1978 

National Cancer Institute 

CARCINOGENESIS 

Technical Report Series 

No. 94 

BIOASSAY OF 

4-AMINO-2-NITROPHENOL 

FOR POSSIBLE CARCINOGENICITY 

CAS No. 119-34-6 

NCI-CG-TR-94 

U.S. DEPARTMENT OF HEALTH, EDUCATION, AND WELFARE 

Public Health Service 

National Institutes of Health 
BIOASSAY OF
 

4-AMIN0-2-NITROPHENOL
 

FOR POSSIBLE CARCINOGENICITY
 

Carcinogenesis Testing Program
 

Division of Cancer Cause and Prevention
 

National Cancer Institute
 

National Institutes of Health
 

Bethesda, Maryland 20014
 

U.S. DEPARTMENT OF HEALTH, EDUCATION, AND WELFARE
 

Public Health Service
 

National Institutes of Health
 

DHEW Publication No. (NIH) 78-1344
 

BIOASSAY OF
 

4-AMINO-2-NITROPHENOL
 

FOR POSSIBLE CARCINOGENICITY
 

Carcinogenesis Testing Program
 

Division of Cancer Cause and Prevention
 

National Cancer Institute
 

National Institutes of Health
 

FOREWORD; This report presents the results of the bioassay of
 

4-amino-2-nitrophenol conducted for the Carcinogenesis Testing
 

Program, Division of Cancer Cause and Prevention, National Cancer
 

Institute (NCI), National Institutes of Health, Bethesda,
 

Maryland. This is one of a series of experiments designed to
 

determine whether selected chemicals have the capacity to produce
 

cancer in animals. Negative results, in which the test animals
 

do not have a greater incidence of cancer than control animals,
 

do not necessarily mean that the test chemical is not a
 

carcinogen, inasmuch as the experiments are conducted under a
 

limited set of circumstances. Positive results demonstrate that
 

the test chemical is carcinogenic for animals under the
 

conditions of the test and indicate that exposure to the chemical
 

is a potential risk to man. The actual determination of the risk
 

to man from animal carcinogens requires a wider analysis.
 

CONTRIBUTORS: The bioassay of 4-amino-2-nitrophenol was
 

conducted by Litton Bionetics, Inc., Kensington, Maryland,
 

initially under direct contract to NCI and currently under a 

subcontract to Tracer Jitco, Inc., the prime contractor for the 

NCI Carcinogenesis Testing Program. 

The experimental design and doses were chosen by Drs. E. K.
 

Weisburger1, J. H. Weisburger1 >2 , and N. P. Page1'3, the NCI
 

project officers; Dr. F. M. Garner^ was the principal
 

investigator, and Mr. S. Johnson4

, co-principal investigator.
 

The administration of the test chemical and the observation of
 

the animals were supervised by Dr. Garner, and technical
 

assistance with the bioassay was provided by Mr. R. Cypher4

. Mr.
 

H. D. Thornett4, and Mr. D. J. Howard4. Ms. J. Blalock^ was
 

responsible for data assembly.
 

iii
 
Histopathologic examination was performed by Drs. P. K.
 

Hildebrandt, N. J. . F. M. Garner and Dr. B. C. Zook Dr. 

R. Montali4 reviewed the diagnoses and prepared the interpretive 

pathology narrative. 

Animal pathology tables and survival tables were compiled at EG&G
 

Mason Research Institute-'. The statistical analyses were
 

performed by Dr. J. R. Joiner", using methods selected for the
 

bioassay program by Dr. J. J. Gart . Chemicals used in this
 

bioassay were analyzed under the direction of Dr. E. Murrill",
 

dosed feed mixtures were analyzed by Mr. H. Paulin^, and the 

results of the analyses were reviewed by Dr. S. S. Olin". The 

chemical structure was supplied by NCI. 

This report was prepared at Tracer Jitco^ under the direction of
 

NCI. Those responsible for the report at Tracer Jitco were Dr.
 

L. A. Campbell, Director of the Bioassay Program; Dr. S. S. Olin,
 

Deputy Director for Science; Dr. J. F. Robens, toxicologist; Dr.
 

R. L. Schueler, pathologist; Dr. G. L. Miller, Ms. L. A. Waitz,
 

and Mr. W. D. Reichardt, bioscience writers; and Dr. E. W.
 

Gunberg, technical editor, assisted by Ms. Y. E. Presley and Ms.
 

P. J. Graboske.
 

The following other scientists at NCI were responsible for
 

evaluating the bioassay, interpreting the results, and reporting
 

the findings: Dr. Kenneth C. Chu, Dr. Cipriano Cueto, Jr., Dr. J.
 

Fielding Douglas, Dr. Dawn G. Goodman, Dr. Richard A. Griesemer,
 

Dr. Harry A. Milman, Dr. Thomas W. Orme, Dr. Robert A. Squire^,
 

Dr. Sherman Stinson, Dr. Jerrold M. Ward, and Dr. Carrie E.
 

Whitmire.
 

^Carcinogenesis Testing Program, Division of Cancer Cause and
 

Prevention, National Cancer Institute, National Institutes of
 

Health, Bethesda, Maryland.
 

with the Naylor Dana Institute for Disease Prevention,
 

American Health Foundation, Hammond House Road, Valhalla,
 

New York.
 

iv
 
-'Now with the Environmental Protection Agency, 401 M Street,
 

S.W., Washington, D.C.
 

^Litton Bionetics, Inc., 5516 Nicholson Lane, Kensington,
 

Maryland.
 

-"EG&G Mason Research Institute, 1530 East Jefferson Street,
 

Rockville, Maryland.
 

"Tracer Jitco, Inc., 1776 East Jefferson Street, Rockville,
 

Maryland.
 

'Mathematical Statistics and Applied Mathematics Section,
 

Biometry Branch, Field Studies and Statistics, Division of
 

Cancer Cause and Prevention, National Cancer Institute, National
 

Institutes of Health, Bethesda, Maryland.
 

"Midwest Research Institute, 425 Volker Boulevard, Kansas City,
 

Missouri.
 

"Now with the Division of Comparative Medicine, Johns Hopkins
 

University, School of Medicine, Traylor Building, Baltimore,
 

Maryland.
 

v
 

SUMMARY
 

A bioassay of 4-amino-2-nitrophenol for possible carcinogenicity
 

was conducted by administering the test chemical in feed to
 

Fischer 344 rats and B6C3F1 mice.
 

Groups of 50 rats and 50 mice of each sex were administered
 

4-amino-2-nitrophenol at one of two doses, either 1,250 or 2,500
 

ppm, for 103 weeks. Matched controls consisted of groups of 20
 

untreated rats and 20 untreated mice of each sex. All dosed and
 

matched-control groups of each species and sex were killed at 105
 

weeks.
 

Mean body weights of dosed rats of each sex were not appreciably
 

affected by administration of the 4-amino-2-nitrophenol, and mean
 

body weights of dosed mice of each sex were only slightly lower
 

than those of corresponding matched controls. Survival of
 

neither rats nor mice was affected by the test chemical, and
 

sufficient numbers of animals in dosed and control groups were at
 

risk for development of late-appearing tumors. Since both male
 

and female mice receiving 4-amino-2-nitrophenol had little or no
 

depression in mean weights and their survival was comparable to
 

that of controls, they may have been able to tolerate a higher
 

dose.
 

In rats, transitional-cell carcinomas of the urinary bladder
 

showed a dose-related trend in the males (P < 0.001) and occurred
 

at a significantly higher incidence (P = 0.018) in the high-dose
 

males than in the matched-control males (controls 0/15, low-dose
 

0/46, high-dose 11/39 [28%]). Carcinomas of the bladder also
 

occurred in one low-dose female and two high-dose females, but in
 

none of the control females. Transitional-cell papillomas of the
 

bladder occurred in two additional high-dose males, and
 

transitional-cell hyperplasia of the bladder occurred in four
 

additional high-dose males, but neither lesion occurred in
 

control males. No tumors of the bladder were found among 220
 

male and 220 female historical-control rats at this laboratory.
 

In mice, no tumors occurred in dosed groups of males or females
 

vii
 
at incidences that were significantly higher than those in the
 

corresponding matched-control groups.
 

Deposition of pigment occurred in the lamina propria of the small
 

intestine in at least 91% of the animals in the dosed groups of
 

rats and in at least 89% of the animals in the dosed groups of
 

mice, but in none of the control groups of either species.
 

It is concluded that under the conditions of the bioassay,
 

4-amino-2-nitrophenol was carcinogenic for male Fischer 344 rats,
 

inducing transitional-cell carcinomas of the urinary bladder; the
 

transitional-cell carcinomas of the urinary bladder observed in
 

three dosed female rats may also have been associated with
 

administration of the 4-amino-2-nitrophenol. The test chemical
 

was not carcinogenic for male or female B6C3F1 mice at the doses
 

tested.
 

viii
 
TABLE OF CONTENTS
 

Page
 

I. Introduction	 1
 

II. Materials and Methods	 3
 

A. Chemical	 3
 

B. Dietary Preparation	 4
 

C. Animals	 4
 

D. Animal Maintenance	 5
 

E. Subchronic Studies	 7
 

F. Chronic Studies	 8
 

G. Clinical and Pathologic Examinations	 8
 

H. Data Recording and Statistical Analyses	 12
 

III. Results - Rats	 17
 

A. Body Weights and Clinical Signs (Rats)	 17
 

B. Survival (Rats)	 17
 

C. Pathology (Rats)	 20
 

D. Statistical Analyses of Results (Rats)	 22
 

IV. Results - Mice	 25
 

A. Body Weights and Clinical Signs (Mice)	 25
 

B. Survival (Mice)	 25
 

C. Pathology (Mice)	 28
 

D. Statistical Analyses of Results (Mice)	 29
 

V. Discussion	 31
 

VI.	 Bibliography 35
 

APPENDIXES
 

Appendix A Summary of the Incidence of Neoplasms in
 

Table Al Summary of the Incidence of Neoplasms in
 

Rats Fed 4-Amino-2-Nitrophenol in the Diet 37
 

Male Rats Fed 4-Amino-2-Nitrophenol in the Diet 39
 

Table A2 Summary of the Incidence of Neoplasms in Female
 

Rats Fed 4-Amino-4-Nitrophenol in the Diet..' 43
 

ix
 
Page 

Appendix B Summary of the Incidence of Neoplasms in 

Mice Fed 4-Amino-2-Nitrophenol in the Diet 47 

Table Bl Summary of the Incidence of Neoplasms in 

Male Mice Fed 4-Amino-2-Nitrophenol in the Diet.... 49 

Table B2 Summary of the Incidence of Neoplasms in Female 

Mice Fed 4-Amino-2-Nitrophenol in the Diet 52 

Appendix C Summary of the Incidence of Nonneoplastic Lesions 

in Rats Fed 4-Amino-2-Nitrophenol in the Diet 57 

Table Cl Summary of the Incidence of Nonneoplastic 

Lesions in Male Rats Fed 4-Amino-2-Nitrophenol 

in the Diet 59 

Table C2 Summary of the Incidence of Nonneoplastic 

Lesions in Female Rats Fed 4-Amino-2-Nitrophenol 

in the Diet 64 

Appendix D Summary of the Incidence of Nonneoplastic 

Lesions in Mice Fed 4-Amino-2-Nitrophenol 

in the Diet 69 

Table Dl Summary of the Incidence of Nonneoplastic 

Lesions in Male Mice Fed 4-Amino-2-Nitrophenol 

in the Diet 71 

Table D2 Summary of the Incidence of Nonneoplastic 

Lesions in Female Mice Fed 4-Amino-2-Nitrophenol 

in the Diet 74 

Appendix E Analyses of the Incidence of Primary Tumors 

in Rats Administered 4-Amino-2-Nitrophenol 

in the Diet 79 

Table El Analyses of the Incidence of Primary Tumors in Male 

Rats Administered 4-Amino-2-Nitrophenol 

In the Diet 81 

Table E2 Analyses of the Incidence of Primary Tumors in Female 

Rats Administered 4-Amino-2-Nitrophenol 

in the Diet 86 
Page 

Appendix F Analyses of the Incidence of Primary Tumors 

in Mice Administered 4-Amino-2-Nitrophenol 

in the Diet 91 

Table Fl Analyses of the Incidence of Primary Tumors in Male 

Mice Administered 4-Amino-2-Nitrophenol 

in the Diet 93 

Table F2 Analyses of the Incidence of Primary Tumors in Female 

Mice Administered 4-Amino-2-Nitrophenol 

in the Diet 97 

TABLES 

Table 1 4-Amino-2-Nitrophenol Chronic 

Feeding Studies in Rats 9 

Table 2 4-Amino-2-Nitrophenol Chronic 

Feeding Studies in Mice 10 

FIGURES 

Figure 1 Growth Curves for Rats Administered 4-Amino-2­

Nitrophenol in the Diet 18 

Figure 2 Survival Curves for Rats Administered 4-Amino-2­

Nitrophenol in the Diet 19 

Figure 3 Growth Curves for Mice Administered 4-Amino-2­

Nitrophenol in the Diet 26 

Figure 4 Survival Curves for Mice Administered 4-Amino-2­

Nitrophenol in the Diet 27 

xi
 

I. INTRODUCTION
 

4-Amino-2-nitrophenol (CAS
 

119-34-6; NCI C03963) is used
 

as an industrial dye
 

intermediate, and as a
 

constituent of "semi- OH
 

permanent" hair dyes.
 4-AMINO-2-IMITROPHEIMOL
 

This compound is designated CI76555 by the Society of Dyers and
 

Colourists, who describe its industrial use as an oxidation base
 

(Oxidation Base 25) in applications involving furs. These bases
 

are applied in conjunction with an oxidizing agent, such as hydrogen peroxide, for the development of the color (Society of Dyers
 

and Colourists, 1971).
 

In contrast, in hair dyes for humans, 4-amino-2-nitrophenol is
 

formulated without an oxidizing agent. It is used in
 

concentrations estimated at 0.1-1.0% in the "semi-permanent" hair
 

dyes, which are applied as shampoos and remain on the hair for
 

20-40 minutes before they are rinsed out (Wall, 1972; Corbett and
 

Menkart, 1973; Burnett et al., 1976; FDA, 1977).
 

4-Amino-2-nitrophenol was one of a group of hair dye constituents
 

selected for the Carcinogenesis Testing Program because several
 

of these chemicals had been shown to be mutagenic in bacterial
 

test systems (work later published by Ames et al., 1975).
 

II. MATERIALS AND METHODS
 

A. Chemical
 

Two batches of 4-amino-2-nitrophenol were obtained from the
 

Aldrich Chemical Company, Milwaukee, Wisconsin. Lot No. 071137
 

was used in the subchronic studies; Lot No. 100737 was used in
 

the chronic studies.
 

The identity of both lots was established through elemental
 

analyses (C, H, N), and spectral data (ultraviolet, infrared, and
 

nuclear magnetic resonance). Lot No. 071137 had a purity of 99.0
 

+ 0.3% as determined by perchloric acid titration of the amine
 

function, and Lot No. 100737 had a purity of 99.6 + 0.3% by the
 

same method. Vapor-phase chromatography and thin-layer chromatography showed several trace impurities in Lot No. 071137 and one
 

impurity in Lot No. 100737. Lot No. 071137 contained 0.18 +
 

0.01% water, and Lot No. 100737, less than 0.2% water, as
 

determined by Karl Fischer analysis. Melting ranges were similar
 

for each lot (Lot No. 100737: 128-130°C [visual, capillary]; Lot
 

No. 671137: 126.5-129°C [visual, capillary]) and corresponded to
 

values in the literature (127-128°C) (Verkade et al., 1946).
 

After the completion of the bioassay, a sample from Lot No.
 

100737 was reanalyzed. The infrared spectrum of this lot was
 
identical with that obtained in the original analysis, and
 

perchloric acid titration indicated 98.7 + 0.2% purity.
 

The bulk chemical was stored at 4°C.
 

B. Dietary Preparation
 

A 6-kilogram diet was prepared three times per week for the rats
 

and two times per week for the mice. To obtain each dietary
 

concentration, the appropriate weight of the compound was mixed
 

with a small portion of Wayne® Lab Blox animal meal (Allied
 

Mills, Inc., Chicago, 111.) in a mortar. This premix was then
 

added to the remaining feed and blended for 15 minutes in a
 

Patterson-Kelly twin-shell blender equipped with an intensifier
 

bar. Dosed feed preparations were stored at 4°C for up to 1
 

week.
 

As a quality control measure, selected samples from 14 freshly
 

prepared diets were analyzed during the chronic studies. The
 

compound was extracted from feed with 0.1 N ammonium hydroxide in
 

1:1 water:methanol, diluted with methanol, centrifuged, and the
 

absorbance of the supernatant read at 445 nm. Concentrations
 

were found to be within 25% of theoretical concentrations.
 

C. Animals
 

Fischer 344 rats and B6C3F1 mice were obtained from the Frederick
 
Cancer Research Center, Frederick, Maryland, under a contract
 

with the Division of Cancer Treatment, NCI.
 

The animals were 28 days of age when received at the laboratory
 

and were quarantined for 2 weeks prior to the start of the
 

bioassay. Any animals with clinical signs of disease and any
 

runts were destroyed. The remaining animals were segregated into
 

equal weight groups and assigned to control or dosed groups in
 

such a way that the mean weights of animals in each cage within a
 

particular group were approximately the same.
 

D. Animal Maintenance
 

All animals were housed in temperature- and humidity-controlled
 

rooms. The temperature was maintained at 21-25°C and the
 

relative humidity at 45-55%. There were 15 changes of room air
 

per hour, and the incoming and exhaust air was filtered through
 

high efficiency particulate air (HEPA) filters. The animal rooms
 

were positively pressurized with respect to the exit hall and
 

negatively pressurized with respect to the entrance hall. Rooms
 

were illuminated with cool white fluorescent lighting for 8 hours
 

per day.
 

Rats were housed four per cage and mice five per cage in solid
 

polycarbonate cages (Lab Products, Inc., Garfield, N. J.). Each
 

cage was covered with a wire mesh screen and a sheet of filter
 
paper, and contained a heat-treated hardwood chip bedding
 

(Absorb-Dri®, Lab Products, Inc., Garfield, N. J.) in the bottom.
 

Cages were washed and furnished with fresh bedding two times per
 

week. Water bottles, sipper tubes and stoppers were also washed
 

twice per week while feed hoppers were washed once per week. All
 

of this equipment was cleaned at 82°C with detergent, rinsed, and
 

steamed.
 

Control animals were fed Wayne® Lab Blox animal meal, and dosed
 

animals received the same product mixed with the test chemical.
 

The feed hoppers were filled three times per week. Acidified tap
 

water (pH 2.5) was available ad libitum in water bottles.
 

Rats and mice were housed in separate rooms. Control and dosed
 

animals were housed in the same room as the respective dosed
 

animals. Animals fed 4-amino-2-nitrophenol were maintained in
 

the same rooms as animals of the same species being administered
 

one of the following chemicals:
 

Rats
 

Feed Studies
 

(CAS 624-18-0) p-phenylenediamine dihydrochloride
 

(CAS 18662-53-8) nitrilotriacetic acid, trisodium salt
 

(CAS 101-61-1) 4,4'-methylene bis(N,N'-dimethylaniline)
 

(CAS 105-11-3) p-quinone dioxime
 
Gavage Studies
 

(CAS 4377-33-7) 2-(chloromethyl)pyridine hydrochloride
 

(CAS 100-42-5) styrene
 

Mice
 

Feed Studies
 

(CAS 76-78-9) triphenyltin hydroxide
 

(CAS 91-93-0) 3-,3'-dimethoxybenzidine diisocyanate
 

(CAS 77-65-6) alpha-bromo-alpha-ethylbutyryl carbamide
 

(CAS 105-55-5) N,N'-diethylthiourea
 

(CAS 1596-84-5) succinic acid, 2,2-dimethylhydrazide
 

(CAS 126-31-8) iodomethanesulfonic acid, sodium salt
 

(CAS 105-11-3) p-quinone dioxime
 

(CAS 150-38-9) ethylenediaminetetraacetic acid, trisodium salt
 

trihydrate
 

Gavage Studies
 

(CAS 434-13-9) lithocholic acid
 

E. Subchronic Studies
 

Subchronic studies were conducted with Fischer 344 rats and
 

B6C3F1 mice of each sex to estimate the maximum tolerated doses
 

of 4-amino-2-nitrophenol, on the basis of which two
 

concentrations (hereinafter referred to as "low" and "high"
 

doses) were selected for the chronic studies.
 

4-Amino-2-nitrophenol was administered in the diet to rats at one
 

of 11 doses, either 147, 215, 316, 464, 681, 1,000, 1,470, 2,150,
 

3,160, 4,640, or 6,810 ppm and to mice at one of 11 doses, either
 

100, 147, 215, 316, 464, 681, 1,000, 1,470, 2,150, 3,160, or
 

4,640 ppm. Groups of five males and five females were tested at
 
each dose, and groups of equal size were used as matched
 

controls. The compound was administered for 6 weeks, and the
 

animals were observed for the following 2 weeks.
 

After the 6 weeks of compound administration, there were no
 

deaths in the rats or the mice, nor were there any changes in the
 

mean body weights of the dosed animals in comparison with the
 

controls, other than an approximate 20% weight depression in
 

female rats fed concentrations of 3,160 ppm and greater. No
 

gross pathologic changes were reported.
 

The doses selected for the chronic studies in both species were
 

1,250 and 2,500 ppm.
 

F. Chronic Studies
 

The test groups, doses administered, and times on study of the
 

chronic feeding studies are shown in tables 1 and 2.
 

G. Clinical and Pathologic Examinations
 

All animals were observed twice daily for signs of toxicity, and
 

animals that were moribund were killed and necropsied. Rats and
 

mice were weighed at regular intervals. Palpation for masses was
 

carried out at each weighing.
 

The pathologic evaluation consisted of gross and microscopic
 
Table 1. 4-Amino-2-Nitrophenol Chronic Feeding Studies in Rats
 

4-Amino-2­

Sex and Initial Nitrophenol Time on Study 

Test No. of Dose° Dosed Observed0 

Group Animals3 (ppm) (weeks) (weeks) 

Male 

Matched-Control 20 0 105 

Low-Dose 50 1,250 103 2 

High-Dose 50 2,500 103 2 

Female 

Matched-Control 20 0 105 

Low-Dose 50 1,250 103 2 

High-Dose 50 2,500 103 2 

a

All animals were approximately 6 weeks of age when placed on
 

study.
 

t>Rats were fed the diet preparations ad libitum, 7 days per week.
 

c

Control diet was fed during the observation period.
 
Table 2. 4-Amino-2-Nitrophenol Chronic Feeding Studies in Mice
 

4-Amino-2­

Sex and 

Test 

Group 

Initial 

No. of 

Animalsa 

Nitrophenol 

Dose° 

(ppm) 

Time 

Dosed 

(weeks) 

on Study 

Observed0 

(weeks) 

Male 

Matched-Control 

Low-Dose 

High-Dose 

20 

50 

50 

0 

1,250 

2,500 

103 

103 

105 

2 

2 

Female 

Matched-Control 

Low-Dose 

High-Dose 

20 

50 

50 

0 

1,250 

2,500 

103 

103 

105 

2 

2 

a

All animals were approximately 6 weeks of age when placed on
 

study.
 

°Mice were fed the diet preparations ad_ libitum, 7 days per week*
 

G

Control diet was fed during the observation period.
 

10
 
examination of major tissues, major organs, and all gross lesions
 

from killed animals and from animals found dead. All animals
 

were killed with carbon dioxide. The following tissues were
 

examined microscopically: skin, lymph nodes, mammary gland,
 

salivary gland, bone marrow, trachea, lungs and bronchi, heart,
 

thyroid, parathyroid, esophagus, stomach, small intestine, large
 

intestine, liver, gallbladder (mice), pancreas, spleen, kidney,
 

adrenal, urinary bladder, prostate or uterus, testis or ovary,
 

brain, and pituitary. Occasionally, additional tissues were also
 

examined microscopically. The different tissues were preserved
 

in 10% buffered formalin, embedded in paraffin, sectioned, and
 

stained with hematoxylin and eosin. An occasional section was
 

subjected to special staining techniques for more definitive
 

diagnosis.
 

A few tissues from some animals were not examined, particularly
 

from those animals that died early. Also, some animals may have
 

been missing, cannibalized, or judged to be in such an advanced
 

state of autolysis as to preclude histopathologic evaluation.
 

Thus, the number of animals from which particular organs or
 

tissues were examined microscopically varies, and does not
 

necessarily represent the number of animals that were placed on
 

study in each group.
 

11
 
H. Data Recording and Statistical Analyses
 

Pertinent data on this experiment have been recorded in an automatic data processing system, the Carcinogenesis Bioassay Data
 

System (Linhart et al., 1974). The data elements include descriptive information on the chemicals, animals, experimental design,
 

clinical observations, survival, body weight, and individual
 

pathologic results, as recommended by the International Union
 

Against Cancer (Berenblum, 1969). Data tables were generated for
 

verification of data transcription and for statistical review.
 

These data were analyzed using the statistical techniques
 

described in this section. Those analyses of the experimental
 

results that bear on the possibility of carcinogenicity are
 

discussed in the statistical narrative sections.
 

Probabilities of survival were estimated by the product-limit
 

procedure of Kaplan and Meier (1958) and are presented in this
 

report in the form of graphs. Animals were statistically
 

censored as of the time that they died of other than natural
 

causes or were found to be missing; animals dying from natural
 

causes were not statistically censored. Statistical analyses for
 

a possible dose-related effect on survival used the method of Cox
 

(1972) for testing two groups for equality and Tarone's (1975)
 

extensions of Cox's methods for testing for a dose-related trend.
 

12
 
One-tailed P values have been reported for all tests except the
 

departure from linearity test, which is only reported when its
 

two-tailed P value is less than 0.05.
 

The incidence of neoplastic or nonneoplastic lesions has been
 

given as the ratio of the number of animals bearing such lesions
 

at a specific anatomic site (numerator) to the number of animals
 

in which that site is examined (denominator). In most instances,
 

the denominators included only those animals for which that site
 

was examined histologically. However, when macroscopic examination was required to detect lesions prior to histologic sampling
 

(e.g., skin or mammary tumors), or when lesions could have
 

appeared at multiple sites (e.g., lymphomas), the denominators
 

consist of the numbers of animals necropsied.
 

The purpose of the statistical analyses of tumor incidence is to
 

determine whether animals receiving the test chemical developed a
 

significantly higher proportion of tumors than did the control
 

animals. As a part of these analyses, the one-tailed Fisher
 

exact test (Cox, 1970) was used to compare the tumor incidence of
 

a control group with that of a group of dosed animals at each
 

dose level. When results for a number of dosed groups (k) are
 

compared simultaneously with those for a control group, a
 

correction to ensure an overall significance level of 0.05 may be
 

made. The Bonferroni inequality (Miller, 1966) requires that the
 

13
 
P value for any comparison be less than or equal to 0.05/k. In
 

cases where this correction was used, it is discussed in the
 

narrative section. It is not, however, presented in the tables,
 

where the Fisher exact P values are shown.
 

The Cochran-Armitage test for linear trend in proportions, with
 

continuity correction (Armitage, 1971), was also used. Under the
 

assumption of a linear trend, this test determines if the slope
 

of the dose-response curve is different from zero at the one-


tailed 0.05 level of significance. Unless otherwise noted, the
 

direction of the significant trend is a positive dose relationship. This method also provides a two-tailed test of departure
 

from linear trend.
 

A time-adjusted analysis was applied when numerous early deaths
 

resulted from causes that were not associated with the formation
 

of tumors. In this analysis, deaths that occurred before the
 

first tumor was observed were excluded by basing the statistical
 

tests on animals that survived at least 52 weeks, unless a tumor
 

was found at the anatomic site of interest before week 52. When
 

such an early tumor was found, comparisons were based exclusively
 

on animals that survived at least as long as the animal in which
 

the first tumor was found. Once this reduced set of data was
 

obtained, the standard procedures for analyses of the incidence
 

14
 
of tumors (Fisher exact tests, Cochran-Armitage tests, etc.) were
 

followed.
 

When appropriate, life-table methods were used to analyze the
 

incidence of tumors. Curves of the proportions surviving without
 

an observed tumor were computed as in Saffiotti et al. (1972).
 

The week during which an animal died naturally or was sacrificed
 

was entered as the time point of tumor observation. Cox's
 

methods of comparing these curves were used for two groups;
 

Tarone's extension to testing for linear trend was used for three
 

groups. The statistical tests for the incidence of tumors which
 

used life-table methods were one-tailed and, unless otherwise
 

noted, in the direction of a positive dose relationship.
 

Significant departures from linearity (P < 0.05, two-tailed test)
 

were also noted.
 

The approximate 95 percent confidence interval for the relative
 

risk of each dosed group compared to its control was calculated
 

from the exact interval on the odds ratio (Gart, 1971). The
 

relative risk is defined as Pt/Pc where pj- is the true binomial
 

probability of the incidence of a specific type of tumor in a
 

dosed group of animals and pc is the true probability of the
 

spontaneous incidence of the same type of tumor in a control
 

group. The hypothesis of equality between the true proportion of
 

a specific tumor in a dosed group and the proportion in a control
 

15
 
group corresponds to a relative risk of unity. Values in excess
 

of unity represent the condition of a larger proportion in the
 

dosed group than in the control.
 

The lower and upper limits of the confidence interval of the
 

relative risk have been included in the tables of statistical
 

analyses. The interpretation of the limits is that in
 

approximately 95% of a large number of identical experiments, the
 

true ratio of the risk in a dosed group of animals to that in a
 

control group would be within the interval calculated from the
 

experiment. When the lower limit of the confidence interval is
 

greater than one, it can be inferred that a statistically
 

significant result (P < 0.025 one-tailed test when the control
 

incidence is not zero, P < 0.050 when the control incidence is
 

zero) has occurred. When the lower limit is less than unity, but
 

the upper limit is greater than unity, the lower limit indicates
 

the absence of a significant result while the upper limit
 

indicates that there is a theoretical possibility of the
 

induction of tumors by the test chemical, which could not be
 

detected under the conditions of this test.
 

16
 
III. RESULTS - RATS
 

A. Body Weights and Clinical Signs (Rats)
 

Mean body weights of dosed male and female rats were not
 

appreciably affected by the administration of 4-amino-2­

nitrophenol (figure 1). Fluctuation in the growth curve may be
 

due to mortality; as the size of a group diminishes, the mean
 

body weight may be subject to variation. No other clinical signs
 

were reported on the animals administered the test chemical.
 

B. Survival (Rats)
 

The Kaplan and Meier curves estimating the probabilities of
 

survival for male and female rats administered 4-amino-2­

nitrophenol in the diet at the doses of this bioassay, together
 

with those of the matched controls, are shown in figure 2. The
 

result of the Tarone test for dose-related trend in mortality is
 

not significant in either sex.
 

In male rats, 33/50 (66%) of the high-dose group, 37/50 (74%) of
 

the low-dose group, and 12/20 (60%) of the matched-control group
 

lived to termination of the study. In females, 37/50 (74%) of
 

the high-dose group, 36/49 (74%) of the low-dose group, and 14/20
 

(70%) of the matched-control group survived to termination of the
 

study.
 

17
 
MEAN BODY 

MEAN BODY WEIGHT (G) 

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TIME ON STUDY (WEEKS) 

Figure 2. Survival Curves for Rats Fed 4-Amino-2-Witrophenol in the Diet 

19
 
Sufficient numbers of rats of each sex were at risk for
 

development of late-appearing tumors.
 

C. Pathology (Rats)
 

Histopathologic findings on neoplasms in rats are summarized in
 

Appendix A, tables Al and A2; findings on nonneoplastic lesions
 

are summarized in Appendix C, tables Cl and C2.
 

Tumors of the urinary bladder occurred only in the rats administered 4-amino-2-nitrophenol, as follows:
 

RATS
 

Males Females
 

Matched Low High Matched Low High
 

Control Dose Dose Control Dose Dose
 

Number of Animals with
 

Tissues Examined
 

Microscopically (15) (46) (39) (15) (42) (46)
 

Transitional-cell
 

carcinoma 11(28%) 1(2%) 2(4%)
 

Transitional-cell
 

papilloma 2(5%)
 

Transitional-cell
 

hyperplasia 4(10%)
 

Microscopically, the transitional-cell tumors varied from
 

papillary structures packed with hyperchromatic epithelial cells,
 

pleomorphic nuclei, and mitotic figures to subepithelial, dome-


shaped solid masses of similar tumor cells. The masses often
 

20
 
protruded into the bladder lumen. There was invasion of the
 

bladder wall, and in one case metastases appeared in the lungs.
 

Other remaining tumors that occurred in the control and dosed
 

rats were considered spontaneous. Most of them had incidences as
 

expected for this age and Fischer 344 strain of rat. In these
 

cases there were approximate equivalent frequencies and expected
 

sex predilections for tumors and hyperplasias, including
 

testicular interstitial-cell tumors in the males, pituitary
 

chromophobe tumors mainly in females, and C-cell tumors and
 

hyperplasias of the thyroid in both sexes.
 

There were a few randomly distributed malignant tumors in the
 

dosed rats that did not occur in the controls, including an
 

epidermoid (squamous-cell) carcinoma of the salivary gland (1/47
 

high-dose males); a fibrosarcoma (1/50 high-dose males); an osteosarcoma (1/50 high-dose females); and a chondrosarcoma (1/50
 

high-dose females) that metastasized to the lung, but because of
 

the single occurrences they were not considered significant.
 

There occurred also a variety of nonneoplastic lesions that are
 

commonly observed in Fischer 344 rats.
 

Pigmentary changes occurred in the lamina propria of the small
 

intestines in 44/45 low-dose males, 43/45 high-dose males, 43/44
 

low-dose females, and 43/47 high-dose females. The pigment was
 

21
 
dark brown and finely granular. It was within macrophages of the
 

lamina propria and usually oriented more towards the tips of the
 

villi. The pigment was not birefringent under polarized light
 

and was negative for iron (Prussian-blue). It was melanin-like,
 

and appeared to be more abundant in the high-dose group.
 

There were degenerative and inflammatory conditions usually
 

encountered in aging rats.
 

The results of the histopathologic examination indicate that the
 

transitional-cell tumors of the urinary bladder occurring in the
 

high-dose male rats and in the low- and high-dose females, the
 

transitional-cell papillomas and hyperplasias occurring in the
 

dosed males, and the pigmentary intestinal changes occurring in
 

all dosed groups were induced by the administration of
 

4-amino-2-nitrophenol under the conditions of this bioassay.
 

D. Statistical Analyses of Results (Rats)
 

Tables El and E2 in Appendix E contain the statistical analyses
 

of the incidences of those primary tumors that occurred in at
 

least two animals of one group and at an incidence of at least 5%
 

in one or more than one group.
 

In male rats, the results of the Cochran-Armitage test for positive dose-related trend in the incidence of transitional-cell
 

22
 
carcinomas of the urinary bladder is significant (P < 0.001). A
 

departure from linear trend is indicated (P = 0.030), because of
 

the relatively steep increase in incidence in the high-dose
 

group. The results of the Fisher exact test show that the
 

incidence in the high-dose group is significantly higher (P =
 

0.018) than that in the matched-control group. The statistical
 

conclusion is that the incidence of transitional-cell carcinomas
 

of the urinary bladder in male rats is associated with the
 

administration of 4-amino-2-nitrophenol. No tumors of the
 

urinary bladder were found in 220 male or 220 female historical
 

controls at this laboratory. The results of the statistical
 

tests on the incidence of this tumor in female rats are not
 

significant; however, transitional-cell carcinomas of the urinary
 

bladder were observed in 1/43 (2%) of the low-dose females and
 

2/44 (5%) of the high-dose females compared with 0/15 in the
 

control groups.
 

A significant dose-related trend in the negative direction (P =
 

0.043) is observed in the incidence of thyroid tumors in male
 

rats, due to the higher incidence in the control group than in
 

the dosed groups.
 

23
 

IV. RESULTS - MICE
 

A. Body Weights and Clinical Signs (Mice)
 

Mean body weights of dosed male and female mice were only
 

slightly lower than those of corresponding matched controls
 

(figure 3). Fluctuation in the growth curve may be due to
 

mortality; as the size of a group diminishes, the mean body
 

weight may be subject to variation. No other clinical signs were
 

reported.
 

B. Survival (Mice)
 

The Kaplan and Meier curves estimating the probabilities of
 

survival for male and female mice administered 4-amino-2­

nitrophenol in the diet at the doses of this bioassay, together
 

with those of the matched controls, are shown in figure 4. The
 

result of the Tarone test for dose-related trend in mortality is
 

not significant in either sex.
 

In male mice, 43/50 (86%) of the high-dose group, 41/50 (82%) of
 

the low-dose group, and 17/20 (85%) of the matched-control group
 

lived to termination of the study. In females, 44/50 (88%) of
 

the high-dose group, 46/50 (92%) of the low-dose group, and 19/20
 

(95%) of the matched-control group survived to termination of the
 

study.
 

25
 
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—T" —I— —I— —I— —I— —I— —I— 

10 20 30 40 SOO 60 70 80 90 100 

TIME ON STUDY (WEEKS) 

Figure 3. Growth Curves for Mice Fed 4-Amino-2-Nitrophenol in the Diet 

26 
PROBABILITY OF SURVIVAL PROBABILIT

Y OF 

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Sufficient numbers of mice of each sex were at risk for development of late-appearing tumors.
 

C. Pathology (Mice)
 

Histopathologic findings on neoplasms in mice are summarized in
 

Appendix B, tables Bl and B2; findings on nonneoplastic lesions
 

are summarized in Appendix D, tables Dl and D2.
 

All of the tumors and hyperplasias that occurred in the mice were
 

spontaneous types which occurred in approximately equal
 

incidences in the control and dosed groups. There was a slight
 

increase of hepatic adenomas in the high-dose mice compared with
 

the controls, but an equal percentage of hepatocellular
 

carcinomas occurred in both control and dosed groups.
 

Several nonneoplastic changes were observed and were considered
 

to be either spontaneous or intercurrent disease processes. One
 

change that occurred in the dosed mice but not in the controls
 

consisted of deposits of dark brown, finely granular pigment in
 

the lamina propria of the small intestine. The pigment change
 

occurred in 46/49 low-dose and 43/47 high-dose males and in 43/48
 

low-dose and 42/47 high-dose females. It appeared similar to
 

that described in the rats of this study with respect to its
 

location, dose relationship, and staining characteristics.
 

28
 
The results of the histopathologic examination indicate that
 

4-amino-2-nitrophenol was not carcinogenic in mice but induced
 

the deposition of pigment in the lamina propria of the small
 

intestine under the conditions of this bioassay.
 

D. Statistical Analyses of Results (Mice)
 

Tables Fl and F2 in Appendix F contain the statistical analyses
 

of the incidences of those primary tumors that occurred in at
 

least two animals of one group and at an incidence of at least 5%
 

in one or more than one group.
 

The results of the Cochran-Armitage test for dose-related trend
 

and those of the Fisher exact test comparing the incidences of
 

tumors in each of the dosed groups with that in the control group
 

are not significant in the positive direction in either sex.
 

Significant results in the negative direction are observed in the
 

incidence of alveolar/bronchiolar carcinoma of the lung in male
 

mice; however, when combined incidences of animals with either
 

adenoma or carcinoma of the lungs are analyzed, there is no
 

significant difference between control and dosed groups. In
 

female mice, a significant trend (P = 0.035) in the negative
 

direction in the incidences of follicular-cell adenoma or
 

papillary adenoma of the thyroid is observed, but the results of
 

the Fisher exact tests are not significant.
 

29
 
In each of the 95% confidence intervals of relative risk, shown
 

in the tables, the value of one or less than one is included;
 

this indicates the absence of significant positive results. It
 

should also be noted that each of the intervals (except that for
 

the incidence of alveolar/bronchiolar carcinoma of the lung in
 

male mice) has an upper limit greater than one, indicating the
 

theoretical possibility of the induction of tumors by 4-amino-2­

nitrophenol, which could not be detected under the conditions of
 

this test.
 

30
 
V. PISCUSSION
 

On the basis of rate of growth, mortality, or other clinical
 

signs, there was little evidence of toxicity of 4-amino-2­

nitrophenol in the dosed rats or mice. Mean body weights of the
 

female mice were only slightly lower than those of the controls
 

throughout much of the bioassay. The survival of either rats or
 

mice was not affected by the test chemical, and at the end of the
 

bioassay, the survival of the animals in dosed and control groups
 

of both the rats and the mice was at least 60%. Sufficient
 

numbers of animals were at risk for the development of late-


appearing tumors. Since both male and female mice receiving
 

4-amino-2-nitrophenol had little or no depression in mean weights
 

and their survival was comparable to controls, they may have been
 

able to tolerate a higher dose.
 

In rats, transitional-cell carcinomas of the urinary bladder
 

showed a dose-related trend in the males (P < 0.001) and occurred
 

at a significantly higher incidence (P = 0.018) in the high-dose
 

males than in the matched-control males (controls 0/15, low-dose
 

0/46, high-dose 11/39 [28%]). Carcinomas of the bladder also
 

occurred in one low-dose female and two high-dose females, but in
 

none of the control females. Transitional-cell papillomas of the
 

bladder occurred in two additional high-dose males, and
 

transitional-cell hyperplasia of the bladder occurred in four
 

31
 
additional high-dose males, but neither lesion occurred in
 

control males. No tumors of the bladder were found among 220
 

male and 220 female historical-control Fischer 344 rats at this
 

laboratory.
 

In mice, no tumors occurred in dosed groups of males or females
 

at incidences that were significantly higher than those in the
 

corresponding matched-control groups.
 

Deposition of pigment occurred in the lamina propria of the small
 

intestine in at least 91% of the animals in the dosed groups of
 

rats and in at least 89% of the animals in the dosed groups of
 

mice, but in none of the control groups of either species.
 

The LD^Q of 4-amino-2-nitrophenol in Charles River CD rats has
 

been reported as 3,300 mg/kg when the chemical was administered
 

orally and 302 mg/kg when it was given by intraperitoneal
 

injection (Burnett et al., 1976; Burnett et al., 1977). A hair
 

dye containing the chemical caused no embryotoxic or teratogenic
 

effects in CD rats when it was applied to the skin at 2 ml/kg at
 

intervals during pregnancy (Burnett et al. , 1976) and also did
 

not induce a dominant lethal effect when it was tested in mature
 

male CD rats by intraperitoneal injection at a dose of 20 mg/kg
 

(Burnett et al. , 1977). When a hair dye containing 4-amino-2­

nitrophenol as well as Cl Acid Black 107 was applied to the skin
 

32
 
of DBAf or A strain mice, tumors of lymphoid origin developed at
 

a higher incidence in the DBAf strain and at earlier times in
 

both strains than in corresponding untreated controls (Venitt and
 

Searle, 1976). Two dosed DBAf females developed sarcomas of the
 

genital tract at weeks 66 and 69 (Venitt and Searle, 1976).
 

Positive results were also obtained when the same hair dye
 

(Venitt and Searle, 1976) or the 4-amino-2-nitrophenol alone
 

(Garner and Nutman, 1977) was tested in the Salmonella
 

mutagenicity test (McCann et al., 1975).
 

It is concluded that under the conditions of the bioassay,
 

4-amino-2-nitrophenol was carcinogenic for male Fischer 344 rats,
 

inducing transitional-cell carcinomas of the urinary bladder; the
 

transitional-cell carcinomas of the urinary bladder observed in
 

three dosed female rats may also have been associated with
 

administration of the 4-amino-2-nitrophenol. The test chemical
 

was not carcinogenic for male or female B6C3F1 mice at the doses
 

tested.
 

33
 

VI. BIBLIOGRAPHY
 

Ames, B. N. , Kammen, H. 0., and Yamasaki, E. , Hair dyes are
 

mutagenic: identification of a variety of mutagenic 

ingredients. Proc. Nat. Acad. Sci. U.S.A. 72(6):2423-2427, 

1975. 

Armitage, P. , Statistical Methods in Medical Research, John Wiley
 

& Sons, Inc., New York, 1971, pp. 362-365.
 

Berenblum, I. , ed. , Carcinogenicity Testing: _A Report of the
 

Panel of Carcinogenicity of the Cancer Research Commission
 

of UICC, Vol. _2, International Union Against Cancer, Geneva,
 

1969.
 

Burnett, C., Loehr, R. , and Corbett, J. , Dominant lethal
 

mutagenicity study on hair dyes. J. Toxicol. Environ. Health
 

2^:657-662, 1977.
 

Burnett, C. , Goldenthal, E. I., Harris, S. B., Wazeter, F. X. ,
 

Strausburg, J., Kapp, R. , and Voelker, R. , Teratology and
 

percutaneous toxicity studies on hair dyes. J. Toxicol.
 

Environ. Health 1:1027-1040, 1976.
 

Corbett, J. F. and Menkart, J. , Hair coloring. Cutis 12:190-197,
 

1973.
 

Cox, D. R. , Regression models and life tables. J. R. Statist.
 

Sec. B 34(2):187-220, 1972.
 

Cox, D. R., Analysis of Binary Data, Methuen & Co., Ltd., London,
 

1970, pp. 48-52.
 

Food and Drug Administration, FDA File of Cosmetic Product
 

Ingredient Statements Registered in Accordance with 21 CFR
 

720, August 15, 1977.
 

Garner, R. C. and Nutman, C. A. , Testing of some azo dyes and
 

their reduction products for mutagenicity using Salmonella
 

typhimurium TA 1538. Mutation Res. 44:9-19, 1977.
 

Gart, J. J., The comparison of proportions: a review of
 

significance tests, confidence limits and adjustments for
 

stratification. Rev. Int. Statist. Inst. 39(2):148-169,
 

1971.
 

35
 
Kaplan, E. L. , and Meier, P., Nonparametric estimation from
 

incomplete observations. J_._ Am. Statist. Assoc. 53;457-481,
 

1958.
 

Linhart, M. S. , Cooper J. A., Martin, R. L. , Page, N. P., and
 

Peters, J. A., Carcinogenesis bioassay data system. Comp.
 

and Biomed. Res. 7:230-248, 1974.
 

McCann, J., Choi, E., Yamasaki, E., and Ames, B. N. Detection of
 

carcinogens as mutagens in the Salmonella/microsome test:
 

assay of 300 chemicals. Proc. Nat. Acad. Sci. U.S.A.
 

_7_2:950, 1975.
 

Miller, R. G., Jr., Simultaneous Statistical Inference,
 

McGraw-Hill Book Co., New York, 1966, pp. 6-10.
 

Saffiotti, U., Montesano, R. , Sellakumar, A. R., Cefis, F. , and
 

Kaufman, D. G. , Respiratory tract carcinogenesis in hamsters
 

induced by different numbers of administrations of benzo (a)
 

pyrene and ferric oxide. Cancer Res. 32:1073-1081, 1972.
 

Society of Dyers and Colourists, Oxidation bases. Colour Index,
 

Vol. _4_, The Society of Dyers and Colourists; Yorkshire,
 

England, 1971, pp. 4641, 4648, and 4814.
 

Tarone, R. E. , Tests for trend in life table analysis*
 

Biometrika 62(3):679-682, 1975.
 

Venitt, S. , and Searle, C. E. , Mutagenicity and possible
 

carcinogenicity of hair colourants and constituents. In:
 

Environmental Pollution and Carcinogenic Risks, IARC Sci.
 

Publication No. 13, International Agency Research Cancer,
 

Lyon, 1976, pp. 263-272.
 

Verkade, P. E. , van Dijk, C. P., and Meerburg, W. , Researches on
 

the alkoxy-amino-nitrobenzenes. Rec. trav. chim. 65!346-360,
 

1946.
 

Wall, F. E., Bleaches, hair colorings, and dye removers. In:
 

Cosmetics, Science and Technology, Balsam, M. S. and
 

Sagarin, E. , eds., Wiley-Interscience, New York, 1972, pp.
 

296-300.
 

36
 
APPENDIX A
 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN
 

RATS FED 4-AMINO-2-NITROPHENOL IN THE DIET
 

37
 

TABLE A1.

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE RATS 

FED 4-AMINO-2-NITROPHENOL IN THE DIET 

MATCHED
 

CONTROL LOW DOSE HIGH DOSE
 

ANIMALS INITIALLY IN STUDY 20 50 50
 ANIMALS NECHOPSIED 20 50 50
 ANIMALS EXAMINED HISTOPATHOLOGICALLY 19 50 50
 

INTEGUMENTARY SYSTEM
 

*SKIN (20) (50) (50)
 

SQUAMOUS CELL CARCINOMA 1 (2X)
 

*SUBCUT TISSUE (20) (50) (50)
 

SQUAMOUS JELL CARCINOMA 1 (5*) 1 (2%) 1 (2%}
 

FIBROMA 1 (5X) 1 (2*) 1 (2«)
 

FIEROSARCOMA 1 (2«)
 

LIFOMA 1 (2%)
 

NEUROFIBRUMA 1 (5*)
 

RESPIRATORY SYSTEM
 

ILUNG (19) (50) (48)
 

SQUAMOUS CELL CARCINOMA, METASTA 1 (5%) 1 (2*) 1 (2*)
 

TRANSITIONAL-CELL CARCINOMA, MET 1 (2%)


ALVEOLAR/BRONCHIOLAR ADENOMA 1 (5%) 1 (2%)
 

HEMATOPOIET1C SYSTEM
 

*MULTIPLE ORGANS (20) (50) (50)
 

LEUKEMIA,NOS 3 (6X) 2
 

UNDIFFERENTIATED LEOKEHIA 3 (15%) 7 (14*) 7 (14*)
 

LYRPHOCYTiC LEUKEBIA 1 (2S)
 

GRANULOCYIIC LEUKEMIA 2 (4X) 1 (2X)
 

*SPLEEN (17) (45) (47)
 

MALIG.LYMPHOMA , UNDIFFER-TYPE 1 (2X)
 

CIRCULATORY SYSTEM
 

,,.NQNE
 

« NUMBER OF ANIMALS HITH TISSUE EXAMINED BICROSCOPICA LY
 

* NUMBER OF ANIHALS NECROPSIED
 

39
 
TABLE A1. MALE RATS: NEOPLASMS (CONTINUED) 

i

TM r
 "~~ — '" ~ - - - - '"""- ""~ "~ ' ~ "" ' ••
 

MATCHED
 

CONTROL LOW DOSE HIGH DOSE
 

DIGESTIVE SYSTEM 

tSALIVARY GLAND (16) (49) 

SQUAMOUS CELL CARCINOMA, INVASIV 

ILIVEP. HEPATOCELi,ULAR ADENOHA 

(19) (48) 

1 (2%) 

URINARY SYSTEil
 

*URINARY BLADDER (15) (46)
 

TRANSITIONAL-CELL PAPILLOBA
 

TRANSITIONAL-CELL CARCINOMA
 

HEMANGIOMA
 

ENDOCRINE SYSTEM
 

•PITUITARY	 (15) (»0)
 

CHROHOPHOitE ADENOMA 2 (13%) 4 (10S)
 

tADRENAL (18) (46)
 

CORTICAL CARCINOMA
 

PHEOCHROMOCYTOMA 1 (6%) 1 (2S)
 

tTHYROID (18) (42)
 

FOLLICULAfi-CELL ADENOMA 1 (6%)
 

FOLLICULAH-CELL CARCINOMA
 

C-CELL ADENOMA 2 (11%) 2 (5%)
 

C-CELL CAHCINOMA 1 (6%)
 

tPANCBEATIC ISLETS (18) (46)
 

ISLET-CELL ADENOMA 2 (11*) 1 (2%)
 

BEPRODOCTIVE SYSTEM
 

tTESTIS (") (50)
 

INTERSTITiAL-CELL TOBOR 15 (88*) 50 (100X)
 

NERVOUS SYSTES
 

NONE
 

* NUMBER OF ANIMALS HITH TISSUE EXAHINED HICROSCOPICALLY
 * NUBBER OF ANIMALS NECROPSIED
 

40
 

(47)
 

1 (2%)
 

(48)
 

(39)
 

2 (5%)
 

11	 (28%)
 

1 (3%)
 

(39)
 

8 (21%)
 

(49)
 

1 (2%)
 

2 (<»%)
 

(44)
 

1 (2%)
 

1 (2%)
 

1 (2%)
 

(49)
 

1 (2%)
 

(50)
 

41 (82%)
 
TABLE A1. MALE RATS: NEOPLASMS (CONTINUED) 

MATCHED
 

CONTROL LOW DOSE HIGH DOSE
 

SPECIAL SENSE ORGANS
 

NONE
 

MUSCULOSKELETAL SYSTEM
 

NONE
 

EODY CAVITIES
 

NONE
 

ALL OTHER SYSTEMS
 

NONE
 

*NIMAL DISPOSITION SUMMARY
 

ANIMALS INITIALLY IN STUDY
 

NATURAL DEATHS
 

MORIBUND SACRIFICE
 

SCHEDULED SACRIFICE
 

ACCIDENTALLY KILLED
 

TERMINAL SACRIFICE
 

ANIMAL MISSING
 

d INCLUDES AUIOLYZED ANIMALS
 

20 50 50
 

12 37 33
 

* NUMB3B OF ANIMALS HITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIED
 
TABLE A1. MALE RATS: NEOPLASMS (CONTINUED) 

MATCHED
 

CONTROL LOW DOSE HIGH DOSE
 

TUHOE SUMMARY
 

TOTAL ANIMALS WITH PRIHAHY TUMORS* 16 50 48
 

DIAL PSIdARY TUMOfiS 31 75 86
 

TOTAL ANIMALS WITH BENIGN TUMORS 15 50 46
 

TOTAL BENIGN TUMORS 26 60 60
 

TOTAL ANIMALS WITH MALIGNANT TUMORS 5 14 23
 

TOTAL MALIGNANT TUMOfiS 5 15 26
 

TOTAL ANIMALS WITH SECONDARY TUMORS* 1 1 2
 

TOTAL SECONDARY TUMORS 1 1 3
 

TOTAL ANIMAL S WITH TUMORS DNCERTAIN ­

BENIG N OR MALIGNAN T 

TOTAL UNCERTAI N TUMORS 

TOTAL ANIMAL S WIT H TUMORS UNCERTAIN ­

PRIMAR Y OR ilETASTATIC 

TOTAL UNCERTAI N TUMORS 

* PRIMARY TUMuRS: ALL TUMORS EXCEPT SECONDARY TUBORS
 

* SECONDARY TUMORS: ME1ASTATIC TUMORS OB TUMOKS INVASIVE INTO All ADJACENT ORGAN
 

42
 
TABLE A2.

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE RATS 

FED 4-AMINO-2-NITROPHENOL IN THE DIET 

MATCHED 

CONTROL LOW DOSE HIGH DOSE 

ANIMALS INITIALLY IN STUDY 20 350 50 

ANIMALS NECPOPSIED 20 49 50 

ANIMALS EXAMINED HIS] ^PATHOLOGICALLY 20 49 48 

INTEGUMENTARY SYSTEM
 

*SK1N (20) (1*9) (50)
 

FIBROMA 1 (2%)
 

*SOBCUT TISSUE (20) (49) (50)
 

SQUAMOUS CELL CARCINOMA 1 (2%)
 

FIBROMA 2 (4*) 1 (2%)
 

LIPOMA 1 (2X)
 

OSTEOSARCOMA 1 (2%)
 

CHONDROSARCOMA 1 (2%)
 

RESPIRATORY SiSTEM
 

ITRACHEA (18) (45) (48)
 

CAECINCMA-IN-SITU, NOS 1 (6%)
 

#LUNG (18) (48) (46)
 

CARCINOMA, NOS, METASTATIC 1 (6»)
 

ALVEOLAR/bRONCHIOLAR ADENOMA 1 (6X)
 

ALVEOLAR/oRONCHIOLAR CARCINOMA 1 (6*)
 

CKONDROSAHCOMA, METASTATIC 1 (2%)
 

HEMATOPOIEIIC SYSTEM
 

*MULT1PLE ORGANS (20) (49) (50)
 

MALIG.LYMPHOMA, UN DIPPER-TYPE 1 (2%)
 

LEUKEMIA,NOS 2 (14%)
 

UNBIFFSRENTIATED LEUKEMIA 1 (5X) 4 (8%) 1 (2X)
 

LYBPHOCYTXC LEUKEMIA 1 (5S)
 

GRANULOCYTIC LEUKEMIA 2 (4*)
 

tSPLEEN (19) (48) (47)
 

HEMANGIQMA 1 12%}
 

t NUMBER OP AiKIMALS HITH TISSUE EXAMINED MICROSCOPICALLY
 * NUMBER OF ANIMALS NECROPSIED
 

d 50 ANIMALS WERE INITIALLY IN THE STUDY, BDT ONE ANIMAL HAS POUND TO BB A HALE
 

IN A FEMALE GROUP.
 

43
 
TABLE A2. FEMALE RATS: NEOPLASMS (CONTINUED) 

MATCHED
 

CONTROL LOW DOSE HIGH DOSE
 

•MANDIBULAR	 L. NODE (19) (49) (47)
 

CARCINOMA, N05, METAS1ATIC 1 (2%)
 

CIRCULATORY SYSTEM
 

NONE
 

DIGESTIVE SYSTEM
 

#LIVEE (20) (U8) (48)
 

CARCINOMA, NOS, METAS1ATIC 1 (5%)
 

URINARY SYSTEM
 

KKIDNEY (20) (49) (48)
 

CARCINOMA, NOS, METASTAIIC 1 (5%)
 

#UR1NARY BLADDER (15) (43) (44)
 

TRANSITIONAL-CELL CARCINOM A 1 (2%) 2 (5X)
 

ENDOCRINE SYSTEM
 

• PITUITAR Y (18) (48) 

CHFOMOPHOdi ADENOMA 8 (44*) 26 (54S) 20 (44X) 

•ADRENAL	 (19) (48) (48)
 

PHEOCHROMOCYTOHA 1 (5%)
 

*THYROID (17) (44) (47)
 

ADENOMA, NOS 1 (2%)
 

FOLLICULAB-CELL ADENOMA 1 (2X)
 

C-CELL ADENOMA 1 (6»)
 

REPRODUCTIVE oYSTEM
 

*MAMMAEY GLAWD (20) (49) (50)
 

ADENOMA, NOS 1 (2%) 1 (2%)
 

FIBROADENOMA 1 (5%) 3 (6%) 5 (10X)
 

•UTERUS	 (17) (47) (48)
 

LEIOHYQMA
 

# NUMBE R OF ANIMAL S WITH TISSUE EXAMINE D MICROSCOPICALLY
 

* NUMBER OF ANIMALS NSCROPSIED
 

44
 
TABLE A2. FEMALE RATS: NEOPLASMS (CONTINUED) 

MATCHED 

CONTROL LOW DOSE HIGH DOSE 

NERVOUS SYSTEa 

NONE 

SPECIAL SENSE ORGANS 

NONE 

MUSCULOSKELETAL SYSTEM 

NONE 

BODY CAVITIES 

NONE 

ALL OThER SYSIEMS 

NONE 

ANIMAL DISPOSITION SUHMABY 

ANIMALS INITIALLY IN STUDY

NATURAL DEATHS

MORIBUND SACRIFICE

SCHEDULED SACRIFICE 

ACCIDENTALLY KILLED 

TERMINAL SACRIFICE

ANIMAL MISSING 

ANIMAL DELETED(WRONG SEX)

a INCLUDES AUTOLYZED ANIBALS 

 20

 1 

2 

 14

 50

5 

8 

 36

 1 

 50 

6 

7 

 37 

t NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY * NUMBER OF ANIMALS NECROPSIED 

45
 
TABLE A2. FEMALE RATS: NEOPLASMS (CONTINUED) 

TUMOR SUMMAR Y
 

TOTAL ANIMALS WITH PRIMARY TOMOES*
 

TOTAL PRIMARY TUMORS
 

TOTAL ANIMALS WITH BENIGN TUMORS
 

TOTAL EENIGN TUMORS
 

TOTAL ANIMALS WITH MALIGNANT TUMORS
 

TOTAL MALIGNANT TUMORS
 

TOTAL ANIMALS WITH SECONDARY TUMORS*
 

TOTAL SECONDARY TUM08S
 

TOTAL ANIMALS WITH TUMORS UNCERTAINBENIGN OH MALIGNANT
 

TOTAL UNCERTAIN TUMORS
 

TOTAL ANIMALS WITH TUMORS UNCERTAINPRIMARY OR METASTATIC
 

TOTAL UNCERTAIN TUMORS
 

MATCHED 

CONTROL LOW DOSE HIGH DOSE 

13 31 28 

16 U2 1*1 

11 30 23 

12 31* 33 

U 8 8 

a 8 8 

1 2 

3 2 

* PRIMARY TUMORS: ALL TUMORS EXCEPT SECONDARY TUMORS
 

* SECONDARY TUMORS: MSIASTATIC TUMORS 08 TUMORS INVASIVE INTO AN ADJACENT ORGAH
 

46
 
APPENDIX B
 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN
 

MICE FED 4-ANIMO-2-NITROPHENOL IN THE DIET
 

47
 

TABLE B1.

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE MICE 

FED 4-AMINO-2-NITROPHENOL IN THE DIET 

' 

MATCHED 

CONTROL LOW DOSE 

ANIMALS INITIALLY IN S1UEY 20 50 ANIMALS NECROPSIED 20 50 ANIMALS EXAMINED HI SI ^PATHOLOGICALLY 20 50 

INTEGUMENTARY SYSTEM
 

NONE
 

RESPIRATORY SYSTEM
 

tLUNG/BRONCHUS (20) (U9)
 

ADENOMATOUS POLYP, NOS 1 (2%)
 

«LUNG (20) (49)
 

HEPATOCELiULAR CAHCINOHA, METAST 1 (256)
 

ALVEOLAR/BRONCHIOLAR ADENOMA 2 (10X) 10 (20X)
 

ALVEOLAR/dRONCHIOLAR CARCINOMA 3 (155S)
 

PAPILLARY ADENOCARCINOHA, METAST 1 (5%)
 

HEMATOPOIEIIC SYSTEM
 

*MULTIPLE ORGANS (20) (50)
 

MALIG.LYMPHOMA, LYMPHOCYTIC TYPE 1 (2X)
 

MAIIG.LYMPHOMA, HISTIOCYTIC TYPE 2 (4*)
 

GRANULOCYIIC LEUKEMIA 1 (5S)
 

tLYMPK NODE (19) (50)
 

MALIG.LYMPHOMA, LYMPHOCYTIC TYPE 1 (2«)
 

MALIG.LYMPHOMA, HISTIOCYTIC TYPE
 

#MANDIBl'LAR L. NODE (19) (50)
 

MALIG.LYMPHOMA, LYMPHOCYTIC TYPE 1 (5X)
 

IMESENTERIC L. NODE (19) (50)
 

NEOPLASM, NOS
 

MALIG.LYfiPHOHA, HISTIOCYTIC TYPE
 

•LIVER	 (20) (50)
 

MALIG.LYMPHOMA. HISTIOCtTIC TYPE 1. (2*)
 

f NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECSOPSIED
 

49
 

HIGH DOSE
 

50
 

50
 

50
 

(48)
 

(48)
 

1 (2X)
 

7 <15X)
 

(50)
 

1 (2X)
 

1 (2X)
 

(48)
 

1 (2%)
 

(48)
 

(48)
 

1 12%)
 

1 (2X)
 

(49)
 
TABLE B1. MALE MICE: NEOPLASMS (CONTINUED) 

MATCHED 

CONTROL LOW DOSE HIGH DOSE 

CIRCULATORY S/STEM 

NONE 

DIGESTIVE SYSIEfi 

#LIVEF.

NEOPLASM, NOS

HEPATOCELi-ULAR ADSNOMA

HEPATOCELi-ULAH CARCINOMA

SARCOMA, NOS

 (20)

 3 (15*)

 3 (15S)

 (50)

 13 (26%)

 7 (14%)

 (49) 

1 (2%) 

 12 (24X) 

7 (14X) 

1 (2*) 

URINARY SYSTEd 

NONE 

ENDOCRINE SYSiEM 

ITHYROI D

FOLLICULAB-CELL CABCINOMA

 (18) (44) (45) 

1 (2%) 

REPRODUCTIVE oYSTEM 

NONE 

NERVOUS SYSTEM 

NONE 

SPECIAL SENSE ORGANS 

NONE 

HUSCULOSKELETAL

__NON E 

 SYSTEM 

* NUflBER

* NUMBER

 OF

 OF

 ANIMALS

 ANIMALS

 WITH TISSUE

 NECROPSIED 

 EXAHINE D MICROSCOPICALLY 

50 
TABLE B1. MALE MICE: NEOPLASMS (CONTINUED) 

--T--—• • 

MATCHED 

CONTROL LOW DOSE HIGH DOS!-; 

BODY CAVITIES
 

*MESENTEHY (20) {50} (50)
 

LIPOBA 1 (2%)
 

ALL OTHER SYSTEMS
 

THORAX
 

LIPOSABCOHA 1
 

INIMAL DISPOSITION SUMMARY
 

ANIMALS INIl'IALLY IN STUDY 20 50 50
 

NATURAL DEATHS 2 8 6
 

MORIBUND SACRIFICE 1 1
 

SCHEDULED SACRIFICE
 

ACCIDENTALLY KILLED 1
 

TERMINAL SACRIFICE 17 41 43
 

ANIMAL MISSING
 

a INCLUDES AUTOLYZED ANIMALS
 

TUMOR SUMMARY
 

TOTAL ANIMA1.S HITH PRIMARY TUMORS* 11 32 28
 

TOTAL PRIuARY TUMOES 13 36 36
 

TOTAL ANIMALS HITH BENIGN TUMORS 5 23 18
 

TOTAL BENIGN TUMORS 5 24 20
 

TOTAL ANIMALS WITH MALIGNANT TUMORS 8 12 13
 

TOTAL MALIGNANT TUMORS 8 12 14
 

TOTAL ANIMALS WITH SECONDARY TUMORS* 1 1 1
 

TOTAL SECONDARY TUMOBS 1 1 1
 

TOTAL ANIMALS HITH TUMORS UNCERTAINBENIGN OR MALIGNANT 1
 

TOTAL UNCERTAIN TUMORS 2
 

TOTAL ANIMALS HITH TUMORS ONCERTAINPEIMARY OR METASTATIC
 

TOTAL UNCERTAIN TUMOHS
 

* PRIMARY TUMORS: ALL TUMORS EXCEPT SECONDARY TUMORS
 

* SECONDARY HUMORS: METASTATIC TUMORS OR TUHOBS INVASIVE INTO AN ADJACENT ORGAK
 

51
 
TABLE B2.

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE MICE 

FED 4-AMINO-2-NITROPHENOL IN THE DIET 

MATCHED
 

CONTROL LOW DOSE
 

ANIMALS INITIALLY IN STUDY 20 50
 

ANIMALS NECRCPSIED 20 50
 

ANIMALS EXAMINED HI STO PATHOLOGICALLY 20 50
 

INTEGUMENTARY SYSTEM
 

*SUBCUT TISSUE (20) (50)
 

FIBROSABCoMA 1 (2X)
 

RHABDOMY03A
 

RESPIRATORY SYSTEM
 

*LUNG (20) (U9)
 

ALVEOLA8/ORONCHIOLAB ADENOMA 2 (10%) 3 (6%)
 

HEMATOPOIETIC SYSTEB
 

•MULTIPLE ORGANS (20) (50)
 

MALIGNANT LYMPHOMA, NOS 1 (5*) 1 (2«)
 

MALIG.LYKPHOHA, LYBPHOCYTIC TYPE t (2X)
 

MALIG.LYKPHOHA, HISTIOCYTIC TYPE 1 (5%) 2 (<*%)
 

LYHPHOCYT^C LEUKEMIA 1 12%)
 

*MEDIASTINUK (20) (50)
 

MALIG.LYKfHOBA, LYMPHOCYTIC TYPE
 

tSPLEEN (20) (50)
 

MALIG. LYMPHOMA. LYMPHOCYIIC TYPE
 

fLYMPH NODE (20) (U9)
 

MALIG.LYBPHOMA, LYMPHOCYTIC TYPE 2 (4X)
 

MALIG.LYBPHOBA, HISTIOCYIIC TYPE 1 (2X)
 

fMESENTERIC i, . NODE (20) (U9)
 

BALIG-LYKPHOMA, ONDIFFEB-TYPE 1 (5X)
 

#LIVEP. (20) (50)
 

MALIG.LYBPHOMA, UNDIFFEE-TYPE t 12%)
 

« NUMBER OF ANIMALS WITH TISSUE EXAMINED HICROSCOPICAU.*
 

* NUMBER OF ANIMALS NECROPSIED
 

HIGH DOSE
 

50
 

50
 

50
 

(50)
 

1 (2X)
 

(50)
 

2 (H%)
 

(50)
 

1 (2X)
 

2 (UX)
 

2 (UX)
 

(50)
 

1 (2X)
 

(50)
 

1 (2X)
 

(U8)
 

(U8)
 

(50)
 

52
 
TABLE B2. FEMALE MICE: NEOPLASMS (CONTINUED) 

MATCHED
 

CONTROL LOW DOSE HIGH DOSE
 

MALIS.iYMPHOMA, LYMPHOCYIIC TYPE	 1 (2X)
 

IPEYERS PAlCd {19} (48) (47)
 

MALIG.LYKPHOMA, LYHPHOCYTIC TYPE 1 (2X)
 

CIRCULATORY SiSTEM
 

NONE
 

DIGESTIVE SYSTEM
 

iLIVER (20) (50) (50)
 

HEPATOCELi,ULAR ADENOMA 1 (2X) 2 (4X)
 

HEPATOCEUULAR CARCINOHA 1 (2%)
 

tSTOHACH (19) (49) (47)
 

PAPILLCMA, NOS 1 (2X)
 

URINARY SYSTEM
 

NONE
 

ENDOCRINE SYSTEM
 

fADRENAL (19) (47) (46)
 

CORTICAL ADENOMA 1 (5X) 1 (2X) 1 (2X)
 

fTHYROIE (17) (40) (42)
 

PAPILLARY ADENOMA 1 (3X)
 

FOLLICULAR-CELL ADENOHA 2 (12X)
 

•PANCREATIC XSLETS (20) (49) (49)
 

ISLET-CELL ADENOMA 1 (2X)
 

BEPRODOCTIVE oYSTEM
 

•MAMMARY	 GLAND (20) (50) (50)
 

ADENOMA, NOS 1 (5X)
 

*OTERUS	 (19) (48) (49)
 

EiNDOMETRIAL STBOHAL POLYP 

* NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIED
 

53
 
TABLE B2. FEMALE MICE: NEOPLASMS (CONTINUED) 

MATCHED 

CONTROL LOW DOSE HIGH DOSE 

•OVARY

TERATOMA, BENIGN

 (18) (41) (45) 

1 (2X) 

NERVOUS SYSTEM 

NONE 

SPECIAL SENSi, ORGANS 

NONE 

HUSCULOSKELETAL SYSTEM 

NONE 

BODY CAVITIES 

NONE 

AIL OTHER

NON E 

 SYSTEM S 

ANIMAL DISFCSiTION SUMMAR Y 

ANIMALS IN1IIALLT IN STUDY 

NATURAL DEATHS 

MORIBUND SACRIFICE 

SCHEDULED SACRIFICE 

ACCIDENTALLY KILLED 

TERMINAL SACRIFICE 

ANIMAL MISSING 

3 INCLUDES AUIOLYZED ANIMALS 

20 

19 

50 50 

44 

* NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY * NUMBER OF ANIMALS NECROPSIED 

54
 
TABLE B2. FEMALE MICE: NEOPLASMS (CONTINUED) 

MATCHED
 

CONTROL LOW DOSE HIGH DOSE
 

IOMOE SUMMARY 

TOTAL ANIMALS WITH PRIMARY TUMORS* 

TOTAL PRIMARY TUMORS 

8 

9 

17 

19 

16 

17 

TOTAL ANIMALS WITH 3ENIGN TUMORS 

TOTAL EEN1GN TUMORS 

6 

6 

6 

7 

9 

9 

TOTAL ANIMALS WITH MALIGNANT TUMORS 

TOTAL MALIGNANT TUMORS 

3 

3 

12 

12 

8 

8 

TOTAL ANIMALS WITH SECONDARY TUMORS* 

TOTAL SECONDARY TUMORS 

TOTAL ANIMALS WITH TUMORS UNCERTAINBENIGN OR MALIGNANT 

TOTAL UNCERTAIN TUMORS 

TOTAL ANIMALS WITH TUMORS UNCERTAINPRIMABY OR METASTATIC 

TOTAL UNCERTAIN TUMORS
 

* PRIMARY TUMORS: ALL TUMORS EXCEPT SECONDARY TUMORS
 

I SECONDARY TUMORS: METASTATIC TUMORS OR TUMORS INVASIVE INTO AN ADJACENT ORGAN
 

55
 

APPENDIX C
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS
 

IN RATS FED 4-AMINO-2-NITROPHENOL IN THE DIET
 

57
 

TABLE C1.

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN MALE RATS 

FED 4-AMINO-2-NITROPHENOL IN THE DIET 

ANIMALS INITIALLY IN STUDY
 

ANIMALS NECROPSIED
 

ANIMALS EXAMINED HISTO PATHOLOGICALLY
 

INTEGUMENTARY SYSTEM
 

*SKiN
 

CYST, NOS
 

EPIDERMAL INCLUSION CYST
 

*SUBCUT TISSUE
 

ABSCESS, NOS
 

BESPIRAT08Y SiSTEM 

tTRACHE A 

INFLAMMATION, NOS
 

tLUNG
 

MINERALIZATION
 

CONGESTION, NOS
 

INFLAMMATION, FOCAL
 

BRONCHOPNiUMONIA, ACUTE
 

PNEUMONIA, CHRONIC MURINE
 

HYPERPLAS^A, ADENOMATOUS
 

HYPEEPLASiA, LYMPHOID
 

HEKATOPOIEIIC SYSTEM
 

•SPLEEN
 

CONGESTION, NOS
 

PIGMENTATION, NOS
 

CIRCULATORY SYSTEM
 

•HEART
 

THROHBOSIS. NOS
 

MATCHED 

CONTROL LOW DOSE HIGH DOSE 

20 50 50 

20 50 50 

19 50 50 

(20) (50) (50) 

1 (2X) 

1 (2X) 

(20) (50) (50) 

1 (2X) 

(17) (42) 

1 (2X) 

(19) (50) (48) 

1 (2X) 

4 (21X) 8 (16S) 5 (10%) 

1 <2X) 

11 (58X) 35 (10%) 

2 <4X) 

35 (73X) 

1 (5S) 2 (4X) 1 (2X) 

1 (2X) 

(17) (45) (47) 

1 (2X) 

1 (2%) 

(18) (50) (50) 

1 <2X> 

t NUMBEB OF AblMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 * NUHBER OF ANIMALS NECROPSIED
 

59
 
TABLE C1. MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

^ ' — -J -.1- ... - — • — -•• — •• - - ­

MATCHED
 

CONTROL LOW DOSE HIGH DOSE
 

•HEART/ATRIUa	 (18) (50) (50)
 

THROMBOSIS, NOS 1 (6X) 2 (4X) 1 (2X)
 

•MYOCARDIUM	 (18) (50) (50)
 

FIBBOSIS 15 (83X) 32 (64X) 32 (64X)
 

DEGENERATION, NOS 1 (6X) 2 (4X)
 

*GASTRODUODENAL ARTER (20) (50) (50)
 

PEP.IVASCUi.ITIS 1 (5X)
 

DIGESTIVE SYSTEM
 

•SALIVARY	 GLAND (16) (49) (47)
 

ATROPHY, NOS 1 (2X)
 

ATROPHY, JIFFUSE 1 (6X)
 

•LIVER	 (19) (48) (48)
 

CONGESTION, NOS 1 (2X)
 

GRANULOMA, NOS 1 (2X)
 

DEGENERATION, NOS 3 (16X) 5 (10X) 5 (10X)
 

NECROSIS, FOCAL 1 (2X)
 

METASORPHOSIS FATTY 1 (5X) 6 (13X) 7 (15X)
 

LIPOIDOSIi 1 (5X)
 

FOCAL CELLULAR CHANGE 1 (2X)
 

INCLUSION, CYTOPLASMIC 1 (2X)
 

HYPERPLASTIC NODULE 1 (2X)
 

HYPERPLASiA, FOCAL 11 (58X) 28 (58X) 17 (35X)
 

•LIVER/CENTRILOBULAR	 (19) (48) (48)
 

CONGESTION, NOS 1 (5X) 1 (2X)
 

DEGENERATION, NOS 1 (2X)
 

NECROSIS, NOS 1 (2X)
 

METAMORPHOSIS FATTY 1 (2X)
 

*BILE DUCT (20) (50) (50)
 

HYPESPLASIA, NOS 9 (45X) 26 (52%) 19 (38X)
 

•PANCREAS	 (18) (46) (49)
 

ATEOPHY, FOCAL 6 (33X) 9 (20X) 10 (20X)
 

•PANCREATIC ACINUS (18) (46) (49)
 

HYPERPLASJ.A, FOCAL 1 (2X)
 

•SHALL INTESTINE (15) (45) (45)
 

PIGMEHTAI-iON, NOS 44 (98X) 43 (96X)
 

• NUMBER OF ANIMALS HITH TISSUE EXAMINED MICROSCOPICALLY
 

60 
TABLE C1. MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

MATCHED
 

CONTROL LOW DOSE HIGH DOSE
 

•LARGE	 INTESTINE (15) (42) (44)
 

INFLAMMATION, NOS 1 (2%)
 

NEMATODIA-ilS 3 (20%) 10 (24X) 7 (16X)
 

URINARY SYSIEfl
 

•KIDNEY	 (19) (50) (50)
 

INFLAMMATION, CHRONIC 17 (89%) 48 (96X) 49 (98%)
 

•KIDNEY/TUBULE	 (19) (50) (50)
 

NECROSIS, NOS 1 (5%)
 

PIGMENTATION. NOS 3 (6X)
 

•URINARY	 BLADDER (15) (U6) (39)
 

HYPERPLASiA, EPITHELIAL 4 (10S)
 

ENDOCRINE SYSIEM
 

•PITUITARY	 (15) (40) (39)
 

CYST, NOS 1 (3X) 3 (8X)
 

•ADRENAL	 (18) (46) (49)
 

HEHORRaAGi 1 (6%)
 

LIPOIDOSIJ 2 (4X)
 

•ADRENAL COEXEX (18) (46) (49)
 

HYPERPLAS.LA, FOCAL 2 (4X)
 

•ADRENAL	 MEDULLA (18) (46) (49)
 

CYST, NOS 1 (2%)
 

•THYROID	 (18) (42) (44)
 

HYPERPLASiA, C-CELL 1 (2%) 1 (2X)
 

HYPERPLASiA, FOLLICULAR-CELL 1 (6%) 2 (5X)
 

•THYROID FOLLICLE (18) (42) (44)
 

HYPERTROPHY, FOCAL 1 (2X)
 

•PANCREATIC ISLETS (18) (46) (49)
 

HYPERTROPHY, NOS 1 (2X)
 

HYPERPLASIA. NOS 1 12%) 2 (4X)
 

• NUMBER OF ANIMALS HITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECBOPSIED
 

61
 
TABLE C1.MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

MATCHED 

CONTROL LOW DOSE HIGH DOSE 

REPRODUCTIVE SYSTEM 

*SEMINiL VESICLE (20) (50) (50) 

INFLAMMATION, ACUTE 1 (5X) 2 (4X) 

INFLAMMATION, ACUTE FOCAL 1 (2X) 

INFLAMMATION, CHRONIC 1 (2X) 

GRANULOMA , NOS 1 (2%) 

4TESTIS (17) (50) (50)
 

NECROSIS, NOS 2 (4X)
 

NECROSIS, FAT 1 (2%)
 

ATROPHY, NOS 1 (2X) 2 (4X)
 

NERVOUS SYSTEM
 

•BRAIK	 (18) (46) (48)
 

ABSCESS, NOS 1 (2%)
 

SPECIAL SENSE ORGANS
 

NONE
 

HUSCULOSKELETAL SYSTEM
 

*SKELETAL MUSCLE (20) (50) (50)
 

HEMORRHAGE 1 (2X)
 

BODY CAVITIES
 

*ABDOHINAL CAVITY (20) (50) (50)
 

NECROSIS, FAT 1 (2X)
 

*PLEURA (20) (50) (50)
 

FOAM-CELL 2 (4X)
 

ALL OTHER SYSTEMS 

, NONE , 

* NUMBER OF ANIMALS HITH TISSUE EXAMINED MICROSCOPICALLY
 * NUMBER OF ANIMALS NBCROPSIED
 

62
 
TABLE C1. MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

MATCHED
 

CONTROL LOW DOSE HIGH DOSE
 

SPECIAL HOEPBOLOGY SUMHABY
 

AUTO/NECROPSY/NO HiSTO
 

* NUHBEB OF ANIMALS WITH TISSUE EXAMINED BICEOSCOPICALLY
 * NUMBER OF ANIMALS NECEOPSIED
 

63
 
TABLE C2.

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN FEMALE RATS 

FED 4-AMINO-2-NITROPHENOL IN THE DIET 

MATCHED
 

CONTROL LOW DOSE HIGH DOSE
 

ANIMALS INITIALLY IN STUDY 20 950	 50
 

ANIMALS NECROt'SIED	 20 49 50
 

ANIMALS EXAMINED HIS1 ^PATHOLOGICALLY 20	 49 48
 

INTEGUMENTAR Y SYSTEH
 

*SKIN (20) (49) (50)
 

EPIDERMAL INCLUSION CYST 1 (5%)
 

INFLAMMATION, NOS 1 (5*)
 

NECROSIS, NOS 1 (5X)
 

*SUBCUT TISSUE (20) (49) (50)
 

EPIDERMAL INCLUSION CYST 1 (2*)
 

DERMAL INCLUSION CYST 1 (2%)
 

RESPIRATORY SYSTEM 

#LUNG 

CONGESTION, NOS 

BRONCHOPN£UMONIA, NOS 

PNEUMONIA, CHRONIC MURINE 

FOAM-CELL 

HYPERPLASxA, ADENOMATODS 

(18) 

12 (67%) 

(48) 

2 (4X) 

37 (77%) 

2 (4%) 

4 (8%) 

(46) 

1 (2X) 

34 (74X) 

HEHATOPOIETIC SYSTEM
 

•SPLEEN	 (19) (48) (47)
 

HEMOHRHAGiC CYST 1 (2%)
 

HEMOSIDEROSIS 1 (2%)
 

tMESENTERIC	 L. NODE (19) (49) (47)
 

CYTOLOGIC ALTERATION, NOS 1 (5X)
 

CIECULATOBY SYSTEM
 

•HEART/ATRIUM	 (20) (49) (48)
 

THBOMBQSIa, NOS 1 (2%)
 

#	 NUMBE R OF ANIMAL S WITH TISSUE EXAMINED MICROSCOPICALLY
 

*	 NUMBE R OF ANIMAL S NECROPSIED
 

3 50	 ANIMAL S WERE INITIALLY IN THE STUDY, BUT ONE ANIHAL HAS FOUND TO BE A HALE
 

IN A FEMAL E GROUP.
 

64
 
TABLE C2. FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

•MYOCARDIUM
 

INFLAMMATION, ACUTE FOCAL
 

FIBROSIS
 

*PULMONARY ARTERY
 

MINERALIZATION
 

DIGESTIVE SYSTEM
 

tSALIVARY GLAND
 

ATROPHY, jJOS
 

ATROPHY, uIFFUSE
 

*LIVEE
 

ABSCESS, NOS
 

GRANULOMA, NOS
 

DEGENERATION, NOS
 

NECROSIS, FOCAL
 

METAMORPHOSIS FATTY
 

HEFATOCYUHEGALY
 

GLYCOGENIC CELL
 

HYPERPLASiA, NODULAR
 

HYFEHPLASIA, NOS
 

HYPERPLASIA, FOCAL
 

ANGIECTASj.5
 

fLIVER/KUPFFER CELL
 

CYTOPLASMiC VACUOLIZ ATION
 

*BILE DUCT
 

HYPERPLASIA, NOS
 

tPANCEEAS
 

ATROPHY, NOS
 

AT&OPHY. FOCAL
 

ATROPHY, DIFFUSE
 

HYPERPLASIIC NODULE
 

*SMALL INTESTINE
 

PIGMENTATION, NOS
 

•LARGE	 IN1ESI1NE
 

NEM1TODIA3IS
 

MATCHED
 

CONTROL
 

(20)
 

12 (60S)
 

(20)
 

(18)
 

1 (6%)
 

(20)
 

3 (15X)
 

3 (15X)
 

12	 (60%)
 

(20)
 

1 (5%)
 

(20)
 

7 (35%)
 

(18)
 

(17)
 

(18)
 

6 (33%)
 

LOW DOSE
 

(49)
 

28 (57%)
 

(49)
 

1 (2%)
 

(48)
 

(48)
 

1 (2X)
 

3 (6%)
 

3 (6«)
 

2 {<**)
 

31 (65%)
 

(48)
 

(49)
 

1 (2%)
 

(48)
 

6 (13%)
 

1 (2%)
 

(44)
 

43 (98*)
 

(46)
 

6 (13X)
 

HIGH DOSE
 

(48)
 

1 (2%)
 

18 (39%)
 

(50)
 

(45)
 

2 (4%)
 

(48)
 

1 (2%)
 

4(8%)
 

1 (2*)
 

3 (6%)
 

1 (2%)
 

1(2%)
 

1 (2%)
 

1 (2%)
 

29 (60S)
 

1 (2%)
 

(48)
 

(50)
 

3 (6%)
 

(46)
 

1 (2X)
 

4 (9%)
 

1 (2X)
 

(47)
 

43 (91X)
 

(47)
 

8 J17X)
 

I NUMBEH OF ANIMALS KITH 1ISSUE EXAMINED HICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIED
 

65
 
TABLE C2. FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

— ————— . , ._,__,_. _ ,. „, , ._ ,_ ._ _ ..,,- I—.,.
 

MATCHED
 

CONTROL LOW DOSE HIGH DOSE
 

URINARY SYSTEM
 

•KIDNEY	 (20) (49) (48)
 

INFLAMMATION, NOS 1 (2%)
 

ABSCESS, NOS 1 (2%)
 

INFLAMMATION, CHRONIC 18 (90S) 45 (92X) 46 (96X)
 

DEGENERATION, HYALINE 1 (2*)
 

•KIDNEY/TUEULE	 (20) (49) (48)
 

NEPHROSIS, NOS 1 (2X)
 

NECROSIS, NOS 1 (5%) 1 (2%)
 

PIGMENTATION, NOS 2 («)
 

•URINARY	 BLADDER (15) (43) (44)
 

METAMORPHOSIS FATTY 1 (2X)
 

LIPOIDOSIi 1 (2%)
 

ENDOCRINE SYSiEB
 

•PITUITARY	 (18) (48) (45)
 

CYST, NOS 2 (11%) 4 (8X) 9 (20X)
 

HEMORRHAGiC CYST 2 (4X)
 

•ADRENAL	 (19) (48) (48)
 

HEMOHRHAGri 1 (2X)
 

LIPOIDOSIo 1 (5X) 1 (2%) 1 (2X)
 

•ADRENAL CORiEX (19) (48) (48)
 

LIPOIDOSIS 1 (2X)
 

HYPERPLASj-A, FOCAL 1 (2X)
 

•THYROID	 (17) (44) (47)
 

HYPERPLAiUA, FOCAL 1 (2X)
 

HYPERPLASIA, C-CELL 1 (6X) 3 (7X) 1 (2X)
 

HYPERPLASIA, FOLLICULAR-CELL 2 (4X)
 

•PANCREATIC iSLETS (18) (48) (46)
 

ATROPHY, FOCAL 1 (6%)
 

HYPERPLASxA, NOS 1 (2%)
 

REPRODUCTIVE SYSTEM
 

*MAMMARY GLAND (20) (49) (50)
 

DILATATION/DUCTS 4 (8%) 2 <4X)
 

• NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 * NUMBER OF ANIMALS NECROPSIED
 

66
 
TABLE C2. FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

f UTERUS
 

THROMBUS, ORGANIZED
 

BLOOD CLOI, POSTMORTEM
 

PYOMETRA
 

NECROSIS, NOS
 

tCERVIX UTERI
 

INFLAMMATION, NOS
 

« UTERUS/ EN DOM ETRIUM
 

INFLAMMATION, NOS
 

INFLAMMATION, ACUTE
 

HYPERPLAEIA, NOS
 

HYPERPLASxA, CYSTIC
 

*OVARY
 

FOLLICULA* CYST, NOS
 

INFLAMMATION, ACUTE
 

DESENERATiON, NOS
 

NECROSIS, NOS
 

NERVOUS SYSTEM
 

#BRAIB/«ENINuES
 

INFLAMMATION, ACUTE FOCAL
 

tBRAIN
 

DEKYELINIZATION
 

SPECIAL SENSE ORGANS
 

NONE
 

MUSCULOSKELETAL SYSTEM
 

NONE
 

BODY CAVITIES
 

*PLEUP.A
 

FOAM-CEIL
 

MATCHED 

CONTROL LOW DOSE HIGH DOSE 

(17) (47) (48) 

1 (2%) 

(17) (47) (48) 

1 (2%) 

(17) (47) (48) 

1 (6X) 

1 (2*) 

2 (4X) 

2 (H%) 

2 (4X) 

1 16%) 2 (<*X) 1 (2X) 

(17) (47) (48) 

1 (2X) 

1 (2X) 

1 (2%) 

(19) (49) (47) 

1 (2X) 

(19) (49) (47) 

1 (5X) 

(20) (49) (50)
 

2
 

* NUMBER OF ANIMALS WITH TISSUE EX&HINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIBD
 

67
 
TABLE C2. FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

MATCHED 

CONTROL LOW DOSE HIGH DOSE 

*MESENTER Y 

ARTERIOSCLEROSIS, NOS 

(20) (U9)

1 (2X) 

 (50) 

A LL OTHER SYSIEMS 

NON E 

SPECIAL MORPHOLOGY SUMMARY 

AUTO/NECROPSY/NO H1STO 

# NUMBER OF ANIMALS HUH TISSUE EXAMINED

* NUMBER OF ANIMALS NECROPSIED 

 MICROSCOPICALLY 

68
 
APPENDIX D
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS
 

IN MICE FED 4-AMINO-2-NITROPHENOL IN THE DIET
 

69
 

TABLE 01.

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN MALE MICE 

FED 4-AMINO-2-NITROPHENOL IN THE DIET 

ANIMALS INITIALLY IN STUDY
 

JNIMAL3 NECEOPSIED
 

ANIMALS EXAMINED HISTOPATHOLOGICALLY
 

INTEGUMENTARY SYSTEM
 

NONE
 

RESPIRATORY SYSTEM
 

tLUNG
 

CONGESTION. CHRONIC PASSIVE
 

INFLAMMATION, INTERSTITIAL
 

PNEUMONIA, ASPIRATION
 

PNSUHONIA, CHRONIC MURINE
 

HYPERPLASJ.A, ADENOHATOUS
 

HEMATOPOIETIC SYSTEM
 

•SPLEiN
 

DEGENERATION, HYALINE
 

NECROSIS, CASEOUS
 

ANGIECTASj.5
 

HYPERPLAS1A. LYBPHOID
 

HErtATOPOIiSIS
 

#MESENTERIC L. NODE
 

INFLAMMATION, HEHORRHAGIC
 

INFLAMMATION, GRANULOMATOOS
 

CIHCOLATORY 3YST2H
 

*PULMONAEY AHTERY
 

FIBROSIS
 

D1GEST.IVE SYSfEM
 

•LIVER
 

NECROSIS^ _FOCAL
 

MATCHED
 

CONTROL
 

20
 

20
 

20
 

(20)
 

2 (10X)
 

(20)
 

1 (5X)
 

1 (5X)
 

1 (5%)
 

(19)
 

1 (5S)
 

(20)
 

(20)
 

1 (5X)
 

LOW DOSE HIGH DOSE
 

50 50
 

50 50
 

50 50
 

(49)	 (48)
 

1 (2*)
 

1 <2X)
 

1 (2X) 1 (2X)
 

11	 (22X) 2 (4X)
 

1 (2X) 1 (2%)
 

(48) (47)
 

1 (2X)
 

<2X)
 

(50)	 (48)
 

1 (2X)
 

(50)	 (50)
 

1 (2X)
 

(50)	 (49)
 _ 3 (6X1
 

*	 NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

*	 NUMBER OF ANIMALS NECROPSIED
 

71
 
TABLE 01. MALE MICE: NONNEOPLASTIC LESIONS (CONTINUED) 

MATCHED 

CONTROL LOW DOSE HIGH DOSE 

NECROSIS, CASEOUS 1 (5X)

 

INFARCT, NOS 1 (5X) 3 (6X)

 

METAHOEPHuSIS FATTY 1 (5X) 4 (8X) 2 (4X)

 

NUCLEAfi ENLARGEMENT 1 (2X) 

CYrOPLASIUC VACUOLIZATION 1 (5X)

 

HEPATOCYTOMEGAL Y 1 (2X)

 

HEMATOPOIiSIS 2 (<*X)

 

tLIVER/PERIPORTA L (20) (50) (49) 

MONOCYTOSiS 1 (2X) 

#SMAL L INTESTIN E (20) (49) (47) 

PIGMENTATION , NOS 46 (94X) 43 (91X) 

#PEYE3S PATCJ (20) (49) (47) 

HYPBRPLASiA , NOS 1 (2X) 1 12*) 

ICOLON (20) (50) (48) 

NEMATODIASIS 2 (10X) 9 (18X) 10 (21X) 

PAHASIlISil 1 (5X) 

URINARY SYSTEM
 

*KIDNSY (20) (50) (49)
 

INFLAMMATION, CHRONIC 1 (5X) 2 (UX) 3 (6X)
 

INFAHCT, HOS 1 (2X)
 

#URINARY BLADDER (18) (46) (48)
 

INFLAMMATION, CHRONIC 1 (2X)
 

ENDOCRINE SYSIEM
 

tPANCHBATIC J.SLETS . (19) («9) (47)
 

HYPERTROPdY, NOS 1 (2X)
 

HYPBRPLASxA, NOS 3 (16X) 8 (16X)
 

REPRODUCTIVE JYSTEM
 

tTESTIS (18) (50) (49)
 

CALCIFICATION, NOS 1 (6X)
 

NERVOUS SYSTEM
 

t BRAIN (19) (50) (50)
 

HINEBALIZATION 1 (5X) 1 (2X1 1 (2X1
 

* NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* NUMBSB OF ANIMALS NECROPSIED
 

72
 
TABLE 01. MALE MICE: NONNEOPLASTIC LESIONS (CONTINUED) 

MATCHED 

CONTROL LOW DOSE HIGH DOSE 

SPECIAL

NON E 

 SENSE ORGAN S 

HUSCULOSKELETA L

NON E 

 SYSTEM 

EODY CAVITIES 

•MESENTER Y

NECROSIS , FAT

 (20) (50)

 1 (2«)

 (50) 

2 (H%) 

ALL OTHER SYSTEMS 

NONE 

SPECIAL MORPHOLOGY SUMMARY 

NO LESICN REPORTED 3

* NUMBKR OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY * NUMBER OF ANIMALS NECROPSIED 

2 

73
 
TABLE 02.

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN FEMALE MICE 

FED 4-AMINO-2-NITROPHENOL IN THE DIET 

MATCHED
 

CONTROL
 

iNIMALS INITIALLY IN STUD* 20
 

{NIHALS NECROPSIED 20
 

ANIMALS EXAMINED HI SO CO PATHOLOGICALLY 20
 

INTEGUMENTARY SYSTEM
 

NONE
 

BESPIRATORY SiSTEM
 

ILUNG
 

HEMORRHAGE
 

INFLAMMATION, INTERSTITIAL
 

PNEUMONIA, CHRONIC HURINE
 

PERIVASCULAR CUFFING
 

HEflATOPOIETIC SYSTEM
 

ISPLEUN
 

1NFARCI, NOS
 

HYPERPLASIA, NOS
 

HYPERPLASJ.A, LYMPHOID
 

*LYMPH NODE
 

HYPEBPLASIA, NOS
 

*MANDIBULAR L. NODE
 

HYPEKPLASJ.A, LYBPHOID
 

tMESENTEBIC *.. NODE
 

INFLAMMATION, GflANOLOMATOtJS
 

CIRCULA.TOBY SiSTEM
 

•CARDIAC	 VALVE
 

INFLAMMATION. NOS
 

(20)
 

1 (5X)
 

5 (25X)
 

(20)
 

(20)
 

(20)
 

(20)
 

1 (5X)
 

(19)
 

LOW DOSE
 

50
 

50
 

50
 

(49)
 

2 (4%)
 

15 (31X)
 

2 (UX)
 

(50)
 

1 (2X)
 

1 (2X)
 

1 (2X)
 

(49)
 

1 (2X)
 

(49)
 

1 (2X)
 

(49)
 

(48)
 

HIGH DOSE
 

50
 

50
 

50
 

(50)
 

10 (20X)
 

1 (2X)
 

(50)
 

(48)
 

1 (2X)
 

(48)
 

(48)
 

(50)
 

1 (2X1
 

* NUMBER OF ANIMALS KITH TISSUE EXAMINED MICROSCOPICALLY
 * NUMBER OF ANIMALS NECROPSIED
 

74
 
TABLE 02. FEMALE MICE: NONNEOPLASTIC LESIONS (CONTINUED) 

MATCHED
 

CONTROL LOW DOSE
 

*PULMONARY AHTSRY (20) (50)
 

HYPERPLASiA, LYMPHOID 1 (2%)
 

DIGESTIVE SYSiEM
 

•SALIVARY	 GLAND (19) (49)
 

PERIVASCULAR CUFFING 1 (2%)
 

*LIVER (20) (50)
 

INFLAMMATION, ACUTE FOCAL 1 (5%)
 

PERIVASCULAR CUFFING 1 (2%)
 

NECROSIS, FOCAL 1 (2%)
 

METAMORPHOSIS FATTY 1 (5%)
 

•PANCREAS	 (20) (49)
 

CYST, NOS 1 (2%)
 

INFLAMMATION, SUPPURATIVE 1 (5X)
 

•SBALL INTESTINE (19) (48)
 

PIGMENTATION, NOS 43 (90*)
 

IPEYEBS PATCd (19) (48)
 

HYPERPLASiA, LYMPHOID
 

•COLON	 (19) (48)
 

NEMAlODIAalS 1 (5X) 2 (4X)
 

URINARY SYSTEM
 

•KIDNEY	 (20) (49)
 

INFLAMMATION, CHRONIC 1 (2X)
 

PEBIVASCUi,AR CUFFING
 

NEPHROSIS, HEHOGLOBINURIC
 

METAPLASIA, OSSEOUS
 

ENDOCRINE SYSIEM
 

•PITUITARY	 (17) (38)
 

ANGIECTASIS
 

•ADRENAL	 (19)
 

CYST. NOS 1 (2X)
 

* NUMBER OF AKIMALS WITH TISSDE EXAMINED HICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIED
 

HIGH DOSE
 

(50)
 

(46)
 

(50)
 

1 (2X)
 

(49)
 

(47)
 

42 (89X)
 

(47)
 

1 (2X)
 

(49)
 

(50)
 

3 (6X)
 

1 (2X)
 

(44)
 

1 (2X)
 

(46)
 

75
 
TABLE D2. FEMALE MICE: NONNEOPLASTIC LESIONS (CONTINUED) 

MATCHED
 

CONTROL LOW DOSE HIGH DOSE
 

L1POIDOSIS 1 (2%)
 

tPANCREATIC ISLETS (20) (49) (49)
 

HYPERTROPHY, DOS 1 (2%)
 

REPRODUCTIVE SYSTEM
 

f UTERUS (19) (48) (49)
 

HYDROMEIRA 4 (21X) 8 (17%) 13 (27%)
 

CYST, NOS 1 (2%)
 

THROMBUS, ORGANIZED 1 (2%)
 

INFLAMHAl^ON, NOS 1 (2%)
 

PYOME1HA 1 (2%)
 

tUTEHUS/ENDOi-JETRIUM (19) (48) (49)
 

HYPERPLASJ.A, NOS 2 (4%)
 

HYPERPLASIA, CYSTIC 1 (5%) 5 (10*) 1 (2X)
 

#OVARY (18) (41) (45)
 

CYST, NOS 2 (11%) 2 (5S) 2 (4%)
 

FOLLICULA8 CYST, NOS 1 (6%) 1 (2%)
 

NERVOUS SYSTEM
 

# BRAIN (19) (49) (49)
 

MINERALIZATION 3 (6%)
 

SPECIAL SENSE ORGANS
 

NONE
 

MUSCOLOSKELETAL SYSTEM
 

NONE
 

EODY CAVITIES
 

*MZSENTERY (20) (50) (50)
 

NECROSIS, FAT 1 (2%)
 

# NUMBER OF ANIMALS WITH TISSUE EXABINED MIC80SCOPICALLY
 

* NUMBER OF ANIMALS NBCROPSIED
 

76
 
TABLE 02. FEMALE MICE: NONNEOPLASTIC LESIONS (CONTINUED) 

MATCHED 

CONTROL LOW DOSE HIGH DOSE 

ALL OTHER SYSTEMS 

*HULTIPLE ORGAN S (20) (50) (50) 

POSTMORTEM CHANG E 1 (5X)

 

SPECIAL MORPHOLOGY SUMMARY
 

NO LESION REPORTED
 

* NUMBER OF ANIMALS WITH TISSUE EXAHINED MICROSCOPICALLY
 

* NUMBER OF ANIMALS NECROPSIED
 

77
 

APPENDIX E
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS IN
 

RATS ADMINISTERED 4-AMINO-2-NITROPHENOL IN THE DIET
 

79
 

00 

Table El. Analyses of

Administered

Topography: Morphology


Hematopoietic System: tialignant
 

Lymphoma, Lymphocytic Leukemia,
 

Undif f erentiated Leukemia, or
 

Leukemia, NOSb


P Valuesc'd


Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor


Hematopoietic System:
 

All Lymphomas or Leukemias*3


P Valuesc>d


Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor


 the Incidence of Primary Tumors in Hale Rats 

 4-Amino-2-Nitrophenol in the Diet3 

Matched Low High
 

 Control Dose Dose
 

 3/20 (15) 11/50 (22) 10/50 (20)
 

 N.S. N.S. N.S.
 

1.467 1.333
 

0.450 0.398
 

7.594 7.002
 

 105 86 83
 

 3/20 (15) 13/50 (26) 11/50 (22)
 

 N.S. N.S. N.S.
 

1.733 1,467
 

0.556 0.450
 

8.773 7.594
 

 105 86 73
 
Table El. Analyses of the Incidence of Primary Tumors in Male Rats
 

Administered 4-Amino-2-Nitrophenol in the Dieta
 

(continued)
 

Topography: Morphology


Urinary Bladder: Transitional-cell
 

Carcinomab


P Valuesc»d


Departure from Linear Trend6


Relative Riskf
 

Lower Limit
 

Upper Limit
 

00
 

NJ	 Weeks to First Observed Tumor


Urinary Bladder: Transitional-cell
 

Papillomab


P Valuesc»d


Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor


Matched
 

 Control
 

 0/15 (0)
 

 P < 0.001
 

P = 0.030
 

—
 

 0/15	 (0)
 

 N.S.
 

—
 

Low High 

Dose Dose 

0/46 (0) 11/39 (28) 

N.S. P = 0.018 

— Infinite 

1.367 

— Infinite 

90 

0/46 (0) 2/39 (5) 

N.S. N.S. 

Infinite 

0.120 

— Infinite 

90 
Table El. Analyses of the Incidence of Primary Tumors in Male Rats
 

Administered 4-Amino-2-Nitrophenol in the Dieta
 

(continued)
 

Matched Low High
 

Topography; Morphology Control Dose Dose
 

Pituitary: Chromophobe Adenoma" 2/15 (13) 4/40 (10) 8/39 (21)
 

P Values0>d N.S. N.S. N.S.
 

Relative Riskf 0.750 1.538
 

Lower Limit 0.125 0.366
 

Upper Limit 7.797 13.883
 

Weeks to First Observed Tumor 96 105 70
 

oo
 Thyroid: C-cell Adenoma or
 

Carcinoma^ 3/18 (17) 2/42 (5) 1/44 (2)
 

P Valuesc»d P = 0.043(N) N.S. N.S.
 

Relative Riskf 0.286 0.136
 

Lower Limit 0.026 0.003
 

Upper Limit 2.323 1.592
 

Weeks to First Observed Tumor 99 105 105
 
Table El. Analyses of

Administered

 the Incidence of Primary Tumors in

 4-Amino-2-Nitrophenol in the Dieta 

 Male Rats 

(continued) 

Topography: Morphology

Matched 

 Control 

Low 

Dose 

High 

Dose 

Pancreatic Lslets:

Adenomab

 Islet-cell 

 2/18 (11) 1/46 (2) 1/49 (2) 

P Valuesc»d N.S. N.S. N.S. 

Relative Riskf 

Lower Limit 

Upper Limit 

0.196 

0.004 

3.586 

0.184 

0.003 

3.372 

Weeks to First Observed Tumor 105 105 105 

oo 

Testis: Interstitial-cell Tumorb 15/17 (88) 50/50 (100) 41/50 (82) 

P Valuesc»d N.S. N.S. N.S. 

Departure from Linear Trend6 P = 0.011 

Relative Riskf 

Lower Limit 

Upper Limit 

1.133 

0.996 

Infinite 

0.929 

0.798 

1.283 

Weeks to First Observed Tumor 90 86 78 
Table El. Analyses of the Incidence of Primary Tumors in Male Rats
 

Administered 4-Amno-2-Nitrophenol in the '3ieta
 

(continued)
 

a

Dosed groups received 1,250 or 2,500 ppm.
 

^Number of tumor-bearing animals /number of animals examined at site (percent).
 

c

Beneath the incidence of tumors in the control group is the probability level for the Cochran-


Armitage test when P < 0.05; otherwise, not significant (N.S.) is indicated. Beneath the
 

incidence of tumors in a dosed group is the probability level for the Fisher exact test for
 

the comparison of that dosed group with the matched-control group when P < 0.05; otherwise,
 

not significant (N.S.) is indicated.
 

"A negative trend (N) indicates a lower incidence in a dosed group than in a control group.
 

o° eThe probability level for departure from linear trend is given when P < 0.05 for any comparison.
 

95% confidence interval of the relative risk between each dosed group and the control
 

group.
 
Table E2. Analyses of the Incidence of Primary Tumors in Female Rats
 

Administered 4-Amino-2-Nitrophenol in the Diet3
 

Topography: Morphology
 

Hematopoietic System: Malignant
 

Lymphoma, Lymphocytic Leukemia,
 

Undif ferentiated Leukemia, or
 

Leukemia, NOSb
 

P Values0>d
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

CD
 

Ov
 Weeks to First Observed Tumor
 

Hematopoietic System: All
 

Lymphomas or Leukemias^
 

P Valuesc»d
 

Relative Risk^
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Matched


Control


2/20 (10)


N.S.


100


2/20 (10)


N.S.


100


 Low
 

 Dose
 

 6/49 (12)
 

 N.S.
 

1.224
 

0.248
 

11.802
 

 86
 

 6/49 (12)
 

 N.S.
 

1.224
 

0.248
 

11.802
 

- 86
 

High
 

Dose
 

2/50 (4)
 

N.S.
 

0.400
 

0.032
 

5.277
 

63
 

4/50 (8)
 

N.S.
 

0.800
 

0.128
 

8.436
 

63
 
Table E2. Analyses of the Incidence of Primary Tumors in Female Rats 

Administered 4-Amino-2-Nitrophenol in the Dieta 

(continued) 

Matched Low High
 

Topography: Morphology Control Dose Dojse
 

Urinary Bladder: Transitional-cell
 

Carcinomab ' 0/15 (0) 1/43 (2) 2/44 (5)
 

P Values0 >d N.S. N.S. N.S.
 

Relative Risk^ Infinite Infinite
 

Lower Limit 0.020 0.107
 

Upper Limit Infinite Infinite
 

Weeks to First Observed Tumor 79 61
 00 

Pituitary: Chromop'nobe Adenoma0 8/18 (44) 26/48 (54) 20/45 (44)
 

P Valuesc»d N.S. N.S. N.S.
 

Relative Risk^ 1.219 1.000
 

Lower Limit 0.695 0.546
 

Upper Limit 2.571 2.192
 

Weeks to First Observed Tumor 72 76 92
 
Table E2. Analyses of

Administered

(continued) 

Topography: Morphology

Thyroid: C-cell Adenomab

>d P Valuesc

Relative Riskf

 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor

oo 

CO Mammary Gland: Fibroadenoma or 

Adenoma, NOSb

P Valuesc»d

Relative Riskf

 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor

 the Incidence of Primary Tumors in Female Rats 

 4-Amino-2-Nitrophenol in the Dieta 

Matched Low High 

 Control Dose Dose 

 1/17 (6) 2/44 (5) 0/47 (0) 

 N.S. N.S. N.S. 

0.773 0.000 

0.044 0.000 

44.565 6.754 

 105 105 — 

 1/20 (5) 4/49 (8) 6/50 (12) 

 N.S. N.S. N.S. 

1.633 2.400 

0.179 0.325 

78.704 108.021 

 105 105 92 
Table E2. Analyses of the Incidence of Primary Tumors in Female Rats
 

Administered 4-Amino-2-Nitrophenol in the Dieta
 

(continued)
 

Matched Low High
 

Topography: Morphology Control Dose Dose
 

Mammary Gland: Fibroadenoma'3 1/20 (5) 3/49 (6) 5/50 (10)
 

P Values0 »d N.S. N.S. N.S.
 

Relative Risk^ 1.224 2.000
 

Lower Limit 0.108 0.249
 

Upper Limit 62.958 92.596
 

Weeks to First Observed Tumor 105 105 92
 

oo aDosed groups received 1,250 or 2,500 ppm.
 VO
 

"Number of tumor-bearing animals /number of animals examined at site (percent).
 

c

Beneath the incidence of tumors in the control group is the probability level for the Cochran-


Armitage test when P < 0.05; otherwise, not significant (N.S.) is indicated. Beneath the
 

incidence of tumors in a dosed group is the probability level for the Fisher exact test for
 

the comparison of that dosed group with the matched-control group when P < 0.05; otherwise,
 

not significant (N.S.) is indicated.
 

"A negative trend (N) indicates a lower incidence in a dosed group than in a control group.
 

e

The probability level for departure from linear trend is given when P < 0.05 for any comparison.
 

95% confidence interval of the relative risk between each dosed group and the control
 

group.
 

APPENDIX F
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS IN
 

MICE ADMINISTERED 4-AMINO-2-NITROPHENOL IN THF, DIET
 

91
 

Table Fl. Analyses of the Incidence of Primary Tumors in Male Mice
 

Administered 4-Amino-2-Nitrophenol in the Diet3
 

Topography: Morphology
 

Lung: Alveolar/Bronchiolar Adenoma^
 

P Values0 >d
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Lung: Alveolar/Bronchiolar
 

Carcinoma"
 

P Valuesc'd
 

Departure from Linear Trend6
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor


Matched
 

Control
 

2/20 (10)
 

N.S.
 

105
 

3/20 (15)
 

P = 0.005(N)
 

P = 0.015
 

 105
 

Low
 

Dose
 

10/49 (20)
 

N.S.
 

2.041
 

0.498
 

18.154
 

70
 

0/49 (0)
 

P = 0.022(N)
 

0.000
 

0.000
 

0.673
 

High
 

Dose
 

7/48 (15)
 

N.S.
 

1.458
 

0.316
 

13.664
 

105
 

0/48 (0)
 

P = 0.023(N)
 

0.000
 

0.000
 

0.686
 
Table Fl. Analyses of the Incidence of Primary Tumors in Male Mice
 

Adiiinistered 4-Amino-2-Nitrophenol in the Diet3
 

(continued)
 

Topography: Morphology


Lung: Alveolar /Bronchiolar
 

Adenoma or Carcinoma*3


P Valuesc»d


Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor


rlematopoietic System: Lymphoraa^


P Valuesc'd


Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor


Matched Low 

 Control Dose 

 5/20 (25) 10/49 (20) 

 N.S. N.S. 

0.816 

0.302 

2.740 

 105 70 

 1/20 (5) 5/50 (10) 

 N.S. N.S. 

2.000 

0.249 

92.596 

 105 83 

High
 

Dose
 

7/48 (15)
 

N.S.
 

0.583
 

0.187
 

2.109
 

105
 

4/50 (8)
 

N.S.
 

1.600
 

0.175
 

77.169
 

93
 
Table Fl. Analyses of

Administered

 the Incidence of Primary Tumors in

 4-Amino-2-Nitrophenol in the Diet3 

 Male Mice 

(continued) 

Topography: Morphology

Matched 

 Control 

Low 

Dose 

High 

Dose 

Hematopoietic System: 

Lymphoma or Leukemia" 2/20 (10) 5/50 (10) 4/50 (8) 

P Valuesc » d ' N.S. N.S. N.S. 

Relative Riskf 

Lower Limit 

Upper Limit 

1.000 

0.184 

10.007 

0.800 

0.128 

8.436 

Weeks to First Observed Tumor 69 83 93 

<-" Liver: Hepatocellular Carcinoma^3 3/20 (15) 7/50 (14) 7/49 (14) 

P Valuesc'd N.S. N.S. N.S. 

Relative Riskf 

Lower Limit 

Upper Limit 

0.933 

0.245 

5.215 

0.952 

0.250 

5.317 

Weeks to First Observed Tumor 95 87 93 
Table Fl. Analyses of the Incidence of Primary Tumors in Male Mice

 

Administered 4-Amino-2-Nitrophenol in the Diet3

 

(continued) 

Matched Low High 

Topography: Morphology Control Dose Dose 

Liver: Hepatocellular Adenoma
 

or Carcinoma'-1 6/20 (30) 18/50 (36) 19/49 (39)
 

P Valuesc>d N.S. N.S. N.S.
 

Relative Riskf 1.200 1.293
 

Lower Limit 0.557 0.607
 

Upper Limit 3.238 3.452
 

Weeks to First Observed Tumor 95 78 93
 

cr> aDosed groups received 1,250 or 2,500 ppm.
 

^Number of tumor-bearing animals/number of animals examined at site (percent).
 

G

Beneath the incidence of tumors in the control group is the probability level for the Cochran-


Armitage test when P < 0.05; otherwise, not significant (N.S.) is indicated. Beneath the
 

incidence of tumors in a dosed group is the probability level for the Fisher exact test for
 

the comparison of that dosed group with the matched-control group when P < 0.05; otherwise,
 

not significant (N.S.) is indicated.
 

"A negative trend (N) indicates a lower incidence in a dosed group than in a control group.
 

e

The probability level for departure from linear trend is given when P < 0.05 for any comparison.
 

•'-The 95% confidence interval of the relative risk between each dosed group and the control
 

group.
 
Table F2. Analyses of

Administered

Topography: Morphology


Lung: Alveolar /Bronchiolar
 

Adenomab


P Values0 >d


Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor


Hematopoietic System: Malignant
 

Lymphoma or Lymphocytic Leukemia"


P Value sc'd


Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor


 the Incidence of Primary Tumors in Female Mice 

 4-Amino-2-Nitrophenol in the Diet3 

Matched Low 

 Control Dose 

 2/20 (10) 3/49 (6) 

 N.S. N.S. 

0.612 

0.078 

6.996 

 105 105 

 3/20 (15) 10/50 (20) 

 N.S. N.S. 

1.333 

0.398 

7.002 

 85 76 

High
 

Dose
 

2/50 (4)
 

N.S.
 

0.400
 

0.032
 

5.277
 

105
 

8/50 (16)
 

N.S.
 

1.067
 

0.295
 

5.813
 

64
 
Table F2. Analyses of the Incidence of Primary Tumors in Female Mice 

Administered 4-Amino-2-Nitrophenol in the Diet3 

(continued) 

Matched Low High
 

Topography: Morphology Control Dose Dose
 

Thyroid: Follicular-cell Adenoma
 

or Papillary Adenoma'3 2/17 (12) 1/40 (3) 0/42 (0)
 

c d P Values » P = 0.035(N) N.S.
 N.S.
 

Relative Riskf 0.213 0.000
 

Lower Limit 0.004 0.000
 

Upper Limit 3.873 1.353
 

Weeks to First Observed Tumor 105 105
 —
 

o? aDosed groups received 1,250 or 2,500 ppm.
 

^Number of tumor-bearing animals/number of animals examined at site (percent).
 

G

Beneath the incidence of tumors in the control group is the probability level for the Cochran-


Armitage test when P < 0.05; otherwise, not significant (N.S.) is indicated. Beneath the
 

incidence of tumors in a dosed group is the probability level for the Fisher exact test for
 

the comparison of that dosed group with the matched-control group when P < 0.05; otherwise,
 

not significant (N.S.) is indicated.
 

d

A negative trend (N) indicates a lower incidence in a dosed group than in a control group.
 

e

The probability level for departure from linear trend is given when P < 0.05 for any comparison.
 

95% confidence interval of the relative risk between each dosed group and the control
 

group.
 
Review of the Bioassay of 4-Amino-2-Nitrophenol*
 

for Carcinogenicity
 

by the Data Evaluation/Risk Assessment Subgroup of the
 

Clearinghouse on Environmental Carcinogens
 

April 26, 1978
 

The Clearinghouse on Environmental Carcinogens was
 

established in May, 1978, in compliance with DHEW Committee
 

Regulations and the Provisions of the Federal Advisory
 

Committee Act. The purpose of the Clearinghouse is to
 

advise the Director of the National Cancer Institute (NCI)
 

on its bioassay program to identify and to evaluate chemical
 

carcinogens in the environment to which humans may be
 

exposed. The members of the Clearinghouse have been drawn
 

from academia, industry, organized labor, public interest
 

groups, State health officials, and quasi-public health and
 

research organizations. Members have been selected on the
 

basis of their experience in carcinogenesis or related
 

fields and, collectively, provide expertise in chemistry,
 biochemistry, biostatistics, toxicology, pathology, and
 

epidemiology. Representatives of various Governmental
 

agencies participate as ad hoc members. The Data Evaluation/
 

Risk Assessment Subgroup of the Clearinghouse is charged
 

with the responsibility of providing a peer review of
 

reports prepared on NCI-sponsored bioassays of chemicals
 

studied for carcinogenicity. It is in this context that
 

the below critique is given on the bioassay of 4-Amino-2­

Nitrophenol for carcinogenicity.
 

The primary reviewer said that the compound induced
 

bladder cancer in treated male rats and that the evidence
 

was suggestive for a similar effect in females. No carcin­ ogenic effect was observed among treated mice. After a
 

brief description of the experimental design, he opined that
 

the incidence of bladder cancer was not dose related, as
 

indicated in the report. He noted that the compound was
 

the only phenol he was aware of which was (systemically)
 

carcinogenic. Based on the relatively low incidence of
 

bladder cancer and long latent period, the primary reviewer
 

concluded that-the compound did not pose a carcinogenic
 

risk to humans.
 

The secondary reviewer agreed with the conclusion that
 

the compound was carcinogenic in rats. He said that a
 

conclusion on the carcinogenicity of il-Amino-2-Nitrophenol
 

in mice could not be made since it appeared that a maximum
 

tolerated dose was not achieved. Had higher doses been
 

99
 
administered, the elevated incidence of liver tumors in
 

treated male mice may have increased to a statistically
 

significant number. The secondary reviewer concluded
 

that 4-Amino-2-Nitrophenol poses a carcinogenic risk to
 

humans.
 

A Subgroup member disagreed with the primary reviewer's
 

conclusion with respect to human risk. Unlike the results
 

of this study, the primary reviewer argued that human
 

carcinogens induce a high yield of cancer in a relatively
 

short time in experimental animals.
 

In response to a question, a Program staff pathologist
 

said that the presence of bladder calculi in the rats were
 

not noted by the testing laboratories' pathologists. In
 

his experience, he added, bladder parasites have not been
 

found in rats used in bioassay studies. It was noted that
 

only one of the bladder tumors metastasized. The Program
 

staff pathologist continued that the finding was not unusual
 

since bladder tumors normally do not metastasize even when
 

induced by strong carcinogens. A Subgroup member suggested
 

that the conclusion on the carcinogenicity of 4-Amino-2­

Nitrophenol should be qualified, since the urine was not
 

analyzed for crystals.
 

A motion was approved unanimously that the report
 

on the bioassay of 4-Amino-2-Nitrophenol be accepted as
 

written.
 

Members present were:
 

Michael Shimkin (Acting Chairman), University of California
 

at San Diego
 

Joseph Highland, Environmental Defense Fund
 

George Roush, Jr., Monsanto Company
 

Louise Strong, University of Texas Health Sciences Center
 

John Weisburger, American Health Foundation
 

(Sidney Wolfe, Health Research Group, submitted a written review)
 

Subsequent to this review, changes may have been made
 

in the bioassay report either as a result of the review
 

or other reasons. Thus, certain comments and criticisms
 

reflected in the review may no longer be appropriate.
 

100
 

GOVERNMEN T PRINTIN G OFFICE : 1978-261-324/10 3
 
DHEW Publication No. (NIH) 78-1344