivocal with S9; positive in strain TA98 without S9,

negative with S9 

Sister chromatid exchanges

Cultured Chinese hamster ovary cells in vitro: Positive with and without S9 (combined results from testing in two laboratories) 

Chromosomal aberrations

Cultured Chinese hamster ovary cells in vitro: Weakly positive without S9, negative with S9 
1-Amino-2,4-dibromoanthraquinone, NTP TR 383 13

EXPLANATION OF LEVELS OF EVIDENCE OF CARCINOGENIC ACTIVITY

The National Toxicology Program describes the results of individual experiments on a chemical agent and notes the strength of the

evidence for conclusions regarding each study. Negative results, in which the study animals do not have a greater incidence of

neoplasia than control animals, do not necessarily mean that a chemical is not a carcinogen, inasmuch as the experiments are

conducted under a limited set of conditions. Positive results demonstrate that a chemical is carcinogenic for laboratory animals under

the conditions of the study and indicate that exposure to the chemical has the potential for hazard to humans. Other organizations,

such as the International Agency for Research on Cancer, assign a strength of evidence for conclusions based on an examination of all

available evidence, including animal studies such as those conducted by the NTP, epidemiologic studies, and estimates of exposure. 

Thus, the actual determination of risk to humans from chemicals found to be carcinogenic in laboratory animals requires a wider

analysis that extends beyond the purview of these studies.

Five categories of evidence of carcinogenic activity are used in the Technical Report series to summarize the strength of the evidence

observed in each experiment: two categories for positive results (clear evidence and some evidence); one category for uncertain

findings (equivocal evidence); one category for no observable effects (no evidence); and one category for experiments that cannot

be evaluated because of major flaws (inadequate study). These categories of interpretative conclusions were first adopted in June

1983 and then revised in March 1986 for use in the Technical Report series to incorporate more specifically the concept of actual

weight of evidence of carcinogenic activity. For each separate experiment (male rats, female rats, male mice, female mice), one of the

following five categories is selected to describe the findings. These categories refer to the strength of the experimental evidence and

not to potency or mechanism.

• Clear evidence of carcinogenic activity is demonstrated by studies that are interpreted as showing a dose-related 

(i) increase of malignant neoplasms, (ii) increase of a combination of malignant and benign neoplasms, or (iii) marked

increase of benign neoplasms if there is an indication from this or other studies of the ability of such tumors to progress to

malignancy.

• Some evidence of carcinogenic activity is demonstrated by studies that are interpreted as showing a

chemical-related increased incidence of neoplasms (malignant, benign, or combined) in which the strength of the

response is less than that required for clear evidence.

• Equivocal evidence of carcinogenic activity is demonstrated by studies that are interpreted as showing a marginal

increase of neoplasms that may be chemical related.

• No evidence of carcinogenic activity is demonstrated by studies that are interpreted as showing no chemical-related

increases in malignant or benign neoplasms.

• Inadequate study of carcinogenic activity is demonstrated by studies that, because of major qualitative or

quantitative limitations, cannot be interpreted as valid for showing either the presence or absence of carcinogenic

activity.

When a conclusion statement for a particular experiment is selected, consideration must be given to key factors that would extend the

actual boundary of an individual category of evidence. Such consideration should allow for incorporation of scientific experience and

current understanding of long-term carcinogenesis studies in laboratory animals, especially for those evaluations that may be on the

borderline between two adjacent levels. These considerations should include:

• adequacy of the experimental design and conduct;

• occurrence of common versus uncommon neoplasia;

• progression (or lack thereof) from benign to malignant neoplasia as well as from preneoplastic to neoplastic lesions;

• some benign neoplasms have the capacity to regress but others (of the same morphologic type) progress. At present,

it is impossible to identify the difference. Therefore, where progression is known to be a possibility, the most prudent

course is to assume that benign neoplasms of those types have the potential to become malignant;

• combining benign and malignant tumor incidence known or thought to represent stages of progression in the same

organ or tissue;

• latency in tumor induction;

• multiplicity in site-specific neoplasia;

• metastases;

• supporting information from proliferative lesions (hyperplasia) in the same site of neoplasia or in other experiments

(same lesion in another sex or species);

• presence or absence of dose relationships;

• statistical significance of the observed tumor increase;

• concurrent control tumor incidence as well as the historical control rate and variability for a specific neoplasm;

• survival-adjusted analyses and false positive or false negative concerns;

• structure-activity correlations; and

• in some cases, genetic toxicology.
14 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

NATIONAL TOXICOLOGY PROGRAM BOARD OF SCIENTIFIC COUNSELORS

TECHNICAL REPORTS REVIEW SUBCOMMITTEE

The members of the Technical Reports Review Subcommittee who evaluated the draft NTP Technical Report on 1-amino2,4-dibromoanthraquinone on June 21, 1994, are listed below. Subcommittee members serve as independent scientists, not as

representatives of any institution, company, or governmental agency. In this capacity, subcommittee members have five major

responsibilities in reviewing NTP studies:

! to ascertain that all relevant literature data have been adequately cited and interpreted,

! to determine if the design and conditions of the NTP studies were appropriate,

! to ensure that the Technical Report presents the experimental results and conclusions fully and clearly,

! to judge the significance of the experimental results by scientific criteria, and

! to assess the evaluation of the evidence of carcinogenic activity and other observed toxic responses.

Arnold L. Brown, M.D., Chair

University of Wisconsin Medical School

Madison, WI

Paul T. Bailey, Ph.D.

Environmental and Health Sciences Laboratory

Mobil Oil Corporation

Princeton, NJ

Meryl H. Karol, Ph.D.

Department of Environmental Occupational Health

University of Pittsburgh

Pittsburgh, PA

Curtis D. Klaassen, Ph.D.

Department of Pharmacology and Toxicology

University of Kansas Medical Center

Kansas City, KS

Claudia S. Miller, M.D.

University of Texas Health Sciences Center

San Antonio, TX

Janardan K. Reddy, M.D., Principal Reviewer

Department of Pathology

Northwestern University Medical School

Chicago, IL

Irma Russo, M.D.

Fox Chase Cancer Center

Philadelphia, PA

Louise Ryan, Ph.D.

Division of Biostatistics

Harvard School of Public Health and

Dana-Farber Cancer Institute

Boston, MA

Robert E. Taylor, M.D., Ph.D.

Department of Pharmacology

Howard University College of Medicine

Washington, DC

Matthew J. van Zwieten, D.V.M., Ph.D., Principal Reviewer

Department of Safety Assessment

Merck Research Laboratories

West Point, PA

Mary Jo Vodicnik, Ph.D.

Lilly MSG Development Center

Belgium

Jerrold Ward, D.V.M., Ph.D., Principal Reviewer

National Cancer Institute

Frederick, MD 
1-Amino-2,4-dibromoanthraquinone, NTP TR 383 15

SUMMARY OF TECHNICAL REPORTS REVIEW SUBCOMMITTEE COMMENTS

On June 21, 1994, the draft Technical Report on the

toxicology and carcinogenesis studies of 1-amino--

2,4-dibromoanthraquinonereceivedpublicreviewbythe

National Toxicology Program's Board of Scientific

Counselors'TechnicalReportsReviewSubcommittee.

The reviewmeetingwasheldattheNationalInstituteof

EnvironmentalHealthSciences,ResearchTrianglePark,

NC.

Dr. J.E. Huff, NIEHS, introduced the toxicology and

carcinogenesisstudiesof1-amino-2,4-dibromoanthraquinone bydiscussingthe uses and rationale forstudy,

includingitbeingapartof a classstudyof anthraquinone

derivatives. He described the experimental design,

reportedonsurvivalandbodyweight effects, and commentedonchemical-relatedneoplasms andnonneoplastic

lesionsinmale and femaleratsandmice. The proposed

conclusions for the studies were clear evidence of

carcinogenic activityinmale and female F344/N rats

and in male and female B6C3F1 mice.

Dr.Huffreviewed the carcinogenicresponsesin other

anthraquinonederivativesthathadbeenstudied,noting

thatthe liverseemedtobeamajorsiteandthat1-amino2,4-dibromoanthraquinonewasthemostactive asfar as

thenumberofsites. Interpretive conclusionsthat could

be drawn from the cumulative National Toxicology

Programstudiesonthisclassofinsolubledyeswerethat

anthraquinonesare typicallymutagenicandclastogenic,

theyare carcinogenictomaleandfemale rats andmice,

and theyarepredicted to representlikely carcinogenic

hazardsto humansexposed to these agents, especially

occupationally. Dr.J.R.Bucher,NIEHS,reportedthat

the toxicology and carcinogenesis studies on

anthraquinone were in progress. 

Dr.vanZwieten, a principal reviewer,agreedwiththe

proposedconclusions. He thoughtthereshouldbemore

discussionofthe findingsfromthe stop-exposuregroups

ofrats. (Stop-exposure groupswere evaluatedat 9and

15months as part of an attempt to gain insightintothe

progressionorregressionof chemical-inducedlesions.)

Dr.vanZwietennotedthehigh impuritylevelsinthe first

lotofthe chemicalused forthe 13-weekstudies andfor

the first 2 months of the 2-year studies and said that a

 

statementindicatingthatthisdidnot affectthe integrity

ofthe studiesmight behelpful. Dr.Huffrespondedthat

the impuritieshadbeencharacterized(page 20;Arneson

et al., 1996).

Dr.Ward,the secondprincipalreviewer,agreedwiththe

proposedconclusions. He commented that no hyaline

droplets were reported in the kidney of rats after

9months,andsince1-amino-2,4-dibromoanthraquinone

might cause accumulation of "2:-globulin, the report

shouldindicatethat dropletswere searched for but not

found or found but not reported (page 83). Dr. Ward

objectedtocharacterizingcholangiofibrosisfoundinthe

liverofratsin a 13-week study as "premalignant." He

statedthatthislesionisusuallyinducedbylivercarcinogens but does not typically progress to bile duct neoplasms. Dr.M.R.Elwell,NIEHS,saidthisinterpretation

was from the literature and the wording on neoplastic

potential would be revised to also reflect Dr. Ward's

experience.

Dr.Reddy, the thirdprincipalreviewer,alsoagreedwith

the proposedconclusions. He said it would have been

usefultocharacterize the chemicalnatureofthepigment

that accumulatedinthe liver,kidney, andotherorgans,as

well as in the fur and tail. Dr. Huff responded that,

logically, the pigmentwas eitherthe chemicaloroneof

its metabolites, but the feasibility of going back and

defining it better would have to be determined.

Dr.Russohad observedevidenceof chronicinflammationinoneoftheplates andwonderedwhetherthe liver

lesionswere associatedwithhepatitis. Dr.Karol asked

iftherewasinflammationofthe eosinophilic foci,which

would suggest a hypersensitivity-type reaction.

Dr. Elwellsaidtherewassome inflammation with the

cholangiofibrosis,butthiswasreallylimitedtothe focal

lesionswhere therewasfibrosisandtocysticbileducts,

andtherewasnot aneosinophilicinflammation;the term

"eosinophilic foci" referredtoa focal cellular alteration

of hepatocytes. 

Dr.Baileycited a statement from the use, production,

and human exposure sections that "no individualized

information waslocated regardingamounts produced
16 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

or specific uses of 1-amino-2,4-dibromoanthraquinone," leading him to wonder if this chemical is currently used. Dr. Huffsaid this was a valid

question for 1-amino-2,4-dibromoanthraquinoneand

the other anthraquinone derivatives. He said proprietary information was difficult to obtain, although he

was hopeful that a request to the AmericanPharmaceuticalAssociationconcerninganthraquinonedyesin

over-the-counter or prescription items might yield

some data on human exposure. There was some

discussionthatprimaryexposure tothesedyeswouldbe

from topical application or exposure.

Dr. van Zwieten moved that the Technical Report on

1-amino-2,4-dibromoanthraquinonebeacceptedwiththe

revisions discussed and the conclusions as written for

male and female rats and mice, clear evidence of

carcinogenicactivity. Dr.Reddysecondedthemotion,

which was accepted unanimously with eleven votes.
17

INTRODUCTION

1-AMINO-2,4-DIBROMOANTHRAQUINONE

CAS No. 81-49-2

Chemical Formula: C14H7Br2NO2 Molecular Weight: 381.04

Synonym: ADBAQ 

CHEMICAL AND PHYSICAL PROPERTIES

1-Amino-2,4-dibromoanthraquinone, a reddish brown

to orange powder,isan anthraquinone-derived vatdye.

Anthraquinone (9,10-anthraquinone:CAS No.84-65-

1),whichdoes notoccur naturally,wasfirstsynthesized

by Laurent in 1835 as an oxidation product of anthracene and nitric acid (Chung, 1978). "Anthra" comes

fromtheGreekwordfor coal,from which anthracene

was originally obtained.

USE, PRODUCTION,

AND HUMAN EXPOSURE

Anthraquinone is an important and widelyused starting

materialforthemanufactureof vat dyes(Merck Index,

1989). Classhomologuesof anthraquinonecomprise a

greater numberof dyes having outstanding"fastness"

properties than any other group of commercial dyes

(Chung, 1978;Chung and Farris,1979). No informationwaslocated regarding amounts produced orspecific uses of 1-amino-2,4-dibromoanthraquinone. The

2-alkyl derivatives of anthraquinonewith alkyl chains

rangingfromone to fivecarbons aremost often used in

the dye industry (Chung, 1978).

Vat dyes area classofwater-insoluble dyesthat can be

easily reduced (i.e., vatted) to a water-soluble and

usuallycolorlessleucoforminwhich they can readily

impregnate fibers and textiles. Subsequent oxidation

thenproducesthe insoluble colored formthatisremarkably "fast" to washing, light, and chemicals. The

reducingagentsareusuallyalkalinesolutionsofsodium

hydrosulfite; oxidation takes place in the presence of

air,perborate, dichromate, and other agents(Hawley,

1981). Vat dyes areused typicallyforcotton, wool,and

cellulose acetate. Production of vat dyesin theUnited

States totaled 14,000,000kg (30.8 millionpounds)in

1991 (USITC,1993); these figuresdo not accountfor

the "large" amounts extracted frombotanical species

containing naturally occurring anthraquinones used

therapeutically and for other purposes.

ABSORPTION, DISTRIBUTION,

METABOLISM, AND EXCRETION

No informationonthe absorption,distribution,metabolism, and excretion of 1-amino-2,4-dibromoanthraquinone in experimental animals or in humans was

found in a search of the available literature.
18 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TOXICITY

No information on the toxicity of 1-amino-2,4-

dibromoanthraquinone in experimental animals or in

humanswasfound ina search ofthe available literature.

REPRODUCTIVE

AND DEVELOPMENTAL TOXICITY

No informationonthe reproductive and developmental

toxicity of 1-amino-2,4-dibromoanthraquinone in

experimental animals or in humans was found in a

search of the available literature.

CARCINOGENICITY

Experimental Animals

Chemicalsbelongingto the anthraquinone classofdyes

are carcinogenic to rodents(IARC, 1987;Sendelbach,

1989) and consistently induce neoplasms ofthe liver

(Huff et al., 1991). However, each anthraquinone

derivativeappears to induce cancer in other organs or

tissue sites as well (Huff et al., 1991).

For the five anthraquinonesevaluatedand reported by

NCI/NTP, the 2-year exposure concentrations in the

feed varied from a low of 300 ppm (0.03%)

for 2-methyl-1-nitroanthraquinone to a high of

20,000 ppm(2%)for1-amino-2,4-dibromoanthraquinone (Table 1).

Humans

No information on the potential carcinogenicity of

1-amino-2,4-dibromoanthraquinone in humans was

found in a search of the available literature.

GENETIC TOXICITY

All five anthraquinones evaluated and reported by

NCI/NTP induced mutations in Salmonella

typhimurium (Brown and Brown, 1976; Haworth

et al., 1983; Dunkel et al., 1985; Zeiger et al., 1988).

Each also caused sister chromatid exchanges and

chromosomalaberrationsin culturedChinesehamster

ovary cells (Anderson et al., 1990; Loveday et al.,

1990; NTP, unpublished). S9 activation was not

required for 1-amino-2,4-dibromoanthraquinone to

produce theseeffects. The parentcompound,anthraquinone, is also mutagenic in S.typhimurium;significant

increases in mutant colonies were observed with 

strains TA98andTA100with and without S9 (Zeiger

et al., 1988). In addition, anthraquinone and

1-aminoanthraquinone (250 mg/kg) werereported to

induceDNAstrand breaksin liver and kidneytissueof

male Swiss (CD-1®)mice following intraperitoneal

injection (Cesarone et al., 1982).

STUDY RATIONALE

The NCI selected and evaluated several of the

anthraquinone-derived dyes for a classstudyto determinewhetherthesedyes have anyinherentpotentialfor

carcinogenicity in laboratory rodents and, if so, in

humans aswell. The firstthree studieswereconducted

with 2-aminoanthraquinone (NCI, 1978a),

1-amino-2-methylanthraquinone (NCI, 1978b), and

2-methyl-1-nitroanthraquinone(NCI,1978c). Afourth

substance, 1,4,5,8-tetraaminoanthraquinone (C.I.

DisperseBlue1)wasselectedand evaluatedfor carcinogenicitybytheNTP(NTP,1986a). This Technical

Report addresses the fifth chemical in this class,

1-amino-2,4-dibromoanthraquinone. In addition,the

parent chemical, anthraquinone,has been selectedfor

study to complete the overall effort on these dyes.

Anthraquinone and the five substituted anthraquinones

(Figure1), representative of a large group of amino-,

alkyl-, and nitro-, or halogen-containing anthraquinones,werechosenfortoxicologic characterizationand

to establish some predictivestructure-activityrelationships that could be used on other dyesin this category

ratherthan testingeach and everyone. Other chemical

classes that have been likewise evaluated by the

NCI/NTP to reduce the need for "one-by-one" testing

include benzidine-based dyes (Morgan et al.,1994),

phthalates(Kluwe et al.,1982;Huff and Kluwe, 1984;

Kluwe et al., 1985), benzene and metabolites (Huff,

1992), dioxins (Huff, 1992), anilines (Weisburger

et al., 1984; Lamb et al., 1986), naturally occurring

"gums" (Melnick et al., 1983), chlorinated paraffins

(Bucher et al., 1987), 1,3-butadiene and derivatives

(Melnick and Huff, 1992), pesticides (Yang et al.,

1989; Huff and Haseman,1991), and penicillins and

tetracyclines(Dunnicket al., 1989;Dietzet al., 1991).

The bases for selection of anthraquinones(and other

chemical classes as well) centered mainly on four

criteria: 1) lack of available or adequate data on

carcinogenicity, 2) magnitudeof production and use 
Introduction 19

TABLE 1

Exposure Concentrations in the NCI/NTP 2-Year Feed Studies of Anthraquinone Derivativesa

Anthraquinone Derivative Low Dose High Dose Carcinogenic Response

(ppm) (ppm)

Rats

Male

2-Aminoanthraquinoneb 3,500 6,900 liver

1-Amino-2,4-dibromoanthraquinone 2,000 10,000 liver, large intestine, kidney, urinary

bladder

1-Amino-2-methylanthraquinoneb 1,000 2,000 liver, kidney

2-Methyl-1-nitroanthraquinone 600 1,200 liver, skin

1,4,5,8-Tetraaminoanthraquinone 1,250 5,000 urinary bladder, pancreas

Female

2-Aminoanthraquinonec 2,000

1-Amino-2,4-dibromoanthraquinone 2,000 10,000 liver, large intestine, kidney, urinary

bladder

1-Amino-2-methylanthraquinone 1,000 2,000 liver

2-Methyl-1-nitroanthraquinone 600 1,200 skin

1,4,5,8-Tetraaminoanthraquinone 1,250 5,000 urinary bladder

Mice

Male

2-Aminoanthraquinone 5,000 10,000 liver

1-Amino-2,4-dibromoanthraquinone 10,000 20,000 liver, forestomach, lung

1-Amino-2-methylanthraquinone 600 _ d

2-Methyl-1-nitroanthraquinone 300 600 hemangiosarcoma

1,4,5,8-Tetraaminoanthraquinone 600 2,500 liver, lung

e

Female

2-Aminoanthraquinone 5,000 10,000 liver, lymphoma

1-Amino-2,4-dibromoanthraquinone 10,000 20,000 liver, forestomach, lung

1-Amino-2-methylanthraquinone 600 _ d liver

2-Methyl-1-nitroanthraquinone 300 600 hemangiosarcoma

1,4,5,8-Tetraaminoanthraquinone 600 2,500

a Data from NCI, 1978a, 1978b, 1978c; NTP, 1986a

b Exposure concentrations in this study were time-weighted averages.

c

Inadequate study

d Two dosage regimens were used, but the time-weighted average concentrations were the same.

e

"Equivocal evidence" for both organs
20 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

patterns, 3) awareness of potential and actual human

exposure, and 4)representation of asbroad a spec-trum

ofstructural diversity within this class as possible. 1-

Amino-2,4-dibromoanthraquinonewasselectedfrom a

group of36 environmentallysignificantaryl bromides.

Because everyother anthraquinonederivativetested so

far for carcinogenic activity had been shown to be

carcinogenic in rodents, 1-amino-2,4--

dibromoanthraquinone was also expected to be

carcinogenic in laboratory animals. Thus, the

experimental design, while being consistent with a

"core protocol" (Huff et al., 1988), contains 

severalmodificationssuchas "stop-exposure" groupsto

better characterize this chemical. Additionally,

chemicaldispositionstudieswereaccomplishedpriorto

the 2-year exposures to permit optimal selection of

exposure concentrationsforthis water-insoluble dye.

Because these chemicals may and often do contain

considerable quantitiesoftheparentchemicaland other

anthraquinone derivatives, an extensive chemical

analysis was undertaken on these five chemicals to

quantitate their purity and to identify the major

impurities of 1-amino-2,4-dibromoanthraquinone

(Arneson et al., 1996).

1-Amino-2,4-dibromoanthraquinone

2-Aminoanthraquinone

1-Amino-2-methylanthraquinone 2-Methyl-1-nitroanthraquinone

1,4,5,8-Tetraaminoanthraquinone

FIGURE 1

Five Anthraquinone Derivatives Evaluated by the NCI/NTP
21

MATERIALS AND METHODS

PROCUREMENT

AND CHARACTERIZATION OF

1-AMINO-2,4-DIBROMOANTHRAQUINONE

1-Amino-2,4-dibromoanthraquinone was

obtained from American Color and Chemical

Corporation (Charlotte, NC; lot 1076-C) and

Mobay Corporation (Pittsburgh, PA). The

second lot was procured from Mobay

CorporationsinceAmerican Color and Chemical

Corporation had stopped production.

Lot 1076-C was used in the 13-week studies and

for 2 months of the 2-year studies. The lot from

Mobay Corporation was assigned lot number

M061583 and was used throughout the

remainder of the 2-year studies. Identity, purity,

and stability analyses were conducted by the

analytical chemistry laboratory, Midwest

Research Institute (Kansas City, MO)

(Appendix I). Reports on analyses performed in

suppo r t of the 1-ami n o -

2,4-dibromoanthraquinone studies are on file at

the National Institute of Environmental Health

Sciences (NIEHS).

The two lots of the chemical, a reddish brown to

orange powder, were identified as 1-amino2,4-dibromoanthraquinone by infrared,

ultraviolet/visible, and nuclear magnetic

resonance spectroscopy. The purity of each lot

was determined by elemental analyses, Karl

F ischer water an alysis, thin -laye r

chromatography, and high-performance liquid

chromatography. 

For lot 1076-C, elemental analyses for carbon,

hydrogen, nitrogen, and bromine were in general

agreement with theoretical values for 1-amino2,4-dibromoanthraquinone. Karl Fischer water

analysis indicated approximately 0.21% water.

Thin-layer and high-performance liquid

chromatography indicated a major peak and

eight impurities. Five of the impurities had peak

areas of less than 0.3%. The three major

impurities were identified as anthraquinone,

1-amino-2-bromoanthraquinone, and 2-amino1,3-dibromoanthraquinone. By highperformance liquid chromatography,

anthraquinone was found to be present at a

concentration of approximately 5.0%. 1-Amino2-bromoanthraquinone and 2-amino-1,3-

dibromoanthraquinone were found to be present

at concentrations of approximately 4.3% and

2.2%, respectively. The overall purity of

lot 1076-C was determined to be approximately

87%. 

For lot M061583, elemental analyses for carbon,

hydrogen, nitrogen, and bromine were in general

agreement with theoretical values for 1-amino2,4-dibromoanthraquinone. Karl Fischer water

analysis indicated approximately 0.32% water.

Thin-layer and high-performance liquid

chromatography indicated a major peak and six

impurities with the same retention times as found

for lot 1076-C. A total impurity area of 3% of the

total chromatographic peak area was found. The

overall purity of lot M061583 was determined to

be approximately 97%.

Stability studies performed using highperformance liquid chromatography indicated

that 1-amino-2,4-dibromoanthraquinone, when

stored protected from light, was stable as a bulk

chemical for at least 2 weeks at temperatures up

to 60" C. To ensure stability, the bulk chemical was

stored in the dark at 4"± 3" C throughout the

studies. During the 2-year studies, the stability of the

bulk chemical was monitored periodically by the

study laboratory using high-performance liquid

chromatography; no degradation of 1-amino2,4-dibromoanthraquinone was observed throughout

the studies.

PREPARATION AND ANALYSIS 

OF DOSE FORMULATIONS

The dose formulationswere prepared weekly by

mixing 1-amino-2,4-dibromoanthraquinonewith

feed (Table I1). Homogeneity and at least 2-

week stability at 25

B C were confirmed by the

analytical chemistry laboratory using

spectrophotometry and high-performance liquid

chromatography, respectively. During the

13-week and 2-year feed studies, the dose

formulations were stored in the dark for no

longer than 2 weeks.

The study laboratory conducted periodic

a n alyses o f the 1-amin o -

2,4-dibromoanthraquinone dose formulations

using a spectrophotometric method. For the

13-week feed studies, dose formulations were
22 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

analyzed at the beginning, midpoint, and end of the

studies (Table I2). During the 2-year feed studies, the

dose formulations were analyzed every 6 to 10 weeks

(Table I3). All dose formulations for rats and mice

were within 10% of the target concentrations during the

13-week and 2-year studies. Results of periodic referee

analyses performed by the analytical chemistry

laboratory indicated good agreement with the results

obtained by the study laboratory (Table I4).

13-WEEK STUDIES

The 13-week studies were conducted to evaluate the

cumulative toxic effects of repeated exposure to

1-amino-2,4-dibromoanthraquinone and to determine

the appropriate exposure concentrations to be used in

the 2-year studies.

Male and female F344/N rats and B6C3F1 mice were

obtained from Charles River Breeding Laboratories

(Portage, MI). Upon receipt, the animals were 5 weeks

old. The rats and mice were quarantined for 15 days

before the studies began. 

Groups of 10 male and 10 female rats and 10 male and

10 female mice received 0, 2,500, 5,000, 10,000,

25,000, or 50,000 ppm 1-amino -

2,4-dibromoanthraquinone in feed for 13 weeks. Males

and females were housed five per cage; water and feed

were available ad libitum, and feed consumption was

measured weekly. Clinical findings were recorded

twice daily. Animals were weighed at study initiation,

weekly, and at the end of the studies. Further details of

study design and animal maintenance are summarized

in Table 2. 

A necropsy was performed on all animals. The brain,

heart, right kidney, liver, lungs, right testis, and thymus

of all animals were weighed. Tissues for microscopic

examination were fixed and preserved in 10% neutral

buffered formalin, processed and trimmed, embedded in

paraffin, sectioned to a thickness of 4 to 6 :m, and

stained with hematoxylin and eosin. A complete

histopathologic examination was performed on all

animals that died prior to the end of the studies, control

animals, and animals administered 50,000 ppm.

Table 2 lists the tissues and organs examined.

2-YEAR STUDIES

Study Design

Groups of 70 male and 70 female rats received 0, 5,000,

or 10,000 ppm 1-amino-2,4-dibromoanthraquinone in

feed, and a group of 50 male and 50 female rats

received 2,000 ppm 1-amino-2,4-dibromoanthraquinone

in feed for 104 weeks. Ten male and 10 female rats

from the 0, 2,000, 5,000, and 10,000 ppm groups were

designated for an interim evaluation after 9 months.

Ten male and 10 female rats from the 0 and 10,000 ppm

groups were designated for an interim evaluation after

15 months. Groups of 60 male and 60 female mice

received 0, 10,000, or 20,000 ppm 1-amino2,4-dibromoanthraquinone in feed for 104 weeks. Ten

male and 10 female mice per group were evaluated after

15 months.

Stop-Exposure Evaluation

Groups of 40 male and 40 female rats received

20,000ppm 1-amino-2,4-dibromoanthraquinone in feed

for 9 months, when 10 males and 10 females were

evaluated. At 9 months, the dosed feed was replaced

with a control diet for 10 male and 10 female rats,

which were then necropsied and evaluated at

15 months. Twenty male and 20 female rats continued

t o r eceive 20, 000 ppm 1-amino -

2,4-dibromoanthraquinone in feed and were also

evaluated at 15 months. 

Source and Specification of Animals

Male and female F344/N rats and B6C3F1 mice were

obtained from Frederick Cancer Research Facility

(Frederick, MD). Rats were quarantined 12 to 14 days

(males) or 9 days (females) and mice were quarantined

12 days (males) or 15 days (females) before the

beginning of the studies. Five male and five female rats

and mice were selected and evaluated for evidence of

parasites and gross observation of disease. Serology

samples were collected for viral screening. Rats and

mice were approximately 6 weeks old at the beginning

of the 2-year studies. The health of the animals was

monitored during the studies according to the protocols

of the NTP Sentinel Animal Program (Appendix L).

Animal Maintenance

Males and females were housed five per cage. Feed and

water were available ad libitum. Feed consumption

was measured monthly (Appendix J). Cages and
Materials and Methods 23

racks were rotated every 2 weeks during the studies.

Further details of animal maintenance are given in

Table 2. Information on feed composition and

contaminants is provided in Appendix K.

Clinical Examinations and Pathology

All animals were observed twice daily. Clinical

findings were recorded weekly for 14 weeks then

monthly until the end of the studies. Animals were

weighed at study initiation, weekly for 14 weeks, and

monthly thereafter.

Animals were killed with CO2, and a complete

necropsy was performed on all animals. The

right kidney and liver of rats and mice were weighed at

the interim evaluations (Appendix H). At necropsy, all

organs and tissues were examined for gross lesions, and

all major tissues were fixed and preserved in 10%

neutral buffered formalin, processed and trimmed,

embedded in paraffin, sectioned, and stained with

hematoxylin and eosin for microscopic examination.

Histopathologic examinations were performed on all

tissues with grossly visible lesions. For all paired

organs (i.e., adrenal gland, kidney, ovary), samples

from each organ were examined. Tissues examined are

listed in Table 2.

Microscopic evaluations were completed by the study

laboratory pathologist, and the pathology data were

entered into the Toxicology Data Management System.

The microscope slides, paraffin blocks, and residual wet

tissues were sent to the NTP Archives for inventory,

slide/block match, and wet-tissue audit. The slides,

individual animal data records, and pathology tables

were evaluated by an independent pathology quality

assessment laboratory. The individual animal records

and tables were compared for accuracy, the slide and

tissue counts were verified, and the histotechnique was

evaluated by the quality assessment laboratory. The

quality assessment pathologist microscopically

reviewed selected neoplasms or nonneoplastic lesions.

The quality assessment report and slides were submitted

to the NTP Pathology Working Group (PWG) chair,

who reviewed the selected slides and any other tissues

for which a disagreement in diagnosis between the

laboratory and quality assessment pathologists existed.

Representative histopathology slides containing

examples of lesions related to chemical administration,

examples of disagreements in diagnoses between the

laboratory and quality assessment pathologists, or

lesions of general interest were presented by the chair

to the PWG for review. Tissues examined included the

adrenal cortex (female rats), ear (rats), kidney (rats),

large intestine (rats), liver, lung (mice), skin (rats),

forestomach, thyroid gland (rats), and urinary bladder

(rats). The PWG consisted of the quality assessment

pathologist and other pathologists experienced in rodent

toxicologic pathology. This group examined the tissues

without any knowledge of exposure groups or

previously rendered diagnoses. When the PWG

consensus differed from the opinion of the laboratory

pathologist, the diagnosis was changed. Thus, the final

diagnoses represent a consensus of contractor

pathologists and the PWG. Details of these review

procedures have been described by Maronpot and

Boorman (1982) and Boorman et al. (1985). For

subsequent analysis of pathology data, the diagnosed

lesions for each tissue type were evaluated separately or

combined according to the guidelines of McConnell

et al. (1986).

STATISTICAL METHODS

Survival Analyses

The probability of survival was estimated by the

product-limit procedure of Kaplan and Meier (1958)

and is presented in the form of graphs. Animals found

dead of other than natural causes or missexed animals

were censored from the survival analyses; animals

dying from natural causes were not censored.

Statistical analyses for possible dose-related effects on

survival used Cox's (1972) method for testing two

groups for equality and Tarone's (1975) life table test to

identify dose-related trends. All reported P values for

the survival analyses are two sided.

Calculation of Incidence

The incidences of neoplasms or nonneoplastic lesions

as presented in Tables A1, A5, B1, B5, C1, C5, D1, D5,

E1, E3, F1, and F3 are given as the number of animals

bearing such lesions at a specific anatomic site and the

number of animals with that site examined

microscopically. For calculation of statistical

significance, the incidences of most neoplasms (Tables

A3, B3, C3, D3, E2a, E2b, F2a, and F2b) and of all

nonneoplastic lesions are given as the numbers of

animals affected at each site examined microscopically.

However, when macroscopic examination was required

to detect neoplasms in certain tissues (e.g., harderian

gland, intestine, mammary gland, and skin) before

microscopic evaluation or when neoplasms 
24 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

had multiple potential sites of occurrence (e.g.,

leukemia or lymphoma), the denominators consist of

the number of animals on which a necropsy was

performed. Tables A3, B3, C3, D3, E2a, E2b, F2a, and

F2b also give the survival-adjusted neoplasm rate for

each group and each site-specific neoplasm, i.e., the

Kaplan-Meier estimate of the neoplasm incidence that

would have been observed at the end of the study in the

absence of mortality from all other competing risks

(Kaplan and Meier, 1958).

Analysis of Neoplasm Incidence

The majority of neoplasms in these studies were

considered to be incidental to the cause of death or not

rapidly lethal. Thus, the primary statistical method

used was logistic regression analysis, which assumed

that the diagnosed neoplasms were discovered as the

result of death from an unrelated cause and thus did not

affect the risk of death. In this approach, neoplasm

prevalence was modeled as a logistic function of

chemical exposure and time. Both linear and quadratic

terms in time were incorporated initially, and the

quadratic term was eliminated if the fit of the model

was not significantly enhanced. The neoplasm

incidences of exposed and control groups were

compared on the basis of the likelihood score test for

the regression coefficient of dose. This method of

adjusting for intercurrent mortality is the prevalence

analysis of Dinse and Lagakos (1983), further described

and illustrated by Dinse and Haseman (1986). When

neoplasms are incidental, this comparison of the

time-specific neoplasm prevalences also provides a

comparison of the time-specific neoplasm incidences

(McKnight and Crowley, 1984).

In addition to logistic regression, other methods of

statistical analysis were used, and the results of these

tests are summarized in the appendixes. These methods

include the life table test (Cox, 1972; Tarone, 1975),

appropriate for rapidly lethal neoplasms, and the Fisher

exact test and the Cochran-Armitage trend test (Armitage, 1971; Gart et al., 1979), procedures based on the

overall proportion of lesion-bearing animals.

Tests of significance included pairwise comparisons of

each dosed group with controls and a test for an overall

dose-related trend. Continuity-corrected tests were

used in the analysis of neoplasm incidence, and

reported P values are one sided. The procedures

described in the preceding paragraphs were also used

 to evaluate selected nonneoplastic lesions. For further

discussion of these statistical methods, refer to Haseman

(1984).

Analysis of Nonneoplastic Lesion Incidences

Because all nonneoplastic lesions in this study were

considered to be incidental to the cause of death or not

rapidly lethal, the primary statistical analysis used was

a logistic regression analysis in which nonneoplastic

lesion prevalence was modeled as a logistic function of

chemical exposure and time. For lesions detected at the

interim evaluation, the Fisher exact test, a procedure

based on the overall proportion of affected animals, was

used.

Analysis of Continuous Variables

Organ and body weight data, which have approximately

normal distributions, were analyzed using the

parametric multiple comparison procedures of Dunnett

(1955) and Williams (1971, 1972). Jonckheere's test

(Jonckheere, 1954) was used to assess the significance

of the dose-related trends and to determine whether a

trend-sensitive test (Williams' test) was more

appropriate for pairwise comparisons than a test that

does not assume a monotonic dose-related trend

(Dunnett's test). Prior to analysis, extreme values

identified by the outlier test of Dixon and Massey

(1951) were examined by NTP personnel, and

implausible values were eliminated from the analysis.

Average severity values were analyzed for significance

using the Mann-Whitney U test (Hollander and Wolfe,

1973).

Historical Control Data

Although the concurrent control group is always the

first and most appropriate control group used for

evaluation, historical control data can be helpful in the

overall assessment of lesion incidence in certain

instances. Consequently, neoplasm incidences from the

NTP historical control database (Haseman et al., 1984,

1985) are included in the NTP reports for neoplasms

appearing to show compound-related effects.

QUALITY ASSURANCE METHODS

The 13-week and 2-year studies were conducted in

compliance with Food and Drug Administration Good

Laboratory Practice Regulations (21 CFR, Part 58). In

addition, as records from the 2-year studies were

submitted to the NTP Archives, these
Materials and Methods 25

studies were audited retrospectively by an

independent quality assurance contractor.

Separate audits covering completeness and

accuracy of the pathology data, pathology

specimens, final pathology tables, and a draft of

this NTP Technical Report were conducted.

Audit procedures and findings are presented in

the reports and are on file at NIEHS. The audit

findings were reviewed and assessed by NTP

staff, so all discrepancies had been resolved or

were otherwise addressed during the preparation

of this Technical Report.

GENETIC TOXICOLOGY

The genetic t oxicity o f 1-amino2,4-dibromoanthraquinone was assessed by

testing the ability of the chemical to induce

mutations in various strains of Salmonella

typhimurium cells and sister chromatid

exchanges and chromosomal aberrations in

cultured Chinese hamster ovary cells. The

protocols for these studies and the results are

given in Appendix G.

The genetic toxicity studies of 1-amino2,4-dibromoanthraquinone are part of a larger

effort by the NTP to develop a database that

would permit the evaluation of carcinogenicity

in experimental animals from the structure and

responses of the chemical in short-term in vitro

and in vivo genetic toxicity tests. These

genetic toxicity testswere originally developed to

study mechanisms of chemically induced DNA

damage and to predict carcinogenicity in

animals, based on the electrophilic theory of

chemical carcinogenesis and the somatic

mutation theory (Miller and Miller, 1977;

Straus, 1981; Crawford, 1985).

There is a strong correlation between a

chemical's potential electrophilicity (structural

alert to DNA reactivity), mutagenicity in

Salmonella, and carcinogenicity in rodents. The

combination of electrophilicity and Salmonella

mutagenicity is highly correlated with the

induction of carcinogenicity in rats and mice

and/or at multiple tissue sites (Ashby and

Tennant, 1991). Other in vitro genetic toxicity

tests do not correlate well with rodent

carcinogenicity (Tennant et al., 1987; Zeiger

et al., 1990), although these other tests can

provide information on the types of DNA and

chromosome effects that can be induced by the

chemical being investigated. Data from NTP

studies show that a positive response in

Salmonella is currently the most predictive in

vitro test for rodent carcinogenicity (89% of the

Salmonella mutagenswere rodent carcinogens),

and that there is no complementarity among the

in vitro genetic toxicity tests. That is, no battery

of tests that included the Salmonella test

improved the predictivity of the Salmonella test

alone.
26 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE 2

Experimental Design and Materials and Methods in the Feed Studies 

of 1-Amino-2,4-dibromoanthraquinone

13-Week Studies 2-Year Studies Stop-Exposure

Evaluation

Study Laboratory

EG&G Mason Research Institute (Worcester,

MA)

Same as 13-week studies Same as 13-week studies

Strain and Species

Rats: F344/N

Mice: B6C3F1

Rats: F344/N

Mice: B6C3F1

Rats: F344/N

Animal Source

Charles River Breeding Laboratories

(Portage, MI)

Frederick Cancer Research Facility

(Frederick, MD)

Same as 2-year studies

Time Held Before Studies

15 days Rats: 12-14 days (males) or

9 days (females)

Mice: 12 days (males) or

15 days (females)

12-14 days (males) or 

9 days (females)

Average Age When Studies Began

7 weeks 6 weeks 6 weeks

Date of First Dose

Rats: 22 April (males) or

29 April (females) 1982

Mice: 6 May (males) or

13 May (females) 1982

Rats: 13 July (males) or 

4 August (females) 1983

Mice: 20 June (males) or

30 June (females) 1983

13 July (males) or 

4 August (females) 1983

Duration of Dosing

13 weeks 104 weeks 9-Month stop-exposure group:

39 weeks (males) or 40 weeks (females)

followed by control feed for remainder of

study

15-Month exposure group:

66 weeks

Date of Last Dose

Rats: 21-23 July (males) or 

28-30 July (females) 1982

Mice: 4-6 August (males) or 

11-13 August (females) 1982

Rats: 3 July (males) or 

25 July (females) 1985

Mice: 10 June (males) or 

20 June (females) 1985

9-Month stop-exposure group:

10-13 April (males) or 8-10 May (females)

1984

15-Month exposure group:

10-12 October (males) or 7-9 November

(females) 1984
Materials and Methods 27

TABLE 2

Experimental Design and Materials and Methods in the Feed Studies 

of 1-Amino-2,4-dibromoanthraquinone (continued)

13-Week Studies 2-Year Studies Stop-Exposure

Evaluation

Necropsy Dates

Rats: 21-23 July (males) or 

28-30 July (females) 1982

Mice: 4-6 August (males) or

11-13 August (females) 1982

Rats: 

9-Month interim evaluation: 

10-13 April (males) or

8-10 May (females) 1984

15-Month interim evaluation: 

10-12 October (males) or 7-9 November

(females) 1984

Terminal:

10-16 July (males) or

1-8 August (females) 1985

Mice:

15-Month interim evaluation:

19-21 September (males) or

26-28 September (females) 1984

Terminal:

17-20 June (males) or 27 June - 2 July

(females) 1985

9-Month interim evaluation:

10-13 April (males) or 8-10 May (females)

1984

15-Month terminal:

10-12 October (males) or 7-9 November

(females) 1984

Average Age at Necropsy

20 weeks 9-Month interim evaluation:

45-46 weeks

15-Month interim evaluation:

72 weeks 

Terminal:

110-112 weeks

9-Month interim evaluation:

45-46 weeks

15-Month terminal:

72 weeks

Size of Study Groups

10 males and 10 females Rats: 70 males and 70 females in the

0, 5,000, and 10,000 ppm groups; 50 males

and 50 females in the 2,000 ppm group

Mice: 60 males and 60 females

40 males and 40 females

Method of Animal Distribution

Animals were caged by 1-gram weight

classes and then distributed into treatment

groups such that within a given sex, all cage

weights were approximately equal (± 2 g).

Animals were distributed randomly into

groups of approximately equal initial mean

body weight.

Same as 2-year studies

Animals per Cage

5 5 5

Method of Animal Identification

Ear punch Ear punch Ear punch
28 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE 2

Experimental Design and Materials and Methods in the Feed Studies 

of 1-Amino-2,4-dibromoanthraquinone (continued)

13-Week Studies 2-Year Studies Stop-Exposure

Evaluation

Diet

NIH-07 open formula meal diet (Zeigler

Brothers, Inc., Gardners, PA), Available

ad libitum, changed weekly

Same as 13-week studies Same as 13-week studies

Water

Tap water (City of Worcester) available

ad libitum via automatic watering system

(Edstrom Industries, Inc., Waterford, WI)

Same as 13-week studies Same as 13-week studies

Cages

Polycarbonate cage (Lab Products, Inc.,

Rochelle Park, NJ), changed twice weekly

Same as 13-week studies Same as 13-week studies

Bedding

Aspen Bed® heat-treated hardwood chips

(American Excelsior, Baltimore, MD),

changed twice weekly

Same as 13-week studies; BetaChips®

hardwood chips (Northeastern Products,

Warrensburg, NY) were used if necessary.

Same as 2-year studies

Cage Filters

Nonwoven fiber filters (Lab Products,

Rochelle Park, NJ; or Snow Filtration,

Cincinnati, OH); changed every 2 weeks

Nonwoven fiber filters (Snow Filtration Co.,

Cincinnati, OH), changed every 2 weeks

Same as 2-year studies

Racks

Stainless steel racks (Lab Products, Inc.,

Maywood, NY), changed every 2 weeks

Same as 13-week studies Same as 13-week studies

Animal Room Environment

Average temperature: 

22° to 26° C

Relative humidity: 

24% to 66% (rats),

28% to 66% (mice)

Fluorescent light: 

12 hours/day

Room air:

12 to 15 changes/hour

Average temperature: 

19° to 26° C 

Relative humidity: 

16% to 76% 

Fluorescent light: 

12 hours/day

Room air:

12 to 15 changes/hour

Same as 2-year studies
Materials and Methods 29

TABLE 2

Experimental Design and Materials and Methods in the Feed Studies 

of 1-Amino-2,4-dibromoanthraquinone (continued)

13-Week Studies 2-Year Studies Stop-Exposure

Evaluation

Doses

0, 2,500, 5,000, 10,000, 25,000, or

50,000 ppm in feed, available ad libitum

Rats: 0, 2,000, 5,000, or 

10,000 ppm in feed, available ad libitum

Mice: 0, 10,000, or

20,000 ppm in feed, available ad libitum

20,000 ppm in feed, available ad libitum

Type and Frequency of Observation

Observed twice daily; animals weighed

initially, weekly, and at end of studies;

clinical observations recorded twice daily;

feed consumption measured weekly

Observed twice daily; animals weighed

initially, weekly for 14 weeks, and monthly

thereafter; clinical observations recorded

weekly for 14 weeks, then monthly until end

of the studies; feed consumption measured

monthly

Same as 2-year studies

Method of Sacrifice

CO2 asphyxiation CO2 asphyxiation CO2 asphyxiation

Necropsy

Necropsy performed on all animals. Organs

weighed were brain, heart, right kidney, liver,

lungs, right testis, and thymus.

Necropsy was performed on all animals. 

Organs weighed at the 9- and 15-month

interim evaluations were right kidney and

liver.

Necropsy was performed on all animals. 

Organs weighed at 9 months and 15 months

were right kidney and liver.

Histopathology

Complete histopathologic examinations were

performed on all animals that died during the

study, control animals, and 50,000 ppm

animals. In addition to tissue masses, gross

lesions, and associated lymph nodes, the

tissues examined included: adrenal gland,

bone and marrow, brain, clitoral gland,

epididymis, esophagus, heart, kidney, large

intestine (cecum, colon, rectum), liver, lung,

lymph nodes (mandibular and mesenteric), 

mammary gland, nose, ovary, pancreas,

parathyroid gland, pituitary gland, preputial

gland, prostate gland, salivary gland, seminal

vesicle, skin, small intestine (duodenum,

jejunum, ileum), spleen, stomach, testis,

thymus, thyroid gland, trachea, urinary

bladder, and uterus. The kidney, liver, spleen

(rats), thymus (rats), and uterus (rats) of all

other exposed animals were examined.

Complete histopathologic examinations were

performed on all animals. In addition to

tissue masses, gross lesions, and associated

lymph nodes, the tissues examined included:

adrenal gland, bone and marrow, brain,

clitoral gland, epididymis, esophagus,

gallbladder (mice), heart, kidney, large

intestine (cecum, colon, rectum), liver, lung,

lymph nodes (mandibular and mesenteric),

mammary gland, nose, ovary, pancreas,

parathyroid gland, pituitary gland, preputial

gland, prostate gland, salivary gland, seminal

vesicle, skin, small intestine (duodenum,

jejunum, ileum), spleen, stomach, testis,

thymus, thyroid gland, trachea, urinary

bladder, and uterus.

Complete histopathologic examinations were

performed on all animals. In addition to

tissue masses, gross lesions, and associated

lymph nodes, the tissues examined included: 

adrenal gland, bone and marrow, brain,

clitoral gland, epididymis, esophagus, heart,

kidney, large intestine (cecum, colon,

rectum), liver, lung, lymph nodes (mandibular

and mesenteric), mammary gland, nose,

ovary, pancreas, parathyroid gland, pituitary

gland, preputial gland, prostate gland, salivary

gland, seminal vesicle, skin, small intestine

(duodenum, jejunum, ileum), spleen, stomach,

testis, thymus, thyroid gland, trachea, urinary

bladder, and uterus.
30 1-Amino-2,4-dibromoanthraquinone, NTP TR 383
31

RESULTS

RATS

13-WEEK STUDY

One male (week 13) and one female (week 8) in the

50,000 ppmgroups died during the study (Table 3).

The deaths of one 5,000 ppmmale (week 4) and two

additional 50,000 ppm males (week 13) were not

chemicalrelated. The final mean body weights and

body weight gains of all exposed rat groups were

significantlylowerthan those of the controls. Feed

consumption by all exposed groups was less than that

by the controls throughout the study and generally

decreased with increasing exposure concentration

(Table 3). The greatest differences in feed consumption fromthat by the controls occurred in the 25,000

and 50,000 ppmmales and females. Feed consumption by these groups ranged from45% to 79% that by

the controls at week 1 and from64% to 82% that by

the controls at week 13. Dietary levels of 2,500,

5,000,10,000,25,000, and 50,000ppmdelivered daily

doses of approximately 150, 300, 620, 1,600, and

3,200mg1-amino-2,4-dibromoanthraquinone/kgbody

TABLE 3

Surviva l, Mean Body Weights, and Feed Consumption of Rats in the 13-Week Feed Study

of 1-Amino-2,4-dibromoanthraquinone

Final Weight

Mean Body Weightb

(g) Relative Feed

Dose SurvivalaInitial Final Change to Controls Consumptionc

(ppm) (%) Week 1 Week 13

Male

0 10/10 180 ± 3 358 ± 3 179 ± 3 14.9 18.1

2,500 10/10 180 ± 3 325 ± 3** 145 ± 3** 91 14.3 16.7

5,000 9/10d

180 ± 3 328 ± 3** 148 ± 6** 92 13.9 17.1

10,000 10/10 181 ± 3 310 ± 3** 129 ± 3** 86 13.5 17.0

25,000 10/10 181 ± 3 232 ± 3** 52 ± 4** 65 11.7 14.9

50,000 7/10e180 ± 3 164 ± 6** -17 ± 6** 46 10.3 11.6

Female

0 10/10 140 ± 2 211 ± 3 71 ± 2 13.0 15.7

2,500 10/10 140 ± 2 197 ± 3** 57 ± 3** 93 10.5 12.7

5,000 10/10 140 ± 2 188 ± 3** 47 ± 3** 89 10.2 12.1

10,000 10/10 140 ± 2 185 ± 2** 45 ± 2** 88 9.6 11.9

25,000 10/10 140 ± 2 159 ± 2** 19 ± 2** 75 7.0 10.6

50,000 9/10f140 ± 2 130 ± 4** -10 ± 4** 61 5.9 11.5

** Significantly different (P#0.01) from the control group by Williams' or Dunnett's test

a Number of animals surviving at 13 weeks/number initially in group

b Weights and weight changes are given as mean ± standard error. Subsequent calculations are based on animals surviving to the end of the study.

c

Feed consumption is expressed as grams of feed consumed per animal per day.

d Week of death: 4

e Week of death: 13, 13, 13 (2 were accidental deaths)

f Week of death: 8
32 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

weight to males and 170, 340, 660, 1,500, and

3,200 mg/kg to females. Pink-red staining of the fur and

tail was observed in all exposed groups of rats. The

bedding of all exposed groups except the 2,500 ppm

groups was stained pink-red from day 2 of the study.

Lethargy and emaciation were noted in all 50,000 ppm

males. Female rats in the 25,000 and 50,000 ppm groups

were lethargic and staggered, and 50,000 ppm females

exhibited hunched posture.

The relative liver weights of exposed groups of males

and the absolute and relative liver weights of exposed

groupsoffemalesweresignificantlygreaterthanthose

of the controls(Table H1). The absolute and relative

thymus weights of exposed males and females were

significantly lower than those of controls. The lower

absolute brain, heart, kidney, lung, and testis weights

of exposedmale and female ratswereprobably related

to the lower final mean body weights of these groups.

Observations at necropsyincludedred or pink staining

of the gastrointestinal tract contents and/or mucosa,

kidneys, and urine. In addition, regional lymph nodes

and livers were dark in color, and capsularsurfaces of

the liversweregranularinappearance. These findings

were most common in the 25,000 and 50,000 ppm

groups.

Chemical-related lesions were present in the liver,

kidney, and spleen of male and female rats. In the

liver, a spectrum of nonneoplastic degenerative and

proliferative lesions occurred in males and femalesin

the 25,000 and 50,000 ppm groups (Table 4).

Hepatocellular cytomegaly(hypertrophy) waspresent

in all ratsin the 25,000 and 50,000 ppm groups and in

most females in the 10,000 ppm group. This lesion

consisted of enlarged hepatocytes with eosinophilic

cytoplasmandmarkedvariation in nuclearsize. In the

centrilobular areas of a few ratsfrom exposed groups,

there was a minimal to mild cytoplasmic vacuolation

(vacuolar degeneration). The incidence of vacuolar

degeneration was not dose-related, but at the higher

exposure concentrations, minimal hepatocellular

necrosis was sometimes associated with vacuolar

change. Focal hepatocellular alterations including

basophilic, eosinophilic, or clear cell foci were also

present in the 25,000 and 50,000 ppm groups. In the

periportal region of the hepatic lobules, there was an

increased number of inflammatory cells around the

bile d ucts. Bile duct hyperplasi a

consistedof proliferation of oval cellsinthe periportal

area as well as proliferation of larger bile ducts lined

by hyperchromatic, pleomorphic biliary epithelium.

Focal necrosis of biliary epitheliumand acute inflammation (necrotizing cholangitis) in some hyperplastic

bile ducts were associated with periportal fibrosis.

The spectrumof proliferative bileduct lesions(hyperplasia, necrotizing cholangitis, and fibrosis) was

morphologically consistent with the lesion described

as cholangiofibrosis. Abrown pigment was present in

the cytoplasm of hepatocytes. The pigment was

negative foriron, PAS, bile, and acid-faststaining; did

not polarize light or fluoresce; and was considered to

represent1-amino-2,4-dibromoanthraquinone and/orits

metabolites.

In the kidneyof exposedgroups of males and females,

there were chemical-related increases in the incidences of a brown, granular pigment in the tubule

epithelium (Table 4). This brown pigment had the

same staining features asthe pigment that was present

in the liver. In both males and females, there were

renal tubule cellswithenlargednuclei. Inmales,there

was a hyaline droplet nephropathy characterized by an

increase in eosinophilic protein droplets (hyaline

droplet accumulation) in the cytoplasm of the renal

tubule epithelium as well as in the lumen of the

tubules. There was no evidence of increased severity

of tubule regeneration in males or females.

Chemical-related effects in the spleen of all exposed

groups of males and females consisted of a slight

increase in the amount of hematopoiesis relative to

that normally present in controls.

Other nonspecific changes included lymphoid depletion in the thymus and decreased uterus size. These

findings were attributed to the markedly lower body

weight gain in ratsfrom the higher exposure groups.

Dose SelectionRationale: Based on chemical disposition studies,meanbodyweights,and chemical-related

lesions of the liver, kidney, and spleen present mainly

in the 25,000 and 50,000 ppm groups, exposure concentrations selected for the 2-year feed study of

1-amino-2,4-dibromoanthraquinone in rats were 0,

2,000, 5,000, and 10,000 ppm. Much of the differencesin mean body weightsrecorded for the 13-week

studies were more likely due to decreased feed palatability than to any overt toxicity. Nonetheless, if
Results 33

TABLE 4

Incidences of Selected Nonneoplastic Lesions in Rats in the 13-Week Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

0 ppm 2,500 ppm 5,000 ppm 10,000 ppm 25,000 ppm 50,000 ppm

Male

Livera10 10 10 10 10 10

Basophilic Focusb0 0 0 0 4* (1.5)d9** (2.3)

Clear Cell Focus 0 0 0 0 6** (1.0) 0

Eosinophilic Focus 0 0 0 0 4* (1.5) 0

Cytomegaly 0 0 0 0 10** (3.2) 10** (4.0)

Bile Duct Hyperplasia 0 0 0 0 8** (2.3) 10** (3.1)

Inflammation 0 0 0 0 10** (2.1) 10** (3.0)

Fibrosisc0 0 0 0 10** (1.9) 10** (3.1)

Necrotizing Cholangitisc0 0 0 0 7** (1.3) 10** (2.8)

Vacuolar Degenerationc0 0 4* (1.5) 5* (1.8) 3 (1.3) 4* (1.5)

Pigmentation 0 0 0 0 10** (1.5) 9** (1.0)

Kidney 10 10 10 10 10 10

Renal Tubule Pigmentation 0 10** (1.0) 9** (1.0) 10** (1.1) 10** (2.2) 10** (1.8)

Hyaline Droplet Accumulation 0 10** (1.7) 9** (1.7) 10** (2.0) 2 (1.0) 0

Female

Liver 10 10 10 10 10 10

Basophilic Focus 0 0 0 0 1 (1.0) 9** (2.6)

Eosinophilic Focus 0 0 0 0 1 (1.0) 0

Cytomegaly 0 0 0 8** (1.0) 10** (2.2) 10** (4.0)

Bile Duct Hyperplasia 0 0 0 4* (1.0) 9** (1.6) 10** (2.4)

Inflammation 0 0 0 0 10** (1.6) 9** (2.3)

Fibrosisc0 0 0 0 2 (2.0) 9** (2.7)

Necrotizing Cholangitisc0 0 0 0 0 9** (2.6)

Vacuolar Degenerationc0 0 0 1 (1.0) 2 (2.0) 8** (1.4)

Pigmentation 0 0 1 (1.0) 7** (1.0) 9** (1.2) 10** (1.5)

Kidney 10 10 10 10 10 10

Renal Tubule Pigmentation 0 10** (1.0) 9** (1.0) 10** (1.2) 10** (1.7) 10** (1.9)

* Significantly different (P#0.05) from the control group by the Fisher exact test

** P#0.01

a Number of animals with organ examined microscopically

b Number of animals with lesion

c Data from Fleischman et al., 1986

d Average severity of lesions in affected rats: 1 = minimal; 2 = mild; 3 = moderate; 4 = marked

exposure selection werebased onmeanbodyweights

alone formale rats,the10,000ppmexposureconcentration could have been considered slightly high. Moreover,consideringthe lackoflivertoxicityat exposures

of 10,000 ppm and below, this exposure 

concentrationwas predicted notto adverselyaffectthe

health or survivalofthese animals. Higher exposure

concentrations(20,000 ppm)werechosen forthe startstop, progression/regressionexperiments(stop-exposure

evaluation).
34 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

2-YEAR STUDY

Survival

Estimatesof 2-yearsurvivalprobabilitiesformale and

female rats are shown in Table 5 and in the

Kaplan-Meiersurvival curvesinFigure 2. Survivalof

male and female rats in the 10,000 ppm groups was

significantly lower than that of the controls.

Body Weights, Feed and Compound

Consumption, and Clinical Findings

Mean body weights of exposed male and female rats

were lower than those of the controls after week 2

(Tables6 and 7,Figure 3). Finalmeanbodyweights of

exposed males were 14% to 30% lower than that

of the controls; final mean body weights of exposed

females were 20% to 46% lower than that of the

controls. Feed consumption by exposed males and

females was similar among exposed groups and was

slightlylowerthan that by the controls (TablesJ3 and

J4). Dietary levels of 2,000, 5,000, and 10,000 ppm

delivered average daily doses of approximately 90,

240, and 490 mg 1-amino-2,4-dibromoanthraquinone/kg bodyweight to males and 110, 285, and 600

mg/kg to females. Discoloration of the furand urine

was evidentin all exposed groups as earlyas day8 and

was observed throughout the study. Emaciation

occurred in a dose-relatedmannerinmale and female

rats and occurred in over 50% of the rats exposed to

10,000 ppm.

TABLE 5

Survival of Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Male

Animals initially in study 70 50 70 70

9-Month interim evaluationa10 10 10 10

15-Month interim evaluationa10 0 0 10

Moribund 19 15 34 33

Natural deaths 5 1 5 7

Animals surviving to study termination 26e24 21 10

Percent probability of survival at end of studyb53 60 35 20

Mean survival (days)c586 618 615 547

Survival analysesd P<0.001 P=0.467N P=0.141 P<0.001

Female

Animals initially in study 70 50 70 70

9-Month interim evaluationa10 10 10 10

15-Month interim evaluationa10 0 0 10

Missexeda0 0 0 1

Moribund 8 5 15 29

Natural deaths 4 3 7 8

Animals surviving to study termination 38f32 38 12

Percent probability of survival at end of study 76 80 63 25

Mean survival (days) 610 626 643 569

Survival analyses P<0.001 P=0.857N P=0.216 P<0.001

a Censored from survival analyses b Kaplan-Meier determinations based on the number of animals alive on first day of terminal sacrifice

c Mean of all deaths (uncensored, censored, and terminal sacrifice) d The result of the life table trend test (Tarone, 1975) is in the control column, and the results of the life table pairwise comparisons (Cox, 1972)

with the controls are in the exposed columns. A lower mortality in an exposure group is indicated by N.

e

Includes three males that died during the last week of the study. f

Includes one female that died during the last week of the study.
Results 35 

FIGURE 2

Kaplan-Meier Survival Curves for Rats Administered l-Amino-2,4-dibromoanthraquinone in Feed 

for 2 Years
36 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE 6

Mean Body Weights and Survival of Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

Weeks 0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

on Av. Wt. No. of Av. Wt.Wt. (% of No. of Av. Wt.Wt. (% of No. of Av. Wt.Wt. (% of No. of

Study (g) Survivors (g) controls)Survivors (g) controls)Survivors (g) controls)Survivors

1 139 70 136 98 50 136 98 70 134 97 70

2 161 70 165 103 50 164 102 70 155 97 70

 3 206 70 203 99 50 198 96 70 184 89 70

4 235 70 225 96 50 222 95 70 204 87 70

5 240 70 233 97 50 230 96 70 215 90 70

6 269 70 258 96 50 257 96 70 237 88 70

7 287 70 271 94 50 269 94 70 248 86 70

8 302 70 285 94 50 280 93 70 260 86 70

9 312 70 295 95 50 287 92 70 268 86 70

10 325 70 308 95 50 303 93 70 281 86 70

11 333 70 316 95 50 313 94 70 293 88 70

12 338 70 323 96 50 318 94 70 300 89 70

13 332 70 321 97 50 309 93 70 298 90 70

14 356 70 339 95 50 330 93 70 314 88 70

17 387 70 363 94 50 350 91 70 335 87 70

21 406 70 382 94 50 368 91 70 356 88 70

25 423 70 398 94 50 383 90 70 369 87 70

29 435 70 403 93 50 384 88 70 376 86 70

33 445 70 413 93 50 398 89 70 383 86 70

37 453 70 420 93 50 401 89 70 388 86 70

 41a 468 60 433 92 40 415 89 60 397 85 59

45 473 60 440 93 40 424 90 60 403 85 59

49 479 60 452 94 40 428 89 60 409 84 57

53 489 60 457 93 40 439 90 60 415 85 57

57 486 60 453 93 40 430 88 60 409 84 57

61 484 59 445 92 40 430 89 60 405 84 57

65 484 57 448 93 40 426 88 60 408 84 55

 69a 479 47 442 92 40 417 87 59 398 83 44

72 472 46 429 91 40 407 86 58 389 82 42

77 460 46 412 90 40 392 85 56 381 83 41

80 462 44 418 91 40 390 84 56 373 81 38

85 467 40 419 90 40 388 83 52 365 78 34

89 455 40 405 89 39 378 83 49 357 79 29

93 445 39 396 89 36 363 82 43 347 78 24

97 432 35 380 88 32 355 82 37 332 77 22

101 422 30 380 90 26 343 81 28 301 71 18

103 406 29 349 86 24 341 84 22 283 70 13

Mean for weeks

1-13 268 257 96 253 94 237 89

14-52 433 404 93 388 90 373 86

53-103 460 417 91 393 85 369 80

a

Interim evaluations occurred during week 39 for all groups and week 66 for the 0 and 10,000 ppm groups.
Results 37

TABLE 7

Mean Body Weights and Survival of Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

Weeks 0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

on Av. Wt. No. of Av. Wt.Wt. (% of No. of Av. Wt.Wt. (% of No. of Av. Wt.Wt. (% of No. of

Study (g) Survivors (g) controls)Survivors (g) controls)Survivors (g) controls)Survivors

1 93 70 93 100 50 94 101 70 94 101 70

2 114 70 110 96 50 107 94 70 102 90 70

3 133 70 128 96 50 121 91 70 116 87 70

4 147 70 141 96 50 136 93 70 128 87 70

5 157 70 151 96 50 146 93 70 137 87 70

6 166 70 159 96 50 152 91 70 145 87 70

7 173 70 169 97 50 161 93 70 150 87 69

8 180 70 174 97 50 167 93 70 158 88 69

9 186 70 178 96 50 173 93 70 167 90 69

10 191 70 185 97 50 179 94 70 173 90 69

11 196 70 189 96 50 184 94 70 177 90 69

12 203 70 194 95 50 188 93 70 181 89 69

13 208 70 195 94 50 192 93 70 185 89 69

14 212 70 201 95 50 196 93 70 189 89 69

17 222 70 209 94 50 205 92 70 200 90 69

21 228 70 216 95 50 213 93 70 205 90 69

25 237 70 219 93 50 216 91 70 208 88 69

29 246 70 223 91 50 219 89 70 212 86 69

33 251 70 227 90 50 222 88 70 213 85 69

37 258 70 233 90 50 230 89 70 218 85 69

 41a265 59 233 88 40 227 86 60 217 82 59

45 272 59 239 88 40 231 85 60 221 81 59

49 284 59 246 87 40 237 84 60 227 80 59

53 299 59 257 86 40 245 82 60 232 78 59

57 311 59 264 85 40 249 80 60 238 77 58

61 315 59 267 85 40 251 80 60 238 76 57

65 328 59 275 84 40 257 78 60 242 74 57

 68a 333 49 277 83 40 257 77 60 241 72 47

73 343 49 286 83 40 263 77 60 245 71 46

77 347 49 289 83 40 269 78 60 243 70 45

81 351 49 296 84 40 270 77 60 239 68 43

85 354 49 295 83 40 268 76 58 234 66 39

89 354 47 298 84 37 262 74 55 229 65 34

93 356 45 299 84 37 251 71 53 224 63 28

97 358 43 298 83 33 250 70 46 213 60 23

100 361 42 293 81 32 243 67 43 202 56 20

103 362 38 290 80 32 234 65 39 194 54 13

Mean for weeks

1-13 165 159 96 154 93 147 89

14-52 248 225 91 220 89 211 85

53-103 341 285 84 255 75 230 67

a

Interim evaluations occurred du ring week 40 for all groups and week 66 for the 0 and 10,000 ppm groups.
38 l-Ajnino-2,4-dibromoanthraquinone, NTP TR 383

FIGURE 3

Growth Curves for Rats Administered 1-Amino-2,4-dibromoanthraquinone in Feed for 2 Years
Results 39

Pathology and Statistical Evaluation

This section describes the statistically significant or

biologically noteworthy changes in the incidences of

mononuclear cell leukemia; neoplasms of the liver,

large intestine, kidney, urinary bladder, and other

organs; and nonneoplastic lesionsofthe liver, kidney,

urinarybladder,forestomach, and seminal vesicles of

rats. Summaries of the incidences of neoplasms and

nonneoplastic lesions, individual animal tumor

diagnoses, statistical analyses of primary neoplasms

that occurred with an incidence of atleast5%inatleast

one exposuregroup, and historical incidences for the

neoplasms mentioned in this section are presented in

Appendix A formale rats and Appendix B for female

rats.

Liver: Atthe 9-monthinterimevaluation,the absolute

and relative liverweightsof exposed groups of males

and femalesweresignificantlygreaterthan those ofthe

controls(TableH2). One hepatocellular adenomawas

observed in a 5,000 ppm male at 9 months (Tables 8

and A1). Incidencesoffoci of hepatocellularalteration

were increased inmalesin the 10,000ppmgroup, and

aminimalaccumulationofpigmentinhepatocyteswas

present in males and females from the 10,000 ppm

groups and in females from the 5,000 ppm group

(Tables 8, A5, and B5). At the 15-month interim

evaluation, the absolute and relative liver weights of

exposed groups of females were significantly greater

than those of the controls (Table H3). Incidences of

single and multiple hepatocellular adenomas and

carcinomaswereincreasedat15monthsin10,000 ppm

malesand females(Tables8,A1,andB1). Incidences

offoci of hepatocellular alterationand accumulation of

pigmentin hepatocyteswerealsoincreased in exposed

groups of males and females (Tables 8, A5, and B5).

Atthe end ofthe 2-yearstudy, the incidencesofhepatocellular adenoma, hepatocellular carcinoma, and

hepatocellularadenoma or carcinoma (combined)were

significantlyincreasedin all exposed groups of males

and females (Tables 8, A3, and B3). 

The incidences of multiple hepatocellular adenomas

andmultiple hepatocellular carcinomasinexposedmale

and female groups were greater than those of the

controls (Tables 8, A1, and B1). Incidences of

hepatocellular adenoma or carcinoma (combined)inall

exposed groupsofmales and femalesexceeded theNTP

historical ranges(males: 0%-10%;females: 0%-6%)

forfeed studycontrols (Tables 8, A4a, andB4a). The

majority of the benign and malignantliver neoplasms

consisted ofwell-differentiatedneoplastic hepatocytes

with cellular atypia and increasednumbers of mitoses.

Carcinomashad trabecular,glandular,orsolid growth

patterns(Plate1)withareasofnecrosis,cavitation,and

fibrosis. Metastases were common in the lungs

(Plates2 and 3), butmetastatic fociwerealso presentin

the stomach,pancreas,adrenalgland,lymphnode, and

spleen.

The incidences ofsingle andmultiple hepatocholangiocarcinoma were significantly increased in 5,000ppm

malesand femalesand in 10,000ppmfemales(Tables

8, A3, and B3). These neoplasms consisted of a

mixture of malignant hepatocytes and welldifferentiated cuboidal epithelium forming distinct

ductular structures (Plate4). Both hepatocellular and

biliary components of this neoplasm were present in

metastatic foci. In addition, several other benign

(cholangioma and hepatocholangioma) andmalignant

(cholangiocarcinoma)liverneoplasms occurred only in

exposed groups of males and females (Tables 8, A1,

and B1). 

Duringthe2-yearstudy,the incidences ofpigmentation

and foci of hepatocellular alteration (clear cell,

basophilic,and eosinophilic)wereincreased inexposed

groups of males and females (Tables 8,A5, and B5).

Cellsinsome focihad intensely eosinophilic cytoplasm

and hepatocellular atypia similarto the appearance of

cellsinthehepatocellular neoplasms. The pigmentwas

considered to be1-amino-2,4-dibromoanthraquinone or

a metabolite based on the results ofthe histochemical

procedures performed duringthe13-weekstudyand the

15-month interim evaluation.
40 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE 8

Incidences of Neoplasms and Nonneoplastic Lesions of the Liver in Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Male

9-Month Interim Evaluation

Number Examined 10 10 10 10

Basophilic Focusa0 0 1 (1.0)b1 (1.0)

Clear Cell Focus 0 0 0 4* (1.0)

Eosinophilic Focus 0 0 0 1 (1.0)

Pigmentation 0 1 (1.0) 0 6** (1.0)

Hepatocellular Adenoma 0 0 1 0

15-Month Interim Evaluation

Number Examined 10

-

c

- 10

Basophilic Focus 1 (1.0) 5 (1.0)

Clear Cell Focus 0 7** (1.0)

Eosinophilic Focus 1 (1.0) 0

Pigmentation 0 10** (1.0)

Hepatocellular Adenoma (Multiple) 0 2

Hepatocellular Adenoma

(Single or Multiple) 0 4*

Hepatocellular Carcinoma (Multiple) 0 3

Hepatocellular Carcinoma

(Single or Multiple) 0 7**

Hepatocellular Adenoma or Carcinoma 0 10**

2-Year Study

Number Examined 50 40 59 50

Basophilic Focus 9 (1.1) 12 (1.7) 24** (1.8) 22** (1.5)

Clear Cell Focus 3 (1.0) 26** (1.8) 39** (1.8) 27** (1.8)

Eosinophilic Focus 1 (1.0) 13** (2.9) 14** (2.1) 6 (2.6)

Pigmentation 3 (1.0) 19** (1.1) 48** (1.1) 39** (1.1)

Hepatocellular Adenoma (Multiple) 0 10** 23** 24**

Hepatocellular Adenoma (Single or Multiple)

Overall rated1/50 (2%) 20/40 (50%) 40/59 (68%) 34/50 (68%)

Terminal ratee1/26 (4%) 16/24 (67%) 18/21 (86%) 9/10 (90%)

Adjusted ratef3.8% 71.3% 92.3% 97.0%

First incidence (days) 729 (T) 675 521 435

Logistic regression testgP<0.001 P<0.001 P<0.001 P<0.001

Hepatocellular Carcinoma (Multiple) 0 1 43** 37**

Hepatocellular Carcinoma (Single or Multiple)

Overall rate 1/50 (2%) 12/40 (30%) 55/59 (93%) 46/50 (92%)

Terminal rate 0/26 (0%) 9/24 (38%) 21/21 (100%) 10/10 (100%)

Adjusted rate 2.7% 43.5% 100.0% 100.0%

First incidence (days) 666 650 465 436

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

(continued)
Results 41

TABLE 8

Incidences of Neoplasms and Nonneoplastic Lesions of the Liver in Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Male (continued)

2-Year Study (continued)

Hepatocellular Adenoma or Carcinomah

Overall rate 2/50 (4%) 25/40 (63%) 57/59 (97%) 47/50 (94%)

Terminal rate 1/26 (4%) 19/24 (79%) 21/21 (100%) 10/10 (100%)

Adjusted rate 6.4% 83.1% 100.0% 100.0%

First incidence (days) 666 650 465 435

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Number Examined 50 40 59 50

Hepatocholangioma 0 0 1 1

Hepatocholangiocarcinoma 0 0 6* 2

Cholangioma 0 0 2 0

Cholangiocarcinoma 0 0 0 1

Female

9-Month Interim Evaluation

Number Examined 10 10 10 10

Basophilic Focus 1 (1.0) 0 0 1 (1.0)

Clear Cell Focus 0 0 0 1 (1.0)

Pigmentation 0 2 (1.0) 6** (1.0) 6** (1.0)

15-Month Interim Evaluation

Number Examined 10

- - 10

Basophilic Focus 8 (1.0) 9 (1.4)

Clear Cell Focus 0 5* (1.6)

Pigmentation 1 (1.0) 10** (1.1)

Hepatocellular Adenoma (Multiple) 0 5*

Hepatocellular Adenoma

(Single or Multiple) 0 8**

Hepatocellular Carcinoma (Multiple) 0 3

Hepatocellular Carcinoma

(Single or Multiple) 0 6**

Hepatocellular Adenoma or Carcinoma 0 9**

2-Year Study

Number Examined 50 40 60 48

Basophilic Focus 39 (1.3) 15** (1.6) 22** (1.7) 16** (1.4)

Clear Cell Focus 3 (1.3) 28** (1.6) 39** (2.0) 17** (1.6)

Eosinophilic Focus 7 (1.4) 23** (2.0) 12 (2.5) 1 (4.0)

Pigmentation 1 (1.0) 19** (1.1) 51** (1.4) 45** (1.1)

(continued)
42 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE 8

Incidences of Neoplasms and Nonneoplastic Lesions of the Liver in Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Female (continued)

2-Year Study (continued)

Number Examined 50 40 60 48

Hepatocellular Adenoma (Multiple) 0 18** 39** 22**

Hepatocellular Adenoma (Single or Multiple)

Overall rate 0/50 (0%) 28/40 (70%) 47/60 (78%) 29/48 (60%)

Terminal rate 0/38 (0%) 23/32 (72%) 29/38 (76%) 8/12 (67%)

Adjusted rate 0.0% 75.5% 83.7% 83.6%

First incidence (days)_ -

i

600 575 418

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Hepatocellular Carcinoma (Multiple) 0 7** 51** 41**

Hepatocellular Carcinoma (Single or Multiple)

Overall rate 0/50 (0%) 12/40 (30%) 57/60 (95%) 45/48 (94%)

Terminal rate 0/38 (0%) 12/32 (38%) 37/38 (97%) 12/12 (100%)

Adjusted rate 0.0% 37.5% 98.3% 100.0%

First incidence (days) - 729 (T) 575 460

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Hepatocellular Adenoma or Carcinomaj

Overall rate 0/50 (0%) 33/40 (83%) 59/60 (98%) 47/48 (98%)

Terminal rate 0/38 (0%) 28/32 (88%) 38/38 (100%) 12/12 (100%)

Adjusted rate 0.0% 89.1% 100.0% 100.0%

First incidence (days) - 600 575 418

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Hepatocholangioma 0 0 2 0

Hepatocholangiocarcinoma 0 0 11** 13**

Cholangioma 0 0 0 1

* Significantly different (P#0.05) from the control group by the Fisher exact test (9-month and 15-month interim evaluations) or the logistic

regression test (2-year study)

** P#0.01

(T)Terminal sacrifice

a Number of animals with lesion

b Average severity grade of lesions in affected rats: 1 = minimal; 2 = mild; 3 = moderate; 4 = marked

c Liver not microscopically examined in this group

d Number of animals with neoplasm per number of animals with liver examined microscopically

e Observed incidence in animals surviving until the end of the study

f Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality.

g

In the control column are the P values associated with the trend test. In the exposed group columns are the P values corresponding to

pairwise comparisons between the controls and that exposure group. The logistic regression test regards these lesions as nonfatal.

h Historical incidence for 2-year NTP feed studies with untreated control groups (mean ± standard deviation): 45/1,350 (3.3% ± 3.6%); range,

0%-10% i Not applicable; no neoplasms in animal group

j Historical incidence: 9/1,351 (0.7% ± 1.5%); range, 0%-6%
Results 43

Large Intestine: Adenomatous polyps (adenomas)

were observed in the large intestine of 10,000 ppm

males and females at the 15-month interim evaluation (Tables 9, A1, and B1).

At 2 years, the incidences of adenomatous polyps

(adenomas) in the rectum were significantly

increased in all exposed groups of males and

females (Tables 9, A3, and B3). The incidence of

carcinom a of the 

TABLE 9

Incidences of Neoplasms of the Large Intestine in Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Male

15-Month Interim Evaluation

Rectuma9

-

c

- 10

Adenomatous Polyp (Adenoma)b0 6**

2-Year Study

Colon

Adenomatous Polyp (Adenoma)

Overall rated0/50 (0%) 1/40 (3%) 1/59 (2%) 3/50 (6%)

Terminal ratee0/26 (0%) 1/24 (4%) 0/21 (0%) 1/10 (10%)

Adjusted ratef0.0% 4.2% 4.3% 19.9%

First incidence (days) -

h

729 (T) 720 590

Logistic regression testgP=0.027 P=0.484 P=0.494 P=0.081 

Carcinomai

Overall rate 0/50 (0%) 0/40 (0%) 1/59 (2%) 4/50 (8%)

Terminal rate 0/26 (0%) 0/24 (0%) 1/21 (5%) 0/10 (0%)

Adjusted rate 0.0% 0.0% 4.8% 20.4%

First incidence (days) - - 729 (T) 590

Logistic regression test P=0.003 - P=0.457 P=0.046

Rectum

Adenomatous Polyp (Adenoma)

Overall rate 0/50 (0%) 13/40 (33%) 51/59 (86%) 40/50 (80%)

Terminal rate 0/26 (0%) 9/24 (38%) 21/21 (100%) 10/10 (100%)

Adjusted rate 0.0% 45.8% 100.0% 100.0%

First incidence (days) - 659 478 352

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Carcinomai

Overall rate 0/50 (0%) 1/40 (3%) 10/59 (17%) 15/50 (30%)

Terminal rate 0/26 (0%) 0/24 (0%) 5/21 (24%) 4/10 (40%)

Adjusted rate 0.0% 3.8% 32.4% 63.0%

First incidence (days) - 718 608 493

Logistic regression test P<0.001 P=0.480 P=0.003 P<0.001

Large Intestine (All Sites) 50 40 59 50

Adenomatous Polyp (Adenoma) (Multiple) 0 1 34** 32**

Adenomatous Polyp (Adenoma) 

(Single or Multiple) 0 13** 51** 40**

Carcinoma (Multiple) 0 0 0 3

Carcinoma (Single or Multiple) 0 1 11** 17**

(continued)
44 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE 9

Incidences of Neoplasms of the Large Intestine in Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Female

15-Month Interim Evaluation

Rectum 10

- - 10 Adenomatous Polyp (Adenoma) 0 2

2-Year Study

Colon 50 40 60 49

Adenomatous Polyp (Adenoma) 0 1 2 2

Carcinomaj0 1 2 1

Rectum

Adenomatous Polyp (Adenoma)

Overall rate 0/50 (0%) 27/40 (68%) 53/60 (88%) 43/49 (88%)

Terminal rate 0/38 (0%) 23/32 (72%) 38/38 (100%) 12/12 (100%)

Adjusted rate 0.0% 75.0% 100.0% 100.0%

First incidence (days) - 616 582 512

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Carcinomaj

Overall rate 0/50 (0%) 1/40 (3%) 19/60 (32%) 7/49 (14%)

Terminal rate 0/38 (0%) 1/32 (3%) 13/38 (34%) 4/12 (33%)

Adjusted rate 0.0% 3.1% 41.7% 41.9%

First incidence (days) - 729 (T) 606 625

Logistic regression test P<0.001 P=0.466 P<0.001 P=0.001

Large Intestine (All Sites) 50 40 60 49

Adenomatous Polyp (Adenoma) (Multiple) 0 18** 46** 32**

Adenomatous Polyp (Adenoma) 

(Single or Multiple) 0 28** 53** 43**

Carcinoma (Multiple) 0 1 1 1

Carcinoma (Single or Multiple) 0 2 21** 8**

** Significantly different (P#0.01) from the control group by the Fisher exact test (15-month interim evaluation) or the logistic regression test

(2-year study)

(T)Terminal sacrifice

a Number of animals with large intestine examined microscopically

b Number of animals with lesion

c Large intestine not microscopically examined in this group

d Number of animals with neoplasm per number of animals necropsied

e Observed incidence in animals surviving until the end of the study

f Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality.

g

In the control column are the P values associated with the trend test. In the exposed group columns are the P values corresponding to

pairwise comparisons between the controls and that exposure group. The logistic regression test regards these lesions as nonfatal.

h Not applicable; no neoplasms in animal group

i Historical incidence for 2-year NTP feed studies with untreated control groups (mean ± standard deviation): 1/1,353 (0.1% ± 0.4%); range,

0%-2% (includes all carcinomas of the large intestine)

j Historical incidence: 0/1,351 (includes all carcinomas of the large intestine)
Results 45

colon was significantly increased in 10,000 ppm males,

and the incidences of rectal carcinoma were significantly

increased in 5,000 and 10,000 ppm males and females

(Tables 9, A3, and B3). The intestinal neoplasms that

occurred in the distal colon and rectum of rats were

morphologically similar.

Adenomatous polyps ( adenomas) con sisted of

pedunculated, exophytic masses (Plate 5) of well-differentiated, columnar epithelium with prominent, hyperchromatic nuclei. Carcinoma (adenocarcinoma) was

generally similar to adenoma, except that invasion of the

stromal stalk of the neoplasm and extension into the

submucosa, and occasionally into the muscular wall, were

evident microscopically. In the malignant neoplasms,

irregular glandular structures or cords of atypical epithelial

cells were present in the submucosa (Plate 6) and were

frequently associated with a scirrhous response. In males,

metastatic colon carcinoma was observed in the lung and

mesenteric lymph nodes, and metastatic rectal carcinoma

was observed in the lumbar lymph nodes and pancreas

(Table A1). Incidences of carcinoma (colon and rectum

combined) in exposed groups of males and females

exceeded the NTP historical ranges (males: 0%-2%;

females: 0%) for feed study controls (Tables 9, A4b, and

B4b).

Kidney: At the 9-month interim evaluation, the relative

kidney weights of exposed groups of males and females

were significantly greater than those of the controls (Table

H2). Pigmentation and hyaline droplet accumulation were

present in the kidneys of all exposed males (Tables 10 and

A5). The sizes of some renal tubule nuclei were minimally increased in males and females from all exposure

groups, and the severity of the nephropathy (tubule

epithelial regeneration, mononuclear inflammation, and

renal tubule dilation with protein casts) was slightly more

severe than that observed in the controls (Table 10). At

the 15-month interim evaluation, the relative kidney

weights of 10,000 ppm males and females were significantly greater than those of the controls (Table H3). The

severity of nephropathy was increased in 10,000 ppm

males and females compared to controls (Table 10). This

was characterized by an increase in the foci of tubule

epithelial regeneration; nuclear enlargement in some

tubule epithelium, similar to that in the 13-week studies

and at the 9-month interim evaluation, was also present.

At 15 months in the 10,000 ppm groups, pigmentation of

renal tubule epithelium was present in all rats; renal tubule

epithelial hyperplasia was observed in two males

and three females; and adenomas were observed in two

males (Tables 10, A1, A5, and B5).

At 2 years, there was a significant dose-related increase in

the incidences of renal tubule adenoma in exposed groups

of males and females (Tables 10, A3, and B3). Multiple

adenomas were observed in all exposed groups of males

and in the 5,000 and 10,000 ppm females. Adenomas

were expansile lesions involving one or more adjacent

tubules and were generally five or more times the diameter

of the normal renal tubule. The cells within the adenomas

were generally similar in morphology to those in the focal

hyperplastic lesions. Carcinomas were larger than adenomas and frequently had more cellular atypia, necrosis,

and local invasion. One carcinoma in a male rat metastasized to the lung, and one in a female rat metastasized to

the adrenal gland. Renal tubule carcinomas occurred in

two 5,000 ppm males, one 10,000 ppm male, and two

10,000 ppm females. The combined incidences of renal

tubule adenoma or carcinoma were significantly increased

in exposed males and females and exceeded the NTP

historical ranges (males: 0%-6%; females: 0%-2%) for

feed study controls (Tables 10, A4c, and B4c).

Incidences of renal tubule hyperplasia were significantly

increased in exposed males and females (Tables 10, A5,

and B5). Hyperplasia consisted of a tubule lined by two

or more layers of renal tubule epithelium; these were most

often located in the cortex or outer stripe of the outer

medulla. Foci of hyperplasia were distinguished from the

more basophilic foci of tubule epithelial regeneration

typically associated with nephropathy. There was no clear

dose-related increase in the incidence or severity of

nephropathy in rats at 2 years (Table 10). The incidences

and severity of transitional cell hyperplasia in the renal

pelvis were increased in exposed groups of males and

females (Tables 10, A5, and B5); there were no increases

in the incidences of transitional cell papilloma or carcinoma of the renal pelvis (Tables 10, A1, and B1). The

incidence of a reddish brown pigment within the renal

tubule epithelium and lumina of exposed rats was increased at the 9- and 15-month interim evaluations and at

2 years. The pigment was characterized in the 13-week

study and at the 15-month interim evaluation as PASnegative; resistant to digestion by diastase; isotropic; and

negative for melanin, hemosiderin, hematoidin, bile,

lipofuscin, or ceroid staining methods. The pigment was

presumed to be 1-amino-2,4-dibromoanthraquinone or one

of its metabolites.
46 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE 10

Incidences of Neoplasms and Nonneopla stic Lesions of the Kidney in Rats in the 2-Year Feed Study

of 1-Amino-2,4-dibromoanthraquinone

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Male

9-Month Interim Evaluation

Number Examined 10 10 10 10 Renal Tubule Hyaline Droplet

Accumulationa0 10** (2.0)b10** (2.0) 10** (1.9)

Renal Tubule Pigmentation 0 10** (1.1) 10** (1.4) 10** (1.9)

15-Month Interim Evaluation

Number Examined 10 _c _ 10

Nephropathy 10 (2.0) 10 (2.5)

Renal Tubule Hyperplasia 0 2 (1.0)

Renal Tubule Pigmentation 0 10** (2.0)

Transitional Cell Hyperplasia 0 4* (1.3)

Renal Tubule Adenoma 0 2

2-Year Study

Number Examined 50 40 59 50

Nephropathy 50 (2.9) 40 (3.6) 59 (2.9) 49 (2.7)

Renal Tubule Hyperplasia 9 (2.2) 30** (2.9) 25** (2.4) 19** (2.9)

Renal Tubule Pigmentation 5 (2.0) 40** (1.9) 58** (2.0) 49** (1.9)

Transitional Cell Hyperplasia 30 (1.4) 40** (2.1) 51** (1.9) 35* (1.6)

Transitional Cell Papilloma 0 0 1 0

Renal Tubule Adenoma (Multiple) 0 4* 4 5*

Renal Tubule Adenoma (Single or Multiple)

Overall rated2/50 (4%) 10/40 (25%) 11/59 (19%) 14/50 (28%)

Terminal ratee2/26 (8%) 6/24 (25%) 4/21 (19%) 5/10 (50%)

Adjusted ratef7.7% 33.6% 34.6% 68.3%

First incidence (days) 729 (T) 618 636 588

Logistic regression testgP<0.001 P=0.007 P=0.014 P<0.001

Renal Tubule Carcinoma 0 0 2 1

Renal Tubule Adenoma or Carcinomah

Overall rate 2/50 (4%) 10/40 (25%) 13/59 (22%) 15/50 (30%)

Terminal rate 2/26 (8%) 6/24 (25%) 4/21 (19%) 5/10 (50%)

Adjusted rate 7.7% 33.6% 39.4% 69.1%

First incidence (days) 729 (T) 618 636 497

Logistic regression test P<0.001 P=0.007 P=0.005 P<0.001

(continued)
Results 47

TABLE 10

Incidences of Neoplasms and Nonneoplastic Lesions of the Kidney in Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Female

15-Month Interim Evaluation

Number Examined 10

- - 10

Nephropathy 10 (1.7) 10 (2.5)

Renal Tubule Hyperplasia 0 3 (1.0)

Renal Tubule Pigmentation 0 10** (2.0)

Transitional Cell Hyperplasia 3 (1.0) 4 (1.8)

2-Year Study

Number Examined 50 40 60 48

Nephropathy 50 (1.9) 39 (2.6) 60 (2.7) 46 (2.7)

Renal Tubule Hyperplasia 1 (3.0) 12** (2.5) 23** (2.7) 27** (2.7)

Renal Tubule Pigmentation 0 40** (2.0) 60** (2.0) 48** (2.0)

Transitional Cell Hyperplasia 10 (1.2) 16* (1.8) 44** (1.5) 21** (1.6)

Transitional Cell Papilloma 0 0 0 1

Transitional Cell Carcinoma 0 1 0 0

Renal Tubule Adenoma (Multiple) 0 0 5* 5*

Renal Tubule Adenoma (Single or Multiple)

Overall rate 0/50 (0%) 3/40 (8%) 16/60 (27%) 16/48 (33%)

Terminal rate 0/38 (0%) 1/32 (3%) 11/38 (29%) 6/12 (50%)

Adjusted rate 0.0% 8.0% 36.0% 69.7%

First incidence (days) -

i

600 601 625

Logistic regression test P<0.001 P=0.049 P<0.001 P<0.001

Renal Tubule Carcinoma 0 0 0 2

Renal Tubule Adenoma or Carcinomaj

Overall rate 0/50 (0%) 3/40 (8%) 16/60 (27%) 16/48 (33%)

Terminal rate 0/38 (0%) 1/32 (3%) 11/38 (29%) 6/12 (50%)

Adjusted rate 0.0% 8.0% 36.0% 69.7%

First incidence (days) - 600 601 625

Logistic regression test P<0.001 P=0.049 P<0.001 P<0.001

* Significantly different (P#0.05) from the control group by the Fischer exact test (9-month and 15-month interim evaluations) or the logistic

regression test (2-year study) 

** P#0.01

(T)Terminal sacrifice

a Number of animals with lesion

b Average severity grade of lesions in affected rats: 1 = minimal; 2 = mild; 3 = moderate; 4 = marked

c Kidney not microscopically examined in this group

d Number of animals with neoplasm per number of animals with kidney examined microscopically

e Observed incidence in animals surviving until the end of the study

f Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality.

g

In the control column are the P values associated with the trend test. In the exposed group columns are the P values corresponding to

pairwise comparisons between the controls and that exposure group. The logistic regression test regards these lesions as nonfatal.

h Historical incidence for 2-year NTP feed studies with untreated control groups (mean ± standard deviation): 15/1,350 (1.1% ± 1.7%); range,

0%-6% i Not applicable; no neoplasms in animal group

j Historical incidence: 1/1,348 (0.1% ± 0.4%); range, 0%-2%
48 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

Urinary Bladder: Hyperplasiaofthe transitional cell

epithelium of the urinary bladder was observed in

10,000 ppmfemales at 9 months and in 10,000 ppm

males and females at 15months(Tables 11, A5, and

B5). Transitional cellhyperplasiawas presentinmost

5,000 and 10,000 ppm females at 2 years, and the

incidences of this lesion were significantly increased

in 5,000and10,000ppmmales. Hyperplasia consisted

of a diffuse or focal increase in thickness of the

transitional epithelium; minimal cellular

pleomorphismand increased numbersofmitotic cells

were sometimespresent. Other nonneoplastic lesions

that occurredonly in exposed rats included squamous

metaplasia of the transitional epithelium and fatty

metaplasia (fat proliferation) of the stroma of the

bladder wall.

Two transitional cell carcinomas and one papilloma

occurred in 10,000 ppm females at the 15-month

interimevaluation. At 2 years,incidences of transitional cell papilloma, carcinoma, and papilloma or

carcinoma (combined)were significantlyincreased in

10,000ppmmales and 5,000 and 10,000 ppm females

(Tables 11,A3, andB3) andexceeded theNTPhistoricalranges(males: 0%-2%; females: 0%-2%)forfeed

study controls (Tables 11, A4d, and B4d). Transitional cell papilloma consisted of a pedunculated or

broad-basedmass of transitional epitheliumwith a

central fibrovascular stroma; there was squamous

metaplasia of the surface epitheliumin some papillomas. Transitional cell carcinoma was characterized

by an exophytic or endophytic growth pattern and

invasion of the lamina propria or muscularis of the

bladder wall (Plates 7 and 8). There was cellular

atypia and squamous or mucous metaplasia of transitional epithelium in some carcinomas.
Results 49

TABLE 11

Incidences of Neoplasms and Nonneopla stic Lesions of the Urinary Bladder in Rats

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Male

15-Month Interim Evaluation

Number Examined 10 _b _ 10

Transitional Cell Hyperplasia

a

0 3 (1.3)

c

Metaplasia, Squamous 0 1 (2.0)

2-Year Study

Number Examined 50 38 58 50

Transitional Cell Hyperplasia 1 (2.0) 5 (2.0) 17** (1.9) 30** (2.1)

Metaplasia, Squamous 0 0 0 3 (1.7)

Transitional Cell Papilloma

Overall rated0/50 (0%) 1/38 (3%) 2/58 (3%) 8/50 (16%)

Terminal ratee0/26 (0%) 0/22 (0%) 2/21 (10%) 2/10 (20%)

Adjusted ratef0.0% 3.7% 9.5% 40.3%

First incidence (days) _h700 729 (T) 493

Logistic regression testgP<0.001 P=0.459 P=0.192 P=0.004

Transitional Cell Carcinoma

Overall rate 0/50 (0%) 0/38 (0%) 1/58 (2%) 4/50 (8%)

Terminal rate 0/26 (0%) 0/22 (0%) 0/21 (0%) 1/10 (10%)

Adjusted rate 0.0% 0.0% 4.3% 24.5%

First incidence (days) _ _720 674

Logistic regression test P=0.001 _ P=0.491 P=0.022

Transitional Cell Papilloma or Carcinomai

Overall rate 0/50 (0%) 1/38 (3%) 3/58 (5%) 12/50 (24%)

Terminal rate 0/26 (0%) 0/22 (0%) 2/21 (10%) 3/10 (30%)

Adjusted rate 0.0% 3.7% 13.5% 56.2%

First incidence (days) ) 700 720 493

Logistic regression test P<0.001 P=0.459 P=0.096 P<0.001

Female

9-Month Interim Evaluation

Number Examined 10 10 10 10

Transitional Cell Hyperplasia 0 0 0 2 (1.5)

15-Month Interim Evaluation

Number Examined 10 _ _

10

Transitional Cell Hyperplasia 0 9** (2.6)

Transitional Cell Papilloma 0 1

Transitional Cell Carcinoma 0 2

Squamous Cell Carcinoma 0 2

(continued)
50 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE 11

Incidences of Neoplasms and Nonneopla stic Lesions of the Urinary Bladder in Rats

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Female (continued)

2-Year Study

Number Examined 50 40 60 46

Transitional Cell Hyperplasia 1 (1.0) 2 (3.0) 41** (2.0) 41** (2.3)

Metaplasia, Squamous 0 1 (1.0) 4 (2.3) 8** (2.9)

Fat Proliferation 0 0 4 (2.3) 2 (2.5)

Transitional Cell Papilloma

Overall rate 0/50 (0%) 2/40 (5%) 7/60 (12%) 9/46 (20%)

Terminal rate 0/38 (0%) 2/32 (6%) 6/38 (16%) 1/12 (8%)

Adjusted rate 0.0% 6.3% 17.6% 39.5%

First incidence (days) _729 (T) 691 637

Logistic regression test P<0.001 P=0.201 P=0.012 P=0.003

Transitional Cell Carcinoma

Overall rate 0/50 (0%) 0/40 (0%) 8/60 (13%) 16/46 (35%)

Terminal rate 0/38 (0%) 0/32 (0%) 6/38 (16%) 4/12 (33%)

Adjusted rate 0.0% 0.0% 19.5% 55.8%

First incidence (days) _ _670 367

Logistic regression test P<0.001 _ P=0.008 P<0.001

Transitional Cell Papilloma or Carcinomaj

Overall rate 0/50 (0%) 2/40 (5%) 17/60 (28%) 26/46 (57%)

Terminal rate 0/38 (0%) 2/32 (6%) 14/38 (37%) 6/12 (50%)

Adjusted rate 0.0% 6.3% 40.9% 78.1%

First incidence (days) _729 (T) 670 367

Logistic regression test P<0.001 P=0.201 P<0.001 P<0.001

Squamous Cell Papilloma

(Single or Multiple) 0 0 1 2

Squamous Cell Carcinoma 0 0 1 0

** Significantly different (P#0.01) from the control group by the Fisher exact test (9-month and 15-month interim evaluations) or the logistic regression

test (2-year study)

(T)Terminal sacrifice

a Number of animals with lesion

b Urinary bladder not microscopically examined in this group

c Average severity grade of lesions in affected rats: 1 = minimal; 2 = mild; 3 = moderate; 4 = marked

d Number of animals with neoplasm per number of animals necropsied or examined microscopically

e Observed incidence in animals surviving until the end of the study

f Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality.

g

In the control column are the P values associated with the trend test. In the exposed group columns are the P values corresponding to pairwise

comparisons between the controls and that exposure group. The logistic regression test regards these lesions as nonfatal.

h Not applicable; no neoplasms in animal group

i Historical incidence for 2-year NTP feed studies with untreated control groups (mean ± standard deviation): 3/1,329 (0.2% ± 0.6%); range, 0%-2%

j Historical incidence: 3/1,334 (0.2% ± 0.6%); range, 0%-2%
Results 51

Forestomach: Several proliferative and degenerative

lesions occurredwithincreasedincidencesin the forestomach of exposed males and females(Table 12).

These mucosallesions frequently occurred together

and consisted of thickening (hyperplasia) of the

squamous epithelium and an increase

in the surface keratin layers (hyperkeratosis)

(Tables 12, A5, andB5). Focal areas of hyperplasia

were sometimes adjacentto ulceration and inflammation ofthe squamousmucosa. Therewas no significant increase in incidences of neoplasms of the

forestomach (Tables 12, A1, and B1).

TABLE 12

Incidences of Neoplasms and Nonneopla stic Lesions of the Forestomach in Rats

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Male

Number Examined 49 39 59 49

Hyperkeratosis

a

5 (2.4)

b

18** (1.8) 21** (2.0) 20** (2.0)

Hyperplasia, Squamous 3 (1.7) 19** (3.0) 25** (3.2) 26** (3.0)

Inflammation, Chronic Active 3 (2.0) 12** (2.4) 11 (2.0) 11* (2.2)

Ulcer 3 (2.7) 10** (3.2) 15** (2.7) 16** (2.6)

Squamous Cell Papillomac0 2 0 1

Squamous Cell Carcinoma 0 0 0 1

Squamous Cell Papilloma or Carcinomad0 2 0 2

Female

Number Examined 49 40 60 47

Hyperkeratosis 2 (2.0) 7* (1.4) 23** (2.1) 28** (1.9)

Hyperplasia, Squamous 2 (2.0) 7* (1.9) 26** (2.9) 33** (3.0)

Inflammation, Chronic Active 0 1 (2.0) 13** (2.2) 10** (2.2)

Ulcer 1 (2.0) 2 (2.5) 7 (1.7) 17** (2.9)

Squamous Cell Papilloma 0 0 0 1

Squamous Cell Carcinoma 0 1 1 1

Squamous Cell Papilloma or Carcinomae0 1 1 2

* Significantly different (P#0.05) from the control group by the logistic regression test

** P#0.01

a Number of animals with lesion

b Average severity grade of lesions in affected animals: 1 = minimal; 2 = mild; 3 = moderate; 4 = marked

c Number of animals with neoplasm per number of animals necropsied

d Historical incidence for 2-year NTP feed studies with untreated control groups (mean ± standard deviation): 4/1,353 (0.3% ± 0.7%); range, 0%-2% e Historical incidence: 2/1,351 (0.2% ± 0.5%); range, 0%-2%

MiscellaneousNeoplasms andNonneoplasticLesions:

In exposed males and females, incidences of

mononuclear cell leukemia occurred with significant

negative trends(Tables 13, A3, and B3). The incidences of pituitary gland adenoma in males and

females (males: 0 ppm, 21/48; 2,000 ppm, 14/40;

5,000ppm, 10/56;10,000ppm, 10/49;females: 32/50,

19/39, 32/60, 13/47; Tables A3 and B3) and the

incidence of mammarygland fibroadenoma (21/50,

10/40, 9/60, 5/49; Table B3)in females also occurred

withsignificantnegative trends. These decreasesmay

have been related to lower body weights.
52 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE 13

Incidences of Mononuclear Cell Leukemia in Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone 

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Male

15-Month Interim Evaluation

Mononuclear Cell Leukemia

a

0/10 _b _2/10

2-Year Study

Mononuclear Cell Leukemia

Overall ratea25/50 (50%) 5/40 (13%) 3/59 (5%) 1/50 (2%)

Terminal ratec9/26 (35%) 4/24 (17%) 2/21 (10%) 0/10 (0%)

Adjusted rated59.0% 18.8% 11.7% 2.9%

First incidence (days) 514 604 650 590

Life table testeP<0.001N P<0.001N P<0.001N P<0.001N

Logistic regression test P<0.001N P<0.001N P<0.001N P<0.001N

Female

2-Year Study

Mononuclear Cell Leukemia

Overall rate 9/50 (18%) 1/40 (3%) 5/60 (8%) 1/49 (2%)

Terminal rate 6/38 (16%) 0/32 (0%) 1/38 (3%) 0/12 (0%)

Adjusted rate 21.5% 3.0% 10.0% 3.7%

First incidence (days) 620 689 601 662

Life table test P=0.112N P=0.026N P=0.177N P=0.162N

Logistic regression test P=0.011N P=0.023N P=0.111N P=0.024N

a Number of animals with neoplasm per number of animals necropsied 

b Animals not microscopically examined in this group

c Observed incidence in animals surviving until the end of the study

d Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality.

e

In the control column are the P values associated with the trend test. In the exposed group columns are the P values corresponding to

pairwise comparisons between the controls and that exposure group. The life table analysis regards neoplasms in animals dying prior to

terminal kill as being (directly or indirectly) the cause of death. The logistic regression test regards these lesions as nonfatal. A negative

trend or a lower incidence in an exposure group is indicated by N.

In males, there was a chemical-related increased

incidence of atrophy of the seminal vesicles (1/49,

30/40, 35/59, 23/50; Table A5). This was not

evident at the 9- or 15-month interim evaluation,

but was present in most males at the end of the 2-

year study. 

Atrophy of the seminal vesicles of exposed males

was characterized by a reduction in the size of the

secretory epithelium from a tall columnar shape to

a low cuboidal shape and by an increase in the

amount of connective tissue stroma in the gland. 
Results 53

STOP-EXPOSURE EVALUATION

Stop-exposure groups of male and female rats were

included in the NTP 2-year study to evaluate the

potential for progression or regression of chemicalrelated liver,largeintestine,kidney, urinarybladder,and

forestomachlesionsduringa recoveryperiod. Tenmale

and 10 female rats were exposed to 20,000 ppm

1-amino-2,4-dibromoanthraquinone infeed for9months

followedbyadministrationofundosed feed untilthe end

ofthe15-monthperiod (9-monthstop-exposuregroups).

In addition, 30 males and 30 femaleswereexposed to

20,000ppm1-amino-2,4-dibromoanthraquinoneinfeed

for15months(15-monthexposuregroups). Tenmales

and 10 females from the 15-month exposure groups

were evaluated at the 9-month interim evaluation

(9-month interim evaluation groups).

Survival

Estimatesof 2-year survivalprobabilitiesformale and

female rats are shown in the Kaplan-Meier survival

curvesin Figure 4. All malessurvived9months;three

malesfromthe 9-monthstop-exposuregroup and three

malesfromthe15-monthexposuregroup died beforethe

end ofthe 15-monthevaluation (Table14). Allfemales

in the 9-monthstop-exposuregroup survived until the

end ofthe 15-monthevaluation. One female from the

15-monthexposuregroup died duringthe first9months;

an additionalseven femalesdied betweenmonth9 and

the end of the 15-month evaluation (Table 15).

Body Weights

and Feed and Compound Consumption

Themeanbodyweightsof male and female ratsin the

9-monthstop-exposure and 15-monthexposuregroups

are compared with the controls from the 2-year core

study in Tables 14 and 15, and the growth curves for

exposed rats in the 15-month exposure groups are

shown in Figure 5. The mean body weights of males

and femalesin the 9-monthstop-exposure groupswere

20% to 22% lower than those of the controls at the

9-monthinterimevaluationofthe 2-yearcore studyand

were20%to 21%lowerthanthose ofthe controlsatthe

15-monthinterimevaluation of the 2-year core study.

The mean body weights of males and females in the

15-month exposure groups were 19% to 21% lower

than those of the controls at the 9-month interim

evaluation of the 2-year core study and were 25% to

33% lower than those ofthe controls at the 15-month

interim evaluation of the 2-year core study. Feed

consumption by9-monthstop-exposure and 15-month

exposuremalesand females was generally lowerthan

that bythe controlsthroughout the study(TablesJ1 and

J2). The dietary level of 20,000 ppm delivered daily

doses of approximately 1,300 mg 1-amino2,4-dibromoanthraquinone/kgbodyweighttomalesand

1,800mg/kg to femalesin the 9-monthstop-exposure

groups and daily doses of approximately1,100mg/kg

tomales and 1,400 mg/kg to femalesin the 15-month

exposure groups.
54 l-Amino-2,4-dibromoanthraquinone, NTP TR 383

FIGURE 4

Kaplan-Meier Survival Curves for Rats Administered l-Amino-2,4-dibromoanthraqninone in Feed 

in the 15-Month Stop-Exposure Evaluation 
Results 55 

FIGURE 5

Growth Curves for Rats Administered l-Amino-2,4-dibromoanthraquinone in Feed 

in the 15-Month Stop-Exposure Evaluation 
56 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE 14

Mean Body Weights and Survival of Male Rats in the Stop-Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone

Weeks 0 ppm 20,000 ppm (9-Month Stop-Exposure) 20,000 ppm (15-Month Exposure)

on Av. Wt.Number of Av. Wt. Wt. (% of Number of Av. Wt. Wt. (% of Number of

Study (g) Survivors (g) controls) Survivors (g) controls) Survivors

 1 139 70 130 94 10 136 98 30

 2 161 70 144 90 10 149 93 30

 3 206 70 160 78 10 167 81 30

 4 235 70 169 72 10 173 74 30

 5 240 70 169 71 10 177 74 30

 6 269 70 193 72 10 201 75 30

 7 287 70 204 71 10 211 74 30

 8 302 70 211 70 10 217 72 30

 9 312 70 225 72 10 226 73 30

10 325 70 239 73 10 247 76 30

11 333 70 255 76 10 264 79 30

12 338 70 267 79 10 277 82 30

13 332 70 267 81 10 271 82 30

14 356 70 284 80 10 292 82 30

17 387 70 305 79 10 318 82 30

21 406 70 327 81 10 334 82 30

25 423 70 341 81 10 351 83 30

29 435 70 338 78 10 355 82 30

33 445 70 351 79 10 364 82 30

37 453 70 352 78 10 364 80 30

41 468 60a 374 80 10b 377 81 20a

45 473 60 384 81 10 381 81 20

49 479 60 395 83 10 383 80 20

53 489 60 398 81 10 389 80 20

57 486 60 393 81 10 377 78 20

61 484 59 379 78 9 370 77 19

65 484 57 388 80 7 364 75 17

Mean for weeks

1-13 268 203 76 209 78

14-37 415 328 79 340 82

41-65 480 387 81 377 79

a

Interim evaluation occurred during week 39. b Animals switched to undosed feed
Results 57

TABLE 15

Mean Body Weights and Survival of Female Rats in the Stop-Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone

Weeks 0 ppm 20,000 ppm (9-Month Stop-Exposure) 20,000 ppm (15-Month Exposure)

on Av. Wt.Number of Av. Wt. Wt. (% of Number of Av. Wt. Wt. (% of Number of

Study (g) Survivors (g) controls) Survivors (g) controls) Survivors

 1 93 70 90 96 10 94 101 30

 2 114 70 93 82 10 97 85 30

 3 133 70 103 78 10 104 78 30

 4 147 70 113 77 10 114 78 30

 5 157 70 117 75 10 121 77 30

 6 166 70 119 72 10 129 78 30

 7 173 70 129 75 10 141 81 30

 8 180 70 141 78 10 150 84 30

 9 186 70 146 79 10 157 84 30

10 191 70 159 83 10 163 85 30

11 196 70 163 83 10 166 85 29

12 203 70 168 83 10 171 84 29

13 208 70 172 83 10 175 84 29

14 212 70 177 84 10 179 85 29

17 222 70 188 85 10 190 85 29

21 228 70 194 85 10 196 86 29

25 237 70 198 84 10 200 85 29

29 246 70 200 82 10 203 83 29

33 251 70 201 80 10 203 81 29

37 258 70 207 80 10 211 82 29

41 265 59a 207 78 10 10b 210 79 19a

45 272 59 213 78 16

49 284 59 231 81 10 213 75 15

53 299 59 248 83 10 219 73 15

57 311 59 252 81 10 220 71 15

61 315 59 217 69 15

65 328 59 258 79 10 220 67 12

Mean for weeks

1-13 165 132 80 137 83

14-37 236 195 83 197 84

41-65 296 239 81 216 73

a

Interim evaluation occurred during week 40. b Animals switched to undosed feed
58 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

Pathology and Statistical Analysis of Results

Summariesofthe incidences of neoplasms and nonneoplastic lesionsare shown inTablesE1 andE3 formale

rats and Tables F1 and F3 for female rats. For

statisticalanalyses,the incidencesin the 9-monthstopexposuregroups and the15-monthexposuregroups at

the end of 15months are comparedwith the 15-month

interimevaluation controls ofthe2-year core studyfor

male rats(TableE2a) and female rats(Table F2a). The

incidences in the 15-month exposure groups are

comparedwith the 9-monthstop-exposuregroups after

6 months of recovery for male rats (Table E2b) and

fem ale rats (Table F2b) .

Progression or Regression

of Chemical-Induced Lesions

Liver: Rats in the stop-exposure study exposed to

20,000ppm1-amino-2,4-dibromoanthraquinone inthe

feed for 9 or 15 months had chemical-related 

effects similar to those observed in rats exposed to

concentrations up to 10,000 ppm in the 2-year core

study. The absolute and relative liver weights of

exposedmalesand femalesweresignificantly greater

than those of the controls at both the 9-month interim

and 15-month evaluations of the 15-month exposure

groups (Tables H2 and H3). With respect to both

neoplasms and nonneoplastic lesionsin the liver,there

was no evidence of regression in the incidence or

severityof chemical-relatedpigmentation,focalhepatocellular alteration, or hepatocellular adenoma or

carcinoma when administration of 1-amino--

2,4-dibromoanthraquinone was discontinued after

9 months. The incidences and severityofliver lesions

after 9 months of exposure were similar with and

without a 6-monthrecoveryperiod.The incidences of

liver lesions in the 15-month exposure groups were

greater than in the 9-monthstop-exposure groups but

the severities were comparable (Tables 16, E2a, and

F2a).
Results 59

TABLE 16

Incidences of Neoplasms and Nonneoplastic Lesions of the Liver in Rats

in the 9-Month, 9-Month with 6-Month Recovery, and 15-Month Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone

9-Month Evaluationa 15-Month Evaluationb

0 ppm 20,000 ppm 0 ppm 20,000 ppm 20,000 ppm

(9-Month (9-Month (15-Month 

Exposure) Exposure Plus Exposure)

6-Month Recovery)

Male

Number Examined 10 10 10 10 20

Basophilic Focus

c

0 6** (1.5)d1 (1.0) 4 (1.0) 13** (1.5)

Clear Cell Focus 0 4* (1.0) 0 6** (1.6) 13** (1.2)

Eosinophilic Focus 0 0 1 (1.0) 0 2 (1.5)

Bile Duct Hyperplasia 1 (1.0) 7** (1.3) 10 (1.1) 7 (1.0) 19 (1.3)

Ch ronic an d Chronic Active

Periportal Inflammation 9 (1.0) 10 (1.0) 10 (1.1) 7 (1.7) 18 (1.9)

Pigmentation 0 10**(1.0) 0 8** (1.0) 18** (1.0)

Hepatocellular Adenoma 0 2 0 7** 8*

Hepatocellular Carcinoma 0 2 0 7** 19**

Hepatocellular Adenoma or 

Carcinoma 0 2 0 9** 20**

Female

Number Examined 10 10 10 10 18

Basophilic Focus 1 (1.0) 3 (1.3) 8 (1.0) 6 (1.2) 13 (1.5)

Clear Cell Focus 0 1 (1.0) 0 5* (1.0) 13** (1.2)

Eosinophilic Focus 0 0 2 (1.5) 2 (1.0) 3 (1.7)

Bile Duct Hyperplasia 1 (1.0) 9** (1.1) 7 (1.1) 10 (1.5) 18* (1.6)

Ch ronic an d Chronic Active

Periportal Inflammation 6 (1.0) 10* (1.7) 10 (1.0) 9 (1.6) 18 (1.7)

Pigmentation 0 10**(1.2) 1 (1.0) 10** (1.0) 17** (1.2)

Hepatocellular Adenoma 0 2 0 6** 10**

Hepatocellular Carcinoma 0 1 0 6** 15**

Hepatocellular Adenoma or

Carcinoma 0 2 0 8** 16**

* Significantly different (P#0.05) from the control group by the Fisher exact test

** P#0.01

a

Controls from the 9-month interim evaluation of the 2-year core study were used for statistical comparison.

b

Controls from the 15-month interim evaluation of the 2-year core study were used for statistical comparison.

c Number of animals with lesion

d Average severity grade of lesions in affected rats: 1 = minimal; 2 = mild; 3 = moderate; 4 = marked

Large Intestine: The stop-exposure regimen had no

effect on the development of adenomatous polyps

(adenomas) of the large intestine. An adenomatous

polypwasobserved in thecolonofone exposedmale at

9 months. After 15 months, the incidence of

adenomatous polyps of the rectum was significantly

increased in the 9-month stop-exposure

females (Tables 17 and F2a). At the 15-month

evaluation, adenomatous polypswereobserved in the

rectums of three males in the 9-monthstop-exposure

group, seven males in the 15-monthexposure group,

five femalesin the 9-monthstop-exposure group, and

three females in the 15-month exposure group. No

carcinomas of the colon or rectum were observed.
60 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

Kidney: Atthe9-monthand 15-monthevaluations,the

relative kidney weights of males and females in the

15-monthexposuregroupsweresignificantly greater

than those ofthe controls (Tables H2 and H3). In the

exposedmalesand females at9months,kidneychanges

included pigmentation and minimal enlargement of

some renal tubule cell nuclei (karyomegaly). In

exposed males, there was also hyaline droplet

accumulation and a slight increase in the severity of

nephropathy. At 15 months, renal tubule 

epithelial pigmentation, karyomegaly, and increased

severityof nephropathyand transitionalcellhyperplasia

ofthe renalpelvis wereobserved in exposed groups of

malesand females. The severities ofthese lesionswere

similar or slightly less severe in the stop-exposure

groups than in those exposed continuously for

15 months. Atthe 15-month evaluation, renal tubule

adenomas wereobserved inthe9-monthstop-exposure

and 15-month exposuregroups ofmales and females

(Tables 17, E1, and F1). In the 9-month 

TABLE 17

Incidences of Neoplasms and Nonneoplastic Lesions of the Large Intestine and Kidney in Rats 

in the 9-Month, 9-Month with 6-Month Recovery, and 15-Month Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone

9-Month Evaluationa15-Month Evaluationb

0 ppm 20,000 ppm 0 ppm 20,000 ppm 20,000 ppm

(9-Month (9-Month (15-Month 

Exposure) Exposure Plus Exposure)

6-Month Recovery)

Male

Large Intestine, Colonc10 10 10 10 20

Aden omatous Polyp (Aden oma)d0 1 0 0 0

Large Intestine, Rectum 10 10 9 10 20

Aden omatous Polyp (Aden oma) 0 0 0 3 7*

Kidney 10 10 10 10 20

Nephropathy 10 (1.0)e10 (1.8) 10 (2.0) 10 (2.0) 20 (2.5)

Transition al Cell Hyperplasia 0 0 0 1 (1.0) 11** (1.3)

Hyaline Droplet Accumulation 0 10** (2.1) 0 0 0

Pigmentation 0 10** (2.0) 0 9** (1.2) 20** (2.0)

Renal Tubule Hyperplasia 0 0 0 1 (1.0) 1 (1.0)

Renal Tubule Adenoma 0 0 0 3 2

Female

Large Intestine, Rectum 10 10 10 10 17

Aden omatous Polyp (Aden oma) 0 0 0 5* 3

Kidney 10 10 10 10 18

Nephropathy 4 (1.0) 7 (1.0) 10 (1.7) 10 (2.2) 18 (2.1)

Transition al Cell Hyperplasia 0 0 3 (1.0) 1 (1.0) 5 (1.6)

Pigmentation 0 10** (2.0) 0 10** (1.9) 18** (2.0)

Renal Tubule Hyperplasia 0 0 0 2 (2.0) 2 (1.5)

Renal Tubule Adenoma 0 0 0 3 2

* Significantly different (P#0.05) from the control group by the Fisher exact test

** P#0.01

a

Controls from the 9-month interim evaluation of the 2-year core study were used for statistical comparison.

b

Controls from the 15-month interim evaluation of the 2-year core study were used for statistical comparison.

c Number of animals with organ examined microscopically

d Number of animals with lesion

e Average severity grade of lesions in affected rats: 1 = minimal; 2 = mild; 3 = moderate; 4 = marked
Results 61

stop-exposure and 15-monthexposure groups, renal

tubule hyperplasia and adenomas were observed in

males and females. At 15 months, renal tubule

adenomas were observed in three males and three

femalesinthe9-monthstop-exposuregroups and intwo

males and two females in the 15-month exposure

groups. No renal tubule carcinomas were found.

Urinary Bladder: A transitional cell papilloma was

present in the urinary bladder of one exposed male at

9 months (Tables 18 and E1). When exposure was

discontinued at 9 months, no chemical-related

nonneoplastic lesions or neoplasms were present at

15months. Withcontinuoustreatment,transitionalcell

hyperplasia and neoplasms of the urinary bladder

developed by15months. In females,aminimaltomild

transitional cell hyperplasia wasobserved at 9months

and did not completely regress with the cessation of

exposure; a squamous cell papilloma was observed in

one female at 15 months (Tables 18 and

F1). In contrast, continuous exposure for 15 months

resulted in an increased incidence and severity of

transitional cell hyperplasia, as well as squamous

metaplasia of the transitional epithelium and a fatty

metaplasia (fat proliferation) of the stroma of the

bladder wall.

Forestomach: Chemical-related lesions of the forestomach were not present in males or females at the

9-monthinterim evaluation. Hyperplasia,hyperkeratosis, inflammation, and ulceration were present in

approximately 20% of exposed males, but not in the

controls at 15 months (Table 18). A squamous cell

papillomawaspresentin onemale fromthe 15-month

exposuregroup (TableE1). Infemale rats,forestomach

lesionswerenotpresentinthe controlor 9-monthstopexposuregroups. Hyperplasia, hyperkeratosis,and/or

ulceration were observed in a few females in the

15-monthexposuregroup; no neoplasms werepresent

in the forestomach.
62 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE 18

Incidences of Neoplasms and Nonneoplastic Lesions of the Urinary Bladder and Forestomach in Rats

in the 9-Month, 9-Month with 6-Month Recovery, and 15-Month Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone

9-Month Evaluationa15-Month Evaluationb

0 ppm 20,000 ppm 0 ppm 20,000 ppm 20,000 ppm

(9-Month (9-Month (15-Month 

Exposure) Exposure Plus Exposure)

6-Month Recovery)

Male

Urinary Bladder

c

10 9 10 10 19

Fat Proliferationd0 0 0 0 1 (3.0)e

Transitional Epithelial

Hyperplasia 0 0 0 0 9** (1.9)

Squamous Cell Carcinoma 0 0 0 0 1

Transitional Epithelial Papilloma 0 1 0 0 3

Transitional Epithelial Carcinoma 0 0 0 0 1

Forestomach 10 10 10 10 20

Hyperkeratosis 0 0 0 2 (2.5) 1 (1.0)

Hyperplasia 0 0 0 2 (2.0) 3 (1.3)

Inflammation 0 0 0 1 (3.0) 1 (3.0)

Ulceration 0 0 0 2 (2.5) 0

Female

Urinary Bladder 10 10 10 10 18

Fat Proliferation 0 0 0 0 2 (3.0)

Transitional Epithelial

Hyperplasia 0 4* (1.5) 0 4* (1.8) 17** (2.6)

Transitional Epithelial

Squamou s Metaplasia 0 0 0 0 3 (2.7)

Squamous Cell Papilloma 0 0 0 0 1

Squamous Cell Carcinoma 0 0 0 1 4

Transitional Epithelial Papilloma 0 0 0 0 1

Transitional Epithelial Carcinoma 0 0 0 0 1

Forestomach 10 10 10 10 18

Hyperkeratosis 0 0 0 0 1 (1.0)

Hyperplasia 0 0 0 0 6* (1.0)

Ulceration 0 0 0 0 1 (3.0)

* Significantly different (P#0.05) from the control group by the Fisher exact test

** P#0.01

a

Controls from the 9-month interim evaluation of the 2-year core study were used for statistical comparison.

b

Controls from the 15-month interim evaluation of the 2-year core study were used for statistical comparison.

c Number of animals with organ examined microscopically

d Number of animals with lesion

e Average severity grade of lesions in affected rats: 1 = minimal; 2 = mild; 3 = moderate; 4 = marked
Results 63

MICE

13-WEEK STUDY

One 25,000 ppmmale (week 11) and one 5,000 ppm

male (week 13) died during the study (Table 19).

Neitherdeathwaschemicalrelated. One 10,000 ppm

femalewasaccidentlykilled. Finalmeanbodyweights

of exposed groups of male and female mice were

similar to those of the controls. Mean body weight

gains of exposed groups of males and females were

generally greater than those of the controls. Feed

consumption by exposed mice was similar to

that by the controls. Dietary levels of 2,500, 5,000,

10,000,25,000,and50,000 ppmdeliveredaveragedaily

doses of approximately500,1,080, 1,850, 6,200,and

10,600mg1-amino-2,4-dibromoanthraquinone/kgbody

weighttomalesand approximately660,1,150,2,600,

5,900, and 11,700mg/kgto females. Reddened furwas

observed in 10,000,25,000, and 50,000 ppm mice as

early as day 4 in males and day 5 in females and was

observed throughout the study. No other clinical

observations were attributed to 1-amino2,4-dibromoanthraquinone.

TABLE 19

Survival, Mean Body Weights, and Feed Consumption of Mice in the 13-Week Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

Final Weight

Mean Body Weightb

 (g) Relative Feed

Dose SurvivalaInitial Final Change to Controls Consumption

c

(ppm) (%) Week 1 Week 13

Male

0 10/10 23.7 ± 0.4 30.5 ± 0.6 6.9 ± 0.7 5.8 6.0

2,500 10/10 23.5 ± 0.3 30.6 ± 0.6 7.1 ± 0.6 100 5.3 5.6

5,000 9/10

d

 23.5 ± 0.4 30.7 ± 0.7 7.3 ± 0.5 101 6.1 5.6

10,000 10/10 23.7 ± 0.5 32.1 ± 0.4 8.4 ± 0.4 105 5.5 4.8

25,000 9/10 e 23.4 ± 0.3 30.9 ± 0.6 7.5 ± 0.5 101 6.3 7.1

50,000 10/10 23.4 ± 0.3 31.5 ± 0.4 8.1 ± 0.4 103 6.1 5.5

Female

0 10/10 18.2 ± 0.3 24.0 ± 0.2 5.8 ± 0.2 4.8 6.2

2,500 10/10 18.1 ± 0.3 24.7 ± 0.6 6.6 ± 0.4 103 5.0 6.4

5,000 10/10 18.4 ± 0.3 25.0 ± 0.6 6.5 ± 0.4 104 4.3 5.7

10,000 9/10

f

 18.3 ± 0.3 25.0 ± 0.4 6.5 ± 0.3 104 4.5 6.7

25,000 10/10 18.2 ± 0.2 23.6 ± 0.3 5.4 ± 0.3 98 3.8 6.1

50,000 10/10 18.4 ± 0.3 24.7 ± 0.4 6.3 ± 0.2 103 4.0 6.1

a Number of animals surviving at 13 weeks/number initially in group

b Weigh ts an d weigh t changes are given as mean ± standard error. Subsequent calcu lations are based on animals surviving to the end of the study.

c

Feed consumption is expressed as grams of feed consumed per animal per day.

d Week of death: 13

e Week of death: 11

f Week of death: 2 (accidental)
64 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

Absolute and relative liver weights of 5,000, 10,000,

25,000, and 50,000 ppm male and female mice were

significantly greater than those of the controls

(Table H4). Absolute and relative kidneyweights of

25,000and 50,000 ppmmalesweresignificantlylower

than those of the controls. Observations at necropsy

included red staining ofthegastrointestinaltract and its

contents in all exposed male mice exceptthose in the

2,500 ppm group and red staining in the kidney and

urine. These findingswereobserved lessfrequentlyin

females than in males.

Chemical-related lesions were present in the liver

(Table20). Therewereincreasedincidences of centrilobular hypertrophy in the 10,000, 25,000, and

50,000ppmmaleswithadose-related increasedseverity. Minimal gold to brown pigment granules were

presentin the cytoplasm of hepatocytesof all exposed

groups ofmales. Pigment was generally located in the

centrilobular portion of the hepatic lobule. Pigment

similar to that in male mice was present in just a few

hepatocytes in the liver of one female in the 25,000-

 ppm group and one 50,000 ppm female.

TABLE 20

Incidences of Nonneoplastic Lesions of the Liver in Mice in the 13-Week Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

0 ppm 2,500 ppm 5,000 ppm 10,000 ppm 25,000 ppm 50,000 ppm

Male

Number Examined 10 10 10 10 10 10

Centrilobular Hypertrophya0 0 0 8** (1.8)b8** (2.0) 10** (2.8)

Pigmentation 0 5* (1.0) 8** (1.0) 8** (1.0) 8** (1.0) 10** (1.1)

Female

Number Examined 10 10 10 10 10 10

Pigmentation 0 0 0 0 1 (1.0) 1 (1.0)

* Significantly different (P#0.05) from the control group by the Fisher exact test

** P#0.01

a Number of animals with lesion

b Average severity grade of lesions in affected mice: 1 = minimal; 2 = mild; 3 = moderate; 4 = marked

Dose Selection Rationale: As no 1-amino2,4-dibromoanthraquinone-related adverse effectswere

observed in feed consumption,meanbodyweights,or

survival, exposureconcentrations chosenforthe2-year

study were based mainly on the frequency and especially the severity of centrilobularhypertrophy of the

liver in male mice. Because only lesions of mild

severitywereobserved in the 10,000and 25,000 ppm

groups, and lesionsofmoderate severitywereobserved

in the 50,000 ppm group, and these were 

notlife-jeopardizinglesions,exposureconcentrations

selected for the 2-year study of 1-amino2,4-dibromoanthraquinone inmicewere0,10,000,and

20,000ppm. Other considerationsinclude consistency

among males and females (since females could have

been givenhigherexposureconcentrations) and correspondence to the exposureconcentrationsselectedfor

the start-stop,progression/regressionexperiments(stopexposure evaluation) in rats.
Results 65

2-YEAR STUDY

Survival

Estimatesof 2-yearsurvivalprobabilitiesformale and

femalemiceare shown inTable 21 and in the KaplanMeiersurvivalcurvesinFigure 6. Survivalof exposed

male mice was significantly lower than that of the

controls;survivalof exposed female mice was similar

to that of the controls.

Body Weights, Feed and Compound

Consumption, and Clinical Findings

Mean body weights of exposed groups of male mice

were lowerthan that ofthe controls afterweek9;mean

body weights of exposed groups of females were

lowerthan that ofthe controls afterweek17 (Figure7,

Tables 22 and 23). Final mean body weights of exposed groups of males were 15% to 17% lower than

thatofthe controls;finalmeanbodyweightsof exposed

females were 14% to 19% lower than that of the controls. Feed consumption byexposedmales and females

wasgenerally similar to that bythe controls(TablesJ5

and J6). Dietarylevels of10,000 and 20,000ppmwere

estimatedto deliverdaily doses of approximately 1,700

and 3,500mg1-amino-2,4-dibromoanthraquinone/kg

body weight to males and 2,000 and 4,400 mg/kg to

females. Discolorationof the fur,urine,and feceswas

evident in all exposed groups as early as day 8.

TABLE 21

Survival of Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

0 ppm 10,000 ppm 20,000 ppm

Male

Animals initially in study 60 60 60

15-Month interim evaluationa10 9 10

Accidental deatha0 1 0

Moribund 7 23 21

Natural deaths 3 5 6

Animals surviving to study termination 40 22 23

Percent probability of survival at end of studyb81 45 47

Mean survival (days)c656 620 609

Survival analysesdP=0.001 P<0.001 P<0.001

Female

Animals initially in study 60 60 60

15-Month interim evaluationa10 10 10

Moribund 5 11 11

Natural deaths 6 5 6

Animals surviving to study termination 39e34e33

Percent probability of survival at end of study 78 69 66

Mean survival (days) 659 649 657

Survival analyses P=0.234 P=0.381 P=0.259

a

Censored from survival analyses

b Kaplan-Meier determinations based on the number of animals alive on first day of terminal sacrifice

c Mean of all deaths (uncensored, censored, and terminal sacrifice)

d

The resu lt of the life table trend test (Tarone, 197 5) is in the control column , and the results of the life table pairwise comparison s (Cox, 1 972 ) with the controls

are in the exposed column s. 

e

Includes one female that died during the last week of the study.
66 l-Amino-2,4-dibromoanthraquinone, NTP TR 383

FIGURE 6

Kaplan-Meier Survival Curves for Mice Administered l-Amino-2,4-dibromoanthraquinone in Feed 

for 2 Years 
Results 67

FIGURE 7

Growth Curves for Mice Administered l-Amino-2,4-dibromoanthraquinone in Feed for 2 Years
68 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE 22

Mean Body Weights and Survival of Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

Weeks 0 ppm 10,000 ppm 20,000 ppm

on Av. Wt. Number of Av. Wt. Wt. (% of Number of Av. Wt. Wt. (% of Number of

Study (g) Survivors (g) controls) Survivors (g) controls) Survivors

1 21.7 60 22.0 101 60 21.2 98 60

2 23.5 60 23.2 99 60 23.9 102 60

3 25.2 60 24.6 98 60 25.5 101 60

4 26.3 60 25.2 96 60 26.2 100 60

5 27.3 60 26.9 99 60 27.3 100 60

6 28.7 60 28.1 98 60 28.1 98 60

7 29.5 60 29.2 99 60 29.6 100 60

8 29.2 60 29.3 100 60 29.9 102 60

9 30.2 60 30.1 100 60 30.5 101 60

10 31.7 60 30.7 97 60 31.4 99 60

11 32.6 60 31.1 95 60 31.9 98 60

12 33.3 60 32.1 96 60 32.2 97 60

13 33.7 60 32.5 96 60 33.0 98 60

14 34.1 60 33.0 97 60 33.1 97 60

17 36.3 60 34.4 95 60 34.3 95 59

21 38.8 60 34.0 88 60 34.6 89 58

25 39.2 60 37.0 94 59 35.6 91 58

29 39.9 60 35.6 89 59 36.2 91 58

33 41.3 60 38.5 93 59 36.4 88 58

37 42.1 60 39.6 94 59 37.9 90 56

41 42.8 60 39.8 93 59 38.2 89 56

45 42.1 59 39.5 94 59 37.4 89 56

49 43.5 59 40.1 92 59 38.9 89 56

53 44.4 59 41.8 94 59 39.6 89 56

57 44.0 59 41.6 95 59 39.3 89 56

61 44.8 58 42.3 94 58 40.2 90 56

65 45.2 57 41.6 92 58 39.4 87 56

69a43.6 47 40.7 93 46 37.7 87 46

73 42.1 46 39.8 95 45 39.2 93 45

77 44.2 45 40.0 91 44 38.7 88 42

81 43.0 45 39.5 92 43 38.4 89 41

85 44.1 45 38.9 88 39 38.0 86 41

89 44.0 45 37.7 86 36 37.1 84 38

93 42.3 44 38.1 90 32 36.2 86 34

97 43.8 44 37.4 85 29 36.8 84 30

101 43.7 44 37.4 86 26 36.7 84 27

104 43.6 42 37.0 85 22 36.3 83 26

Mean for weeks

1-13 28.7 28.1 98 28.5 99

14-52 40.0 37.2 93 36.3 91

53-104 43.8 39.6 90 38.1 87

a

Interim evaluation occurred during week 66.
Results 69

TABLE 23

Mean Body Weights and Survival of Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

Weeks 0 ppm 10,000 ppm 20,000 ppm

on Av. Wt. Number of Av. Wt. Wt. (% of Number of Av. Wt. Wt. (% of Number of

Study (g) Survivors (g) controls) Survivors (g) controls) Survivors

1 17.1 60 17.4 102 60 17.1 100 60

2 18.1 60 18.4 102 60 18.0 99 60

3 19.3 60 19.8 103 60 19.6 102 60

4 19.9 60 20.7 104 60 20.4 103 60

5 20.8 60 21.3 102 60 20.8 100 60

6 21.6 60 22.1 102 60 21.1 98 60

7 21.8 60 22.5 103 60 22.4 103 60

8 21.9 60 22.9 105 60 22.8 104 60

9 22.9 60 22.9 100 60 23.3 102 60

10 23.2 60 23.6 102 60 24.0 103 60

11 23.6 60 24.0 102 60 24.3 103 60

12 24.4 59 24.4 100 60 24.7 101 60

13 24.8 59 24.8 100 60 25.0 101 60

14 25.2 59 25.5 101 60 25.4 101 60

17 26.0 59 26.6 102 60 26.0 100 60

21 27.7 59 27.3 99 60 27.3 99 60

25 29.2 59 28.6 98 60 28.1 96 60

29 30.2 59 28.8 95 60 28.9 96 60

33 32.1 59 30.7 96 59 28.8 90 60

37 32.9 59 31.9 97 59 30.5 93 60

41 34.2 59 33.0 97 59 30.1 88 60

45 35.2 59 32.8 93 59 31.2 89 60

49 36.6 59 34.5 94 59 32.5 89 60

53 36.8 59 35.0 95 59 32.5 88 60

56 38.2 59 36.3 95 58 33.1 87 60

61 39.1 59 36.2 93 58 33.8 86 60

65 38.4 59 36.1 94 57 33.5 87 60

69a37.6 49 35.4 94 47 32.4 86 50

73 37.3 48 35.7 96 46 32.7 88 49

77 39.3 48 35.8 91 46 33.1 84 49

81 39.9 48 36.4 91 45 33.8 85 47

85 39.5 48 36.7 93 44 33.6 85 45

89 41.1 46 36.1 88 44 33.7 82 44

93 41.4 45 35.3 85 44 33.3 80 41

97 40.0 44 33.6 84 41 31.9 80 39

101 38.5 43 34.1 89 40 31.5 82 37

104 39.2 39 33.6 86 35 31.9 81 35

Mean for weeks

1-13 21.5 21.9 102 21.8 101

14-52 30.9 30.0 97 28.8 93

53-104 39.0 35.5 91 32.9 84

a

Interim evaluation occurred during week 65.
70 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

Pathology and Statistical Evaluation

This section describes the statistically significant

or biologically noteworthy changes in the incidences of neoplasms and/or nonneoplastic lesions

of the liver, forestomach, lung, kidney, uterus, and

pituitary gland of mice. Summaries of the incidences of neoplasms and nonneoplastic lesions,

individual animal tumor diagnoses, statistical

analyses of primary neoplasms that occurred with

an incidence of at least 5% in at least one exposure

group, and historical incidences for the neoplasms

mentioned in this section are presented in Appendix C for male mice and Appendix D for female

mice.

Liver: At the 15-month interim evaluation, absolute and relative liver weights of exposed groups

of females were significantly greater than those of

the controls (Table H5). Hepatocellular adenomas

and carcinomas were observed in exposed groups

of males and females; none were present in the

controls (Tables 24, C1, and D1). At the end of

the 2-year study, the incidences of hepatocellular

adenoma and hepatocellular carcinoma in exposed

groups of males and females were significantly

increased (Tables 24, C3, and D3). The incidences of multiple hepatocellular adenomas or

multiple hepatocellular carcinomas in exposed

groups of males and females were also increased

(Tables 24, C1, and D1). Incidences of

hepatocellular adenomas or carcinomas (combined)

 

in exposed groups of males and females exceeded

the NTP historical ranges for feed study controls

(Tables 24, C4a, and D4a). In addition to

hepatocellular adenomas and carcinomas, a small

number of hepatoblastomas occurred in exposed

groups of males and females (Tables 24, C1, and

D1). These malignant hepatocellular neoplasms

contained areas resembling hepatocellular carcinoma; in addition, there were prominent lobules or

nodular foci separated by vascular channels and

composed of undifferentiated cells (Plates 9 and

10). The neoplastic cells were elongated with a

scant amount of darkly staining cytoplasm and

oval hyperchromatic nuclei. Cellular

pleomorphism and mitotic figures were commonly

present.

At the 15-month interim evaluation and at the end

of the 2-year study, the incidences of centrilobular

hypertrophy of hepatocytes in exposed groups of

males were significantly increased, and the incidences of hepatocellular pigmentation were significantly increased in exposed groups of males and

females (Tables 24, C5, and D5). This brown,

granular pig-ment resembled that found in the

13-week studies of 1-amino-2,4-dibromoanthraquinone; histochemical procedures were not

repeated during this 2-year study. The incidences

of clear cell focus in exposed groups of female

mice were significantly increased at the end of the

2-year study.
Results 71

TABLE 24

Incidences of Neoplasms and Nonneoplastic Lesions of the Liver in Mice in the 2-Year Feed Study

of 1-Amino-2,4-dibromoanthraquinone 

0 ppm 10,000 ppm 20,000 ppm

Male

15-Month Interim Evaluation

Number Examined 10 9 10

Centrilobular Hepatocyte Hypertrophya0 9** (2.9)b8** (2.9)

Pigmentation 0 9** (1.1) 10** (1.2)

Hepatocellular Adenoma 0 2 4*

Hepatocellular Adenoma or Carcinoma 0 3 4*

2-Year Study

Number Examined 50 51 50

Basophilic Focus 0 4 (1.0) 3 (1.0)

Centrilobular Hepatocyte Hypertrophy 0 17** (2.0) 13** (2.0)

Clear Cell Focus 4 (1.3) 4 (1.0) 2 (1.0)

Eosinophilic Focus 0 6** (1.5) 1 (1.0)

Pigmentation 1 (3.0) 50** (1.1) 47** (1.4)

Hepatocellular Adenoma (Multiple) 6 29** 31**

Hepatocellular Adenoma (Single or Multiple)

Overall ratec10/50 (20%) 38/51 (75%) 39/50 (78%)

Terminal rated9/40 (23%) 20/22 (91%) 21/23 (91%)

Adjusted ratee24.3% 94.7% 95.0%

First incidence (days) 723 451 484

Logistic regression testfP<0.001 P<0.001 P<0.001

Hepatocellular Carcinoma (Multiple) 1 3 9**

Hepatocellular Carcinoma (Single or Multiple)

Overall rate 9/50 (18%) 18/51 (35%) 21/50 (42%)

Terminal rate 7/40 (18%) 10/22 (45%) 9/23 (39%)

Adjusted rate 21.1% 58.1% 58.4%

First incidence (days) 445 505 535

Logistic regression test P=0.002 P=0.017 P=0.003

Hepatocellular Adenoma or Carcinomag

Overall rate 18/50 (36%) 43/51 (84%) 42/50 (84%)

Terminal rate 15/40 (38%) 21/22 (95%) 22/23 (96%)

Adjusted rate 41.7% 97.7% 97.7%

First incidence (days) 445 451 484

Logistic regression test P<0.001 P<0.001 P<0.001

Hepatoblastoma 0 3 5*

(continued)

* Significantly different (P#0.05) from the control group by the Fisher exact test (15-month interim evaluation) or the logistic regression test

(2-year study)

** P#0.01

(T)Terminal sacrifice

a Number of animals with lesion

b Average severity grade of lesions in affected mice: 1 = minimal; 2 = mild; 3 = moderate; 4 = marked

c Number of animals with neoplasm per number of animals with liver examined microscopically
72 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE 24

Incidences of Neoplasms and Nonneoplastic Lesions of the Liver in Mice in the 2-Year Feed Study

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

Female

15-Month Interim Evaluation

Number Examined 10 10 10

Pigmentation 0 10** (1.0) 9** (1.0)

Hepatocellular Adenoma (Multiple) 0 1 4*

Hepatocellular Adenoma (Single or Multiple) 0 2 7**

Hepatocellular Adenoma or Carcinoma 0 2 8**

2-Year Study

Number Examined 50 50 50

Basophilic Focus 0 4 (1.3) 5* (1.2)

Clear Cell Focus 0 10** (1.2) 9** (1.6)

Eosinophilic Focus 0 4* (1.5) 2 (2.5)

Pigmentation 0 44** (1.1) 49** (1.6)

Hepatocellular Adenoma (Multiple) 0 40** 45**

Hepatocellular Adenoma (Single or Multiple)

Overall rate 6/50 (12%) 45/50 (90%) 49/50 (98%)

Terminal rate 6/39 (15%) 32/34 (94%) 33/33 (100%)

Adjusted rate 15.4% 95.7% 100.0%

First incidence (days) 729 (T) 442 501

Logistic regression test P<0.001 P<0.001 P<0.001

Hepatocellular Carcinoma (Multiple) 0 13** 13**

Hepatocellular Carcinoma (Single or Multiple)

Overall rate 0/50 (0%) 23/50 (46%) 27/50 (54%)

Terminal rate 0/39 (0%) 17/34 (50%) 16/33 (48%)

Adjusted rate 0.0% 57.2% 60.8%

First incidence (days) )

h

503 538

Logistic regression test P<0.001 P<0.001 P<0.001

Hepatocellular Adenoma or Carcinomai

Overall rate 6/50 (12%) 46/50 (92%) 50/50 (100%)

Terminal rate 6/39 (15%) 33/34 (97%) 33/33 (100%)

Adjusted rate 15.4% 97.9% 100.0%

First incidence (days) 729 (T) 442 501

Logistic regression test P<0.001 P<0.001 P<0.001

Hepatoblastoma 0 0 2

d Observed incidence in animals surviving until the end of the study

e Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality.

f

In the control column are the P values associated with the trend test. In the exposed group columns are the P values corresponding to

pairwise comparisons between the controls and that exposure group. The logistic regression test regards these lesions as nonfatal.

g Historical incidence for 2-year NTP feed studies with untreated control groups (mean ± standard deviation): 531/1,466 (36.2% ± 14.1%);

range, 10%-68% h Not applicable; no neoplasms in animal group

i Historical incidence: 247/1,462 (16.9% ± 10.7%); range, 3%-42%
Results 73

Forestomach: Squamouscellpapillomaswereobserved

in 20,000 ppm males and in 10,000 and 20,000 ppm

females atthe 15-monthinterimevaluation(Tables 25,

C1, and D1). At the end of the 2-year study, the incidences ofsquamouscellpapilloma and squamous cell

carcinoma inexposedgroupsofmalesandfemaleswere

significantly increased (Tables 25, C3, and D3). The

incidences of multiple squamous cell papilloma in

20,000ppmmales and femaleswere also significantly

increased in the 2-year study(Tables 25, C1, and D1).

Incidences of squamous cell papilloma or carcinoma

(combined)inexposedgroupsofmalesandfemaleswere

significantly greater than those in the controls and

exceeded the NTP historical ranges for feed study

controls(Tables 25, C4b, and D4b). Compared to the

exophyticmasses with well-differentiated squamous

epitheliumtypicalofthe squamouscellpapillomas,the

squamous cell carcinomas were locally invasive

neoplasms that sometimes resulted in perforation of

the forestomach(Plate11). Frequently, a squamouscell

carcinoma appeared to arise atthe base of a squamous

cell papilloma. Metastatic neoplasms arising from

squamous cell carcinomas of the forestomach were

observedin the coagulatingglands, colon, duodenum,

epididymis,gallbladder,glandular stomach,jejunum,

kidney, liver, lung, ovary, pancreas, prostate gland,

spleen,testis, and thymus of exposed mice (TablesC1

and D1).

Nonneoplastic lesions of the forestomach included

acanthosis,hyperkeratosis,and basal cell hyperplasia

(Tables 25, C5, and D5). At the 15-month interim

evaluation,therewereexposure-relatedincreasesinthe

incidences and severities of acanthosis and

hyperkeratosisinexposedgroupsofmalesandfemales.

At the end of the 2-year study, the incidences and

severitiesoftheselesionsinexposedgroupsofmalesand

femalesweregenerallygreaterthanthoseinthe controls.
74 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE 25

Incidences of Neoplasms and Nonneoplastic Lesions of the Forestomach in Mice in the 2-Year Feed Study

of 1-Amino-2,4-dibromoanthraquinone 

0 ppm 10,000 ppm 20,000 ppm

Male

15-Month Interim Evaluation

Number Examined 9

9 10

Acanthosis (Hyperplasia)a0 2 (1.0)b3 (2.0)

Basal Cell Hyperplasia 0 1 (1.0) 1 (1.0)

Hyperkeratosis 1 (2.0) 2 (1.0) 3 (2.0)

Inflammation 0 0 3 (1.2)

Squamous Cell Papilloma 0 0 5*

2-Year Study

Number Examined 50 50 50

Acanthosis 1 (1.0) 9** (1.1) 4 (2.0)

Basal Cell Hyperplasia 0 0 2 (1.5)

Hyperkeratosis 1 (1.0) 7* (1.0) 6 (1.8)

Inflammation 2 (1.5) 6 (1.2) 13** (1.5)

Squamous Cell Papilloma (Multiple) 0 2 5*

Squamous Cell Papilloma (Single or Multiple)

Overall ratec0/50 (0%) 13/51 (25%) 16/50 (32%)

Terminal rated0/40 (0%) 10/22 (45%) 11/23 (48%)

Adjusted ratee0.0% 51.0% 55.6%

First incidence (days) )

g

613 606

Logistic regression testfP<0.001 P<0.001 P<0.001

Squamous Cell Carcinoma

Overall rate 0/50 (0%) 12/51 (24%) 13/50 (26%)

Terminal rate 0/40 (0%) 4/22 (18%) 4/23 (17%)

Adjusted rate 0.0% 36.5% 37.7%

First incidence (days) ) 505 523

Logistic regression test P<0.001 P<0.001 P<0.001

Squamous Cell Papilloma or Carcinomah

Overall rate 0/50 (0%) 19/51 (37%) 27/50 (54%)

Terminal rate 0/40 (0%) 11/22 (50%) 14/23 (61%)

Adjusted rate 0.0% 61.2% 73.9%

First incidence (days) ) 505 523

Logistic regression test P<0.001 P<0.001 P<0.001

(continued)

* Significantly different (P#0.05) from the control group by the Fisher exact test (15-month interim evaluation) or the logistic regression test

(2-year study)

** P#0.01

(T)Terminal sacrifice
Results 75

TABLE 25

Incidences of Neoplasms and Nonneoplastic Lesions of the Forestomach in Mice in the 2-Year Feed Study

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

Female

15-Month Interim Evaluation

Number Examined 10 10 10

Acanthosis (Hyperplasia) 0 1 (1.0) 8** (2.0)

Basal Cell Hyperplasia 0 0 2 (1.5)

Hyperkeratosis 0 2 (1.0) 7** (2.0)

Inflammation 0 1 (1.0) 5* (1.6)

Squamous Cell Papilloma 0 4* 2

2-Year Study

Number Examined 48 50 50

Acanthosis (Hyperplasia) 9 (1.7) 15 (1.7) 19* (1.6)

Basal Cell Hyperplasia 0 7* (1.4) 3 (1.7)

Hyperkeratosis 10 (1.4) 14 (1.4) 17 (1.5)

Inflammation 7 (1.4) 10 (1.4) 21** (1.7)

Squamous Cell Papilloma (Multiple) 0 4 14**

Squamous Cell Papilloma (Single or Multiple)

Overall rate 2/50 (4%) 16/50 (32%) 27/50 (54%)

Terminal rate 2/39 (5%) 12/34 (35%) 23/33 (70%)

Adjusted rate 5.1% 41.7% 72.4%

First incidence (days) 729 (T) 671 538

Logistic regression test P<0.001 P<0.001 P<0.001

Squamous Cell Carcinoma

Overall rate 0/50 (0%) 12/50 (24%) 11/50 (22%)

Terminal rate 0/39 (0%) 8/34 (24%) 5/33 (15%)

Adjusted rate 0.0% 30.9% 27.3%

First incidence (days) ) 587 501

Logistic regression test P=0.002 P<0.001 P<0.001

Squamous Cell Papilloma or Carcinomai

Overall rate 2/50 (4%) 25/50 (50%) 34/50 (68%)

Terminal rate 2/39 (5%) 18/34 (53%) 25/33 (76%)

Adjusted rate 5.1% 60.7% 80.5%

First incidence (days) 729 587 501

Logistic regression test P<0.001 P<0.001 P<0.001

a Number of animals with lesion

b Average severity grade of lesions in affected mice: 1 = minimal; 2 = mild; 3 = moderate; 4 = marked

c Number of animals with neoplasm per number of animals necropsied

d Observed incidence in animals surviving until the end of the study

e Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality.

f

In the control column are the P values associated with the trend test. In the exposed group columns are the P values corresponding to

pairwise comparisons between the controls and that exposure group. The logistic regression test regards these lesions as nonfatal.

g Not applicable; no neoplasms in animal group

h Historical incidence for 2-year NTP feed studies with untreated control groups (mean ± standard deviation): 22/1,474 (1.5% ± 2.0%); range,

0%-6% i Historical incidence: 33/1,470 (2.2% ± 3.1%); range, 0%-14%
76 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

Lung: Alveolar/bronchiolar adenomas were

observed in exposed groups of males and females

at the 15-month interim evaluation (Tables 26, C1,

and D1). During the 2-year study, the incidences

of alveolar/bronchiolar adenomas in exposed

groups of males and females were significantly

increased (Tables 26, C3, and D3). In male mice,

the incidences of multiple alveolar/bronchiolar

adenomas in exposed groups were significantly

greater than that in the controls (Tables 26, C1,

and D1). The incidences of alveolar/ 

bronchiolar adenoma in exposed groups of males

and females exceeded the NTP historical ranges

for feed study controls (Tables 26, C4c, and D4c).

The alveolar/bronchiolar adenomas were generally

well-circumscribed, expansile masses that slightly

compressed the surrounding normal pulmonary

alveolar tissue (Plate 12). Well-differentiated

cuboidal to columnar epithelial cells formed

papillary structures or solid foci that filled alveolar

spaces.

TABLE 26

Incidences of Neoplasms of the Lung in Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone 

0 ppm 10,000 ppm 20,000 ppm

Male

15-Month Interim Evaluation 

Number Examined 10 9 10

Alveolar/bronchiolar Adenomaa0 3 5*

2-Year Study

Number Examined 50 51 50

Alveolar/bronchiolar Hyperplasia 1 (1.0)b0 4 (1.3)

Alveolar/bronchiolar Adenoma (Multiple) 0 6** 9**

Alveolar/bronchiolar Adenoma (Single or Multiple)

Overall ratec7/50 (14%) 26/51 (51%) 24/50 (48%)

Terminal rated6/40 (15%) 12/22 (55%) 12/23 (52%)

Adjusted ratee16.8% 71.0% 66.5%

First incidence (days) 445 578 248

Logistic regression testfP<0.001 P<0.001 P<0.001

Alveolar/bronchiolar Carcinoma

Overall rate 3/50 (6%) 4/51 (8%) 1/50 (2%)

Terminal rate 2/40 (5%) 2/22 (9%) 0/23 (0%)

Adjusted rate 6.9% 15.9% 3.0%

First incidence (days) 393 673 648

Logistic regression test P=0.259N P=0.512 P=0.251N

Alveolar/bronchiolar Adenoma or Carcinomag

Overall rate 10/50 (20%) 28/51 (55%) 25/50 (50%)

Terminal rate 8/40 (20%) 13/22 (59%) 12/23 (52%)

Adjusted rate 23.3% 75.0% 67.5%

First incidence (days) 393 578 248

Logistic regression test P<0.001 P<0.001 P=0.002

(continued)
Results 77

TABLE 26

Incidences of Neoplasms of the Lung in Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

Female

15-Month Interim Evaluation

Number Examined 10 10 10

Alveolar/bronchiolar Adenoma 0 3 2

2-Year Study

Number Examined 50 50 49

Alveolar/bronchiolar Hyperplasia 0 0 1 (1.0)

Alveolar/bronchiolar Adenoma (Multiple) 0 2 1

Alveolar/bronchiolar Adenoma (Single or Multiple)

Overall rate 4/50 (8%) 17/50 (34%) 13/49 (27%)

Terminal rate 3/39 (8%) 14/34 (41%) 9/33 (27%)

Adjusted rate 9.8% 45.6% 33.5%

First incidence (days) 685 587 538

Logistic regression test P=0.017 P=0.001 P=0.015

Alveolar/bronchiolar Carcinoma 0 0 2

Alveolar/bronchiolar Adenoma or Carcinomah

Overall rate 4/50 (8%) 17/50 (34%) 15/49 (31%)

Terminal rate 3/39 (8%) 14/34 (41%) 10/33 (30%)

Adjusted rate 9.8% 45.6% 37.9%

First incidence (days) 685 587 538

Logistic regression test P=0.006 P=0.001 P=0.005

* Significantly different (P#0.05) from the control group by the Fisher exact test (15-month interim evaluation) or the logistic regression test

(2-year study)

** P#0.01

a Number of animals with lesion

b Average severity grade of lesions in affected mice: 1 = minimal; 2 = mild; 3 = moderate; 4 = marked

c Number of animals with neoplasm per number of animals with lung examined microscopically

d Observed incidence in animals surviving until the end of the study

e Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality.

f

In the control column are the P values associated with the trend test. In the exposed group columns are the P values corresponding to

pairwise comparisons between the controls and that exposure group. The logistic regression test regards these lesions as nonfatal. A negative

trend or a lower incidence in an exposure group is indicated by N.

g Historical incidence for 2-year NTP feed studies with untreated control groups (mean ± standard deviation): 265/1,469 (18.0% ± 7.6%);

range, 4%-32% h Historical incidence: 110/1,469 (7.5% ± 5.0%); range, 2%-26%
78 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

Kidney: Pigmentationwaspresentin the kidneys ofmost

mice after 2 years of exposure to 1-amino2,4-dibromoanthraquinone (males: 0 ppm, 0/50;

10,000ppm, 42/51; 20,000ppm, 43/50;females: 0/50,

43/50,43/50;TablesC5andD5). Thisbrown,granular

pigmentinthe renaltubuleepitheliumandtubulelumina

resembledthepigmentdescribedinthe liver.Therewere

no other chemical-related lesions in the kidney.

Uterus: There was a significant, but not exposurerelated, increase in the incidence of uterine polyps or

sarcomas(combined)(0/50,5/50,0/50;TableD1)inthe

10,000ppmfemales. Althoughthisincidence (10%)was

above the average for historical controls (3.5%), the

combined incidence was within the historical control

range (0%-16%;TableD4d). Thismarginalincreasewas

not considered to be chemical related. There were no

chemical-relatednonneoplasticlesionsinthe reproductive tract (Table D5).

PituitaryGland: Asignificant,but not exposure-related,

increase in the incidence of adenoma of the pituitary

gland (pars distalis) was observed in the 10,000 ppm

females(1/43,9/45, 4/43;TableD1). The incidenceof

adenoma (20%) in the 10,000 ppm females is slightly

above the average forhistorical controls(15.2%),butis

withinthe historicalrange (2%-36%;TableD4e). This

marginal increase was not considered to be chemical

related. The incidenceofhyperplasiaoftheparsdistalis

ofthe pituitaryglandwas also increased in the 10,000-

 ppm females (7/43, 22/45, 7/43; Table D5).

DISPOSITION

AND METABOLISM STUDIES

AdultmaleF344/Nratsreceived[

14C]-labeled1-amino2,4-dibromoanthraquinoneas a singleintravenousdose

of0.4mg1-amino-2,4-dibromoanthraquinone/kg body

weight or as a single oral dose of 2, 23, 118, 814, or

1,473mg/kg. After excretawerecollectedfor72hours,

the animalswerekilled, and tissues were removed for

analysis. Additionalanimalsthatreceivedintravenous

dosesof1-amino-2,4-dibromoanthraquinonewerekilled

0.25, 0.75, 2, 6, or 24 hours after chemical administration, and their tissues were analyzed. A 6-hour bile

cannulation study was also performed.

Fromday0 throughday3afterintravenousadministrationof[

14C]-1-amino-2,4-dibromoanthraquinone, about

50% of the 14C was excreted in the feces, 15% in the

urine, and 6%in expired air. Unmetabolized1-amino2,4-dibromoanthraquinoneaccountedforlessthan3%of

the excreted 14Cafter intravenousadministration. The

amountof anoraldosethatwasabsorbedwas calculated

fromthepercentofthedose thatwas excretedinexpired

airorurineafteroral administrationversusthepercentof

the dose excreted after intravenous administration.

Excretionof 14Cinexpired air yieldedthemost consistent results. For oral doses greater than or equal to

2mg/kg,the amountofthedose thatwasabsorbedfitted

the equation: absorbed dose = 6.6 log(dose), with

both doses expressed in mg/kg. While 90% of the

2mg/kg dose was absorbed,only2%ofthe 814mg/kg

dose was absorbed.

Twohoursafterintravenousadministration,lessthan3%

of the circulating

14Cwas attributed to the parent compound. The metabolites of 1-amino2,4-dibromoanthraquinoneinbloodwereprimarilyinthe

plasma fraction (blood:plasma ratioof approximately

0.5:1). The highest concentrations of 14C in tissues

15minutes after intravenous dosingwere in excretory

organs,lung, kidney,smallintestine,liver,adiposetissue,

andadrenalgland. Tissue:blood ratios(TBR)forthese

tissuesweregreaterthan or equalto 3:1. Onlythe liver

and kidney had TBRs significantly greater than 1:1 at

72 hours. The terminal half-life of 14C was approximately40hoursinthe liver andapproximately90hours

in the kidney. Adipose tissue contained primarily

unmetabolized1-amino-2,4-dibromoanthraquinone at

24 hours; liver, muscle, and skin contained mostly

metabolitesof1-amino-2,4-dibromoanthraquinone. The

elimination half -life of 1-amino -

2,4-dibromoanthraquinoneinadiposetissuewasapproximately 11 hours.

GENETIC TOXICOLOGY

1-Amino-2,4-dibromoanthraquinone (100 to

10,000:g/plate)wastestedforinductionofgenemutations in four strains of Salmonella typhimurium in a

preincubation protocol with and without

Aroclor 1254-induced male Sprague-Dawley rat or

Syrian hamster liver S9 (Table G1; Haworth et al.,

1983). 1-Amino-2,4-dibromoanthraquinone was
Results 79

positive in the absence of S9 in the frameshift

strains TA98 and TA1537; with S9, an equivocal

response was obtained in TA1537, and TA98 was

negative. In TA100, 1-amino-2,4-dibromoanthraquinone gave equivocal responses with and without S9, and all trials were negative with TA1535.

The equivocal calls were the results of positive or

weakly positive responses that were not duplicated

in a second trial. Precipitation of 1-amino2,4-dibromoanthraquinone occurred at concentrations of 100 :g/plate and above, and this may have

been a factor in the nonreproducibility of the

results.

1-Amino-2,4-dibromoanthraquinonewas tested in

two laboratories for induction of sister chromatid

exchanges and chromosomal aberrations in cultured Chinese hamster ovary cells, with and

without Aroclor 1254-induced male SpragueDawley rat liver S9. In the sister chromatid

exchange test, one 

laboratory observed a significant increase in sister

chromatid exchanges only in the absence of S9,

while the second laboratory recorded positive

responses with and without S9 (Table G2; Loveday et al., 1990). This discrepancy cannot be

explained by a difference in dose levels employed

at the two laboratories because the positive responses with S9 were observed at 3, 10, 15, and

30 :g/mL at the second laboratory, whereas

negative trials resulted from testing doses up to

100 :g/mL at the first laboratory. In the chromosomal aberrations test, one laboratory observed a

weakly positive response only in the absence of S9

(Table G3). Another laboratory obtained a positive response in the first trial without S9 but did

not duplicate the positive response in the second

trial, and the overall call without S9 was concluded to be equivocal (Loveday et al., 1990).

Neither laboratory observed an increase in chromosomal aberrations with 1-amin o -

2,4-dibromoanthraquinone in the presence of S9.
80 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

 
PLATE 1

An hepatocellular carcinoma in a female F344/N rat exposed to 10,000 ppm 1-

amino-2,4-dibromoanthraquinone in feed for 2 years. Note compression of 

normal liver (small arrows) by neoplastic hepatocytes. Carcinoma embolus (large 

arrow) is in an hepatic vein. H&E; 90x

PLATE 2

Multiple metastatic foci (*) of an hepatocellular carcinoma in the lung of a female 

F344/N rat exposed to 5,000 ppm l-amino-2,4-dibromoanthraquinone in feed for 

2 years. H&E; 15x

 

PLATE 3

Detail of a metastatic focus of the hepatocellular carcinoma shown in Plate 2 

shows the solid and acinar growth patterns of the well-differentiated neoplastic 

hepatocytes. H&E; 90x

PLATE 4

An hepatocholangiocarcinoma in a female F344/N rat exposed to 10,000 ppm lamino-2,4-dibromoanthraquinone in feed for 2 years. Note the welldifferentiated hepatocyte (solid areas) and biliary components within the neoplasm. 

H&E; 140x
 

PLATE 5

An adenoma (adenomatous polyp) in the colon of a female F344/N rat 

exposed to 10,000 ppm l-amino-2,4-dibromoanthraquinone in feed for 

2 years forms an exophytic mass (arrows) that partially occludes the 

intestinal lumen (L). H&E; 25*

PLATE 6

Detail of a carcinoma in the colon of a female F344/N rat exposed to 

10,000 ppm l-amino-2,4-dibromoanthraquinone in feed for 2 years. Note 

the disruption of the muscularis mucosa (M) layer at right with formation 

of irregular-shaped neoplastic glands (arrows), inflammation, and fibrosis in 

the submucosa. H&E; 160x

 

PLATE 7

A transitional cell carcinoma in the urinary bladder of a female F344/N rat 

exposed to 10,000 ppm l-amino-2,4-dibromoanthraquinone in feed for 

2 years. Note the thickened neoplastic mucosal surface and a papillary 

projection of the neoplasm extending into the bladder lumen (L). The 

mucosal surface consists of a thickened layer of neoplastic transitional cells 

(arrows); a large nodule of transitional epithelium invades the wall of the 

urinary bladder. H&E; 160x

PLATE 8

Detail of the transitional cell carcinoma shown in Plate 7 shows cellular 

atypia and an increased number of mitotic cells. H&E; 320x
 

PLATE 9

An hepatoblastoma in the liver of a female B6C3F1 mouse exposed to 

20,000 ppm l-amino-2,4-dibromoanthraquinone in feed for 2 years consists 

of prominent neoplasm lobules separated by vascular channels. H&E; 80x

PLATE 10

Detail of the hepatoblastoma shown in Plate 9 shows closely packed 

undifferentiated neoplastic cells with scant cytoplasm, oval nuclei, and 

numerous mitotic cells (arrows). H&E; 320x

 

PLATE 11

A squamous cell carcinoma of the forestomach in a male B6C3F1 mouse 

exposed to 20,000 ppm l-amino-2,4-dibromoanthraquinone in feed for 

2 years. Note the thickened keratin (K) layer on the mucosal surface and 

invasion of the wall by nodules (arrows) of neoplastic squamous cells that 

have extended through the peritoneal surface of the stomach. H&E; 40x

PLATE 12

An alveolar/bronchiolar adenoma of the lung in a male B6C3F1 mouse 

exposed to 20,000 ppm l-amino-2,4-dibromoanthraquinone in feed for 

2 years forms a non-encapsulated subpleural nodule. H&E; 60*

81

DISCUSSION AND CONCLUSIONS

Anthraquinonesrepresentthe largestgroupof naturallyoccurringquinones. Bothnatural andsynthetic

anthraquinoneshave been and continue to bewidely

usedas colorantsinfood,drugs,cosmetics,hairdyes,

andtextiles. Dantron(1,8-dihydroxyanthraquinone)

andemodin(1,3,8-trihydroxy-6-methylanthraquinone)

are also usedtherapeuticallyas cathartics and purgatives. Chrysophanol (1,8-dihydroxy3-methylanthraquinone)occursin cascara sagrada,

senna, and various species of Rumex and Rheum

(rhubarb).

Anthraquinoneand five substitutedanthraquinones

wereselected fortoxicologiccharacterizationfrom a

large group of amino-, alkyl-, nitro-, or halogencontaining anthraquinones. The basisforselection

centeredmainlyon four criteria: 1)lackof available

dataoncarcinogenicity, 2)magnitudeof production

andusepatterns,3) awareness of potential andactual

humanexposure,4) and representationof as broad a

spectrum of structural diversity within this class as

possible. 1-Amino-2,4-dibromoanthraquinonewas

selected froma groupof 36environmentallysignificant aryl bromides. Since all other substituted

anthraquinonechemicalsalreadyevaluatedforlongtermeffectsinducedcarcinogenicresponsesinlaboratoryanimals(NCI,1978a,1978b,1978c;IARC,1982,

1987; NTP, 1986a ), 1-amino-2,4-

dibromoanthraquinonewas predicted tobe carcinogenic in laboratory animals as well (Fung et al.,

1993).

1-Amino-2,4-dibromoanthraquinonewasstudiedfor

long-termtoxicityand carcinogenesis usinga "startstop" experimentaldesign. Oneofthe first chemicals

to be studied by the National Toxicology Program

(NTP) with a start-stop protocol, 1-amino2,4-dibromoanthraquinonewaspredictedtobe carcinogenic,sothe experimentaldesign wasselectedinan

attempt to gain some insight into the progression

and/orregressionof chemical-inducedlesions aswell

asto perhapsgainknowledge aboutpotentialmechanisms of action.

Because1-amino-2,4-dibromoanthraquinone caused

significantcarcinogenic responsesinmaleandfemale

rats and mice and in several organs and tissues, the

discussionoflesionsthatfollows hasbeengroupedby

organ.

Liver: 1-Amino-2,4-dibromoanthraquinone differs

fromother chemicalsstudied bythe NTP because it

induced greater than 90% incidences of multiple

benignandmalignanthepatocellularneoplasmsinrats

withfrequentmetastases(almost50%)ofthemalignant liver neoplasms. Although other chemicals

including 3,3N-dimethylbenzidinedihydrochloride

(NTP, 1991) and furan (NTP, 1993a) have caused

significant increases in benign and malignant liver

neoplasmsthat approacha100%incidence,metastases

occurred in only one or two instances in a group of

50 rats. Only in the 18-month study of methyl

carbamate(NTP,1988)hasa similarlyhighincidence

ofmalignant,metastatic liver neoplasmsoccurredin

male rats.

The liverlesionspresent after13weeksinthe25,000

and50,000ppmgroups ofratsincludedproliferative

bile duct lesions (cholangiofibrosis) and foci of

hepatocellular alteration. Based on morphological

features of this proliferative bile duct lesion and

resultsoftransplantationandstop-exposure studies,

cholangiofibrosis has been considered a

"premalignant" lesion which is autonomous and

progressive and not qualitatively different from

cholangiocarcinoma (Maronpot etal., 1991). While

thereissome disagreementonthebiologicalbehavior

of cholangiofibrosis,thishas generallybeenconsidered to be a preneoplastic lesion (Bannasch and

Massner, 1976; Ohshima et al., 1984).

Cholangiofibrosis has been described in toxicity

studies of methapyrilene (Ohshima et al., 1984),

aflatoxin (Wilson et al., 1985), and furan (NTP,

1993a) in rats. 

Exposure concentrations of 1-amino2,4-dibromoanthraquinone administered in the

2-year study were below those which produced

cholangiofibrosis at 13 weeks; however,manyof

the benign and malignant liver neoplasms in the

2-year study in rats were composed of a mixed

growth pattern of both hepatocytes and bile duct

formation. The incidences of foci
82 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

of hepatocellular alteration were increased with

1-amino-2,4-dibromoanthraquinone exposure at

13 weeks and all scheduled intervals examined

during the 2-year study. A detailed analysis of foci

of hepatocellular alteration in the liver of rats from

thisstudyhas been reported byHarada et al.(1989).

In addition to overall exposure-related increased

incidences of eosinophilic and clear cell foci of

alteration,there were increasesin size,number, and

volume fractionof atypicaleosinophilic, basophilic,

and clear cellfociinratsthat correlatedwithconcentration and d ur ation o f 1-amino2,4-dibromoanthraquinone exposure.

Although foci of hepatocellular alteration in rats are

believed to be precursors of liver neoplasms, their

biological nature and potential for progression to

neoplasms is uncertain (Popp and Goldsworthy,

1989;Squire,1989). Some ofthisuncertaintyresults

from a considerable variation in phenotypes of

hepatocellularfoci andthedifferentbiomarkersused

in their classification. Inmany studies, the conversion rateoffocito neoplasmshasbeenconsidered to

be extremelylow, and,in some instances, increases

in the incidences of basophilic foci have not been

associatedwithliverneoplasms(MacDonald et al.,

1988;Harada et al., 1989; Squire, 1989). Clear and

acidophilic cell foci have been suggested to be

important in the development of some chemicalinduced liver neoplasms (Bannasch et al., 1989;

Bannasch and Zerban, 1992). The atypical

eosinophilic foci that occurred in rats administered

1-amino-2,4-dibromoanthraquinone were rarely

observed in controls orin groups ofratsreceiving

other hepatocarcinogens (Harada et al., 1989).

Adenomasin the liversofratstreatedwith1-amino2,4-dibromoanthraquinone often contained cells

morphologically identical to those in the atypical

eosinophilic foci,suggestingthatsome ofthese foci

mayhave beenprecursorsforthehepaticneoplasms.

Liver effectsinmalemice administered 1-amino2,4-dibromoanthraquinonefor13weeks consistedof

pigmentationandhypertrophythatpersistedthroughoutthe 2-yearstudy. Although hypertrophydidnot

occurin female mice during this early period, pigmentation in the liver was present by 15 months.

Foci of hepatocellular alterationwerenot presentin

the 13-weekstudy, and after 2 years, the incidences

were only slightlyincreased inmice. After 2 years,

therewereincreasedincidencesofliverneoplasmsin

all groups of mice exposed to 1-amino2,4-dibromoanthraquinone in feed. This response

was more prominent in females that also had a

greater number of hepatocellular carcinomas and

more multiple liver neoplasms than male mice.

Unlike the highly metastatic liver neoplasms observed in rats, only a few neoplasms in mice had

detectable metastatic foci. The incidences of

hepatoblastomaswere alsoincreasedinthe exposed

groups of male and female mice. These distinctive

liver neoplasms rarelyoccur in control animals but

have been induced in mice administered

acetylaminofluorene (Nonoyama et al., 1988) or

N-nitrosodiethylamine (Ward et al., 1983).

Large Intestine: Adenomatous polyps(adenomas)

and carcinomas of the large intestine (distal colon

and rectum) in rats were generally observed after

15 months of exposure to 1-amino2,4-dibromoanthraquinone, although one adenomatous polyp (adenoma) was observed as early as 9

monthsin the 20,000 ppm group of male rats in the

"stop-exposure" study. Further, these lesions were

often grossly visible. Even when exposure was

stoppedafter 9months,the percentage of chemicalinducedrectalneoplasms was equaltoorgreaterthan

that observed with continuous exposure for 15

months. Inmanyrats,theseneoplasmsweremultiple

and malignant, based upon local invasion and/or

metastases. Neoplasms of the large intestine have

not been observed for other previously tested

anthraquinones. One other chemical studied bythe

NTP,bromodichloromethane(NTP,1987a),resulted

in similarlyhigh incidencesofbenign andmalignant

neoplasms of the large intestine.

Kidney: Accumulationofpigmentin the kidneywas

observed in both male and female rats by 13 weeks

and throughout the 2-year study. 1-Amino2,4-dibromoanthraquinone (ormetabolite) pigment

in the kidney of mice was not evident until after

the 15-month evaluation; there were no increased

incidencesofothernonneoplasticlesionsorneoplasms

of the kidney in mice. In the kidney of rats, several

changes in addition to pigment were present at 13

weeks. In male rats, increased accumulation of

hyaline droplets was observed in the cytoplasm of

the renaltubuleepithelium, yetno chemical-related

exacerbationofrenaltubuleepithelial regeneration

was observed at 13 weeks. There was a slight

enlargement(karyomegaly)ofsome nucleiinthe renal

tubule epithelium of male and female rats. At
Discussion and Conclusions 83

the 15-monthevaluation,thisslightnuclear enlargement was still evident, and the severity of

nephropathy(tubuleepithelialregeneration;transitional cell hyperplasia of the renal pelvis) was

increasedinexposedmaleandfemale rats. Accumulationofhyalinedropletswas not presentinexposed

malerats afterthe9-monthevaluation. Themorphological appearance of hyaline droplets and their

presenceonlyinmalesissuggestiveof accumulation

of "2:-globulin in the renal tubule epithelium, although the identity ofthe proteindroplets was not

determined. Theirabsence inthekidneytubulecells

of exposedmalerats afterthe 9-month evaluation is

consistentwiththenormallydecreasedproductionof

"2:-globulinby the liver beginning at 5 months of

age (Baetcke et al., 1991). Chemicalsthat cause a

hyaline droplet nephropathy syndrome are often

empirically associated with increasesin the incidences of benign and malignant renal tubule

neoplasms,linearfoci ofmineralizationofthe renal

medulla,andenhancednephropathyinmaleratsafter

2 years (Baetcke et al., 1991);however,other alternative mechanistic explanations exist that do not

show a dominant role for "2:-globulin. The key to

thisviewcenterson severalchemicalsthatincitethe

"hyalinedropletsyndrome," yetdonotinduce tubule

cellneoplasms ofthekidney(BarrettandHuff,1991;

Huff,1992, 1993; Melnick, 1992). Another strong

neoplastic response in the kidney of female rats

showsthatmechanisms other than those associated

with hyaline droplet nephropathy were involved in

the renaltubuleneoplasmresponse in rats administered1-amino-2,4-dibromoanthraquinone. Increased

incidencesofbenign andmalignantneoplasms ofthe

kidney occurred inmaleand female rats in the NTP

study of bromodichloromethane (NTP, 1987a).

Urinary Bladder: Chemical-related increased

incidencesof proliferative lesions (hyperplasia and

neoplasia) of the transitional cell epithelium of the

urinarybladderoccurredinmaleandfemale ratswith

a greater number of neoplasms observed in female

rats(45/146,31%)than in male rats (16/146, 11%).

In the stop-exposure groups,transitional cellhyperplasiawas present in four female rats at 9 months,

and, with the absence of continued chemical exposure, hyperplasia did not develop in male rats atthe

15-monthevaluation. Atransitional cell carcinoma

occurred in one female ratfromthe 15-monthexposure group. With continuous exposure to 1-amino2,4-dibromoanthraquinone, benign and malignant

neoplasms of the urinary bladder

developed by 15monthsin female rats and in both

male and female rats after 2 years of exposure. In

rats following chronic administration of

1,4,5,8-tetraaminoanthraquinone, a spectrum of

nonneoplastic lesions and neoplasms ofthe urinary

bladder was observed with similar morphologic

featuresincludingsquamousmetaplasia,squamous

cell carcinoma, and proliferation of fat (fatty

metaplasia)in thewallof the urinarybladder(NTP,

1986a). In that study, calculi were present in the

urinarybladder ofmostrats, yet there was a significant increase in the incidence of smooth muscle

neoplasms ofthe wall of the urinarybladder. The

hypothesis of cell proliferation and development of

urinary bladder neoplasms has been described

(Greenfield et al., 1984; Cohen et al., 1991). The

mechanismforformationofneoplasmsoftheurinary

bladder is uncertain. Increased cell proliferation

evidenced by transitional cell hyperplasia in the

urinarybladderdidnot occurbefore9monthsinrats.

Potential local irritant effects and associated neoplasmformationin the urinary bladder attributed to

calculusformation (Okumara et al., 1992)werenot

identified in thisstudy. Mostmice in that study had

calculi of the urinary bladder, yet did not have any

evidence of carcinogenic activity. No scientific

consensus existsthat endorsesthenotionthat calculi

or stones cause cancer; there may be some cocarcinogenicorpromotionactivity, yet eventhisdoes

not occur consistently (Huff, 1992, 1993).

Forestomach: In both rats and mice, there were

severalnonneoplasticproliferative andinflammatory

lesionsin the forestomach at the end of the 2-year

studies. These forestomach lesions were not observedin either species in the 13-week studies or in

rats at the 9- and 15-monthevaluations. In the stopexposure evaluation,rats exposed to 20,000 ppm

developednonneoplasticlesions ofthe forestomach

by 15 months. However, in the 9-month stopexposure group,forestomachlesionswereobserved

in a few males by the 15-month evaluation, but the

incidenceswere higher than those observedinmale

rats with the continuous 15-month exposure. In

female rats, forestomach lesions were present at

15 months with continuous exposure but were not

observed at 9 months or after 6 months of

nonexposure. Chemical-related lesions consistedof

hyperplasia,hyperkeratosisandassociatedinflammation, and focal erosionorulcerationofthe squamous

mucosa. The inflammatory and ulcerative lesions

were generallymore severe and more common in
84 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

rats, butsignificant increasesin the incidences of

benign andmalignantforestomachneoplasmswere

limitedtomice. These dataprovide further evidence

that inflammation or ulceration does not always

resultin neoplasia (Berenblum, 1944; Huff, 1992,

1993;Melnicketal.,1993a,1993b). The absenceof

forestomachneoplasmsinratsmayhavebeenrelated

to lower exposure concentrations. Many of the

malignant forestomach neoplasms of mice

metastasized or invaded adjacent organs. 

Increasesinthe incidencesofforestomachneoplasms

have not been observed in mice or rats following

long-term administration of four other structurally

related anthraquinones. However,administration of

1-amino-2-methylanthraquinone(NCI, 1978b)torats

for 62 weeksfollowed by a 6-month nonexposure

periodwas associatedwithanincreasedincidence in

hyperplasia of the forestomach. Administration of

2-methyl-1-nitroanthraquinone (NCI, 1978c)torats

for 78 weeksfollowed by a 6-month nonexposure

period was also associated with an increased incidence of proliferative lesions ofthe forestomach.

Marked increases in the incidences of forestomach

neoplasms have been reported for some chemicals

that caused a sustainedproliferative response in the

squamousmucosathatwas evidentwithinthe first 2

to 13 weeks of chemical administration (NTP,

1987b). However, a number of other chemicals

causingforestomachneoplasms in rodentshave not

been associated with an early,sustainedincrease in

the incidence of hyperplasia (NTP, 1990a, 1990b,

1990c).

Lung: The incidences of alveolar/bronchiolar

adenoma andmultiplealveolar/bronchiolar adenoma

(malesonly) ofthe lungwere significantlyincreased

in mice in the 10,000 and 20,000 ppm groups.

Althoughtherewasnoevidencefor anincrease inthe

incidence of hyperplasia or for a progression of the

lung neoplasms to malignancy, the incidence of

adenoma in all four exposuregroups exceeded the

NTP historical control ranges. Administration of a

structurally related anthraquinone, C.I. Disperse

Blue 1 (1,4,5,8-tetraaminoanthraquinone)(NTP,

1986a), resulted in a marginal increase in the

incidenceof alveolar/bronchiolar adenoma in male

micewithno associatedincrease in the incidence of

alveolar/bronchiolar hyperplasia. Other chemicals

tested by the NTP have also caused increased

incidences of lung neoplasms without increased

incidencesof alveolar/bronchiolarhyperplasia(NTP,

1994),butmore commonly an increase in the incidence of alveolar/bronchiolar hyperplasiaorinflammation is also present with increased incidences of

lung neoplasms (NTP,1986b, 1989, 1990a, 1992).

Although the incidence of alveolar/bronchiolar

carcinoma was not increased in the lungs of mice

administered 1-amino-2,4-dibromoanthraquinone,a

number of chemicals have induced both alveolar/

bronchiolar adenomasandcarcinomas(NTP,1990b,

1990c, 1993b; Huff, 1994).

CONCLUSIONS

Under the conditions of these 2-yearfeed studies,

there was clear evidence of carcinogenic activity*

of 1-amino-2,4-dibromoanthraquinone inmaleand

femaleF344/Nratsbasedonincreasedincidences of

neoplasms in the liver, large intestine, kidney, and

urinary bladder. There was clear evidence of

carcinogenic activity of 1-ami n o -

2,4-dibromoanthraquinone in male and female

B6C3F1 mice based on increased incidences of

neoplasms in the liver, forestomach, and lung.

Exposure of male and female rats to 1-amino2,4-dibromoanthraquinonefor2yearswasassociated

withbasophilic focus (males only), clear cellfocus,

eosinophilic focus, and pigmentation in the liver;

renaltubulehyperplasia, renaltubulepigmentation,

and transitional cellhyperplasiainthekidney; transitional cell hyperplasia,squamousmetaplasia, and

stromal metaplasia (females only) in the urinary

bladder; squamous hyperplasia, hyperkeratosis,

ulceration, and inflammation of the forestomach

mucosa; and seminal vesicle atrophy. Exposure of

male and female mice to 1-amino2,4-dibromoanthraquinonefor2yearswasassociated

withcentrilobularhepatocellularhypertrophy(males

only), basophilic focus, clear cell focus (females

only),eosinophilic focus, and pigmentationin the

liver; pigmentation in the kidney; and hyperplasia,

basal cellhyperplasia,hyperkeratosis,andinflammation of the forestomach mucosa.

__________

* Explanation of Levels of Evidence of Carcinogenic Activity is on page 13. A summary of the Technical Reports Review Subcommittee

 comments and the public discussion on this Technical Report appears on page 15.
85

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Sendelbach, L.E. (1989). A review of the toxicity

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Toxicology 57, 227-240.

Squire, R.A. (1989). Evaluation and grading of rat

liver foci in carcinogenicity tests. Toxicol. Pathol.

17, 685-689.
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Tennant, R.W., Margolin, B.H., Shelby, M.D.,

Zeiger, E., Haseman, J.K., Spalding, J., Caspary, W.,

Resnick, M., Stasiewicz, S., Anderson, B., and

Minor, R. (1987). Prediction of chemical carcinogenicity in rodents from in vitro genetic toxicity

assays. Science 236, 933-941.

United States International Trade Commission

(USITC) (1993). Synthetic organic chemicals:

United States production and sales, 1991. USITC

Publication 2607. U.S. Government Printing Office,

Washington, DC.

Ward, J.M., Rice, J.M., Creasia, D., Lynch, P., and

Riggs, C. (1983). Dissimilar patterns of promotion

by di(2-ethylhexyl)pthalate and phenobarbital of

hepatocellular neoplasia initiated by

diethylnitrosamine in B6C3F1 mice.

Carcinogenesis 4, 1021-1029.

Weisburger, E.K., Murthy, A.S.K., Lilja, H.S., and

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intermediates 4,4N-methylenebis(N,N-dimethyl)-

benzenamine, 4,4N-oxydianiline, and

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27, 103-117.

Williams, D.A. (1972). The comparison of several

dose levels with a zero dose control. Biometrics

28, 519-531.

Wilson, T.M., Nelson, P.E., and Knepp, C.R. (1985).

Hepatic neoplastic nodules, adenofibrosis, and

cholangiocarcinomas in male Fischer 344 rats fed

corn naturally contaminated with Fusarium

moniliforme. Carcinogenesis 6, 1155-1160.

Yang, R.S.H., Huff, J., Germolec, D.R.,

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Issues, and Relationships(N.N. Ragsdale and R.E.

Menzer, Eds.), pp. 142-163. American Chemical

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Washington, DC.

Zeiger, E., Anderson, B., Haworth, S., Lawlor, T.,

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300 chemicals. Environ. Mol. Mutagen. 11

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Zeiger, E., Haseman, J.K., Shelby, M.D.,

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Environ. Mol. Mutagen. 16 (Suppl. 18), 1-14.
92 1-Amino-2,4-dibromoanthraquinone, NTP TR 383
93

APPENDIX A

SUMMARY OF LESIONS IN MALE RATS

IN THE 2-YEAR FEED STUDY

OF 1-AMINO-2,4-DIBROMOANTHRAQUINONE

TABLE A1 Summary of the Incidence of Neoplasms in Male Rats 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 95

TABLE A2 Individual Animal Tumor Pathology of Male Rats 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 102

TABLE A3 Statistical Analysis of Primary Neoplasms in Male Rats 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 126

TABLE A4a Historical Incidence of Hepatocellular Neoplasms 

in Untreated Male F344/N Rats . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 135

TABLE A4b Historical Incidence of Large Intestine Neoplasms

in Untreated Male F344/N Rats . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 135

TABLE A4c Historical Incidence of Renal Tubule Neoplasms 

in Untreated Male F344/N Rats . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 136

TABLE A4d Historical Incidence of Urinary Bladder Neoplasms 

in Untreated Male F344/N Rats . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 136

TABLE A4e Historical Incidence of Forestomach Squamous Cell Neoplasms

in Untreated Male F344/N Rats . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 137

TABLE A5 Summary of the Incidence of Nonneoplastic Lesions in Male Rats 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 138
94 1-Amino-2,4-dibromoanthraquinone, NTP TR 383
Lesions in Male Rats 95

TABLE A1

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Disposition Summary

Animals initially in study 70 50 70 70

9-Month interim evaluation 10 10 10 10

15-Month interim evaluation 10 10

Early deaths

Moribund 19 15 34 33

Natural deaths 5 1 5 7

Survivors

Died last week of study 3

Terminal sacrifice 23 24 21 10

Animals examined microscopically 70 50 69b 70

9-Month Interim Evaluation

Alimentary System

Intestine large, colon (10) (10) (10) (10)

Liver (10) (10) (10) (10)

Hepatocellular adenoma 1 (10%)

Endocrine System

Adrenal medulla (10) (10) (10) (10)

Ganglioneuroma 1 (10%) 

Thyroid gland (10) (10) (10) (10)

Nervous System

Brain (10) (10) (10) (10)

Cerebrum, meningioma benign 1 (10%)

Systems Examined With No Neoplasms Observed 

Cardiovascular System

General Body System

Genital System

Hematopoietic System

Integumentary System

Musculoskeletal System

Respiratory System

Special Senses System

Urinary System
96 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A1

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

(continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

15-Month Interim Evaluation

Alimentary System

Intestine large, rectum (9) (10)

Polyp adenomatous 6 (60%)

Liver (10) (10)

Hepatocellular carcinoma 4 (40%)

Hepatocellular carcinoma, multiple 3 (30%)

Hepatocellular adenoma 2 (20%)

Hepatocellular adenoma, multiple 2 (20%)

Pancreas (10)

Adenoma 1 (10%)

Stomach, forestomach (10) (10)

Endocrine System

Pituitary gland (8)

Pars distalis, adenoma 1 (13%)

Thyroid gland (10) (1)

Adenoma 1 (10%) 

C-cell, adenoma 1 (10%) 

Genital System

Preputial gland (9)

Testes (10) (8)

Adenoma 1 (10%)

Interstitial cell, adenoma 2 (20%) 5 (63%)

Respiratory System

Lung (10) (3)

Alveolar/bronchiolar adenoma 1 (10%) 1 (33%)

Urinary System

Kidney (10) (10)

Renal tubule, adenoma 2 (20%)

Urinary bladder (10) (10)

Systemic Lesions

Multiple organsc(10) (10)

Leukemia mononuclear 2 (20%)

 

Mesothelioma malignant 1 (10%)

 
Lesions in Male Rats 97

TABLE A1

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

(continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

15-Month Interim Evaluation (continued)

Systems Examined With No Neoplasms Observed

Cardiovascular System

General Body System

Hematopoietic System

Integumentary System

Musculoskeletal System

Nervous System

Special Senses System

2-Year Study

Alimentary System

Intestine large, colon (47) (40) (59) (49)

Adenocarcinoma 1 (2%) 4 (8%)

Polyp adenomatous 1 (3%)

Polyp adenomatous, multiple 1 (2%) 3 (6%) 

Intestine large, rectum (46) (40) (58) (49)

Adenocarcinoma 1 (3%) 10 (17%) 12 (24%)

Adenocarcinoma, multiple 3 (6%)

Polyp adenomatous 12 (30%) 17 (29%) 10 (20%) 

Polyp adenomatous, multiple 1 (3%) 34 (59%) 30 (61%)

Intestine large, cecum (48) (40) (59) (50)

Intestine small, duodenum (48) (40) (59) (50)

Intestine small, jejunum (48) (38) (57) (48)

Intestine small, ileum (48) (40) (57) (49)

Liver (50) (40) (59) (50)

Cholangiocarcinoma 1 (2%)

Cholangioma 2 (3%)

Hepatocellular carcinoma 1 (2%) 11 (28%) 12 (20%) 9 (18%) 

Hepatocellular carcinoma, multiple 1 (3%) 43 (73%) 37 (74%) 

Hepatocellular adenoma 1 (2%) 10 (25%) 17 (29%) 10 (20%) 

Hepatocellular adenoma, multiple 10 (25%) 23 (39%) 24 (48%) 

Hepatocholangiocarcinoma 5 (8%) 2 (4%) 

Hepatocholangiocarcinoma, multiple 1 (2%)

Hepatocholangioma 1 (2%) 1 (2%) 

Myxoma 1 (2%) 

Mesentery (3) (2) (4) (4)

Pancreas (50) (40) (59) (50)

Adenocarcinoma, metastatic, intestine large, 

 rectum 1 (2%) 

Adenoma 2 (4%) 

Salivary glands (50) (40) (58) (49)

Stomach, forestomach (49) (39) (59) (49)

Squamous cell carcinoma 1 (2%) 

Squamous cell papilloma 2 (5%) 1 (2%)

Stomach, glandular (50) (40) (59) (50)

Cardiovascular System

Heart (50) (40) (59) (49)
98 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A1

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

(continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Endocrine System

Adrenal cortex (50) (40) (58) (50)

Hepatocellular carcinoma, metastatic, liver 1 (2%)

Adrenal medulla (50) (40) (58) (50)

Pheochromocytoma malignant 1 (2%) 2 (5%) 1 (2%)

Pheochromocytoma benign 11 (22%) 12 (30%) 11 (19%) 5 (10%) 

Bilateral, pheochromocytoma benign 1 (2%) 3 (8%) 2 (3%) 2 (4%) 

Islets, pancreatic (50) (40) (58) (50)

Adenoma 2 (4%) 2 (5%) 1 (2%) 

Parathyroid gland (43) (35) (55) (44)

Adenoma 1 (2%)

Pituitary gland (48) (40) (56) (49)

Pars distalis, adenoma 20 (42%) 12 (30%) 9 (16%) 10 (20%) 

Pars distalis, adenoma, multiple 1 (2%) 2 (5%) 1 (2%)

Pars distalis, carcinoma 1 (3%) 

Pars intermedia, adenoma 1 (3%) 

Thyroid gland (49) (40) (59) (50)

Adenoma 1 (2%) 

C-cell, adenoma 9 (18%) 5 (13%) 3 (5%) 3 (6%) 

Follicular cell, adenoma 2 (3%) 1 (2%) 

Follicular cell, carcinoma 2 (4%) 3 (8%) 1 (2%) 3 (6%) 

General Body System

None 

Genital System

Coagulating gland (2) (2) 

Epididymis (50) (40) (59) (50)

Preputial gland (49) (39) (58) (47)

Adenocarcinoma 1 (2%) 1 (2%)

Adenoma 3 (6%) 2 (5%) 1 (2%) 1 (2%) 

Prostate (50) (40) (59) (49)

Seminal vesicle (49) (40) (59) (50)

Testes (50) (40) (59) (50)

Bilateral, interstitial cell, adenoma 40 (80%) 34 (85%) 49 (83%) 38 (76%)

Interstitial cell, adenoma 3 (6%) 3 (8%) 6 (10%) 5 (10%) 

Hematopoietic System

Bone marrow (50) (40) (59) (50)

Lymph node (17) (5) (12) (20)

Lumbar, adenocarcinoma, metastatic, 

 intestine large, rectum 1 (8%) 

Lymph node, mandibular (50) (40) (54) (48)

Lymph node, mesenteric (48) (40) (57) (49)

Adenocarcinoma, metastatic, intestine large, 

 colon 1 (2%)
Lesions in Male Rats 99

TABLE A1

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

(continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Hematopoietic System (continued)

Spleen (50) (40) (58) (50)

Fibroma 1 (2%) 

Sarcoma 1 (3%) 

Thymus (37) (32) (41) (34)

Thymoma benign 1 (2%) 

Integumentary System

Mammary gland (27) (22) (29) (24)

Adenocarcinoma 1 (3%) 

Fibroadenoma 1 (5%) 

Fibroadenoma, multiple 1 (4%)

Skin (50) (38) (58) (50)

Basal cell adenoma 1 (2%) 1 (2%)

Basal cell carcinoma 2 (4%) 

Keratoacanthoma 1 (2%) 1 (3%) 1 (2%) 

Squamous cell papilloma 1 (2%) 3 (5%) 1 (2%) 

Trichoepithelioma 1 (2%) 

Pinna, squamous cell papilloma 2 (5%) 2 (3%) 1 (2%) 

Subcutaneous tissue, fibroma 2 (4%) 3 (5%) 1 (2%) 

Subcutaneous tissue, fibrosarcoma 1 (2%) 1 (2%) 

Subcutaneous tissue, lipoma 1 (2%) 

Subcutaneous tissue, sarcoma 1 (2%) 3 (8%) 1 (2%) 

Musculoskeletal System

Skeletal muscle (2) (1) (1)

Nervous System

Brain (50) (40) (59) (50)

Carcinoma, metastatic, pituitary gland 1 (3%) 

Oligodendroglioma benign 1 (2%) 

Meninges, granular cell tumor benign 1 (2%) 

Peripheral nerve (2) 

Squamous cell carcinoma, metastatic, 

 uncertain primary site 1 (50%)

Spinal cord (5) (1) (1) (4)

Respiratory System

Lung (50) (40) (59) (49)

Adenocarcinoma, metastatic, kidney 1 (2%) 

Adenocarcinoma, metastatic, intestine large, 

 colon 1 (2%) 

Alveolar/bronchiolar adenoma 2 (5%) 2 (3%) 3 (6%) 

Alveolar/bronchiolar carcinoma 1 (3%) 2 (3%) 

Hepatocellular carcinoma, metastatic, liver 1 (3%) 18 (31%) 19 (39%) 

Sarcoma, metastatic, kidney 1 (2%) 

Sarcoma, metastatic, skin 1 (2%) 

Mediastinum, alveolar/bronchiolar carcinoma 1 (2%) 
100 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A1

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

(continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Respiratory System (continued)

Nose (48) (40) (59) (50)

Trachea (50) (40) (59) (50)

Special Senses System

Ear (3) (7) (2) (1)

Fibrosarcoma 1 (33%) 

Eye (6) (3) (1) (6)

Zymbal's gland (1) (1) (1) (2)

Carcinoma 1 (100%) 1 (100%) 1 (50%) 

Squamous cell carcinoma 1 (100%) 1 (50%) 

Urinary System

Kidney (50) (40) (59) (50)

Hepatocellular carcinoma, metastatic, liver 1 (3%) 

Pelvis, transitional epithelium, papilloma 1 (2%) 

Renal tubule, adenoma 2 (4%) 6 (15%) 7 (12%) 9 (18%) 

Renal tubule, adenoma, multiple 4 (10%) 4 (7%) 5 (10%) 

Renal tubule, carcinoma 2 (3%) 1 (2%) 

Urinary bladder (50) (38) (58) (50)

Transitional epithelium, carcinoma 1 (2%) 4 (8%) 

Transitional epithelium, papilloma 1 (3%) 2 (3%) 8 (16%) 

Systemic Lesions

Multiple organs (50) (40) (59) (50)

Leukemia mononuclear 25 (50%) 5 (13%) 3 (5%) 1 (2%) 

Mesothelioma malignant 1 (2%) 1 (3%) 1 (2%) 1 (2%) 

Neoplasm Summary

Total animals with primary neoplasmsd

9-Month interim evaluation 1 2

15-Month interim evaluation 7 10

2-Year study 49 40 59 48

Total primary neoplasms

9-Month interim evaluation 1 2

15-Month interim evaluation 8 28

2-Year study 140 160 297 258

Total animals with benign neoplasms

9-Month interim evaluation 1 2

15-Month interim evaluation 7 10

2-Year study 49 40 58 47

Total benign neoplasms

9-Month interim evaluation 1 2

15-Month interim evaluation 8 18

2-Year study 103 129 209 176
Lesions in Male Rats 101

TABLE A1

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

(continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Neoplasm Summary (continued)

Total animals with malignant neoplasms

15-Month interim evaluation 8

2-Year study 29 24 57 47

Total malignant neoplasms

15-Month interim evaluation 10

2-Year study 37 31 88 82

Total animals with metastatic neoplasms

2-Year study 2 3 20 22

Total metastatic neoplasms

2-Year study 2 3 23 22

Total animals with malignant neoplasms

 of uncertain primary site

2-Year study 1

a Number of animals examined microscopically at site and number of animals with neoplasm b One animal not examined microscopically c Number of animals with any tissue examined microscopically d Primary neoplasms: all neoplasms except metastatic neoplasms
102 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm

4445555555666666666677777 

Number of Days on Study 2231456778345677779902223 

1754962347164614492671171

1100101110000000100000101

Carcass ID Number 1278134321835361081927153

5255435445424154532334331

Alimentary System 

Esophagus + + + + + + + + + + M + + + + + + + + + + + + + +

Intestine large, colon A + + + + + + + + + A + M + + + + + + + + + + + +

Intestine large, rectum A + + + + + + + + + A + + + + + + + + + + + M + +

Intestine large, cecum A + + + + + + + + + A + + + + + + + + + + + + + +

Intestine small, duodenum A + + + + + + + + + A + + + + + + + + + + + + + +

Intestine small, jejunum A + + + + + + + + + A + + + + + + + + + + + + + +

Intestine small, ileum A + + + + + + + + + A + + + + + + + + + + + + + +

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 

Hepatocellular carcinoma X 

Hepatocellular adenoma 

Mesentery + + + 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + +

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + +

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + +

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + +

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + +

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma malignant X 

Pheochromocytoma benign X X X X X 

Bilateral, pheochromocytoma benign 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma 

Parathyroid gland + + + M + + + + + + + + + M + M + + + + + M + + M

Pituitary gland + + M + + + + + + + + + + + + + + + + + + + + + + 

Pars distalis, adenoma X X X X X X X X X X 

Pars distalis, adenoma, multiple X 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

C-cell, adenoma X X X X X 

Follicular cell, carcinoma X

General Body System 

None 

Genital System

Coagulating gland + 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + +

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenocarcinoma X 

Adenoma X 

+: Tissue examined microscopically M: Missing tissue X: Lesion present

A: Autolysis precludes examination I: Insufficient tissue Blank: Not examined
Lesions in Male Rats 103

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

7777777777777777777777777

Number of Days on Study 3333333333333333333333333

1134444444444444444455555

1 1 1 0 0 0 0 0 0 0 0 0 0 1 1 1 1 1 1 1 0 0 0 0 1 Total

Carcass ID Number 3 4 4 2 2 4 5 5 6 6 7 8 9 0 0 0 1 2 2 4 1 6 7 8 4 Tissues/

2 2 3 1 2 2 1 2 1 3 2 1 1 1 3 4 2 1 3 1 1 4 3 2 4 Tumors

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine large, rectum + + + + + + + + + + + + M + + + + + + + + + + + + 46 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hepatocellular carcinoma 1

Hepatocellular adenoma X 1

Mesentery 3

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X X 2

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + M + + + + + + + + + + + + + + + + + 49 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pheochromocytoma malignant 1

Pheochromocytoma benign X X X X X X 11 

Bilateral, pheochromocytoma benign X 1

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X X 2

Parathyroid gland + + + + + + + + + + + + + M + + + + + + + + + M + 43 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + I + 48 

Pars distalis, adenoma X X X X X X X X X X 20 

Pars distalis, adenoma, multiple 1

Thyroid gland + + + + + + + + + + + + + A + + + + + + + + + + + 49 

C-cell, adenoma X X X X 9

Follicular cell, carcinoma X 2

General Body System 

None 

Genital System 

Coagulating gland + 2

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Preputial gland + + + + + + + + + + + + + + M + + + + + + + + + + 49 

Adenocarcinoma 1

Adenoma X X 3
104 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

4445555555666666666677777 

Number of Days on Study 2231456778345677779902223 

1754962347164614492671171

1100101110000000100000101 

Carcass ID Number 1278134321835361081927153 

5255435445424154532334331

Genital System (continued) 

Prostate + + + + + + + + + + + + + + + + + + + + + + + + +

Seminal vesicle + + + M + + + + + + + + + + + + + + + + + + + + +

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 

Bilateral, interstitial cell, adenoma X X X X X X X X X X X X X X X X X 

Interstitial cell, adenoma X X 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node + + + + + + + + + + + 

Lymph node, mandibular + + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node, mesenteric + + + + + M + + + + + + + + + + + M + + + + + + +

Spleen + + + + + + + + + + + + + + + + + + + + + + + + +

Thymus + + M + + M + + + + + M + + M + + + M M + + M + +

Integumentary System 

Mammary gland M + M + + + M M + M + + M + + M M + + + M + M M M 

Fibroadenoma, multiple X 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 

Basal cell carcinoma 

Keratoacanthoma X 

Squamous cell papilloma 

Trichoepithelioma 

Subcutaneous tissue, fibroma X 

Subcutaneous tissue, fibrosarcoma X 

Subcutaneous tissue, lipoma X 

Subcutaneous tissue, sarcoma X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + +

Skeletal muscle + 

Nervous System

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 

Oligodendroglioma benign X 

Spinal cord + + + + + 

Respiratory System 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma malignant, 

metastatic, adrenal medulla X 

Sarcoma, metastatic, skin X 

Nose + + + + + + + + + + + + + + + + + + + + + + + +

Trachea + + + + + + + + + + + + + + + + + + + + + + + + +
Lesions in Male Rats 105

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

7777777777777777777777777

Number of Days on Study 3333333333333333333333333

1134444444444444444455555

1 1 1 0 0 0 0 0 0 0 0 0 0 1 1 1 1 1 1 1 0 0 0 0 1 Total

Carcass ID Number 3 4 4 2 2 4 5 5 6 6 7 8 9 0 0 0 1 2 2 4 1 6 7 8 4 Tissues/

2 2 3 1 2 2 1 2 1 3 2 1 1 1 3 4 2 1 3 1 1 4 3 2 4 Tumors

Genital System (continued) 

Prostate + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Bilateral, interstitial cell, adenoma X X X X X X X X X X X X X X X X X X X X X X X 40 

Interstitial cell, adenoma X 3

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymph node + + + + + + 17 

Lymph node, mandibular + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Thymus M M M + M + + + + M + + + + + + + + M + + + + + + 37 

Integumentary System 

Mammary gland M M + + + M M + + + + MMMM+M++MM + + + + 27 

Fibroadenoma, multiple 1

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Basal cell carcinoma X X 2

Keratoacanthoma 1

Squamous cell papilloma X 1

Trichoepithelioma X 1

Subcutaneous tissue, fibroma X 2

Subcutaneous tissue, fibrosarcoma 1

Subcutaneous tissue, lipoma 1

Subcutaneous tissue, sarcoma 1

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Skeletal muscle + 2

Nervous System

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Oligodendroglioma benign 1

Spinal cord 5

Respiratory System 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pheochromocytoma malignant, 

metastatic, adrenal medulla 1

Sarcoma, metastatic, skin 1

Nose + + M + + + + + + + + + + + + + + + + + + + + + + 48 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 50 
106 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

4445555555666666666677777 

Number of Days on Study 2231456778345677779902223 

1754962347164614492671171

1100101110000000100000101 

Carcass ID Number 1278134321835361081927153 

5255435445424154532334331

Special Senses System 

Ear + + + 

Fibrosarcoma X 

Eye + + + + + 

Zymbal's gland 

Carcinoma 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 

Renal tubule, adenoma 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + +

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Leukemia mononuclear X X X X X X X X X X X X X X X X X 

Mesothelioma malignant 
Lesions in Male Rats 107

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

7777777777777777777777777

Number of Days on Study 3333333333333333333333333

1134444444444444444455555

1 1 1 0 0 0 0 0 0 0 0 0 0 1 1 1 1 1 1 1 0 0 0 0 1 Total

Carcass ID Number 3 4 4 2 2 4 5 5 6 6 7 8 9 0 0 0 1 2 2 4 1 6 7 8 4 Tissues/

2 2 3 1 2 2 1 2 1 3 2 1 1 1 3 4 2 1 3 1 1 4 3 2 4 Tumors

Special Senses System 

Ear 3

Fibrosarcoma 1

Eye I + 6

Zymbal's gland + 1

Carcinoma X 1

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Renal tubule, adenoma X X 2

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Leukemia mononuclear X X X X X X X X 25 

Mesothelioma malignant X 1
108 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

2,000 ppm

6666666666666777777777777 

Number of Days on Study 0124555677799012333333333 

4890039834534080000111111

1112212212212211222111111 

Carcass ID Number 5682260450174067333558889 

4542144333452234345121231

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 

Polyp adenomatous 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenocarcinoma X 

Polyp adenomatous X X X X X X X 

Polyp adenomatous, multiple 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + M + + M + +

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + +

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 

Hepatocellular carcinoma X X X X X X X 

Hepatocellular carcinoma, multiple 

Hepatocellular adenoma X X X X X 

Hepatocellular adenoma, multiple X X X X X X

Mesentery + 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + +

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + +

Stomach, forestomach + + + M + + + + + + + + + + + + + + + + + + + + + 

Squamous cell papilloma X X 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + +

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + +

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + +

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma malignant X X 

Pheochromocytoma benign X X X X X X X X 

Bilateral, pheochromocytoma benign X X 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma X X 

Parathyroid gland + + + + + + + + + + + + + + + + + M + M + + + + +

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Pars distalis, adenoma X X X X X X X X X 

Pars distalis, adenoma, multiple X 

Pars distalis, carcinoma X 

Pars intermedia, adenoma 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

C-cell, adenoma X X X 

Follicular cell, carcinoma X X X 

General Body System 

None 
Lesions in Male Rats 109

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

2,000 ppm (continued)

777777777777777 

Number of Days on Study 333333333333333 

111111444444444 

1 1 1 2 2 2 1 1 1 1 1 2 2 2 2 Total

Carcass ID Number 9 9 9 0 3 3 6 6 7 7 7 1 1 1 4 Tissues/

2 3 4 1 1 2 1 2 1 2 3 1 2 3 1 Tumors

Alimentary System 

Esophagus + + + + + + + + + + + + + + + 40 

Intestine large, colon + + + + + + + + + + + + + + + 40 

Polyp adenomatous X 1

Intestine large, rectum + + + + + + + + + + + + + + + 40 

Adenocarcinoma 1

Polyp adenomatous X X X X X 12 

Polyp adenomatous, multiple X 1

Intestine large, cecum + + + + + + + + + + + + + + + 40 

Intestine small, duodenum + + + + + + + + + + + + + + + 40 

Intestine small, jejunum + + + + + + + + + + + + + + + 38 

Intestine small, ileum + + + + + + + + + + + + + + + 40 

Liver + + + + + + + + + + + + + + + 40 

Hepatocellular carcinoma X X X X 11 

Hepatocellular carcinoma, multiple X 1

Hepatocellular adenoma X X X X X 10 

Hepatocellular adenoma, multiple X X X X 10 

Mesentery + 2

Pancreas + + + + + + + + + + + + + + + 40 

Salivary glands + + + + + + + + + + + + + + + 40 

Stomach, forestomach + + + + + + + + + + + + + + + 39 

Squamous cell papilloma 2

Stomach, glandular + + + + + + + + + + + + + + + 40 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + 40 

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + 40 

Adrenal medulla + + + + + + + + + + + + + + + 40 

Pheochromocytoma malignant 2

Pheochromocytoma benign X X X X 12 

Bilateral, pheochromocytoma benign X 3

Islets, pancreatic + + + + + + + + + + + + + + + 40 

Adenoma 2

Parathyroid gland M + + + + + + + M + M + + + + 35 

Pituitary gland + + + + + + + + + + + + + + + 40 

Pars distalis, adenoma X X X 12 

Pars distalis, adenoma, multiple X 2

Pars distalis, carcinoma 1

Pars intermedia, adenoma X 1

Thyroid gland + + + + + + + + + + + + + + + 40 

C-cell, adenoma X X 5

Follicular cell, carcinoma 3

General Body System 

None 
110 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

2,000 ppm (continued)

6666666666666777777777777 

Number of Days on Study 0124555677799012333333333 

4890039834534080000111111

1112212212212211222111111 

Carcass ID Number 5682260450174067333558889 

4542144333452234345121231

Genital System 

Coagulating gland + + 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + +

Preputial gland + + + + + + + + + + + + + + + + + + + M + + + + + 

Adenoma X X 

Prostate + + + + + + + + + + + + + + + + + + + + + + + + +

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + +

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 

Bilateral, interstitial cell, adenoma X X X X X X X X X X X X X X X X X X X 

Interstitial cell, adenoma X X X

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node + + + + + 

Lymph node, mandibular + + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + +

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 

Sarcoma 

Thymus M M M M +++M+++M+++M+++++++M+

Integumentary System 

Mammary gland + + + M + M + M + M M + + + M M + MMM+M+++ 

Fibroadenoma 

Skin + + + + + + + + + + + + + + + + + + + + + I + + + 

Keratoacanthoma X 

Pinna, squamous cell papilloma X 

Subcutaneous tissue, sarcoma X X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + +

Skeletal muscle + 

Nervous System

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 

Carcinoma, metastatic, pituitary 

gland X 

Spinal cord + 

Respiratory System 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma X 

Hepatocellular carcinoma, metastatic, 

liver 

Nose + + + + + + + + + + + + + + + + + + + + + + + + +

Trachea + + + + + + + + + + + + + + + + + + + + + + + + +
Lesions in Male Rats 111

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

2,000 ppm (continued)

777777777777777 

Number of Days on Study 333333333333333 

111111444444444 

1 1 1 2 2 2 1 1 1 1 1 2 2 2 2 Total

Carcass ID Number 9 9 9 0 3 3 6 6 7 7 7 1 1 1 4 Tissues/

2 3 4 1 1 2 1 2 1 2 3 1 2 3 1 Tumors

Genital System 

Coagulating gland 2

Epididymis + + + + + + + + + + + + + + + 40 

Preputial gland + + + + + + + + + + + + + + + 39 

Adenoma 2

Prostate + + + + + + + + + + + + + + + 40 

Seminal vesicle + + + + + + + + + + + + + + + 40 

Testes + + + + + + + + + + + + + + + 40 

Bilateral, interstitial cell, adenoma X X X X X X X X X X X X X X X 34 

Interstitial cell, adenoma 3

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + 40 

Lymph node 5

Lymph node, mandibular + + + + + + + + + + + + + + + 40 

Lymph node, mesenteric + + + + + + + + + + + + + + + 40 

Spleen + + + + + + + + + + + + + + + 40 

Sarcoma X 1

Thymus + + + + + + + + + + + + + + + 32 

Integumentary System 

Mammary gland + M + + + + M + M + M M M M + 22 

Fibroadenoma X 1

Skin + + + + + + + + + I + + + + + 38 

Keratoacanthoma 1

Pinna, squamous cell papilloma X 2

Subcutaneous tissue, sarcoma X 3

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + 40 

Skeletal muscle 1

Nervous System

Brain + + + + + + + + + + + + + + + 40 

Carcinoma, metastatic, pituitary 

gland 1

Spinal cord 1

Respiratory System 

Lung + + + + + + + + + + + + + + + 40 

Alveolar/bronchiolar adenoma X X 2

Alveolar/bronchiolar carcinoma 1

Hepatocellular carcinoma, metastatic, 

liver X 1

Nose + + + + + + + + + + + + + + + 40 

Trachea + + + + + + + + + + + + + + + 40 
112 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

2,000 ppm (continued)

6666666666666777777777777 

Number of Days on Study 0124555677799012333333333 

4890039834534080000111111

1112212212212211222111111 

Carcass ID Number 5682260450174067333558889 

4542144333452234345121231

Special Senses System 

Ear + + + + + + + 

Eye I + 

Zymbal's gland + 

Carcinoma X 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 

Hepatocellular carcinoma, metastatic, 

liver X 

Renal tubule, adenoma X X X 

Renal tubule, adenoma, multiple X X X X 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + M + 

Transitional epithelium, papilloma X 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Leukemia mononuclear X X X 

Mesothelioma malignant X 
Lesions in Male Rats 113

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

2,000 ppm (continued)

777777777777777 

Number of Days on Study 333333333333333 

111111444444444 

1 1 1 2 2 2 1 1 1 1 1 2 2 2 2 Total

Carcass ID Number 9 9 9 0 3 3 6 6 7 7 7 1 1 1 4 Tissues/

2 3 4 1 1 2 1 2 1 2 3 1 2 3 1 Tumors

Special Senses System 

Ear 7

Eye + + 3

Zymbal's gland 1

Carcinoma 1

Urinary System 

Kidney + + + + + + + + + + + + + + + 40 

Hepatocellular carcinoma, metastatic, 

liver 1

Renal tubule, adenoma X X X 6

Renal tubule, adenoma, multiple 4

Urinary bladder + + + + M + + + + + + + + + + 38 

Transitional epithelium, papilloma 1

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + 40 

Leukemia mononuclear X X 5

Mesothelioma malignant 1
114 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

5,000 ppm

445555556666666666666666666667

Number of Days on Study 672267880002223335555567778890

581136245893996670023914445730

332332333322333332332322333323

Carcass ID Number 526507328799154438285457135870

533445425543432335145144322333

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + M + + + + + +

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Adenocarcinoma 

Polyp adenomatous, multiple 

Intestine large, rectum + + + + + M + + + + + + + + + + + + + + + + + + + + + + + +

Adenocarcinoma X X X X X 

Polyp adenomatous X X X X X X X X X X X 

Polyp adenomatous, multiple X X X X X X X X X X X

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, jejunum + + A + + + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + M + + + + +

Liver + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Cholangioma X 

Hepatocellular carcinoma X X X X X X 

Hepatocellular carcinoma, multiple X X X X X X X X X X X X X X X X X X X X X

Hepatocellular adenoma X X X X X X X X X 

Hepatocellular adenoma, multiple X X X X X X X X

Hepatocholangiocarcinoma X 

Hepatocholangiocarcinoma, multiple X 

Hepatocholangioma X 

Myxoma 

Mesentery + 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Salivary glands + + + + + + M + + + + + + + + + + + + + + + + + + + + + + +

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Tooth + + 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Pheochromocytoma malignant X 

Pheochromocytoma benign X X X X X X

Bilateral, pheochromocytoma benign 

Islets, pancreatic + A + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Parathyroid gland + + + + + + + + + M + + + + + + + + + + + + + + + M + + + +

Adenoma 

Pituitary gland + + + + + + + + + + + + M + + + + + + + + + + + + M + + + +

Pars distalis, adenoma X X 

Pars distalis, adenoma, multiple X 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

C-cell, adenoma 

Follicular cell, adenoma X 

Follicular cell, carcinoma X 
Lesions in Male Rats 115

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

5,000 ppm (continued)

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7

Number of Days on Study 0 0 0 1 1 1 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 3 3 3 3 3 3 3 3

0 2 9 2 4 5 0 8 9 9 9 9 9 9 9 9 9 9 9 9 9 0 0 0 0 0 0 0 0

3 3 3 2 2 3 2 3 2 2 2 2 2 2 2 2 2 2 3 3 3 3 3 3 3 3 3 3 3 Total

Carcass ID Number 7 1 6 6 8 6 5 6 5 5 6 7 7 8 8 8 9 9 0 0 1 3 5 6 6 7 7 8 8 Tissues/

3 2 5 2 4 4 3 3 1 2 1 1 2 1 2 3 1 2 1 2 1 1 1 1 2 1 2 1 2 Tumors

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 58

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Adenocarcinoma X 1

Polyp adenomatous, multiple X 1

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 58

Adenocarcinoma X X X X X 10

Polyp adenomatous X X X X X X 17 

Polyp adenomatous, multiple X X X X X X X X X X X X X X X X X X X X X X X 34

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + M + + + + 57

Intestine small, ileum + + + M + + + + + + + + + + + + + + + + + + + + + + + + + 57

Liver + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Cholangioma X 2

Hepatocellular carcinoma X X X X X X 12

Hepatocellular carcinoma, multiple X X X X X X X X X X X X X X X X X X X X X X 43

Hepatocellular adenoma X X X X X X X X 17

Hepatocellular adenoma, multiple X X X X X X X X X X X X X X X 23

Hepatocholangiocarcinoma X X X X 5

Hepatocholangiocarcinoma, multiple 1

Hepatocholangioma 1

Myxoma X 1

Mesentery + + + 4

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 58

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Tooth 2

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + M + + + + 58

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + M + + + + 58

Pheochromocytoma malignant 1

Pheochromocytoma benign X X X X X 11

Bilateral, pheochromocytoma benign X X 2

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 58

Parathyroid gland + + + + M + + + + + + + + + + + + M + + + + + + + + + + + 55

Adenoma X 1

Pituitary gland + + + + + M + + + + + + + + + + + + + + + + + + + + + + + 56

Pars distalis, adenoma X X X X X X X 9

Pars distalis, adenoma, multiple 1

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

C-cell, adenoma X X X 3

Follicular cell, adenoma X 2

Follicular cell, carcinoma 1
116 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

5,000 ppm (continued)

445555556666666666666666666667

Number of Days on Study 672267880002223335555567778890

581136245893996670023914445730

332332333322333332332322333323

Carcass ID Number 526507328799154438285457135870

533445425543432335145144322333

General Body System 

None 

 

Genital System

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Adenocarcinoma X

Adenoma X 

Prostate + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Testes + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Bilateral, interstitial cell, adenoma X X X X X X X X X X X X X X X X X X X X X X X X X

Interstitial cell, adenoma X X 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node + + + + + 

Lumbar, adenocarcinoma, metastatic, 

intestine large, rectum X 

Lymph node, mandibular + + + + + + M M + + + + + + + + + + + + + + + + + + M + + +

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + + + + + M

Adenocarcinoma, metastatic, 

intestine large, colon 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Fibroma 

Thymus + M + + M + M M M M ++++++++ MMM++M++M+++

Thymoma benign 

Integumentary System 

Mammary gland M M + + M + + + + + + + + M + M + MMMM ++++ MMM++

Adenocarcinoma X 

Skin + I + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Basal cell adenoma 

Squamous cell papilloma X 

Pinna, squamous cell papilloma 

Subcutaneous tissue, fibroma 

Subcutaneous tissue, fibrosarcoma X 

Subcutaneous tissue, sarcoma X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Skeletal muscle + 
Lesions in Male Rats 117

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

5,000 ppm (continued)

77777777777777777777777777777

Number of Days on Study 00011122222222222222233333333

02924508999999999999900000000

3 3 3 2 2 3 2 3 2 2 2 2 2 2 2 2 2 2 3 3 3 3 3 3 3 3 3 3 3 Total

Carcass ID Number 7 1 6 6 8 6 5 6 5 5 6 7 7 8 8 8 9 9 0 0 1 3 5 6 6 7 7 8 8 Tissues/

3 2 5 2 4 4 3 3 1 2 1 1 2 1 2 3 1 2 1 2 1 1 1 1 2 1 2 1 2 Tumors

General Body System 

None 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Preputial gland + M + + + + + + + + + + + + + + + + + + + + + + + + + + + 58

Adenocarcinoma 1

Adenoma 1

Prostate + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Testes + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Bilateral, interstitial cell, adenoma X X X X X X X X X X X X X X X X X X X X X X X X 49

Interstitial cell, adenoma X X X X 6

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Lymph node + + + + + + + 12

Lumbar, adenocarcinoma, metastatic, 

intestine large, rectum 1

Lymph node, mandibular + + M + + + + + + + + + + + + + + + + + + M + + + + + + + 54

Lymph node, mesenteric M + + + + + + + + + + + + + + + + + + + + + + + + + + + + 57

Adenocarcinoma, metastatic, 

intestine large, colon X 1

Spleen + + + + + + + M + + + + + + + + + + + + + + + + + + + + + 58

Fibroma X 1

Thymus M + M M + + M M + + M + + + + + + + + + + + + + + + + + M 41

Thymoma benign X 1

Integumentary System 

Mammary gland M M M M M M M + M M M M M M + M M M + M + + + + + M + + + 29

Adenocarcinoma 1

Skin + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 58

Basal cell adenoma X 1

Squamous cell papilloma X X 3

Pinna, squamous cell papilloma X X 2

Subcutaneous tissue, fibroma X X X 3

Subcutaneous tissue, fibrosarcoma 1

Subcutaneous tissue, sarcoma 1

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Skeletal muscle 1
118 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

5,000 ppm (continued)

445555556666666666666666666667

Number of Days on Study 672267880002223335555567778890

581136245893996670023914445730

332332333322333332332322333323

Carcass ID Number 526507328799154438285457135870

533445425543432335145144322333

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Meninges, granular cell tumor benign 

Peripheral nerve + 

Squamous cell carcinoma, metastatic, 

uncertain primary site 

Spinal cord + 

Respiratory System 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Adenocarcinoma, metastatic, kidney X 

Adenocarcinoma, metastatic, 

intestine large, colon 

Alveolar/bronchiolar adenoma X 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, 

liver X X X X X X X X X X X X X

Nose + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Special Senses System 

Ear + 

Eye + 

Zymbal's gland + 

Squamous cell carcinoma X 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Pelvis, transitional epithelium, 

papilloma X 

Renal tubule, adenoma X X X 

Renal tubule, adenoma, multiple X X 

Renal tubule, carcinoma X 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + + + + M +

Transitional epithelium, carcinoma 

Transitional epithelium, papilloma 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Leukemia mononuclear X 

Mesothelioma malignant 
Lesions in Male Rats 119

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

5,000 ppm (continued)

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7

Number of Days on Study 0 0 0 1 1 1 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 3 3 3 3 3 3 3 3

0 2 9 2 4 5 0 8 9 9 9 9 9 9 9 9 9 9 9 9 9 0 0 0 0 0 0 0 0

3 3 3 2 2 3 2 3 2 2 2 2 2 2 2 2 2 2 3 3 3 3 3 3 3 3 3 3 3 Total

Carcass ID Number 7 1 6 6 8 6 5 6 5 5 6 7 7 8 8 8 9 9 0 0 1 3 5 6 6 7 7 8 8 Tissues/

3 2 5 2 4 4 3 3 1 2 1 1 2 1 2 3 1 2 1 2 1 1 1 1 2 1 2 1 2 Tumors

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Meninges, granular cell tumor benign X 1

Peripheral nerve + 2

Squamous cell carcinoma, metastatic, 

uncertain primary site X 1

Spinal cord 1

Respiratory System 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Adenocarcinoma, metastatic, kidney 1

Adenocarcinoma, metastatic, 

intestine large, colon X 1

Alveolar/bronchiolar adenoma X 2

Alveolar/bronchiolar carcinoma X X 2

Hepatocellular carcinoma, metastatic, 

liver X X X X X 18

Nose + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Special Senses System 

Ear + 2

Eye 1

Zymbal's gland 1

Squamous cell carcinoma 1

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Pelvis, transitional epithelium, 

papilloma 1

Renal tubule, adenoma X X X X 7

Renal tubule, adenoma, multiple X X 4

Renal tubule, carcinoma X 2

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 58

Transitional epithelium, carcinoma X 1

Transitional epithelium, papilloma X X 2

Systemic Lesions

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Leukemia mononuclear X X 3

Mesothelioma malignant X 1
120 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm

2334444455555555555666666 

Number of Days on Study 8563379924557788999012223 

1245663772233718016502451

4454443444534444443544444 

Carcass ID Number 4904939758098526359124193 

5544445553242432332523431

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenocarcinoma X X 

Polyp adenomatous, multiple X 

Intestine large, rectum + + + + + + + + + M + + + + + + + + + + + + + + + 

Adenocarcinoma X X X X 

Adenocarcinoma, multiple X X 

Polyp adenomatous X X X X X X X X 

Polyp adenomatous, multiple X X X X X X X

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, jejunum + + + + + + + + + + + A + + + + A + + + + + + + +

Intestine small, ileum + + + + + + + + + + + + + + + + A + + + + + + + +

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 

Cholangiocarcinoma 

Hepatocellular carcinoma X X X X X X X X 

Hepatocellular carcinoma, multiple X X X X X X X X X X X X X 

Hepatocellular adenoma X X X X 

Hepatocellular adenoma, multiple X X X X X X X 

Hepatocholangiocarcinoma X 

Hepatocholangioma X 

Mesentery + + 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenocarcinoma, metastatic, 

intestine large, rectum X 

Pharynx + 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + +

Stomach, forestomach + + + + M + + + + + + + + + + + + + + + + + + + + 

Squamous cell carcinoma 

Squamous cell papilloma 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + +

Tongue 

Cardiovascular System 

Heart + + + + + + + + + + + + M + + + + + + + + + + + +

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 

Hepatocellular carcinoma, metastatic, 

liver 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma benign X 

Bilateral, pheochromocytoma benign X

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + +

Adenoma 
Lesions in Male Rats 121

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

6666666777777777777777777

Number of Days on Study 3477778000112222222222222

1614499239451449999999999

5 5 5 5 5 4 4 5 4 3 4 4 4 4 4 4 4 4 4 4 4 4 5 5 5 Total

Carcass ID Number 2 0 2 1 2 0 7 1 9 9 5 7 2 7 8 0 1 1 4 4 6 9 1 1 2 Tissues/

4 1 3 4 2 4 4 3 2 1 2 3 1 1 1 1 2 3 1 2 1 1 1 2 1 Tumors

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, colon + + + + + + + + + + + + + + + + + + M + + + + + + 49 

Adenocarcinoma X X 4

Polyp adenomatous, multiple X X 3

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Adenocarcinoma X X X X X X X X 12 

Adenocarcinoma, multiple X 3

Polyp adenomatous X X 10 

Polyp adenomatous, multiple X X X X X X X X X X X X X X X X X X X X X X X 30 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Cholangiocarcinoma X 1

Hepatocellular carcinoma X 9

Hepatocellular carcinoma, multiple X X X X X X X X X X X X X X X X X X X X X X X X 37 

Hepatocellular adenoma X X X X X X 10 

Hepatocellular adenoma, multiple X X X X X X X X X X X X X X X X X 24 

Hepatocholangiocarcinoma X 2

Hepatocholangioma 1

Mesentery + + 4

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenocarcinoma, metastatic, 

intestine large, rectum 1

Pharynx 1

Salivary glands + + M + + + + + + + + + + + + + + + + + + + + + + 49 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Squamous cell carcinoma X 1

Squamous cell papilloma X 1

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Tongue + 1

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hepatocellular carcinoma, metastatic, 

liver X 1

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pheochromocytoma benign X X X X 5

Bilateral, pheochromocytoma benign X 2

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X 1
122 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

2334444455555555555666666 

Number of Days on Study 8563379924557788999012223 

1245663772233718016502451

4454443444534444443544444 

Carcass ID Number 4904939758098526359124193 

5544445553242432332523431

Endocrine System (continued) 

Parathyroid gland + + + + + + + + + + M + + M + + + + M + + + + + +

Pituitary gland + + + M + + + + + + + + + + + + + + + + + + + + + 

Pars distalis, adenoma X X X X X 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma X 

C-cell, adenoma 

Follicular cell, adenoma 

Follicular cell, carcinoma X 

General Body System 

None 

Genital System

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + +

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma X 

Prostate + + + + + M + + + + + + + + + + + + + + + + + + +

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + +

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 

Bilateral, interstitial cell, adenoma X X X X X X X X X X X X X X X X X 

Interstitial cell, adenoma 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node + + + + + + +

Lymph node, mandibular + + + + + + M + + + + + + + + + + + + + + + + + +

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + M + +

Spleen + + + + + + + + + + + + + + + + + + + + + + + + +

Thymus M + + M + + + M + + + + M + + M + M + + + M + M +

Integumentary System 

Mammary gland M + M M M M +++++M++++M++++++MM

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 

Basal cell adenoma 

Keratoacanthoma X 

Squamous cell papilloma 

Pinna, squamous cell papilloma 

Subcutaneous tissue, fibroma 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + +

Nervous System

Brain + + + + + + + + + + + + + + + + + + + + + + + + +

Spinal cord + + + 
Lesions in Male Rats 123

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

6666666777777777777777777

Number of Days on Study 3477778000112222222222222

1614499239451449999999999

5 5 5 5 5 4 4 5 4 3 4 4 4 4 4 4 4 4 4 4 4 4 5 5 5 Total

Carcass ID Number 2 0 2 1 2 0 7 1 9 9 5 7 2 7 8 0 1 1 4 4 6 9 1 1 2 Tissues/

4 1 3 4 2 4 4 3 2 1 2 3 1 1 1 1 2 3 1 2 1 1 1 2 1 Tumors

Endocrine System (continued) 

Parathyroid gland + + + + + M + + + + M + + + + M + + + + + + + + + 44 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Pars distalis, adenoma X X X X X 10 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma 1

C-cell, adenoma X X X 3

Follicular cell, adenoma X 1

Follicular cell, carcinoma X X 3

General Body System 

None 

Genital System

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Preputial gland M + + + + + M + + + + + + + + + + + M + + + + + + 47 

Adenoma 1

Prostate + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Bilateral, interstitial cell, adenoma X X X X X X X X X X X X X X X X X X X X X 38 

Interstitial cell, adenoma X X X X X 5

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymph node + + + + + + + + + + + + + 20 

Lymph node, mandibular + + + + + + + + + + + + + + + + + + M + + + + + + 48 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Thymus M M + + M + M + M + + + M + M M + + + + + + + + + 34 

Integumentary System 

Mammary gland M + M + M M M + + M M M + M M M M M M M M + + + M 24 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Basal cell adenoma X 1

Keratoacanthoma 1

Squamous cell papilloma X 1

Pinna, squamous cell papilloma X 1

Subcutaneous tissue, fibroma X 1

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Nervous System

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Spinal cord + 4
124 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

2334444455555555555666666 

Number of Days on Study 8563379924557788999012223 

1245663772233718016502451

4454443444534444443544444 

Carcass ID Number 4904939758098526359124193 

5544445553242432332523431

Respiratory System 

Lung + + + + + + + + + + + + M + + + + + + + + + + + + 

Alveolar/bronchiolar adenoma X 

Hepatocellular carcinoma, metastatic, 

liver X X X X X X X X X 

Sarcoma, metastatic, kidney X 

Mediastinum, alveolar/bronchiolar 

carcinoma X 

Nose + + + + + + + + + + + + + + + + + + + + + + + + +

Trachea + + + + + + + + + + + + + + + + + + + + + + + + +

Special Senses System 

Ear 

Eye M 

Zymbal's gland + 

Carcinoma X 

Squamous cell carcinoma 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 

Renal tubule, adenoma X 

Renal tubule, adenoma, multiple X 

Renal tubule, carcinoma X 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 

Transitional epithelium, carcinoma 

Transitional epithelium, papilloma X X X 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Leukemia mononuclear X 

Mesothelioma malignant X 
Lesions in Male Rats 125

TABLE A2

Individual Animal Tumor Pathology of Male Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

6666666777777777777777777

Number of Days on Study 3477778000112222222222222

1614499239451449999999999

5 5 5 5 5 4 4 5 4 3 4 4 4 4 4 4 4 4 4 4 4 4 5 5 5 Total

Carcass ID Number 2 0 2 1 2 0 7 1 9 9 5 7 2 7 8 0 1 1 4 4 6 9 1 1 2 Tissues/

4 1 3 4 2 4 4 3 2 1 2 3 1 1 1 1 2 3 1 2 1 1 1 2 1 Tumors

Respiratory System 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Alveolar/bronchiolar adenoma X X 3

Hepatocellular carcinoma, metastatic, 

liver X X X X X X X X X X 19 

Sarcoma, metastatic, kidney 1

Mediastinum, alveolar/bronchiolar 

carcinoma 1

Nose + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Special Senses System 

Ear + 1

Eye + + + + + + 6

Zymbal's gland + 2

Carcinoma 1

Squamous cell carcinoma X 1

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Renal tubule, adenoma X X X X X X X X 9

Renal tubule, adenoma, multiple X X X X 5

Renal tubule, carcinoma 1

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Transitional epithelium, carcinoma X X X X 4

Transitional epithelium, papilloma X X X X X 8

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Leukemia mononuclear 1

Mesothelioma malignant 1
126 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A3

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Adrenal Medulla: Benign Pheochromocytoma

Overall ratea 12/50 (24%) 15/40 (38%) 13/58 (22%) 7/50 (14%)

Adjusted rateb 36.6% 45.3% 38.7% 39.0%

Terminal ratec 7/26 (27%) 7/24 (29%) 4/20 (20%) 2/10 (20%)

First incidence (days) 574 629 521 590

Life table testd P=0.496 P=0.257 P=0.407 P=0.475 

Logistic regression testd P=0.130N P=0.160 P=0.536N P=0.357N

Cochran-Armitage testd P=0.041N

Fisher exact testd P=0.124 P=0.512N P=0.154N

Adrenal Medulla: Malignant Pheochromocytoma

Overall rate 1/50 (2%) 2/40 (5%) 1/58 (2%) 0/50 (0%)

Adjusted rate 3.1% 6.9% 3.2% 0.0%

Terminal rate 0/26 (0%) 1/24 (4%) 0/20 (0%) 0/10 (0%)

First incidence (days) 692 653 693 )e

Life table test P=0.359N P=0.470 P=0.757 P=0.615N

Logistic regression test P=0.250N P=0.429 P=0.738N P=0.557N

Cochran-Armitage test P=0.207N

Fisher exact test P=0.416 P=0.714N P=0.500N

Adrenal Medulla: Benign or Malignant Pheochromocytoma

Overall rate 13/50 (26%) 17/40 (43%) 14/58 (24%) 7/50 (14%)

Adjusted rate 38.6% 50.0% 40.7% 39.0%

Terminal rate 7/26 (27%) 8/24 (33%) 4/20 (20%) 2/10 (20%)

First incidence (days) 574 629 521 590

Life table test P=0.504N P=0.200 P=0.409 P=0.547 

Logistic regression test P=0.075N P=0.231 P=0.525N P=0.277N

Cochran-Armitage test P=0.020N

Fisher exact test P=0.077 P=0.499N P=0.105N

Kidney (Renal Tubule): Adenoma

Overall rate 2/50 (4%) 10/40 (25%) 11/59 (19%) 14/50 (28%)

Adjusted rate 7.7% 33.6% 34.6% 68.3%

Terminal rate 2/26 (8%) 6/24 (25%) 4/21 (19%) 5/10 (50%)

First incidence (days) 729 (T) 618 636 588

Life table test P<0.001 P=0.012 P=0.007 P<0.001

Logistic regression test P<0.001 P=0.007 P=0.014 P<0.001

Cochran-Armitage test P=0.008

Fisher exact test P=0.004 P=0.017 P<0.001

Kidney (Renal Tubule): Adenoma or Carcinoma

Overall rate 2/50 (4%) 10/40 (25%) 13/59 (22%) 15/50 (30%)

Adjusted rate 7.7% 33.6% 39.4% 69.1%

Terminal rate 2/26 (8%) 6/24 (25%) 4/21 (19%) 5/10 (50%)

First incidence (days) 729 (T) 618 636 497

Life table test P<0.001 P=0.012 P=0.002 P<0.001

Logistic regression test P<0.001 P=0.007 P=0.005 P<0.001

Cochran-Armitage test P=0.004

Fisher exact test P=0.004 P=0.006 P<0.001
Lesions in Male Rats 127

TABLE A3

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Large Intestine (Colon): Adenomatous Polyp

Overall rate 0/50 (0%) 1/40 (3%) 1/59 (2%) 3/50 (6%)

Adjusted rate 0.0% 4.2% 4.3% 19.9%

Terminal rate 0/26 (0%) 1/24 (4%) 0/21 (0%) 1/10 (10%)

First incidence (days) ) 729 (T) 720 590

Life table test P=0.009 P=0.484 P=0.454 P=0.037

Logistic regression test P=0.027 P=0.484 P=0.494 P=0.081 

Cochran-Armitage test P=0.065 

Fisher exact test P=0.444 P=0.541 P=0.121 

Large Intestine (Colon): Carcinoma

Overall rate 0/50 (0%) 0/40 (0%) 1/59 (2%) 4/50 (8%)

Adjusted rate 0.0% 0.0% 4.8% 20.4%

Terminal rate 0/26 (0%) 0/24 (0%) 1/21 (5%) 0/10 (0%)

First incidence (days) ) ) 729 (T) 590

Life table test P<0.001 ) P=0.457 P=0.018

Logistic regression test P=0.003 ) P=0.457 P=0.046

Cochran-Armitage test P=0.007

Fisher exact test ) P=0.541 P=0.059 

Large Intestine (Rectum): Adenomatous Polyp

Overall rate 0/50 (0%) 13/40 (33%) 51/59 (86%) 40/50 (80%)

Adjusted rate 0.0% 45.8% 100.0% 100.0%

Terminal rate 0/26 (0%) 9/24 (38%) 21/21 (100%) 10/10 (100%)

First incidence (days) ) 659 478 352

Life table test P<0.001 P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001 P<0.001

Large Intestine (Rectum): Carcinoma

Overall rate 0/50 (0%) 1/40 (3%) 10/59 (17%) 15/50 (30%)

Adjusted rate 0.0% 3.8% 32.4% 63.0%

Terminal rate 0/26 (0%) 0/24 (0%) 5/21 (24%) 4/10 (40%)

First incidence (days) ) 718 608 493

Life table test P<0.001 P=0.478 P=0.001 P<0.001

Logistic regression test P<0.001 P=0.480 P=0.003 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P=0.444 P=0.001 P<0.001

Large Intestine (All Sites): Adenomatous Polyp

Overall rate 0/50 (0%) 13/40 (33%) 51/59 (86%) 40/50 (80%)

Adjusted rate 0.0% 45.8% 100.0% 100.0%

Terminal rate 0/26 (0%) 9/24 (38%) 21/21 (100%) 10/10 (100%)

First incidence (days) ) 659 478 352

Life table test P<0.001 P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001 P<0.001
128 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A3

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Large Intestine (All Sites): Carcinoma

Overall rate 0/50 (0%) 1/40 (3%) 11/59 (19%) 17/50 (34%)

Adjusted rate 0.0% 3.8% 36.6% 67.1%

Terminal rate 0/26 (0%) 0/24 (0%) 6/21 (29%) 4/10 (40%)

First incidence (days) ) 718 608 493

Life table test P<0.001 P=0.478 P<0.001 P<0.001

Logistic regression test P<0.001 P=0.480 P=0.002 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P=0.444 P<0.001 P<0.001

Liver: Hepatocellular Adenoma

Overall rate 1/50 (2%) 20/40 (50%) 40/59 (68%) 34/50 (68%)

Adjusted rate 3.8% 71.3% 92.3% 97.0%

Terminal rate 1/26 (4%) 16/24 (67%) 18/21 (86%) 9/10 (90%)

First incidence (days) 729 (T) 675 521 435

Life table test P<0.001 P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001 P<0.001

Liver: Hepatocellular Carcinoma

Overall rate 1/50 (2%) 12/40 (30%) 55/59 (93%) 46/50 (92%)

Adjusted rate 2.7% 43.5% 100.0% 100.0%

Terminal rate 0/26 (0%) 9/24 (38%) 21/21 (100%) 10/10 (100%)

First incidence (days) 666 650 465 436

Life table test P<0.001 P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001 P<0.001

Liver: Hepatocellular Adenoma or Carcinoma

Overall rate 2/50 (4%) 25/40 (63%) 57/59 (97%) 47/50 (94%)

Adjusted rate 6.4% 83.1% 100.0% 100.0%

Terminal rate 1/26 (4%) 19/24 (79%) 21/21 (100%) 10/10 (100%)

First incidence (days) 666 650 465 435

Life table test P<0.001 P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001 P<0.001

Liver: Hepatocholangiocarcinoma

Overall rate 0/50 (0%) 0/40 (0%) 6/59 (10%) 2/50 (4%)

Adjusted rate 0.0% 0.0% 19.1% 12.1%

Terminal rate 0/26 (0%) 0/24 (0%) 2/21 (10%) 1/10 (10%)

First incidence (days) ) ) 563 527

Life table test P=0.025 ) P=0.019 P=0.133 

Logistic regression test P=0.110 ) P=0.029 P=0.250 

Cochran-Armitage test P=0.117 

Fisher exact test ) P=0.022 P=0.247 
Lesions in Male Rats 129

TABLE A3

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Lung: Alveolar/bronchiolar Adenoma

Overall rate 0/50 (0%) 2/40 (5%) 2/59 (3%) 3/49 (6%)

Adjusted rate 0.0% 8.3% 7.2% 15.5%

Terminal rate 0/26 (0%) 2/24 (8%) 1/21 (5%) 1/10 (10%)

First incidence (days) ) 729 (T) 653 527

Life table test P=0.024 P=0.220 P=0.219 P=0.052 

Logistic regression test P=0.103 P=0.220 P=0.269 P=0.124 

Cochran-Armitage test P=0.130 

Fisher exact test P=0.195 P=0.291 P=0.117 

Lung: Alveolar/bronchiolar Adenoma or Carcinoma

Overall rate 0/50 (0%) 3/40 (8%) 4/59 (7%) 4/49 (8%)

Adjusted rate 0.0% 12.0% 16.5% 17.3%

Terminal rate 0/26 (0%) 2/24 (8%) 3/21 (14%) 1/10 (10%)

First incidence (days) ) 720 653 435

Life table test P=0.010 P=0.105 P=0.046 P=0.028

Logistic regression test P=0.089 P=0.099 P=0.068 P=0.094 

Cochran-Armitage test P=0.100 

Fisher exact test P=0.084 P=0.082 P=0.056 

Pancreatic Islets: Adenoma

Overall rate 2/50 (4%) 2/40 (5%) 0/58 (0%) 1/50 (2%)

Adjusted rate 7.7% 7.4% 0.0% 10.0%

Terminal rate 2/26 (8%) 1/24 (4%) 0/21 (0%) 1/10 (10%)

First incidence (days) 729 (T) 675 ) 729 (T)

Life table test P=0.519N P=0.664 P=0.286N P=0.671 

Logistic regression test P=0.415N P=0.660 P=0.286N P=0.671 

Cochran-Armitage test P=0.255N

Fisher exact test P=0.603 P=0.212N P=0.500N

Pituitary Gland (Pars Distalis): Adenoma

Overall rate 21/48 (44%) 14/40 (35%) 10/56 (18%) 10/49 (20%)

Adjusted rate 56.3% 40.8% 33.0% 51.1%

Terminal rate 10/25 (40%) 6/24 (25%) 4/21 (19%) 4/10 (40%)

First incidence (days) 549 604 608 577

Life table test P=0.298N P=0.190N P=0.041N P=0.430N

Logistic regression test P=0.006N P=0.155N P=0.005N P=0.035N

Cochran-Armitage test P=0.004N

Fisher exact test P=0.269N P=0.004N P=0.012N

Pituitary Gland (Pars Distalis): Adenoma or Carcinoma

Overall rate 21/48 (44%) 15/40 (38%) 10/56 (18%) 10/49 (20%)

Adjusted rate 56.3% 42.3% 33.0% 51.1%

Terminal rate 10/25 (40%) 6/24 (25%) 4/21 (19%) 4/10 (40%)

First incidence (days) 549 604 608 577

Life table test P=0.273N P=0.248N P=0.041N P=0.430N

Logistic regression test P=0.004N P=0.222N P=0.005N P=0.035N

Cochran-Armitage test P=0.003N

Fisher exact test P=0.354N P=0.004N P=0.012N
130 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A3

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Preputial Gland: Adenoma

Overall rate 3/49 (6%) 2/39 (5%) 1/58 (2%) 1/47 (2%)

Adjusted rate 11.0% 7.2% 2.4% 2.5%

Terminal rate 2/25 (8%) 1/23 (4%) 0/21 (0%) 0/9 (0%)

First incidence (days) 696 659 650 552

Life table test P=0.368N P=0.543N P=0.332N P=0.557N

Logistic regression test P=0.198N P=0.554N P=0.264N P=0.384N

Cochran-Armitage test P=0.179N

Fisher exact test P=0.608N P=0.248N P=0.324N

Preputial Gland: Adenoma or Carcinoma

Overall rate 3/49 (6%) 2/39 (5%) 2/58 (3%) 1/47 (2%)

Adjusted rate 11.0% 7.2% 5.6% 2.5%

Terminal rate 2/25 (8%) 1/23 (4%) 0/21 (0%) 0/9 (0%)

First incidence (days) 696 659 650 552

Life table test P=0.432N P=0.543N P=0.525N P=0.557N

Logistic regression test P=0.247N P=0.554N P=0.448N P=0.384N

Cochran-Armitage test P=0.212N

Fisher exact test P=0.608N P=0.420N P=0.324N

Skin: Squamous Cell Papilloma

Overall rate 1/50 (2%) 2/40 (5%) 5/59 (8%) 2/50 (4%)

Adjusted rate 3.8% 7.7% 18.9% 10.5%

Terminal rate 1/26 (4%) 1/24 (4%) 2/21 (10%) 0/10 (0%)

First incidence (days) 729 (T) 700 693 674

Life table test P=0.117 P=0.469 P=0.079 P=0.278 

Logistic regression test P=0.351 P=0.466 P=0.100 P=0.366 

Cochran-Armitage test P=0.413 

Fisher exact test P=0.416 P=0.146 P=0.500 

Skin: Trichoepithelioma, Basal Cell Adenoma, or Basal Cell Carcinoma

Overall rate 3/50 (6%) 0/40 (0%) 1/59 (2%) 1/50 (2%)

Adjusted rate 11.5% 0.0% 4.3% 5.0%

Terminal rate 3/26 (12%) 0/24 (0%) 0/21 (0%) 0/10 (0%)

First incidence (days) 729 (T) ) 720 679

Life table test P=0.564N P=0.134N P=0.386N P=0.627N

Logistic regression test P=0.462N P=0.134N P=0.351N P=0.501N

Cochran-Armitage test P=0.294N

Fisher exact test P=0.167N P=0.249N P=0.309N

Skin: Squamous Cell Papilloma, Keratoacanthoma, Trichoepithelioma, Basal Cell Adenoma, or Basal Cell Carcinoma

Overall rate 5/50 (10%) 3/40 (8%) 6/59 (10%) 4/50 (8%)

Adjusted rate 18.0% 10.1% 22.5% 17.0%

Terminal rate 4/26 (15%) 1/24 (4%) 2/21 (10%) 0/10 (0%)

First incidence (days) 692 629 693 527

Life table test P=0.205 P=0.403N P=0.394 P=0.377 

Logistic regression test P=0.448 P=0.421N P=0.494 P=0.629 

Cochran-Armitage test P=0.483N

Fisher exact test P=0.488N P=0.616 P=0.500N
Lesions in Male Rats 131

TABLE A3

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Skin (Subcutaneous Tissue): Fibroma

Overall rate 2/50 (4%) 0/40 (0%) 3/59 (5%) 1/50 (2%)

Adjusted rate 6.3% 0.0% 12.9% 3.8%

Terminal rate 1/26 (4%) 0/24 (0%) 1/21 (5%) 0/10 (0%)

First incidence (days) 646 ) 715 631

Life table test P=0.389 P=0.258N P=0.437 P=0.707N

Logistic regression test P=0.568 P=0.283N P=0.529 P=0.543N

Cochran-Armitage test P=0.549N

Fisher exact test P=0.306N P=0.579 P=0.500N

Skin (Subcutaneous Tissue): Sarcoma

Overall rate 1/50 (2%) 3/40 (8%) 1/59 (2%) 0/50 (0%)

Adjusted rate 2.3% 11.1% 3.0% 0.0%

Terminal rate 0/26 (0%) 1/24 (4%) 0/21 (0%) 0/10 (0%)

First incidence (days) 562 675 685 )

Life table test P=0.283N P=0.272 P=0.732N P=0.529N

Logistic regression test P=0.156N P=0.187 P=0.741N P=0.416N

Cochran-Armitage test P=0.152N

Fisher exact test P=0.229 P=0.709N P=0.500N

Skin (Subcutaneous Tissue): Fibrosarcoma or Sarcoma

Overall rate 2/50 (4%) 3/40 (8%) 2/59 (3%) 0/50 (0%)

Adjusted rate 4.7% 11.1% 4.7% 0.0%

Terminal rate 0/26 (0%) 1/24 (4%) 0/21 (0%) 0/10 (0%)

First incidence (days) 562 675 465 )

Life table test P=0.207N P=0.456 P=0.638N P=0.304N

Logistic regression test P=0.062N P=0.321 P=0.703 P=0.175N

Cochran-Armitage test P=0.103N

Fisher exact test P=0.394 P=0.626N P=0.247N

Skin (Subcutaneous Tissue): Fibroma, Fibrosarcoma, or Sarcoma

Overall rate 4/50 (8%) 3/40 (8%) 5/59 (8%) 1/50 (2%)

Adjusted rate 10.7% 11.1% 17.0% 3.8%

Terminal rate 1/26 (4%) 1/24 (4%) 1/21 (5%) 0/10 (0%)

First incidence (days) 562 675 465 631

Life table test P=0.412N P=0.550N P=0.493 P=0.351N

Logistic regression test P=0.136N P=0.637 P=0.601 P=0.160N

Cochran-Armitage test P=0.148N

Fisher exact test P=0.624N P=0.605 P=0.181N

Stomach (Forestomach): Squamous Cell Papilloma

Overall rate 0/50 (0%) 2/40 (5%) 0/59 (0%) 1/50 (2%)

Adjusted rate 0.0% 7.7% 0.0% 5.6%

Terminal rate 0/26 (0%) 1/24 (4%) 0/21 (0%) 0/10 (0%)

First incidence (days) ) 700 ) 702

Life table test P=0.427 P=0.220 ) P=0.398 

Logistic regression test P=0.487 P=0.215 ) P=0.426 

Cochran-Armitage test P=0.604 

Fisher exact test P=0.195 ) P=0.500 
132 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A3

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Stomach (Forestomach): Squamous Cell Papilloma or Squamous Cell Carcinoma

Overall rate 0/50 (0%) 2/40 (5%) 0/59 (0%) 2/50 (4%)

Adjusted rate 0.0% 7.7% 0.0% 15.0%

Terminal rate 0/26 (0%) 1/24 (4%) 0/21 (0%) 1/10 (10%)

First incidence (days) ) 700 ) 702

Life table test P=0.117 P=0.220 ) P=0.099 

Logistic regression test P=0.157 P=0.215 ) P=0.130 

Cochran-Armitage test P=0.299 

Fisher exact test P=0.195 ) P=0.247 

Testes: Adenoma

Overall rate 43/50 (86%) 37/40 (93%) 55/59 (93%) 42/50 (84%)

Adjusted rate 97.7% 100.0% 100.0% 100.0%

Terminal rate 25/26 (96%) 24/24 (100%) 21/21 (100%) 10/10 (100%)

First incidence (days) 421 604 521 476

Life table test P<0.001 P=0.404N P=0.022 P<0.001

Logistic regression test P=0.068 P=0.471N P=0.235 P=0.150 

Cochran-Armitage test P=0.341N

Fisher exact test P=0.265 P=0.177 P=0.500N

Thyroid Gland (C-cell): Adenoma

Overall rate 9/49 (18%) 5/40 (13%) 3/59 (5%) 3/50 (6%)

Adjusted rate 26.8% 20.8% 11.6% 23.5%

Terminal rate 4/25 (16%) 5/24 (21%) 1/21 (5%) 2/10 (20%)

First incidence (days) 427 729 (T) 702 671

Life table test P=0.231N P=0.243N P=0.088N P=0.348N

Logistic regression test P=0.052N P=0.332N P=0.030N P=0.085N

Cochran-Armitage test P=0.026N

Fisher exact test P=0.324N P=0.030N P=0.056N

Thyroid Gland (Follicular Cell): Carcinoma

Overall rate 2/49 (4%) 3/40 (8%) 1/59 (2%) 3/50 (6%)

Adjusted rate 8.0% 12.5% 2.1% 18.7%

Terminal rate 2/25 (8%) 3/24 (13%) 0/21 (0%) 1/10 (10%)

First incidence (days) 729 (T) 729 (T) 623 577

Life table test P=0.201 P=0.481 P=0.519N P=0.205 

Logistic regression test P=0.396 P=0.481 P=0.445N P=0.369 

Cochran-Armitage test P=0.546 

Fisher exact test P=0.404 P=0.430N P=0.510 

Thyroid Gland (Follicular Cell): Adenoma or Carcinoma

Overall rate 2/49 (4%) 3/40 (8%) 3/59 (5%) 4/50 (8%)

Adjusted rate 8.0% 12.5% 8.8% 27.7%

Terminal rate 2/25 (8%) 3/24 (13%) 1/21 (5%) 2/10 (20%)

First incidence (days) 729 (T) 729 (T) 623 577

Life table test P=0.051 P=0.481 P=0.482 P=0.079 

Logistic regression test P=0.178 P=0.481 P=0.576 P=0.192 

Cochran-Armitage test P=0.332 

Fisher exact test P=0.404 P=0.588 P=0.349 
Lesions in Male Rats 133

TABLE A3

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Urinary Bladder: Papilloma

Overall rate 0/50 (0%) 1/38 (3%) 2/58 (3%) 8/50 (16%)

Adjusted rate 0.0% 3.7% 9.5% 40.3%

Terminal rate 0/26 (0%) 0/22 (0%) 2/21 (10%) 2/10 (20%)

First incidence (days) ) 700 729 (T) 493

Life table test P<0.001 P=0.479 P=0.192 P<0.001

Logistic regression test P<0.001 P=0.459 P=0.192 P=0.004

Cochran-Armitage test P<0.001

Fisher exact test P=0.432 P=0.286 P=0.003

Urinary Bladder: Carcinoma

Overall rate 0/50 (0%) 0/38 (0%) 1/58 (2%) 4/50 (8%)

Adjusted rate 0.0% 0.0% 4.3% 24.5%

Terminal rate 0/26 (0%) 0/22 (0%) 0/21 (0%) 1/10 (10%)

First incidence (days) ) ) 720 674

Life table test P<0.001 ) P=0.454 P=0.012

Logistic regression test P=0.001 ) P=0.491 P=0.022

Cochran-Armitage test P=0.007

Fisher exact test ) P=0.537 P=0.059 

Urinary Bladder: Papilloma or Carcinoma

Overall rate 0/50 (0%) 1/38 (3%) 3/58 (5%) 12/50 (24%)

Adjusted rate 0.0% 3.7% 13.5% 56.2%

Terminal rate 0/26 (0%) 0/22 (0%) 2/21 (10%) 3/10 (30%)

First incidence (days) ) 700 720 493

Life table test P<0.001 P=0.479 P=0.087 P<0.001

Logistic regression test P<0.001 P=0.459 P=0.096 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P=0.432 P=0.151 P<0.001 

All Organs: Mononuclear Cell Leukemia

Overall rate 25/50 (50%) 5/40 (13%) 3/59 (5%) 1/50 (2%)

Adjusted rate 59.0% 18.8% 11.7% 2.9%

Terminal rate 9/26 (35%) 4/24 (17%) 2/21 (10%) 0/10 (0%)

First incidence (days) 514 604 650 590

Life table test P<0.001N P<0.001N P<0.001N P<0.001N

Logistic regression test P<0.001N P<0.001N P<0.001N P<0.001N

Cochran-Armitage test P<0.001N

Fisher exact test P<0.001N P<0.001N P<0.001N

All Organs: Benign Neoplasms

Overall rate 49/50 (98%) 40/40 (100%) 58/59 (98%) 47/50 (94%)

Adjusted rate 100.0% 100.0% 100.0% 100.0%

Terminal rate 26/26 (100%) 24/24 (100%) 21/21 (100%) 10/10 (100%)

First incidence (days) 421 604 478 352

Life table test P<0.001 P=0.263N P=0.074 P<0.001

Logistic regression test P=0.543 )f P=0.486N P=0.727 

Cochran-Armitage test P=0.102N

Fisher exact test P=0.556 P=0.709 P=0.309N
134 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A3

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

All Organs: Malignant Neoplasms

Overall rate 29/50 (58%) 24/40 (60%) 57/59 (97%) 47/50 (94%)

Adjusted rate 67.0% 71.7% 100.0% 100.0%

Terminal rate 12/26 (46%) 15/24 (63%) 21/21 (100%) 10/10 (100%)

First incidence (days) 514 604 465 435

Life table test P<0.001 P=0.398N P<0.001 P<0.001

Logistic regression test P<0.001 P=0.583 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P=0.510 P<0.001 P<0.001

All Organs: Benign or Malignant Neoplasms

Overall rate 49/50 (98%) 40/40 (100%) 59/59 (100%) 48/50 (96%)

Adjusted rate 100.0% 100.0% 100.0% 100.0%

Terminal rate 26/26 (100%) 24/24 (100%) 21/21 (100%) 10/10 (100%)

First incidence (days) 421 604 465 352

Life table test P<0.001 P=0.263N P=0.061 P<0.001

Logistic regression test P=0.296 ) P=0.943 P=0.501 

Cochran-Armitage test P=0.252N

Fisher exact test P=0.556 P=0.459 P=0.500N

(T)Terminal sacrifice

a Number of lesion-bearing animals/number of animals examined. Denominator is number of animals examined microscopically for adrenal gland, kidney, liver, lung,

pancreatic islets, pituitary gland, preputial gland, stomach, testes, thyroid gland, and urinary bladder; for other tissues, denominator is number of animals necropsied. b Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality c Observed incidence at terminal kill d Beneath the control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values corresponding to pairwise comparisons

between the controls and that exposed group. The life table analysis regards neoplasms in animals dying prior to terminal kill as being (directly or indirectly) the cause of

death. The logistic regression test regards these lesions as nonfatal. The Cochran-Armitage and Fisher exact tests compare directly the overall incidence rates. For all tests,

a negative trend or a lower incidence in an exposure group is indicated by N. e Not applicable; no neoplasms in animal group f Value of statistic cannot be computed.
Lesions in Male Rats 135

TABLE A4a

Historical Incidence of Hepatocellular Neoplasms in Untreated Male F344/N Ratsa

Incidence in Controls

Study Adenoma Carcinoma Adenoma or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 1/50 1/50 2/50

Acetaminophen 0/50 0/50 0/50

HC Yellow 4 1/50 0/50 1/50

Methylphenidate Hydrochloride 0/50 0/50 0/50

Pentaerythritol Tetranitrate 2/49 1/49 3/49

Quercetin 3/50 0/50 3/50

Turmeric Oleoresin 0/50 1/50 1/50

Overall Historical Incidence

Total 35/1,350 (2.6%) 14/1,350 (1.0%) 45/1,350 (3.3%)

Standard deviation 3.2% 1.8% 3.6%

Range 0%-10% 0%-6% 0%-10%

a Data as of 31 March 1993

TABLE A4b

Historical Incidence of Large Intestine Neoplasms in Untreated Male F344/N Ratsa

Incidence in Controls

Study Adenomatous Polyp Carcinoma Adenomatous Polyp

(Adenoma) (Adenoma) or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 0/50 0/50 0/50

Acetaminophen 0/50 0/50 0/50

HC Yellow 4 0/50 0/50 0/50

Methylphenidate Hydrochloride 0/50 0/50 0/50

Pentaerythritol Tetranitrate 0/50 0/50 0/50

Quercetin 0/50 0/50 0/50

Turmeric Oleoresin 0/50 1/50 1/50

Overall Historical Incidence

Total 0/1,353 (0.0%) 1/1,353 (0.1%) 1/1,353 (0.1%)

Standard deviation 0.4% 0.4%

Range 0%-2% 0%-2%

a Data as of 31 March 1993; the data include incidences for the colon and rectum.
136 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A4c

Historical Incidence of Renal Tubule Neoplasms in Untreated Male F344/N Ratsa

Incidence in Controls

Study Adenoma Carcinoma Adenoma or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 2/50 0/50 2/50

Acetaminophen 3/50 0/50 3/50

HC Yellow 4 1/50 0/50 1/50

Methylphenidate Hydrochloride 0/49 0/49 0/49

Pentaerythritol Tetranitrate 0/49 0/49 0/49

Quercetin 0/50 0/50 0/50

Turmeric Oleoresin 0/50 0/50 0/50

Overall Historical Incidence

Total 9/1,350 (0.7%) 6/1,350 (0.4%) 15/1,350 (1.1%)

Standard deviation 1.5% 1.0% 1.7%

Range 0%-6% 0%-4% 0%-6%

a Data as of 31 March 1993

TABLE A4d

Historical Incidence of Urinary Bladder Neoplasms in Untreated Male F344/N Ratsa

Incidence in Controls

Study Papilloma Carcinoma Papilloma or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 0/50 0/50 0/50

Acetaminophen 0/49 0/49 0/49

HC Yellow 4 0/48 0/48 0/48

Methylphenidate Hydrochloride 0/43 0/43 0/43

Pentaerythritol Tetranitrate 0/46 0/46 0/46

Quercetin 0/50 0/50 0/50

Turmeric Oleoresin 0/49 0/49 0/49

Overall Historical Incidence

Total 3/1,329 (0.2%) 0/1,329 (0.0%) 3/1,329 (0.2%)

Standard deviation 0.6% 0.6%

Range 0%-2% 0%-2%

a Data as of 31 March 1993
Lesions in Male Rats 137

TABLE A4e

Historical Incidence of Forestomach Squamous Cell Neoplasms in Untreated Male F344/N Ratsa

Incidence in Controls

Study Papilloma Carcinoma Papilloma or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 0/50 0/50 0/50

Acetaminophen 0/50 0/50 0/50

HC Yellow 4 0/50 1/50 1/50

Methylphenidate Hydrochloride 0/50 0/50 0/50

Pentaerythritol Tetranitrate 0/50 0/50 0/50

Quercetin 0/50 0/50 0/50

Tumeric Oleoresin 0/50 0/50 0/50

Overall Historical Incidence

Total 1/1,353 (0.1%) 3/1,353 (0.2%) 4/1,353 (0.3%)

Standard deviation 0.4% 0.6% 0.7%

Range 0%-2% 0%-2% 0%-2%

a Data as of 31 March 1993
138 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A5

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Disposition Summary

Animals initially in study 70 50 70 70

9-Month interim evaluation 10 10 10 10

15-Month interim evaluation 10 10

Early deaths

Moribund 19 15 34 33

Natural deaths 5 1 5 7

Survivors

Died last week of study 3

Terminal sacrifice 23 24 21 10

Animals examined microscopically 70 50 69b 70

9-Month Interim Evaluation

Alimentary System

Intestine large, colon (10) (10) (10) (10)

Parasite metazoan 2 (20%) 3 (30%) 3 (30%) 2 (20%)

Intestine large, rectum (10) (10) (10) (10)

Parasite metazoan 2 (20%)

Intestine large, cecum (10) (10) (10) (10)

Parasite metazoan 1 (10%)

Lymphoid tissue, hemorrhage 1 (10%)

Liver (10) (10) (10) (10)

Basophilic focus 1 (10%) 1 (10%)

Clear cell focus 4 (40%)

Developmental malformation 1 (10%)

Eosinophilic focus 1 (10%)

Fatty change 3 (30%) 7 (70%) 6 (60%)

Inflammation, chronic 1 (10%)

Inflammation, chronic active 1 (10%) 1 (10%)

Necrosis, coagulative 1 (10%) 2 (20%) 2 (20%)

Pigmentation 1 (10%) 6 (60%)

Bile duct, hyperplasia 1 (10%) 1 (10%) 5 (50%)

Periportal, inflammation 1 (10%)

Periportal, inflammation, chronic active 9 (90%) 10 (100%) 10 (100%) 8 (80%)

Mesentery (1) (1)

Hemorrhage 1 (100%)

Inflammation, chronic, granulomatous 1 (100%)

Inflammation, chronic active 1 (100%)

Necrosis 1 (100%)

Necrosis, coagulative 1 (100%)

Pancreas (10) (10) (10) (10)

Atrophy 2 (20%) 1 (10%) 1 (10%)

Infiltration cellular, mononuclear cell 3 (30%) 5 (50%) 2 (20%) 3 (30%)

Infiltration cellular, mixed cell 1 (10%)

Inflammation, chronic 1 (10%)

Inflammation, chronic active 1 (10%)

a Number of animals examined microscopically at site and number of animals with lesion b One animal not examined microscopically
Lesions in Male Rats 139

TABLE A5

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

9-Month Interim Evaluation (continued)

Alimentary System (continued)

Pancreas (continued) (10) (10) (10) (10)

Necrosis, coagulative 2 (20%)

Pigmentation 1 (10%)

Acinus, atrophy 1 (10%)

Cardiovascular System

Heart (10) (10) (10) (10)

Cardiomyopathy 9 (90%) 8 (80%) 8 (80%) 6 (60%)

Infiltration cellular, mononuclear cell 1 (10%)

Endocrine System

Adrenal cortex (10) (10) (10) (10)

Adrenal medulla (10) (10) (10) (10)

Hyperplasia 1 (10%)

Pituitary gland (9) (10) (9) (9)

Cyst 1 (11%)

Pars distalis, cyst 1 (11%) 1 (11%) 2 (22%)

Pars distalis, hyperplasia 3 (33%) 4 (40%) 1 (11%) 5 (56%)

Pars nervosa, cyst 1 (11%)

Thyroid gland (10) (10) (10) (10)

Infiltration cellular, mononuclear cell 1 (10%)

Ultimobranchial cyst 1 (10%) 1 (10%) 1 (10%)

Genital System

Preputial gland (9) (9) (8) (9)

Inflammation, acute 1 (13%)

Inflammation, chronic 2 (22%) 1 (13%) 2 (22%)

Inflammation, chronic active 7 (78%) 6 (67%) 2 (25%) 2 (22%)

Prostate (10) (10) (9) (10)

Inflammation, acute 1 (10%) 2 (20%)

Inflammation, chronic active 4 (40%) 1 (11%) 1 (10%)

Testes (10) (10) (10) (10)

Abscess, chronic 1 (10%)

Atrophy 1 (10%) 1 (10%)

Infarct 1 (10%) 1 (10%)

Infarct, chronic 1 (10%)

Mineralization 1 (10%)

Interstitial cell, hyperplasia 1 (10%) 9 (90%) 5 (50%) 5 (50%)

Hematopoietic System

Lymph node (2) (1) (6) (2)

Lumbar, pigmentation 1 (17%)

Mediastinal, hemorrhage 1 (50%) 4 (67%) 2 (100%)

Mediastinal, pigmentation 1 (50%) 2 (100%)

Pancreatic, hemorrhage 1 (17%)
140 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A5

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

9-Month Interim Evaluation (continued)

Hematopoietic System (continued)

Lymph node (continued) (2) (1) (6) (2)

Pancreatic, infiltration cellular, histiocyte 1 (17%) 1 (50%)

Pancreatic, inflammation, chronic active 1 (17%)

Pancreatic, pigmentation 1 (50%)

Renal, hemorrhage 1 (50%) 1 (100%)

Renal, pigmentation 1 (50%) 1 (100%)

Lymph node, mandibular (10) (10) (10) (10)

Hemorrhage 9 (90%) 5 (50%) 7 (70%) 7 (70%)

Infiltration cellular, histiocyte 1 (10%)

Lymph node, mesenteric (10) (10) (10) (10)

Hemorrhage 1 (10%)

Infiltration cellular, histiocyte 10 (100%) 10 (100%) 10 (100%) 10 (100%)

Pigmentation 1 (10%) 6 (60%)

Spleen (10) (10) (10) (10)

Depletion lymphoid 1 (10%)

Pigmentation 2 (20%)

Thymus (9) (9) (10) (10)

Depletion lymphoid 3 (33%) 4 (44%) 5 (50%) 7 (70%)

Hemorrhage 1 (11%) 1 (10%) 5 (50%)

Integumentary System

Mammary gland (8) (7) (5) (5)

Hyperplasia 7 (88%) 7 (100%) 5 (100%) 5 (100%)

Respiratory System

Lung (10) (10) (10) (10)

Hemorrhage 1 (10%)

Infiltration cellular, histiocyte 1 (10%) 1 (10%) 2 (20%) 1 (10%)

Alveolar epithelium, hyperplasia 1 (10%)

Artery, mineralization 6 (60%) 3 (30%) 5 (50%) 4 (40%)

Nose (10) (10) (10) (10)

Special Senses System

Ear (1)

Inflammation, chronic active 1 (100%)

Ulcer 1 (100%)

Urinary System

Kidney (10) (10) (10) (10)

Granuloma 1 (10%)

Infiltration cellular, mononuclear cell 9 (90%) 9 (90%) 9 (90%)

Inflammation, chronic 4 (40%)

Renal tubule, degeneration, hyaline 10 (100%) 10 (100%) 10 (100%)

Renal tubule, pigmentation 10 (100%) 10 (100%) 10 (100%)

Renal tubule, regeneration 10 (100%) 10 (100%) 10 (100%) 9 (90%)
Lesions in Male Rats 141

TABLE A5

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

9-Month Interim Evaluation (continued)

Urinary System (continued)

Urinary bladder (10) (10) (9) (10)

Calculus, microscopic observation only 2 (20%) 1 (11%)

Serosa, mineralization 1 (10%)

Systems Examined With No Lesions Observed

General Body System

Musculoskeletal System

Nervous System

15-Month Interim Evaluation

Alimentary System

Intestine large, colon (10) (2)

Parasite metazoan 4 (40%) 2 (100%)

Intestine large, rectum (9) (10)

Parasite metazoan 1 (11%)

Intestine large, cecum (10)

Parasite metazoan 3 (30%)

Intestine small, ileum (10)

Liver (10) (10)

Basophilic focus 1 (10%) 5 (50%)

Clear cell focus 7 (70%)

Cytomegaly 1 (10%)

Degeneration 2 (20%) 1 (10%)

Eosinophilic focus 1 (10%)

Fatty change 6 (60%) 9 (90%)

Hematopoietic cell proliferation 2 (20%)

Inflammation, chronic, granulomatous 2 (20%) 1 (10%)

Inflammation, chronic active 6 (60%)

Mixed cell focus 1 (10%)

Necrosis, coagulative 6 (60%) 2 (20%)

Pigmentation 10 (100%)

Bile duct, hyperplasia 10 (100%) 10 (100%)

Periportal, inflammation, chronic 10 (100%) 10 (100%)

Pancreas (10)

Atrophy 4 (40%)

Inflammation, chronic 8 (80%)

Salivary glands (10)

Stomach, forestomach (10) (10)

Arteriole, mineralization 1 (10%)

Muscularis, mineralization 1 (10%) 1 (10%)

Stomach, glandular (10) (10)

Inflammation, chronic 1 (10%)

Muscularis, mineralization 1 (10%)
142 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A5

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

15-Month Interim Evaluation (continued)

Cardiovascular System

Heart (10)

Cardiomyopathy 10 (100%)

Endocrine System

Adrenal cortex (10)

Hyperplasia 1 (10%)

Bilateral, vacuolization cytoplasmic 1 (10%)

Adrenal medulla (9)

Bilateral, hyperplasia 1 (11%)

Islets, pancreatic (4)

Hemorrhage 1 (25%)

Pituitary gland (8)

Pars distalis, hyperplasia 8 (100%)

Thyroid gland (10) (1)

C-cell, hyperplasia 1 (10%) 1 (100%)

Genital System

Epididymis (10)

Preputial gland (9)

Inflammation, chronic 5 (56%)

Inflammation, chronic active 4 (44%)

Prostate (10)

Inflammation, acute 1 (10%)

Inflammation, chronic active 6 (60%)

Testes (10) (8)

Interstitial cell, hyperplasia 10 (100%) 8 (100%)

Seminiferous tubule, atrophy 2 (20%) 3 (38%)

Hematopoietic System

Lymph node (3)

Mediastinal, pigmentation 1 (33%)

Pancreatic, infiltration cellular, histiocyte 3 (100%)

Pancreatic, pigmentation 3 (100%)

Lymph node, mandibular (10) (1)

Hemorrhage 3 (30%) 1 (100%)

Lymph node, mesenteric (10) (1)

Depletion lymphoid 1 (10%)

Hemorrhage 1 (10%)

Infiltration cellular, histiocyte 10 (100%) 1 (100%)

Pigmentation 9 (90%) 1 (100%)

Thymus (10) (9)

Cyst 1 (10%)

Depletion lymphoid 3 (33%)
Lesions in Male Rats 143

TABLE A5

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

15-Month Interim Evaluation (continued)

Integumentary System

Mammary gland (4)

Hyperplasia 2 (50%)

Skin (10)

Respiratory System

Lung (10) (3)

Hemorrhage 1 (33%)

Infiltration cellular, histiocyte 2 (20%) 2 (67%)

Alveolar epithelium, hyperplasia 3 (30%)

Alveolus, mineralization 1 (10%)

Artery, mineralization 8 (80%) 2 (67%)

Nose (10)

Glands, inflammation, acute 3 (30%)

Glands, inflammation, chronic active 1 (10%)

Nasolacrimal duct, inflammation, chronic 

active 1 (10%)

Urinary System

Kidney (10) (10)

Nephropathy 10 (100%) 10 (100%)

Renal tubule, hyperplasia 2 (20%)

Renal tubule, pigmentation 10 (100%)

Transitional epithelium, hyperplasia 4 (40%)

Urinary bladder (10) (10)

Metaplasia, squamous 1 (10%)

Transitional epithelium, hyperplasia 3 (30%)

Systems Examined With No Lesions Observed

General Body System

Musculoskeletal System

Nervous System

Special Senses System

2-Year Study

Alimentary System

Intestine large, colon (47) (40) (59) (49)

Autolysis 1 (2%) 1 (3%) 4 (7%) 5 (10%)

Hyperplasia, lymphoid 1 (2%)

Parasite metazoan 13 (28%) 14 (35%) 9 (15%) 3 (6%)

Polyarteritis 1 (3%) 1 (2%)

Ulcer 1 (2%)
144 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A5

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Alimentary System (continued)

Intestine large, rectum (46) (40) (58) (49)

Atypia cellular 2 (3%)

Autolysis 1 (2%) 5 (10%)

Inflammation, chronic active 1 (2%)

Parasite metazoan 6 (13%) 6 (15%) 9 (16%) 2 (4%)

Artery, neovascularization 2 (3%)

Artery, thrombosis 1 (2%)

Epithelium, hyperplasia 1 (2%) 1 (2%)

Intestine large, cecum (48) (40) (59) (50)

Atypia cellular 1 (2%)

Autolysis 2 (4%) 1 (3%) 3 (5%) 5 (10%)

Hyperplasia, lymphoid 1 (2%)

Inflammation, chronic active 1 (3%) 1 (2%)

Parasite metazoan 4 (8%) 8 (20%) 10 (17%) 3 (6%)

Ulcer, acute 1 (3%)

Intestine small, duodenum (48) (40) (59) (50)

Autolysis 2 (4%) 1 (3%) 3 (5%) 5 (10%)

Ectopic tissue 1 (2%)

Inflammation, chronic active 1 (2%)

Intestine small, jejunum (48) (38) (57) (48)

Autolysis 2 (4%) 1 (3%) 5 (9%) 6 (13%)

Intestine small, ileum (48) (40) (57) (49)

Autolysis 3 (6%) 1 (3%) 4 (7%) 9 (18%)

Hyperplasia, lymphoid 1 (2%) 2 (4%)

Liver (50) (40) (59) (50)

Angiectasis 3 (6%) 3 (8%)

Autolysis 2 (4%) 2 (4%)

Basophilic focus 9 (18%) 12 (30%) 24 (41%) 22 (44%)

Clear cell focus 3 (6%) 26 (65%) 39 (66%) 27 (54%)

Cyst 1 (2%) 2 (4%)

Degeneration 10 (20%) 3 (8%) 12 (20%) 3 (6%)

Eosinophilic focus 1 (2%) 13 (33%) 14 (24%) 6 (12%)

Fatty change 7 (14%) 4 (10%) 7 (12%) 2 (4%)

Fibrosis 1 (2%)

Hematopoietic cell proliferation 4 (7%)

Hemorrhage 1 (2%) 1 (2%)

Hepatodiaphragmatic nodule 3 (6%) 2 (5%)

Hyperplasia 1 (2%)

Infarct 1 (3%) 1 (2%)

Inflammation, chronic active 3 (6%) 1 (3%) 4 (7%) 6 (12%)

Mixed cell focus 2 (4%) 17 (43%) 8 (14%) 1 (2%)

Necrosis, coagulative 3 (6%) 6 (15%) 8 (14%) 6 (12%)

Pigmentation 3 (6%) 19 (48%) 48 (81%) 39 (78%)

Regeneration 4 (8%)

Bile duct, cyst 1 (3%)

Bile duct, hyperplasia 45 (90%) 9 (23%) 54 (92%) 45 (90%)

Periportal, inflammation, chronic active 2 (4%) 2 (5%) 51 (86%) 36 (72%)
Lesions in Male Rats 145

TABLE A5

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Alimentary System (continued)

Mesentery (3) (2) (4) (4)

Fibrosis 2 (100%) 2 (50%) 2 (50%)

Hemorrhage 1 (25%) 1 (25%)

Inflammation, chronic active 1 (33%) 2 (100%) 3 (75%) 3 (75%)

Necrosis, coagulative 1 (50%) 2 (50%) 1 (25%)

Polyarteritis 1 (50%)

Pancreas (50) (40) (59) (50)

Atrophy 11 (22%) 26 (65%) 29 (49%) 20 (40%)

Autolysis 2 (3%) 2 (4%)

Cytoplasmic alteration 1 (2%)

Ectopic tissue 1 (2%)

Inflammation, chronic active 10 (20%) 28 (70%) 45 (76%) 26 (52%)

Vacuolization cytoplasmic 17 (34%) 2 (5%) 6 (10%) 15 (30%)

Acinus, hyperplasia 1 (2%) 1 (2%)

Artery, fibrosis 1 (2%) 1 (2%)

Artery, polyarteritis 4 (8%) 8 (20%) 4 (7%) 3 (6%)

Duct, hyperplasia 1 (2%) 1 (2%)

Pharynx (1)

Palate, hyperkeratosis 1 (100%)

Salivary glands (50) (40) (58) (49)

Duct, submandibular gland, inflammation,

acute 1 (2%)

Parotid gland, inflammation, chronic 1 (2%)

Submandibular gland, inflammation, chronic 2 (3%)

Submandibular gland, metaplasia, squamous 1 (2%)

Stomach, forestomach (49) (39) (59) (49)

Erosion 1 (2%) 1 (2%)

Fibrosis 1 (2%) 1 (2%)

Hyperkeratosis 5 (10%) 18 (46%) 21 (36%) 20 (41%)

Hyperplasia, basal cell 11 (22%) 18 (46%) 23 (39%) 23 (47%)

Hyperplasia, squamous 3 (6%) 19 (49%) 25 (42%) 26 (53%)

Infarct 1 (2%)

Inflammation, chronic active 3 (6%) 12 (31%) 11 (19%) 11 (22%)

Ulcer 3 (6%) 10 (26%) 15 (25%) 16 (33%)

Stomach, glandular (50) (40) (59) (50)

Autolysis 2 (3%) 2 (4%)

Inflammation, chronic active 3 (6%) 2 (5%) 2 (3%) 3 (6%)

Mucosa, hyperplasia 1 (2%)

Mucosa, necrosis, coagulative 1 (2%)

Cardiovascular System

Heart (50) (40) (59) (49)

Autolysis 1 (3%)

Cardiomyopathy 47 (94%) 38 (95%) 57 (97%) 48 (98%)

Metaplasia, osseous 1 (2%)

Polyarteritis 1 (3%)

Thrombosis 2 (4%) 1 (2%)
146 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A5

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Endocrine System

Adrenal cortex (50) (40) (58) (50)

Angiectasis 2 (4%) 5 (13%) 13 (22%) 15 (30%)

Autolysis 2 (4%)

Hematopoietic cell proliferation 2 (4%) 1 (2%)

Hyperplasia 4 (8%) 1 (3%) 2 (3%) 1 (2%)

Necrosis, coagulative 1 (2%)

Vacuolization cytoplasmic 23 (46%) 18 (45%) 19 (33%) 11 (22%)

Adrenal medulla (50) (40) (58) (50)

Autolysis 2 (4%)

Fibrosis 1 (2%)

Hematopoietic cell proliferation 1 (2%)

Hyperplasia 17 (34%) 25 (63%) 24 (41%) 21 (42%)

Necrosis, coagulative 2 (4%)

Islets, pancreatic (50) (40) (58) (50)

Autolysis 1 (2%) 1 (2%)

Hyperplasia 1 (2%) 2 (4%)

Parathyroid gland (43) (35) (55) (44)

Hyperplasia 4 (9%) 12 (34%) 1 (2%)

Pituitary gland (48) (40) (56) (49)

Autolysis 1 (2%)

Pars distalis, cyst 6 (13%) 4 (10%) 7 (13%) 7 (14%)

Pars distalis, hyperplasia 12 (25%) 11 (28%) 23 (41%) 27 (55%)

Pars intermedia, cyst 2 (4%) 1 (3%) 2 (4%)

Thyroid gland (49) (40) (59) (50)

Autolysis 2 (3%) 2 (4%)

Inflammation, chronic active 2 (4%) 1 (3%) 1 (2%)

Ultimobranchial cyst 2 (5%) 3 (5%) 1 (2%)

C-cell, hyperplasia 10 (20%) 8 (20%) 5 (8%) 3 (6%)

Follicle, cyst 2 (4%) 1 (3%) 7 (12%) 3 (6%)

Follicular cell, hyperplasia 3 (6%) 1 (3%) 2 (3%)

General Body System

None 

Genital System

Coagulating gland (2) (2)

Inflammation, chronic active 1 (50%) 2 (100%)

Epididymis (50) (40) (59) (50)

Aspermia 1 (2%)

Autolysis 2 (3%)

Inflammation, chronic active 3 (6%) 2 (5%) 10 (17%) 3 (6%)

Vacuolization cytoplasmic 1 (2%) 1 (2%)

Preputial gland (49) (39) (58) (47)

Abscess 6 (12%) 1 (3%) 1 (2%)

Cyst 2 (4%) 1 (2%) 1 (2%)

Hyperplasia 2 (3%)

Inflammation, chronic active 46 (94%) 38 (97%) 57 (98%) 43 (91%)
Lesions in Male Rats 147

TABLE A5

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Genital System (continued)

Prostate (50) (40) (59) (49)

Abscess 3 (8%) 1 (2%) 1 (2%)

Fibrosis 1 (2%)

Inflammation, chronic active 11 (22%) 18 (45%) 24 (41%) 23 (47%)

Metaplasia, squamous 1 (2%) 11 (28%) 6 (10%) 1 (2%)

Polyarteritis 1 (3%)

Epithelium, hyperplasia 8 (16%) 4 (10%) 2 (3%) 1 (2%)

Seminal vesicle (49) (40) (59) (50)

Atrophy 1 (2%) 30 (75%) 35 (59%) 23 (46%)

Autolysis 1 (2%)

Inflammation, chronic active 3 (8%) 2 (3%) 1 (2%)

Metaplasia, squamous 1 (3%)

Testes (50) (40) (59) (50)

Infarct 1 (3%)

Inflammation, chronic 1 (2%)

Polyarteritis 1 (2%)

Interstitial cell, hyperplasia 16 (32%) 3 (8%) 27 (46%) 19 (38%)

Seminiferous tubule, atrophy 41 (82%) 37 (93%) 55 (93%) 36 (72%)

Seminiferous tubule, mineralization 8 (16%) 2 (3%) 6 (12%)

Hematopoietic System

Bone marrow (50) (40) (59) (50)

Sternal, autolysis 1 (2%)

Sternal, hypoplasia 1 (2%)

Sternal, myelofibrosis 1 (2%)

Lymph node (17) (5) (12) (20)

Deep cervical, hyperplasia 1 (5%)

Lumbar, hyperplasia 1 (20%) 1 (5%)

Mediastinal, hemorrhage 1 (6%)

Mediastinal, infiltration cellular,

histiocyte 1 (20%) 2 (10%)

Mediastinal, sinus, ectasia 1 (6%)

Pancreatic, hyperplasia 1 (20%) 1 (8%) 2 (10%)

Pancreatic, infiltration cellular, histiocyte 2 (40%) 9 (75%) 19 (95%)

Renal, infiltration cellular, histiocyte 1 (6%) 1 (8%)

Lymph node, mandibular (50) (40) (54) (48)

Hyperplasia 11 (22%) 7 (18%) 7 (13%) 4 (8%)

Infiltration cellular, histiocyte 2 (4%) 1 (2%) 8 (17%)

Inflammation, chronic active 1 (2%)

Lymph node, mesenteric (48) (40) (57) (49)

Autolysis 1 (2%)

Hyperplasia 1 (2%) 2 (4%)

Infiltration cellular, histiocyte 44 (92%) 40 (100%) 56 (98%) 49 (100%)

Pigmentation 1 (2%)
148 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A5

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Hematopoietic System (continued)

Spleen (50) (40) (58) (50)

Autolysis 1 (2%) 2 (4%)

Depletion lymphoid 12 (24%) 2 (5%) 2 (4%)

Fibrosis 6 (12%) 3 (8%) 2 (3%) 5 (10%)

Hyperplasia, RE cell 1 (2%)

Infiltration cellular, histiocyte 1 (2%)

Necrosis, coagulative 2 (4%)

Thymus (37) (32) (41) (34)

Cyst 2 (5%)

Integumentary System

Mammary gland (27) (22) (29) (24)

Galactocele 3 (11%) 1 (5%) 1 (3%)

Hyperplasia 16 (59%) 18 (82%) 17 (59%) 12 (50%)

Skin (50) (38) (58) (50)

Abscess 2 (5%) 1 (2%)

Cyst epithelial inclusion 1 (2%) 3 (5%)

Fibrosis 1 (2%)

Inflammation, chronic active 1 (2%) 2 (3%) 2 (4%)

Musculoskeletal System

Bone (50) (40) (59) (50)

Cranium, abscess 1 (2%)

Joint, tarsal, hyperostosis 1 (2%)

Joint, tarsal, inflammation, chronic active 1 (2%)

Rib, hyperostosis 1 (3%)

Skeletal muscle (2) (1) (1)

Fibrosis 1 (100%)

Polyarteritis 1 (100%)

Nervous System

Brain (50) (40) (59) (50)

Infarct 3 (6%)

Spinal cord (5) (1) (1) (4)

Infarct 1 (20%) 1 (100%)

Respiratory System

Lung (50) (40) (59) (49)

Crystals 1 (2%)

Fibrosis 3 (5%)

Foreign body 1 (2%)

Infiltration cellular, histiocyte 21 (42%) 22 (55%) 32 (54%) 23 (47%)

Inflammation, chronic active 1 (3%) 5 (8%) 3 (6%)

Metaplasia, osseous 1 (2%)

Respiratory epithelium, hyperplasia 2 (4%) 1 (2%) 2 (4%)
Lesions in Male Rats 149

TABLE A5

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Respiratory System (continued)

Lung (continued) (50) (40) (59) (49)

Necrosis, coagulative 1 (2%)

Alveolar epithelium, hyperplasia 4 (8%) 1 (3%) 7 (12%) 1 (2%)

Artery, mineralization 22 (44%) 14 (35%) 36 (61%) 28 (57%)

Artery, thrombosis 1 (2%)

Bronchiole, epithelium, hyperplasia 1 (2%)

Mediastinum, polyarteritis 2 (5%)

Nose (48) (40) (59) (50)

Autolysis 1 (2%)

Foreign body 2 (5%) 7 (12%) 1 (2%)

Hemorrhage 3 (6%)

Inflammation, chronic active 26 (54%) 25 (63%) 27 (46%) 18 (36%)

Metaplasia, squamous 10 (21%) 7 (18%) 7 (12%) 6 (12%)

Ulcer 2 (4%)

Glands, hyperplasia 9 (19%) 3 (6%)

Respiratory epithelium, hyperplasia 2 (4%) 1 (2%) 2 (4%)

Trachea (50) (40) (59) (50)

Autolysis 1 (2%)

Inflammation, chronic active 1 (3%) 3 (5%) 2 (4%)

Special Senses System

Ear (3) (7) (2) (1)

Acanthosis 1 (33%) 1 (14%) 1 (50%)

Hyperkeratosis 1 (33%)

Hyperplasia, basal cell 1 (33%) 1 (14%) 1 (50%)

Inflammation, chronic active 1 (33%)

Submucosa, abscess 1 (100%)

Eye (6) (3) (1) (6)

Cataract 1 (100%)

Synechia 1 (17%)

Anterior, synechia 1 (17%)

Cornea, inflammation, chronic active 1 (17%)

Lens, cataract 3 (50%)

Retina, degeneration 1 (17%) 1 (100%) 3 (50%)

Urinary System

Kidney (50) (40) (59) (50)

Autolysis 2 (4%) 1 (2%) 4 (8%)

Cyst 4 (8%) 3 (8%) 1 (2%)

Cyst, multiple 1 (2%)

Fibrosis, focal 1 (3%)

Hydronephrosis 1 (2%)

Nephropathy 50 (100%) 40 (100%) 59 (100%) 49 (98%)

Collecting tubule, mineralization 1 (2%)

Renal tubule, degeneration, hyaline 1 (2%) 1 (2%)

Renal tubule, hyperplasia 9 (18%) 30 (75%) 25 (42%) 19 (38%)

Renal tubule, hyperplasia, oncocytic 1 (3%) 1 (2%)
150 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE A5

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Urinary System (continued)

Kidney (continued) (50) (40) (59) (50)

Renal tubule, inflammation, chronic active 14 (28%) 10 (25%) 22 (37%) 14 (28%)

Renal tubule, mineralization 1 (2%) 2 (5%) 13 (22%) 4 (8%)

Renal tubule, pigmentation 5 (10%) 40 (100%) 58 (98%) 49 (98%)

Transitional epithelium, hyperplasia 30 (60%) 40 (100%) 51 (86%) 35 (70%)

Urinary bladder (50) (38) (58) (50)

Autolysis 1 (2%) 1 (2%)

Edema 1 (3%)

Fibrosis 1 (3%)

Hemorrhage 2 (4%)

Hyperplasia, lymphoid 4 (8%)

Inflammation, chronic 1 (2%)

Inflammation, chronic active 1 (2%) 3 (8%) 5 (10%)

Polyarteritis 1 (3%)

Transitional epithelium, hyperplasia 1 (2%) 5 (13%) 17 (29%) 30 (60%)

Transitional epithelium, metaplasia, squamous 3 (6%)
151

APPENDIX B

SUMMARY OF LESIONS IN FEMALE RATS

IN THE 2-YEAR FEED STUDY

OF 1-AMINO-2,4-DIBROMOANTHRAQUINONE

TABLE B1 Summary of the Incidence of Neoplasms in Female Rats 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 153

TABLE B2 Individual Animal Tumor Pathology of Female Rats 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 160

TABLE B3 Statistical Analysis of Primary Neoplasms in Female Rats 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 182

TABLE B4a Historical Incidence of Hepatocellular Neoplasms 

in Untreated Female F344/N Rats . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 189

TABLE B4b Historical Incidence of Large Intestine Neoplasms

in Untreated Female F344/N Rats . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 189

TABLE B4c Historical Incidence of Renal Tubule Neoplasms 

in Untreated Female F344/N Rats . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 190

TABLE B4d Historical Incidence of Urinary Bladder Neoplasms

in Untreated Female F344/N Rats . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 190

TABLE B4e Historical Incidence of Forestomach Squamous Cell Neoplasms

in Untreated Female F344/N Rats . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 191

TABLE B5 Summary of the Incidence of Nonneoplastic Lesions in Female Rats 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 192
152 1-Amino-2,4-dibromoanthraquinone, NTP TR 383
Lesions in Female Rats 153

TABLE B1

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Disposition Summary

Animals initially in study 70 50 70 70

9-Month interim evaluation 10 10 10 10

15-Month interim evaluation 10 10

Early deaths

Moribund 8 5 15 29

Natural deaths 4 3 7 8

Survivors

Died last week of study 1

Terminal sacrifice 37 32 38 12

Missexed 1

Animals examined microscopically 70 50 70 69

9-Month Interim Evaluation

Genital System

Uterus (10) (10) (10) (10)

Polyp stromal 1 (10%)

Systems Examined With No Neoplasms Observed

Alimentary System

Cardiovascular System

Endocrine System

General Body System

Hematopoietic System

Integumentary System

Musculoskeletal System

Nervous System

Respiratory System

Special Senses System

Urinary System

15-Month Interim Evaluation

Alimentary System

Intestine large, rectum (10) (10)

Polyp adenomatous 2 (20%)

Liver (10) (10)

Hepatocellular carcinoma 3 (30%)

Hepatocellular carcinoma, multiple 3 (30%)

Hepatocellular adenoma 3 (30%)

Hepatocellular adenoma, multiple 5 (50%)

Endocrine System

Adrenal medulla (10) (1)

Pheochromocytoma benign 1 (100%)

Pituitary gland (10)

Pars distalis, adenoma 2 (20%)
154 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B1

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

15-Month Interim Evaluation (continued)

Genital System

Uterus (10) (3)

Polyp stromal 1 (10%) 1 (33%)

Integumentary System

Mammary gland (7)

Fibroadenoma 1 (14%)

Skin (10) (1)

Subcutaneous tissue, fibroma 1 (100%)

Urinary System

Kidney (10) (10)

Urinary bladder (10) (10)

Carcinoma 2 (20%) 

Papilloma 1(10%)

Squamous cell carcinoma 2 (20%)

Systems Examined With No Neoplasms Observed

Cardiovascular System

General Body System

Hematopoietic System

Musculoskeletal System

Nervous System

Respiratory System

Special Senses System

2-Year Study

Alimentary System

Esophagus (50) (40) (57) (48)

Intestine large, colon (49) (40) (59) (47)

Adenocarcinoma 2 (3%) 1 (2%)

Adenocarcinoma, multiple 1 (3%)

Polyp adenomatous 1 (2%) 1 (2%)

Polyp adenomatous, multiple 1 (3%) 1 (2%) 1 (2%)

Intestine large, rectum (49) (40) (60) (47)

Adenocarcinoma 1 (3%) 18 (30%) 6 (13%)

Adenocarcinoma, multiple 1 (2%) 1 (2%)

Polyp adenomatous 10 (25%) 8 (13%) 12 (26%)

Polyp adenomatous, multiple 17 (43%) 45 (75%) 31 (66%)

Intestine large, cecum (50) (40) (60) (47)

Polyp adenomatous 1 (2%)

Intestine small, jejunum (48) (40) (59) (46)

Intestine small, ileum (49) (39) (59) (44)

Leiomyoma 1 (2%)
Lesions in Female Rats 155

TABLE B1

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Alimentary System (continued)

Liver (50) (40) (60) (48)

Cholangioma 1 (2%)

Hepatocellular carcinoma 5 (13%) 6 (10%) 4 (8%)

Hepatocellular carcinoma, multiple 7 (18%) 51 (85%) 41 (85%)

Hepatocellular adenoma 10 (25%) 8 (13%) 7 (15%)

Hepatocellular adenoma, multiple 18 (45%) 39 (65%) 22 (46%)

Hepatocholangiocarcinoma 10 (17%) 8 (17%)

Hepatocholangiocarcinoma, multiple 1 (2%) 5 (10%)

Hepatocholangioma 1 (2%)

Hepatocholangioma, multiple 1 (2%)

Mesentery (4) (6) (1) (4)

Hepatocellular carcinoma, metastatic, liver 3 (75%)

Pancreas (50) (40) (60) (47)

Adenoma 2 (3%)

Hepatocellular carcinoma, metastatic, liver 1 (2%)

Pharynx (1)

Palate, squamous cell papilloma, multiple 1 (100%)

Stomach, forestomach (49) (40) (60) (47)

Squamous cell carcinoma 1 (3%) 1 (2%) 1 (2%)

Squamous cell papilloma 1 (2%)

Stomach, glandular (50) (40) (60) (48)

Hepatocellular carcinoma, metastatic, liver 1 (2%)

Tongue (1)

Squamous cell papilloma 1 (100%)

Tooth (1) (1) (1)

Gingiva, squamous cell carcinoma 1 (100%)

Cardiovascular System

Heart (50) (40) (60) (49)

Hepatocellular carcinoma, metastatic, liver 1 (2%) 2 (4%)

Endocrine System

Adrenal cortex (47) (40) (59) (47)

Adenoma 2 (4%) 1 (3%)

Carcinoma, metastatic, kidney 1 (2%)

Adrenal medulla (47) (40) (59) (47)

Pheochromocytoma complex 1 (2%)

Pheochromocytoma benign 2 (4%) 3 (8%) 1 (2%) 1 (2%)

Bilateral, pheochromocytoma benign 1 (2%)

Islets, pancreatic (50) (40) (60) (47)

Adenoma 1 (3%)

Hepatocellular carcinoma, metastatic, liver 1 (2%)

Parathyroid gland (43) (37) (57) (38)

Adenoma 1 (2%)

Pituitary gland (50) (39) (60) (47)

Pars distalis, adenoma 27 (54%) 16 (41%) 25 (42%) 13 (28%)

Pars distalis, adenoma, multiple 5 (10%) 3 (8%) 7 (12%)

Pars intermedia, adenoma 1 (3%)
156 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B1

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Endocrine System (continued)

Thyroid gland (50) (40) (60) (49)

C-cell, adenoma 5 (10%) 5 (13%) 5 (8%)

C-cell, adenoma, multiple 1 (2%)

C-cell, carcinoma 1 (2%) 1 (2%)

Follicular cell, adenoma 1 (3%) 1 (2%) 2 (4%)

General Body System

Tissue NOS (2) (1) (1)

Basosquamous tumor malignant 1 (100%)

Neoplasm NOS 1 (100%)

Sarcoma 2 (100%)

Genital System

Clitoral gland (45) (36) (60) (45)

Adenoma 5 (11%) 3 (8%) 3 (5%) 2 (4%)

Ovary (50) (40) (60) (47)

Granulosa cell tumor benign 1 (2%) 1 (3%)

Hepatocellular carcinoma, metastatic, liver 1 (2%) 1 (2%)

Uterus (50) (40) (60) (47)

Adenocarcinoma 1 (2%)

Hemangioma 1 (2%)

Polyp stromal 7 (14%) 12 (30%) 7 (12%) 5 (11%)

Polyp stromal, multiple 1 (2%) 3 (8%)

Sarcoma stromal 1 (2%)

Hematopoietic System

Bone marrow (50) (40) (60) (49)

Lymph node (3) (5) (25) (14)

Mediastinal, hepatocellular carcinoma,

metastatic, liver 1 (7%)

Lymph node, mandibular (50) (39) (56) (45)

Lymph node, mesenteric (50) (40) (59) (46)

Spleen (50) (40) (60) (48)

Hepatocellular carcinoma, metastatic, liver 1 (2%)

Lipoma 1 (3%)

Thymus (42) (31) (51) (38)

Hepatocellular carcinoma, metastatic, liver 1 (3%)

Integumentary System

Mammary gland (49) (34) (50) (41)

Adenocarcinoma 2 (4%)

Fibroadenoma 17 (35%) 8 (24%) 9 (18%) 5 (12%)

Fibroadenoma, multiple 4 (8%) 2 (6%)

Fibroma 1 (2%)
Lesions in Female Rats 157

TABLE B1

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Integumentary System (continued)

Skin (50) (39) (60) (49)

Squamous cell papilloma 1 (2%) 1 (2%) 1 (2%)

Pinna, granular cell tumor benign 1 (2%)

Pinna, squamous cell papilloma 2 (3%) 2 (4%)

Subcutaneous tissue, fibroma 2 (4%) 2 (5%)

Subcutaneous tissue, sarcoma 1 (2%)

Musculoskeletal System

Bone (50) (40) (60) (49)

Basosquamous tumor malignant, metastatic,

tissue NOS 1 (3%)

Squamous cell carcinoma, metastatic,

uncertain primary site 1 (2%)

Skeletal muscle (1)

Rhabdomyosarcoma 1 (100%)

Nervous System

Brain (50) (40) (60) (49)

Oligodendroglioma benign 1 (2%)

Respiratory System

Lung (50) (40) (60) (49)

Alveolar/bronchiolar adenoma 1 (2%) 2 (5%) 1 (2%)

Alveolar/bronchiolar adenoma, multiple 1 (3%)

Alveolar/bronchiolar carcinoma 1 (2%)

Hepatocellular carcinoma, metastatic, liver 1 (3%) 22 (37%) 24 (49%)

Neoplasm NOS, metastatic, uncertain primary

site 1 (2%)

Sarcoma, metastatic, thymus 1 (2%)

Squamous cell carcinoma 1 (3%)

Nose (50) (40) (60) (49)

Special Senses System

Ear (8) (3)

Zymbal's gland (1) (2)

Adenoma 1 (50%)

Carcinoma 1 (50%)

Squamous cell carcinoma 1 (100%)
158 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B1

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Urinary System

Kidney (50) (40) (60) (48)

Squamous cell carcinoma, metastatic,

urinary bladder 1 (2%)

Pelvis, transitional epithelium, carcinoma 1 (3%)

Pelvis, transitional epithelium, papilloma 1 (2%)

Renal tubule, adenoma 3 (8%) 11 (18%) 11 (23%)

Renal tubule, adenoma, multiple 5 (8%) 5 (10%)

Renal tubule, carcinoma 2 (4%)

Urinary bladder (50) (40) (60) (46)

Transitional epithelium, carcinoma 8 (13%) 16 (35%)

Transitional epithelium, papilloma 2 (5%) 6 (10%) 9 (20%)

Transitional epithelium, papilloma, multiple 1 (2%)

Transitional epithelium, squamous cell

carcinoma 1 (2%)

Transitional epithelium, squamous cell

papilloma 1 (2%) 1 (2%)

Transitional epithelium, squamous cell

papilloma, multiple 1 (2%)

Systemic Lesions

Multiple organsb (50) (40) (60) (49)

Leukemia mononuclear 9 (18%) 1 (3%) 5 (8%) 1 (2%)

Lymphoma malignant histiocytic 1 (2%)

Neoplasm Summary

Total animals with primary neoplasmsc

9-Month interim evaluation 1

15-Month interim evaluation 2 9

2-Year study 46 40 60 48

Total primary neoplasms

9-Month interim evaluation 1

15-Month interim evaluation 4 24

2-Year study 100 148 306 228

Total animals with benign neoplasms

9-Month interim evaluation 1

15-Month interim evaluation 2 8

2-Year study 43 40 60 46

Total benign neoplasms

9-Month interim evaluation 1

15-Month interim evaluation 4 14

2-Year study 83 128 197 139

Total animals with malignant neoplasms

15-Month interim evaluation 9

2-Year study 16 15 60 46

Total malignant neoplasms

15-Month interim evaluation 10

2-Year study 17 20 108 89
Lesions in Female Rats 159

TABLE B1

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Neoplasm Summary (continued)

Total animals with metastatic neoplasms

2-Year study 1 2 22 27

Total metastatic neoplasms

2-Year study 1 2 26 38

Total animals with malignant neoplasms

 of uncertain primary site

2-Year study 2

Total animals with uncertain neoplasms -

 benign or malignant

2-Year study 1

Total uncertain neoplasms

2-Year study 1

a Number of animals examined microscopically at site and number of animals with neoplasm b Number of animals with any tissue examined microscopically c Primary neoplasms: all neoplasms except metastatic neoplasms
160 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm

2666666677777777777777777 

Number of Days on Study 6002455700113333333333333 

8160268548691444444444444

6667777777777666667777777 

Carcass ID Number 8687323016843667773445556 

5343433442452121231121231

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine large, colon + + + + + + + + + + M + + + + + + + + + + + + + +

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, jejunum + + + + + + + + M + + + + + + + + + + + + + + + +

Intestine small, ileum + + + + + + + + + + + + A + + + + + + + + + + + +

Liver + + + + + + + + + + + + + + + + + + + + + + + + +

Mesentery + + 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + +

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + +

Stomach, forestomach + + + + + + + + + + M + + + + + + + + + + + + + +

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + +

Tooth + 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + +

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma X 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma benign 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + +

Parathyroid gland + + M + M + + + M + + + + + + + + + + + M + M + + 

Adenoma 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Pars distalis, adenoma X X X X X X X X X X X X X 

Pars distalis, adenoma, multiple 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

C-cell, adenoma X X X 

C-cell, carcinoma X 

General Body System 

Tissue NOS + + 

Sarcoma X X 

Genital System

Clitoral gland + M M + + + + + M + + + + + + + + + + + + + + + + 

Adenoma X X X X 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 

Granulosa cell tumor benign 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenocarcinoma X 

Hemangioma X 

Polyp stromal X X 

Polyp stromal, multiple 

+: Tissue examined microscopically M: Missing tissue X: Lesion present

A: Autolysis precludes examination I: Insufficient tissue Blank: Not examined
Lesions in Female Rats 161

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

7777777777777777777777777

Number of Days on Study 3333333333333333333333333

5555555555566666666666666

6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 Total

Carcass ID Number 8 8 8 9 9 4 4 5 7 9 9 0 0 0 1 1 1 2 2 7 8 8 8 9 9 Tissues/

1 2 3 1 2 3 4 4 2 3 4 1 2 3 1 2 3 1 2 1 1 2 3 1 2 Tumors

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine large, rectum + + + + + + + + + + + + + + + + + M + + + + + + + 49 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine small, jejunum + + M + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Mesentery + + 4

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Tooth 1

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System

Adrenal cortex + + + + + + + M + M + M + + + + + + + + + + + + + 47 

Adenoma X 2

Adrenal medulla + + + + + + + M + M + M + + + + + + + + + + + + + 47 

Pheochromocytoma benign X X 2

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Parathyroid gland + + + + + + + M + + + + + + + + + + + + M + + + + 43 

Adenoma X 1

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pars distalis, adenoma X X X X X X X X X X X X X X 27 

Pars distalis, adenoma, multiple X X X X X 5

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

C-cell, adenoma X X 5

C-cell, carcinoma 1

General Body System 

Tissue NOS 2

Sarcoma 2

Genital System

Clitoral gland + + + + + + + + + + + M + + + + + + + + M + + + + 45 

Adenoma X 5

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Granulosa cell tumor benign X 1

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenocarcinoma 1

Hemangioma 1

Polyp stromal X X X X X 7

Polyp stromal, multiple X 1
162 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

2666666677777777777777777 

Number of Days on Study 6002455700113333333333333 

8160268548691444444444444

6667777777777666667777777 

Carcass ID Number 8687323016843667773445556 

5343433442452121231121231

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node + 

Lymph node, mandibular + + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + +

Spleen + + + + + + + + + + + + + + + + + + + + + + + + +

Thymus + + + + + + M + + + M + + + + I M + + + + + + + M

Integumentary System 

Mammary gland + + + + + + + + + + + + + + + + + + + + M + + + + 

Adenocarcinoma X 

Fibroadenoma X X X X X 

Fibroadenoma, multiple X X X 

Fibroma 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 

Squamous cell papilloma 

Subcutaneous tissue, fibroma 

Subcutaneous tissue, sarcoma X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + +

Nervous System

Brain + + + + + + + + + + + + + + + + + + + + + + + + +

Respiratory System

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 

Alveolar/bronchiolar adenoma X 

Sarcoma, metastatic, thymus X 

Nose + + + + + + + + + + + + + + + + + + + + + + + + +

Trachea + + + + + + + + + + + + + + + + + + + + + + + + +

Special Senses System 

Eye + + + + + + + + + +

Zymbal's gland + 

Squamous cell carcinoma X 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + +

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + +

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Leukemia mononuclear X X X X X 
Lesions in Female Rats 163

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

7777777777777777777777777

Number of Days on Study 3333333333333333333333333

5555555555566666666666666

6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 Total

Carcass ID Number 8 8 8 9 9 4 4 5 7 9 9 0 0 0 1 1 1 2 2 7 8 8 8 9 9 Tissues/

1 2 3 1 2 3 4 4 2 3 4 1 2 3 1 2 3 1 2 1 1 2 3 1 2 Tumors

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymph node + + 3

Lymph node, mandibular + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Thymus + + + + M + + + + + + + + M + + + + M + + + + + + 42 

Integumentary System 

Mammary gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Adenocarcinoma X 2

Fibroadenoma X X X X X X X X X X X X 17 

Fibroadenoma, multiple X 4

Fibroma X 1

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Squamous cell papilloma X 1

Subcutaneous tissue, fibroma X X 2

Subcutaneous tissue, sarcoma 1

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Nervous System

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Respiratory System

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Alveolar/bronchiolar adenoma 1

Sarcoma, metastatic, thymus 1

Nose + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Special Senses System 

Eye + + + 13 

Zymbal's gland 1

Squamous cell carcinoma 1

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Leukemia mononuclear X X X X 9
164 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

2,000 ppm

6666666677777777777777777 

Number of Days on Study 0115667833333333333333333 

0169140900333333333333334

8888888888888888888888888 

Carcass ID Number 0103476433677778888899990 

4435354234312341234512341

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenocarcinoma, multiple 

Polyp adenomatous, multiple X 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenocarcinoma 

Polyp adenomatous X X X X X X 

Polyp adenomatous, multiple X X X X X X X X X X

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + M + +

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 

Hepatocellular carcinoma X 

Hepatocellular carcinoma, multiple X X X X 

Hepatocellular adenoma X X X X X X X 

Hepatocellular adenoma, multiple X X X X X X X X X X X X X

Mesentery + + + + + 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + +

Pharynx + 

Palate, squamous cell papilloma, 

multiple X 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + +

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 

Squamous cell carcinoma 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + +

Tooth + 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + +

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma benign X 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma X 

Parathyroid gland + + + + + + + + + + + + + + M + + + M + + + + + +

Pituitary gland + M + + + + + + + + + + + + + + + + + + + + + + + 

Pars distalis, adenoma X X X X X X X X X X 

Pars distalis, adenoma, multiple X X 

Pars intermedia, adenoma X 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + +

C-cell, adenoma X 

Follicular cell, adenoma X 
Lesions in Female Rats 165

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

2,000 ppm (continued)

777777777777777 

Number of Days on Study 333333333333333 

444444444444444 

8 8 8 8 8 8 8 8 8 8 8 8 8 8 8 Total

Carcass ID Number 0 1 1 1 2 2 2 3 3 4 5 5 5 6 6 Tissues/

2 1 2 3 1 2 3 1 2 1 1 2 3 1 2 Tumors

Alimentary System 

Esophagus + + + + + + + + + + + + + + + 40 

Intestine large, colon + + + + + + + + + + + + + + + 40 

Adenocarcinoma, multiple X 1

Polyp adenomatous, multiple 1

Intestine large, rectum + + + + + + + + + + + + + + + 40 

Adenocarcinoma X 1

Polyp adenomatous X X X X 10 

Polyp adenomatous, multiple X X X X X X X 17 

Intestine large, cecum + + + + + + + + + + + + + + + 40 

Intestine small, duodenum + + + + + + + + + + + + + + + 40 

Intestine small, jejunum + + + + + + + + + + + + + + + 40 

Intestine small, ileum + + + + + + + + + + + + + + + 39 

Liver + + + + + + + + + + + + + + + 40 

Hepatocellular carcinoma X X X X 5

Hepatocellular carcinoma, multiple X X X 7

Hepatocellular adenoma X X X 10 

Hepatocellular adenoma, multiple X X X X X 18 

Mesentery + 6

Pancreas + + + + + + + + + + + + + + + 40 

Pharynx 1

Palate, squamous cell papilloma, 

multiple 1

Salivary glands + + + + + + + + + + + + + + + 40 

Stomach, forestomach + + + + + + + + + + + + + + + 40 

Squamous cell carcinoma X 1

Stomach, glandular + + + + + + + + + + + + + + + 40 

Tooth 1

Cardiovascular System 

Heart + + + + + + + + + + + + + + + 40 

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + 40 

Adenoma X 1

Adrenal medulla + + + + + + + + + + + + + + + 40 

Pheochromocytoma benign X X 3

Islets, pancreatic + + + + + + + + + + + + + + + 40 

Adenoma 1

Parathyroid gland + + + + + + + M + + + + + + + 37 

Pituitary gland + + + + + + + + + + + + + + + 39 

Pars distalis, adenoma X X X X X X 16 

Pars distalis, adenoma, multiple X 3

Pars intermedia, adenoma 1

Thyroid gland + + + + + + + + + + + + + + + 40 

C-cell, adenoma X X X X 5

Follicular cell, adenoma 1
166 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

2,000 ppm (continued)

6666666677777777777777777 

Number of Days on Study 0115667833333333333333333 

0169140900333333333333334

8888888888888888888888888 

Carcass ID Number 0103476433677778888899990 

4435354234312341234512341

General Body System 

Tissue NOS 

Basosquamous tumor malignant 

Genital System

Clitoral gland + + + + + M M + + + + + + + + + + + + + + + + + + 

Adenoma X X 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 

Granulosa cell tumor benign X 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 

Polyp stromal X X X X X X X X X 

Polyp stromal, multiple X X 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node + + 

Lymph node, mandibular M + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + +

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 

Lipoma 

Thymus M + + + M + M + + + M + + + + + + + + + + + + + +

Integumentary System 

Mammary gland + + + + + + + + + + + M M + + + + + M + + + + M + 

Fibroadenoma X X X X X 

Fibroadenoma, multiple X X

Skin + + + + + + + + + + + + M + + + + + + + + + + + + 

Subcutaneous tissue, fibroma X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Basosquamous tumor malignant, 

metastatic, tissue NOS 

Skeletal muscle 

Rhabdomyosarcoma 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + +

Respiratory System

Lung + + + + + + + + + + + + + + + + + + + + + + + + +

Alveolar/bronchiolar adenoma X 

Alveolar/bronchiolar adenoma, 

multiple X 

Hepatocellular carcinoma, metastatic, 

liver X 

Squamous cell carcinoma 

Nose + + + + + + + + + + + + + + + + + + + + + + + + +

Trachea + + + + + + + + + + + + + + + + + + + + + + + + +
Lesions in Female Rats 167

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

2,000 ppm (continued)

777777777777777 

Number of Days on Study 333333333333333 

444444444444444 

8 8 8 8 8 8 8 8 8 8 8 8 8 8 8 Total

Carcass ID Number 0 1 1 1 2 2 2 3 3 4 5 5 5 6 6 Tissues/

2 1 2 3 1 2 3 1 2 1 1 2 3 1 2 Tumors

General Body System 

Tissue NOS + 1

Basosquamous tumor malignant X 1

Genital System

Clitoral gland + + + + + + + + + + M + M + + 36 

Adenoma X 3

Ovary + + + + + + + + + + + + + + + 40 

Granulosa cell tumor benign 1

Uterus + + + + + + + + + + + + + + + 40 

Polyp stromal X X X 12 

Polyp stromal, multiple X 3

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + 40 

Lymph node + + + 5

Lymph node, mandibular + + + + + + + + + + + + + + + 39 

Lymph node, mesenteric + + + + + + + + + + + + + + + 40 

Spleen + + + + + + + + + + + + + + + 40 

Lipoma X 1

Thymus + M + + + M + + + M M + M + + 31 

Integumentary System 

Mammary gland + + M + + + + + + + + M + + + 34 

Fibroadenoma X X X 8

Fibroadenoma, multiple 2

Skin + + + + + + + + + + + + + + + 39 

Subcutaneous tissue, fibroma X 2

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + 40 

Basosquamous tumor malignant, 

metastatic, tissue NOS X 1

Skeletal muscle + 1

Rhabdomyosarcoma X 1

Nervous System 

Brain + + + + + + + + + + + + + + + 40 

Respiratory System

Lung + + + + + + + + + + + + + + + 40 

Alveolar/bronchiolar adenoma X 2

Alveolar/bronchiolar adenoma, 

multiple 1

Hepatocellular carcinoma, metastatic, 

liver 1

Squamous cell carcinoma X 1

Nose + + + + + + + + + + + + + + + 40 

Trachea + + + + + + + + + + + + + + + 40 
168 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

2,000 ppm (continued)

6666666677777777777777777 

Number of Days on Study 0115667833333333333333333 

0169140900333333333333334

8888888888888888888888888 

Carcass ID Number 0103476433677778888899990 

4435354234312341234512341

Special Senses System 

Eye + + + + + + 

Urinary System

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 

Pelvis, transitional epithelium, 

carcinoma 

Renal tubule, adenoma X X X 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 

Transitional epithelium, papilloma X 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Leukemia mononuclear X 
Lesions in Female Rats 169

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

2,000 ppm (continued)

777777777777777 

Number of Days on Study 333333333333333 

444444444444444 

8 8 8 8 8 8 8 8 8 8 8 8 8 8 8 Total

Carcass ID Number 0 1 1 1 2 2 2 3 3 4 5 5 5 6 6 Tissues/

2 1 2 3 1 2 3 1 2 1 1 2 3 1 2 Tumors

Special Senses System 

Eye + 7

Urinary System

Kidney + + + + + + + + + + + + + + + 40 

Pelvis, transitional epithelium, 

carcinoma X 1

Renal tubule, adenoma 3

Urinary bladder + + + + + + + + + + + + + + + 40 

Transitional epithelium, papilloma X 2

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + 40 

Leukemia mononuclear 1
170 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

5,000 ppm

5566666666666666677777777777777

Number of Days on Study 7800034444666799901112222222222

5216922799007015684695999999999

1010100101010001011001000000000

Carcass ID Number 0909099090909990900990999999999

0726098342830671610532000011112

5355454444335424333443123412341

Alimentary System 

Esophagus + + + + + + + M + + + + + + + + + + + + + + + + + + + + + + +

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + M + + + + +

Adenocarcinoma X 

Polyp adenomatous 

Polyp adenomatous, multiple X 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Adenocarcinoma X X X X X X X 

Adenocarcinoma, multiple 

Polyp adenomatous X X X X X X 

Polyp adenomatous, multiple X X X X X X X X X X X X X X X X X X

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Polyp adenomatous 

Intestine small, duodenum + + + + + + + + + + + + M + + + + + + + + + + + + + + + + + +

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + M + + + + + + + + + +

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + M + + + + + + + + +

Leiomyoma 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Hepatocellular carcinoma X X X X X 

Hepatocellular carcinoma, multiple X X X X X X X X X X X X X X X X X X X X X X X

Hepatocellular adenoma X X X X X 

Hepatocellular adenoma, multiple X X X X X X X X X X X X X X X X X X

Hepatocholangiocarcinoma X X X X X

Hepatocholangiocarcinoma, multiple X 

Hepatocholangioma X 

Hepatocholangioma, multiple 

Mesentery + 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Adenoma X 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Squamous cell carcinoma X 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Hepatocellular carcinoma, metastatic, 

liver X 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Pheochromocytoma complex X 

Pheochromocytoma benign 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +
Lesions in Female Rats 171

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

5,000 ppm (continued)

77777777777777777777777777777

Number of Days on Study 22222223333333333333333333333

99999990000000000000000000000

00011110000000000000000001111

Carcass ID Number 9 9 9 0 0 0 0 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 0 0 0 0 Total

2 2 2 0 0 3 3 3 3 3 4 4 4 5 5 5 6 6 7 8 8 9 9 9 9 1 1 2 2 Tissues/

2 3 4 1 2 1 2 1 2 3 1 2 3 1 2 3 1 2 1 1 2 1 2 3 4 1 2 1 2 Tumors

Alimentary System 

Esophagus + + + + + + + + + + + M + + + + + + + + + + M + + + + + + 57

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Adenocarcinoma X 2

Polyp adenomatous X 1

Polyp adenomatous, multiple 1

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Adenocarcinoma X X X X X X X X X X X 18

Adenocarcinoma, multiple X 1

Polyp adenomatous X X 8

Polyp adenomatous, multiple X X X X X X X X X X X X X X X X X X X X X X X X X X X 45

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Polyp adenomatous X 1

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Leiomyoma X 1

Liver + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Hepatocellular carcinoma X 6

Hepatocellular carcinoma, multiple X X X X X X X X X X X X X X X X X X X X X X X X X X X X 51

Hepatocellular adenoma X X X 8

Hepatocellular adenoma, multiple X X X X X X X X X X X X X X X X X X X X X 39

Hepatocholangiocarcinoma X X X X X 10

Hepatocholangiocarcinoma, multiple 1

Hepatocholangioma 1

Hepatocholangioma, multiple X 1

Mesentery 1

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Adenoma X 2

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Squamous cell carcinoma 1

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Hepatocellular carcinoma, metastatic, 

liver 1

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + M + + + + + + + + + + + + + + 59

Adrenal medulla + + + + + + + + + + + + + + M + + + + + + + + + + + + + + 59

Pheochromocytoma complex 1

Pheochromocytoma benign X 1

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60
172 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

5,000 ppm (continued)

5566666666666666677777777777777

Number of Days on Study 7800034444666799901112222222222

5216922799007015684695999999999

1010100101010001011001000000000

Carcass ID Number 0909099090909990900990999999999

0726098342830671610532000011112

5355454444335424333443123412341

Endocrine System (continued) 

Parathyroid gland + + + + + + + + + + + + + + + + + + + + + M + + + + + + M + +

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Pars distalis, adenoma X X X X X X X X X X X X X X X 

Pars distalis, adenoma, multiple X X X

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

C-cell, adenoma X X X 

C-cell, carcinoma X 

Follicular cell, adenoma X 

General Body System 

Tissue NOS 

Neoplasm NOS 

Genital System

Clitoral gland + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Adenoma X X 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Hepatocellular carcinoma, metastatic, 

liver 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Polyp stromal X X X X 

Sarcoma stromal 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node + + + + + + + + +

Lymph node, mandibular + + + + + M + + + + + + + + + + M + + + + + M + + + + + + + +

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + M + + + + +

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Hepatocellular carcinoma, metastatic, 

liver X 

Thymus + M M + I M + M + M + + + M + + + + + + M + + + + + + + + + +

Integumentary System 

Mammary gland + + + M + + + + + + + + + + + M + + + + + + + + + + + + + M +

Fibroadenoma X X X X X 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Squamous cell papilloma X 

Pinna, granular cell tumor benign 

Pinna, squamous cell papilloma 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Nervous System

Brain + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +
Lesions in Female Rats 173

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

5,000 ppm (continued)

77777777777777777777777777777

Number of Days on Study 22222223333333333333333333333

99999990000000000000000000000

00011110000000000000000001111

Carcass ID Number 9 9 9 0 0 0 0 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 0 0 0 0 Total

2 2 2 0 0 3 3 3 3 3 4 4 4 5 5 5 6 6 7 8 8 9 9 9 9 1 1 2 2 Tissues/

2 3 4 1 2 1 2 1 2 3 1 2 3 1 2 3 1 2 1 1 2 1 2 3 4 1 2 1 2 Tumors

Endocrine System (continued)

Parathyroid gland + + + + + + + + + + + + M + + + + + + + + + + + + + + + + 57

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Pars distalis, adenoma X X X X X X X X X X 25

Pars distalis, adenoma, multiple X X X X 7

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

C-cell, adenoma X X 5

C-cell, carcinoma 1

Follicular cell, adenoma 1

General Body System

Tissue NOS + 1

Neoplasm NOS X 1

Genital System

Clitoral gland + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Adenoma X 3

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Hepatocellular carcinoma, metastatic,

liver X 1

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Polyp stromal X X X 7

Sarcoma stromal X 1

Hematopoietic System

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Lymph node + + + + + + + + + + + + + + + + 25

Lymph node, mandibular + + + + M + + + + + + + + + + + + + + + + + + + + + + + + 56

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 59

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Hepatocellular carcinoma, metastatic,

liver 1

Thymus + + M + + + + + + + + + + + + + + + + + + + + + + + + + + 51

Integumentary System

Mammary gland M + + + + M + M + M + + + M M + + + + + + M + + + + + + + 50

Fibroadenoma X X X X 9

Skin + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Squamous cell papilloma 1

Pinna, granular cell tumor benign X 1

Pinna, squamous cell papilloma X X2

Musculoskeletal System

Bone + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Nervous System

Brain + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60
174 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

5,000 ppm (continued)

5566666666666666677777777777777

Number of Days on Study 7800034444666799901112222222222

5216922799007015684695999999999

1010100101010001011001000000000

Carcass ID Number 0909099090909990900990999999999

0726098342830671610532000011112

5355454444335424333443123412341

Respiratory System 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Alveolar/bronchiolar adenoma X 

Hepatocellular carcinoma, metastatic, 

liver X X X X X X X X X 

Nose + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Special Senses System 

Ear + + + 

Eye + 

Zymbal's gland + 

Adenoma 

Carcinoma X 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Squamous cell carcinoma, metastatic, 

urinary bladder X 

Renal tubule, adenoma X X X X X X

Renal tubule, adenoma, multiple X X 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Transitional epithelium, carcinoma X X 

Transitional epithelium, papilloma X X X X

Transitional epithelium, papilloma, 

multiple 

Transitional epithelium, squamous 

cell carcinoma X 

Transitional epithelium, squamous 

cell papilloma 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + +

Leukemia mononuclear X X X X 
Lesions in Female Rats 175

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

5,000 ppm (continued)

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7

Number of Days on Study 2 2 2 2 2 2 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3

9 9 9 9 9 9 9 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

0 0 0 1 1 1 1 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 1 1 1 1

Carcass ID Number 9 9 9 0 0 0 0 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 9 0 0 0 0 Total

2 2 2 0 0 3 3 3 3 3 4 4 4 5 5 5 6 6 7 8 8 9 9 9 9 1 1 2 2 Tissues/

2 3 4 1 2 1 2 1 2 3 1 2 3 1 2 3 1 2 1 1 2 1 2 3 4 1 2 1 2 Tumors

Respiratory System 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Alveolar/bronchiolar adenoma 1

Hepatocellular carcinoma, metastatic,

liver X X X X X X X X X X X X X 22

Nose + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Special Senses System

Ear + + + + + 8

Eye + + + + + + 7

Zymbal's gland + 2

Adenoma X 1

Carcinoma 1

Urinary System

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Squamous cell carcinoma, metastatic,

urinary bladder 1

Renal tubule, adenoma X X X X X 11

Renal tubule, adenoma, multiple X X X 5

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Transitional epithelium, carcinoma X X X X X X 8

Transitional epithelium, papilloma X X 6

Transitional epithelium, papilloma,

multiple X 1

Transitional epithelium, squamous

cell carcinoma 1

Transitional epithelium, squamous

cell papilloma X 1

Systemic Lesions

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 60

Leukemia mononuclear X 5
176 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm

344555555556666666666666 

Number of Days on Study 616145667890111233333566 

780239696080046556678822

111111111111111111111111 

Carcass ID Number 101101001011010010010101 

774593475813805909948266 

353545444434344331222133

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + +

Intestine large, colon + + + + + + + + + + + + + + + + M + + + + + + + 

Adenocarcinoma 

Polyp adenomatous 

Polyp adenomatous, multiple 

Intestine large, rectum + + + + + + + + + + + + + + + + M + + + + + + + 

Adenocarcinoma 

Adenocarcinoma, multiple X 

Polyp adenomatous X X X X X X X X X X 

Polyp adenomatous, multiple X X X X X X X X X

Intestine large, cecum + + + + + + + + + A + + + + + + M + + + + + + +

Intestine small, duodenum + + + + M + + + + + + + + + + + M + + + + + + A

Intestine small, jejunum + + + + + + + + + A + + + + + + M + + + + + + A

Intestine small, ileum + + + + + + + + + A + + + M + + M + + + + + + A

Liver + + + + + + + + + + + + + + + + M + + + + + + + 

Cholangioma 

Hepatocellular carcinoma X X X 

Hepatocellular carcinoma, multiple X X X X X X X X X X X X X X X X X 

Hepatocellular adenoma X X X X 

Hepatocellular adenoma, multiple X X X X X X X X X X X X X 

Hepatocholangiocarcinoma X X X 

Hepatocholangiocarcinoma, multiple X X 

Mesentery + 

Hepatocellular carcinoma, metastatic, 

liver 

Pancreas + + + + + + + + + + + + + + + + M + + + + + + + 

Hepatocellular carcinoma, metastatic, 

liver 

Salivary glands + + + M + + + + + + + + + + + + + + + + + + + +

Stomach, forestomach + + + + + + + + + + + + + + + + M + + + + + + + 

Squamous cell carcinoma X 

Squamous cell papilloma 

Stomach, glandular + + + + + + + + + + + + + + + + M + + + + + + + 

Hepatocellular carcinoma, metastatic, 

liver 

Tongue + 

Squamous cell papilloma X 

Tooth 

Gingiva, squamous cell carcinoma 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + 

Hepatocellular carcinoma, metastatic, 

liver 
Lesions in Female Rats 177

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

6666677777777777777777777

Number of Days on Study 6778900011122222222222222

4345522545902999999999999

1111111111111111111111111

Carcass ID Number 1 1 0 0 0 0 0 1 1 1 1 0 0 0 0 0 0 0 1 1 1 1 1 1 1 Total

6 5 7 6 4 5 8 3 1 3 4 6 7 4 4 5 5 7 0 0 1 3 5 5 6 Tissues/

2 3 3 2 3 3 1 3 2 2 1 1 2 1 2 1 2 1 1 2 1 1 1 2 1 Tumors

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + M + + + + + + 48 

Intestine large, colon + + + + + + + + + + M + + + + + + + + + + + + + + 47 

Adenocarcinoma X 1

Polyp adenomatous X 1

Polyp adenomatous, multiple X 1

Intestine large, rectum + + + + + M + + + + + + + + + + + + + + + + + + + 47 

Adenocarcinoma X X X X X X 6

Adenocarcinoma, multiple 1

Polyp adenomatous X X 12 

Polyp adenomatous, multiple X X X X X X X X X X X X X X X X X X X X X X 31 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 46 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 46 

Intestine small, ileum + + + + + M + + + + + + + + + + + + + + + + + + + 44 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Cholangioma X 1

Hepatocellular carcinoma X 4

Hepatocellular carcinoma, multiple X X X X X X X X X X X X X X X X X X X X X X X X 41 

Hepatocellular adenoma X X X 7

Hepatocellular adenoma, multiple X X X X X X X X X 22 

Hepatocholangiocarcinoma X X X X X 8

Hepatocholangiocarcinoma, multiple X X X 5

Mesentery + + + 4

Hepatocellular carcinoma, metastatic, 

liver X X X 3

Pancreas + + + + + + + M + + + + + + + + + + + + + + + + + 47 

Hepatocellular carcinoma, metastatic, 

liver X 1

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Stomach, forestomach + + + + + + + + + + + + + + + + + M + + + + + + + 47 

Squamous cell carcinoma 1

Squamous cell papilloma X 1

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Hepatocellular carcinoma, metastatic, 

liver X 1

Tongue 1

Squamous cell papilloma 1

Tooth + 1

Gingiva, squamous cell carcinoma X 1

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Hepatocellular carcinoma, metastatic, 

liver X X 2
178 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

344555555556666666666666 

Number of Days on Study 616145667890111233333566 

780239696080046556678822

111111111111111111111111 

Carcass ID Number 101101001011010010010101 

774593475813805909948266 

353545444434344331222133

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + M + + + + + + + 

Carcinoma, metastatic, kidney 

Adrenal medulla + + + + + + + + + + + + + + + + M + + + + + + + 

Pheochromocytoma benign 

Bilateral, pheochromocytoma benign X 

Islets, pancreatic + + + + + + + + + + + + + + + + M + + + + + + + 

Hepatocellular carcinoma, metastatic, 

liver 

Parathyroid gland + + M + + M + M + M + + M + + + M + + + + + M +

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + 

Pars distalis, adenoma X X X X X X

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + 

C-cell, adenoma, multiple 

Follicular cell, adenoma 

General Body System 

None 

Genital System

Clitoral gland + + M + + + + + + + + + + + + + M + + M + + + + 

Adenoma X 

Ovary + + + + + + + + + + + + + + + + M + + + + + + + 

Hepatocellular carcinoma, metastatic, 

liver 

Uterus + + + + + + + + + + + + + + + + M + + + + + + + 

Polyp stromal X X X 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node + + + + + + 

Mediastinal, hepatocellular 

carcinoma, metastatic, liver 

Lymph node, mandibular + + + + + + + + + + + + + + + M M + + + + + + +

Lymph node, mesenteric + + + + + + + + + + + + + + + + M + + + + + + +

Spleen + + + + + + + + + + + + + + + + M + + + + + + +

Thymus + + + M + M + + + + + M + + + + M + + M + + + + 

Hepatocellular carcinoma, metastatic, 

liver 

Integumentary System 

Mammary gland + + + + + + + + + + + + + + + M M + + + + + + + 

Fibroadenoma X X X 

Skin + + + + + + + + + + + + + + + + + + + + + + + + 

Squamous cell papilloma X 

Pinna, squamous cell papilloma 
Lesions in Female Rats 179

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

6666677777777777777777777

Number of Days on Study 6778900011122222222222222

4345522545902999999999999

1111111111111111111111111

Carcass ID Number 1 1 0 0 0 0 0 1 1 1 1 0 0 0 0 0 0 0 1 1 1 1 1 1 1 Total

6 5 7 6 4 5 8 3 1 3 4 6 7 4 4 5 5 7 0 0 1 3 5 5 6 Tissues/

2 3 3 2 3 3 1 3 2 2 1 1 2 1 2 1 2 1 1 2 1 1 1 2 1 Tumors

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + M + + + 47 

Carcinoma, metastatic, kidney X 1

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + M + + + 47 

Pheochromocytoma benign X 1

Bilateral, pheochromocytoma benign 1

Islets, pancreatic + + + + + + + M + + + + + + + + + + + + + + + + + 47 

Hepatocellular carcinoma, metastatic, 

liver X 1

Parathyroid gland + M + + + + + + + + + + M + + + + + + + M M + + + 38 

Pituitary gland + + + + + + + + + + + M + + + + M + + + + + + + + 47 

Pars distalis, adenoma X X X X X X X 13 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

C-cell, adenoma, multiple X 1

Follicular cell, adenoma X X 2

General Body System 

None 

Genital System

Clitoral gland + + + + + M + + + + + + + + + + + + + + + + + + + 45 

Adenoma X 2

Ovary + + + + + M + + + + + + + + + + + + + + + + + + + 47 

Hepatocellular carcinoma, metastatic, 

liver X 1

Uterus + + + + + M + + + + + + + + + + + + + + + + + + + 47 

Polyp stromal X X 5

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Lymph node + + + + + + + + 14 

Mediastinal, hepatocellular 

carcinoma, metastatic, liver X 1

Lymph node, mandibular + + M + + + + + + + + + M + + + + + + + + + + + + 45 

Lymph node, mesenteric + + + + + + + + + + M + + + M + + + + + + + + + + 46 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Thymus + + + + M M + + M + + + M + + M + + + + + + + + M 38 

Hepatocellular carcinoma, metastatic, 

liver X 1

Integumentary System 

Mammary gland + + + + M + + M + + + + + + M + M + + + M + + + M 41 

Fibroadenoma X X 5

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Squamous cell papilloma 1

Pinna, squamous cell papilloma X X 2
180 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

344555555556666666666666 

Number of Days on Study 616145667890111233333566 

780239696080046556678822

111111111111111111111111 

Carcass ID Number 101101001011010010010101 

774593475813805909948266 

353545444434344331222133

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + 

Squamous cell carcinoma, metastatic, 

uncertain primary site X 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + 

Oligodendroglioma benign X 

Spinal cord + 

Respiratory System 

Lung + + + + + + + + + + + + + + + + + + + + + + + + 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, 

liver X X X X X X 

Neoplasm NOS, metastatic, uncertain 

primary site X 

Nose + + + + + + + + + + + + + + + + + + + + + + + +

Trachea + + + + + + + + + + + + + + + + + + + + + + + +

Special Senses System 

Ear 

Eye + 

Urinary System

Kidney + + + + + + + + + + + + + + + + M + + + + + + + 

Pelvis, transitional epithelium, 

papilloma 

Renal tubule, adenoma X X X X 

Renal tubule, adenoma, multiple 

Renal tubule, carcinoma 

Urinary bladder + + + + + + + + + + + + + + + + M + + + + + + A 

Transitional epithelium, carcinoma X X X X X X X X X 

Transitional epithelium, papilloma X X 

Transitional epithelium, squamous 

cell papilloma 

Transitional epithelium, squamous 

cell papilloma, multiple 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + +

Leukemia mononuclear X 

Lymphoma malignant histiocytic 
Lesions in Female Rats 181

TABLE B2

Individual Animal Tumor Pathology of Female Rats in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

6666677777777777777777777

Number of Days on Study 6778900011122222222222222

4345522545902999999999999

1111111111111111111111111

Carcass ID Number 1 1 0 0 0 0 0 1 1 1 1 0 0 0 0 0 0 0 1 1 1 1 1 1 1 Total

6 5 7 6 4 5 8 3 1 3 4 6 7 4 4 5 5 7 0 0 1 3 5 5 6 Tissues/

2 3 3 2 3 3 1 3 2 2 1 1 2 1 2 1 2 1 1 2 1 1 1 2 1 Tumors

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Squamous cell carcinoma, metastatic, 

uncertain primary site 1

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Oligodendroglioma benign 1

Spinal cord 1

Respiratory System 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Alveolar/bronchiolar carcinoma X 1

Hepatocellular carcinoma, metastatic, 

liver X X X X X X X X X X X X X X X X X X 24 

Neoplasm NOS, metastatic, uncertain 

primary site 1

Nose + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Special Senses System 

Ear + + + 3

Eye + + + + + + 7

Urinary System

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Pelvis, transitional epithelium, 

papilloma X 1

Renal tubule, adenoma X X X X X X X 11 

Renal tubule, adenoma, multiple X X X X X 5

Renal tubule, carcinoma X X 2

Urinary bladder + + + + + M + + + + + + + + + + + + + + + + + + + 46 

Transitional epithelium, carcinoma X X X X X X X 16 

Transitional epithelium, papilloma X X X X X X X 9

Transitional epithelium, squamous 

cell papilloma X 1

Transitional epithelium, squamous 

cell papilloma, multiple X 1

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Leukemia mononuclear 1

Lymphoma malignant histiocytic X 1
182 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B3

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Adrenal Medulla: Benign Pheochromocytoma

Overall ratea 2/47 (4%) 3/40 (8%) 1/59 (2%) 2/47 (4%)

Adjusted rateb 5.7% 9.4% 2.7% 7.7%

Terminal ratec 2/35 (6%) 3/32 (9%) 1/37 (3%) 0/11 (0%)

First incidence (days) 729 (T) 729 (T) 729 (T) 636

Life table testd P=0.396 P=0.459 P=0.481N P=0.379 

Logistic regression testd P=0.602N P=0.459 P=0.481N P=0.624 

Cochran-Armitage testd P=0.441N

Fisher exact testd P=0.423 P=0.415N P=0.692N

Adrenal Medulla: Benign or Complex Pheochromocytoma

Overall rate 2/47 (4%) 3/40 (8%) 2/59 (3%) 2/47 (4%)

Adjusted rate 5.7% 9.4% 4.6% 7.7%

Terminal rate 2/35 (6%) 3/32 (9%) 1/37 (3%) 0/11 (0%)

First incidence (days) 729 (T) 729 (T) 660 636

Life table test P=0.354 P=0.459 P=0.665N P=0.379 

Logistic regression test P=0.583N P=0.459 P=0.617N P=0.624 

Cochran-Armitage test P=0.467N

Fisher exact test P=0.423 P=0.601N P=0.692N

Clitoral Gland: Adenoma

Overall rate 5/45 (11%) 3/36 (8%) 3/60 (5%) 2/45 (4%)

Adjusted rate 13.3% 10.0% 7.4% 10.8%

Terminal rate 4/36 (11%) 3/30 (10%) 2/38 (5%) 1/12 (8%)

First incidence (days) 708 729 (T) 696 610

Life table test P=0.524N P=0.465N P=0.330N P=0.664 

Logistic regression test P=0.265N P=0.488N P=0.263N P=0.365N

Cochran-Armitage test P=0.137N

Fisher exact test P=0.488N P=0.212N P=0.217N

Kidney (Renal Tubule): Adenoma

Overall rate 0/50 (0%) 3/40 (8%) 16/60 (27%) 16/48 (33%)

Adjusted rate 0.0% 8.0% 36.0% 69.7%

Terminal rate 0/38 (0%) 1/32 (3%) 11/38 (29%) 6/12 (50%)

First incidence (days) )e 600 601 625

Life table test P<0.001 P=0.093 P<0.001 P<0.001

Logistic regression test P<0.001 P=0.049 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P=0.084 P<0.001 P<0.001

Kidney (Renal Tubule): Adenoma or Carcinoma

Overall rate 0/50 (0%) 3/40 (8%) 16/60 (27%) 16/48 (33%)

Adjusted rate 0.0% 8.0% 36.0% 69.7%

Terminal rate 0/38 (0%) 1/32 (3%) 11/38 (29%) 6/12 (50%)

First incidence (days) ) 600 601 625

Life table test P<0.001 P=0.093 P<0.001 P<0.001

Logistic regression test P<0.001 P=0.049 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P=0.084 P<0.001 P<0.001
Lesions in Female Rats 183

TABLE B3

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Large Intestine (Rectum): Adenomatous Polyp

Overall rate 0/50 (0%) 27/40 (68%) 53/60 (88%) 43/49 (88%)

Adjusted rate 0.0% 75.0% 100.0% 100.0%

Terminal rate 0/38 (0%) 23/32 (72%) 38/38 (100%) 12/12 (100%)

First incidence (days) ) 616 582 512

Life table test P<0.001 P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001 P<0.001

Large Intestine (Rectum): Carcinoma

Overall rate 0/50 (0%) 1/40 (3%) 19/60 (32%) 7/49 (14%)

Adjusted rate 0.0% 3.1% 41.7% 41.9%

Terminal rate 0/38 (0%) 1/32 (3%) 13/38 (34%) 4/12 (33%)

First incidence (days) ) 729 (T) 606 625

Life table test P<0.001 P=0.466 P<0.001 P<0.001

Logistic regression test P<0.001 P=0.466 P<0.001 P=0.001

Cochran-Armitage test P=0.005

Fisher exact test P=0.444 P<0.001 P=0.006

Large Intestine (All Sites): Adenomatous Polyp

Overall rate 0/50 (0%) 28/40 (70%) 53/60 (88%) 43/49 (88%)

Adjusted rate 0.0% 77.7% 100.0% 100.0%

Terminal rate 0/38 (0%) 24/32 (75%) 38/38 (100%) 12/12 (100%)

First incidence (days) ) 616 582 512

Life table test P<0.001 P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001 P<0.001

Large Intestine (All Sites): Carcinoma

Overall rate 0/50 (0%) 2/40 (5%) 21/60 (35%) 8/49 (16%)

Adjusted rate 0.0% 6.3% 45.1% 46.3%

Terminal rate 0/38 (0%) 2/32 (6%) 14/38 (37%) 4/12 (33%)

First incidence (days) ) 729 (T) 606 625

Life table test P<0.001 P=0.201 P<0.001 P<0.001

Logistic regression test P<0.001 P=0.201 P<0.001 P<0.001

Cochran-Armitage test P=0.004

Fisher exact test P=0.195 P<0.001 P=0.003

Liver: Hepatocellular Adenoma

Overall rate 0/50 (0%) 28/40 (70%) 47/60 (78%) 29/48 (60%)

Adjusted rate 0.0% 75.5% 83.7% 83.6%

Terminal rate 0/38 (0%) 23/32 (72%) 29/38 (76%) 8/12 (67%)

First incidence (days) ) 600 575 418

Life table test P<0.001 P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001 P<0.001
184 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B3

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Liver: Hepatocellular Carcinoma

Overall rate 0/50 (0%) 12/40 (30%) 57/60 (95%) 45/48 (94%)

Adjusted rate 0.0% 37.5% 98.3% 100.0%

Terminal rate 0/38 (0%) 12/32 (38%) 37/38 (97%) 12/12 (100%)

First incidence (days) ) 729 (T) 575 460

Life table test P<0.001 P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001 P<0.001

Liver: Hepatocellular Adenoma or Carcinoma

Overall rate 0/50 (0%) 33/40 (83%) 59/60 (98%) 47/48 (98%)

Adjusted rate 0.0% 89.1% 100.0% 100.0%

Terminal rate 0/38 (0%) 28/32 (88%) 38/38 (100%) 12/12 (100%)

First incidence (days) ) 600 575 418

Life table test P<0.001 P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001 P<0.001

Liver: Hepatocholangiocarcinoma

Overall rate 0/50 (0%) 0/40 (0%) 11/60 (18%) 13/48 (27%)

Adjusted rate 0.0% 0.0% 27.1% 56.9%

Terminal rate 0/38 (0%) 0/32 (0%) 9/38 (24%) 5/12 (42%)

First incidence (days) ) ) 670 600

Life table test P<0.001 ) P<0.001 P<0.001

Logistic regression test P<0.001 ) P=0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test ) P<0.001 P<0.001

Lung: Alveolar/bronchiolar Adenoma

Overall rate 1/50 (2%) 3/40 (8%) 1/60 (2%) 0/49 (0%)

Adjusted rate 2.5% 9.4% 1.9% 0.0%

Terminal rate 0/38 (0%) 3/32 (9%) 0/38 (0%) 0/12 (0%)

First incidence (days) 716 729 (T) 647 )

Life table test P=0.338N P=0.244 P=0.739N P=0.695N

Logistic regression test P=0.196N P=0.229 P=0.732N P=0.616N

Cochran-Armitage test P=0.154N

Fisher exact test P=0.229 P=0.705N P=0.505N

Lung: Alveolar/bronchiolar Adenoma or Carcinoma

Overall rate 1/50 (2%) 3/40 (8%) 1/60 (2%) 1/49 (2%)

Adjusted rate 2.5% 9.4% 1.9% 5.0%

Terminal rate 0/38 (0%) 3/32 (9%) 0/38 (0%) 0/12 (0%)

First incidence (days) 716 729 (T) 647 702

Life table test P=0.572 P=0.244 P=0.739N P=0.559 

Logistic regression test P=0.472N P=0.229 P=0.732N P=0.682 

Cochran-Armitage test P=0.380N

Fisher exact test P=0.229 P=0.705N P=0.747 
Lesions in Female Rats 185

TABLE B3

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Mammary Gland: Fibroadenoma

Overall rate 21/50 (42%) 10/40 (25%) 9/60 (15%) 5/49 (10%)

Adjusted rate 52.2% 29.1% 20.7% 22.3%

Terminal rate 19/38 (50%) 8/32 (25%) 6/38 (16%) 1/12 (8%)

First incidence (days) 606 659 582 610

Life table test P=0.058N P=0.055N P=0.008N P=0.210N

Logistic regression test P=0.001N P=0.065N P=0.002N P=0.008N

Cochran-Armitage test P<0.001N

Fisher exact test P=0.071N P=0.002N P<0.001N

Mammary Gland: Fibroma, Fibroadenoma, or Carcinoma

Overall rate 23/50 (46%) 10/40 (25%) 9/60 (15%) 5/49 (10%)

Adjusted rate 57.2% 29.1% 20.7% 22.3%

Terminal rate 21/38 (55%) 8/32 (25%) 6/38 (16%) 1/12 (8%)

First incidence (days) 606 659 582 610

Life table test P=0.030N P=0.024N P=0.003N P=0.147N

Logistic regression test P<0.001N P=0.029N P<0.001N P=0.004N

Cochran-Armitage test P<0.001N

Fisher exact test P=0.033N P<0.001N P<0.001N

Pituitary Gland (Pars Distalis): Adenoma

Overall rate 32/50 (64%) 19/39 (49%) 32/60 (53%) 13/47 (28%)

Adjusted rate 74.3% 55.6% 62.3% 55.7%

Terminal rate 27/38 (71%) 17/32 (53%) 19/38 (50%) 4/11 (36%)

First incidence (days) 642 616 575 512

Life table test P=0.285 P=0.060N P=0.510N P=0.431 

Logistic regression test P=0.015N P=0.081N P=0.208N P=0.013N

Cochran-Armitage test P<0.001N

Fisher exact test P=0.109N P=0.175N P<0.001N

Skin: Squamous Cell Papilloma

Overall rate 1/50 (2%) 0/40 (0%) 3/60 (5%) 3/49 (6%)

Adjusted rate 2.6% 0.0% 7.9% 19.4%

Terminal rate 1/38 (3%) 0/32 (0%) 3/38 (8%) 2/12 (17%)

First incidence (days) 729 (T) ) 729 (T) 637

Life table test P=0.007 P=0.534N P=0.305 P=0.056 

Logistic regression test P=0.033 P=0.534N P=0.305 P=0.171 

Cochran-Armitage test P=0.109 

Fisher exact test P=0.556N P=0.381 P=0.301 

Skin (Subcutaneous Tissue): Fibroma

Overall rate 2/50 (4%) 2/40 (5%) 0/60 (0%) 0/49 (0%)

Adjusted rate 5.3% 6.0% 0.0% 0.0%

Terminal rate 2/38 (5%) 1/32 (3%) 0/38 (0%) 0/12 (0%)

First incidence (days) 729 (T) 670 ) )

Life table test P=0.148N P=0.625 P=0.238N P=0.513N

Logistic regression test P=0.089N P=0.616 P=0.238N P=0.513N

Cochran-Armitage test P=0.068N

Fisher exact test P=0.603 P=0.204N P=0.253N
186 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B3

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Skin (Subcutaneous Tissue): Fibroma or Sarcoma

Overall rate 3/50 (6%) 2/40 (5%) 0/60 (0%) 0/49 (0%)

Adjusted rate 7.5% 6.0% 0.0% 0.0%

Terminal rate 2/38 (5%) 1/32 (3%) 0/38 (0%) 0/12 (0%)

First incidence (days) 704 670 ) )

Life table test P=0.080N P=0.591N P=0.122N P=0.355N

Logistic regression test P=0.039N P=0.599N P=0.100N P=0.247N

Cochran-Armitage test P=0.031N

Fisher exact test P=0.606N P=0.091N P=0.125N

Thyroid Gland (C-cell): Adenoma

Overall rate 5/50 (10%) 5/40 (13%) 5/60 (8%) 1/49 (2%)

Adjusted rate 13.2% 15.6% 12.7% 6.7%

Terminal rate 5/38 (13%) 5/32 (16%) 4/38 (11%) 0/12 (0%)

First incidence (days) 729 (T) 729 (T) 714 719

Life table test P=0.405N P=0.519 P=0.627N P=0.509N

Logistic regression test P=0.303N P=0.519 P=0.604N P=0.421N

Cochran-Armitage test P=0.058N

Fisher exact test P=0.481 P=0.509N P=0.107N

Thyroid Gland (C-cell): Adenoma or Carcinoma

Overall rate 5/50 (10%) 5/40 (13%) 6/60 (10%) 1/49 (2%)

Adjusted rate 13.2% 15.6% 15.2% 6.7%

Terminal rate 5/38 (13%) 5/32 (16%) 5/38 (13%) 0/12 (0%)

First incidence (days) 729 (T) 729 (T) 714 719

Life table test P=0.469N P=0.519 P=0.503 P=0.509N

Logistic regression test P=0.360N P=0.519 P=0.530 P=0.421N

Cochran-Armitage test P=0.069N

Fisher exact test P=0.481 P=0.622N P=0.107N

Urinary Bladder: Papilloma

Overall rate 0/50 (0%) 2/40 (5%) 7/60 (12%) 9/46 (20%)

Adjusted rate 0.0% 6.3% 17.6% 39.5%

Terminal rate 0/38 (0%) 2/32 (6%) 6/38 (16%) 1/12 (8%)

First incidence (days) ) 729 (T) 691 637

Life table test P<0.001 P=0.201 P=0.010 P<0.001

Logistic regression test P<0.001 P=0.201 P=0.012 P=0.003

Cochran-Armitage test P<0.001

Fisher exact test P=0.195 P=0.012 P<0.001

Urinary Bladder: Carcinoma

Overall rate 0/50 (0%) 0/40 (0%) 8/60 (13%) 16/46 (35%)

Adjusted rate 0.0% 0.0% 19.5% 55.8%

Terminal rate 0/38 (0%) 0/32 (0%) 6/38 (16%) 4/12 (33%)

First incidence (days) ) ) 670 367

Life table test P<0.001 ) P=0.006 P<0.001

Logistic regression test P<0.001 ) P=0.008 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test ) P=0.006 P<0.001
Lesions in Female Rats 187

TABLE B3

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Urinary Bladder: Papilloma, Squamous Cell Papilloma, Carcinoma, or Squamous Cell Carcinoma

Overall rate 0/50 (0%) 2/40 (5%) 17/60 (28%) 26/46 (57%)

Adjusted rate 0.0% 6.3% 40.9% 78.1%

Terminal rate 0/38 (0%) 2/32 (6%) 14/38 (37%) 6/12 (50%)

First incidence (days) ) 729 (T) 670 367

Life table test P<0.001 P=0.201 P<0.001 P<0.001

Logistic regression test P<0.001 P=0.201 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P=0.195 P<0.001 P<0.001

Uterus: Stromal Polyp

Overall rate 8/50 (16%) 15/40 (38%) 7/60 (12%) 5/49 (10%)

Adjusted rate 20.2% 41.4% 16.6% 17.4%

Terminal rate 7/38 (18%) 11/32 (34%) 5/38 (13%) 0/12 (0%)

First incidence (days) 656 600 601 512

Life table test P=0.498N P=0.030 P=0.482N P=0.393 

Logistic regression test P=0.041N P=0.019 P=0.371N P=0.295N

Cochran-Armitage test P=0.038N

Fisher exact test P=0.019 P=0.350N P=0.290N

Uterus: Stromal Polyp or Stromal Sarcoma 

Overall rate 8/50 (16%) 15/40 (38%) 8/60 (13%) 5/49 (10%)

Adjusted rate 20.2% 41.4% 19.2% 17.4%

Terminal rate 7/38 (18%) 11/32 (34%) 6/38 (16%) 0/12 (0%)

First incidence (days) 656 600 601 512

Life table test P=0.540N P=0.030 P=0.589N P=0.393 

Logistic regression test P=0.050N P=0.019 P=0.477N P=0.295N

Cochran-Armitage test P=0.042N

Fisher exact test P=0.019 P=0.449N P=0.290N

All Organs: Mononuclear Cell Leukemia

Overall rate 9/50 (18%) 1/40 (3%) 5/60 (8%) 1/49 (2%)

Adjusted rate 21.5% 3.0% 10.0% 3.7%

Terminal rate 6/38 (16%) 0/32 (0%) 1/38 (3%) 0/12 (0%)

First incidence (days) 620 689 601 662

Life table test P=0.112N P=0.026N P=0.177N P=0.162N

Logistic regression test P=0.011N P=0.023N P=0.111N P=0.024N

Cochran-Armitage test P=0.016N

Fisher exact test P=0.019N P=0.110N P=0.009N

All Organs: Benign Neoplasms

Overall rate 43/50 (86%) 40/40 (100%) 60/60 (100%) 46/49 (94%)

Adjusted rate 91.5% 100.0% 100.0% 100.0%

Terminal rate 34/38 (89%) 32/32 (100%) 38/38 (100%) 12/12 (100%)

First incidence (days) 606 600 575 418

Life table test P<0.001 P=0.223 P=0.007 P<0.001

Logistic regression test P=0.009 P=0.025 P=0.004 P=0.015

Cochran-Armitage test P=0.164 

Fisher exact test P=0.013 P=0.003 P=0.167 
188 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B3

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

All Organs: Malignant Neoplasms

Overall rate 16/50 (32%) 15/40 (38%) 60/60 (100%) 46/49 (94%)

Adjusted rate 33.9% 45.5% 100.0% 100.0%

Terminal rate 8/38 (21%) 14/32 (44%) 38/38 (100%) 12/12 (100%)

First incidence (days) 268 689 575 367

Life table test P<0.001 P=0.424 P<0.001 P<0.001

Logistic regression test P<0.001 P=0.341 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P=0.373 P<0.001 P<0.001

All Organs: Benign or Malignant Neoplasms

Overall rate 46/50 (92%) 40/40 (100%) 60/60 (100%) 49/49 (100%)

Adjusted rate 92.0% 100.0% 100.0% 100.0%

Terminal rate 34/38 (89%) 32/32 (100%) 38/38 (100%) 12/12 (100%)

First incidence (days) 268 600 575 367

Life table test P<0.001 P=0.435 P=0.035 P<0.001

Logistic regression test P=0.049 P=0.086 P=0.063 P=0.289 

Cochran-Armitage test P=0.021

Fisher exact test P=0.090 P=0.040 P=0.061 

(T)Terminal sacrifice

a Number of lesion-bearing animals/number of animals examined. Denominator is number of animals examined microscopically for adrenal gland, clitoral gland, kidney, liver,

lung, pituitary gland, thyroid gland, urinary bladder, and uterus; for other tissues, denominator is number of animals necropsied. b Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality c Observed incidence at terminal kill d Beneath the control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values corresponding to pairwise comparisons

between the controls and that exposed group. The life table analysis regards neoplasms in animals dying prior to terminal kill as being (directly or indirectly) the cause of

death. The logistic regression test regards these lesions as nonfatal. The Cochran-Armitage and Fisher exact tests compare directly the overall incidence rates. For all tests,

a negative trend or a lower incidence in an exposure group is indicated by N. e Not applicable; no neoplasms in animal group
Lesions in Female Rats 189

TABLE B4a

Historical Incidence of Hepatocellular Neoplasms in Untreated Female F344/N Ratsa

Incidence in Controls

Study Adenoma Carcinoma Adenoma or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 0/50 0/50 0/50

Acetaminophen 0/50 0/50 0/50

HC Yellow 4 0/50 1/50 1/50

Methylphenidate Hydrochloride 0/50 0/50 0/50

Pentaerythritol Tetranitrate 0/50 0/50 0/50

Quercetin 0/50 0/50 0/50

Turmeric Oleoresin 1/50 0/50 1/50

Overall Historical Incidence

Total 8/1,351 (0.6%) 1/1,351 (0.1%) 9/1,351 (0.7%)

Standard deviation 1.5% 0.4% 1.5%

Range 0%-6% 0%-2% 0%-6%

a Data as of 31 March 1993

TABLE B4b

Historical Incidence of Large Intestine Neoplasms in Untreated Female F344/N Ratsa

Incidence in Controls

Study Adenomatous Polyp Carcinoma Adenomatous Polyp

(Adenoma) (Adenoma) or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 0/50 0/50 0/50

Acetaminophen 0/50 0/50 0/50

HC Yellow 4 0/50 0/50 0/50

Methylphenidate Hydrochloride 0/50 0/50 0/50

Pentaerythritol Tetranitrate 0/50 0/50 0/50

Quercetin 0/50 0/50 0/50

Turmeric Oleoresin 0/50 0/50 0/50

Overall Historical Incidence

Total 0/1,351 (0.0%) 0/1,351 (0.0%) 0/1,351 (0.0%)

a Data as of 31 March 1993; the data include incidences for the colon and rectum.
190 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B4c

Historical Incidence of Renal Tubule Neoplasms in Untreated Female F344/N Ratsa

Incidence in Controls

Study Adenoma Carcinoma Adenoma or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 0/50 0/50 0/50

Acetaminophen 0/50 0/50 0/50

HC Yellow 4 0/49 0/49 0/49

Methylphenidate Hydrochloride 0/50 0/50 0/50

Pentaerythritol Tetranitrate 0/50 0/50 0/50

Quercetin 0/49 0/49 0/49

Turmeric Oleoresin 0/50 0/50 0/50

Overall Historical Incidence

Total 1/1,348 (0.1%) 0/1,348 (0.0%) 1/1,348 (0.1%)

Standard deviation 0.4% 0.4%

Range 0%-2% 0%-2%

a Data as of 31 March 1993

TABLE B4d

Historical Incidence of Urinary Bladder Neoplasms in Untreated Female F344/N Ratsa

Incidence in Controls

Study Papilloma Carcinoma Papilloma or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 0/50 0/50 0/50

Acetaminophen 0/44 0/44 0/44

HC Yellow 4 0/48 0/48 0/48

Methylphenidate Hydrochloride 0/47 0/47 0/47

Pentaerythritol Tetranitrate 0/49 0/49 0/49

Quercetin 1/50 0/50 1/50

Turmeric Oleoresin 0/50 0/50 0/50

Overall Historical Incidence

Total 3/1,334 (0.2%) 0/1,334 (0.0%) 3/1,334 (0.2%)

Standard deviation 0.6% 0.6%

Range 0%-2% 0%-2%

a Data as of 31 March 1993
Lesions in Female Rats 191

TABLE B4e

Historical Incidence of Forestomach Squamous Cell Neoplasms in Untreated Female F344/N Ratsa

Incidence in Controls

Study Papilloma Carcinoma Papilloma or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 0/50 0/50 0/50

Acetaminophen 0/50 0/50 0/50

HC Yellow 4 0/50 0/50 0/50

Methylphenidate Hydrochloride 0/50 0/50 0/50

Pentaerythritol Tetranitrate 0/50 0/50 0/50

Quercetin 0/50 0/50 0/50

Tumeric Oleoresin 0/50 1/50 1/50

Overall Historical Incidence

Total 0/1,351 (0%) 2/1,351 (0.2%) 2/1,351 (0.2%)

Standard deviation 0.5% 0.5%

Range 0%-2% 0%-2%

a Data as of 31 March 1993
192 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B5

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Disposition Summary

Animals initially in study 70 50 70 70

9-Month interim evaluation 10 10 10 10

15-Month interim evaluation 10 10

Early deaths

Moribund 8 5 15 29

Natural deaths 4 3 7 8

Survivors

Died last week of study 1

Terminal sacrifice 37 32 38 12

Missexed 1

Animals examined microscopically 70 50 70 69

9-Month Interim Evaluation

Alimentary System

Intestine large, colon (10) (10) (10) (10)

Parasite metazoan 1 (10%) 4 (40%) 2 (20%) 1 (10%)

Lymphoid tissue, hemorrhage 1 (10%)

Intestine large, rectum (10) (10) (10) (10)

Parasite metazoan 2 (20%) 1 (10%)

Intestine large, cecum (10) (10) (10) (10)

Parasite metazoan 1 (10%)

Liver (10) (10) (10) (10)

Basophilic focus 1 (10%) 1 (10%)

Clear cell focus 1 (10%)

Developmental malformation 1 (10%)

Fatty change 1 (10%) 1 (10%)

Inflammation, chronic active 2 (20%) 2 (20%)

Necrosis, coagulative 4 (40%)

Pigmentation 2 (20%) 6 (60%) 6 (60%)

Bile duct, hyperplasia 1 (10%) 5 (50%) 9 (90%) 3 (30%)

Periportal inflammation, chronic active 6 (60%) 10 (100%) 10 (100%) 8 (80%)

Pancreas (10) (10) (10) (10)

Atrophy 1 (10%) 2 (20%)

Ectopic tissue 1 (10%)

Infiltration cellular, mononuclear cell 3 (30%) 7 (70%) 1 (10%) 1 (10%)

Infiltration cellular, mixed cell 1 (10%)

Salivary glands (10) (10) (10) (10)

Infiltration cellular, mononuclear cell 2 (20%)

Infiltration cellular, lymphocyte 1 (10%)

Stomach, forestomach (10) (10) (10) (10)

Stomach, glandular (10) (10) (10) (10)

Muscularis, mineralization 1 (10%)

Cardiovascular System

Heart (10) (10) (10) (10)

Cardiomyopathy 7 (70%) 8 (80%) 6 (60%) 3 (30%)

a Number of animals examined microscopically at site and number of animals with lesion
Lesions in Female Rats 193

TABLE B5

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

9-Month Interim Evaluation (continued)

Endocrine System

Adrenal cortex (10) (10) (10) (10)

Angiectasis 3 (30%) 1 (10%)

Hyperplasia 1 (10%)

Capsule, fibrosis 1 (10%)

Capsule, inflammation, chronic 1 (10%)

Zona reticularis, hyperplasia 1 (10%)

Pituitary gland (10) (10) (10) (10)

Pars distalis, angiectasis 3 (30%)

Pars distalis, cyst 2 (20%) 1 (10%) 4 (40%) 1 (10%)

Pars distalis, hemorrhage 1 (10%)

Pars distalis, hyperplasia 1 (10%) 1 (10%)

Pars distalis, pars intermedia, cyst 2 (20%)

Pars intermedia, cyst 1 (10%) 2 (20%)

Thyroid gland (10) (10) (10) (10)

Infiltration cellular, mononuclear cell 1 (10%)

Ultimobranchial cyst 1 (10%)

C-cell, hyperplasia 1 (10%)

Follicular cell, hyperplasia 1 (10%)

General Body System

Tissue NOS (1)

Hemorrhage 1 (100%)

Genital System

Clitoral gland (10) (10) (8) (10)

Infiltration cellular, mononuclear cell 1 (10%)

Inflammation, chronic 3 (30%)

Inflammation, chronic active 2 (20%) 3 (30%) 4 (50%)

Ovary (10) (10) (10) (10)

Congestion 1 (10%)

Cyst 1 (10%)

Pigmentation 1 (10%)

Periovarian tissue, cyst 4 (40%) 3 (30%)

Uterus (10) (10) (10) (10)

Decidual reaction 1 (10%)

Hydrometra 1 (10%) 2 (20%) 1 (10%) 5 (50%)

Hematopoietic System

Lymph node (3) (6) (3) (7)

Pigmentation 1 (14%)

Lumbar, hemorrhage 1 (14%)

Lumbar, infiltration cellular, histiocyte 1 (14%)

Lumbar, pigmentation 1 (14%)

Mediastinal, hemorrhage 3 (100%) 1 (17%) 3 (100%) 3 (43%)

Mediastinal, infiltration cellular,

histiocyte 1 (17%) 1 (14%)

Mediastinal, pigmentation 1 (17%) 2 (67%) 2 (29%)
194 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B5

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

9-Month Interim Evaluation (continued)

Hematopoietic System (continued)

Lymph node (continued) (3) (6) (3) (7)

Pancreatic, depletion lymphoid 1 (17%)

Pancreatic, hemorrhage 2 (33%) 1 (33%) 3 (43%)

Pancreatic, infiltration cellular, histiocyte 3 (50%)

Pancreatic, pigmentation 3 (50%) 1 (33%) 1 (14%)

Renal, hemorrhage 1 (17%) 1 (14%)

Renal, infiltration cellular, histiocyte 1 (14%)

Renal, pigmentation 1 (17%) 1 (14%)

Lymph node, mandibular (9) (10) (10) (8)

Hemorrhage 7 (78%) 3 (30%) 5 (50%) 4 (50%)

Infiltration cellular, histiocyte 1 (10%)

Pigmentation 1 (10%)

Lymph node, mesenteric (10) (10) (10) (10)

Hemorrhage 4 (40%) 4 (40%) 2 (20%)

Infiltration cellular, histiocyte 10 (100%) 10 (100%) 10 (100%) 10 (100%)

Pigmentation 7 (70%) 8 (80%)

Spleen (10) (10) (10) (10)

Congestion 1 (10%)

Depletion lymphoid 2 (20%)

Thymus (10) (10) (10) (8)

Congestion 1 (10%)

Depletion lymphoid 1 (10%)

Hemorrhage 1 (10%) 1 (10%)

Respiratory System

Lung (10) (10) (10) (10)

Hemorrhage 1 (10%)

Infiltration cellular, histiocyte 1 (10%) 2 (20%) 3 (30%) 2 (20%)

Alveolar epithelium, hyperplasia 1 (10%)

Artery, mineralization 4 (40%) 3 (30%) 3 (30%) 2 (20%)

Urinary System

Kidney (10) (10) (10) (10)

Hydronephrosis 1 (10%)

Infiltration cellular 1 (10%)

Infiltration cellular, mononuclear cell 9 (90%) 7 (70%) 3 (30%)

Inflammation, chronic 4 (40%)

Mineralization 3 (30%) 1 (10%)

Pigmentation 1 (10%)

Renal tubule, pigmentation 10 (100%) 10 (100%) 10 (100%)

Renal tubule, regeneration 4 (40%) 5 (50%) 7 (70%) 4 (40%)

Transitional epithelium, mineralization 1 (10%) 1 (10%)

Urinary bladder (10) (10) (10) (10)

Infiltration cellular, mononuclear cell 1 (10%)

Transitional epithelium, hyperplasia 2 (20%)

Transitional epithelium, infiltration

 cellular, mononuclear cell 1 (10%)
Lesions in Female Rats 195

TABLE B5

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

9-Month Interim Evaluation (continued)

Systems Examined With No Lesions Observed

Integumentary System

Musculoskeletal System

Nervous Sytem

Special Senses System

15-Month Interim Evaluation

Alimentary System

Intestine large, colon (10)

Parasite metazoan 2 (20%)

Intestine large, rectum (10) (10)

Parasite metazoan 1 (10%)

Intestine large, cecum (10)

Parasite metazoan 2 (20%)

Liver (10) (10)

Abscess 1 (10%)

Basophilic focus 8 (80%) 9 (90%)

Clear cell focus 5 (50%)

Eosinophilic focus 2 (20%)

Fatty change 8 (80%)

Hepatodiaphragmatic nodule 2 (20%) 1 (10%)

Inflammation, chronic, granulomatous 6 (60%) 6 (60%)

Inflammation, chronic active 1 (10%)

Necrosis, coagulative 1 (10%) 4 (40%)

Pigmentation 1 (10%) 10 (100%)

Bile duct, hyperplasia 7 (70%) 10 (100%)

Periportal, inflammation, chronic 10 (100%) 10 (100%)

Mesentery (1)

Inflammation, chronic 1 (100%)

Pigmentation 1 (100%)

Pancreas (10) (1)

Atrophy 1 (10%)

Ectopic tissue 1 (10%)

Inflammation, chronic 9 (90%) 1 (100%)

Pharynx (1)

Hemorrhage 1 (100%)

Salivary glands (10)

Stomach, forestomach (10) (10)

Hyperplasia, basal cell 1 (10%)

Inflammation, chronic 1 (10%)

Stomach, glandular (10) (10)

Muscularis, mineralization 1 (10%)

Cardiovascular System

Heart (10)

Cardiomyopathy 10 (100%)
196 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B5

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

15-Month Interim Evaluation (continued)

Endocrine System

Adrenal cortex (10) (1)

Angiectasis 5 (50%)

Vacuolization cytoplasmic 1 (100%)

Adrenal medulla (10) (1)

Pituitary gland (10)

Pars distalis, cyst 8 (80%)

Pars distalis, hyperplasia 1 (10%)

Pars intermedia, cyst 3 (30%)

Thyroid gland (10)

C-cell, hyperplasia 4 (40%)

Genital System

Clitoral gland (10) (3)

Inflammation, chronic 7 (70%) 3 (100%)

Inflammation, chronic active 1 (10%)

Ovary (10) (4)

Cyst 2 (50%)

Periovarian tissue, cyst 4 (100%)

Uterus (10) (3)

Hydrometra 2 (20%) 3 (100%)

Endometrium, hyperplasia 1 (10%)

Hematopoietic System

Lymph node (1) (3)

Mediastinal, hemorrhage 1 (100%)

Mediastinal, pigmentation 1 (100%)

Pancreatic, infiltration cellular, histiocyte 3 (100%)

Pancreatic, pigmentation 3 (100%)

Lymph node, mandibular (7)

Hemorrhage 1 (14%)

Lymph node, mesenteric (10) (4)

Hemorrhage 1 (25%)

Infiltration cellular, histiocyte 10 (100%) 4 (100%)

Pigmentation 10 (100%) 4 (100%)

Thymus (10) (9)

Cyst 1 (10%) 1 (11%)

Depletion lymphoid 4 (44%)

Pigmentation 1 (11%)

Integumentary System

Mammary gland (7)

Hyperplasia 6 (86%)

Musculoskeletal System

Bone (10)

Osteopetrosis 1 (10%)
Lesions in Female Rats 197

TABLE B5

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

15-Month Interim Evaluation (continued)

Respiratory System

Lung (10) (5)

Hemorrhage 1 (10%)

Infiltration cellular, histiocyte 6 (60%) 5 (100%)

Inflammation, acute 1 (20%)

Alveolus, mineralization 1 (10%)

Artery, mineralization 4 (40%) 2 (40%)

Nose (10)

Submucosa, inflammation, chronic 1 (10%)

Urinary System

Kidney (10) (10)

Nephropathy 10 (100%) 10 (100%)

Renal tubule, hyperplasia 3 (30%)

Renal tubule, mineralization 1 (10%)

Renal tubule, pigmentation 10 (100%)

Transitional epithelium, hyperplasia 3 (30%) 4 (40%)

Transitional epithelium, mineralization 2 (20%) 1 (10%)

Urinary bladder (10) (10)

Inflammation, chronic 1 (10%)

Transitional epithelium, hyperplasia 9 (90%)

Systems Examined With No Lesions Observed

General Body System

Nervous System

Special Senses System

2-Year Study

Alimentary System

Esophagus (50) (40) (57) (48)

Autolysis 1 (2%)

Intestine large, colon (49) (40) (59) (47)

Autolysis 3 (6%) 5 (8%) 3 (6%)

Parasite metazoan 14 (29%) 9 (23%) 7 (12%) 6 (13%)

Intestine large, rectum (49) (40) (60) (47)

Atypia cellular 1 (3%) 6 (10%)

Autolysis 4 (8%) 6 (10%) 1 (2%)

Diverticulum 1 (2%)

Fibrosis 1 (3%)

Inflammation, acute, necrotizing 3 (6%)

Parasite metazoan 10 (20%) 4 (10%) 8 (13%) 8 (17%)

Ulcer 1 (2%)

Epithelium, hyperplasia 3 (5%)
198 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B5

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Alimentary System (continued)

Intestine large, cecum (50) (40) (60) (47)

Autolysis 4 (8%) 1 (3%) 6 (10%) 4 (9%)

Hemorrhage 1 (3%)

Mineralization 1 (2%)

Parasite metazoan 4 (8%) 6 (15%) 5 (8%) 2 (4%)

Intestine small, duodenum (50) (40) (59) (46)

Autolysis 2 (4%) 5 (8%) 1 (2%)

Intestine small, jejunum (48) (40) (59) (46)

Autolysis 3 (6%) 1 (3%) 8 (14%) 3 (7%)

Intestine small, ileum (49) (39) (59) (44)

Autolysis 3 (6%) 1 (3%) 8 (14%) 4 (9%)

Parasite metazoan 1 (3%)

Ulcer 1 (2%)

Liver (50) (40) (60) (48)

Angiectasis 1 (2%) 1 (3%)

Autolysis 4 (7%)

Basophilic focus 39 (78%) 15 (38%) 22 (37%) 16 (33%)

Clear cell focus 3 (6%) 28 (70%) 39 (65%) 17 (35%)

Degeneration 3 (5%)

Eosinophilic focus 7 (14%) 23 (58%) 12 (20%) 1 (2%)

Fatty change 8 (16%) 3 (5%) 5 (10%)

Hematopoietic cell proliferation 10 (17%) 2 (4%)

Hemorrhage 1 (2%)

Hepatodiaphragmatic nodule 8 (16%) 4 (10%) 1 (2%)

Hepatodiaphragmatic nodule, multiple 1 (2%)

Hyperplasia, focal 1 (3%)

Infarct 1 (2%)

Inflammation, chronic active 21 (42%) 16 (40%) 23 (38%) 5 (10%)

Mixed cell focus 1 (2%) 11 (28%) 5 (8%) 4 (8%)

Necrosis, coagulative 3 (6%) 4 (10%) 11 (18%) 9 (19%)

Pigmentation 1 (2%) 19 (48%) 51 (85%) 45 (94%)

Thrombosis 1 (2%) 1 (2%)

Bile duct, hyperplasia 33 (66%) 5 (13%) 56 (93%) 42 (88%)

Centrilobular, hemorrhage 1 (2%)

Centrilobular, necrosis, coagulative 1 (2%)

Periportal, inflammation, chronic active 25 (50%) 2 (5%) 54 (90%) 39 (81%)

Mesentery (4) (6) (1) (4)

Fibrosis 2 (50%) 5 (83%) 1 (100%) 1 (25%)

Inflammation, chronic active 3 (75%) 3 (50%) 1 (100%) 1 (25%)

Necrosis, coagulative 2 (50%) 5 (83%) 1 (100%)

Polyarteritis 1 (17%) 1 (25%)

Pancreas (50) (40) (60) (47)

Atrophy 20 (40%) 17 (43%) 16 (27%) 15 (32%)

Autolysis 1 (2%) 1 (2%) 1 (2%)

Cytoplasmic alteration 1 (3%) 2 (3%) 1 (2%)

Ectopic liver 2 (3%)

Ectopic tissue 1 (2%)

Inflammation, chronic active 29 (58%) 24 (60%) 30 (50%) 22 (47%)

Polyarteritis 2 (5%)

Vacuolization cytoplasmic 5 (10%) 12 (20%) 3 (6%)

Acinus, hyperplasia 1 (2%) 1 (2%)

Duct, hyperplasia 1 (2%)
Lesions in Female Rats 199

TABLE B5

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Alimentary System (continued)

Salivary glands (50) (40) (60) (48)

Parotid gland, inflammation, chronic active 2 (3%) 1 (2%)

Sublingual gland, inflammation, chronic

active 1 (2%)

Submandibular gland, inflammation, chronic

active 1 (2%) 1 (2%)

Stomach, forestomach (49) (40) (60) (47)

Autolysis 1 (2%)

Cyst epithelial inclusion 1 (2%)

Erosion 2 (3%) 3 (6%)

Hyperkeratosis 2 (4%) 7 (18%) 23 (38%) 28 (60%)

Hyperplasia, basal cell 2 (4%) 7 (18%) 35 (58%) 28 (60%)

Hyperplasia, squamous 2 (4%) 7 (18%) 26 (43%) 33 (70%)

Inflammation, chronic active 1 (3%) 13 (22%) 10 (21%)

Ulcer 1 (2%) 2 (5%) 7 (12%) 17 (36%)

Stomach, glandular (50) (40) (60) (48)

Autolysis 2 (4%) 3 (5%) 1 (2%)

Fibrosis 1 (2%)

Inflammation, chronic active 3 (6%) 1 (3%) 6 (10%) 6 (13%)

Necrosis, coagulative 1 (2%) 1 (2%)

Ulcer 1 (2%)

Tongue (1)

Foreign body 1 (100%)

Inflammation, chronic 1 (100%)

Tooth (1) (1) (1)

Cyst 1 (100%)

Cardiovascular System

Heart (50) (40) (60) (49)

Cardiomyopathy 46 (92%) 38 (95%) 54 (90%) 40 (82%)

Endocrine System

Adrenal cortex (47) (40) (59) (47)

Angiectasis 25 (53%) 29 (73%) 40 (68%) 26 (55%)

Autolysis 1 (2%) 3 (6%)

Hyperplasia 3 (6%) 4 (10%) 3 (5%) 1 (2%)

Hypertrophy 1 (2%)

Necrosis, coagulative 1 (2%)

Thrombosis, multiple 1 (2%)

Vacuolization cytoplasmic 17 (36%) 16 (40%) 35 (59%) 19 (40%)

Capsule, hyperplasia 1 (2%)

Adrenal medulla (47) (40) (59) (47)

Autolysis 1 (2%) 1 (2%)

Hyperplasia 11 (23%) 5 (13%) 11 (19%) 14 (30%)

Islets, pancreatic (50) (40) (60) (47)

Autolysis 1 (2%)

Parathyroid gland (43) (37) (57) (38)

Hyperplasia 3 (8%) 1 (2%)
200 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B5

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Endocrine System (continued)

Pituitary gland (50) (39) (60) (47)

Thrombosis 1 (2%)

Pars distalis, angiectasis 1 (2%)

Pars distalis, autolysis 2 (3%) 1 (2%)

Pars distalis, cyst 14 (28%) 20 (51%) 19 (32%) 17 (36%)

Pars distalis, cyst, multiple 3 (6%)

Pars distalis, hyperplasia 12 (24%) 9 (23%) 15 (25%) 8 (17%)

Pars distalis, necrosis, coagulative 1 (2%)

Pars intermedia, cyst 1 (2%)

Rathke's cleft, cyst 1 (2%)

Thyroid gland (50) (40) (60) (49)

Autolysis 2 (4%) 1 (2%) 4 (8%)

Inflammation, chronic 1 (3%)

Inflammation, chronic active 1 (2%)

Ultimobranchial cyst 1 (2%)

C-cell, hyperplasia 7 (14%) 2 (5%) 3 (5%) 1 (2%)

Follicle, cyst 1 (2%)

Follicular cell, hyperplasia 1 (2%)

General Body System

None

Genital System

Clitoral gland (45) (36) (60) (45)

Abscess 2 (4%) 3 (5%) 1 (2%)

Cyst 1 (2%) 1 (3%)

Hyperplasia 1 (3%)

Inflammation, chronic active 34 (76%) 18 (50%) 47 (78%) 31 (69%)

Duct, metaplasia, squamous 1 (2%)

Ovary (50) (40) (60) (47)

Cyst 11 (22%) 2 (5%) 5 (8%) 4 (9%)

Uterus (50) (40) (60) (47)

Abscess 6 (15%) 8 (13%)

Autolysis 1 (2%)

Cyst 1 (2%) 5 (13%) 3 (6%)

Fibrosis 3 (6%) 1 (2%) 1 (2%)

Hydrometra 5 (10%) 4 (10%) 8 (13%) 4 (9%)

Inflammation, acute 1 (2%)

Inflammation, chronic active 1 (2%)

Endometrium, hyperplasia 4 (8%) 9 (23%) 2 (3%) 1 (2%)

Hematopoietic System

Bone marrow (50) (40) (60) (49)

Autolysis 1 (2%)

Myelofibrosis 1 (3%)
Lesions in Female Rats 201

TABLE B5

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Hematopoietic System (continued)

Lymph node (3) (5) (25) (14)

Iliac, infiltration cellular, histiocyte 1 (7%)

Lumbar, infiltration cellular, histiocyte 1 (7%)

Mediastinal, hyperplasia 1 (33%)

Mediastinal, infiltration cellular,

histiocyte 1 (33%) 2 (8%) 2 (14%) 

Pancreatic, hyperplasia 2 (67%) 5 (20%) 1 (7%)

Pancreatic, infiltration cellular, histiocyte 3 (100%) 5 (100%) 21 (84%) 8 (57%)

Renal, infiltration cellular, histiocyte 1 (20%)

Lymph node, mandibular (50) (39) (56) (45)

Autolysis 1 (2%)

Depletion lymphoid 1 (2%)

Hyperplasia 1 (3%) 2 (4%) 1 (2%)

Infiltration cellular, histiocyte 1 (2%) 1 (3%) 1 (2%) 9 (20%)

Inflammation, chronic active 1 (2%)

Lymph node, mesenteric (50) (40) (59) (46)

Autolysis 1 (2%) 2 (4%)

Depletion lymphoid 2 (4%) 1 (3%) 2 (3%) 8 (17%)

Infiltration cellular, histiocyte 49 (98%) 39 (98%) 57 (97%) 45 (98%)

Spleen (50) (40) (60) (48)

Autolysis 1 (2%)

Depletion lymphoid 8 (16%) 1 (3%) 3 (5%) 5 (10%)

Fibrosis 1 (3%) 1 (2%) 1 (2%)

Hematopoietic cell proliferation 1 (2%)

Hyperplasia 1 (2%)

Infiltration cellular, histiocyte 1 (2%)

Thymus (42) (31) (51) (38)

Cyst 1 (3%)

Hemorrhage 1 (3%)

Integumentary System

Mammary gland (49) (34) (50) (41)

Galactocele 1 (2%)

Hyperplasia 41 (84%) 29 (85%) 41 (82%) 26 (63%)

Inflammation, chronic active 1 (2%)

Skin (50) (39) (60) (49)

Abscess 1 (2%) 1 (2%)

Cyst epithelial inclusion 2 (3%)

Foreign body 1 (2%)

Inflammation, chronic active 2 (4%) 6 (10%) 2 (4%)

Foot, acanthosis 1 (2%)

Foot, hyperkeratosis 1 (2%)

Foot, inflammation, chronic active 1 (3%) 1 (2%)

Musculoskeletal System

Bone (50) (40) (60) (49)

Cranium, osteopetrosis 1 (2%)

Femur, osteopetrosis 1 (2%)

Sternum, osteopetrosis 1 (2%) 3 (6%)
202 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE B5

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Nervous System

Brain (50) (40) (60) (49)

Hydrocephalus 1 (2%) 2 (5%)

Infarct 1 (2%)

Respiratory System

Lung (50) (40) (60) (49)

Autolysis 1 (2%)

Fibrosis 2 (3%)

Infiltration cellular, histiocyte 31 (62%) 31 (78%) 50 (83%) 36 (73%)

Inflammation, chronic active 4 (8%) 3 (5%) 2 (4%)

Polyarteritis 1 (2%)

Alveolar epithelium, hyperplasia 1 (2%)

Artery, mineralization 23 (46%) 6 (15%) 20 (33%) 14 (29%)

Bronchiole, epithelium, hyperplasia 1 (2%)

Mediastinum, polyarteritis 2 (5%)

Nose (50) (40) (60) (49)

Autolysis 1 (2%)

Foreign body 1 (2%) 4 (7%)

Inflammation, chronic active 9 (18%) 8 (20%) 13 (22%) 12 (24%)

Metaplasia, squamous 1 (2%) 4 (10%) 4 (7%) 2 (4%)

Arteriole, thrombosis 1 (2%)

Trachea (50) (40) (60) (49)

Autolysis 1 (2%)

Special Senses System

Ear (8) (3)

External ear, inflammation, acute 2 (25%)

Internal ear, inflammation, acute 1 (13%)

Eye (13) (7) (7) (7)

Phthisis bulbi 1 (14%) 1 (14%)

Synechia 1 (8%) 1 (14%)

Cornea, inflammation, acute 1 (14%)

Lens, cataract 1 (8%) 1 (14%)

Retina, degeneration 1 (8%) 3 (43%) 1 (14%)

Zymbal's gland (1) (2)

Autolysis 1 (100%)
Lesions in Female Rats 203

TABLE B5

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

2-Year Study (continued)

Urinary System

Kidney (50) (40) (60) (48)

Abscess 1 (2%)

Autolysis 2 (4%) 3 (5%) 3 (6%)

Cyst 1 (3%) 2 (3%)

Fibrosis 1 (2%)

Hydronephrosis 2 (4%) 2 (3%)

Infarct 1 (3%)

Inflammation, chronic active 1 (2%)

Nephropathy 50 (100%) 39 (98%) 60 (100%) 46 (96%)

Polyarteritis 1 (3%)

Artery, thrombosis 1 (2%)

Papilla, necrosis, coagulative 2 (4%)

Pelvis, inflammation, chronic active 1 (2%) 1 (3%) 4 (7%) 6 (13%)

Pelvis, metaplasia, squamous 3 (5%) 2 (4%)

Renal tubule, hyperplasia 1 (2%) 12 (30%) 23 (38%) 27 (56%)

Renal tubule, hyperplasia, oncocytic 1 (2%)

Renal tubule, inflammation, acute 1 (2%) 7 (18%) 3 (5%)

Renal tubule, inflammation, chronic active 2 (3%) 7 (15%)

Renal tubule, pigmentation 40 (100%) 60 (100%) 48 (100%)

Transitional epithelium, hyperplasia 10 (20%) 16 (40%) 44 (73%) 21 (44%)

Transitional epithelium, mineralization 4 (10%)

Urinary bladder (50) (40) (60) (46)

Autolysis 1 (2%) 2 (3%)

Calculus, gross observation 2 (5%) 1 (2%)

Calculus, microscopic observation only 1 (2%)

Fibrosis 1 (2%)

Hemorrhage 2 (4%)

Hyperplasia, lymphoid 15 (30%) 2 (5%) 3 (5%)

Inflammation, chronic active 1 (2%) 1 (3%) 3 (5%) 1 (2%)

Necrosis 1 (2%)

Fat, proliferation 4 (7%) 2 (4%)

Transitional epithelium, hyperplasia 1 (2%) 2 (5%) 41 (68%) 41 (89%)

Transitional epithelium, metaplasia, squamous 1 (3%) 4 (7%) 8 (17%)
204 1-Amino-2,4-dibromoanthraquinone, NTP TR 383
205

APPENDIX C

SUMMARY OF LESIONS IN MALE MICE

IN THE 2-YEAR FEED STUDY

OF 1-AMINO-2,4-DIBROMOANTHRAQUINONE

TABLE C1 Summary of the Incidence of Neoplasms in Male Mice 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 207

TABLE C2 Individual Animal Tumor Pathology of Male Mice 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 214

TABLE C3 Statistical Analysis of Primary Neoplasms in Male Mice 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 232

TABLE C4a Historical Incidence of Hepatocellular Neoplasms 

in Untreated Male B6C3F1 Mice . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 237

TABLE C4b Historical Incidence of Forestomach Squamous Cell Neoplasms 

in Untreated Male B6C3F1 Mice . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 237

TABLE C4c Historical Incidence of Alveolar/bronchiolar Neoplasms

in Untreated Male B6C3F1 Mice . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 238

TABLE C5 Summary of the Incidence of Nonneoplastic Lesions in Male Mice 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 239
206 1-Amino-2,4-dibromoanthraquinone, NTP TR 383
Lesions in Male Mice 207

TABLE C1

Summary of the Incidence of Neoplasms in Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 10,000 ppm 20,000 ppm

Disposition Summary

Animals initially in study 60 60 60

15-Month interim evaluation 10 9 10

Early deaths

Accidental death 1

Moribund 7 23 21

Natural deaths 3 5 6

Survivors

Terminal sacrifice 40 22 23

Animals examined microscopically 60 60 60

15-Month Interim Evaluation

Alimentary System

Liver (10) (9) (10)

Hepatocellular carcinoma 1 (11%)

Hepatocellular adenoma 2 (22%) 4 (40%)

Stomach, forestomach (9) (9) (10)

Squamous cell carcinoma 1 (10%)

Squamous cell papilloma 5 (50%)

Genital System

Preputial gland (5) (1)

Squamous cell carcinoma 1 (20%)

Respiratory System

Lung (10) (9) (10)

Alveolar/bronchiolar adenoma 3 (33%) 5 (50%)

Special Senses System

Lacrimal gland (1)

Adenoma 1 (100%)

Urinary System

Urinary bladder (10) (9) (10)

Papilloma 1 (10%)

Systems Examined With No Neoplasms Observed

Cardiovascular System

Endocrine System

General Body System

Hematopoietic system

Integumentary System

Musculoskeletal System

Nervous System
208 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C1

Summary of the Incidence of Neoplasms in Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

(continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study

Alimentary System

Gallbladder (46) (47) (42)

Sarcoma, metastatic, stomach, forestomach 1 (2%)

Squamous cell carcinoma, metastatic, stomach, 

 forestomach 1 (2%) 1 (2%)

Intestine large, colon (50) (49) (50)

Mast cell tumor malignant 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 1 (2%)

Intestine large, rectum (48) (49) (48)

Squamous cell carcinoma, metastatic, stomach,

forestomach 1 (2%)

Intestine large, cecum (49) (51) (50)

Adenocarcinoma 1 (2%)

Mast cell tumor malignant 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 1 (2%)

Intestine small, duodenum (50) (50) (46)

Mast cell tumor malignant 1 (2%)

Polyp adenomatous 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 1 (2%)

Intestine small, jejunum (50) (47) (48)

Mast cell tumor malignant 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 1 (2%)

Intestine small, ileum (50) (49) (47)

Adenocarcinoma 1 (2%) 2 (4%)

Histiocytic sarcoma 1 (2%)

Mast cell tumor malignant 1 (2%)

Liver (50) (51) (50)

Fibrosarcoma, metastatic, stomach,

forestomach 1 (2%)

Hemangiosarcoma 2 (4%) 1 (2%)

Hemangiosarcoma, multiple 3 (6%)

Hepatoblastoma 3 (6%) 5 (10%)

Hepatocellular carcinoma 8 (16%) 15 (29%) 12 (24%)

Hepatocellular carcinoma, multiple 1 (2%) 3 (6%) 9 (18%)

Hepatocellular adenoma 4 (8%) 9 (18%) 8 (16%)

Hepatocellular adenoma, multiple 6 (12%) 29 (57%) 31 (62%)

Hepatocholangiocarcinoma, multiple 1 (2%)

Histiocytic sarcoma 1 (2%) 1 (2%)

Sarcoma, metastatic, stomach, forestomach 1 (2%)

Squamous cell carcinoma, metastatic, 

tissue NOS 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 4 (8%) 4 (8%)
Lesions in Male Mice 209

TABLE C1

Summary of the Incidence of Neoplasms in Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

(continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Alimentary System (continued)

Mesentery (2) (7) (9)

Sarcoma, metastatic, stomach, forestomach 1 (14%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 2 (29%) 5 (56%)

Pancreas (50) (50) (48)

Histiocytic sarcoma 1 (2%)

Sarcoma, metastatic, stomach, forestomach 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 1 (2%) 1 (2%)

Salivary glands (50) (51) (50)

Stomach, forestomach (50) (50) (50)

Leiomyosarcoma 1 (2%)

Mast cell tumor malignant 1 (2%)

Squamous cell carcinoma 12 (24%) 13 (26%)

Squamous cell papilloma 11 (22%) 11 (22%)

Squamous cell papilloma, multiple 2 (4%) 5 (10%)

Stomach, glandular (50) (50) (49)

Histiocytic sarcoma 1 (2%)

Mast cell tumor malignant 1 (2%)

Sarcoma 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 3 (6%) 4 (8%)

Tooth (4) (7)

Cardiovascular System

Heart (50) (51) (50)

Endocrine System

Adrenal cortex (50) (51) (50)

Adenoma 2 (4%) 1 (2%)

Sarcoma, metastatic, stomach, forestomach 1 (2%)

Adrenal medulla (50) (50) (50)

Pheochromocytoma benign 1 (2%)

Pituitary gland (43) (45) (47)

Pars distalis, adenoma 2 (4%)

Thyroid gland (49) (50) (49)

C-cell, adenoma 1 (2%)

General Body System

Tissue NOS (1) (1) (1)

Squamous cell carcinoma 1 (100%)

Genital System

Coagulating gland (1) (1) (1)

Squamous cell carcinoma, metastatic, stomach,

forestomach 1 (100%) 1 (100%)
210 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C1

Summary of the Incidence of Neoplasms in Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

(continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Genital System (continued)

Epididymis (49) (51) (50)

Fibrosarcoma, metastatic, stomach,

forestomach 1 (2%)

Histiocytic sarcoma 1 (2%)

Sarcoma, metastatic, stomach, forestomach 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 1 (2%) 1 (2%)

Preputial gland (16) (16) (12)

Squamous cell carcinoma 1 (6%)

Prostate (47) (46) (48)

Histiocytic sarcoma 1 (2%)

Squamous cell carcinoma, metastatic,

preputial gland 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 1 (2%)

Seminal vesicle (49) (50) (45)

Fibrosarcoma, metastatic, stomach,

forestomach 1 (2%)

Histiocytic sarcoma 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 1 (2%)

Testes (50) (50) (50)

Sarcoma, metastatic, stomach, forestomach 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 1 (2%)

Hematopoietic System

Bone marrow (49) (50) (49)

Mast cell tumor malignant 1 (2%)

Lymph node (8) (8) (11)

Lumbar, histiocytic sarcoma 1 (13%)

Mediastinal, histiocytic sarcoma 1 (13%)

Mediastinal, squamous cell carcinoma,

metastatic, stomach, forestomach 2 (25%) 1 (9%)

Pancreatic, histiocytic sarcoma 1 (13%)

Pancreatic, squamous cell carcinoma,

metastatic, stomach, forestomach 1 (9%)

Renal, histiocytic sarcoma 1 (13%)

Lymph node, mandibular (32) (34) (26)

Histiocytic sarcoma 1 (3%)

Mast cell tumor malignant 1 (3%)

Lymph node, mesenteric (46) (47) (47)

Histiocytic sarcoma 1 (2%)

Mast cell tumor malignant 1 (2%)

Sarcoma, metastatic, stomach, forestomach 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 2 (4%)
Lesions in Male Mice 211

TABLE C1

Summary of the Incidence of Neoplasms in Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

(continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Hematopoietic System (continued)

Spleen (50) (51) (50)

Hemangiosarcoma 1 (2%) 1 (2%)

Histiocytic sarcoma 1 (2%)

Mast cell tumor malignant 2 (4%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 1 (2%)

Thymus (37) (35) (33)

Mast cell tumor malignant 1 (3%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 1 (3%)

Integumentary System

Skin (50) (48) (48)

Mast cell tumor malignant 1 (2%)

Squamous cell carcinoma 1 (2%)

Squamous cell papilloma 1 (2%)

Subcutaneous tissue, fibroma 2 (4%) 2 (4%) 1 (2%)

Subcutaneous tissue, fibroma, multiple 1 (2%)

Subcutaneous tissue, fibrosarcoma 3 (6%) 8 (17%) 2 (4%)

Subcutaneous tissue, sarcoma 1 (2%)

Musculoskeletal System

Skeletal muscle (3) (3) (3)

Abdominal, squamous cell carcinoma,

metastatic, stomach, forestomach 2 (67%)

Diaphragm, sarcoma, metastatic, stomach,

forestomach 1 (33%)

Diaphragm, squamous cell carcinoma,

metastatic, stomach, forestomach 2 (67%) 1 (33%)

Nervous System

Brain (50) (51) (50)

Respiratory System

Lung (50) (51) (50)

Alveolar/bronchiolar adenoma 7 (14%) 20 (39%) 15 (30%)

Alveolar/bronchiolar adenoma, multiple 6 (12%) 9 (18%)

Alveolar/bronchiolar carcinoma 3 (6%) 3 (6%) 1 (2%)

Alveolar/bronchiolar carcinoma, multiple 1 (2%)

Hepatocellular carcinoma, metastatic 1 (2%) 2 (4%)

Hepatocellular carcinoma, metastatic, liver 2 (4%)

Histiocytic sarcoma 1 (2%) 1 (2%)

Sarcoma, metastatic, stomach, forestomach 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 3 (6%)

Mediastinum, hemangiosarcoma, metastatic,

spleen 1 (2%)
212 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C1

Summary of the Incidence of Neoplasms in Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

(continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Special Senses System

Ear (1) (1) (1)

Fibrosarcoma 1 (100%)

Harderian gland (2) (2) (1)

Adenoma 2 (100%) 2 (100%) 1 (100%)

Urinary System

Kidney (50) (51) (50)

Histiocytic sarcoma 1 (2%)

Mast cell tumor malignant 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

forestomach 2 (4%)

Urinary bladder (49) (50) (49)

Histiocytic sarcoma 1 (2%)

Mast cell tumor malignant 1 (2%)

Systemic Lesions

Multiple organsb (50) (51) (50)

Histiocytic sarcoma 1 (2%) 1 (2%)

Lymphoma malignant histiocytic 1 (2%) 1 (2%)

Lymphoma malignant lymphocytic 1 (2%) 1 (2%)

Lymphoma malignant mixed 7 (14%) 3 (6%) 5 (10%)

Lymphoma malignant undifferentiated cell 2 (4%) 2 (4%) 1 (2%)

Mesothelioma malignant 1 (2%)

Neoplasm Summary

Total animals with primary neoplasmsc

15-Month interim evaluation 1 4 8

2-Year study 37 49 47

Total primary neoplasms

15-Month interim evaluation 1 6 12

2-Year study 57 162 139

Total animals with benign neoplasms

15-Month interim evaluation 4 7

2-Year study 20 43 43

Total benign neoplasms

15-Month interim evaluation 5 11

2-Year study 25 85 83
Lesions in Male Mice 213

TABLE C1

Summary of the Incidence of Neoplasms in Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone

(continued)

0 ppm 10,000 ppm 20,000 ppm

Neoplasm Summary (continued)

Total animals with malignant neoplasms

15-Month interim evaluation 1 1 1

2-Year study 25 39 39

Total malignant neoplasms

15-Month interim evaluation 1 1 1

2-Year study 32 77 56

Total animals with metastatic neoplasms

2-Year study 4 7 9

Total metastatic neoplasms

2-Year study 4 37 36

a Number of animals examined microscopically at site and number of animals with neoplasm b Number of animals with any tissue examined microscopically c Primary neoplasms: all neoplasms except metastatic neoplasms
214 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm

2344567777777777777777777 

Number of Days on Study 9948220022333333333333333 

2350115533001111111111111

0000000000000000000000000 

Carcass ID Number 2343314636222445555566677 

5554353524134141234512323

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + +

Gallbladder + + A + + + + + + M + + + + + + + + + + + + + + +

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine large, rectum + M + M + + + + + + + + + + + + + + + + + + + + +

Intestine large, cecum + + + M + + + + + + + + + + + + + + + + + + + + +

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 

Polyp adenomatous X 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + +

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 

Hemangiosarcoma, multiple 

Hepatocellular carcinoma X X X X 

Hepatocellular carcinoma, multiple X 

Hepatocellular adenoma X X 

Hepatocellular adenoma, multiple X 

Mesentery + + 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + +

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + +

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + +

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + +

Tooth 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + +

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma X 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + +

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + +

Parathyroid gland + + + + MMMMM++MM +++++++M++MM

Pituitary gland + + I + M + + + + MMM+M +++++++++++

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + +

General Body System 

Tissue NOS 

Genital System

Coagulating gland + 

Epididymis + + I + + + + + + + + + + + + + + + + + + + + + +

Preputial gland M + + + + + + + + + + 

Squamous cell carcinoma 

Prostate + + + + + M + + + + + + + + + + + + + + + + + + + 

Squamous cell carcinoma, metastatic, 

preputial gland 

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + +

Testes + + + + + + + + + + + + + + + + + + + + + + + + +

+: Tissue examined microscopically M: Missing tissue X: Lesion present

A: Autolysis precludes examination I: Insufficient tissue Blank: Not examined
Lesions in Male Mice 215

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

7777777777777777777777777

Number of Days on Study 3333333333333333333333333

1111111122222222222222222

0 0 0 1 1 1 1 1 0 0 0 0 0 0 0 0 0 0 0 1 1 1 1 1 1 Total

Carcass ID Number 7 7 9 0 0 1 2 2 1 1 1 3 8 8 8 8 9 9 9 0 1 1 1 2 2 Tissues/

4 5 5 3 4 4 4 5 1 2 4 1 1 2 3 5 1 3 4 1 1 2 3 2 3 Tumors

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Gallbladder + + + + + M + + + + + + + + + + + + + + + + + M + 46 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Polyp adenomatous 1

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma, multiple X X X 3

Hepatocellular carcinoma X X X X 8

Hepatocellular carcinoma, multiple 1

Hepatocellular adenoma X X 4

Hepatocellular adenoma, multiple X X X X X 6

Mesentery 2

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Tooth + + + + 4

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X 2

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Parathyroid gland + + M + M M + + M + M M + + + + M M + M M + + + + 30 

Pituitary gland + + + + + + + + + + + + + + + + M + + + + + + + + 43 

Thyroid gland + + + + + + + + + + M + + + + + + + + + + + + + + 49 

General Body System 

 

Tissue NOS + 1

Genital System 

Coagulating gland 1

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Preputial gland + + + + + + 16 

Squamous cell carcinoma X 1

Prostate + + + + + M + + + + + + + + + + + + + + + + + + M 47 

Squamous cell carcinoma, metastatic, 

preputial gland X 1

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 50 
216 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

2344567777777777777777777 

Number of Days on Study 9948220022333333333333333 

2350115533001111111111111

0000000000000000000000000 

Carcass ID Number 2343314636222445555566677 

5554353524134141234512323

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node + + + 

Lymph node, mandibular M + + + M + + M M + M + + + + MMM+MM+M++

Lymph node, mesenteric + + M + + + + + + + + + + + + M + + + + + + + + +

Spleen + + + + + + + + + + + + + + + + + + + + + + + + +

Thymus M + + + M M + M + M M M M M + M ++++++++I

Integumentary System 

Mammary gland M M M M + M M M M M M M M M M M M M M M M M M M M

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 

Squamous cell papilloma 

Subcutaneous tissue, fibroma X X 

Subcutaneous tissue, fibrosarcoma X X 

Subcutaneous tissue, sarcoma X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + +

Skeletal muscle + + + 

Nervous System

Brain + + + + + + + + + + + + + + + + + + + + + + + + +

Respiratory System

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 

Alveolar/bronchiolar adenoma X X X 

Alveolar/bronchiolar carcinoma X X X 

Hepatocellular carcinoma, metastatic 

Hepatocellular carcinoma, metastatic, 

liver X 

Nose + + + + + + + M + + + + + + + + + + + M + + +

Trachea + + + + + + + + + + + + + + + + + + + + + + + + +

Special Senses System 

 

Ear 

Fibrosarcoma 

Eye + 

Harderian gland + 

Adenoma X 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + +

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + M +

Systemic Lesions

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Lymphoma malignant histiocytic X 

Lymphoma malignant lymphocytic 

Lymphoma malignant mixed X X X X 

Lymphoma malignant undifferentiated 

cell type X X 
Lesions in Male Mice 217

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

7777777777777777777777777

Number of Days on Study 3333333333333333333333333

1111111122222222222222222

0 0 0 1 1 1 1 1 0 0 0 0 0 0 0 0 0 0 0 1 1 1 1 1 1 Total

Carcass ID Number 7 7 9 0 0 1 2 2 1 1 1 3 8 8 8 8 9 9 9 0 1 1 1 2 2 Tissues/

4 5 5 3 4 4 4 5 1 2 4 1 1 2 3 5 1 3 4 1 1 2 3 2 3 Tumors

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Lymph node + + + + + 8

Lymph node, mandibular M + M M + + + + M + + + + + M M + + + + M + + + + 32 

Lymph node, mesenteric + + + + + + + + M + + + + + + M + + + + + + + + + 46 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Thymus + + + + + + I + + + + + + + + + + + + + + + + M + 37 

Integumentary System 

Mammary gland M M M M M M M M M M M M M M M M M M M M M M M M M 1

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Squamous cell papilloma X 1

Subcutaneous tissue, fibroma 2

Subcutaneous tissue, fibrosarcoma X 3

Subcutaneous tissue, sarcoma 1

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Skeletal muscle 3

Nervous System

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Respiratory System

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Alveolar/bronchiolar adenoma X X X X 7

Alveolar/bronchiolar carcinoma 3

Hepatocellular carcinoma, metastatic X 1

Hepatocellular carcinoma, metastatic, 

liver X 2

Nose + + + + + + M + + + + + + + + + + + + + M + + M + 43 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Special Senses System 

Ear + 1

Fibrosarcoma X 1

Eye + 2

Harderian gland + 2

Adenoma X 2

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymphoma malignant histiocytic 1

Lymphoma malignant lymphocytic X 1

Lymphoma malignant mixed X X X 7

Lymphoma malignant undifferentiated 

cell type 2
218 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm

1444455555556666666666666 

Number of Days on Study 5155600577881111334445778 

2417415028231238332784339

1212121212212111112111111 

Carcass ID Number 8360903162270648594985334 

5551555445454254545333344

Alimentary System 

Esophagus + + + + M + + M + + + + + + + + + + M + + + + +

Gallbladder + + + + + + M + + M + + + + + + + + + + M + + + + 

Sarcoma, metastatic, stomach, 

forestomach X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + M 

Mast cell tumor malignant X 

Intestine large, rectum + + + + M + + + + M + + + + + + + + + + + + + + +

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 

Mast cell tumor malignant X 

Intestine small, duodenum + + + + + + + + + + + + + + A + + + + + + + + + + 

Mast cell tumor malignant X 

Intestine small, jejunum A + + + + + + + + + + + + M A + + + + + + + + + M 

Mast cell tumor malignant X 

Intestine small, ileum + + + + + + + + + + + + + + A M + + + + + + + + + 

Adenocarcinoma 

Histiocytic sarcoma X 

Mast cell tumor malignant X 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 

Hemangiosarcoma X 

Hepatoblastoma X X X 

Hepatocellular carcinoma X X X X X 

Hepatocellular carcinoma, multiple X 

Hepatocellular adenoma X X X X X X 

Hepatocellular adenoma, multiple X X X X X X X X X X 

Histiocytic sarcoma X 

Sarcoma, metastatic, stomach, 

forestomach X 

Squamous cell carcinoma, metastatic, 

tissue NOS X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X X X

Mesentery + + + + + + 

Sarcoma, metastatic, stomach, 

forestomach X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X 

Pancreas + + + + + + + + + + M + + + + + + + + + + + + + + 

Histiocytic sarcoma X 

Sarcoma, metastatic, stomach, 

forestomach X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + +
Lesions in Male Mice 219

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

77777777777777777777777777 

Number of Days on Study 00112233333333333333333333 

69579900000000000000000000 

2 2 2 2 1 1 1 1 1 1 1 1 1 1 1 1 1 2 2 2 2 2 2 2 2 2 Total

Carcass ID Number 1 0 3 2 3 3 4 4 5 5 6 7 7 7 8 9 9 0 2 2 3 3 4 4 4 4 Tissues/

3 3 4 3 1 2 1 3 1 2 1 1 2 4 2 1 2 2 1 2 1 3 1 2 3 4 Tumors

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Gallbladder + + + + + + + + + + + + + + + + M + + + + + + + + + 47 

Sarcoma, metastatic, stomach, 

forestomach 1

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Mast cell tumor malignant 1

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + + 51 

Mast cell tumor malignant 1

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Mast cell tumor malignant 1

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Mast cell tumor malignant 1

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Adenocarcinoma X 1

Histiocytic sarcoma 1

Mast cell tumor malignant 1

Liver + + + + + + + + + + + + + + + + + + + + + + + + + + 51 

Hemangiosarcoma X 2

Hepatoblastoma 3

Hepatocellular carcinoma X X X X X X X X X X 15 

Hepatocellular carcinoma, multiple X X 3

Hepatocellular adenoma X X X 9

Hepatocellular adenoma, multiple X X X X X X X X X X X X X X X X X X X 29 

Histiocytic sarcoma 1

Sarcoma, metastatic, stomach, 

forestomach 1

Squamous cell carcinoma, metastatic, 

tissue NOS 1

Squamous cell carcinoma, metastatic, 

stomach, forestomach 4

Mesentery + 7

Sarcoma, metastatic, stomach, 

forestomach 1

Squamous cell carcinoma, metastatic, 

stomach, forestomach 2

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1

Sarcoma, metastatic, stomach, 

forestomach 1

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + + 51 
220 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

1444455555556666666666666 

Number of Days on Study 5155600577881111334445778 

2417415028231238332784339

1212121212212111112111111 

Carcass ID Number 8360903162270648594985334 

5551555445454254545333344

Alimentary System (continued) 

Stomach, forestomach + + + + + + + + + + + + M + + + + + + + + + + + + 

Leiomyosarcoma X 

Mast cell tumor malignant X 

Squamous cell carcinoma X X X X X X X 

Squamous cell papilloma X X 

Squamous cell papilloma, multiple 

Stomach, glandular + + + + + + + + + + + + I + + + + + + + + + + + + 

Histiocytic sarcoma X 

Mast cell tumor malignant X 

Sarcoma X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X X

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + +

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma 

Sarcoma, metastatic, stomach, 

forestomach X 

Adrenal medulla + + + + + M + + + + + + + + + + + + + + + + + + +

Islets, pancreatic + + + + + + + + + + M + + + + + + + + + + + + + +

Parathyroid gland M + M + + M + + M + + + + M + + M M + M M + + +

Pituitary gland + + + + + + + M + + + + + + + + + + + + + + + M 

Pars distalis, adenoma X 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + +

General Body System 

Tissue NOS + 

Squamous cell carcinoma X 

Genital System

Coagulating gland + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X 

Sarcoma, metastatic, stomach, 

forestomach X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X 

Penis 

Preputial gland + + + + + + + + + + 

Prostate + + + + + + + + + + + + MMM+++++++++

Histiocytic sarcoma X 

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 

Sarcoma, metastatic, stomach, 

forestomach X 
Lesions in Male Mice 221

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7

Number of Days on Study 0 0 1 1 2 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3

6 9 5 7 9 9 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

2 2 2 2 1 1 1 1 1 1 1 1 1 1 1 1 1 2 2 2 2 2 2 2 2 2 Total

Carcass ID Number 1 0 3 2 3 3 4 4 5 5 6 7 7 7 8 9 9 0 2 2 3 3 4 4 4 4 Tissues/

3 3 4 3 1 2 1 3 1 2 1 1 2 4 2 1 2 2 1 2 1 3 1 2 3 4 Tumors

Alimentary System (continued)

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Leiomyosarcoma 1

Mast cell tumor malignant 1

Squamous cell carcinoma X X X X X 12 

Squamous cell papilloma X X X X X X X X X 11 

Squamous cell papilloma, multiple X X 2

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1

Mast cell tumor malignant 1

Sarcoma 1

Squamous cell carcinoma, metastatic,

stomach, forestomach 3

Cardiovascular System

Heart + + + + + + + + + + + + + + + + + + + + + + + + + + 51 

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + + 51 

Adenoma X 1

Sarcoma, metastatic, stomach,

forestomach 1

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Parathyroid gland + + + + + + + M + + + + + + M + M + + + + + + M + + 37 

Pituitary gland + + + + + + + + + + + + + M M + + + + + M + + + + + 45 

Pars distalis, adenoma X 2

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + + 50 

General Body System

Tissue NOS 1

Squamous cell carcinoma 1

Genital System

Coagulating gland 1

Squamous cell carcinoma, metastatic,

stomach, forestomach 1

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + + 51 

Histiocytic sarcoma 1

Sarcoma, metastatic, stomach,

forestomach 1

Squamous cell carcinoma, metastatic,

stomach, forestomach 1

Penis + 1

Preputial gland + M + + + + + 16 

Prostate + + + I + + + + + + + + + + + + + + + + + + + + + + 46 

Histiocytic sarcoma 1

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Sarcoma, metastatic, stomach,

forestomach 1
222 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

1 4 4 4 4 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 

Number of Days on Study 5 1 5 5 6 0 0 5 7 7 8 8 1 1 1 1 3 3 4 4 4 5 7 7 8 

2 4 1 7 4 1 5 0 2 8 2 3 1 2 3 8 3 3 2 7 8 4 3 3 9

1 2 1 2 1 2 1 2 1 2 2 1 2 1 1 1 1 1 2 1 1 1 1 1 1 

Carcass ID Number 8 3 6 0 9 0 3 1 6 2 2 7 0 6 4 8 5 9 4 9 8 5 3 3 4 

5 5 5 1 5 5 5 4 4 5 4 5 4 2 5 4 5 4 5 3 3 3 3 4 4

Hematopoietic System 

Bone marrow M + + + + + + + + + + + + + + + + + + + + + + + + 

Mast cell tumor malignant

Lymph node + + + + + + 

Lumbar, histiocytic sarcoma X 

Mediastinal, histiocytic sarcoma X 

Mediastinal, squamous cell carcinoma, 

metastatic, stomach, forestomach X X 

Pancreatic, histiocytic sarcoma X 

Renal, histiocytic sarcoma X

Lymph node, mandibular + + + + M + M + + + + + M + + M M M + M M M + + M 

Histiocytic sarcoma X 

Mast cell tumor malignant X

Lymph node, mesenteric M + + + + + + + + + + + + + + + + + + + + + + + M 

Histiocytic sarcoma X 

Mast cell tumor malignant X 

Sarcoma, metastatic, stomach, 

forestomach X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 

Hemangiosarcoma 

Histiocytic sarcoma X 

Mast cell tumor malignant X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X

Thymus + M + M + M M M + + + M M + + M + + + + M + M 

Mast cell tumor malignant 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X

Integumentary System 

Mammary gland M M M M M M M M M M M M M M M M M M M M M M M M

Skin M M M + + + + + + + + + + + + + + + + + + + + + + 

Mast cell tumor malignant X 

Squamous cell carcinoma X 

Subcutaneous tissue, fibroma

Subcutaneous tissue, fibroma, 

multiple 

Subcutaneous tissue, fibrosarcoma X X X X X

Musculoskeletal System 

Bone M + + + + + + + + + + + + + + + + + + + + + + + +

Skeletal muscle + + + 

Diaphragm, sarcoma, metastatic, 

stomach, forestomach X 

Diaphragm, squamous cell carcinoma, 

metastatic, stomach, forestomach X X
Lesions in Male Mice 223

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7

Number of Days on Study 0 0 1 1 2 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3

6 9 5 7 9 9 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

2 2 2 2 1 1 1 1 1 1 1 1 1 1 1 1 1 2 2 2 2 2 2 2 2 2 Total

Carcass ID Number 1 0 3 2 3 3 4 4 5 5 6 7 7 7 8 9 9 0 2 2 3 3 4 4 4 4 Tissues/

3 3 4 3 1 2 1 3 1 2 1 1 2 4 2 1 2 2 1 2 1 3 1 2 3 4 Tumors

Hematopoietic System

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Mast cell tumor malignant X 1

Lymph node + + 8

Lumbar, histiocytic sarcoma 1

Mediastinal, histiocytic sarcoma 1

Mediastinal, squamous cell carcinoma,

metastatic, stomach, forestomach 2

Pancreatic, histiocytic sarcoma 1

Renal, histiocytic sarcoma 1

Lymph node, mandibular M + M + M M + + + + M I + + + M + + + + + + + + + + 34 

Histiocytic sarcoma 1

Mast cell tumor malignant 1

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + M + + 47 

Histiocytic sarcoma 1

Mast cell tumor malignant 1

Sarcoma, metastatic, stomach,

forestomach 1

Squamous cell carcinoma, metastatic,

stomach, forestomach 2

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + + 51 

Hemangiosarcoma X 1

Histiocytic sarcoma 1

Mast cell tumor malignant X 2

Squamous cell carcinoma, metastatic,

stomach, forestomach 1

Thymus M + M + + + + + + + + + + M + + + + + + + + + + + M 35 

Mast cell tumor malignant X 1

Squamous cell carcinoma, metastatic,

stomach, forestomach 1

Integumentary System

Mammary gland M M M M M M M M M M M M M M M M M M M + M M M M M M 1

Skin + + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Mast cell tumor malignant 1

Squamous cell carcinoma 1

Subcutaneous tissue, fibroma X X 2

Subcutaneous tissue, fibroma,

multiple X 1

Subcutaneous tissue, fibrosarcoma X X X 8

Musculoskeletal System

Bone + + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Skeletal muscle 3

Diaphragm, sarcoma, metastatic,

stomach, forestomach 1

Diaphragm, squamous cell carcinoma,

metastatic, stomach, forestomach 2
224 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

1 4 4 4 4 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 

Number of Days on Study 5 1 5 5 6 0 0 5 7 7 8 8 1 1 1 1 3 3 4 4 4 5 7 7 8 

2 4 1 7 4 1 5 0 2 8 2 3 1 2 3 8 3 3 2 7 8 4 3 3 9

1 2 1 2 1 2 1 2 1 2 2 1 2 1 1 1 1 1 2 1 1 1 1 1 1 

Carcass ID Number 8 3 6 0 9 0 3 1 6 2 2 7 0 6 4 8 5 9 4 9 8 5 3 3 4 

5 5 5 1 5 5 5 4 4 5 4 5 4 2 5 4 5 4 5 3 3 3 3 4 4

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + +

Respiratory System 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 

Alveolar/bronchiolar adenoma X X X X X X X X X X 

Alveolar/bronchiolar adenoma, 

multiple X X 

Alveolar/bronchiolar carcinoma 

Alveolar/bronchiolar carcinoma, 

multiple X 

Histiocytic sarcoma X 

Sarcoma, metastatic, stomach, 

forestomach X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X X

Nose M + + + + + + + + + + + + + + + + + + + + + M + +

Trachea + + + + + + + + + + + + + + + + + + + + + + + +

Special Senses System 

Ear

Eye

Harderian gland + 

Adenoma X

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X 

Mast cell tumor malignant X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X

Urinary bladder + + + + I + + + + + + + + + + + + + + + + + + + +

Histiocytic sarcoma X 

Mast cell tumor malignant X

Systemic Lesions

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X 

Lymphoma malignant histiocytic 

Lymphoma malignant mixed X 

Lymphoma malignant undifferentiated 

cell type X 

Mesothelioma malignant X
Lesions in Male Mice 225

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

10,000 ppm (continued)

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7

Number of Days on Study 0 0 1 1 2 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3

6 9 5 7 9 9 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0

2 2 2 2 1 1 1 1 1 1 1 1 1 1 1 1 1 2 2 2 2 2 2 2 2 2 Total

Carcass ID Number 1 0 3 2 3 3 4 4 5 5 6 7 7 7 8 9 9 0 2 2 3 3 4 4 4 4 Tissues/

3 3 4 3 1 2 1 3 1 2 1 1 2 4 2 1 2 2 1 2 1 3 1 2 3 4 Tumors

Nervous System

Brain + + + + + + + + + + + + + + + + + + + + + + + + + + 51 

Respiratory System

Lung + + + + + + + + + + + + + + + + + + + + + + + + + + 51 

Alveolar/bronchiolar adenoma X X X X X X X X X X 20 

Alveolar/bronchiolar adenoma,

multiple X X X X 6

Alveolar/bronchiolar carcinoma X X X 3

Alveolar/bronchiolar carcinoma, 

multiple 1

Histiocytic sarcoma 1

Sarcoma, metastatic, stomach,

forestomach 1

Squamous cell carcinoma, metastatic,

stomach, forestomach 3

Nose + + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Special Senses System

Ear + 1

Eye + 1

Harderian gland + 2

Adenoma X 2

Urinary System

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + + 51 

Histiocytic sarcoma 1

Mast cell tumor malignant 1

Squamous cell carcinoma, metastatic,

stomach, forestomach 2

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1

Mast cell tumor malignant 1

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + + 51 

Histiocytic sarcoma 1

Lymphoma malignant histiocytic X 1

Lymphoma malignant mixed X X 3

Lymphoma malignant undifferentiated

cell type X 2

Mesothelioma malignant 1
226 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

20,000 ppm

1122455555666666666666677 

Number of Days on Study 0344811239011123446688902 

9689427353638813582599692

3233233332223323223323333 

Carcass ID Number 5635642126973694862385026 

5454351552554544514341534

Alimentary System 

Esophagus + + + + + M + + + + + + + + + + + + + + + + + + +

Gallbladder A + M + + M + + + + + + + M + + + + + + M + + + + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + M + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenocarcinoma X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X

Intestine small, duodenum M A + + + + + + + + + + + + + + + + + A + + M + + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X

Intestine small, jejunum A A + + + + + + + + + + + + + + + + + + + + + + + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X

Intestine small, ileum M A + + + + + M + + + + + + + + + + + + + + + + + 

Adenocarcinoma X 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 

Fibrosarcoma, metastatic, stomach, 

forestomach X 

Hemangiosarcoma X 

Hepatoblastoma X X X 

Hepatocellular carcinoma X X X X X X X X 

Hepatocellular carcinoma, multiple X X 

Hepatocellular adenoma X X X X X 

Hepatocellular adenoma, multiple X X X X X X X X X X X 

Hepatocholangiocarcinoma, multiple 

Histiocytic sarcoma X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X X

Mesentery + + + + + + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X X

Pancreas + + + + + + + + + + + M + + + + + + + + + + + + + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + +

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 

Squamous cell carcinoma X X X X X X X X

Squamous cell papilloma X X X 

Squamous cell papilloma, multiple X 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X X
Lesions in Male Mice 227

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

20,000 ppm (continued)

7777777777777777777777777

Number of Days on Study 2222222222222222222222222

4699999999999999999999999

3 3 2 2 2 2 2 2 2 2 2 2 2 2 3 3 3 3 3 3 3 3 3 3 3 Total

Carcass ID Number 4 0 5 5 5 5 7 7 7 7 8 8 9 9 0 0 1 1 1 1 2 3 4 6 6 Tissues/

3 4 2 3 4 5 1 2 3 4 1 3 1 2 1 3 1 2 3 4 2 1 1 1 2 Tumors

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Gallbladder + + + + + + + + + + + + M + + + M + + + + + + + 42 

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Intestine large, rectum M + + + + + + + + + + + + + + + + + + + + + + + + 48 

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenocarcinoma 1

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 46 

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Adenocarcinoma X 2

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Fibrosarcoma, metastatic, stomach, 

forestomach 1

Hemangiosarcoma 1

Hepatoblastoma X X 5

Hepatocellular carcinoma X X X X 12 

Hepatocellular carcinoma, multiple X X X X X X X 9

Hepatocellular adenoma X X X 8

Hepatocellular adenoma, multiple X X X X X X X X X X X X X X X X X X X X 31 

Hepatocholangiocarcinoma, multiple X 1

Histiocytic sarcoma 1

Squamous cell carcinoma, metastatic, 

stomach, forestomach X 4

Mesentery + + + 9

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X 5

Pancreas M + + + + + + + + + + + + + + + + + + + + + + + + 48 

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Squamous cell carcinoma X X X X X 13 

Squamous cell papilloma X X X X X X X X 11 

Squamous cell papilloma, multiple X X X X 5

Stomach, glandular M + + + + + + + + + + + + + + + + + + + + + + + + 49 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X 4
228 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

20,000 ppm (continued)

1122455555666666666666677 

Number of Days on Study 0344811239011123446688902 

9689427353638813582599692

3233233332223323223323333 

Carcass ID Number 5635642126973694862385026 

5454351552554544514341534

Alimentary System (continued) 

Tongue + 

Tooth + + +

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + +

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + +

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma benign X 

Islets, pancreatic + + + + + + + + + + + M + + + + + + + + + + + + +

Parathyroid gland + + + + M + M + + + + M + + + + + + + M M + + + M

Pituitary gland + + + + + + + + + + + + + + + + + + M + + + + M +

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + M 

C-cell, adenoma 

General Body System 

Tissue NOS 

Genital System

Coagulating gland + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 

Fibrosarcoma, metastatic, stomach, 

forestomach X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X

Penis 

Preputial gland + + + + + +

Prostate + + + + + + + + + M + + + + + + + + + + + + + + + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + 

Fibrosarcoma, metastatic, stomach, 

forestomach X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X

Testes + + + + + + + + + + + + + + + + + + + + + + + + +

Squamous cell carcinoma, metastatic, 

stomach, forestomach X

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node + + + + + + 

Mediastinal, squamous cell carcinoma, 

metastatic, stomach, forestomach X 

Pancreatic, squamous cell carcinoma, 

metastatic, stomach, forestomach 

Lymph node, mandibular M M + M + + M M M M M ++++M+MM+MM+++

Lymph node, mesenteric M + + + + + + + + + + + + + + + + + + + + + + + +
Lesions in Male Mice 229

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

20,000 ppm (continued)

7777777777777777777777777

Number of Days on Study 2222222222222222222222222

4699999999999999999999999

3 3 2 2 2 2 2 2 2 2 2 2 2 2 3 3 3 3 3 3 3 3 3 3 3 Total

Carcass ID Number 4 0 5 5 5 5 7 7 7 7 8 8 9 9 0 0 1 1 1 1 2 3 4 6 6 Tissues/

3 4 2 3 4 5 1 2 3 4 1 3 1 2 1 3 1 2 3 4 2 1 1 1 2 Tumors

Alimentary System (continued) 

Tongue 1

Tooth + + + + 7

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pheochromocytoma benign 1

Islets, pancreatic M + + + + + + + + + + + + + + + + + + + + + + + + 48 

Parathyroid gland M M + + + + + + M + + + M + + + + M + + M + + + + 38 

Pituitary gland + + + + + + + + + + + + M + + + + + + + + + + + + 47 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

C-cell, adenoma X 1

General Body System 

Tissue NOS + 1

Genital System

Coagulating gland 1

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Fibrosarcoma, metastatic, stomach, 

forestomach 1

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Penis + + 2

Preputial gland + + + + M + + 12 

Prostate + M + + + + + + + + + + + + + + + + + + + + + + + 48 

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Seminal vesicle M M + + + + + + + + + + + + + + + + + + + + + + + 45 

Fibrosarcoma, metastatic, stomach, 

forestomach 1

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + 49 

Lymph node + + + + + 11 

Mediastinal, squamous cell carcinoma, 

metastatic, stomach, forestomach 1

Pancreatic, squamous cell carcinoma, 

metastatic, stomach, forestomach X 1

Lymph node, mandibular + + + + + + M M + M + M M + + + M I M M M + + + M 26 

Lymph node, mesenteric M + + + + + + + + + + + + + + + + + + + + + + + M 47 
230 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

20,000 ppm (continued)

1122455555666666666666677 

Number of Days on Study 0344811239011123446688902 

9689427353638813582599692

3233233332223323223323333 

Carcass ID Number 5635642126973694862385026 

5454351552554544514341534

Hematopoietic System (continued) 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 

Hemangiosarcoma X 

Thymus + M + + M + M + M + + + + + + MMMM++MM+M

Integumentary System 

Mammary gland M M M M M M M M M M M M M M M M M M M M M M M M M

Skin + + + + + + + + + + + + + + + + + + + + + + M + + 

Subcutaneous tissue, fibroma X 

Subcutaneous tissue, fibrosarcoma X X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + +

Skeletal muscle + + + 

Abdominal, squamous cell carcinoma, 

metastatic, stomach, forestomach X X 

Diaphragm, squamous cell carcinoma, 

metastatic, stomach, forestomach X

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + +

Respiratory System

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 

Alveolar/bronchiolar adenoma X X X X X X X X 

Alveolar/bronchiolar adenoma, 

multiple X X X 

Alveolar/bronchiolar carcinoma X 

Hepatocellular carcinoma, metastatic X 

Histiocytic sarcoma X 

Mediastinum, hemangiosarcoma, 

metastatic, spleen X 

Nose + + M + + + + + + + + + + + + + + + + + + + + + +

Trachea + + + + + + + + + + + + + + + + + + + + + + + + +

Special Senses System 

Ear 

Harderian gland 

Adenoma 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + +

Urinary bladder + M + + + + + + + + + + + + + + + + + + + + + + +

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X 

Lymphoma malignant lymphocytic X 

Lymphoma malignant mixed X 

Lymphoma malignant undifferentiated 

cell type X 
Lesions in Male Mice 231

TABLE C2

Individual Animal Tumor Pathology of Male Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

20,000 ppm (continued)

7777777777777777777777777

Number of Days on Study 2222222222222222222222222

4699999999999999999999999

3 3 2 2 2 2 2 2 2 2 2 2 2 2 3 3 3 3 3 3 3 3 3 3 3 Total

Carcass ID Number 4 0 5 5 5 5 7 7 7 7 8 8 9 9 0 0 1 1 1 1 2 3 4 6 6 Tissues/

3 4 2 3 4 5 1 2 3 4 1 3 1 2 1 3 1 2 3 4 2 1 1 1 2 Tumors

Hematopoietic System (continued) 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma 1

Thymus M + + + M + + + + + + + + I + + + M M + M + + + + 33 

Integumentary System 

Mammary gland M M M M M M M M M M M M M M M M M M M M M M M M M 

Skin + + + + + + + + + + + + + + + + + + + + + + M + + 48 

Subcutaneous tissue, fibroma 1

Subcutaneous tissue, fibrosarcoma 2

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + 49 

Skeletal muscle 3

Abdominal, squamous cell carcinoma, 

metastatic, stomach, forestomach 2

Diaphragm, squamous cell carcinoma, 

metastatic, stomach, forestomach 1

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Respiratory System

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Alveolar/bronchiolar adenoma X X X X X X X 15 

Alveolar/bronchiolar adenoma, 

multiple X X X X X X 9

Alveolar/bronchiolar carcinoma 1

Hepatocellular carcinoma, metastatic X 2

Histiocytic sarcoma 1

Mediastinum, hemangiosarcoma, 

metastatic, spleen 1

Nose + + + + + + + M + + + + + + + + + + + + + + + + + 48 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Special Senses System 

Ear + 1

Harderian gland + 1

Adenoma X 1

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1

Lymphoma malignant lymphocytic 1

Lymphoma malignant mixed X X X X 5

Lymphoma malignant undifferentiated 

cell type 1
232 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C3

Statistical Analysis of Primary Neoplasms in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

0 ppm 10,000 ppm 20,000 ppm

Liver: Hemangiosarcoma

Overall ratea 3/50 (6%) 2/51 (4%) 1/50 (2%)

Adjusted rateb 7.5% 6.5% 2.2%

Terminal ratec 3/40 (8%) 1/22 (5%) 0/23 (0%)

First incidence (days) 729 (T) 414 484

Life table testd P=0.366N P=0.658 P=0.461N

Logistic regression testd P=0.189N P=0.481N P=0.301N

Cochran-Armitage testd P=0.221N

Fisher exact testd P=0.491N P=0.309N

Liver: Hepatocellular Adenoma

Overall rate 10/50 (20%) 38/51 (75%) 39/50 (78%)

Adjusted rate 24.3% 94.7% 95.0%

Terminal rate 9/40 (23%) 20/22 (91%) 21/23 (91%)

First incidence (days) 723 451 484

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001

Liver: Hepatocellular Carcinoma

Overall rate 9/50 (18%) 18/51 (35%) 21/50 (42%)

Adjusted rate 21.1% 58.1% 58.4%

Terminal rate 7/40 (18%) 10/22 (45%) 9/23 (39%)

First incidence (days) 445 505 535

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P=0.002 P=0.017 P=0.003

Cochran-Armitage test P=0.007

Fisher exact test P=0.040 P=0.008

Liver: Hepatocellular Adenoma or Carcinoma

Overall rate 18/50 (36%) 43/51 (84%) 42/50 (84%)

Adjusted rate 41.7% 97.7% 97.7%

Terminal rate 15/40 (38%) 21/22 (95%) 22/23 (96%)

First incidence (days) 445 451 484

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001

Liver: Hepatoblastoma

Overall rate 0/50 (0%) 3/51 (6%) 5/50 (10%)

Adjusted rate 0.0% 8.2% 16.8%

Terminal rate 0/40 (0%) 0/22 (0%) 2/23 (9%)

First incidence (days) )e 572 633

Life table test P=0.011 P=0.090 P=0.011

Logistic regression test P=0.021 P=0.151 P=0.026

Cochran-Armitage test P=0.022

Fisher exact test P=0.125 P=0.028
Lesions in Male Mice 233

TABLE C3

Statistical Analysis of Primary Neoplasms in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

Liver: Hepatocellular Carcinoma or Hepatoblastoma

Overall rate 9/50 (18%) 20/51 (39%) 24/50 (48%)

Adjusted rate 21.1% 60.2% 65.3%

Terminal rate 7/40 (18%) 10/22 (45%) 11/23 (48%)

First incidence (days) 445 505 535

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P=0.007 P<0.001

Cochran-Armitage test P=0.001

Fisher exact test P=0.016 P=0.001

Liver: Hepatocellular Adenoma, Hepatocellular Carcinoma, or Hepatoblastoma

Overall rate 18/50 (36%) 43/51 (84%) 42/50 (84%)

Adjusted rate 41.7% 97.7% 97.7%

Terminal rate 15/40 (38%) 21/22 (95%) 22/23 (96%)

First incidence (days) 445 451 484

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001

Lung: Alveolar/bronchiolar Adenoma

Overall rate 7/50 (14%) 26/51 (51%) 24/50 (48%)

Adjusted rate 16.8% 71.0% 66.5%

Terminal rate 6/40 (15%) 12/22 (55%) 12/23 (52%)

First incidence (days) 445 578 248

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001

Lung: Alveolar/bronchiolar Carcinoma

Overall rate 3/50 (6%) 4/51 (8%) 1/50 (2%)

Adjusted rate 6.9% 15.9% 3.0%

Terminal rate 2/40 (5%) 2/22 (9%) 0/23 (0%)

First incidence (days) 393 673 648

Life table test P=0.454N P=0.259 P=0.442N

Logistic regression test P=0.259N P=0.512 P=0.251N

Cochran-Armitage test P=0.252N

Fisher exact test P=0.511 P=0.309N

Lung: Alveolar/bronchiolar Adenoma or Carcinoma

Overall rate 10/50 (20%) 28/51 (55%) 25/50 (50%)

Adjusted rate 23.3% 75.0% 67.5%

Terminal rate 8/40 (20%) 13/22 (59%) 12/23 (52%)

First incidence (days) 393 578 248

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P=0.002

Cochran-Armitage test P=0.002

Fisher exact test P<0.001 P=0.002
234 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C3

Statistical Analysis of Primary Neoplasms in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

Skin (Subcutaneous Tissue): Fibroma

Overall rate 2/50 (4%) 3/51 (6%) 1/50 (2%)

Adjusted rate 5.0% 11.9% 3.8%

Terminal rate 2/40 (5%) 1/22 (5%) 0/23 (0%)

First incidence (days) 729 (T) 706 722

Life table test P=0.578 P=0.262 P=0.679N

Logistic regression test P=0.547N P=0.330 P=0.650N

Cochran-Armitage test P=0.399N

Fisher exact test P=0.509 P=0.500N

Skin (Subcutaneous Tissue): Fibrosarcoma

Overall rate 3/50 (6%) 8/51 (16%) 2/50 (4%)

Adjusted rate 6.9% 22.8% 5.1%

Terminal rate 1/40 (3%) 2/22 (9%) 0/23 (0%)

First incidence (days) 621 501 606

Life table test P=0.526 P=0.037 P=0.638N

Logistic regression test P=0.381N P=0.136 P=0.488N

Cochran-Armitage test P=0.429N

Fisher exact test P=0.106 P=0.500N

Skin (Subcutaneous Tissue): Fibrosarcoma or Sarcoma

Overall rate 4/50 (8%) 8/51 (16%) 2/50 (4%)

Adjusted rate 9.3% 22.8% 5.1%

Terminal rate 2/40 (5%) 2/22 (9%) 0/23 (0%)

First incidence (days) 621 501 606

Life table test P=0.501N P=0.066 P=0.508N

Logistic regression test P=0.268N P=0.222 P=0.339N

Cochran-Armitage test P=0.303N

Fisher exact test P=0.188 P=0.339N

Skin (Subcutaneous Tissue): Fibroma, Fibrosarcoma, or Sarcoma

Overall rate 6/50 (12%) 10/51 (20%) 3/50 (6%)

Adjusted rate 14.1% 29.5% 8.7%

Terminal rate 4/40 (10%) 3/22 (14%) 0/23 (0%)

First incidence (days) 621 501 606

Life table test P=0.494N P=0.053 P=0.476N

Logistic regression test P=0.233N P=0.220 P=0.280N

Cochran-Armitage test P=0.225N

Fisher exact test P=0.220 P=0.243N

Stomach (Forestomach): Squamous Cell Papilloma

Overall rate 0/50 (0%) 13/51 (25%) 16/50 (32%)

Adjusted rate 0.0% 51.0% 55.6%

Terminal rate 0/40 (0%) 10/22 (45%) 11/23 (48%)

First incidence (days) ) 613 606

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001
Lesions in Male Mice 235

TABLE C3

Statistical Analysis of Primary Neoplasms in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

Stomach (Forestomach): Squamous Cell Carcinoma

Overall rate 0/50 (0%) 12/51 (24%) 13/50 (26%)

Adjusted rate 0.0% 36.5% 37.7%

Terminal rate 0/40 (0%) 4/22 (18%) 4/23 (17%)

First incidence (days) ) 505 523

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001

Stomach (Forestomach): Squamous Cell Papilloma or Squamous Cell Carcinoma

Overall rate 0/50 (0%) 19/51 (37%) 27/50 (54%)

Adjusted rate 0.0% 61.2% 73.9%

Terminal rate 0/40 (0%) 11/22 (50%) 14/23 (61%)

First incidence (days) ) 505 523

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001

All Organs: Hemangiosarcoma

Overall rate 3/50 (6%) 2/51 (4%) 2/50 (4%)

Adjusted rate 7.5% 6.5% 4.4%

Terminal rate 3/40 (8%) 1/22 (5%) 0/23 (0%)

First incidence (days) 729 (T) 414 484

Life table test P=0.560N P=0.658 P=0.650N

Logistic regression test P=0.340N P=0.481N P=0.455N

Cochran-Armitage test P=0.406N

Fisher exact test P=0.491N P=0.500N

All Organs: Malignant Lymphoma (Histiocytic, Lymphocytic, Mixed, or Undifferentiated Cell Type)

Overall rate 10/50 (20%) 6/51 (12%) 7/50 (14%)

Adjusted rate 21.9% 20.8% 23.3%

Terminal rate 5/40 (13%) 2/22 (9%) 4/23 (17%)

First incidence (days) 480 612 512

Life table test P=0.517 P=0.565N P=0.567 

Logistic regression test P=0.282N P=0.218N P=0.306N

Cochran-Armitage test P=0.243N

Fisher exact test P=0.195N P=0.298N

All Organs: Benign Neoplasms

Overall rate 20/50 (40%) 43/51 (84%) 44/50 (88%)

Adjusted rate 47.4% 100.0% 100.0%

Terminal rate 18/40 (45%) 22/22 (100%) 23/23 (100%)

First incidence (days) 445 451 248

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001
236 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C3

Statistical Analysis of Primary Neoplasms in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

All Organs: Malignant Neoplasms

Overall rate 26/50 (52%) 39/51 (76%) 39/50 (78%)

Adjusted rate 53.1% 82.9% 88.3%

Terminal rate 17/40 (43%) 14/22 (64%) 18/23 (78%)

First incidence (days) 393 414 484

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P=0.002 P=0.012 P=0.003

Cochran-Armitage test P=0.003

Fisher exact test P=0.009 P=0.006

All Organs: Benign or Malignant Neoplasms

Overall rate 37/50 (74%) 49/51 (96%) 48/50 (96%)

Adjusted rate 75.5% 100.0% 100.0%

Terminal rate 28/40 (70%) 22/22 (100%) 23/23 (100%)

First incidence (days) 393 414 248

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P=0.002 P=0.002

(T)Terminal sacrifice

a Number of lesion-bearing animals/number of animals examined. Denominator is number of animals examined microscopically for liver, lung, and stomach; for other tissues,

denominator is number of animals necropsied. b Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality c Observed incidence at terminal kill d Beneath the control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values corresponding to pairwise comparisons

between the controls and that exposed group. The life table analysis regards neoplasms in animals dying prior to terminal kill as being (directly or indirectly) the cause of

death. The logistic regression test regards these lesions as nonfatal. The Cochran-Armitage and Fisher exact tests compare directly the overall incidence rates. For all tests,

a negative trend or a lower incidence in an exposure group is indicated by N. e Not applicable; no neoplasms in animal group
Lesions in Male Mice 237

TABLE C4a

Historical Incidence of Hepatocellular Neoplasms in Untreated Male B6C3F1 Micea

Incidence in Controls

Study Adenoma Carcinoma Adenoma or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 10/50 9/50 18/50

Acetaminophen 11/50 7/50 16/50

HC Yellow 4 8/49 5/49 13/49

Methylphenidate Hydrochloride 18/50 10/50 24/50

Pentaerythritol Tetranitrate 9/48 3/48 11/48

Turmeric Oleoresin 25/50 12/50 30/50

Overall Historical Incidence

Total 347/1,466 (23.7%) 241/1,466 (16.4%) 531/1,466 (36.2%)

Standard deviation 13.6% 7.0% 14.1%

Range 4%-60% 3%-29% 10%-68%

a Data as of 31 March 1993

TABLE C4b

Historical Incidence of Forestomach Squamous Cell Neoplasms in Untreated Male B6C3F1 Micea

Incidence in Controls

Study Papilloma Carcinoma Papilloma or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 0/50 0/50 0/50

Acetaminophen 1/50 0/50 1/50

HC Yellow 4 3/50 0/50 3/50

Methylphenidate Hydrochloride 1/50 0/50 1/50

Pentaerythritol Tetranitrate 0/49 0/49 0/49

Turmeric Oleoresin 2/50 0/50 2/50

Overall Historical Incidence

Total 20/1,474 (1.4%) 2/1,474 (0.1%) 22/1,474 (1.5%)

Standard deviation 2.0% 0.5% 2.0%

Range 0%-6% 0%-2% 0%-6%

a Data as of 31 March 1993
238 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C4c

Historical Incidence of Alveolar/bronchiolar Neoplasms in Untreated Male B6C3F1 Micea

Incidence in Controls

Study Adenoma Carcinoma Adenoma or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 7/50 3/50 10/50

Acetaminophen 8/50 0/50 8/50

HC Yellow 4 7/50 2/50 8/50

Methylphenidate Hydrochloride 14/50 4/50 16/50

Pentaerythritol Tetranitrate 11/47 1/47 11/47

Turmeric Oleoresin 11/50 4/50 14/50

Overall Historical Incidence

Total 201/1,469 (13.7%) 73/1,469 (5.0%) 265/1,469 (18.0%)

Standard deviation 6.2% 4.0% 7.6%

Range 4%-28% 0%-14% 4%-32%

a Data as of 31 March 1993
Lesions in Male Mice 239

TABLE C5

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 10,000 ppm 20,000 ppm

Disposition Summary

Animals initially in study 60 60 60

15-Month interim evaluation 10 9 10

Early deaths

Accidental death 1

Moribund 7 23 21

Natural deaths 3 5 6

Survivors

Terminal sacrifice 40 22 23

Animals examined microscopically 60 60 60

15-Month Interim Evaluation

Alimentary System

Gallbladder (10) (9) (9)

Inflammation, chronic 1 (11%)

Inflammation, chronic active 1 (10%)

Intestine large, cecum (10) (9) (10)

Ulcer, acute 1 (10%)

Liver (10) (9) (10)

Basophilic focus 1 (10%)

Fatty change 7 (70%) 3 (33%)

Inflammation, acute 1 (10%) 9 (100%) 3 (30%)

Inflammation, chronic active 4 (40%) 5 (50%)

Necrosis, coagulative 4 (40%) 8 (89%) 8 (80%)

Pigmentation 9 (100%) 10 (100%)

Centrilobular, cytoplasmic alteration 9 (100%) 8 (80%)

Mesentery (1)

Inflammation, chronic 1 (100%)

Necrosis, coagulative 1 (100%)

Pancreas (10) (9) (10)

Atrophy 1 (10%)

Cytoplasmic alteration 1 (10%)

Inflammation, chronic 2 (20%)

Artery, inflammation, chronic active 1 (10%)

Salivary glands (10) (8) (10)

Submandibular gland, inflammation, chronic 8 (80%) 4 (40%)

Stomach, forestomach (9) (9) (10)

Acanthosis 2 (22%) 3 (30%)

Hyperkeratosis 1 (11%) 2 (22%) 3 (30%)

Hyperplasia, basal cell 1 (11%) 1 (10%)

Inflammation, acute 1 (10%)

Inflammation, chronic active 2 (20%)

Stomach, glandular (9) (9) (10)

Inflammation, chronic 1 (11%) 5 (56%) 2 (20%)

Inflammation, chronic active 1 (11%) 1 (10%)

Muscularis, mineralization 1 (11%)

a Number of animals examined microscopically at site and number of animals with lesion
240 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C5

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

15-Month Interim Evaluation (continued)

Cardiovascular System

Heart (10) (9) (10)

Cardiomyopathy 3 (30%) 1 (10%)

Endocrine System

Islets, pancreatic (1) (1)

Hyperplasia 1 (100%)

Inflammation, chronic 1 (100%)

Pituitary gland (9) (9) (9)

Pars distalis, hyperplasia 2 (22%)

Thyroid gland (10) (9) (10)

Follicle, cyst 1 (10%)

Genital System

Epididymis (10) (9) (10)

Inflammation, chronic active 1 (10%)

Penis (1)

Inflammation, acute 1 (100%)

Preputial gland (5) (1)

Abscess 1 (100%)

Inflammation, chronic 2 (40%)

Inflammation, chronic active 2 (40%)

Duct, dilatation 1 (20%)

Prostate (10) (9) (9)

Inflammation, chronic 5 (50%) 2 (22%)

Inflammation, chronic active 1 (10%)

Artery, inflammation, chronic active 1 (10%)

Testes (10) (9) (10)

Atrophy 2 (20%)

Seminiferous tubule, atrophy 6 (60%) 2 (20%)

Hematopoietic System

Lymph node (2)

Inguinal, hyperplasia, lymphoid 1 (50%)

Lumbar, hyperplasia 1 (50%)

Lymph node, mesenteric (8) (9) (8)

Hemorrhage 3 (38%)

Infiltration cellular, histiocyte 1 (13%)

Pigmentation 1 (13%)

Spleen (10) (9) (8)

Hematopoietic cell proliferation 1 (10%)

Thymus (7) (9) (8)

Cyst 3 (38%)
Lesions in Male Mice 241

TABLE C5

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

15-Month Interim Evaluation (continued)

Integumentary System

Skin (10) (9) (10)

Inflammation, acute 1 (10%)

Inflammation, chronic active 1 (10%)

Subcutaneous tissue, inflammation, acute 2 (20%)

Subcutaneous tissue, inflammation, chronic active 1 (10%)

Musculoskeletal System

Bone (2) (2) (10)

Joint, tarsal, hyperostosis 2 (100%) 2 (100%)

Nervous System

Brain (10) (9) (10)

Thalamus, mineralization 9 (90%) 6 (60%)

Respiratory System

Lung (10) (9) (10)

Congestion 1 (10%)

Hemorrhage 1 (10%) 1 (10%)

Inflammation, chronic active 1 (11%)

Peribronchiolar, inflammation, chronic 1 (10%)

Perivascular, inflammation, chronic 1 (10%)

Nose (10) (9) (10)

Crystals 1 (10%)

Inflammation, acute 2 (20%)

Glands, inflammation, acute 2 (20%)

Respiratory epithelium, necrosis 1 (10%)

Urinary System

Kidney (10) (1) (10)

Inflammation, chronic 10 (100%) 1 (100%) 8 (80%)

Renal tubule, regeneration 4 (40%)

Urinary bladder (10) (9) (10)

Inflammation, chronic 3 (30%) 7 (78%) 5 (50%)

Inflammation, chronic active 1 (10%)

Systems Examined With No Lesions Observed

General Body System

Special Senses System
242 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C5

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study

Alimentary System

Gallbladder (46) (47) (42)

Autolysis 1 (2%) 2 (4%)

Inflammation, chronic active 11 (24%) 12 (26%) 8 (19%)

Epithelium, pigmentation 1 (2%)

Intestine large, colon (50) (49) (50)

Autolysis 1 (2%) 2 (4%)

Inflammation, chronic active 1 (2%) 1 (2%) 2 (4%)

Peyer's patch, hyperplasia 1 (2%)

Intestine large, rectum (48) (49) (48)

Autolysis 2 (4%) 2 (4%)

Intestine large, cecum (49) (51) (50)

Autolysis 2 (4%) 2 (4%)

Hemorrhage 1 (2%)

Hyperplasia, lymphoid 1 (2%)

Epithelium, hyperplasia 1 (2%)

Peyer's patch, hyperplasia 1 (2%) 1 (2%)

Serosa, inflammation, chronic active 1 (2%)

Intestine small, duodenum (50) (50) (46)

Autolysis 2 (4%) 1 (2%)

Mesothelium, hyperplasia 1 (2%)

Peyer's patch, inflammation, chronic,

granulomatous 1 (2%)

Serosa, fibrosis 1 (2%)

Intestine small, jejunum (50) (47) (48)

Autolysis 2 (4%) 1 (2%)

Hyperplasia, lymphoid 1 (2%)

Inflammation, acute 1 (2%)

Epithelium, pigmentation 1 (2%)

Peyer's patch, inflammation, chronic,

granulomatous 1 (2%)

Intestine small, ileum (50) (49) (47)

Autolysis 2 (4%) 1 (2%)

Hyperplasia, lymphoid 1 (2%)

Inflammation, acute, necrotizing 1 (2%)

Inflammation, chronic active 1 (2%)

Peyer's patch, hyperplasia 1 (2%) 2 (4%) 1 (2%)

Peyer's patch, hyperplasia, lymphoid 1 (2%)

Peyer's patch, inflammation, chronic,

granulomatous 1 (2%)

Liver (50) (51) (50)

Angiectasis 1 (2%)

Basophilic focus 4 (8%) 3 (6%)

Basophilic focus, focal 1 (2%)

Clear cell focus 4 (8%) 4 (8%) 2 (4%)

Cytoplasmic alteration 1 (2%)

Eosinophilic focus 6 (12%) 1 (2%)

Fatty change 4 (8%) 3 (6%) 7 (14%)

Fibrosis 1 (2%)

Hematopoietic cell proliferation 1 (2%) 5 (10%) 10 (20%)

Infarct 1 (2%)
Lesions in Male Mice 243

TABLE C5

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Alimentary System (continued)

Liver (continued) (50) (51) (50)

Inflammation, chronic active 11 (22%) 10 (20%) 22 (44%)

Karyomegaly 1 (2%)

Mineralization 1 (2%) 1 (2%)

Necrosis, coagulative 5 (10%) 12 (24%) 20 (40%)

Nuclear alteration 1 (2%)

Pigmentation 1 (2%) 50 (98%) 47 (94%)

Thrombosis 1 (2%)

Bile duct, hyperplasia 1 (2%) 2 (4%) 1 (2%)

Hepatocyte, centrilobular, hypertrophy 17 (33%) 13 (26%)

Mesentery (2) (7) (9)

Abscess 1 (11%)

Fibrosis 1 (11%)

Inflammation, chronic active 4 (57%) 5 (56%)

Mineralization 1 (11%)

Necrosis, coagulative 2 (22%)

Thrombosis 1 (14%)

Pancreas (50) (50) (48)

Atrophy 2 (4%)

Autolysis 1 (2%) 1 (2%)

Cytoplasmic alteration 4 (8%) 1 (2%)

Ectopic liver 1 (2%)

Inflammation, chronic active 19 (38%) 12 (24%) 8 (17%)

Vacuolization cytoplasmic 20 (40%) 18 (36%) 4 (8%)

Acinus, atrophy 1 (2%)

Salivary glands (50) (51) (50)

Inflammation, chronic 2 (4%)

Duct, parotid gland, mineralization 1 (2%)

Parotid gland, inflammation, chronic 2 (4%)

Sublingual gland, inflammation, chronic 1 (2%)

Sublingual gland, submandibular gland,

inflammation, chronic 1 (2%)

Submandibular gland, atrophy 1 (2%)

Submandibular gland, inflammation, chronic 37 (74%) 29 (57%) 36 (72%)

Stomach, forestomach (50) (50) (50)

Abscess 1 (2%)

Acanthosis 1 (2%) 9 (18%) 4 (8%)

Autolysis 1 (2%)

Cyst 1 (2%)

Diverticulum 1 (2%)

Edema 1 (2%)

Erosion 1 (2%)

Hyperkeratosis 1 (2%) 7 (14%) 6 (12%)

Hyperplasia, basal cell 2 (4%)

Inflammation, chronic active 2 (4%) 6 (12%) 12 (24%)

Inflammation, chronic active, necrotizing 1 (2%)

Ulcer 2 (4%)
244 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C5

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Alimentary System (continued)

Stomach, glandular (50) (50) (49)

Autolysis 1 (2%)

Edema 1 (2%)

Infiltration cellular, mast cell 1 (2%)

Inflammation, chronic active 14 (28%) 24 (48%) 22 (45%)

Mineralization 4 (8%) 3 (6%)

Tooth (4) (7)

Dysplasia 4 (100%) 6 (86%)

Cardiovascular System

Heart (50) (51) (50)

Inflammation, chronic 7 (14%) 17 (33%) 5 (10%)

Mineralization 1 (2%)

Necrosis, coagulative 1 (2%)

Coronary artery, thrombosis 1 (2%)

Endocrine System

Adrenal cortex (50) (51) (50)

Hematopoietic cell proliferation 1 (2%)

Hyperplasia 3 (6%) 2 (4%)

Inflammation, chronic active 3 (6%)

Pigmentation 2 (4%)

Vacuolization cytoplasmic 1 (2%)

Capsule, inflammation, chronic active 1 (2%)

Adrenal medulla (50) (50) (50)

Hyperplasia 1 (2%) 1 (2%)

Inflammation, acute 1 (2%)

Pigmentation 1 (2%)

Islets, pancreatic (50) (50) (48)

Hyperplasia 8 (16%) 1 (2%) 1 (2%)

Pituitary gland (43) (45) (47)

Pars distalis, cyst 5 (12%) 4 (9%) 1 (2%)

Pars distalis, hyperplasia 13 (30%) 9 (20%) 4 (9%)

Thyroid gland (49) (50) (49)

Inflammation, chronic 1 (2%) 1 (2%) 1 (2%)

Ultimobranchial cyst 1 (2%) 1 (2%)

Follicular cell, hyperplasia 2 (4%) 3 (6%) 3 (6%)

General Body System

None
Lesions in Male Mice 245

TABLE C5

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Genital System

Coagulating gland (1) (1) (1)

Inflammation, chronic active 1 (100%)

Epididymis (49) (51) (50)

Inflammation, chronic active 29 (59%) 21 (41%) 26 (52%)

Epithelium, hyperplasia 1 (2%)

Serosa, hyperplasia 1 (2%)

Penis (1) (2)

Hemorrhage 1 (100%)

Preputial gland (16) (16) (12)

Abscess 1 (6%) 3 (19%) 1 (8%)

Inflammation, chronic active 12 (75%) 7 (44%) 11 (92%)

Duct, dilatation 11 (69%) 10 (63%) 2 (17%)

Prostate (47) (46) (48)

Inflammation 1 (2%)

Inflammation, chronic active 31 (66%) 29 (63%) 20 (42%)

Epithelium, hyperplasia 1 (2%)

Seminal vesicle (49) (50) (45)

Atrophy 1 (2%) 1 (2%)

Cyst 1 (2%)

Fibrosis 3 (6%)

Inflammation, chronic active 4 (8%) 14 (28%) 13 (29%)

Artery, thrombosis 1 (2%)

Testes (50) (50) (50)

Inflammation, chronic active 2 (4%)

Interstitial cell, hyperplasia 1 (2%)

Seminiferous tubule, atrophy 1 (2%) 2 (4%)

Seminiferous tubule, mineralization 2 (4%) 1 (2%) 3 (6%)

Hematopoietic System

Bone marrow (49) (50) (49)

Myeloid cell, sternal, hyperplasia 1 (2%) 2 (4%) 1 (2%)

Sternal, infiltration cellular, mast cell 1 (2%)

Sternal, inflammation, granulomatous 1 (2%)

Lymph node (8) (8) (11)

Hyperplasia, lymphoid 1 (9%)

Lumbar, hyperplasia, lymphoid 1 (13%) 1 (13%)

Lumbar, hyperplasia, plasma cell 2 (18%)

Lumbar, inflammation, chronic active 1 (13%)

Mediastinal, infiltration cellular,

 histiocyte 1 (13%)

Mediastinal, inflammation, chronic active 1 (9%)

Mediastinal, pigmentation 1 (13%)

Pancreatic, angiectasis 1 (9%)

Pancreatic, hyperplasia, lymphoid 2 (25%)

Pancreatic, infiltration cellular, histiocyte 1 (13%)

Pancreatic, pigmentation 1 (13%)

Renal, angiectasis 1 (13%)

Renal, hyperplasia, lymphoid 1 (13%)

Renal, inflammation, chronic active 1 (13%)
246 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C5

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Hematopoietic System (continued)

Lymph node, mandibular (32) (34) (26)

Depletion lymphoid 3 (9%) 1 (3%) 1 (4%)

Hyperplasia, lymphoid 3 (9%) 1 (3%) 1 (4%)

Hyperplasia, plasma cell 1 (3%)

Infiltration cellular, histiocyte 3 (9%) 8 (31%)

Inflammation, chronic active 1 (3%) 1 (3%) 1 (4%)

Necrosis, coagulative 1 (4%)

Pigmentation 5 (16%) 4 (12%) 9 (35%)

Lymph node, mesenteric (46) (47) (47)

Angiectasis 24 (52%) 11 (23%) 21 (45%)

Congestion 1 (2%)

Depletion lymphoid 1 (2%) 1 (2%)

Hematopoietic cell proliferation 2 (4%)

Hemorrhage 1 (2%) 1 (2%)

Hyperplasia 1 (2%)

Hyperplasia, lymphoid 6 (13%) 9 (19%) 4 (9%)

Hyperplasia, plasma cell 1 (2%)

Infiltration cellular, histiocyte 24 (52%) 22 (47%) 21 (45%)

Inflammation, chronic active 7 (15%) 4 (9%)

Pigmentation 23 (50%) 22 (47%) 19 (40%)

Spleen (50) (51) (50)

Angiectasis 1 (2%)

Congestion 1 (2%)

Depletion lymphoid 1 (2%) 3 (6%) 3 (6%)

Fibrosis 1 (2%)

Hematopoietic cell proliferation 5 (10%) 14 (27%) 12 (24%)

Hyperplasia, lymphoid 2 (4%) 1 (2%)

Inflammation, chronic active 1 (2%) 1 (2%) 2 (4%)

Necrosis, coagulative 1 (2%)

Thymus (37) (35) (33)

Cyst 2 (5%) 6 (17%) 9 (27%)

Cyst, multiple 2 (5%)

Depletion lymphoid 2 (5%) 8 (23%) 6 (18%)

Hyperplasia, lymphoid 1 (3%)

Infiltration cellular, histiocyte 1 (3%)

Necrosis, coagulative 1 (3%) 1 (3%)

Pigmentation 1 (3%) 1 (3%)

Integumentary System

Skin (50) (48) (48)

Inflammation, chronic active 4 (8%) 9 (19%) 12 (25%)

Subcutaneous tissue, abscess 1 (2%)

Subcutaneous tissue, cyst 1 (2%)

Subcutaneous tissue, fibrosis 2 (4%) 1 (2%)
Lesions in Male Mice 247

TABLE C5

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Musculoskeletal System

Bone (49) (50) (49)

Cartilage, tarsal, hyperplasia 1 (2%)

Joint, tarsal, hyperostosis 10 (20%) 11 (22%) 12 (24%)

Skeletal muscle (3) (3) (3)

Fibrosis 1 (33%)

Intercostal, inflammation, chronic active 1 (33%)

Nervous System

Brain (50) (51) (50)

Cyst epithelial inclusion 1 (2%)

Cerebellum, necrosis 1 (2%)

Cerebrum, necrosis 1 (2%)

Thalamus, mineralization 17 (34%) 20 (39%) 13 (26%)

Respiratory System

Lung (50) (51) (50)

Bronchiectasis 1 (2%)

Congestion 1 (2%)

Crystals 1 (2%)

Foreign body 1 (2%)

Granuloma 1 (2%)

Hemorrhage 8 (16%) 9 (18%) 9 (18%)

Infiltration cellular, histiocyte 2 (4%) 2 (4%) 6 (12%)

Inflammation, acute 2 (4%)

Inflammation, chronic active 1 (2%) 2 (4%) 1 (2%)

Mineralization 1 (2%)

Pigmentation 1 (2%)

Alveolar epithelium, hyperplasia 1 (2%) 4 (8%)

Bronchiole, hyperplasia 1 (2%)

Bronchiole, metaplasia, squamous 1 (2%)

Nose (43) (49) (48)

Inflammation, acute 11 (26%) 11 (22%) 6 (13%)

Glands, crystals 2 (5%)

Nasolacrimal duct, inflammation, acute 2 (5%)

Trachea (50) (50) (50)

Inflammation, acute 2 (4%) 3 (6%)

Special Senses System

Eye (2) (1)

Anterior, synechia 1 (100%)

Cornea, fibrosis 1 (100%)

Cornea, inflammation, chronic active 1 (50%)

Harderian gland (2) (2) (1)

Inflammation, chronic active 1 (50%)
248 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE C5

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Urinary System

Kidney (50) (51) (50)

Autolysis 1 (2%)

Congestion 1 (2%)

Cyst 1 (2%) 1 (2%)

Glomerulosclerosis 3 (6%)

Infiltration cellular, mononuclear cell 1 (2%)

Inflammation, acute, necrotizing 1 (2%)

Inflammation, chronic active 47 (94%) 42 (82%) 47 (94%)

Metaplasia, osseous 1 (2%)

Mineralization 1 (2%)

Necrosis, coagulative 1 (2%)

Artery, necrosis, fibrinoid 1 (2%)

Pelvis, transitional epithelium, hyperplasia 1 (2%)

Proximal convoluted renal tubule, hyperplasia 1 (2%)

Proximal convoluted renal tubule,

 regeneration 2 (4%) 11 (22%) 1 (2%)

Proximal convoluted renal tubule,

 vacuolization cytoplasmic 1 (2%)

Renal tubule, mineralization 1 (2%) 4 (8%) 1 (2%)

Renal tubule, pigmentation 42 (82%) 43 (86%)

Renal tubule, regeneration 4 (8%) 1 (2%)

Urinary bladder (49) (50) (49)

Autolysis 2 (4%) 3 (6%)

Calculus, microscopic observation only 5 (10%) 2 (4%)

Inflammation, chronic active 36 (73%) 31 (62%) 34 (69%)

Transitional epithelium, hyperplasia 2 (4%)
249

APPENDIX D

SUMMARY OF LESIONS IN FEMALE MICE

IN THE 2-YEAR FEED STUDY

OF 1-AMINO-2,4-DIBROMOANTHRAQUINONE

TABLE D1 Summary of the Incidence of Neoplasms in Female Mice 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 251

TABLE D2 Individual Animal Tumor Pathology of Female Mice 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 256

TABLE D3 Statistical Analysis of Primary Neoplasms in Female Mice 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 274

TABLE D4a Historical Incidence of Hepatocellular Neoplasms 

in Untreated Female B6C3F1 Mice . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 278

TABLE D4b Historical Incidence of Forestomach Squamous Cell Neoplasms 

in Untreated Female B6C3F1 Mice . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 278

TABLE D4c Historical Incidence of Alveolar/bronchiolar Neoplasms

in Untreated Female B6C3F1 Mice . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 279

TABLE D4d Historical Incidence of Uterine Neoplasms in Untreated Female B6C3F1 Mice . . . . . . . . . . . . . . . . . . . . . . . 279

TABLE D4e Historical Incidence of Pituitary Gland Pars Distalis Neoplasms

in Untreated Female B6C3F1 Mice . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 280

TABLE D5 Summary of the Incidence of Nonneoplastic Lesions in Female Mice 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 281
250 1-Amino-2,4-dibromoanthraquinone, NTP TR 383
Lesions in Female Mice 251

TABLE D1

Summary of the Incidence of Neoplasms in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 10,000 ppm 20,000 ppm

Disposition Summary

Animals initially in study 60 60 60

15-Month interim evaluation 10 10 10

Early deaths

Moribund 5 11 11

Natural deaths 6 5 6

Survivors

Died last week of study 1 1

Terminal sacrifice 38 33 33

Animals examined microscopically 60 60 60

15-Month Interim Evaluation

Alimentary System

Liver (10) (10) (10)

Hepatocellular carcinoma 1 (10%) 

Hepatocellular adenoma 1 (10%) 3 (30%)

Hepatocellular adenoma, multiple 1 (10%) 4 (40%) 

Stomach, forestomach (10) (10) (10)

Squamous cell papilloma 4 (40%) 2 (20%) 

Respiratory System

Lung (10) (10) (10)

Alveolar/bronchiolar adenoma 3 (30%) 2 (20%) 

Systems Examined With No Neoplasms Observed

Cardiovascular System

Endocrine System

General Body System

Genital System

Hematopoietic System

Integumentary System

Musculoskeletal System

Nervous System

Special Senses System

Urinary System

2-Year Study

Alimentary System

Esophagus (50) (45) (47)

Basosquamous tumor malignant, metastatic, 

uterus 1 (2%)

Gallbladder (49) (47) (44)

Basosquamous tumor malignant, metastatic,

 uterus 1 (2%)
252 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D1

Summary of the Incidence of Neoplasms in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Alimentary System (continued)

Intestine large, colon (50) (50) (48)

Intestine small, duodenum (48) (50) (46)

Intestine small, jejunum (47) (50) (46)

Intestine small, ileum (48) (50) (46)

Liver (50) (50) (50)

Fibrosarcoma, metastatic, skin 1 (2%)

Hemangioma 1 (2%)

Hepatoblastoma 2 (4%)

Hepatocellular carcinoma 10 (20%) 14 (28%) 

Hepatocellular carcinoma, multiple 13 (26%) 13 (26%)

Hepatocellular adenoma 6 (12%) 5 (10%) 4 (8%) 

Hepatocellular adenoma, multiple 40 (80%) 45 (90%)

Squamous cell carcinoma, metastatic, stomach,

 forestomach 3 (6%) 4 (8%)

Mesentery (6) (9) (8)

Basosquamous tumor malignant, metastatic,

 uterus 1 (17%)

Fibrosarcoma, metastatic, skin 1 (11%)

Sarcoma, metastatic, stomach, forestomach 1 (13%)

Squamous cell carcinoma, metastatic, stomach,

 forestomach 2 (22%) 3 (38%)

Pancreas (50) (50) (49)

Basosquamous tumor malignant, metastatic,

 uterus 1 (2%)

Fibrosarcoma, metastatic, skin 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

 forestomach 1 (2%) 2 (4%)

Salivary glands (49) (50) (47)

Stomach, forestomach (48) (50) (50)

Sarcoma 1 (2%)

Squamous cell carcinoma 12 (24%) 11 (22%)

Squamous cell papilloma 2 (4%) 12 (24%) 13 (26%)

Squamous cell papilloma, multiple 4 (8%) 14 (28%)

Stomach, glandular (49) (48) (48)

Sarcoma 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

 forestomach 3 (6%) 3 (6%)

Cardiovascular System

Heart (50) (50) (49)

Endocrine System

Adrenal cortex (50) (49) (48)

Carcinoma 1 (2%)

Squamous cell carcinoma, metastatic 1 (2%)

Adrenal medulla (49) (49) (48)

Islets, pancreatic (49) (50) (48)

Adenoma 2 (4%) 1 (2%)
Lesions in Female Mice 253

TABLE D1

Summary of the Incidence of Neoplasms in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Endocrine System (continued)

Pituitary gland (43) (45) (43)

Pars distalis, adenoma 1 (2%) 8 (18%) 4 (9%)

Pars distalis, adenoma, multiple 1 (2%) 

Pars intermedia, adenoma 1 (2%)

Pars nervosa, adenoma 1 (2%)

Thyroid gland (50) (50) (48)

Follicular cell, adenoma 3 (6%) 1 (2%) 2 (4%)

General Body System

Tissue NOS (1)

Genital System

Ovary (49) (49) (47)

Basosquamous tumor malignant, metastatic,

 uterus 1 (2%)

Granulosa cell tumor benign 1 (2%)

Hemangioma 1 (2%)

Luteoma 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

 forestomach 1 (2%) 3 (6%)

Uterus (49) (50) (49)

Leiomyoma 2 (4%)

Polyp stromal 2 (4%)

Sarcoma stromal 3 (6%)

Cervix, basosquamous tumor malignant 1 (2%)

Vagina (1)

Squamous cell carcinoma 1 (100%)

Hematopoietic System

Bone marrow (50) (49) (49)

Lymph node (7) (12) (12)

Bronchial, sarcoma, metastatic, stomach,

 forestomach 1 (8%)

Mediastinal, squamous cell carcinoma,

 metastatic, stomach, forestomach 1 (8%) 1 (8%)

Renal, basosquamous tumor malignant,

 metastatic, uterus 1 (14%)

Lymph node, mandibular (32) (45) (31)

Lymph node, mesenteric (49) (46) (44)

Basosquamous tumor malignant, metastatic,

 uterus 1 (2%)

Sarcoma, metastatic, stomach, forestomach 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

 forestomach 1 (2%) 1 (2%)
254 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D1

Summary of the Incidence of Neoplasms in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Hematopoietic System (continued)

Spleen (50) (50) (50)

Hemangioma 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

 forestomach 1 (2%)

Thymus (40) (38) (34)

Osteosarcoma, metastatic, bone 1 (3%)

Integumentary System

Mammary gland (25) (25) (20)

Fibroadenoma 1 (4%)

Skin (50) (50) (50)

Squamous cell papilloma 1 (2%)

Subcutaneous tissue, fibrosarcoma 2 (4%) 1 (2%)

Subcutaneous tissue, hemangioma 1 (2%)

Musculoskeletal System

Bone (50) (48) (49)

Rib, osteosarcoma 1 (2%)

Skeletal muscle (4) (2) (2)

Abdominal, fibrosarcoma, metastatic, skin 1 (50%)

Diaphragm, basosquamous tumor malignant,

 metastatic, uterus 1 (25%)

Diaphragm, squamous cell carcinoma,

 metastatic, stomach, forestomach 1 (50%) 1 (50%)

Nervous System

Brain (49) (50) (50)

Respiratory System

Lung (50) (50) (49)

Alveolar/bronchiolar adenoma 4 (8%) 15 (30%) 12 (24%)

Alveolar/bronchiolar adenoma, multiple 2 (4%) 1 (2%)

Alveolar/bronchiolar carcinoma 2 (4%)

Basosquamous tumor malignant, metastatic,

 uterus 1 (2%)

Hemangioma 1 (2%)

Hepatocellular carcinoma, metastatic, liver 1 (2%)

Osteosarcoma, metastatic, bone 1 (2%)

Squamous cell carcinoma, metastatic, stomach,

 forestomach 1 (2%)

Nose (48) (45) (44)
Lesions in Female Mice 255

TABLE D1

Summary of the Incidence of Neoplasms in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Special Senses System

Harderian gland (1) (3) (2)

Adenoma 1 (100%) 2 (67%) 2 (100%)

Carcinoma 1 (33%)

Urinary System

Kidney (50) (50) (50)

Osteosarcoma, metastatic, bone 1 (2%)

Urinary bladder (49) (50) (50)

Systemic Lesions

Multiple organsb (50) (50) (50)

Lymphoma malignant histiocytic 1 (2%)

Lymphoma malignant lymphocytic 1 (2%)

Lymphoma malignant mixed 9 (18%) 13 (26%) 13 (26%)

Lymphoma malignant undifferentiated cell 2 (4%)

Neoplasm Summary

Total animals with primary neoplasmsc

15-Month interim evaluation 5 9

2-Year study 32 50 50

Total primary neoplasms

15-Month interim evaluation 9 12

2-Year study 45 149 157

Total animals with benign neoplasms

15-Month interim evaluation 5 8

2-Year study 24 46 50

Total benign neoplasms

15-Month interim evaluation 9 11

2-Year study 29 94 99

Total animals with malignant neoplasms

15-Month interim evaluation 1

2-Year study 14 40 37

Total malignant neoplasms

15-Month interim evaluation 1

2-Year study 16 55 58

Total animals with metastatic neoplasms

2-Year study 2 4 6

Total metastatic neoplasms

2-Year study 12 18 24

a Number of animals examined microscopically at site and number of animals with neoplasm b Number of animals with any tissue examined microscopically c Primary neoplasms: all neoplasms except metastatic neoplasms
256 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm

0456666777777777777777777 

Number of Days on Study 7791258000033333333333333 

6820835116823333333333333

4333443444433333444444444 

Carcass ID Number 4989667088678899000111222 

5554545454342312123145123

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 

Basosquamous tumor malignant, 

metastatic, uterus X 

Gallbladder + + + + + + + + + M + + + + + + + + + + + + + + + 

Basosquamous tumor malignant, 

metastatic, uterus X 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, duodenum A + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, jejunum A + + A + + + + + + A + + + + + + + + + + + + + +

Intestine small, ileum A + + A + + + + + + + + + + + + + + + + + + + + +

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 

Hemangioma 

Hepatocellular adenoma X X X 

Mesentery + + + + + 

Basosquamous tumor malignant, 

metastatic, uterus X 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 

Basosquamous tumor malignant, 

metastatic, uterus X 

Salivary glands + + M + + + + + + + + + + + + + + + + + + + + + +

Stomach, forestomach + + + M + + + + + + + + + + + + + + + + + + + + + 

Squamous cell papilloma 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + +

Tooth + 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + +

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 

Carcinoma 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + +

Islets, pancreatic + + + + + + + + + + + + + + + + + M + + + + + + + 

Adenoma X 

Parathyroid gland M + M + + + + + + + + + + + + MMM+M++M++

Pituitary gland M + + + + + M + + + + + + M + + + + + + + + + + + 

Pars distalis, adenoma X 

Pars intermedia, adenoma X 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Follicular cell, adenoma 

General Body System 

None 

+: Tissue examined microscopically M: Missing tissue X: Lesion present

A: Autolysis precludes examination I: Insufficient tissue Blank: Not examined
Lesions in Female Mice 257

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

7777777777777777777777777

Number of Days on Study 3333333333333333333333333

3333333333333333444444444

4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 3 3 4 4 4 4 4 4 4 Total

Carcass ID Number 2 2 3 3 3 3 3 4 4 4 5 5 6 7 7 7 7 8 4 5 5 6 8 8 8 Tissues/

4 5 1 2 3 4 5 1 2 3 1 3 1 1 3 4 3 4 4 4 5 2 1 2 3 Tumors

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Basosquamous tumor malignant, 

metastatic, uterus 1

Gallbladder + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Basosquamous tumor malignant, 

metastatic, uterus 1

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + M + 48 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangioma X 1

Hepatocellular adenoma X X X 6

Mesentery + 6

Basosquamous tumor malignant, 

metastatic, uterus 1

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Basosquamous tumor malignant, 

metastatic, uterus 1

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Stomach, forestomach + + + + + + + + + + + + M + + + + + + + + + + + + 48 

Squamous cell papilloma X X 2

Stomach, glandular + + + + + + + + + + + + M + + + + + + + + + + + + 49 

Tooth 1

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma X 1

Adrenal medulla + + + + + + + M + + + + + + + + + + + + + + + + + 49 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Adenoma X 2

Parathyroid gland + M M M + + + + M + M + + M M + M + + M + M + + + 33 

Pituitary gland + + M + M + + + + + + + + + + + + M M + + + + + + 43 

Pars distalis, adenoma 1

Pars intermedia, adenoma 1

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Follicular cell, adenoma X X X 3

General Body System 

None 
258 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

0456666777777777777777777 

Number of Days on Study 7791258000033333333333333 

6820835116823333333333333

4333443444433333444444444 

Carcass ID Number 4989667088678899000111222 

5554545454342312123145123

Genital System 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + M 

Basosquamous tumor malignant, 

metastatic, uterus X 

Granulosa cell tumor benign 

Hemangioma 

Uterus M + + + + + + + + + + + + + + + + + + + + + + + + 

Leiomyoma 

Cervix, basosquamous tumor malignant X 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node + + + + + + 

Renal, basosquamous tumor malignant, 

metastatic, uterus X 

Lymph node, mandibular M + M M + + + + + + M + + M + + + + M M + + M + M

Lymph node, mesenteric M + + + + + + + + + + + + + + + + + + + + + + + + 

Basosquamous tumor malignant, 

metastatic, uterus X 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 

Hemangioma 

Thymus M + + M M + I + M M + + + + + + + + + + + + + + + 

Osteosarcoma, metastatic, bone X 

Integumentary System 

Mammary gland M + M + + + + M + MMM++M++++M++M++ 

Fibroadenoma X 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 

Squamous cell papilloma X 

Subcutaneous tissue, fibrosarcoma X X 

Subcutaneous tissue, hemangioma X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Rib, osteosarcoma X 

Skeletal muscle + + + + 

Diaphragm, basosquamous tumor 

malignant, metastatic, uterus X 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + +

Respiratory System

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 

Alveolar/bronchiolar adenoma X 

Basosquamous tumor malignant, 

metastatic, uterus X 

Osteosarcoma, metastatic, bone X 

Nose + + + + + + + + + + + + + + M + + + + + + + + + +

Trachea + + + + + + + + + + + + + + + + + + + + + + + + +
Lesions in Female Mice 259

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

7777777777777777777777777

Number of Days on Study 3333333333333333333333333

3333333333333333444444444

4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 3 3 4 4 4 4 4 4 4 Total

Carcass ID Number 2 2 3 3 3 3 3 4 4 4 5 5 6 7 7 7 7 8 4 5 5 6 8 8 8 Tissues/

4 5 1 2 3 4 5 1 2 3 1 3 1 1 3 4 3 4 4 4 5 2 1 2 3 Tumors

Genital System 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Basosquamous tumor malignant, 

metastatic, uterus 1

Granulosa cell tumor benign X 1

Hemangioma X 1

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Leiomyoma X X 2

Cervix, basosquamous tumor malignant 1

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymph node + 7

Renal, basosquamous tumor malignant, 

metastatic, uterus 1

Lymph node, mandibular + + M M + + + + I + + M + M + M M + + + + M M + + 32 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Basosquamous tumor malignant, 

metastatic, uterus 1

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangioma X 1

Thymus + + + + + + + M + + + + M + M + + + + + + + + + M 40 

Osteosarcoma, metastatic, bone 1

Integumentary System 

Mammary gland M M + + M M M M + M + + M + M M + M M M + M M M + 25 

Fibroadenoma 1

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Squamous cell papilloma 1

Subcutaneous tissue, fibrosarcoma 2

Subcutaneous tissue, hemangioma 1

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Rib, osteosarcoma 1

Skeletal muscle 4

Diaphragm, basosquamous tumor 

malignant, metastatic, uterus 1

Nervous System 

Brain + + + + + + + + + + M + + + + + + + + + + + + + + 49 

Respiratory System

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Alveolar/bronchiolar adenoma X X X 4

Basosquamous tumor malignant, 

metastatic, uterus 1

Osteosarcoma, metastatic, bone 1

Nose + M + + + + + + + + + + + + + + + + + + + + + + + 48 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 50 
260 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

0456666777777777777777777 

Number of Days on Study 7791258000033333333333333 

6820835116823333333333333

4333443444433333444444444 

Carcass ID Number 4989667088678899000111222 

5554545454342312123145123

Special Senses System 

Ear + 

Eye 

Harderian gland 

Adenoma 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 

Osteosarcoma, metastatic, bone X 

Urinary bladder + + + + + + A + + + + + + + + + + + + + + + + + +

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Lymphoma malignant histiocytic 

Lymphoma malignant lymphocytic 

Lymphoma malignant mixed X X X X X X 
Lesions in Female Mice 261

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

0 ppm (continued)

7777777777777777777777777

Number of Days on Study 3333333333333333333333333

3333333333333333444444444

4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 3 3 4 4 4 4 4 4 4 Total

Carcass ID Number 2 2 3 3 3 3 3 4 4 4 5 5 6 7 7 7 7 8 4 5 5 6 8 8 8 Tissues/

4 5 1 2 3 4 5 1 2 3 1 3 1 1 3 4 3 4 4 4 5 2 1 2 3 Tumors

Special Senses System 

Ear 1

Eye + 1

Harderian gland + 1

Adenoma X 1

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Osteosarcoma, metastatic, bone 1

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymphoma malignant histiocytic X 1

Lymphoma malignant lymphocytic X 1

Lymphoma malignant mixed X X X 9
262 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

10,000 ppm

2345556666777777777777777 

Number of Days on Study 0740585577011112333333333 

2223572719633367000000000

5455655455555555444555555 

Carcass ID Number 6976040977613935999000011 

5554545443355545123123412

Alimentary System 

Esophagus + + + + M + + M + + + + M + + + + + + + + + + + +

Gallbladder + + + M + + + + + + + + + + + + + + + + + + + + +

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine large, cecum + + + + + + M + + + + + + + + + + + + + + + + + +

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + +

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + +

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 

Fibrosarcoma, metastatic, skin X 

Hepatocellular carcinoma X X X X 

Hepatocellular carcinoma, multiple X X X X X X 

Hepatocellular adenoma X X X X 

Hepatocellular adenoma, multiple X X X X X X X X X X X X X X X X X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X 

Mesentery + + + + + + 

Fibrosarcoma, metastatic, skin X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 

Fibrosarcoma, metastatic, skin X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + +

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 

Squamous cell carcinoma X X X X X X 

Squamous cell papilloma X X X X X 

Squamous cell papilloma, multiple X 

Stomach, glandular + + + + + + + + + + + + + + + + + + M + + + + + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + +

Endocrine System

Adrenal cortex + + + + + + + + + + + + + M + + + + + + + + + + +

Adrenal medulla + + + + + + + + + + + + + M + + + + + + + + + + +

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma 

Parathyroid gland M + + + + + + + + + + M M + + + + + + + + M + +

Pituitary gland + + + + + + + + + + I + + + + + + + I M + + + + + 

Pars distalis, adenoma X X X 

Pars distalis, adenoma, multiple 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Follicular cell, adenoma 
Lesions in Female Mice 263

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

10,000 ppm (continued)

7777777777777777777777777

Number of Days on Study 3333333333333333333333333

0000000000000002333333333

5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 6 6 Total

Carcass ID Number 1 2 2 2 2 3 3 4 4 4 5 5 6 6 7 8 7 8 8 8 9 9 9 0 0 Tissues/

3 1 2 3 4 1 2 1 2 3 3 4 1 2 2 5 1 1 3 4 1 2 3 3 4 Tumors

Alimentary System 

Esophagus + + + + + + + + + + + + M + + + + + + M + + + + + 45 

Gallbladder + M + + + + + + + + + + + + M + + + + + + + + + + 47 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Fibrosarcoma, metastatic, skin 1

Hepatocellular carcinoma X X X X X X 10 

Hepatocellular carcinoma, multiple X X X X X X X 13 

Hepatocellular adenoma X 5

Hepatocellular adenoma, multiple X X X X X X X X X X X X X X X X X X X X X X X 40 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X 3

Mesentery + + + 9

Fibrosarcoma, metastatic, skin 1

Squamous cell carcinoma, metastatic, 

stomach, forestomach 2

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Fibrosarcoma, metastatic, skin 1

Squamous cell carcinoma, metastatic, 

stomach, forestomach X 1

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Squamous cell carcinoma X X X X X X 12 

Squamous cell papilloma X X X X X X X 12 

Squamous cell papilloma, multiple X X X 4

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X 3

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X 1

Parathyroid gland + + M + + + + M + M + + M + + M + + + M + + + + + 39 

Pituitary gland + + + I + + + + I + + + + + + + + + + + + + + + + 45 

Pars distalis, adenoma X X X X X 8

Pars distalis, adenoma, multiple X 1

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Follicular cell, adenoma X 1
264 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

10,000 ppm (continued)

2345556666777777777777777 

Number of Days on Study 0740585577011112333333333 

2223572719633367000000000

5455655455555555444555555 

Carcass ID Number 6976040977613935999000011 

5554545443355545123123412

General Body System 

Tissue NOS + 

Genital System

Clitoral gland M 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + 

Luteoma X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 

Polyp stromal X X 

Sarcoma stromal X 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + +

Lymph node + + + + + + 

Mediastinal, squamous cell carcinoma, 

metastatic, stomach, forestomach X 

Lymph node, mandibular + + + + + + + M + + + + + + + + + + + + + + + + M

Lymph node, mesenteric + + + M + + M + + + + + + + + + + + + + + + + + + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + +

Thymus + + + + + + M M + M + M + M + M + + + + + + + + +

Integumentary System 

Mammary gland M + + M + + M + M M M + + M + + + + + M + MMM+

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 

Subcutaneous tissue, fibrosarcoma X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + +

Skeletal muscle + + 

Abdominal, fibrosarcoma, metastatic, 

skin X 

Diaphragm, squamous cell carcinoma, 

metastatic, stomach, forestomach X 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + +

Respiratory System

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 

Alveolar/bronchiolar adenoma X X X X X X X X 

Alveolar/bronchiolar adenoma, 

multiple 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X 

Nose + + + + + + + + + + + + + + + + + + + + + + + +

Trachea + + + + + + + + + + + + + + + + + + + + + + + + +
Lesions in Female Mice 265

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

10,000 ppm (continued)

7777777777777777777777777

Number of Days on Study 3333333333333333333333333

0000000000000002333333333

5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 6 6 Total

Carcass ID Number 1 2 2 2 2 3 3 4 4 4 5 5 6 6 7 8 7 8 8 8 9 9 9 0 0 Tissues/

3 1 2 3 4 1 2 1 2 3 3 4 1 2 2 5 1 1 3 4 1 2 3 3 4 Tumors

General Body System 

Tissue NOS 1

Genital System

Clitoral gland 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Luteoma 1

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Polyp stromal 2

Sarcoma stromal X X 3

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Lymph node + + + + + + 12 

Mediastinal, squamous cell carcinoma, 

metastatic, stomach, forestomach 1

Lymph node, mandibular + + I + + + + + + + + M + + + M + + + + + + + + + 45 

Lymph node, mesenteric M + + + + + + + + + + + + + + + M + + + + + + + + 46 

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Thymus + + M M + + + + M + + + M + + M + + + + M + + + + 38 

Integumentary System 

Mammary gland M M M M M M + M M + + M + + M +++M++ M M M + 25 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Subcutaneous tissue, fibrosarcoma 1

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + 48 

Skeletal muscle 2

Abdominal, fibrosarcoma, metastatic, 

skin 1

Diaphragm, squamous cell carcinoma, 

metastatic, stomach, forestomach 1

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Respiratory System

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Alveolar/bronchiolar adenoma X X X X X X X 15 

Alveolar/bronchiolar adenoma, 

multiple X X 2

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Nose + + + + M + + M + + + + + M + + + + + + + + + + 45 

Trachea + + + + + + M + + + + + + + + + + + + + + + + + + 49 
266 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

10,000 ppm (continued)

2345556666777777777777777 

Number of Days on Study 0740585577011112333333333 

2223572719633367000000000

5455655455555555444555555 

Carcass ID Number 6976040977613935999000011 

5554545443355545123123412

Special Senses System 

Harderian gland + + 

Adenoma X 

Carcinoma X 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + +

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + +

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Lymphoma malignant mixed X X X X X 

Lymphoma malignant undifferentiated 

cell type X X 
Lesions in Female Mice 267

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

10,000 ppm (continued)

7777777777777777777777777

Number of Days on Study 3333333333333333333333333

0000000000000002333333333

5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 6 6 Total

Carcass ID Number 1 2 2 2 2 3 3 4 4 4 5 5 6 6 7 8 7 8 8 8 9 9 9 0 0 Tissues/

3 1 2 3 4 1 2 1 2 3 3 4 1 2 2 5 1 1 3 4 1 2 3 3 4 Tumors

Special Senses System 

Harderian gland + 3

Adenoma X 2

Carcinoma 1

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymphoma malignant mixed X X X X X X X X 13 

Lymphoma malignant undifferentiated 

cell type 2
268 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

20,000 ppm

5555556666666777777777777 

Number of Days on Study 0356692236689112222222222 

1832863312407383499999999

6766776677766666766666666 

Carcass ID Number 3179113320169113211122223 

4545433555244542412312341

Alimentary System 

Esophagus + + M + + + + + + M + + + + + + + + + + + + + + +

Gallbladder + A M + + + + + + M + + + + + + + + + + + + + + +

Intestine large, colon + + + + + + + + + + + + + + A + + + + + + + + + +

Intestine large, rectum + + + + + + + + + + + M + + A + + + + + + + + + +

Intestine large, cecum + + + + + + + + + + + M + + A + + + + + + + + + +

Intestine small, duodenum + A A + + + + + + + + + + + A + + + + + + + + + +

Intestine small, jejunum + A A + + + + + + + + + + + A + + + + + + + + + +

Intestine small, ileum A + A + + + + + + + + + + + A + + + + + + + + + +

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 

Hepatoblastoma X X 

Hepatocellular carcinoma X X X X X X X X X X X 

Hepatocellular carcinoma, multiple X X X X X X 

Hepatocellular adenoma X X X 

Hepatocellular adenoma, multiple X X X X X X X X X X X X X X X X X X X X X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X X X

Mesentery + + + + + + + + 

Sarcoma, metastatic, stomach, 

forestomach X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X X

Pancreas + + + + + M + + + + + + + + + + + + + + + + + + + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X 

Salivary glands + + M + + + + + + M + + + + + + + + + + + + + + +

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 

Sarcoma X 

Squamous cell carcinoma X X X X X X X X X X 

Squamous cell papilloma X X X X 

Squamous cell papilloma, multiple X X X X X X 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + 

Sarcoma X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X X

Cardiovascular System 

Heart + + M + + + + + + + + + + + + + + + + + + + + + +

Endocrine System

Adrenal cortex + + + + + + + + + + + + + M + + + + + + + + + + + 

Squamous cell carcinoma, metastatic X 

Adrenal medulla + + + + + + + + + + + + + M + + + + + + + + + + +

Islets, pancreatic + + + + + M + + + + + + + + A + + + + + + + + + +

Parathyroid gland M M M + + + + M + M + + + M M + + + + + + + M M +

Pituitary gland + + + + + + + + + M + M + M + + + M + + + + + + + 

Pars distalis, adenoma X X 

Pars nervosa, adenoma X 

Thyroid gland + + M + + + + + + M + + + + + + + + + + + + + + + 

Follicular cell, adenoma 
Lesions in Female Mice 269

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

20,000 ppm (continued)

7777777777777777777777777

Number of Days on Study 2222222222333333333333333

9999999999000000000000000

6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 7 7 7 7 Total

Carcass ID Number 4 4 4 4 5 8 8 8 8 8 5 5 5 6 6 6 7 7 7 9 9 0 0 1 2 Tissues/

1 2 4 5 5 1 2 3 4 5 2 3 4 1 2 3 1 2 3 1 3 3 4 1 3 Tumors

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + M + + + + + + + + 47 

Gallbladder + + + + + + M + + + + M + + + + + + + + + + M + + 44 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + 46 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + 46 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + 46 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hepatoblastoma 2

Hepatocellular carcinoma X X X 14 

Hepatocellular carcinoma, multiple X X X X X X X 13 

Hepatocellular adenoma X 4

Hepatocellular adenoma, multiple X X X X X X X X X X X X X X X X X X X X X X X X 45 

Squamous cell carcinoma, metastatic, 

stomach, forestomach 4

Mesentery 8

Sarcoma, metastatic, stomach, 

forestomach 1

Squamous cell carcinoma, metastatic, 

stomach, forestomach 3

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Squamous cell carcinoma, metastatic, 

stomach, forestomach 2

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + M 47 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Sarcoma 1

Squamous cell carcinoma X 11 

Squamous cell papilloma X X X X X X X X X 13 

Squamous cell papilloma, multiple X X X X X X X X 14 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + 48 

Sarcoma 1

Squamous cell carcinoma, metastatic, 

stomach, forestomach 3

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Endocrine System

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + 48 

Squamous cell carcinoma, metastatic 1

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + 48 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Parathyroid gland + M + M + + M + + + + + M + M + M + + M + + + M + 33 

Pituitary gland + + + M + + + + + + M + M + + + + + + + + + + + + 43 

Pars distalis, adenoma X X 4

Pars nervosa, adenoma 1

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Follicular cell, adenoma X X 2
270 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

20,000 ppm (continued)

5555556666666777777777777 

Number of Days on Study 0356692236689112222222222 

1832863312407383499999999

6766776677766666766666666 

Carcass ID Number 3179113320169113211122223 

4545433555244542412312341

General Body System 

None 

Genital System

Clitoral gland + 

Ovary + + + + + + + + + + + + + M M + + + + + + + + + + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X X X

Uterus + + + + + + + + + + + + + + A + + + + + + + + + +

Vagina + 

Squamous cell carcinoma X 

Hematopoietic System 

Bone marrow + + M + + + + + + + + + + + + + + + + + + + + + +

Lymph node + + + + + + + + 

Bronchial, sarcoma, metastatic, 

stomach, forestomach X 

Mediastinal, squamous cell carcinoma, 

metastatic, stomach, forestomach X 

Lymph node, mandibular + + M + + M + M + M M + + + + + + + + + + + + + I

Lymph node, mesenteric + + M + M + + + + M + + + + M + + + + + + M + + + 

Sarcoma, metastatic, stomach, 

forestomach X 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 

Squamous cell carcinoma, metastatic, 

stomach, forestomach X 

Thymus M + M + M M + + + M M M M M M + M M + + M ++++

Integumentary System 

Mammary gland + + M M + M + + M + M M + M M M M M + + M + M M M

Skin + + + + + + + + + + + + + + + + + + + + + + + + +

Musculoskeletal System

Bone + + M + + + + + + + + + + + + + + + + + + + + + +

Skeletal muscle + + 

Diaphragm, squamous cell carcinoma, 

metastatic, stomach, forestomach X 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + +

Spinal cord + 
Lesions in Female Mice 271

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

20,000 ppm (continued)

7777777777777777777777777

Number of Days on Study 2222222222333333333333333

9999999999000000000000000

6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 7 7 7 7 Total

Carcass ID Number 4 4 4 4 5 8 8 8 8 8 5 5 5 6 6 6 7 7 7 9 9 0 0 1 2 Tissues/

1 2 4 5 5 1 2 3 4 5 2 3 4 1 2 3 1 2 3 1 3 3 4 1 3 Tumors

General Body System 

None 

Genital System

Clitoral gland 1

Ovary + M + + + + + + + + + + + + + + + + + + + + + + + 47 

Squamous cell carcinoma, metastatic, 

stomach, forestomach 3

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Vagina 1

Squamous cell carcinoma 1

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Lymph node + + + + 12 

Bronchial, sarcoma, metastatic, 

stomach, forestomach 1

Mediastinal, squamous cell carcinoma, 

metastatic, stomach, forestomach 1

Lymph node, mandibular M + M + M M I + + M M M + + + M M + + + + I M + M 31 

Lymph node, mesenteric + + + + + + + + + + M + + + + + + + + + + + + + + 44 

Sarcoma, metastatic, stomach, 

forestomach 1

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Squamous cell carcinoma, metastatic, 

stomach, forestomach 1

Thymus + + + + + + + + + + + + + + + + + + + M M M + + + 34 

Integumentary System 

Mammary gland + M M M M + M M + + M + M + + M M M + M M M + + M 20 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Musculoskeletal System

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Skeletal muscle 2

Diaphragm, squamous cell carcinoma, 

metastatic, stomach, forestomach 1

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Spinal cord 1
272 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone:

20,000 ppm (continued)

5555556666666777777777777 

Number of Days on Study 0356692236689112222222222 

1832863312407383499999999

6766776677766666766666666 

Carcass ID Number 3179113320169113211122223 

4545433555244542412312341

Respiratory System 

Lung + + M + + + + + + + + + + + + + + + + + + + + + + 

Alveolar/bronchiolar adenoma X X X X X X 

Alveolar/bronchiolar adenoma, 

multiple X 

Alveolar/bronchiolar carcinoma X 

Hemangioma 

Hepatocellular carcinoma, metastatic, 

liver 

Nose + + + + + + + + + + + + + + M + + + + + + + + M +

Trachea + + M + + + + + + M + + + + + + + I + + + + + + +

Special Senses System 

Eye + 

Harderian gland + 

Adenoma X 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + +

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + +

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Lymphoma malignant mixed X X X X X X 
Lesions in Female Mice 273

TABLE D2

Individual Animal Tumor Pathology of Female Mice in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinone: 

20,000 ppm (continued)

7777777777777777777777777

Number of Days on Study 2222222222333333333333333

9999999999000000000000000

6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 7 7 7 7 Total

Carcass ID Number 4 4 4 4 5 8 8 8 8 8 5 5 5 6 6 6 7 7 7 9 9 0 0 1 2 Tissues/

1 2 4 5 5 1 2 3 4 5 2 3 4 1 2 3 1 2 3 1 3 3 4 1 3 Tumors

Respiratory System 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Alveolar/bronchiolar adenoma X X X X X X 12 

Alveolar/bronchiolar adenoma, 

multiple 1

Alveolar/bronchiolar carcinoma X 2

Hemangioma X 1

Hepatocellular carcinoma, metastatic, 

liver X 1

Nose + + + + + + + M + + + M M + + + M + + + + + + + + 44 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Special Senses System 

Eye 1

Harderian gland + 2

Adenoma X 2

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymphoma malignant mixed X X X X X X X 13 
274 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D3

Statistical Analysis of Primary Neoplasms in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

0 ppm 10,000 ppm 20,000 ppm

Harderian Gland: Adenoma or Carcinoma

Overall ratea 1/50 (2%) 3/50 (6%) 2/50 (4%)

Adjusted rateb 2.6% 7.8% 6.1%

Terminal ratec 1/39 (3%) 2/34 (6%) 2/33 (6%)

First incidence (days) 729 (T) 372 729 (T)

Life table testd P=0.348 P=0.271 P=0.442 

Logistic regression testd P=0.393 P=0.314 P=0.442 

Cochran-Armitage testd P=0.399 

Fisher exact testd P=0.309 P=0.500 

Liver: Hepatocellular Adenoma

Overall rate 6/50 (12%) 45/50 (90%) 49/50 (98%)

Adjusted rate 15.4% 95.7% 100.0%

Terminal rate 6/39 (15%) 32/34 (94%) 33/33 (100%)

First incidence (days) 729 (T) 442 501

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001

Liver: Hepatocellular Carcinoma

Overall rate 0/50 (0%) 23/50 (46%) 27/50 (54%)

Adjusted rate 0.0% 57.2% 60.8%

Terminal rate 0/39 (0%) 17/34 (50%) 16/33 (48%)

First incidence (days) )e 503 538

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001

Liver: Hepatocellular Adenoma or Carcinoma

Overall rate 6/50 (12%) 46/50 (92%) 50/50 (100%)

Adjusted rate 15.4% 97.9% 100.0%

Terminal rate 6/39 (15%) 33/34 (97%) 33/33 (100%)

First incidence (days) 729 (T) 442 501

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001

Lung: Alveolar/bronchiolar Adenoma

Overall rate 4/50 (8%) 17/50 (34%) 13/49 (27%)

Adjusted rate 9.8% 45.6% 33.5%

Terminal rate 3/39 (8%) 14/34 (41%) 9/33 (27%)

First incidence (days) 685 587 538

Life table test P=0.010 P<0.001 P=0.009

Logistic regression test P=0.017 P=0.001 P=0.015

Cochran-Armitage test P=0.018

Fisher exact test P=0.001 P=0.014
Lesions in Female Mice 275

TABLE D3

Statistical Analysis of Primary Neoplasms in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

Lung: Alveolar/bronchiolar Adenoma or Carcinoma

Overall rate 4/50 (8%) 17/50 (34%) 15/49 (31%)

Adjusted rate 9.8% 45.6% 37.9%

Terminal rate 3/39 (8%) 14/34 (41%) 10/33 (30%)

First incidence (days) 685 587 538

Life table test P=0.003 P<0.001 P=0.003

Logistic regression test P=0.006 P=0.001 P=0.005

Cochran-Armitage test P=0.006

Fisher exact test P=0.001 P=0.004

Pituitary Gland (Pars Distalis): Adenoma

Overall rate 1/43 (2%) 9/45 (20%) 4/43 (9%)

Adjusted rate 2.2% 27.2% 12.2%

Terminal rate 0/34 (0%) 7/30 (23%) 3/29 (10%)

First incidence (days) 628 671 553

Life table test P=0.145 P=0.007 P=0.151 

Logistic regression test P=0.181 P=0.009 P=0.216 

Cochran-Armitage test P=0.191 

Fisher exact test P=0.009 P=0.180 

Stomach (Forestomach): Squamous Cell Papilloma

Overall rate 2/50 (4%) 16/50 (32%) 27/50 (54%)

Adjusted rate 5.1% 41.7% 72.4%

Terminal rate 2/39 (5%) 12/34 (35%) 23/33 (70%)

First incidence (days) 729 (T) 671 538

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001

Stomach (Forestomach): Squamous Cell Carcinoma

Overall rate 0/50 (0%) 12/50 (24%) 11/50 (22%)

Adjusted rate 0.0% 30.9% 27.3%

Terminal rate 0/39 (0%) 8/34 (24%) 5/33 (15%)

First incidence (days) ) 587 501

Life table test P=0.001 P<0.001 P<0.001

Logistic regression test P=0.002 P<0.001 P<0.001

Cochran-Armitage test P=0.002

Fisher exact test P<0.001 P<0.001

Stomach (Forestomach): Squamous Cell Papilloma or Squamous Cell Carcinoma

Overall rate 2/50 (4%) 25/50 (50%) 34/50 (68%)

Adjusted rate 5.1% 60.7% 80.5%

Terminal rate 2/39 (5%) 18/34 (53%) 25/33 (76%)

First incidence (days) 729 587 501

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001
276 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D3

Statistical Analysis of Primary Neoplasms in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

Thyroid Gland (Follicular Cell): Adenoma

Overall rate 3/50 (6%) 1/50 (2%) 2/48 (4%)

Adjusted rate 7.7% 2.9% 6.1%

Terminal rate 3/39 (8%) 1/34 (3%) 2/33 (6%)

First incidence (days) 729 (T) 729 (T) 729 (T)

Life table test P=0.467N P=0.355N P=0.576N

Logistic regression test P=0.467N P=0.355N P=0.576N

Cochran-Armitage test P=0.415N

Fisher exact test P=0.309N P=0.520N

Uterus: Stromal Sarcoma

Overall rate 0/50 (0%) 3/50 (6%) 0/50 (0%)

Adjusted rate 0.0% 8.1% 0.0%

Terminal rate 0/39 (0%) 2/34 (6%) 0/33 (0%)

First incidence (days) ) 671 )

Life table test P=0.597 P=0.105 )

Logistic regression test P=0.634 P=0.116 )

Cochran-Armitage test P=0.640 

Fisher exact test P=0.121 )

Uterus: Stromal Polyp or Stromal Sarcoma 

Overall rate 0/50 (0%) 5/50 (10%) 0/50 (0%)

Adjusted rate 0.0% 12.4% 0.0%

Terminal rate 0/39 (0%) 2/34 (6%) 0/33 (0%)

First incidence (days) ) 587 )

Life table test P=0.562 P=0.029 )

Logistic regression test P=0.608 P=0.034 )

Cochran-Armitage test P=0.610 

Fisher exact test P=0.028 )

All Organs: Hemangioma

Overall rate 4/50 (8%) 0/50 (0%) 1/50 (2%)

Adjusted rate 9.6% 0.0% 3.0%

Terminal rate 3/39 (8%) 0/34 (0%) 1/33 (3%)

First incidence (days) 478 ) 729 (T)

Life table test P=0.105N P=0.080N P=0.222N

Logistic regression test P=0.082N P=0.060N P=0.182N

Cochran-Armitage test P=0.082N

Fisher exact test P=0.059N P=0.181N

All Organs: Malignant Lymphoma (Histiocytic, Lymphocytic, Mixed, or Undifferentiated Cell Type)

Overall rate 11/50 (22%) 15/50 (30%) 13/50 (26%)

Adjusted rate 25.5% 37.1% 35.7%

Terminal rate 7/39 (18%) 10/34 (29%) 10/33 (30%)

First incidence (days) 653 202 680

Life table test P=0.237 P=0.181 P=0.268 

Logistic regression test P=0.364 P=0.254 P=0.339 

Cochran-Armitage test P=0.366 

Fisher exact test P=0.247 P=0.408 
Lesions in Female Mice 277

TABLE D3

Statistical Analysis of Primary Neoplasms in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

All Organs: Benign Neoplasms

Overall rate 24/50 (48%) 46/50 (92%) 50/50 (100%)

Adjusted rate 54.1% 95.8% 100.0%

Terminal rate 19/39 (49%) 32/34 (94%) 33/33 (100%)

First incidence (days) 478 442 501

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001

All Organs: Malignant Neoplasms

Overall rate 14/50 (28%) 40/50 (80%) 38/50 (76%)

Adjusted rate 30.9% 84.9% 80.7%

Terminal rate 8/39 (21%) 27/34 (79%) 24/33 (73%)

First incidence (days) 478 202 501

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001

All Organs: Benign or Malignant Neoplasms

Overall rate 32/50 (64%) 50/50 (100%) 50/50 (100%)

Adjusted rate 66.7% 100.0% 100.0%

Terminal rate 23/39 (59%) 34/34 (100%) 33/33 (100%)

First incidence (days) 478 202 501

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P<0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001

(T)Terminal sacrifice

a Number of lesion-bearing animals/number of animals examined. Denominator is number of animals examined microscopically for liver, lung, pituitary gland, stomach, thyroid

gland, and uterus; for other tissues, denominator is number of animals necropsied. b Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality c Observed incidence at terminal kill d Beneath the control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values corresponding to pairwise comparisons

between the controls and that exposed group. The life table analysis regards neoplasms in animals dying prior to terminal kill as being (directly or indirectly) the cause of

death. The logistic regression test regards these lesions as nonfatal. The Cochran-Armitage and Fisher exact tests compare directly the overall incidence rates. For all tests,

a negative trend or a lower incidence in an exposure group is indicated by N. e Not applicable; no neoplasms in animal group
278 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D4a

Historical Incidence of Hepatocellular Neoplasms in Untreated Female B6C3F1 Micea

Incidence in Controls

Study Adenoma Carcinoma Adenoma or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 6/50 0/50 6/50

Acetaminophen 3/49 0/49 3/49

HC Yellow 4 5/50 1/50 6/50

Methylphenidate Hydrochloride 6/49 5/49 9/49

Pentaerythritol Tetranitrate 5/49 1/49 6/49

Turmeric Oleoresin 7/50 7/50 13/50

Overall Historical Incidence

Total 176/1,462 (12.0%) 89/1,462 (6.1%) 247/1,462 (16.9%)

Standard deviation 8.2% 5.4% 10.7%

Range 0%-33% 0%-20% 3%-42%

a Data as of 31 March 1993

TABLE D4b

Historical Incidence of Forestomach Squamous Cell Neoplasms in Untreated Female B6C3F1 Micea

Incidence in Controls

Study Papilloma Carcinoma Papilloma or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 2/50 0/50 2/50

Acetaminophen 0/50 0/50 0/50

HC Yellow 4 3/50 0/50 3/50

Methylphenidate Hydrochloride 1/49 0/49 1/49

Pentaerythritol Tetranitrate 1/50 0/50 1/50

Turmeric Oleoresin 0/50 0/50 0/50

Overall Historical Incidence

Total 31/1,470 (2.1%) 2/1,470 (0.1%) 33/1,470 (2.2%)

Standard deviation 2.9% 0.5% 3.1%

Range 0%-14% 0%-2% 0%-14%

a Data as of 31 March 1993
Lesions in Female Mice 279

TABLE D4c

Historical Incidence of Alveolar/bronchiolar Neoplasms in Untreated Female B6C3F1 Micea

Incidence in Controls

Study Adenoma Carcinoma Adenoma or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 4/50 0/50 4/50

Acetaminophen 1/50 0/50 1/50

HC Yellow 4 3/50 1/50 4/50

Methylphenidate Hydrochloride 1/48 0/48 1/48

Pentaerythritol Tetranitrate 2/50 1/50 3/50

Turmeric Oleoresin 4/50 1/50 5/50

Overall Historical Incidence

Total 82/1,469 (5.6%) 30/1,469 (2.1%) 110/1,469 (7.5%)

Standard deviation 4.8% 2.2% 5.0%

Range 0%-24% 0%-8% 2%-26%

a Data as of 31 March 1993

TABLE D4d

Historical Incidence of Uterine Neoplasms in Untreated Female B6C3F1 Micea

Incidence in Controls

Study Stromal Polyp Stromal Sarcoma Stromal Polyp or

Stromal Sarcoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 0/50 0/50 0/50

Acetaminophen 1/50 0/50 1/50

HC Yellow 4 2/50 0/50 2/50

Methylphenidate Hydrochloride 2/49 0/49 2/49

Pentaerythritol Tetranitrate 3/50 0/50 3/50

Tumeric Oleoresin 1/50 0/50 1/50

Overall Historical Incidence

Total 44/1,470 (3.0%) 7/1,470 (0.5%) 51/1,470 (3.5%)

Standard deviation 3.2% 0.9% 3.1%

Range 0%-16% 0%-2% 0%-16%

a Data as of 31 March 1993
280 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D4e

Historical Incidence of Pituitary Gland Pars Distalis Neoplasms in Untreated Female B6C3F1 Micea

Incidence in Controls

Study Adenoma Carcinoma Adenoma or Carcinoma

Historical Incidence at EG&G Mason Research Institute

1-Amino-2,4-dibromoanthraquinone 1/43 0/431/43

Acetaminophen 14/46 1/46 15/46

HC Yellow 4 5/42 0/425/42

Methylphenidate Hydrochloride 7/48 0/487/48

Pentaerythritol Tetranitrate 8/45 1/459/45

Tumeric Oleoresin 0/46 0/460/46

Overall Historical Incidence

Total 212/1,392 (15.2%) 8/1,392 (0.6%) 220/1,392 (15.8%)

 Standard deviation 9.9% 1.0% 10.3%

Range 0%-36% 0%-4% 0%-36%

a Data as of 31 March 1993
Lesions in Female Mice 281

TABLE D5

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 10,000 ppm 20,000 ppm

Disposition Summary

Animals initially in study 60 60 60

15-Month interim evaluation 10 10 10

Early deaths

Moribund 5 11 11

Natural deaths 6 5 6

Survivors

Died last week of study 1 1

Terminal sacrifice 38 33 33

Animals examined microscopically 60 60 60

15-Month Interim Evaluation

Alimentary System

Gallbladder (10) (10) (10)

Inflammation, chronic 1 (10%) 1 (10%)

Inflammation, chronic active 1 (10%)

Liver (10) (10) (10)

Basophilic focus 1 (10%) 1 (10%)

Fatty change 1 (10%) 1 (10%) 3 (30%)

Inflammation, acute 5 (50%) 4 (40%)

Inflammation, chronic active 9 (90%) 4 (40%) 6 (60%)

Necrosis, coagulative 7 (70%) 8 (80%) 9 (90%)

Pigmentation 10 (100%) 9 (90%)

Bile duct, hyperplasia, focal 1 (10%)

Pancreas (10) (10) (10)

Inflammation, chronic 5 (50%) 1 (10%)

Vacuolization cytoplasmic 1 (10%)

Salivary glands (10) (10) (10)

Inflammation, chronic 5 (50%)

Submandibular gland, inflammation, chronic 7 (70%) 4 (40%)

Stomach, forestomach (10) (10) (10)

Acanthosis 1 (10%) 8 (80%)

Hyperkeratosis 2 (20%) 7 (70%)

Hyperplasia, basal cell 2 (20%)

Inflammation, chronic 2 (20%)

Inflammation, chronic active 1 (10%) 3 (30%)

Stomach, glandular (10) (10) (10)

Inflammation, chronic 2 (20%) 1 (10%)

Inflammation, chronic active 1 (10%)

Cardiovascular System

Heart (10) (10) (10)

Cardiomyopathy 2 (20%)

Endocrine System

Pituitary gland (8) (10) (9)

Pars distalis, cyst 1 (11%)

Pars distalis, hyperplasia 1 (13%) 1 (11%)

a Number of animals examined microscopically at site and number of animals with lesion
282 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D5

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

15-Month Interim Evaluation (continued)

Genital System

Ovary (10) (2) (10)

Cyst 1 (10%) 1 (50%) 2 (20%)

Periovarian tissue, cyst 1 (50%)

Periovarian tissue, inflammation, chronic 2 (20%)

Uterus (10) (10) (10)

Hydrometra 8 (80%) 3 (30%)

Endometrium, hyperplasia 6 (60%) 10 (100%)

Hematopoietic System

Bone marrow (10) (10) (10)

Myelofibrosis 2 (20%) 1 (10%)

Lymph node, mandibular (6) (10) (10)

Pigmentation 1 (17%)

Lymph node, mesenteric (7) (10) (8)

Depletion lymphoid 1 (14%)

Infiltration cellular, histiocyte 4 (57%) 7 (88%)

Pigmentation 3 (43%) 7 (88%)

Spleen (10) (10) (9)

Depletion lymphoid 1 (11%)

Thymus (9) (10) (8)

Cyst 6 (67%) 6 (75%)

Nervous System

Brain (10) (10) (10)

Thalamus, mineralization 3 (30%) 4 (40%)

Respiratory System

Nose (10) (10) (9)

Glands, inflammation, acute 2 (20%) 1 (11%)

Urinary System

Kidney (10) (4) (10)

Inflammation, chronic 10 (100%) 4 (100%) 9 (90%)

Renal tubule, regeneration 2 (20%) 1 (10%)

Urinary bladder (10) (10) (10)

Inflammation, chronic 7 (70%) 7 (70%) 8 (80%)

Systems Examined With No Lesions Observed

General Body System

Integumentary System

Musculoskeletal System

Special Senses System
Lesions in Female Mice 283

TABLE D5

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study

Alimentary System

Esophagus (50) (45) (47)

Autolysis 1 (2%)

Gallbladder (49) (47) (44)

Autolysis 3 (6%) 2 (4%) 2 (5%)

Inflammation, chronic active 5 (10%) 6 (13%) 8 (18%)

Epithelium, hyperplasia 1 (2%) 

Intestine large, colon (50) (50) (48)

Autolysis 3 (6%) 1 (2%)

Inflammation, acute 1 (2%)

Peyer's patch, hyperplasia 1 (2%) 1 (2%)

Intestine large, rectum (50) (50) (47)

Autolysis 2 (4%) 1 (2%)

Peyer's patch, epithelium, proliferation 1 (2%)

Intestine large, cecum (50) (49) (47)

Autolysis 4 (8%) 1 (2%)

Peyer's patch, hyperplasia 1 (2%) 2 (4%)

Intestine small, duodenum (48) (50) (46)

Angiectasis 1 (2%)

Autolysis 2 (4%) 2 (4%) 2 (4%)

Intestine small, jejunum (47) (50) (46)

Autolysis 1 (2%) 2 (4%) 2 (4%)

Peyer's patch, hyperplasia 2 (4%)

Intestine small, ileum (48) (50) (46)

Autolysis 2 (4%) 2 (4%) 2 (4%)

Inflammation, acute 1 (2%)

Peyer's patch, hyperplasia 4 (8%) 3 (6%) 1 (2%)

Peyer's patch, inflammation, acute 1 (2%)

Liver (50) (50) (50)

Autolysis 2 (4%) 1 (2%)

Basophilic focus 4 (8%) 5 (10%)

Clear cell focus 10 (20%) 9 (18%)

Eosinophilic focus 4 (8%) 2 (4%)

Fatty change 2 (4%) 3 (6%) 15 (30%)

Fibrosis 1 (2%)

Hematopoietic cell proliferation 3 (6%) 6 (12%) 6 (12%)

Hemorrhage 1 (2%)

Inflammation, chronic active 27 (54%) 24 (48%) 23 (46%)

Mixed cell focus 2 (4%)

Necrosis, coagulative 9 (18%) 18 (36%) 16 (32%)

Pigmentation 44 (88%) 49 (98%)

Bile duct, hyperplasia 1 (2%)

Mesentery (6) (9) (8)

Fibrosis 4 (44%)

Inflammation, chronic active 3 (50%) 5 (56%) 4 (50%)

Mineralization 1 (17%)

Necrosis, coagulative 1 (17%) 3 (33%)

Pigmentation 1 (17%)
284 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D5

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Alimentary System (continued)

Pancreas (50) (50) (49)

Atrophy 1 (2%) 2 (4%)

Autolysis 2 (4%)

Cytoplasmic alteration 3 (6%) 1 (2%) 

Fibrosis 1 (2%)

Inflammation 1 (2%)

Inflammation, chronic active 25 (50%) 24 (48%) 18 (37%)

Vacuolization cytoplasmic 19 (38%) 13 (26%) 21 (43%)

Duct, dilatation 1 (2%) 1 (2%) 1 (2%)

Duct, hyperplasia 1 (2%)

Salivary glands (49) (50) (47)

Inflammation, chronic 1 (2%)

Parotid gland, autolysis 1 (2%)

Sublingual gland, inflammation, chronic 1 (2%) 2 (4%)

Submandibular gland, autolysis 1 (2%)

Submandibular gland, inflammation, chronic 40 (82%) 32 (64%) 29 (62%)

Stomach, forestomach (48) (50) (50)

Acanthosis 9 (19%) 15 (30%) 19 (38%)

Autolysis 1 (2%)

Erosion 1 (2%) 1 (2%)

Hyperkeratosis 10 (21%) 14 (28%) 17 (34%)

Hyperplasia, basal cell 7 (14%) 3 (6%)

Hyperplasia, squamous 2 (4%)

Inflammation, chronic active 7 (15%) 10 (20%) 21 (42%)

Ulcer 2 (4%) 1 (2%) 2 (4%)

Ulcer, multiple 1 (2%)

Stomach, glandular (49) (48) (48)

Autolysis 2 (4%) 1 (2%)

Inflammation, chronic 1 (2%)

Inflammation, chronic active 20 (41%) 19 (40%) 21 (44%)

Mineralization 1 (2%) 1 (2%)

Tooth (1)

Abscess 1 (100%)

Dysplasia 1 (100%)

Cardiovascular System

Heart (50) (50) (49)

Autolysis 1 (2%)

Inflammation, chronic 9 (18%) 6 (12%) 8 (16%)

Mineralization 1 (2%) 1 (2%)

Thrombosis 1 (2%)
Lesions in Female Mice 285

TABLE D5

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Endocrine System

Adrenal cortex (50) (49) (48)

Angiectasis 2 (4%)

Autolysis 1 (2%)

Cyst 1 (2%)

Degeneration, fatty 1 (2%)

Hematopoietic cell proliferation 1 (2%) 3 (6%) 1 (2%)

Hyperplasia 1 (2%) 1 (2%)

Inflammation, chronic active 2 (4%) 1 (2%)

Vacuolization cytoplasmic 1 (2%)

Adrenal medulla (49) (49) (48)

Autolysis 1 (2%)

Hematopoietic cell proliferation 1 (2%) 1 (2%)

Hyperplasia 1 (2%)

Islets, pancreatic (49) (50) (48)

Hyperplasia 3 (6%) 1 (2%) 1 (2%)

Parathyroid gland (33) (39) (33)

Cyst 1 (3%)

Pituitary gland (43) (45) (43)

Autolysis 1 (2%)

Pars distalis, angiectasis 1 (2%)

Pars distalis, cyst 3 (7%)

Pars distalis, hyperplasia 7 (16%) 22 (49%) 7 (16%)

Pars distalis, pigmentation 1 (2%)

Thyroid gland (50) (50) (48)

Autolysis 1 (2%)

Inflammation, chronic active 1 (2%) 2 (4%) 3 (6%)

C-cell, hyperplasia 1 (2%) 1 (2%) 1 (2%)

Follicle, cyst 2 (4%) 2 (4%)

Follicular cell, hyperplasia 3 (6%) 19 (38%) 18 (38%)

General Body System

None 

Genital System

Clitoral gland (1)

Duct, dilatation 1 (100%)

Ovary (49) (49) (47)

Abscess 3 (6%) 2 (4%)

Angiectasis 2 (4%) 4 (8%) 4 (9%)

Cyst 12 (24%) 11 (22%) 7 (15%)

Cyst, multiple 1 (2%) 3 (6%) 1 (2%)

Hemorrhage 1 (2%)

Inflammation, acute, necrotizing 1 (2%)

Mineralization 2 (4%)

Pigmentation 1 (2%)
286 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D5

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Genital System (continued)

Ovary (continued) (49) (49) (47)

Periovarian tissue, cyst 1 (2%) 1 (2%)

Periovarian tissue, cyst, multiple 1 (2%)

Periovarian tissue, hemorrhage 1 (2%)

Periovarian tissue, inflammation, chronic

 active 29 (59%) 24 (49%) 28 (60%)

Periovarian tissue, mineralization 1 (2%)

Periovarian tissue, pigmentation 1 (2%)

Uterus (49) (50) (49)

Angiectasis 1 (2%)

Fibrosis 2 (4%) 1 (2%)

Hydrometra 14 (29%) 3 (6%)

Inflammation, acute 1 (2%)

Inflammation, chronic active 4 (8%) 6 (12%) 7 (14%)

Endometrium, hyperplasia 38 (78%) 36 (72%) 35 (71%)

Epithelium, metaplasia, squamous 2 (4%) 1 (2%) 1 (2%)

Hematopoietic System

Bone marrow (50) (49) (49)

Myelofibrosis 5 (10%)

Myeloid cell, sternal, hyperplasia 2 (4%) 1 (2%) 2 (4%)

Sternal, autolysis 1 (2%)

Sternal, myelofibrosis 38 (76%) 32 (65%) 34 (69%)

Lymph node (7) (12) (12)

Lumbar, hyperplasia, lymphoid 1 (14%) 1 (8%)

Lumbar, inflammation, acute 1 (14%)

Mediastinal, hyperplasia, lymphoid 1 (8%)

Mediastinal, hyperplasia, plasma cell 1 (8%)

Mediastinal, inflammation, chronic active 1 (14%)

Pancreatic, hyperplasia, lymphoid 1 (8%) 1 (8%)

Pancreatic, infiltration cellular, histiocyte 1 (8%)

Pancreatic, pigmentation 1 (8%)

Renal, hyperplasia, plasma cell 2 (17%)

Renal, sinus, ectasia 1 (8%)

Lymph node, mandibular (32) (45) (31)

Autolysis 1 (3%)

Congestion 1 (3%)

Depletion lymphoid 1 (2%)

Hyperplasia, lymphoid 3 (9%) 1 (2%) 2 (6%)

Hyperplasia, plasma cell 1 (3%) 1 (3%)

Inflammation, acute 1 (2%)

Pigmentation 2 (6%) 2 (4%) 3 (10%)

Sinus, ectasia 2 (4%)
Lesions in Female Mice 287

TABLE D5

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Hematopoietic System (continued)

Lymph node, mesenteric (49) (46) (44)

Angiectasis 7 (14%) 4 (9%) 3 (7%)

Depletion lymphoid 1 (2%) 1 (2%)

Hemorrhage 2 (4%)

Hyperplasia, lymphoid 2 (4%) 6 (13%) 10 (23%)

Infiltration cellular, histiocyte 32 (65%) 24 (52%) 27 (61%)

Inflammation, chronic active 1 (2%) 3 (7%) 2 (5%)

Mineralization 1 (2%)

Necrosis, coagulative 1 (2%)

Pigmentation 31 (63%) 24 (52%) 27 (61%)

Sinus, ectasia 1 (2%)

Spleen (50) (50) (50)

Autolysis 1 (2%) 1 (2%)

Depletion lymphoid 4 (8%) 2 (4%) 2 (4%)

Fibrosis 1 (2%)

Hematopoietic cell proliferation 6 (12%) 13 (26%) 11 (22%)

Hyperplasia, lymphoid 15 (30%) 14 (28%) 8 (16%)

Inflammation, chronic 1 (2%)

Thymus (40) (38) (34)

Angiectasis 1 (3%)

Cyst 14 (35%) 16 (42%) 22 (65%)

Depletion lymphoid 4 (10%) 2 (5%) 6 (18%)

Hemorrhage 1 (3%) 1 (3%)

Hyperplasia, lymphoid 3 (8%) 1 (3%) 1 (3%)

Integumentary System

Mammary gland (25) (25) (20)

Hyperplasia 2 (8%) 9 (36%) 3 (15%)

Inflammation, chronic 1 (4%)

Skin (50) (50) (50)

Acanthosis 1 (2%)

Autolysis 1 (2%)

Inflammation, acute 1 (2%)

Epidermis, inflammation, acute 1 (2%)

Subcutaneous tissue, cyst epithelial

 inclusion 1 (2%)

Subcutaneous tissue, fibrosis 1 (2%)

Subcutaneous tissue, inflammation, chronic

 active 1 (2%) 2 (4%) 2 (4%)

Subcutaneous tissue, mineralization 1 (2%)

Musculoskeletal System

None 

Nervous System

Brain (49) (50) (50)

Perivascular, inflammation, chronic 1 (2%)

Thalamus, mineralization 21 (43%) 15 (30%) 21 (42%)
288 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE D5

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 10,000 ppm 20,000 ppm

2-Year Study (continued)

Respiratory System

Lung (50) (50) (49)

Autolysis 1 (2%)

Congestion 1 (2%)

Hemorrhage 4 (8%) 4 (8%) 8 (16%)

Infiltration cellular, histiocyte 1 (2%)

Inflammation, chronic active 3 (6%) 4 (8%) 6 (12%)

Leukocytosis 1 (2%) 1 (2%)

Alveolar epithelium, hyperplasia 1 (2%)

Pleura, inflammation, acute 1 (2%)

Nose (48) (45) (44)

Inflammation, acute 21 (44%) 9 (20%) 7 (16%)

Nasolacrimal duct, hyperplasia 1 (2%)

Nasolacrimal duct, inflammation, acute 1 (2%)

Trachea (50) (49) (47)

Autolysis 1 (2%)

Special Senses System

Eye (1) (1)

Cornea, inflammation, chronic active 1 (100%)

Cornea, neovascularization 1 (100%)

Harderian gland (1) (3) (2)

Inflammation, chronic 1 (100%) 2 (100%)

Urinary System

Kidney (50) (50) (50)

Autolysis 1 (2%)

Glomerulosclerosis 2 (4%) 3 (6%) 4 (8%)

Hydronephrosis 1 (2%) 1 (2%)

Infarct 1 (2%)

Inflammation, chronic 43 (86%) 42 (84%) 45 (90%)

Metaplasia, osseous 3 (6%) 1 (2%)

Papilla, mineralization 1 (2%)

Pelvis, transitional epithelium, hyperplasia 1 (2%)

Proximal convoluted renal tubule,

 degeneration, hyaline 1 (2%)

Renal tubule, atrophy 1 (2%) 3 (6%)

Renal tubule, casts protein 1 (2%)

Renal tubule, mineralization 3 (6%)

Renal tubule, pigmentation 43 (86%) 43 (86%)

Renal tubule, regeneration 1 (2%) 7 (14%) 3 (6%)

Transitional epithelium, mineralization 1 (2%)

Autolysis 3 (6%) 1 (2%) 2 (4%)

Inflammation, chronic active 42 (86%) 43 (86%) 46 (92%)

Arteriole, necrosis, fibrinoid 1 (2%)

Submucosa, proliferation 1 (2%)
289

APPENDIX E

SUMMARY OF LESIONS IN MALE RATS

IN THE STOP-EXPOSURE EVALUATION

OF 1-AMINO-2,4-DIBROMOANTHRAQUINONE

TABLE E1 Summary of the Incidence of Neoplasms in Male Rats 

in the Stop-Exposure Evaluation of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . 290

TABLE E2a Statistical Analysis of Primary Neoplasms in Male Rats 

in the Stop-Exposure Evaluation of 1-Amino-2,4-dibromoanthraquinone: 

15-Month Interim Evaluation Control Group versus

9-Month and 15-Month 20,000 ppm Groups at the 15-Month Evaluation . . . . . . . . . . . . . 294

TABLE E2b Statistical Analysis of Primary Neoplasms in Male Rats 

in the Stop-Exposure Evaluation of 1-Amino-2,4-dibromoanthraquinone: 

9-Month 20,000 ppm Stop-Exposure Group versus

15-Month 20,000 ppm Exposure Group at the 15-Month Evaluation . . . . . . . . . . . . . . . . 297

TABLE E3 Summary of the Incidence of Nonneoplastic Lesions in Male Rats 

in the Stop-Exposure Evaluation of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . 300
290 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE E1

Summary of the Incidence of Neoplasms in Male Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

Disposition Summary

Animals initially in study 70 10 30

9-Month interim evaluation 10 10

Early deaths

Moribund 3 2

Natural death 1

Survivors 7 17

Animals examined microscopically 10 10 20

9-Month Interim Evaluationb

Alimentary System

Intestine large, colon (10) (10)

Polyp adenomatous 1 (10%)

Liver (10) (10)

Hepatocellular carcinoma 2 (20%)

Hepatocellular adenoma 2 (20%)

Endocrine System

Adrenal medulla (10) (10)

Thyroid gland (10) (10)

Follicular cell, adenoma 1 (10%)

Nervous System

Brain (10) (10)

 Cerebrum, meningioma benign 1 (10%)

Respiratory System

Lung (10) (10)

Alveolar/bronchiolar adenoma 1 (10%)

Urinary System

Urinary bladder (10) (9)

Papilloma 1 (11%)

Systems Examined With No Neoplasms Observed

Cardiovascular System

General Body System

Genital System

Hematopoietic System

Integumentary System

Musculoskeletal System

Special Senses System
Lesions in Male Stop-Exposure Rats 291

TABLE E1

Summary of the Incidence of Neoplasms in Male Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

15-Month Evaluationc

Alimentary System

Intestine large, rectum (9) (10) (20)

Adenocarcinoma 2 (10%)

Polyp adenomatous 2 (20%) 6 (30%)

Polyp adenomatous, multiple 1 (10%) 1 (5%)

Liver (10) (10) (20)

Hepatocellular carcinoma 4 (40%) 6 (30%)

Hepatocellular carcinoma, multiple 3 (30%) 13 (65%)

Hepatocellular adenoma 7 (70%) 3 (15%)

Hepatocholangiocarcinoma 1 (10%) 5 (25%)

Pancreas (10) (10)

Adenoma 1 (10%)

Stomach, forestomach (10) (10) (20)

Squamous cell papilloma 1 (5%)

Cardiovascular System

Heart (10) (10) (20)

Hepatocholangiocarcinoma, metastatic, liver 1 (10%)

Endocrine System

Pituitary gland (8) (10) (19)

Pars distalis, adenoma 1 (13%) 1 (5%)

Thyroid gland (10) (10)

Adenoma 1 (10%) 

C-cell, adenoma 1 (10%) 

General Body System

None

Genital System

Epididymis (20)

Preputial gland (9) (19)

Carcinoma 1 (5%)

Testes (10) (10) (20)

Adenoma 1 (10%)

Bilateral, interstitial cell, adenoma 1 (10%)

Interstitial cell, adenoma 2 (20%) 3 (30%) 3 (15%)

Hematopoietic System

Lymph node (3)

Mediastinal, hepatocellular carcinoma,

metastatic, liver 1 (33%)
292 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE E1

Summary of the Incidence of Neoplasms in Male Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

15-Month Evaluation (continued)

Integumentary System

None

Musculoskeletal System

None

Nervous System

None

Respiratory System

Lung (10) (10) (20)

Alveolar/bronchiolar adenoma 1 (10%)

Hepatocellular carcinoma, metastatic, liver 5 (50%) 7 (35%)

Hepatocholangiocarcinoma, metastatic, liver 1 (10%)

Special Senses System

None

Urinary System

Kidney (10) (10) (20)

Hepatocellular carcinoma, metastatic, liver 1 (5%)

Renal tubule, adenoma 3 (30%) 2 (10%)

Urinary bladder (10) (10) (19)

Transitional epithelium, papilloma 3 (16%)

Squamous cell carcinoma 1 (5%)

Transitional epithelium, carcinoma 1 (5%)

Systemic Lesions

Multiple organsd (10) (10) (20)

Mesothelioma malignant 1 (5%)
Lesions in Male Stop-Exposure Rats 293

TABLE E1

Summary of the Incidence of Neoplasms in Male Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

Neoplasm Summary

Total animals with primary neoplasmse 7 10 20

Total primary neoplasms 8 25 50

Total animals with benign neoplasms 7 10 15

Total benign neoplasms 8 17 25

Total animals with malignant neoplasms 8 19

Total malignant neoplasms 8 25

Total animals with metastatic neoplasms 6 7

Total metastatic neoplasms 7 9

a Number of animals examined microscopically at site and number of animals with neoplasm (includes interim and moribund animals) b Controls from the 9-month interim evaluation of the 2-year core study were used for comparison. c Controls from the 15-month interim evaluation of the 2-year core study were used for comparison. d Number of animals with any tissue examined microscopically e Primary neoplasms: all neoplasms except metastatic neoplasms
294 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE E2a

Statistical Analysis of Primary Neoplasms in Male Rats in the Stop-Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone: 15-Month Interim Evaluation Control Group versus 9-Month 

and 15-Month 20,000 ppm Groups at the 15-Month Evaluation

0 ppm 20,000 ppm 20,000 ppm

(15-month (9-month stop-exposure) (15-month exposure)

interim evaluation)

Kidney (Renal Tubule): Adenoma

Overall ratea 0/10 (0%) 3/10 (30%) 2/20 (10%)

Adjusted rateb 0.0% 60.0% 18.2%

Interim evaluation 0/10 (0%) 3/7 (43%) 2/17 (12%)

First incidence (days) )d 458 (I) 457 (I)

Life table testc P=0.336 P=0.056 P=0.360

Logistic regression testc P=0.336 P=0.056 P=0.360

Cochran-Armitage testc P=0.385

Fisher exact testc P=0.105 P=0.437

Large Intestine (Rectum): Adenomatous Polyp 

Overall rate 0/10 (0%) 3/10 (30%) 7/20 (35%)

Adjusted rate 0.0% 38.3% 66.3%

Interim evaluation 0/10 (0%) 2/7 (29%) 7/17 (41%)

First incidence (days) ) 414 456 (I)

Life table test P=0.030 P=0.080 P=0.031

Logistic regression test P=0.034 P=0.232 

P=0.031 

Cochran-Armitage test P=0.037

Fisher exact test P=0.105 P=0.038

Large Intestine (Rectum): Carcinoma

Overall rate 0/10 (0%) 0/10 2/20 (10%)

Adjusted rate 0.0% 0.0% 29.2%

Interim evaluation 0/10 (0%) 0/7 (0%) 1/17 (6%)

First incidence (days) ) ) 427

Life table test P=0.229 P=0.369

Logistic regression test P=0.284 P=0.502

Cochran-Armitage test P=0.239

Fisher exact test P=0.437

Liver: Hepatocellular Adenoma

Overall rate 0/10 (0%) 7/10 (70%) 8/20 (40%)

Adjusted rate 0.0% 86.3% 69.6%

Interim evaluation 0/10 (0%) 5/7 (71%) 7/17 (41%)

First incidence (days) ) 414 423

Life table test P=0.038 P=0.002 P=0.023

Logistic regression test P=0.049 P=0.006 P=0.026

Cochran-Armitage test P=0.043

Fisher exact test P=0.002 P=0.022

Liver: Hepatocellular Carcinoma

Overall rate 0/10 (0%) 7/10 (70%) 19/20 (95%)

Adjusted rate 0.0% 77.8% 100.0%

Interim evaluation 0/10 (0%) 5/7 (71%) 16/17 (94%)

First incidence (days) ) 442 400

Life table test P<0.001 P=0.001 P<0.001

Logistic regression test P<0.001 P=0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P=0.002 P<0.001
Lesions in Male Stop-Exposure Rats 295

TABLE E2a

Statistical Analysis of Primary Neoplasms in Male Rats in the Stop-Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone: 15-Month Interim Evaluation Control Group versus 9-Month 

and 15-Month 20,000 ppm Groups at the 15-Month Evaluation (continued)

0 ppm 20,000 ppm 20,000 ppm

(15-month (9-month stop-exposure) (15-month exposure)

interim evaluation)

Liver: Hepatocellular Adenoma or Carcinoma

Overall rate 0/10 (0%) 9/10 (90%) 20/20 (100%)

Adjusted rate 0.0% 90.0% 100.0%

Interim evaluation 0/10 (0%) 6/7 (86%) 17/17 (100%)

First incidence (days) ) 414 400

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P=0.001 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001

Pituitary Gland (Pars Distalis): Adenoma

Overall rate 1/8 (13%) 0/10 (0%) 1/19 (5%)

Adjusted rate 10.0% 0.0% 10.0%

Interim evaluation 1/8 (13%) 0/7 (0%) 1/16 (6%)

First incidence (days) 456 (I) ) 457 (I)

Life table test P=0.498N P=0.527N P=0.601N

Logistic regression test P=0.498N P=0.527N P=0.601N

Cochran-Armitage test P=0.462N

Fisher exact test P=0.444N P=0.513N

Testes: Adenoma

Overall rate 3/10 (30%) 4/10 (40%) 3/20 (15%)

Adjusted rate 46.7% 65.0% 35.8%

Interim evaluation 3/10 (30%) 3/7 (43%) 3/17 (18%)

First incidence (days) 456 (I) 447 456 (I)

Life table test P=0.328N P=0.300 P=0.397N

Logistic regression test P=0.306N P=0.395 P=0.397N

Cochran-Armitage test P=0.238N

Fisher exact test P=0.500 P=0.306N

Urinary Bladder: Papilloma

Overall rate 0/10 (0%) 0/10 (0%) 3/19 (16%)

Adjusted rate 0.0% 0.0% 38.6%

Interim evaluation 0/10 (0%) 0/7 (0%) 3/17 (18%)

First incidence (days) ) ) 457 (I)

Life table test P=0.126 P=0.223

Logistic regression P=0.126 P=0.223

Cochran-Armitage test P=0.128

Fisher exact test P=0.265

Urinary Bladder: Carcinoma

Overall rate 0/10 (0%) 0/10 (0%) 1/19 (5%)

Adjusted rate 0.0% 0.0% 25.0%

Interim evaluation 0/10 (0%) 0/7 (0%) 1/17 (6%)

First incidence (days) ) ) 458 (I)

Life table test P=0.430 P=0.606

Logistic regression P=0.430 P=0.606

Cochran-Armitage test P=0.442

Fisher exact test P=0.655
296 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE E2a

Statistical Analysis of Primary Neoplasms in Male Rats in the Stop-Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone: 15-Month Interim Evaluation Control Group versus 9-Month 

and 15-Month 20,000 ppm Groups at the 15-Month Evaluation (continued)

0 ppm 20,000 ppm 20,000 ppm

(15-month (9-month stop-exposure) (15-month exposure)

interim evaluation)

All Organs: Benign Neoplasms

Overall rate 7/10 (70%) 10/10 (100%) 15/20 (75%)

Adjusted rate 73.3% 100.0% 93.4%

Interim evaluation 7/10 (70%) 7/7 (100%) 14/17 (82%)

First incidence (days) 456 (I) 414 423

Life table test P=0.289 P=0.024 P=0.296

Logistic regression test P=0.420 P=0.178 P=0.388 

Cochran-Armitage test P=0.538

Fisher exact test P=0.105 P0.548 

All Organs: Malignant Neoplasms

Overall rate 0/10 (0%) 8/10 (80%) 19/20 (95%)

Adjusted rate 0.0% 80.0% 100.0%

Interim evaluation 0/10 (0%) 5/7 (71%) 16/17 (94%)

First incidence (days) ) 414 400

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P=0.005 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001

All Organs: Benign or Malignant Neoplasms

Overall rate 7/10 (70%) 10/10 (100%) 20/20 (100%)

Adjusted rate 73.3% 100.0% 100.0%

Interim evaluation 7/10 (70%) 7/7 (100%) 17/17 (100%)

First incidence (days) 456 (I) 414 400

Life table test P=0.020 P=0.024

P=0.018

Logistic regression test P=0.015 P=0.178 P=0.042 

Cochran-Armitage test P=0.008

Fisher exact test P=0.105 P=0.030

(I) Interim evaluation

a Number of lesion-bearing animals/number of animals examined (includes interim and moribund animals). Denominator is number of animals examined microscopically for

kidney, liver, pituitary gland, testes, and urinary bladder; for other tissues, denominator is number of animals necropsied. b Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality c Beneath the exposed group incidence are the P values corresponding to pairwise comparisons between the controls and that exposed group. The life table analysis regards

neoplasms in animals dying prior to terminal kill as being (directly or indirectly) the cause of death. The logistic regression test regards these lesions as nonfatal. The Fisher

exact test compares directly the overall incidence rates. For all tests, a lower incidence in an exposure group is indicated by N. d Not applicable; no neoplasms in animal group
Lesions in Male Stop-Exposure Rats 297

TABLE E2b

Statistical Analysis of Primary Neoplasms in Male Rats in the Stop-Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone: 9-Month 20,000 ppm Stop-Exposure Group

versus 15-Month 20,000 ppm Exposure Group at the 15-Month Evaluation 

20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

Kidney (Renal Tubule): Adenoma

Overall ratea 3/10 (30%) 2/20 (10%)

Adjusted rateb 60.0% 18.2%

Interim evaluation 3/7 (43%) 2/17 (12%)

First incidence (days) 458 (I) 457 (I)

Life table testc P=0.130N

Logistic regression testc P=0.130N

Fisher exact testc P=0.191N

Large Intestine (Rectum): Adenomatous Polyp

Overall rate 3/10 (30%) 7/20 (35%)

Adjusted rate 38.3% 66.3%

Interim evaluation 2/7 (29%) 7/17 (41%)

First incidence (days) 414 456 (I)

Life table test P=0.652N

Logistic regression test P=0.561

Fisher exact test P=0.560

Large Intestine (Rectum): Carcinoma

Overall rate 0/10 (0%) 2/20 (10%)

Adjusted rate 0.0% 29.2%

Interim evaluation 0/7 (0%) 1/17 (6%)

First incidence (days) )d 427

Life table test P=0.424

Logistic regression test P=0.400

Fisher exact test P=0.437

Liver: Hepatocellular Adenoma

Overall rate 7/10 (70%) 8/20 (40%)

Adjusted rate 86.3% 69.6%

Interim evaluation 5/7 (71%) 7/17 (41%)

First incidence (days) 414 423

Life table test P=0.078N

Logistic regression test P=0.122N

Fisher exact test P=0.123N

Liver: Hepatocellular Carcinoma

Overall rate 7/10 (70%) 19/20 (95%)

Adjusted rate 77.8% 100.0%

Interim evaluation 5/7 (71%) 16/17 (94%)

First incidence (days) 442 400

Life table test P=0.415

Logistic regression test P=0.091

Fisher exact test P=0.095
298 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE E2b

Statistical Analysis of Primary Neoplasms in Male Rats in the Stop-Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone: 9-Month 20,000 ppm Stop-Exposure Group

versus 15-Month 20,000 ppm Exposure Group at the 15-Month Evaluation (continued)

20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

Liver: Hepatocellular Adenoma or Carcinoma

Overall rate 9/10 (90%) 20/20 (100%)

Adjusted rate 90.0% 100.0%

Interim evaluation 6/7 (86%) 17/17 (100%)

First incidence (days) 414 400

Life table test P=0.566N

Logistic regression test P=0.323

Fisher exact test P=0.333

Pituitary Gland (Pars Distalis): Adenoma

Overall rate 0/10 (0%) 1/19 (5%)

Adjusted rate 0.0% 10.0%

Interim evaluation 0/7 (0%) 1/16 (6%)

First incidence (days) ) 457 (I)

Life table test P=0.665

Logistic regression test P=0.665

Fisher exact test P=0.655

Testes: Adenoma

Overall rate 4/10 (40%) 3/20 (15%)

Adjusted rate 65.0% 35.8%

Interim evaluation 3/7 (43%) 3/17 (18%)

First incidence (days) 447 456 (I)

Life table test P=0.095N

Logistic regression test P=0.120N

Fisher exact test P=0.143N

Urinary Bladder: Papilloma

Overall rate 0/10 (0%) 3/19 (16%)

Adjusted rate 0.0% 38.6%

Interim evaluation 0/7 (0%) 3/17 (18%)

First incidence (days) ) 457 (I)

Life table test P=0.309

Logistic regression test P=0.309

Fisher exact test P=0.265

Urinary Bladder: Carcinoma

Overall rate 0/10 (0%) 1/19 (5%)

Adjusted rate 0.0% 25.0%

Interim evaluation 0/7 (0%) 1/17 (6%)

First incidence (days) ) 458 (I)

Life table test P=0.677

Logistic regression test P=0.677

Fisher exact test P=0.655
Lesions in Male Stop-Exposure Rats 299

TABLE E2b

Statistical Analysis of Primary Neoplasms in Male Rats in the Stop-Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone: 9-Month 20,000 ppm Stop-Exposure Group 

versus 15-Month 20,000 ppm Exposure Group at the 15-Month Evaluation (continued)

20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

All Organs: Benign Neoplasms

Overall rate 10/10 (100%) 15/20 (75%)

Adjusted rate 100.0% 93.4%

Interim evaluation 7/7 (100%) 14/17 (82%)

First incidence (days) 414 423

Life table test P=0.045N

Logistic regression test P=0.100N

Fisher exact test P=0.109N

All Organs: Malignant Neoplasms

Overall rate 8/10 (80%) 19/20 (95%)

Adjusted rate 80.0% 100.0%

Interim evaluation 5/7 (71%) 16/17 (94%)

First incidence (days) 414 400

Life table test P=0.608

Logistic regression test P=0.203

Fisher exact test P=0.251

All Organs: Benign or Malignant Neoplasms

Overall rate 10/10 (100%) 20/20 (100%)

Adjusted rate 100.0% 100.0%

Interim evaluation 7/7 (100%) 17/17 (100%)

First incidence (days) 414 400

Life table test P=0.332N

Logistic regression test )e

Fisher exact test P=1.000

(I) Interim evaluation

a Number of lesion-bearing animals/number of animals examined. Denominator is number of animals examined microscopically for kidney, liver, pituitary gland, testes, and

urinary bladder; for other tissues, denominator is number of animals necropsied. b Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality c Beneath the exposed group incidence are the P values corresponding to pairwise comparisons between both exposed groups. The life table analysis regards neoplasms in

animals dying prior to terminal kill as being (directly or indirectly) the cause of death. The logistic regression test regards these lesions as nonfatal. The Fisher exact test

compares directly the overall incidence rates. For all tests, a lower incidence in an exposure group is indicated by N. d Not applicable; no neoplasms in animal group e Value of the statistic cannot be computed.
300 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE E3

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

Disposition Summary

Animals initially in study 70 10 30

9-Month interim evaluation 10 10

Early deaths

Moribund 3 2

Natural death 1

Survivors 7 17

Animals examined microscopically 10 10 20

9-Month Interim Evaluationb

Alimentary System

Intestine large, colon (10) (10)

Parasite metazoan 2 (20%)

Intestine large, rectum (10) (10)

Intestine large, cecum (9)

Parasite metazoan (10) 1 (11%)

Liver (10) (10)

Basophilic focus 6 (60%)

Clear cell focus 4 (40%)

Cytomegaly 2 (20%)

Fatty change 3 (30%)

Inflammation, chronic active 1 (10%)

Mixed cell focus 1 (10%)

Necrosis, coagulative 1 (10%) 2 (20%)

Pigmentation 10 (100%)

Bile duct, hyperplasia 1 (10%) 7 (70%)

Periportal, inflammation, chronic active 9 (90%) 10 (100%)

Pancreas (10) (10)

Atrophy 2 (20%)

Ectopic tissue 1 (10%)

Infiltration cellular, mononuclear cell 3 (30%) 1 (10%)

Infiltration cellular, mixed cell 1 (10%)

Inflammation, chronic 1 (10%)

Inflammation, chronic active 1 (10%)

Necrosis, coagulative 2 (20%)

Pigmentation 1 (10%) 1 (10%)

Acinus, atrophy 1 (10%)

Salivary glands (10) (10)

Sublingual gland, atrophy 1 (10%)

Sublingual gland, pigmentation 1 (10%)

Cardiovascular System

Heart (10) (10)

Cardiomyopathy 9 (90%) 6 (60%)
Lesions in Male Stop-Exposure Rats 301

TABLE E3

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

9-Month Interim Evaluation (continued)

Endocrine System

Adrenal cortex (10) (10)

Angiectasis 1 (10%)

Adrenal medulla (10) (10)

Hyperplasia 1 (10%)

Pituitary gland (9) (10)

Pars distalis, cyst 1 (11%)

Pars distalis, hyperplasia 3 (33%) 4 (40%)

Thyroid gland (10) (10)

Genital System

Preputial gland (9) (10)

Inflammation, chronic 2 (22%) 2 (20%)

Inflammation, chronic active 7 (78%) 8 (80%)

Prostate (10) (10)

Inflammation, acute 1 (10%) 6 (60%)

Inflammation, chronic active 1 (10%)

Testes(10) (10)

Atrophy 1 (10%)

Infarct 1 (10%)

Inflammation, chronic active 1 (10%)

Interstitial cell, hyperplasia 1 (10%) 2 (20%)

Hematopoietic System

Lymph node (2) (3)

Mediastinal, hemorrhage 1 (50%) 1 (33%)

Mediastinal, pigmentation 1 (50%) 1 (33%)

Pancreatic, pigmentation 1 (33%)

Renal, hemorrhage 1 (50%)

Renal, pigmentation 1 (50%)

Lymph node, mandibular (10) (10)

Hemorrhage 9 (90%) 1 (10%)

Infiltration cellular, histiocyte 1 (10%)

Lymph node, mesenteric (10) (10)

Hemorrhage 1 (10%)

Infiltration cellular, histiocyte 10 (100%) 10 (100%)

Pigmentation 8 (80%)

Spleen(10) (10)

Thymus (9) (10)

Depletion lymphoid 3 (33%) 8 (80%)

Hemorrhage 1 (10%)

Integumentary System

Mammary gland (8) (5)

Hyperplasia 7 (88%) 5 (100%)
302 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE E3

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

9-Month Interim Evaluation (continued)

Respiratory System

Lung (10) (10)

Infiltration cellular, histiocyte 1 (10%)

Artery, mineralization 6 (60%) 4 (40%)

Nose (10) (10)

Glands, inflammation, acute 1 (10%)

Nasolacrimal duct, inflammation, chronic active 1 (10%)

Special Senses System

Ear (1)

Inflammation, chronic active 1 (100%)

Ulcer 1 (100%)

Urinary System

Kidney (10) (10)

Granuloma 1 (10%)

Infiltration cellular, mononuclear cell 7 (70%)

Infiltration cellular, mixed cell 1 (10%)

Inflammation, chronic 4 (40%)

Renal tubule, degeneration, hyaline 10 (100%)

Renal tubule, pigmentation 10 (100%)

Renal tubule, regeneration 10 (100%) 10 (100%)

Urinary bladder (10) (9)

Calculus microscopic observation only 2 (20%) 2 (22%)

Serosa, mineralization 1 (10%)

Systems Examined With No Lesions Observed

General Body System

Musculoskeletal System

Nervous System

15-Month Evaluationc

Alimentary System

Intestine large, colon (10) (10) (20)

Parasite metazoan 4 (40%) 1 (5%)

Intestine large, rectum (9) (10) (20)

Parasite metazoan 1 (11%) 1 (10%) 

Intestine large, cecum (10) (10) (19)

Inflammation, chronic active 1 (5%)

Parasite metazoan 3 (30%) 3 (30%)

Intestine small, ileum (19)

Inflammation, chronic active 1 (5%)
Lesions in Male Stop-Exposure Rats 303

TABLE E3

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

15-Month Evaluation (continued)

Alimentary System (continued)

Liver (10) (10) (20)

Anisokaryosis 1 (10%)

Basophilic focus 1 (10%) 4 (40%) 13 (65%) 

Clear cell focus 6 (60%) 13 (65%)

Degeneration 2 (20%) 4 (20%)

Eosinophilic focus 1 (10%) 2 (10%)

Fatty change 6 (60%) 9 (90%) 10 (50%) 

Hemmorrhage 1 (5%)

Hematopoietic cell proliferation 2 (20%)

Hepatodiaphragmatic nodule 1 (10%) 

Inflammation, chronic, granulomatous 2 (20%)

Inflammation, chronic active 6 (60%) 2 (10%)

Necrosis, coagulative 6 (60%) 1 (10%) 2 (10%)

Pigmentation 8 (80%) 18 (90%)

Bile duct, hyperplasia 10 (100%) 7 (70%) 19 (95%)

Periportal, inflammation, chronic 10 (100%) 7 (70%) 17 (85%)

Periportal, inflammation, chronic active 1 (5%)

Mesentery (1) (1)

Hemorrhage 1 (100%)

Inflammation, chronic 1 (100%)

Inflammation, chronic active 1 (100%)

Necrosis, coagulative 1 (100%)

Thrombosis 1 (100%)

Pancreas (10) (10) (20)

Atrophy 4 (40%) 2 (20%) 4 (20%)

Hyperplasia 1 (5%)

Inflammation, chronic 8 (80%) 5 (50%) 11 (55%)

Inflammation, chronic active 1 (5%)

Vacuolization cytoplasmic 1 (10%) 

Artery, inflammation, chronic active 1 (5%)

Salivary glands (19)

Duct, parotid gland, mineralization 1 (5%)

Stomach, forestomach (10) (10) (20)

Acanthosis 2 (10%)

Hyperkeratosis 2 (20%) 1 (5%)

Hyperplasia, basal cell 2 (20%) 3 (15%)

Hyperplasia, squamous 2 (20%)

Inflammation, chronic active 1 (10%) 1 (5%)

Ulcer 2 (20%)

Muscularis, mineralization 1 (10%)

Stomach, glandular (10) (10) (20)

Inflammation, chronic 1 (10%) 1 (10%) 1 (5%)

Inflammation, chronic active 1 (10%)

Arteriole, mineralization 1 (5%)

Muscularis, mineralization 1 (10%) 1 (5%)
304 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE E3

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

15-Month Evaluation (continued)

Cardiovascular System

Heart (10) (10) (20)

Cardiomyopathy 10 (100%) 8 (80%) 17 (85%)

Endocrine System

Adrenal cortex (10) (10) (20)

Hyperplasia 1 (10%) 1 (10%)

Bilateral, vacuolization cytoplasmic 1 (10%) 6 (30%)

Adrenal medulla (9) (10) (20)

Bilateral, hyperplasia 1 (11%) 1 (5%)

Islets, pancreatic (4) (3)

Hemorrhage 1 (25%)

Pituitary gland (8) (10) (19)

Pars distalis, cyst 1 (10%) 3 (16%)

Pars distalis, hyperplasia 8 (100%) 6 (60%) 16 (84%)

Pars nervosa, cyst 1 (5%)

Pars nervosa, hyperplasia 1 (5%)

Thyroid gland (10) (10) (20)

Ultimobranchial cyst 1 (10%) 1 (5%)

C-cell, hyperplasia 1 (10%) 1 (10%) 1 (5%)

General Body System

None

Genital System

Epididymis (10) (10) (20)

Inflammation, chronic 1 (5%)

Vacuolization cytoplasmic 1 (10%)

Preputial gland (9) (10) (19)

Inflammation, chronic 5 (56%) 2 (20%) 6 (32%)

Inflammation, chronic active 4 (44%) 7 (70%) 9 (47%)

Prostate (10) (10) (20)

Inflammation, acute 1 (10%) 4 (40%) 5 (25%)

Inflammation, chronic 1 (5%)

Inflammation, chronic active 6 (60%) 4 (40%) 6 (30%)

Epithelium, hyperplasia 1 (5%)

Testes (10) (10) (20)

Abscess 1 (10%)

Abscess, chronic 1 (5%) 

Inflammation, chronic 2 (10%)

Interstitial cell, hyperplasia 10 (100%) 8 (80%) 17 (85%)

Seminiferous tubule, atrophy 2 (20%) 3 (30%) 7 (35%)

Seminiferous tubule, mineralization 2 (20%)
Lesions in Male Stop-Exposure Rats 305

TABLE E3

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

15-Month Evaluation (continued)

Hematopoietic System

Bone marrow (10) (10) (20)

Myelofibrosis 1 (10%)

Lymph node (2) (3)

Mediastinal, hemorrhage 1 (50%) 2 (67%)

Pancreatic, hemorrhage 1 (50%) 

Pancreatic, infiltration cellular, histiocyte 1 (50%) 2 (67%)

Pancreatic, pigmentation 2 (67%)

Lymph node, mandibular (10) (8) (17)

Hemorrhage 3 (30%) 1 (13%) 3 (18%)

Hyperplasia, plasma cell 1 (6%)

Infiltration cellular, histiocyte 1 (6%)

Pigmentation 1 (6%)

Lymph node, mesenteric (10) (9) (20)

Depletion lymphoid 1 (10%)

Hemorrhage 1 (10%) 1 (5%)

Infiltration cellular, histiocyte 10 (100%) 9 (100%) 20 (100%)

Pigmentation 9 (90%) 7 (78%) 15 (75%)

Spleen (20)

Depletion lymphoid 1 (5%)

Thymus (10) (9) (16)

Cyst 1 (10%)

Depletion lymphoid 4 (44%) 12 (75%)

Integumentary System

Mammary gland (4) (7) (10)

Hyperplasia 2 (50%) 7 (100%) 9 (90%)

Skin (19)

Inflammation, chronic 1 (5%)

Musculoskeletal System

None

Nervous System

None

Respiratory System

Lung (10) (10) (20)

Infiltration cellular, histiocyte 2 (20%) 2 (20%) 7 (35%)

Alveolar epithelium, hyperplasia 3 (30%) 1 (5%)

Alveolus, mineralization 1 (10%) 1 (5%)

Artery, mineralization 8 (80%) 7 (70%) 9 (45%)
306 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE E3

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

15-Month Evaluation (continued)

Respiratory System (continued)

Nose (10) (10) (20)

Glands, inflammation, acute 3 (30%) 3 (30%) 7 (35%)

Glands, inflammation, chronic active 1 (10%) 3 (15%)

Lumen, inflammation, acute 1 (10%)

Nasolacrimal duct, inflammation, chronic active 1 (10%)

Respiratory epithelium, metaplasia, squamous 1 (5%)

Special Senses System

Eye (1)

Conjunctiva, inflammation, chronic active 1 (100%)

Cornea, fibrosis 1 (100%)

Cornea, neovascularization 1 (100%)

Urinary System

Kidney (10) (10) (20)

Autolysis 1 (5%)

Cyst 1 (10%)

Nephropathy 10 (100%) 10 (100%) 20 (100%)

Renal tubule, hyperplasia 1 (10%) 1 (5%)

Renal tubule, inflammation, chronic active 1 (5%)

Renal tubule, pigmentation 9 (90%) 20 (100%)

Transitional epithelium, hyperplasia 1 (10%) 11 (55%)

Urinary bladder (10) (10) (19)

Inflammation, chronic 1 (5%)

Fat, proliferation 1 (5%)

Muscularis, mineralization 1 (5%)

Transitional epithelium, hyperplasia 9 (47%)

a Number of animals examined microscopically at site and number of animals with lesion (includes interim and moribund animals)

b Controls from the 9-month interim evaluation of the 2-year core study were used for comparison.

c Controls from the 15-month interim evaluation of the 2-year core study were used for comparison. 
307

APPENDIX F

SUMMARY OF LESIONS IN FEMALE RATS

IN THE STOP-EXPOSURE EVALUATION

OF 1-AMINO-2,4-DIBROMOANTHRAQUINONE

TABLE F1 Summary of the Incidence of Neoplasms in Female Rats

in the Stop-Exposure Evaluation of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . 308

TABLE F2a Statistical Analysis of Primary Neoplasms in Female Rats 

in the Stop-Exposure Evaluation of 1-Amino-2,4-dibromoanthraquinone: 

15-Month Interim Evaluation Control Group versus

9-Month and 15-Month 20,000 ppm Groups at the 15-Month Evaluation . . . . . . . . . . . . . . . . . . . . . . . . . . . 311

TABLE F2b Statistical Analysis of Primary Neoplasms in Female Rats 

in the Stop-Exposure Evaluation of 1-Amino-2,4-dibromoanthraquinone: 

9-Month 20,000 ppm Stop-Exposure Group versus

15-Month 20,000 ppm Exposure Group at the 15-Month Evaluation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 314

TABLE F3 Summary of the Incidence of Nonneoplastic Lesions in Female Rats

in the Stop-Exposure Evaluation of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . 317
308 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE F1

Summary of the Incidence of Neoplasms in Female Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

Disposition Summary

Animals initially in study 70 10 30

9-Month interim evaluation 10 10

Early deaths

Moribund 1 5

Natural deaths 3

Survivors 9 11

Missexed 1

Animals examined microscopically 10 10 18b

9-Month Interim Evaluationc

Alimentary System

Liver (10) (10)

Hepatocellular carcinoma 1 (10%)

Hepatocellular adenoma 2 (20%)

Endocrine System

Pituitary gland (10) (10)

Pars distalis, adenoma 1 (10%)

Urinary System

Kidney (10) (10)

Adenoma 1 (10%)

Systems Examined With No Neoplasms Observed

Cardiovascular System

General Body System

Genital System

Hematopoietic System

Integumentary System

Musculoskeletal System

Nervous System

Respiratory System

Special Senses System

15-Month Evaluationd

Alimentary System

Intestine large, rectum (10) (10) (17)

Polyp adenomatous 5 (50%) 2 (12%)

Polyp adenomatous, multiple 1 (6%)
Lesions in Female Stop-Exposure Rats 309

TABLE F1

Summary of the Incidence of Neoplasms in Female Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

15-Month Evaluation (continued)

Alimentary System (continued)

Liver (10) (10) (18)

Hepatocellular carcinoma 3 (30%) 7 (39%)

Hepatocellular carcinoma, multiple 3 (30%) 8 (44%)

Hepatocellular adenoma 5 (50%) 3 (17%)

Hepatocellular adenoma, multiple 1 (10%) 7 (39%)

Cardiovascular System

None

Endocrine System

Pituitary gland (10) (10) (18)

Pars distalis, adenoma 2 (20%) 2 (20%) 1 (6%)

Thyroid gland (10) (10) (18)

Adenoma 1 (10%)

General Body System

None 

Genital System

Uterus (10) (10) (18)

Polyp stromal 1 (10%) 1 (10%) 1 (6%)

Hematopoietic System

None 

Integumentary System

Mammary gland (7) (7)

Fibroadenoma 1 (14%)

Musculoskeletal System

None 

Nervous System

None

Respiratory System

Lung (10) (10) (18)

Hepatocellular carcinoma, metastatic, liver 1 (10%) 1 (6%)
310 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE F1

Summary of the Incidence of Neoplasms in Female Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

15-Month Evaluation (continued)

Special Senses System

None 

Urinary System

Kidney (10) (10) (18)

Renal tubule, adenoma 3 (30%) 1 (6%)

Renal tubule, adenoma, multiple 1 (6%)

Urinary bladder (10) (10) (18)

Squamous cell, carcinoma 4 (22%)

Squamous cell, papilloma 1 (6%)

Transitional epithelium, carcinoma 1 (6%)

Transitional epithelium, papilloma 1 (6%)

Neoplasm Summary

Total animals with primary neoplasmse 2 9 17

Total primary neoplasms 4 25 39

Total animals with benign neoplasms 2 9 13

Total benign neoplasms 4 18 19

Total animals with malignant neoplasms 6 16

Total malignant neoplasms 7 20

Total animals with metastatic neoplasms 1 1

Total metastatic neoplasms 1 1

a Number of animals examined microscopically at the site and the number of animals with neoplasm (includes interim and moribund animals) b One animal not examined microscopically c Controls from the 9-month interim evaluation of the 2-year core study were used for comparison. d Controls from the 15-month interim evaluation of the 2-year core study were used for comparison. e Primary neoplasms: all neoplasms except metastatic neoplasms
Lesions in Female Stop-Exposure Rats 311

TABLE F2a

Statistical Analysis of Primary Neoplasms in Female Rats in the Stop-Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone: 15-Month Interim Evaluation Control Group versus 9-Month 

and 15-Month 20,000 ppm Groups at the 15-Month Evaluation

0 ppm 20,000 ppm 20,000 ppm

(15-month (9-month stop-exposure) (15-month exposure)

interim evaluation)

Kidney (Renal Tubule): Adenoma

Overall ratea 0/10 (0%) 3/10 (30%) 2/18 (11%)

Adjusted rateb 0.0% 40.7% 24.2%

Interim evaluation 0/10 (0%) 3/9 (33%) 2/11 (18%)

First incidence (days) )d 462 (I) 462 (I)

Life table testc P=0.189 P=0.093 P=0.256 

Logistic regression testc P=0.189 P=0.093 P=0.256

Cochran-Armitage testc P=0.345

Fisher exact testc P=0.105 P=0.405 

Large Intestine (Rectum): Adenomatous Polyp

Overall rate 0/10 (0%) 5/10 (50%) 3/18 (17%)

Adjusted rate 0.0% 61.1% 20.5%

Interim evaluation 0/10 (0%) 5/9 (56%) 1/11 (9%)

First incidence (days) ) 462 (I) 306

Life table test P=0.136 P=0.015 P=0.186

Logistic regression test P=0.320 P=0.015 P=0.367

Cochran-Armitage test P=0.266

Fisher exact test P=0.016 P=0.249

Liver: Hepatocellular Adenoma

Overall rate 0/10 (0%) 6/10 (60%) 10/18 (56%)

Adjusted rate 0.0% 65.0% 100.0%

Interim evaluation 0/10 (0%) 5/9 (56%) 9/11 (82%)

First incidence (days) ) 456 306

Life table test P<0.001 P=0.009 P<0.001

Logistic regression test P=0.002 P=0.015 P=0.003

Cochran-Armitage test P=0.005

Fisher exact test P=0.005 P=0.003

Liver: Hepatocellular Carcinoma

Overall rate 0/10 (0%) 6/10 (60%) 15/18 (83%)

Adjusted rate 0.0% 66.7% 100.0%

Interim evaluation 0/10 (0%) 6/9 (67%) 11/11 (100%)

First incidence (days) ) 462 (I) 426

Life table test P<0.001 P=0.005 P<0.001

Logistic regression test P<0.001 P=0.005 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P=0.005 P<0.001

Liver: Hepatocellular Adenoma or Carcinoma

Overall rate 0/10 (0%) 8/10 (80%) 16/18 (89%)

Adjusted rate 0.0% 80.0% 100.0%

Interim evaluation 0/10 (0%) 7/9 (78%) 11/11 (100%)

First incidence (days) ) 456 306

Life table test P<0.001 P<0.001 P<0.001

Logistic regression test P<0.001 P=0.002 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P<0.001 P<0.001
312 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE F2a

Statistical Analysis of Primary Neoplasms in Female Rats in the Stop-Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone: 15-Month Interim Evaluation Control Group versus 9-Month 

and 15-Month 20,000 ppm Groups at the 15-Month Evaluation (continued)

0 ppm 20,000 ppm 20,000 ppm

(15-month (9-month stop-exposure) (15-month exposure)

interim evaluation)

Pituitary Gland (Pars Distalis): Adenoma

Overall rate 2/10 (20%) 2/10 (20%) 1/18 (6%)

Adjusted rate 20.0% 25.0% 8.3%

Interim evaluation 2/10 (20%) 1/9 (11%) 0/11 (0%)

First incidence (days) 462 (I) 456 461

Life table test P=0.387N P=0.670 P=0.456N 

Logistic regression test P=0.319N P=0.539N P=0.427N

Cochran-Armitage test P=0.222N

Fisher exact test P=0.709N P=0.284N

Urinary Bladder: Papilloma

Overall rate 0/10 (0%) 0/10 (0%) 2/18 (11%)

Adjusted rate 0.0% 0.0% 22.2%

Interim evaluation 0/10 (0%) 0/9 (0%) 1/11 (9%)

First incidence (days) ) ) 426

Life table test P=0.174 P=0.295

Logistic regression test P=0.215 P=0.357

Cochran-Armitage test P=0.220

Fisher exact test P=0.405

Urinary Bladder: Carcinoma

Overall rate 0/10 (0%) 1/10 (10%) 5/18 (28%)

Adjusted rate 0.0% 11.1% 48.7%

Interim evaluation 0/10 (0%) 1/9 (11%) 4/11 (36%)

First incidence (days) ) 462 (I) 299

Life table test P=0.022 P=0.479 P=0.044

Logistic regression test P=0.061 P=0.479 P=0.105

Cochran-Armitage test P=0.051

Fisher exact test P=0.500 P=0.087

All Organs: Benign Neoplasms

Overall rate 2/10 (20%) 9/10 (90%) 13/18 (72%)

Adjusted rate 20.0% 90.0% 100.0%

Interim evaluation 2/10 (20%) 8/9 (89%) 10/11 (91%)

First incidence (days) 462 (I) 456 306

Life table test P<0.001 P=0.004 P<0.001

Logistic regression test P=0.001 P=0.007 P=0.004

Cochran-Armitage test P=0.008

Fisher exact test P=0.003 P=0.011

All Organs: Malignant Neoplasms

Overall rate 0/10 (0%) 6/10 (60%) 16/18 (89%)

Adjusted rate 0.0% 66.7% 100.0%

Interim evaluation 0/10 (0%) 6/9 (67%) 11/11 (100%)

First incidence (days) ) 462 (I) 299

Life table test P<0.001 P=0.005 P<0.001

Logistic regression test P<0.001 P=0.005 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P=0.005 P<0.001
Lesions in Female Stop-Exposure Rats 313

TABLE F2a

Statistical Analysis of Primary Neoplasms in Female Rats in the Stop-Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone: 15-Month Interim Evaluation Control Group versus 9-Month 

and 15-Month 20,000 ppm Groups at the 15-Month Evaluation (continued)

0 ppm 20,000 ppm 20,000 ppm

(15-month (9-month stop-exposure) (15-month exposure)

interim evaluation)

All Organs: Benign or Malignant Neoplasms

Overall rate 2/10 (20%) 9/10 (90%) 17/18 (94%)

Adjusted rate 20.0% 90.0% 100.0%

Interim evaluation 2/10 (20%) 8/9 (89%) 11/11 (100%)

First incidence (days) 462 456 299

Life table test P<0.001 P=0.004 P<0.001

Logistic regression test P<0.001 P=0.007 P<0.001

Cochran-Armitage test P<0.001

Fisher exact test P=0.003 P<0.001

(I) Interim evaluation

a Number of lesion-bearing animals/number of animals examined (includes interim and moribund animals). Denominator is number of animals examined microscopically for

kidney, liver, pituitary gland, and urinary bladder; for other tissues, denominator is number of animals necropsied. b Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality c Beneath the exposed group incidence are the P values corresponding to pairwise comparisons between the controls and that exposed group. The life table analysis regards

neoplasms in animals dying prior to terminal kill as being (directly or indirectly) the cause of death. The logistic regression test regards these lesions as nonfatal. The Fisher

exact test compares directly the overall incidence rates. For all tests, a lower incidence in an exposure group is indicated by N. d Not applicable; no neoplasms in animal group
314 1-Amino-2,4-Dibromoanthraquinone, NTP TR 383

TABLE F2b

Statistical Analysis of Primary Neoplasms in Female Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone: 9-Month 20,000 ppm Stop-Exposure Group

versus 15-Month 20,000 ppm Exposure Group at the 15-Month Evaluation

20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

Kidney (Renal Tubule): Adenoma

Overall ratesa 3/10 (30%) 2/18 (11%)

Adjusted ratesb 40.7% 24.2%

Interim evaluation 3/9 (33%) 2/11 (18%)

First incidence (days) 462 (I) 462 (I)

Life table testsc P=0.400N

Logistic regression testsc P=0.400N

Fisher exact testc P=0.228N

Large Intestine (Rectum): Adenomatous Polyp

Overall rates 5/10 (50%) 3/18 (17%)

Adjusted rates 61.1% 20.5%

Interim evaluation 5/9 (56%) 1/11 (9%)

First incidence (days) 462 (I) 306

Life table tests P=0.209N

Logistic regression tests P=0.057N

Fisher exact test P=0.077N

Liver: Hepatocellular Adenoma

Overall rates 6/10 (60%) 10/18 (56%)

Adjusted rates 65.0% 100.0%

Interim evaluation 5/9 (56%) 9/11 (82%)

First incidence (days) 456 306

Life table tests P=0.305

Logistic regression tests P=0.595

Fisher exact test P=0.570N

Liver: Hepatocellular Carcinoma

Overall rates 6/10 (60%) 15/18 (83%)

Adjusted rates 66.7% 100.0%

Interim evaluation 6/9 (67%) 11/11 (100%)

First incidence (days) 462 (I) 426

Life table tests P=0.014

Logistic regression tests P=0.010

Fisher exact test P=0.181

Liver: Hepatocellular Adenoma or Carcinoma

Overall rates 8/10 (80%) 16/18 (89%)

Adjusted rates 80.0% 100.0%

Interim evaluation 7/9 (78%) 11/11 (100%)

First incidence (days) 456 306

Life table tests P=0.063

Logistic regression tests P=0.126

Fisher exact test P=0.452
Lesions in Female Stop-Exposure Rats 315

TABLE F2b

Statistical Analysis of Primary Neoplasms in Female Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone: 9-Month 20,000 ppm Stop-Exposure Group

versus 15-Month 20,000 ppm Exposure Group at the 15-Month Evaluation (continued)

20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

Pituitary Gland (Pars Distalis): Adenoma

Overall rates 2/10 (20%) 1/18 (6%)

Adjusted rates 25.0% 8.3%

Interim evaluation 1/9 (11%) 0/11 (0%)

First incidence (days) 456 461

Life table tests P=0.423N

Logistic regression tests P=0.385N

Fisher exact test P=0.284N

Urinary Bladder: Papilloma

Overall rates 0/10 (0%) 2/18 (11%)

Adjusted rates 0.0% 22.2%

Interim evaluation 0/9 (0%) 1/11 (9%)

First incidence (days) )d 426

Life table tests P=0.308

Logistic regression tests P=0.357

Fisher exact test P=0.405

Urinary Bladder: Carcinoma

Overall rates 1/10 (10%) 5/18 (28%)

Adjusted rates 11.1% 48.7%

Interim evaluation 1/9 (11%) 4/11 (36%)

First incidence (days) 462 (I) 299

Life table tests P=0.154

Logistic regression tests P=0.292

Fisher exact test P=0.277

All Organs: Benign Neoplasms

Overall rates 9/10 (90%) 13/18 (72%)

Adjusted rates 90.0% 100.0%

Interim evaluation 8/9 (89%) 10/11 (91%)

First incidence (days) 456 306

Life table tests P=0.412

Logistic regression tests P=0.539N

Fisher exact test P=0.277N

All Organs: Malignant Neoplasms

Overall rates 6/10 (60%) 16/18 (89%)

Adjusted rates 66.7% 100.0%

Interim evaluation 6/9 (67%) 11/11 (100%)

First incidence (days) 462 (I) 299

Life table tests P=0.010

Logistic regression tests P=0.015

Fisher exact test P=0.098
316 1-Amino-2,4-Dibromoanthraquinone, NTP TR 383

TABLE F2b

Statistical Analysis of Primary Neoplasms in Female Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone: 9-Month 20,000 ppm Stop-Exposure Group

versus 15-Month 20,000 ppm Exposure Group at the 15-Month Evaluation (continued)

20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

All Organs: Benign or Malignant Neoplasms

Overall rates 9/10 (90%) 17/18 (94%)

Adjusted rates 90.0% 100.0%

Interim evaluation 8/9 (89%) 11/11 (100%)

First incidence (days) 456 299

Life table tests P=0.082

Logistic regression tests P=0.396

Fisher exact test P=0.595

(I) Interim evaluation

a Number of lesion-bearing animals/number of animals examined (includes interim and moribund animals). Denominator is number of animals examined microscopically for

kidney, liver, pituitary gland, and urinary bladder; for other tissues, denominator is number of animals necropsied. b Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality c Beneath the exposed group incidence are the P values corresponding to pairwise comparisons between both exposed groups. The life table analysis regards neoplasms in

animals dying prior to terminal kill as being (directly or indirectly) the cause of death. The logistic regression test regards these lesions as nonfatal. The Fisher exact test

compares directly the overall incidence rates. For all tests, a lower incidence in an exposure group is indicated by N. d Not applicable; no neoplasms in animal group
Lesions in Female Stop-Exposure Rats 317

TABLE F3

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

Disposition Summary

Animals initially in study 70 10 30

9-Month interim evaluation 10 10

Early deaths

Moribund 1 5

Natural deaths 3

Survivors 9 11

Missexed 1

Animals examined microscopically 10 10 18b

9-Month Interim Evaluationc

Alimentary System

Intestine large, colon (10) (10)

Parasite metazoan 1 (10%)

Intestine large, rectum (10) (10)

Parasite metazoan 2 (20%)

Intestine large, cecum (10) (10)

Liver (10) (10)

Angiectasis 2 (20%)

Basophilic focus 1 (10%) 3 (30%)

Clear cell focus 1 (10%)

Fatty change 7 (70%)

Inflammation, chronic active 2 (20%)

Necrosis, coagulative 8 (80%)

Pigmentation 10 (100%)

Bile duct, hyperplasia 1 (10%) 9 (90%)

Periportal, inflammation, chronic active 6 (60%) 10 (100%)

Pancreas (10) (10)

Atrophy 1 (10%) 2 (20%)

Infiltration cellular, mononuclear cell 3 (30%) 4 (40%)

Salivary glands (10) (10)

Stomach, forestomach (10) (10)

Muscularis, mineralization 1 (10%)

Stomach, glandular (10) (10)

Muscularis, mineralization 1 (10%) 1 (10%)

Cardiovascular System

Heart (10) (10)

Cardiomyopathy 7 (70%) 5 (50%)

Endocrine System

Adrenal cortex (10) (10)

Capsule, fibrosis 1 (10%)

Capsule, inflammation, chronic 1 (10%)
318 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE F3

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

9-Month Interim Evaluation (continued)

Endocrine System (continued)

Pituitary gland (10) (10)

Pars distalis, angiectasis 1 (10%)

Pars distalis, cyst 2 (20%) 3 (30%)

Pars distalis, hemorrhage 1 (10%)

Pars distalis, hyperplasia 1 (10%)

Pars intermedia, cyst 1 (10%) 1 (10%)

Thyroid gland (10) (10)

Infiltration cellular, mononuclear cell 1 (10%)

Genital System

Clitoral gland (10) (9)

Infiltration cellular, mononuclear cell 1 (10%)

Inflammation, chronic 3 (30%) 1 (11%)

Inflammation, chronic active 2 (20%) 2 (22%)

Ovary (10) (10)

Periovarian tissue 1 (10%)

Uterus (10) (10)

Hydrometra 1 (10%) 1 (10%)

Hematopoietic System

Lymph node (3) (5)

Mediastinal, hemorrhage 3 (100%)

Mediastinal, pigmentation 3 (60%)

Pancreatic, infiltration cellular, histiocyte 4 (80%)

Pancreatic, pigmentation 5 (100%)

Lymph node, mandibular (9) (10)

Hemorrhage 7 (78%) 2 (20%)

Hyperplasia 1 (10%)

Infiltration cellular, histiocyte 1 (10%)

Lymph node, mesenteric (10) (10)

Hemorrhage 5 (50%)

Infiltration cellular, histiocyte 10 (100%) 10 (100%)

Pigmentation 1 (10%)

Spleen (10) (10)

Congestion 1 (10%)

Thymus (10) (9)

Congestion 1 (10%)

Hemorrhage 1 (10%) 2 (22%)

Respiratory System

Lung (10) (10)

Infiltration cellular, histiocyte 1 (10%) 5 (50%)

Artery, mineralization 4 (40%) 2 (20%)
Lesions in Female Stop-Exposure Rats 319

TABLE F3

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the Stop-Exposure Evaluaton 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

9-Month Interim Evaluation (continued)

Special Senses System

Eye (1)

Anterior chamber, inflammation, acute 1 (100%)

Posterior chamber, inflammation, acute 1 (100%)

Retina, degeneration 1 (100%)

Harderian gland (1)

Hyperplasia 1 (100%)

Urinary System

Kidney (10) (10)

Fibrosis, focal 1 (10%)

Hydronephrosis 1 (10%)

Infiltration cellular, mononuclear cell 7 (70%)

Inflammation, chronic 4 (40%)

Mineralization 3 (30%) 1 (10%)

Mineralization, focal 1 (10%)

Renal tubule, pigmentation 10 (100%)

Renal tubule, regeneration 4 (40%) 7 (70%)

Transitional epithelium, mineralization 1 (10%)

Urinary bladder (10) (10)

Transitional epithelium, hyperplasia 4 (40%)

Systems Examined With No Lesions Observed

General Body System

Integumentary System

Musculoskeletal System

Nervous System

15-Month Evaluationd

Alimentary System

Intestine large, colon (10) (10) (18)

Parasite metazoan 2 (20%) 1 (10%)

Intestine large, rectum (10) (10) (17)

Parasite metazoan 1 (10%)

Intestine large, cecum (10) (10) (18)

Parasite metazoan 2 (20%) 2 (20%)
320 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE F3

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

15-Month Evaluation (continued)

Alimentary System (continued)

Liver (10) (10) (18)

Basophilic focus 8 (80%) 6 (60%) 13 (72%)

Clear cell focus 5 (50%) 13 (72%)

Cyst 1 (10%)

Cytomegaly 1 (10%) 2 (11%)

Cytoplasmic alteration 1 (10%)

Eosinophilic focus 2 (20%) 2 (20%) 3 (17%)

Fatty change 10 (56%)

Hematopoietic cell proliferation 1 (6%)

Hepatodiaphragmatic nodule 2 (20%)

Inflammation, chronic, granulomatous 6 (60%) 2 (20%) 1 (6%)

Inflammation, chronic active 1 (10%) 1 (6%)

Mixed cell focus 2 (20%) 3 (17%)

Necrosis, coagulative 1 (10%) 4 (22%)

Pigmentation 1 (10%) 10 (100%) 17 (94%)

Bile duct, hyperplasia 7 (70%) 10 (100%) 18 (100%)

Periportal, inflammation, chronic 10 (100%) 9 (90%) 11 (61%)

Periportal, inflammation, chronic active 1 (10%) 7 (39%)

Mesentery (1)

Inflammation, chronic, granulomatous 1 (100%)

Necrosis, coagulative 1 (100%)

Pancreas (10) (10) (18)

Atrophy 1 (10%) 3 (30%) 3 (17%)

Ectopic tissue 1 (10%) 1 (6%)

Inflammation, chronic 9 (90%) 6 (60%) 6 (33%)

Inflammation, chronic, active 2 (11%)

Salivary glands (10) (10) (18)

Duct, submandibular gland, dilatation 1 (6%)

Parotid gland, atrophy 1 (6%)

Parotid gland, inflammation, chronic 2 (11%)

Parotid gland, inflammation, chronic active 2 (11%)

Submandibular gland, atrophy 1 (6%)

Submandibular gland, inflammation, chronic 1 (6%)

Submandibular gland, inflammation, chronic

active 5 (28%)

Submandibular gland, metaplasia, squamous 2 (11%)

Submandibular gland, pigmentation 1 (6%)

Stomach, forestomach (10) (10) (18)

Acanthosis 1 (6%)

Erosion 1 (6%)

Hyperkeratosis 1 (6%)

Hyperplasia, basal cell 6 (33%)

Hyperplasia, squamous 2 (11%)

Ulcer 1 (6%)

Stomach, glandular (10) (10) (18)

Inflammation, chronic 1 (6%)

Mineralization 1 (10%)

Artery, mineralization 1 (6%)

Muscularis, mineralization 1 (10%) 1 (6%)
Lesions in Female Stop-Exposure Rats 321

TABLE F3

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

15-Month Evaluation (continued)

Cardiovascular System

Blood vessel (1)

Aorta, inflammation, chronic 1 (100%)

Heart (10) (10) (18)

Cardiomyopathy 10 (100%) 10 (100%) 13 (72%)

Endocrine System

Adrenal cortex (10) (10) (17)

Angiectasis 5 (50%) 2 (20%) 3 (18%)

Hyperplasia 1 (10%)

Vacuolization, cytoplasmic 1 (6%)

Adrenal medulla (10) (10) (17)

Hyperplasia 1 (10%) 2 (12%)

Parathyroid gland (10) (10) (15)

Hyperplasia 1 (10%)

Pituitary gland (10) (10) (18)

Pars distalis, angiectasis 5 (50%)

Pars distalis, cyst 8 (80%) 6 (60%) 8 (44%)

Pars distalis, hyperplasia 1 (10%) 2 (11%)

Pars intermedia, cyst 3 (30%) 1 (10%) 3 (17%)

Thyroid gland (10) (10)

C-cell, hyperplasia 4 (40%)

General Body System

None 

Genital System

Clitoral gland (10) (10) (18)

Cyst 1 (6%)

Inflammation, chronic 7 (70%) 6 (60%) 3 (17%)

Inflammation, chronic active 1 (10%) 1 (10%) 8 (44%)

Ovary (18)

Periovarian tissue, cyst 3 (17%)

Uterus (10) (10) (18)

Hydrometra 2 (20%) 2 (20%) 5 (28%)

Inflammation, chronic active 1 (6%)

Endometrium, hyperplasia 1 (10%) 1 (10%) 1 (6%)

Submucosa, hyperplasia 1 (6%)

Hematopoietic System

Lymph node (1) (1) (4)

Mediastinal, hemorrhage 1 (100%) 1 (100%)

Mediastinal, pigmentation 1 (100%)

Pancreatic, infiltration cellular, histiocyte 1 (100%) 4 (100%)

Pancreatic, pigmentation 1 (100%) 2 (50%)
322 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE F3

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

15-Month Evaluation (continued)

Hematopoietic System (continued)

Lymph node, mandibular (7) (9) (17)

Hemorrhage 1 (14%) 3 (33%) 1 (6%)

Infiltration cellular, hystiocyte 6 (35%)

Lymph node, mesenteric (10) (9) (18)

Depletion lymphoid 1 (6%)

Hemorrhage 2 (22%)

Infiltration cellular, histiocyte 10 (100%) 9 (100%) 18 (100%)

Pigmentation 10 (100%) 8 (89%) 11 (61%)

Spleen (18)

Fibrosis 1 (6%)

Thymus (10) (10) (13)

Cyst 1 (10%)

Depletion lymphoid 1 (10%) 6 (46%)

Hemorrhage 1 (10%)

Integumentary System

Mammary gland (7) (7) (14)

Hyperplasia 6 (86%) 4 (57%) 5 (36%)

Musculoskeletal System

Bone (10) (10)

Osteopetrosis 1 (10%)

Nervous System

None 

Respiratory System

Lung (10) (10) (18)

Congestion 1 (10%)

Hemorrhage 1 (10%)

Infiltration cellular, histiocyte 6 (60%) 7 (70%) 12 (67%)

Inflammation, chronic active 1 (6%)

Alveolus, mineralization 1 (10%)

Artery, mineralization 4 (40%) 5 (50%) 10 (56%)

Nose (10) (10) (18)

Inflammation, chronic active 2 (11%)

Submucosa, inflammation, chronic 1 (10%)

Special Senses System

None 
Lesions in Female Stop-Exposure Rats 323

TABLE F3

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 20,000 ppm 20,000 ppm

(9-month stop-exposure) (15-month exposure)

15-Month Evaluation (continued)

Urinary System

Kidney (10) (10) (18)

Fibrosis 1 (6%)

Inflammation, chronic active 1 (6%)

Nephropathy 10 (100%) 10 (100%) 18 (100%)

Papilla, necrosis, coagulative 2 (11%)

Pelvis, inflammation, chronic active 1 (6%)

Renal tubule, hyperplasia 2 (20%) 2 (11%)

Renal tubule, inflammation, chronic active 1 (10%)

Renal tubule, mineralization 1 (10%)

Renal tubule, pigmentation 10 (100%) 18 (100%)

Transitional epithelium, hyperplasia 3 (30%) 1 (10%) 5 (28%)

Transitional epithelium, mineralization 2 (20%) 1 (10%)

Urinary bladder (10) (10) (18)

Hemorrhage 4 (22%)

Necrosis 1 (6%)

Fat, proliferation 2 (11%)

Transitional epithelium, hyperplasia 4 (40%) 17 (94%)

Transitional epithelium, metaplasia, squamous 3 (17%)

a Number of animals examined microscopically at site and number of animals with lesion (includes interim and moribund animals) b One animal not examined microscopically c Controls from the 9-month interim evaluation of the 2-year core study were used for comparison. d Controls from the 15-month interim evaluation of the 2-year core study were used for comparison.
324 1-Amino-2,4-dibromoanthraquinone, NTP TR 383
325

APPENDIX G

GENETIC TOXICOLOGY

SALMONELLA TYPHIMURIUM MUTAGENICITY TEST PROTOCOL . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 326

CHINESE HAMSTER OVARY CELL CYTOGENETICS PROTOCOLS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 326

RESULTS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 327

TABLE G1 Mutagenicity of 1-Amino-2,4-dibromoanthraquinone in Salmonella typhimurium . . . . . . . 329

TABLE G2 Induction of Sister Chromatid Exchanges in Chinese Hamster Ovary Cells

by 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 330

TABLE G3 Induction of Chromosomal Aberrations in Chinese Hamster Ovary Cells

by 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 332
326 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

GENETIC TOXICOLOGY

SALMONELLA TYPHIMURIUM MUTAGENICITY TEST PROTOCOL

Testing was performed as reported by Haworth et al. (1983). 1-Amino-2,4-dibromoanthraquinone was

sent to the laboratory as a coded aliquot from Radian Corporation (Austin, TX). It was incubated with

the Salmonella typhimurium tester strains TA98, TA100, TA1535, and TA1537 either in buffer or S9 mix

(metabolic activation enzymes and cofactors from Aroclor 1254-induced male Sprague-Dawley rat or

Syrian hamster liver) for 20 minutes at 37° C. Top agar supplemented with l-histidine and d-biotin was

added, and the contents of the tubes were mixed and poured onto the surfaces of minimal glucose agar

plates. Histidine-independent mutant colonies arising on these plates were counted following

incubation for 2 days at 37° C.

Each trial consisted of triplicate plates of concurrent positive and negative controls and of five doses of

1-amino-2,4-dibromoanthraquinone. The high dose was limited to 10,000 :g/plate. All positive trials

were repeated under the conditions that elicited the positive response. If no positive responses were

seen, all negative trials were repeated.

In this assay, a positive response is defined as a reproducible, dose-related increase in histidineindependent (revertant) colonies in any one strain/activation combination. An equivocal response is

defined as an increase in revertants that is not dose related, not reproducible, or is of insufficient

magnitude to support a determination of mutagenicity. A negative response was obtained when no

increase in revertant colonies was observed following chemical treatment. There was no minimum

percentage or fold increase required for a chemical to the judged positive or weakly positive.

CHINESE HAMSTER OVARY CELL CYTOGENETICS PROTOCOLS

Testing was performed as reported by Loveday et al. (1990). 1-Amino-2,4-dibromoanthraquinone was

sent to each of two testing laboratories as a coded aliquot by Radian Corporation. The aliquots were

tested in cultured Chinese hamster ovary (CHO) cells for induction of sister chromatid exchanges

(SCEs) and chromosomal aberrations (Abs), both in the presence and absence of Aroclor 1254-induced

male Sprague-Dawley rat liver S9 and cofactor mix. Cultures were handled under gold lights to prevent

photolysis of bromodeoxyuridine-substituted DNA. Each test consisted of concurrent solvent and

positive controls and of at least three doses of 1-amino-2,4-dibromoanthraquinone. A single flask per

dose was used, and tests yielding equivocal or positive results were repeated.

Sister Chromatid Exchange Test: In the SCE test without S9, CHO cells were incubated for 26 hours

with 1-amino-2,4-dibromoanthraquinone in McCoy's 5A medium supplemented with fetal bovine

serum, l-glutamine, and antibiotics. Bromodeoxyuridine (BrdU) was added 2 hours after culture

initiation. After 26 hours, the medium containing 1-amino-2,4-dibromoanthraquinone was removed

and replaced with fresh medium plus BrdU and Colcemid, and incubation was continued for 2 hours.

Cells were then harvested by mitotic shake-off, fixed, and stained with Hoechst 33258 and Giemsa.

In the SCE test with S9, cells were incubated with 1-amino-2,4-dibromoanthraquinone, serum-free

medium, and S9 for 2 hours. The medium was then removed and replaced with medium containing

serum and BrdU and no 1-amino-2,4-dibromoanthraquinone and incubation proceeded for an

additional 26 hours, with Colcemid present for the final 2 hours. Harvesting and staining were the

same as for cells treated without S9. All slides were scored blind and those from a single test were

read by the same person. Usually, 50 second-division metaphase cells were scored for frequency of

SCEs/cell from each dose level. The high dose was limited by toxicity.
Genetic Toxicology 327

Statistical analyses were conducted on the slopes of the dose-response curves and the individual dose points

(Galloway et al., 1987). An SCE frequency 20% above the concurrent solvent control value was chosen as a

statistically conservative positive response. The probability of this level of difference occurring by chance at

one dose point is less than 0.01; the probability for such a chance occurrence at two dose points is less than

0.001. An increase of 20% or greater at any single dose was considered weak evidence of activity; increases at

two or more doses resulted in a determination that the trial was positive. A statistically significant trend

(P#0.005) in the absence of any responses reaching 20% above background led to a call of equivocal.

Chromosomal Aberrations Test: In the Abs test without S9, cells were incubated in McCoy's 5A medium with

1-amino-2,4-dibromoanthraquinone for 8 to 10 hours; Colcemid was added and incubation continued for 2

hours. The cells were then harvested by mitotic shake-off, fixed, and stained with Giemsa. For the Abs test

with S9, cells were treated with 1-amino-2,4-dibromoanthraquinone and S9 for 2 hours, after which the

treatm ent medium was removed and the cells incubated for 10 to 11 hours in fresh medium, with Colcemid

present for the final 2 hours. Cells were harvested in the same manner as for the treatment without S9. The

high dose was limited to 100 :g/mL.

Cells were selected for scoring on the basis of good morphology and completeness of karyotype

(21 ± 2 chromosomes). All slides were scored blind and those from a single test were read by the same

person. Two hundred first-division metaphase cells were scored at each dose level. Classes of aberrations

included simple (breaks and terminal deletions), complex (rearrangements and translocations), and other

(pulverized cells, despiralized chromosomes, and cells containing 10 or more aberrations).

Chromosomal aberration data are presented as percentage of cells with aberrations. To arrive at a statistical

call for a trial, analyses were conducted on both the dose response curve and individual dose points. For a

single trial, a statistically significant (P#0.05) difference for one dose point and a significant trend (P#0.015)

are considered weak evidence for a positive response; significant differences for two or more doses indicated

the trial was positive. A positive trend test in the absence of a statistically significant increase at any one dose

resulted in an equivocal call (Galloway et al., 1987). Ultimately, the trial calls were based on a consideration

of the statistical analyses as well as the biological information available to the reviewers.

RESULTS

1-Amino-2,4-dibromoanthraquinone (100 to 10,000 :g/plate) was tested for induction of gene mutations in

four strains of Salmonella typhimurium in a preincubation protocol with and without Aroclor 1254-induced

male Sprague-Dawley rat or Syrian hamster liver S9 (Table G1; Haworth et al., 1983). 1-Amino2,4-dibromoanthraquinone was positive in the absence of S9 in the frameshift strains TA98 and TA1537;

with S9, an equivocal response was obtained in TA1537, and TA98 was negative. In TA100, 1-amino2,4-dibromoanthraquinone gave equivocal responses with and without S9, and all trials were negative in

TA1535. The equivocal calls were the results of positive or weakly positive responses that were not

duplicated in a second trial. Precipitation of 1-amino-2,4-dibromoanthraquinone occurred at

concentrations of 100 :g/plate and above, and this may have been a factor in the nonreproducibility of the

results.

1-Amino-2,4-dibromoanthraquinone was tested in two laboratories for induction of SCEs and Abs in cultured

CHO cells, with and without Aroclor 1254-induced male Sprague-Dawley rat liver S9. In the SCE test,

Environmental Health Research & Testing observed a significant increase in SCEs only in the absence of S9,

while Bioassay Systems Corporation recorded positive responses with and without S9 (Table G2; Loveday et

al., 1990). The discrepancy with S9 cannot be explained by a difference in dose levels employed at the two

laboratories because positive responses were seen at 3, 10, 15, and 30 :g/mL at Bioassay Systems Corporation,

whereas negative trials resulted from testin g doses u p t o 100 : g /m L a t
328 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

Environmental Health Research & Testing. In the Abs test, Environmental Health Research &

Testing observed a weakly positive response only in the absence of S9 (Table G3). Bioassay Systems

Corporation obtained a positive response in the first trial without S9 but did not duplicate the positive

response in the second trial (Table G3), and the overall call without S9 was concluded to be equivocal

(Loveday et al., 1990). Neither laboratory observed an increase in Abs with 1-amino2,4-dibromoanthraquinone in the presence of S9.
329 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE G1

Mutagenicity of 1-Amino-2,4-dibromoanthraquinone in Salmonella typhimuriuma

Revertants/plateb

Strain Dose !S9 +10 hamster S9 +10 rat S9

:g/plate Trial 1 Trial 2 Trial 1 Trial 2 Trial 1 Trial 2

TA100 0 119 ± 4.2 170 ± 2.6 112 ± 6.7 138 ± 10.6 121 ± 7.8 156 ± 12.0

100 115 ± 5.0 146 ± 6.2c143 ± 11.8 157 ± 10.2 202 ± 8.0 182 ± 3.5

333 136 ± 7.5c140 ± 13.0c136 ± 2.9c218 ± 11.8 194 ± 3.8c184 ± 10.7

1,000 154 ± 6.4c177 ± 6.4c151 ± 8.5c209 ± 7.2c191 ± 4.4c181 ± 9.1c

3,333 158 ± 6.2c 184 ± 8.8c190 ± 18.4c194 ± 12.9c185 ± 7.8c159 ± 5.4c

10,000 218 ± 24.5c 233 ± 27.0c178 ± 12.5c215 ± 10.7c205 ± 14.0c184 ± 11.0c

Trial summary Weakly Positive Equivocal Weakly Positive Equivocal Equivocal Negative

Positive controld420 ± 7.0 365 ± 4.7 1,027 ± 43.5 1,978 ± 31.5 612 ± 14.7 1,703 ± 202.1

TA1535 0 29 ± 0.9 26 ± 6.4 11 ± 1.8 16 ± 0.6 13 ± 0.6 16 ± 0.9

100 22 ± 2.1 25 ± 3.8c9 ± 2.3 14 ± 2.7 11 ± 3.0 13 ± 2.7

333 27 ± 1.5c22 ± 3.1c10 ± 0.9c11 ± 0.9 13 ± 1.7c16 ± 3.0

1,000 28 ± 5.6c30 ± 7.6c13 ± 2.1c14 ± 0.7c8 ± 1.5c12 ± 1.9c

3,333 25 ± 3.5c30 ± 3.5c11 ± 1.7c14 ± 3.0c10 ± 2.0c13 ± 2.4c

10,000 26 ± 6.0c25 ± 5.0c10 ± 2.3c11 ± 0.9c7 ± 1.5c11 ± 0.9c

Trial summary Negative Negative Negative Negative Negative Negative

Positive control 554 ± 6.7 359 ± 12.5 377 ± 16.4 606 ± 23.6 346 ± 24.7 528 ± 24.8

TA1537 0 5 ± 0.7 5 ± 1.2 9 ± 1.8 16 ± 2.1 8 ± 1.0 12 ± 3.2

100 11 ± 1.2 9 ± 1.2c8 ± 0.9 13 ± 3.2 10 ± 1.2 12 ± 2.5

333 11 ± 0.6c8 ± 1.2c15 ± 2.0c13 ± 1.2 16 ± 2.7c19 ± 4.5

1,000 13 ± 1.8c8 ± 1.2c17 ± 2.7c12 ± 2.6c20 ± 1.7c18 ± 2.5c

3,333 19 ± 5.3c17 ± 2.3c21 ± 2.9c21 ± 2.1c24 ± 3.8c19 ± 2.7c

10,000 28 ± 0.7c32 ± 0.9c30 ± 3.0c27 ± 4.0c28 ± 0.3c30 ± 3.5c

Trial summary Positive Positive Positive Negative Positive Equivocal

Positive control 141 ± 3.2 147 ± 18.7 324 ± 8.8 367 ± 4.4 234 ± 14.0 308 ± 36.0

TA98 0 23 ± 4.9 19 ± 1.5 28 ± 2.0 29 ± 0.7 36 ± 3.3 36 ± 3.2

100 20 ± 3.2 18 ± 3.6c24 ± 3.2 30 ± 1.8 40 ± 1.5 44 ± 7.4

333 19 ± 2.1c29 ± 1.5c33 ± 8.3c21 ± 3.2 41 ± 3.5c36 ± 6.1

1,000 33 ± 1.7c24 ± 4.6c30 ± 4.6c33 ± 3.8c38 ± 4.2c46 ± 3.3c

3,333 46 ± 6.0c37 ± 1.2c43 ± 4.4c47 ± 1.8c38 ± 4.4c50 ± 2.7c

10,000 73 ± 10.6c86 ± 9.5c53 ± 3.8c46 ± 0.7c45 ± 2.9c41 ± 2.9c

Trial summary Positive Positive Equivocal Negative Negative Negative

Positive control 536 ± 43.2 591 ± 76.8 1,048 ± 40.5 1,503 ± 69.9 507 ± 21.0 1,080 ± 15.6

a The detailed protocol and these data are presented in Haworth et al. (1983). 

b Revertants are presented as mean ± the standard error from three plates.

c

Precipitate on plate

d The positive controls in the absence of metabolic activation were sodium azide (TA1535 and TA100), 9-aminoacridine (TA1537), and 4-nitroo-phenylenediamine (TA98). The positive control for metabolic activation with all strains was 2-aminoanthracene.
Genetic Toxicology 330

TABLE G2

Induction of Sister Chromatid Exchanges in Chinese Hamster Ovary Cells

by 1-Amino-2,4-dibromoanthraquinonea

No. of SCEs/ Relative

Dose Total Chromo- No. of Chromo- SCEs/ Hrs Change of SCEs/

Compound (g/mL) Cells somes SCEs some Cell in BrdU Chromosomeb

(%)

Study Performed at Environmental Health Research & Testing

S9

Trial 1

Summary: Positive

Dimethylsulfoxide 50 1,049 435 0.41 8.7 26.0

Mitomycin-C 0.0008 50 1,039 591 0.56 11.8 26.0 37.17

0.0050 10 210 194 0.92 19.4 26.0 122.77

1-Amino-2,4-dibromoanthraquinone

1.6 50 1,043 516 0.49 10.3 26.0 19.30

5.0 50 1,041 526 0.50 10.5 26.0 21.85*

16.0 50 1,048 565 0.53 11.3 26.0 30.01*

50.0 14c 290 171 0.58 12.2 26.0 42.19*

P<0.001d

Trial 2

Summary: Weakly Positive

Dimethylsulfoxide 50 1,047 468 0.44 9.4 26.0

Mitomycin-C 0.0005 50 1,043 574 0.55 11.5 26.0 23.12

0.0050 10 210 273 1.30 27.3 26.0 190.83

1-Amino-2,4-dibromoanthraquinone

1.6 50 1,046 515 0.49 10.3 26.0 10.15

5.0 50 1,044 538 0.51 10.8 26.0 15.29

16.0 50 1,047 554 0.52 11.1 26.0 18.38

50.0 13c 267 147 0.55 11.3 26.0 23.17*

P=0.001

+S9

Summary: Negative

Dimethylsulfoxide 50 1,051 432 0.41 8.6 26.0

Cyclophosphamide 0.1 50 1,046 546 0.52 10.9 26.0 26.99

0.6 10 210 193 0.91 19.3 26.0 123.59

1-Amino-2,4-dibromoanthraquinone

5.0 50 1,049 412 0.39 8.2 26.0 4.45

16.0 50 1,050 465 0.44 9.3 26.0 7.74

50.0 50 1,040 489 0.47 9.8 26.0 14.39

100.0 50 1,050 480 0.45 9.6 26.0 11.22

P=0.003

* Significant positive response (P0.01) a A detailed description of the protocol and these data are presented by Loveday et al. (1990). SCE = sister chromatid exchange; BrdU =

bromodeoxyuridine.
331 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE G2

Induction of Sister Chromatid Exchanges in Chinese Hamster Ovary Cells

by 1-Amino-2,4-dibromoanthraquinone (continued)

No. of SCEs/ Relative

Dose Total Chromo- No. of Chromo- SCEs/ Hrs Change of SCEs/

Compound (g/mL) Cells somes SCEs some Cell in BrdU Chromosome

(%)

Study Performed at Bioassay Systems Corporation

S9

Summary: Weakly Positive

Dimethylsulfoxide 50 1,046 335 0.32 6.7 26.0

Mitomycin-C 0.002 50 1,041 567 0.54 11.3 26.0 70.07

0.010 10 208 243 1.16 24.3 26.0 264.78

1-Amino-2,4-dibromoanthraquinone 

2.5 50 1,037 355 0.34 7.1 26.0 6.89

5.0 50 1,040 395 0.37 7.9 26.0 18.59

10.0 50 1,043 436 0.41 8.7 26.0 30.53*

P<0.001

+S9

Trial 1

Summary: Positive

Dimethylsulfoxide 50 1,047 420 0.40 8.4 26.0

Cyclophosphamide 0.5 50 1,038 642 0.61 12.8 26.0 54.18

2.5 10 206 291 1.41 29.1 26.0 252.15

1-Amino-2,4-dibromoanthraquinone 

3.01 50 1,044 528 0.50 10.6 26.0 26.08*

10.10 50 1,047 581 0.55 11.6 26.0 38.33*

30.10 50 1,048 597 0.56 11.9 26.0 42.01*

P<0.001

Trial 2

Summary: Weakly Positive

Dimethylsulfoxide 50 1,032 402 0.38 8.0 26.0

Cyclophosphamide 0.5 50 1,044 593 0.56 11.9 26.0 45.82

2.5 10 207 248 1.19 24.8 26.0 207.57

1-Amino-2,4-dibromoanthraquinone

7.5 50 1,041 440 0.42 8.8 26.0 8.51

10.0 50 1,041 452 0.43 9.0 26.0 11.47

15.0 50 1,043 501 0.48 10.0 26.0 23.31*

P<0.001

b SCEs/chromosome in treated cells versus SCEs/chromosome in solvent control cells c Decreased number of metaphases available for evaluation due to the cytostatic nature of 1-amino-2,4-dibromoanthraquinone d Significance of relative SCEs/chromosome tested by the linear regression trend test vs. log of the dose
Genetic Toxicology 332

TABLE G3

Induction of Chromosomal Aberrations in Chinese Hamster Ovary Cells

by 1-Amino-2,4-dibromoanthraquinonea

S9 +S9 

Dose Total No. of Abs/ Cells with Dose Total No. of Abs/ Cells with

(g/mL) Cells Abs Cell Abs (%) (g/mL) Cells Abs Cell Abs (%)

Study Performed at Environmental Health Research & Testing

Trial 1  Harvest Time: 12.0 hours Harvest Time: 13.0 hours

Summary: Weakly Positive Summary: Negative

Dimethylsulfoxide Dimethylsulfoxide

200 3 0.02 1.5 200 4 0.02 2.0

Mitomycin-C Cyclophosphamide

0.0625 200 23 0.12 10.0 5.0 200 15 0.08 7.0

0.2500 50 18 0.36 32.0 7.5 50 19 0.38 36.0

1-Amino-2,4-dibromoanthraquinone 1-Amino-2,4-dibromoanthraquinone

5 200 7 0.04 3.5 16 200 3 0.02 0.5

16 200 9 0.05 4.0 50 200 2 0.01 1.0

50 200 12 0.06 5.5* 100 200 4 0.02 2.0

P=0.017b P=0.467

Trial 2  Harvest Time: 12.0 hours

Summary: Weakly Positive

Dimethylsulfoxide

200 4 0.02 2.0

Mitomycin-C

0.0625 200 25 0.13 12.5

0.2500 50 21 0.42 36.0

1-Amino-2,4-dibromoanthraquinone

16 200 12 0.06 5.0

30 200 9 0.05 4.5

50 200 13 0.07 6.0

P=0.039
333 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE G3

Induction of Chromosomal Aberrations in Chinese Hamster Ovary Cells

by 1-Amino-2,4-dibromoanthraquinone (continued)

!S9 +S9 

Dose Total No. of Abs/ Cells with Dose Total No. of Abs/ Cells with

(:g/mL) Cells Abs Cell Abs (%) (:g/mL) Cells Abs Cell Abs (%)

Study Performed at Bioassay Systems Corporation

Trial 1 ) Harvest Time: 10.5 hours Harvest Time: 12.0 hours

Summary: Positive Summary: Negative

Dimethylsulfoxide Dimethylsulfoxide

200 0 0.00 0.0 200 4 0.02 2.0

Mitomycin-C Cyclophosphamide

1 200 51 0.26 20.5 50 50 25 0.50 34.0

5 50 47 0.94 56.0

1-Amino-2,4-dibromoanthraquinone

1-Amino-2,4-dibromoanthraquinone 3.02 200 9 0.05 4.0

3.02 200 7 0.04 3.0* 10.10 200 13 0.07 5.5

10.10 200 5 0.03 2.5* 30.20 200 7 0.04 3.5

30.20 200 4 0.02 1.5

P=0.153

P=0.164

Trial 2 ) Harvest Time: 10.0 hours

Summary: Negative

Dimethylsulfoxide

200 5 0.03 2.0

Mitomycin-C

1 200 171 0.86 40.5

5 50 41 0.82 56.0

1-Amino-2,4-dibromoanthraquinone

1 200 12 0.06 5.0

3 200 18 0.09 6.0

10 200 11 0.06 4.5

30 200 6 0.03 2.0

P=0.550

* Significant positive response (P<0.05) 

a The detailed protocol and these data are presented in Loveday et al. (1990). Abs = aberrations.

b

Significance of percent cells with aberrations tested by the linear regression trend test vs. log of the dose
Genetic Toxicology 334

 
335

APPENDIX H

ORGAN WEIGHTS 

AND ORGAN-WEIGHT-TO-BODY-WEIGHT RATIOS

TABLE H1 Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats 

in the 13-Week Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . 336

TABLE H2 Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats 

at the 9-Month Interim Evaluation in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 338

TABLE H3 Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats 

at the 15-Month Interim Evaluation in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 339

TABLE H4 Organ Weights and Organ-Weight-to-Body-Weight Ratios for Mice 

in the 13-Week Feed Study of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . 340

TABLE H5 Organ Weights and Organ-Weight-to-Body-Weight Ratios for Mice 

at the 15-Month Interim Evaluation in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 342
336 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE H1

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats in the 13-Week Feed Study 

of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 2,500 ppm 5,000 ppm 10,000 ppm 25,000 ppm 50,000 ppm

Male

n 10 10 9 9 10 7

Necropsy body wt 359 ± 4 327 ± 4** 332 ± 4** 305 ± 2** 232 ± 3** 161 ± 6**

Brain

Absolute 1.924 ± 0.021 1.942 ± 0.014 1.945 ± 0.016 1.914 ± 0.019 1.798 ± 0.016** 1.708 ± 0.015** 

Relative 5.36 ± 0.08 5.95 ± 0.06** 5.86 ± 0.07** 6.26 ± 0.05** 7.77 ± 0.07** 10.69 ± 0.37** 

Heart

Absolute 0.953 ± 0.019 0.964 ± 0.042 0.923 ± 0.026 0.895 ± 0.019 0.683 ± 0.008**b 0.579 ± 0.050** 

Relative 2.65 ± 0.05 2.95 ± 0.12 2.78 ± 0.08 2.90 ± 0.05 2.94 ± 0.06b 3.59 ± 0.26** 

R. Kidney

Absolute 1.102 ± 0.017 1.094 ± 0.023 1.121 ± 0.018 1.096 ± 0.015c 0.892 ± 0.011** 0.708 ± 0.026** 

Relative 3.07 ± 0.04 3.35 ± 0.06** 3.37 ± 0.06** 3.58 ± 0.03**c 3.86 ± 0.04** 4.40 ± 0.09** 

Liver

Absolute 12.513 ± 0.288 14.374 ± 0.381** 16.172 ± 0.309** 16.295 ± 0.370** 13.661 ± 0.206 12.333 ± 0.562 

Relative 34.84 ± 0.71 43.96 ± 0.95** 48.64 ± 0.67** 52.67 ± 1.02** 59.03 ± 0.76** 76.58 ± 1.70** 

Lung

Absolute 1.322 ± 0.042 1.338 ± 0.026 1.329 ± 0.047 1.259 ± 0.011 1.067 ± 0.038** 0.861 ± 0.014** 

Relative 3.68 ± 0.12 4.10 ± 0.07 4.00 ± 0.14 4.13 ± 0.05* 4.62 ± 0.18** 5.40 ± 0.22** 

R. Testis

Absolute 1.468 ± 0.020b 1.506 ± 0.020 1.508 ± 0.014 1.479 ± 0.020 1.390 ± 0.026* 1.082 ± 0.037** 

Relative 4.08 ± 0.08b 4.61 ± 0.05** 4.54 ± 0.06** 4.82 ± 0.07** 6.01 ± 0.11** 6.73 ± 0.12** 

Thymus

Absolute 0.235 ± 0.014 0.193 ± 0.009* 0.163 ± 0.018** 0.146 ± 0.008** 0.112 ± 0.010**b 0.065 ± 0.010** 

Relative 0.65 ± 0.04 0.59 ± 0.03 0.49 ± 0.05* 0.48 ± 0.03** 0.48 ± 0.04**b 0.42 ± 0.07** 
Organ Weight Analyses 337

TABLE H1

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats in the 13-Week Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,500 ppm 5,000 ppm 10,000 ppm 25,000 ppm 50,000 ppm

Female

n 10 10 10 10 10 9

Necropsy body wt 209 ± 3 195 ± 3** 188 ± 3** 184 ± 2** 157 ± 2** 129 ± 4** 

Brain

Absolute 1.809 ± 0.019 1.757 ± 0.014* 1.727 ± 0.016**b 1.710 ± 0.012** 1.683 ± 0.018** 1.598 ± 0.017** 

Relative 8.66 ± 0.13 9.02 ± 0.19 9.20 ± 0.10b 9.32 ± 0.11* 10.71 ± 0.15** 12.49 ± 0.39** 

Heart

Absolute 0.616 ± 0.010 0.605 ± 0.015 0.580 ± 0.016 0.557 ± 0.014** 0.473 ± 0.012**b 0.439 ± 0.016** 

Relative 2.94 ± 0.03 3.10 ± 0.09 3.09 ± 0.08 3.04 ± 0.07 3.03 ± 0.06b 3.42 ± 0.11** 

R. Kidney

Absolute 0.629 ± 0.013 0.684 ± 0.015 0.667 ± 0.012 0.656 ± 0.014 0.590 ± 0.007* 0.532 ± 0.011** 

Relative 3.01 ± 0.04 3.51 ± 0.07** 3.55 ± 0.04** 3.57 ± 0.05** 3.75 ± 0.06** 4.15 ± 0.15** 

Liver

Absolute 6.561 ± 0.095 7.674 ± 0.202** 8.335 ± 0.213** 8.509 ± 0.159** 8.090 ± 0.158** 8.243 ± 0.236** 

Relative 31.39 ± 0.41 39.28 ± 0.77** 44.35 ± 1.06** 46.33 ± 0.60** 51.45 ± 0.77** 64.12 ± 1.45** 

Lung

Absolute 1.001 ± 0.018 1.010 ± 0.043b 0.937 ± 0.029 0.930 ± 0.020 0.855 ± 0.023**b 0.702 ± 0.022** 

Relative 4.79 ± 0.06 5.15 ± 0.17b 4.98 ± 0.12 5.07 ± 0.11 5.47 ± 0.16**b 5.46 ± 0.12** 

Thymus

Absolute 0.221 ± 0.007 0.179 ± 0.008**b 0.155 ± 0.006** 0.145 ± 0.008** 0.130 ± 0.016**b 0.062 ± 0.006** 

Relative 1.06 ± 0.03 0.91 ± 0.04b 0.82 ± 0.03** 0.79 ± 0.04** 0.84 ± 0.11**b 0.48 ± 0.04** 

* Significantly different (P#0.05) from the control group by Williams' or Dunnett's test

** P#0.01 a Organ weights and body weights are given in grams; organ-weight-to-body-weight ratios are given as mg organ weight/g body weight (mean ± standard error) b n=9 c n=8
338 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE H2

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats at the 9-Month Interim Evaluation 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm 20,000 ppm

(15-month exposure)

Male

n 10 10 10 10 10

Necropsy body wt 435 ± 11 424 ± 9 398 ± 4** 388 ± 7** 372 ± 10** 

R. Kidney

Absolute 1.247 ± 0.040 1.362 ± 0.047 1.300 ± 0.022 1.259 ± 0.026 1.294 ± 0.044 

Relative 2.87 ± 0.06 3.21 ± 0.07** 3.27 ± 0.06** 3.25 ± 0.04** 3.47 ± 0.05** 

Liver

Absolute 13.934 ± 0.391 16.876 ± 0.506** 16.578 ± 0.441** 18.009 ± 0.594** 17.925 ± 0.901** 

Relative 32.04 ± 0.54 39.80 ± 0.72** 41.60 ± 0.85** 46.37 ± 0.80** 48.06 ± 1.92** 

Female

n 10 10 10 10 10

Necropsy body wt 251 ± 7 236 ± 4* 229 ± 5** 218 ± 3** 208 ± 3** 

R. Kidney

Absolute 0.710 ± 0.014b 0.786 ± 0.020* 0.770 ± 0.019 0.775 ± 0.016* 0.739 ± 0.016

Relative 2.90 ± 0.06b 3.35 ± 0.09** 3.37 ± 0.07** 3.56 ± 0.06** 3.56 ± 0.09**

Liver

Absolute 7.378 ± 0.400 8.753 ± 0.240** 9.361 ± 0.253** 9.313 ± 0.124** 9.651 ± 0.241** 

Relative 29.17 ± 0.95 37.20 ± 0.88** 40.89 ± 0.74** 42.83 ± 0.43** 46.38 ± 0.94** 

* Significantly different (P#0.05) from the control group by Williams' or Dunnett's test

** P#0.01 a Organ weights and body weights are given in grams; organ-weight-to-body-weight ratios are given as mg organ weight/g body weight (mean ± standard error). b n=9
Organ Weight Analyses 339

TABLE H3

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Rats at the 15-Month Interim Evaluation 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 10,000 ppm 20,000 ppm 20,000 ppm

(15-month exposure) (9-month stop-exposure)

Male

n 10 10 10 7

Necropsy body wt 481 ± 6 394 ± 7** 361 ± 11** 374 ± 22** 

R. Kidney

Absolute 1.431 ± 0.032 1.472 ± 0.044 1.543 ± 0.041 1.319 ± 0.032 

Relative 2.98 ± 0.07 3.73 ± 0.07** 4.34 ± 0.25** 3.61 ± 0.26** 

Liver

Absolute 16.348 ± 0.522 20.110 ± 1.296 26.448 ± 2.340** 25.495 ± 5.628* 

Relative 34.00 ± 1.01 50.88 ± 2.84 75.40 ± 8.94** 74.37 ± 20.34** 

Female

n 10 10 8 9

Necropsy body wt 318 ± 6 234 ± 11** 216 ± 4** 260 ± 9** 

R. Kidney

Absolute 0.941 ± 0.028 0.971 ± 0.035 0.994 ± 0.037 0.917 ± 0.035 

Relative 2.96 ± 0.06 4.18 ± 0.08** 4.67 ± 0.17** 3.55 ± 0.13** 

Liver

Absolute 9.394 ± 0.276 11.986 ± 0.644** 13.738 ± 0.358** 10.804 ± 0.679** 

Relative 29.59 ± 0.85 51.31 ± 1.29** 64.66 ± 2.34** 41.65 ± 2.43** 

* Significantly different (P#0.05) from the control group by Williams' or Dunnett's test

** P#0.01 a Organ weights and body weights are given in grams; organ-weight-to-body-weight ratios are given as mg organ weight/g body weight (mean ± standard error).
340 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE H4

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Mice in the 13-Week Feed Study 

of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 2,500 ppm 5,000 ppm 10,000 ppm 25,000 ppm 50,000 ppm

Male

n 10 10 9 10 9 10

Necropsy body wt 30.0 ± 0.6 30.2 ± 0.5 30.0 ± 0.7 31.2 ± 0.5 30.4 ± 0.6 30.5 ± 0.4 

Brain

Absolute 0.454 ± 0.005b 0.450 ± 0.006 0.452 ± 0.004 0.446 ± 0.007 0.446 ± 0.007 0.437 ± 0.017b

Relative 15.26 ± 0.28b 14.98 ± 0.38 15.12 ± 0.32 14.33 ± 0.28 14.68 ± 0.27 14.35 ± 0.56b Heart

Absolute 0.146 ± 0.004 0.141 ± 0.005 0.148 ± 0.004 0.147 ± 0.004 0.139 ± 0.003 0.148 ± 0.004 

Relative 4.88 ± 0.10 4.68 ± 0.10 4.92 ± 0.10 4.70 ± 0.10 4.57 ± 0.06 4.85 ± 0.10 

R. Kidney

Absolute 0.266 ± 0.005 0.257 ± 0.010 0.250 ± 0.004 0.251 ± 0.008 0.231 ± 0.006* 0.250 ± 0.008* 

Relative 8.90 ± 0.20 8.51 ± 0.28 8.35 ± 0.17 8.03 ± 0.20** 7.59 ± 0.17** 8.19 ± 0.24** 

Liver

Absolute 1.593 ± 0.041 1.738 ± 0.039 1.765 ± 0.052* 2.011 ± 0.077**b 2.002 ± 0.069** 2.381 ± 0.061** 

Relative 53.11 ± 0.94 57.60 ± 0.65* 58.94 ± 1.59** 63.92 ± 1.94**b 65.73 ± 1.79** 78.05 ± 1.84** 

Lung

Absolute 0.215 ± 0.006 0.201 ± 0.006b 0.195 ± 0.005 0.196 ± 0.008 0.197 ± 0.004 0.205 ± 0.011 

Relative 7.17 ± 0.10 6.71 ± 0.21b 6.55 ± 0.25 6.26 ± 0.21 6.47 ± 0.09 6.75 ± 0.40 

R. Testis

Absolute 0.114 ± 0.002 0.113 ± 0.008b 0.110 ± 0.002 0.110 ± 0.003b 0.108 ± 0.002c 0.111 ± 0.002b

Relative 3.81 ± 0.11 3.74 ± 0.22b 3.69 ± 0.08 3.51 ± 0.09b 3.53 ± 0.04c 3.66 ± 0.09b

 

Thymus

Absolute 0.031 ± 0.001 0.038 ± 0.003d 0.031 ± 0.004 0.034 ± 0.003 0.033 ± 0.002 0.029 ± 0.002 

Relative 1.05 ± 0.04 1.24 ± 0.07d 1.05 ± 0.14 1.09 ± 0.07 1.09 ± 0.06 0.96 ± 0.06 
Organ Weight Analyses 341

TABLE H4

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Mice in the 13-Week Feed Study 

of 1-Amino-2,4-dibromoanthraquinone (continued)

0 ppm 2,500 ppm 5,000 ppm 10,000 ppm 25,000 ppm 50,000 ppm

Female

n 10 10 10 9 10 10

Necropsy body wt 23.8 ± 0.3 24.6 ± 0.6 24.0 ± 0.7 24.5 ± 0.4 23.1 ± 0.3 24.2 ± 0.4 

Brain

Absolute 0.461 ± 0.005 0.456 ± 0.006 0.452 ± 0.004 0.446 ± 0.005 0.450 ± 0.004 0.438 ± 0.006** 

Relative 19.37 ± 0.24 18.65 ± 0.36 19.01 ± 0.62 18.20 ± 0.31 19.50 ± 0.30 18.14 ± 0.31 

Heart

Absolute 0.109 ± 0.002 0.112 ± 0.002 0.111 ± 0.003 0.117 ± 0.003 0.110 ± 0.002 0.116 ± 0.003 

Relative 4.56 ± 0.12 4.59 ± 0.07 4.63 ± 0.08 4.75 ± 0.12 4.78 ± 0.10 4.81 ± 0.11 

R. Kidney

Absolute 0.152 ± 0.004 0.166 ± 0.004* 0.156 ± 0.004 0.163 ± 0.003 0.158 ± 0.003 0.158 ± 0.003 

Relative 6.36 ± 0.11 6.77 ± 0.13 6.51 ± 0.15 6.64 ± 0.15 6.84 ± 0.14* 6.55 ± 0.07 

Liver

Absolute 1.146 ± 0.022 1.453 ± 0.053** 1.529 ± 0.055** 1.666 ± 0.030** 1.613 ± 0.043** 1.904 ± 0.043** 

Relative 48.16 ± 1.27 59.10 ± 1.30** 63.92 ± 2.11** 67.94 ± 1.16** 69.83 ± 1.49** 78.72 ± 1.02** 

Lung

Absolute 0.184 ± 0.008 0.175 ± 0.007 0.186 ± 0.006 0.185 ± 0.009 0.174 ± 0.006 0.187 ± 0.007 

Relative 7.73 ± 0.35 7.15 ± 0.32 7.81 ± 0.31 7.52 ± 0.30 7.55 ± 0.25 7.77 ± 0.32 

Thymus

Absolute 0.036 ± 0.001 0.037 ± 0.002 0.038 ± 0.003 0.038 ± 0.002 0.029 ± 0.002 0.034 ± 0.003 

Relative 1.51 ± 0.06 1.52 ± 0.08 1.60 ± 0.11 1.55 ± 0.08 1.25 ± 0.09 1.39 ± 0.12 

* Significantly different (P#0.05) from the control group by Williams' or Dunnett's test

** P#0.01 a Organ weights and body weights are given in grams; organ-weight-to-body-weight ratios are given as mg organ weight/g body weight (mean ± standard error). b n=9 c n=7 d n=8
342 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE H5

Organ Weights and Organ-Weight-to-Body-Weight Ratios for Mice at the 15-Month Interim Evaluation 

in the 2-Year Feed Study of 1-Amino-2,4-dibromoanthraquinonea

0 ppm 10,000 ppm 20,000 ppm

Male

n 10 9 10

Necropsy body wt 42.9 ± 1.2 39.9 ± 0.9 35.5 ± 1.1** 

R. Kidney

Absolute 0.334 ± 0.014 0.289 ± 0.011* 0.288 ± 0.011* 

Relative 7.81 ± 0.32 7.26 ± 0.23 8.18 ± 0.40 

Liver

Absolute 1.979 ± 0.146 2.022 ± 0.051 1.932 ± 0.092 

Relative 46.69 ± 4.43 50.85 ± 1.59 54.45 ± 2.13 

Female

n 10 10 10

Necropsy body wt 38.5 ± 1.5 34.5 ± 1.9 33.6 ± 0.9* 

R. Kidney

Absolute 0.215 ± 0.005 0.209 ± 0.005 0.208 ± 0.006b

Relative 5.66 ± 0.25 6.21 ± 0.36 6.24 ± 0.23b

 

Liver

Absolute 1.373 ± 0.038 1.657 ± 0.042** 1.805 ± 0.043** 

Relative 35.96 ± 1.06 48.92 ± 2.20** 53.86 ± 1.13** 

* Significantly different (P#0.05) from the control group by Williams' or Dunnett's test

** P#0.01 a Organ weights and body weights are given in grams; organ-weight-to-body-weight ratios are given as mg organ weight/g body weight (mean ± standard error). b n=9
343

APPENDIX I

CHEMICAL CHARACTERIZATION AND

DOSE FORMULATION STUDIES

PROCUREMENT AND CHARACTERIZATION OF 1-AMINO-2,4-DIBROMOANTHRAQUINONE . . . . . . . . . . . . . . . . . 344

PREPARATION AND ANALYSIS OF DOSE FORMULATIONS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 345

FIGURE I1 Infrared Absorption Spectrum of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . 346

FIGURE I2 Nuclear Magnetic Resonance Spectrum of 1-Amino-2,4-dibromoanthraquinone . . . . . . . 347

TABLE I1 Preparation and Storage of Dose Formulations in the Feed Studies 

of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 348

TABLE I2 Results of Analysis of Dose Formulations Administered to Rats and Mice 

in the 13-Week Feed Studies of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . 349

TABLE I3 Results of Analysis of Dose Formulations Administered to Rats and Mice

in the 2-Year Feed Studies of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . 350

TABLE I4 Results of Referee Analysis of Dose Formulations in the 13-Week

and 2-Year Feed Studies of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . 352
344 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

CHEMICAL CHARACTERIZATION AND

DOSE FORMULATION STUDIES

PROCUREMENT AND CHARACTERIZATION 

OF 1-AMINO-2,4-DIBROMOANTHRAQUINONE

1-Amino-2,4-dibromoanthraquinone was obtained from American Color and Chemical Corporation (Charlotte, NC) (lot

1076-C) and Mobay Corporation (Pittsburgh, PA). Lot 1076-C was used in the 13-week studies and for 2 months of the

2-year studies. For tracking purposes, the lot from Mobay Corporation was assigned the number M061583 and was used

throughout the remainder of the 2-year studies. Identity, purity, and stability analyses were conducted by the analytical

chemistry laboratory, Midwest Research Institute (MRI) (Kansas City, MO). MRI reports on analyses performed in support of

the 1-amino-2,4-dibromoanthraquinone studies are on file at the National Institute of Environmental Health Sciences.

The two lots of the study chemical, a reddish brown to orange powder, were identified as 1-amino-2,4-dibromoanthraquinone

by infrared, ultraviolet/visible, and nuclear magnetic resonance (NMR) spectroscopy. All spectra were consistent with those

expected for the structure or with the literature spectra, as shown in Figures I1 and I2.

The purity of both lots was determined by elemental analysis, Karl Fischer water analysis, thin-layer chromatography (TLC),

and high-performance liquid chromatography (HPLC). TLC was performed with two systems: A) aluminum oxide F-254

Type E plates using a solvent system of hexanes:diethylether (50:50) and B) Whatman RP KC18 F-254 plates using methanol

as the solvent. Visualization was accomplished with visible light and shortwave (254 nm) ultraviolet light for both lots;

longwave (366 nm) ultraviolet light was also used for visualization of lot M061583. HPLC was performed with a Waters

:Bondapak C18 column with ultraviolet detection at 254 nm using a solvent system of water: tetrahydrofuran (65:35 for lot

1076-C and 61:39 for lot M061583) and a flow rate of 1.0 mL/minute.

For lot 1076-C, elemental analyses for hydrogen and nitrogen were in agreement with theoretical values; the elemental

analysis for carbon was high and the analysis for bromine was low. Karl Fischer water analysis indicated 0.21% ± 0.05%

water. TLC indicated a major spot and two slight trace impurities by system A and a major spot and one trace impurity by

system B. HPLC indicated a major peak and six impurities, with a total impurity area of approximately 20% relative to the

major peak area. 

The impurities in lot 1076-C were further identified and quantified using HPLC and direct-inlet mass spectrometry. A filtered

solution of 0.57 mg/mL 1-amino-2,4-dibromoanthraquinone in N,N-dimethylformamide was analyzed by HPLC with a Waters

:Bondapak C18 column with a solvent system of water:acetonitrile (52:48) at a flow rate of 1.0 mL/minute. A major peak

and eight impurities with areas of 0.1% or larger were detected by ultraviolet light (254 nm). No additional impurities were

observed when the ratio of the solvent was increased linearly from 48% to 100%. Three impurities had areas larger than 1%

of the total peak area. Aliquots of 5.0 mg/mL 1-amino-2,4-dibromoanthraquinone in N,N-dimethylformamide were analyzed

using the same HPLC system as was used in the impurity profile analysis, but with a solvent ratio of 53:47. The fractions

were then evaporated to dryness, reconstituted in 5 mL n-hexane, and concentrated under purified nitrogen to a volume of

200 :L. The samples were then analyzed using direct-inlet mass spectrometry. The three impurities with areas greater than

1% of the major peak area were identified as anthraquinone, a monoamino-monobromoanthraquinone, and an isomer of the

major component. Further analysis of the second and third of these impurities using proton Fourier transform NMR

spectroscopy indicated that the structure of the second impurity, a monoamino-monobromoanthraquinone, was consistent with

that of 1-amino-2-bromoanthraquinone, and that the isomer was probably 2-amino-1,3-dibromoanthraquinone. Quantitation

of the impurity identified as anthraquinone using HPLC with a Dupont Zorbax ODS column using a solvent system of water:

acetonitrile (30:70) and 0.3 mg/mL butyrophenone as an internal standard indicated approximately 5.0% anthraquinone. 

Quantitation of the impurities identified as 1-amino-2-bromoanthraquinone and an isomer of the major component by the

HPLC system described for the impurity profile analysis indicated approximately 4.3% 1-amino-2-bromoanthraquinone and

approximately 2.2% isomer. The overall purity of lot 1076-C was determined to be approximately 87%.

For lot M061583, elemental analyses for carbon, hydrogen, nitrogen, and bromine were in agreement with theoretical values. 

Karl Fischer water analysis indicated 0.32% ± 0.04% water. TLC indicated a major spot and one trace and one slight trace

impurity by system A and a major spot and one slight trace impurity by system B. HPLC indicated a major peak and six

impurities with the same retention times as found for lot 1076-C and a total impurity area of 3% relative to the major peak

area. Additionally, when the tetrahydrofuran in the solvent was increased to 60%, two impurity peaks with areas less than 1%

of the peak area were observed. A concomitant analysis of lot 1076-C with lot M061583 with the same high-performance

liquid chromatography system described for the purity analyses, but with a solvent ratio of 50:50 and with octanophenone as

an internal standard, indicated a 1-amino-2,4-dibromoanthraquinone concentration of approximately 112% in lot M061583

relative to lot 1076-C. The overall purity of lot M061583 was determined to be approximately 97%.

Stability studies were performed using the same HPLC system described for the purity determination of lot 1076-C but with a

solvent ratio of 55:45 and with octanophenone added as an internal standard. The studies indicated that 1-amino-2,4-
Chemical Characterization and Dose Formulations 345

dibromoanthraquinone, when stored protected from light, was stable as a bulk chemical for 2 weeks at temperatures up to

60° C. To ensure stability, the bulk chemical was stored in the dark at 4° ± 3° C throughout the studies. During the 2-year

studies, the stability of the bulk chemical was monitored periodically by the study laboratory using the same HPLC system; no

degradation of 1-amino-2,4-dibromoanthraquinone was seen throughout the studies.

PREPARATION AND ANALYSIS OF DOSE FORMULATIONS

The dose formulations were prepared by mixing appropriate quantities of 1-amino-2,4-dibromoanthraquinone with feed (Table

I1). Dosed feed formulations for chronic toxicity testing were made by preparing a 1-amino-2,4-dibromoanthraquinone/feed

premix by hand, which was then blended with plain feed in a Patterson-Kelly twin-shell blender for 15 minutes using an

intensifier bar. Dose formulations were prepared weekly.

Homogeneity and stability analyses of the dosed feed preparations were conducted by the analytical chemistry laboratory. For

the homogeneity analyses, the formulations were extracted with 100 mL of acetonitrile, centrifuged, and then further diluted

with acetonitrile. The absorbance of the samples was determined versus acetonitrile using ultraviolet spectroscopy at 249 nm. 

For the stability studies, 2,000 ppm feed samples were extracted with 100 mL of an acetonitrile hydrochloride solution

(999:1) and centrifuged; the extracts were then mixed with 0.5 mg/mL octanophenone (internal standard) in aceto-nitrile and

further diluted with acetonitrile. The samples were injected into an HPLC system equipped with a :Bondapak C18 column

and a 254 nm filtered detector, with a mobile phase of water:acetonitrile (57:43) at 1 mL/minute. Homogeneity of these

formulations was confirmed; stability was established for at least 2 weeks when dose formulations were stored in the dark at

temperatures up to 25° C.

Periodic analyses of the dose formulations of 1-amino-2,4-dibromoanthraquinone were conducted at the study laboratory and

at the analytical chemistry laboratory using ultraviolet/visible spectroscopy at 460 nm. For the 13-week studies, dose

formulations were analyzed at the beginning, midpoint, and end of the studies (Table I2). During the 2-year studies, the first

set of dose formulations and one randomly selected preparation every 8 weeks were analyzed (Table I3). All dose

formulations for rats and mice were within 10% of the target concentrations during the 13-week and 2-year studies. Results

of periodic referee analyses performed by the analytical chemistry laboratory indicated good agreement with the results

obtained by the study laboratory (Table I4).
346 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

FIGURE I1

Infrared Absorption Spectrum of 1-Amino-2,4-dibromoanthraquinone
Chemical Characterization and Dose Formulations 347

FIGURE I2

Nuclear Magnetic Resonance Spectrum of 1-Amino-2,4-dibromoanthraquinone
348 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE I1

Preparation and Storage of Dose Formulations in the Feed Studies of 1-Amino-2,4-dibromoanthraquinone

13-Week Studies 2-Year Studies

Preparation

A premix with feed and 1-amino-2,4-dibromoanthraquinone was

prepared with a mortar and pestle; premix and remainder of feed were

layered into a blender with a intensifier bar and mixed for 15 minutes

with the bar on. Doses were prepared weekly.

Same as 13-week studies

Chemical Lot Number

1076-C 1076-C and M061583

Maximum Storage Time

14 days 14 days

Storage Conditions

In double, clear plastic bags, in the dark, at 4° C In double, clear plastic bags, in the dark, at 4° ± 3° C

Study Laboratory

EG&G Mason Research Institute

(Worcester, MA)

EG&G Mason Research Institute

(Worcester, MA)

Referee Laboratory

Midwest Research Institute

(Kansas City, MO)

Midwest Research Institute

(Kansas City, MO)
Chemical Characterization and Dose Formulations 349

TABLE I2

Results of Analysis of Dose Formulations Administered to Rats and Mice in the 13-Week Feed Studies 

of 1-Amino-2,4-dibromoanthraquinone

Target Determined % Difference

Date Prepared Date Analyzed Concentration Concentrationa from Target

(ppm) (ppm)

14 April 1982 16 April 1982 2,500b 2,500 0

2,500 c 2,620 +5

2,500d 2,400 !4

5,000 4,780 !4

20 April 1982 10,000 10,200 +2

25,000 24,800 !1

50,000b 50,100 0

50,000 c 49,500 !1

50,000d 48,600 !3

9 June 1982 11 June 1982 2,500 2,500 0

5,000 4,690 !6

10,000 9,700 !3

25,000 25,000 0

50,000 50,400 +1

21 July 1982 22 July 1982 2,500 2,400 !4

5,000 4,670 !7

10,000 10,000 0

25,000 24,500 !2

50,000 50,200 0

a Results of duplicate analyses b Sample selection from top left of twin-shell blender c Sample selection from top right of twin-shell blender d Sample selection from bottom of twin-shell blender
350 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE I3

Results of Analysis of Dose Formulations Administered to Rats and Mice in the 2-Year Feed Studies 

of 1-Amino-2,4-dibromoanthraquinone

Target Determined % Difference

Date Prepared Date Analyzed Concentration Concentrationa from Target

(ppm) (ppm)

13 June 1983 14 June 1983b 10,000 9,920 !1

20,000 19,700 !2

6 July 1983 7 July 1983 2,000 1,980 !1

5,000 4,950 !1

10,000 9,760 !2

20,000 20,100 +1

15 August 1983 16 August 1983 2,000 1,990 !1

5,000 5,000 0

10,000 9,900 !1

20,000 19,600 !2

17 October 1983 19 October 1983 2,000 1,910 !5

5,000 4,920 !2

10,000 9,800 !2

20,000 20,000 0

19 December 1983 21 December 1983 2,000 1,980 !1

5,000 4,870 !3

10,000 9,800 !2

20,000 20,700 +4

21 February 1984 22 February 1984 2,000 1,960 !2

5,000 4,780 !4

10,000 10,100 +1

20,000 19,900 !1

9 April 1984 10 April 1984 2,000 1,900 !5

5,000 4,980 0

10,000 9,800 !2

20,000 19,400 !3

29 May 1984 30 May 1984 2,000 1,970 !2

5,000 4,870 !3

10,000 10,100 +1

20,000 20,000 0

9 July 1984 10 July 1984 2,000 1,880 !6

5,000 4,930 !1

10,000 10,000 0

20,000 20,100 +1

17 September 1984 18 September 1984 2,000 1,910 !5

5,000 5,120 +2

10,000 10,100 +1

20,000 19,800 !1
Chemical Characterization and Dose Formulations 351

TABLE I3

Results of Analysis of Dose Formulations Administered to Rats and Mice in the 2-Year Feed Studies 

of 1-Amino-2,4-dibromoanthraquinone (continued)

Target Determined % Difference

Date Prepared Date Analyzed Concentration Concentration from Target

(ppm) (ppm)

19 November 1984 20 November 1984 2,000 1,910 !5

5,000 4,900 !2

10,000 10,200 +2

20,000 19,900 !1

21 January 1985 23 January 1985 2,000 1,930 !4

5,000 4,970 !1

10,000 10,200 +2

20,000 20,100 +1

1 April 1985 2 April 1985 2,000 1,920 !4

5,000 5,150 +3

10,000 10,200 +2

20,000 20,200 +1

3 June 1985 4 June 1985 2,000 1,970 !2

5,000 4,830 !3

10,000 10,100 +1

20,000 19,500 !3

a Results of duplicate analyses b Doses mixed on this date were administered to mice only.
352 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE I4

Results of Referee Analysis of Dose Formulations in the 13-Week and 2-Year Feed Studies 

of 1-Amino-2,4-dibromoanthraquinone

Determined Concentration (ppm)

Target Concentration Study Referee

Date Prepared (ppm) Laboratorya Laboratoryb

13-Week Studies

14 April 1982 5,000 4,780 5,050 ± 130

2-Year Studies

13 June 1983 10,000 9,920 10,500 ± 100

19 December 1983 2,000 1,980 1,960 ± 20

9 April 1984 20,000 19,400 19,600 ± 100

17 September 1984 5,000 5,120 5,250 ± 110

1 April 1985 2,000 1,920 2,030 ± 40

a Results of duplicate analyses b Results of triplicate analyses (mean ± standard error)
353

APPENDIX J

FEED AND COMPOUND CONSUMPTION

IN THE 2-YEAR FEED STUDIES

OF 1-AMINO-2,4-DIBROMOANTHRAQUINONE

TABLE J1 Feed and Compound Consumption by Male Rats in the Stop-Exposure 

Evaluation of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 354

TABLE J2 Feed and Compound Consumption by Female Rats in the Stop-Exposure 

Evaluation of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 355

TABLE J3 Feed and Compound Consumption by Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 356

TABLE J4 Feed and Compound Consumption by Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 357

TABLE J5 Feed and Compound Consumption by Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 358

TABLE J6 Feed and Compound Consumption by Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 359
354 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE J1

Feed and Compound Consumption by Male Rats in the Stop-Exposure Evaluation 

of 1-Amino-2,4-dibromoanthraquinone

20,000 ppm (9-month stop-exposure) 20,000 ppm (15-month exposure)

Feed Body Dose/ Feed Body Dose/

(g/day)a Weight Dayb

(g/day) Weight Day

Week (g) (mg/kg) (g) (mg/kg)

1 11.2 130 1,718 10.9 136 1,605

2 15.1 144 2,096 15.9 149 2,136

4 22.8 169 2,700 20.0 173 2,303

5 17.7 169 2,095 17.0 177 1,922

8 20.3 211 1,924 20.8 217 1,919

12 20.2 267 1,511 18.5 277 1,336

13 17.8 267 1,328 17.7 271 1,308

14 19.6 284 1,381 18.9 292 1,292

17 16.9 305 1,110 17.8 318 1,117

21 17.9 327 1,092 18.9 334 1,133

25 14.1 341 827 14.5 351 829

29 15.8 338 932 15.9 355 898

33 16.8 351 957 15.4 364 844

37 18.0 352 1,019 18.9 364 1,038

41c 17.6 374 0 15.6 377 825

45 16.0 384 0 15.2 381 799

49 17.3 395 0 15.4 383 805

53 15.5 398 0 16.3 389 840

57 15.3 393 0 15.7 377 833

61 14.9 379 0 15.1 370 817

65 15.3 364 841

Mean for weeks

1-13 17.9 194 1,910 17.3 200 1,790

14-40 17.0 328 1,045 17.1 340 1,022

41-61 or 65 16.1 387 0 15.5 377 823

a Grams of feed consumed per animal per day b Milligrams of compound consumed per kilogram body weight per day c 9-Month stop-exposure animals switched to undosed feed
Feed and Compound Consumption 355

TABLE J2

Feed and Compound Consumption by Female Rats in the Stop-Exposure Evaluation

of 1-Amino-2,4-dibromoanthraquinone

20,000 ppm (9-month stop-exposure) 20,000 ppm (15-month exposure)

Feed Body Dose/ Feed Body Dose/

(g/day)a Weight Dayb

(g/day) Weight Day

Week (g) (mg/kg) (g) (mg/kg)

1 8.1 90 1,810 7.3 94 1,543

2 10.6 93 2,259 10.0 97 2,079

4 14.6 113 2,591 14.6 114 2,563

5 13.2 117 2,240 13.8 121 2,296

8 14.0 141 1,982 14.7 150 1,955

9 12.9 146 1,764 11.6 157 1,474

12 15.1 168 1,794 12.8 171 1,497

13 14.6 172 1,688 14.2 175 1,628

14 15.0 177 1,694 15.8 179 1,764

17 18.8 188 2,000 16.8 190 1,770

21 17.1 194 1,771 16.4 196 1,666

25 14.5 198 1,462 12.9 200 1,290

29 16.5 200 1,653 14.9 203 1,469

33 21.0 201 2,083 21.0 203 2,071

37 10.9 207 1,052 11.0 211 1,048

41c

 9.7 207 0 10.9 210 1,040

49 12.1 231 0 10.2 213 959

53 11.0 248 0 11.6 219 1,062

57 11.9 252 0 11.2 220 1,022

61 11.1 258 0 10.3 217 953

65 10.9 220 992

Mean for weeks

1-13 12.9 130 2,016 12.4 135 1,879

14-40 16.3 195 1,673 15.5 197 1,582

41-61 or 65 11.2 239 0 10.9 217 1,005

a Grams of feed consumed per animal per day b Milligrams of compound consumed per kilogram body weight per day c 9-Month stop-exposure animals switched to undosed feed
356 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE J3

Feed and Compound Consumption by Male Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Feed Body Feed Body Dose/ Feed Body Dose/ Feed Body Dose/

(g/day)a Weight (g/day) Weight Dayb (g/day) Weight Day (g/day) Weight Day

Week (g) (g) (mg/kg) (g) (mg/kg) (g) (mg/kg)

1 20.1 139 16.2 136 238 15.0 136 549 12.6 134 937

2 17.2 161 16.6 165 202 16.5 164 504 14.9 155 961

4 21.4 235 20.3 225 180 20.7 222 466 19.3 204 943

5 17.5 240 16.9 233 145 17.0 230 369 15.9 215 742

8 22.5 302 20.7 285 145 21.3 280 380 20.0 260 769

12 18.5 338 18.8 323 116 20.2 318 317 16.9 300 563

13 17.9 332 17.8 321 111 17.8 309 288 17.6 298 589

14 22.4 356 19.9 339 117 18.5 330 281 17.7 314 561

17 22.9 387 20.1 363 111 19.3 350 275 19.7 335 589

21 20.9 406 20.8 382 109 18.7 368 254 17.3 356 486

25 19.0 423 18.0 398 91 15.8 383 206 15.8 369 427

29 20.8 435 18.5 403 92 17.9 384 232 19.5 376 520

33 19.9 445 17.6 413 85 16.8 398 210 16.9 383 441

37 21.8 453 21.6 420 103 21.9 401 273 19.9 388 514

41 17.1 468 16.7 433 77 16.2 415 196 15.5 397 392

45 16.2 473 16.0 440 73 16.1 424 190 15.7 403 389

49 15.5 479 17.4 452 77 16.7 428 195 15.6 409 382

53 16.7 489 17.8 457 78 17.7 439 201 16.6 415 399

57 17.4 486 16.9 453 75 16.8 430 195 15.9 409 389

61 16.3 484 15.3 445 69 15.4 430 179 14.3 405 352

65 15.8 484 16.8 448 75 15.5 426 182 16.5 408 405

69 15.6 479 15.1 442 68 15.7 417 188 14.6 398 368

72 16.0 472 15.1 429 70 14.8 407 182 15.8 389 407

77 15.1 460 13.8 412 67 14.3 392 183 15.2 381 400

80 15.3 462 14.9 418 71 14.5 390 187 14.7 373 393

85 14.3 467 14.8 419 71 14.2 388 183 13.9 365 382

89 13.8 455 13.8 405 68 14.7 378 194 14.8 357 414

93 14.4 445 14.9 396 75 14.6 363 202 16.0 347 462

97 12.9 432 15.7 380 82 16.4 355 231 16.9 332 508

101 14.5 422 16.2 380 85 15.6 343 227 17.0 301 565

103 14.7 406 14.7 349 84 16.6 341 244 18.6 283 656

Mean for weeks 

1-13 19.3 249 18.2 241 163 18.4 237 411 16.7 224 786

14-52 19.6 433 18.7 404 93 17.8 388 231 17.4 373 470

53-103 15.2 460 15.4 417 74 15.5 393 198 15.8 369 436

a Grams of feed consumed per animal per day b Milligrams of compound consumed per kilogram body weight per day 
Feed and Compound Consumption 357

TABLE J4

Feed and Compound Consumption by Female Rats in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

0 ppm 2,000 ppm 5,000 ppm 10,000 ppm

Feed Body Feed Body Dose/ Feed Body Dose/ Feed Body Dose/

(g/day)a Weight (g/day) Weight Dayb (g/day) Weight Day (g/day) Weight Day

Week (g) (g) (mg/kg) (g) (mg/kg) (g) (mg/kg)

1 12.4 93 10.2 93 219 9.3 94 493 8.5 94 911

2 14.6 114 11.8 110 216 12.2 107 571 11.1 102 1,091

4 15.5 147 14.0 141 198 14.5 136 534 14.3 128 1,119

5 15.7 157 14.5 151 192 15.0 146 514 13.4 137 984

8 16.4 180 15.2 174 175 18.4 167 552 13.9 158 882

9 16.9 186 11.8 178 133 12.5 173 362 12.5 167 749

12 17.0 203 13.3 194 137 13.3 188 352 13.5 181 747

13 19.2 208 14.6 195 149 15.2 192 396 14.0 185 758

14 19.2 212 14.8 201 147 14.6 196 372 14.5 189 767

17 20.0 222 14.7 209 141 16.7 205 409 16.9 200 846

21 17.1 228 15.0 216 139 14.7 213 346 14.8 205 723

25 15.3 237 11.0 219 100 12.1 216 280 11.3 208 545

29 18.6 246 13.7 223 123 13.7 219 312 13.1 212 619

33 19.8 251 18.7 227 165 19.3 222 434 20.1 213 943

37 10.4 258 10.3 233 89 12.9 230 282 10.7 218 492

41 9.8 265 9.6 233 82 9.1 227 201 9.2 217 425

45 10.6 272 10.4 239 87 10.0 231 216 10.2 221 461

49 11.8 284 11.4 246 93 10.8 237 228 10.7 227 474

53 11.0 299 11.3 257 88 10.7 245 218 10.8 232 467

57 12.4 311 11.5 264 88 11.0 249 222 10.9 238 457

61 12.0 315 12.0 267 90 11.4 251 228 10.9 238 456

65 12.6 328 11.6 276 84 11.6 257 226 10.9 242 452

68 12.0 333 11.8 277 85 11.5 257 224 11.4 241 471

73 12.4 343 12.4 286 87 12.1 263 230 11.8 245 481

77 11.7 347 11.2 289 78 11.2 269 208 10.9 243 449

81 11.9 351 11.7 296 79 11.2 270 206 10.9 239 457

85 11.7 354 11.2 295 76 10.8 268 201 10.8 234 463

89 12.3 354 11.8 298 79 11.2 262 213 11.5 229 500

93 11.6 356 10.9 299 73 10.4 251 208 11.6 224 518

97 11.9 358 12.5 298 84 11.7 250 233 11.6 213 543

100 11.7 361 12.0 293 82 11.5 243 237 12.3 202 607

103 12.3 362 12.6 290 87 11.6 234 249 12.9 194 663

Mean for weeks

1-13 16.0 161 13.2 154 177 13.8 150 472 12.7 144 905

14-52 15.3 248 13.0 225 117 13.4 219 308 13.2 211 629

53-103 12.0 341 11.8 285 83 11.3 255 222 11.4 230 499

a Grams of feed consumed per animal per day b Milligrams of compound consumed per kilogram body weight per day 
358 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE J5

Feed and Compound Consumption by Male Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

0 ppm 10,000 ppm 20,000 ppm

Feed Body Feed Body Dose/ Feed Body Dose/

(g/day)a Weight (g/day) Weight Dayb(g/day) Weight Day

Week (g) (g) (mg/kg) (g) (mg/kg)

1 6.6 21.2 6,260

2 5.8 23.5 5.9 23.2 2,525 6.0 23.9 5,000

5 6.0 27.3 6.2 26.9 2,297 6.0 27.3 4,380

9 6.2 30.2 5.8 30.1 1,916 5.4 30.5 3,561

13 5.7 33.7 5.6 32.5 1,732 5.5 33.0 3,326

17 5.9 36.3 5.8 34.4 1,689 5.7 34.4 3,339

21 5.8 38.8 6.4 34.0 1,890 5.7 34.6 3,292

25 6.0 39.2 6.1 37.0 1,653 6.1 35.6 3,445

29 5.5 39.9 5.8 35.6 1,617 5.5 36.2 3,064

33 5.7 41.3 5.8 38.5 1,496 5.0 36.4 2,757

37 5.3 42.1 5.1 39.6 1,281 5.1 37.9 2,697

41 5.3 42.8 4.9 39.8 1,242 5.2 38.2 2,703

45 5.9 42.1 5.9 39.5 1,492 5.8 37.4 3,097

49 5.5 43.5 5.5 40.1 1,373 6.1 38.9 3,119

53 6.2 44.4 5.1 41.8 1,215 5.5 39.6 2,792

57 5.4 44.0 5.4 41.6 1,294 5.1 39.3 2,630

61 6.6 44.8 5.8 42.3 1,383 6.4 40.2 3,165

65 6.8 45.2 5.5 41.6 1,323 6.0 39.4 3,046

69 6.5 43.6 6.1 40.7 1,501 6.5 37.7 3,468

73 4.1 42.1 6.4 39.8 1,600 6.0 39.2 3,052

77 6.3 44.2 6.0 40.0 1,507 6.3 38.7 3,257

81 5.1 43.0 6.0 39.5 1,530 6.1 38.4 3,200

85 5.6 44.1 6.2 38.9 1,584 6.3 38.0 3,306

89 6.2 44.0 6.4 37.7 1,694 6.7 37.1 3,600

93 4.7 42.3 6.3 38.1 1,656 5.8 36.2 3,191

97 7.8 43.8 9.4 37.4 2,505 9.5 36.8 5,179

101 8.3 43.7 8.9 37.4 2,394 8.5 36.7 4,668

104 9.1 43.6 11.2 37.0 3,016 9.6 36.3 5,315

Mean for weeks

1-13 5.9 28.7 5.9 28.2 2,117 5.9 27.2 4,506

14-52 5.6 40.7 5.7 37.6 1,526 5.6 36.6 3,057

53-104 6.3 43.8 6.7 39.6 1,702 6.7 38.1 3,519

a Grams of feed consumed per animal per day 

b Milligrams of compound consumed per kilogram body weight per day 
Feed and Compound Consumption 359

TABLE J6

Feed and Compound Consumption by Female Mice in the 2-Year Feed Study 

of 1-Amino-2,4-dibromoanthraquinone

0 ppm 10,000 ppm 20,000 ppm

Feed Body Feed Body Dose/ Feed Body Dose/

(g/day)a Weight (g/day) Weight Dayb(g/day) Weight Day

Week (g) (g) (mg/kg) (g) (mg/kg)

2 5.4 18.1 4.8 18.4 2,628 4.8 18.0 5,385

5 6.4 20.8 5.5 21.3 2,597 5.7 20.8 5,487

9 5.9 22.9 5.9 22.9 2,579 5.1 23.3 4,402

13 6.0 24.8 5.0 24.8 2,023 6.7 25.0 5,339

17 5.8 26.0 5.8 26.6 2,190 6.2 26.0 4,789

21 5.6 27.7 5.1 27.3 1,866 5.7 27.3 4,189

25 6.0 29.2 5.6 28.6 1,941 6.3 28.1 4,507

29 6.4 30.2 5.8 28.8 2,002 6.8 28.9 4,737

33 6.0 32.1 5.7 30.7 1,854 5.3 28.8 3,681

37 6.5 32.9 5.5 31.9 1,726 6.1 30.5 4,029

41 6.7 34.2 6.3 33.0 1,910 6.6 30.1 4,365

45 5.9 35.2 5.5 32.8 1,687 5.6 31.2 3,588

49 6.8 36.6 6.4 34.5 1,844 6.7 32.5 4,124

53 5.3 36.8 4.9 35.0 1,389 5.2 32.5 3,190

56 6.7 38.2 6.8 36.3 1,881 6.7 33.1 4,086

61 6.9 39.1 6.2 36.2 1,704 6.4 33.8 3,796

65 6.7 38.4 6.2 36.1 1,722 6.3 33.5 3,754

69 6.6 37.6 6.4 35.4 1,798 6.6 32.4 4,075

73 5.1 37.3 7.4 35.7 2,062 7.2 32.7 4,374

77 5.1 39.3 5.7 35.8 1,581 5.7 33.1 3,427

81 5.5 39.9 5.7 36.4 1,575 6.1 33.8 3,597

85 6.3 39.5 6.4 36.7 1,754 6.6 33.6 3,905

89 7.0 41.1 7.2 36.1 1,985 7.3 33.7 4,347

93 7.8 41.4 7.5 35.3 2,137 8.5 33.3 5,121

97 8.3 40.0 6.8 33.6 2,019 7.8 31.9 4,902

101 8.6 38.5 9.6 34.1 2,833 10.2 31.5 6,521

104 9.1 39.2 10.2 33.6 3,021 11.1 31.9 6,985

Mean for weeks

1-13 5.9 21.7 5.3 21.9 2,457 5.6 21.8 5,153

14-52 6.2 31.6 5.5 30.5 1,798 6.2 29.3 4,223

53-104 6.8 39.0 6.8 35.5 1,935 7.2 32.9 4,415

a Grams of feed consumed per animal per day 

b Milligrams of compound consumed per kilogram body weight per day 
360 1-Amino-2,4-dibromoanthraquinone, NTP TR 383
361

APPENDIX K

INGREDIENTS, NUTRIENT COMPOSITION,

AND CONTAMINANT LEVELS

IN NIH-07 RAT AND MOUSE RATION

TABLE K1 Ingredients of NIH-07 Rat and Mouse Ration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 362

TABLE K2 Vitamins and Minerals in NIH-07 Rat and Mouse Ration . . . . . . . . . . . . . . . . . . . . . . . . . 362

TABLE K3 Nutrient Composition of NIH-07 Rat and Mouse Ration . . . . . . . . . . . . . . . . . . . . . . . . . . 363

TABLE K4 Contaminant Levels in NIH-07 Rat and Mouse Ration . . . . . . . . . . . . . . . . . . . . . . . . . . . 364
362 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE K1

Ingredients of NIH-07 Rat and Mouse Rationa

Ingredientsb Percent by Weight

Ground #2 yellow shelled corn 24.50

Ground hard winter wheat 23.00

Soybean meal (49% protein) 12.00

Fish meal (60% protein) 10.00

Wheat middlings 10.00

Dried skim milk 5.00

Alfalfa meal (dehydrated, 17% protein) 4.00

Corn gluten meal (60% protein) 3.00

Soy oil 2.50

Dried brewer's yeast 2.00

Dry molasses 1.50

Dicalcium phosphate 1.25

Ground limestone 0.50

Salt 0.50

Premixes (vitamin and mineral) 0.25

a NCI, 1976; NIH, 1978 b Ingredients were ground to pass through a U.S. Standard Screen No. 16 before being mixed.

TABLE K2

Vitamins and Minerals in NIH-07 Rat and Mouse Rationa

Amount Source

Vitamins

A 5,500,000 IU Stabilized vitamin A palmitate or acetate

D3 4,600,000 IU D-activated animal sterol

K3 2.8 g Menadione

d-"-Tocopheryl acetate 20,000 IU

Choline 560.0 g Choline chloride

Folic acid 2.2 g

Niacin 30.0 g

d-Pantothenic acid 18.0 g d-Calcium pantothenate

Riboflavin 3.4 g

Thiamine 10.0 g Thiamine mononitrate

B12 4,000 :g

Pyridoxine 1.7 g Pyridoxine hydrochloride

Biotin 140.0 mg d-Biotin

Minerals

Iron 120.0 g Iron sulfate

Manganese 60.0 g Manganous oxide

Zinc 16.0 g Zinc oxide

Copper 4.0 g Copper sulfate

Iodine 1.4 g Calcium iodate

Cobalt 0.4 g Cobalt carbonate

a Per ton (2,000 lb) of finished product
Feed Analyses 363

TABLE K3

Nutrient Composition of NIH-07 Rat and Mouse Ration

Mean ± Standard

Nutrient Deviation Range Number of Samples

Protein (% by weight) 22.29 ± 0.86 21.00 ) 24.30 22

Crude fat (% by weight) 5.38 ± 0.59 4.40 ) 6.30 22

Crude fiber (% by weight) 3.72 ± 0.47 3.10 ) 5.40 22

Ash (% by weight) 6.67 ± 0.29 5.96 ) 7.27 22

Amino Acids (% of total diet)

Arginine 1.308 ± 0.060 1.210 ) 1.390 8

Cystine 0.306 ± 0.084 0.181 ) 0.400 8

Glycine 1.150 ± 0.047 1.060 ) 1.210 8

Histidine 0.576 ± 0.024 0.531 ) 0.607 8

Isoleucine 0.917 ± 0.029 0.881 ) 0.944 8

Leucine 1.946 ± 0.055 1.850 ) 2.040 8

Lysine 1.270 ± 0.058 1.200 ) 1.370 8

Methionine 0.448 ± 0.128 0.306 ) 0.699 8

Phenylalanine 0.987 ± 0.140 0.665 ) 1.110 8

Threonine 0.877 ± 0.042 0.824 ) 0.940 8

Tryptophan 0.236 ± 0.176 0.107 ) 0.671 8

Tyrosine 0.676 ± 0.105 0.564 ) 0.794 8

Valine 1.103 ± 0.040 1.050 ) 1.170 8

Essential Fatty Acids (% of total diet)

Linoleic 2.393 ± 0.258 1.830 ) 2.570 7

Linolenic 0.280 ± 0.040 0.210 ) 0.320 7

Vitamins

Vitamin A (IU/kg) 10,459 ± 4,285 4,200 ) 17,800 22

Vitamin D (IU/kg) 4,450 ± 1,382 3,000 ) 6,300 4

")Tocopherol (ppm) 37.95 ± 9.406 22.5 ) 48.90 8

Thiamine (ppm) 20.73 ± 5.08 12.0 ) 37.0 22

Riboflavin (ppm) 7.92 ± 0.87 6.10 ) 9.00 8

Niacin (ppm) 103.38 ± 26.59 65.0 ) 150.0 8

Pantothenic acid (ppm) 29.54 ± 3.60 23.0 ) 34.0 8

Pyridoxine (ppm) 9.55 ± 3.48 5.60 ) 14.0 8

Folic acid (ppm) 2.25 ± 0.73 1.80 ) 3.70 8

Biotin (ppm) 0.25 ± 0.04 0.19 ) 0.32 8

Vitamin B12 (ppb) 38.45 ± 22.01 10.6 ) 65.0 8

Choline (ppm) 3,089 ± 328 2,400 ) 3,430 8

Minerals

Calcium (%) 1.20 ± 0.14 0.91 ) 1.43 22

Phosphorus (%) 0.94 ± 0.06 0.84 ) 1.10 22

Potassium (%) 0.883 ± 0.078 0.772 ) 0.971 6

Chloride (%) 0.526 ± 0.092 0.380 ) 0.635 8

Sodium (%) 0.313 ± 0.390 0.258 ) 0.371 8

Magnesium (%) 0.168 ± 0.010 0.151 ) 0.181 8

Sulfur (%) 0.280 ± 0.064 0.208 ) 0.420 8

Iron (ppm) 360.54 ± 100 255.0 ) 523.0 8

Manganese (ppm) 91.97 ± 6.01 81.70 ) 99.40 8

Zinc (ppm) 54.72 ± 5.67 46.10 ) 64.50 8

Copper (ppm) 11.06 ± 2.50 8.09 ) 15.39 8

Iodine (ppm) 3.37 ± 0.92 1.52 ) 4.13 6

Chromium (ppm) 1.79 ± 0.36 1.04 ) 2.09 8

Cobalt (ppm) 0.681 ± 0.14 0.490 ) 0.780 4
364 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE K4

Contaminant Levels in NIH-07 Rat and Mouse Rationa

Mean ± Standard

Deviationb Range Number of Samples

Contaminants

Arsenic (ppm) 0.56 ± 0.19 0.18 ) 0.80 22

Cadmium (ppm)c 0.11 ± 0.03 0.10 ) 0.20 22

Lead (ppm) 0.57 ± 0.19 0.24 ) 1.00 22

Mercury (ppm) <0.05 22

Selenium (ppm) 0.33 ± 0.06 0.23 ) 0.45 22

Aflatoxins (ppb) <5.00 22

Nitrate nitrogen (ppm)d 11.57 ± 5.85 2.50 ) 22.0 22

Nitrite nitrogen (ppm)d 0.69 ± 1.42 <0.10 ) 6.10 22

BHA (ppm)e <2 22

BHT (ppm)e 2.36 ± 1.00 <1.00 ) 4.00 22

Aerobic plate count (CFU/g) 144,259 ± 157,664 6,200 ) 443,800 22

Coliform (MPN/g) 317 ± 567 <3.00 ) 2,400 22

Escherichia coli (MPN/g)f 9.73 ± 31.33 <3.00 ) 150 22

Escherichia coli (MPN/g)g 3.04 ± 0.22 <3.00 ) 4.0 21

Total nitrosoamines (ppb)h 6.44 ± 6.26 0.80 ) 30.30 22

N)Nitrosodimethylamine (ppb)h 5.91 ± 6.21 0.50 ) 30.00 22

N)Nitrosopyrrolidine (ppb)h 0.53 ± 0.58 0.30 ) 2.70 22

Pesticides (ppm)

")BHC <0.01 22

$)BHC <0.02 22

()BHC <0.01 22

*)BHC <0.01 22

Heptachlor <0.01 22

Aldrin <0.01 22

Heptachlor epoxide <0.01 22

DDE <0.01 22

DDD <0.01 22

DDT <0.01 22

HCB <0.01 22

Mirex <0.01 22

Methoxychlor <0.05 22

Dieldrin <0.01 22

Endrin <0.01 22

Telodrin <0.01 22

Chlordane <0.05 22

Toxaphene <0.1 22

Estimated PCBs <0.2 22

Ronnel <0.01 22

Ethion <0.02 22

Trithion <0.05 22

Diazinon <0.1 22

Methyl parathion <0.02 22

Ethyl parathion <0.02 22

Malathioni

 0.32 ± 0.68 0.05 ) 3.20 22

Endosulfan I <0.01 22

Endosulfan II <0.01 22

Endosulfan sulfate <0.03 22
Feed Analyses 365

TABLE K4

Contaminant Levels in NIH-07 Rat and Mouse Ration (continued)

a CFU = colony forming units, MPN = most probable number, BHC is hexachlorocyclohexane or benzene hexachloride b For values less than the limit of detection, the detection limit is given as the mean. c Three lots milled 22 February 1984, 14 March 1984, and 9 May 1984 contained 0.20 ppm. All other lots measured less than or equal to the detection

limit. d Sources of contamination: alfalfa, grains, and fish meal e Sources of contamination: soy oil and fish meal f Mean, standard deviation, and range include one high value of 150 MPN/g from the lot milled on 17 October 1984. g Mean, standard deviation, and range exclude the high value of 150 MPN/g from the lot milled on 17 October 1984. h All values were corrected for percent recovery. i Mean, standard deviation, and range include one high value of 3.20 ppm.
366 1-Amino-2,4-dibromoanthraquinone, NTP TR 383
367

APPENDIX L

SENTINEL ANIMAL PROGRAM 

METHODS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 368

TABLE L1 Murine Virus Antibody Determinations for Rats in the 13-Week

and 2-Year Feed Studies of 1-Amino-2,4-dibromoanthraquinone . . . . . . . . . . . . . . . . . . . . 370
368 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

SENTINEL ANIMAL PROGRAM

METHODS

Rodents used in the Carcinogenesis Program of the National Toxicology Program are produced in optimally clean facilities to

eliminate potential pathogens that may affect study results. The Sentinel Animal Program is part of the periodic monitoring of

animal health that occurs during the toxicologic evaluation of chemical compounds. Under this program, the disease state of

the rodents is monitored via serology on sera from extra (sentinel) animals in the study rooms. These animals and the study

animals are all subject to identical environmental conditions. The sentinel animals come from the same production source and

weanling groups as the animals used for the studies of chemical compounds.

Serum samples were collected from randomly selected rats and mice during the 13-week and 2-year studies. Blood from each

animal was collected, allowed to clot, and the serum separated. The samples were processed appropriately and sent to

Microbiological Associates, Inc. (Bethesda, MD) for determination of antibody titers. The laboratory serology methods and

viral agents for which testing was performed are tabulated below; the times at which blood was collected during the studies are

also listed.

Method of Analysis Time of Analysis

Rats

13-Week Study

Complement Fixation

RCV (rat coronavirus) Study termination

Sendai Study termination

Hemagglutination Inhibition

H-1 (Toolan's H-1 virus) Study termination

KRV (Kilham rat virus) Study termination

PVM (pneumonia virus of mice) Study termination

2-Year Study

ELISA

Mycoplasma arthritidis 18 and 24 months

Mycoplasma pulmonis 18 and 24 months

PVM 18 and 24 months

RCV/SDA (rat coronavirus/

 sialodacryoadenitis virus) 6, 12, 18, and 24 months

Sendai 18 and 24 months

Hemagglutination Inhibition

H-1

6, 12, 18, and 24 months

KRV 6, 12, 14, 15, 18, and 24 months

PVM 6 and 12 months

Sendai 6 and 12 months

 
Sentinel Animal Program 369

Mice

13-Week Study

Complement Fixation

LCM (lymphocytic choriomeningitis virus) Study termination

Mouse adenoma virus Study termination

Sendai Study termination

ELISA

MHV (mouse hepatitis virus) Study termination

Hemagglutination Inhibition

Ectromelia virus Study termination

GDVII (mouse encephalomyelitis virus) Study termination

MVM (minute virus of mice) Study termination

PVM Study termination

Polyoma virus Study termination

Reovirus 3 Study termination

2-Year Study

Complement Fixation

LCM 6, 12, 18, and 24 months

Mouse adenoma virus 6 and 12 months

ELISA

Ectromelia virus 18 and 24 months

GDVII 12, 18, and 24 months

Mouse adenoma virus 18 and 24 months

MHV 6, 12, 18, and 24 months

M. arthritidis 18 and 24 months

M. pulmonis 18 and 24 months

PVM 18 and 24 months

Reovirus 3 18 and 24 months

Sendai 18 and 24 months

Hemagglutination Inhibition

Ectromelia virus 6 and 12 months

GDVII 6 months

K (papovavirus) 18 and 24 months

MVM 6, 12, 18, and 24 months

PVM 6 and 12 months

Polyoma virus 6, 12, 18, and 24 months

Reovirus 3 6 and 12 months

Sendai 6 and 12 months

Immunofluorescence Assay

EDIM (epizootic diarrhea of infant mice) 18 and 24 months

Results of serology tests for rats are presented in Table L1. All test results for mice were negative.
370 1-Amino-2,4-dibromoanthraquinone, NTP TR 383

TABLE L1

Murine Virus Antibody Determinations for Rats in the 13-Week and 2-Year Feed Studies

of 1-Amino-2,4-dibromoanthraquinone

Interval Incidence of Antibody Positive Serologic

in Sentinel Animals Reaction for

13-Week Study

Study termination 0/10 None positive

2-Year Study

Male

6 Months 0/5 None positive

12 Months 0/5 None positive

18 Months 0/4 None positive

24 Months 0/5 None positive

Female

6 Months 5/5 Sendai

12 Months 1/5 KRV

5/5 Sendai

14 Months 1/5 KRV

15 Months 0/10 None positive

18 Months 1/5 KRV

5/5 Sendai

24 Months 5/5 Sendai