National Cancer Institute 

CARCINOGENESIS 

Technical Report Series 

No. 89 

1978 

BIOASSAY OF 

o-ANISIDINE HYDROCHLORIDE 

FOR POSSIBLE CARCINOGENICITY 

CAS No. 134-29-2 

NCI-CG-TR-89 

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

Public Health Service 

National Institutes of Health 


BIOASSAY OF
 

o-ANISIDINE HYDROCHLORIDE
 

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) 78-1339
 

BIOASSAY OF
 

o-ANISIDINE HYDROCHLORIDE
 

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
 

o-anisidine hydrochloride 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 environmental 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 o-anisidine hydrochloride was
 

conducted by EG&G Mason Research Institute, Worcester,
 

Massachusetts, initially under direct contract to NCI and
 

currently under a subcontract to Tracer Jitco, Inc., prime
 

contractor for the NCI Carcinogenesis Testing Program.
 

The bioassay was conducted under the supervision of Drs. A.
 

Handler^- and E. Smith^ and Mr. G. Wade^. NCI project officers
 

were Drs. E. Weisburger^, T. Cameron^, and N. P. Page^»^. The
 

program manager was Mr. J. Baker-'. Ms. A. Good-' supervised the
 

technicians in charge of animal care, and Ms. E. Zepp-* supervised
 

the preparation of the feed mixtures and collected samples of the
 

diets for analysis. Ms. D. Bouthot^ kept all daily records of
 

the test, and Ms. R. Monson^ prepared a draft of the experimental
 

design based on these records. Histopathologic examinations on
 

rats and mice were performed by Drs. D. S. Wyand-* and A.
 

ill
 
Russfield^, and the diagnoses included in this report represent
 

their interpretations.
 

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

Mason Research Institute, Rockville, Maryland". 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^, and dosed feed mixtures were analyzed by Dr.
 

M. Hagopian^. 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 Tracor 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. Squire10,
 

Dr. Jerrold M. Ward, and Dr. Carrie E. Whitmire.
 

with Abcor, Inc., 850 Main Street, Wilmington,
 

Massachusetts.
 

Now with the University of Massachusetts Medical Center,
 

55 Lake Avenue, Worcester, Massachusetts.
 

Mason Research Institute, 57 Union Street, Worcester,
 

Massachusetts.
 

iv
 
^Carcinogenesis Testing Program, Division of Cancer Cause and
 

Prevention, National Cancer Institute, National Institutes of
 

Health, Bethesda, Maryland.
 

with the Environmental Protection Agency, 401 M Street,
 

S.W., Washington, B.C.
 

6

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

Rockville, Maryland.
 

7

Tracor 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 Instiutes of Health, Bethesda, Maryland.
 

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

Missouri.
 

with the Division of Comparative Medicine, Johns Hopkins
 

University School of Medicine, Traylor Building, Baltimore,
 

Maryland.
 

V 

SUMMARY
 

A bioassay of o-anisidine hydrochloride for possible carcinogenicity was conducted by administering the test chemical in feed
 

to Fischer 344 rats and B6C3F1 mice.
 

Groups of 55 rats of each sex and 55 mice of each sex were
 

administered o-anisidine hydrochloride at one of the following
 

doses, either 5,000 or 10,000 ppm for rats and either 2,500 or
 

5,000 ppm for mice, for 103 weeks, then observed for 1 or 2
 

additional weeks. Controls consisted of groups of 55 untreated
 

rats of each sex and 55 untreated mice of each sex. All
 

surviving rats were killed at 103-107 weeks, and all surviving
 

mice at 104 or 105 weeks.
 

Mean body weights of the dosed male and female rats and mice were
 

lower than those of the corresponding controls throughout the
 

bioassay. Bloody exudates and stained fur in the urogenital area
 

were noted in many dosed animals. Sufficient numbers of animals
 

were at risk in the mice, but not in the rats, for development of
 

late-appearing tumors; however, survival in the rats was 80% or
 

more at week 52.
 

Transitional-cell carcinomas or papillomas of the urinary bladder
 

occurred at statistically significant incidences (P < 0.001) in
 

the low- and high-dose groups of rats (males: controls 0/51,
 

low-dose 52/54, high-dose 52/52; females: controls 0/49, low-dose
 

46/49, high-dose 50/51) and in high-dose groups of mice (males:
 

controls 0/48, low-dose 2/55, high-dose 22/53; females: controls
 

0/50, low-dose 1/51, high-dose 22/50); the incidences also had
 

significant dose-related trends (P < 0.001) in both species.
 

These lesions were observed as early as week 36 in female rats,
 

week 40 in male rats, and week 45 in male mice. Transitional-


cell carcinomas of the pelvis of the kidney occurred with a
 

significant dose-related trend (P = 0.005) in the male rats, and
 

the incidence in the high-dose group was significantly higher
 

(P = 0.006) than that in the control group (controls 0/53, low-


dose 3/55, high-dose 7/53); all rats having this tumor also had a
 

transitional-cell carcinoma of the urinary bladder. Only one
 

animal in the control groups of rats or mice had any tumor of the
 

vii
 
urinary system (a transitional-cell papilloma of the pelvis of
 

the kidney in a male mouse).
 

Follicular-cell tumors of the thyroid (carcinomas, cystadenocarcinomas, adenomas, cystadenomas, and papillary cystadenomas)
 

occurred at statistically significant incidences (P _< 0.005) in
 

low- and high-dose groups of male rats (controls 0/53, low-dose
 

7/40, high-dose 6/40); the incidences also had a dose-related
 

trend (P = 0.009). These tumors did not occur at significant
 

incidences in dosed groups of female rats.
 

It is concluded that under the conditions of this bioassay,
 

o-anisidine hydrochloride was carcinogenic for Fischer 344 rats
 

and B6C3F1 mice, inducing transitional-cell carcinomas or
 

papillomas of the bladder in both rats and mice and in both sexes
 

of each species, transitional-cell carcinomas of the pelvis of
 

the kidney in male rats, and follicular-cell tumors of the
 

thyroid in male rats.
 

viii
 
TABLE OF CONTENTS
 

Page
 

I. Introduction 1
 

II. Materials and Methods 3
 

A. Chemical 3
 

B. Dietary Preparation 4
 

C. Animals 4
 

0. Animal Maintenance 5
 

E. Subchronic Studies 8
 

F. Chronic Studies 9
 

G. Clinical and Pathologic Examinations 9
 

H. Data Recording and Statistical Analyses 13
 

III. Results - Rats 19
 

A. Body Weights and Clinical Signs (Rats) 19
 

B. Survival (Rats) 21
 

C. Pathology (Rats) 21
 

D. Statistical Analyses of Results (Rats) 25
 

IV. Results - Mice 29
 

A. Body Weights and Clinical Signs (Mice) 29
 

B. Survival (Mice) 31
 

C. Pathology (Mice) 31
 

D. Statistical Analyses of Results (Mice) 34
 

V. Discussion 37
 

VI. Bibliography 41
 

APPENDIXES
 

Appendix A Summary of the Incidence of Neoplasms in
 

Rats Fed o-Anisidine Hydrochloride in the Diet 43
 

Table Al Summary of the Incidence of Neoplasms in Male
 

Rats Fed o-Anisidine Hydrochloride in the Diet 45
 

Table A2 Summary of the Incidence of Neoplasms in Female
 

Rats Fed o-Anisidine Hydrochloride in the Diet 50
 

ix
 
Page 

Appendix B Summary of the Incidence of Neoplasms in Mice 

Fed o-Anisidine Hydrochloride in the Diet.. 55 

Table Bl Summary of the Incidence of Neoplasms in Male 

Mice Fed o-Anisidine Hydrochloride in the Diet 57 

Table B2 Summary of the Incidence of Neoplasms in Female 

Mice Fed o-Anisidine Hydrochloride in the Diet 61 

Appendix C Summary of the Incidence of Nonneoplastic Lesions 

in Rats Fed o-Anisidine Hydrochloride in the Diet.. 67 

Table Cl Summary of the Incidence of Nonneoplastic Lesions 

in Male Rats Fed o-Anisidine Hydrochloride 

in the Diet 69 

Table C2 Summary of the Incidence of Nonneoplastic 

Lesions in Female Rats Fed o-Anisidine 

Hydrochloride in the Diet 75 

Appendix D Summary of the Incidence of Nonneoplastic 

Lesions in Mice Fed o-Anisidine Hydrochloride 

in the Diet 81 

Table Dl Summary of the Incidence of Nonneoplastic 

Lesions in Male Mice Fed o-Anisidine 

Hydrochloride in the Diet 83 

Table D2 Summary of the Incidence of Nonneoplastic 

Lesions in Female Mice Fed o-Anisidine 

Hydrochloride in the Diet 86 

Appendix E Analyses of the Incidence of Primary Tumors in 

Rats Fed o-Anisidine Hydrochloride in the Diet 91 

Table El Analyses of the Incidence of Primary Tumors 

in Male Rats Fed o-Anisidine Hydrochloride 

in the Diet 93 

Table E2 Analyses of the Incidence of Primary Tumors 

in Female Rats Fed o-Anisidine Hydrochloride 

in the Diet 103 

Appendix F Analyses of the Incidence of Primary Tumors 

in Mice Fed o-Anisidine Hydrochloride 

in the Diet 113 

X 
Page 

Table Fl Analyses of the Incidence of Primary Tumors 

in Male Mice Fed o-Anisidine Hydrochloride 

in the Diet.... 115 

Table F2 Analyses of the Incidence of Primary Tumors 

in Female Mice Fed o-Anisidine Hydrochloride 

in the Diet 120 

Appendix G Analysis of Formulated Diets for o-Anisidine 

Hydrochloride Concentration 125 

TABLES 

Table 1 o-Anisidine Hydrochloride Chronic Feed 

Studies in Rats 10 

Table 2 o-Anisidine Hydrochloride Chronic Feeding 

Studies in Mice 11 

FIGURES 

Figure 1 Growth Curves for Rats Fed o-Anisidine 

Hydrochloride in the Diet 20 

Figure 2 Survival Curves for Rats Fed o-Anisidine 

Hydrochloride in the Diet 22 

Figure 3 Growth Curves for Mice Fed o-Anisidine 

Hydrochloride in the Diet 30 

Figure 4 Survival Curves for Mice Fed o-Anisidine 

Hydrochloride in the Diet 32 

xi
 

I. INTRODUCTION
 

Ortho-anisidine (CAS 134-29-0;
 

NCI COS747) is the common name
 

for 2-methoxyaniline and has
 

been marketed in Germany under
 

the trade name Fast Red BB
 

Base (Society of Dyers and
 

Colourists, 1971).
 

The chemical is used chiefly in the manufacture of dyes, one
 

method being the diazotization of o-anisidine and coupling with
 

other aromatic amines or phenols to yield a large number of the
 

azo dyes (Noller, 1965). Chloro, nitro, alkyl, and aryl
 

derivatives of o-anisidine exist and are used similarly in the
 

synthesis of other azo dyes. o-Anisidine is listed as a possible
 

ingredient in permanent oxidation hair dyes (Hall, 1972) although
 

it is not currently used in any hair dyes in the United States
 

(FDA, 1977). Another use of o-anisidine is as a starting
 

material in the synthesis of guaiacol (o-methoxyphenol) (Stecher,
 

1968).
 

o-Anisidine was selected for study in the Carcinogenesis Testing
 

Program because of its industrial importance, which indicated
 

that there was long-term exposure to the chemical among many
 

persons employed in the dye manufacturing industry. The earlier
 

1 
finding that o-toluidine, an aromatic amine analog of
 

o-anisidine, was carcinogenic in long-term feeding studies
 

(Homburger et al., 1972; Russfield et al., 1973) was a further
 

reason for testing o-anisidine.
 

2 
II. MATERIALS AND METHODS
 

A. Chemical
 

o-Anisidine hydrochloride was obtained from Pfaltz and Bauer,
 

Flushing, New York, in two batches. When the identity and purity
 

of Lot No. M7024-4, which was used during the subchronic studies,
 

and of Lot No. M7270, used during the chronic studies, were
 

analytically determined, both lots showed a single homogeneous
 

peak by vapor-phase chromatography and a trace impurity at the
 

origin in two different solvent systems by thin-layer
 

chromatography. Nonaqueous titration of the amine group with
 

perchloric acid was 99.5% + 0.2% of the theoretical value for Lot
 

No. M7024-4 and 100.1 + 0.3% for Lot No. M7270. By Karl Fischer
 

analysis, 0.23 + 0.01% water was detected in Lot No. M7024-4 and
 

less than 0.5% water in Lot No. M7270. Elemental analyses for
 

both lots were consistent with CyHjoNOCl, ^e molecular formula
 

of o-anisidine hydrochloride. Infrared, ultraviolet, and nuclear
 

magnetic resonance spectra were consistent with expectations
 

based on the structure and spectra in the literature for
 

o-anisidine (Sadtler, 1966, 1970; Ungnade, 1954; Pawlawski,
 

1967).
 

The bulk chemical was stored at 4°C in a plastic-lined drum. A
 

small quantity was stored in an amber glass bottle for daily use.
 

3 
B. Dietary Preparation
 

Diets were prepared once per week by first mixing a weighed
 

amount of chemical with an aliquot of ground Wayne® Lab Blox
 

animal meal (Allied Mills, Inc., Chicago, 111.) in a mortar.
 

When this premix appeared homogeneous, it was placed in a
 

Patterson-Kelly twin-shell blender with the remaining feed and
 

mixed for 20 minutes. Formulated diets were stored in double
 

plastic bags at 4°C and used within 1 week of preparation.
 

As a quality control test on the accuracy of preparation of the
 

diets, the concentration of o-anisidine hydrochloride was
 

determined at Midwest Research Institute in selected batches of
 

formulated diets during the chronic studies. The results are
 

summarized in Appendix G. At each dietary concentration, the
 

mean of the analytical concentrations for the checked samples was
 

within 12% of the theoretical concentration, and the coefficient
 

of variation ranged from 9.5% to 24.4%. In temperature-dependent
 

stability studies performed at Midwest, some loss of the test
 

chemical in feed was observed after 2 weeks at 25°C, which may
 

account in part for the unusually large variations.
 

C. Animals
 

For the subchronic studies, Fischer 344 rats and B6C3F1 mice of
 

each sex were obtained from Charles River Laboratories, Inc.,
 

4 
Wilmington, Massachusetts. For the chronic studies, Fischer 344
 

rats and B6C3F1 mice of each sex were obtained from Frederick
 

Cancer Research Center, Frederick, Maryland. Control rats and
 

mice were received earlier than, and placed on test prior to,
 

animals in the dosed groups. All animals were approximately 28
 

days of age when received at the laboratory and were quarantined
 

for 2 weeks prior to the start of the bioassay. Animals in a
 

random sample were found to be free of parasites and signs of
 

disease. At the end of the quarantine period, animals were
 

assigned to control or dosed groups in such a way that the mean
 

weights of animals in each cage were approximately the same
 

within a given group.
 

D. Animal Maintenance
 

Animal rooms were maintained at temperatures ranging from
 

23-34°C. Air was filtered through Tri-Dek® 15/40 denier Dacron
 

filters and air flow was maintained at a velocity permitting six
 

changes of room air per hour. Rooms were illuminated by
 

fluorescent lighting for 12 hours per day.
 

Rats were housed five per cage in galvanized steel wire mesh
 

cages (Fenco Cage Products, Boston, Mass.), suspended over drop
 

trays lined with newspaper. Cages and racks were sanitized every
 

week and paper in the drop trays was replaced daily. After 48
 

5 
weeks on study, rats were transferred to suspended solid
 

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

of which housed five animals per cage. These cages were equipped
 

with disposable nonwoven fiber filter sheets. The polycarbonate
 

cages were sanitized and supplied with fresh bedding two times
 

per week. A hardwood chip bedding (Aspen-bed®, American
 

Excelsior, Sommerville, Mass.) was used in the polycarbonate
 

cages.
 

Mice were housed five per cage in solid polycarbonate cages
 

fitted with perforated stainless steel lids and disposable filter
 

bonnets. A hardwood chip bedding (Aspen-Bed , American
 

Excelsior) or a corn cob bedding (Bed-o-Cobs®, Anderson Cob
 

Mills, Inc., Maumee, Ohio) was used in the mouse cages. Cages
 

were sanitized and furnished with fresh bedding two times per
 

week. Cage racks were sanitized every 2 weeks. All equipment
 

that was sanitized was washed with detergents and rinsed at 82°C.
 

Tap water (0.75-1.0 ppm chlorine) was provided in 250-ml polycarbonate bottles which were sanitized two times per week and
 

refilled as necessary. Sipper tubes and stoppers were soaked in
 

a disinfectant (Environ, -Vestal Laboratories, St. Louis, Mo.) and
 

rinsed before use, once per week. Control animals were fed
 

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

product which had been mixed with the test chemical. All diets
 

6 
were available ad libitum 7 days per week in Alpine aluminum
 

feed cups (Curtin Matheson Scientific, Inc., Woburn, Mass.) for 8
 

weeks in the mice and for 48 weeks in the rats. Thereafter, feed
 

was placed in stainless steel hoppers (Scientific Cages, Inc.,
 

Bryan, Texas). The Alpine® cups were emptied and filled with
 

fresh feed every day. The stainless steel hoppers were changed
 

two times per week and were filled with fresh feed and any that
 

had not been consumed in the earlier part of the week. However,
 

no feed was more than 1 week old when presented to the animals.
 

Rats and mice were housed in separate rooms. Control animals
 

matched with o-anisidine hydrochloride animals were in the same
 

room as the respective dosed animals. Rats on study with
 

o-anisidine hydrochloride were housed in the same room with other
 

rats being fed the following chemicals:
 

Rats
 

(CAS 615-66-7) 2-chloro-p-phenylenediamine sulfate
 

(CAS 20265-97-8) p-anisidine hydrochloride
 

(CAS 126-72-7) tris (2,3-dibromopropyl)phosphate
 

Mice were housed in a room with other mice being fed the
 

following chemicals:
 

Mice
 

(CAS 615-66-7) 2-chloro-p-phenylenediamine sulfate
 

(CAS 126-72-7) tris (2,3-dibromopropyl)phosphate
 

(CAS 20265-97-8) p-anisidine hydrochloride
 

(CAS 2438-88-2) 2,3,5,6-tetrachloro-4-nitroanisole
 

7 
(CAS 1465-25-4) N-1-naphthylethylenediamine dihydrochloride
 

(CAS 142-04-1) aniline hydrochloride
 

E. Subchronic Studies
 

Subchronic feeding studies were conducted with Fischer 344 rats
 

and B6C3F1 mice to estimate the maximum tolerated doses of
 

o-anisidine hydrochloride, on the basis of which two
 

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

doses") were determined for use in the chronic studies. In the
 

subchronic studies, the chemical was administered in feed for 7
 

weeks at doses of 1,000, 3,000, 10,000, or 30,000 ppm. Five
 

males and five females of each species were administered each
 

dose, and five males and five females of each species were given
 

basal diets. All animals were killed by inhalation of carbon
 

dioxide and necropsied 1 week after the end of the administration
 

of the test chemical.
 

The rats at doses of 1,000 and 3,000 ppm showed weight
 

depressions of less than 10%. At 10,000 ppm, weight depressions
 

were 21% for males and 11% for females; at 30,000 ppm, weight
 

depressions were 52% for males and 27% for females. No deaths
 

occurred among the rats. However, on gross pathology examination, all animals administered 10,000 or 30,000 ppm had
 

moderately enlarged spleens which were black and granular in
 

appearance. Spleens of males administered 1,000 or 3,000 ppm
 

8 
were granular, whereas in the females administered the chemical
 

at these doses, no effects were noted. Based on these data, the
 

low and high doses for the chronic studies using rats were set at
 

5,000 and 10,000 ppm.
 

In the male mice, mean body weights were depressed 6% at 1,000
 

ppm, 14% at 3,000 ppm, 28% at 10,000, and 40% at 30,000 ppm. In
 

the female mice, mean body weights were depressed 9% at 1,000
 

ppm, 19% at 3,000 ppm, 23% at 10,000 ppm, and 37% at 30,000 ppm.
 

One female died during the study at 30,000 ppm. Spleens were
 

black and enlarged in both the males and the females administered
 

10,000 and 30,000 ppm. Based on these data, the low and high
 

doses for the chronic studies using mice were set at 2,500 and
 

5,000 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
 

Inspections for mortality and morbidity were carried out twice
 

daily. Body weights were recorded every 2 weeks for the first 12
 

weeks and monthly thereafter. Clinical observations were
 

recorded every month.
 

9 
Table 1. o-Anisidine Hydrochloride Chronic Feeding Studies in Rats
 

o-Anisidine 

Sex and Initial Hydrochloride Time on Study 

Test No. of in Dietb Dosed Observed 

Group Animals3 (ppm) (weeks) (weeks) 

Male 

Control0 55 0 106 

Low-Dose 55 5,000 103 1 

High-Dose 55 10,000 88d 

Female 

Control0 55 0 107 

Low-Dose 55 5,000 103 

High-Dose 55 10,000 83d 

a

Rats were 41 days of age when placed on study.
 

^Diets were available ad libiturn 7 days per week.
 

c

Controls were placed on study 3 weeks earlier than the dosed
 

groups.
 

"Period of administration terminated at time indicated, due to
 

death of all animals.
 

10
 
Table 2. o-Anisidine Hydrochloride Chronic Feeding Studies in Mice
 

o-Anisidine 

Sex and Initial Hydrochloride Time on Study 

Test No. of in Dietb Dosed Observed 

Group Animals3 (ppm) (weeks) (weeks) 

Male 

Control0 55 0 105 

Low-Dose 55 2,500 103 1-2 

High-Dose 55 5,000 103 2 

Female 

Control0 55 0 105 

Low -Dose 55 2,500 103 2 

High-Dose 55 5,000 103 2 

a

Mice were 41 days of age when placed on study.
 

"Diets were available ad libitum 7 days per week.
 

°Controls were placed on study 3 weeks earlier than the dosed
 

groups.
 

11
 
Moribund animals and animals that survived to the end of the
 

bioassay were killed using CC>2 anesthesia and necropsied.
 

Necropsies were also performed on all animals found dead, unless
 

precluded by autolysis or severe cannibalization. The following
 

tissues were examined where possible: tissue masses, abnormal
 

regional lymph nodes, skin, mandibular lymph nodes, mammary
 

gland, salivary gland, thigh muscle, sciatic nerve, bone marrow,
 

costochondral junction (rib), thymus, larynx, trachea, lungs and
 

bronchi, heart, thyroid, parathyroid, esophagus, stomach,
 

duodenum, jejunum, ileum, colon, mesenteric lymph nodes, liver,
 

gall bladder (mice), pancreas, spleen, kidney, adrenal, bladder,
 

seminal vesicles/prostate/testis (males), ovary/uterus (females),
 

nasal cavity, brain, pituitary, eyes, external and middle ear,
 

and spinal cord. Peripheral blood smears were prepared from each
 

animal whenever possible. The different tissues were preserved
 

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

stained with hematoxylin and eosin. Special staining techniques
 

were utilized when indicated for more definite 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
 

12
 
tissues were examined microscopically varies, and does not
 

necessarily represent the number of animals that were placed on
 

study in each group.
 

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
 

13
 
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.
 

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
 

14
 
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
 

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
 

15
 
the first tumor was found. Once this reduced set of data was
 

obtained, the standard procedures for analyses of the incidence
 

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 p^/Pc where p^ 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
 

16
 
group. The hypothesis of equality between the true proportion of
 

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

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.
 

17
 

III. RESULTS - RATS
 

A. Body Weights and Clinical Signs (Rats)
 

Mean body weights of dosed male and female rats were lower than
 

those of corresponding controls throughout the bioassay, and the
 

depressions in weight were dose related (figure 1). Fluctuations
 

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.
 

Palpable subcutaneous masses were seen in 4 control males, 17
 

control females, 8 low-dose males, and 7 low-dose females.
 

Cutaneous lesions and/or growths developed in five control males,
 

four control females, five low-dose males, seven low-dose
 

females, and three high-dose females. Discoloration, reddening,
 

and crusting of the eyes was noted in 2 control males, 19
 

low-dose males, 17 low-dose females, and 1 high-dose male.
 

Rectal prolapses occurred in three control males and one
 

high-dose female. Two control males displayed jaundice and one
 

had a distended scrotal sac. Nine control females had facial
 

alopecia. Emaciation was noted in two control females, two
 

low-dose males, five low-dose females, two high-dose males, and
 

one high-dose female. Abdominal distention occurred in one
 

low-dose male, two low-dose females, and two high-dose males.
 

19
 
Figure 1. Growth Curves For Rats Fed o-Anisidine Hydrochloride In The Diet 

20 
Two low-dose males displayed posterior ataxia, and three
 

high-dose males were badly hunched over prior to death.
 

Blood/exudates in the urogenital area and/or black/brown stained
 

fur in the urogenital region were seen in 45 low-dose males, 18
 

low-dose females, 5 high-dose males, and 4 high-dose females.
 

B. Survival (Rats)
 

The Kaplan and Meier curves estimating the probabilities of
 

survival for male and female rats fed o-anisidine hydrochloride
 

in the diet at the doses of this bioassay, together with those of
 

the controls, are shown in figure 2.
 

The result of the Tarone test for positive dose-related trend in
 

mortality is significant (P < 0.001) in each sex. A departure
 

from linear trend is observed (P < 0.001), because of the steep
 

decrease in survival in the dosed groups. In the high-dose group
 

of each sex, no animal survived to termination of the study;
 

however, 49/55 (89%) of the high-dose males and 44/55 (80%) of
 

the high-dose females were still alive at week 52. All 55 of the
 

low-dose animals and all 55 of the control animals of each sex
 

lived beyond week 52 on study.
 

C. Pathology (Rats)
 

Histopathologic findings on neoplasms in rats are summarized in
 

21
 
Figure 2. Survival Curves For Rats Fed o-Anisidine Hydrochloride In The Diet 

22 
Appendix A, tables Al and A2; findings on nonneoplastic lesions
 

are summarized in Appendix C, tables Cl and C2.
 

Transitional-cell carcinomas of the urinary bladder were found at
 

a high incidence in both males (controls 0/51, low-dose 50/54,
 

high-dose 51/52) and females (controls 0/49, low-dose 41/49,
 

high-dose 50/51). Transitional—cell carcinomas of the bladder
 

had varying morphological patterns. The more well-differentiated
 

tumors consisted of transitional cells growing in solid sheets
 

and serpentine strands two or three cells thick in a fibrous
 

stroma. There were scattered foci of necrosis and mitotic
 

figures were uncommon. The bladder wall was usually extensively
 

invaded and the tumor often nearly filled the lumen. In some
 

tumors, there was a marked desmoplasia and a mucinous degeneration of the stroma. Many tumors were very pleomorphic with
 

spindle cells, tumor giant cells, bizarre hyperchromatic nuclei,
 

and numerous abnormal mitotic figures. Areas of squamous
 

metaplasia were common. In a few tumors, extensive formation of
 

cystic spaces occurred. The tumor appeared to have a multi-


centric origin in a few instances with transitional-cell
 

carcinomas appearing in the bladder and one or more additional
 

sites such as renal pelvis, prostatic urethra, and kidney. One
 

low-dose and one high-dose rat had two types of malignant primary
 

tumors involving the bladder. A transitional-cell carcinoma
 

23
 
filled the bladder lumen and a leiomyosarcoma originated from the
 

muscularis. Although distant metastases were not seen in rats
 

with bladder carcinomas, the bladder was heavily invaded, and in
 

one case, the tumor extended to the serosa of the spleen and
 

prostate. Diffuse and focal transitional-cell hyperplasias were
 

seen in many bladders with carcinomas.
 

A variety of other neoplasms were observed with approximately
 

equal frequency in the control and dosed animals. There were
 

instances in this bioassay, as noted in the summary tables, where
 

these neoplastic lesions occurred only in dosed animals or with
 

increased frequency when compared with the control group. The
 

nature and incidence of these neoplasms are similar to those
 

known to occur spontaneously in aged Fischer 344 rats. However,
 

follicular-cell tumors of the thyroid in males may be related to
 

administration of the test chemical.
 

Nonneoplastic lesions which commonly occur in rats of this strain
 

were seen. These were not considered to be compound induced.
 

Based on the histopathologic examination, there was evidence that
 

under the conditions of this bioassay o-anisidine hydrochloride
 

was carcinogenic when fed to Fischer 344 rats, inducing
 

transitional-cell carcinomas of the urinary system.
 

24
 
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 each sex, the incidence of transitional-cell carcinomas or
 

papillomas of the urinary bladder is high in either dosed group
 

(males: low-dose 52/54 [96%], high-dose 52/52 [100%]; females:
 

low-dose 46/49 [94%], high-dose 50/51 [98%]). These lesions were
 

observed as early as week 36 in female and week 40 in male rats;
 

none were observed in the control groups. The results of the
 

Cochran-Armitage test for positive dose-related trend are
 

significant (P < 0.001) in each sex. A departure from linear
 

trend is observed (P < 0.001), because of the steep increase in
 

incidence in the dosed groups. The results of the Fisher exact
 

test comparing the incidence of these tumors in each dosed group
 

with the incidence in the control group are significant (P <
 

0.001) in each sex. The statistical conclusion is that the
 

incidence of these transitional-cell tumors of the urinary
 

bladder in rats of each sex is dose associated.
 

In male rats, the results of the Cochran-Armitage test are
 

significant (P = 0.005) for the incidence of transitional-cell
 

25
 
carcinomas of the kidney or kidney pelvis, and the results of the
 

Fisher exact test show that the incidence of this tumor is
 

significantly higher (P = 0.006) in the high-dose group than in
 

the control group. The statistical conclusion is that the occurrence of transitional-cell carcinomas of the kidney or kidney
 

pelvis in the male rats was associated with the administration of
 

o-anisidine hydrochloride. In female rats the incidence of this
 

tumor was 1/54 in the high-dose group and 0/52 in both the low-


dose and the control groups.
 

The results of the Cochran-Armitage test are also significant in
 

the male rats for the incidence of follicular-cell adenomas,
 

cystadenomas, or papillary cystadenomas of the thyroid or thyroid
 

follicle (P = 0.030). The corresponding results of the Fisher
 

exact test show P values of 0.031 for each dosed group; however,
 

these values are not significant when the Bonferroni inequality
 

criterion for multiple comparisons is applied. The results of
 

the Cochran-Armitage test on the combined incidence of all
 

follicular-cell tumors of the thyroid (carcinomas, adenomas, cystadenomas, papillary cystadenomas, or papillary cystadenocarcinomas) show an increased significant trend (P = 0.009), and the
 

Fisher exact comparisons of the combined incidence in each dosed
 

group with that in the control group also show an increased
 

significance (P <_ 0.005). The incidence of follicular-cell
 

26
 
tumors in female rats is not significant. The historical
 

controls at this laboratory indicate an incidence of follicularcell thyroid tumors in male rats of 3/250 (1.2%) and in female
 

rats of 2/249 (0.8%).
 

In each sex, significant results in the negative direction are
 

observed in incidences of several of the tumors, due to higher
 

incidences in the control groups than in the dosed groups. This
 

negative significance may be caused by the fact that the dosed
 

animals did not live as long as the controls.
 

In summary, the results of the statistical tests show that the
 

administration of o-anisidine hydrochloride is associated with
 

the occurrence of transitional-cell carcinomas or papillomas of
 

the urinary bladder in both sexes of Fischer 344 rats, and with
 

the occurrence of transitional-cell carcinomas of the kidney or
 

kidney pelvis and follicular-cell tumors of the thyroid in male
 

rats.
 

27
 

IV. RESULTS - MICE
 

A. Body Weights and Clinical Signs (Mice)
 

Mean body weights of dosed male and female mice were lower than
 

those of corresponding controls throughout the bioassay, and the
 

depressions in weight were dose related (figure 3). Fluctuations
 

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.
 

Subcutaneous palpable masses were recorded in four control males,
 

three control females, and one low-dose female. Urogenital
 

distention, nodules and/or bleeding was seen in three control
 

males and one control female. Cutaneous growths developed in two
 

control males, one low-dose male, and two high-dose females.
 

Swelling of the eyes was seen in two low-dose females, two high-


dose males, and two high-dose females. Alopecia, at one time or
 

another during the bioassay, was noted in 42 control males, 54
 

control females, 49 low-dose males, 34 low-dose females, 25 high-


dose males, and 26 high-dose females. One high-dose female had
 

labored breathing. These signs were not considered to be related
 

to administration of the test chemical.
 

29
 
Figure 3. Growth Curves For Mice Fed o-Anisidine Hydrochloride In The Diet 

30 
B. Survival (Mice)
 

The Kaplan and Meier curves estimating the probabilities of
 

survival for male and female mice fed o-anisidine hydrochloride
 

in the diet at the doses of this bioassay, together with those of
 

the 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, 43/55 (78%) of
 

the high-dose group, 43/55 (78%) of the low-dose group, and 44/55
 

(80%) of the controls lived to the end of the bioassay. In
 

females, the proportions which survived to termination of the
 

study were 42/55 (76%) of the high-dose group, 38/55 (69%) of the
 

low-dose group, and 44/55 (80%) of the controls. Sufficient
 

numbers of mice of each sex were at risk for the development of
 

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.
 

Transitional-cell papillomas of the urinary bladder were significantly elevated in dosed male mice (controls 0/48, low-dose 2/55,
 

high-dose 7/53). Transitional-cell carcinomas of the bladder
 

31
 
Figure 4. Survival Curves For Mice Fed o-Anisidine Hydrochloride In The Diet 

32 
were found to be significantly elevated in both males (high-dose
 

15/53) and females (high-dose 18/50). No transitional-cell
 

carcinomas were seen in male and female low-dose or control
 

groups. The earliest bladder changes recognized were focal
 

hyperplasias (males: controls 1/48, low-dose 2/55, high-dose
 

21/53; females: controls 0/50, low-dose 1/51, high-dose 12/50)
 

consisting of a few transitional cells with slight atypia piling
 

up and projecting into the lumen and diffuse mild thickening of
 

the epithelium with occasional piling up of cells and short
 

papillary formations. There was normal cellular polarity and an
 

intact basement membrane. The transitional-cell carcinomas
 

invaded the bladder wall with strands and finger-like sheets of
 

transitional cells. In some tumors, many small cystic spaces
 

were formed, giving a pseudoacinar appearance. Extensive
 

invasion of the bladder wall was often seen. Occasionally, a
 

relatively small transitional-cell carcinoma invaded to the
 

serosa. Some of the invasive tumors had extensive lymphocytic
 

and plasma-cell infiltrates both surrounding and within the
 

tumor. There were no distant metastases.
 

A variety of other neoplasms were observed with approximately
 

equal frequency in the control and dosed mice. There were
 

instances in this study, as noted in the summary tables, where
 

these neoplastic lesions occurred only in dosed animals or with
 

33
 
increased frequency when compared with the control group. The
 

nature and incidence of these neoplasms are similar to those
 

known to occur spontaneously in aged mice of this strain.
 

A variety of inflammatory and degenerative lesions which commonly
 

occur in aging B6C3F1 mice were seen. None of these lesions were
 

considered to be compound related.
 

Based on the histopathologic examination, there was evidence that
 

o-anisidine hydrochloride was carcinogenic when fed to both male
 

and female B6C3F1 mice, inducing transitional-cell carcinomas,
 

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 in one group and with an incidence of at least
 

5% in one or more than one group.
 

In each sex, transitional-cell carcinomas or papillomas of the
 

urinary bladder occur in significant incidences (P < 0.001)
 

exclusively in the high-dose groups (high-dose males 22/53 [42%]
 

and high-dose females 22/50 [44%]), compared with the controls.
 

These lesions were observed as early as week 45 in male mice.
 

The results of the Cochran-Armitage test are also significant
 

34
 
(P < 0.001) in each sex. A departure from linear trend is
 

observed (P = 0.005 in male mice and P = 0.001 in female mice),
 

because of the steep increase in the incidence of tumors in the
 

high-dose group. The statistical conclusion is that the
 

incidence of transitional-cell carcinomas or papillomas of the
 

urinary bladder in mice is dose associated.
 

Significant results in the negative direction are observed in
 

several of the incidences of tumors in each sex, where the
 

incidences in the control group exceed those in the dosed groups.
 

In summary, the statistical conclusion indicates that the
 

incidence of transitional-cell tumors of the urinary bladder in
 

B6C3F1 mice is associated with the administration of o-anisidine
 

hydrochloride.
 

35
 

V. DISCUSSION
 

The toxicity of o-anisidine hydrochloride for Fischer 344 rats
 

and B6C3F1 mice was shown by consistently lowered mean body
 

weights of all dosed groups when compared with corresponding
 

controls throughout the bioassay; further, the data indicated
 

dose-related effects of the test chemical on the mean body
 

weights. Bloody exudates and stained fur in the urogenital area
 

were noted in many dosed animals.
 

Survival of the rats at the end of the bioassay was low (males:
 

controls 71%, low-dose 13%, high-dose 0%; females: controls 65%,
 

low-dose 0%, high-dose 0%); however, survival was 80% or greater
 

at week 52, and early development of tumors established the
 

carcinogenicity of the test chemical, as given below. Survival
 

of the mice at the end of the bioassay was high (males 78-80%;
 

females 76-80%), and sufficient numbers of animals were at risk
 

for the development of late-appearing tumors.
 

Transitional-cell carcinomas or papillomas of the urinary bladder
 

occurred at statistically significant incidences (P < 0.001) in
 

the low- and high-dose groups of rats (males: controls 0/51,
 

low-dose 52/54, high-dose 52/52; females: controls 0/49,
 

low-dose 46/49, high-dose 50/51) and in high-dose groups of mice
 

(males: controls 0/48, low-dose 2/55, high-dose 22/53; females:
 

37
 
controls 0/50, low-dose 1/51, high-dose 22/50); the incidences
 

also had significant dose-related trends (P < 0.001) in both
 

species. These lesions were observed as early as week 36 in
 

female rats, week 40 in male rats, and week 45 in male mice.
 

Transitional—cell carcinomas of the pelvis of the kidney occurred
 

with a significant dose-related trend (P = 0.005) in the male
 

rats, and the incidence in the high-dose group was significantly
 

higher (P = 0.006) than in the control group (controls 0/53,
 

low-dose 3/55, high-dose 7/53); all rats having this tumor also
 

had a transitional-cell carcinoma of the urinary bladder. A
 

transitional-cell carcinoma also occurred in the pelvis of the
 

kidney in one high-dose female rat. Only one animal in the
 

control groups of rats or mice had any tumor of the urinary
 

system (a transitional-cell papilloma of the pelvis of the kidney
 

in a male mouse).
 

Follicular-cell tumors of the thyroid (carcinomas, cystadenocarcinomas, adenomas, cystadenomas, and papillary cystadenomas)
 

occurred at statistically significant incidences (P _£ 0.005) in
 

low- and high-dose groups of male rats (controls 0/53, low-dose
 

7/40, high-dose 6/40); the incidences also had a dose-related
 

trend (P = 0.009). These tumors did not occur at significant
 

incidences in dosed groups of female rats.
 

In humans, o-anisidine can be absorbed through the skin and is an
 

38
 
irritant and a sensitizer (Stecher, 1968). No previous work has
 

been reported on long-term studies of toxicity or carcinogenicty
 

of o-anisidine.
 

It is concluded that under the conditions of the bioassay,
 

o-anisidine hydrochloride was carcinogenic for Fischer 344 rats
 

and B6C3F1 mice, inducing transitional-cell carcinomas or
 

papillomas of the bladder in both rats and mice and in both sexes
 

of each species, transitional-cell carcinomas of the pelvis of
 

the kidney in male rats, and follicular-cell tumors of the
 

thyroid in male rats.
 

39
 

VI. BIBLIOGRAPHY
 

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

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

Berenblum, I., ed. , Carcinogenic!ty Testing; A. Report on the
 

Panel on Carcinogenicity of the Cancer Research Commission
 

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

Geneva, 1969.
 

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

Soc. 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 Products.
 

Ingredient Statements Registered in Accordance with jl CFR
 

720, August 15, 1977.
 

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

significance tests, confidence limits and adjustments for
 

stratification. Rev. Int. Statist. Inst. 39;148-169, 1971.
 

Homburger, F., Friedell, G. H., Weisburger, E. K., and
 

Weisburger, J. H., Carcinogenicity of simple aromatic amine
 

derivatives in mice and rats. Toxicol. Appl. Pharmacol.
 

.22:280-281, 1972.
 

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

incomplete observations. J. Am. Statist. Assoc. _53:457-481,
 

1958.
 

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

J., Carcinogenesis bioassay data system. Comp. and Biomed.
 

Res. 7;230-248, 1974.
 

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

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

Noller, C. R., Chemistry of Organic Compounds, W. B. Saunders
 

Co., Philadelphia, 1965, pp. 561, 564, 565, and 743-744.
 

41
 
Pawlowski, W., Molar absorptivity of aromatic amines in 

ultraviolet (Region of Spectrum). Chernia Analityczna 12 

(4):893-896, 1967. 

Russfield, A. B., Homburger, F., Weisburger, E. K. , and
 

Weisburger, J. H., Further studies on carcinogenicity of
 

environmental chemicals including simple aromatic amines.
 

Toxicol. Appl. Pharmacol. 25; 446-447, 1973.
 

Sadtler Standard Spectra, NMR No. 192M and 1235M, Sadtler
 

Research Laboratories, Philadelphia, 1966.
 

Sadtler Standard Spectra, IR No. 3696, Sadtler Research
 

Laboratories, Philadelphia, 1970.
 

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, The Colour Index, Vol. k_, Dean
 

House, Yorkshire, England, 1971 pp. 4349 and 4705.
 

Stecher, P. G. , ed. , The Merck Index, Merck Co. Rahway, N. J.,
 

1968, p. 508.
 

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

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

Ungnade, H. E., Ultraviolet absorption spectra of acetanilides.
 

J. Am. Chem. Soc. 7J>;5133-5135, 1954.
 

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

Cosmetics Science and Technology, Vol. 2., Balsam, M. S. and
 

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

306-308.
 

42
 
APPENDIX A
 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN
 

RATS FED o-ANISIDINE HYDROCHLORIDE IN THE DIET
 

43
 

TABLE A1.

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE RATS 

FED o-ANISIDINE HYDROCHLORIDE IN THE DIET 

CONTROL LOW DOSE HIGH DOSE 

AHIHALS INITIALLY IN STUDY 55 55 55 

ANIMALS NECFOPSIBP 5<» 55 53 

ANIMALS EXAMINE D HISTOPATHOLOGICALLY 54 55 53 

INTEGUMENTARY SYSTEM 

*SKIN (5U) (55) (53)

 

SQOAHOOS CF.LL PAPILLOJ1A 2 (4X) 

BASAL-CBLL TTIMO5 1 (2X> 

FIBROMA 2 (U%)

 

*SOBCUT TISSUE (5U ) (55) (53)

 

FIBROMA 2 (US)

 

RESPIRATORY SYSTEM 

*NASAL CAVITY (5«) (55) (53)

 

UNDIPFEP?NTIATEO CAPCINOM A 2 (UX|

 

tl.UNG (5U) (55) (51)

 

M.VOLA?/3?OHCHIOT H P AD5HOSA 1 (2%)

 

HEBATOPOIFTIC SVST5!1 

tBRAIN/MFNING^ S (54) (53) (51) 

MALIGNAN T LYMPHOM^ , NO? 1 (2«) 

*MULTIPLS ORGANS (5U) (55) (53)

 

MALIGNANT LYBPHOMA, NOS 1 (2*)

 

USDIF'ESENTIATEr LFJKFMI A 13 (2«*,)

 

LYMPHOCYTIC LEUKH>1IA "» (7X)

 

*BONF MARRO W (52) (55) (47)

 

OSTFOSARCOMA , INVASIV^ 1 1 <2S)

 

tSPLENIC CAPSUL2 (5U) (55) (52) 

!EUISlllSa4.Lz£2LL_£lESLLH2ttA 1_12SL_ 

I NUMBFR OF ANIMAL S WIT H TISSUE EXAMINE D MICROSCOPICALLY 

* NUMBFR OF ANIMAL S N7.CPOPSIED 

45 
TABLE A1. MALE RATS: NEOPLASMS (CONTINUED) 

fHANDIBOLAR I. MODE 

MALIGNAN T LYHPHOMA , NO S 

CIRCULATOR Y SYSTEM 

NONF 

DIGESTIVE SYSTEM 

tLTV'R 

NEOPLASTIC NODU11: 

HEPATOCHLLOIAR CARCINOM A 

LEIOMYOSA8COMA , S^TS.STATIC 

• PANCREA S 

LEIOMYOSAPCONA , K'TA STATIC 

•STOMACH 

SQOAMOO S CFII. CAfCINOa A 

LFIOMYOSAPCOil A 

iDUOPFNA L SEFOS A 

LFIOMYOSAPCOMA , «'?A STATIC 

URINAR Y SYSTE M 

• KIDNE Y 

TPANSITTONAL-C?r.I . CAPCINOI'i A 

#KIDNEY/PFt.VI S 

TRANSITIONAL-CFL l CARCINOM A 

tURINAR Y BLADDS 8 

T R A NSTTIONAL-CFIL PIPILLO^ A 

TRANSITIONAL-CEL L CARCINOM A 

LEIOMYOSAP.COMA 

HEMANGIOM A 

*PPOSTATIC IIHRTHF A 

T3ANSITIONAL-CFLL CARCINOM A 

t NUMBE R OF ANIMAL S HIT^ TISSU F

* NUMBF B OF ANISAI S N^C^OPSIS D 

CONTROL LOW DOSE 

(53) C*7) 

1 (2X) 

(5U) (55) 

2 (4*) 

2 (4X) 

(53) (54) 

(53) (55) 

2 (41?) 

(52) (53) 

(53) (55) 

1 (2!?) 

(53) (55) 

2 (4%) 

(51 ) (54) 

2 (4%) 

SO (93X ) 

1 (2%) 

(54) (55) 

 ^XAMINF D MICROSCOPICALL Y 

HIGH DOSE 

(3«) 

(52) 

1 (2X) 

(51) 

1 (2*) 

(52) 

1 (2X) 

(51) 

1 (2«) 

(53) 

3 (6%) 

(53) 

4 (8X) 

(52) 

1 (2X ) 

51 (9835) 

1 (2«) 

1 (2X) 

(53) 

2 J4J1 

46 
TABLE A1. MALE RATS: NEOPLASMS (CONTINUED) 

CONTROL LOW DOSE 

ENDOCRIN E SYST2M 

#PITUITAP Y (U8 ) (49) 

ADENOMA , NOS 2 (<**) 

CHROMOPHOS E ADENOM A U (3%) 

ACIDOPHI L ADENOW A 1 (2% ) 

f AD3?NU. (51) (55)

 

CORTICAL ADENOK A 1 (2%)

 

PHEOCHRO«OCYTOM» 12 (22*)

 

PHPOCHROMOCYTOMA , MALIGNAN T 2 (i*%) 

LEIOMYOSAPCOMA , MFTASTATI C 

tTHYROI D (53) (HO ) 

FOLLICULAE-CELL ADENOM A 3 (835) 

FOLLICDLAR-CELL CARCINOM A 2 (5%) 

C-CELL ADENOM A 3 (6«) 1 (3X ) 

C-CELL CARCINOM A 2 (5%) 

CYSTADENOKA , NO S 1 (3K) 

tTHYHOID FOLLICLE (53) C»0) 

PAPILLAR Y CYSTADENOMA , NO S 

PAPILLAR Y CYSTADENOCA8CINOHA.NO S 1 (3X ) 

#PANC8EATIC ISLETS (53) (54) 

ISLET-CELL ADENOM A 1 (2%) 1 (2«) 

ISLET-CELL CARCINOMA 1 (2%) 

REPRODUCTIVE SYSTEM 

*HAHHAR Y GLAN D (5<» ) (55)

 

INTRADUCTAL PAPILLOMA 1 (2%)

 

"IBRD ADENOM A 1 (2%) 

*PREPUTIAL GLAS D (54) (55) 

CARCINOMA , NOS 1 (2X) 2 (4S) 

•PROSTATE (52) (52)

 

TRANSITIONAL-CELL CARCINOM A 

#TESTIS (5U) (52) 

_ IflT_EHS_miA.L-C_£LL TUMOP _ . _53 J9B?1_ _ i»3_J83XL 

t NUHBER OF ANIMAL S WIT H TISSUE EXAMINE D MICRDSCOPICALL Y 

* NUMBER OF ANIMAL S NECPOPSIED 

HIGH DOSE 

(47 ) 

(53) 

1 (2*) 

(tO) 

1 (3%) 

2 (5%) 

(40) 

3 (8S) 

(51) 

(53) 

(53) 

2 (4X) 

(52) 

1 (2%) 

(51) 

ii_i8SL 

47 
TABLE A1. MALE RATS: NEOPLASMS (CONTINUED) 

CONTROL LOW DOSE 

NEPVOfJ S SYSTE M 

IBP A IN (54) (53) 

CFRUMINOU S CAPCIN01A , i*l HT A STATIC 1 (2% ) 

ASTPOCYTOI A 2 («) 

SPECIA L SENS E ORGAN S 

*EYP (54) (55)

 

SQUAMOU S CFL L CA PC I N OH A 1 (2% )

 

*EAR (54) (55)

 

CERUMINOU S CAPCINOM» . 1 (2% ) 

*EAS CANA L (54) (55)

 

CEPUttlNOU S CARCINOM A 1 (2% )

 

MOSCULOSKELKTAL SYSTFM 

*VERTEBR A (54) (55) 

OSTFOSAPCOP1A 1 (2%) 

BODY CAVITIES 

*BODY CAVITIE S (54 ) (55) 

MESOTHELIOMA , NOS 2 (i*«) 4 (7%) 

MFSOTtTELIOMA , MALIGNAN T 2 (US) 

A LL OTHER SYSTEMS 

NONE 

ANIMA L DISPOSITION SUMMAR Y 

ANIMAL S INITIALL Y I N STUDY 55 55 

NATURA L DFATH3 ) 6 21 

MORIBUN D SACRIFICE 10 27 

SCHEDOLFD SACRIFICE 

ACCIDENTALL Y KILLED 

TERMINA L SACRIFICE 39 7 

ANIMA L MISSING 

9 INCLUDES AUTOLYZE D ANIMAL S 

* NUMBER OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICAL!,!
 

* NUHBEB OF ANIMALS NECBOPSIED
 

HIGH DOSE 

(51) 

(53) 

(53) 

(53) 

(53) 

(53) 

1 (2%) 

55 

37 

18 

48 
TABLE A1. MALE RATS: NEOPLASMS (CONTINUED) 

CONTROL LOW DOSE HIGH DOSE 

TUBOB SUMMAR Y 

TOTAL ANIMALS WITH PRIMAR Y TUMORS* 54 53 53 

TOTAL PRIMAR Y TUMORS 112 132 80 

TOTAL ANIMAL S WITH B2NIT, N TUMORS 53 43 8 

TOTAL BENIGN TUMORS 83 55 10 

TOTAL ANIMAL S WITH MALIGNAN T TUMORS 26 51 53 

^OTAL MALIGNAN T TUMORS 27 71 69 

TOTAL ANIMAL S WITH SECONDAR Y TUMORS* 1 1 1 

TOTA L SECONDAR Y TUMOP S 1 1 4 

TOTAL ANIMAL S WITH TUMOH S UNCFRTAIN ­

BENIG N O R MALIGNAN T 2 6 1 

TOTAL UNCERTAI N TUMOR S 2 6 1 

TOTAL ANIMAL S WITH '"UflOSS UNCERTAIN ­

PRIMAR Y OR MFTftSTATIC 

TOTAL UNCERTAI N TUMD? S 

* PRIMARY TMP1CRS: ALL TUMORS EXCFPT SECONDARY TUHORS
 

* SECONDARY TUMORS: qVTASTATIC TMOPS OB TUMORS INVASIV E INTO AN ADJACENT ORGAN
 

49 
TABLE A2.

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE RATS 

FED o-ANISIDINE HYDROCHLORIDE IN THE DIET 

CONTROL LOW DOSE 

ANIMALS INITIALLY IN STUDY 55 55 

ANIMAL S NECBOPSIE D 54 53 

ANIMAL S PVAMINE D HISTOP A THOLOGICALL Y 54 53 

INTEGUMENTAR Y SYSTE M 

*SKIN (54) (53) 

SQUAaOU S CEL L PAPILtOM A 2 (UX ) 

SQtlAHOOS CEL L CARCINOM A 2 (US ) 

*SUBCUT TISSFJK (54) (53) 

SQUAHOU S C^T L CARCINOM A 

ADENOCARCIN01A ,

"I3POMA 

 NCS 

1 (2?!) 

1

2

 (2%) 

 (4JS) 

"IBRO ADENOM A 1 (255) 

RESPIRATOR Y SYSTE M 

*NASA L CAVIT Y (54) (53) 

CARCINOMA , NOS 1 (28) 

#I.UN G (53 ) (53) 

ALVPOLAR/GaONCHIO ! A F ADflNO.I A 1 (25!) 

HEMATOPOI^TTC : oYSTF M 

*«ULTIP1=' ORGAN S (54) (53) 

1ALIGMAN T TYMPHOI" , KOS 1 (2%) 

FIND I^'ERENTI ATE.": LKL' K ^?1IA 9 (15%) 

LYMPHOCYTI C L£ 'JK^ M T« 1 (2?,) 

*SPLEFN (52) (52) 

NE'JROFIBPOSAFCOKA , UN C ?EI M 03 .1 1 (2!?) 

IMEDIASTINA L L. NOD^ (51) (46) 

!JNDIFFEREKTIA"-F D CAPCIN03 A META S 1 (2%) 

*KES:1NT7PIC L. NOHE (51) (46) 

UN£IFfEPEN^IA2ID_C:APCINOMA_2Fl!A S l_12Sl 

* NUMBF.P OF ANIMAL S KITK TISSUE EXAMINED MICBOSCOPICALLY
 * NU3SF-F OF ANIMAL S N"?FO?SI fit) 

HIGH DOSE 

55 

54 

54 

(54) 

(54) 

1 (2%) 

(54) 

(54) 

(54) 

(51) 

(36) 

(36) 

50 
TABLE A2. FEMALE RATS: NEOPLASMS (CONTINUED) 

CONTROL LOW DOSE HIGH DOSE 

CIRCULATOR Y SYSTE M 

tHHAP T 

N?rjFOFI3ROSASCO?.A, USC ?ET N

DIGESTIVE SYST3M 

tSALIVAR Y GLAN D 

ADENOMA, NOS 

fLIVER 

NKOPLASTIC NODULE 

NEUROFIBBOSABCOMA , U NC PRI M

tSTOMACH 

SQUAHOU S CELL PAPILLOM 

SQUAHOUS CELL CARCINOK A 

ADENOCAPCINOMA , NOS 

UPINAR Y SYSTEM 

tKIDNEY/PELVI S 

TRANSITIONAL-CELL

tURINAR Y BLADDER 

TRANSITIONAL-CELL

TRANSITIONAL-CELL

"IBROMA 

LEIOMYOSARCOM A 

HEBANGIOM A 

ENDOCRINE SYSTEM 

f PITUITARY 

CARCINOMA, NOS 

ADENOMA , NOS 

 CARCINOM A 

 PAPILLOHA 

 CARCINOM A 

CHROHOPHOBE ADENOM A 

CHROMOPHOBE CARCINOM A 

ACIDOPHIL ADENOM A 

tADBENAL 

CQBT.IEAI ABENOMA 

t NUHBER OF ANIMAL S WITH TISSUE

* NUMBER OF ANIHALS NECROPSISD 

(53) 

(47) 

(53) 

(49) 

(54) 

1 (2X) 

(51) 

50 (98X) 

1 (2X) 

(45) 

1 (2X) 

(54) 

(53)

 OR « 1 (2%) 

(52) 

3 (6%) 

(53) 

1 (2% ) 

 OR M 1 (2X) 

(51) 

1 (2%) 

1 (2X) 

1 (2%) 

(52) 

(49) 

(48) 

3 (6X) 

1 (2%) 

15 (31X) 

1 (2X) 1 (2X) 

(53) 

112 Jl

(53) 

(51) 

(53) 

(51) 

(52) 

C*9) 

5 (10X) 

41 (84X) 

1 (2X) 

1 (2X) 

1 (2X) 

(51) 

9 (18JS) 

(53) _ . 

 EXAMINE D MICROSCOPICALLY 

51 
TABLE A2. FEMALE RATS: NEOPLASMS (CONTINUED) 

PH?OCHROBOCYTO:i» . 

ANGIOLIPOB A 

tTHYROID 

USDIFFFRFNTIATF.!) CA7CINOM A 

POLLICULAR-CEL L AD? NO W A 

"OLLIC'TLAP-CELL CATINOM A 

C-CELL ADENOB A 

C-CELL CARCIMOB A 

#THYROI D FOLLICLF 

PAPILLAR Y CYSTADENOMA , SO S 

*PANCRFATIC ISLETS 

ISLET-CELL ADENOM A 

REPRODUCTIV E SYSTE M 

*BABBAR Y GLAN D 

ADENOMA , NO S 

ADENOCARCINOBA , NOS 

"IBPO ADENOM A 

*CLITORAL GLAN D 

CARCINOBA.NO S 

iOTEPtJS 

BNDOBETHIA L STROBAL POLYP 

HEMANGIOM A 

fOVARY 

GRANULOSA-CEL L TUMO R 

TUBULA R ADENOB A 

NEFVOOS SYSTKB 

*OIFACTOHY SYSTEB 

HENINGIOMA 

#BRAIN/BENIHGES 

CARCINOMA , NOS, INVASIV E 

tBRAIN 

CARCINQBA , NOSj, BETA STATIC 

* NOBBEF OF ANIBAL S WITH TISSUE

* NUMBER OF ANIMAL S NECROPSIE D 

CONTROL LOW DOSE 

3 (6%) 1 (2%) 

1 (2X) 

(49) (45) 

1 (2«) 

3 (7X) 

1 (2%) 

3 (6X) 1 (2X) 

(49 ) (45) 

1 (2X) 

(52) (50) 

1 (2X) 

(5«*) (53) 

1 (25) 

2 (4X) 

16 (30X) 1 (2X) 

(54) (53) 

2 (4X) 2 (4X) 

(52) (50) 

16 (31X) 5 (12X) 

1 (2X) 

(53) (1,9) 

1 (2*) 

2 (4X) 

(54) (53) 

1 (2X) 

(52) (53) 

1 (2X) 

(52) (53) 

Z-IIZL- . 

 FXABIHED BICBOSCOPICALLY 

HIGH DOSE 

1 (2X) 

(46) 

2 (4X) 

(46) 

1 (2X) 

(43) 

(54) 

(54) 

1 (2X) 

(50) 

(45) 

(54) 

(52) 

(52) „. 

52 
TABLE A2. FEMALE RATS: NEOPLASMS (CONTINUED) 

CONTROL LOW DOSE 

CHBOHOPHOBE CARCINOMA, METASTATI 1 (2%) 

iSTPOCYTOBA 

SPECIAL SENSE OP CANS 

*EYE 

SQUAHODS CELL CA?CINOM A 

(5<*) 

2 (UX ) 

(53) 

*FAB CAHA L 

CERUHINOUS CABCINOHA 

(5i») 

1 (2%) 

(53| 

HUSCULOSKtLETAL SYSTKf. 

NONF 

BODY CAVITIFS 

NONF 

A LL OTHER SYSTEMS 

*KtJLTIPL? ORGAN S (5«) (53) 

AP'JNOCAPCINim, N''S, ''^ASTATIC 1 (2%) 

ANIfl^ L DISPOSITIO N SIMSAP Y 

ANIMAT S INI"'IPLIY T N STUDY 55 55 

NATURA L DLATK.J 6 2U 

SOEIBUNO SACKIFIC^ 1 13 31 

SCHEDOtFD SACPIFIC1; 

ACCID5STAILY KILLED 

TERMINA L SACRI'ICT •?f> ANTKU L MISSING 

»_I]JSI"EBS_A,Ul2II£I2_ftNiaiLS 

» NUHBE9 OF ANIHAL S WITH TISSUE FXAMINE D MICROSCOPICALLY 

* NUMB? ? OF P.NIMAt S N'CPOPSIK D 

HIGH DOSE 

1 (2X) 

(54) 

(54) 

(5«) 

55 

U1 

14 

53 
TABLE A2. FEMALE RATS: NEOPLASMS (CONTINUED) 

CONTROL LOW DOSE HIGH DOSE 

TOHOR SUMMAR Y 

TOTAL ANIMAL S WITH PRIMAR Y TUM03S* 52 50 51 

TOTAL PRIMAR Y TUMORS 103 81 60 

TOTAL ANIMAL S WITH BTNIG N TUMORS as 2U 5 

TOTAL BENIGN TUMORS 67 29 5 

TOTAL ANIMAL S WIT H MALIGNAN T TUMOP S 21 H'4 51 

TOTAL MALIGNAN T TUMORS 29 52 55 

TOTAL ANIKAL S WTTH SECONDAR Y TU.103S* 5 1

 

TOTAL SFCONDAPY TUMORS 6 1

 

TOTAL ANIMAL S WITH TU»ORS UNC'RTAIN ­

BFNIG N O R MALIGNAN T 2

 

TOTAL UNCERTAI N T'HOPS 2

 

TOTAL ANIMAL S WIT H TUMOR S ONCERTAIN ­

PRIMAR Y O R METASTATI C 1

 

TOTAL UNCFHTAI N T;)M)~ ; 3 

* PRIMAR Y TIJ.10RS: »I.T T'J«:)ES FXCE?T SECONDAR Y TUMORS 

» S"CONDAP Y T'JMORi : M^T A STATI C TMJ^OPS 3E TUMOR S INVASIV E INT O A N ADJACEN T ORGA N 

54 
APPENDIX B
 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN
 

MICE FED o-ANISIDINE HYDROCHLORIDE IN THE DIET
 

55
 

TABLE B1.

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN MALE MICE 

FED o-ANISIDINE HYDROCHLORIDE IN THE DIET 

1CONTROL LOW DOSE HIGH DOSE 

ANIMAL S INITIALL Y I N STUD Y 55 55 55 

ANIMAL S MISSIN G 1 

ANIMAL S N^CPO^SIE D 55 55 53 

ANIMAL S "vnNiKS D iIIS1U?!.TKOLOGICALLY 55 55 53 

INTESUfl^NTAF Y SYSTEM 

*SKIR (55) (55) (53)

 

SQUAMOU S CELL PAPILLOM A 1 (2*)

 

*SUBCUT TISSUE (55) (55) (53) 

SQUAMOOS CKLL PAPILLOMA 1 (2*) 

"I3ROMA 2 (i*%) 

"IBROSARCOHA 1 (2%) 1 (2*) 

RESPIRATOR Y SYSTF M 

*NASA L CAVIT Y (55) (55) (53) 

SQUABOUS CELL CARCINOM A 1 (2*) 

#LUN G (5U) (54) (52) 

HEPATOCELL0LAR CAECINOMA , MSTAST U (7X) 1 (2%) 

ALVFOLAP/BRONCHIOLA P ADENOM A 6 (11X) 3 (6X) 2 (IX) 

ALVFOLAR/BRONCHIOLA R CARCINOM A 6 (11X) 6 (11X) 

HSMATOPOIETIC SYSTEM 

*MULTIPLS ORGAN S (55) (55) (53)

 

MALIGNAN T LYMPHOMA , NO S 3 (5X) 

BALIG.LYHPHOHA, HISTIDCYTIC TYPE 1 (2%)

 

MALIGNAN T LYMPHOSA , MTXH D TYPE 2 (US)

 

nHDIFFEREMTIATKD LEUK2MIA 1 (2%)

 

tSPLEEN (51) (55) (52)

 

HEMAHGIOM A 1 (2%)

 

#HESEHTEHIC L. NODE (U8) (4U) (47) 

_ BEP4T3CSLLOIAR CAFCINOMAi . H^Il^T . 1 J2?l 

* NUMBER OF ANIMAL S WITH TISSUE EXAMINE D MICROSCOPICALLY 

* NUMBER OF ANIMALS NECROPSIED 

57 
TABLE B1. MALE MICE: NEOPLASMS (CONTINUED) 

CONTROL LOW DOSE 

tP^NA L LYMP H NOD' (48 ) (44)

 

MALIGNAN T LYMPIiOIA , NJ S 

tJF.JUN'IM (50) (54)

 

MALIGNAN T LYMPHOMA , MIXK D TYPE 1 (2% ) 

CIRCULATOR Y SYSTE M 

NON F 

DIGESTIVE SYSTEM 

tLIVFR (54) (54) 

HUPATOCELLUIA R ADENOM A 4 (7%) 

HSPATOCELLULA R CARCINOM A 24 (44%) 13 (24X) 

"ISROSARCOM A 1 (2X ) 

#PANCPFA S (49) (53) 

FIBPOSAPCOM A 1 (2») 

f STOMAC H (51) (53) 

FIBROSAPCOMA 1 (2%) 

URINAR Y SYSTFM 

#KIDN5Y/P5LVI S (54) (55)

 

TRANSITIONAL-CFL L PAPILLOM A 1 (2%)

 

#URINAP Y BLADDE R (48) (55) 

NF.OPLASM, NOS 1 (2%) 

TRANSITIONAL-CEL L PAPILLOM A 2 (4X) 

TRANSITIONAL-CELL CARCINOM A 

ENDOCRINE SYSTEM 

#ADRENA L (50) (53)

 

CORTICAL ADENOM A 1 (2%) 

tADRENAL/CAPSUL E (50) (53)

 

iD_E.HQJll«- H°S _ 5 J10SI _. 

I NUMBER OF ANIMAL S WIT H TISSUE EXAMINE D MICROSCOPICALL Y 

* NUMBER OF ANIMAL S NECROPSIE D 

HIGH DOSE 

(47) 

1 (255) 

(53) 

(52) 

7 (13X) 

(53) 

(51) 

(51) 

(53) 

7 (13X) 

15 (28X) 

(52) 

(52) 

58 
TABLE B1. MALE MICE: NEOPLASMS (CONTINUED) 

fTHYBOI D 

FOLLICUtAR-CELL ADENOM A 

FOLIICULAB-CELL CARCINOM A 

tPANCBEATIC ISLSTS 

ISLET-CFLL ADFNOMS 

RFPBODUCTIVE SYSTEM 

SON? 

NEPVO'JS SYSTEM. 

NON' 

SPFCIAI. S^NSE OFGRN S 

*HAR r >ERIA N GLAN D 

CYSTADENOKA , NOT 

HUSCOLOSKSLErAI. SYST*"1! 

*SKEL':'TAL MUSCL E 

FIBROSABCCHA 

BODY CAVITIES 

NON=: 

A LT OTHF P SYSTEMS 

OM^NTO M 

"IBROSARCOM A 

* NUMBE R O F ANIKAL S HTT P TISSU E

* NUBBFB OF ANIMAI S N'FCROPSI^ D 

CONTROL LOW DOSE HIGH DOSE 

(i»8) C*9) (39) 

1 (2X) 

1 (2X) 

(«9 ) (53) (53) 

2 (i»X) 

(55 ) (55) (53) 

1 (2%) 1 (2X) 

(55) (55) (53 ) 

1 (2X) 

1 

 EXAMINE D MICROSCOPICALLY 

59 
TABLE B1. MALE MICE: NEOPLASMS (CONTINUED) 

CONTROL LOW DOSE HIGH DOSE 

ANIMA L DISPOSITION SUMMAR Y 

ANIMAL S INITIALT.Y I N STUDY 55 55 55

 

NATUPA L DEATH S 9 U 6

 

MORIBUN D SACRIFICE 2 8 4

 

SCHEDULE D SACRIFICE 

ACCIDENTALL Y KILLE D 1

 

TERMINA L SACRIFICE U U 43 43

 

ANIMA L MISSIN G 1

 

3 INCLUDE S AUTOLY7FT 1 ANIMAL S 

TUMO R SUMMAR Y 

TOTA L ANIMAL S WIT H PRIMAR Y TUMORS * 43 27 30 

TOTA L PRIMAR Y TUMO? S 59 37 35 

TOTAL ANIMAL S WIT H FTNIS N TUMOR S 22 5 11 

TOTAL BENIG N TUMOR S 21* f> 11 

TOTAL ANIMAL S WITH MALIGNAN T TUMOR S 29 25 23 

TOTA L MALIGNAN T TU10RS 35 30 24 

TOTAL ANIMAL S WIT' 1 S?CCND»R Y TUMORS * U 1 

TOTA L SECONDAR Y TU*OP S 5 1

 

TOTA L ANIMAL S KIT H VJK03S IINC' STAINBENIG N O P MALIGNAN T 1

 

TOTAL UNCKPTAI N THMOE S 1

 

TOTA L ANIMAL S WIT H TUMOR S iJNC^P T 'INPRIMAR Y O R MTASTATIC

 

TOTAL UNCERTAI N T?IMJR S 

* PRIMAR Y TUMORS : AL L T'JMOR S 5XCFPT SECONDAR Y TOMORS 

f SFCONDAH Y TUKOPS : KFT?. STATIC TUMOR S 3R TU!1oaS INVASIV F INTO AN ADJACEN T 03GA N 

60 
TABLE B2.

 

SUMMARY OF THE INCIDENCE OF NEOPLASMS IN FEMALE MICE 

FED o-ANISIDINE HYDROCHLORIDE IN THE DIET 

CONTROL LOW DOSE HIGH DOSE 

ANIMAL S INITIALL Y IN STUD Y 55 55 55 

ANIMALS NECROPSIES 55 52 54 

ANIMAL S EXAMINE D HISTOPATHOLOGICAL L Y 55 52 5U 

INTEGUMFNTAR Y SYSTEM 

*SUBCOT TISSUE (55) (52) (5U) 

CARCINOMA.NO S 1 (2X) 

SQOAHOUS CELL CARCINOM A 1 (2X) 

ADENOCARCINOHA , NOS 1 (2X) 

LEIOMYOSARCOM A 1 (2X ) 

MEHANGIOH A 1 (2%) 

HEMANGIOS'ARCOM A 1 (2X) 

RESPIRATOR Y SYSTE M 

*LUNG (55) (51) (52) 

ADENOCARCINOHA , NOS, METASTATIC 1 (2X) 

HEPATOCELLULAR CARCINOMA , METAST 2 (4%) 

ALVEOLAR/BRONCHIOLA P ADENOM A 3 (555) 2 (««) 1 (2X) 

ALVBOLAR/SRONCHIOLAS CARCINOM A 1 (2%) 

HEMATOPOIFTIC SYSTEM 

•MULTIPLE ORGAN S (55) (52) (5«) 

MALIGNAN T LYMPHOHA , HOS 1 (2%) 9 (17X) 5 (9X) 

MALIG.IYMPHOHA , LYMPHOCYTIC TYPE 2 («*) 

MALIG.LYMPHOMA , HISIIOCYTIC TYPE 1 (2%) 

MALIGNAN T LYMPHOMA , MIXE D TYPE 6 (11S) 

LYHPHOCYTIC LEUKEMIA K (7X) 

tSPtSEN (53) (52) (51) 

NEOPLASH, NOS 1 (2X) 

HEMANGIOSARCON A 2 (4X) 1 (2X) 

MALIGNAN T LYMPHOMA , NOS 1 (2X)

 

MALIGNAN T LYMPHOMA , MIXE D TYPE 1 (255) 

tNANDIBULAR L. NODE (U7) (U5) (47) 

A.DE.fiQ£AIC.INQflA.i_MQSt US1AS2A1IC _ 1_12XL_ 

# NUMBER OF ANIMAL S WITH TISSUE EXAHINB D MICRDSCOPICH.LY 

* NUMBER OF ANIMAL S NECPOPSIBD 

61 
TABLE B2. FEMALE MICE: NEOPLASMS (CONTINUED) 

HIGH DOSE 

(47) 

(53) 

(»1) 

1 (2%) 

(51) 

(53) 

4 (8X) 

(52) 

(53) 

(52) 

(54) 

1 (2X) 

(50) 

IMES^NTFPIC L. NODr! 

MALIG.LYtiPHOSA , HISTIOCYTIC TYPF 

SALIGNANT LYMPHOHA , MIXE D TY ??. 

tLIVFB 

MALIGNAN T LYMPHO?"» , NOS 

#THYMU S 

THYMOM A 

1ALIGNAN T LYHPHOHA . NOS 

CIRCULATOR Y SYSTE M 

tH^AFT 

HEMANGIOM A 

DIGESTIVE SYSTEM 

•LITER 

HEPATOCELLULA P AD'NOH A 

NEOPLASTIC NODULE 

H5PATOCELLITLA S CARCINOH? . 

HEMANGIOM A 

tPANCPEA S 

NEOPLASM , NO S 

•STOMACH 

NFOPLASH, NOS 

SOUAHOO S CELL PAPILLOM A 

*DOODENUn 

NEOPLASH , NOS 

*ANO S 

SOUAHOU S CELL PAPILI OMA 

URINAR Y SYSTEM 

*URINAP Y BLADDER 

8£2ElA5fl*-l!03 ___ 

I NUHB5R OP ANIMAL S WITH TISSUE

* NUMBER OF ANIHAL S NECROPSIED 

CONTROL LOW DOSE 

(47) («5) 

1 (2%) 

2 (H%) 

(5«) (52)

 

1 (2%) 

(35) (31) 1 (3*) 

1 (33S) 

(55) (51)

 

1 (2%) 

(5U) (52) 

H (7%) 

1 (2X) 7 (13«) 

1 (2%) 

(«9) (52) 

1 (28) 

(53) (52) 

1 (2X) 2 (US) 

(52) (52) 

1 (2X) 

(55) (52) 

(50) (51) 

1 12XL 

 EXAMINE D MICROSCOPICALLY 

62 
TABLE B2. FEMALE MICE: NEOPLASMS (CONTINUED) 

CONTROL LOW DOSE 

TRUNSITIONAL-C^I.I, PAPILLOM A 1 (2%) 

TRANSITIO N AL-CELL CARCINOM A 

ENDOCRINE SYSTEM 

tPITOITARY (42 ) (UO )

 

CHROHOPHOBE- ADENOM A 2 (5%)

 

BASOPHIL ADENOMA . 1 (2?)

 

tADRENA L (50) (52) 

PHEOCHROKOCYTOH A 1 (2% )

 

ITHYROI D (48) (39) 

FOLLICULAR-CELI. ADENOM A 1 (2%) 1 <3X ) 

PE PRODUCT IV SYSTF M 

*MAMHAP Y GLAN D (55) (52) 

ADENOCAPCINOMA , NO S 1 (2%) 

ACINAR-CEL L CAKCIJiOK A 1 (2%} 

"IBPO ADEN01 A 1 (2% ) 

tUTEPUS (54) (51) 

NFOPLASH , NOS, ^AT.ir.NAN T 1 (2% ) 

?NnOM"TPIA L STPOMS L POLY P 1 (2X ) 

* OVAR Y (50) (46) 

TEPATOMA , NO S 1 (2%) 

HEflANGIOM A 1 (2») 

NTPVOri S SYSTFfl 

NONF 

SPECIAL SENSE ORGAN S 

*HARDERTA N GLAN D (55) (52)

 

ADFNOHA , NO S 

*EXTFRNAI. FA R (55) (52) 

'IBROSAECOM A 1 (2%) 

MUSCULOSKELFTA ! SYS^F M 

NON F 

« NUMBE R O F ANIMAL S WIT H TISSU E EXAMINE D MICROSCOPICALL Y 

* NUMBE P OF ANIMAL S NECPOPSIE D 

HIGH DOSE 

4 (8%) 

18 (36X) 

(43) 

(52) 

(38) 

(54) 

(50) 

(52) 

(54) 

2 (4X) 

(54) 

63 
TABLE B2. FEMALE MICE: NEOPLASMS (CONTINUED) 

BODY CAVTTIFS 

*BO^Y CAVITIE S 

NESOTHELIOMA , NOS 

A LL OTHER SYSTEMS 

* MULTIPLE ORGAN S 

NEOPLASM, NOS 

CSINTUM 

NEOPLASM, NOS 

ANTNA L DISPOSITION SIIKMS.R Y 

ANIMAL S INITIALL Y IS <iTUn Y 

NATURA L DEATH S 

«0?IBMNr SACRIFICE 

SCHEDULFD SACPIFIC P 

ACCIDENTALL Y KILLF ? 

TSPBINA L SACBfTC" 

ANIMAT , NISSIH G 

1 UJ£1"DES AUTOLY? 7 " " ANT HAL 3 

• NU1BEP O F ANIMAL S WIT H TISSU E

* NTIMP"? OF ANIMAL S N^CFOPSIF D 

CONTROL LOW DOSE 

(55) (52) 

1 (2%) 

(55) (52) 

1 (2X) 

1 

55 55 

7 12 

4 5 

l*U 38 

 EXAHINF D HICROSCDPICALL Y 

HIGH DOSE 

(5U) 

(5U) 

55 

5 

7 

1 

«2 

64 
TABLE B2. FEMALE MICE: NEOPLASMS (CONTINUED) 

CONTROL LOW DOSE HIGH DOSE 

TUHO R SUMMAB Y 

TOTAL ANIHAI S WIT H PRIMAR Y TUMORS * 34 20 33

 

TOTA L PRIMAR Y TUMOP S 53 30 41

 

TOTAL ANIMAL S WIT H BENIG N TUMOR S 15 6 7

 

TOTAL BENIGN TUMORS 19 6 8 

TOTAL ANIMAL S WIT H MALIGNAN T TUMOR S 27 13 29 

TOTAL MALIGNAN T TUMORS 30 15 33

 

TOTA L ANIMAL S WIT H SECONDAR Y TUMORS * 2 1

 

TOTA L SECONDAR Y TUMOR S 2 2 

TOTAL ANIMAL S WIT H TUMOR S UNCERTAIN ­

BFNIG N OF MALIGNAN T 1 4

 

TOTAL UNCERTAIN TUMORS 1 9 

TOTAL ANIHAL S WITH TUMOR S UNCERTAIN ­

PRIMAR Y OR HSTASTATIC 

TOTAL UNCERTAI N T1IM3RS 

* PRIMARY TUMORS: ALL TUMORS ^XCEPT SECONDARY TUMORS
 

t SECONDARY TUMORS: .1KTASTATIC TUMORS OR TUMORS INVASIVE IHTO AN ADJACENT ORGAH
 

65 

APPENDIX C
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS
 

IN RATS FED o-ANISIDINE HYDROCHLORIDE IN THE DIET
 

67
 

TABLE C1.

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN MALE RATS 

FED o-ANISIDINE HYDROCHLORIDE IN THE DIET 

CONTROL LOW DOSE HIGH DOSE 

ANIMALS INITIALLY IN STUDY 55 55 55 

ANIMAL S NECPOPSIED 5U 55 53 

ANIMAL S EXAMINE D HISTOF ATH3I.OGIC ALL Y 5U 55 53 

INTEGUMENTAR Y SYSTEM 

*SKIN (5U) (55) (53) 

INFLAMMATION , NrTCPOTTZIN' ? 1 (2X) 

*SUBCUT TISSUE (5«) (55) (53) 

ABSCESS, NOS 1 (2X) 

RESPIRATOR Y SYSTEM 

tTRACHF A (51) (53) (U7 ) 

INFLAMMATION , CHRONIC 1 (2X ) 

METAPLASIA , SQUAMOU S 1 (2X ) 

tlUNG/BRONCHUS (5U) (55) (51) 

BHONCHIECTASIS 1 (2% ) 

tLUNG (5U) (55) (51) 

BFONCHCPNEUMONIA, NOS 2 (4X) 

BRONCHOPNEUBONIA , ACUTE 1 (2X) 

PNEUMONIA , CHRONIC MORIN E 2 (HV> 2 (<*X) 

HEMATOPOIETIC SYSTEM 

•BONE MARPOW (52) (55) (47) 

FIEROSIS, FOCAL 1 (2X) 

HYPERPLASIA , NOS 7 (13*) 

ISPLEEN (54) (55) (52) 

FIBROSIS, FOCAL 1 (2«) 

METAMORPHOSI S FATTY 1 (2») 

HEMOSIDEROSIS 1 (2%) 

HEHATOPOIESIS 1_12SL_. 

f NOMBFP OF ANIMAL S HITH TISSUE EXAMINE D MICROSCOPICALLY 

* NUMBER OF ANIMAL S NECEOPSIED 

69 
TABLE C1. MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

CONTROL LOW DOSE HIGH DOSE
 

ERYTHROPOIESIS 1 (2%)
 

tlYHPH NODE (53) (ft?) (3ft)
 

HYPERPLASIA, NOS 1 (2%)
 

fHANDIB'JLAR L. NODE (53) (ft?) (3ft)
 

HYPERPLASIA, NOS 1 (2X)
 

HYPFRDLASIA, PLASMA CELL 1 (2X)
 

*LOHBAR LYHPH NODE (53) (ft?) (3ft)
 

HYPERPLASIA, PLASMA CELL 1 (2<)
 

CIRCULATORY SYSTEM
 

tHEART (5U) (55) (51)
 

THROMBUS, MORAL 1 (2%)
 

PFPIARTFBITIS 1 (2«)
 

NECROSIS, POCAL 2 ((»%)
 

CALCIFICATION, NOS 3 (6X)
 

*!1YOCAPDIU« (5<*) (55) (51)
 

INFLASMATION, FOCAL 1 (2%)
 

DEGENERATION, NOS 16 (33X)
 

tCARDIAC VALV F (5H) (55) (51)
 

INFLA1MATION, CHRONIC 1 (2X)
 

*AORTA (51) (55) (53)
 

MEDIAL CALCIFICATION 1 (2X) 3 (6X)
 

*CORONARY ARTEPY (5ft) (55) (53)
 

«FDIAL CALCIFICATION 2 (ftX)
 

*CELIAC ARTEPY (5ft) (55) (53)
 

THROMBOSIS, NOS 1 (2*)
 

DIGESTIVF SYSTEM
 

ILIVER (5ft) (55) (52)
 

CHOLANGIOFIPROSI^ 9 (17*)
 

NECROSIS, FOCAL 1 (2X)
 

NFCROSIS, FAT 1 (2%)
 

aSTAaORPEOSIS "ATIY 1 12SL__.
 

t NUSEER OF ANIMALS WITH TISSUE EXAMINED RICRDSCOPICALLY
 

* NDMBSP OF ANIMALS NFCFOPSIED
 

70
 
TABLE C1. MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

CONTROL LOW DOSE HIGH DOSE 

BASOPHILIC CYTO CHANG S 2 IUX) 1 (2X) 

CLEAR-CEL L CHANG ? 1 (2<) 

*BILE DUCT 

INFLAMMATION , NOS (5t)1 (2?) 

(55) (53) 

tPANCREA S (53) (50) (51) 

DILATATION/DacTS 

INFLAMMATION , CHRONI C 

1 (2%) 

1 (2*) 

"IBROSIS 1 (2Jt( 

•STOMAC H (53) (55) (52) 

ULCF.P, NOS 

EROSIO N 

HYPEPPLASIA , BAS« I "5LL 

2

1

1H

 (U% ) 

 (2% ) 

 (26*) 

tGASTSIC MUCOSA (53) (55) (52) 

ULCFR , NOS 1 (2X) 

CALCIFICATION ,

CALCIFICATION ,

 NO S POCAL 

1 (2St) 2

1

 (Hit) 

 (2«) 

URINAP Y SYSTEM 

t KIDNE Y 

HYDKONFPHFOSI S 

(53) (55) 

2 (45S) 

(53) 

3 (63t) 

PYELONEPHRITIS , N'35 1 (235) 

PYELONEPHRITIS . AC'ITL 

N?PHROSIS, NOS 

NFPHPOSIS , CHOL'-""TC 

OLOMEPULOSCLE3CST3 , SOS 

26

2

 (49 S) 

C*1"!) 

U3 (78S| 

1

5

2

 (21) 

 (9)1) 

 (4*) 

tKIDNFY/MEDULL A (53) (55) (53) 

CALCIFICATION , NO T 1 (2?,) 

tPFNA L PAPILL A (53) (55) (53) 

I NFL A MMAT I ON , S'J? ?-U~ A TI V* 

NECROSIS , NOS 5 (9*) 

1

3

 (2% ) 

 (6%) 

CALCIFICATION , UOS 1 (2X| 3 (6?) 

tKIDNFY/TUB'JIr (53) (55) (53) 

NF.PHROSTS, NOS 1 (2") 

NSCPOSIS, NOS 

CALCI'TCATIOS, ??03 

1 (295) 

2 (4X| 

1

2

 (2<) 

 (4% ) 

tKTONFY/PFLVI S (53) (55) (53) 

* NUMBF " OF ANIKAL S VI? ;? 'ISS'IE FXAMIK5 D KIC8DSC3? 1C ALL Y * NUMBE R OF ANIMAL S H'C"OPSr3D 

71 
TABLE C1. MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

CONTROL LOW DOSE 

INFLAMMATION , ACIJT? FOCA L 

HYPE^PLASIA , 5PITHELIA L 

*URTNAP Y BLADDE R (51) (54)

 

HYPFPPLASIA , EPITHELIA L 1 (2%) 

• TJ.BLADDEP/SUBMUCOS A (51) <5«)

 

HYP^EPLASIA , NO S 1 (2%)

 

*PROSTATIC URETHR A (5U) (55)

 

INFLAMMATION , SUPPURATIV E 

ENDOCRIN E SYSTE M 

IPITUITAR Y C»8) (49) 

HEMORRHAG E 1 (2%)

 

HYPFRPLASIA, POCAL 1 (255) 

HYPEPPLASIA , BASOPHILIC 2 (H%)

 

• ADRENA L (54) (55)

 

CYST, NOS 1 (255) 

• ADRENA L CORTE X (5U) (55)

 

HYPERPLASIA , NOS 1 (2%) 

HYPEPPLASIA , FOCAL 1 (2X)

 

tTHYFOID (53) (40)

 

CYSTIC FOLLICLES 10 (25X)

 

FOLLICULAR CYST, NOS 

CALCIFICATION, FOCAL 

HYPEEPLASIA , POLLIC'ILAR-CELL 

tPAHCREATIC ISLETS (53) (54)

 

HYPEPPLASIA , NOS 1 (2*)

 

REPRODUCTIVE SYSTE M 

*PFEPOTIAL GLAN D (5U) (55)

 

INFLAMMATION , ACUT E 1 (2X|

 

ABSCESS, NOS 1 (2%) 

INFLAMMATION , CHRONI C 1 (2*)

 

•PROSTATE (52) (52) 

INFLlflflillQH*. SU.PPUBA.TLVS __ 6 J12JL . 

* NOHBEE OF ANIMALS WITH TISSUE EXAMINED MICROSCOPICALLY
 

* HOHBBE OF ANIMALS NSCROPSIED
 

HIGH DOSE 

1 (2%) 

2 (4X) 

(52) 

(52) 

(53) 

1 (2X) 

(47) 

(53) 

(53) 

(40) 

3 (8*) 

3 (8*) 

1 (3«) 

3 (8X) 

(51) 

(53) 

(52) 

_Z_ill«L_ 

72
 
TABLE C1. MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

CONTROL LOW DOSE 

INFLAMMATION , SCUT? 

INFLAMMATIO N AC'JTE ^N D C^ONIC 1 (2%) 

•T^STIS (:,4> (52) 

ATROPHY , NOS 

*SCROTUM (S«) (55) 

ULCER, AC 071? 1 (2X) 

NEFVOUS SYSTFM 

tBPAIN/HENINGE S (54) (53) 

INFLAMMATION , NOS 1 (2X) 

#CESEBRA L VENTFICLS (54) (53) 

HSMOESHAG ? 1 (2?)

 

tBBAT N (S4) (53) 

HKMOPBHAG F 1 (2X )

 

SPFCIAI SENSE ORGAN S 

*EYS (5«) (55) 

HF.MOPRHAG E 1 (2K ) 

*EYE/IRIS ( c

i U) (55)

INFIABKATION , CHRONIC 1 (2X) 

*FYF/CRYSTALLIN E LFNS (54) (55) 

CALCIFICATION, NOS 1 (2K) 

*FYE/LACRIHAL GLAN D (54) (55) 

INFLAMMATION , NOS 1 (2%)

 

MUSCULOSKEtSTAL SYSTFH 

NONE 

BODY CAVITIES 

*ABDOMINA L CAVITY (54) (55) 

NECROSIS, FAT ,.9 J1ZXL _

 

t NUMBE R OF ANIHAL S WITH TISSUE EXAHIHED HICRDSCaPICALLY 

* NUMBER OF ANIMAL S NECHOPSIED 

HIGH DOSE 

1 (2S) 

(51) 

3 (6*) 

(53) 

(51) 

(51) 

(51) 

(53) 

(53) 

(53) 

(53) 

(53) 

73 
TABLE C1. MALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

CONTROL LOW DOSE HIGH DOSE 

ftLL OTHFR SYSTFM S 

NON E 

SPECIAL MORPHOLOG Y SUMMAR Y 

NO LESION REPORTE D 1 

MJTOLYSIS/NO NECROPS Y 1 2 

« NHMBF P Ov ANIKAL S WIT H TISSU E EXAMINE D MICROSCOPICALL Y 

* NUMBE R O F AKIMAL S N^CPOPSI^ D 

74 
TABLE C2. 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN FEMALE RATS 

FED o-ANISIDINE HYDROCHLORIDE IN THE DIET 

COMTROL LOW DOSE HIGH DOSE 

ANIMAL S INITIALL Y IN ST'.'DY 5 '5 55 55 

ANIMAL S NECROPSISD 54 53 5« 

ANIMAL S EXAMINE D HIS70PATHOLOGICALL Y 5'* 53 5U 

INTEGUMENTAR Y SYSTEM 

*SKIN (S4 ) (53) (5«) 

ULCFR, "OCAL 1 (2«) 

RHSPIRATO» Y SYSTE M 

#LUNG/BRONCKU S (33) (53) (5U) 

INFLA1MATION , S"P?UF AT I V F 2 (HI) 

fLMN G (51) (53) (5H) 

BRONCHOPN2UBONTA , NOS 1 (2«) 

PNEUMONIA , CHRONI C MU&IN E 3 (6%) 

MKTAPLASIf , NO S 1 (2«) 

tALVFOLA P WAL L (S3) (53) (5U) 

CALCIFICATION , NO ? 1 (2%) 1 (2^) 

HEHATOOOIFTIC SYST^K 

tBON F MAPRO K (53) («9) (51) 

HISTIOCYTOSI S 1 (2? ) 

#SPLEE N (52) (52) (51) 

H^HATOPOIFSI S 1 (2"-,) 

EPYTHROPOIFSI S U <8X) 

tCFRVICA L LYMP H NOD ^ (51 ) (46) (36) 

HYPEBPIASIA , PTASM A C^L L 1 (2«) 

tLIMEJV R LYMP H NOD ^ 

INFLAMHATTON . CHPOSTC 

aiE^EPiiSIA,. NOS _ . 

(51 ) 

1 (2«) 

. ­

(<*6) 

_ 1 l&L 

(36) 

t NUM3F R OF ANIMAL S WI.T M TISS'I F FXASINS D MIC33SCOP 1C AL LY 

* NUMEFB OF ANIMAL S Nrc°0?SIFD 

75 
TABLE C2. FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED)

 

*PSNAL LYMP H NOD E 

INFLAMMATION , CHEONI C 

HYPERPLASIA , NO S

 

CIPCULATOP. Y SYSTE M 

#HEAP T 

CALCIFICATION , NOS 

*MYOCAPDIU M 

DEGENERATION , NO S 

*AORT A 

MEDIA L CALCIFICATIO N 

*CORONAP Y ARTER Y 

MEDIA L CALCIFICATIO N

 

DIGESTIVE SYSTEM 

tLIVER 

CHOLANGIOFIBROSI S 

NECROSIS, NOS 

METAMORPHOSI S "ATTY 

BASOPHILIC CYTO CHANG E 

ANGIECTASI S 

tPANCREA S 

ATROPHY , FOCAL 

ISTOMACH 

INFLAMMATION , NOS 

OLCER, NOS 

HLCEF, ACUTE 

CALCIFICATION, NOS 

HYPERPLASIA , BASAL CELL 

tGASTRIC MUCOSA 

OLCER, FOCAL 

CALCIFICATION, NOS 

URINAR Y SYSTEM 

fKIDNEY 

^MBBYOMftl DUCT CYST 

t BUHBER OF ANIMAL S WIT H TISSUE

* NUMBER OF ANIMAL S NECROPSIED 

CONTROL LOW DOSE 

(51) C*6) 

1 (2%) 

1 (2X) 

(53) (53) 

1 (2%) 

(53) (53) 

6 (11X) 1 (2X) 

(54) (53) 

2 (4X) 

(54) (53) 

2 (4X) 

(53) (53) 

2 (4%) 

1 (2X) 

6 (11X) 

10 (19X) 10 (19X) 

1 (2X| 

(52) (50) 

1 (2X) 

(51) (51) 

1 (2X) 

1 (2X) 

1 (2X) 

1 (2X) 

11 (22X) 

(51) (51) 

1 (2X) 

1 (2X) 

(52) (52) 

1.J2IL 

 EXAHIHBD HICR3SCOPICALLY 

HIGH DOSE 

(36) 

(53) 

(53) 

(5tt) 

1 (2X) 

(54) 

(53) 

(<*3) 

(49) 

(U9) 

1 (2X) 

(54) 

76 
TABLE C2. FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

CONTROL LOW DOSE 

HYDRONEPHROSI S 7 (13X) 

PYELONEPHRITIS , NO S 

NEPHSOSTS , NOS 2 (H%) 3 (6S) 

NFPHPOSIS, CHOLHMIC 1 (2%) 

SLOHFRULOSCLEROSTS , NO S 1 (2% ) 

CALCI»ICATION , NO S 1 (2X) 

CALCIFICATION , 'or>t 5 (101?) 

HYPBRPLASIA , TUBULA? CELL 

tRFNA L PAPILL A (52) (52) 

INFLAMMATION , SUPPURAT I VF , 1 (2X) 

INFLAMMATION , ACtJTF 

N'CPOSIS, NOS 6 (12X) 

CALCIFICATION, NOS 5 (10*) 

*KIDNFY/TU3ULS (52) (52) 

CALCIFICATION , NOS 3 (63t) 

#KTDNFY/PELVI S (52) (52) 

INFLAMHATION , ACUTS 1 (2*) 

HYPEBPLASIA , "PITH^LIA L 2 (4Jt) 

IURINAB Y BLADDER (49 ) (49) 

CALCIFICATION, NOS 1 (2X> 

EBDOCFINF SYSTEM 

tPITUITARY 

HYPERPtASIA , NOS 

l|l»fl) 

1 (2«) 

(51) 

ANGI'-ICTASIS 1 (2X» 

tADRFHA L CORTEX f53 ) (53) 

HYPERPLASIA , NOS 1 (2%) 

*ADSFNAL MEDULL A 

NTCPOSIS, NOS 

(53) 

1 (255) 

(53) 

•THYPOID I[I49) (45) 

CYSTIC FOLLICLFS 3 (7X) 

HYPEEPLASIA , C-CELL 

HYPERPtASIA , FOLLICOLAR-CELL 

tPARATHYROID (15) (12) 

_ -.MEEimSIA.x.-.N.SS . 1 IIS) . 

* NOHBEF OF ANIMAL S WITH TISSUE EXAMINE D MICROSCOPICALLY 

» NUMBER OF ANIMAL S NECROPSIED 

HIGH DOSE 

14 (26%) 

3 (6S) 

1 (2X) 

(54) 

1 (2«) 

20 (37X) 

5 (9«) 

(54) 

2 (UX) 

(54) 

(51) 

(45) 

(54) 

(54) 

(46) 

3 (7X) 

2 (4X) 

2 (4X) 

(17) 

77 
TABLE C2. FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED) 

•PANCREATIC ISLETS 

HYPERPLASIA , NOS 

REPRODUCTIVE SYSTE M 

*MANMAP Y GLAN D 

GALACTOCEL^ l 

*CLITOPAL 61 AN D 

INFLAMMATION , ACU"F 

tUTBRtJS 

HYDPOMETRA 

THROMBOSIS , NOS 

ABSCESS, NOS 

tC^RVIX UTFRI 

POLYP, TNM.AHM'LT'OF Y 

*UT^RT?S/3NDO?IETRII"1 

INFLAMMATION , Sl'PP UP A TI V £ 

INFI AHHATION , ACTTF 

ABSCESS, NOS 

HYPF.RPLAS I A, CYSTIC 

#OVA3Y/OVIDUCT 

INFLAKMATION , SttPPUSATIV? 

ABSCESS, NOS 

tOVARY 

CYST, NOS 

INFLAMMATION , SUPPURATIV 2 

IN"LA«!MATION , CHRONI C 

NERVOU S SYSTFK 

NON ^ 

SPFCIAL SENSE ORGAN S 

* EYE/COR N "A 

INFLAMMATION . ACU^ F 

f NUM3E P OF AN'IMAL S WIT H TISSU E

* NUMBER OF ANIMAL S K?C30?SI*D 

CONTROL LOW DOSE 

(52) (50) 

1 (2%) 

(5U) (53) 

6 (11*) 

(5U ) (53) 

H (8X) 

(52) (50) 

2 (H%) 

1 (2%) 

1 (2X| 

(52) (50) 

1 (2< ) 

(52) (50) 

1 (2X ) 

1 (2X) 

1 (2«) 

3 (6X) 

(52) (50) 

1 (2X ) 

(53) (09) 

1 (2*) 

1 (2%) 

1 (2?) 

(5U) (53) 

1 12»L_

 EXAMINE D MICROSCOPI- A LLY 

HIGH DOSE 

(43) 

(54) 

(54) 

1 (2%) 

(50) 

(50) 

(50) 

1 (2X ) 

(50) 

1 (2X) 

(«5) 

(54) 

. 

78
 
TABLE C2. FEMALE RATS: NONNEOPLASTIC LESIONS (CONTINUED)

 

COPJTROL LOW DOSE HIGH DOSE 

MUSCULOSKELETA L SYSTE M 

*SKI>LETAL MUSCLE (54) (53) (5i*) 

METAPLASIA , OSSEOUS 1 (2%) 

BODY CAVITIFS 

*A3DOMINA L CAVIT Y (54) (53) (54) 

NECROSIS , FAT 6 (111?) 

A L L OTHFP SYSTEM S 

NON" 

SPECIA L MORPHOLOG Y SUMMAR Y 

MO LESION PEPORTrT) 

A'TTOLYSIS/NO NFCEO !'

l

" Y 1 2 

1 

1 

* NUMBE R O F ANIMAL S V L'l'i TISSU E £XAKINE D MICROSCOPICALL Y 

* NUMBF P OF ANIBAI. S MICFOPS I tD 

79 

APPENDIX D
 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS
 

IN MICE FED o-ANISIDINE HYDROCHLORIDE IN THE DIET
 

81
 

TABLE D1. 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN MALE MICE 

FED o-ANISIDINE HYDROCHLORIDE IN THE DIET 

CONTROL LOW DOSE HIGH DOSE 

ANIMM S INITIALL Y I N STUD Y 55 55 55 

ANIMAL S MISSIN G 1 

ANIMAL S NFCROPSIKD 55 55 53 

ANIKHL S EXAMINE D HISTOPATHOLOGICALL Y 55 55 53 

INTEGUMENTAR Y SYSTFR 

*SKIN (55) (55) (53) 

EPIDERMA L INCLUSION CYS T 1 (2%) 

POLYP, INFLAMMATOR Y 1 (2%) 

RESPIRATOR Y SYSTEM 

NON E 

HEBATOPOIETIC SYSTEM 

fSPLEEN (51) (55) (52) 

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

tNESENTERIC L. NODE (i»8) (<K») (47) 

CONGESTION, NOS 6 (13%) 

HYPERPLASIA , NOS 1 (2S) 2 (5X) 

HISTIOCYTOSIS 1 (2K) 

HYPERPLASIA , HEM ATOP3IETIC 1 (2X) 

CIRCULATORY SYSTEM 

NOME 

DIGESTIVE SYSTEM 

tLIVER (54) (5U) (52) 

NECROSIS, NOS 

METAMORPHOSI S FATTY 

1 (2J) 

3 (6X) 

BESiLQCXIQSIg .. .. _ _3_J[6|L 

t NUMBER OF ANIMALS WITH TISSUE EXAMINED HICR3SCOP1CALLY 

* NDHBEE OF ANIMALS NECFOPSIED 

83
 
TABLE 01. MALE MICE: NONNEOPLASTIC LESIONS (CONTINUED)

 

HYPERPLASTIC NOPUI"
 

HYPEPPLASI A, NOS
 

ANGIECTASIS
 

HFMATOPOIESIS
 

tPANCPEAS
 

CYSTIC DUCTS
 

ATROPHY, NOS
 

#STOMACH
 

INFLAMMATION, ACUTF.
 ATYPIA, NOS
 

HYPERPLASTIC NODULE
 

fPEYPRS PATCH
 

HYPERPLASIA, NOS
 

UPINAPY SYSTFM
 

IKIDNEY
 

HYDRONEPHROSIS
 

CYST, NOS
 

PYELONEPHRITIS, ACUTE
 

PYELONEPHRITIS, CHRONIC
 

tKIDNEY/GLOMERULUS
 

AMYLOIDOSTS
 

tURINARY BLADDER
 

CALCUtOS, NOS
 

HYPURPLASIA, EPITHELIAL
 

ENDOCRINE SYSTEM
 

*PANC8EATIC ISLETS
 

HYPERPLASIA, NOS
 

REPRODUCTIVE SYSTFM
 

*PR5POTIAL GLAND
 

CALCULUS, NOS
 

tPROSTATE
 

OELiJUAIIoi!,, ACUTE
 

* NUMBER OF ANIMAL S WITH TISSUE

* NUMBER OF ANIMAL S NBCBOPSIED 

CONTROL LOW DOSE
 

1 (2S)
 1	 (2T.)
 

(1*9) (53)
 

1 (2X)
 

1 (2*)
 

(51)	 (53)
 

1 (2%)
 

1 (2%)
 

1	 (2X)
 

(50)	 (5«)
 

1 (2%)
 

(50)	 (55)
 

1 (2*)
 

1 (2X)
 

1 (2X)
 

1 (2%)
 

(54) (55)
 

(48)	 (55)
 

1 (2%)
 

1 (2S) 2 (IX)
 

(U9) (53)
 

12 (2H%)
 

(55)	 (55)
 

1 (2%)
 

(52)	 (53)
 

1
 .111,1
 

 EXAMINED MICROSCOPICALLY 

HIGH DOSE
 

1 (2S)
 

1 (2R)
 

2 («*)
 

(53)
 

(51)
 

(53)
 

(51)
 

(51)
 

1 (2«)
 

(53)
 

21 (UOX)
 

(53)
 

(53)
 

(51)
 

84 
TABLE 01. MALE MICE: NONNEOPLASTIC LESIONS (CONTINUED) 

NERVOUS SYSTEM 

NONE 

SPECIAL SENSE 03GAN S 

* EY3 

INFLAMMATION , ACUT? 

CATARAC T 

*HARDEPIA N GLAN D 

INFLAMMATION , ACt)"1?: 

MUSCULOSKELETA L SVST'1 

NON E 

BODY CAVITIES 

*ABDOMINA L CAVIT Y 

NECROSIS, FAT 

*MESFNTEPY 

INFLAMMATION , ACUT E FOCAL 

A LL OTHER SYSTEM S 

OMENTU M 

HFMATOMA , NO S 

SP'CIAL MORPHOLOG V SMMNA8 Y 

NO LFSION REPOPTT D 

ANIMA L MISSING/N O NECROPS Y 

AUTO/N"CROPSY/HISTO PTP F 

AOTOLYSTS/NO NECROPS Y 

f NUMBE R O F ANIMAL S WITH TISSU E

* NUMBE R OF ANIMAL S N*CFO?SI?D 

CONTROL LOW DOSE 

(55) (55) 

1 (2%) 

1 (2%) 

(55) (55) 

(55) (55) 

5 (9S) 

(55) (55) 

1 (2%) 

1 

8 23 

1

1 

 EXAMINE D MICROSCOPI~ALL Y 

HIGH DOSE 

(53) 

(53) 

1 (2%) 

(53) 

(53) 

6 

1 

1 

85 
TABLE D2.

 

SUMMARY OF THE INCIDENCE OF NONNEOPLASTIC LESIONS IN FEMALE MICE 

FED o-ANISIDINE HYDROCHLORIOE IN THE DIET 

CONTROL LOW DOSE HIGH DOSE 

ANIMAL S INITIALL Y IN STUD Y 55 55 55 

ANIMAL S NECPOPSIED 55 52 54 

ANIMAL S EXAMINE D HISTO? ATHOLOGICALL Y 55 52 54 

INTSGHSENTAR Y SYSTEM 

NON F 

RFSPIRATOSY SYSTE M 

UlUN G (55) (51) (52)

 

ATFLECTASIS 1 (2*)

 

HEMATOPOTETIC SYSTEM 

fBON E HARRO W (52) (51) (51)

 

MY^LOFIBROSI S 31 (60%)

 

HYPERPLASI" , RFMATOPOIETI C 1 (2*)

 

f SPLE3N (53) (52) (51)

HYPERPLASIA , LY1PHOT D 1 (2«) 

HEHATOPOI F S I S 1 (2«) 1 (2% ) 

ISFS^HTERIC I. NOH^ (!»?) (45) (47) 

CONGESTION, NOS 1 (2S) 

HYPEPPLASIA , NOS 2 (455) 

HYPERPLR3IA , LYKPHOI D 2 (4«) 

CIRCULATOR Y SYSTE H 

* H" AST (55) (51) (51) 

PFRIART^PITT S 1 (2«)

 

DIGESTIV SYSTE M 

tLIVEP (54) (52) (53) jsoaaiE 1 J2«L_

 

t NUNBF P OF ANTMAt S »'Tf TISSU E FXAMINE D MIC33SCOP 1C ALL Y 

* NOH3F.R OF ANIHAI S KECTOPSIfi D 

86 
TABLE D2. FEMALE MICE: NONNEOPLASTIC LESIONS (CONTINUED) 

CONTROL LOW DOSE
 

METAMORPHOSIS FATTY 2 (4S) 

MEGALOCYTOSTS 1 ( 2X) 

HSMATOPOIF.SIS 1 (2*) 

IPANCRFAS 

DILATATION/DUCTS 

C*9) 

1 {2".) 

(52) 

CYSTIC DUCTS 

INFLAMMATION , CHRONIC 2 (4%) 

METAMORPHOSIS "ATTY 

ATROPHY, N05 1 (2?,) 

•STOHACH (53) (52) 

ULCER, NOS 1 (2%) 

EROSION 1 (2%) 

HYPERPLASIA, EPITHELIAL 2 (US) 

*GA3TRIC SEP.OSA 

STEATITIS 

(53) (52)

i ;:" 

NECROSIS, FAT 1 (2X) 

IPFYERS PATCH (52) (52) 

HYPERPLASIA, LYMPHOID 

URINAR Y SYSTF.K
 

f KIDNEY (55) (52)
 

HEMATOMA, ORGANIZED
 

STEATITTS 1 (2*)
 

PYELONEPHPITIS, CHPONIC 1 (2%)
 

NECROSIS, FAT 1 (2*|
 

tURINAPY BLADDER (50) (51)
 

INFLAMMATION, ACUTP 1 (2*)
 

INFLAMMATION, ACUTE/CHRONIC 1 (2/J)
 

INFLAMMATION, CHRONIC 1 (2%)
 

INFLAMMATION , CHRONIC DIFFUSE 1 (2X)
 

HYPERPLASIA, EPITHELIAL 1 (2X)
 

ENDOCRINE SYSTEM
 

IADRENAL (50) (52)
 

INFLAMMATION, ACUTE 1 (2S)
 

tTHYROID (48) (39)
 

CYSIIC. FOLLICLES
 

* NUMBER OF ANIMALS WITH TISSUE EXAMINED MICR3SCOP 1CALLY
 

* NUMBER OF ANIMAL S NFCPOPSIED
 

HIGH DOSE
 

(52)
 

1 (2X)
 

1 (2X)
 

(53)
 

(53)
 

(52)
 

1 (2%)
 

(54)
 

1 (2%)
 

(50)
 

3 (6X)
 

12 (2i*X)
 

(52)
 

(38)
 

1 13%L
 

87
 
TABLE 02. FEMALE MICE: NONNEOPLASTIC LESIONS (CONTINUED) 

CONTROL LOW DOSE 

*PANCR*ATIC ISLETS {"9) (52)

 

HYPFRPLASIA , NO S 3 (6% )

 

PPPPOPUCTTV? SYSTFM 

#UT5ROS (5<t) (51)

 

HYDROMETR A 3 (6%)

 

f UTEPTJS/ENDOKETRI DM (5<t) (51) 

HYPFRPLASIA , CYSTIC 15 (28%) 36 (7U) 

#OVAR Y (50) (U6 ) 

CYST, NOS 7 (1'4% ) 6 (13X) 

HEMORRHAGIC CYST 1 (2%) 

INFLAMMATION , ACUT E 1 (2% ) 

ABSCESS, NO S 1 (2«) 

NERVOU S SYSTFM 

*BRATN/MENINGE S (55) ((»8) 

INFLAMMATION , NO S 1 (2%)

 

FIBROSIS, FOCAL 

#BPAI N (55) (48)

 

HYDPOCEPHALUS , NOS 2 (US ) 

SPECIAL SENSE ORGAN S 

NON* 

flUSCULOSKELETAL SYSTEM 

NON E 

BODY CAVITIES 

*ABDOMINA L CAVIT Y (55) (52)

 

NECROSIS, FAT 7 (13%)

 

*MESENTERY (55) (52)

 

C!S_T,._flO.S_ _ .. 1 .12*1 _.

 

t NUMBER OF ANIMAL S WITH TISSUE 'EXAMINE D MICROSCOPICALL Y 

* NUMBER OF ANIMAL S NECROPSIED 

HIGH DOSE 

(52) 

(50) 

1 (255) 

(50) 

39 (7855) 

(52) 

5 (10%) 

(52) 

1 (2%) 

(52) 

(54) 

(54) 

88 
TABLE D2. FEMALE MICE: NONNEOPLASTIC LESIONS (CONTINUED) 

CONTROL LOW DOSE 

A LL OTHER SYSTEM S 

*«ULTIPLE ORGAN S 

PERIAPTFRITI S 

(55) 

1 (2% ) 

(52) 

OKSNTU M 

STEATITIS 

NECROSIS, FAT 

1 

1 

SPFCIAI. MORPHOLOG Y SUMMAR Y 

N O LESION REPORTE D 

AUTO/NECROPSY/HI3TO p^RF 

AOTOLYSIS/NO NECROPS Y 

1 14 

3 

t NUNBKR OF ANIMAL S WITH TISSUE EXAMINE D MICROSCOPICALLY 

* NUHBK P OF ANIMAL S NFCROPSIK D 

HIGH DOSE 

(5«) 

1 

1 

1 

89 

APPENDIX E
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUMORS
 

IN RATS FED o-ANISIDINE HYDROCHLORIDE IN THE DIET
 

91
 

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

 

Fed o-Anisidine Hydrochloride in the Diet3

 

Topography: Morphology Control 

Low 

Dose 

High 

Dose 

Integumentary System: Fibroma^ 4/54 (7) 0/55 (0) 0/53 (0) 

P Values0 >d P = 0.015(N) N.S. N.S. 

Relative Riskf 

Lower Limit 

Upper Limit 

0.000 

0.000 

1.062 

0.000 

0.000 

1.101 

Weeks to First Observed Tumor 106 __ __ 

Hematopoietic System: 

Lymphoma or Leukemia'5 18/54 (33) 1/55 (2) 1/53 (2) 

P Values

c

»d P < O.OOl(N) P < O.OOl(N) P < O.OOl(N) 

Departure from Linear Trend6 P = 0.004 

Relative Riskf 0.055 0.057 

Lower Limit 0.001 0.001 

Upper Limit 0.324 0.336 

Weeks to First Observed Tumor 84 98 40 
__ __ 

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

Fed o-Anisidine Hydrochloride in the Diet3 

(continued) 

Topography: Morphology 

Liver: Neoplastic Nodule or 

Hepatocellular Carcinoma^
 

P Values0 »d
 

Departure from Linear Trend6
 

Relative Risk^
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Kidney or Kidney Pelvis:
 

Transitional-cell Carcinoma*5
 

P Values0 »d
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Low
 

Control Dose
 

0/54 (0) 4/55 (7)
 

N.S. N.S.
 

P = 0.005
 

Infinite
 

0.908
 

Infinite
 

85
 

0/53 (0) 3/55 (5)
 

P = 0.005 N.S.
 

Infinite
 

0.578
 

Infinite
 

64
 —
 

High
 

Dose
 

0/52 (0)
 

N.S.
 

...
 

—
 

—
 

7/53 (13)
 

P = 0.006
 

Infinite
 

1.937
 

Infinite
 

51 
Table El. Analyses of the Incidence of Primary Tumors in Male Rats
 

Fed o-Anisidine Hydrochloride in the Dieta
 

(continued)
 

Topography: Morphology
 

Urinary Bladder: Transitional-cell
 

Carcinoma^5
 

P Values0 »d
 

Departure from Linear Trend6
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Urinary Bladder: Transitional-cell
 

Papilloma or Carcinoma*3
 

P Values0 »d
 

Departure from Linear Trend6
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Control
 

0/51 (0)
 

P < 0.001
 

P < 0.001
 

—
 

0/51 (0)
 

P < 0.001
 

P < 0.001
 

—
 

Low
 

Dose
 

50/54 (93)
 

P < 0.001
 

Infinite
 

17.045
 

Infinite
 

64
 

52/54 (96)
 

P <0.001
 

Infinite
 

18.455
 

Infinite
 

64
 

High
 

Dose
 

51/52 (98)
 

P < 0.001
 

Infinite
 

19.525
 

Infinite
 

48
 

52/52 (100)
 

P < 0.001
 

Infinite
 

21.590
 

Infinite
 

40 
Table El. Analyses of the Incidence of Primary Tumors in Male Rats

 

Fed o-Anisidine Hydrochloride in the Diet3

 

(continued) 

Low High 

Topography: Morphology Control Dose Dose 

Pituitary: Chromophobe Adenoma^ 4/48 (8) 0/49 (0) 0/47 (0)
 

P Values0 »d P = 0.015(N) N.S. N.S.
 

Relative Riskf 0.000 0.000
 

Lower Limit 0.000 0.000
 

Upper Limit 1.056 1.100
 

Weeks to First Observed Tumor 90
 — —
 

Pituitary: Adenoma, NOS, or
 

Chromophobe Adenoma^ 4/48 (8) 2/49 (4) 0/47 (0)
 

P Valuesc»d P = 0.038(N) N.S. N.S.
 

Relative Riskf 0.490 0.000
 

Lower Limit 0.046 0.000
 

Upper Limit 3.246 1.100
 

Weeks to First Observed Tumor 90 103
 —
 
Table El. Analyses of the Incidence of Primary Tumors in Male Rats
 

Fed o-Anisidine Hydrochloride in the Diet3
 

(continued)
 

High
 

Dose
 

0/53 (0)
 

P < O.OOl(N)
 

0.000
 

0.000
 

0.234
 

—
 

0/40 (0)
 

N.S.
 

—
 

—
 

—
 

—
 

Topography: Morphology
 

Adrenal : Pheochromocytoma or
 

Pheochromocytoma, Malignant^
 

P Values0 »d
 

Departure from Linear Trend6
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Thyroid: C-cell Carcinoma^*
 

P Values0 »d
 

Departure from Linear Trend6
 

Relative Risk^
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Low
 

Control Dose
 

14/54 (26) 0/55 (0)
 

P < O.OOl(N) P < O.OOl(N)
 

P = 0.005
 

0.000
 

0.000
 

0.226
 

73
 —
 

0/53 (0) 2/40 (5)
 

N.S. N.S.
 

P = 0.030
 

Infinite
 

0.392
 

Infinite
 

87
 —
 
Table El.


(continued)
 

Topography: Morphology
 

Thyroid: C-cell
 

Adenoma or Carcinoma"
 

P Values0 >d
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Thyroid: Follicular-cell
 

Carcinoma^
 

P Values0 >d
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

 Analyses of the Incidence of Primary Tumors in Male Rats
 

Fed o-Anisidine Hydrochloride in the Diet3
 

Control
 

3/53 (6)
 

N.S.
 

106
 

0/53(0)
 

N.S.
 

—
 

Low
 

Dose
 

3/40 (8)
 

N.S.
 

1.325
 

0.186
 

9.370
 

87
 

2/40 (5)
 

N.S.
 

Infinite
 

0.392
 

Infinite
 

104
 

High
 

Dose
 

0/40 (0)
 

N.S.
 

0.000
 

0.000
 

2.190
 

—
 

2/40 (5)
 

N.S.
 

Infinite
 

0.392
 

Infinite
 

78 
Table El. Analyses of the Incidence of Primary Tumors in Male Rats
 

Fed o-Anisidine Hydrochloride in the Diet3
 

(continued)
 

Low High
 

Topography: Morphology Control Dose Dose
 

Thyroid or Thyroid Follicle:
 

Follicular-cell Carcinoma or
 

Papillary Cystadenocarcinoma, NOS^ 0/53 (0) 3/40 (8) 2/40 (5)
 

P Values0 »d N.S. N.S. N.S.
 

Relative Riskf Infinite Infinite
 

Lower Limit 0.797 0.392
 

Upper Limit Infinite Infinite
 

Weeks to First Observed Tumor 91 78
 —
 

Thyroid or Thyroid Follicle:
 

Follicular-cell Adenoma,
 

Cystadenoma, or Papillary
 

Cystadenoma, NOSb 0/53 (0) 4/40 (10) 4/40 (10)
 

d
 P Values0

 » P = 0.030 P - 0.031 P = 0.031
 

Relative Riskf Infinite Infinite
 

Lower Limit 1.229 1.229
 

Upper Limit Infinite Infinite
 

Weeks to First Observed Tumor 92 55
 —
 
Table El. Analyses of the Incidence of Primary Tumors in Male Rats
 

Fed o-Anisidine Hydrochloride in the Diet3
 

(continued)
 

Topography : Morphology
 

Thyroid: All Follicular-cell TumorsS
 

P Values0 »d
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Testis: Interstitial-cell
 

Tumor*5
 

P Values0 »d
 

Departure from Linear Trend6
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Control
 

0/53 (0)
 

P = 0.009
 

—
 

53/54 (98)
 

P < O.OOl(N)
 

P < 0.001
 

73
 

Low
 

Dose
 

7/40 (17)
 

P - 0.002
 

Infinite
 

2.574
 

Infinite
 

91
 

43/52 (83)
 

P - 0.007(N)
 

0.843
 

0.809
 

0.968
 

71
 

High
 

Dose
 

6/40 (15)
 

P = 0.005
 

Infinite
 

2.121
 

Infinite
 

55
 

4/51 (8)
 

P < O.OOl(N)
 

0.080
 

0.060
 

0.161
 

62
 
Table El. Analyses of the Incidence of Primary Tumors in Male Rats
 

Fed o-Anisidine Hydrochloride in the Diet3
 

(continued) 

Low High 

Topography: Morphology Control Dose Dose 

Body Cavities: Mesothelioma or
 

Mesothelioma, Malignant" 2/54 (4) 6/55 (11) 1/53 (2)
 

P Values0 »d N.S. N.S. N.S.
 

Departure from Linear Trend6 P = 0.033
 

Relative Riskf 2.945 0.509
 

Lower Limit 0.556 0.009
 

Upper Limit 28.827 9.485
 

Weeks to First Observed Tumor 87 77 61
 

a

Dosed groups received 5,000 or 10,000 ppm in feed.
 

"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 control group when P < 0.05; otherwise,
 

not significant (N.S.) is indicated.
 
Table El. Analyses of the Incidence of Primary Tumors in Male Rats
 

Fed o-Anisidine Hydrochloride in the Dieta
 

(continued)
 

^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.
 

§These tumors include follicular-cell adenoma, follicular-cell carcinoma, cystadenoma, NOS,
 

papillary cystadenoma, NOS and papillary cystadenocarcinoma, NOS.
 
Table E2. Analyses of the Incidence of Primary Tumors in Female Rats

 

Fed o-Anisidine Hydrochloride in the Diet3

 

Topography: Morphology Control 

Low 

Dose 

High 

Dose 

Hematopoietic System: 

Lymphoma or Leukemia" 9/54 (17) 1/53 (2) 0/54 (0) 

P Values

c

»

d P < O.OOl(N) P = 0.009(N) P = O.OOl(N) 

Relative Riskf 

Lower Limit 

Upper Limit 

0.113 

0.003 

0.773 

0.000 

0.000 

0.382 

Weeks to First Observed Tumor 91 78 __ 

Salivary Gland: Adenoma, NOSb 3/52 (6) 0/51 (0) 0/47 (0) 

P Values

c

»

d P = 0.043(N) N.S. N.S. 

Relative Riskf 

Lower Limit 

Upper Limit 

0.000 

0.000 

1.697 

0.000 

0.000 

1.837 

Weeks to First Observed Tumor 107 — —— 
__ _ _ 

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

Fed o-Anisidine Hydrochloride in the Dieta
 

(continued)
 

Low High
 

Topography: Morphology Control Dose Dose
 

Urinary Bladder: Transitional-cell
 

Papillomab 0/49 (0) 5/49 (10) 0/51 (0)
 

P Values0 »d N.S. P = 0.028 N.S.
 

Departure from Linear Trend6 P = 0.002
 

_._
 Relative Riskf Infinite
 

Lower Limit 1.262
 

Upper Limit Infinite
 —
 

—
 

Weeks to First Observed Tumor 74
 

Urinary Bladder: Transitional-cell
 

Carcinoma*5 0/49 (0) 41/49 (84) 50/51 (98)
 

P Valuesc»d P < 0.001 P < 0.001 P < 0.001
 

Departure from Linear Trend6 P < 0.001
 

Relative Riskf Infinite Infinite
 

Lower Limit 14.250 18.754
 

Upper Limit Infinite Infinite
 

Weeks to First Observed Tumor 61
 36
 —
 
Table E2. Analyses of the Incidence of Primary Tumors in Female Rats
 

Fed o-Anisidine Hydrochloride in the Dieta
 

(continued)
 

Topography: Morphology
 

Urinary Bladder: Transitional-cell
 

Papilloma or Carcinoma"
 

P Values0 »d
 

Departure from Linear Trend6
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Pituitary: Carcinoma, NOSb
 

P Value sc>d
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Control
 

0/49 (0)
 

P < 0.001
 

P < 0.001
 

—
 

3/48 (6)
 

P = 0.039(N)
 

89
 

Low High 

Dose Dose 

46/49 (94) 50/51 (98) 

P < 0.001 P < 0.001 

Infinite Infinite 

16.833 18.754 

Infinite Infinite 

61 36 

0/51 (0) 0/45 (0) 

N.S. N.S. 

0.000 0.000 

0.000 0.000 

1.565 1.768 

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

Fed o-Anisidine Hydrochloride in the Dieta
 

(continued)
 

Topography: Morphology
 

Pituitary: Chromophobe Adenomab
 

P Valuesc»d
 

Departure from Linear Trend6
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Pituitary : Chromophobe Carcinoma
 

or Carcinoma, NOS^
 

P Values0 »d
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Low
 

Control Dose
 

15/48 (31) 0/51 (0)
 

P < O.OOl(N) P < O.OOl(N)
 

P = 0.003
 

0.000
 

0.000
 

0.200
 

74
 —
 

4/48 (8) 0/51 (0)
 

P = 0.016(N) N.S.
 

0.000
 

0.000
 

1.016
 

89
 —
 

High
 

Dose
 

0/45 (0)
 

P < O.OOl(N)
 

0.000
 

0.000
 

0.226
 

—
 

0/45 (0)
 

N.S.
 

0.000
 

0.000
 

1.147
 

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

(continued)
 

Topography: Morphology
 

Pituitary: Adenoma, NOS,
 

Chromophobe Adenoma,
 

or Acidophil Adenoma*3
 

P Values0 »d
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Adrenal: Pheochromocytoma"
 

P Values0 »d
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Fed o-Anisidine Hydrochloride in the Diet3 

Low High 

Control Dose Dose 

17/48 (35) 9/51 (18) 1/45 (2) 

P < O.OOl(N) P = 0.037(N) P < O.OOl(N) 

0.498 0.063 

0.219 0.002 

1.060 0.372 

74 72 66 

3/53 (6) 1/53 (2) 1/54 (2) 

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

0.333 0.327 

0.006 0.006 

3.993 3.921 

107 97 55 
Table E2. Analyses of the Incidence of Primary Tumors in Female Rats 

(continued) 

Topography: Morphology
 

Thyroid: C-cell Carcinomab
 

P Values0 >d
 

Relative Rlskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Thyroid C-cell Adenoma
 

or Carcinoma^
 

P Values0 »d
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Fed o-Anisidine Hydrochloride

Control
 

3/49 (6)
 

N.S.
 

107
 

4/49 (8)
 

P = 0.03KN)
 

107
 

 in the Dieta 

Low
 

Dose
 

1/45 (2)
 

N.S.
 

0.363
 

0.007
 

4.321
 

95
 

1/45 (2)
 

N.S.
 

0.272
 

0.006
 

2.615
 

95
 

High
 

Dose
 

0/46 (0)
 

N.S.
 

0.000
 

0.000
 

1.767
 

—
 

0/46 (0)
 

N.S.
 

0.000
 

0.000
 

1.146
 

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

Fed o-Anisidine Hydrochloride in the Diet3
 

(continued)
 

Topography: Morphology
 

Thyroid: Follicular-cell Carcinoma*1
 

P Valuesc»d
 

Departure from Linear Trend6
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Thyroid: All Follicular-cell Tumor s8
 

P Value sc>d
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Low High 

Control Dose Dose 

0/49 (0) 3/45 (7) 0/46 (0) 

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

P = 0.011 

__ 

Infinite 

0.656 

Infinite — 

—— 

79 — — 

1/49 (2) 4/45 (9) 3/46 (7) 

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

4.356 3.196 

0.453 0.268 

209.417 164.012 

77 79 57 
Table E2. Analyses of the Incidence of Primary Tumors in Female Rats
 

Fed o-Anisidine Hydrochloride in the Diet3
 

(continued)
 

Topography: Morphology
 

Subcutaneous Tissue or
 

Mammary Gland : Fibroadenomab
 

P Values0 >d
 

Departure from Linear Trend6
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Uterus: Endometrial Stromal Polyp'5
 

P Values0 »d
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Control
 

16/54 (30)
 

P < O.OOl(N)
 

P = 0.033
 

99
 

16/52 (31)
 

P < O.OOl(N)
 

68
 

Low
 

Dose
 

2/53 (4)
 

P < O.OOl(N)
 

0.127
 

0.015
 

0.505
 

90
 

6/50 (12)
 

P = 0.019(N)
 

0.390
 

0.136
 

0.955
 

66
 

High
 

Dose
 

0/54 (0)
 

P < O.OOl(N)
 

0.000
 

0.000
 

0.198
 

—
 

0/50 (0)
 

P < O.OOl(N)
 

0.000
 

0.000
 

0.206
 

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

Fed o-Anisidine Hydrochloride in the Diet3
 

(continued)
 

a

Dosed groups received 5,000 or 10,000 ppm in feed.
 

^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 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.
 

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

group.
 

&These tumors include undifferentiated carcinoma, follicular-cell adenoma, follicular-cell
 

carcinoma or papillary cystadenoma, NOS.
 
\ 
APPENDIX F
 

ANALYSES OF THE INCIDENCE OF PRIMARY TUIIOUS
 

IN MICE FED o-ANISIDINE HYDROCliLORIDE IN THE DIET
 

113
 

__ 

Table Fl.


Topography: Morphology 

Lung: Alveolar/Bronchiolar 

Carcinoma'3 

P Values0 »d 

Relative Risk^ 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Lung: Alveolar/Bronchiolar 

Adenoma or Carcinoma^3 

P Values c»d 

Relative Riskf 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

 Analyses of the Incidence of Primary Tumors in Male Mice
 

Fed o-Anisidine Hydrochloride in the Diet3
 

Control 

6/54 (11) 

P = 0.024(N) 

105 

12/54 (22) 

P = 0.006(N) 

79 

Low 

Dose 

6/54 (11) 

N.S. 

1.000 

0.285 

3.508 

104 

9/54 (17) 

N.S. 

0.750 

0.304 

1.774 

104 

High 

Dose 

0/52 (0) 

P = 0.015(N) 

0.000 

0.000 

0.650 

2/52 (4) 

P = 0.005(N) 

0.173 

0.020 

0.727 

105 
Table Fl. Analyses of the Incidence of Primary Tumors in Male Mice

 

Fed o-Anisidine Hydrochloride in the Diet3

 

(continued) 

Topography: Morphology 

Hematopoietic System: 

Lymphoma or Leukemia^ 

P Values0 »d 

Relative Riskf 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Liver: Hepatocellular Carcinoma" 

P Values

c

»d 

Relative Riskf 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Control 

4/55 (7) 

u.s. 

105 

24/54 (44) 

P < O.OOl(N) 

53 

Low 

Dose 

4/55 (7) 

N.S. 

1.000 

0.196 

5.110 

71 

13/54 (24) 

P = 0.021(N) 

0.542 

0.288 

0.980 

88 

High 

Dose 

1/53 (2) 

N.S. 

0.259 

0.005 

2.510 

88 

7/52 (13) 

P < O.OOl(N) 

0.303 

0.122 

0.651 

83 
Table Fl.


(continued)
 

Topography: Morphology
 

Liver: Hepatocellular
 

Adenoma or Carcinoma^
 

P Valuesc»d
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

 Analyses of the Incidence of Primary Tumors in Male Mice
 

Fed o-Anisidine Hydrochloride in the Diet3
 

Low
 

Control Dose
 

28/54 (52) 13/54 (24)
 

P < O.OOl(N) P = 0.003(N)
 

0.464
 

0.254
 

0.815
 

Weeks to First Observed Tumor 53 88 

Urinary Bladder:

Carcinoma^ 

 Transitional-cell 

0/48 (0) 0/55 (0) 

P Values0 » d P < 0.001 N.S. 

Departure from Linear Trend6 P = 0.004 

Relative Riskf 

Lower Limit 

Upper Limit 

— 

— 

— — 

Weeks to First Observed Tumor — — 

High
 

Dose
 

7/52 (13)
 

P < O.OOl(N)
 

0.260
 

0.108
 

0.543
 

83
 

15/53 (28)
 

P < 0.001
 

Infinite
 

4.267
 

Infinite
 

45
 
Table Fl. Analyses of the Incidence of Primary Tumors in Male Mice
 

Fed o-Anisidine Hydrochloride in the Dieta
 

(continued)
 

Topography: Morphology
 

Urinary Bladder: Transitional-cell
 

Papilloma or Carcinoma*3
 

P Values0 »d
 

Departure from Linear Trend6
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Adrenal/Capsule: Adenoma, NOS*3
 

P Values0 »d
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Control
 

0/48 (0)
 

P < 0.001
 

P = 0.005
 

—
 

5/50 (10)
 

P = 0.006(N)
 

105
 

Low
 

Dose
 

2/55 (4)
 

N.S.
 

Infinite
 

0.258
 

Infinite
 

104
 

0/53 (0)
 

P = 0.024(N)
 

0.000
 

0.000
 

0.749
 

—
 

High
 

Dose
 

22/53 (42)
 

P < 0.001
 

Infinite
 

6.486
 

Infinite
 

45
 

0/52 (0)
 

P = 0.025(N)
 

0.000
 

0.000
 

0.763
 

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

Fed o-Anisidine Hydrochloride in the Diet3
 

(continued)
 

a

Dosed groups received 2,500 or 5,000 ppm in feed. 

^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 CochranArmitage 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 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.


Topography: Morphology 

Lung: Alveolar/Bronchiolar 

Adenoma or Carcinoma^ 

P Values0 »d 

Relative Riskf 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

Hematopoietic System: 

Lymphoma or Leukemia" 

P Valuesc»d 

Relative Riskf 

Lower Limit 

Upper Limit 

Weeks to First Observed Tumor 

 Analyses of the Incidence of Primary Tumors in Female Mice
 

Fed o-Anisidine Hydrochloride in the Diet3
 

Low 

Control Dose 

4/55 (7) 2/51 (4) 

N.S. N.S. 

0.539 

0.050 

3.586 

105 105 

18/55 (33) 12/52 (23) 

P = 0.005(N) N.S. 

0.705 

0.345 

1.385 

86 78 

High 

Dose 

1/52 (2) 

N.S. 

0.264 

0.005 

2.557 

105 

6/54 (11) 

P = 0.006(N) 

0.340 

0.120 

0.815 

34 
Table F2. Analyses of the Incidence of Primary Tumors in Female Mice
 

Fed o-Anisidine Hydrochloride in the Diet3
 

(continued) 

Low High 

Topography: Morphology Control Dose Dose 

Liver: Hepatocellular Carcinoma^ 7/54 (13) 0/52 (0) 4/53 (8) 

P Values0 »d N.S. P = 0.007(N) N.S. 

Departure from Linear Trend6 P = 0.017 

Relative Riskf 0.000 0.582 

Lower Limit 0.000 0.132 

Upper Limit 0.536 2.148 

Weeks to First Observed Tumor 101 81 — 

Liver: Hepatocellular Adenoma, 

Carcinoma, or Neoplastic Nodule° 11/54 (20) 1/52 (2) 4/53 (8) 

P Values0 »d P = 0.020(N) P = 0.002(N) N.S. 

Departure from Linear Trend6 P = 0.018 

Relative Risk^ 0.094 0.370 

Lower Limit 0.002 0.091 

Upper Limit 0.&13 1.161 

Weeks to First Observed Tumor 59 105 81 
Table F2. Analyses of the Incidence of Primary Tumors in Female Mice
 

Fed o-Anisidine Hydrochloride in the Dieta
 

(continued)
 

Topography: Morphology
 

Urinary Bladder: Transitional-cell
 

Carcinoma^
 

P ValuesC,d
 

Departure from Linear Trend6
 

Relative Riskf'
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Urinary Bladder: Transitional-cell
 

Papilloma or Carcinoma*1
 

P Values0 »d
 

Departure from Linear Trend6
 

Relative Riskf
 

Lower Limit
 

Upper Limit
 

Weeks to First Observed Tumor
 

Low High 

Control Dose Dose 

0/50 (0) 0/51 (0) 18/50 (36) 

P < 0.001 N.S. P < 0.001 

P = 0.001 

__ 

Infinite 

5.758 — Infinite — 

81 — — 

0/50 (0) 1/51 (2) 22/50 (44) 

P < 0.001 N.S. P < 0.001 

P = 0.001 

Infinite Infinite 

0.053 7.163 

Infinite Infinite 

105 81 — 
Table F2. Analyses of the Incidence of Primary Tumors in Female Mice
 

Fed o-Anisidine Hydrochloride in the Diet3
 

(continued) 

Low High 

Topography: Morphology Control Dose Dose 

Pituitary: Chromophobe Adenoma'3 2/42 (5) 0/40 (0) 0/43 (0) 

P Valuesc»d N.S. N.S. N.S. 

Relative Risk> 0.000 0.000 

Lower Limit 0.000 0.000 

Upper Limit 3.526 3.287 

Weeks to First Observed Tumor 105 — — 

a

Treated groups received doses of 2,500 or 5,000 ppm in feed.
 

"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 treated group is the probability level for the Fisher exact test for
 

the comparison of that treated group with the control group when P < 0.05; otherwise,
 

not significant (N.S.) is indicated.
 

d-A negative trend (N) indicates a lower incidence in a treated 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 treated group and the control
 

group.
 

APPENDIX G
 

ANALYSIS OF FORMULATED DIETS FOR
 

o-ANISIDINE HYDROCIILORIDE CONCENTRATION
 

125
 

APPENDIX G
 

Analysis of Formulated Diets for
 

o-Anisidine Hydrochloride Concentration
 

Duplicate 2-g samples of the diet mixtures were each shaken with
 

50 ml of 95% ethanol for 15 minutes. The mixture was allowed to
 

settle overnight, and the absorbance of the supernatant, after
 

appropriate dilution, was measured at 270 nm against a "blank"
 

extracted from 2 g of the same feed used to prepare the diet
 

mixtures. Concentrations were determined by comparison with
 

standard solutions. Recoveries were determined from duplicate
 

spiked feed samples worked up simultaneously with each set of
 

diet samples. The average recovery from the 0.5% spiked feed
 

samples was
 

Theoretical No. of Sample Coefficient of Range 

Concentrations Samples Analytical Variation (%) (% in feed) 

In Diet Mean 

(% in Feed) (% in Feed) 

0.25 6 0.26 24.4 0.16-0.33 

0.5 6 0.44 11.8 0.36-0.51 

1.0 5 0.97 9.5 0.86-1.11 

127
 

Review of the Bioassay of o-Anisidine Hydrochloride*
 

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, 1976, 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 o-Anisidine
 

Hydrochloride for carcinogenicity.
 

The primary reviewer said that the compound was carcinogenic in both the treated rats and mice, under the
 

conditions of test. Although a different batch of the
 

test chemical was used for the subchronic and chronic
 

phases, each was reported to be more than 99% pure.
 Transitional-cell carcinomas and papillomas of the urinary
 

bladder were induced, in a dose-related fashion, in both
 

species. Transitional-cell carcinomas of the renal pelvis
 

and follicular-cell tumors of the thyroid also were induced
 

in treated male rats. Although there were a number of
 

experimental shortcomings, including poor survival, the
 

primary reviewer said that the conclusion on the carcino­ genicity of o-Anisidine Hydrochloride was still valid.
 

The secondary reviewer also agreed that o-Anisidine
 

Hydrochloride was carcinogenic under the conditions of test.
 He added that the chemical may pose a carcinogenic risk to
 

humans, especially in occupational situations.
 

129
 
It was moved that the report on the bioassay of
 

o-Anisidine Hydrochloride be accepted as written. The
 

motion was seconded and approved unanimously.
 

Members present were:
 

Michael Shlmkln (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
 

*	 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.
 

«U.S. GOVERNMENT PRINTING OFFICE: 1978 260-899/3167 1-3 130 

DHEW Publication No. (NIH) 78-1339