NTP TECHNICAL REPORT
 

ON THE 

TOXICOLOGY AND CARCINOGENESIS
 

STUDIES OF
 

COBALT SULFATE HEPTAHYDRATE
 

(CAS NO. 10026-24-1) 

IN F344/N RATS AND B6C3F1 MICE

(INHALATION STUDIES) 

NATIONAL TOXICOLOGY PROGRAM
 

P.O. Box 12233
 

Research Triangle Park, NC 27709
 

August 1998 

NTP TR 471
 

NIH Publication No. 98-3961
 

U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES
 

Public Health Service
 

National Institutes of Health
 
FOREWORD 

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

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

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

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

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

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

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

research and to biological assay development and validation. 

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

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

prevention of disease. 

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

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

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

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

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

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

before being presented for public review. 

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

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

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

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

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

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

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

Listings of all published NTP reports and ongoing studies are available from NTP Central Data 

Management, NIEHS, P.O. Box 12233, MD E1-02, Research Triangle Park, NC 27709 (919-541-3419). 

The Abstracts and other study information for 2-year studies are also available at the NTP’s World Wide 

Web site: http://ntp-server.niehs.nih.gov. 
NTP TECHNICAL REPORT
 

ON THE
 

TOXICOLOGY AND CARCINOGENESIS
 

STUDIES OF
 

COBALT SULFATE HEPTAHYDRATE
 

(CAS NO. 10026-24-1) 

IN F344/N RATS AND B6C3F1 MICE

(INHALATION STUDIES) 

NATIONAL TOXICOLOGY PROGRAM
 

P.O. Box 12233
 

Research Triangle Park, NC 27709
 

August 1998 

NTP TR 471
 

NIH Publication No. 98-3961
 

U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES
 

Public Health Service
 

National Institutes of Health
 
2 Cobalt Sulfate Heptahydrate, NTP TR 471 

CONTRIBUTORS
 

National Toxicology Program 

Evaluated and interpreted results and reported findings 

J.R. Bucher, Ph.D., Study Scientist 

D.A. Bridge, B.S. 

R.E. Chapin, Ph.D. 

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

J.K. Haseman, Ph.D. 

R.R. Maronpot, D.V.M. 

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

J.H. Roycroft, Ph.D. 

C.S. Smith, Ph.D. 

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

D.B. Walters, Ph.D. 

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

Battelle Pacific Northwest Laboratories 

Conducted studies, evaluated pathology findings 

B.J. Chou, D.V.M., Ph.D., Principal Investigator 

J.A. Dill, Ph.D. 

S.L. Grumbein, D.V.M., Ph.D. 

P.W. Mellick, D.V.M., Ph.D. 

S.E. Rowe, D.V.M., M.S. 

Experimental Pathology Laboratories, Inc. 

Provided pathology quality assurance 

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

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

C.C. Shackelford, D.V.M., M.S., Ph.D. 

Dynamac Corporation 

Prepared quality assurance audits 

S. Brecher, Ph.D., Principal Investigator 

NTP Pathology Working Group 

Evaluated slides, prepared pathology report on rats 

(26 April 1996) 

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

Pathology Associates International 

D. Dixon, D.V.M., Ph.D. 

National Toxicology Program 

J. Everitt, D.V.M. 

Chemical Industry Institute of Toxicology 

S.L. Grumbein, D.V.M., Ph.D. 

Battelle Pacific Northwest Laboratories 

F.F. Hahn, D.V.M., Ph.D. 

IIT Research Institute 

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

National Toxicology Program 

R.A. Herbert, D.V.M., Ph.D. 

National Toxicology Program 

R.R. Maronpot, D.V.M. 

National Toxicology Program 

A. Nyska, D.V.M. 

National Toxicology Program 

C.C. Shackelford, D.V.M., M.S., Ph.D. 

Experimental Pathology Laboratories, Inc. 

Evaluated slides, prepared pathology report on mice 

(29 February 1996) 

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

Pathology Associates International 

D. Dixon, D.V.M., Ph.D. 

National Toxicology Program 

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

Experimental Pathology Laboratories, Inc. 

J. Everitt, D.V.M. 

Chemical Industry Institute of Toxicology 

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

National Toxicology Program 

R.A. Herbert, D.V.M., Ph.D. 

National Toxicology Program 

R.R. Maronpot, D.V.M. 

National Toxicology Program 

A. Nyska, D.V.M. 

National Toxicology Program 

A. Radovsky, D.V.M., Ph.D. 

National Toxicology Program 
Cobalt Sulfate Heptahydrate, NTP TR 471 3 

Analytical Sciences, Inc. Biotechnical Services, Inc. 

Provided statistical analyses Prepared Technical Report 

R.W. Morris, M.S., Principal Investigator S.R. Gunnels, M.A., Principal Investigator 

S.R. Lloyd, M.S. L.M. Harper, B.S. 

N.G. Mintz, B.S. A.M. Macri-Hanson, M.A., M.F.A. 
4 

CONTENTS
 

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

EXPLANATION OF LEVELS OF EVIDENCE OF CARCINOGENIC ACTIVITY ............. 10
 

TECHNICAL REPORTS REVIEW SUBCOMMITTEE ................................ 11
 

SUMMARY OF TECHNICAL REPORTS REVIEW SUBCOMMITTEE COMMENTS .......... 12
 

INTRODUCTION ......................................................... 13
 

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

RESULTS ............................................................... 29
 

DISCUSSION AND CONCLUSIONS ............................................ 51
 

REFERENCES ........................................................... 55
 

APPENDIX A Summary of Lesions in Male Rats in the 2-Year Inhalation Study
 

of Cobalt Sulfate Heptahydrate ..................................... 63
 

APPENDIX B Summary of Lesions in Female Rats in the 2-Year Inhalation Study
 

of Cobalt Sulfate Heptahydrate ..................................... 105
 

APPENDIX C Summary of Lesions in Male Mice in the 2-Year Inhalation Study
 

of Cobalt Sulfate Heptahydrate ..................................... 139
 

APPENDIX D Summary of Lesions in Female Mice in the 2-Year Inhalation Study
 

of Cobalt Sulfate Heptahydrate ..................................... 173
 

APPENDIX E Genetic Toxicology ............................................. 209
 

APPENDIX F Chemical Characterization and Generation of Chamber Concentrations ......... 213
 

APPENDIX G Ingredients, Nutrient Composition, and Contaminant Levels 

in NIH-07 Rat and Mouse Ration .................................... 225
 

APPENDIX H Sentinel Animal Program ......................................... 229
 

APPENDIX I K-ras Mutation Frequency and Spectra in Lung Neoplasms
 

from B6C3F Mice Exposed to Cobalt Sulfate Heptahydrate for 2 Years 1 ......... 233
 

APPENDIX J Impact of Helicobacter Hepaticus Infection in B6C3F Mice 1
 

from 12 NTP 2-Year Carcinogenesis Studies ............................ 241
 
 

 

5 

ABSTRACT
 

CoSO 4 2 C 7H O

COBALT SULFATE HEPTAHYDRATE 

CAS No. 10026-24-1 

Molecular Weight: 281.13 

Synonyms: Bieberite; cobalt(II) sulfate (1:1) heptahydrate; cobalt monosulfate heptahydrate; cobalt(II) sulphate heptahydrate; sulfuric acid, 

cobalt(2+) salt (1:1) heptahydrate 

Cobalt sulfate is used in the electroplating and electrochemical industries. It is also used as a coloring agent 

for ceramics and as a drying agent in inks, paints, 

varnishes, and linoleum. Cobalt sulfate may be added 

to animal feed as a mineral supplement and has been 

used as a top dressing on pasture lands. Cobalt sulfate 

was nominated by the National Cancer Institute for 

study based on a lack of information on the toxicity of 

soluble salts. Male and female F344/N rats and 

B6C3F mice were exposed to cobalt sulfate 1 

heptahydrate (approximately 99% pure) by inhalation 

for 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium. The results of 

prechronic inhalation toxicity studies were reported 

previously (Bucher et al., 1990; NTP, 1991). 

2-YEAR STUDY IN RATS 

Groups of 50 male and 50 female rats were exposed to 

aerosols containing 0, 0.3, 1.0, or 3.0 mg/m3

 cobalt 

sulfate heptahydrate 6 hours per day, 5 days per week, 

for 105 weeks. 

Survival and Body Weights 

Survival of exposed males and females was similar to 

that of the chamber controls. Mean body weights of 

exposed male and female rats were similar to those of 

the chamber controls throughout the study. 

Pathology Findings 

The incidences and severities of proteinosis, alveolar 

epithelial metaplasia, granulomatous alveolar inflammation, and interstitial fibrosis were markedly greater 

in all exposed groups of male and female rats than in 

the chamber controls. The incidences of alveolar 

epithelial hyperplasia in all groups of exposed males 

and in females exposed to 3.0 mg/m3

 were significantly greater than those in the chamber control 

groups, as were the incidences of squamous metaplasia in 1.0 mg/m3 females and atypical alveolar 

epithelial hyperplasia in 3.0 mg/m3 females. In 

3 3.0 mg/m males, the combined incidence of alveolar/

bronchiolar neoplasms (adenoma and/or carcinoma) 

was significantly greater than in the chamber controls. 

In female rats exposed to 1.0 or 3.0 mg/m , the 3 
 

6 Cobalt Sulfate Heptahydrate, NTP TR 471 

incidences of alveolar/bronchiolar neoplasms were 

significantly greater than those in the chamber control 

group and exceeded the NTP historical control ranges. 

A squamous cell carcinoma was observed in one 

3 3 1.0 mg/m and one 3.0 mg/m female. 

The incidences of benign, complex, or malignant 

pheochromocytoma (combined) in 1.0 mg/m3 males 

3 and in 3.0 mg/m females were significantly greater 

than those in the chamber controls and exceeded the 

historical control ranges. 

Hyperplasia of the lateral wall of the nose, atrophy of 

the olfactory epithelium, and squamous metaplasia of 

the epiglottis were observed in all exposed groups of 

males and females, and the severities of these lesions 

increased with increasing exposure concentration. The 

incidences of squamous metaplasia of the lateral wall 

of the nose and metaplasia of the olfactory epithelium 3 were increased in 3.0 mg/m males and females. 

2-YEAR STUDY IN MICE 

Groups of 50 male and 50 female mice were exposed 3 to aerosols containing 0, 0.3, 1.0, or 3.0 mg/m cobalt

sulfate heptahydrate 6 hours per day, 5 days per week, 

for 105 weeks. 

Survival and Body Weights 

Survival of exposed males and females was similar to 

that of the chamber controls. Mean body weights of 

3.0 mg/m3 male mice were less than those of the 

chamber controls from week 96 until the end of the 

study. The mean body weights of all exposed groups 

of female mice were generally greater than those of the 

chamber controls from week 20 until the end of the 

study. 

Pathology Findings 

The incidences of diffuse histiocytic cell infiltration in 

3.0 mg/m3

 males and of focal histiocytic cell infil­ 3 tration in 3.0 mg/m females were significantly greater

than those in the chamber controls. The incidences of 

alveolar/bronchiolar neoplasms in 3.0 mg/m3

 males 

and females were significantly greater than those in the 

chamber control groups. The combined incidences 

of alveolar/bronchiolar adenoma or carcinoma and the 

incidences of alveolar/bronchiolar carcinoma in 

3.0 mg/m3 males and females and the incidence of 

alveolar/bronchiolar adenoma in 3.0 mg/m3 females 

exceeded the NTP historical control ranges for inhalation studies. 

The incidences of atrophy of the olfactory epithelium 3 in 1.0 and 3.0 mg/m males and females and hyper­ 3 plasia of the olfactory epithelium in 3.0 mg/m males 

and females were significantly greater than in the 

chamber controls. Squamous metaplasia of the larynx 

was observed in all exposed groups of males and 

females. 

Male mice had a pattern of nonneoplastic liver lesions 

along with silver-staining helical organisms within 

the liver, characteristic of an infection with 

Helicobacter hepaticus. In NTP studies with 

H. hepaticus-associated hepatitis, increased incidences 

of heman-giosarcoma were seen in the liver of male 

mice. In this study of cobalt sulfate heptahydrate, 

incidences of hemangiosarcoma were increased in 

exposed groups of male mice. Because of the above 

association, interpretation of the increased incidences 

of hemangiosarcoma in the livers of male mice was 

confounded. Incidences of lesions at other sites in this 

study of cobalt sulfate heptahydrate were not considered to have been significantly impacted by the 

infection with H. hepaticus or its associated hepatitis. 

GENETIC TOXICOLOGY 

Cobalt sulfate heptahydrate was mutagenic in 

S. typhimurium strain TA100 with and without liver 

S9 metabolic activation enzymes; no mutagenic 

activity was detected in strain TA98 or TA1535, with 

or without S9. 

CONCLUSIONS 

Under the conditions of these 2-year inhalation 

studies, there was some evidence of carcinogenic 

activity* of cobalt sulfate heptahydrate in male F344/N 

rats based on increased incidences of alveolar/ 

bronchiolar neoplasms. Marginal increases in 

incidences of pheochromocytomas of the adrenal 
__________ 

Cobalt Sulfate Heptahydrate, NTP TR 471 7 

medulla may have been related to exposure to cobalt 

sulfate heptahydrate. There was clear evidence of 

carcinogenic activity in female F344/N rats based on 

increased incidences of alveolar/bronchiolar neoplasms and pheochromocytomas of the adrenal medulla 

in groups exposed to cobalt sulfate heptahydrate. 

There was clear evidence of carcinogenic activity of 

cobalt sulfate heptahydrate in male and female 

B6C3F mice based on increased incidences of 1 

alveolar/bronchiolar neoplasms. 

Exposure to cobalt sulfate heptahydrate caused a 

spectrum of inflammatory, fibrotic, and proliferative 

lesions in the respiratory tract of male and female rats 

and mice. 

* Explanation of Levels of Evidence of Carcinogenic Activity is on page 10. A summary of the Technical Reports Review Subcommittee 

comments and the public discussion on this Technical Report appears on page 12. 
8 Cobalt Sulfate Heptahydrate, NTP TR 471 

Summary of the 2-Year Carcinogenesis and Genetic Toxicology Studies of Cobalt Sulfate Heptahydrate 

Male Female Male Female
 

F344/N Rats F344/N Rats B6C3F Mice B6C3F Mice
 1 1 

Concentrations 

Body weights 

Survival rates 

Nonneoplastic 

effects 

Chamber control, 0.3, 

1.0, or 3.0 mg/m3 Chamber control, 0.3, 1.0, or 3.0 mg/m3 Chamber control, 0.3, 1.0, or 3.0 mg/m3 Chamber control, 0.3, 1.0, or 3.0 mg/m3 

Exposed groups Exposed groups 3.0 mg/m group 3 Exposed groups 

similar to chamber similar to chamber slightly less than slightly greater than 

controls controls chamber controls chamber controls 

17/50, 15/50, 21/50, 28/50, 25/49, 26/50, 22/50, 31/50, 24/50, 34/50, 37/50, 32/50, 

15/50 30/50 20/50 28/50 

Lung: proteinosis Lung: proteinosis Lung: diffuse Lung: focal 

(0/50, 16/50, 40/48, (0/50, 36/49, 49/50, histiocytic cell histiocytic cell 

47/50); alveolar 49/50); alveolar infiltrate (1/50, 2/50, infiltrate (2/50, 5/50, 

epithelial metaplasia epithelial metaplasia 4/50, 10/50) 7/50, 10/50) 

(0/50, 50/50, 48/48, (2/50, 47/49, 50/50, 

49/50); granulomatous 49/50); granulomatous Nose: olfactory Nose: olfactory 

alveolar inflammation alveolar inflammation epithelial atrophy epithelial atrophy 

(2/50, 50/50, 48/48, (9/50, 47/49, 50/50, (0/50, 0/50, 29/48, (0/50, 2/50, 12/49, 

50/50); interstitial 49/50); interstitial 48/49); olfactory 46/48); olfactory 

fibrosis (1/50, 50/50, fibrosis (7/50, 47/49, epithelial hyperplasia epithelial hyperplasia 

48/48, 49/50); 50/50, 49/50); (0/50, 0/50, 0/48, (0/50, 0/50, 0/49, 

alveolar epithelial alveolar epithelial 10/49) 30/48) 

hyperplasia (9/50, hyperplasia (15/50, 

20/50, 20/48, 23/50) 7/49, 20/50, 33/50); Larynx: squamous Larynx: squamous 

squamous metaplasia metaplasia (0/48, metaplasia (0/50, 

Nose: lateral wall (0/50, 1/49, 8/50, 37/49, 48/48, 44/49) 45/49, 40/47, 50/50) 

hyperplasia (2/50, 3/50); atypical 

14/50, 21/49, 20/50); alveolar epithelial 

olfactory epithelial hyperplasia (0/50, 

atrophy (8/50, 24/50, 0/49, 3/50, 5/50) 

42/49, 48/50); lateral 

wall squamous Nose: lateral wall 

metaplasia (1/50, hyperplasia (1/50, 

3/50, 5/49, 8/50); 8/49, 26/50, 38/50); 

olfactory epithelial olfactory epithelial 

metaplasia (5/50, atrophy (5/50, 29/49, 

1/50, 5/49, 30/50) 46/50, 47/50); lateral 

wall squamous 

Larynx: epiglottis metaplasia (1/50, 

squamous metaplasia 1/49, 4/50, 10/50); 

(0/50, 10/49, 37/48, olfactory epithelial 

50/50) metaplasia (2/50, 

2/49, 3/50, 40/50) 

Larynx: epiglottis 

squamous metaplasia 

(1/50, 22/49, 39/50, 

48/50) 
Cobalt Sulfate Heptahydrate, NTP TR 471 9 

Summary of the 2-Year Carcinogenesis and Genetic Toxicology Studies of Cobalt Sulfate Heptahydrate 

Male Female Male Female
 

F344/N Rats F344/N Rats B6C3F Mice B6C3F Mice
 1 1 

Neoplastic effects Lung: alveolar/ Lung: alveolar/ Lung: alveolar/ Lung: alveolar/ 

bronchiolar adenoma bronchiolar adenoma bronchiolar adenoma bronchiolar adenoma 

(1/50, 4/50, 1/48, (0/50, 1/49, 10/50, (9/50, 12/50, 13/50, (3/50, 6/50, 9/50, 

6/50); alveolar/ 9/50); alveolar/ 18/50); alveolar/ 10/50); alveolar/ 

bronchiolar carcinoma bronchiolar carcinoma bronchiolar carcinoma bronchiolar carcinoma 

(0/50, 0/50, 3/48, (0/50, 2/49, 6/50, (4/50, 5/50, 7/50, (1/50, 1/50, 4/50, 

1/50); alveolar/ 6/50); alveolar/ 11/50); alveolar/ 9/50); alveolar/ 

bronchiolar adenoma bronchiolar adenoma, bronchiolar adenoma bronchiolar adenoma 

or carcinoma (1/50, alveolar/bronchiolar or carcinoma (11/50, or carcinoma (4/50, 

4/50, 4/48, 7/50) carcinoma, or 14/50, 19/50, 28/50) 7/50, 13/50, 18/50) 

squamous cell 

carcinoma (0/50, 3/49, 

16/50, 16/50) 

Adrenal medulla: 

benign, complex, or 

malignant 

pheochromocytoma 

(2/48, 1/49, 4/50, 

10/48) 

Uncertain findings Adrenal medulla: None None None

benign, complex, or
 

malignant
 

pheochromocytoma
 

(15/50, 19/50, 25/49,
 

20/50)
 

Level of evidence Some evidence Clear evidence Clear evidence Clear evidence

of carcinogenic 

activity 

Genetic toxicology 

Salmonella typhimurium gene mutations: Positive in strain TA100 with and without S9 

Negative in strains TA98 and TA1535 with and without S9 
10 Cobalt Sulfate Heptahydrate, NTP TR 471 

EXPLANATION OF LEVELS OF EVIDENCE OF CARCINOGENIC ACTIVITY 

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

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

animals, do not necessarily mean that a chemical is not a carcinogen, inasmuch as the experiments are conducted under a limited set of 

conditions. Positive results demonstrate that a chemical is carcinogenic for laboratory animals under the conditions of the study and indicate 

that exposure to the chemical has the potential for hazard to humans. Other organizations, such as the International Agency for Research on 

Cancer, assign a strength of evidence for conclusions based on an examination of all available evidence, including animal studies such as 

those conducted by the NTP, epidemiologic studies, and estimates of exposure. Thus, the actual determination of risk to humans from 

chemicals found to be carcinogenic in laboratory animals requires a wider analysis that extends beyond the purview of these studies. 

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

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

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

because of major flaws (inadequate study). These categories of interpretative conclusions were first adopted in June 1983 and then revised 

in March 1986 for use in the Technical Report series to incorporate more specifically the concept of actual weight of evidence of 

carcinogenic activity. For each separate experiment (male rats, female rats, male mice, female mice), one of the following five categories is 

selected to describe the findings. These categories refer to the strength of the experimental evidence and not to potency or mechanism. 

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

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

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

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

incidence of neoplasms (malignant, benign, or combined) in which the strength of the response is less than that required for clear 

evidence. 

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

neoplasms that may be chemical related. 

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

malignant or benign neoplasms. 

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

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

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

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

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

between two adjacent levels. These considerations should include: 

• adequacy of the experimental design and conduct;
 

• occurrence of common versus uncommon neoplasia;
 

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

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

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

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

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

tissue; 

•	 latency in tumor induction; 

•	 multiplicity in site-specific neoplasia; 

•	 metastases; 

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

in another sex or species); 

•	 presence or absence of dose relationships; 

•	 statistical significance of the observed tumor increase; 

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

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

•	 structure-activity correlations; and 

•	 in some cases, genetic toxicology. 
__________ 

Cobalt Sulfate Heptahydrate, NTP TR 471 11 

NATIONAL TOXICOLOGY PROGRAM BOARD OF SCIENTIFIC COUNSELORS
 

TECHNICAL REPORTS REVIEW SUBCOMMITTEE
 

The members of the Technical Reports Review Subcommittee who evaluated the draft NTP Technical Report on cobalt sulfate heptahydrate 

on 11 December 1996 are listed below. Subcommittee members serve as independent scientists, not as representatives of any institution, 

company, or governmental agency. In this capacity, subcommittee members have five major responsibilities in reviewing the NTP studies: 

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

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

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

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

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

Gary P. Carlson, Ph.D., Chairperson Irma Russo, M.D., Principal Reviewer 

School of Health Sciences Fox Chase Cancer Center 

Purdue University Philadelphia, PA 

West Lafayette, IN 

Louise Ryan, Ph.D. Arnold L. Brown, M.D. Division of Biostatistics 

University of Wisconsin Medical School Dana-Farber Cancer Institute 

Madison, WI Boston, MA 

Thomas L. Goldsworthy, Ph.D.* Robert E. Taylor, M.D., Ph.D. 

Department of Experimental Pathology and Toxicology Department of Pharmacology 

Chemical Industry Institute of Toxicology Howard University College of Medicine 

Research Triangle Park, NC Washington, DC 

Robert LeBoeuf, Ph.D. Frederick L. Tyson, Ph.D., Principal Reviewer 

Corporate Professional and Regulatory Services St. Mary’s Hospital and Medical Center 

Human Safety Department Cancer Research Institute 

The Procter & Gamble Company Grand Junction, CO 

Cincinnati, OH 

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

Janardan K. Reddy, M.D. National Cancer Institute 

Department of Pathology Frederick, MD 

Northwestern University Medical School 

Chicago, IL 

* Did not attend 
 

12 Cobalt Sulfate Heptahydrate, NTP TR 471 

SUMMARY OF TECHNICAL REPORTS REVIEW SUBCOMMITTEE COMMENTS
 

On 11 December 1996, the draft Technical Report on 

the toxicology and carcinogenesis studies of cobalt 

sulfate heptahydrate received public review by the 

National Toxicology Program’s Board of Scientific 

Counselors’ Technical Reports Review Subcommittee. 

The review meeting was held at the National Institute 

of Environmental Health Sciences, Research Triangle 

Park, NC. 

Dr. J.R. Bucher, NIEHS, introduced the toxicology 

and carcinogenesis studies of cobalt sulfate heptahydrate by discussing the uses of the chemical and 

rationale for study, describing the experimental 

design, reporting on survival and body weight effects, 

and commenting on the chemical-related neoplastic and 

nonneoplastic lesions in male and female rats and 

mice. The proposed conclusions were some evidence 

of carcinogenic activity in male F344/N rats and clear 

evidence of carcinogenic activity in female F344/N rats 

and male and female B6C3F mice. 1 

Dr. Tyson, a principal reviewer, agreed with the proposed conclusions. Concerning the genetic mechanisms involved in murine lung tumorigenesis, he said 

that although a comprehensive study of K-ras activation was done in lung neoplasms, other molecular 

markers could have been assessed as well. Loss of 

heterozygosity or homozygous deletions on regions of 

chromosome 4, which are syntenic to regions of 

human chromosome 9p21 where frequent deletions are 

observed in human lung cancer, could have been 

studied to determine if similar mechanisms are at work 

in both murine and human lung tumorigenesis via 

exposure to this chemical. Dr. R.C. Sills, NIEHS, 

reported that further studies were planned with the 

next step being to look at loss of heterozygosity not 

only on chromosome 4, but also to look at chromosomes 6 and 11, where the p53 genes are located. 

Dr. Ward, the second principal reviewer, agreed with 

the proposed conclusions. He agreed with the rationale for the exposure concentrations chosen for the 

2-year studies but because there was no concentrationrelated body weight gain depression, he thought that 

rats and mice could have tolerated higher concentrations. With regard to the extensive lesions in the 

nasal cavity and larynx, he stated that this was a 

classic case showing the association between toxic 

and regenerative/reparative lesions resulting in no 

neoplasms. 

Dr. Russo, the third principal reviewer, agreed with 

the proposed conclusions. 

Dr. Tyson moved that the Technical Report on cobalt 

sulfate heptahydrate be accepted with the revisions 

discussed and with the conclusions as written for male 

F344/N rats, some evidence of carcinogenic activity 

and for female F344/N rats and male and female 

B6C3F mice, clear evidence of carcinogenic activity. 1 

Dr. Russo seconded the motion, which was accepted 

unanimously with eight votes. 
 

 

13 

INTRODUCTION
 

CoSO 4 2 C 7H O

COBALT SULFATE HEPTAHYDRATE 

CAS No. 10026-24-1 

Molecular Weight: 281.13 

Synonyms: Bieberite; cobalt(II) sulfate (1:1) heptahydrate; cobalt monosulfate heptahydrate; cobalt(II) sulphate heptahydrate; sulfuric acid, 

cobalt(2+) salt (1:1) heptahydrate 

CHEMICAL AND PHYSICAL PROPERTIES 

Cobalt sulfate is a reddish, crystalline, water-soluble 

powder. It is usually produced as cobalt(II) sulfate but 

can also exist in the cobalt(III) sulfate form with a 

formula of Co (SO ) • 18H O. The heptahydrate salt 2 4 3 2 

is reported to have a structure of [Co(H 2 6 O) ] •

[H SO ] (Merck Index, 1983). Cobalt(II) salts are 2 5 

stable to autoxidation in air or in solution (Smith and 

Carson, 1981). 

PRODUCTION, USE, 

AND HUMAN EXPOSURE 

The production of cobalt sulfate in the United States in 

1983 was estimated to be 450,000 pounds 

(204,000 kg) (J.V. Gandhi, Hall Chemical Co., 

personal communication); more recent production 

estimates are not available. Seven companies were 

listed as producing or handling cobalt sulfate at 10 

facilities in the United States (USDHHS, 1992). 

Cobalt sulfate has been widely used in the electroplating and electrochemical industries. It is used as a 

coloring agent for ceramics and as a drying agent in 

inks, paints, varnishes, and linoleum. Cobalt sulfate 

may be added to animal feed as a mineral supplement 

and has been used as a top dressing on pasture lands 

(De Bie and Doyen, 1962). 

Cobalt is an essential trace element because it is an 

integral part of vitamin B . The human body burden 12 

is approximately 1.1 mg, and the daily intake is about 

0.3 mg, primarily via food (Hammond and Beliles, 

3 1980). Cobalt is found in urban air (0.5 to 60 ng/m )

(Morgan et al., 1970) and has been identified in trace 

amounts in natural waters; concentrations in excess of 

10 μg/L are rare (NRC, 1977). Ocean water contains 

about 0.3 μg/L (Hamilton, 1994). Cobalt has been 

identified in chemical waste dumps (Barrett, 1983). 

In the 1960s, several breweries added cobalt sulfate to 

beer at a level of about 1 ppm to counteract the 

antifoaming activity of detergent residues left on 

poorly rinsed glasses (Morin and Daniel, 1967). Soon 

after this, an epidemic of “beer-drinkers’ cardiomyopathy” occurred, and cobalt was identified as the 

causative agent. The addition of cobalt salts to beer 

was discontinued, and the epidemic ceased. Doses of 

cobalt chloride of up to 200 to 300 mg per day were 

given orally to patients as treatment for various types 

of anemia in the 1950s (Finch, 1980). This practice 
 

 

14 Cobalt Sulfate Heptahydrate, NTP TR 471 

has largely stopped because of associated 

toxicity (gastrointestinal upset, goiter, cardiomyopathies) and the development of less hazardous 

therapies. 

It has been estimated that over 1 million workers in 

the United States are exposed to cobalt or cobalt 

compounds (Jensen and Tüchsen, 1990). Occupational exposure to cobalt occurs principally in refining 

processes, in the production of alloys, and in the 

tungsten carbide hard metal industry (Kazantzis, 

1981). Exposure under these conditions is primarily 

dermal or via inhalation of cobalt metal dusts or 

fumes, often in combination with other elements such as nickel, arsenic, or tungsten; adverse 

respiratory effects (such as pneumoconiosis) have been 

reported at cobalt concentrations between 0.1 and 

3 2 mg/m (Domingo, 1989). The threshold limit valuetimeweighted average for elemental cobalt is 

0.02 mg/m3 (ACGIH, 1996). Airborne levels of 

cobalt dust from spray painting in a Danish porcelain 3 factory in 1981 were as high as 8.6 mg/m (Jensen and

Tüschen, 1990). 

ABSORPTION, DISTRIBUTION, 

METABOLISM, AND EXCRETION 

The absorption of cobalt salts after oral administration 

is variable and is influenced by the nature of the salt, 

the size of the dose, and the presence of food in the 

gastrointestinal tract (Murdock, 1959; Smith et al., 

1972). Clearance of inhaled soluble cobalt salts from 

the lung has not been studied but is expected to be 

rapid (Kerfoot et al., 1975). Several processes could 

contribute to this effect. The water-soluble salts 

dissolve directly, and certain insoluble salts and cobalt 

metal powder appear to have an appreciable solubility 

in protein-containing fluids (Harding, 1950). Clearance by phagocytic alveolar macrophages may also 

occur (Kerfoot et al., 1975). Cobalt is distributed to 

all tissues after administration by the oral or inhalation 

route or by injection (Smith and Carson, 1981). 

Tissue retention is not marked, but higher concentrations have been noted in the liver, kidney, spleen, and 

heart than in other organs (Domingo et al., 1984a,b; 

Llobet et al., 1986). 

Experimental Animals 

In an unspecified strain of rabbits administered cobalt 

sulfate at doses of 0.25 mg/kg per day orally or by 

injection for 2 months, some accumulation of cobalt 

occurred in the liver, small intestine, lung, blood, 

kidney, and stomach (Kichina, 1974). Excretion is 

primarily via the urine and secondarily via the feces. 

The cobalt content of bile collected for 2 hours 57 after intravenous administration of [ Co] cobalt

chloride to Sprague-Dawley rats totaled about 2% to 

5% of the dose over a thirty-fold dose range 

(0.03 mg/kg to 1 mg/kg of Co2+) (Gregus and 

Klaassen, 1986). Several studies have shown that a 

small portion of cobalt, given in several forms by 

parenteral or inhalation routes, is retained in tissues 

with a biological half-time of several years (IARC, 

1991). The form of these materials has not been 

determined, but this could represent uptake into 

vitamin B (Edel et al., 1990). 12 

Humans 

A recent report has demonstrated significant dermal 

absorption of cobalt by humans exposed to mixed 

cobalt-tungsten carbide powders (Scansetti et al., 

1994). The concentration of cobalt in the blood and 

urine of nonoccupationally exposed humans is 0.2 to 

2.0 μg/L (Hamilton, 1994). Cobalt concentrations in 

the urine of workers in the Italian hard metal industry 

were between 10 and 100 μg/L at the beginning of the 

work shift and increased to between 16 and 210 μg/L 

at the end of the work shift (Sabbioni et al., 1994). 

TOXICITY 

Experimental Animals 

Exposure to cobalt results in a wide spectrum of 

toxicities in mammals. The ionic radius of cobalt is 

2+ 2+ between that of Mg and Ca , so cobalt can replace 

or mimic these ions and also may influence reactions 2+ 2+ 2+ 2+ normally involving Fe , Zn , Cu , or Mn 

(Jennette, 1981). For example, cobalt can bind to 

Ca2+-binding proteins in or near microtubules 

(Phillips, 1980) and has been shown to block Ca2+ 

channels in squid axons (Baker et al., 1973). Cobalt 

promotes aberrant microtubule assembly (Buttlaire 

et al., 1980) and can alter the activity of metalloenzymes such as carboxypeptidase (Jennette, 1981). 

Cobalt also inhibits the activity of DNA polymerase I 

from Micrococcus luteus (Korman et al., 1978). 

Cobalt binds to sulfhydryl groups, including those of 

glutathione and cysteine, and through its binding to 

lipoic acid inhibits pyruvate dehydrogenase and 

"-ketoglutarate dehydrogenase, effectively stopping 

oxidative metabolism (Dingle et al., 1962). 
Cobalt Sulfate Heptahydrate, NTP TR 471 15 

A 250 μmol/kg (approximately 60 mg/kg) dose of A single injection of 35 mg/kg cobalt chloride caused 

cobalt chloride heptahydrate administered by sub- degranulation and disintegration of the " cells of the 

cutaneous injection to male Sprague-Dawley rats pancreatic islets in rabbits (Telib, 1972). This was 

caused a rapid increase in biliary excretion of both followed by degranulation of the ß cells. 

reduced and oxidized glutathione, but total hepatic 

glutathione tended to increase after cobalt exposure Although exposure to cobalt affects a wide variety of 

(Stelzer and Klaassen, 1985). enzymatic processes, the acute toxicity of cobalt is not 

as great as might be expected. The oral LD50 for 

A dose of 60 mg cobalt/kg body weight given to an anhydrous cobalt sulfate is 420 mg/kg in male and 

unspecified strain of rats was found to inhibit heme female Wistar rats (Speijers et al., 1982). 

synthesis in the liver (De Matteis and Gibbs, 1977). 

This apparently results from the formation of cobalt Krasovskii and Fridlyand (1971) administered 0.5 or 

protoporphyrin by ferrochelatase and feedback inhi- 2.5 mg/kg cobalt chloride by gavage to rats six times 

bition of *-aminolevulinic acid synthetase activity by per week for 7 months. These investigators found 

the abnormal protoporphyrin (Sinclair et al., 1982). polycythemia and a suppression of leukocyte function. 

Cobalt also induces heme oxygenase (Maines and Myocardial histologic changes were seen in 26 of 

Kappas, 1976), and the combined effect of these 30 rats given 26 mg/kg cobalt sulfate by gavage once 

actions is to rapidly decrease the cytochrome P daily for 8 weeks (Grice et al., 1969). This study is 450 

concentrations in the liver. Other cytochromes appear representative of a large number of animal studies 

to be less affected (Tephly and Hibbeln, 1971). designed to examine beer-drinkers’ cardiomyopathy 

(cited in Smith and Carson, 1981, and USDHHS, 

In contrast to its actions on heme synthesis in the 1992). Overall, these studies indicated that rather 

liver, cobalt administration promotes polycythemia. large doses of cobalt could mimic the cardiomyopathy 

This effect is more pronounced in humans than in caused by cobalt-treated beer, but that cobalt probably 

rodents (Smith and Carson, 1981) and is the basis for acted synergistically in humans with thiamine defithe use of cobalt chloride to treat anemia. The oral ciency and an insufficient intake of sulfur-containing 

administration of 10 mg cobalt/kg body weight given amino acids. Deficits in thyroid function have been 

as cobalt chloride to male rats of unspecified strain shown in l-day-old chicks and guinea pigs but not in 

five times per week for 150 days resulted in an young chicks, rats, mice, or rabbits given cobalt 

increase in the erythrocyte count, hematocrit value, (Sederholm et al., 1968). 

and hemoglobin concentration of the blood; however, 

the mean cell volume and hemoglobin concentration A variety of cobalt dusts and aerosols have been 

per cell were unchanged, indicating a simple poly- administered to animals via inhalation. Results of 

cythemic effect (Murdock, 1959). This response is these studies indicate that lung compliance is decreased 

mediated by an increase in circulating erythropoietin, and that electrical properties of the heart are affected 

postulated to be a secondary response to a central as in beer-drinkers’ cardiomyopathy (Kerfoot et al., 

nervous system effect of cobalt which results in 1975; Smith, 1980). In general, similar toxicity has 

respiratory alkalosis. Alkalosis increases the affinity been elicited by cobalt whether administered orally or 

of heme for oxygen, which is interpreted by tissue by inhalation. These effects have been seen after 

“sensors” as hypoxia (Miller et al., 1974). exposure of rats to atmospheres containing 0.05 or 

0.5 mg/m3 cobalt for 3 months (Popov, 1977). In 

A second effect of cobalt administration on the blood addition, specific pulmonary effects in male rabbits 

3 is an increase in triglycerides, cholesterol, and free exposed to 0.5 mg/m cobalt (as cobalt chloride) by 

fatty acids (Taylor and Marks, 1978). This may be inhalation for 6 hours per day, 5 days per week, for 4 

caused by inhibition of tissue lipoprotein lipase, to 6 weeks included a change in the growth pattern of 

resulting in failure to clear very low-density lipo- alveolar type II cells, resulting in clusters of cells 

protein (Taylor and Marks, 1978), and perhaps by projecting into the alveolar lumen, and changes in 

stimulation of lipoprotein synthesis in the liver oxidative metabolism of lung macrophages (Johansson 

(Eaton, 1972). et al., 1984, 1986). 
 

16 Cobalt Sulfate Heptahydrate, NTP TR 471 

Sixteen-day and 13-week inhalation studies with cobalt 

sulfate heptahydrate in F344/N rats and B6C3F mice 1 

have been reported (Bucher et al., 1990; NTP, 1991). 

In the 13-week studies, groups of 10 male and 

10 female rats and mice were exposed to cobalt sulfate 

heptahydrate concentrations ranging from 0 to 

3 30 mg/m , 6 hours per day, 5 days per week. Two 

male mice exposed to 30 mg/m3 died. All groups at 

this concentration initially lost weight, but then gained 

weight at rates similar to controls. At the end of the 

studies, lung weights were generally increased in rats 

and mice exposed to 1.0 mg/m3 and higher, and 

polycythemia was observed in exposed rats but not in 

mice. Lesions observed in the respiratory tract of rats 

and mice included degeneration of the olfactory 

epithelium, squamous metaplasia of the respiratory 

epithelium, and inflammation in the nose; inflammation, necrosis, squamous metaplasia, ulcers (rats), 

and inflammatory polyps (rats) of the larynx; squamous metaplasia of the trachea (mice); and histiocytic 

infiltrates, bronchiolar regeneration, peribronchiolar 

and septal fibrosis, and epithelial hyperplasia in the 

alveoli of the lung. A no-observed-adverse-effectlevel (NOAEL) was not reached in these studies as 

lesions, particularly in the larynx, were observed at 

3 the lowest exposure (0.3 mg/m ) used. 

In other NTP studies (unpublished, available upon 

request), cobalt sulfate elicited contact hypersensitivity. Female Hartley guinea pigs received dermal 

applications of 100 μL of an aqueous 6% solution 

once per day for 14 days. A dose-related increase in 

contact hypersensitivity, as measured by retention of 

labeled inflammatory cells in the skin, was observed 

upon challenge application of solutions of 0.3%, 1%, 

or 3% aqueous cobalt sulfate to a site distant from the 

induction site 7 days after the last induction dose. 

Erythema and edema in the ears and paws of rats 

resulted from the administration of 5 mg cobalt sulfate 

by injection (Jasmin, 1974). 

Humans 

Besides myocardial toxicity, as noted above, a second 

effect of cobalt observed in victims of beer-drinkers’ 

cardiomyopathy was hypothyroidism (Taylor and 

Marks, 1978). Thyroid function tests, including 

uptake of [131I]iodide, were also depressed in patients 

receiving 0.17 to 3.9 mg/kg cobalt per day for treatment of anemia (Paley et al., 1958). It has been 

proposed that cobalt interferes with binding of inorganic iodide to tyrosine in the thyroid gland. 

Hypersensitivity reactions have been observed in 

patients who received prosthetic implants made of a 

cobalt alloy and in industrial workers exposed to 

cobalt dusts (Smith and Carson, 1981). Asthma 

related to cobalt exposure has also been described 

(Cirla, 1994). 

Most inhalation of cobalt is by workers in the refining 

and alloy production industries (NIOSH, 1981). The 

dusts may be in the form of the metal, its alloys, or its 

salts, but most often the oxide form is present. Consequently, no toxicity studies exist on exposure to pure 

cobalt metal or to cobalt sulfate. Exposure appears to 

cause pulmonary fibrosis, splenic enlargement, dermatitis, and losses of appetite and sense of smell 

(Dorsit et al., 1970). Cobalt is used in the cemented 

tungsten carbide industry and is thought to be primarily responsible for pulmonary “hard metal disease,” consisting of upper respiratory tract irritation, 

pneumonitis, and pulmonary fibrosis (NIOSH, 1981). 

However, the actual role of inhaled cobalt versus an 

interaction of cobalt and other inhaled particles 

remains a subject of debate (Swennen et al., 1993). 

REPRODUCTIVE 

AND DEVELOPMENTAL EFFECTS 

Experimental Animals 

Sprague-Dawley rats maintained on diets containing 

265 ppm cobalt for 98 days showed degenerative 

changes in the testis; these changes were considered 

secondary to hypoxia (Mollenhaur et al., 1985). 

Decreases in sperm motility and/or increased abnormal 

sperm were noted in mice, but not in rats, exposed to 

3 3 mg/m or higher in 13-week inhalation studies with

cobalt sulfate (NTP, 1991). Following 13 weeks of 

chronic exposure to 100 to 400 ppm cobalt chloride in 

drinking water, male CD-1 mice showed marked doserelated decreases in fertility, testicular weight, and 

sperm concentration and motility, and increases in 

circulating levels of testosterone (Pedigo et al., 1988). 

Cobalt has been shown to cross the placenta; cobalt 

chloride and nitrite salt solutions induced fetal cleft 
Cobalt Sulfate Heptahydrate, NTP TR 471 17 

palates when injected alone into mouse dams, but 

inhibited cleft formation caused by cortisone or phenytoin (Kasirsky et al., 1969; Mitala et al., 1978). 

Oral exposure of rats to cobalt chloride at daily doses 

of 5.4 or 21.8 mg cobalt/kg body weight from gestation day 14 through lactation day 21 resulted in 

stunted growth and/or decreased pup survival, although adverse effects were also evident in the dams 

at both doses (Domingo et al., 1985). In contrast, 

Paternain et al. (1988) reported that doses of up to 

100 mg/kg cobalt chloride administered by gavage to 

pregnant Sprague-Dawley rats once per day on days 6 

to 15 of gestation did not result in significant fetotoxicity or teratogenicity. Similarly, Seidenberg et al. 

(1986) reported no effect on mouse fetal growth or 

mortality in dams given daily doses of 81.7 mg 

cobalt/kg on days 8 to 12 of pregnancy. 

Humans 

Cobalt has not been shown to cause significant teratogenic or reproductive effects in humans (Smith and 

Carson, 1981). No clinical effects were noted in the 

babies of women who had taken cobalt chloride to 

counter anemia while pregnant (Jacobziner and 

Raybin, 1961). 

CARCINOGENICITY 

Experimental Animals 

There have been no reports of adequate chronic 

inhalation toxicity or carcinogenicity studies with 

soluble or insoluble cobalt salts or metal powders 

(IARC, 1991). Wehner et al. (1977) found no increase in tumors in Syrian golden hamsters exposed to 

10 mg/m3 cobalt oxide dust for 7 hours per day, 

5 days per week, for life; however, the study was 

faulted for poor survival (IARC, 1991). Cobalt oxide 

has been studied by intratracheal administration to 

groups of 50 male and 50 female Sprague-Dawley rats 

(Steinhoff and Mohr, 1991). Doses of 2 or 10 mg/kg 

were given in 19 treatments at 2-week intervals and in 

10 treatments at 4-week intervals over 2 years. Two 

groups of 50 male and 50 female controls received 

saline or no treatment. Approximately 80% of the 

material was within the particle size range of 5 to 

40 μm. At the end of the study an unspecified 

bronchioalveolar proliferation was noted in 51 of 100 

low-dose rats (male and females combined), in 70 of 

100 high-dose rats, and in no controls. One male and 

one female from the low-dose groups developed a 

benign lung tumor, and one high-dose female had a 

bronchioalveolar carcinoma. Three adenocarcinomas 

and two bronchioalveolar adenomas were observed in 

high-dose males. No lung tumors occurred in the 

controls. In a similar but smaller study by the same 

group, cobalt oxide was found to enhance the lung 

tumor yield of benzo[a]pyrene treatment (Steinhoff and 

Mohr, 1991). 

Sarcomas in rats have been observed at the site of 

injection of cobalt salts or cobalt metal powder (IARC, 

1991). Heath (1956, 1960) gave an unspecified strain 

of rats a single injection of 0.28 mg cobalt metal 

powder in fowl serum into the thigh muscle. Within 

2 weeks, atypical myoblasts were observed (Heath, 

1960), and between 5 and 12 months, malignant 

neoplasms developed at the injection site in 17 of 

30 rats; 11 were rhabdomyosarcomas (Heath, 1956). 

Gilman (1962) reported a similar neoplastic response 

to injections of cobalt sulfide and cobalt oxide in an 

unspecified strain of rats but saw no neoplasms in an 

unspecified strain of mice. These materials are 

relatively insoluble, and Abbracchio et al. (1982) 

suggested that intracellular solubilization of relatively 

insoluble cobalt salts would favor cellular transformation. Heath and Webb (1967) determined that 

cobalt is bound intracellularly in primary rhabdomyosarcomas induced by intramuscular injection of 

metallic cobalt, with 70% to 90% of the bound cobalt 

found in the nucleus. Further fractionation studies 

demonstrated that 50% of the nuclear cobalt is bound 

in the nucleolus (Webb et al., 1972). Similar injection 

studies have given little evidence of cobalt-induced 

cancer in mice, hamsters, or guinea pigs (Christensen 

and Poulsen, 1994). 

There is only one report of the formation of neoplasms 

after injection of a soluble cobalt salt. Shabaan et al. 

(1977) observed fibrosarcomas in 14 of 40 male 

Wistar rats 8 months to 1 year after administration of 

40 mg/kg cobalt chloride by subcutaneous injection 

once per day for 10 days. Four of these neoplasms 

were not at the site of injection. 

Humans 

Cobalt has been used in hundreds of patients as part 

of an alloy with chromium and molybdenum in 

prosthetic implants. During the first 14 years of its 

use for this purpose, no fibrosarcomas were identified 

in the recipients (McKee, 1971); however, a number 

of cases of malignant neoplasia have been reported 
18 Cobalt Sulfate Heptahydrate, NTP TR 471 

since that time at the sites of metal-containing fracture 

plates or joint prostheses, some of which contained 

cobalt (IARC, 1991). 

The IARC (1991) considered the available data 

inadequate to establish an association between cancer 

and cobalt exposure to humans. At that time there 

were two epidemiological studies that were considered 

adequate for evaluation (Mur et al., 1987; Hogstedt 

and Alexandersson, 1990). The Mur et al. (1987) 

cohort study was composed of 1,143 workers who 

were employed for at least a year between 1950 and 

1980 in a French electrochemical plant producing 

cobalt and sodium. For workers employed only in 

cobalt production, the standard mortality ratio for lung 

cancer was 466 (95% confidence interval from 146 

to 1,064) based on four cases. Hogstedt and 

Alexandersson (1990) studied a cohort of 3,163 male 

Swedish workers with at least 1 year of exposure to 

cobalt-containing, hard-metal dust ore between 1940 

and 1982. There were 17 cases of lung cancer versus 

12.7 expected (SMR, 134; 95% CI 77 to 213). 

Interpretation of both studies was made difficult by 

concurrent exposures to other substances including 

arsenic and nickel in the French plant and tungsten 

carbide in the Swedish facility. 

Since the IARC evaluation, a follow-up study of the 

French electrochemical plant workers was completed 

which extended the period of observation from 1981 

to 1988. No additional lung cancers were observed. 

Based on this and other factors, the authors concluded 

that the data no longer supported an association of 

cobalt exposure with lung cancer (Moulin et al., 

1993). In contrast, Lasfargues et al. (1994) reported 

on a cohort mortality study carried out on workers at 

a French hard-metal plant. The study specifically 

addressed lung cancer risks in relation to cobalt 

exposure and included 709 male workers who had at 

least 1 year of employment at the plant and who died 

between the years 1956 and 1989. While overall 

mortality was not increased, death due to lung cancer 

was significantly elevated (SMRù213), with 10 cases 

observed. This excess was associated with high cobalt 

exposure, but no effect of employment duration was 

noted. Smoking did not account for the observed 

incidence of lung cancer. 

GENETIC TOXICITY 

Genetic toxicity data for cobalt sulfate heptahydrate 

are limited to a single publication. Zeiger et al. 

(1992) reported the results of a mutagenicity study 

with cobalt sulfate heptahydrate which showed a 

weakly positive response in Salmonella typhimurium 

strain TA100 in the absence of exogenous metabolic 

activation as well as with hamster or rat liver S9; the 

authors reported no induction of mutations in strain 

TA98 or TA1535, with or without S9. 

Few studies with other cobalt compounds have been 

reported. The literature on genetic and related effects 

of cobalt compounds was reviewed by Beyersmann 

and Hartwig (1992). Most of the bacterial mutagenicity test results included in this review were 

negative. However, some positive results were 

reported for mammalian cell DNA damage studies, 

including the observation of DNA strand breaks in 

human cells (McLean et al., 1982; Hamilton-Koch 

et al., 1986; Hartwig et al., 1990) and sister chromatid exchange induction in human (Anderson, 1983) 

and hamster cells (Hartwig et al., 1991) treated 

in vitro with cobalt chloride in the absence of exogenous metabolic systems. The authors discussed the 

possible role of hydroxyl and superoxide radical formation in the generation of DNA breaks (Beyersmann 

and Hartwig, 1992). Morita et al. (1991) reported a 

weak response in an in vitro test designed to detect 

increased frequencies of 6-thioguanine-resistant mutant 

FM3A cell colonies. At a concentration of 

2 × 10-4 M cobalt chloride (which induced a 50% 

decrease in cell survival), an increased number of 

mutant colonies (approximately four to five times the 

control number) was observed. At concentrations 

higher and lower than 2 × 10-4 M, the mutagenic 

response was weaker. The authors suggested, based 

upon results from the testing of other known mutagens 

in this assay, that metal ions such as cobalt require 

relatively high concentrations and long exposure 

periods to induce an effect and that the induced 

mutagenic response obtained is weak and seen over a 

narrow dose range. In the Drosophila wing spot test, 

cobalt chloride was demonstrated to induce a significant, dose-dependent increase in somatic recombination in third instar larvae exposed to cobalt 

chloride concentrations of 2 to 10 mM during development to the adult stage (Ogawa et al., 1994). 
Cobalt Sulfate Heptahydrate, NTP TR 471 19 

STUDY RATIONALE 

Cobalt sulfate was nominated by the National Cancer 

Institute for study based on a lack of information on 

the toxicity of soluble cobalt salts. The more common 

cobalt(II) form and the inhalation route were selected 

for study to mimic worker exposure. Prechronic 

studies were previously reported (Bucher et al., 1990; 

NTP, 1991) with a spectrum of lesions noted in the 

respiratory tract of rats and mice. Polycythemia was 

also observed in rats. A NOAEL was not reached in 

3 these studies using doses as low as 0.3 mg/m . This 

report documents the findings of 2-year inhalation 

exposure studies with cobalt sulfate heptahydrate in 

F344/N rats and B6C3F mice. 1 
20 Cobalt Sulfate Heptahydrate, NTP TR 471 
21 

MATERIALS AND METHODS
 

PROCUREMENT 

AND CHARACTERIZATION 

OF COBALT SULFATE HEPTAHYDRATE 

Cobalt sulfate heptahydrate was obtained from Curtin 

Matheson Scientific (Kansas City, MO) in one lot 

(412092). Identity and purity analyses were conducted 

by the analytical chemistry laboratory, Midwest 

Research Institute (Kansas City, MO) (Appendix F). 

Reports on analyses performed in support of the cobalt 

sulfate heptahydrate studies are on file at the National 

Institute of Environmental Health Sciences. 

The chemical, a red, crystalline solid, was identified as 

cobalt sulfate heptahydrate by infrared, ultraviolet, 

and/or visible spectroscopy. The purity of lot 412092 

was determined by elemental analysis, Karl Fischer 

water analysis, and spark source mass spectroscopy. 

Elemental analyses for sulfur and hydrogen were in 

agreement with the theoretical values for cobalt sulfate 

heptahydrate, but results for cobalt were slightly low. 

Karl Fischer water analysis indicated 44.6% ± 0.5% 

water. Spark source mass spectroscopy indicated 

140 ppm nickel present as an impurity; all other 

impurities had a combined total of less than 175 ppm. 

The overall purity was determined to be approximately 

99%. 

Literature references indicate that cobalt sulfate 

heptahydrate is stable as a bulk chemical when stored 

protected from light at normal temperatures. The 

heptahydrate dehydrates to the hexahydrate at 41.5E C 

and to the monohydrate when heated to 71E C, with 

no further changes expected below the decomposition 

temperature (708E C). Therefore, an accelerated 

stability study was not conducted. To ensure stability, 

the bulk chemical was stored in its original shipping 

containers, metal cans, at room temperature. Stability 

was monitored during the studies using elemental 

analysis by inductively coupled plasma/atomic emission spectroscopy (ICP/AES) normalized against a 

cobalt standard (National Institute of Standards and 

Technology, Gaithersburg, MD); no degradation of 

the bulk chemical was detected. 

AEROSOL GENERATION 

AND EXPOSURE SYSTEM 

Cobalt sulfate heptahydrate was generated and delivered from an aqueous solution by a system composed 

of three main components: a compressed-air-driven 

nebulizer (Model PN7002; RETEC Development 

Laboratory, Portland, OR), an aerosol charge neutralizer, and an aerosol distribution system. Cobalt 

sulfate heptahydrate in deionized water was siphoned 

from the bulk reservoir to the nebulizer reservoir and 

then aspirated into the nebulizer chamber and expelled 

as a stream. Shear forces broke the stream into 

droplets that were evaporated to leave dry particles of 

cobalt sulfate heptahydrate. The aerosol generation 

and delivery system included primary and secondary 

compressed-air-driven nebulizers. The aerosol generated by the compressed-air-driven nebulizer was 

passed through the aerosol charge neutralizer to 

remove static charge that formed on the aerosol 

particles during generation. Detailed descriptions of 

the inhalation chambers and the vapor generation 

system are provided in Appendix F. 

A distribution line carried aerosol to the Hazleton 

2000 inhalation exposure chambers (Harford Systems 

Division of Lab Products, Inc., Aberdeen, MD) on 

both sides of the exposure room. At each chamber, 

aerosol moving through the chamber inlet was further 

diluted with HEPA-filtered air to the appropriate 

concentration for the chamber. 

AEROSOL CONCENTRATION 

MONITORING 

The chamber concentrations of cobalt sulfate heptahydrate were monitored by computer-controlled realtime aerosol monitors (Model RAM-1; MIE, Inc., 
22 Cobalt Sulfate Heptahydrate, NTP TR 471 

Bedford, MA). Chamber aerosol concentrations were 

sampled at least once per hour during each exposure 

day. Throughout the studies, the background concentrations of total suspended particles in the control 

chambers were less than the limit of detection. The 

RAM-1 voltage output was calibrated against cobalt 

sulfate heptahydrate concentrations of chamber filter 

samples. Solutions of filter samples in 2% nitric acid 

were analyzed quantitatively for cobalt sulfate heptahydrate by ICP/AES. The ICP/AES was calibrated 

with a solution of standard cobalt diluted with nitric 

acid. Stability studies performed with X-ray diffraction analyses of samples from the 0.3 and 3.0 mg/m3 

chambers indicated that cobalt sulfate hexahydrate was 

the primary species delivered to the chambers. Chamber concentration uniformity was maintained throughout the 2-year studies. A summary of chamber concentrations is presented in Table F1. 

CHAMBER ATMOSPHERE 

CHARACTERIZATION 

The time required for the chamber concentration to 

reach 90% of the target value following the beginning 

of exposure (T ) and the time required for the cham­ 90 

ber concentration to reach 10% of the target value 

following termination of the exposure (T ) were 10 

determined for each exposure chamber. Without 

animals present, T values ranged from 9 to 90 

11 minutes for rats and from 7 to 12 minutes for mice; 

T10 ranged from 8 to 9 minutes for rats and mice. 

With animals present, T90 values ranged from 11 to 

16 minutes for rats and from 8 to 12 minutes for mice; 

T10 ranged from 12 to 13 minutes for rats and from 11 

to 12 minutes for mice. A T90 of 12 minutes was 

selected for the 2-year studies. 

Aerosol size distribution was determined monthly for 

each exposure chamber with a Mercer-style sevenstage impactor (In-Tox Products, Albuquerque, NM). 

Samples were analyzed for cobalt sulfate heptahydrate 

with ICP/AES. The relative mass on each impactor 

stage was analyzed by probit analysis; the mass 

median aerodynamic diameter for the aerosol was 

within the specified range of 1 to 3 μm (Tables F2 

and F3). 

Studies of cobalt sulfate heptahydrate degradation and 

monitoring for impurities were conducted throughout 

the 2-year studies with ICP/AES. No degradation of 

cobalt sulfate heptahydrate was observed during the 

studies. Cageboards were used after the first 8 weeks 

of the studies to control ammonia in the exposure 

chambers. 

2-YEAR STUDIES 

Study Design 

Groups of 50 male and 50 female rats and mice were 

exposed to aqueous aerosols containing 0, 0.3, 1.0, or 3 3.0 mg/m cobalt sulfate heptahydrate for 6 hours plus

T (12 minutes) per day, 5 days per week, for 90 

105 weeks. 

The exposure concentrations for the 2-year cobalt 

sulfate heptahydrate studies were based on the findings 

of 16-day and 13-week studies reported previously 

(NTP, 1991). The most sensitive tissue was the 

larynx, with squamous metaplasia observed in rats and 

mice at the lowest exposure concentration of 

3 0.3 mg/m . A NOAEL was not reached for this 

tissue. Inflammatory polyps, some nearly obstructing 3 the esophagus, were observed at 10 and 30 mg/m in

rats, while these lesions at the 0.3 and 1.0 mg/m3 

exposure concentrations were composed of mild or 

minimal squamous metaplasia and/or chronic inflammation in both rats and mice. The severity of the 

laryngeal changes and other lesions in the respiratory 

tract at 3.0 mg/m3 was not considered life threatening, 

and, therefore, exposure concentrations of 0.3, 1.0, 3 and 3.0 mg/m were chosen for the 2-year study with

rats and mice. 

Source and Specification of Animals 

Male and female F344/N rats and B6C3F1 mice were

obtained from Simonsen Laboratories (Gilroy, CA) for 

use in the 2-year studies. Rats and mice were quarantined for 14 days before the beginning of the studies. Five male and five female rats and mice were 

selected for parasite evaluation and gross observation 

of disease. Serology samples were collected for viral 

screening. Rats and mice were approximately 6 weeks 

old at the beginning of the studies. The health of the 

animals was monitored during the studies according to 

the protocols of the NTP Sentinel Animal Program 

(Appendix H). 

Animal Maintenance 

Rats and mice were housed individually. Feed was 

available ad libitum except during exposure periods, 

and water was available ad libitum. Cages and racks 
Cobalt Sulfate Heptahydrate, NTP TR 471 23 

were rotated weekly. Further details of animal maintenance are given in Table 1. Information on feed 

composition and contaminants is provided in 

Appendix G. 

Clinical Examinations and Pathology 

All animals were observed twice daily. Clinical findings and body weights were recorded initially, at 

weeks 5, 9, and 13 (clinical findings) or weekly for 

13 weeks (body weights), monthly through week 92, 

every 2 weeks thereafter, and at the end of the studies. 

A complete necropsy and microscopic examination 

were performed on all rats and mice. At necropsy, all 

organs and tissues were examined for grossly visible 

lesions, and all major tissues were fixed and preserved 

in 10% neutral buffered formalin, processed and 

trimmed, embedded in paraffin, sectioned to a thickness of 5 to 6 μm, and stained with hematoxylin and 

eosin for microscopic examination. For all paired 

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

from each organ were examined. Tissues examined 

microscopically are listed in Table 1. 

Microscopic evaluations were completed by the study 

laboratory pathologist, and the pathology data were 

entered into the Toxicology Data Management System. 

The slides, paraffin blocks, and residual wet tissues 

were sent to the NTP Archives for inventory, 

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

individual animal data records, and pathology tables 

were evaluated by an independent quality assessment 

laboratory. The individual animal records and tables 

were compared for accuracy, the slide and tissue 

counts were verified, and the histotechnique was 

evaluated. For the 2-year studies, a quality assessment 

pathologist evaluated slides from all tumors and all 

potential target organs which included the adrenal 

medulla, lung, larynx, nose, and all neoplasms in all 

groups except testicular neoplasms for male and 

female rats. For male and female mice, the quality 

assessment pathologist evaluated slides from all tumors 

and all potential target organs which included the 

larynx, liver, lung, nose, and trachea, and all neoplasms in all organs. Additionally, all thyroid glands 

were reviewed for incidences of proliferative lesions of 

the follicular cells. Renal and iliac lymph nodes of 

male mice were reviewed when the diagnosis of 

lymphoid hyperplasia occurred. Ovaries of female 

mice were reviewed when the diagnoses of cyst, 

bilateral cyst, or corpus luteum cyst occurred. 

The quality assessment report and the reviewed slides 

were submitted to the NTP Pathology Working Group 

(PWG) chairperson, who reviewed the selected tissues 

and addressed any inconsistencies in the diagnoses 

made by the laboratory and quality assessment pathologists. Representative histopathology slides containing examples of lesions related to chemical administration, examples of disagreements in diagnoses 

between the laboratory and quality assessment pathologist, or lesions of general interest were presented by 

the chairperson to the PWG for review. The PWG 

consisted of the quality assessment pathologist and 

other pathologists experienced in rodent toxicologic 

pathology. This group examined the tissues usually 

without any knowledge of dose groups or previously 

rendered diagnoses. When the PWG consensus 

differed from the opinion of the laboratory pathologist, 

the diagnosis was changed. Final diagnoses for 

reviewed lesions represent a consensus between the 

laboratory pathologist, reviewing pathologist(s), and 

the PWG. Details of these review procedures have 

been described, in part, by Maronpot and Boorman 

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

analyses of the pathology data, the decision of whether 

to evaluate the diagnosed lesions for each tissue type 

separately or combined was generally based on the 

guidelines of McConnell et al. (1986). 
24 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE 1 

Experimental Design and Materials and Methods in the 2-Year Inhalation Studies 

of Cobalt Sulfate Heptahydrate 

Study Laboratory 

Battelle Pacific Northwest Laboratories 

(Richland, WA) 

Strain and Species 

Rats: F344/N 

Mice: B6C3F1 

Animal Source 

Simonsen Laboratories 

(Gilroy, CA) 

Time Held Before Studies 

14 days 

Average Age When Studies Began 

6 weeks 

Date of First Exposure 

Rats: 30 August 1990 

Mice: 23 August 1990 

Duration of Exposure 

6 hours plus T90 (12 minutes) per day, 5 days per week, for 105 weeks 

Date of Last Exposure 

Rats: 28 August 1992 

Mice: 21 August 1992 

Necropsy Dates 

Rats: 1-4 September 1992 

Mice: 24-27 August 1992 

Average Age at Necropsy 

111 weeks 

Size of Study Groups 

50 males and 50 females 

Method of Distribution 

Animals were distributed randomly into groups of approximately equal initial mean body weights; cages were distributed randomly into 

groups from another computer-generated list of random numbers. 

Animals per Cage 

1 
Cobalt Sulfate Heptahydrate, NTP TR 471 25 

TABLE 1 

Experimental Design and Materials and Methods in the 2-Year Inhalation Studies 

of Cobalt Sulfate Heptahydrate 

Method of Animal Identification 

Tail tattoo 

Diet 

NIH-07 open formula pellet diet (Zeigler Brothers, Inc., Gardners, PA), available ad libitum except during exposure periods, changed 

weekly 

Water Distribution 

Tap water (Richland municipal supply) via automatic watering system (Edstrom Industries, Waterford, WI), available ad libitum 

Cages 

Stainless-steel wire-bottom (Hazleton System, Inc., Aberdeen, MD), changed weekly 

Bedding 

Cageboard (Bunzl Cincinnati Paper Co., Cincinnati, OH), changed daily (15 October 1990 to study termination) 

Chamber Air Supply Filters 

Single HEPA (Flanders Filters, Inc., San Rafael, CA) 

Chambers 

Stainless-steel with excreta pan suspended below each cage unit (Harford System Division of Lab Products, Inc., Aberdeen, MD), changed 

weekly 

Chamber Environment 

Temperature: 21.3E–26.6E C (rats); 19.5E–27.1E C (mice)
 

Relative humidity: 31%–89% (rats); 28%–93% (mice)
 

Room fluorescent light: 12 hours/day
 

Chamber air changes: 9-23/hour
 

Exposure Concentrations 

0, 0.3, 1.0, or 3.0 mg/m3 

Type and Frequency of Observation 

Observed twice daily; animals were weighed and clinical findings were recorded initially, at weeks 5, 9, and 13 (clinical findings) or weekly 

for 13 weeks (body weights), monthly through week 92, every 2 weeks thereafter, and at the end of the studies. 

Method of Sacrifice 

CO2 anesthetization

Necropsy 

Necropsy performed on all animals 

Histopathology 

Complete histopathology was performed on all rats and mice. In addition to gross lesions and tissue masses, the tissues examined included: 

adrenal gland, bone with marrow, brain, clitoral gland, esophagus, gallbladder (mice), harderian gland (rats), heart, large intestine (cecum, 

colon, rectum), small intestine (duodenum, jejunum, ileum), kidney, larynx, liver, lungs/bronchi, lymph nodes (mandibular, mesenteric, 

bronchial, mediastinal), mammary gland (except male mice), nose, oral cavity (rats), ovary, pancreas, pancreatic islets, parathyroid gland, 

pituitary gland, preputial gland, prostate gland, salivary gland, sciatic nerve, seminal vesicle, skin, spinal cord, spleen, stomach (forestomach 

and glandular), testes/epididymides, thymus, thyroid gland, trachea, urinary bladder, and uterus. 
26 Cobalt Sulfate Heptahydrate, NTP TR 471 

STATISTICAL METHODS 

Survival Analyses 

The probability of survival was estimated by the 

product-limit procedure of Kaplan and Meier (1958) 

and is presented in the form of graphs. Animals found 

dead of other than natural causes or pregnant were 

censored from the survival analyses; animals dying 

from natural causes were not censored. Statistical 

analyses for possible dose-related effects on survival 

used Cox’s (1972) method for testing two groups for 

equality and Tarone’s (1975) life table test to identify 

dose-related trends. All reported P values for the 

survival analyses are two sided. 

Calculation of Incidence 

The incidences of neoplasms or nonneoplastic lesions 

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

and D5 are given as the number of animals bearing 

such lesions at a specific anatomic site and the number 

of animals with that site examined microscopically. 

For calculation of statistical significance, the incidences of most neoplasms (Tables A3, B3, C3, and 

D3) and all nonneoplastic lesions are given as the 

numbers of animals affected at each site examined 

microscopically. However, when macroscopic 

examination was required to detect neoplasms in 

certain tissues (e.g., harderian gland, intestine, mammary gland, and skin) before microscopic evaluation, 

or when neoplasms had multiple potential sites of 

occurrence (e.g., leukemia or lymphoma), the 

denominators consist of the number of animals on 

which a necropsy was performed. Tables A3, B3, C3, 

and D3 also give the survival-adjusted neoplasm rate 

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

the Kaplan-Meier estimate of the neoplasm incidence 

that would have been observed at the end of the study 

in the absence of mortality from all other competing 

risks (Kaplan and Meier, 1958). 

Analysis of Neoplasm Incidences 

The majority of neoplasms in these studies were 

considered to be incidental to the cause of death or not 

rapidly lethal. Thus, the primary statistical method 

used was logistic regression analysis, which assumed 

that the diagnosed neoplasms were discovered as the 

result of death from an unrelated cause and thus did 

not affect the risk of death. In this approach, neoplasm prevalence was modeled as a logistic function 

of chemical exposure and time. Both linear and 

quadratic terms in time were incorporated initially, and 

the quadratic term was eliminated if the fit of the 

model was not significantly enhanced. The neoplasm 

incidences of exposed and control groups were compared on the basis of the likelihood score test for the 

regression coefficient of dose. This method of 

adjusting for intercurrent mortality is the prevalence 

analysis of Dinse and Lagakos (1983), further described and illustrated by Dinse and Haseman (1986). 

When neoplasms are incidental, this comparison of the 

time-specific neoplasm prevalences also provides a 

comparison of the time-specific neoplasm incidences 

(McKnight and Crowley, 1984). 

In addition to logistic regression, other methods of 

statistical analysis were used, and the results of these 

tests are summarized in the appendixes. These methods include the life table test (Cox, 1972; Tarone, 

1975), appropriate for rapidly lethal neoplasms, and 

the Fisher exact test and the Cochran-Armitage trend 

test (Armitage, 1971; Gart et al., 1979), procedures 

based on the overall proportion of neoplasm-bearing 

animals. 

Tests of significance included pairwise comparisons of 

each exposed group with controls and a test for an 

overall dose-related trend. Continuity-corrected tests 

were used in the analysis of neoplasm incidence, and 

reported P values are one sided. The procedures 

described in the preceding paragraphs were also used 

to evaluate selected nonneoplastic lesions. For further 

discussion of these statistical methods, refer to 

Haseman (1984). 

Analysis of Nonneoplastic Lesion Incidences 

Because all nonneoplastic lesions in this study were 

considered to be incidental to the cause of death or not 

rapidly lethal, the primary statistical analysis used was 

a logistic regression analysis in which nonneoplastic 

lesion prevalence was modeled as a logistic function of 

chemical exposure and time. 

Analysis of Continuous Variables 

Average severity values were analyzed for significance 

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

1973). 
Cobalt Sulfate Heptahydrate, NTP TR 471 27 

Historical Control Data 

Although the concurrent control group is always the 

first and most appropriate control group used for 

evaluation, historical control data can be helpful in the 

overall assessment of neoplasm incidence in certain 

instances. Consequently, neoplasm incidences from 

the NTP historical control database, which is updated 

yearly, are included in the NTP reports for neoplasms 

appearing to show compound-related effects. 

QUALITY ASSURANCE METHODS 

The studies were conducted in compliance with Food 

and Drug Administration Good Laboratory Practice 

Regulations (21 CFR, Part 58). In addition, as 

records from the studies were submitted to the NTP 

Archives, these studies were audited retrospectively by 

an independent quality assurance contractor. Separate 

audits covering completeness and accuracy of the 

pathology data, pathology specimens, final pathology 

tables, and a draft of this NTP Technical Report were 

conducted. Audit procedures and findings are presented in the reports and are on file at NIEHS. The 

audit findings were reviewed and assessed by NTP 

staff, so all comments had been resolved or were 

otherwise addressed during the preparation of this 

Technical Report. 

GENETIC TOXICOLOGY 

The genetic toxicity of cobalt sulfate heptahydrate was 

assessed by testing the ability of the chemical to 

induce mutations in various strains of Salmonella 

typhimurium. The protocols for these studies and the 

results are given in Appendix E. 

The genetic toxicity studies of cobalt sulfate heptahydrate are part of a larger effort by the NTP to 

develop a database that would permit the evaluation of 

carcinogenicity in experimental animals from the 

molecular structure and the effects of the chemical in 

short-term in vitro and in vivo genetic toxicity tests. 

These genetic toxicity tests were originally developed 

to study mechanisms of chemical-induced DNA 

damage and to predict carcinogenicity in animals, 

based on the electrophilicity theory of chemical 

mutagenesis and the somatic mutation theory of cancer 

(Miller and Miller, 1977; Straus, 1981; Crawford, 

1985). 

There is a strong correlation between a chemical’s 

potential electrophilicity (structural alert to DNA 

reactivity), mutagenicity in Salmonella, and carcinogenicity in rodents. The combination of electrophilicity and Salmonella mutagenicity is highly correlated with the induction of carcinogenicity in rats 

and mice and/or at multiple tissue sites (Ashby and 

Tennant, 1991). Other in vitro genetic toxicity tests 

correlate less well with rodent carcinogenicity 

(Tennant et al., 1987; Zeiger et al., 1990), although 

these other tests can provide information on the types 

of DNA and chromosome effects that can be induced 

by the chemical being investigated. Data from NTP 

studies show that a positive response in Salmonella is 

the most predictive in vitro test for rodent carcinogenicity (89% of the Salmonella mutagens are rodent 

carcinogens), and that there is no complementarity 

among the in vitro genetic toxicity tests. That is, no 

battery of tests that included the Salmonella test 

improved the predictivity of the Salmonella test alone. 
28 Cobalt Sulfate Heptahydrate, NTP TR 471 
 29 

RESULTS
 

RATS Body Weights and Clinical Findings 

Survival Mean body weights of exposed male and female rats 

Estimates of 2-year survival probabilities for male and were similar to those of the chamber controls throughfemale rats are shown in Table 2 and in the Kaplan- out the study (Figure 2 and Tables 3 and 4). Irregular 

Meier survival curves (Figure 1). Survival of exposed breathing was observed more frequently in female rats 3 males and females was similar to that of the chamber exposed to 3.0 mg/m than in the chamber controls or

controls. other exposed groups. 

TABLE 2 

Survival of Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Male 

Animals initially in study 50 50 50 50 

Moribund 30 34 26 34 

Natural deaths 3 1 3 1 

Animals surviving to study termination 

Percent probability of survival at end of studya 

Mean survival (days)b 

17 

34 

648 

15 

30 

655 

21 

42 

663 

15 

30 

643 

Survival analysisc Pù0.723 Pù1.000N Pù0.292N Pù0.876 

Female 

Animals initially in study 50 50 50 50 

Moribund 19 20 20 17 

Natural deaths 3 4 4 3 

Pregnantd 0 1 0 0 

Animals surviving to study termination 28 25 26 30 

Percent probability of survival at end of study 56 51 52 60 

Mean survival (days) 699 677 691 684 

Survival analysis Pù0.642N Pù0.583 Pù0.756 Pù0.959N 

a Kaplan-Meier determinations b	 Mean of all deaths (uncensored, censored, and terminal sacrifice) 

The result of the life table trend test (Tarone, 1975) is in the chamber control column, and the results of the life table pairwise 

comparisons (Cox, 1972) with the chamber controls are in the exposed group columns. A negative trend or lower mortality in an 

exposure group is indicated by N. d	 Censored from survival analyses 

c 
30 Cobalt Sulfate Heptahydrate, NTP TR 471 

FIGURE 1
 

Kaplan-Meier Survival Curves for Male and Female Rats 


Exposed to Cobalt Sulfate Heptahydrate by Inhalation for 2 Years
 
Cobalt Sulfate Heptahydrate, NTP TR 471 31 

FIGURE 2
 

Growth Curves for Male and Female Rats
 

Exposed to Cobalt Sulfate Heptahydrate by Inhalation for 2 Years
 
32 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE 3 

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

of Cobalt Sulfate Heptahydrate 

Weeks Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

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

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

1 124 50 124 100 50 122 99 50 122 99 50 

2 149 50 148 100 50 145 97 50 146 98 50 

3 177 50 177 100 50 172 97 50 172 97 50 

4 202 50 202 100 50 200 99 50 196 97 50 

5 225 50 226 101 50 221 99 50 219 97 50 

6 247 50 247 100 50 244 99 50 240 97 50 

7 264 50 263 99 50 260 98 50 255 97 50 

8 277 50 278 100 50 279 101 50 273 98 50 

9 291 50 292 100 50 291 100 50 284 98 50 

10 303 50 304 100 50 306 101 50 299 98 50 

11 315 50 317 101 50 320 102 50 310 98 50 

12 324 50 328 101 50 329 102 50 319 99 50 

13 332 50 336 101 50 336 101 50 327 98 50 

16 358 50 364 102 50 363 101 50 352 98 50 

20 387 50 390 101 50 390 101 50 377 98 50 

24 409 50 413 101 50 412 101 50 398 97 50 

28 425 50 428 101 50 429 101 50 413 97 50 

32 437 50 439 100 50 440 101 50 424 97 50 

36 450 50 452 101 50 455 101 50 438 97 50 

40 458 50 462 101 50 463 101 49 448 98 50 

44 464 50 468 101 50 470 101 49 454 98 50 

48 473 50 476 101 50 482 102 49 461 98 50 

52 479 50 483 101 50 488 102 49 468 98 50 

56 483 50 487 101 50 492 102 49 472 98 50 

60 489 50 493 101 49 498 102 49 474 97 49 

64 491 49 497 101 49 499 102 48 479 98 49 

68 497 48 498 100 49 503 101 47 485 98 47 

72 495 48 505 102 46 506 102 47 491 99 47 

76 493 48 506 103 46 506 103 45 491 100 46 

80 498 43 505 102 45 512 103 43 493 99 43 

84 493 40 505 103 42 507 103 42 485 98 38 

88 490 32 501 102 39 509 104 41 476 97 33 

92 495 27 497 100 33 512 103 36 471 95 29 

94 483 24 486 101 28 502 104 34 468 97 25 

96 488 23 485 99 26 506 104 31 463 95 24 

98 482 21 472 98 23 503 104 27 462 96 21 

100 487 18 467 96 19 498 102 26 462 95 18 

102 481 18 462 96 17 486 101 24 460 96 16 

104 476 18 454 95 16 481 101 22 459 96 15 

Mean for weeks 

1-13 248 249 100 248 100 243 98 

14-52 434 438 101 439 101 423 97 

53-104 489 489 100 501 102 474 97 
Cobalt Sulfate Heptahydrate, NTP TR 471 33 

TABLE 4 

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

of Cobalt Sulfate Heptahydrate 

Weeks Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

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

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

1 100 50 100 100 50 99 98 50 99 98 50 

2 112 50 112 100 50 110 99 50 111 99 49 

3 126 50 125 99 50 123 98 50 123 98 49 

4 138 50 136 99 50 134 98 50 134 98 49 

5 147 50 145 99 50 143 97 50 144 98 49 

6 156 50 154 99 50 151 97 50 154 99 49 

7 162 50 158 98 50 156 97 50 158 98 49 

8 166 50 164 99 50 163 98 50 166 100 49 

9 171 50 168 98 50 168 98 50 170 99 49 

10 176 50 173 98 50 173 98 50 176 100 49 

11 179 50 178 99 50 179 100 50 181 101 49 

12 184 50 181 99 49 181 99 50 184 100 49 

13 186 50 184 99 49 186 100 50 187 100 49 

16 195 50 193 99 49 193 99 50 196 101 49 

20 207 50 204 99 49 205 99 50 208 101 49 

24 219 50 216 99 49 215 99 50 219 100 49 

28 224 50 223 99 49 222 99 50 225 100 49 

32 230 50 230 100 49 231 100 50 233 101 49 

36 238 50 240 101 49 241 101 50 244 103 49 

40 247 50 246 100 49 249 101 50 251 102 49 

44 255 50 254 100 49 260 102 50 259 101 49 

48 267 50 269 101 49 273 102 50 273 102 49 

52 276 50 275 100 49 279 101 49 282 102 49 

56 286 49 288 101 48 290 102 49 293 103 49 

60 293 49 294 100 48 299 102 49 297 101 48 

64 302 49 304 101 47 307 102 49 306 101 48 

68 308 49 310 100 47 313 102 49 312 101 48 

72 314 49 314 100 45 316 101 49 318 102 47 

76 321 49 320 100 44 322 100 48 323 101 47 

80 323 49 328 102 42 326 101 46 325 101 47 

84 324 47 331 102 42 329 101 45 329 102 47 

88 326 44 331 102 41 331 102 42 327 101 46 

92 334 42 339 102 38 337 101 40 335 100 43 

94 327 41 333 102 38 331 101 40 328 100 41 

96 330 41 336 102 37 334 101 38 328 99 39 

98 333 39 336 101 34 334 100 38 333 100 37 

100 328 38 335 102 33 333 102 36 333 102 33 

102 328 36 336 103 31 334 102 34 333 102 33 

104 326 32 337 103 27 331 102 32 334 102 31 

Mean for weeks 

1-13 154 152 99 151 98 153 99 

14-52 236 235 100 237 100 239 101 

53-104 319 323 101 323 101 322 101 
34 Cobalt Sulfate Heptahydrate, NTP TR 471 

Pathology and Statistical Analyses 

This section describes the statistically significant or 

biologically noteworthy changes in the incidences of 

neoplasms and/or nonneoplastic lesions of the lung, 

adrenal medulla, nose, and larynx. Summaries of the 

incidences of neoplasms and nonneoplastic lesions, 

individual animal tumor diagnoses, statistical analyses 

of primary neoplasms that occurred with an incidence 

of at least 5% in at least one animal group, and 

historical incidences for the neoplasms mentioned in 

this section are presented in Appendix A for male rats 

and Appendix B for female rats. 

Lung: In all exposed groups of male and female rats, 

the incidences of proteinosis, alveolar epithelial 

metaplasia, granulomatous alveolar inflammation, and 

interstitial fibrosis were significantly greater than those 

in the chamber controls (Tables 5, A5, and B5). In 

general, these lung lesions increased in incidence and 

severity with increased exposure to cobalt sulfate 

heptahydrate. The incidence of squamous metaplasia 3 in 1.0 mg/m females was significantly greater than in

the chamber control group. Multifocally, throughout 

the lungs, pulmonary architecture was distorted by a 

combination of inflammatory cells, fibrosis, and 

epithelial metaplasia. Lesions tended to be subpleural, 

peripheral, and/or along larger blood vessels and 

airways. Granulomatous inflammation was characterized by accumulations of alveolar macrophages 

with foamy cytoplasm, occasional multinucleated giant 

cells and cholesterol clefts, cell debris and few neutrophils. In these areas, the alveolar interstitium and 

occasionally the overlying pleura were variably 

thickened by dense fibrous connective tissue which 

often effaced alveoli (Plates 1 and 2). Although a 

diffuse change, aggregates of homogeneous to granular eosinophilic material within alveolar lumens 

(alveolar proteinosis) were often pronounced within 

the areas of chronic inflammation. Metaplasia of the 

alveolar epithelium in alveoli within and at the 

periphery of foci of inflammation was characterized by 

replacement of normal Type I epithelial cells with 

plump cuboidal or ciliated columnar epithelial cells. 

The incidences of alveolar epithelial hyperplasia in all 

groups of exposed males and in females exposed to 

3.0 mg/m3

 and atypical alveolar epithelial hyperplasia 

3 in 3.0 mg/m females were significantly greater than

those in the chamber control groups. 

The combined incidence of alveolar/bronchiolar 

neoplasms (adenoma and/or carcinoma) was significantly greater in 3.0 mg/m3

 males than that in the 

chamber controls and exceeded the historical control 

range (Tables 5 and A3). In females exposed to 1.0 or 3 3.0 mg/m , the incidences of alveolar/bronchiolar

neoplasms were significantly greater than those in the 

chamber control group and exceeded the historical 

control ranges (Tables 5, B3, and B4a). Although the 

incidences of alveolar/bronchiolar adenoma in 

3 3.0 mg/m males and alveolar/bronchiolar carcinoma

3 in 1.0 mg/m males were not significantly increased,

they exceeded the historical control ranges for inhalation studies (Tables 5, A3, and A4a). 

The spectrum of alveolar/bronchiolar neoplasms and 

nonneoplastic proliferative lesions observed within the 

lungs of exposed rats was broad. While many of these 

lesions were highly cellular and morphologically 

similar to those observed spontaneously, others were 

predominantly fibrotic, squamous, or mixtures of 

alveolar/bronchiolar epithelium and squamous or 

fibrous components. Hyperplasia generally represented an increase in numbers of epithelial cells along 

alveolar walls with maintenance of normal alveolar 

architecture (Plates 3 and 4). Multiple hyperplastic 

lesions were often observed in animals receiving 

higher concentrations of cobalt sulfate heptahydrate. 

The benign neoplasms typical of those observed 

spontaneously were generally distinct masses that often 

compressed surrounding tissue (Plates 5 and 6). 

Component epithelial cells were often arranged in 

acinar and/or irregular papillary structures and occasionally in a solid cellular pattern. These epithelial 

cells were typically uniform and similar to hyperplastic 

counterparts. Malignant alveolar/bronchiolar neoplasms had similar cellular patterns but were generally 

larger and had one or more of the following histologic 

features: heterogeneous growth pattern, cellular 

pleomorphism and/or atypia, and local invasion or 

metastasis (Plate 7). 

In addition to these more typical proliferative lesions, 

there were “fibroproliferative” lesions ranging from 

less than 1 mm to greater than 1 cm in diameter. 

Generally, these lesions had a rounded outline and 

a central fibrous core containing dispersed glandular 

(alveolar) structures lined by uniformly cuboidal 

epithelial cells. Aggregates of mostly necrotic 
 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 35 

TABLE 5 

Incidences of Neoplasms and Nonneoplastic Lesions of the Lung in Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Male 

Number Examined Microscopically 

Alveolar Epithelium, Hyperplasiaa 50 9 (1.8)b 

50 

20* (2.0) 

48 

20* (2.1) 

50 

23**(2.0) 

Alveolar Epithelium, Hyperplasia, Atypical 0 1 (2.0) 2 (3.0) 2 (4.0) 

Metaplasia, Squamous 0 1 (1.0) 4 (2.0) 2 (3.0) 

Alveolar Epithelium, Metaplasia 0 50**(1.9) 48**(3.1) 49**(3.7) 

Inflammation, Granulomatous 2 (1.0) 50**(1.9) 48**(3.1) 50**(3.7) 

Interstitium, Fibrosis 1 (1.0) 50**(1.9) 48**(3.1) 49**(3.7) 

Proteinosis 0 16**(1.4) 40**(2.3) 47**(3.4) 

Cyst 0 0 0 1 (4.0) 

Alveolar/bronchiolar Adenomac 

Overall rated 

Adjusted ratee 

Terminal ratef 

1/50 (2%) 

2.3% 

0/17 (0%) 

4/50 (8%) 

17.7% 

2/15 (13%) 

1/48 (2%) 

2.4% 

0/21 (0%) 

6/50 (12%) 

28.4% 

2/15 (13%) 

First incidence (days) 

Logistic regression testg 

568 

Pù0.051 

589 

Pù0.179 

611 

Pù0.753 

638 

Pù0.055 

Alveolar/bronchiolar Carcinomah 

Overall rate 0/50 (0%) 0/50 (0%) 3/48 (6%) 1/50 (2%) 

Adjusted rate 0.0% 0.0% 11.3% 6.7% 

Terminal rate 

First incidence (days) 

Logistic regression test 

0/17 (0%) 

—i 

Pù0.360 

0/15 (0%) 

— 

— 

1/21 (5%) 

652 

Pù0.136 

1/15 (7%) 

734 (T) 

Pù0.475 

Alveolar/bronchiolar Adenoma or Carcinomaj 

Overall rate 1/50 (2%) 4/50 (8%) 4/48 (8%) 7/50 (14%) 

Adjusted rate 2.3% 17.7% 13.4% 33.9% 

Terminal rate 0/17 (0%) 2/15 (13%) 1/21 (5%) 3/15 (20%) 

First incidence (days) 

Logistic regression test 

568 

Pù0.032 

589 

Pù0.179 

611 

Pù0.163 

638 

Pù0.029 

Female 

Number Examined Microscopically 50 49 50 50 

Alveolar Epithelium, Hyperplasia 15 (1.4) 7 (1.6) 20 (1.8) 33**(2.0) 

Alveolar Epithelium, Hyperplasia, Atypical 0 0 3 (3.7) 5* (3.2) 

Metaplasia, Squamous 0 1 (2.0) 8**(2.3) 3 (1.7) 

Alveolar Epithelium, Metaplasia 2 (1.0) 47**(2.0) 50**(3.6) 49**(3.9) 

Inflammation, Granulomatous 9 (1.0) 47**(2.0) 50**(3.6) 49**(3.9) 

Interstitium, Fibrosis 7 (1.0) 47**(2.0) 50**(3.6) 49**(3.9) 

Proteinosis 0 36**(1.2) 49**(2.8) 49**(3.9) 

Cyst 0 0 1 (4.0) 0 

Alveolar/bronchiolar Adenomak 

Overall rate 0/50 (0%) 1/49 (2%) 10/50 (20%) 9/50 (18%) 

Adjusted rate 0.0% 3.4% 36.4% 30.0% 

Terminal rate 0/28 (0%) 0/25 (0%) 9/26 (35%) 9/30 (30%) 

First incidence (days) 

Logistic regression test 

— 

Pù0.001 

714 

Pù0.480 

692 

P<0.001 

735 (T) 

Pù0.003 
36 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE 5 

Incidences of Neoplasms and Nonneoplastic Lesions of the Lung in Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Female (continued) 

Alveolar/bronchiolar Carcinomal 

Overall rate 0/50 (0%) 2/49 (4%) 6/50 (12%) 6/50 (12%) 

Adjusted rate 0.0% 8.0% 20.2% 17.5% 

Terminal rate 0/28 (0%) 2/25 (8%) 4/26 (15%) 4/30 (13%) 

First incidence (days) — 735 (T) 694 610 

Logistic regression test Pù0.023 Pù0.213 Pù0.015 Pù0.017 

Alveolar/bronchiolar Adenoma or Carcinomam 

Overall rate 0/50 (0%) 3/49 (6%) 15/50 (30%) 15/50 (30%) 

Adjusted rate 0.0% 11.2% 50.6% 46.1% 

Terminal rate 0/28 (0%) 2/25 (8%) 12/26 (46%) 13/30 (43%) 

First incidence (days) — 714 692 610 

Logistic regression test P<0.001 Pù0.096 P<0.001 P<0.001 

Squamous Cell Carcinoma 

Overall rate 0/50 (0%) 0/49 (0%) 1/50 (2%) 1/50 (2%) 

Alveolar/bronchiolar Adenoma, Alveolar/bronchiolar Carcinoma, or Squamous Cell Carcinoma 

Overall rate 0/50 (0%) 3/49 (6%) 16/50 (32%) 16/50 (32%) 

Adjusted rate 0.0% 11.2% 54.1% 49.2% 

Terminal rate 0/28 (0%) 2/25 (8%) 13/26 (50%) 14/30 (47%) 

First incidence (days) — 714 692 610 

Logistic regression test P<0.001 Pù0.096 P<0.001 P<0.001 

* Significantly different (P#0.05) from the chamber control by the logistic regression test 

** P#0.01 

(T)Terminal sacrifice 

a Number of animals with lesion 

b Average severity grade of lesions in affected animals: 1 ù minimal, 2 ù mild, 3 ù moderate, 4 ù marked c Historical incidence for 2-year inhalation studies with chamber controls (mean ± standard deviation): 17/654 (2.6% ± 3.6%); range 

0%-10% 

d	 Number of animals with neoplasm per number of animals with lung examined microscopically e Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality f	 Observed incidence in animals surviving until the end of the study g	 In the chamber control column are the P values associated with the trend test. In the exposed group columns are the P values 

corresponding to the pairwise comparisons between the chamber controls and that exposed group. The logistic regression test regards 

lesions in animals dying prior to terminal kill as nonfatal. h	 Historical incidence: 6/654 (0.9% ± 1.0%); range 0%-2% i	 Not applicable; no neoplasms in animal group j Historical incidence: 23/654 (3.5% ± 3.7%); range 0%-10% k	 Historical incidence: 7/650 (1.1% ± 1.6%); range 0%-4% l	 Historical incidence: 0/650 m Historical incidence: 7/650 (1.1% ± 1.6%); range 0%-4% 
 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 37 

inflammatory cells were also present in adjacent 

alveoli and often within the glandular structures. 

Peripherally, the fibroproliferative lesions had one to 

several layers of epithelium which coursed along and 

often extended into adjacent alveoli, frequently 

forming papillary projections (Plates 8, 9, and 10). 

These epithelial cells were often slightly pleomorphic 

with occasional mitotic figures. The smallest of these 

lesions were usually observed adjacent to areas of 

chronic inflammation. Small lesions with modest 

amounts of peripheral epithelial proliferation were 

diagnosed as atypical hyperplasia, while larger lesions 

with florid epithelial proliferation, marked cellular 

pleomorphism, and/or local invasion were diagnosed 

as alveolar/bronchiolar carcinomas (Plate 11). 

While squamous epithelium is not normally observed 

within the lung, squamous metaplasia of alveolar/ 

bronchiolar epithelium is a relatively common response to pulmonary injury and occurred in a number 

of rats in this study (Table 5). Squamous metaplasia 

was a minor change consisting of a small cluster of 

alveoli in which the normal epithelium was replaced 

by multiple layers of flattened squamous epithelial 

cells (Plate 12) that occasionally formed keratin. One 

3 3 3.0 mg/m male and one 1.0 mg/m female had a large 

cystic squamous lesion rimmed by a variably thick (a 

few to many cell layers) band of viable squamous 

epithelium with a large central core of keratin 

(Plate 13). These were diagnosed as cysts. In one 

3 3 1.0 mg/m and one 3.0 mg/m female, proliferative 

squamous lesions had cystic areas but also more solid 

areas of pleomorphic cells and invasion into the 

adjacent lung; these lesions were considered to be 

squamous cell carcinomas (Plate 14). In general, 

diagnoses of squamous lesions were made only when 

the lesion composition was almost entirely squamous 

epithelium. However, squamous metaplasia/ differentiation was a variable component of other 

alveolar/bronchiolar proliferative lesions (Plate 15), 

including the fibroproliferative lesions, and was clearly a part of the spectrum of lesions resulting from 

exposure to cobalt sulfate heptahydrate. 

Adrenal Medulla: The incidence of benign pheo­ 3 chromocytoma in 3.0 mg/m females was significantly 

greater than that in the chamber controls and exceeded 

the historical range for inhalation studies (Tables 6, 

B3, and B4b). The incidences of benign, complex, 

or malignant pheochromocytoma (combined) in 

1.0 mg/m3 males and in 3.0 mg/m3 females were 

significantly greater than those in the chamber controls 

and exceeded the historical control ranges (Tables 6, 

A3, A4b, B3, and B4b). 

The incidences of bilateral pheochromocytoma in 

exposed males slightly exceeded that in the chamber 

control group. The incidence of hyperplasia was not 

significantly increased in exposed males or females. 

Focal hyperplasia and pheochromocytoma are considered to constitute a morphological continuum in the 

adrenal medulla. Focal hyperplasia consisted of 

irregular, small foci of small- to normal-sized medullary cells arranged in packets or solid clusters 

slightly larger than normal; compression of surrounding parenchyma was minimal or absent. Benign 

pheochromocytomas were well-delineated masses often 

with altered architecture and variable compression of 

surrounding parenchyma. Neoplastic cells were 

arranged in variably sized aggregates, clusters, and/or 

variably thick trabecular cords. Larger neoplasms 

usually exhibited greater cellular pleomorphism and 

atypia than smaller neoplasms. Malignant pheochromocytomas were identified when there was 

invasion of or beyond the adrenal capsule or when 

distant metastases were observed. Although a very 

common spontaneous neoplasm in male F344/N rats, 

pheochromocytomas have a lower spontaneous occurrence in females. In this study, the incidence of 

pheochromocytoma in 3.0 mg/m3 females was considered related to the administration of cobalt sulfate 

heptahydrate. The marginally increased incidence of 

pheochromocytoma in males was considered an 

uncertain finding because it occurred only in the 

1.0 mg/m3 group and was not supported by increased 

incidence or severity of hyperplasia. 
38 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE 6 

Incidences of Neoplasms and Nonneoplastic Lesions of the Adrenal Medulla in Rats 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Male 

Number Examined Microscopically 50 50 49 50 

Hyperplasiaa 34 (2.0)b 23* (2.5) 29 (2.1) 30 (2.1) 

Benign Bilateral Pheochromocytoma 

Overall rate 1/50 (2%) 4/50 (8%) 6/49 (12%) 5/50 (10%) 

Benign Pheochromocytoma (includes benign bilateral pheochromocytoma)

c 

Overall rated 14/50 (28%) 19/50 (38%) 23/49 (47%) 20/50 (40%) e Adjusted rate 51.0% 70.0% 71.9% 71.4% 

Terminal ratef 6/17 (35%) 8/15 (53%) 13/21 (62%) 8/15 (53%) 

First incidence (days) 534 541 526 526 

g Logistic regression test	 Pù0.172 Pù0.226 Pù0.069 Pù0.126 

Benign, Complex, or Malignant Pheochromocytoma (includes benign bilateral pheochromocytoma)

h 

Overall rate 15/50 (30%) 19/50 (38%) 25/49 (51%) 20/50 (40%) 

Adjusted rate 52.1% 70.0% 74.1% 71.4% 

Terminal rate 6/17 (35%) 8/15 (53%) 13/21 (62%) 8/15 (53%) 

First incidence (days) 534 541 526 526 

Logistic regression test Pù0.218 Pù0.295 Pù0.045 Pù0.180 

Female 

Number Examined Microscopically 48 49 50 48 

Hyperplasia 8 (1.6) 7 (2.3) 11 (2.1) 13 (2.0) 

Benign Pheochromocytomai 

Overall rate 2/48 (4%) 1/49 (2%) 3/50 (6%) 8/48 (17%) 

Adjusted rate 5.1% 3.1% 9.3% 26.4% 

Terminal rate 0/27 (0%) 0/25 (0%) 1/26 (4%) 7/29 (24%) 

First incidence (days) 666 702 694 709 

Logistic regression test Pù0.004 Pù0.498N Pù0.512 Pù0.043 

Benign, Complex, or Malignant Pheochromocytomaj 

Overall rate 2/48 (4%) 1/49 (2%) 4/50 (8%) 10/48 (21%) 

Adjusted rate 5.1% 3.1% 11.7% 31.5% 

Terminal rate 0/27 (0%) 0/25 (0%) 1/26 (4%) 8/29 (28%) 

First incidence (days) 666 702 685 663 

Logistic regression test P<0.001 Pù0.498N Pù0.323 Pù0.014 

* Significantly different (P#0.05) from the chamber control by the logistic regression test a Number of animals with lesion 

b Average severity grade of lesions in affected animals: 1 ù minimal, 2 ù mild, 3 ù moderate, 4 ù marked c Historical incidence for 2-year inhalation studies with chamber controls (mean ± standard deviation): 163/623 (26.2% ± 13.2%); range 

0%-50% 

d	 Number of animals with neoplasm per number of animals with adrenal medulla examined microscopically e Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality f	 Observed incidence in animals surviving until the end of the study g	 In the chamber control column are the P values associated with the trend test. In the exposed group columns are the P values 

corresponding to the pairwise comparisons between the chamber controls and that exposed group. The logistic regression test regards 

lesions in animals dying prior to terminal kill as nonfatal. A lower incidence in an exposure group is indicated by N. h	 Historical incidence: 176/623 (28.3% ± 12.0%); range 8%-50% i	 Historical incidence: 35/608 (5.8% ± 4.9%); range 0%-14% j Historical incidence: 39/608 (6.4% ± 4.4%); range 2%-14% 
 

Cobalt Sulfate Heptahydrate, NTP TR 471 39 

Nose: The incidences of hyperplasia of the lateral wall squamous metaplasia were minimal to mild, unilateral 

of the nose and atrophy of the olfactory epithelium in or bilateral, and involved the transitional epithelium 

all exposed groups of males and females were along the walls and turbinates of the anterior nasal 

significantly greater than those in the chamber con- passage. Hyperplasia was characterized by an increase 

trols, and the severities of these lesions increased with in thickness of the epithelium from the normal one to 

increasing exposure concentration (Tables 7, A5, and two layers to two or more layers, while squamous 

B5). The incidences of squamous metaplasia of the metaplasia represented areas where the normal 

lateral wall of the nose and metaplasia of the olfactory transitional epithelium was replaced by multiple layers 

epithelium in 3.0 mg/m3 males and females were of flattened epithelial cells. More posterior in the 

significantly greater than those in the chamber nose, along the dorsal meatus, atrophy of the olfactory 

controls. epithelium was characterized by loss of cell layers and 

disorganization of remaining epithelium, and in some 

Although the incidence and severity of nasal lesions instances, increased prominence of sensory cell nuclei. 

increased with increased exposure to cobalt sulfate Metaplasia was characterized by replacement of 

heptahydrate, they involved limited portions of nasal olfactory epithelium with respiratory-type ciliated 

epithelium and none were severe. Hyperplasia and columnar epithelium. 

TABLE 7 

Incidences of Nonneoplastic Lesions of the Nose and Larynx in Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Male 

Nosea 50 50 49 50 

Lateral Wall, Hyperplasiab 2 (1.5)c 14**(1.4) 21**(1.5) 20**(1.6) 

Lateral Wall, Metaplasia, Squamous 1 (1.0) 3 (1.3) 5 (1.4) 8* (2.0) 

Olfactory Epithelium, Atrophy 8 (1.1) 24**(1.4) 42**(1.5) 48**(2.5) 

Olfactory Epithelium, Metaplasia 5 (1.2) 1 (3.0) 5 (1.8) 30**(1.9) 

Larynx 50 49 48 50 

Epiglottis, Metaplasia, Squamous 0 10**(1.3) 37**(1.8) 50**(2.8) 

Female 

Nose 50 49 50 50 

Lateral Wall, Hyperplasia 1 (1.0) 8* (1.3) 26**(1.4) 38**(1.7) 

Lateral Wall, Metaplasia, Squamous 1 (1.0) 1 (3.0) 4 (1.3) 10**(1.4) 

Olfactory Epithelium, Atrophy 5 (1.4) 29**(1.2) 46**(1.6) 47**(2.9) 

Olfactory Epithelium, Metaplasia 2 (2.0) 2 (1.5) 3 (1.7) 40**(2.3) 

Larynx 50 49 50 50 

Epiglottis, Metaplasia, Squamous 1 (1.0) 22**(1.1) 39**(1.4) 48**(2.6) 

* Significantly different (P#0.05) from the chamber control by the logistic regression test 

** P#0.01 a Number of animals with tissue examined microscopically b Number of animals with lesion 

Average severity grade of lesions in affected animals: 1 ù minimal, 2 ù mild, 3 ù moderate, 4 ù marked c 
40 

Larynx: The incidences of squamous metaplasia of the 

epiglottis in all exposed groups of males and females 

were significantly greater than those in the chamber 

controls, and the severity of this lesion increased with 

increasing exposure concentration (Tables 7, A5, and 

B5). Squamous metaplasia was limited to the base 

Cobalt Sulfate Heptahydrate, NTP TR 471 

of the epiglottis and was not a severe lesion in exposed 

rats. It was characterized by replacement of the 

ciliated respiratory epithelium by one or more layers 

of flattened epithelial cells overlying a basal layer 

of cuboidal cells. Keratinization was sometimes 

observed. 
Cobalt Sulfate Heptahydrate, NTP TR 471 41 

MICE 

Survival 

Estimates of 2-year survival probabilities for male and were less than those of the chamber controls from 

female mice are shown in Table 8 and in the Kaplan- week 96 until the end of the study. The mean body 

Meier survival curves (Figure 3). Survival of exposed weights of all exposed female mice were generally 

males and females was similar to that of the chamber greater than those of the chamber controls from week 

controls. 20 until the end of the study. Irregular breathing was 

observed slightly more frequently in female mice 

3 Body Weights and Clinical Findings exposed to 1.0 mg/m than in the chamber controls or

Mean body weights are given in Figure 4 and Tables 9 other exposed groups. 

3 and 10. Mean body weights of 3.0 mg/m male mice

TABLE 8 

Survival of Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Male 

Animals initially in study	 50 50 50 50 

Accidental deathsa 1 0 0 1 

Moribund 19 16 17 23 

Natural deaths 8 3 9 6 

Animals surviving to study termination 22 31 24 20 

Percent probability of survival at end of studyb 46 62 48 42 c Mean survival (days)	 662 695 670 643 

d Survival analysis	 Pù0.104 Pù0.088N Pù0.861N Pù0.577 

Female 

Animals initially in study	 50 50 50 50 

Moribund 11 10 13 16 

Natural deaths 5 3 5 6 

Animals surviving to study termination 34 37 32 28 

Percent probability of survival at end of study 68 74 64 56 

Mean survival (days) 694 713 685 680 

Survival analysis	 Pù0.102 Pù0.529N Pù0.855 Pù0.327 

a Censored from survival analyses b	 Kaplan-Meier determinations 

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

(Cox, 1972) with the chamber controls are in the exposed group columns. A lower mortality in an exposure group is indicated by N. 

c 
42 Cobalt Sulfate Heptahydrate, NTP TR 471 

FIGURE 3
 

Kaplan-Meier Survival Curves for Male and Female Mice 


Exposed to Cobalt Sulfate Heptahydrate by Inhalation for 2 Years
 
Cobalt Sulfate Heptahydrate, NTP TR 471 43 

FIGURE 4
 

Growth Curves for Male and Female Mice Exposed 


to Cobalt Sulfate Heptahydrate by Inhalation for 2 Years
 
44 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE 9 

Mean Body Weights and Survival of Male Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Weeks Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

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

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

1 23.7 50 23.5 99 50 23.4 99 50 23.4 99 50 

2 25.8 50 25.5 99 50 25.4 98 50 25.7 100 50 

3 27.0 50 26.9 100 50 26.6 99 50 27.1 100 50 

4 28.0 50 27.9 100 50 27.8 99 50 28.3 101 50 

5 28.8 50 28.8 100 50 28.7 100 50 29.1 101 50 

6 29.3 50 29.6 101 50 29.2 100 50 29.6 101 50 

7 29.8 50 30.0 101 50 30.0 101 50 30.3 102 50 

8 30.5 50 30.2 99 50 30.4 100 50 30.9 101 50 

9 31.0 50 30.8 99 50 30.8 99 50 31.4 101 50 

10 31.6 50 31.3 99 50 31.4 99 50 31.9 101 50 

11 32.0 50 31.9 100 50 31.8 99 50 32.7 102 50 

12 32.2 50 32.2 100 50 32.2 100 50 33.2 103 50 

13 33.1 50 33.0 100 50 32.7 99 50 33.9 102 50 

16 34.7 50 34.4 99 50 34.5 99 50 35.3 102 50 

20 36.1 50 36.4 101 50 36.6 101 50 37.2 103 50 

24 38.3 50 38.4 100 50 38.3 100 50 39.6 103 50 

28 40.3 50 40.3 100 50 40.0 99 50 41.4 103 50 

32 41.8 50 41.7 100 50 41.4 99 50 43.1 103 50 

36 43.1 50 43.1 100 50 42.5 99 50 44.3 103 50 

40 44.3 50 44.1 100 50 43.9 99 50 45.9 104 50 

44 44.6 50 44.9 101 50 44.7 100 50 45.9 103 50 

48 45.5 50 45.6 100 50 45.0 99 50 46.5 102 50 

52 45.8 50 45.4 99 50 44.8 98 50 46.6 102 50 

56 45.9 50 46.2 101 50 45.5 99 50 46.5 101 50 

60 45.8 50 46.8 102 50 45.6 100 50 47.1 103 48 

64 45.8 50 47.4 104 50 46.6 102 49 47.8 104 48 

68 47.1 48 47.8 102 49 47.2 100 49 47.2 100 48 

72 46.4 46 48.3 104 48 46.8 101 48 46.5 100 48 

76 46.9 45 48.5 103 48 47.2 101 45 46.5 99 46 

80 46.6 42 48.3 104 47 46.9 101 43 46.1 99 40 

84 45.6 39 47.2 104 47 46.7 102 40 45.7 100 34 

88 45.5 37 46.1 101 45 46.0 101 38 43.4 95 32 

92 44.1 36 45.6 103 42 44.4 101 38 42.8 97 28 

94 44.2 32 45.7 103 41 44.4 101 35 42.9 97 26 

96 44.3 30 45.2 102 39 43.5 98 33 41.5 94 24 

98 43.0 29 44.7 104 37 42.0 98 32 40.7 95 22 

100 43.3 25 44.7 103 34 42.0 97 29 40.3 93 21 

102 42.7 25 43.7 102 34 41.5 97 26 39.5 93 21 

104 42.6 23 43.2 101 32 41.6 98 24 38.5 90 20 

Mean for weeks 

1-13 29.4 29.4 100 29.3 100 29.8 101 

14-52 41.5 41.4 100 41.2 99 42.6 103 

53-104 45.0 46.2 103 44.9 100 43.9 98 
Cobalt Sulfate Heptahydrate, NTP TR 471 45 

TABLE 10 

Mean Body Weights and Survival of Female Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Weeks Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

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

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

1 19.2 50 18.9 98 50 19.2 100 50 18.7 97 50 

2 21.2 50 21.1 100 50 21.1 100 50 20.8 98 50 

3 22.0 50 22.3 101 50 22.2 101 49 22.1 101 50 

4 23.2 50 23.5 101 50 23.4 101 49 23.5 101 50 

5 24.0 50 24.4 102 50 24.3 101 49 24.1 100 50 

6 24.8 50 25.3 102 50 25.1 101 49 25.1 101 50 

7 25.4 50 25.6 101 50 25.5 100 49 25.8 102 50 

8 26.0 50 25.9 100 50 26.5 102 49 25.7 99 50 

9 25.6 50 26.2 102 50 26.6 104 49 26.4 103 50 

10 26.2 50 26.9 103 50 26.9 103 49 26.8 102 50 

11 26.7 50 27.5 103 50 27.6 103 49 27.6 103 50 

12 26.9 50 27.9 104 50 28.0 104 49 28.2 105 50 

13 27.3 50 28.0 103 50 28.6 105 49 28.6 105 50 

16 29.1 50 30.1 103 50 30.9 106 49 30.5 105 50 

20 29.7 50 32.3 109 50 33.0 111 49 33.2 112 50 

24 33.1 50 35.3 107 50 36.3 110 49 35.8 108 50 

28 35.2 50 36.6 104 50 37.3 106 49 37.3 106 50 

32 36.3 50 38.4 106 50 39.7 109 49 39.8 110 50 

36 37.3 49 40.3 108 50 42.1 113 49 42.1 113 50 

40 39.4 49 42.4 108 50 43.3 110 49 43.4 110 50 

44 41.3 49 43.4 105 50 44.7 108 49 44.5 108 50 

48 42.5 49 45.0 106 50 46.0 108 49 46.2 109 50 

52 43.0 49 44.8 104 50 46.9 109 49 46.3 108 49 

56 44.2 49 46.5 105 50 47.9 108 49 47.1 107 49 

60 45.0 49 47.7 106 50 48.5 108 49 48.5 108 49 

64 46.7 49 48.6 104 50 49.9 107 49 50.1 107 47 

68 47.5 49 49.5 104 50 51.3 108 49 50.7 107 46 

72 48.2 49 51.6 107 49 51.0 106 48 52.3 109 45 

76 48.8 49 52.8 108 49 51.0 105 46 52.3 107 45 

80 48.9 48 52.6 108 49 51.4 105 45 53.0 108 43 

84 48.1 46 53.5 111 49 51.8 108 44 53.5 111 43 

88 48.1 44 52.1 108 47 50.0 104 44 51.9 108 41 

92 48.2 42 51.5 107 47 49.7 103 43 51.0 106 40 

94 47.9 40 51.2 107 46 48.8 102 41 50.8 106 39 

96 47.4 38 50.6 107 44 47.3 100 40 49.7 105 39 

98 47.9 37 50.8 106 41 47.8 100 36 48.7 102 37 

100 47.6 37 50.5 106 40 47.2 99 36 49.0 103 34 

102 46.7 35 49.8 107 40 47.0 101 35 48.5 104 33 

104 46.9 34 49.7 106 39 46.9 100 33 47.7 102 30 

Mean for weeks 

1-13 24.5 24.9 102 25.0 102 24.9 102 

14-52 36.7 38.9 106 40.0 109 39.9 109 

53-104 47.4 50.6 107 49.2 104 50.3 106 
46 Cobalt Sulfate Heptahydrate, NTP TR 471 

Pathology and Statistical Analyses 

This section describes the statistically significant or 

biologically noteworthy changes in the incidences of 

neoplasms and/or nonneoplastic lesions of the lung, 

nose, larynx, thyroid gland, and liver. Summaries of 

the incidences of neoplasms and nonneoplastic lesions, 

individual animal tumor diagnoses, statistical analyses 

of primary neoplasms that occurred with an incidence 

of at least 5% in at least one animal group, and 

historical incidences for the neoplasms mentioned in 

this section are presented in Appendix C for male mice 

and Appendix D for female mice. 

Lung: In all exposed groups of males and females, the 

incidences of cytoplasmic vacuolization of the bronchi 

were significantly greater than those in the chamber 

control groups (Tables 11, C5, and D5). The incidences of diffuse histiocytic cell infiltration in 

3.0 mg/m3

 males and of focal histiocytic cell infil­ 3 tration in 3.0 mg/m females were significantly greater

than those in the chamber controls. 

Cytoplasmic vacuolization of the bronchial epithelium 

was a minimal change of unknown biological significance confined to the epithelial cells lining the apex of 

the bronchial bifurcation. The affected cells were 

somewhat larger than normal with a diffusely clear to 

finely vacuolated cytoplasm. Histiocyte infiltration 

was characterized by one or more histiocytes with 

foamy cytoplasm within variable numbers of alveolar 

lumens. Focal infiltrate was a localized accumulation 

of histiocytes, while diffuse infiltrate was more widely 

scattered. The histiocyte infiltrate was very commonly 

seen in lungs with alveolar/bronchiolar neoplasms, and 

the increased incidences of infiltrate in the lungs of 

exposed animals were considered to reflect the higher 

incidences of lung neoplasms in these animals rather 

than a primary effect of cobalt sulfate heptahydrate 

exposure. 

The incidences of alveolar/bronchiolar neoplasms 

3 (adenoma and/or carcinoma) in 3.0 mg/m males and

females and the combined incidence of alveolar/ 

bronchiolar neoplasms in 1.0 mg/m3 females were 

significantly greater than those in the chamber control 

groups and generally exceeded the historical control 

ranges for inhalation studies (Tables 11, C3, C4a, D3, 

and D4a). In exposed males and females, the incidences of all lung neoplasms occurred with positive 

trends. 

Unlike in the rat, all the alveolar/bronchiolar proliferative lesions observed within the lungs of exposed 

mice were typical of those observed spontaneously. 

Hyperplasia generally represented an increase in 

numbers of epithelial cells along alveolar walls which 

retained normal alveolar structure. Adenomas generally were distinct masses that often compressed 

surrounding tissue (Plate 16). Component cells were 

arranged in acinar and/or irregular papillary structures 

and occasionally in a solid cellular pattern. These 

cells were typically uniform and similar to hyperplastic 

counterparts. Malignant alveolar/bronchiolar neoplasms had similar cellular patterns but were generally 

larger (Plate 17) and had one or more of the following: heterogeneous growth pattern, cellular pleomorphism, and/or atypia and local invasion or 

metastasis. 

Although similar in appearance to “spontaneous” lung 

neoplasms in chamber controls, alveolar/ bronchiolar 

neoplasms in mice exposed to cobalt sulfate 

heptahydrate had different molecular lesions in the 

Kras gene (Appendix I). Of the K-ras mutations 

detected at the second base of codon 12, a higher 

frequency (5/9, 55%) of G to T transversions was 

detected compared to concurrent (0/1) and historical 

control lung neoplasms (1/24, 4%). K-ras codon 61 

CTA or CGA mutations were not present in cobalt 

sulfate heptahydrate-induced lung neoplasms. 
Cobalt Sulfate Heptahydrate, NTP TR 471 47 

TABLE 11 

Incidences of Neoplasms and Nonneoplastic Lesions of the Lung in Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Male 

Number Examined Microscopically 

Infiltration Cellular, Diffuse, Histiocytea 50 1 (3.0)b 

50 

2 (3.0)

50 

4 (2.3) 

50 

10**(1.5) 

Infiltration Cellular, Focal, Histiocyte 10 (2.7) 5 (2.6) 8 (3.0) 17 (2.7) 

Bronchus, Cytoplasmic Vacuolization 0 18**(1.0) 34**(1.0) 38**(1.0) 

Alveolar Epithelium Hyperplasia 0 4 (2.3) 4 (1.8) 4 (2.3) 

Alveolar/bronchiolar Adenomac 

Overall rated 

Adjusted ratee 

Terminal ratef 

9/50 (18%) 

30.4% 

4/22 (18%) 

12/50 (24%) 

30.9% 

6/31 (19%) 

13/50 (26%) 

41.1% 

7/24 (29%) 

18/50 (36%) 

54.6% 

7/20 (35%) 

First incidence (days) 

Logistic regression testg 

600 

Pù0.018 

460 

Pù0.353 

548 

Pù0.256 

524 

Pù0.027 

Alveolar/bronchiolar Carcinomah 

Overall rate 4/50 (8%) 5/50 (10%) 7/50 (14%) 11/50 (22%) 

Adjusted rate 13.2% 16.1% 25.3% 43.7% 

Terminal rate 2/22 (9%) 5/31 (16%) 4/24 (17%) 7/20 (35%) 

First incidence (days) 

Logistic regression test 

449 

Pù0.006 

733 (T) 

Pù0.528 

687 

Pù0.273 

552 

Pù0.033 

Alveolar/bronchiolar Adenoma or Carcinomai 

Overall rate 11/50 (22%) 14/50 (28%) 19/50 (38%) 28/50 (56%) 

Adjusted rate 35.5% 36.5% 56.5% 78.8% 

Terminal rate 5/22 (23%) 8/31 (26%) 10/24 (42%) 13/20 (65%) 

First incidence (days) 

Logistic regression test 

449 

P<0.001 

460 

Pù0.345 

548 

Pù0.071 

524 

P<0.001 

Female 

Number Examined Microscopically 50 50 50 50 

Infiltration Cellular, Diffuse, Histiocyte 0 0 0 4 (3.3) 

Infiltration Cellular, Focal, Histiocyte 2 (2.0) 5 (1.8) 7 (2.9) 10* (2.4) 

Bronchus, Cytoplasmic Vacuolization 0 6* (1.0) 31**(1.0) 43**(1.0) 

Alveolar Epithelium Hyperplasia 2 (1.5) 3 (1.3) 0 5 (2.0) 

Alveolar/bronchiolar Adenomaj 

Overall rate 3/50 (6%) 6/50 (12%) 9/50 (18%) 10/50 (20%) 

Adjusted rate 8.8% 15.0% 25.2% 32.8% 

Terminal rate 3/34 (9%) 4/37 (11%) 6/32 (19%) 8/28 (29%) 

First incidence (days) 

Logistic regression test 

734 (T) 

Pù0.024 

664 

Pù0.287 

649 

Pù0.057 

706 

Pù0.024 

Alveolar/bronchiolar Carcinomak 

Overall rate 1/50 (2%) 1/50 (2%) 4/50 (8%) 9/50 (18%) 

Adjusted rate 2.9% 2.7% 9.2% 25.3% 

Terminal rate 1/34 (3%) 1/37 (3%) 1/32 (3%) 4/28 (14%) 

First incidence (days) 

Logistic regression test 

734 (T) 

P<0.001 

734 (T) 

Pù0.743N 

495 

Pù0.201 

536 

Pù0.009 
 

48 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE 11 

Incidences of Neoplasms and Nonneoplastic Lesions of the Lung in Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Female (continued) 

Alveolar/bronchiolar Adenoma or Carcinomal 

Overall rate 4/50 (8%) 7/50 (14%) 13/50 (26%) 18/50 (36%) 

Adjusted rate 11.8% 17.5% 32.6% 50.2% 

Terminal rate 4/34 (12%) 5/37 (14%) 7/32 (22%) 11/28 (39%) 

First incidence (days) 

Logistic regression test 

734 (T) 

P<0.001 

664 

Pù0.318 

495 

Pù0.016 

536 

P<0.001 

* Significantly different (P#0.05) from the chamber control by the logistic regression test 

** P#0.01 

(T)Terminal sacrifice 

a Number of animals with lesion 

b Average severity grade of lesions in affected animals: 1 ù minimal, 2 ù mild, 3 ù moderate, 4 ù marked c Historical incidence for 2-year NTP inhalation studies with chamber control groups (mean ± standard deviation): 141/947 

(14.9% ± 7.0%); range 6%-36% d Number of animals with neoplasm per number of animals with lung examined microscopically e Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality f Observed incidence in animals surviving until the end of the study g In the chamber control column are the P values associated with the trend test. In the exposed group columns are the P values 

corresponding to the pairwise comparisons between the chamber controls and that exposed group. The logistic regression test regards 

lesions in animals dying prior to terminal kill as nonfatal. A lower incidence in an exposure group is indicated by N. h Historical incidence: 75/947 (7.9% ± 5.7%); range 0%-16% i Historical incidence: 205/947 (21.7% ± 8.0%); range 10%-42% j Historical incidence: 61/939 (6.5% ± 3.2%); range 0%-14% k Historical incidence: 38/939 (4.1% ± 3.2%); range 0%-12% l Historical incidence: 97/939 (10.3% ± 3.7%); range 0%-16% 

Nose: The incidences of atrophy of the olfactory The suppurative inflammation involved only a few 

3 epithelium in 1.0 and 3.0 mg/m males and females animals and was a very mild change. It primarily 

and hyperplasia of the olfactory epithelium in involved animals that died prior to the end of the study 

3.0 mg/m3 males and females were significantly and consisted of a focal aggregate of inflammatory 

greater than those in the chamber controls. The inci- cells. 

dences of suppurative inflammation in 3.0 mg/m3 

males and in 1.0 mg/m3 females were significantly 

greater than those in the chamber controls (Tables 12, 

C5, and D5). The nasal lesions in mice were 

less severe than in the rats and involved limited seg- Larynx: The incidences of squamous metaplasia in all 

ments of the olfactory epithelium located further back exposed groups of males and females were signifin the nasal passage. Atrophy of the olfactory epi- icantly greater than those in the chamber controls 

thelium was characterized by loss of cell layers (Tables 12, C5, and D5). Squamous metaplasia was 

(sensory cells) and a decrease in the number of axons limited to the base of the epiglottis and was not a 

in the lamina propria. Hyperplasia of the olfactory severe lesion in exposed mice. It was characterized by 

epithelium was observed only in animals exposed to replacement of the ciliated respiratory epithelium by 3 3.0 mg/m and was characterized by increased num- one or more layers of flattened epithelial cells overbers of sensory cells that were usually arranged in lying a basal layer of cuboidal cells. Keratinization 

nests or rosettes. was sometimes observed. 
 

 

 

49 

c 

Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE 12 

Incidences of Nonneoplastic Lesions of the Nose and Larynx in Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Male 

Nosea 50 50 48 49 

Olfactory Epithelium, Atrophyb 0 0 29**(1.2)c 48**(1.8) 

Olfactory Epithelium, Hyperplasia 0 0 0 10**(1.0) 

Inflammation, Suppurative 0 1 (3.0) 0 6* (2.2) 

Larynx 48 49 48 49 

Metaplasia, Squamous 0 37**(1.0) 48**(1.0) 44**(1.0) 

Female 

Nose 50 50 49 48 

Olfactory Epithelium, Atrophy 0 2 (1.5) 12**(1.0) 46**(1.5) 

Olfactory Epithelium, Hyperplasia 0 0 0 30**(1.3) 

Inflammation, Suppurative 0 1 (1.0) 5* (1.6) 4 (1.5) 

Larynx 50 49 47 50 

Metaplasia, Squamous 0 45**(1.0) 40**(1.0) 50**(1.1) 

* Significantly different (P#0.05) from the chamber control by the logistic regression test 

** P#0.01 a Number of animals with tissue examined microscopically b Number of animals with lesion 

Average severity grade of lesions in affected animals: 1 ù minimal, 2 ù mild, 3 ù moderate, 4 ù marked 

Thyroid Gland: The incidences of follicular cell in severity and observed throughout the liver (usually 

hyperplasia in all exposed groups of males were not within proliferative lesions), but they appeared 

significantly greater than the incidence in the chamber most pronounced in the portal regions. Similar lesions 

3 controls (chamber control, 3/49; 0.3 mg/m , 17/50; were observed in only a few females, and the severity 3 3 1.0 mg/m , 11/50; 3.0 mg/m , 10/50; Table C5). was also much less than that observed in most males 

Minimal hyperplasias are commonly observed in (Tables 13 and D5). This spectrum of lesions is 

untreated male and female mice, suggesting that the consistent with those observed with Helicobacter 

rate in the concurrent chamber control group is low. hepaticus infection (Appendix J). Liver sections from 

The severity of most hyperplasias in these mice was four of five male mice with liver lesions were positive 

minimal to mild and did not differ between chamber for bacterial organisms consistent with H. hepaticus 

control and exposed groups. The incidence of hyper- when examined using Steiner’s modification of the 

plasia did not increase with exposure to cobalt sulfate Warthin Starry silver stain. 

heptahydrate, nor was the incidence of neoplasms of 

the follicular cells increased. 

The incidences of hemangiosarcoma in all exposed

3 Liver: High incidences of chronic inflammation, groups of male mice and in 1.0 mg/m in female mice 

karyomegaly, oval cell hyperplasia, and regeneration exceeded the range observed in historical controls for 

occurred in all groups of male mice and were usually inhalation studies (Tables 13, C3, and C4b). In 

observed together in the same liver (Tables 13 and addition, the incidence of hemangiosarcoma in 

3 C5). These changes were generally mild to moderate 1.0 mg/m males was significantly greater than that in 
50 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE 13 

Incidences of Neoplasms and Nonneoplastic Lesions of the Liver of Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Male 

Number Examined Microscopically 

Inflammation, Chronica 50 33 (1.3)b 

50 

36 (1.6) 

50 

40 (1.7) 

50 

39 (1.3) 

Karyomegaly 39 (2.3) 35 (2.8) 39 (2.7) 43 (2.7) 

Regeneration 32 (2.3) 30 (2.7) 35 (2.4) 38 (2.8) 

Bile Duct, Hyperplasia 0 3 (1.3) 6* (1.7) 4 (2.5) 

Oval Cell, Hyperplasia 38 (2.6) 36 (2.8) 40 (2.7) 44 (2.7) 

Hemangiosarcomac 

Overall rated 

Adjusted ratee 

Terminal ratef 

2/50 (4%) 

9.1% 

2/22 (9%) 

4/50 (8%) 

11.5% 

2/31 (6%) 

8/50 (16%) 

23.5% 

2/24 (8%) 

7/50 (14%) 

25.0% 

3/20 (15%) 

First incidence (days) 

Logistic regression test g 

733 (T) 

Pù0.078 

685 

Pù0.441 

523 

Pù0.050 

502 

Pù0.069 

Female 

Number Examined Microscopically 50 50 50 49 

Inflammation, Chronic 6 (1.7) 1 (1.0) 1 (1.0) 2 (2.0) 

Karyomegaly 4 (2.8) 2 (1.5) 0 1 (2.0) 

Oval Cell, Hyperplasia 2 (2.0) 1 (2.0) 0 0 

Hemangiosarcomah 

Overall rate 1/50 (2%) 0/50 (0%) 3/50 (6%) 0/49 (0%) 

Adjusted rate 2.9% 0.0% 7.3% 0.0% 

Terminal rate 

First incidence (days) 

Logistic regression test 

1/34 (3%) 

734 (T) 

Pù0.431N 

0/37 (0%) 

—i 

Pù0.483N 

1/32 (3%) 

524 

Pù0.318 

0/28 (0%) 

— 

Pù0.539N 

* Significantly different (P#0.05) from the chamber control by the logistic regression test 

(T)Terminal sacrifice 

a Number of animals with lesion 

b Average severity grade of lesions in affected animals: 1 ù minimal, 2 ù mild, 3 ù moderate, 4 ù marked c Historical incidence for 2-year NTP inhalation studies with chamber control groups (mean ± standard deviation): 12/947 (1.3% ± 

1.7%); range 0%-6% d	 Number of animals with neoplasm per number of animals with liver examined microscopically e Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality f	 Observed incidence in animals surviving until the end of the study g	 In the chamber control column are the P values associated with the trend test. In the exposed group columns are the P values 

corresponding to the pairwise comparisons between the chamber controls and that exposed group. The logistic regression test regards 

lesions in animals dying prior to terminal kill as nonfatal. A negative trend or lower incidence in an exposure group is indicated by N. h	 Historical incidence: 5/937 (0.5% ± 1.0%); range 0%-3% i	 Not applicable; no neoplasms in animal group 

the chamber controls. Hemangiosarcomas were GENETIC TOXICOLOGY 

morphologically similar to those observed spon- Cobalt sulfate heptahydrate (3 to 10,000 μg/mL) was taneously and consisted of multiple variably sized mutagenic in Salmonella typhimurium strain TA100 in blood-filled spaces that were separated by cords of the absence of S9 metabolic activation, and with 5% hepatocytes and lined by plump endothelial cells. hamster or rat liver S9; no mutagenicity was detected 

in strain TA98 or TA1535, with or without S9 (Zeiger 

et al., 1992; Table E1). 
PLATE 1 

Low magnification of a typical area of chronic inflammation (arrows) 

in the lung of a female F344/N rat exposed to 3.0mg/m3 cobalt sulfate 

heptahydrate by inhalation for 2 years. H&E; 20× 

PLATE 3 

Hyperplasia (arrow) in the lung of a female F344/N rat exposed to 3.0 

mg/m3 cobalt sulfate heptahydrate by inhalation for 2 years. H&E; 

20× 

PLATE 2 

Higher magnification of an area of chronic inflammation. Note the 

areas of fibrosis (large arrow), foamy alveolar macrophages (large 

curved arrow), necrotic cellular debris (small arrow), and epithelial 

hyperplasia (curved arrows) in the lung of a female F344/N rat 

exposed to 3.0 mg/m3 cobalt sulfate heptahydrate by inhalation for 2 

years. H&E; 160× 

PLATE 4 

Higher magnification of Plate 3. Note the proliferation of cells along 

the alveolar walls, but normal alveolar strucvture is maintained. 

H&E; 100× 
PLATE 5 PLATE 6 

Alveolar/bronchiolar adenoma in the lung of a male F344/N rat Higher magnification of Plate 5. Component cells are arranged in 

exposed to 3.0 mg/m3 cobalt sulfate heptahydrate by inhalation for 2 acini (small arrows) and papillary projection s (large arrow). H&E; 

years. H&E; 26× 66× 

PLATE 7 PLATE 8 

Alveolar/bronchiolar carcinoma in the lung of a female F344/N rat Atypical hyperplasia (arrows) in the lung of a female F344/N rat 

exposed to 3.0 mg/m3 cobalt sulfate heptahydrate by inhalation for 2 exposed to 1.0 mg/m3 cobalt sulfate heptahydrate by inhalation for 2 

years. Note the variation in the size of the cells comprising acini at years. The lesion is located within an area of chronic inflammation 

this high magnification. H&E; 200× (curved arrows). H&E; 16× 
PLATE 9 PLATE 10 

Higher magnification of Plate 8. Note the glandular structures High magnification of the border of an atypical hyperplasia in the 

(arrows) lined by cuboidal epithelium within the fibrotic core. H&E; lung of a male F344/N rat exposed to 3.0 mg/m3 cobalt sulfate 

80× heptahydrate by inhalation for 2 years. Note the necrotic debris within 

the glandular structure (arrows) and the proliferative epithelium at the 

periphery (curved arrows). H&E; 80× 

PLATE 11 

Alveolar/bronchiolar carcinoma with abundant fibrous connective 

tissue (arrows) in the lung of a male F344/N rat exposed to 1.0 mg/m3 

cobalt sulfate heptahydrate by inhalation for 2 years. H&E; 10× 

PLATE 12 

Squamous metaplasia along the alveolar wall consisting of several 

layers of squamous epithelium (arrows) in the lung of a female 

F344/N rat exposed to 3.0 mg/m3 cobalt sulfate heptahydrate by 

inhalation for 2 years. H&E; 100× 
PLATE 13 

Squamous cyst rimmed by a variably thick wall of squamous 

epithelium (large arrows) and filled with keratinous material (curved 

arrows) in the lung of a male F344/N rat exposed to 3.0 mg/m3 cobalt 

sulfate heptahydrate by inhalation for 2 years. H&E; 66×
 

PLATE 14 

High magnification of a squamous cell carcinoma in the lung of a
 

female F344/N rat exposed to 3.0 mg/m3 cobalt sulfate heptahydrate
 

by inhalation for 2 years. H&E; 40×
 

PLATE 15 

Alveolar/bronchiolar carcinoma with an area of alveolar/bronchiolar 

epithelium to the right (curved arrows) and squamous differentiation 

to the left (arrow) in the lung of a male F344/N rat exposed to 1.0 

mg/m3 cobalt sulfate heptahydrate by inhalation for 2 years. H&E; 

66× 
PLATE 16 

Alveolar/bronchiolar adenoma in the lung of a female B6C3F1 mouse 

exposed to 3.0 mg/m3 cobalt sulfate heptahydrate by inhalation for 2 

years. H&E; 40× 

PLATE 17 

Section of an alveolar/bronchiolar carcinoma with irregular and 

variably sized acinar structures in a female B6C3F1 mouse exposed to 

3.0 mg/m3 cobalt sulfate heptahydrate by inhalation for 2 years. 

H&E; 26× 
51 

DISCUSSION AND CONCLUSIONS
 

This report presents the findings and conclusions of mation in the alveoli of rats was much more severe 

2-year inhalation studies with cobalt sulfate hepta- and occurred at lower concentrations than in the 

hydrate. A companion report (NTP, 1991) discusses prechronic studies, and proteinosis was moderate to 

the findings of 16-day and 13-week inhalation studies marked in the 2-year study rats and not noted in the 

conducted prior to the 2-year studies at the same prechronic study. Interstitial fibrosis is known to be 

laboratory. In all studies, the respiratory tract was the a rather slowly developing lesion, but the extent of 

primary site of nonneoplastic lesions and neoplasms. this lesion and its occurrence in essentially all rats at 

In the 13-week studies, laryngeal lesions ranged from all exposure concentrations was not predicted based on 

mild squamous metaplasia with or without chronic the findings of the 13-week study. The alveolar 

inflammation at concentrations ultimately selected for epithelium of rats also displayed a spectrum of prolifthe 2-year studies, to large inflammatory polyps erative changes ranging from metaplasia through 

present in rats exposed to higher concentrations. hyperplasia and atypical hyperplasia, and extending to 

Although other respiratory tract lesions were present, neoplasia. 

the larynx appeared to be the most sensitive to cobalt 

sulfate heptahydrate exposure, and lesions in this The spectrum of proliferative lung lesions observed in 

tissue were the determining factor in exposure concen- rats in the 2-year study ranged from highly cellular 

tration selection for the 2-year studies. proliferations (typical of spontaneous lesions) to 

fibroproliferative and squamous lesions not typical of 3 The highest concentration (3.0 mg/m ) chosen for the spontaneous lesions, and morphologic variants in 

2-year studies did not affect survival or body weight between. The biological behavior of “typical” lung 

gains of rats or survival of mice in either the 13-week lesions, and to a lesser extent, squamous lesions, is 

or 2-year studies. The polycythemia noted in rats in fairly well documented. However, little is known 

3 the 13-week study was very mild at 3.0 mg/m , and about the biology of fibroproliferative lesions. In this 

there was no indication that this effect worsened to the study, many of the smaller lesions were identified 

point of causing clinical effects with longer exposure, within and/or adjacent to areas of chronic inflamalthough no hematologic measures were performed mation and fibrosis; however, it was clear that these 

during the 2-year study. Similarly, there was no lesions represented proliferative lesions distinct from 

indication that the lesions observed in rats and mice in the inflammation. Based upon the morphologic 

the 13-week studies in the larynx progressed in extent spectrum observed, it appears that their growth is 

or changed in character with the prolonged exposures. progressive. There was, however, no clear morphoThere was no evidence of laryngeal polyp formation in logic correlate signaling autonomy of growth (i.e., 

rats, and the metaplastic and inflammatory changes in consistent with a benign neoplasm) for these fibrorats remained greater than in mice. proliferative lesions. Therefore, unless growth alterations consistent with a malignant neoplasm were 

In contrast to the findings in the larynx, prolonged present, all fibroproliferative lesions were diagnosed 

exposure to cobalt sulfate heptahydrate aerosol ap- as atypical hyperplasia. There were several animals 

peared to cause a progressive injury to the nose of rats that had malignant neoplasms with a very prominent 

and mice and to the lung of rats. Olfactory epithelial fibrous component; presumably, some of these prodegeneration occurred primarily in rats and mice gressed from atypical hyperplasias. In many respects, 

exposed to 10 and 30 mg/m3 in the 13-week studies, the range of proliferative lesions within the lungs 

but olfactory epithelial atrophy was increased at even of exposed rats resembled those observed in NTP 

3 the lowest concentration (0.3 mg/m ) in rats and at studies of particulates (talc and the nickel compounds; 3 1.0 mg/m in mice in the 2-year studies. Lesions in NTP, 1993, 1996a,b,c), and it is clear that all the 

the lungs of rats changed markedly in character with morphologic variants of proliferative lesions represent 

the prolonged exposure in the 2-year study. Inflam- a response to cobalt sulfate heptahydrate. 
52 Cobalt Sulfate Heptahydrate, NTP TR 471 

Nonneoplastic lesions in the lungs of mice exposed to much greater in females, and the increase in males was 

cobalt sulfate heptahydrate did not appear to differ judged equivocal. In the NTP database of chemical 

appreciably from those expected in mice based on the carcinogenesis studies of nearly 450 chemicals, 

results of the prechronic study. The lesions were con- pheochromocytomas were part of a carcinogenic 

fined primarily to histiocytic infiltration, and there was response in only 13 rat studies, five of which were 

an absence of fibrosis and only minimal evidence of inhalation studies. Although the historical control 

the nonneoplastic proliferative lesions noted in rates of pheochromocytomas do not appreciably differ 

exposed rats. Most of the diagnoses of histiocytic between inhalation and dosed feed studies, a positive 

infiltration were noted in animals that also had an response is more likely to occur in inhalation studies 

alveolar/bronchiolar neoplasm; this is a frequent than in studies using other routes of exposure. The 

observation in mice with lung neoplasms and is not reasons for this are not clear. Of the five other 

necessarily related to exposure to cobalt sulfate hepta- positive inhalation studies, two were with nickel 

hydrate. Thus, it is not possible to clearly attribute compounds (oxide and subsulfide) and one with the 

the presence of histiocytic infiltration to cobalt sul- particulate, talc. 

fate heptahydrate exposure. Nonetheless, the lung 

changes were clearly much less severe than those seen Although the mechanisms responsible for induction of 

in rats and differed markedly in character. pheochromocytomas in rats are not understood, it is 

worth considering whether the adrenal gland and the 

While rats and mice exhibited quite different nonneo- pulmonary responses to cobalt sulfate heptahydrate in 

plastic pulmonary responses to cobalt sulfate hepta- the rat might represent nonspecific responses to the 

hydrate, exposed male and female rats and mice physical inhalation and pulmonary accumulation of a 

developed alveolar/bronchiolar adenomas and car- particle, rather than a chemical-specific response. 

cinomas. The distinction between these neoplasms is Measures of the possible accumulation of cobalt in the 

largely based on size, and both categories of this lung were not taken during these studies, although 

neoplasm were increased in exposed male and female urinary cobalt concentrations have demonstrated doserats and mice. In all groups, the neoplasms appeared related absorption in the prechronic studies. Nickel 

with a significant positive trend, and the incidences in sulfate hexahydrate is a highly water-soluble salt, as is 3 the 3.0 mg/m groups exceeded the historical control cobalt sulfate heptahydrate. In similar studies, nickel 

ranges in the respective groups. The magnitude of the sulfate hexahydrate did not show evidence of 

neoplastic response was somewhat less in male rats exposure-concentration-related accumulation in the 

than in the other groups. lung of rats or mice exposed to concentrations as high 

as 30 mg/m3 (NTP, 1996c). In contrast, the less 

The incidences of follicular cell hyperplasia of the soluble nickel subsulfide (NTP, 1996b) and the highly 

thyroid gland were moderately increased in all ex- insoluble nickel oxide (NTP, 1996a) did accumulate in 

posed groups of male mice, although no dose response the lung. Thus, given the similar solubility and use of 

was observed. Hypothyroidism has been noted in exposure concentrations ten-fold lower than those used 

humans who also exhibited cardiomyopathy associated with nickel sulfate hexahydrate, it is unlikely that 

with consumption of cobalt-contaminated beer (Taylor cobalt would accumulate in the lung unless there was 

and Marks, 1978). specific toxicity to pulmonary clearance mechanisms. 

The absence of nonneoplastic changes associated with 

Incidences of pheochromocytoma of the adrenal cobalt sulfate heptahydrate inhalation by mice would 

medulla were increased in female rats exposed to argue against impaired clearance. The rather extensive 

cobalt sulfate heptahydrate. Pheochromocytomas are and progressive pulmonary toxicity in the rat could 

relatively common in male F344/N rats, occurring have resulted in impaired clearance of cobalt, but it is 

with an historical rate of about 30% in inhalation unlikely that the toxicity represented a simple 

studies. The historical inhalation chamber control rate inflammatory and fibrotic response to an “overload” 

in females is much lower (6%), and the incidence in situation as has been postulated with chemically inert 

the concurrent chamber control was 4%. While the particles (Morrow et al., 1991). The fact that the 

incidences of this neoplasm were increased in exposed entire respiratory tract demonstrated a toxic response 

males and females, the strength of the response was to cobalt sulfate heptahydrate argues convincingly that 
Cobalt Sulfate Heptahydrate, NTP TR 471 53 

the chemical has inherent toxicity and is not acting The mechanisms of cobalt-induced carcinogenesis are 

through secondary mechanisms related to its inhalation not well understood. The genotoxicity of cobalt comas a particle. pounds has been established in a variety of eukaryotic 

test systems (reviewed in IARC, 1991), and cobalt has 

A number of factors need to be considered to properly been shown under certain conditions to catalyze the 

address the relationship of these findings to typical production of oxygen-based free radicals that may 

human exposures to cobalt. The segments of the underlie some of the observed adverse genetic events 

human population with the highest potential exposure (Shi et al., 1993). The observation of a larger than 

to significant airborne cobalt concentrations are usual number of G to T transversions at the second 

workers in the hard metal industry, coal mining, and base of codon 12 of those mouse lung neoplasms 

those involved in ore processing (USDHHS, 1992). carrying a mutated K-ras gene (Appendix I) is also 

In these situations cobalt may exist in various forms, consistent with oxidative injury. Similar increases in 

primarily as cobalt powder or cobalt oxide. These G to T transversions were seen in lung neoplasms 

agents are less soluble than cobalt sulfate hepta- from mice exposed to ozone (NTP, 1994) 

hydrate, and the toxic response of the respiratory 

system would likely depend on the combination of The potential contribution of the sulfate moiety to the 

inherent toxicity, solubility in biological fluids, and carcinogenic response is worthy of consideration in 

residence time in the tissue. The carcinogenic poten- that exposures of humans to concentrated inorganic 

tial of various cobalt compounds has been perhaps best acid mists are recognized as causing respiratory tract 

demonstrated in injection studies in experimental neoplasms, primarily in the larynx (IARC, 1992). 

animals (reviewed in IARC, 1991), and both insoluble There are no experimental animal carcinogenicity 

and soluble forms have been shown to produce studies with sulfuric acid mists per se (IARC, 1992), 

injection-site neoplasms. but nickel sulfate hexahydrate was studied by inhalation as mentioned earlier (NTP, 1996c). In this 

The present demonstration of alveolar/bronchiolar instance, there was no evidence of carcinogenicity of 

neoplasms in rats and mice exposed to cobalt sulfate nickel sulfate hexahydrate to the respiratory tract or 

heptahydrate by inhalation confirms the findings of the other tissues despite the fact that other nickel salts are 

injection studies and suggests that cobalt is inherently carcinogenic. Additionally, nickel sulfate hexahydrate 

carcinogenic. These findings also lend credence to the was studied at an equivalent exposure concentration to 

epidemiological investigations of Mur et al. (1987), that which caused significant increases in lung neoHogstedt and Alexandersson (1990), and Lasfargues plasms in mice exposed to cobalt sulfate heptahydrate 3 et al. (1994) that reported increased risks for lung (1.0 mg/m ). Thus, there seems to be little evidence 

cancer among workers producing cobalt and exposed to suggest that the sulfate moiety contributed signifto cobalt in the hard metal industry. Cobalt con- icantly to the carcinogenic response. 

centrations in the urine of workers in the Italian hard 

metal industry were found to be as high as Based on retrospective analyses, Helicobacter 

0.21 μg/mL at the end of the work shift (Sabbioni hepaticus was determined to have infected mice in 

et al., 1994). Ichikawa et al. (1985) reported even 12 recent NTP 2-year studies (Appendix J). Of the 

higher concentrations (0.39 μg/mL) in Japanese 12 studies, mice (primarily males) from nine studies 

workers. In prechronic inhalation studies reported (including this study of cobalt sulfate heptahydrate) 

previously (NTP, 1991), average urinary cobalt had a H. hepaticus-associated hepatitis. Qualitatively, 

concentrations in rats exposed to 0.3, 1.0, and the hepatitis and silver-staining organisms within the 

3 3.0 mg/m , respectively, were 0.14, 0.32, and liver were similar among the nine studies. In a 

1.77 μg/mL. If urine cobalt concentrations roughly polymerase chain reaction-restriction fragment length 

approximate relative inhalation exposures to cobalt, polymorphism (PCR-RFLP) based assay, H. hepaticus 

then the results from the current 2-year rat and mouse was identified in studies from which adequately 

studies appear similar to occupational exposure levels preserved (frozen) liver tissue was available. In 

and suggest that humans and rodents may be similarly general, efforts to identify H. hepaticus from tissue 

sensitive to cobalt carcinogenesis. fixed in formalin for over a week were not successful 
__________ 

54 Cobalt Sulfate Heptahydrate, NTP TR 471 

(Malarkey et al., 1997), which was the case for this 

study of cobalt sulfate heptahydrate. However, because of the presence of the typical liver lesions and 

silver-staining helical organisms, mice from the study 

were presumed to be infected with H. hepaticus. 

Increases in the incidences of hepatocellular neoplasms 

in male mice have been shown to be associated with 

H. hepaticus infection when hepatitis is also present 

(Ward et al., 1994; Fox et al., 1996; Appendix J). 

Additionally, in NTP studies with H. hepaticusassociated hepatitis, increased incidences of hemangiosarcoma were seen in the livers of male mice 

(Appendix J). Because of the latter association, 

interpretation of the increased incidences of hemangiosarcoma in the liver of male mice was confounded. 

Incidences of lesions at other sites in this study of 

cobalt sulfate heptahydrate were not considered to 

have been significantly impacted by the infection with 

H. hepaticus or its associated hepatitis (Appendix J). 

CONCLUSIONS 

Under the conditions of these 2-year inhalation studies, there was some evidence of carcinogenic activity* 

of cobalt sulfate heptahydrate in male F344/N rats 

based on increased incidences of alveolar/bronchiolar 

neoplasms. Marginal increases in incidences of 

pheochromocytomas of the adrenal medulla may have 

been related to exposure to cobalt sulfate heptahydrate. 

There was clear evidence of carcinogenic activity in 

female F344/N rats based on increased incidences of 

alveolar/bronchiolar neoplasms and pheochromocytomas of the adrenal medulla in groups 

exposed to cobalt sulfate heptahydrate. There was 

clear evidence of carcinogenic activity of cobalt sulfate 

heptahydrate in male and female B6C3F1 mice based 

on increased incidences of alveolar/bronchiolar neoplasms. 

Exposure to cobalt sulfate heptahydrate caused a 

spectrum of inflammatory, fibrotic, and proliferative 

lesions in the respiratory tract of male and female rats 

and mice. 

* Explanation of Levels of Evidence of Carcinogenic Activity is on page 10. A summary of the Technical Reports Review Subcommittee 

comments and the public discussion on this Technical Report appears on page 12. 
55 

REFERENCES
 

Abbracchio, M.P., Heck, J.D., and Costa, M. (1982). 

The phagocytosis and transforming activity of 

crystalline metal sulfide particles are related to their 

negative surface charge. Carcinogenesis 3, 175-180. 

American Conference of Governmental Industrial 

Hygienists (ACGIH) (1996). 1996 Threshold Limit 

Values for Chemical Substances and Physical Agents 

and Biological Exposure Indices. Cincinnati, OH. 

Anderson, O. (1983). Effects of coal combustion 

products and metal compounds on sister chromatid 

exchanges in a macrophagelike cell line. Environ. 

Health Perspect. 47, 239-253. 

Armitage, P. (1971). Statistical Methods in Medical 

Research, pp. 362-365. John Wiley and Sons, New 

York. 

Ashby, J., and Tennant, R.W. (1991). Definitive 

relationships among chemical structure, carcinogenicity and mutagenicity for 301 chemicals tested by 

the U.S. NTP. Mutat. Res. 257, 229-306. 

Baker, P.F., Meves, H., and Ridgway, E.B. (1973). 

Effects of manganese and other agents on the calcium 

uptake that follows depolarization of squid axons. J. 

Physiol. 231, 511-526. 

Barrett, K.W. (1983). Report from the Mitre 

Corporation to the U.S. Environmental Protection 

Agency. 

Beyersmann, D., and Hartwig, A. (1992). The 

genetic toxicology of cobalt. Toxicol. Appl. 

Pharmacol. 115, 137-145. 

Boorman, G.A., Montgomery, C.A., Jr., Eustis, S.L., 

Wolfe, M.J., McConnell, E.E., and Hardisty, J.F. 

(1985). Quality assurance in pathology for rodent 

carcinogenicity studies. In Handbook of Carcinogen 

Testing (H.A. Milman and E.K. Weisburger, Eds.), 

pp. 345-357. Noyes Publications, Park Ridge, NJ. 

Bucher, J.R., Elwell, M.R., Thompson, M.B., 

Chou, B.J., Renne, R., and Ragan, H.A. (1990). 

Inhalation toxicity studies of cobalt sulfate in F344/N 

rats and B6C3F1 mice. Fundam. Appl. Toxicol. 15, 

357-372. 

Buttlaire, D.H., Czuba, B.A., Stevens, T.H., 

Lee, Y.C., and Himes, R.H. (1980). Manganous ion 

binding to tubulin. J. Biol. Chem. 255, 2164-2168. 

Christensen, J.M., and Poulsen, O.M. (1994). A 

1982-1992 surveillance programme on Danish pottery 

painters. Biological levels and health effects following 

exposure to soluble or insoluble cobalt compounds in 

cobalt blue dyes. Sci. Total Environ. 150, 95-104. 

Cirla, A.M. (1994). Cobalt-related asthma: Clinical 

and immunological aspects. Sci. Total Environ. 150, 

85-94. 

Code of Federal Regulations (CFR). 21, Part 58. 

Cox, D.R. (1972). Regression models and life-tables. 

J. R. Stat. Soc. B34, 187-220. 

Crawford, B.D. (1985). Perspectives on the somatic 

mutation model of carcinogenesis. In Advances in 

Modern Environmental Toxicology. Mechanisms and 

Toxicity of Chemical Carcinogens and Mutagens 

(M.A. Mehlman, W.G. Flamm, and R.J. Lorentzen, 

Eds.), pp. 13-59. Princeton Scientific Publishing Co., 

Princeton, NJ. 

De Bie, E., and Doyen, P. (1962). Cobalt oxides and 

salts. Cobalt 15, 3-13. 

De Matteis, F., and Gibbs, A.H. (1977). Inhibition of 

haem synthesis caused by cobalt in rat liver. Evidence 

for two different sites of action. Biochem. J. 162, 

213-216. 
56 Cobalt Sulfate Heptahydrate, NTP TR 471 

Dingle, J.T., Heath, J.C., Webb, M., and Daniel, M. 

(1962). The biological action of cobalt and other 

metals. II. The mechanism of the respiratory inhibition produced by cobalt in mammalian tissues. 

Biochim. Biophys. Acta 65, 34-46. 

Dinse, G.E., and Haseman, J.K. (1986). Logistic 

regression analysis of incidental-tumor data from 

animal carcinogenicity experiments. Fundam. Appl. 

Toxicol. 6, 44-52. 

Dinse, G.E., and Lagakos, S.W. (1983). Regression 

analysis of tumour prevalence data. Appl. Stat. 32, 

236-248. 

Domingo, J.L. (1989). Cobalt in the environment and 

its toxicological implications. Rev. Environ. Contam. 

Toxicol. 108, 105-123. 

Domingo, J.L., Llobet, J.M., and Bernat, R. (1984a). 

Nutritional and toxicological study of cobalt administered to rats in their drinking water. Rev. Toxicol. 1, 

43-54. 

Domingo, J.L., Llobet, J.M., and Bernat, R. (1984b). 

A study of the effects of cobalt administered orally to 

rats. Arch. Farmacol. Toxicol. 10, 13-20. 

Domingo, J.L., Paternain, J.L., Llobet, J.M., and 

Corbella, J. (1985). Effects of cobalt on postnatal 

development and late gestation in rats upon oral 

administration. Rev. Esp. Fisiol. 41, 293-298. 

Dorsit, G., Girard, R., Rousset, H., Brune, J., 

Wiesendanger, T., Tolot, F., Bourret, J., and Galy, P. 

(1970). Fibrose pulmonaire chez 3 sujects d’une 

meme usine exposes aux poussieres de cobalt et de 

carbure de tungstene. Les troubles pulmonaires de 

l’industrie des metaux durs. Sem. Hop. 46, 

3363-3376. 

Eaton, R.P. (1972). Cobalt chloride-induced hyperlipemia in the rat: Effects on intermediary metabolism. Am. J. Physiol. 222, 1550-1557. 

Edel, J., Sabbioni, E., Pietra, R., Rossi, A., 

Torre, M., Rizzato, G., and Fraioli, P. (1990). Trace 

metal lung disease: In vitro interaction of hard metals 

with human lung and plasma components. Sci. Total 

Environ. 95, 107-117. 

Finch, C.A. (1980). Drugs effective in irondeficiency 

and other hypochromic anemias. In Goodman and 

Gilman’s The Pharmacological Basis of Therapeutics, 

6th ed. (A.G. Gilman, L.S. Goodman, A. Gilman, 

S.E. Mayer, and K.L. Melmon, Eds.), pp. 1315-1330. 

Macmillan Publishing Co., New York. 

Fox, J.G., Li, X., Yan, L., Cahill, R.J., Hurley, R., 

Lewis, R., and Murphy, J.C. (1996). Chronic 

proliferative hepatitis in A/JCr mice associated with 

persistent Helicobacter hepaticus infection: A model 

of Helicobacter-induced carcinogenesis. Infect. 

Immun. 64, 1548-1558. 

Gart, J.J., Chu, K.C., and Tarone, R.E. (1979). 

Statistical issues in interpretation of chronic bioassay 

tests for carcinogenicity. JNCI 62, 957-974. 

Gilman, J.P.W. (1962). Metal carcinogenesis. II. A 

study on the carcinogenic activity of cobalt, copper, 

iron, and nickel compounds. Cancer Res. 22, 

158-162. 

Gregus, Z., and Klaassen, C.D. (1986). Disposition 

of metals in rats: A comparative study of fecal, 

urinary, and biliary excretion and tissue distribution of 

eighteen metals. Toxicol. Appl. Pharmacol. 85, 

24-38. 

Grice, H.C., Goodman, T., Munro, I.C., 

Wiberg, G.S., and Morrison, A.B. (1969). Myocardial toxicity of cobalt in the rat. Ann. N. Y. Acad. Sci. 

156, 189-194. 

Hamilton, E.I. (1994). The geobiochemistry of 

cobalt. Sci. Total Environ. 150, 7-39. 

Hamilton-Koch, W., Snyder, R.D., and Lavelle, J.M. 

(1986). Metal-induced DNA damage and repair in 

human diploid fibroblasts and Chinese hamster ovary 

cells. Chem. Biol. Interact. 59, 17-28. 

Hammond, P.B., and Beliles, R.P. (1980). Metals. 

In Casarett and Doull’s Toxicology, 2nd ed. (J. Doull, 

C.D. Klaassen, and M.O. Amdur, Eds.), pp. 442-443. 

Macmillan Publishing Co., New York. 

Harding, H.E. (1950). Notes on the toxicology of 

cobalt metal. Br. J. Ind. Med. 7, 76-78. 
 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 57 

Hartwig, A., Kasten, U., Boakye-Dankwa, K., 

Schlepegrell, R., and Beyersmann, D. (1990). Uptake and genotoxicity of micromolar concentrations of 

cobalt chloride in mammalian cells. Toxicol. Environ. 

Chem. 28, 205-215. 

Hartwig, A., Snyder, R.D., Schlepegrell, R., and 

Beyersmann, D. (1991). Modulation by Co(II) of 

UV-induced DNA repair, mutagenesis and sisterchromatid exchanges in mammalian cells. Mutat. Res. 

248, 177-185. 

Haseman, J.K. (1984). Statistical issues in the design, 

analysis and interpretation of animal carcinogenicity 

studies. Environ. Health Perspect. 58, 385-392. 

Heath, J.C. (1956). The production of malignant 

tumours by cobalt in the rat. Br. J. Cancer 10, 

668-673. 

Heath, J.C. (1960). The histogenesis of malignant 

tumours induced by cobalt in the rat. Br. J. Cancer 

14, 478-482. 

Heath, J.C., and Webb, M. (1967). Content and 

intracellular distribution of the inducing metal in the 

primary rhabdomyosarcomata induced in the rat by 

cobalt, nickel and cadmium. Br. J. Cancer 21, 

768-779. 

Hogstedt, C., and Alexandersson, R. (1990). Mortality among hard metal workers [in German, English 

summary]. Arbete och Hälsa 21, 1-26. 

Hollander, M., and Wolfe, D.A. (1973). Nonparametric Statistical Methods, pp. 120-123. John 

Wiley and Sons, New York. 

Ichikawa, Y., Kusaka, Y., and Goto, S. (1985). 

Biological monitoring of cobalt exposure, based on 

cobalt concentrations in blood and urine. Int. Arch. 

Occup. Environ. Health 55, 269-276. 

International Agency for Research on Cancer (IARC) 

(1991). IARC Monographs on the Evaluation of 

Carcinogenic Risks to Humans. Chlorinated Drinkingwater; Chlorination By-products; Some Other 

Halogenated Compounds; Cobalt and Cobalt Compounds, Vol. 52. IARC, Lyon, France. 

International Agency for Research on Cancer (IARC) 

(1992). IARC Monographs on the Evaluation of 

Carcinogenic Risks to Humans. Occupational Exposures to Mists and Vapours from Strong Inorganic 

Acids; and Other Industrial Chemicals, Vol. 54. 

IARC, Lyon, France. 

Jacobziner, H., and Raybin, H.W. (1961). Poison 

control. Accidental cobalt poisoning. Arch. Pediatr. 

78, 200-205. 

Jasmin, G. (1974). Anaphylactoid edema induced in 

rats by nickel and cobalt salts. Proc. Soc. Exp. Biol. 

Med. 147, 289-292. 

Jennette, K.W. (1981). The role of metals in carcinogenesis: Biochemistry and metabolism. Environ. 

Health Perspect. 40, 233-252. 

Jensen, A.A., and Tüchsen, F. (1990). Cobalt exposure and cancer risk. Crit. Rev. Toxicol. 20, 

427-437. 

Johansson, A., Curstedt, T., Robertson, B., and 

Camner, P. (1984). Lung morphology and phospholipids after experimental inhalation of soluble 

cadmium, copper, and cobalt. Environ. Res. 34, 

295-309. 

Johansson, A., Lundborg, M., Wiernik, A., 

Jarstrand, C., and Camner, P. (1986). Rabbit alveolar 

macrophages after long-term inhalation of soluble 

cobalt. Environ. Res. 41, 488-496. 

Kaplan, E.L., and Meier, P. (1958). Nonparametric 

estimation from incomplete observations. J. Am. Stat. 

Assoc. 53, 457-481. 

Kasirsky, G., Sherman, W.T., Gautieri, R.F., and 

Mann, D.E., Jr. (1969). Cobalt-cortisone interrelationships in the induction and inhibition of cleft palate 

in mice. J. Pharm. Sci. 58, 766-767. 

Kazantzis, G. (1981). Role of cobalt, iron, lead, 

manganese, mercury, platinum, selenium, and titanium in carcinogenesis. Environ. Health Perspect. 40, 

143-161. 
58 Cobalt Sulfate Heptahydrate, NTP TR 471 

Kerfoot, E.J., Fredrick, W.G., and Domeier, E. 

(1975). Cobalt metal inhalation studies on miniature 

swine. Am. Ind. Hyg. Assoc. J. 36, 17-25. 

Kichina, M.M. (1974). Cobalt and titanium levels in 

animals under the influence of cobalt sulfate. Sb. Rab. 

Leningr. Vet. Inst. 38, 83-87. 

Korman, E.F., Ward, J.F., and Myers, L.S. (1978). 

Development of toxicology of energy-related pollutants. DOE Symposium Series 47, 383-395. 

Krasovskii, G.N., and Fridlyand, S.A. (1971). 

Experimental data for the validation of the maximum 

permissible concentration of cobalt in water bodies. 

Hyg. Sanit. 36, 277-279. 

Lasfargues, G., Wild, P., Moulin, J.J., Hammon, B., 

Rosmorduc, B., du Noyer, C.R., Lavandier, M., and 

Moline, J. (1994). Lung cancer mortality in a French 

cohort of hard-metal workers. Am. J. Ind. Med. 26, 

585-595. 

Llobet, J.M., Domingo, J.L., and Corbella, J. (1986). 

Comparison of the effectiveness of several chelators 

after single administration on the toxicity, excretion 

and distribution of cobalt. Arch. Toxicol. 58, 

278-281. 

McConnell, E.E., Solleveld, H.A., Swenberg, J.A., 

and Boorman, G.A. (1986). Guidelines for combining neoplasms for evaluation of rodent carcinogenesis 

studies. JNCI 76, 283-289. 

McKee, G.K. (1971). Carcinogenic properties of wear 

particles from prostheses made in cobalt-chromium 

alloy. Lancet 1, 750. 

McKnight, B., and Crowley, J. (1984). Tests for 

differences in tumor incidence based on animal 

carcinogenesis experiments. J. Am. Stat. Assoc. 79, 

639-648. 

McLean, J.R., McWilliams, R.S., Kaplan, J.G., and 

Birnboim, H.C. (1982). Rapid detection of DNA 

strand breaks in human peripheral blood cells and 

animal organs following treatment with physical and 

chemical agents. In Progress in Mutation Research: 

Chemical Mutagenesis, Human Population Monitoring 

and Genetic Risk Assessment (K.C. Bora, 

G.R. Douglas, and E.R. Nestmann, Eds.), Vol. 3, 

pp. 137-141. Elsevier Biomedical Press, Amsterdam. 

Maines, M.D., and Kappas, A. (1976). Studies on the 

mechanism of induction of haem oxygenase by cobalt 

and other metal ions. Biochem. J. 154, 125-131. 

Malarkey, D.E., Ton, T.-V., Hailey, J.R., and 

Devereux, T.R. (1997). A PCR-RFLP method for the 

detection of Helicobacter hepaticus in frozen or fixed 

liver from B6C3F1 mice. Toxicol. Pathol. 25, 

606-612. 

Maronpot, R.R., and Boorman, G.A. (1982). Interpretation of rodent hepatocellular proliferative alterations and hepatocellular tumors in chemical safety 

assessment. Toxicol. Pathol. 10, 71-80. 

The Merck Index (1983). 10th ed. (M. Windholz, 

Ed.), p. 347. Merck and Company, Rahway, NJ. 

The Merck Index (1989). 11th ed. (S. Budavari, Ed.), 

p. 2439. Merck and Company, Rahway, NJ. 

Miller, J.A., and Miller, E.C. (1977). Ultimate 

chemical carcinogens as reactive mutagenic 

electrophiles. In Origins of Human Cancer 

(H.H. Hiatt, J.D. Watson, and J.A. Winsten, Eds.), 

pp. 605-627. Cold Spring Harbor Laboratory, Cold 

Spring Harbor, NY. 

Miller, M.E., Howard, D., Stohlman, F., Jr., and 

Flanagan, P. (1974). Mechanism of erythropoietin 

production by cobaltous chloride. Blood 44, 339-346. 
Cobalt Sulfate Heptahydrate, NTP TR 471 59 

Mitala, J.J., Mann, D.E., Jr., and Gautieri, R.F. 

(1978). Influence of cobalt (dietary), cobalamins, and 

inorganic cobalt salts on phenytoin- and cortisoneinduced teratogenesis in mice. J. Pharm. Sci. 67, 

377-380. 

Mollenhauer, H.H., Corrier, D.E., Clark, D.E., 

Hare, M.F., and Elissalde, M.H. (1985). Effects of 

dietary cobalt on testicular structure. Virchows Arch. 

Cell Pathol. 49, 241-248. 

Morgan, G.B., Ozolins, G., and Tabor, E.C. (1970). 

Air pollution surveillance systems. Science 170, 

289-296. 

Morin, Y., and Daniel, P. (1967). Quebec beerdrinkers’ cardiomyopathy: Etiological considerations. 

Can. Med. Assoc. J. 97, 926-928. 

Morita, H., Umeda, M., and Ogawa, H.I. (1991). 

Mutagenicity of various chemicals including nickel 

and cobalt compounds in cultured mouse FM3A cells. 

Mutat. Res. 261, 131-137. 

Morrow, P.E., Muhle, H., and Mermelstein, R. 

(1991). Chronic inhalation study findings as a basis 

for proposing a new occupational dust exposure limit. 

J. Am. Coll. Toxicol. 10, 279-290. 

Moulin, J.J., Wild, P., Mur, J.M., Fournier Betz, M., 

and Mercier-Gallay, M. (1993). A mortality study of 

cobalt production workers: An extension of the 

follow-up. Am. J. Ind. Med. 23, 281-288. 

Mur, J.M., Moulin, J.J., Charruyer-Seinerra, M.P., 

and Lafitte, J. (1987). A cohort mortality study 

among cobalt and sodium workers in an electrochemical plant. Am. J. Ind. Med. 11, 75-81. 

Murdock, H.R., Jr. (1959). Studies on the pharmacology of cobalt chloride. J. Am. Pharm. Assoc. 48, 

140-142. 

National Cancer Institute (NCI) (1976). Guidelines for 

Carcinogen Bioassay in Small Rodents. Technical 

Report Series No. 1. NIH Publication No. 76-801. 

U.S. Department of Health, Education, and Welfare, 

Public Health Service, National Institutes of Health, 

Bethesda, MD. 

National Institute for Occupational Safety and Health 

(NIOSH) (1981). Criteria for Controlling 

Occupational Exposure to Cobalt. Occupational 

Hazard Assessment. DHHS (NIOSH) Publication 

No. 82-107. U.S. Department of Health and Human 

Services, National Institute for Occupational Safety 

and Health, Washington, DC. 

National Institutes of Health (NIH) (1978). Open 

Formula Rat and Mouse Ration (NIH-07). Specification NIH-11-1335. U.S. Department of Health, 

Education, and Welfare, Public Health Service, 

National Institutes of Health, Bethesda, MD. 

National Research Council (NRC) (1977). Drinking 

Water and Health. National Academy of Sciences, 

Washington, DC. 

National Toxicology Program (NTP) (1991). Toxicity 

Studies of Cobalt Sulfate Heptahydrate (CAS 

No. 10026-24-1) in F344/N Rats and B63C3F Mice 1 

(Inhalation Studies). Toxicity Study Report Series 

No. 5. NIH Publication No. 91-3124. U.S. Department of Health and Human Services, Public Health 

Service, National Institutes of Health, Research 

Triangle Park, NC. 

National Toxicology Program (NTP) (1993). 

Toxicology and Carcinogenesis Studies of Talc (CAS 

No. 14807-96-6) in F344/N Rats and B6C3F1 Mice 

(Inhalation Studies). Technical Report Series No. 421. 

NIH Publication No. 93-3152. U.S. Department of 

Health and Human Services, Public Health Service, 

National Institutes of Health, Research Triangle 

Park, NC. 
 

60 Cobalt Sulfate Heptahydrate, NTP TR 471 

National Toxicology Program (NTP) (1994). 

Toxicology and Carcinogenesis Studies of Ozone 

(CAS No. 10028-15-6/64091-91-4) in F344/N Rats 

and B6C3F1 Mice (Inhalation Studies). Technical 

Report Series No. 440. NIH Publication 

No. 95-3371. U.S. Department of Health and Human 

Services, Public Health Service, National Institutes of 

Health, Research Triangle Park, NC. 

National Toxicology Program (NTP) (1996a). 

Toxicology and Carcinogenesis Studies of Nickel 

Oxide (CAS No. 1313-99-1) in F344/N Rats and 

B6C3F Mice (Inhalation Studies). Technical Report 1 

Series No. 451. NIH Publication No. 96-3367. U.S. 

Department of Health and Human Services, Public 

Health Service, National Institutes of Health, Research 

Triangle Park, NC. 

National Toxicology Program (NTP) (1996b). 

Toxicology and Carcinogenesis Studies of Nickel 

Subsulfide (CAS No. 12035-72-2) in F344/N Rats 

and B6C3F1 Mice (Inhalation Studies). Technical 

Report Series No. 453. NIH Publication 

No. 96-3369. U.S. Department of Health and Human 

Services, Public Health Service, National Institutes of 

Health, Research Triangle Park, NC. 

National Toxicology Program (NTP) (1996c). 

Toxicology and Carcinogenesis Studies of Nickel 

Sulfate Hexahydrate (CAS No. 10101-97-0) in F344/N 

Rats and B6C3F Mice (Inhalation Studies). Technical 1 

Report Series No. 454. NIH Publication 

No. 96-3370. U.S. Department of Health and Human 

Services, Public Health Service, National Institutes of 

Health, Research Triangle Park, NC. 

Ogawa, H.I., Shibahara, T., Iwata, H., Okada, T., 

Tsuruta, S., Kakimoto, K., Sakata, K., Kato, Y., 

Ryo, H., Itoh, T., and Fujikawa, K. (1994). Genotoxic activities in vivo of cobaltous chloride and other 

metal chlorides as assayed in the Drosophila wing spot 

test. Mutat. Res. 320, 133-140. 

Paley, K.R., Sobel, E.S., and Yalow, R.S. (1958). 

Effect of oral and intravenous cobaltous chloride on 

thyroid function. J. Clin. Endocrinol. Metab. 18, 

850-859. 

Paternain, J.L., Domingo, J.L., and Corbella, J. 

(1988). Developmental toxicity of cobalt in the rat. 

J. Toxicol. Environ. Health 24, 193-200. 

Pedigo, N.G., George, W.J., and Anderson, M.B. 

(1988). Effects of acute and chronic exposure to 

cobalt on male reproduction in mice. Reprod. Toxicol. 

2, 45-53. 

Phillips, C.E. (1980). Intracellularly injected 

cobaltous ions accumulate at synaptic densities. 

Science 207, 1477-1479. 

Popov, L.N. (1977). Izuchenie vliianiia malykh 

konsentratsii aeorosolia metallicheskogo kobal’ta na 

organizm zhivotnykh v gigiencheskom eksperimente. 

Gig. Sanit. 4, 97-98. 

Sabbioni, E., Minoia, C., Pietra, R., Mosconi, G., 

Forni, A., and Scansetti, G. (1994). Metal determinations in biological specimens of diseased and nondiseased hard metal workers. Sci. Total Environ. 150, 

41-54. 

Scansetti, G., Botta, G.C., Spinelli, P., 

Reviglione, L., and Ponzetti, C. (1994). Absorption 

and excretion of cobalt in the hard metal industry. 

Sci. Total Environ. 150, 141-144. 

Sederholm, T., Kouvalainen, K., and Lamberg, B.A. 

(1968). Cobalt-induced hypothyroidism and polycythemia in lipoid nephrosis. Acta Med. Scand. 184, 

301-306. 

Seidenberg, J.M., Anderson, D.G., and Becker, R.A. 

(1986). Validation of an in vivo developmental 

toxicity screen in the mouse. Teratog. Carcinog. 

Mutagen. 6, 361-374. 

Shabaan, A.A., Marks, V., Lancaster, M.C., and 

Dufeu, G.N. (1977). Fibrosarcomas induced by 

cobalt chloride (CoCl ) in rats. Lab. Anim. 11, 43-46. 2 

Shi, X., Dalal, N.S., and Kasprzak, K.S. (1993). 

Generation of free radicals from model lipid 

hydroperoxides and H O by Co(II) in the presence of 2 2 

cysteinyl and histidyl chelators. Chem. Res. Toxicol. 

6, 277-283. 
Cobalt Sulfate Heptahydrate, NTP TR 471 61 

Sinclair, P.R., Sinclair, J.F., Bonkowsky, H.L., 

Gibbs, A.H., and De Matteis, F. (1982). Formation 

of cobalt protoporphyrin by chicken hepatocytes 

in culture. Relationship to decrease of 

5-aminolaevulinate synthase caused by cobalt. 

Biochem. Pharmacol. 31, 993-999. 

Smith, I.C., and Carson, B.L., Eds. (1981). Cobalt. 

Trace Metals in the Environment, Vol. 6. Ann Arbor 

Science Publishers, Ann Arbor, MI. 

Smith, R.P. (1980). Toxic responses of the blood. In 

Casarett and Doull’s Toxicology, 2nd ed. (J. Doull, 

C.D. Klaassen, and M.O. Amdur, Eds.), pp. 311-331. 

Macmillan Publishing Co., New York. 

Smith, T., Edmonds, C.J., and Barnaby, C.F. (1972). 

Absorption and retention of cobalt in man by wholebody counting. Health Phys. 22, 359-367. 

Speijers, G.J., Krajnc, E.I., Berkvens, J.M., and 

van Logten, M.J. (1982). Acute oral toxicity of 

inorganic cobalt compounds in rats. Food Chem. 

Toxicol. 20, 311-314. 

Stelzer, K.J., and Klaassen, C.D. (1985). Effect of 

cobalt on biliary excretion of bilirubin and glutathione. 

J. Toxicol. Environ. Health 15, 813-822. 

Steinhoff, D., and Mohr, U. (1991). On the question 

of a carcinogenic action of cobalt-containing compounds. Exp. Pathol. 41, 169-174. 

Straus, D.S. (1981). Somatic mutation, cellular 

differentiation, and cancer causation. JNCI 67, 

233-241. 

Swennen, B., Buchet, J.-P., Stánescu, D., Lison, D., 

and Lauwerys, R. (1993). Epidemiological survey of 

workers exposed to cobalt oxides, cobalt salts, and 

cobalt metal. Br. J. Ind. Med. 50, 835-842. 

Tarone, R.E. (1975). Tests for trend in life table 

analysis. Biometrika 62, 679-682. 

Taylor, A., and Marks, V. (1978). Cobalt: A review. 

J. Hum. Nutr. 32, 165-177. 

Telib, M. (1972). Effects of cobaltous chloride in 

laboratory animals: I. The histological and electron 

microscopical changes in the islets of rabbits. 

Endokrinologie 60, 81-102. 

Tennant, R.W., Margolin, B.H., Shelby, M.D., 

Zeiger, E., Haseman, J.K., Spalding, J., Caspary, W., 

Resnick, M., Stasiewicz, S., Anderson, B., and 

Minor, R. (1987). Prediction of chemical 

carcinogenicity in rodents from in vitro genetic 

toxicity assays. Science 236, 933-941. 

Tephly, T.R., and Hibbeln, P. (1971). The effect of 

cobalt chloride administration on the synthesis of 

hepatic microsomal cytochrome P-450. Biochem. 

Biophys. Res. Commun. 42, 589-595. 

U.S. Department of Health and Human Services 

(USDHHS) (1992). Toxicological Profile for Cobalt. 

U.S. Department of Health and Human Services, 

Public Health Service, Agency for Toxic Substances 

and Disease Registry. 

Ward, J.M., Fox, J.G., Anver, M.R., Haines, D.C., 

George, C.V., Collins, M.J., Jr., Gorelick, P.L., 

Nagashima, K.., Gonda, M.A., Gilden, R.V., 

Tully, J.G., Russell, R.J., Benveniste, R.E., 

Paster, B.J., Dewhirst, F.E., Donovan, J.C., 

Anderson, L.M., and Rice, J.M. (1994). Chronic 

active hepatitis and associated liver tumors in mice 

caused by a persistent bacterial infection with a novel 

Helicobacter species. J. Natl. Cancer Inst. 86, 

1222-1227. 

Webb, M., Heath, J.C., and Hopkins, T. (1972). 

Intranuclear distribution of the inducing metal in 

primary rhabdomyosarcomata induced in the rat by 

nickel, cobalt, and cadmium. Br. J. Cancer 26, 

274-278. 

Wehner, A.P., Busch, R.H., Olson, R.J., and 

Craig, D.K. (1977). Chronic inhalation of cobalt 

oxide and cigarette smoke by hamsters. Am. Ind. 

Hyg. Assoc. J. 38, 338-346. 
62 Cobalt Sulfate Heptahydrate, NTP TR 471 

Zeiger, E., Haseman, J.K., Shelby, M.D., Zeiger, E., Anderson, B., Haworth, S., Lawlor, T.,
 

Margolin, B.H., and Tennant, R.W. (1990). Eval- and Mortelmans, K. (1992). Salmonella mutagenicity
 

uation of four in vitro genetic toxicity tests for pre- tests: V. Results from the testing of 311 chemicals.
 

dicting rodent carcinogenicity: Confirmation of earlier Environ. Mol. Mutagen. 19 (Suppl. 21), 2-141.
 

results with 41 additional chemicals. Environ. Mol.
 

Mutagen. 16 (Suppl. 18), 1-14.
 
63 

APPENDIX A 

SUMMARY OF LESIONS IN MALE RATS 

IN THE 2-YEAR INHALATION STUDY 

OF COBALT SULFATE HEPTAHYDRATE 

TABLE A1 

TABLE A2 

TABLE A3 

TABLE A4a 

TABLE A4b 

TABLE A5 

Summary of the Incidence of Neoplasms in Male Rats 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 

Individual Animal Tumor Pathology of Male Rats 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 

Statistical Analysis of Primary Neoplasms in Male Rats 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 

Historical Incidence of Alveolar/bronchiolar Neoplasms 

in Chamber Control Male F344/N Rats .................................. 

Historical Incidence of Neoplasms of the Adrenal Medulla 

in Chamber Control Male F344/N Rats .................................. 

Summary of the Incidence of Nonneoplastic Lesions in Male Rats 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 

64 

68 

92 

99 

99 

100 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

64 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A1 

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydratea 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Moribund 30 34 26 34 

Natural deaths 3 1 3 1 

Survivors 

Terminal sacrifice 17 15 21 15 

Animals examined microscopically 50 50 50 50 

Alimentary System 

Intestine large, colon (49) (50) (49) (49) 

Intestine large, cecum (48) (49) (48) (49) 

Intestine small, duodenum (50) (50) (48) (49) 

Intestine small, jejunum (49) (49) (47) (49) 

Intestine small, ileum (50) (49) (47) (48) 

Liver (50) (50) (48) (50) 

Carcinoma, metastatic, seminal vesicle 1 (2%) 

Hepatocellular carcinoma 1 (2%) 

Hepatocellular adenoma 1 (2%) 1 (2%) 1 (2%) 

Sarcoma, metastatic, uncertain primary site 1 (2%) 

Mesentery (21) (14) (11) (14) 

Carcinoma, metastatic, seminal vesicle 1 (7%) 

Sarcoma, metastatic, uncertain primary site 1 (7%) 

Fat, lipoma 1 (9%) 

Pancreas (50) (50) (48) (50) 

Adenoma 1 (2%) 1 (2%) 

Carcinoma 1 (2%) 

Carcinoma, metastatic, seminal vesicle 1 (2%) 

Mixed tumor malignant 1 (2%) 

Sarcoma, metastatic, uncertain primary site 1 (2%) 

Salivary glands (50) (50) (50) (50) 

Stomach, forestomach (50) (50) (49) (50) 

Stomach, glandular (50) (50) (48) (50) 

Schwannoma malignant 1 (2%) 

Cardiovascular System 

Heart (50) (50) (50) (50) 

Carcinoma, metastatic, seminal vesicle 1 (2%) 

Endocrine System 

Adrenal cortex (50) (50) (48) (50) 

Adenoma 1 (2%) 1 (2%) 

Carcinoma 1 (2%) 1 (2%) 

Adrenal medulla (50) (50) (49) (50) 

Pheochromocytoma malignant 2 (4%) 2 (4%) 2 (4%) 

Pheochromocytoma complex 1 (2%) 1 (2%) 

Pheochromocytoma benign 13 (26%) 15 (30%) 17 (35%) 15 (30%) 

Bilateral, pheochromocytoma benign 1 (2%) 4 (8%) 6 (12%) 5 (10%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 65 

TABLE A1 

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Endocrine System (continued) 

Islets, pancreatic (50) (50) (48) (50) 

Adenoma 1 (2%) 4 (8%) 5 (10%) 5 (10%) 

Adenoma, multiple 1 (2%) 1 (2%) 

Carcinoma 3 (6%) 4 (8%) 4 (8%) 7 (14%) 

Pituitary gland (49) (49) (50) (49) 

Pars distalis, adenoma 43 (88%) 40 (82%) 42 (84%) 41 (84%) 

Thyroid gland (49) (50) (48) (50) 

Bilateral, C-cell, adenoma 1 (2%) 

C-cell, adenoma 4 (8%) 2 (4%) 3 (6%) 3 (6%) 

C-cell, carcinoma 3 (6%) 3 (6%) 4 (8%) 3 (6%) 

Follicular cell, adenoma 1 (2%) 1 (2%) 

Follicular cell, carcinoma 1 (2%) 2 (4%) 

General Body System 

Peritoneum (2) (1) (1) 

Sarcoma, metastatic, uncertain primary site 1 (50%) 

Genital System 

Epididymis (50) (50) (50) (50) 

Preputial gland (50) (50) (49) (50) 

Adenoma 1 (2%) 1 (2%) 

Carcinoma 2 (4%) 2 (4%) 3 (6%) 

Bilateral, adenoma 1 (2%) 

Bilateral, carcinoma 1 (2%) 

Prostate (50) (50) (49) (50) 

Adenoma 1 (2%) 2 (4%) 4 (8%) 1 (2%) 

Seminal vesicle (50) (50) (49) (49) 

Carcinoma 1 (2%) 

Sarcoma, metastatic, uncertain primary site 1 (2%) 

Testes (50) (50) (50) (50) 

Bilateral, interstitial cell, adenoma 26 (52%) 18 (36%) 26 (52%) 19 (38%) 

Interstitial cell, adenoma 9 (18%) 13 (26%) 7 (14%) 10 (20%) 

Hematopoietic System 

Bone marrow (50) (50) (48) (50) 

Lymph node (8) (9) (9) (9) 

Lymph node, bronchial (45) (30) (41) (49) 

Lymph node, mandibular (46) (47) (47) (49) 

Lymph node, mesenteric (50) (50) (48) (50) 

Lymph node, mediastinal (47) (46) (44) (49) 

Carcinoma, metastatic, seminal vesicle 1 (2%) 

Spleen (50) (50) (49) (50) 

Carcinoma, metastatic, seminal vesicle 1 (2%) 

Fibroma 1 (2%) 

Sarcoma, metastatic, uncertain primary site 1 (2%) 

Thymus (45) (42) (47) (48) 

Thymoma benign 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

66 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A1 

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Integumentary System 

Mammary gland (30) (34) (36) (38) 

Fibroadenoma 3 (10%) 1 (3%) 2 (6%) 3 (8%) 

Fibroadenoma, multiple 1 (3%) 

Skin (50) (48) (50) (50) 

Basal cell adenoma 2 (4%) 

Basal cell carcinoma 1 (2%) 

Keratoacanthoma 2 (4%) 5 (10%) 2 (4%) 

Keratoacanthoma, multiple 1 (2%) 

Sarcoma, metastatic, uncertain primary site 1 (2%) 

Squamous cell papilloma 1 (2%) 

Sebaceous gland, adenoma 1 (2%) 1 (2%) 

Subcutaneous tissue, fibroma 1 (2%) 2 (4%) 2 (4%) 

Subcutaneous tissue, fibrosarcoma 2 (4%) 1 (2%) 2 (4%) 

Subcutaneous tissue, lipoma 1 (2%) 1 (2%) 1 (2%) 

Subcutaneous tissue, sarcoma 1 (2%) 

Subcutaneous tissue, schwannoma malignant 1 (2%) 

Musculoskeletal System 

Bone (50) (50) (50) (50) 

Chordoma 1 (2%) 1 (2%) 

Osteosarcoma 3 (6%) 

Skeletal muscle (2) (1) (1) 

Carcinoma, metastatic, seminal vesicle 1 (50%) 

Sarcoma 1 (100%) 

Sarcoma, metastatic, uncertain primary site 1 (50%) 

Nervous System 

Brain (50) (50) (50) (50) 

Astrocytoma malignant 1 (2%) 1 (2%) 

Spinal cord (1) 

Respiratory System 

Larynx (50) (49) (48) (50) 

Lung (50) (50) (48) (50) 

Alveolar/bronchiolar adenoma 1 (2%) 4 (8%) 1 (2%) 5 (10%) 

Alveolar/bronchiolar adenoma, multiple 1 (2%) 

Alveolar/bronchiolar carcinoma 3 (6%) 1 (2%) 

Carcinoma, metastatic, seminal vesicle 1 (2%) 

Chordoma, metastatic, bone 1 (2%) 

Osteosarcoma, metastatic, bone 1 (2%) 

Sarcoma, metastatic, skeletal muscle 1 (2%) 

Nose (50) (50) (49) (50) 

Nasopharyngeal duct, squamous cell carcinoma 1 (2%) 

Pleura (1) 

Trachea (50) (50) (48) (50) 

Special Senses System 

Zymbal’s gland (1) (2) 

Carcinoma 1 (100%) 2 (100%) 
 

 

 

67 

c 

Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A1 

Summary of the Incidence of Neoplasms in Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Urinary System 

Kidney 

Sarcoma, metastatic, uncertain primary site

Renal tubule, adenoma

Urinary bladder 

(50) 

 1 (2%) 

(50) 

(50) 

 1 (2%) 

(50) 

(48) 

 1 (2%) 

(48) 

(50) 

(50) 

Systemic Lesions 

Multiple organsb 

Leukemia mononuclear

Mesothelioma malignant

(50) 

 30 (60%) 

 3 (6%) 

(50) 

 32 (64%) 

 2 (4%) 

(50) 

 29 (58%) 

 2 (4%) 

(50) 

 28 (56%) 

 2 (4%) 

Neoplasm Summary 

Total animals with primary neoplasmsc 

Total primary neoplasms 

Total animals with benign neoplasms 

Total benign neoplasms 

Total animals with malignant neoplasms 

Total malignant neoplasms 

Total animals with metastatic neoplasms 

Total metastatic neoplasms 

Total animals with malignant neoplasms

 of uncertain primary site 

50 

159 

48 

111 

38 

48 

1 

1 

50 

169 

46 

113 

41 

56 

3 

18 

1 

50 

173 

47 

124 

35 

49 

50 

178 

48 

122 

38 

56 

1 

1 

a Number of animals examined microscopically at the site and the number of animals with neoplasm b Number of animals with any tissue examined microscopically 

Primary neoplasms: all neoplasms except metastatic neoplasms 
 

 

 

 

 

 

 

 

 

 

 

 

 

68 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

Number of Days on Study 

4 4 5 5 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 

3 5 2 2 2 3 4 6 7 7 8 8 9 0 0 0 0 1 1 1 1 3 3 4 4 

5 3 6 6 7 4 8 8 6 9 2 9 3 2 2 3 5 0 0 0 9 1 2 3 5 

Carcass ID Number 

0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 

2 1 4 4 1 1 3 3 4 3 0 1 4 2 4 2 4 2 3 4 3 0 1 4 3 

2 5 4 5 6 4 3 8 8 7 4 2 7 1 1 4 9 9 6 6 5 3 7 3 0 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hepatocellular adenoma 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Tongue 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + A + + + + + 

+ A + + + + + + M + + + + + + + + + + A + + + + + 

+ A + + + + + + + + + + + + + + + + + A + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ A + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma complex X 

Pheochromocytoma benign X X X X X 

Bilateral, pheochromocytoma benign X 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma 

Adenoma, multiple 

Carcinoma X X 

Parathyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X X X X X 

Thyroid gland + A + + + + + + + + + + + + + + + + + + + + + + + 

C-cell, adenoma X 

C-cell, carcinoma X 

Follicular cell, carcinoma 

General Body System 

None 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 

Penis + + + 

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Carcinoma 

Bilateral, carcinoma 

+: Tissue examined microscopically M: Missing tissue X: Lesion present 

A: Autolysis precludes examination I: Insufficient tissue Blank: Not examined 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 69 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 4 6 7 7 8 9 9 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

9 4 3 9 0 1 2 7 4 4 4 4 4 4 4 4 4 4 4 4 5 5 5 5 5 

0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 Total 

Carcass ID Number 2 2 2 1 2 0 4 4 0 0 0 0 0 1 1 1 2 3 3 3 0 1 2 3 5 Tissues/ 

7 6 8 8 3 8 0 2 1 5 6 7 9 0 1 9 0 1 2 9 2 3 5 4 0 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hepatocellular adenoma X 1 

Mesentery + + + + + + + + + 21 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Tongue + 1 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pheochromocytoma complex 1 

Pheochromocytoma benign X X X X X X X X 13 

Bilateral, pheochromocytoma benign 1 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X 1 

Adenoma, multiple X 1 

Carcinoma X 3 

Parathyroid gland + + + + + + + + + + + + + + M + + + + + + M + + + 48 

Pituitary gland + + + + + + + + + + + + + + + + + + M + + + + + + 49 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X X X X 43 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

C-cell, adenoma X X X 4 

C-cell, carcinoma X X 3 

Follicular cell, carcinoma X 1 

General Body System 

None 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Penis 3 

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma X X 2 

Bilateral, carcinoma X 1 

50 
 

 

 

 

 

 

 

 

 

 

 

 

 

70 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

4 4 5 5 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 

Number of Days on Study 3 

5 

5 

3 

2 

6 

2 

6 

2 

7 

3 

4 

4 

8 

6 

8 

7 

6 

7 

9 

8 

2 

8 

9 

9 

3 

0 

2 

0 

2 

0 

3 

0 

5 

1 

0 

1 

0 

1 

0 

1 

9 

3 

1 

3 

2 

4 

3 

4 

5 

0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 

Carcass ID Number 2 

2 

1 

5 

4 

4 

4 

5 

1 

6 

1 

4 

3 

3 

3 

8 

4 

8 

3 

7 

0 

4 

1 

2 

4 

7 

2 

1 

4 

1 

2 

4 

4 

9 

2 

9 

3 

6 

4 

6 

3 

5 

0 

3 

1 

7 

4 

3 

3 

0 

Genital System (continued) 

Prostate 

Adenoma 

Seminal vesicle 

Testes 

Bilateral, interstitial cell, adenoma 

Interstitial cell, adenoma 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X X X 

X X X X X X 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Fibroma 

Thymus 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + 

+ + M + + + + + + M + M + + + + + M + + + + + + + 

+ + M + + + + + M + + + + + + + + M M + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + M + + M + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

+ + + M + + + + + + M + + + + + + + + + + + M + M 

Integumentary System 

Mammary gland 

Fibroadenoma 

Skin 

Basal cell adenoma 

Keratoacanthoma 

Sebaceous gland, adenoma 

Subcutaneous tissue, schwannoma, malignant 

M + M M M + M M + + + M M + + M + M M + M M + + M 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

Musculoskeletal System 

Bone 

Chordoma 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Nervous System 

Brain 

Astrocytoma malignant 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Chordoma, metastatic, bone 

Nose 

Pleura 

Trachea 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Special Senses System 

Eye 

Zymbal’s gland 

Carcinoma 

+ 

+ 

X 

+ 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 71 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 4 

9 

6 

4 

7 

3 

7 

9 

8 

0 

9 

1 

9 

2 

2 

7 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 Total 

Carcass ID Number 2 

7 

2 

6 

2 

8 

1 

8 

2 

3 

0 

8 

4 

0 

4 

2 

0 

1 

0 

5 

0 

6 

0 

7 

0 

9 

1 

0 

1 

1 

1 

9 

2 

0 

3 

1 

3 

2 

3 

9 

0 

2 

1 

3 

2 

5 

3 

4 

5 

0 

Tissues/ 

Tumors 

Genital System (continued) 

Prostate 

Adenoma 

Seminal vesicle 

Testes 

Bilateral, interstitial cell, adenoma 

Interstitial cell, adenoma 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X X X X X X X X X X X X X X X 

X X X

50 

1 

50 

50 

26 

9 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Fibroma

Thymus 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ +

+ + M + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + M + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + M + + + + + + + + + + + + + + + + + + + + + 

50 

8 

45 

46 

50 

47 

50 

1 

45 

Integumentary System 

Mammary gland 

Fibroadenoma 

Skin 

Basal cell adenoma 

Keratoacanthoma 

Sebaceous gland, adenoma 

Subcutaneous tissue, schwannoma, malignant 

+ + + + + M M + + + + M + + + + + + M + + + + M M 

X X X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X

X X

X

30 

3 

50 

2 

2 

1 

1 

Musculoskeletal System 

Bone 

Chordoma 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

50 

1 

Nervous System 

Brain 

Astrocytoma malignant 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

50 

1 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma

Chordoma, metastatic, bone 

Nose 

Pleura 

Trachea 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+

+ + + + + + + + + + + + + + + + + + + + + + + + + 

50 

50 

1 

1 

50 

1 

50 

Special Senses System 

Eye 

Zymbal’s gland

Carcinoma

+ + + 5 

1 

1 
 

 

 

72 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

4 4 5 5 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 

Number of Days on Study 3 5 2 2 2 3 4 6 7 7 8 8 9 0 0 0 0 1 1 1 1 3 3 4 4 

5 3 6 6 7 4 8 8 6 9 2 9 3 2 2 3 5 0 0 0 9 1 2 3 5 

0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 

Carcass ID Number 2 1 4 4 1 1 3 3 4 3 0 1 4 2 4 2 4 2 3 4 3 0 1 4 3 

2 5 4 5 6 4 3 8 8 7 4 2 7 1 1 4 9 9 6 6 5 3 7 3 0 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 

Renal tubule, adenoma X 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Leukemia mononuclear X X X X X X X X X X X X X X X 

Mesothelioma malignant X X 
 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 73 

TABLE A2 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 4 6 7 7 8 9 9 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

9 4 3 9 0 1 2 7 4 4 4 4 4 4 4 4 4 4 4 4 5 5 5 5 5 

0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 Total 

Carcass ID Number 2 2 2 1 2 0 4 4 0 0 0 0 0 1 1 1 2 3 3 3 0 1 2 3 5 Tissues/ 

7 6 8 8 3 8 0 2 1 5 6 7 9 0 1 9 0 1 2 9 2 3 5 4 0 Tumors 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Renal tubule, adenoma 1 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Leukemia mononuclear X X X X X X X X X X X X X X X 30 

Mesothelioma malignant X 3 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

74 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

Number of Days on Study 

4 4 4 4 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

0 7 7 9 4 5 7 7 8 8 0 1 2 2 3 3 3 3 3 4 4 4 5 6 6 

8 1 5 5 1 4 6 9 9 9 0 0 9 9 1 1 2 8 8 5 6 9 2 3 9 

Carcass ID Number 

2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 

3 2 0 4 3 0 2 0 2 4 3 3 2 2 3 4 1 1 4 5 0 3 4 1 4 

3 0 7 6 7 6 9 5 7 9 6 0 2 6 9 0 9 1 7 0 4 2 5 5 8 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Carcinoma, metastatic, seminal vesicle 

Hepatocellular carcinoma 

Sarcoma, metastatic, uncertain primary site 

Mesentery 

Carcinoma, metastatic, seminal vesicle 

Sarcoma, metastatic, uncertain primary site 

Pancreas 

Carcinoma, metastatic, seminal vesicle 

Sarcoma, metastatic, uncertain primary site 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + A 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + A 

+ + + + + + + + + + + + + + + + + + + + + + + + A 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

X 

X 

+ + + + + + 

X 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 

Carcinoma, metastatic, seminal vesicle X 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 

Carcinoma X 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma malignant 

Pheochromocytoma benign X X X X X 

Bilateral, pheochromocytoma benign X 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma X X 

Adenoma, multiple 

Carcinoma 

Parathyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

C-cell, adenoma 

C-cell, carcinoma X 

Follicular cell, adenoma 

Follicular cell, carcinoma 

General Body System 

Peritoneum + 

Sarcoma, metastatic, uncertain primary site X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 75 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 7 7 8 8 8 9 9 0 1 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

1 8 0 0 1 1 4 1 9 2 4 4 4 4 4 4 4 4 4 4 5 5 5 5 5 

2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 Total 

Carcass ID Number 1 3 1 2 4 2 2 1 1 1 0 0 1 1 2 2 3 3 4 4 0 0 0 3 4 Tissues/ 

0 1 3 4 2 5 8 8 6 7 2 8 2 4 1 3 5 8 3 4 1 3 9 4 1 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma, metastatic, seminal vesicle 1 

Hepatocellular carcinoma 1 

Sarcoma, metastatic, uncertain primary site 1 

Mesentery + + + + + + + + 14 

Carcinoma, metastatic, seminal vesicle 1 

Sarcoma, metastatic, uncertain primary site 1 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma, metastatic, seminal vesicle 1 

Sarcoma, metastatic, uncertain primary site 1 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma, metastatic, seminal vesicle 1 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma 1 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pheochromocytoma malignant X X 2 

Pheochromocytoma benign X X X X X X X X X X 15 

Bilateral, pheochromocytoma benign X X X 4 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X X 4 

Adenoma, multiple X 1 

Carcinoma X X X X 4 

Parathyroid gland + + + + + + + + + + + + + + + + + M + + + + M + + 48 

Pituitary gland M + + + + + + + + + + + + + + + + + + + + + + + + 49 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X X X X X X 40 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

C-cell, adenoma X X 2 

C-cell, carcinoma X X 3 

Follicular cell, adenoma X 1 

Follicular cell, carcinoma X X 2 

General Body System 

Peritoneum + 2 

Sarcoma, metastatic, uncertain primary site 1 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

76 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

4 4 4 4 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

Number of Days on Study 0 7 7 9 4 5 7 7 8 8 0 1 2 2 3 3 3 3 3 4 4 4 5 6 6 

8 1 5 5 1 4 6 9 9 9 0 0 9 9 1 1 2 8 8 5 6 9 2 3 9 

2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 

Carcass ID Number 3 2 0 4 3 0 2 0 2 4 3 3 2 2 3 4 1 1 4 5 0 3 4 1 4 

3 0 7 6 7 6 9 5 7 9 6 0 2 6 9 0 9 1 7 0 4 2 5 5 8 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Carcinoma 

Prostate + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma 

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + + 

Carcinoma X 

Sarcoma, metastatic, uncertain primary site X 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 

Bilateral, interstitial cell, adenoma X X X X X X X X 

Interstitial cell, adenoma X X X X 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 

Lymph node + + + + + + 

Lymph node, bronchial M + + + M M + + M + + M + + + + + M + + + M + M + 

Lymph node, mandibular + + + + + + M + + + + + + + + + + + + + M + + + + 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + 

Lymph node, mediastinal + + + + + + + + + + + M + + + + + + M + + + + + + 

Carcinoma, metastatic, seminal vesicle X 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 

Carcinoma, metastatic, seminal vesicle X 

Sarcoma, metastatic, uncertain primary site X 

Thymus + + + + + + + + + + + + + + M + + M + + + + + + + 

Integumentary System 

Mammary gland + M M + + + + M + M + + + + M M M + M + M + + M + 

Fibroadenoma 

Skin + + + + + + + I + + + + + + + + + + + + + + + I + 

Keratoacanthoma X X X 

Sarcoma, metastatic, uncertain primary site X 

Sebaceous gland, adenoma 

Subcutaneous tissue, fibroma 

Subcutaneous tissue, fibrosarcoma X 

Subcutaneous tissue, lipoma 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Chordoma X 

Osteosarcoma X 

Skeletal muscle + + 

Carcinoma, metastatic, seminal vesicle X 

Sarcoma, metastatic, uncertain primary site X 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 77 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 7 7 8 8 8 9 9 0 1 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

1 8 0 0 1 1 4 1 9 2 4 4 4 4 4 4 4 4 4 4 5 5 5 5 5 

2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 Total 

Carcass ID Number 1 3 1 2 4 2 2 1 1 1 0 0 1 1 2 2 3 3 4 4 0 0 0 3 4 Tissues/ 

0 1 3 4 2 5 8 8 6 7 2 8 2 4 1 3 5 8 3 4 1 3 9 4 1 Tumors 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma X X 2 

Prostate + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X X 2 

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma 1 

Sarcoma, metastatic, uncertain primary site 1 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Bilateral, interstitial cell, adenoma X X X X X X X X X X 18 

Interstitial cell, adenoma X X X X X X X X X 13 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymph node + + + 9 

Lymph node, bronchial M + + + + M M M + M + M M + + M + M M + + M + M + 30 

Lymph node, mandibular + + + + + M + + + + + + + + + + + + + + + + + + + 47 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymph node, mediastinal + + + + + + + + + + + M + + + M + + + + + + + + + 46 

Carcinoma, metastatic, seminal vesicle 1 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma, metastatic, seminal vesicle 1 

Sarcoma, metastatic, uncertain primary site 1 

Thymus + M + M + + + + M + + + + + + + + + + M + + + M M 42 

Integumentary System 

Mammary gland + + + + M M + M + + + + + + M M + + + + + + + + M 34 

Fibroadenoma X 1 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Keratoacanthoma X X 5 

Sarcoma, metastatic, uncertain primary site 1 

Sebaceous gland, adenoma X 1 

Subcutaneous tissue, fibroma X 1 

Subcutaneous tissue, fibrosarcoma X 2 

Subcutaneous tissue, lipoma X 1 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Chordoma 1 

Osteosarcoma X X 3 

Skeletal muscle 2 

Carcinoma, metastatic, seminal vesicle 1 

Sarcoma, metastatic, uncertain primary site 1 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 
 

 

 

 

 

 

 

78 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

Number of Days on Study 

4 

0 

8 

4 

7 

1 

4 

7 

5 

4 

9 

5 

5 

4 

1 

5 

5 

4 

5 

7 

6 

5 

7 

9 

5 

8 

9 

5 

8 

9 

6 

0 

0 

6 

1 

0 

6 

2 

9 

6 

2 

9 

6 

3 

1 

6 

3 

1 

6 

3 

2 

6 

3 

8 

6 

3 

8 

6 

4 

5 

6 

4 

6 

6 

4 

9 

6 

5 

2 

6 

6 

3 

6 

6 

9 

Carcass ID Number 

2 

3 

3 

2 

2 

0 

2 

0 

7 

2 

4 

6 

2 

3 

7 

2 

0 

6 

2 

2 

9 

2 

0 

5 

2 

2 

7 

2 

4 

9 

2 

3 

6 

2 

3 

0 

2 

2 

2 

2 

2 

6 

2 

3 

9 

2 

4 

0 

2 

1 

9 

2 

1 

1 

2 

4 

7 

2 

5 

0 

2 

0 

4 

2 

3 

2 

2 

4 

5 

2 

1 

5 

2 

4 

8 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Carcinoma, metastatic, seminal vesicle 

Osteosarcoma, metastatic, bone 

Nose 

Trachea 

Special Senses System 

None 

+ + + + + + + + + + + + + + + + + + + + + + + + A 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X 

X 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Urinary System 

Kidney 

Sarcoma, metastatic, uncertain primary site 

Urinary bladder 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Mesothelioma malignant 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X X X X X X X X X X X X X 
 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 79 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 7 7 8 8 8 9 9 0 1 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

1 8 0 0 1 1 4 1 9 2 4 4 4 4 4 4 4 4 4 4 5 5 5 5 5 

2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 Total 

Carcass ID Number 1 3 1 2 4 2 2 1 1 1 0 0 1 1 2 2 3 3 4 4 0 0 0 3 4 Tissues/ 

0 1 3 4 2 5 8 8 6 7 2 8 2 4 1 3 5 8 3 4 1 3 9 4 1 Tumors 

Respiratory System 

Larynx + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Alveolar/bronchiolar adenoma X X 4 

Carcinoma, metastatic, seminal vesicle 1 

Osteosarcoma, metastatic, bone 1 

Nose + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Special Senses System 

None 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Sarcoma, metastatic, uncertain primary site 1 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Leukemia mononuclear X X X X X X X X X X X X X X X 32 

Mesothelioma malignant X X 2 
 

 

 

 

 

 

 

 

 

 

 

 

 

80 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

2 4 4 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

Number of Days on Study 7 3 6 0 1 2 3 8 8 1 1 3 3 3 4 4 5 5 5 7 7 7 7 8 9 

2 5 7 9 8 6 4 1 2 1 1 0 1 2 2 9 2 4 7 1 3 3 8 1 4 

4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 

Carcass ID Number 2 0 3 2 4 2 1 0 0 0 4 4 3 3 4 5 1 1 1 3 2 2 2 3 0 

0 6 8 1 4 2 2 3 1 9 6 8 7 3 1 0 1 7 3 9 5 7 4 4 7 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 

Intestine large, colon + + + + A + + + + + + + + + + + + + + + + + + + + 

Intestine large, rectum + + + + A + + + + + + + + + + + + + + + + + + + + 

Intestine large, cecum + + + + A + + A + + + + + + + + + + + + + + + + + 

Intestine small, duodenum + + + + A + + A + + + + + + + + + + + + + + + + + 

Intestine small, jejunum A + + + A + + A + + + + + + + + + + + + + + + + + 

Intestine small, ileum A + + + A + + A + + + + + + + + + + + + + + + + + 

Liver + + + + A + + A + + + + + + + + + + + + + + + + + 

Hepatocellular adenoma X 

Mesentery + + + + + + 

Fat, lipoma X 

Pancreas + + + + A + + A + + + + + + + + + + + + + + + + + 

Adenoma X 

Carcinoma 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 

Stomach, forestomach + + + + A + + + + + + + + + + + + + + + + + + + + 

Stomach, glandular + + + + A + + A + + + + + + + + + + + + + + + + + 

Tooth 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 

Endocrine System 

Adrenal cortex + + + + A + + A + + + + + + + + + + + + + + + + + 

Adenoma X 

Carcinoma X 

Adrenal medulla + + + + A + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma malignant X 

Pheochromocytoma complex 

Pheochromocytoma benign X X X X X X X 

Bilateral, pheochromocytoma benign X X 

Islets, pancreatic + + + + A + + A + + + + + + + + + + + + + + + + + 

Adenoma X X 

Carcinoma X X X 

Parathyroid gland + + + + + + + + + + + + + + + + + + + M + + + + + 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X X X X X 

Thyroid gland + + + + A + + A + + + + + + + + + + + + + + + + + 

C-cell, adenoma X 

C-cell, carcinoma X X 

General Body System 

Peritoneum + 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 81 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 9 0 1 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

9 9 3 9 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 5 5 5 5 

4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 Total 

Carcass ID Number 4 1 0 3 0 0 0 1 1 1 1 2 2 3 3 3 3 4 4 4 4 1 2 2 4 Tissues/ 

0 8 5 2 2 4 8 0 4 5 9 6 9 0 1 5 6 2 3 5 9 6 3 8 7 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Hepatocellular adenoma 1 

Mesentery + + + + + 11 

Fat, lipoma 1 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Adenoma 1 

Carcinoma X 1 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Tooth + 1 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Adenoma 1 

Carcinoma 1 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Pheochromocytoma malignant X 2 

Pheochromocytoma complex X 1 

Pheochromocytoma benign X X X X X X X X X X 17 

Bilateral, pheochromocytoma benign X X X X 6 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Adenoma X X X 5 

Carcinoma X 4 

Parathyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X X X 42 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 48 

C-cell, adenoma X X 3 

C-cell, carcinoma X X 4 

General Body System 

Peritoneum 1 
 

 

 

 

 

 

 

 

 

 

 

82 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

2 4 4 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

Number of Days on Study 7 3 6 0 1 2 3 8 8 1 1 3 3 3 4 4 5 5 5 7 7 7 7 8 9 

2 5 7 9 8 6 4 1 2 1 1 0 1 2 2 9 2 4 7 1 3 3 8 1 4 

4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 

Carcass ID Number 2 0 3 2 4 2 1 0 0 0 4 4 3 3 4 5 1 1 1 3 2 2 2 3 0 

0 6 8 1 4 2 2 3 1 9 6 8 7 3 1 0 1 7 3 9 5 7 4 4 7 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 

Preputial gland + + + + A + + + + + + + + + + + + + + + + + + + + 

Adenoma X 

Bilateral, adenoma X 

Prostate + + + + A + + + + + + + + + + + + + + + + + + + + 

Adenoma X 

Seminal vesicle + + + + A + + + + + + + + + + + + + + + + + + + + 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 

Bilateral, interstitial cell, adenoma X X X X X X X X X X X 

Interstitial cell, adenoma X X 

Hematopoietic System 

Bone marrow + + + + A + + A + + + + + + + + + + + + + + + + + 

Lymph node + + + + 

Lymph node, bronchial M + + + A + + + + + + + + + + + + M M + + + + + M 

Lymph node, mandibular + + + + + M + + + + + + + + + + + + + + M + + + + 

Lymph node, mesenteric + + + + A + + + + + + + + + + + + + + + + + + + + 

Lymph node, mediastinal + + + + A M + A + + + + + + + + + + M + + + + + M 

Spleen + + + + A + + + + + + + + + + + + + + + + + + + + 

Thymus + + + + A M + + + + + + + + + + + + + + + M + + + 

Thymoma benign 

Integumentary System 

Mammary gland + + + + + + M M + M M + M + + + + + + + M + + M + 

Fibroadenoma X 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 

Subcutaneous tissue, fibroma X 

Subcutaneous tissue, fibrosarcoma X 

Subcutaneous tissue, lipoma X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Skeletal muscle + 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 

Astrocytoma malignant X 

Spinal cord + 

Respiratory System 

Larynx I + + + + + + A + + + + + + + + + + + + + + + + + 

Lung + + + + A + + A + + + + + + + + + + + + + + + + + 

Alveolar/bronchiolar adenoma X 

Alveolar/bronchiolar carcinoma X 

Nose + + + + A + + + + + + + + + + + + + + + + + + + + 

Trachea + + + + A + + A + + + + + + + + + + + + + + + + + 

Special Senses System 

Eye 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 83 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 9 

9 

0 

9 

1 

3 

2 

9 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

5 

3 

5 

3 

5 

3 

5 

4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 Total 

Carcass ID Number 4 

0 

1 

8 

0 

5 

3 

2 

0 

2 

0 

4 

0 

8 

1 

0 

1 

4 

1 

5 

1 

9 

2 

6 

2 

9 

3 

0 

3 

1 

3 

5 

3 

6 

4 

2 

4 

3 

4 

5 

4 

9 

1 

6 

2 

3 

2 

8 

4 

7 

Tissues/ 

Tumors 

Genital System 

Epididymis 

Preputial gland 

Adenoma

Bilateral, adenoma

Prostate 

Adenoma 

Seminal vesicle 

Testes 

Bilateral, interstitial cell, adenoma 

Interstitial cell, adenoma 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X X X X X X X X X X X 

X X X X X

50 

49 

1 

1 

49 

4 

49 

50 

26 

7 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Thymus 

Thymoma benign 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + +

+ + + + + + + M + + M + + + + + + M + + + + + M + 

+ + + + + + + + + + + + + + + + + + + + + + + + M 

+ + + + + + + + + + + + + + + + + + + + + M + + + 

+ + + + + + + + + + + + + + + + + + M + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

48 

9 

41 

47 

48 

44 

49 

47 

1 

Integumentary System 

Mammary gland 

Fibroadenoma 

Skin 

Subcutaneous tissue, fibroma 

Subcutaneous tissue, fibrosarcoma

Subcutaneous tissue, lipoma

+ + M + M + + M + + + + M M + + M + + + + + + + M 

X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

36 

2 

50 

2 

1 

1 

Musculoskeletal System 

Bone 

Skeletal muscle

+ + + + + + + + + + + + + + + + + + + + + + + + + 50 

1 

Nervous System 

Brain 

Astrocytoma malignant

Spinal cord

+ + + + + + + + + + + + + + + + + + + + + + + + + 50 

1 

1 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma

Alveolar/bronchiolar carcinoma 

Nose 

Trachea 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

48 

48 

1 

3 

49 

48 

Special Senses System 

Eye + 1 
 

 

 

84 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

2 4 4 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

Number of Days on Study 7 3 6 0 1 2 3 8 8 1 1 3 3 3 4 4 5 5 5 7 7 7 7 8 9 

2 5 7 9 8 6 4 1 2 1 1 0 1 2 2 9 2 4 7 1 3 3 8 1 4 

4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 

Carcass ID Number 2 0 3 2 4 2 1 0 0 0 4 4 3 3 4 5 1 1 1 3 2 2 2 3 0 

0 6 8 1 4 2 2 3 1 9 6 8 7 3 1 0 1 7 3 9 5 7 4 4 7 

Urinary System 

Kidney + + + + A + + A + + + + + + + + + + + + + + + + + 

Renal tubule, adenoma X 

Urinary bladder + + + + A + + A + + + + + + + + + + + + + + + + + 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Leukemia mononuclear X X X X X X X X X X X X X X 

Mesothelioma malignant X X 
 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 85 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 9 0 1 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

9 9 3 9 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 5 5 5 5 

4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 Total 

Carcass ID Number 4 1 0 3 0 0 0 1 1 1 1 2 2 3 3 3 3 4 4 4 4 1 2 2 4 Tissues/ 

0 8 5 2 2 4 8 0 4 5 9 6 9 0 1 5 6 2 3 5 9 6 3 8 7 Tumors 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Renal tubule, adenoma 1 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Leukemia mononuclear X X X X X X X X X X X X X X X 29 

Mesothelioma malignant 2 
 

 

 

 

 

 

 

 

 

 

 

 

86 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

Number of Days on Study 

4 4 4 5 5 5 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 

0 6 6 1 2 3 4 6 6 6 7 7 8 8 9 0 0 1 1 1 3 3 3 4 4 

1 3 3 1 6 4 5 2 2 7 4 9 6 9 3 3 4 1 1 1 2 8 8 3 6 

Carcass ID Number 

6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

2 1 1 0 1 2 0 2 3 4 3 1 3 0 2 3 3 4 4 4 1 2 3 2 1 

8 2 5 7 8 1 4 0 0 1 7 9 4 1 5 5 3 5 7 9 4 4 9 3 3 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hepatocellular adenoma 

Mesentery 

Pancreas 

Adenoma 

Mixed tumor malignant 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Schwannoma malignant 

Tongue 

Tooth 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + A + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + A + + + + + + + + + 

+ + + + + + + + + + + + + + + A + + + + + + + + + 

+ + + + + + + + + + + + + + + A + + + + + + + + + 

+ + + + + + + + + + + + + + + A + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Cardiovascular System 

Blood vessel + 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma X 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma malignant 

Pheochromocytoma benign X X X X 

Bilateral, pheochromocytoma benign X X 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma X X 

Carcinoma X 

Parathyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Pituitary gland + + M + + + + + + + + + + + + + + + + + + + + + + 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X X 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Bilateral, C-cell, adenoma 

C-cell, adenoma X 

C-cell, carcinoma X 

Follicular cell, adenoma 

General Body System 

Peritoneum 
 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 87 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 6 6 6 7 8 9 9 9 0 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

3 6 9 3 0 2 3 4 6 0 4 4 4 4 4 4 4 4 4 4 5 5 5 5 5 

6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 Total 

Carcass ID Number 4 3 0 4 1 5 4 0 0 2 0 1 1 1 2 3 3 4 4 4 0 0 2 2 3 Tissues/ 

3 1 8 4 0 0 8 5 6 2 9 1 6 7 9 2 8 0 2 6 2 3 6 7 6 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine small, ileum + + + + I + + + + + + + + + + + + + + + + + + + + 48 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hepatocellular adenoma X 1 

Mesentery + + + + + + + 14 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X 1 

Mixed tumor malignant X 1 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Schwannoma malignant X 1 

Tongue + + 2 

Tooth + 1 

Cardiovascular System 

Blood vessel 1 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma 1 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pheochromocytoma malignant X X 2 

Pheochromocytoma benign X X X X X X X X X X X 15 

Bilateral, pheochromocytoma benign X X X 5 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X X X 5 

Carcinoma X X X X X X 7 

Parathyroid gland + + + M + + + + + + + + + + + + + + + + + + + + + 49 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X X X X X 41 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Bilateral, C-cell, adenoma X 1 

C-cell, adenoma X X 3 

C-cell, carcinoma X X 3 

Follicular cell, adenoma X 1 

General Body System 

Peritoneum + 1 
 

 

 

 

 

 

 

 

 

 

88 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

4 4 4 5 5 5 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 

Number of Days on Study 0 6 6 1 2 3 4 6 6 6 7 7 8 8 9 0 0 1 1 1 3 3 3 4 4 

1 3 3 1 6 4 5 2 2 7 4 9 6 9 3 3 4 1 1 1 2 8 8 3 6 

6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

Carcass ID Number 2 1 1 0 1 2 0 2 3 4 3 1 3 0 2 3 3 4 4 4 1 2 3 2 1 

8 2 5 7 8 1 4 0 0 1 7 9 4 1 5 5 3 5 7 9 4 4 9 3 3 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 

Penis + + 

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma X 

Carcinoma X 

Prostate + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma 

Seminal vesicle + + + + + + + + + + + + + + + A + + + + + + + + + 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 

Bilateral, interstitial cell, adenoma X X X X X 

Interstitial cell, adenoma X X X X 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 

Lymph node + + + + + 

Lymph node, bronchial + + + M + + + + + + + + + + + + + + + + + + + + + 

Lymph node, mandibular + + + + + + + + + + + + + + + + + + + + + + + M + 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + 

Lymph node, mediastinal + + + + + + + + + + + + + + + + + + + + + + + + + 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 

Thymus + + + + + + + + + + + + + + + M + + + + + + + + + 

Integumentary System 

Mammary gland + M M M + + M M + + M + + M + + + + + + M + + + M 

Fibroadenoma X 

Fibroadenoma, multiple 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 

Basal cell carcinoma 

Keratoacanthoma 

Keratoacanthoma, multiple 

Squamous cell papilloma 

Subcutaneous tissue, fibroma 

Subcutaneous tissue, fibrosarcoma 

Subcutaneous tissue, lipoma 

Subcutaneous tissue, sarcoma 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Skeletal muscle + 

Sarcoma X 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 
 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 89 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 6 6 6 7 8 9 9 9 0 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

3 6 9 3 0 2 3 4 6 0 4 4 4 4 4 4 4 4 4 4 5 5 5 5 5 

6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 Total 

Carcass ID Number 4 3 0 4 1 5 4 0 0 2 0 1 1 1 2 3 3 4 4 4 0 0 2 2 3 Tissues/ 

3 1 8 4 0 0 8 5 6 2 9 1 6 7 9 2 8 0 2 6 2 3 6 7 6 Tumors 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Penis 2 

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma 1 

Carcinoma X X 3 

Prostate + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X 1 

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Bilateral, interstitial cell, adenoma X X X X X X X X X X X X X X 19 

Interstitial cell, adenoma X X X X X X 10 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymph node + + + + 9 

Lymph node, bronchial + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Lymph node, mandibular + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymph node, mediastinal + + + + + + + M + + + + + + + + + + + + + + + + + 49 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Thymus + + + + + + + M + + + + + + + + + + + + + + + + + 48 

Integumentary System 

Mammary gland + + + + + + + + M + M + + + + + + + + + + + + + M 38 

Fibroadenoma X X 3 

Fibroadenoma, multiple X 1 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Basal cell carcinoma X 1 

Keratoacanthoma X X 2 

Keratoacanthoma, multiple X 1 

Squamous cell papilloma X 1 

Subcutaneous tissue, fibroma X X 2 

Subcutaneous tissue, fibrosarcoma X X 2 

Subcutaneous tissue, lipoma X 1 

Subcutaneous tissue, sarcoma X 1 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Skeletal muscle 1 

Sarcoma 1 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 
 

 

 

 

 

 

 

 

90 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

4 4 4 5 5 5 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 

Number of Days on Study 0 

1 

6 

3 

6 

3 

1 

1 

2 

6 

3 

4 

4 

5 

6 

2 

6 

2 

6 

7 

7 

4 

7 

9 

8 

6 

8 

9 

9 

3 

0 

3 

0 

4 

1 

1 

1 

1 

1 

1 

3 

2 

3 

8 

3 

8 

4 

3 

4 

6 

6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

Carcass ID Number 2 

8 

1 

2 

1 

5 

0 

7 

1 

8 

2 

1 

0 

4 

2 

0 

3 

0 

4 

1 

3 

7 

1 

9 

3 

4 

0 

1 

2 

5 

3 

5 

3 

3 

4 

5 

4 

7 

4 

9 

1 

4 

2 

4 

3 

9 

2 

3 

1 

3 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma 

Sarcoma, metastatic, skeletal muscle 

Nose 

Nasopharyngeal duct, squamous cell carcinoma 

Trachea 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Special Senses System 

Eye 

Zymbal’s gland 

Carcinoma 

+ 

X 

+ 

X 

Urinary System 

Kidney 

Urinary bladder 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Mesothelioma malignant 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X X X X X X X X X X X 

X 
 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 91 

TABLE A2
 

Individual Animal Tumor Pathology of Male Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 6 6 6 7 8 9 9 9 0 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

3 6 9 3 0 2 3 4 6 0 4 4 4 4 4 4 4 4 4 4 5 5 5 5 5 

6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 Total 

Carcass ID Number 4 3 0 4 1 5 4 0 0 2 0 1 1 1 2 3 3 4 4 4 0 0 2 2 3 Tissues/ 

3 1 8 4 0 0 8 5 6 2 9 1 6 7 9 2 8 0 2 6 2 3 6 7 6 Tumors 

Respiratory System 

Larynx + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Alveolar/bronchiolar adenoma X X X X 5 

Alveolar/bronchiolar adenoma, multiple X 1 

Alveolar/bronchiolar carcinoma X 1 

Sarcoma, metastatic, skeletal muscle 1 

Nose + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Nasopharyngeal duct, squamous cell carcinoma 1 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Special Senses System 

Eye + 1 

Zymbal’s gland 2 

Carcinoma 2 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Leukemia mononuclear X X X X X X X X X X X X X 28 

Mesothelioma malignant X 2 
 

92 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Adrenal Medulla: Benign Pheochromocytoma a Overall rate 14/50 (28%) 19/50 (38%) 23/49 (47%) 20/50 (40%) 

Adjusted rateb 51.0% 70.0% 71.9% 71.4% c Terminal rate 6/17 (35%) 8/15 (53%) 13/21 (62%) 8/15 (53%) 

First incidence (days) 534 541 526 526 

d Life table test Pù0.166 Pù0.220 Pù0.214 Pù0.134 

d Logistic regression test Pù0.172 Pù0.226 Pù0.069 Pù0.126 

d Cochran-Armitage test Pù0.229 

d Fisher exact test Pù0.198 Pù0.041 Pù0.146 

Adrenal Medulla: Benign, Complex, or Malignant Pheochromocytoma 

Overall rate 15/50 (30%) 19/50 (38%) 25/49 (51%) 20/50 (40%) 

Adjusted rate 52.1% 70.0% 74.1% 71.4% 

Terminal rate 6/17 (35%) 8/15 (53%) 13/21 (62%) 8/15 (53%) 

First incidence (days) 534 541 526 526 

Life table test Pù0.206 Pù0.285 Pù0.188 Pù0.182 

Logistic regression test Pù0.218 Pù0.295 Pù0.045 Pù0.180 

Cochran-Armitage test Pù0.279 

Fisher exact test Pù0.263 Pù0.027 Pù0.201 

Bone: Osteosarcoma 

Overall rate 0/50 (0%) 3/50 (6%) 0/50 (0%) 0/50 (0%) 

Adjusted rate 0.0% 11.4% 0.0% 0.0% 

Terminal rate 0/17 (0%) 0/15 (0%) 0/21 (0%) 0/15 (0%) e First incidence (days) — 631 — —
 

Life table test Pù0.258N Pù0.146 — —
 

Logistic regression test Pù0.257N Pù0.123 — —
 

Cochran-Armitage test Pù0.255N
 

Fisher exact test Pù0.121 — —
 

Lung: Alveolar/bronchiolar Adenoma 

Overall rate 1/50 (2%) 4/50 (8%) 1/48 (2%) 6/50 (12%) 

Adjusted rate 2.3% 17.7% 2.4% 28.4% 

Terminal rate 0/17 (0%) 2/15 (13%) 0/21 (0%) 2/15 (13%) 

First incidence (days) 568 589 611 638 

Life table test Pù0.042 Pù0.187 Pù0.726N Pù0.056 

Logistic regression test Pù0.051 Pù0.179 Pù0.753 Pù0.055 

Cochran-Armitage test Pù0.055 

Fisher exact test Pù0.181 Pù0.742 Pù0.056 

Lung: Alveolar/bronchiolar Carcinoma 

Overall rate 0/50 (0%) 0/50 (0%) 3/48 (6%) 1/50 (2%)
 

Adjusted rate 0.0% 0.0% 11.3% 6.7%
 

Terminal rate 0/17 (0%) 0/15 (0%) 1/21 (5%) 1/15 (7%)
 

First incidence (days) — — 652 734 (T)
 

Life table test Pù0.355 — Pù0.181 Pù0.475 

Logistic regression test Pù0.360 — Pù0.136 Pù0.475 

Cochran-Armitage test Pù0.382 

Fisher exact test — Pù0.114 Pù0.500 
Cobalt Sulfate Heptahydrate, NTP TR 471 93 

TABLE A3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Lung: Alveolar/bronchiolar Adenoma or Carcinoma 

Overall rate 1/50 (2%) 4/50 (8%) 4/48 (8%) 7/50 (14%) 

Adjusted rate 2.3% 17.7% 13.4% 33.9% 

Terminal rate 0/17 (0%) 2/15 (13%) 1/21 (5%) 3/15 (20%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

568 

Pù0.027 

Pù0.032 

Pù0.038 

589 

Pù0.187 

Pù0.179 

Pù0.181 

611 

Pù0.263 

Pù0.163 

Pù0.168 

638 

Pù0.030 

Pù0.029 

Pù0.030 

Mammary Gland: Fibroadenoma 

Overall rate 3/50 (6%) 1/50 (2%) 2/50 (4%) 4/50 (8%) 

Adjusted rate 17.6% 6.7% 7.6% 21.2% 

Terminal rate 3/17 (18%) 1/15 (7%) 1/21 (5%) 2/15 (13%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

734 (T) 

Pù0.199 

Pù0.203 

Pù0.240 

734 (T) 

Pù0.346N 

Pù0.346N 

Pù0.309N 

652 

Pù0.391N 

Pù0.398N 

Pù0.500N 

611 

Pù0.449 

Pù0.475 

Pù0.500 

Pancreatic Islets: Adenoma 

Overall rate 2/50 (4%) 5/50 (10%) 5/48 (10%) 5/50 (10%) 

Adjusted rate 10.2% 21.3% 18.3% 20.5% 

Terminal rate 1/17 (6%) 2/15 (13%) 3/21 (14%) 1/15 (7%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

679 

Pù0.278 

Pù0.304 

Pù0.316 

471 

Pù0.217 

Pù0.224 

Pù0.218 

509 

Pù0.301 

Pù0.226 

Pù0.201 

611 

Pù0.222 

Pù0.208 

Pù0.218 

Pancreatic Islets: Carcinoma 

Overall rate 3/50 (6%) 4/50 (8%) 4/48 (8%) 7/50 (14%) 

Adjusted rate 11.4% 19.8% 14.3% 33.2% 

Terminal rate 1/17 (6%) 1/15 (7%) 1/21 (5%) 2/15 (13%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

526 

Pù0.089 

Pù0.091 

Pù0.110 

680 

Pù0.501 

Pù0.515 

Pù0.500 

657 

Pù0.611 

Pù0.489 

Pù0.477 

638 

Pù0.149 

Pù0.149 

Pù0.159 

Pancreatic Islets: Adenoma or Carcinoma 

Overall rate 5/50 (10%) 9/50 (18%) 9/48 (19%) 11/50 (22%) 

Adjusted rate 20.7% 37.6% 30.6% 44.7% 

Terminal rate 2/17 (12%) 3/15 (20%) 4/21 (19%) 3/15 (20%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

526 

Pù0.101 

Pù0.107 

Pù0.127 

471 

Pù0.202 

Pù0.205 

Pù0.194 

509 

Pù0.328 

Pù0.192 

Pù0.172 

611 

Pù0.088 

Pù0.077 

Pù0.086 
94 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Pituitary Gland (Pars Distalis): Adenoma 

Overall rate 43/49 (88%) 40/49 (82%) 42/50 (84%) 41/49 (84%) 

Adjusted rate 97.4% 100.0% 89.3% 95.1% 

Terminal rate 15/16 (94%) 15/15 (100%) 16/21 (76%) 13/15 (87%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

435 

Pù0.455 

Pù0.485N 

Pù0.474N 

471 

Pù0.340N 

Pù0.269N 

Pù0.288N 

467 

Pù0.121N 

Pù0.376N 

Pù0.403N 

401 

Pù0.509N 

Pù0.386N 

Pù0.387N 

Preputial Gland: Carcinoma 

Overall rate 3/50 (6%) 2/50 (4%) 0/49 (0%) 3/50 (6%) 

Adjusted rate 14.7% 11.1% 0.0% 12.8% 

Terminal rate 1/17 (6%) 0/15 (0%) 0/21 (0%) 1/15 (7%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

679 

Pù0.472 

Pù0.486 

Pù0.500 

701 

Pù0.512N 

Pù0.229N 

Pù0.500N 

— 

Pù0.087N 

Pù0.093N 

Pù0.125N 

511 

Pù0.632 

Pù0.657 

Pù0.661N 

Preputial Gland: Adenoma or Carcinoma 

Overall rate 3/50 (6%) 2/50 (4%) 2/49 (4%) 4/50 (8%) 

Adjusted rate 14.7% 11.1% 6.8% 15.8% 

Terminal rate 1/17 (6%) 0/15 (0%) 0/21 (0%) 1/15 (7%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

679 

Pù0.292 

Pù0.305 

Pù0.320 

701 

Pù0.512N 

Pù0.229N 

Pù0.500N 

654 

Pù0.380N 

Pù0.329N 

Pù0.510N 

511 

Pù0.478 

Pù0.494 

Pù0.500 

Prostate Gland: Adenoma 

Overall rate 1/50 (2%) 2/50 (4%) 4/49 (8%) 1/50 (2%) 

Adjusted rate 5.9% 10.7% 17.1% 6.7% 

Terminal rate 1/17 (6%) 1/15 (7%) 3/21 (14%) 1/15 (7%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

734 (T) 

Pù0.537N 

Pù0.533N 

Pù0.500N 

680 

Pù0.481 

Pù0.508 

Pù0.500 

673 

Pù0.250 

Pù0.251 

Pù0.175 

734 (T) 

Pù0.736 

Pù0.736 

Pù0.753N 

Skin: Keratoacanthoma 

Overall rate 2/50 (4%) 5/50 (10%) 0/50 (0%) 3/50 (6%) 

Adjusted rate 11.1% 17.8% 0.0% 20.0% 

Terminal rate 1/17 (6%) 1/15 (7%) 0/21 (0%) 3/15 (20%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

727 

Pù0.602 

Pù0.608N 

Pù0.582N 

589 

Pù0.231 

Pù0.227 

Pù0.218 

— 

Pù0.196N 

Pù0.183N 

Pù0.247N 

734 (T) 

Pù0.437 

Pù0.445 

Pù0.500 
Cobalt Sulfate Heptahydrate, NTP TR 471 95 

TABLE A3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Skin: Squamous Cell Papilloma or Keratoacanthoma 

Overall rate 2/50 (4%) 5/50 (10%) 0/50 (0%) 4/50 (8%)
 

Adjusted rate 11.1% 17.8% 0.0% 26.7%
 

Terminal rate 1/17 (6%) 1/15 (7%) 0/21 (0%) 4/15 (27%)
 

First incidence (days) 727 589 — 734 (T)
 

Life table test Pù0.385 Pù0.231 Pù0.196N Pù0.272 

Logistic regression test Pù0.400 Pù0.227 Pù0.183N Pù0.271 

Cochran-Armitage test Pù0.429 

Fisher exact test Pù0.218 Pù0.247N Pù0.339 

Skin: Squamous Cell Papilloma, Keratoacanthoma, Basal Cell Adenoma, or Basal Cell Carcinoma 

Overall rate 4/50 (8%) 5/50 (10%) 0/50 (0%) 5/50 (10%)
 

Adjusted rate 22.2% 17.8% 0.0% 33.3%
 

Terminal rate 3/17 (18%) 1/15 (7%) 0/21 (0%) 5/15 (33%) 

First incidence (days) 727 589 — 734 (T) 

Life table test Pù0.406 Pù0.493 Pù0.040N Pù0.413 

Logistic regression test Pù0.420 Pù0.526 Pù0.033N Pù0.414 

Cochran-Armitage test Pù0.458 

Fisher exact test Pù0.500 Pù0.059N Pù0.500 

Skin (Subcutaneous Tissue): Fibrosarcoma or Sarcoma 

Overall rate 0/50 (0%) 2/50 (4%) 1/50 (2%) 3/50 (6%) 

Adjusted rate 0.0% 9.8% 3.3% 16.9% 

Terminal rate 0/17 (0%) 1/15 (7%) 0/21 (0%) 1/15 (7%) 

First incidence (days) — 646 673 692 

Life table test Pù0.113 Pù0.245 Pù0.553 Pù0.113 

Logistic regression test Pù0.113 Pù0.246 Pù0.509 Pù0.109 

Cochran-Armitage test Pù0.132 

Fisher exact test Pù0.247 Pù0.500 Pù0.121 

Skin (Subcutaneous Tissue): Fibroma, Fibrosarcoma, or Sarcoma 

Overall rate 0/50 (0%) 3/50 (6%) 2/50 (4%) 5/50 (10%) 

Adjusted rate 0.0% 15.4% 7.9% 25.9% 

Terminal rate 0/17 (0%) 1/15 (7%) 1/21 (5%) 2/15 (13%) 

First incidence (days) — 646 673 663 

Life table test Pù0.033 Pù0.120 Pù0.293 Pù0.031 

Logistic regression test Pù0.031 Pù0.126 Pù0.272 Pù0.028 

Cochran-Armitage test Pù0.044 

Fisher exact test Pù0.121 Pù0.247 Pù0.028 

Testes: Adenoma 

Overall rate 35/50 (70%) 31/50 (62%) 33/50 (66%) 29/50 (58%) 

Adjusted rate 94.3% 90.5% 88.7% 92.7% 

Terminal rate 15/17 (88%) 12/15 (80%) 17/21 (81%) 13/15 (87%) 

First incidence (days) 568 475 534 526 

Life table test Pù0.369N Pù0.321N Pù0.093N Pù0.319N 

Logistic regression test Pù0.236N Pù0.142N Pù0.185N Pù0.145N 

Cochran-Armitage test Pù0.179N 

Fisher exact test Pù0.263N Pù0.415N Pù0.149N 
96 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Thyroid Gland (C-cell): Adenoma 

Overall rate 4/49 (8%) 2/50 (4%) 3/48 (6%) 4/50 (8%) 

Adjusted rate 18.8% 13.3% 12.1% 20.3% 

Terminal rate 2/17 (12%) 2/15 (13%) 2/21 (10%) 2/15 (13%) 

First incidence (days) 645 734 (T) 649 586 

Life table test Pù0.391 Pù0.358N Pù0.368N Pù0.602 

Logistic regression test Pù0.406 Pù0.316N Pù0.426N Pù0.634 

Cochran-Armitage test Pù0.447 

Fisher exact test Pù0.329N Pù0.512N Pù0.631N 

Thyroid Gland (C-cell): Carcinoma 

Overall rate 3/49 (6%) 3/50 (6%) 4/48 (8%) 3/50 (6%) 

Adjusted rate 12.4% 13.6% 13.6% 15.3% 

Terminal rate 1/17 (6%) 1/15 (7%) 1/21 (5%) 2/15 (13%) 

First incidence (days) 435 576 582 562 

Life table test Pù0.569 Pù0.647 Pù0.597 Pù0.631 

Logistic regression test Pù0.587N Pù0.656N Pù0.479 Pù0.655N 

Cochran-Armitage test Pù0.582N 

Fisher exact test Pù0.651N Pù0.488 Pù0.651N 

Thyroid Gland (C-cell): Adenoma or Carcinoma 

Overall rate 7/49 (14%) 5/50 (10%) 6/48 (13%) 7/50 (14%) 

Adjusted rate 29.5% 25.9% 22.3% 34.1% 

Terminal rate 3/17 (18%) 3/15 (20%) 3/21 (14%) 4/15 (27%) 

First incidence (days) 435 576 582 562 

Life table test Pù0.394 Pù0.408N Pù0.349N Pù0.558 

Logistic regression test Pù0.430 Pù0.353N Pù0.479N Pù0.609 

Cochran-Armitage test Pù0.466 

Fisher exact test Pù0.365N Pù0.516N Pù0.597N 

Thyroid Gland (Follicular Cell): Adenoma or Carcinoma 

Overall rate 1/49 (2%) 3/50 (6%) 0/48 (0%) 1/50 (2%)
 

Adjusted rate 5.9% 16.0% 0.0% 6.7%
 

Terminal rate 1/17 (6%) 1/15 (7%) 0/21 (0%) 1/15 (7%)
 

First incidence (days) 734 (T) 680 — 734 (T)
 

Life table test Pù0.453N Pù0.288 Pù0.458N Pù0.736 

Logistic regression test Pù0.448N Pù0.309 Pù0.458N Pù0.736 

Cochran-Armitage test Pù0.423N
 

Fisher exact test Pù0.316 Pù0.505N Pù0.747N
 

All Organs: Mononuclear Cell Leukemia 

Overall rate 30/50 (60%) 32/50 (64%) 29/50 (58%) 28/50 (56%) 

Adjusted rate 77.3% 80.6% 74.9% 75.6% 

Terminal rate 9/17 (53%) 8/15 (53%) 12/21 (57%) 8/15 (53%) 

First incidence (days) 453 475 435 463 

Life table test Pù0.468N Pù0.501 Pù0.199N Pù0.507N 

Logistic regression test Pù0.313N Pù0.433 Pù0.467N Pù0.420N 

Cochran-Armitage test Pù0.297N 

Fisher exact test Pù0.418 Pù0.500N Pù0.420N 
Cobalt Sulfate Heptahydrate, NTP TR 471 97 

TABLE A3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

All Organs: Malignant Mesothelioma 

Overall rate 3/50 (6%) 2/50 (4%) 2/50 (4%) 2/50 (4%) 

Adjusted rate 10.2% 11.1% 6.5% 9.3% 

Terminal rate 0/17 (0%) 1/15 (7%) 0/21 (0%) 1/15 (7%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

589 

Pù0.523N 

Pù0.498N 

Pù0.491N 

681 

Pù0.477N 

Pù0.494N 

Pù0.500N 

649 

Pù0.397N 

Pù0.507N 

Pù0.500N 

603 

Pù0.517N 

Pù0.501N 

Pù0.500N 

All Organs: Osteosarcoma 

Overall rate 0/50 (0%) 3/50 (6%) 0/50 (0%) 0/50 (0%) 

Adjusted rate 0.0% 11.4% 0.0% 0.0% 

Terminal rate 0/17 (0%) 0/15 (0%) 0/21 (0%) 0/15 (0%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

— 

Pù0.258N 

Pù0.257N 

Pù0.255N 

631 

Pù0.146 

Pù0.123 

Pù0.121 

— 

— 

— 

— 

— 

— 

— 

— 

All Organs: Benign Neoplasms 

Overall rate 48/50 (96%) 46/50 (92%) 47/50 (94%) 48/50 (96%) 

Adjusted rate 97.9% 100.0% 97.9% 100.0% 

Terminal rate 16/17 (94%) 15/15 (100%) 20/21 (95%) 15/15 (100%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

435 

Pù0.317 

Pù0.385 

Pù0.434 

471 

Pù0.419N 

Pù0.319N 

Pù0.339N 

467 

Pù0.125N 

Pù0.517N 

Pù0.500N 

401 

Pù0.437 

Pù0.687 

Pù0.691N 

All Organs: Malignant Neoplasms 

Overall rate 38/50 (76%) 42/50 (84%) 35/50 (70%) 38/50 (76%) 

Adjusted rate 89.9% 93.1% 78.9% 87.5% 

Terminal rate 13/17 (76%) 12/15 (80%) 12/21 (57%) 10/15 (67%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

435 

Pù0.490 

Pù0.396N 

Pù0.395N 

408 

Pù0.386 

Pù0.241 

Pù0.227 

272 

Pù0.115N 

Pù0.329N 

Pù0.326N 

401 

Pù0.469 

Pù0.592N 

Pù0.592N 
 

98 

c 

Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A3 

Statistical Analysis of Primary Neoplasms in Male Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

All Organs: Benign or Malignant Neoplasms 

Overall rate 50/50 (100%) 50/50 (100%) 50/50 (100%) 50/50 (100%) 

Adjusted rate 100.0% 100.0% 100.0% 100.0% 

Terminal rate 17/17 (100%) 15/15 (100%) 21/21 (100%) 15/15 (100%) 

First incidence (days) 

Life table test 

435 

Pù0.361 

408 

Pù0.510N 

272 

Pù0.146N 

401 

Pù0.438 

Logistic regression test —f — — — 

Cochran-Armitage test 

Fisher exact test 

— 

Pù1.000N Pù1.000N Pù1.000N 

(T)Terminal sacrifice 

a Number of neoplasm-bearing animals/number of animals examined. Denominator is number of animals examined microscopically for adrenal gland, bone, 

lung, pancreatic islets, pituitary gland, preputial gland, prostate gland, testes, and thyroid gland; for other tissues, denominator is number of animals 

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

Observed incidence at terminal kill 

d	 Beneath the chamber control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values corresponding 

to pairwise comparisons between the chamber controls and that exposed group. The life table test regards neoplasms in animals dying prior to terminal kill 

as being (directly or indirectly) the cause of death. The logistic regression test regards these lesions as nonfatal. The Cochran-Armitage and Fisher exact 

tests compare directly the overall incidence rates. For all tests, a negative trend or a lower incidence in an exposure group is indicated by N. e Not applicable; no neoplasms in animal group f Value of statistic cannot be computed. 
99 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A4a 

Historical Incidence of Alveolar/bronchiolar Neoplasms in Chamber Control Male F344/N Ratsa 

Incidence in Controls 

Study Adenoma Carcinoma Adenoma or Carcinoma 

Historical Incidence at Battelle Pacific Northwest Laboratories 

o-Chlorobenzalmalononitrile (CS2) 4/50 0/50 4/50 

Acetonitrile 1/48 1/48 2/48 

2-Chloroacetophenone 1/49 1/49 2/49 

l-Epinephrine Hydrochloride 4/50 1/50 5/50 

Chloroethane 0/50 0/50 0/50 

Hexachlorocyclopentadiene 5/50 0/50 5/50 

Ozone 1/50 1/50 2/50 

Overall Historical Incidence 

Total 17/654 (2.6%) 6/654 (0.9%) 23/654 (3.5%) 

Standard deviation 3.6% 1.0% 3.7% 

Range 0%-10% 0%-2% 0%-10% 

a Data as of 12 May 1995 

TABLE A4b 

Historical Incidence of Neoplasms of the Adrenal Medulla in Chamber Control Male F344/N Ratsa 

Incidence in Controls 

Study Benign Pheochromocytoma Benign, Complex, 

or Malignant Pheochromocytomab 

Historical Incidence at Battelle Pacific Northwest Laboratories 

o-Chlorobenzalmalononitrile (CS2) 18/42 20/42 

Acetonitrile 4/48 4/48 

2-Chloroacetophenone 14/46 15/46 

l-Epinephrine Hydrochloride 11/50 11/50 

Chloroethane 8/36 8/36 

Hexachlorocyclopentadiene 15/50 16/50 

Ozone 17/50 17/50 

Overall Historical Incidence 

Total 163/623 (26.2%) 176/623 (28.3%) 

Standard deviation 13.2% 12.0% 

Range 0%-50% 8%-50% 

a Data as of 12 May 1995 b Seven unspecified pheochromocytomas are included in the overall incidence. 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

100 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A5 

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydratea 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Moribund 30 34 26 34 

Natural deaths 3 1 3 1 

Survivors 

Terminal sacrifice 17 15 21 15 

Animals examined microscopically 50 50 50 50 

Alimentary System 

Esophagus (50) (50) (50) (50) 

Inflammation, suppurative 1 (2%) 

Intestine small, jejunum (49) (49) (47) (49) 

Inflammation, chronic active 1 (2%) 

Liver (50) (50) (48) (50) 

Angiectasis 5 (10%) 1 (2%) 1 (2%) 

Basophilic focus 13 (26%) 16 (32%) 23 (48%) 21 (42%) 

Clear cell focus 8 (16%) 5 (10%) 8 (17%) 6 (12%) 

Degeneration, cystic 10 (20%) 11 (22%) 16 (33%) 10 (20%) 

Degeneration, fatty 6 (12%) 5 (10%) 5 (10%) 6 (12%) 

Eosinophilic focus 2 (4%) 3 (6%) 5 (10%) 

Hepatodiaphragmatic nodule 1 (2%) 4 (8%) 4 (8%) 6 (12%) 

Mixed cell focus 3 (6%) 3 (6%) 4 (8%) 5 (10%) 

Necrosis 2 (4%) 1 (2%) 1 (2%) 

Regeneration 3 (6%) 1 (2%) 1 (2%) 1 (2%) 

Thrombosis 1 (2%) 1 (2%) 

Bile duct, hyperplasia 41 (82%) 42 (84%) 34 (71%) 35 (70%) 

Centrilobular, necrosis 17 (34%) 19 (38%) 5 (10%) 11 (22%) 

Mesentery (21) (14) (11) (14) 

Fat, hemorrhage 1 (5%) 1 (9%) 

Fat, necrosis 20 (95%) 11 (79%) 9 (82%) 13 (93%) 

Pancreas (50) (50) (48) (50) 

Angiectasis 1 (2%) 

Atrophy 25 (50%) 25 (50%) 20 (42%) 28 (56%) 

Basophilic focus 3 (6%) 2 (4%) 6 (13%) 3 (6%) 

Hyperplasia 2 (4%) 1 (2%) 4 (8%) 1 (2%) 

Metaplasia, hepatocyte 1 (2%) 

Salivary glands (50) (50) (50) (50) 

Atrophy 2 (4%) 2 (4%) 1 (2%) 

Stomach, forestomach (50) (50) (49) (50) 

Hyperplasia, basal cell 1 (2%) 

Hyperplasia, squamous 1 (2%) 

Necrosis 6 (12%) 5 (10%) 10 (20%) 3 (6%) 

Stomach, glandular (50) (50) (48) (50) 

Inflammation, acute 1 (2%) 

Mineralization 1 (2%) 2 (4%) 1 (2%) 1 (2%) 

Necrosis 3 (6%) 2 (4%) 5 (10%) 2 (4%) 

Tongue (1) (2) 

Hyperplasia, squamous 1 (100%) 1 (50%) 

Epithelium, cyst 1 (50%) 

Tooth (1) (1) 

Developmental malformation 1 (100%) 1 (100%) 

a Number of animals examined microscopically at the site and the number of animals with lesion 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 101 

TABLE A5 

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Cardiovascular System 

Blood vessel (1) 

Aorta, mineralization 1 (100%) 

Heart (50) (50) (50) (50) 

Cardiomyopathy 38 (76%) 46 (92%) 45 (90%) 38 (76%) 

Atrium, thrombosis 2 (4%) 4 (8%) 1 (2%) 4 (8%) 

Endocrine System 

Adrenal cortex (50) (50) (48) (50) 

Hyperplasia 26 (52%) 24 (48%) 26 (54%) 24 (48%) 

Hypertrophy 2 (4%) 5 (10%) 5 (10%) 5 (10%) 

Necrosis 2 (4%) 1 (2%) 3 (6%) 

Thrombosis 1 (2%) 

Vacuolization cytoplasmic 6 (12%) 4 (8%) 3 (6%) 3 (6%) 

Adrenal medulla (50) (50) (49) (50) 

Cyst 1 (2%) 

Hyperplasia 34 (68%) 23 (46%) 29 (59%) 30 (60%) 

Islets, pancreatic (50) (50) (48) (50) 

Hyperplasia 1 (2%) 

Parathyroid gland (48) (48) (49) (49) 

Hyperplasia 1 (2%) 1 (2%) 3 (6%) 1 (2%) 

Pituitary gland (49) (49) (50) (49) 

Pars distalis, hyperplasia 4 (8%) 5 (10%) 4 (8%) 5 (10%) 

Thyroid gland (49) (50) (48) (50) 

C-cell, hyperplasia 31 (63%) 32 (64%) 32 (67%) 34 (68%) 

Follicular cell, hyperplasia 2 (4%) 1 (2%) 1 (2%) 

General Body System 

None 

Genital System 

Epididymis (50) (50) (50) (50) 

Granuloma sperm 1 (2%) 3 (6%) 

Preputial gland (50) (50) (49) (50) 

Hyperplasia 1 (2%) 

Inflammation, chronic active 11 (22%) 10 (20%) 3 (6%) 12 (24%) 

Prostate (50) (50) (49) (50) 

Hyperplasia 12 (24%) 8 (16%) 5 (10%) 9 (18%) 

Inflammation, chronic active 3 (6%) 4 (8%) 1 (2%) 1 (2%) 

Necrosis 1 (2%) 

Seminal vesicle (50) (50) (49) (49) 

Inflammation, chronic active 1 (2%) 

Necrosis 1 (2%) 

Testes (50) (50) (50) (50) 

Atrophy 2 (4%) 4 (8%) 5 (10%) 

Artery, inflammation, chronic active 1 (2%) 1 (2%) 2 (4%) 3 (6%) 

Interstitial cell, hyperplasia 13 (26%) 10 (20%) 6 (12%) 10 (20%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

102 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE A5 

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Hematopoietic System 

Bone marrow (50) (50) (48) (50) 

Necrosis 1 (2%) 

Lymph node (8) (9) (9) (9) 

Iliac, ectasia 1 (11%) 

Iliac, hemorrhage 1 (11%) 

Pancreatic, ectasia 1 (11%) 

Renal, hemorrhage 1 (11%) 

Lymph node, bronchial (45) (30) (41) (49) 

Inflammation, suppurative 1 (2%) 

Lymph node, mandibular (46) (47) (47) (49) 

Infiltration cellular, plasma cell 1 (2%) 

Lymph node, mediastinal (47) (46) (44) (49) 

Hemorrhage 1 (2%) 1 (2%) 

Spleen (50) (50) (49) (50) 

Accessory spleen 1 (2%) 1 (2%) 1 (2%) 1 (2%) 

Depletion cellular 1 (2%) 

Fibrosis 16 (32%) 16 (32%) 12 (24%) 13 (26%) 

Hematopoietic cell proliferation 1 (2%) 1 (2%) 1 (2%) 

Hyperplasia, focal 1 (2%) 

Necrosis 2 (4%) 1 (2%) 2 (4%) 

Integumentary System 

Mammary gland (30) (34) (36) (38) 

Galactocele 2 (7%) 1 (3%) 1 (3%) 

Hyperplasia, atypical 1 (3%) 

Skin (50) (48) (50) (50) 

Cyst epithelial inclusion 1 (2%) 

Inflammation, chronic 3 (6%) 1 (2%) 1 (2%) 1 (2%) 

Inflammation, chronic active 1 (2%) 2 (4%) 3 (6%) 

Subcutaneous tissue, edema 1 (2%) 

Musculoskeletal System 

Bone (50) (50) (50) (50) 

Fibrous osteodystrophy 2 (4%) 

Hyperostosis 2 (4%) 2 (4%) 

Nervous System 

Brain (50) (50) (50) (50) 

Developmental malformation 1 (2%) 

Gliosis 1 (2%) 1 (2%) 1 (2%) 

Mineralization 1 (2%) 

Necrosis 2 (4%) 1 (2%) 

Respiratory System 

Larynx (50) (49) (48) (50) 

Epiglottis, metaplasia, squamous 10 (20%) 37 (77%) 50 (100%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 103 

TABLE A5 

Summary of the Incidence of Nonneoplastic Lesions in Male Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Respiratory System (continued) 

Lung (50) (50) (48) (50) 

Cyst 1 (2%) 

Foreign body 1 (2%) 

Hemorrhage 1 (2%) 

Hyperplasia, atypical 1 (2%) 1 (2%) 

Infiltration cellular, histiocyte 1 (2%) 1 (2%) 1 (2%) 

Inflammation, suppurative 1 (2%) 

Metaplasia, squamous 1 (2%) 4 (8%) 2 (4%) 

Mineralization 1 (2%) 

Proteinosis 16 (32%) 40 (83%) 47 (94%) 

Thrombosis 1 (2%) 1 (2%) 

Alveolar epithelium, hyperplasia 9 (18%) 20 (40%) 20 (42%) 23 (46%) 

Alveolar epithelium, hyperplasia, atypical 1 (2%) 2 (4%) 2 (4%) 

Alveolar epithelium, metaplasia 50 (100%) 48 (100%) 49 (98%) 

Alveolus, inflammation, granulomatous 2 (4%) 50 (100%) 48 (100%) 50 (100%) 

Artery, mediastinum, mineralization 1 (2%) 

Bronchiole, inflammation, chronic 1 (2%) 

Interstitium, fibrosis 1 (2%) 50 (100%) 48 (100%) 49 (98%) 

Mediastinum, inflammation, suppurative 1 (2%) 

Nose (50) (50) (49) (50) 

Inflammation, chronic active 1 (2%) 

Inflammation, suppurative 4 (8%) 15 (30%) 5 (10%) 6 (12%) 

Metaplasia, squamous 1 (2%) 

Thrombosis 14 (28%) 18 (36%) 3 (6%) 9 (18%) 

Glands, cyst 1 (2%) 

Lateral wall, hyperplasia 2 (4%) 14 (28%) 21 (43%) 20 (40%) 

Lateral wall, inflammation 1 (2%) 

Lateral wall, metaplasia, squamous 1 (2%) 3 (6%) 5 (10%) 8 (16%) 

Olfactory epithelium, atrophy 8 (16%) 24 (48%) 42 (86%) 48 (96%) 

Olfactory epithelium, metaplasia 5 (10%) 1 (2%) 5 (10%) 30 (60%) 

Respiratory epithelium, hyperplasia, focal 6 (12%) 1 (2%) 3 (6%) 2 (4%) 

Respiratory epithelium, inflammation 4 (8%) 1 (2%) 

Respiratory epithelium, metaplasia, squamous 1 (2%) 1 (2%) 

Pleura (1) 

Trachea (50) (50) (48) (50) 

Mineralization 1 (2%) 

Special Senses System 

Eye (5) (1) (1) 

Cataract 5 (100%) 1 (100%) 1 (100%) 

Retina, atrophy 5 (100%) 1 (100%) 1 (100%) 

Urinary System 

Kidney (50) (50) (48) (50) 

Cyst 1 (2%) 1 (2%) 

Hydronephrosis 1 (2%) 1 (2%) 

Infarct 4 (8%) 1 (2%) 1 (2%) 3 (6%) 

Nephropathy 49 (98%) 49 (98%) 48 (100%) 50 (100%) 

Thrombosis 1 (2%) 

Papilla, necrosis 1 (2%) 

Renal tubule, hyperplasia 2 (4%) 4 (8%) 2 (4%) 2 (4%) 

Urinary bladder (50) (50) (48) (50) 

Necrosis 1 (2%) 

Transitional epithelium, hyperplasia 1 (2%) 
104 Cobalt Sulfate Heptahydrate, NTP TR 471 
 

 

105 

APPENDIX B 

SUMMARY OF LESIONS IN FEMALE RATS 

IN THE 2-YEAR INHALATION STUDY 

OF COBALT SULFATE HEPTAHYDRATE 

TABLE B1 

TABLE B2 

TABLE B3 

TABLE B4a 

TABLE B4b 

TABLE B5 

Summary of the Incidence of Neoplasms in Female Rats 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 

Individual Animal Tumor Pathology of Female Rats 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 

Statistical Analysis of Primary Neoplasms in Female Rats 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 

Historical Incidence of Alveolar/bronchiolar Neoplasms 

in Chamber Control Female F344/N Rats ................................ 

Historical Incidence of Neoplasms of the Adrenal Medulla 

in Chamber Control Female F344/N Rats ................................ 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 

106 

110 

128 

134 

134 

135 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

106 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B1 

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydratea 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Moribund 19 20 20 17 

Natural deaths 3 4 4 3 

Survivors 

Terminal sacrifice 28 25 26 30 

Pregnant 1 

Animals examined microscopically 50 49 50 50 

Alimentary System 

Intestine large, colon (48) (45) (48) (48) 

Intestine large, cecum (48) (45) (47) (47) 

Intestine small, duodenum (49) (46) (48) (48) 

Intestine small, ileum (49) (45) (47) (47) 

Liver (50) (49) (50) (49) 

Hemangiosarcoma 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Pheochromocytoma malignant, metastatic, 

adrenal medulla 1 (2%) 

Mesentery (5) (8) (6) (13) 

Sarcoma stromal, metastatic, uterus 1 (13%) 

Oral mucosa (1) (2) 

Pharyngeal, squamous cell papilloma 1 (100%) 2 (100%) 

Pancreas (49) (49) (50) (48) 

Salivary glands (50) (49) (50) (50) 

Adenoma 1 (2%) 

Sarcoma, metastatic, eye 1 (2%) 

Stomach, forestomach (50) (49) (50) (50) 

Leiomyoma 1 (2%) 

Stomach, glandular (49) (49) (49) (48) 

Carcinoid tumor benign 1 (2%) 

Cardiovascular System 

Heart (50) (49) (50) (50) 

Carcinoma, metastatic, lung 1 (2%) 

Carcinoma, metastatic, thyroid gland 1 (2%) 

Schwannoma benign 1 (2%) 2 (4%) 2 (4%) 1 (2%) 

Endocrine System 

Adrenal cortex (49) (49) (50) (49) 

Adenoma 1 (2%) 1 (2%) 

Carcinoma, metastatic, kidney 1 (2%) 

Adrenal medulla (48) (49) (50) (48) 

Pheochromocytoma malignant 1 (2%) 1 (2%) 

Pheochromocytoma complex 1 (2%) 

Pheochromocytoma benign 2 (4%) 1 (2%) 3 (6%) 7 (15%) 

Bilateral, pheochromocytoma benign 1 (2%) 

Islets, pancreatic (49) (49) (50) (48) 

Adenoma 4 (8%) 1 (2%) 

Carcinoma 2 (4%) 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 107 

TABLE B1 

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Endocrine System (continued) 

Pituitary gland 

Pars distalis, adenoma

Thyroid gland 

C-cell, adenoma

C-cell, carcinoma

Follicular cell, adenoma

(50) 

 40 (80%) 

(49) 

 6 (12%) 

 3 (6%) 

(49) 

 39 (80%) 

(49) 

 6 (12%) 

 1 (2%) 

(50) 

 38 (76%) 

(50) 

 7 (14%) 

 2 (4%) 

(49) 

 40 (82%) 

(48) 

 3 (6%) 

 3 (6%) 

 1 (2%) 

General Body System 

None 

Genital System 

Clitoral gland (50) (48) (48) (49) 

Adenoma 5 (10%) 7 (15%) 3 (6%) 3 (6%) 

Carcinoma 2 (4%) 3 (6%) 4 (8%) 3 (6%) 

Bilateral, adenoma 2 (4%) 

Ovary (50) (49) (50) (49) 

Granulosa cell tumor benign 1 (2%) 

Bilateral, granulosa-theca tumor benign 1 (2%) 

Uterus (50) (49) (50) (49) 

Histiocytic sarcoma 1 (2%) 

Leiomyoma 1 (2%) 

Polyp stromal 8 (16%) 7 (14%) 10 (20%) 4 (8%) 

Polyp stromal, multiple 1 (2%) 

Sarcoma stromal 1 (2%) 2 (4%) 

Hematopoietic System 

Bone marrow (49) (49) (50) (50) 

Lymph node (2) (3) (3) (2) 

Lymph node, bronchial (30) (30) (37) (37) 

Lymph node, mandibular (44) (40) (47) (45) 

Sarcoma, metastatic, eye 1 (2%) 

Sarcoma, metastatic, skin 1 (2%) 

Lymph node, mesenteric (49) (49) (50) (48) 

Lymph node, mediastinal (43) (38) (43) (45) 

Spleen (49) (48) (50) (49) 

Histiocytic sarcoma 1 (2%) 

Thymus (42) (45) (45) (44) 

Integumentary System 

Mammary gland (50) (49) (50) (49) 

Adenoma 2 (4%) 

Carcinoma 2 (4%) 2 (4%) 5 (10%) 4 (8%) 

Carcinoma, multiple 1 (2%) 1 (2%) 

Fibroadenoma 18 (36%) 18 (37%) 13 (26%) 20 (41%) 

Fibroadenoma, multiple 4 (8%) 4 (8%) 7 (14%) 7 (14%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

108 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B1 

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Integumentary System (continued) 

Skin (50) (49) (50) (50) 

Basal cell adenoma 1 (2%) 

Squamous cell papilloma 1 (2%) 

Sebaceous gland, adenoma 1 (2%) 1 (2%) 

Subcutaneous tissue, fibroma 1 (2%) 

Subcutaneous tissue, fibrosarcoma 1 (2%) 

Subcutaneous tissue, lipoma 2 (4%) 

Subcutaneous tissue, sarcoma 1 (2%) 

Musculoskeletal System 

Bone (50) (49) (50) (50) 

Sarcoma, metastatic, eye 1 (2%) 

Skeletal muscle (1) (1) 

Carcinoma, metastatic, lung 1 (100%) 

Rhabdomyosarcoma 1 (100%) 

Nervous System 

Brain (50) (49) (50) (50) 

Astrocytoma benign 1 (2%) 

Astrocytoma malignant 1 (2%) 

Oligodendroglioma benign 1 (2%) 

Respiratory System 

Larynx (50) (49) (50) (50) 

Lung (50) (49) (50) (50) 

Alveolar/bronchiolar adenoma 1 (2%) 9 (18%) 7 (14%) 

Alveolar/bronchiolar adenoma, multiple 1 (2%) 2 (4%) 

Alveolar/bronchiolar carcinoma 2 (4%) 6 (12%) 6 (12%) 

Carcinoma, metastatic, mammary gland 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Pheochromocytoma malignant, metastatic, 

adrenal medulla 1 (2%) 

Sarcoma, metastatic, eye 1 (2%) 

Sarcoma, metastatic, skin 1 (2%) 

Sarcoma, metastatic, uncertain primary site 1 (2%) 

Squamous cell carcinoma 1 (2%) 1 (2%) 

Mediastinum, alveolar/bronchiolar carcinoma, 

metastatic, lung 1 (2%) 

Nose (50) (49) (50) (50) 

Pleura (1) 

Special Senses System 

Eye (1) (5) (3) (3) 

Sarcoma 1 (33%) 

Zymbal’s gland (1) (1) 

Carcinoma 1 (100%) 1 (100%) 
 

 

 

 

 

 

 

 

109 

c 

Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B1 

Summary of the Incidence of Neoplasms in Female Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Urinary System 

Kidney (49) (49) (50) (48) 

Histiocytic sarcoma 1 (2%) 

Lipoma 1 (2%) 1 (2%) 

Mesenchymal tumor benign 1 (2%) 

Bilateral, renal tubule, carcinoma, multiple 1 (2%) 

Urinary bladder (49) (49) (50) (47) 

Systemic Lesions 

Multiple organsb (50) (49) (50) (50) 

Histiocytic sarcoma 1 (2%) 1 (2%) 

Leukemia mononuclear 15 (30%) 16 (33%) 19 (38%) 10 (20%) 

Lymphoma malignant 1 (2%) 

Mesothelioma malignant 1 (2%) 

Neoplasm Summary 

Total animals with primary neoplasmsc 48 47 50 46 

Total primary neoplasms 123 122 141 139 

Total animals with benign neoplasms 45 44 47 44 

Total benign neoplasms 96 94 97 105 

Total animals with malignant neoplasms 25 23 32 27 

Total malignant neoplasms 27 28 44 34 

Total animals with metastatic neoplasms 3 3 1 3 

Total metastatic neoplasms 3 3 2 9 

Total animals with malignant neoplasms

 of uncertain primary site 1 

a Number of animals examined microscopically at the site and the number of animals with neoplasm b Number of animals with any tissue examined microscopically 

Primary neoplasms: all neoplasms except metastatic neoplasms 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

110 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

3 5 5 5 5 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 8 6 7 8 8 0 1 3 3 6 7 9 9 0 0 0 1 2 2 2 3 3 3 3 3 

5 9 2 2 2 2 0 2 8 6 9 2 4 5 9 9 9 1 7 7 3 3 5 5 5 

1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 

Carcass ID Number 4 5 0 1 4 1 1 2 3 1 0 3 3 0 0 4 2 2 2 4 2 4 0 0 1 

8 0 7 6 2 8 4 6 6 0 2 1 8 1 4 3 8 2 1 1 3 4 3 8 5 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 

Intestine large, colon + + + + + + + + + + + + + + + + + A + + A + + + + 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + A + + + + 

Intestine large, cecum + + + + + + + + + + + + + + + + + A + + A + + + + 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + A + + + + 

Intestine small, jejunum + + + + + + + + + + + + + + + + + A + A A + + + + 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + A + + + + 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 

Mesentery + + 

Oral mucosa 

Pharyngeal, squamous cell papilloma 

Pancreas + + + + + + + + + + + + + + + + + + + + A + + + + 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + A + + + + 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 

Carcinoma, metastatic, thyroid gland 

Schwannoma benign X 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + A + + + + + 

Carcinoma, metastatic, kidney 

Adrenal medulla + + + + + + + + + + + + + + + + + + + A + + + + + 

Pheochromocytoma benign X X 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + A + + + + 

Adenoma 

Carcinoma X 

Parathyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X X X 

Thyroid gland + + + + + + + + + + + + + + + + + + + + A + + + + 

C-cell, adenoma X 

C-cell, carcinoma X 

General Body System 

Peritoneum + 

Genital System 

Clitoral gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma X X X X 

Carcinoma 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 

Granulosa cell tumor benign 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 

Polyp stromal X X X X 

Sarcoma stromal X 

+: Tissue examined microscopically M: Missing tissue X: Lesion present 

A: Autolysis precludes examination I: Insufficient tissue Blank: Not examined 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 111 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 

1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 Total 

Carcass ID Number 2 2 3 4 4 4 0 0 1 1 1 2 3 3 3 3 3 4 0 1 1 2 2 3 4 Tissues/ 

5 7 4 0 5 9 5 6 2 3 9 9 0 2 5 7 9 6 9 1 7 0 4 3 7 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Mesentery + + + 5 

Oral mucosa + 1 

Pharyngeal, squamous cell papilloma X 1 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma, metastatic, thyroid gland X 1 

Schwannoma benign 1 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Carcinoma, metastatic, kidney X 1 

Adrenal medulla + + + + + + + + + + + + + + M + + + + + + + + + + 48 

Pheochromocytoma benign 2 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Adenoma X X X X 4 

Carcinoma X 2 

Parathyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X X X 40 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

C-cell, adenoma X X X X X 6 

C-cell, carcinoma X X 3 

General Body System 

Peritoneum 1 

Genital System 

Clitoral gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X 5 

Carcinoma X X 2 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Granulosa cell tumor benign X 1 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Polyp stromal X X X X 8 

Sarcoma stromal 1 
 

 

 

 

 

 

 

 

 

 

 

112 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

3 5 5 5 5 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 8 6 7 8 8 0 1 3 3 6 7 9 9 0 0 0 1 2 2 2 3 3 3 3 3 

5 9 2 2 2 2 0 2 8 6 9 2 4 5 9 9 9 1 7 7 3 3 5 5 5 

1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 

Carcass ID Number 4 5 0 1 4 1 1 2 3 1 0 3 3 0 0 4 2 2 2 4 2 4 0 0 1 

8 0 7 6 2 8 4 6 6 0 2 1 8 1 4 3 8 2 1 1 3 4 3 8 5 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + A + + + + + 

Lymph node + + 

Lymph node, bronchial M M + M + + M M + + M + M + + M M + + + M M M + M 

Lymph node, mandibular M + + + M + M + + + + + + + + M M + M + + + + + + 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + A + + + + 

Lymph node, mediastinal + + + + + M + + + + + + + + + + + M + + + + + + + 

Spleen + + + + + + + + + + + + + + + + + + + + A + + + + 

Thymus + + + + + M + + + + + + + + + + + M + + M M + + + 

Integumentary System 

Mammary gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma X X 

Carcinoma X 

Carcinoma, multiple X 

Fibroadenoma X X X X X X X X X X 

Fibroadenoma, multiple X X 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 

Squamous cell papilloma 

Subcutaneous tissue, lipoma X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Nervous System 

Brain 

Respiratory System 

Larynx 

Lung 

Carcinoma, metastatic, mammary gland 

Nose 

Trachea 

Special Senses System 

Eye 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Urinary System 

Kidney 

Lipoma 

Bilateral, renal tubule, carcinoma, multiple 

Urinary bladder 

+ + + + + + + + + + + + + + + + + + + A + + + + + 

+ + + + + + + + + + + + + + + + + + + + A + + + + 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X X X X X X X 
 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 113 

TABLE B2 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 

1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 Total 

Carcass ID Number 2 2 3 4 4 4 0 0 1 1 1 2 3 3 3 3 3 4 0 1 1 2 2 3 4 Tissues/ 

5 7 4 0 5 9 5 6 2 3 9 9 0 2 5 7 9 6 9 1 7 0 4 3 7 Tumors 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Lymph node 2 

Lymph node, bronchial M + M + M M + + M M + + + + + + M + + + + + + + + 30 

Lymph node, mandibular + + + + + + + + + + + + + + + + + + + + + + + + + 44 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Lymph node, mediastinal M M + + + M + + + + + + + + + + + + + + + + M M + 43 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Thymus M M + + + + + + + + + + + M + + + + + + + + + + M 42 

Integumentary System 

Mammary gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma 2 

Carcinoma X 2 

Carcinoma, multiple 1 

Fibroadenoma X X X X X X X X 18 

Fibroadenoma, multiple X X 4 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Squamous cell papilloma X 1 

Subcutaneous tissue, lipoma X 2 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 

Respiratory System 

Larynx + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma, metastatic, mammary gland 1 

Nose + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Special Senses System 

Eye +

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Lipoma X 1 

Bilateral, renal tubule, carcinoma, multiple X 1 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Leukemia mononuclear X X X X 15 

50 

50 

 1 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

114 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B2
 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

Number of Days on Study 

3 4 4 4 5 5 5 5 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 

6 2 8 9 1 2 2 8 1 1 1 5 7 7 7 8 9 0 0 0 1 2 2 3 

2 0 3 1 7 6 6 5 0 0 0 2 0 3 3 7 4 2 9 9 4 2 8 1 

Carcass ID Number 

3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

3 2 1 5 2 1 4 1 0 0 1 0 1 0 4 3 1 0 2 3 1 3 4 0 

6 7 0 0 2 8 0 1 3 7 2 4 5 6 8 1 3 8 3 5 9 4 2 1 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Mesentery 

Sarcoma stromal, metastatic, uterus 

Oral mucosa 

Pharyngeal, squamous cell papilloma 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Carcinoid tumor benign 

Tooth 

+ + + + + + + + + + + + + + + + + + + + + + + + 

A A + + + + + + + + + + + + + + + + + + A + + A 

A + + + + + + + + + + + + + + + + + + + A + + + 

A A + + + + + + + + + + + + + + + + + + A + + A 

+ + + + + + + + + + + + + + + + + + + + A + + A 

A A + + + + + + + + + + + + + + + + + + A + + A 

A A + + + + + + + + + + + + + + + + + + A + + A 

+ + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + 

X 

+ 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + 

Schwannoma benign X 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma X 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma benign X 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma 

Parathyroid gland M + + + + + + + + + + + + + + + + + + + + + + + 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + 

Pars distalis, adenoma X X X X X X X X X X X X X X X X 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + 

C-cell, adenoma X X X 

Follicular cell, adenoma X 

General Body System 

None 

Genital System 

Clitoral gland + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma X X X 

Carcinoma X X 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + 

Bilateral, granulosa-theca tumor, benign X 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + 

Polyp stromal X X X X 

Sarcoma stromal X X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 115 

TABLE B2
 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

5 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 

3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 Total 

Carcass ID Number 0 2 2 2 3 3 3 4 4 4 0 1 2 2 3 3 3 4 4 4 0 1 2 2 4 Tissues/ 

2 4 5 6 0 7 8 1 4 9 9 6 0 8 2 3 9 5 6 7 5 7 1 9 3 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 45 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 45 

Intestine small, duodenum + + I + + + + + + + + + + + + + + + + + + + + + + 46 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 45 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 45 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Mesentery + + + 8 

Sarcoma stromal, metastatic, uterus 1 

Oral mucosa + + 2 

Pharyngeal, squamous cell papilloma X X 2 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Carcinoid tumor benign 1 

Tooth + 2 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Schwannoma benign X 2 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Adenoma 1 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Pheochromocytoma benign 1 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Adenoma X 1 

Parathyroid gland + + + + + M + + + + + + + + + + + + + + + + + + + 47 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X X X X X X 39 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

C-cell, adenoma X X X 6 

Follicular cell, adenoma 1 

General Body System 

None 

Genital System 

Clitoral gland M + + + + + + + + + + + + + + + + + + + + + + + + 48 

Adenoma X X X X 7 

Carcinoma X 3 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Bilateral, granulosa-theca tumor, benign 1 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Polyp stromal X X X 7 

Sarcoma stromal 2 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

116 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B2
 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

3 4 4 4 5 5 5 5 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 

Number of Days on Study 6 

2 

2 

0 

8 

3 

9 

1 

1 

7 

2 

6 

2 

6 

8 

5 

1 

0 

1 

0 

1 

0 

5 

2 

7 

0 

7 

3 

7 

3 

8 

7 

9 

4 

0 

2 

0 

9 

0 

9 

1 

4 

2 

2 

2 

8 

3 

1 

3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

Carcass ID Number 3 2 1 5 2 1 4 1 0 0 1 0 1 0 4 3 1 0 2 3 1 3 4 0 

6 7 0 0 2 8 0 1 3 7 2 4 5 6 8 1 3 8 3 5 9 4 2 1 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Thymus 

+ + + + + + + + + + + + + + + + + + + + + + + + 

+ + + 

M + + M M M M + + + + + M M M + + + M + M + M M 

M + + + M + + + M M + + M + + + + + + + + + M + 

+ + + + + + + + + + + + + + + + + + + + + + + + 

M + + + + + M + + + + M + + + + + + + + + + M + 

+ + + + + + + + + + + + + + + + + + + + + + + A 

+ + + + + + + + + + + + + + + + + + + + + + + + 

Integumentary System 

Mammary gland 

Carcinoma 

Fibroadenoma 

Fibroadenoma, multiple 

Skin 

Sebaceous gland, adenoma 

+ + + + + + + + + + + + + + + + + + + + + + + + 

X X 

X X X X X 

X X 

+ + + + + + + + + + + + + + + + + + + + + + + + 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + 

Nervous System 

Brain 

Astrocytoma malignant 

+ + + + + + + + + + + + + + + + + + + + + + + + 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Mesothelioma malignant, metastatic, pleura 

Sarcoma, metastatic, uncertain primary site 

Nose 

Pleura 

Trachea 

+ + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + 

X 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + 

+ 

+ + + + + + + + + + + + + + + + + + + + + + + + 

Special Senses System 

Eye 

Zymbal’s gland 

Carcinoma 

+ + 

+ 

X 

+ 

Urinary System 

Kidney 

Mesenchymal tumor benign 

Urinary bladder 

+ + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Mesothelioma malignant 

+ + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X X X X X X X 

X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 117 

TABLE B2
 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

7 

3 

7 

3 

7 

3 

7 

3 

7 

3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 Total 

Carcass ID Number 0 

2 

2 

4 

2 

5 

2 

6 

3 

0 

3 

7 

3 

8 

4 

1 

4 

4 

4 

9 

0 

9 

1 

6 

2 

0 

2 

8 

3 

2 

3 

3 

3 

9 

4 

5 

4 

6 

4 

7 

0 

5 

1 

7 

2 

1 

2 

9 

4 

3 

Tissues/ 

Tumors 

Hematopoietic System 

Bone marrow 

Lymph node

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Thymus 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

M M + + + + M + + M + + + + + M + M + + + + + + M 

M + + + + + + + + M + + + + + + + M + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ M + + + + M + + + M + + M M + + + + + + + + M M 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + M + M M + + + + + M + + + + 

49 

3 

30 

40 

49 

38 

48 

45 

Integumentary System 

Mammary gland 

Carcinoma

Fibroadenoma 

Fibroadenoma, multiple 

Skin 

Sebaceous gland, adenoma 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X X X X X X X X X 

X X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

49 

2 

18 

4 

49 

1 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Nervous System 

Brain 

Astrocytoma malignant 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

49 

1 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma

Alveolar/bronchiolar carcinoma 

Mesothelioma malignant, metastatic, pleura

Sarcoma, metastatic, uncertain primary site 

Nose 

Pleura

Trachea 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X

X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

49 

49 

1 

2 

1 

1 

49 

1 

49 

Special Senses System 

Eye 

Zymbal’s gland

Carcinoma

+ + 5 

1 

1 

Urinary System 

Kidney 

Mesenchymal tumor benign 

Urinary bladder 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

49 

1 

49 

Systemic Lesions 

Multiple organs 

Leukemia mononuclear 

Mesothelioma malignant

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X 

49 

16 

1 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

118 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B2
 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

Number of Days on Study 

3 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 

5 1 2 4 6 9 0 0 1 3 5 5 8 9 9 9 0 2 2 2 2 2 2 3 3 

5 0 6 1 8 6 0 3 9 2 2 9 5 2 4 5 9 2 2 2 3 7 9 2 5 

Carcass ID Number 

5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 

2 4 3 2 1 1 2 4 4 0 3 2 1 1 0 0 4 2 2 3 3 4 3 3 0 

5 6 0 7 4 9 6 8 7 7 1 3 8 2 8 3 1 1 8 6 9 0 7 5 2 

Alimentary System 

Esophagus 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hemangiosarcoma 

Histiocytic sarcoma 

Pheochromocytoma malignant, metastatic,

 adrenal medulla 

Mesentery 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + A + + + + + + + + + A + + + + + + + 

+ + + + + + + A + + + + + + + + + + + + + + + + + 

+ + + + + + + A + + + A + + + + + A + + + + + + + 

+ + + + + + + A + + + A + + + + + + + + + + + + + 

+ + + + + + + A + + + A + + + + + A + + + + + + + 

+ + + + + + + A + + + A + + + + + A + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

+ + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + A + + + + + + + + + + + + + + + + + 

Cardiovascular System 

Blood vessel 

Heart 

Schwannoma benign 

+ 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma malignant X 

Pheochromocytoma benign X X 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 

Carcinoma X 

Parathyroid gland + + + + + + + + + + M + + + + + + + + + + + + M + 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

C-cell, adenoma X X 

C-cell, carcinoma X X 

General Body System 

None 

Genital System 

Clitoral gland + + + + + + + M + + + + + + + + + + + + M + + + + 

Adenoma X X 

Carcinoma X X 

Bilateral, adenoma 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 

Polyp stromal X X X 

Polyp stromal, multiple 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 119 

TABLE B2
 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 

5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 Total 

Carcass ID Number 1 1 4 5 0 0 1 1 2 2 3 3 3 4 4 4 0 0 0 1 1 2 2 3 4 Tissues/ 

1 7 9 0 4 5 3 5 4 9 2 3 8 2 3 4 1 6 9 0 6 0 2 4 5 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma X 1 

Histiocytic sarcoma X 1 

Pheochromocytoma malignant, metastatic,

 adrenal medulla 1 

Mesentery + + + + 6 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Cardiovascular System 

Blood vessel 1 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Schwannoma benign X X 2 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pheochromocytoma malignant 1 

Pheochromocytoma benign X 3 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma 1 

Parathyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X X X X 38 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

C-cell, adenoma X X X X X 7 

C-cell, carcinoma 2 

General Body System 

None 

Genital System 

Clitoral gland + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Adenoma X 3 

Carcinoma X X 4 

Bilateral, adenoma X X 2 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Polyp stromal X X X X X X X 10 

Polyp stromal, multiple X 1 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

120 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B2
 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

3 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 

Number of Days on Study 5 1 2 4 6 9 0 0 1 3 5 5 8 9 9 9 0 2 2 2 2 2 2 3 3 

5 0 6 1 8 6 0 3 9 2 2 9 5 2 4 5 9 2 2 2 3 7 9 2 5 

5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 

Carcass ID Number 2 4 3 2 1 1 2 4 4 0 3 2 1 1 0 0 4 2 2 3 3 4 3 3 0 

5 6 0 7 4 9 6 8 7 7 1 3 8 2 8 3 1 1 8 6 9 0 7 5 2 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 

Lymph node + + + 

Lymph node, bronchial + M + + + + + + M + M + + + + + + + + + M M + M + 

Lymph node, mandibular + M + + + + + + + + + + + + + + + + + + + + + + + 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + 

Lymph node, mediastinal + + + + + + + M + + + + + + + + + + + + + + + + M 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 

Thymus + + + + + + + + M + + + + + + + M M + + + + + + + 

Integumentary System 

Mammary gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Carcinoma X X X 

Fibroadenoma X X X X X 

Fibroadenoma, multiple X X X 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 

Subcutaneous tissue, fibrosarcoma X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Skeletal muscle + 

Rhabdomyosarcoma X 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 

Oligodendroglioma benign X 

Respiratory System 

Larynx + + + + + + + + + + + + + + + + + + + + + + + + + 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 

Alveolar/bronchiolar adenoma X 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma X X 

Pheochromocytoma malignant, metastatic,

 adrenal medulla X 

Squamous cell carcinoma 

Nose + + + + + + + + + + + + + + + + + + + + + + + + + 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 

Special Senses System 

Eye + 

Zymbal’s gland + 

Carcinoma X 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma 

Leukemia mononuclear X X X X X X X X X X X X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 121 

TABLE B2
 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 

5 

3 

5 

3 

5 

3 

5 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

7 

3 

7 

3 

7 

3 

7 

3 

7 

3 

7 

3 

7 

3 

7 

3 

7 

5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 Total 

Carcass ID Number 1 

1 

1 

7 

4 

9 

5 

0 

0 

4 

0 

5 

1 

3 

1 

5 

2 

4 

2 

9 

3 

2 

3 

3 

3 

8 

4 

2 

4 

3 

4 

4 

0 

1 

0 

6 

0 

9 

1 

0 

1 

6 

2 

0 

2 

2 

3 

4 

4 

5 

Tissues/ 

Tumors 

Hematopoietic System 

Bone marrow 

Lymph node

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Thymus 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + M M + + + + + + + + M M M + + + + + + + + M M 

+ + + M + + + + + + + + M + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + M + + + M + + + + + M + + + + + + M + + + M + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + M + + M + + + + + + + + + + + 

50 

3 

37 

47 

50 

43 

50 

45 

Integumentary System 

Mammary gland 

Carcinoma 

Fibroadenoma 

Fibroadenoma, multiple 

Skin 

Subcutaneous tissue, fibrosarcoma

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X

X X X X X X X X 

X X X X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

50 

5 

13 

7 

50 

1 

Musculoskeletal System 

Bone 

Skeletal muscle

Rhabdomyosarcoma

+ + + + + + + + + + + + + + + + + + + + + + + + + 50 

1 

1 

Nervous System 

Brain 

Oligodendroglioma benign

+ + + + + + + + + + + + + + + + + + + + + + + + + 50 

1 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma 

Pheochromocytoma malignant, metastatic,

 adrenal medulla 

Squamous cell carcinoma 

Nose 

Trachea 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X X X X

X 

X X X X

X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

50 

50 

9 

1 

6 

1 

1 

50 

50 

Special Senses System 

Eye 

Zymbal’s gland

Carcinoma

+ + 3 

1 

1 

Urinary System 

Kidney 

Urinary bladder 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

50 

50 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Leukemia mononuclear 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

X X X X X X X 

50 

1 

19 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

122 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B2
 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

0 3 4 5 6 6 6 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 

Number of Days on Study 0 8 7 8 1 1 3 4 5 5 6 7 7 8 8 9 9 0 0 2 3 3 3 3 3 

6 6 0 5 0 1 2 2 0 2 3 3 9 0 6 2 3 9 9 2 5 5 5 5 5 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Carcass ID Number 4 3 4 0 4 2 3 1 0 2 4 3 1 4 0 0 0 1 3 2 1 1 1 1 2 

3 0 2 1 9 1 6 7 7 6 7 3 1 4 8 4 9 0 9 9 2 6 8 9 8 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 

Intestine large, colon A + + A + + + + + + + + + + + + + + + + + + + + + 

Intestine large, rectum A + + + + + + + + + + + + + + + + + + + + + + + + 

Intestine large, cecum A + + A + + + A + + + + + + + + + + + + + + + + + 

Intestine small, duodenum A + + A + + + + + + + + + + + + + + + + + + + + + 

Intestine small, jejunum A + + A + + + A + + + + + + + + + + + + + + + + + 

Intestine small, ileum A + + A + + + A + + + + + + + + + + + + + + + + + 

Liver + + + A + + + + + + + + + + + + + + + + + + + + + 

Mesentery + + + + + 

Pancreas A + + A + + + + + + + + + + + + + + + + + + + + + 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma 

Sarcoma, metastatic, eye X 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 

Leiomyoma 

Stomach, glandular A + + A + + + + + + + + + + + + + + + + + + + + + 

Tooth + 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 

Carcinoma, metastatic, lung X 

Schwannoma benign X 

Endocrine System 

Adrenal cortex + + + A + + + + + + + + + + + + + + + + + + + + + 

Adenoma 

Adrenal medulla + + + A + + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma malignant X 

Pheochromocytoma complex 

Pheochromocytoma benign X X 

Bilateral, pheochromocytoma benign 

Islets, pancreatic A + + A + + + + + + + + + + + + + + + + + + + + + 

Parathyroid gland M + + A + + + M + + + + + + + + + + + + + + + + + 

Pituitary gland + + + A + + + + + + + + + + + + + + + + + + + + + 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X 

Thyroid gland + + + A + + + A + + + + + + + + + + + + + + + + + 

C-cell, adenoma X X 

C-cell, carcinoma X 

Follicular cell, adenoma X 

General Body System 

None 

Genital System 

Clitoral gland + + + + + + + + + + + + + + + M + + + + + + + + + 

Adenoma X X 

Carcinoma 

Ovary + + + A + + + + + + + + + + + + + + + + + + + + + 

Uterus A + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X 

Leiomyoma 

Polyp stromal X X X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 123 

TABLE B2
 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 Total 

Carcass ID Number 3 3 0 0 0 0 1 2 2 2 2 3 3 4 4 5 1 1 2 2 3 3 4 4 4 Tissues/ 

5 8 2 3 5 6 5 0 3 4 5 1 2 5 6 0 3 4 2 7 4 7 0 1 8 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Mesentery + + + + + + + + 13 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X 1 

Sarcoma, metastatic, eye 1 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Leiomyoma X 1 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Tooth 1 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Carcinoma, metastatic, lung 1 

Schwannoma benign 1 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Adenoma X 1 

Adrenal medulla + + + + + + + + + + + + + + + + + + M + + + + + + 48 

Pheochromocytoma malignant 1 

Pheochromocytoma complex X 1 

Pheochromocytoma benign X X X X X 7 

Bilateral, pheochromocytoma benign X 1 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Parathyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Pars distalis, adenoma X X X X X X X X X X X X X X X X X X X X X X 40 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 48 

C-cell, adenoma X 3 

C-cell, carcinoma X X 3 

Follicular cell, adenoma 1 

General Body System 

None 

Genital System 

Clitoral gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Adenoma X 3 

Carcinoma X X X 3 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Histiocytic sarcoma 1 

Leiomyoma X 1 

Polyp stromal X 4 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

124 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B2
 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

0 3 4 5 6 6 6 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 

Number of Days on Study 0 8 7 8 1 1 3 4 5 5 6 7 7 8 8 9 9 0 0 2 3 3 3 3 3 

6 6 0 5 0 1 2 2 0 2 3 3 9 0 6 2 3 9 9 2 5 5 5 5 5 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Carcass ID Number 4 3 4 0 4 2 3 1 0 2 4 3 1 4 0 0 0 1 3 2 1 1 1 1 2 

3 0 2 1 9 1 6 7 7 6 7 3 1 4 8 4 9 0 9 9 2 6 8 9 8 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 

Lymph node + + 

Lymph node, bronchial M + + + + + + M + + M + + M + + M + M + M M + + + 

Lymph node, mandibular + + + + + + + + + + + + + + + + + + + + + + + M + 

Sarcoma, metastatic, eye X 

Sarcoma, metastatic, skin X 

Lymph node, mesenteric A + + A + + + + + + + + + + + + + + + + + + + + + 

Lymph node, mediastinal + + + + + + + + + + + + + + + + M + M + M M + + + 

Spleen A + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X 

Thymus + + + A M + + + + + + + + + + + + + + M + M + + + 

Integumentary System 

Mammary gland M + + + + + + + + + + + + + + + + + + + + + + + + 

Carcinoma 

Carcinoma, multiple X 

Fibroadenoma X X X X X X X X X X 

Fibroadenoma, multiple X X 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 

Basal cell adenoma X 

Sebaceous gland, adenoma X 

Subcutaneous tissue, fibroma 

Subcutaneous tissue, sarcoma X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Sarcoma, metastatic, eye X 

Skeletal muscle + 

Carcinoma, metastatic, lung X 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 

Astrocytoma benign X 

Respiratory System 

Larynx + + + + + + + + + + + + + + + + + + + + + + + + + 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 

Alveolar/bronchiolar adenoma X X 

Alveolar/bronchiolar adenoma, multiple X 

Alveolar/bronchiolar carcinoma X X 

Histiocytic sarcoma X 

Sarcoma, metastatic, eye X 

Sarcoma, metastatic, skin X 

Squamous cell carcinoma 

Mediastinum, alveolar/bronchiolar

 carcinoma, metastatic, lung X 

Nose + + + + + + + + + + + + + + + + + + + + + + + + + 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 

Special Senses System 

Eye + + 

Sarcoma X 

Lacrimal gland 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 125 

TABLE B2
 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 Total 

Carcass ID Number 3 3 0 0 0 0 1 2 2 2 2 3 3 4 4 5 1 1 2 2 3 3 4 4 4 Tissues/ 

5 8 2 3 5 6 5 0 3 4 5 1 2 5 6 0 3 4 2 7 4 7 0 1 8 Tumors 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lymph node 2 

Lymph node, bronchial + + M + + + + + + + + M + + + + + M M + + + + + M 37 

Lymph node, mandibular + + M + + + + M + + + + + + + + + M + + + M + + + 45 

Sarcoma, metastatic, eye 1 

Sarcoma, metastatic, skin 1 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Lymph node, mediastinal + + + + + + + + + + + + + + + + M + + + + + + + + 45 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Histiocytic sarcoma 1 

Thymus + + M + + + + + + + + + + + + + M + + + + + + + + 44 

Integumentary System 

Mammary gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Carcinoma X X X X 4 

Carcinoma, multiple 1 

Fibroadenoma X X X X X X X X X X 20 

Fibroadenoma, multiple X X X X X 7 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Basal cell adenoma 1 

Sebaceous gland, adenoma 1 

Subcutaneous tissue, fibroma X 1 

Subcutaneous tissue, sarcoma 1 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Sarcoma, metastatic, eye 1 

Skeletal muscle 1 

Carcinoma, metastatic, lung 1 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Astrocytoma benign 1 

Respiratory System 

Larynx + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Alveolar/bronchiolar adenoma X X X X X 7 

Alveolar/bronchiolar adenoma, multiple X 2 

Alveolar/bronchiolar carcinoma X X X X 6 

Histiocytic sarcoma 1 

Sarcoma, metastatic, eye 1 

Sarcoma, metastatic, skin 1 

Squamous cell carcinoma X 1 

Mediastinum, alveolar/bronchiolar

 carcinoma, metastatic, lung 1 

Nose + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Special Senses System 

Eye + 3 

Sarcoma 1 

Lacrimal gland + + 2 
 

 

 

 

126 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B2
 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

0 3 4 5 6 6 6 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 

Number of Days on Study 0 8 7 8 1 1 3 4 5 5 6 7 7 8 8 9 9 0 0 2 3 3 3 3 3 

6 6 0 5 0 1 2 2 0 2 3 3 9 0 6 2 3 9 9 2 5 5 5 5 5 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Carcass ID Number 4 3 4 0 4 2 3 1 0 2 4 3 1 4 0 0 0 1 3 2 1 1 1 1 2 

3 0 2 1 9 1 6 7 7 6 7 3 1 4 8 4 9 0 9 9 2 6 8 9 8 

Urinary System 

Kidney A + + A + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X 

Lipoma 

Urinary bladder A + + A + + + + + + + + + + + + M + + + + + + + + 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X 

Leukemia mononuclear X X X X X X X X 

Lymphoma malignant X 
 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 127 

TABLE B2
 

Individual Animal Tumor Pathology of Female Rats in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 Total 

Carcass ID Number 3 3 0 0 0 0 1 2 2 2 2 3 3 4 4 5 1 1 2 2 3 3 4 4 4 Tissues/ 

5 8 2 3 5 6 5 0 3 4 5 1 2 5 6 0 3 4 2 7 4 7 0 1 8 Tumors 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Histiocytic sarcoma 1 

Lipoma X 1 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1 

Leukemia mononuclear X X 10 

Lymphoma malignant 1 
 

128 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B3 

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Adrenal Medulla: Benign Pheochromocytoma a Overall rate 2/48 (4%) 1/49 (2%) 3/50 (6%) 8/48 (17%) 

Adjusted rateb 5.1% 3.1% 9.3% 26.4% c Terminal rate 0/27 (0%) 0/25 (0%) 1/26 (4%) 7/29 (24%) 

First incidence (days) 666 702 694 709 

d Life table test Pù0.006 Pù0.546N Pù0.483 Pù0.054 

d Logistic regression test Pù0.004 Pù0.498N Pù0.512 Pù0.043 

d Cochran-Armitage test Pù0.003 

d Fisher exact test Pù0.492N Pù0.520 Pù0.045 

Adrenal Medulla: Benign, Complex, or Malignant Pheochromocytoma 

Overall rate 2/48 (4%) 1/49 (2%) 4/50 (8%) 10/48 (21%) 

Adjusted rate 5.1% 3.1% 11.7% 31.5% 

Terminal rate 0/27 (0%) 0/25 (0%) 1/26 (4%) 8/29 (28%) 

First incidence (days) 666 702 685 663 

Life table test Pù0.001 Pù0.546N Pù0.325 Pù0.019 

Logistic regression test P<0.001 Pù0.498N Pù0.323 Pù0.014 

Cochran-Armitage test P<0.001 

Fisher exact test Pù0.492N Pù0.359 Pù0.014 

Clitoral Gland: Adenoma 

Overall rate 5/50 (10%) 7/48 (15%) 5/48 (10%) 3/49 (6%) 

Adjusted rate 16.0% 23.4% 17.8% 8.8% 

Terminal rate 2/28 (7%) 4/24 (17%) 3/26 (12%) 1/30 (3%) 

First incidence (days) 727 652 727 680 

Life table test Pù0.160N Pù0.289 Pù0.567 Pù0.360N 

Logistic regression test Pù0.182N Pù0.286 Pù0.581 Pù0.384N 

Cochran-Armitage test Pù0.189N 

Fisher exact test Pù0.351 Pù0.603 Pù0.369N 

Clitoral Gland: Carcinoma 

Overall rate 2/50 (4%) 3/48 (6%) 4/48 (8%) 3/49 (6%)
 

Adjusted rate 7.1% 9.3% 13.4% 10.0%
 

Terminal rate 2/28 (7%) 1/24 (4%) 2/26 (8%) 3/30 (10%)
 

First incidence (days) 735 (T) 610 694 735 (T)
 

Life table test Pù0.548 Pù0.439 Pù0.305 Pù0.532 

Logistic regression test Pù0.504 Pù0.459 Pù0.301 Pù0.532 

Cochran-Armitage test Pù0.499 

Fisher exact test Pù0.480 Pù0.319 Pù0.490 

Clitoral Gland: Adenoma or Carcinoma 

Overall rate 7/50 (14%) 10/48 (21%) 8/48 (17%) 6/49 (12%) 

Adjusted rate 22.5% 31.1% 27.0% 18.2% 

Terminal rate 4/28 (14%) 5/24 (21%) 5/26 (19%) 4/30 (13%) 

First incidence (days) 727 610 694 680 

Life table test Pù0.234N Pù0.207 Pù0.429 Pù0.480N 

Logistic regression test Pù0.280N Pù0.205 Pù0.432 Pù0.545N 

Cochran-Armitage test Pù0.291N 

Fisher exact test Pù0.266 Pù0.465 Pù0.516N 
 

Cobalt Sulfate Heptahydrate, NTP TR 471 129 

TABLE B3 

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Lung: Alveolar/bronchiolar Adenoma 

Overall rate 0/50 (0%) 1/49 (2%) 10/50 (20%) 9/50 (18%) 

Adjusted rate 0.0% 3.4% 36.4% 30.0% 

Terminal rate 0/28 (0%) 0/25 (0%) 9/26 (35%) 9/30 (30%) e First incidence (days) — 714 692 735 (T) 

Life table test Pù0.003 Pù0.468 P<0.001 Pù0.003 

Logistic regression test Pù0.001 Pù0.480 P<0.001 Pù0.003 

Cochran-Armitage test Pù0.002 

Fisher exact test Pù0.495 P<0.001 Pù0.001 

Lung: Alveolar/bronchiolar Carcinoma 

Overall rate 0/50 (0%) 2/49 (4%) 6/50 (12%) 6/50 (12%) 

Adjusted rate 0.0% 8.0% 20.2% 17.5% 

Terminal rate 0/28 (0%) 2/25 (8%) 4/26 (15%) 4/30 (13%) 

First incidence (days) — 735 (T) 694 610 

Life table test Pù0.033 Pù0.213 Pù0.015 Pù0.022 

Logistic regression test Pù0.023 Pù0.213 Pù0.015 Pù0.017 

Cochran-Armitage test Pù0.022 

Fisher exact test Pù0.242 Pù0.013 Pù0.013 

Lung: Alveolar/bronchiolar Adenoma or Carcinoma 

Overall rate 0/50 (0%) 3/49 (6%) 15/50 (30%) 15/50 (30%) 

Adjusted rate 0.0% 11.2% 50.6% 46.1% 

Terminal rate 0/28 (0%) 2/25 (8%) 12/26 (46%) 13/30 (43%) 

First incidence (days) — 714 692 610 

Life table test P<0.001 Pù0.101 P<0.001 P<0.001 

Logistic regression test P<0.001 Pù0.096 P<0.001 P<0.001 

Cochran-Armitage test P<0.001 

Fisher exact test Pù0.117 P<0.001 P<0.001 

Lung: Alveolar/bronchiolar Adenoma, Alveolar/bronchiolar Carcinoma, or Squamous Cell Carcinoma 

Overall rate 0/50 (0%) 3/49 (6%) 16/50 (32%) 16/50 (32%) 

Adjusted rate 0.0% 11.2% 54.1% 49.2% 

Terminal rate 0/28 (0%) 2/25 (8%) 13/26 (50%) 14/30 (47%) 

First incidence (days) — 714 692 610 

Life table test P<0.001 Pù0.101 P<0.001 P<0.001 

Logistic regression test P<0.001 Pù0.096 P<0.001 P<0.001 

Cochran-Armitage test P<0.001 

Fisher exact test Pù0.117 P<0.001 P<0.001 

Mammary Gland: Fibroadenoma 

Overall rate 22/50 (44%) 22/49 (45%) 20/50 (40%) 27/50 (54%) 

Adjusted rate 54.6% 67.3% 56.5% 70.5% 

Terminal rate 11/28 (39%) 15/25 (60%) 12/26 (46%) 19/30 (63%) 

First incidence (days) 569 610 510 386 

Life table test Pù0.291 Pù0.386 Pù0.524N Pù0.280 

Logistic regression test Pù0.147 Pù0.439 Pù0.435N Pù0.182 

Cochran-Armitage test Pù0.153 

Fisher exact test Pù0.545 Pù0.420N Pù0.212 
 

130 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B3 

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Mammary Gland: Fibroadenoma or Adenoma 

Overall rate 23/50 (46%) 22/49 (45%) 20/50 (40%) 27/50 (54%) 

Adjusted rate 55.6% 67.3% 56.5% 70.5% 

Terminal rate 11/28 (39%) 15/25 (60%) 12/26 (46%) 19/30 (63%) 

First incidence (days) 569 610 510 386 

Life table test Pù0.334 Pù0.452 Pù0.457N Pù0.343 

Logistic regression test Pù0.179 Pù0.535 Pù0.349N Pù0.244 

Cochran-Armitage test Pù0.186 

Fisher exact test Pù0.537N Pù0.343N Pù0.274 

Mammary Gland: Carcinoma 

Overall rate 3/50 (6%) 2/49 (4%) 5/50 (10%) 5/50 (10%) 

Adjusted rate 8.5% 5.4% 15.9% 15.9% 

Terminal rate 1/28 (4%) 0/25 (0%) 2/26 (8%) 4/30 (13%) 

First incidence (days) 638 526 695 693 

Life table test Pù0.247 Pù0.555N Pù0.333 Pù0.370 

Logistic regression test Pù0.216 Pù0.471N Pù0.341 Pù0.342 

Cochran-Armitage test Pù0.217 

Fisher exact test Pù0.510N Pù0.357 Pù0.357 

Mammary Gland: Adenoma or Carcinoma 

Overall rate 5/50 (10%) 2/49 (4%) 5/50 (10%) 5/50 (10%) 

Adjusted rate 13.2% 5.4% 15.9% 15.9% 

Terminal rate 1/28 (4%) 0/25 (0%) 2/26 (8%) 4/30 (13%) 

First incidence (days) 572 526 695 693 

Life table test Pù0.423 Pù0.280N Pù0.589 Pù0.621N 

Logistic regression test Pù0.390 Pù0.182N Pù0.626 Pù0.622 

Cochran-Armitage test Pù0.393 

Fisher exact test Pù0.226N Pù0.630N Pù0.630N 

Mammary Gland: Fibroadenoma, Adenoma, or Carcinoma 

Overall rate 25/50 (50%) 23/49 (47%) 24/50 (48%) 29/50 (58%) 

Adjusted rate 59.2% 68.1% 65.0% 74.0% 

Terminal rate 12/28 (43%) 15/25 (60%) 14/26 (54%) 20/30 (67%) 

First incidence (days) 569 526 510 386 

Life table test Pù0.324 Pù0.511 Pù0.528 Pù0.348 

Logistic regression test Pù0.160 Pù0.527N Pù0.513N Pù0.242 

Cochran-Armitage test Pù0.168 

Fisher exact test Pù0.459N Pù0.500N Pù0.274 

Pancreatic Islets: Adenoma 

Overall rate 4/49 (8%) 1/49 (2%) 0/50 (0%) 0/48 (0%)
 

Adjusted rate 14.3% 4.0% 0.0% 0.0%
 

Terminal rate 4/28 (14%) 1/25 (4%) 0/26 (0%) 0/30 (0%)
 

First incidence (days) 735 (T) 735 (T) — —
 

Life table test Pù0.049N Pù0.212N Pù0.071N Pù0.053N 

Logistic regression test Pù0.049N Pù0.212N Pù0.071N Pù0.053N 

Cochran-Armitage test Pù0.058N
 

Fisher exact test Pù0.181N Pù0.056N Pù0.061N 
 

Cobalt Sulfate Heptahydrate, NTP TR 471 131 

TABLE B3 

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Pancreatic Islets: Adenoma or Carcinoma 

Overall rate 6/49 (12%) 1/49 (2%) 1/50 (2%) 0/48 (0%) 

Adjusted rate 19.6% 4.0% 3.4% 0.0% 

Terminal rate 5/28 (18%) 1/25 (4%) 0/26 (0%) 0/30 (0%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

572 

Pù0.023N 

Pù0.025N 

Pù0.027N 

735 (T) 

Pù0.078N 

Pù0.069N 

Pù0.056N 

727 

Pù0.073N 

Pù0.059N 

Pù0.053N 

— 

Pù0.015N 

Pù0.019N 

Pù0.014N 

Pituitary Gland (Pars Distalis): Adenoma 

Overall rate 40/50 (80%) 39/49 (80%) 38/50 (76%) 40/49 (82%) 

Adjusted rate 86.8% 95.0% 90.3% 90.8% 

Terminal rate 22/28 (79%) 23/25 (92%) 22/26 (85%) 26/30 (87%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

569 

Pù0.350N 

Pù0.450 

Pù0.450 

420 

Pù0.336 

Pù0.451 

Pù0.579N 

510 

Pù0.530 

Pù0.474N 

Pù0.405N 

386 

Pù0.491N 

Pù0.449 

Pù0.520 

Thyroid Gland (C-cell): Adenoma 

Overall rate 6/49 (12%) 6/49 (12%) 7/50 (14%) 3/48 (6%) 

Adjusted rate 21.4% 19.1% 24.5% 8.3% 

Terminal rate 6/28 (21%) 3/25 (12%) 5/26 (19%) 1/30 (3%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

735 (T) 

Pù0.139N 

Pù0.177N 

Pù0.184N 

517 

Pù0.540 

Pù0.597 

Pù0.620N 

722 

Pù0.449 

Pù0.469 

Pù0.516 

611 

Pù0.220N 

Pù0.254N 

Pù0.254N 

Thyroid Gland (C-cell): Carcinoma 

Overall rate 3/49 (6%) 0/49 (0%) 2/50 (4%) 3/48 (6%) 

Adjusted rate 10.0% 0.0% 6.5% 9.3% 

Terminal rate 2/28 (7%) 0/25 (0%) 0/26 (0%) 2/30 (7%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

721 

Pù0.365 

Pù0.340 

Pù0.323 

— 

Pù0.147N 

Pù0.140N 

Pù0.121N 

709 

Pù0.528N 

Pù0.511N 

Pù0.490N 

686 

Pù0.651N 

Pù0.653 

Pù0.651 

Thyroid Gland (C-cell): Adenoma or Carcinoma 

Overall rate 9/49 (18%) 6/49 (12%) 9/50 (18%) 6/48 (13%) 

Adjusted rate 30.7% 19.1% 29.4% 17.0% 

Terminal rate 8/28 (29%) 3/25 (12%) 5/26 (19%) 3/30 (10%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

721 

Pù0.277N 

Pù0.339N 

Pù0.357N 

517 

Pù0.376N 

Pù0.326N 

Pù0.288N 

709 

Pù0.543 

Pù0.578 

Pù0.584N 

611 

Pù0.263N 

Pù0.303N 

Pù0.303N 
 

132 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B3 

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Uterus: Stromal Polyp 

Overall rate 8/50 (16%) 7/49 (14%) 11/50 (22%) 4/50 (8%) 

Adjusted rate 23.9% 20.0% 35.6% 13.3% 

Terminal rate 5/28 (18%) 3/25 (12%) 8/26 (31%) 4/30 (13%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

569 

Pù0.113N 

Pù0.146N 

Pù0.144N 

483 

Pù0.585N 

Pù0.467N 

Pù0.517N 

600 

Pù0.254 

Pù0.294 

Pù0.306 

735 (T) 

Pù0.159N 

Pù0.189N 

Pù0.178N 

Uterus: Stromal Polyp or Stromal Sarcoma 

Overall rate 9/50 (18%) 9/49 (18%) 11/50 (22%) 4/50 (8%) 

Adjusted rate 26.3% 23.6% 35.6% 13.3% 

Terminal rate 5/28 (18%) 3/25 (12%) 8/26 (31%) 4/30 (13%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

569 

Pù0.059N 

Pù0.071N 

Pù0.074N 

362 

Pù0.507 

Pù0.513N 

Pù0.584 

600 

Pù0.341 

Pù0.387 

Pù0.402 

735 (T) 

Pù0.108N 

Pù0.126N 

Pù0.117N 

All Organs: Mononuclear Cell Leukemia 

Overall rate 15/50 (30%) 16/49 (33%) 19/50 (38%) 10/50 (20%) 

Adjusted rate 37.4% 41.1% 47.7% 25.6% 

Terminal rate 5/28 (18%) 5/25 (20%) 7/26 (27%) 3/30 (10%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

385 

Pù0.107N 

Pù0.091N 

Pù0.095N 

483 

Pù0.367 

Pù0.529 

Pù0.473 

526 

Pù0.242 

Pù0.273 

Pù0.263 

470 

Pù0.217N 

Pù0.166N 

Pù0.178N 

All Organs: Benign Neoplasms 

Overall rate 45/50 (90%) 44/49 (90%) 47/50 (94%) 44/50 (88%) 

Adjusted rate 93.7% 95.6% 100.0% 97.8% 

Terminal rate 25/28 (89%) 23/25 (92%) 26/26 (100%) 29/30 (97%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

569 

Pù0.259N 

Pù0.483N 

Pù0.418N 

420 

Pù0.318 

Pù0.566 

Pù0.617N 

510 

Pù0.275 

Pù0.248 

Pù0.357 

386 

Pù0.420N 

Pù0.636N 

Pù0.500N 

All Organs: Malignant Neoplasms 

Overall rate 25/50 (50%) 23/49 (47%) 32/50 (64%) 27/50 (54%) 

Adjusted rate 58.3% 54.9% 72.0% 65.0% 

Terminal rate 11/28 (39%) 8/25 (32%) 14/26 (54%) 16/30 (53%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

385 

Pù0.472 

Pù0.341 

Pù0.337 

362 

Pù0.513 

Pù0.376N 

Pù0.459N 

355 

Pù0.129 

Pù0.119 

Pù0.113 

470 

Pù0.464 

Pù0.402 

Pù0.421 
 

133 

c 

Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B3 

Statistical Analysis of Primary Neoplasms in Female Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

All Organs: Benign or Malignant Neoplasms 

Overall rate 48/50 (96%) 47/49 (96%) 50/50 (100%) 46/50 (92%) 

Adjusted rate 96.0% 95.9% 100.0% 97.9% 

Terminal rate 26/28 (93%) 23/25 (92%) 26/26 (100%) 29/30 (97%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

385 

Pù0.216N 

Pù0.228N 

Pù0.180N 

362 

Pù0.310 

Pù0.655N 

Pù0.684N 

355 

Pù0.280 

Pù0.254 

Pù0.247 

386 

Pù0.365N 

Pù0.450N 

Pù0.339N 

(T)Terminal sacrifice 

a Number of neoplasm-bearing animals/number of animals examined. Denominator is number of animals examined microscopically for adrenal gland, 

clitoral gland, lung, pancreatic islets, pituitary gland, thyroid gland, and uterus; for other tissues, denominator is number of animals necropsied. b	 Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality 

Observed incidence at terminal kill 

d	 Beneath the chamber control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values corresponding 

to pairwise comparisons between the chamber controls and that exposed group. The life table test regards neoplasms in animals dying prior to terminal kill 

as being (directly or indirectly) the cause of death. The logistic regression test regards these lesions as nonfatal. The Cochran-Armitage and Fisher exact 

tests compare directly the overall incidence rates. For all tests, a negative trend or a lower incidence in an exposure group is indicated by N. e Not applicable; no neoplasms in animal group 
134 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B4a 

Historical Incidence of Alveolar/bronchiolar Neoplasms in Chamber Control Female F344/N Ratsa 

Incidence in Controls 

Study Adenoma Carcinoma Adenoma or Carcinoma 

Historical Incidence at Battelle Pacific Northwest Laboratories 

o-Chlorobenzalmalononitrile (CS2) 2/49 0/49 2/49 

Acetonitrile 0/48 0/48 0/48 

2-Chloroacetophenone 1/49 0/49 1/49 

l-Epinephrine Hydrochloride 0/50 0/50 0/50 

Chloroethane 0/50 0/50 0/50 

Hexachlorocyclopentadiene 1/50 0/50 1/50 

Ozone 0/50 0/50 0/50 

Overall Historical Incidence 

Total 7/650 (1.1%) 0/650 (0.0%) 7/650 (1.1%) 

Standard deviation 1.6% 1.6% 

Range 0%-4% 0%-4% 

a Data as of 12 May 1995 

TABLE B4b 

Historical Incidence of Neoplasms of the Adrenal Medulla in Chamber Control Female F344/N Ratsa 

Incidence in Controls 

Study Benign Pheochromocytoma Benign, Complex, 

or Malignant Pheochromocytomab 

Historical Incidence at Battelle Pacific Northwest Laboratories 

o-Chlorobenzalmalononitrile (CS2) 5/37 5/37 

Acetonitrile 1/48 1/48 

2-Chloroacetophenone 5/49 5/49 

l-Epinephrine Hydrochloride 1/50 1/50 

Chloroethane 1/35 1/35 

Hexachlorocyclopentadiene 6/47 6/47 

Ozone 6/50 6/50 

Overall Historical Incidence 

Total 35/608 (5.8%) 39/608 (6.4%) 

Standard deviation 4.9% 4.4% 

Range 0%-14% 2%-14% 

a Data as of 12 May 1995 b One unspecified pheochromocytoma is included in the overall incidence. 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 135 

TABLE B5 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydratea 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Moribund 19 20 20 17 

Natural deaths 3 4 4 3 

Survivors 

Terminal sacrifice 28 25 26 30 

Pregnant 1 

Animals examined microscopically 50 49 50 50 

Alimentary System 

Esophagus (50) (49) (50) (50) 

Inflammation, suppurative 1 (2%) 

Intestine large, cecum (48) (45) (47) (47) 

Necrosis 1 (2%) 

Liver (50) (49) (50) (49) 

Angiectasis 6 (12%) 4 (8%) 1 (2%) 2 (4%) 

Basophilic focus 39 (78%) 37 (76%) 45 (90%) 41 (84%) 

Clear cell focus 7 (14%) 6 (12%) 5 (10%) 11 (22%) 

Cyst 1 (2%) 

Degeneration, fatty 8 (16%) 9 (18%) 8 (16%) 2 (4%) 

Eosinophilic focus 4 (8%) 1 (2%) 1 (2%) 2 (4%) 

Hepatodiaphragmatic nodule 6 (12%) 6 (12%) 8 (16%) 4 (8%) 

Inflammation, chronic active 1 (2%) 

Mixed cell focus 11 (22%) 11 (22%) 18 (36%) 15 (31%) 

Necrosis 1 (2%) 

Regeneration 1 (2%) 2 (4%) 

Thrombosis 1 (2%) 

Bile duct, hyperplasia 5 (10%) 6 (12%) 9 (18%) 7 (14%) 

Centrilobular, necrosis 5 (10%) 6 (12%) 7 (14%) 5 (10%) 

Mesentery (5) (8) (6) (13) 

Inflammation, chronic active 1 (8%) 

Artery, inflammation, chronic active 1 (13%) 

Fat, necrosis 5 (100%) 6 (75%) 6 (100%) 12 (92%) 

Pancreas (49) (49) (50) (48) 

Atrophy 22 (45%) 11 (22%) 16 (32%) 16 (33%) 

Basophilic focus 3 (6%) 

Hyperplasia 1 (2%) 

Artery, inflammation 1 (2%) 

Salivary glands (50) (49) (50) (50) 

Atrophy 3 (6%) 4 (8%) 1 (2%) 

Stomach, forestomach (50) (49) (50) (50) 

Hyperplasia, basal cell 1 (2%) 

Hyperplasia, squamous 1 (2%) 

Necrosis 3 (6%) 2 (4%) 1 (2%) 1 (2%) 

Stomach, glandular (49) (49) (49) (48) 

Mineralization 1 (2%) 1 (2%) 1 (2%) 1 (2%) 

Necrosis 4 (8%) 1 (2%) 

Tooth (2) (1) 

Developmental malformation 1 (50%) 

Inflammation, chronic active 1 (50%) 1 (100%) 

a Number of animals examined microscopically at the site and the number of animals with lesion 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

136 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B5 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Cardiovascular System 

Blood vessel (1) 

Aorta, inflammation, chronic active 1 (100%) 

Heart (50) (49) (50) (50) 

Cardiomyopathy 29 (58%) 24 (49%) 29 (58%) 23 (46%) 

Atrium, thrombosis 1 (2%) 

Endocrine System 

Adrenal cortex (49) (49) (50) (49) 

Hyperplasia 21 (43%) 21 (43%) 25 (50%) 23 (47%) 

Hypertrophy 5 (10%) 3 (6%) 8 (16%) 9 (18%) 

Necrosis 1 (2%) 

Vacuolization cytoplasmic 11 (22%) 5 (10%) 10 (20%) 12 (24%) 

Adrenal medulla (48) (49) (50) (48) 

Hyperplasia 8 (17%) 7 (14%) 11 (22%) 13 (27%) 

Pituitary gland (50) (49) (50) (49) 

Pars distalis, angiectasis 2 (4%) 

Pars distalis, cyst 1 (2%) 

Pars distalis, hyperplasia 5 (10%) 5 (10%) 8 (16%) 5 (10%) 

Thyroid gland (49) (49) (50) (48) 

C-cell, hyperplasia 37 (76%) 42 (86%) 37 (74%) 38 (79%) 

Follicular cell, hyperplasia 2 (4%) 1 (2%) 

General Body System 

None 

Genital System 

Clitoral gland (50) (48) (48) (49) 

Hyperplasia 2 (4%) 1 (2%) 1 (2%) 2 (4%) 

Inflammation, chronic active 2 (4%) 6 (13%) 2 (4%) 6 (12%) 

Ovary (50) (49) (50) (49) 

Cyst 5 (10%) 1 (2%) 3 (6%) 

Uterus (50) (49) (50) (49) 

Hydrometra 2 (4%) 

Cervix, hypertrophy 1 (2%) 

Hematopoietic System 

Bone marrow (49) (49) (50) (50) 

Atrophy 2 (4%) 1 (2%) 

Hyperplasia, histiocytic 1 (2%) 1 (2%) 

Lymph node (2) (3) (3) (2) 

Renal, hemorrhage 1 (33%) 

Lymph node, mandibular (44) (40) (47) (45) 

Infiltration cellular, plasma cell 1 (2%) 

Lymph node, mesenteric (49) (49) (50) (48) 

Hemorrhage 1 (2%) 1 (2%) 

Infiltration cellular, eosinophil 1 (2%) 

Lymph node, mediastinal (43) (38) (43) (45) 

Hemorrhage 1 (2%) 1 (2%) 

Infiltration cellular, plasma cell 1 (2%) 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 137 

TABLE B5 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Hematopoietic System (continued) 

Spleen (49) (48) (50) (49) 

Accessory spleen 1 (2%) 2 (4%) 

Fibrosis 4 (8%) 2 (4%) 

Hematopoietic cell proliferation 3 (6%) 2 (4%) 4 (8%) 3 (6%) 

Hemorrhage 2 (4%) 3 (6%) 1 (2%) 

Hyperplasia, focal 1 (2%) 

Necrosis 1 (2%) 

Thrombosis 1 (2%) 

Integumentary System 

Mammary gland (50) (49) (50) (49) 

Galactocele 2 (4%) 1 (2%) 1 (2%) 

Inflammation, chronic active 1 (2%) 

Skin (50) (49) (50) (50) 

Hyperkeratosis 1 (2%) 

Inflammation, chronic 1 (2%) 

Inflammation, chronic active 4 (8%) 3 (6%) 3 (6%) 

Musculoskeletal System 

Bone (50) (49) (50) (50) 

Hyperostosis 4 (8%) 5 (10%) 5 (10%) 3 (6%) 

Nervous System 

Brain (50) (49) (50) (50) 

Gliosis 1 (2%) 

Hemorrhage 2 (4%) 

Respiratory System 

Larynx (50) (49) (50) (50) 

Epiglottis, metaplasia, squamous 1 (2%) 22 (45%) 39 (78%) 48 (96%) 

Lung (50) (49) (50) (50) 

Congestion, chronic 1 (2%) 

Cyst 1 (2%) 

Metaplasia, osseous 1 (2%) 

Metaplasia, squamous 1 (2%) 8 (16%) 3 (6%) 

Pigmentation, hemosiderin 1 (2%) 

Proteinosis 36 (73%) 49 (98%) 49 (98%) 

Alveolar epithelium, hyperplasia 15 (30%) 7 (14%) 20 (40%) 33 (66%) 

Alveolar epithelium, hyperplasia, atypical 3 (6%) 5 (10%) 

Alveolar epithelium, metaplasia 2 (4%) 47 (96%) 50 (100%) 49 (98%) 

Alveolus, inflammation, granulomatous 9 (18%) 47 (96%) 50 (100%) 49 (98%) 

Interstitium, fibrosis 7 (14%) 47 (96%) 50 (100%) 49 (98%) 

Perivascular, inflammation, chronic active 1 (2%) 
 

 

 

 

 

 

 

138 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE B5 

Summary of the Incidence of Nonneoplastic Lesions in Female Rats in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Respiratory System (continued) 

Nose (50) (49) (50) (50) 

Inflammation, suppurative 6 (12%) 10 (20%) 2 (4%) 4 (8%) 

Thrombosis 4 (8%) 4 (8%) 3 (6%) 2 (4%) 

Lateral wall, hyperplasia 1 (2%) 8 (16%) 26 (52%) 38 (76%) 

Lateral wall, metaplasia, squamous 1 (2%) 1 (2%) 4 (8%) 10 (20%) 

Nasolacrimal duct, metaplasia, squamous 1 (2%) 

Olfactory epithelium, atrophy 5 (10%) 29 (59%) 46 (92%) 47 (94%) 

Olfactory epithelium, metaplasia 2 (4%) 2 (4%) 3 (6%) 40 (80%) 

Respiratory epithelium, hyperplasia, focal 1 (2%) 

Respiratory epithelium, metaplasia, squamous 2 (4%) 

Pleura (1) 

Special Senses System 

Eye (1) (5) (3) (3) 

Cataract 1 (100%) 4 (80%) 3 (100%) 2 (67%) 

Cornea, edema 1 (20%) 

Retina, atrophy 1 (100%) 4 (80%) 3 (100%) 1 (33%) 

Urinary System 

Kidney (49) (49) (50) (48) 

Hyperplasia, stromal 1 (2%) 

Infarct 1 (2%) 2 (4%) 2 (4%) 

Nephropathy 47 (96%) 48 (98%) 45 (90%) 48 (100%) 

Renal tubule, hyperplasia 1 (2%) 
139 

TABLE C1 

TABLE C2 

TABLE C3 

TABLE C4a 

TABLE C4b 

TABLE C5 

APPENDIX C
 

SUMMARY OF LESIONS IN MALE MICE
 

IN THE 2-YEAR INHALATION STUDY
 

OF COBALT SULFATE HEPTAHYDRATE
 

Summary of the Incidence of Neoplasms in Male Mice 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 140
 

Individual Animal Tumor Pathology of Male Mice 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 144
 

Statistical Analysis of Primary Neoplasms in Male Mice 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 164
 

Historical Incidence of Alveolar/bronchiolar Neoplasms 

in Chamber Control Male B6C3F1
  Mice .................................. 168

Historical Incidence of Hemangiosarcoma of the Liver 

in Chamber Control Male B6C3F1
  Mice .................................. 168

Summary of the Incidence of Nonneoplastic Lesions in Male Mice 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 169
 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

140 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C1 

Summary of the Incidence of Neoplasms in Male Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydratea 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Accidental deaths 1 1 

Moribund 19 16 17 23 

Natural deaths 8 3 9 6 

Survivors 

Terminal sacrifice 22 31 24 20 

Animals examined microscopically 50 50 50 50 

Alimentary System 

Intestine large, cecum (45) (49) (45) (46) 

Intestine small, duodenum (45) (49) (42) (45) 

Polyp adenomatous 1 (2%) 

Intestine small, jejunum (45) (48) (42) (45) 

Carcinoma 1 (2%) 

Intestine small, ileum (45) (49) (43) (44) 

Liver (50) (50) (50) (50) 

Hemangiosarcoma 2 (4%) 4 (8%) 8 (16%) 7 (14%) 

Hepatoblastoma 4 (8%) 2 (4%) 2 (4%) 

Hepatocellular carcinoma 16 (32%) 32 (64%) 29 (58%) 26 (52%) 

Hepatocellular carcinoma, multiple 7 (14%) 1 (2%) 1 (2%) 4 (8%) 

Hepatocellular adenoma 14 (28%) 12 (24%) 16 (32%) 6 (12%) 

Hepatocellular adenoma, multiple 8 (16%) 9 (18%) 9 (18%) 7 (14%) 

Hepatocholangiocarcinoma 1 (2%) 

Histiocytic sarcoma 1 (2%) 1 (2%) 

Sarcoma 1 (2%) 

Mesentery (3) (4) (5) (4) 

Hemangiosarcoma 1 (25%) 1 (25%) 

Hepatocellular carcinoma, metastatic, liver 1 (20%) 

Hepatocholangiocarcinoma, metastatic, liver 1 (33%) 

Sarcoma, metastatic, liver 1 (25%) 

Pancreas (48) (50) (49) (50) 

Histiocytic sarcoma 1 (2%) 

Sarcoma, metastatic, liver 1 (2%) 

Stomach, forestomach (49) (50) (50) (50) 

Sarcoma, metastatic, liver 1 (2%) 

Squamous cell carcinoma 2 (4%) 

Squamous cell papilloma 1 (2%) 

Stomach, glandular (48) (50) (50) (50) 

Histiocytic sarcoma 1 (2%) 

Serosa, hepatocellular carcinoma, metastatic, 

liver 1 (2%) 

Cardiovascular System 

Heart (50) (50) (50) (50) 

Alveolar/bronchiolar carcinoma, metastatic, 

lung 1 (2%) 

Hemangiosarcoma 1 (2%) 

Sarcoma, metastatic, liver 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 141 

TABLE C1 

Summary of the Incidence of Neoplasms in Male Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Endocrine System 

Adrenal cortex (49) (50) (49) (50) 

Hepatocholangiocarcinoma, metastatic, liver 1 (2%) 

Adrenal medulla (48) (50) (49) (50) 

Hepatocholangiocarcinoma, metastatic, liver 1 (2%) 

Pheochromocytoma benign 1 (2%) 1 (2%) 

Islets, pancreatic (48) (50) (49) (50) 

Adenoma 1 (2%) 

Thyroid gland (49) (50) (50) (50) 

Follicular cell, adenoma 1 (2%) 

Follicular cell, carcinoma 1 (2%) 

General Body System 

Peritoneum (1) 

Hepatocholangiocarcinoma, metastatic, liver 1 (100%) 

Tissue NOS (1) (1) 

Hemangioma 1 (100%) 

Genital System 

Epididymis (49) (50) (50) (50) 

Leiomyoma 1 (2%) 

Testes (49) (50) (50) (50) 

Interstitial cell, adenoma 1 (2%) 

Hematopoietic System 

Bone marrow (49) (50) (50) (50) 

Hemangiosarcoma 2 (4%) 1 (2%) 

Lymph node (1) (1) (2) (2) 

Iliac, sarcoma, metastatic, liver 1 (50%) 

Renal, histiocytic sarcoma 1 (100%) 

Lymph node, bronchial (21) (24) (28) (25) 

Alveolar/bronchiolar carcinoma, metastatic, 

lung 1 (4%) 

Histiocytic sarcoma 1 (5%) 

Sarcoma, metastatic, liver 1 (4%) 

Lymph node, mesenteric (49) (48) (43) (47) 

Hemangiosarcoma 1 (2%) 

Hepatoblastoma, metastatic, liver 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Sarcoma, metastatic, liver 1 (2%) 

Lymph node, mediastinal (43) (28) (31) (35) 

Alveolar/bronchiolar carcinoma, metastatic, 

lung 1 (3%) 

Hepatocellular carcinoma, metastatic, liver 1 (2%) 

Hepatocholangiocarcinoma, metastatic, liver 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Sarcoma, metastatic, liver 1 (3%) 

Spleen (47) (50) (49) (50) 

Hemangiosarcoma 1 (2%) 3 (6%) 1 (2%) 2 (4%) 

Histiocytic sarcoma 1 (2%) 1 (2%) 

Thymus (28) (31) (33) (28) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

142 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C1 

Summary of the Incidence of Neoplasms in Male Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Integumentary System 

Skin 

Hemangioma

Hemangiosarcoma

Subcutaneous tissue, carcinoma

Subcutaneous tissue, sarcoma

(49) 

 1 (2%) 

 1 (2%) 

 1 (2%) 

(50) 

 1 (2%) 

(49) (49) 

 2 (4%) 

 1 (2%) 

Musculoskeletal System 

Bone 

Hemangiosarcoma

Skeletal muscle

Alveolar/bronchiolar carcinoma, metastatic, 

lung

Hepatocholangiocarcinoma, metastatic, liver

(50) 

(1)

 1 (100%) 

(50) (50) (50) 

 1 (2%) 

(2) 

 1 (50%) 

Nervous System 

None 

Respiratory System 

Larynx (48) (49) (48) (49) 

Lung (50) (50) (50) (50) 

Alveolar/bronchiolar adenoma 9 (18%) 9 (18%) 11 (22%) 13 (26%) 

Alveolar/bronchiolar adenoma, multiple 3 (6%) 2 (4%) 5 (10%) 

Alveolar/bronchiolar carcinoma 3 (6%) 5 (10%) 5 (10%) 9 (18%) 

Alveolar/bronchiolar carcinoma, multiple 1 (2%) 2 (4%) 2 (4%) 

Hemangiosarcoma, metastatic, liver 1 (2%) 

Hepatoblastoma, metastatic, liver 1 (2%) 

Hepatocellular carcinoma, metastatic, liver 5 (10%) 5 (10%) 6 (12%) 7 (14%) 

Histiocytic sarcoma 1 (2%) 

Nose (50) (50) (48) (49) 

Trachea (49) (50) (50) (50) 

Special Senses System 

Harderian gland (4) (4) (4) (6) 

Adenoma 4 (100%) 2 (50%) 2 (50%) 1 (17%) 

Urinary System 

Kidney (49) (50) (50) (50) 

Alveolar/bronchiolar carcinoma, metastatic, 

lung 1 (2%) 

Histiocytic sarcoma 1 (2%) 

Renal tubule, adenoma 1 (2%) 1 (2%) 

Systemic Lesions 

Multiple organsb (50) (50) (50) (50) 

Histiocytic sarcoma 1 (2%) 1 (2%) 

Lymphoma malignant 1 (2%) 1 (2%) 1 (2%) 
143 

c 

Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C1 

Summary of the Incidence of Neoplasms in Male Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Neoplasm Summary c Total animals with primary neoplasms 49 48 47 46 

Total primary neoplasms 77 94 92 95 

Total animals with benign neoplasms 30 30 30 27 

Total benign neoplasms 38 39 42 35 

Total animals with malignant neoplasms 33 39 41 38 

Total malignant neoplasms 39 55 50 60 

Total animals with metastatic neoplasms 7 6 7 9 

Total metastatic neoplasms 14 6 7 21 

a Number of animals examined microscopically at the site and the number of animals with neoplasm b Number of animals with any tissue examined microscopically 

Primary neoplasms: all neoplasms except metastatic neoplasms 
 

 

 

 

 

 

 

 

 

 

 

 

 

144 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

Number of Days on Study 

4 

4 

9 

4 

7 

0 

4 

8 

2 

4 

8 

6 

5 

1 

6 

5 

3 

3 

5 

4 

1 

5 

4 

1 

5 

5 

5 

5 

7 

5 

5 

7 

9 

6 

0 

0 

6 

0 

0 

6 

1 

1 

6 

4 

0 

6 

4 

3 

6 

4 

9 

6 

5 

1 

6 

5 

2 

6 

5 

2 

6 

7 

6 

6 

8 

0 

6 

8 

5 

6 

8 

6 

6 

8 

6 

Carcass ID Number 

0 

1 

5 

0 

0 

7 

0 

3 

3 

0 

4 

4 

0 

4 

2 

0 

4 

3 

0 

1 

6 

0 

1 

9 

0 

2 

9 

0 

0 

9 

0 

4 

7 

0 

0 

4 

0 

1 

0 

0 

5 

0 

0 

2 

7 

0 

4 

8 

0 

2 

8 

0 

1 

3 

0 

0 

1 

0 

3 

2 

0 

2 

6 

0 

0 

5 

0 

2 

5 

0 

1 

2 

0 

3 

8 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hemangiosarcoma 

Hepatoblastoma 

Hepatocellular carcinoma 

Hepatocellular carcinoma, multiple 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Hepatocholangiocarcinoma 

Histiocytic sarcoma 

Mesentery 

Hepatocholangiocarcinoma, metastatic, liver 

Pancreas 

Histiocytic sarcoma 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

Histiocytic sarcoma 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

A 

A 

A 

A 

A 

A 

A 

A 

+ 

X 

A 

A 

A 

A 

+ 

A 

+ 

+ 

A 

A 

A 

A 

+ 

X 

+ 

+ 

+ 

A 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

A 

+ 

+ 

+ 

A 

A 

A 

+ 

X 

+ 

+ 

+ 

+ 

+ 

A 

A 

A 

A 

A 

A 

A 

+ 

X 

X 

+ 

X 

A 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

A 

+ 

+ 

A 

A 

A 

A 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

A 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

X 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

A 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

X 

+ 

+ 

+ 

+ 

Cardiovascular System 

Heart + 

Endocrine System 

Adrenal cortex + 

Hepatocholangiocarcinoma, metastatic, liver 

Adrenal medulla + 

Hepatocholangiocarcinoma, metastatic, liver 

Pheochromocytoma benign 

Islets, pancreatic + 

Parathyroid gland M 

Pituitary gland + 

Thyroid gland + 

General Body System 

Peritoneum 

Hepatocholangiocarcinoma, metastatic, liver 

Tissue NOS 

Hemangioma 

+ 

+ 

+ 

+ 

I 

+ 

+ 

+ 

A 

A 

A 

A 

A 

A 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

I 

+ 

+ 

+ 

+ 

X 

+ 

X 

A 

M 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+: Tissue examined microscopically M: Missing tissue X: Lesion present 

A: Autolysis precludes examination I: Insufficient tissue Blank: Not examined 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 145 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 0 1 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

8 1 5 3 3 3 3 3 3 3 3 3 3 3 3 4 4 4 4 4 4 4 4 4 4 

0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 Total 

Carcass ID Number 2 3 4 0 1 1 1 2 2 2 2 3 3 4 4 0 0 0 1 3 3 3 3 4 4 Tissues/ 

2 6 0 8 1 4 8 0 1 3 4 0 5 1 5 2 3 6 7 1 4 7 9 6 9 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Gallbladder + + M + + + + + + + + + + + + + M M + + + + + + + 37 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 45 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 45 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 45 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 45 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma X X 2 

Hepatoblastoma X 4 

Hepatocellular carcinoma X X X X X 16 

Hepatocellular carcinoma, multiple X X X X X 7 

Hepatocellular adenoma X X X X X X X X X X X 14 

Hepatocellular adenoma, multiple X X X X 8 

Hepatocholangiocarcinoma 1 

Histiocytic sarcoma X 1 

Mesentery + + 3 

Hepatocholangiocarcinoma, metastatic, liver 1 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Histiocytic sarcoma X 1 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + 46 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Histiocytic sarcoma X 1 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Hepatocholangiocarcinoma, metastatic, liver 1 

Adrenal medulla + + + + + + + + + + + + + + + + + + M + + + + + + 48 

Hepatocholangiocarcinoma, metastatic, liver 1 

Pheochromocytoma benign X 1 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Parathyroid gland M M M M M + M M M + + M M + + M M + + + M + + M + 19 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

General Body System 

Peritoneum 1 

Hepatocholangiocarcinoma, metastatic, liver 1 

Tissue NOS + 1 

Hemangioma X 1 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

146 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

4 4 4 4 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

Number of Days on Study 4 

9 

7 

0 

8 

2 

8 

6 

1 

6 

3 

3 

4 

1 

4 

1 

5 

5 

7 

5 

7 

9 

0 

0 

0 

0 

1 

1 

4 

0 

4 

3 

4 

9 

5 

1 

5 

2 

5 

2 

7 

6 

8 

0 

8 

5 

8 

6 

8 

6 

0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 

Carcass ID Number 1 0 3 4 4 4 1 1 2 0 4 0 1 5 2 4 2 1 0 3 2 0 2 1 3 

5 7 3 4 2 3 6 9 9 9 7 4 0 0 7 8 8 3 1 2 6 5 5 2 8 

Genital System 

Epididymis 

Penis 

+ + A + + + + + + + + + + + + + + + + + + + + + + 

+ 

Preputial gland 

Prostate 

+ + A 

+ + A 

+ + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + A + + + + + + + + + 

Seminal vesicle + + A + + + A + + + + + + + + + + + + + + + + + + 

Testes + + A + + + + + + + + + + + + + + + + + + + + + + 

Hematopoietic System 

Bone marrow + + A + + + + + + + + + + + + + + + + + + + + + + 

Lymph node 

Renal, histiocytic sarcoma 

Lymph node, bronchial 

Histiocytic sarcoma 

Lymph node, mandibular 

Lymph node, mesenteric 

Hepatoblastoma, metastatic, liver 

Histiocytic sarcoma 

Lymph node, mediastinal 

Hepatocellular carcinoma, metastatic, liver 

Hepatocholangiocarcinoma, metastatic, liver 

Histiocytic sarcoma 

Spleen 

Hemangiosarcoma 

Histiocytic sarcoma 

Thymus 

+ + M + M + M M M M + + M I + M M + + M + M M M M 

+ M A M M M M + M M M + + M M + M M + M M + M + + 

+ + A + + + + + + + + + + + + + + + + + + + + + + 

X 

+ M A M + + + + + + + M + + + + + + + + + + + + + 

X 

X 

+ + A A + + + + + + + + + + + + + + + + + + + + + 

X 

M + A + M + + M + M + + + + + M M + + + M I M M M 

Integumentary System 

Mammary gland 

Skin 

M M M M M M M M M M M M M M M M M M M M M M M M M 

+ + A + + + + + + + + + + + + + + + + + + + + + + 

Hemangioma 

Hemangiosarcoma 

Subcutaneous tissue, sarcoma X 

X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Skeletal muscle + 

Hepatocholangiocarcinoma, metastatic, liver X 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 

Respiratory System 

Larynx + + A + + + + + + + + + + + + + + + + + + + + + +
 

Lung + + + + + + + + + + + + + + + + + + + + + + + + +
 

Alveolar/bronchiolar adenoma X X X X 

Alveolar/bronchiolar carcinoma X 

Alveolar/bronchiolar carcinoma, multiple X 

Hepatoblastoma, metastatic, liver X 

Hepatocellular carcinoma, metastatic, liver X X X X 

Histiocytic sarcoma 

Nose + + + + + + + + + + + + + + + + + + + + + + + + +
 

Trachea + + A + + + + + + + + + + + + + + + + + + + + + +
 
 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 147 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 0 1 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

8 1 5 3 3 3 3 3 3 3 3 3 3 3 3 4 4 4 4 4 4 4 4 4 4 

0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 Total 

Carcass ID Number 2 3 4 0 1 1 1 2 2 2 2 3 3 4 4 0 0 0 1 3 3 3 3 4 4 Tissues/ 

2 6 0 8 1 4 8 0 1 3 4 0 5 1 5 2 3 6 7 1 4 7 9 6 9 Tumors 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Penis 1 

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Prostate + + + + + + + M + + + + + + I + + + + + + + + + + 46 

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Lymph node + 1 

Renal, histiocytic sarcoma X 1 

Lymph node, bronchial + + M I M M M + + + M M + I + M M M M + M + + + M 21 

Histiocytic sarcoma X 1 

Lymph node, mandibular M M + + + + + + + M + M + M M M M + M M M + + M + 22 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Hepatoblastoma, metastatic, liver 1 

Histiocytic sarcoma X 1 

Lymph node, mediastinal + + M + + + + + + M + + + + + M + + + + + + + + + 43 

Hepatocellular carcinoma, metastatic, liver 1 

Hepatocholangiocarcinoma, metastatic, liver 1 

Histiocytic sarcoma X 1 

Spleen + + M + + + + + + + + + + + + + + + + + + + + + + 47 

Hemangiosarcoma 1 

Histiocytic sarcoma X 1 

Thymus + M + + M + M + + + M M + I + M + I M + + + M + + 28 

Integumentary System 

Mammary gland M M M M M M M M M M M M M M M M M M M M M M M M M 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Hemangioma X 1 

Hemangiosarcoma 1 

Subcutaneous tissue, sarcoma 1 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Skeletal muscle 1 

Hepatocholangiocarcinoma, metastatic, liver 1 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 

Respiratory System 

Larynx + + + + + + + + + + + + + + + + + + + M + + + + + 48 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Alveolar/bronchiolar adenoma X X X X X 9 

Alveolar/bronchiolar carcinoma X X 3 

Alveolar/bronchiolar carcinoma, multiple 1 

Hepatoblastoma, metastatic, liver 1 

Hepatocellular carcinoma, metastatic, liver X 5 

Histiocytic sarcoma X 1 

Nose + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 49 

50 
 

 

 

 

148 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

4 4 4 4 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

Number of Days on Study 4 7 8 8 1 3 4 4 5 7 7 0 0 1 4 4 4 5 5 5 7 8 8 8 8 

9 0 2 6 6 3 1 1 5 5 9 0 0 1 0 3 9 1 2 2 6 0 5 6 6 

0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 

Carcass ID Number 1 0 3 4 4 4 1 1 2 0 4 0 1 5 2 4 2 1 0 3 2 0 2 1 3 

5 7 3 4 2 3 6 9 9 9 7 4 0 0 7 8 8 3 1 2 6 5 5 2 8 

Special Senses System 

Eye + 

Harderian gland + + + 

Adenoma X X X 

Urinary System 

Kidney + + A + + + + + + + + + + + + + + + + + + + + + + 

Urinary bladder + + A A + + A + + + + + + + + A + + + + + + + + + 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma 

Lymphoma malignant X 
 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 149 

TABLE C2 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 0 1 2 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

8 1 5 3 3 3 3 3 3 3 3 3 3 3 3 4 4 4 4 4 4 4 4 4 4 

0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 Total 

Carcass ID Number 2 3 4 0 1 1 1 2 2 2 2 3 3 4 4 0 0 0 1 3 3 3 3 4 4 Tissues/ 

2 6 0 8 1 4 8 0 1 3 4 0 5 1 5 2 3 6 7 1 4 7 9 6 9 Tumors 

Special Senses System 

Eye 1 

Harderian gland + 4 

Adenoma X 4 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Urinary bladder + + + + + + + + + + + + + + + + + + + + + + + + + 46 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma X 1 

Lymphoma malignant 1 
 

 

 

 

 

 

 

 

 

 

 

 

150 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C2
 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

Number of Days on Study 

4 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 

6 0 4 8 9 1 1 1 4 5 5 6 6 8 8 9 0 2 2 3 3 3 3 3 3 

0 1 1 3 9 1 4 7 9 2 3 6 6 5 7 0 9 0 6 3 3 3 3 3 3 

Carcass ID Number 

2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 

1 2 0 1 4 0 4 1 3 4 4 0 3 3 2 1 2 0 2 0 0 0 0 1 1 

3 5 1 6 2 5 1 0 0 8 7 9 7 2 4 9 6 7 1 2 3 4 6 1 5 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Polyp adenomatous 

Intestine small, jejunum 

Carcinoma 

Intestine small, ileum 

Liver 

Hemangiosarcoma 

Hepatocellular carcinoma 

Hepatocellular carcinoma, multiple 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Hemangiosarcoma 

Pancreas 

Salivary glands 

Stomach, forestomach 

Squamous cell carcinoma 

Stomach, glandular 

Serosa, hepatocellular carcinoma, metastatic, 

liver 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + M A + + M M + + M + + + + + M + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + A + + + + + + + + + + + + + + + + 

+ + + + + + + + A + + + + + + + + + + + + + + + + 

+ + + + A + + + A + + + + + + + + + + + + + + + + 

+ + + + + + + + A + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X 

X X X X X X X X X X X X X X X X X X X X 

X 

X X X X X 

X X X 

+ + + 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 

Hemangiosarcoma 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma 

Parathyroid gland + + M M + M M + + I M M + + + + + + + + + + + + + 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

General Body System 

None 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Prostate + + + + + + + + + + + + + + + + + + + + + + + + + 

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + + 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 

Interstitial cell, adenoma 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 151 

TABLE C2
 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

3 3 3 3 3 3 3 3 3 3 3 3 4 4 4 4 4 4 4 4 4 4 4 4 4 

2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 Total 

Carcass ID Number 2 2 2 3 3 3 3 3 4 4 4 5 0 1 1 1 1 2 2 2 3 3 4 4 4 Tissues/ 

3 8 9 1 3 4 5 8 0 5 9 0 8 2 4 7 8 0 2 7 6 9 3 4 6 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Gallbladder + + + + M + + + + + + + + + + + + + M + + + + M + 41 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Polyp adenomatous X 1 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Carcinoma X 1 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma X X 4 

Hepatocellular carcinoma X X X X X X X X X X X X 32 

Hepatocellular carcinoma, multiple 1 

Hepatocellular adenoma X X X X X X X 12 

Hepatocellular adenoma, multiple X X X X X X 9 

Mesentery + 4 

Hemangiosarcoma 1 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Squamous cell carcinoma X 2 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Serosa, hepatocellular carcinoma, metastatic, 

liver 1 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma X 1 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X 1 

Parathyroid gland M M + M M + + M + + + + + + + M + + + + + M + + + 36 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

General Body System 

None 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Prostate + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Interstitial cell, adenoma X 1 
 

 

 

 

 

 

 

 

 

 

 

 

152 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C2
 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

4 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 

Number of Days on Study 6 

0 

0 

1 

4 

1 

8 

3 

9 

9 

1 

1 

1 

4 

1 

7 

4 

9 

5 

2 

5 

3 

6 

6 

6 

6 

8 

5 

8 

7 

9 

0 

0 

9 

2 

0 

2 

6 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 

Carcass ID Number 1 2 0 1 4 0 4 1 3 4 4 0 3 3 2 1 2 0 2 0 0 0 0 1 1 

3 5 1 6 2 5 1 0 0 8 7 9 7 2 4 9 6 7 1 2 3 4 6 1 5 

Hematopoietic System 

Bone marrow 

Hemangiosarcoma 

Lymph node 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Hemangiosarcoma 

Lymph node, mediastinal 

Spleen 

Hemangiosarcoma 

Thymus 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

+ 

M M + + + + + + + M + + + M + M M + + M M M M M + 

M M M M M + M M M + M + M M M M + + M + + M M + M 

+ + + + + + + + + + + + + + + + + + + + M + + + + 

M + + + + M + + M + M + + M M + M M M + + M + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X 

M M I + + M + + M + + + + M M M M + M + + + + + + 

Integumentary System 

Mammary gland 

Skin 

Hemangiosarcoma 

M M M M M M M M M M M M M M M M M M M M M M M M M 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Nose 

Trachea 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X 

X 

X 

X X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Special Senses System 

Ear 

Harderian gland 

Adenoma 

+ + 

+ 

+ 

X 

Urinary System 

Kidney 

Renal tubule, adenoma 

Urinary bladder 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + M + + + + + + + + + + + + 

Systemic Lesions 

Multiple organs 

Lymphoma malignant 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 
 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 153 

TABLE C2
 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 2 Total 

Carcass ID Number 2 

3 

2 

8 

2 

9 

3 

1 

3 

3 

3 

4 

3 

5 

3 

8 

4 

0 

4 

5 

4 

9 

5 

0 

0 

8 

1 

2 

1 

4 

1 

7 

1 

8 

2 

0 

2 

2 

2 

7 

3 

6 

3 

9 

4 

3 

4 

4 

4 

6 

Tissues/ 

Tumors 

Hematopoietic System 

Bone marrow 

Hemangiosarcoma 

Lymph node

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Hemangiosarcoma 

Lymph node, mediastinal 

Spleen 

Hemangiosarcoma 

Thymus 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

+ M + M M M + + M + M + M + M M M + M M + M + I M 

M M + + + + + + + M M + M M M M + + M M + + + M M 

+ + + + + + + M + + + + + + + + + + + + + + + + + 

X

M M + M M + M M + M M M + + + M M M + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

M + + + + M M + + + + M M + M + + + M + + I + M + 

50 

2 

1 

24 

21 

48 

1 

28 

50 

3 

31 

Integumentary System 

Mammary gland 

Skin 

Hemangiosarcoma 

M M M M M M M M M M M M M M M M M M M M M M M M M 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

50 

1 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Nose 

Trachea 

+ + + + I + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X

X X 

X X X X

X X X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

49 

50 

9 

3 

5 

5 

50 

50 

Special Senses System 

Ear

Harderian gland 

Adenoma 

+

X

 1 

4 

2 

Urinary System 

Kidney 

Renal tubule, adenoma 

Urinary bladder 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

50 

1 

49 

Systemic Lesions 

Multiple organs 

Lymphoma malignant

+ + + + + + + + + + + + + + + + + + + + + + + + + 50 

1 
 

 

 

 

 

 

 

 

 

 

 

 

154 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C2
 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

Number of Days on Study 

4 4 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 7 7 7 

4 8 0 1 2 3 4 5 6 6 9 1 3 4 4 5 5 7 8 8 8 9 0 0 0 

0 3 5 0 3 4 8 5 0 1 7 0 9 5 5 2 3 0 1 1 7 9 5 6 8 

Carcass ID Number 

4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 

3 4 0 0 4 0 1 2 4 4 4 2 3 3 3 3 4 0 1 3 3 4 2 3 1 

7 2 8 4 5 9 6 4 1 6 7 8 8 2 4 5 0 6 3 0 6 8 3 1 9 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hemangiosarcoma 

Hepatoblastoma 

Hepatocellular carcinoma 

Hepatocellular carcinoma, multiple 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Hepatocellular carcinoma, metastatic, liver 

Pancreas 

Salivary glands 

Stomach, forestomach 

Squamous cell papilloma 

Stomach, glandular 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

A A + + A M A + + + + + + + + + A A A + + + M + + 

+ + + + + + A + + + + + + + + + A A A + + + + + + 

+ M + + + + A + + + + + + + + + + A A + I + + + + 

+ A + + + + A + + + + + + + + + A A A + + + + + + 

A A + + M + A + + + + + + + + + A A A A + + + + + 

A A + + A + A + + + + + + + + + A A A A + + + + + 

+ A + + A + A + + + + + + + + + A A A A + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X 

X 

X X X X X X X X X X X X X X X X X X X 

X X X X X X 

X X 

+ + 

+ + + M + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Cardiovascular System 

Blood vessel + 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 

Islets, pancreatic + + + M + + + + + + + + + + + + + + + + + + + + + 

Parathyroid gland M M M + + M M M + M M + M M + M M + + M + + + M M 

Pituitary gland + + + + + + + + + + + + + + + + A + + + + + + + + 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Follicular cell, carcinoma 

General Body System 

Tissue NOS + 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 

Leiomyoma X 

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Prostate + + + + + + + M + + + I + M + + + + A + + + + + + 

Seminal vesicle + + + + + + M + + + + + + + + + + + + + + + + + + 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 
 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 155 

TABLE C2
 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 0 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

9 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 4 4 4 4 4 4 4 4 

4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 Total 

Carcass ID Number 1 0 0 0 1 1 1 1 2 2 2 2 2 2 3 4 5 0 0 1 1 2 3 4 4 Tissues/ 

1 1 2 5 0 2 4 7 0 1 2 5 6 7 3 3 0 3 7 5 8 9 9 4 9 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Gallbladder + + M M + + + + + + + + + + + + + + + + + + + + M 38 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 46 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 45 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 45 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 42 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 42 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 43 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma X X X 8 

Hepatoblastoma X 2 

Hepatocellular carcinoma X X X X X X X X X X 29 

Hepatocellular carcinoma, multiple X 1 

Hepatocellular adenoma X X X X X X X X X X 16 

Hepatocellular adenoma, multiple X X X X X X X 9 

Mesentery + + + 5 

Hepatocellular carcinoma, metastatic, liver X 1 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Squamous cell papilloma X 1 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Cardiovascular System 

Blood vessel + + 3 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + M + + + + 49 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + M + + + + 49 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Parathyroid gland + M + + M + + + + M M + M M M M M M M + + + + + + 24 

Pituitary gland + + + + + + + + + + + + + + + + + + + + M + + + + 48 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Follicular cell, carcinoma X 1 

General Body System 

Tissue NOS 1 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Leiomyoma 1 

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Prostate + + + M + + + M + + + + + + + + + + + + + + + + + 44 

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 50 
 

 

 

 

 

 

 

 

 

 

156 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C2
 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

4 4 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 7 7 7 

Number of Days on Study 4 8 0 1 2 3 4 5 6 6 9 1 3 4 4 5 5 7 8 8 8 9 0 0 0 

0 3 5 0 3 4 8 5 0 1 7 0 9 5 5 2 3 0 1 1 7 9 5 6 8 

4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 

Carcass ID Number 3 4 0 0 4 0 1 2 4 4 4 2 3 3 3 3 4 0 1 3 3 4 2 3 1 

7 2 8 4 5 9 6 4 1 6 7 8 8 2 4 5 0 6 3 0 6 8 3 1 9 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 

Lymph node + + 

Lymph node, bronchial M M M M M + M M M + + + + + + + + A + + + M + + M 

Lymph node, mandibular + M M M M M M + + + + M M M + M + + M + M M M + + 

Lymph node, mesenteric + + + M + M A + + + + + + + + + + A A + + + + + + 

Lymph node, mediastinal + + + + M + + M M M + + + + M + + + + M + + M M M 

Spleen + + + + + + + + + + + + + + + + + A + + + + + + + 

Hemangiosarcoma X 

Thymus M M M M + M M + M M + M + + + + M + M + + I + M M 

Integumentary System 

Mammary gland M M M M M M M M M M M M M M M M M M M M M M M M M 

Skin + + + + + + + + + + + + + + + I + + + + + + + + + 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 

Respiratory System 

Larynx + + + + + + + + + + + + + + + + + A + + + + + + + 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 

Alveolar/bronchiolar adenoma X X X X X X 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma X 

Alveolar/bronchiolar carcinoma, multiple X 

Hepatocellular carcinoma, metastatic, liver X X X X X X 

Nose + + + + + + + + + + + + + + + + A A + + + + + + + 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 

Special Senses System 

Eye 

Harderian gland 

Adenoma 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 

Ureter 

Urinary bladder + + + + + + A + + + + I + + + + A A A + + + + + + 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Lymphoma malignant X 
 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 157 

TABLE C2
 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 0 

9 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 4 Total 

Carcass ID Number 1 

1 

0 

1 

0 

2 

0 

5 

1 

0 

1 

2 

1 

4 

1 

7 

2 

0 

2 

1 

2 

2 

2 

5 

2 

6 

2 

7 

3 

3 

4 

3 

5 

0 

0 

3 

0 

7 

1 

5 

1 

8 

2 

9 

3 

9 

4 

4 

4 

9 

Tissues/ 

Tumors 

Hematopoietic System 

Bone marrow 

Lymph node

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Hemangiosarcoma

Thymus 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

M M + + M M + + + + + M + + M M M M + + + + + M M 

+ M M + + + + + + + + + M M + + + M M + M + + + + 

+ M + + + + + + + + + + + + + M + + + + + + + + + 

+ + + + + M + + M + M M + M M + M I M + + M + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ M + + + + + M + + + + + + + + + + + + M + + + + 

50 

2 

28 

29 

43 

31 

49 

1 

33 

Integumentary System 

Mammary gland 

Skin 

M M M M M M M M M M M M M M M M M M M M M M M M M 

+ + + + + + + + + + + + + + + + + + + + + + + + + 49 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma 

Alveolar/bronchiolar carcinoma, multiple 

Hepatocellular carcinoma, metastatic, liver

Nose 

Trachea 

+ + + + + I + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X 

X X 

X X X X

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

48 

50 

11 

2 

5 

2 

6 

48 

50 

Special Senses System 

Eye 

Harderian gland 

Adenoma 

+

+ + 

X 

+ 

X

+

 1 

4 

2 

Urinary System 

Kidney 

Ureter 

Urinary bladder 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+

+ + + + + + + + + + + + + + + + + + + + + + + + + 

50 

1 

45 

Systemic Lesions 

Multiple organs 

Lymphoma malignant

+ + + + + + + + + + + + + + + + + + + + + + + + + 50 

1 
 

 

 

 

 

 

 

 

 

 

 

 

158 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C2
 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

Number of Days on Study 

3 4 5 5 5 5 5 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 

9 1 0 2 2 3 3 4 5 5 5 5 6 6 6 6 0 1 1 1 1 2 3 4 5 

9 4 2 4 7 3 6 1 2 5 5 5 0 1 1 6 8 1 3 8 8 2 9 9 2 

Carcass ID Number 

6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

4 0 2 1 1 0 2 1 2 1 3 4 1 4 4 4 4 0 0 3 3 4 4 3 3 

7 6 6 6 4 2 2 2 1 8 9 1 3 5 8 4 3 4 5 2 6 0 6 1 0 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hemangiosarcoma 

Hepatoblastoma 

Hepatocellular carcinoma 

Hepatocellular carcinoma, multiple 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Histiocytic sarcoma 

Sarcoma 

Mesentery 

Hemangiosarcoma 

Sarcoma, metastatic, liver 

Pancreas 

Sarcoma, metastatic, liver 

Salivary glands 

Stomach, forestomach 

Sarcoma, metastatic, liver 

Stomach, glandular 

Tooth 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + M + + + + + M + M A + M + M A + + A + M A + 

+ + + + + + + + + + + + A + + + + A + + A + + + + 

+ + + + + + + + + + + + A + + + A A + + + + + + + 

+ + + + + + + + + + + A A + + + + A + + A + + + + 

+ + + + + + + + + + + + A + + + A A + + A + + A + 

+ + + + + + + + + + + + A + + + A A + + A + + A + 

+ + + + + + + + + + + A A + + + A A + + A + + A + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X 

X X X X X X X X X X X X X X X 

X X 

X X X X 

X X 

X 

+ + 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Cardiovascular System 

Heart 

Alveolar/bronchiolar carcinoma, metastatic, lung 

Sarcoma, metastatic, liver 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

X 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma benign 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 

Parathyroid gland + M M M + + + + + + + M + + M + M + M + M M + M + 

Pituitary gland + + + M + + + + + + + + + + + + + M + M + + + + + 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Follicular cell, adenoma 

General Body System 

None 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 

Preputial gland + + + + + + + + + + + + + + + + M + + + + + + + + 

Prostate + + + + + + + + + + + + + + + + + + + + + + + A + 

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + A + 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 159 

TABLE C2
 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

Number of Days on Study 

6 

5 

3 

6 

7 

3 

6 

7 

8 

6 

8 

6 

7 

1 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

4 

7 

3 

4 

7 

3 

4 

7 

3 

4 

7 

3 

4 

7 

3 

4 

7 

3 

4 

7 

3 

4 

Carcass ID Number 

6 

2 

4 

6 

3 

3 

6 

3 

8 

6 

3 

4 

6 

2 

3 

6 

0 

1 

6 

0 

3 

6 

0 

7 

6 

0 

9 

6 

1 

0 

6 

2 

0 

6 

2 

5 

6 

2 

8 

6 

3 

5 

6 

3 

7 

6 

4 

2 

6 

4 

9 

6 

0 

8 

6 

1 

1 

6 

1 

5 

6 

1 

7 

6 

1 

9 

6 

2 

7 

6 

2 

9 

6 

5 

0 

Total 

Tissues/ 

Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Gallbladder + + + + + M + M + + + + + + + + + + + + + + + + + 38 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 46 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 45 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 45 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 44 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma X X X X 7 

Hepatoblastoma X X 2 

Hepatocellular carcinoma X X X X X X X X X X X 26 

Hepatocellular carcinoma, multiple X X 4 

Hepatocellular adenoma X X 6 

Hepatocellular adenoma, multiple X X X X X 7 

Histiocytic sarcoma X 1 

Sarcoma 1 

Mesentery + + 4 

Hemangiosarcoma X 1 

Sarcoma, metastatic, liver 1 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Sarcoma, metastatic, liver 1 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Sarcoma, metastatic, liver 1 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Tooth + 1 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Alveolar/bronchiolar carcinoma, metastatic, lung 1 

Sarcoma, metastatic, liver 1 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pheochromocytoma benign X 1 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Parathyroid gland M M M M M + + + + + + + M M M + M + + + + + M + + 30 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Follicular cell, adenoma X 1 

General Body System 

None 

Genital System 

Epididymis + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Preputial gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Prostate + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Seminal vesicle + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Testes + + + + + + + + + + + + + + + + + + + + + + + + + 50 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

160 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C2
 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

3 4 5 5 5 5 5 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 

Number of Days on Study 9 1 0 2 2 3 3 4 5 5 5 5 6 6 6 6 0 1 1 1 1 2 3 4 5 

9 4 2 4 7 3 6 1 2 5 5 5 0 1 1 6 8 1 3 8 8 2 9 9 2 

6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

Carcass ID Number 4 0 2 1 1 0 2 1 2 1 3 4 1 4 4 4 4 0 0 3 3 4 4 3 3 

7 6 6 6 4 2 2 2 1 8 9 1 3 5 8 4 3 4 5 2 6 0 6 1 0 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 

Hemangiosarcoma 

Lymph node + + 

Iliac, sarcoma, metastatic, liver X 

Lymph node, bronchial M + M + M + + + + M M M M + M M M + M M + M + M M 

Alveolar/bronchiolar carcinoma, metastatic, lung X 

Sarcoma, metastatic, liver X 

Lymph node, mandibular + M + + A + + M M + + M M + M + + + + + + + M + M 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + + + + M + + + + 

Sarcoma, metastatic, liver X 

Lymph node, mediastinal + M + + + + + + + M + M + + + M + I + + + M + + M 

Alveolar/bronchiolar carcinoma, metastatic, lung X 

Sarcoma, metastatic, liver X 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 

Hemangiosarcoma 

Histiocytic sarcoma 

Thymus + M M M M + M + M + + + A + + + M + M + + M M + M 

Integumentary System 

Mammary gland M M M M M M M M M M M M M M M M M M M M M M M M M 

Skin + M + + + + + + + + + + + + + + + + + + + + + + + 

Hemangiosarcoma X 

Subcutaneous tissue, carcinoma 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Hemangiosarcoma 

Skeletal muscle + + 

Alveolar/bronchiolar carcinoma, metastatic, lung X 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 

Spinal cord + 

Respiratory System 

Larynx + + + + + + + + + + + + + + + + + + + + + + + + + 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 

Alveolar/bronchiolar adenoma X X X X X X X X 

Alveolar/bronchiolar adenoma, multiple X 

Alveolar/bronchiolar carcinoma X X 

Alveolar/bronchiolar carcinoma, multiple 

Hemangiosarcoma, metastatic, liver 

Hepatocellular carcinoma, metastatic, liver X X X X 

Nose + + + + + + + + + + + + A + + + + + + + + + + + + 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 161 

TABLE C2
 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 5 7 7 8 1 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

3 3 8 6 3 3 3 3 3 3 3 3 3 3 3 3 3 4 4 4 4 4 4 4 4 

6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 Total 

Carcass ID Number 2 3 3 3 2 0 0 0 0 1 2 2 2 3 3 4 4 0 1 1 1 1 2 2 5 Tissues/ 

4 3 8 4 3 1 3 7 9 0 0 5 8 5 7 2 9 8 1 5 7 9 7 9 0 Tumors 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma X 1 

Lymph node 2 

Iliac, sarcoma, metastatic, liver 1 

Lymph node, bronchial I M + M M M + + + M M + + + + + + + + M M + M + + 25 

Alveolar/bronchiolar carcinoma, metastatic, lung 1 

Sarcoma, metastatic, liver 1 

Lymph node, mandibular + M + + M + + M M M + + + M I M + + + + + M M + + 31 

Lymph node, mesenteric + + + M + + + M + + + + + + + + + + + + + + + + + 47 

Sarcoma, metastatic, liver 1 

Lymph node, mediastinal + + M I + + + M + M + + + M + M M + + + + + + + M 35 

Alveolar/bronchiolar carcinoma, metastatic, lung 1 

Sarcoma, metastatic, liver 1 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma X X 2 

Histiocytic sarcoma X 1 

Thymus M M + + + M + + + + M + M + + + M + + M + M M M + 28 

Integumentary System 

Mammary gland M M M M M M M M M M M M M M M M M M M M M M M M M 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Hemangiosarcoma X 2 

Subcutaneous tissue, carcinoma X 1 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma X 1 

Skeletal muscle 2 

Alveolar/bronchiolar carcinoma, metastatic, lung 1 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Spinal cord 1 

Respiratory System 

Larynx + + + + I + + + + + + + + + + + + + + + + + + + + 49 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Alveolar/bronchiolar adenoma X X X X X 13 

Alveolar/bronchiolar adenoma, multiple X X X X 5 

Alveolar/bronchiolar carcinoma X X X X X X X 9 

Alveolar/bronchiolar carcinoma, multiple X X 2 

Hemangiosarcoma, metastatic, liver X 1 

Hepatocellular carcinoma, metastatic, liver X X X 7 

Nose + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 50 
 

 

 

 

 

 

162 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C2
 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

Number of Days on Study 

3 

9 

9 

4 

1 

4 

5 

0 

2 

5 

2 

4 

5 

2 

7 

5 

3 

3 

5 

3 

6 

5 

4 

1 

5 

5 

2 

5 

5 

5 

5 

5 

5 

5 

5 

5 

5 

6 

0 

5 

6 

1 

5 

6 

1 

5 

6 

6 

6 

0 

8 

6 

1 

1 

6 

1 

3 

6 

1 

8 

6 

1 

8 

6 

2 

2 

6 

3 

9 

6 

4 

9 

6 

5 

2 

Carcass ID Number 

6 

4 

7 

6 

0 

6 

6 

2 

6 

6 

1 

6 

6 

1 

4 

6 

0 

2 

6 

2 

2 

6 

1 

2 

6 

2 

1 

6 

1 

8 

6 

3 

9 

6 

4 

1 

6 

1 

3 

6 

4 

5 

6 

4 

8 

6 

4 

4 

6 

4 

3 

6 

0 

4 

6 

0 

5 

6 

3 

2 

6 

3 

6 

6 

4 

0 

6 

4 

6 

6 

3 

1 

6 

3 

0 

Special Senses System 

Eye 

Harderian gland 

Adenoma 

+ 

+ + + 

Urinary System 

Kidney 

Alveolar/bronchiolar carcinoma, metastatic, lung 

Histiocytic sarcoma 

Renal tubule, adenoma 

Ureter 

Urinary bladder 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

+ + + + + + + + + + + + A + + + + + + + + + + A + 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

+ + + + + + + + + + + + + + + + + + + + + + + + + 
 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 163 

TABLE C2
 

Individual Animal Tumor Pathology of Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

Number of Days on Study 

6 

5 

3 

6 

7 

3 

6 

7 

8 

6 

8 

6 

7 

1 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

3 

7 

3 

4 

7 

3 

4 

7 

3 

4 

7 

3 

4 

7 

3 

4 

7 

3 

4 

7 

3 

4 

7 

3 

4 

Carcass ID Number 

6 

2 

4 

6 

3 

3 

6 

3 

8 

6 

3 

4 

6 

2 

3 

6 

0 

1 

6 

0 

3 

6 

0 

7 

6 

0 

9 

6 

1 

0 

6 

2 

0 

6 

2 

5 

6 

2 

8 

6 

3 

5 

6 

3 

7 

6 

4 

2 

6 

4 

9 

6 

0 

8 

6 

1 

1 

6 

1 

5 

6 

1 

7 

6 

1 

9 

6 

2 

7 

6 

2 

9 

6 

5 

0 

Total 

Tissues/ 

Tumors 

Special Senses System 

Eye

Harderian gland 

Adenoma 

+ + 

X

+

 1 

6 

1 

Urinary System 

Kidney 

Alveolar/bronchiolar carcinoma, metastatic, lung 

Histiocytic sarcoma 

Renal tubule, adenoma 

Ureter 

Urinary bladder 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

X

+

+ + + + + + + + + + + + + + + + + + + + + + + + + 

50 

1 

1 

1 

1 

48 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

50 

1 
164 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C3 

Statistical Analysis of Primary Neoplasms in Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Harderian Gland: Adenoma 

Overall ratea 

Adjusted rateb 

Terminal ratec 

4/50 (8%) 

13.2% 

1/22 (5%) 

2/50 (4%) 

6.5% 

2/31 (6%) 

2/50 (4%) 

8.3% 

2/24 (8%) 

1/50 (2%) 

4.5% 

0/20 (0%) 

First incidence (days) 

Life table testd 

Logistic regression testd 

Cochran-Armitage testd 

Fisher exact testd 

600 

Pù0.270N 

Pù0.229N 

Pù0.185N 

733 (T) 

Pù0.221N 

Pù0.307N 

Pù0.339N 

733 (T) 

Pù0.306N 

Pù0.324N 

Pù0.339N 

686 

Pù0.239N 

Pù0.192N 

Pù0.181N 

Liver: Hemangiosarcoma 

Overall rate 2/50 (4%) 4/50 (8%) 8/50 (16%) 7/50 (14%) 

Adjusted rate 9.1% 11.5% 23.5% 25.0% 

Terminal rate 2/22 (9%) 2/31 (6%) 2/24 (8%) 3/20 (15%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

733 (T) 

Pù0.036 

Pù0.078 

Pù0.096 

685 

Pù0.500 

Pù0.441 

Pù0.339 

523 

Pù0.071 

Pù0.050 

Pù0.046 

502 

Pù0.064 

Pù0.069 

Pù0.080 

Liver: Hepatocellular Adenoma 

Overall rate 22/50 (44%) 21/50 (42%) 25/50 (50%) 13/50 (26%) 

Adjusted rate 71.4% 57.5% 74.0% 42.3% 

Terminal rate 14/22 (64%) 16/31 (52%) 16/24 (67%) 5/20 (25%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

470 

Pù0.192N 

Pù0.061N 

Pù0.026N 

614 

Pù0.102N 

Pù0.290N 

Pù0.500N 

440 

Pù0.473 

Pù0.389 

Pù0.344 

533 

Pù0.123N 

Pù0.067N 

Pù0.046N 

Liver: Hepatocellular Carcinoma 

Overall rate 23/50 (46%) 33/50 (66%) 30/50 (60%) 30/50 (60%) 

Adjusted rate 60.8% 69.5% 66.5% 71.2% 

Terminal rate 9/22 (41%) 17/31 (55%) 10/24 (42%) 9/20 (45%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

482 

Pù0.074 

Pù0.471 

Pù0.303 

460 

Pù0.398 

Pù0.017 

Pù0.035 

440 

Pù0.262 

Pù0.097 

Pù0.115 

502 

Pù0.094 

Pù0.143 

Pù0.115 

Liver: Hepatocellular Adenoma or Carcinoma 

Overall rate 38/50 (76%) 41/50 (82%) 43/50 (86%) 38/50 (76%) 

Adjusted rate 89.8% 85.2% 93.2% 83.9% 

Terminal rate 18/22 (82%) 24/31 (77%) 21/24 (88%) 13/20 (65%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

470 

Pù0.134 

Pù0.375N 

Pù0.399N 

460 

Pù0.161N 

Pù0.284 

Pù0.312 

440 

Pù0.425 

Pù0.155 

Pù0.154 

502 

Pù0.338 

Pù0.591 

Pù0.592N 
Cobalt Sulfate Heptahydrate, NTP TR 471 165 

TABLE C3 

Statistical Analysis of Primary Neoplasms in Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Liver: Hepatoblastoma 

Overall rate 4/50 (8%) 0/50 (0%) 2/50 (4%) 2/50 (4%) 

Adjusted rate 11.8% 0.0% 7.6% 10.0% 

Terminal rate 1/22 (5%) 0/31 (0%) 1/24 (4%) 2/20 (10%) e First incidence (days) 533 — 705 733 (T) 

Life table test Pù0.596 Pù0.043N Pù0.313N Pù0.390N 

Logistic regression test Pù0.567N Pù0.095N Pù0.342N Pù0.345N 

Cochran-Armitage test Pù0.549N 

Fisher exact test Pù0.059N Pù0.339N Pù0.339N 

Liver: Hepatocellular Carcinoma or Hepatoblastoma 

Overall rate 27/50 (54%) 33/50 (66%) 31/50 (62%) 31/50 (62%) 

Adjusted rate 66.6% 69.5% 67.7% 73.8% 

Terminal rate 10/22 (45%) 17/31 (55%) 10/24 (42%) 10/20 (50%) 

First incidence (days) 482 460 440 502 

Life table test Pù0.106 Pù0.458N Pù0.436 Pù0.190 

Logistic regression test Pù0.500N Pù0.062 Pù0.221 Pù0.328 

Cochran-Armitage test Pù0.414 

Fisher exact test Pù0.154 Pù0.272 Pù0.272 

Liver: Hepatocellular Adenoma, Hepatocellular Carcinoma, or Hepatoblastoma 

Overall rate 40/50 (80%) 41/50 (82%) 44/50 (88%) 39/50 (78%) 

Adjusted rate 90.3% 85.2% 93.5% 86.2% 

Terminal rate 18/22 (82%) 24/31 (77%) 21/24 (88%) 14/20 (70%) 

First incidence (days) 470 460 440 502 

Life table test Pù0.134 Pù0.100N Pù0.488 Pù0.383 

Logistic regression test Pù0.340N Pù0.423 Pù0.204 Pù0.494N 

Cochran-Armitage test Pù0.376N 

Fisher exact test Pù0.500 Pù0.207 Pù0.500N 

Lung: Alveolar/bronchiolar Adenoma 

Overall rate 9/50 (18%) 12/50 (24%) 13/50 (26%) 18/50 (36%) 

Adjusted rate 30.4% 30.9% 41.1% 54.6% 

Terminal rate 4/22 (18%) 6/31 (19%) 7/24 (29%) 7/20 (35%) 

First incidence (days) 600 460 548 524 

Life table test Pù0.005 Pù0.589 Pù0.308 Pù0.024 

Logistic regression test Pù0.018 Pù0.353 Pù0.256 Pù0.027 

Cochran-Armitage test Pù0.029 

Fisher exact test Pù0.312 Pù0.235 Pù0.035 

Lung: Alveolar/bronchiolar Carcinoma 

Overall rate 4/50 (8%) 5/50 (10%) 7/50 (14%) 11/50 (22%) 

Adjusted rate 13.2% 16.1% 25.3% 43.7% 

Terminal rate 2/22 (9%) 5/31 (16%) 4/24 (17%) 7/20 (35%) 

First incidence (days) 449 733 (T) 687 552 

Life table test Pù0.004 Pù0.603N Pù0.313 Pù0.031 

Logistic regression test Pù0.006 Pù0.528 Pù0.273 Pù0.033 

Cochran-Armitage test Pù0.021 

Fisher exact test Pù0.500 Pù0.262 Pù0.045 
166 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C3 

Statistical Analysis of Primary Neoplasms in Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Lung: Alveolar/bronchiolar Adenoma or Carcinoma 

Overall rate 11/50 (22%) 14/50 (28%) 19/50 (38%) 28/50 (56%) 

Adjusted rate 35.5% 36.5% 56.5% 78.8% 

Terminal rate 5/22 (23%) 8/31 (26%) 10/24 (42%) 13/20 (65%) 

First incidence (days) 

Life table test 

Logistic regression test 

449 

P<0.001 

P<0.001 

460 

Pù0.544N 

Pù0.345 

548 

Pù0.122 

Pù0.071 

524 

P<0.001 

P<0.001 

Cochran-Armitage test 

Fisher exact test 

P<0.001 

Pù0.322 Pù0.063 P<0.001 

Spleen: Hemangiosarcoma 

Overall rate 1/47 (2%) 3/50 (6%) 1/49 (2%) 2/50 (4%) 

Adjusted rate 3.0% 8.5% 3.1% 10.0% 

Terminal rate 0/22 (0%) 1/31 (3%) 0/24 (0%) 2/20 (10%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

651 

Pù0.454 

Pù0.524 

Pù0.596 

685 

Pù0.419 

Pù0.337 

Pù0.332 

681 

Pù0.739N 

Pù0.754N 

Pù0.742N 

733 (T) 

Pù0.450 

Pù0.465 

Pù0.523 

All Organs: Hemangiosarcoma 

Overall rate 3/50 (6%) 6/50 (12%) 8/50 (16%) 9/50 (18%) 

Adjusted rate 11.8% 17.6% 23.5% 31.1% 

Terminal rate 2/22 (9%) 4/31 (13%) 2/24 (8%) 4/20 (20%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

651 

Pù0.025 

Pù0.061 

Pù0.079 

685 

Pù0.423 

Pù0.344 

Pù0.243 

523 

Pù0.139 

Pù0.103 

Pù0.100 

502 

Pù0.047 

Pù0.054 

Pù0.061 

All Organs: Hemangioma or Hemangiosarcoma 

Overall rate 5/50 (10%) 6/50 (12%) 8/50 (16%) 9/50 (18%) 

Adjusted rate 20.7% 17.6% 23.5% 31.1% 

Terminal rate 4/22 (18%) 4/31 (13%) 2/24 (8%) 4/20 (20%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

651 

Pù0.060 

Pù0.120 

Pù0.160 

685 

Pù0.528N 

Pù0.605N 

Pù0.500 

523 

Pù0.338 

Pù0.289 

Pù0.277 

502 

Pù0.149 

Pù0.162 

Pù0.194 

All Organs: Benign Neoplasms 

Overall rate 30/50 (60%) 30/50 (60%) 30/50 (60%) 27/50 (54%) 

Adjusted rate 84.8% 74.3% 77.7% 73.0% 

Terminal rate 17/22 (77%) 21/31 (68%) 16/24 (67%) 11/20 (55%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

470 

Pù0.276 

Pù0.437N 

Pù0.279N 

460 

Pù0.086N 

Pù0.353N 

Pù0.581N 

440 

Pù0.430N 

Pù0.542N 

Pù0.581N 

524 

Pù0.538 

Pù0.444N 

Pù0.343N 
167 

c 

Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C3 

Statistical Analysis of Primary Neoplasms in Male Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

All Organs: Malignant Neoplasms 

Overall rate 33/50 (66%) 39/50 (78%) 41/50 (82%) 38/50 (76%) 

Adjusted rate 73.8% 79.4% 85.1% 87.8% 

Terminal rate 11/22 (50%) 21/31 (68%) 17/24 (71%) 15/20 (75%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

449 

Pù0.053 

Pù0.412 

Pù0.318 

460 

Pù0.374N 

Pù0.042 

Pù0.133 

440 

Pù0.270 

Pù0.037 

Pù0.055 

502 

Pù0.141 

Pù0.205 

Pù0.189 

All Organs: Benign or Malignant Neoplasms 

Overall rate 49/50 (98%) 48/50 (96%) 47/50 (94%) 46/50 (92%) 

Adjusted rate 100.0% 96.0% 95.9% 95.8% 

Terminal rate 22/22 (100%) 29/31 (94%) 22/24 (92%) 18/20 (90%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

449 

Pù0.114 

Pù0.238N 

Pù0.143N 

460 

Pù0.028N 

Pù0.527N 

Pù0.500N 

440 

Pù0.283N 

Pù0.297N 

Pù0.309N 

502 

Pù0.435 

Pù0.279N 

Pù0.181N 

(T)Terminal sacrifice 

a Number of neoplasm-bearing animals/number of animals examined. Denominator is number of animals examined microscopically for liver, lung, and 

spleen; for other tissues, denominator is number of animals necropsied. b	 Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality 

Observed incidence at terminal kill 

d	 Beneath the chamber control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values corresponding 

to pairwise comparisons between the chamber controls and that exposed group. The life table test regards neoplasms in animals dying prior to terminal kill 

as being (directly or indirectly) the cause of death. The logistic regression test regards these lesions as nonfatal. The Cochran-Armitage and Fisher exact 

tests compare directly the overall incidence rates. For all tests, a negative trend or a lower incidence in an exposure group is indicated by N. e Not applicable; no neoplasms in animal group 
168 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C4a 

Historical Incidence of Alveolar/bronchiolar Neoplasms in Chamber Control Male B6C3F a 1 Mice

Incidence in Controls 

Study Adenoma Carcinoma Adenoma or Carcinoma 

Historical Incidence at Battelle Pacific Northwest Laboratories 

1,3-Butadiene 18/50 5/50 21/50 

Acetonitrile 6/50 4/50 10/50 

Allyl Glycidyl Ether 7/50 0/50 7/50 

2-Chloroacetophenone 7/50 6/50 11/50 

l-Epinephrine Hydrochloride 11/50 5/50 15/50 

Chloroethane 3/50 2/50 5/50 

Hexachlorocyclopentadiene 11/49 0/49 11/49 

o-Chlorobenzalmalononitrile (CS2) 7/49 7/49 14/49 

Ozone 6/50 8/50 14/50 

Overall Historical Incidence 

Total 141/947 (14.9%) 75/947 (7.9%) 205/947 (21.7%) 

Standard deviation 7.0% 5.7% 8.0% 

Range 6%-36% 0%-16% 10%-42% 

a Data as of 12 May 1995 

TABLE C4b 

Historical Incidence of Hemangiosarcoma of the Liver in Chamber Control Male B6C3F a 1 Mice

Study Incidence in Controls 

Historical Incidence at Battelle Pacific Northwest Laboratories 

1,3-Butadiene 0/50 

Acetonitrile 1/50 

Allyl Glycidyl Ether 0/49 

2-Chloroacetophenone 0/50 

l-Epinephrine Hydrochloride 1/50 

Chloroethane 0/50 

Hexachlorocyclopentadiene 0/50 

o-Chlorobenzalmalononitrile (CS2) 0/49 

Ozone 0/50 

Overall Historical Incidence 

Total 12/947 (1.3%) 

Standard deviation 1.7% 

Range 0%-6% 

a Data as of 12 May 1995 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 169 

TABLE C5 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydratea 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Accidental deaths 1 1 

Moribund 19 16 17 23 

Natural deaths 8 3 9 6 

Survivors 

Terminal sacrifice 22 31 24 20 

Animals examined microscopically 50 50 50 50 

Alimentary System 

Gallbladder (37) (41) (38) (38) 

Degeneration, hyaline 1 (3%) 1 (2%) 2 (5%) 

Infiltration cellular, lymphocyte 1 (3%) 

Intestine small, duodenum (45) (49) (42) (45) 

Hyperplasia, lymphoid 1 (2%) 

Inflammation, chronic 1 (2%) 

Necrosis, focal 1 (2%) 

Liver (50) (50) (50) (50) 

Basophilic focus 1 (2%) 

Clear cell focus 3 (6%) 2 (4%) 

Clear cell focus, multiple 1 (2%) 

Cyst 1 (2%) 

Eosinophilic focus 8 (16%) 7 (14%) 9 (18%) 2 (4%) 

Eosinophilic focus, multiple 1 (2%) 1 (2%) 3 (6%) 

Infiltration cellular, lymphocyte 1 (2%) 

Inflammation, chronic 33 (66%) 36 (72%) 40 (80%) 39 (78%) 

Karyomegaly 39 (78%) 35 (70%) 39 (78%) 43 (86%) 

Mineralization 1 (2%) 

Mitotic alteration 1 (2%) 

Necrosis, focal 3 (6%) 3 (6%) 6 (12%) 1 (2%) 

Regeneration 32 (64%) 30 (60%) 35 (70%) 38 (76%) 

Vacuolization cytoplasmic, diffuse 3 (6%) 1 (2%) 

Vacuolization cytoplasmic, focal 1 (2%) 

Bile duct, hyperplasia 3 (6%) 6 (12%) 4 (8%) 

Oval cell, hyperplasia 38 (76%) 36 (72%) 40 (80%) 44 (88%) 

Mesentery (3) (4) (5) (4) 

Inflammation, chronic 1 (33%) 1 (25%) 

Mineralization 1 (20%) 

Artery, fibrosis 1 (33%) 1 (25%) 

Artery, inflammation, chronic 1 (25%) 

Fat, necrosis 1 (33%) 2 (50%) 3 (60%) 

Pancreas (48) (50) (49) (50) 

Acinus, atrophy 1 (2%) 

Stomach, forestomach (49) (50) (50) (50) 

Cyst 1 (2%) 

Hyperplasia, squamous 4 (8%) 2 (4%) 2 (4%) 

Inflammation 1 (2%) 

a Number of animals examined microscopically at the site and the number of animals with lesion 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

170 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C5 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Alimentary System (continued) 

Stomach, glandular (48) (50) (50) (50) 

Mineralization 1 (2%) 

Tooth (1) 

Inflammation 1 (100%) 

Cardiovascular System 

Heart (50) (50) (50) (50) 

Cardiomyopathy 1 (2%) 4 (8%) 2 (4%) 1 (2%) 

Inflammation, chronic 1 (2%) 

Artery, inflammation, chronic 1 (2%) 

Atrium, thrombosis 1 (2%) 2 (4%) 3 (6%) 2 (4%) 

Endocrine System 

Adrenal cortex (49) (50) (49) (50) 

Accessory adrenal cortical nodule 1 (2%) 

Hyperplasia 3 (6%) 4 (8%) 5 (10%) 2 (4%) 

Inflammation 1 (2%) 

Necrosis 1 (2%) 

Necrosis, diffuse 1 (2%) 

Adrenal medulla (48) (50) (49) (50) 

Hyperplasia 1 (2%) 5 (10%) 

Necrosis, diffuse 1 (2%) 

Thyroid gland (49) (50) (50) (50) 

Inflammation 1 (2%) 

Follicular cell, hyperplasia 3 (6%) 17 (34%) 11 (22%) 10 (20%) 

General Body System 

Tissue NOS (1) (1) 

Inflammation, chronic 1 (100%) 

Genital System 

Epididymis (49) (50) (50) (50) 

Degeneration 1 (2%) 

Granuloma sperm 1 (2%) 

Inflammation, chronic 1 (2%) 2 (4%) 

Mineralization 2 (4%) 1 (2%) 

Penis (1) 

Inflammation, chronic 1 (100%) 

Preputial gland (49) (50) (50) (49) 

Cyst 1 (2%) 2 (4%) 3 (6%) 1 (2%) 

Inflammation, chronic 13 (27%) 12 (24%) 9 (18%) 5 (10%) 

Inflammation, suppurative 2 (4%) 2 (4%) 

Prostate (46) (50) (44) (49) 

Inflammation 2 (4%) 2 (5%) 1 (2%) 

Seminal vesicle (48) (50) (49) (49) 

Inflammation, chronic 1 (2%) 

Testes (49) (50) (50) (50) 

Atrophy 2 (4%) 2 (4%) 

Degeneration, focal 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 171 

TABLE C5 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Hematopoietic System 

Lymph node (1) (1) (2) (2) 

Iliac, hyperplasia, lymphoid 1 (100%) 1 (50%) 

Iliac, infiltration cellular, histiocyte 1 (50%) 

Renal, hyperplasia, lymphoid 1 (100%) 

Lymph node, bronchial (21) (24) (28) (25) 

Infiltration cellular, plasma cell 1 (4%) 

Infiltration cellular, histiocyte 1 (5%) 1 (4%) 

Lymph node, mesenteric (49) (48) (43) (47) 

Hematopoietic cell proliferation 1 (2%) 

Hemorrhage 2 (4%) 2 (4%) 4 (9%) 3 (6%) 

Hyperplasia, lymphoid 1 (2%) 2 (4%) 

Inflammation, chronic 1 (2%) 2 (5%) 2 (4%) 

Lymph node, mediastinal (43) (28) (31) (35) 

Hyperplasia, lymphoid 1 (4%) 4 (13%) 

Infiltration cellular, plasma cell 1 (2%) 1 (4%) 

Infiltration cellular, histiocyte 2 (5%) 1 (3%) 

Inflammation, chronic 1 (3%) 1 (3%) 

Spleen (47) (50) (49) (50) 

Angiectasis 1 (2%) 1 (2%) 

Hematopoietic cell proliferation 10 (21%) 6 (12%) 6 (12%) 14 (28%) 

Hyperplasia, lymphoid 4 (8%) 1 (2%) 

Integumentary System 

Skin (49) (50) (49) (49) 

Inflammation, chronic 1 (2%) 2 (4%) 

Inflammation, suppurative 1 (2%) 

Ulcer 1 (2%) 2 (4%) 

Epidermis, hyperplasia 1 (2%) 1 (2%) 

Prepuce, inflammation, chronic 1 (2%) 

Prepuce, ulcer 4 (8%) 1 (2%) 1 (2%) 

Musculoskeletal System 

Bone (50) (50) (50) (50) 

Fracture 1 (2%) 

Skeletal muscle (1) (2) 

Inflammation, chronic, focal 1 (50%) 

Nervous System 

None 

Respiratory System 

Larynx (48) (49) (48) (49) 

Foreign body 1 (2%) 

Metaplasia, squamous 37 (76%) 48 (100%) 44 (90%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

172 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE C5 

Summary of the Incidence of Nonneoplastic Lesions in Male Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Respiratory System (continued) 

Lung (50) (50) (50) (50) 

Hemorrhage 1 (2%) 1 (2%) 1 (2%) 

Infiltration cellular, diffuse, histiocyte 1 (2%) 2 (4%) 4 (8%) 10 (20%) 

Infiltration cellular, focal, histiocyte 10 (20%) 5 (10%) 8 (16%) 17 (34%) 

Inflammation, chronic 1 (2%) 1 (2%) 3 (6%) 

Alveolar epithelium, hyperplasia 4 (8%) 4 (8%) 4 (8%) 

Alveolar epithelium, goblet cell, metaplasia,

 focal 1 (2%) 

Artery, thrombosis 1 (2%) 

Bronchus, hyperplasia 1 (2%) 

Bronchus, vacuolization cytoplasmic 18 (36%) 34 (68%) 38 (76%) 

Nose (50) (50) (48) (49) 

Hemorrhage 1 (2%) 

Inflammation, suppurative 1 (2%) 6 (12%) 

Olfactory epithelium, atrophy 29 (60%) 48 (98%) 

Olfactory epithelium, degeneration, hyaline 2 (4%) 2 (4%) 

Olfactory epithelium, hyperplasia 10 (20%) 

Olfactory epithelium, metaplasia 2 (4%) 

Respiratory epithelium, degeneration, hyaline 1 (2%) 

Trachea (49) (50) (50) (50) 

Inflammation, chronic 1 (2%) 

Special Senses System 

Eye (1) (1) (1) 

Degeneration 1 (100%) 1 (100%) 

Hemorrhage 1 (100%) 

Inflammation 1 (100%) 

Cornea, inflammation 1 (100%) 

Harderian gland (4) (4) (4) (6) 

Inflammation, acute 1 (25%) 

Urinary System 

Kidney (49) (50) (50) (50) 

Cyst 1 (2%) 1 (2%) 

Infiltration cellular, mixed cell 5 (10%) 1 (2%) 5 (10%) 3 (6%) 

Inflammation, chronic 1 (2%) 1 (2%) 

Metaplasia, osseous 1 (2%) 

Mineralization 1 (2%) 

Nephropathy 11 (22%) 13 (26%) 7 (14%) 9 (18%) 

Glomerulus, amyloid deposition 1 (2%) 2 (4%) 

Medulla, inflammation, chronic 1 (2%) 

Renal tubule, hyperplasia 1 (2%) 

Urinary bladder (46) (49) (45) (48) 

Inflammation 3 (7%) 3 (7%) 3 (6%) 
173 

TABLE D1 

TABLE D2 

TABLE D3 

TABLE D4a 

TABLE D4b 

TABLE D5 

APPENDIX D
 

SUMMARY OF LESIONS IN FEMALE MICE
 

IN THE 2-YEAR INHALATION STUDY
 

OF COBALT SULFATE HEPTAHYDRATE
 

Summary of the Incidence of Neoplasms in Female Mice 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 174
 

Individual Animal Tumor Pathology of Female Mice 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 178
 

Statistical Analysis of Primary Neoplasms in Female Mice 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 198
 

Historical Incidence of Alveolar/bronchiolar Neoplasms 

in Chamber Control Female B6C3F1
  Mice ................................ 203

Historical Incidence of Hemangiosarcoma of the Liver 

in Chamber Control Female B6C3F1
  Mice ................................ 203

Summary of the Incidence of Nonneoplastic Lesions in Female Mice 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate .................. 204
 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

174 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D1 

Summary of the Incidence of Neoplasms in Female Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydratea 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Moribund 11 10 13 16 

Natural deaths 5 3 5 6 

Survivors 

Terminal sacrifice 34 37 32 28 

Animals examined microscopically 50 50 50 50 

Alimentary System 

Gallbladder (43) (43) (38) (43) 

Intestine large, cecum (49) (49) (48) (47) 

Leiomyoma 1 (2%) 

Intestine small, duodenum (47) (48) (47) (45) 

Intestine small, jejunum (48) (49) (47) (44) 

Hemangiosarcoma 1 (2%) 

Intestine small, ileum (48) (48) (47) (45) 

Liver (50) (50) (50) (49) 

Hemangiosarcoma 1 (2%) 3 (6%) 

Hepatocellular carcinoma 12 (24%) 9 (18%) 16 (32%) 4 (8%) 

Hepatocellular adenoma 7 (14%) 7 (14%) 11 (22%) 9 (18%) 

Hepatocellular adenoma, multiple 1 (2%) 3 (6%) 2 (4%) 3 (6%) 

Hepatocholangiocarcinoma 2 (4%) 

Histiocytic sarcoma 3 (6%) 2 (4%) 2 (4%) 

Mesentery (10) (12) (8) (7) 

Hemangioma 1 (14%) 

Hemangiosarcoma 1 (8%) 1 (13%) 

Hepatocholangiocarcinoma, metastatic, liver 1 (10%) 

Histiocytic sarcoma 1 (8%) 

Sarcoma 1 (10%) 

Pancreas (50) (50) (49) (49) 

Histiocytic sarcoma 1 (2%) 

Salivary glands (50) (50) (50) (50) 

Stomach, forestomach (50) (50) (49) (50) 

Squamous cell papilloma 1 (2%) 1 (2%) 1 (2%) 

Stomach, glandular (50) (50) (49) (50) 

Muscularis, serosa, sarcoma 1 (2%) 

Cardiovascular System 

Heart (50) (50) (50) (50) 

Hepatocholangiocarcinoma, metastatic, liver 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 175 

TABLE D1 

Summary of the Incidence of Neoplasms in Female Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Endocrine System 

Adrenal cortex (50) (50) (49) (50) 

Histiocytic sarcoma 1 (2%) 

Capsule, adenoma 1 (2%) 

Adrenal medulla (49) (50) (49) (49) 

Pheochromocytoma benign 1 (2%) 

Islets, pancreatic (50) (50) (49) (49) 

Adenoma 1 (2%) 1 (2%) 

Pituitary gland (48) (47) (47) (48) 

Pars distalis, adenoma 11 (23%) 8 (17%) 7 (15%) 8 (17%) 

Thyroid gland (50) (49) (49) (49) 

Follicular cell, adenoma 3 (6%) 5 (10%) 

Follicular cell, carcinoma 1 (2%) 2 (4%) 

General Body System 

None 

Genital System 

Ovary (48) (49) (49) (48) 

Arrhenoblastoma benign 1 (2%) 

Cystadenocarcinoma 1 (2%) 

Cystadenoma 2 (4%) 3 (6%) 3 (6%) 3 (6%) 

Granulosa cell tumor benign 1 (2%) 1 (2%) 

Histiocytic sarcoma 1 (2%) 1 (2%) 1 (2%) 

Luteoma 1 (2%) 1 (2%) 

Teratoma benign 1 (2%) 1 (2%) 1 (2%) 

Yolk sac carcinoma 1 (2%) 

Uterus (50) (50) (49) (49) 

Hemangioma 1 (2%) 2 (4%) 

Hemangiosarcoma 1 (2%) 

Histiocytic sarcoma 1 (2%) 1 (2%) 

Leiomyoma 1 (2%) 

Leiomyosarcoma 1 (2%) 

Polyp stromal 2 (4%) 

Hematopoietic System 

Bone marrow (50) (50) (49) (50) 

Histiocytic sarcoma 1 (2%) 2 (4%) 

Lymph node (3) (6) (3) (4) 

Histiocytic sarcoma 1 (33%) 

Lumbar, histiocytic sarcoma 1 (33%) 

Renal, histiocytic sarcoma 2 (67%) 

Lymph node, bronchial (30) (34) (27) (35) 

Hepatocholangiocarcinoma, metastatic, liver 2 (7%) 

Histiocytic sarcoma 1 (3%) 1 (3%) 1 (3%) 

Lymph node, mandibular (37) (37) (36) (36) 

Histiocytic sarcoma 2 (5%) 2 (5%) 1 (3%) 

Lymph node, mesenteric (46) (45) (46) (44) 

Histiocytic sarcoma 3 (7%) 2 (4%) 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

176 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D1 

Summary of the Incidence of Neoplasms in Female Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Hematopoietic System (continued) 

Lymph node, mediastinal 

Alveolar/bronchiolar carcinoma, metastatic, 

lung

Hepatocholangiocarcinoma, metastatic, liver

Histiocytic sarcoma

Spleen 

Histiocytic sarcoma

Thymus 

Histiocytic sarcoma

(41) 

 2 (5%) 

 3 (7%) 

(50) 

 3 (6%) 

(41) 

 1 (2%) 

(36) 

 2 (6%) 

(50) 

 2 (4%) 

(44) 

 1 (2%) 

(28) 

(49) 

(41) 

(34) 

 1 (3%) 

 1 (3%) 

(49) 

 1 (2%) 

(41) 

Integumentary System 

Mammary gland 

Adenoma

Carcinoma

Skin 

Hemangiosarcoma

Histiocytic sarcoma

Squamous cell carcinoma

Subcutaneous tissue, mast cell tumor benign

Subcutaneous tissue, sarcoma

(47) 

 1 (2%) 

(49) 

 1 (2%) 

 6 (12%) 

(50) 

 1 (2%) 

(50) 

 2 (4%) 

(50) 

 1 (2%) 

(50) 

 1 (2%) 

 1 (2%) 

(50) 

 1 (2%) 

 1 (2%) 

(50) 

 1 (2%) 

Musculoskeletal System 

Skeletal muscle

Sarcoma

 (1) (2)

 1 (50%) 

(1) 

 1 (100%) 

Nervous System 

Brain (50) (50) (50) (50) 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma

Alveolar/bronchiolar adenoma, multiple

Alveolar/bronchiolar carcinoma

Hepatocellular carcinoma, metastatic, liver

Hepatocholangiocarcinoma, metastatic, liver

Histiocytic sarcoma

Sarcoma, metastatic, skin

Nose 

(50) 

(50) 

 3 (6%) 

 1 (2%) 

 3 (6%) 

 2 (4%) 

 3 (6%) 

 1 (2%) 

(50) 

(49) 

(50) 

 5 (10%) 

 1 (2%) 

 1 (2%) 

 2 (4%) 

(50) 

(47) 

(50) 

 8 (16%) 

 1 (2%) 

 4 (8%) 

 5 (10%) 

(49) 

(50) 

(50) 

 8 (16%) 

 2 (4%) 

 9 (18%) 

 2 (4%) 

 1 (2%) 

(48) 

Special Senses System 

Harderian gland

Adenoma

 (2)

 2 (100%)

 (2)

 2 (100%)

 (1) 

 1 (100%) 
 

 

 

177 

c 

Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D1 

Summary of the Incidence of Neoplasms in Female Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Urinary System 

Kidney 

Histiocytic sarcoma

Urinary bladder 

Histiocytic sarcoma

(50) 

 2 (4%) 

(48) 

 1 (2%) 

(50) 

 1 (2%) 

(47) 

(50) 

(45) 

(50) 

 1 (2%) 

(46) 

Systemic Lesions 

Multiple organsb 

Histiocytic sarcoma

Lymphoma malignant

(50) 

 3 (6%) 

 4 (8%) 

(50) 

 2 (4%) 

 7 (14%) 

(50) 

 7 (14%) 

(50) 

 3 (6%) 

 5 (10%) 

Neoplasm Summary 

Total animals with primary neoplasmsc 

Total primary neoplasms 

Total animals with benign neoplasms 

Total benign neoplasms 

Total animals with malignant neoplasms 

Total malignant neoplasms 

Total animals with metastatic neoplasms 

Total metastatic neoplasms 

42 

70 

25 

36 

27 

34 

6 

12 

41 

61 

32 

36 

22 

25 

43 

76 

27 

38 

31 

38 

5 

5 

39 

68 

27 

44 

21 

24 

3 

3 

a Number of animals examined microscopically at the site and the number of animals with neoplasm b Number of animals with any tissue examined microscopically 

Primary neoplasms: all neoplasms except metastatic neoplasms 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

178 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

Number of Days on Study 

2 5 5 5 5 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 

2 3 5 8 9 0 1 2 3 4 5 5 7 9 9 1 3 3 3 3 3 3 3 3 3 

5 1 9 3 6 9 9 0 9 3 2 6 7 9 9 3 4 4 4 4 5 5 5 5 5 

Carcass ID Number 

1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 

3 2 3 1 0 4 0 4 0 2 2 0 0 2 5 4 1 1 1 3 0 0 1 1 1 

6 7 7 5 6 8 2 6 7 8 1 4 9 5 0 5 0 1 9 9 1 8 2 3 4 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hemangiosarcoma 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Hepatocholangiocarcinoma 

Histiocytic sarcoma 

Mesentery 

Hepatocholangiocarcinoma, metastatic, liver 

Sarcoma 

Pancreas 

Histiocytic sarcoma 

Salivary glands 

Stomach, forestomach 

Squamous cell papilloma 

Stomach, glandular 

Muscularis, serosa, sarcoma 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

M + + A + A A A + + + + + + A + + + + + + + + + + 

+ + + + + + A + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + A + + + + + + + + + + + + + + + + + + 

+ + + A + A A + + + + + + + + + + + + + + + + + + 

+ + + A + + A + + + + + + + + + + + + + + + + + + 

+ + + + + + A A + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X X X 

X 

X X 

X X X 

+ + + + + 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 

Hepatocholangiocarcinoma, metastatic, liver X 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 

Adrenal medulla + + + + + + + M + + + + + + + + + + + + + + + + + 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma 

Parathyroid gland + + M + M M + M + M + M M + M + M + M M + M + + + 

Pituitary gland + + + I + + + + + + + + + + + + + + + + + + + + I 

Pars distalis, adenoma X X X X 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Follicular cell, adenoma X 

Follicular cell, carcinoma X 

General Body System 

None 

+: Tissue examined microscopically M: Missing tissue X: Lesion present 

A: Autolysis precludes examination I: Insufficient tissue Blank: Not examined 
 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 179 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

5 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 Total 

Carcass ID Number 1 1 2 2 2 2 3 3 4 4 0 0 1 2 2 3 3 3 3 3 4 4 4 4 4 Tissues/ 

6 7 2 4 6 9 3 5 1 4 3 5 8 0 3 0 1 2 4 8 0 2 3 7 9 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Gallbladder + + + + + + + + + + + + + + + + + M + + + + + + + 43 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma X 1 

Hepatocellular carcinoma X X X X X 12 

Hepatocellular adenoma X X X X X X 7 

Hepatocellular adenoma, multiple X 1 

Hepatocholangiocarcinoma 2 

Histiocytic sarcoma 3 

Mesentery + + + + + 10 

Hepatocholangiocarcinoma, metastatic, liver 1 

Sarcoma X 1 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Squamous cell papilloma X 1 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Muscularis, serosa, sarcoma X 1 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hepatocholangiocarcinoma, metastatic, liver 1 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adenoma X 1 

Parathyroid gland + + + + M M M M M + M M + M M + + + + M + M M + M 25 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Pars distalis, adenoma X X X X X X X 11 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Follicular cell, adenoma X X 3 

Follicular cell, carcinoma 1 

General Body System 

None 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

180 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

2 5 5 5 5 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 2 3 5 8 9 0 1 2 3 4 5 5 7 9 9 1 3 3 3 3 3 3 3 3 3 

5 1 9 3 6 9 9 0 9 3 2 6 7 9 9 3 4 4 4 4 5 5 5 5 5 

1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 

Carcass ID Number 3 2 3 1 0 4 0 4 0 2 2 0 0 2 5 4 1 1 1 3 0 0 1 1 1 

6 7 7 5 6 8 2 6 7 8 1 4 9 5 0 5 0 1 9 9 1 8 2 3 4 

Genital System 

Clitoral gland M + + + + + + + + + + + M M + + + + M + + + M + + 

Ovary + + + + + + + + + + + + + + + + I + + + + + + + + 

Arrhenoblastoma benign 

Cystadenoma 

Granulosa cell tumor benign X 

Histiocytic sarcoma X 

Luteoma X 

Teratoma benign 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X 

Leiomyoma 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X 

Lymph node + + 

Histiocytic sarcoma X 

Lumbar, histiocytic sarcoma X 

Renal, histiocytic sarcoma X X 

Lymph node, bronchial M M M M M M M + M M M M + M + + + M + M + + M + + 

Hepatocholangiocarcinoma, metastatic, liver X X 

Histiocytic sarcoma X 

Lymph node, mandibular M + M + + + + M + M + + + + + + M + + + + + + + + 

Histiocytic sarcoma X X 

Lymph node, mesenteric + + + + + + + + + + + + + + + + + I + + + + + I + 

Histiocytic sarcoma X X X 

Lymph node, mediastinal + + + + + + + + + + + + + + + + + + M + + + + + + 

Hepatocholangiocarcinoma, metastatic, liver X X 

Histiocytic sarcoma X X X 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X X X 

Thymus + + + + M + + + + + + + + + M M + M + M M + + + + 

Histiocytic sarcoma X 

Integumentary System 

Mammary gland M + + + + + I + + + + + + M + + + + + + + + + + + 

Carcinoma X 

Skin + + + + + + + + + + + + + I + + + + + + + + + + + 

Squamous cell carcinoma X 

Subcutaneous tissue, sarcoma X X X X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 

Spinal cord + 
 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 181 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

5 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 Total 

Carcass ID Number 1 1 2 2 2 2 3 3 4 4 0 0 1 2 2 3 3 3 3 3 4 4 4 4 4 Tissues/ 

6 7 2 4 6 9 3 5 1 4 3 5 8 0 3 0 1 2 4 8 0 2 3 7 9 Tumors 

Genital System 

Clitoral gland + + M + + + + + + + + + + + M + + M + + + M + + + 41 

Ovary + + + + + + + + + + + + + + + + + I + + + + + + + 48 

Arrhenoblastoma benign X 1 

Cystadenoma X X 2 

Granulosa cell tumor benign 1 

Histiocytic sarcoma 1 

Luteoma 1 

Teratoma benign X 1 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1 

Leiomyoma X 1 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1 

Lymph node + 3 

Histiocytic sarcoma 1 

Lumbar, histiocytic sarcoma 1 

Renal, histiocytic sarcoma 2 

Lymph node, bronchial + + + + + + + + + M + + + + + + + + + M M + + M M 30 

Hepatocholangiocarcinoma, metastatic, liver 2 

Histiocytic sarcoma 1 

Lymph node, mandibular + + M M + + + M + + + M M + M + + M + + + + + M + 37 

Histiocytic sarcoma 2 

Lymph node, mesenteric + + + + + + + + + + + + + + I + + + + + I + + + + 46 

Histiocytic sarcoma 3 

Lymph node, mediastinal + + + + M + + + + + + M M + M M M M + + + M + + + 41 

Hepatocholangiocarcinoma, metastatic, liver 2 

Histiocytic sarcoma 3 

Spleen + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 3 

Thymus + + + + + M + + + + + + + + + + + + + M + + + M + 41 

Histiocytic sarcoma 1 

Integumentary System 

Mammary gland + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Carcinoma 1 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Squamous cell carcinoma 1 

Subcutaneous tissue, sarcoma X X 6 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Spinal cord 1 
 

 

 

 

 

 

 

 

 

 

182 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

2 5 5 5 5 6 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 2 

5 

3 

1 

5 

9 

8 

3 

9 

6 

0 

9 

1 

9 

2 

0 

3 

9 

4 

3 

5 

2 

5 

6 

7 

7 

9 

9 

9 

9 

1 

3 

3 

4 

3 

4 

3 

4 

3 

4 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 

Carcass ID Number 3 2 3 1 0 4 0 4 0 2 2 0 0 2 5 4 1 1 1 3 0 0 1 1 1 

6 7 7 5 6 8 2 6 7 8 1 4 9 5 0 5 0 1 9 9 1 8 2 3 4 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Hepatocholangiocarcinoma, metastatic, liver 

Histiocytic sarcoma 

Sarcoma, metastatic, skin 

Nose 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X 

X X 

X X 

X X X 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 

Special Senses System 

Eye 

Harderian gland 

Adenoma 

+ 

X 

Urinary System 

Kidney 

Histiocytic sarcoma 

Urinary bladder 

Histiocytic sarcoma 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X 

+ + + + + + A A + + + + + + + + + + + + + + + + + 

X 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma 

Lymphoma malignant 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X 

X 
 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 183 

TABLE D2 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

Chamber Control 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 Total 

Carcass ID Number 1 

6 

1 

7 

2 

2 

2 

4 

2 

6 

2 

9 

3 

3 

3 

5 

4 

1 

4 

4 

0 

3 

0 

5 

1 

8 

2 

0 

2 

3 

3 

0 

3 

1 

3 

2 

3 

4 

3 

8 

4 

0 

4 

2 

4 

3 

4 

7 

4 

9 

Tissues/ 

Tumors 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Hepatocholangiocarcinoma, metastatic, liver

Histiocytic sarcoma

Sarcoma, metastatic, skin

Nose 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

X

X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

50 

50 

3 

1 

3 

2 

3 

1 

50 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Special Senses System 

Eye 

Harderian gland 

Adenoma 

+

+

X

 1 

2 

2 

Urinary System 

Kidney 

Histiocytic sarcoma

Urinary bladder 

Histiocytic sarcoma

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

50 

2 

48 

1 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma

Lymphoma malignant 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X

50 

3 

4 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

184 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D2
 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

Number of Days on Study 

4 5 5 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

8 9 9 4 5 6 6 6 6 9 2 2 2 3 3 3 3 3 3 3 3 3 3 3 3 

1 3 6 9 2 4 6 6 6 3 0 6 7 4 4 4 4 4 4 4 5 5 5 5 5 

Carcass ID Number 

3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

1 3 2 0 0 2 0 3 4 4 3 3 4 2 2 2 2 3 4 4 0 0 0 1 1 

4 4 7 9 8 6 3 3 7 3 2 1 1 0 1 3 9 8 0 4 1 4 6 0 3 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Histiocytic sarcoma 

Mesentery 

Hemangiosarcoma 

Histiocytic sarcoma 

Pancreas 

Salivary glands 

Stomach, forestomach 

Squamous cell papilloma 

Stomach, glandular 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + A + + + + M + A + M A + + + + + + + + + M M + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + A + + + + + + + + + + + + + + + + + + + + + + 

+ + A + + + + + + + M + + + + + + + + + + + + + + 

+ + A + + + + + + + + + + + + + + + + + + + + + + 

+ + A + + + + + + A + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X X 

X X X 

X 

X X 

+ + + + + + + 

X 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X 

Capsule, adenoma 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 

Pheochromocytoma benign 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 

Parathyroid gland + M + M + + M M M M + + + M M M M + M M I + + M + 

Pituitary gland + + + + + + + + + I M + + + + + + + + + + I + + + 

Pars distalis, adenoma X X 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 

General Body System 

None 

Genital System 

Clitoral gland + + + + M + + + + + + + + + + + + M M + + + M M + 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 

Cystadenocarcinoma X 

Cystadenoma X X 

Histiocytic sarcoma X
 

Teratoma benign
 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 

Hemangioma 

Hemangiosarcoma X 

Polyp stromal X 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 185 

TABLE D2
 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

5 5 5 5 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 

3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 Total 

Carcass ID Number 1 1 2 2 3 3 3 3 4 4 4 4 5 0 0 0 1 1 1 1 1 2 2 3 4 Tissues/ 

6 7 2 8 0 5 7 9 2 5 8 9 0 2 5 7 1 2 5 8 9 4 5 6 6 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Gallbladder + + + + + + + + + + + + + + + + + + + + + + + + + 43 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hepatocellular carcinoma X X X 9 

Hepatocellular adenoma X X X X 7 

Hepatocellular adenoma, multiple X X 3 

Histiocytic sarcoma 2 

Mesentery + + + + + 12 

Hemangiosarcoma 1 

Histiocytic sarcoma 1 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Squamous cell papilloma 1 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1 

Capsule, adenoma X 1 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Pheochromocytoma benign X 1 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Parathyroid gland + M M M M M + + M + M M + M M M + M + + M + + + + 23 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Pars distalis, adenoma X X X X X X 8 

Thyroid gland + + + + + + + + + + + + + + + + + I + + + + + + + 49 

General Body System 

None 

Genital System 

Clitoral gland + + + + M + + + + M + + M + + + + + + M M + + + + 40 

Ovary + + + + + + + + + + + + + + + + + + + + + + I + + 49 

Cystadenocarcinoma 1 

Cystadenoma X 3 

Histiocytic sarcoma 1 

Teratoma benign X 1 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangioma X 1 

Hemangiosarcoma 1 

Polyp stromal X 2 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

186 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D2
 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

4 5 5 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 8 

1 

9 

3 

9 

6 

4 

9 

5 

2 

6 

4 

6 

6 

6 

6 

6 

6 

9 

3 

2 

0 

2 

6 

2 

7 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

4 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

Carcass ID Number 1 3 2 0 0 2 0 3 4 4 3 3 4 2 2 2 2 3 4 4 0 0 0 1 1 

4 4 7 9 8 6 3 3 7 3 2 1 1 0 1 3 9 8 0 4 1 4 6 0 3 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 

Lymph node 

Lymph node, bronchial 

Histiocytic sarcoma 

Lymph node, mandibular 

Histiocytic sarcoma 

Lymph node, mesenteric 

Histiocytic sarcoma 

Lymph node, mediastinal 

Histiocytic sarcoma 

Spleen 

Histiocytic sarcoma 

Thymus 

Histiocytic sarcoma 

M 

M 

+ 

+ 

+ 

M 

+ 

+ 

+ 

M 

+ 

+ 

M 

M 

+ 

+ 

+ 

+ 

M 

+ 

X 

+ 

X 

+ 

X 

+ 

X 

+ 

X 

M 

M 

M 

M 

+ 

M 

M 

+ 

+ 

+ 

+ 

+ 

M 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

M 

M 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

M 

M 

+ 

M 

M 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

X 

+ 

X 

+ 

X 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

M 

+ 

M 

+ 

+ 

Integumentary System 

Mammary gland 

Carcinoma 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 

Subcutaneous tissue, sarcoma X X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Skeletal muscle + 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma 

Histiocytic sarcoma 

Nose 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

I 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 

Special Senses System 

Harderian gland 

Adenoma 

+ 

X 

Urinary System 

Kidney + 

Histiocytic sarcoma 

Urinary bladder + 

Systemic Lesions 

Multiple organs + 

Histiocytic sarcoma 

Lymphoma malignant 

+ 

+ 

+ 

+ 

A 

+ 

+ 

X 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

A 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

X 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 
 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 187 

TABLE D2
 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

0.3 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 Total 

Carcass ID Number 1 

6 

1 

7 

2 

2 

2 

8 

3 

0 

3 

5 

3 

7 

3 

9 

4 

2 

4 

5 

4 

8 

4 

9 

5 

0 

0 

2 

0 

5 

0 

7 

1 

1 

1 

2 

1 

5 

1 

8 

1 

9 

2 

4 

2 

5 

3 

6 

4 

6 

Tissues/ 

Tumors 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, bronchial 

Histiocytic sarcoma

Lymph node, mandibular 

Histiocytic sarcoma

Lymph node, mesenteric 

Histiocytic sarcoma

Lymph node, mediastinal 

Histiocytic sarcoma

Spleen 

Histiocytic sarcoma

Thymus 

Histiocytic sarcoma

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + +

+ + M M + + + + + + + + + M + M + + M + M + + + + 

+ + + + + + M M + + M + M + + + + + + + + + + + + 

+ + + + + M + + + + + + + + + + + + I + + + + + + 

+ M + + M + + + + + + + + + M + + + + M M M + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + M + + 

50 

6 

34 

1 

37 

2 

45 

2 

36 

2 

50 

2 

44 

1 

Integumentary System 

Mammary gland 

Carcinoma 

Skin 

Subcutaneous tissue, sarcoma

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

50 

1 

50 

2 

Musculoskeletal System 

Bone 

Skeletal muscle

+ + + + + + + + + + + + + + + + + + + + + + + + + 50 

1 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma 

Histiocytic sarcoma

Nose 

Trachea 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X

X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

49 

50 

5 

1 

1 

2 

50 

50 

Special Senses System 

Harderian gland 

Adenoma 

+

X

 2 

2 

Urinary System 

Kidney 

Histiocytic sarcoma

Urinary bladder 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

M + + + + + + + + + + + + + + + + + + + + + + + + 

50 

1 

47 

Systemic Lesions 

Multiple organs 

Histiocytic sarcoma

Lymphoma malignant 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X

50 

2 

7 
 

 

 

 

 

 

 

 

 

188 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D2
 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

Number of Days on Study 

0 

0 

9 

4 

9 

5 

5 

0 

5 

5 

2 

4 

5 

4 

8 

5 

6 

9 

6 

2 

2 

6 

4 

0 

6 

4 

9 

6 

5 

9 

6 

6 

6 

6 

6 

6 

6 

6 

7 

6 

7 

4 

7 

0 

3 

7 

0 

9 

7 

2 

0 

7 

3 

3 

7 

3 

4 

7 

3 

4 

7 

3 

4 

7 

3 

4 

7 

3 

5 

7 

3 

5 

7 

3 

5 

Carcass ID Number 

5 

2 

0 

5 

0 

7 

5 

1 

9 

5 

2 

2 

5 

3 

7 

5 

4 

4 

5 

3 

8 

5 

4 

8 

5 

4 

3 

5 

1 

0 

5 

1 

3 

5 

3 

5 

5 

3 

0 

5 

0 

5 

5 

2 

3 

5 

2 

9 

5 

1 

4 

5 

0 

9 

5 

0 

3 

5 

1 

1 

5 

1 

8 

5 

3 

9 

5 

0 

1 

5 

0 

2 

5 

0 

4 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Intestine small, duodenum 

Intestine small, jejunum 

Hemangiosarcoma 

Intestine small, ileum 

Liver 

Hemangiosarcoma 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Mesentery 

Hemangiosarcoma 

Pancreas 

Salivary glands 

Stomach, forestomach 

Stomach, glandular 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

A 

+ 

+ 

+ 

A 

A 

A 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

X 

+ 

X 

+ 

+ 

+ 

+ 

+ 

A 

+ 

+ 

A 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

A 

A 

A 

A 

A 

A 

A 

A 

+ 

X 

M 

+ 

A 

A 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

A 

+ 

+ 

+ 

A 

A 

A 

+ 

X 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

Cardiovascular System 

Heart + 

Endocrine System 

Adrenal cortex + 

Adrenal medulla + 

Islets, pancreatic + 

Parathyroid gland + 

Pituitary gland + 

Pars distalis, adenoma 

Thyroid gland + 

Follicular cell, carcinoma 

General Body System 

None 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

X 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

A 

A 

M 

A 

A 

A 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

I 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

Genital System 

Clitoral gland + 

Ovary + 

Cystadenoma 

Granulosa cell tumor benign 

Luteoma 

Teratoma benign 

Yolk sac carcinoma 

Uterus + 

Hemangioma 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

A 

A 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 
 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 189 

TABLE D2
 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

5 5 5 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 

5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 Total 

Carcass ID Number 0 0 1 1 2 2 3 3 4 4 4 5 1 1 2 2 2 2 3 3 3 4 4 4 4 Tissues/ 

6 8 5 6 1 4 1 6 1 2 9 0 2 7 5 6 7 8 2 3 4 0 5 6 7 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Gallbladder + M + + M + + + + + + + + + + + M + + + + + + + + 38 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Hemangiosarcoma 1 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Hemangiosarcoma X 3 

Hepatocellular carcinoma X X X X X X 16 

Hepatocellular adenoma X X 11 

Hepatocellular adenoma, multiple X 2 

Mesentery + + + + 8 

Hemangiosarcoma 1 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Parathyroid gland + + + + + + + + M + + M M + + + + + + M M M + M M 28 

Pituitary gland + I + + + + + + + + + + + + + + + + + + + + + M + 47 

Pars distalis, adenoma X X X 7 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Follicular cell, carcinoma X X 2 

General Body System 

None 

Genital System 

Clitoral gland + M + + + + + + + + + + + + + + + + M M M + + + + 43 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Cystadenoma X X X 3 

Granulosa cell tumor benign X 1 

Luteoma X 1 

Teratoma benign 1 

Yolk sac carcinoma X 1 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Hemangioma X 2 
 

 

 

 

 

 

 

 

 

 

 

190 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D2
 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

0 4 5 5 5 5 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 0 

9 

9 

5 

0 

5 

2 

4 

4 

8 

6 

9 

2 

2 

4 

0 

4 

9 

5 

9 

6 

6 

6 

6 

6 

7 

7 

4 

0 

3 

0 

9 

2 

0 

3 

3 

3 

4 

3 

4 

3 

4 

3 

4 

3 

5 

3 

5 

3 

5 

5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 

Carcass ID Number 2 0 1 2 3 4 3 4 4 1 1 3 3 0 2 2 1 0 0 1 1 3 0 0 0 

0 7 9 2 7 4 8 8 3 0 3 5 0 5 3 9 4 9 3 1 8 9 1 2 4 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Thymus 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

M 

+ 

+ 

M 

+ 

+ 

+ 

M 

I 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

M 

M 

+ 

M 

+ 

+ 

+ 

M 

+ 

M 

+ 

+ 

+ 

+ 

M 

M 

M 

M 

+ 

M 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

M 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

M 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

M 

+ 

+ 

+ 

+ 

+ 

A 

M 

+ 

A 

A 

M 

A 

+ 

M 

M 

+ 

M 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

M 

M 

+ 

M 

+ 

M 

+ 

+ 

M 

+ 

M 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

M 

+ 

M 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

Integumentary System 

Mammary gland 

Carcinoma 

Skin 

Hemangiosarcoma 

Subcutaneous tissue, mast cell tumor benign 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

Musculoskeletal System 

Bone 

Skeletal muscle 

Sarcoma 

+ 

+ 

+ + + + + + + + + + + + + + + + + + + + + + + + 

Nervous System 

Brain + 

Respiratory System 

Larynx + 

Lung + 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver 

Nose + 

Trachea + 

Special Senses System 

None 

Urinary System 

Kidney + 

Urinary bladder + 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

I 

+ 

X 

+ 

+ 

+ 

A 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

I 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

A 

+ 

X 

A 

A 

+ 

A 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

A 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

I 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Lymphoma malignant X X X X 
 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 191 

TABLE D2
 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

1.0 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 5 Total 

Carcass ID Number 0 

6 

0 

8 

1 

5 

1 

6 

2 

1 

2 

4 

3 

1 

3 

6 

4 

1 

4 

2 

4 

9 

5 

0 

1 

2 

1 

7 

2 

5 

2 

6 

2 

7 

2 

8 

3 

2 

3 

3 

3 

4 

4 

0 

4 

5 

4 

6 

4 

7 

Tissues/ 

Tumors 

Hematopoietic System 

Bone marrow 

Lymph node 

Lymph node, bronchial 

Lymph node, mandibular 

Lymph node, mesenteric 

Lymph node, mediastinal 

Spleen 

Thymus 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

M 

+ 

M 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

M 

+ 

+ 

M 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

M 

+ 

+ 

+ 

M 

M 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

M 

+ 

M 

+ 

+ 

+ 

+

+ 

+ 

+ 

M 

+ 

+ 

49 

3 

27 

36 

46 

28 

49 

41 

Integumentary System 

Mammary gland 

Carcinoma 

Skin 

Hemangiosarcoma 

Subcutaneous tissue, mast cell tumor benign 

+ 

+ 

+ 

+ 

+ 

X

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

50 

1 

50 

1 

1 

Musculoskeletal System 

Bone 

Skeletal muscle 

Sarcoma 

+ + + + + + + + + + + + + + + + + + + + 

+

X

+ + + + + 50 

2 

1 

Nervous System 

Brain + 

Respiratory System 

Larynx + 

Lung + 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver

Nose + 

Trachea + 

Special Senses System 

None 

Urinary System 

Kidney + 

Urinary bladder + 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

I 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 50 

+ 47 

+ 50 

X 8 

1 

4 

5 

+ 49 

+ 49 

+ 50 

+ 45 

Systemic Lesions 

Multiple organs + 

Lymphoma malignant 

+ 

X 

+ + + + + 

X 

+ + + + + + + + + + 

X

+ + + + + + + + 50 

7 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

192 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D2
 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

Number of Days on Study 

3 

5 

6 

4 

2 

9 

4 

4 

3 

4 

6 

2 

4 

8 

4 

5 

3 

6 

5 

3 

9 

6 

0 

6 

6 

0 

9 

6 

1 

1 

6 

3 

9 

6 

6 

7 

6 

7 

5 

6 

8 

0 

6 

8 

6 

6 

9 

2 

7 

0 

6 

7 

1 

2 

7 

1 

7 

7 

1 

9 

7 

2 

6 

7 

3 

2 

7 

3 

4 

7 

3 

4 

7 

3 

4 

Carcass ID Number 

7 

0 

7 

7 

3 

1 

7 

1 

3 

7 

2 

0 

7 

2 

2 

7 

2 

6 

7 

0 

3 

7 

0 

8 

7 

2 

7 

7 

4 

0 

7 

4 

6 

7 

4 

9 

7 

3 

3 

7 

0 

2 

7 

1 

8 

7 

1 

1 

7 

0 

4 

7 

1 

7 

7 

4 

3 

7 

2 

3 

7 

4 

2 

7 

1 

0 

7 

1 

4 

7 

3 

0 

7 

3 

8 

Alimentary System 

Esophagus 

Gallbladder 

Intestine large, colon 

Intestine large, rectum 

Intestine large, cecum 

Leiomyoma 

Intestine small, duodenum 

Intestine small, jejunum 

Intestine small, ileum 

Liver 

Hepatocellular carcinoma 

Hepatocellular adenoma 

Hepatocellular adenoma, multiple 

Histiocytic sarcoma 

Mesentery 

Hemangioma 

Pancreas 

Salivary glands 

Stomach, forestomach 

Squamous cell papilloma 

Stomach, glandular 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

A 

+ 

+ 

+ 

A 

A 

A 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

X 

+ 

+ 

+ 

+ 

+ 

A 

+ 

+ 

+ 

+ 

A 

A 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

M 

M 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

A 

A 

A 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

A 

+ 

+ 

A 

A 

A 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

A 

A 

A 

A 

A 

A 

A 

A 

A 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

A 

+ 

A 

A 

A 

A 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

Cardiovascular System 

Heart + 

Endocrine System 

Adrenal cortex + 

Adrenal medulla + 

Islets, pancreatic + 

Adenoma 

Parathyroid gland + 

Pituitary gland + 

Pars distalis, adenoma 

Thyroid gland + 

Follicular cell, adenoma 

General Body System 

None 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

I 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

I 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

I 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

I 

+ 

X 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

A 

+ 

A 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

+ 

+ 

+ 

+ 

I 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

X 

+ 

X 

Genital System 

Clitoral gland + 

Ovary + 

Cystadenoma 

Histiocytic sarcoma 

Uterus + 

Histiocytic sarcoma 

Leiomyosarcoma 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 

X 

+ 

+ 

+ 

+ 

+ 

+ 

M 

+ 

+ 

M 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

I 

+ 

+ 

A 

A 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

+ 

X 

+ 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 193 

TABLE D2
 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

5 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 Total 

Carcass ID Number 0 0 1 1 2 3 3 3 3 5 0 0 1 1 2 2 2 2 3 3 4 4 4 4 4 Tissues/ 

6 9 5 9 1 2 5 6 7 0 1 5 2 6 4 5 8 9 4 9 1 4 5 7 8 Tumors 

Alimentary System 

Esophagus + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Gallbladder + + + + + + + + + + + + + + + + + + + + + + + + + 43 

Intestine large, colon + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Intestine large, rectum + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Intestine large, cecum + + + + + + + + + + + + + + + + + + + + + + + + + 47 

Leiomyoma 1 

Intestine small, duodenum + + + + + + + + + + + + + + + + + + + + + + + + + 45 

Intestine small, jejunum + + + + + + + + + + + + + + + + + + + + + + + + + 44 

Intestine small, ileum + + + + + + + + + + + + + + + + + + + + + + + + + 45 

Liver + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Hepatocellular carcinoma X 4 

Hepatocellular adenoma X X X X 9 

Hepatocellular adenoma, multiple X X 3 

Histiocytic sarcoma 2 

Mesentery + + + + + + 7 

Hemangioma X 1 

Pancreas + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Salivary glands + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Stomach, forestomach + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Squamous cell papilloma X 1 

Stomach, glandular + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Cardiovascular System 

Heart + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Endocrine System 

Adrenal cortex + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Adrenal medulla + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Islets, pancreatic + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Adenoma X 1 

Parathyroid gland + M M M + M M + + M M M I M M M + + + + + M + + M 26 

Pituitary gland + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Pars distalis, adenoma X X X 8 

Thyroid gland + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Follicular cell, adenoma X X X 5 

General Body System 

None 

Genital System 

Clitoral gland + + + M M + + + M M + M + + M + + + + + M + + + M 40 

Ovary + + + + + + + + + + + + + + + + + + + + + + + + + 48 

Cystadenoma X X 3 

Histiocytic sarcoma 1 

Uterus + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Histiocytic sarcoma 1 

Leiomyosarcoma X 1 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

194 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D2
 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

3 4 4 4 4 5 5 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 

Number of Days on Study 5 2 4 6 8 3 3 0 0 1 3 6 7 8 8 9 0 1 1 1 2 3 3 3 3 

6 9 3 2 4 6 9 6 9 1 9 7 5 0 6 2 6 2 7 9 6 2 4 4 4 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Carcass ID Number 0 3 1 2 2 2 0 0 2 4 4 4 3 0 1 1 0 1 4 2 4 1 1 3 3 

7 1 3 0 2 6 3 8 7 0 6 9 3 2 8 1 4 7 3 3 2 0 4 0 8 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X X 

Lymph node + + + 

Lymph node, bronchial + M + M + + M M + M M M M + M M M + M M + M + + + 

Histiocytic sarcoma X 

Lymph node, mandibular M + + + + + + M + + M + M + M + + + + + M + M + + 

Histiocytic sarcoma X 

Lymph node, mesenteric + + + + + + M + + + M + + + + + + + A + + I + + + 

Histiocytic sarcoma X 

Lymph node, mediastinal + + + + + + + + + M M + + + + + + + + + + M + M M 

Alveolar/bronchiolar carcinoma, metastatic, lung X 

Histiocytic sarcoma X 

Spleen + + + + + + + + + + + + + + + + + + A + + + + + + 

Histiocytic sarcoma X 

Thymus + M M + + + + M M + + M M + M + + + + + + M + + + 

Integumentary System 

Mammary gland + + + + + + + + + + + + + + + + + + + + + + + + + 

Adenoma 

Carcinoma X 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 

Skeletal muscle 

Sarcoma 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 

Spinal cord + 

Respiratory System 

Larynx + + + + + + + + + + + + + + + + + + + + + + + + + 

Lung + + + + + + + + + + + + + + + + + + + + + + + + + 

Alveolar/bronchiolar adenoma X 

Alveolar/bronchiolar adenoma, multiple X 

Alveolar/bronchiolar carcinoma X X X X X X 

Hepatocellular carcinoma, metastatic, liver X X 

Histiocytic sarcoma X 

Nose + + + + + + + + + + + + + + + + + + A + + A + + + 

Trachea + + + + + + + + + + + + + + + + + + A + + + + + + 

Special Senses System 

Harderian gland 

Adenoma 

Urinary System 

Kidney + + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X 

Urinary bladder + + A + + I + + + + + + + + + + + + A + + A + + + 
 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 195 

TABLE D2
 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

5 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

3 

6 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 Total 

Carcass ID Number 0 

6 

0 

9 

1 

5 

1 

9 

2 

1 

3 

2 

3 

5 

3 

6 

3 

7 

5 

0 

0 

1 

0 

5 

1 

2 

1 

6 

2 

4 

2 

5 

2 

8 

2 

9 

3 

4 

3 

9 

4 

1 

4 

4 

4 

5 

4 

7 

4 

8 

Tissues/ 

Tumors 

Hematopoietic System 

Bone marrow + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma

Lymph node 

Lymph node, bronchial 

Histiocytic sarcoma

Lymph node, mandibular 

Histiocytic sarcoma

Lymph node, mesenteric 

Histiocytic sarcoma

Lymph node, mediastinal 

Alveolar/bronchiolar carcinoma, metastatic, lung 

Histiocytic sarcoma

Spleen 

Histiocytic sarcoma

Thymus 

+

+ + + + + + + + + + + + + + + + + + + + + + M + + 

+ M M + + + + + + M M I + M + + + + + + M + + + + 

+ + + + + + + + + + + + + M + + + + + M + + + + + 

+ M M M M + + + M + M M + M + M + + + M M + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + M 

2 

4 

35 

1 

36 

1 

44 

1 

34 

1 

1 

49 

1 

41 

Integumentary System 

Mammary gland 

Adenoma 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X

50 

1 

Carcinoma 1 

Skin + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 1 

Musculoskeletal System 

Bone + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Skeletal muscle + 1 

Sarcoma X 1 

Nervous System 

Brain + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Spinal cord 1 

Respiratory System 

Larynx 

Lung 

Alveolar/bronchiolar adenoma 

Alveolar/bronchiolar adenoma, multiple 

Alveolar/bronchiolar carcinoma 

Hepatocellular carcinoma, metastatic, liver

Histiocytic sarcoma

Nose 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

X X X X X X X

X 

X X X

+ + + + + + + + + + + + + + + + + + + + + + + + + 

50 

50 

8 

2 

9 

2 

1 

48 

Trachea + + + + + + + + + + + + + + + + + + + + + + + + + 49 

Special Senses System 

Harderian gland 

Adenoma 

+

X

 1 

1 

Urinary System 

Kidney 

Histiocytic sarcoma

Urinary bladder 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

+ + + + + + + + + + + + + + + + + + + + + + + + + 

50 

1 

46 
 

 

196 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D2
 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

3 4 4 4 4 5 5 6 6 6 6 6 6 6 6 6 7 7 7 7 7 7 7 7 7 

Number of Days on Study 5 2 4 6 8 3 3 0 0 1 3 6 7 8 8 9 0 1 1 1 2 3 3 3 3 

6 9 3 2 4 6 9 6 9 1 9 7 5 0 6 2 6 2 7 9 6 2 4 4 4 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Carcass ID Number 0 3 1 2 2 2 0 0 2 4 4 4 3 0 1 1 0 1 4 2 4 1 1 3 3 

7 1 3 0 2 6 3 8 7 0 6 9 3 2 8 1 4 7 3 3 2 0 4 0 8 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 

Histiocytic sarcoma X X X 

Lymphoma malignant X X X X 
 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 197 

TABLE D2
 

Individual Animal Tumor Pathology of Female Mice in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate: 

3.0 mg/m3 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 

Number of Days on Study 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 3 

5 5 5 5 5 5 5 5 5 5 6 6 6 6 6 6 6 6 6 6 6 6 6 6 6 

7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 7 Total 

Carcass ID Number 0 0 1 1 2 3 3 3 3 5 0 0 1 1 2 2 2 2 3 3 4 4 4 4 4 Tissues/ 

6 9 5 9 1 2 5 6 7 0 1 5 2 6 4 5 8 9 4 9 1 4 5 7 8 Tumors 

Systemic Lesions 

Multiple organs + + + + + + + + + + + + + + + + + + + + + + + + + 50 

Histiocytic sarcoma 3 

Lymphoma malignant X 5 
198 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D3 

Statistical Analysis of Primary Neoplasms in Female Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Liver: Hemangiosarcoma 

Overall ratea 

Adjusted rateb 

Terminal ratec 

First incidence (days) 

Life table testd 

Logistic regression testd 

Cochran-Armitage testd 

Fisher exact testd 

1/50 (2%) 

2.9% 

1/34 (3%) 

734 (T) 

Pù0.499N 

Pù0.431N 

Pù0.455N 

0/50 (0%) 

0.0% 

0/37 (0%) 

—e 

Pù0.483N 

Pù0.483N 

Pù0.500N 

3/50 (6%) 

7.3% 

1/32 (3%) 

524 

Pù0.298 

Pù0.318 

Pù0.309 

0/49 (0%) 

0.0% 

0/28 (0%) 

— 

Pù0.539N 

Pù0.539N 

Pù0.505N 

Liver: Hepatocellular Adenoma 

Overall rate 8/50 (16%) 10/50 (20%) 13/50 (26%) 12/49 (24%) 

Adjusted rate 22.9% 25.0% 34.0% 35.3% 

Terminal rate 7/34 (21%) 8/37 (22%) 8/32 (25%) 8/28 (29%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

713 

Pù0.103 

Pù0.174 

Pù0.221 

593 

Pù0.480 

Pù0.466 

Pù0.398 

622 

Pù0.150 

Pù0.152 

Pù0.163 

539 

Pù0.128 

Pù0.180 

Pù0.212 

Liver: Hepatocellular Carcinoma 

Overall rate 12/50 (24%) 9/50 (18%) 16/50 (32%) 4/49 (8%) 

Adjusted rate 31.7% 22.7% 40.5% 11.8% 

Terminal rate 9/34 (26%) 7/37 (19%) 9/32 (28%) 1/28 (4%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

609 

Pù0.094N 

Pù0.051N 

Pù0.036N 

666 

Pù0.240N 

Pù0.258N 

Pù0.312N 

640 

Pù0.235 

Pù0.238 

Pù0.252 

667 

Pù0.063N 

Pù0.037N 

Pù0.030N 

Liver: Hepatocellular Adenoma or Carcinoma 

Overall rate 18/50 (36%) 18/50 (36%) 24/50 (48%) 16/49 (33%) 

Adjusted rate 48.1% 43.3% 56.7% 43.8% 

Terminal rate 15/34 (44%) 14/37 (38%) 14/32 (44%) 9/28 (32%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

609 

Pù0.444 

Pù0.474N 

Pù0.374N 

593 

Pù0.451N 

Pù0.488N 

Pù0.582N 

622 

Pù0.145 

Pù0.137 

Pù0.156 

539 

Pù0.530 

Pù0.506N 

Pù0.445N 

Lung: Alveolar/bronchiolar Adenoma 

Overall rate 3/50 (6%) 6/50 (12%) 9/50 (18%) 10/50 (20%) 

Adjusted rate 8.8% 15.0% 25.2% 32.8% 

Terminal rate 3/34 (9%) 4/37 (11%) 6/32 (19%) 8/28 (29%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

734 (T) 

Pù0.014 

Pù0.024 

Pù0.045 

664 

Pù0.297 

Pù0.287 

Pù0.243 

649 

Pù0.056 

Pù0.057 

Pù0.061 

706 

Pù0.016 

Pù0.024 

Pù0.036 
Cobalt Sulfate Heptahydrate, NTP TR 471 199 

TABLE D3 

Statistical Analysis of Primary Neoplasms in Female Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Lung: Alveolar/bronchiolar Carcinoma 

Overall rate 1/50 (2%) 1/50 (2%) 4/50 (8%) 9/50 (18%) 

Adjusted rate 2.9% 2.7% 9.2% 25.3% 

Terminal rate 1/34 (3%) 1/37 (3%) 1/32 (3%) 4/28 (14%) 

First incidence (days) 

Life table test 

Logistic regression test 

734 (T) 

P<0.001 

P<0.001 

734 (T) 

Pù0.743N 

Pù0.743N 

495 

Pù0.173 

Pù0.201 

536 

Pù0.007 

Pù0.009 

Cochran-Armitage test 

Fisher exact test 

P<0.001 

Pù0.753N Pù0.181 Pù0.008 

Lung: Alveolar/bronchiolar Adenoma or Carcinoma 

Overall rate 4/50 (8%) 7/50 (14%) 13/50 (26%) 18/50 (36%) 

Adjusted rate 11.8% 17.5% 32.6% 50.2% 

Terminal rate 4/34 (12%) 5/37 (14%) 7/32 (22%) 11/28 (39%) 

First incidence (days) 

Life table test 

Logistic regression test 

734 (T) 

P<0.001 

P<0.001 

664 

Pù0.322 

Pù0.318 

495 

Pù0.016 

Pù0.016 

536 

P<0.001 

P<0.001 

Cochran-Armitage test 

Fisher exact test 

P<0.001 

Pù0.262 Pù0.016 P<0.001 

Ovary: Cystadenoma 

Overall rate 2/48 (4%) 3/49 (6%) 3/49 (6%) 3/48 (6%) 

Adjusted rate 6.3% 8.3% 9.4% 10.7% 

Terminal rate 2/32 (6%) 3/36 (8%) 3/32 (9%) 3/28 (11%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

734 (T) 

Pù0.390 

Pù0.390 

Pù0.487 

734 (T) 

Pù0.554 

Pù0.554 

Pù0.510 

734 (T) 

Pù0.500 

Pù0.500 

Pù0.510 

734 (T) 

Pù0.439 

Pù0.439 

Pù0.500 

Ovary: Cystadenoma or Cystadenocarcinoma 

Overall rate 2/48 (4%) 4/49 (8%) 3/49 (6%) 3/48 (6%) 

Adjusted rate 6.3% 11.1% 9.4% 10.7% 

Terminal rate 2/32 (6%) 4/36 (11%) 3/32 (9%) 3/28 (11%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

734 (T) 

Pù0.390 

Pù0.390 

Pù0.487 

734 (T) 

Pù0.395 

Pù0.395 

Pù0.349 

734 (T) 

Pù0.500 

Pù0.500 

Pù0.510 

734 (T) 

Pù0.439 

Pù0.439 

Pù0.500 

Pituitary Gland (Pars Distalis): Adenoma 

Overall rate 11/48 (23%) 8/47 (17%) 7/47 (15%) 8/48 (17%) 

Adjusted rate 30.0% 22.2% 20.7% 24.7% 

Terminal rate 8/33 (24%) 8/36 (22%) 5/30 (17%) 5/28 (18%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

596 

Pù0.538N 

Pù0.430N 

Pù0.355N 

734 (T) 

Pù0.236N 

Pù0.255N 

Pù0.323N 

659 

Pù0.271N 

Pù0.248N 

Pù0.231N 

675 

Pù0.426N 

Pù0.343N 

Pù0.305N 
200 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D3 

Statistical Analysis of Primary Neoplasms in Female Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

3 3 3 Chamber Control 0.3 mg/m 1.0 mg/m 3.0 mg/m 

Skin (Subcutaneous Tissue): Sarcoma 

Overall rate 6/50 (12%) 2/50 (4%) 0/50 (0%) 0/50 (0%)
 

Adjusted rate 16.0% 4.3% 0.0% 0.0%
 

Terminal rate 4/34 (12%) 0/37 (0%) 0/32 (0%) 0/28 (0%)
 

First incidence (days) 225 593 — —
 

Life table test Pù0.022N Pù0.116N Pù0.022N Pù0.029N
 

Logistic regression test Pù0.012N Pù0.210N Pù0.014N Pù0.014N
 

Cochran-Armitage test Pù0.015N
 

Fisher exact test Pù0.134N Pù0.013N Pù0.013N
 

Thyroid Gland (Follicular Cell): Adenoma 

Overall rate 3/50 (6%) 0/49 (0%) 0/49 (0%) 5/49 (10%) 

Adjusted rate 8.0% 0.0% 0.0% 16.7% 

Terminal rate 2/34 (6%) 0/36 (0%) 0/32 (0%) 4/28 (14%) 

First incidence (days) 619 — — 686 

Life table test Pù0.026 Pù0.113N Pù0.130N Pù0.274 

Logistic regression test Pù0.038 Pù0.126N Pù0.125N Pù0.323 

Cochran-Armitage test Pù0.046 

Fisher exact test Pù0.125N Pù0.125N Pù0.346 

Thyroid Gland (Follicular Cell): Adenoma or Carcinoma 

Overall rate 4/50 (8%) 0/49 (0%) 2/49 (4%) 5/49 (10%) 

Adjusted rate 10.6% 0.0% 6.3% 16.7% 

Terminal rate 2/34 (6%) 0/36 (0%) 2/32 (6%) 4/28 (14%) 

First incidence (days) 619 — 734 (T) 686 

Life table test Pù0.081 Pù0.056N Pù0.360N Pù0.407 

Logistic regression test Pù0.108 Pù0.065N Pù0.351N Pù0.462 

Cochran-Armitage test Pù0.130 

Fisher exact test Pù0.061N Pù0.349N Pù0.487 

All Organs: Hemangiosarcoma 

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

Adjusted rate 2.9% 4.8% 10.3% 0.0%
 

Terminal rate 1/34 (3%) 1/37 (3%) 2/32 (6%) 0/28 (0%)
 

First incidence (days) 734 (T) 652 524 —
 

Life table test Pù0.321N Pù0.536 Pù0.173 Pù0.539N 

Logistic regression test Pù0.250N Pù0.505 Pù0.184 Pù0.539N 

Cochran-Armitage test Pù0.263N
 

Fisher exact test Pù0.500 Pù0.181 Pù0.500N 

All Organs: Hemangioma or Hemangiosarcoma 

Overall rate 1/50 (2%) 3/50 (6%) 6/50 (12%) 1/50 (2%)
 

Adjusted rate 2.9% 7.5% 15.8% 3.6%
 

Terminal rate 1/34 (3%) 2/37 (5%) 3/32 (9%) 1/28 (4%)
 

First incidence (days) 734 (T) 652 524 734 (T)
 

Life table test Pù0.499N Pù0.342 Pù0.057 Pù0.718 

Logistic regression test Pù0.406N Pù0.319 Pù0.059 Pù0.718 

Cochran-Armitage test Pù0.403N
 

Fisher exact test Pù0.309 Pù0.056 Pù0.753N 
Cobalt Sulfate Heptahydrate, NTP TR 471 201 

TABLE D3 

Statistical Analysis of Primary Neoplasms in Female Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

All Organs: Histiocytic Sarcoma 

Overall rate 3/50 (6%) 2/50 (4%) 0/50 (0%) 3/50 (6%) 

Adjusted rate 7.7% 4.8% 0.0% 7.9% 

Terminal rate 0/34 (0%) 1/37 (3%) 0/32 (0%) 0/28 (0%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

656 

Pù0.456 

Pù0.496 

Pù0.500 

649 

Pù0.452N 

Pù0.396N 

Pù0.500N 

— 

Pù0.126N 

Pù0.118N 

Pù0.121N 

639 

Pù0.644 

Pù0.644N 

Pù0.661N 

All Organs: Malignant Lymphoma 

Overall rate 4/50 (8%) 7/50 (14%) 7/50 (14%) 5/50 (10%) 

Adjusted rate 10.8% 16.8% 19.8% 13.4% 

Terminal rate 3/34 (9%) 3/37 (8%) 5/32 (16%) 1/28 (4%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

583 

Pù0.492 

Pù0.524N 

Pù0.528N 

666 

Pù0.327 

Pù0.267 

Pù0.262 

649 

Pù0.240 

Pù0.255 

Pù0.262 

356 

Pù0.429 

Pù0.527 

Pù0.500 

All Organs: Benign Neoplasms 

Overall rate 25/50 (50%) 32/50 (64%) 27/50 (54%) 27/50 (54%) 

Adjusted rate 65.5% 76.0% 68.9% 72.1% 

Terminal rate 21/34 (62%) 27/37 (73%) 20/32 (63%) 18/28 (64%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

596 

Pù0.185 

Pù0.408 

Pù0.465N 

593 

Pù0.242 

Pù0.220 

Pù0.113 

622 

Pù0.336 

Pù0.390 

Pù0.421 

539 

Pù0.163 

Pù0.305 

Pù0.421 

All Organs: Malignant Neoplasms 

Overall rate 27/50 (54%) 22/50 (44%) 31/50 (62%) 21/50 (42%) 

Adjusted rate 58.5% 47.6% 68.3% 49.5% 

Terminal rate 15/34 (44%) 13/37 (35%) 18/32 (56%) 7/28 (25%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

225 

Pù0.530N 

Pù0.187N 

Pù0.199N 

593 

Pù0.154N 

Pù0.266N 

Pù0.212N 

495 

Pù0.268 

Pù0.276 

Pù0.272 

356 

Pù0.368N 

Pù0.147N 

Pù0.158N 
202 

c 

Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D3 

Statistical Analysis of Primary Neoplasms in Female Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

All Organs: Benign or Malignant Neoplasms 

Overall rate 42/50 (84%) 41/50 (82%) 43/50 (86%) 39/50 (78%) 

Adjusted rate 89.3% 87.2% 91.4% 84.8% 

Terminal rate 29/34 (85%) 31/37 (84%) 28/32 (88%) 21/28 (75%) 

First incidence (days) 

Life table test 

Logistic regression test 

Cochran-Armitage test 

Fisher exact test 

225 

Pù0.213 

Pù0.366N 

Pù0.258N 

593 

Pù0.255N 

Pù0.422N 

Pù0.500N 

495 

Pù0.387 

Pù0.459 

Pù0.500 

356 

Pù0.395 

Pù0.353N 

Pù0.306N 

(T)Terminal sacrifice 

a Number of neoplasm-bearing animals/number of animals examined. Denominator is number of animals examined microscopically for liver, lung, ovary, 

pituitary gland, and thyroid gland; for other tissues, denominator is number of animals necropsied. b	 Kaplan-Meier estimated neoplasm incidence at the end of the study after adjustment for intercurrent mortality 

Observed incidence at terminal kill 

d	 Beneath the chamber control incidence are the P values associated with the trend test. Beneath the exposed group incidence are the P values corresponding 

to pairwise comparisons between the chamber controls and that exposed group. The life table test regards neoplasms in animals dying prior to terminal kill 

as being (directly or indirectly) the cause of death. The logistic regression test regards these lesions as nonfatal. The Cochran-Armitage and Fisher exact 

tests compare directly the overall incidence rates. For all tests, a negative trend or a lower incidence in an exposure group is indicated by N. e Not applicable; no neoplasms in animal group 
Cobalt Sulfate Heptahydrate, NTP TR 471 203 

TABLE D4a 

Historical Incidence of Alveolar/bronchiolar Neoplasms in Chamber Control Female B6C3F a 1 Mice

Incidence in Controls 

Study Adenoma Carcinoma Adenoma or Carcinoma 

Historical Incidence at Battelle Pacific Northwest Laboratories 

1,3-Butadiene 4/50 0/50 4/50 

Acetonitrile 7/49 1/49 8/49 

Allyl Glycidyl Ether 0/50 0/50 0/50 

2-Chloroacetophenone 4/50 3/50 6/50 

l-Epinephrine Hydrochloride 3/50 2/50 5/50 

Chloroethane 2/49 3/49 5/49 

Hexachlorocyclopentadiene 4/48 3/48 7/48 

o-Chlorobenzalmalononitrile (CS2) 4/50 1/50 5/50 

Ozone 4/50 2/50 6/50 

Overall Historical Incidence 

Total 61/939 (6.5%) 38/939 (4.1%) 97/939 (10.3%) 

Standard deviation 3.2% 3.2% 3.7% 

Range 0%-14% 0%-12% 0%-16% 

a Data as of 12 May 1995 

TABLE D4b 

Historical Incidence of Hemangiosarcoma of the Liver in Chamber Control Female B6C3F a 1 Mice

Study Incidence in Controls 

Historical Incidence at Battelle Pacific Northwest Laboratories 

1,3-Butadiene 1/49 

Acetonitrile 0/49 

Allyl Glycidyl Ether 0/50 

2-Chloroacetophenone 0/50 

l-Epinephrine Hydrochloride 1/50 

Chloroethane 0/49 

Hexachlorocyclopentadiene 0/49 

o-Chlorobenzalmalononitrile (CS2) 0/50 

Ozone 0/50 

Overall Historical Incidence 

Total 5/937 (0.5%) 

Standard deviation 1.0% 

Range 0%-3% 

a Data as of 12 May 1995 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

204 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D5 

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydratea 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Disposition Summary 

Animals initially in study 50 50 50 50 

Early deaths 

Moribund 11 10 13 16 

Natural deaths 5 3 5 6 

Survivors 

Terminal sacrifice 34 37 32 28 

Animals examined microscopically 50 50 50 50 

Alimentary System 

Intestine large, cecum (49) (49) (48) (47) 

Inflammation 1 (2%) 

Intestine small, ileum (48) (48) (47) (45) 

Peyer's patch, hyperplasia, lymphoid 1 (2%) 

Liver (50) (50) (50) (49) 

Basophilic focus 1 (2%) 

Clear cell focus 2 (4%) 1 (2%) 1 (2%) 1 (2%) 

Clear cell focus, multiple 1 (2%) 

Cyst 1 (2%) 1 (2%) 

Eosinophilic focus 9 (18%) 7 (14%) 8 (16%) 9 (18%) 

Eosinophilic focus, multiple 1 (2%) 

Hematopoietic cell proliferation 1 (2%) 1 (2%) 

Hemorrhage 1 (2%) 

Infiltration cellular, lymphocyte 3 (6%) 2 (4%) 1 (2%) 1 (2%) 

Infiltration cellular, mixed cell 1 (2%) 

Inflammation, chronic 6 (12%) 1 (2%) 1 (2%) 2 (4%) 

Karyomegaly 4 (8%) 2 (4%) 1 (2%) 

Mineralization 1 (2%) 

Necrosis 1 (2%) 

Necrosis, focal 3 (6%) 1 (2%) 2 (4%) 2 (4%) 

Regeneration 1 (2%) 

Thrombosis 1 (2%) 

Vacuolization cytoplasmic 1 (2%) 

Vacuolization cytoplasmic, diffuse 1 (2%) 1 (2%) 

Oval cell, hyperplasia 2 (4%) 1 (2%) 

Serosa, fibrosis 1 (2%) 

Mesentery (10) (12) (8) (7) 

Angiectasis 1 (10%) 

Hemorrhage 1 (14%) 

Infiltration cellular, lymphocyte 1 (8%) 

Inflammation, chronic 1 (8%) 

Fat, necrosis 7 (70%) 8 (67%) 7 (88%) 5 (71%) 

Pancreas (50) (50) (49) (49) 

Infiltration cellular, lymphocyte 1 (2%) 

Inflammation, chronic 1 (2%) 1 (2%) 

Acinus, atrophy 1 (2%) 2 (4%) 3 (6%) 

Duct, cyst 1 (2%) 2 (4%) 1 (2%) 

Stomach, forestomach (50) (50) (49) (50) 

Cyst 1 (2%) 

Hyperplasia, squamous 1 (2%) 3 (6%) 1 (2%) 3 (6%) 

Inflammation 2 (4%) 

a Number of animals examined microscopically at the site and the number of animals with lesion 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 205 

TABLE D5 

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Cardiovascular System 

Heart (50) (50) (50) (50) 

Mineralization 1 (2%) 

Artery, inflammation, chronic 1 (2%) 1 (2%) 

Epicardium, fibrosis 1 (2%) 

Endocrine System 

Adrenal cortex (50) (50) (49) (50) 

Accessory adrenal cortical nodule 1 (2%) 

Angiectasis 1 (2%) 

Cyst, focal 1 (2%) 

Degeneration, cystic, focal 1 (2%) 

Inflammation 1 (2%) 1 (2%) 

Vacuolization cytoplasmic 1 (2%) 1 (2%) 

Capsule, hyperplasia 1 (2%) 

Pituitary gland (48) (47) (47) (48) 

Angiectasis 2 (4%) 5 (11%) 1 (2%) 2 (4%) 

Pars distalis, angiectasis 1 (2%) 

Pars distalis, hyperplasia 12 (25%) 1 (2%) 7 (15%) 6 (13%) 

Thyroid gland (50) (49) (49) (49) 

Cyst 1 (2%) 

Infiltration cellular, lymphocyte 1 (2%) 

Inflammation 1 (2%) 

Follicular cell, hyperplasia 20 (40%) 17 (35%) 14 (29%) 19 (39%) 

General Body System 

None 

Genital System 

Clitoral gland (41) (40) (43) (40) 

Cyst 1 (2%) 

Inflammation 1 (2%) 1 (3%) 

Ovary (48) (49) (49) (48) 

Angiectasis 1 (2%) 1 (2%) 1 (2%) 

Atrophy 2 (4%) 

Cyst 9 (19%) 12 (24%) 16 (33%) 15 (31%) 

Thrombosis 1 (2%) 

Bilateral, cyst 1 (2%) 

Uterus (50) (50) (49) (49) 

Angiectasis 1 (2%) 1 (2%) 1 (2%) 

Hydrometra 3 (6%) 3 (6%) 2 (4%) 

Hyperplasia, cystic 37 (74%) 41 (82%) 38 (78%) 37 (76%) 

Inflammation 2 (4%) 

Inflammation, suppurative 1 (2%) 

Thrombosis 1 (2%) 1 (2%) 1 (2%) 

Cervix, hemorrhage 1 (2%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

206 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE D5 

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Hematopoietic System 

Bone marrow (50) (50) (49) (50) 

Inflammation, chronic 1 (2%) 

Lymph node (3) (6) (3) (4) 

Hyperplasia, lymphoid 1 (17%) 

Iliac, ectasia 1 (33%) 

Renal, hyperplasia, lymphoid 1 (17%) 

Lymph node, bronchial (30) (34) (27) (35) 

Hyperplasia, lymphoid 1 (3%) 2 (6%) 2 (7%) 3 (9%) 

Infiltration cellular, histiocyte 1 (3%) 

Lymph node, mandibular (37) (37) (36) (36) 

Hyperplasia, lymphoid 1 (3%) 

Lymph node, mesenteric (46) (45) (46) (44) 

Hyperplasia, lymphoid 1 (2%) 5 (11%) 2 (4%) 

Infiltration cellular, histiocyte 1 (2%) 

Inflammation, chronic 1 (2%) 

Necrosis 1 (2%) 

Lymph node, mediastinal (41) (36) (28) (34) 

Hemorrhage 1 (3%) 

Hyperplasia, lymphoid 3 (7%) 5 (14%) 1 (4%) 3 (9%) 

Spleen (50) (50) (49) (49) 

Hematopoietic cell proliferation 3 (6%) 11 (22%) 9 (18%) 3 (6%) 

Hyperplasia, lymphoid 1 (2%) 5 (10%) 1 (2%) 2 (4%) 

Thymus (41) (44) (41) (41) 

Hyperplasia, lymphoid 1 (2%) 1 (2%) 

Necrosis 1 (2%) 

Integumentary System 

Mammary gland (47) (50) (50) (50) 

Hyperplasia 3 (6%) 4 (8%) 2 (4%) 

Skin (49) (50) (50) (50) 

Inflammation, chronic 1 (2%) 

Inflammation, suppurative 1 (2%) 

Ulcer 1 (2%) 1 (2%) 1 (2%) 

Subcutaneous tissue, hemorrhage 1 (2%) 

Subcutaneous tissue, inflammation, chronic 1 (2%) 

Musculoskeletal System 

Bone (50) (50) (50) (50) 

Fibrous osteodystrophy 2 (4%) 1 (2%) 1 (2%) 5 (10%) 

Fracture 1 (2%) 1 (2%) 

Maxilla, inflammation, chronic 1 (2%) 

Skeletal muscle (1) (2) (1) 

Hemorrhage 1 (50%) 

Nervous System 

Brain (50) (50) (50) (50) 

Hemorrhage 1 (2%) 

Spinal cord (1) (1) 

Degeneration 1 (100%) 
 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Cobalt Sulfate Heptahydrate, NTP TR 471 207 

TABLE D5 

Summary of the Incidence of Nonneoplastic Lesions in Female Mice in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

Chamber Control 0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Respiratory System 

Larynx (50) (49) (47) (50) 

Inflammation, chronic 1 (2%) 

Metaplasia, squamous 45 (92%) 40 (85%) 50 (100%) 

Lung (50) (50) (50) (50) 

Hemorrhage 2 (4%) 1 (2%) 4 (8%) 

Infiltration cellular, diffuse, histiocyte 4 (8%) 

Infiltration cellular, focal, histiocyte 2 (4%) 5 (10%) 7 (14%) 10 (20%) 

Inflammation, chronic 1 (2%) 1 (2%) 

Alveolar epithelium, hyperplasia 2 (4%) 3 (6%) 5 (10%) 

Bronchus, vacuolization cytoplasmic 6 (12%) 31 (62%) 43 (86%) 

Capillary, thrombosis, diffuse 1 (2%) 

Nose (50) (50) (49) (48) 

Hemorrhage 1 (2%) 

Inflammation, chronic 1 (2%) 

Inflammation, suppurative 1 (2%) 5 (10%) 4 (8%) 

Olfactory epithelium, atrophy 2 (4%) 12 (24%) 46 (96%) 

Olfactory epithelium, degeneration, hyaline 3 (6%) 1 (2%) 2 (4%) 2 (4%) 

Olfactory epithelium, hyperplasia 30 (63%) 

Olfactory epithelium, metaplasia 1 (2%) 1 (2%) 

Respiratory epithelium, degeneration, hyaline 20 (40%) 16 (32%) 14 (29%) 11 (23%) 

Respiratory epithelium, metaplasia, squamous 4 (8%) 

Special Senses System 

Eye (1) 

Degeneration 1 (100%) 

Urinary System 

Kidney (50) (50) (50) (50) 

Cyst 1 (2%) 2 (4%) 

Infarct 1 (2%) 

Infiltration cellular, mixed cell 3 (6%) 1 (2%) 1 (2%) 3 (6%) 

Inflammation, chronic 1 (2%) 1 (2%) 

Metaplasia, osseous 1 (2%) 1 (2%) 

Mineralization 1 (2%) 

Nephropathy 5 (10%) 6 (12%) 7 (14%) 3 (6%) 

Pigmentation, hemosiderin 1 (2%) 

Urinary bladder (48) (47) (45) (46) 

Edema 1 (2%) 
208 Cobalt Sulfate Heptahydrate, NTP TR 471 
209 

APPENDIX E
 

GENETIC TOXICOLOGY
 

SALMONELLA MUTAGENICITY TEST PROTOCOL ........................................ 210
 

RESULTS 210
 

TABLE E1 Mutagenicity of Cobalt Sulfate Heptahydrate in Salmonella typhimurium ............ 211
 
210 Cobalt Sulfate Heptahydrate, NTP TR 471 

GENETIC TOXICOLOGY 

SALMONELLA MUTAGENICITY TEST PROTOCOL 

Testing was performed as reported by Zeiger et al. (1992). Cobalt sulfate heptahydrate was sent to the 

laboratory as a coded aliquot from Radian Corporation (Austin, TX). It was incubated with the Salmonella 

typhimurium tester strains (TA98, TA100, and TA1535) either in buffer or S9 mix (metabolic activation 

enzymes and cofactors from Aroclor 1254-induced male Sprague-Dawley rat or Syrian hamster liver) for 

20 minutes at 37E C. Top agar supplemented with L-histidine and d-biotin was added, and the contents of 

the tubes were mixed and poured onto the surfaces of minimal glucose agar plates. Histidine-independent 

mutant colonies arising on these plates were counted following incubation for 2 days at 37E C. 

Each trial consisted of triplicate plates of concurrent positive and negative controls and of five doses of 

cobalt sulfate heptahydrate. The high dose was limited by experimental design to 10,000 μg/plate. All 

positive assays were repeated under the conditions that elicited the positive response. 

In this assay, a positive response is defined as a reproducible, dose-related increase in histidine-independent 

(revertant) colonies in any one strain/activation combination. An equivocal response is defined as an 

increase in revertants that is not dose related, not reproducible, or not of sufficient magnitude to support a 

determination of mutagenicity. A negative response is obtained when no increase in revertant colonies is 

observed following chemical treatment. There is no minimum percentage or fold increase required for a 

chemical to be judged positive or weakly positive. 

RESULTS 

Cobalt sulfate heptahydrate (3 to 10,000 μg/mL) was mutagenic in S. typhimurium strain TA100 in the 

absence of S9 metabolic activation, and with 5% hamster or rat liver S9; no mutagenicity was detected in 

strain TA98 or TA1535, with or without S9 (Zeiger et al., 1992; Table E1). 
Cobalt Sulfate Heptahydrate, NTP TR 471 211 

TABLE E1 

Mutagenicity of Cobalt Sulfate Heptahydrate in Salmonella typhimuriuma 

Strain Dose 

(μg/plate) Trial 1 

!S9 

Trial 2 

Revertants/plateb 

Trial 3 

+hamster S9 

5% 5% 

TA100 0 

3 

10 

33 

100 

333 

1,000 

3,333 

10,000 

102 ± 6.4 

201 ± 15.3 

217 ± 7.0 

204 ± 14.7 

126 ± 15.3 

101 ± 10.3c 

113 ± 6.8 

143 ± 4.7 

155 ± 1.7 

152 ± 7.9 

208 ± 11.7 

206 ± 17.6 

107 ± 8.5 

134 ± 6.8 

160 ± 10.2 

163 ± 2.2 

204 ± 9.4 

152 ± 9.0 

117 ± 9.6 

118 ± 5.6 

175 ± 1.5 

188 ± 6.2 

187 ± 6.1 

176 ± 7.3 

98 ± 6.4 

119 ± 11.9 

137 ± 13.7 

162 ± 8.4 

176 ± 14.8 

163 ± 1.5 

Trial summary 

Positive controld 

Positive 

429 ± 7.8 

Weakly Positive 

312 ± 2.9 

Weakly Positive 

290 ± 20.1 

Weakly Positive 

922 ± 44.3 

Weakly Positive 

897 ± 57.6 

+hamster S9 +rat S9 

10% 30% 30% 5% 5% 

TA100 0 

(continued) 10 

33 

100 

333 

1,000 

3,333 

10,000 

130 ± 15.0 

134 ± 9.8 

156 ± 2.8 

187 ± 2.5 

159 ± 5.0 

160 ± 3.1 

139 ± 11.2 

194 ± 9.0 

179 ± 6.0 

176 ± 3.0 

188 ± 18.4 

123 ± 3.8 

159 ± 3.9 

168 ± 15.0 

166 ± 0.7 

193 ± 4.0 

160 ± 14.0 

161 ± 5.8 

116 ± 8.7 

143 ± 12.2 

176 ± 8.0 

189 ± 15.9 

168 ± 8.2 

146 ± 9.8 

117 ± 10.6 

133 ± 1.5 

164 ± 4.7 

188 ± 7.5 

201 ± 1.2 

143 ± 24.2 

Trial summary 

Positive control 

Equivocal 

577 ± 10.9 

Equivocal 

462 ± 31.2 

Negative 

457 ± 22.2 

Weakly Positive 

909 ± 16.2 

Weakly Positive 

1,011 ± 25.0 

+rat S9 

10% 30% 30% 

TA100 0 124 ± 10.7 151 ± 12.3 131 ± 3.0 

(continued) 33 122 ± 4.4 123 ± 14.8 

100 142 ± 5.8 179 ± 16.6 144 ± 10.7 

333 154 ± 4.9 223 ± 6.9 138 ± 8.4 

1,000 133 ± 5.5 191 ± 0.0 137 ± 11.2 

3,333 124 ± 4.3 202 ± 1.9 135 ± 5.3 

10,000 176 ± 12.5 

Trial summary Negative Equivocal Negative 

Positive control 556 ± 32.4 244 ± 6.7 521 ± 41.2 
212 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE E1 

Mutagenicity of Cobalt Sulfate Heptahydrate in Salmonella typhimurium 

Revertants/plate 

Strain Dose !S9 +hamster S9 +rat S9 

(μg/plate) 5% 5% 

TA1535 0 16 ± 1.2 12 ± 3.3 11 ± 1.5 

3 10 ± 2.6 

10 17 ± 2.0 9 ± 2.6 8 ± 1.3 

33 14 ± 2.3 8 ± 0.3 10 ± 1.5 

100 9 ± 1.2 5 ± 0.6 8 ± 0.9 

333 8 ± 1.7 4 ± 1.2 9 ± 0.3 

1,000 9 ± 1.3 8 ± 1.5 

Trial summary Negative Negative Negative 

Positive control 213 ± 24.0 64 ± 4.9 186 ± 35.6 

!S9 +hamster S9 

5% 10% 30% 30% 

TA98 0 

10 

33 

100 

333 

1,000 

3,333 

10,000 

22 ± 2.3 

26 ± 0.5 

25 ± 4.0 

19 ± 0.6 

16 ± 3.4 

5 ± 2.7c 

32 ± 2.2 

36 ± 2.7 

31 ± 4.4 

45 ± 4.1 

40 ± 1.2 

42 ± 2.8 

32 ± 0.6 

28 ± 0.9 

38 ± 2.7 

39 ± 4.6 

43 ± 0.0 

48 ± 7.6 

32 ± 3.8 

44 ± 4.7 

53 ± 4.2 

47 ± 7.3 

44 ± 4.7 

31 ± 3.8 

24 ± 1.7 

20 ± 2.3 

24 ± 1.9 

23 ± 3.5 

34 ± 2.6 

29 ± 4.8 

Trial summary 

Positive control 

Negative 

372 ± 12.5 

Negative 

671 ± 34.6 

Negative 

687 ± 24.7 

Equivocal 

328 ± 4.3 

Negative 

542 ± 38.5 

+rat S9 

5% 10% 30% 30% 30% 

TA98 0 

(continued) 10 

33 

100 

333 

1,000 

3,333 

10,000 

36 ± 1.8 

33 ± 4.9 

42 ± 3.0 

41 ± 4.7 

55 ± 5.6 

55 ± 2.9 

28 ± 1.7 

29 ± 2.1 

38 ± 3.8 

45 ± 7.5 

46 ± 4.5 

33 ± 1.8 

32 ± 3.3 

26 ± 1.2 

26 ± 0.6 

14 ± 0.6c 

16 ± 4.0c 

7 ± 2.6c 

32 ± 4.2 

44 ± 3.5 

54 ± 8.5 

28 ± 1.5 

21 ± 2.0 

24 ± 3.7 

34 ± 1.5 

27 ± 1.2 

24 ± 2.3 

37 ± 3.5 

37 ± 3.3 

34 ± 3.8 

Trial summary 

Positive control 

Negative 

742 ± 14.8 

Negative 

451 ± 12.5 

Negative 

104 ± 5.9 

Negative 

105 ± 6.5 

Negative 

208 ± 15.8

a Study performed at SRI International. The detailed protocol and these data are presented in Zeiger et al. (1992). b Revertants are presented as mean ± standard error from three plates. 

Slight toxicity d The positive controls in the absence of metabolic activation were sodium azide (TA100 and TA1535) and 4-nitro-o-phenylenediamine 

(TA98). The positive control for metabolic activation with all strains was 2-aminoanthracene. 

c
213 

APPENDIX F
 

CHEMICAL CHARACTERIZATION AND
 

GENERATION OF CHAMBER CONCENTRATIONS
 

PROCUREMENT AND CHARACTERIZATION OF COBALT SULFATE HEPTAHYDRATE ................... 214
 

AEROSOL GENERATION AND EXPOSURE SYSTEM ........................................ 214
 

AEROSOL CONCENTRATION MONITORING ............................................ 215
 

CHAMBER ATMOSPHERE CHARACTERIZATION ......................................... 215
 

FIGURE F1 Infrared Absorption Spectrum of Cobalt Sulfate Heptahydrate ................... 218
 

FIGURE F2 Schematic of the Aerosol Generation and Delivery System ...................... 219
 

FIGURE F3 Schematic of the RETEC Compressed-Air-Driven Nebulizer ..................... 220
 

FIGURE F4 Inhalation Suite ................................................... 221
 

FIGURE F5 Schematic of the Concentration Monitoring System ........................... 222
 

TABLE F1 Summary of Chamber Concentrations in the 2-Year Inhalation Studies
 

of Cobalt Sulfate Heptahydrate ........................................ 223
 

TABLE F2 Summary of Aerosol Size Measurements for the Rat Exposure Chambers
 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate ................... 223
 

TABLE F3 Summary of Aerosol Size Measurements for the Mouse Exposure Chambers
 

in the 2-Year Inhalation Study of Cobalt Sulfate Heptahydrate ................... 224
 
214 Cobalt Sulfate Heptahydrate, NTP TR 471 

CHEMICAL CHARACTERIZATION AND 

GENERATION OF CHAMBER CONCENTRATIONS 

PROCUREMENT AND CHARACTERIZATION OF COBALT SULFATE HEPTAHYDRATE 

Cobalt sulfate heptahydrate was obtained from Curtin Matheson Scientific (Kansas City, MO) in one 

lot (412092), which was used during the 2-year studies. Identity and purity analyses were conducted by the 

analytical chemistry laboratory, Midwest Research Institute (Kansas City, MO). Reports on analyses 

performed in support of the cobalt sulfate heptahydrate studies are on file at the National Institute of 

Environmental Health Sciences. 

The chemical, a red, crystalline solid, was identified as cobalt sulfate heptahydrate by infrared, ultraviolet 

and/or visible spectroscopy. The spectra were consistent with the structure of cobalt sulfate heptahydrate. 

No literature references were found. The infrared spectrum is presented in Figure F1. 

The purity of lot 412092 was determined by elemental analysis, Karl Fischer water analysis, and spark 

source mass spectroscopy. Elemental analyses for sulfur and hydrogen were in agreement with the 

theoretical values for cobalt sulfate heptahydrate, but values for cobalt were slightly low. Karl Fischer water 

analysis indicated 44.6% ± 0.5% water. Spark source mass spectroscopy indicated 140 ppm nickel present 

as an impurity; all other impurities had a combined total of less than 175 ppm. The overall purity was 

determined to be approximately 99%. 

Literature references indicate that cobalt sulfate heptahydrate is stable as a bulk chemical when stored 

protected from light at normal temperatures (Merck Index, 1989). The heptahydrate dehydrates to the 

hexahydrate at 41.5E C and to the monohydrate at 71E C, with no further changes expected below the 

decomposition temperature (708E C). Therefore, an accelerated stability study was not conducted. To 

ensure stability, the bulk chemical was stored in its original shipping containers, metal cans, at room 

temperature. Stability was monitored during the 2-year studies using elemental analysis by inductively 

coupled plasma/atomic emission spectroscopy, normalized against a cobalt standard (National Institute of 

Standards and Technology, Gaithersburg, MD); no degradation of the bulk chemical was detected. 

AEROSOL GENERATION AND EXPOSURE SYSTEM 

A diagram of the cobalt sulfate heptahydrate aerosol generation and delivery system is shown in Figure F2. 

Cobalt sulfate heptahydrate aerosol was generated and delivered from an aqueous solution by a system 

composed of three main components: a compressed-air-driven nebulizer, an aerosol charge neutralizer, and 

an aerosol distribution system. 

The nebulizer (Model PN7002; RETEC Development Laboratory, Portland, OR), shown in Figure F3, 

consisted of two orifices of different sizes aligned on opposite sides of a small chamber. Compressed air 

entered the chamber through the small orifice and, on entering the larger orifice, induced a negative 

pressure. Cobalt sulfate heptahydrate in deionized water (approximately 400 g/L) was siphoned from the 

bulk reservoir to the nebulizer reservoir and then aspirated into the nebulizer chamber and expelled as a 

stream through the larger orifice. Shear forces broke the stream into droplets that were evaporated to leave 

dry particles of cobalt sulfate heptahydrate. The aerosol generation and exposure system included primary 

and secondary compressed-air-driven nebulizers. The mass concentration of the dry particles in the feed 

solution was used to determine aerosol particle size. The generator output was controlled by adjusting the 

compressed air pressure. 
Cobalt Sulfate Heptahydrate, NTP TR 471 215 

The aerosol generated by the compressed-air-driven nebulizer was passed through the aerosol charge 

neutralizer to remove static charge that formed on the aerosol particles during generation, reducing adhesion 

of the droplets to the walls of the delivery system. This neutralizer consisted of a length of plastic duct with 

two 10-mCi 63Ni-plated foils suspended in the center of the tube. The activity of the foils was matched to the 

diameter of the duct to allow adequate time for the aerosol to approach Boltzmann equilibrium at the system 

flow rate. 

3 A distribution line carried aerosol (20 mg/m ) to exposure chambers on both sides of the exposure room. 

Aerosol was siphoned from the branches of the distribution line by pneumatic pumps (one pump per 

exposure chamber). The flow rate in each branch of the distribution line was controlled by an Air-Vac pump 

(Air-Vac Engineering, Milford, CT) and monitored by a photohelic differential pressure gauge (Dwyer 

Instruments, Inc., Michigan City, IN) coupled to a Venturi tube. At each chamber, aerosol moving through 

the chamber inlet was further diluted with HEPA-filtered air to the appropriate concentration for the 

chamber. A diagram of the inhalation suite is shown in Figure F4. The Hazleton 2000 inhalation exposure 

chambers (Harford Systems Division of Lab Products, Inc., Aberdeen, MD) were designed so that uniform 

aerosol concentrations could be maintained throughout each chamber with the catch pans in place. The total 3 active mixing volume of each chamber was 1.7 m . 

AEROSOL CONCENTRATION MONITORING 

The chamber aerosol concentrations of cobalt sulfate heptahydrate were monitored by real-time aerosol 

monitors (Model RAM-1; MIE, Inc., Bedford, MA) controlled by a Hewlett-Packard HP-85B computer 

(Hewlett-Packard Company, Palo Alto, CA). The RAM-1s detected aerosol particles ranging from 0.1 to 

20 μm in diameter. Three RAM-1s were employed in the monitoring system (Figure F5); these monitors 

were exchanged with different RAM-1s when the on-line monitor performance deteriorated. Chamber 

aerosol concentrations were sampled at least once per hour during each exposure day. Sample lines 

connecting the exposure chambers to the RAM-1s were designed to minimize aerosol particle losses due to 

settling or impaction. Throughout the 2-year studies, the background concentrations of total suspended 

particles in the control chambers were less than the limit of detection. A summary of chamber concentrations 

is presented in Table F1. 

The RAM-1 voltage output was calibrated against cobalt sulfate heptahydrate concentrations of chamber filter 

samples. Samples were collected on Teflon®-coated, glass-fiber filters with a calibrated flow sampler. 

Equations for the calibration curves contained in the HP-85B computer converted the RAM-1 voltages into 

exposure concentrations. Solutions of filter samples in 2% nitric acid were analyzed quantitatively for cobalt 

sulfate heptahydrate by inductively coupled plasma/atomic emission spectroscopy (ICP/AES). Calibration 

samples were collected every 2 weeks. Additional samples for monitoring the accuracy of calibration were 

collected daily from at least one chamber monitored by each RAM-1 and were analyzed two to three times 

per week. The ICP/AES was calibrated with a solution of standard cobalt diluted with nitric acid. 

3 The stability of aerosol concentrations in the 0.3 and 3.0 mg/m chambers was monitored by analyzing

samples collected on Gelman A/E glass fibers using a calibrated flow sampler. X-ray diffraction analyses 

were performed by a Philips 3600 diffraction unit with Cu Ka radiation. Results indicated that cobalt sulfate 

hexahydrate was the primary species delivered to the chambers. 

CHAMBER ATMOSPHERE CHARACTERIZATION 

The time required for the chamber concentration to reach 90% of the target value following the beginning of 

exposure (T 90 ) and the time required for the chamber concentration to reach 10% of the target value

following termination of the exposure (T ) were determined for each exposure chamber. Without animals 10 

present, T90 values ranged from 9 to 11 minutes for the rat chambers and from 7 to 12 minutes for mouse 
 

216 Cobalt Sulfate Heptahydrate, NTP TR 471 

chambers; T10 ranged from 8 to 9 minutes for rats and mice. With animals present, T90 values ranged from 

11 to 16 minutes for rats and from 8 to 12 minutes for mice; T10 ranged from 12 to 13 minutes for rats and 

from 11 to 12 minutes for mice. Variations in these values were considered to be due to differences in 

discrete sampling times, different flow rates for each chamber, fluctuations in generator output, and differing 

transit times for the aerosol through the delivery system. A T90 of 12 minutes was selected for the 2-year 

studies. 

Aerosol size distribution was determined monthly for each exposure chamber with a Mercer-style seven-stage 

impactor (In-Tox Products, Albuquerque, NM). Samples were collected on glass coverslips sprayed with 

silicone and were analyzed for cobalt sulfate heptahydrate with ICP/AES. The relative mass on each 

impactor stage was analyzed by probit analysis; the mass median aerodynamic diameter for the aerosol was 

within the specified range of 1 to 3 μm (Tables F2 and F3). 

The uniformity of aerosol concentration in the inhalation exposure chambers was measured approximately 

every 3 months. Aerosol concentration was determined with the RAM-1s, with an extension tube fitted to 

the sampling lines to allow sampling from ports in the front and back of each chamber. Chamber 

concentration uniformity was acceptable throughout the studies except for measurements taken from the 3 0.3 mg/m mouse exposure chamber during 1 month of the study; however, these measurements were within

the specified 5% variability when repeated. 

At the beginning of the studies and approximately every 90 days thereafter, the persistence of cobalt sulfate 3 heptahydrate aerosol in the 3.0 mg/m rat chamber with and without animals present was determined by

monitoring the concentration overnight with two RAM-1s. The average time for the concentration to 

decrease to 1% of the target concentration was approximately 20 minutes. 

Before the exposures began, a solution of cobalt sulfate heptahydrate was analyzed for purity by ICP/AES; 

mean concentration of cobalt sulfate heptahydrate was found to be 99% of the theoretical value. Another 

sample from this original solution was analyzed after approximately 10 weeks, and the concentration of 

cobalt sulfate heptahydrate was 103% of the theoretical value. Thus, cobalt sulfate heptahydrate in the 

generator reservoir solution was considered to be stable for up to 10 weeks. New formulations were 

prepared at approximately 8-week intervals thereafter. Because the purity information supplied by the 

manufacturer and determined by the analytical chemistry laboratory indicated possible sulfuric acid 

contamination from the manufacturing process, the pH of the cobalt sulfate heptahydrate solution in the 

generator reservoir was analyzed to determine the extent of contamination. The pH was approximately 4.5, 

compared to 6.0 for the deionized water used to prepare the solution. From the pH of 4.5, the sulfuric acid 

content of the solution was calculated to be 0.0004% by weight. This value corresponded to a molar 

concentration of 1.6 × 10!5 sulfuric acid. 

The aerosol stoichiometry was determined by measuring the number of moles of cobalt, sulfate, and water 

associated with samples from the chambers and distribution line. Filters were obtained from the 3.0 mg/m3 

rat and mouse chambers and from the distribution line. The total mass of collected aerosol was determined 

by ICP/AES. The mass of water on each filter was determined gravimetrically, and the masses of cobalt and 

sulfate on each filter were determined as the difference between the net aerosol mass and the combined 

masses of cobalt and sulfate. The results indicated that the average number of moles of cobalt, sulfate, and 

water associated with each mole of aerosol were 1.00 ± 0.00, 1.01 ± 0.01, and 5.9 ± 0.8, respectively. 

These results show that aerosol delivered to the exposure chambers is primarily cobalt sulfate hexahydrate, 

which is in good agreement with results obtained by X-ray diffraction. 

3 Samples from the occupied 0.3 and 3.0 mg/m rat exposure chambers, distribution line, and the generator

reservoir were analyzed for ammonia by an Orion Model 512 ammonia electrode (Orion Research, Beverly, 

MA) with an internal reference. The electrode was calibrated against gravimetrically prepared ammonium 
 

Cobalt Sulfate Heptahydrate, NTP TR 471 217 

chloride solutions ranging from 0.1 to 100 mg ammonia per liter. Samples were collected on Teflon®-coated 

glass fibers with calibrated flow samplers. Filters were extracted with deionized water, and cobalt in the 

samples was quantified by ICP/AES. The filters were then extracted in an ionic strength adjustment buffer 

and ammonia in the sample was quantified with the ammonia-selective electrode. The concentration of 

ammonia relative to the amount of cobalt sulfate heptahydrate determined stoichiometrically from the 3 ICP/AES cobalt measurements was approximately 0.9% by weight in the 0.3 mg/m exposure chamber, 3 approximately 1.8% in the 3.0 mg/m chamber, and below detection limits in the distribution line and

generator reservoir. The ammonia values in the exposure chambers were slightly above allowable impurity 

concentrations; this was attributed to the absence of cageboards during the first 8 weeks of the study. 

Thereafter, cageboards were used and the ammonia concentrations were expected to be significantly lower. 

Due to the possibility of contamination of aerosol with carbon eroded from the organic polymers found in the 

components of the generation and delivery system, samples from the occupied chambers, distribution line, 

and reservoir were analyzed for carbon. Samples were collected from the occupied and unoccupied 0.3 and 3 3.0 mg/m mouse exposure chambers, the distribution line, and the generator reservoir on Gelman A/E glass

fiber filters using calibrated flow samplers. The filters were extracted with deionized water and aliquots 

from the extracts were analyzed for cobalt by ICP/AES. The remainder of the extract was made acidic to 

remove carbonate, and the extract was analyzed for total organic carbon by a Dohrmann Carbon Analyzer 

System (Dohrmann Division, Xertex Corporation, Santa Clara, CA). The instrument was calibrated against 

gravimetric standards prepared from potassium hydrogen phthalate. 

The concentrations of total organic carbon in samples from the occupied exposure chamber were 8% in the 3 3 0.3 mg/m chamber and 2.1% in the 3.0 mg/m chamber. Carbon concentrations in other parts of the 

generation and delivery system and from empty chambers before exposure began were less than 0.5%. The 

high levels of organic carbon detected in the occupied mouse chambers were considered to be due to the 

presence of animals in the chambers. To verify that possibility, the analysis was repeated with samples from 3 the occupied 0.3 and 3.0 mg/m rat exposure chambers, the distribution line, and the generator reservoir. 3 The results (9% and 2.1% total organic carbon in the 0.3 and 3.0 mg/m rat chambers, respectively) were

similar to those found for the mouse exposure chamber analysis, with negligible amounts again found in the 

generation and delivery system components. Thus, the carbon was concluded to be derived from the animals 

rather than from contamination of the aerosol. 
218 Cobalt Sulfate Heptahydrate, NTP TR 471
 Micrometers 

FIGURE F1 

Infrared Absorption Spectrum of Cobalt Sulfate Heptahydrate 
Cobalt Sulfate Heptahydrate, NTP TR 471
 219 

FIGURE F2 

Schematic of the Aerosol Generation and Delivery System 
220 Cobalt Sulfate Heptahydrate, NTP TR 471
 

FIGURE F3 

Schematic of the RETEC Compressed-Air-Driven Nebulizer 
Cobalt Sulfate Heptahydrate, NTP TR 471
 221 

FIGURE F4 

Inhalation Suite 
222 Cobalt Sulfate Heptahydrate, NTP TR 471
 

FIGURE F5 

Schematic of the Concentration Monitoring System 
Cobalt Sulfate Heptahydrate, NTP TR 471 223 

TABLE F1 

Summary of Chamber Concentrations in the 2-Year Inhalation Studies of Cobalt Sulfate Heptahydrate 

Target Concentration Total Number of Readings Average Concentrationa 

3 3 (mg/m ) (mg/m ) 

Rat Chambers 

0.3 4,574 0.31 ± 0.03 

1.0 4,574 1.03 ± 0.10 

3.0 4,580 2.98 ± 0.20 

Mouse Chambers 

0.3 4,571 0.30 ± 0.04 

1.0 4,609 1.02 ± 0.08 

3.0 4,605 3.01 ± 0.19

a Mean ± standard deviation 

TABLE F2 

Summary of Aerosol Size Measurements for the Rat Exposure Chambers in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Mass Median Geometric Mass Median Geometric Mass Median Geometric 

Aerodynamic Standard Aerodynamic Standard Aerodynamic Standard 

Date Diameter (μm) Deviation Diameter (μm) Deviation Diameter (μm) Deviation 

September 1990 1.7 2.4 1.2 2.2 1.6 2.2 

October 1990 1.5 2.0 1.2 2.1 1.6 2.1 

November 1990 1.6 2.1 1.3 2.2 1.6 2.1 

December 1990 1.3 2.1 1.1 2.0 1.1 1.9 

January 1991 1.5 2.1 1.2 2.1 1.6 2.0 

February 1991 1.5 2.2 1.4 2.2 1.5 2.1 

March 1991 1.5 2.2 1.4 2.2 1.6 2.1 

April 1991 1.6 2.1 1.4 2.2 1.6 2.1 

May 1991 1.5 2.0 1.4 2.0 1.7 2.0 

June 1991 1.6 2.1 1.5 2.3 1.7 2.1 

July 1991 1.6 2.1 1.3 2.0 1.6 2.1 

August 1991 1.5 2.0 1.4 1.9 1.7 1.9 

September 1991 1.5 2.1 1.4 1.9 1.6 2.1 

October 1991 1.5 2.2 1.5 2.0 1.7 2.3 

November 1991 1.5 2.6 1.6 2.2 1.8 2.4 

December 1991 1.3 2.2 1.5 2.0 1.7 2.3 

January 1992 1.5 2.1 1.5 2.2 1.7 2.2 

February 1992 1.4 2.3 1.4 2.3 1.6 2.3 

March 1992 1.3 2.3 1.4 2.3 1.6 2.3 

April 1992 1.6 1.9 1.5 2.3 1.6 2.3 

May 1992 1.4 2.2 1.5 2.4 1.6 2.3 

June 1992 1.4 2.2 1.5 2.1 1.6 2.2 

July 1992 1.6 2.2 1.4 2.1 1.6 2.1 

August 1992 1.5 2.2 1.3 2.2 1.6 2.2 

Mean ± standard 

deviation 1.5 ± 0.10 2.2 ± 0.14 1.4 ± 0.12 2.1 ± 0.13 1.6 ± 0.12 2.2 ± 0.13 

TABLE F3 
224 Cobalt Sulfate Heptahydrate, NTP TR 471 

Summary of Aerosol Size Measurements for the Mouse Exposure Chambers in the 2-Year Inhalation Study 

of Cobalt Sulfate Heptahydrate 

0.3 mg/m3 1.0 mg/m3 3.0 mg/m3 

Mass Median Geometric Mass Median Geometric Mass Median Geometric 

Aerodynamic Standard Aerodynamic Standard Aerodynamic Standard 

Date Diameter (μm) Deviation Diameter (μm) Deviation Diameter (μm) Deviation 

September 1990 1.3 3.0 1.3 2.6 1.5 2.4 

October 1990 1.6 2.6 1.7 2.2 1.9 2.1 

November 1990 1.4 2.4 1.6 2.5 1.7 2.3 

December 1990 1.2 2.4 —a —a 1.1 2.2 

January 1991 1.4 2.4 1.6 2.3 1.6 2.2 

February 1991 1.6 2.3 1.5 2.3 1.7 2.2 

March 1991 1.5 2.3 1.5 2.3 1.6 2.1 

April 1991 1.6 2.4 1.5 2.4 1.8 2.3 

May 1991 1.6 2.2 1.6 2.2 1.8 2.1 

June 1991 1.4 2.3 1.4 2.4 1.6 2.2 

July 1991 1.6 2.3 1.4 2.1 1.7 2.1 

August 1991 1.6 2.1 1.5 2.1 1.6 2.2 

September 1991 1.6 2.2 1.5 2.3 1.7 2.2 

October 1991 1.7 2.2 1.7 2.3 1.7 2.3 

November 1991 1.7 2.4 1.6 2.4 1.8 2.2 

December 1991 1.7 2.2 1.6 2.1 1.7 2.3 

January 1992 1.7 2.3 1.5 2.4 1.6 2.3 

February 1992 1.7 2.2 1.7 2.4 1.6 2.3 

March 1992 1.7 2.4 1.4 2.4 1.5 2.3 

April 1992 1.7 2.5 1.6 2.5 1.5 2.6 

May 1992 1.6 2.1 1.7 2.2 1.6 2.5 

June 1992 1.5 2.2 1.5 2.3 1.6 2.2 

July 1992 1.6 2.3 1.3 2.3 1.6 2.2 

August 1992 2.0 2.3 1.5 2.3 1.5 2.2 

Mean ± standard 

deviation 1.6 ± 0.16 2.3 ± 0.19 1.5 ± 0.12 2.3 ± 0.13 1.6 ± 0.15 2.3 ± 0.12 

a No data available due to incomplete stage analysis 
225 

APPENDIX G 

INGREDIENTS, NUTRIENT COMPOSITION, 

AND CONTAMINANT LEVELS 

IN NIH-07 RAT AND MOUSE RATION 

TABLE G1 

TABLE G2 

TABLE G3 

TABLE G4 

Ingredients of NIH-07 Rat and Mouse Ration .............................. 

Vitamins and Minerals in NIH-07 Rat and Mouse Ration ...................... 

Nutrient Composition of NIH-07 Rat and Mouse Ration ...................... 

Contaminant Levels in NIH-07 Rat and Mouse Ration ........................ 

226 

226 

227 

228 
226 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE G1 

Ingredients of NIH-07 Rat and Mouse Rationa 

Ingredientsb Percent by Weight 

Ground #2 yellow shelled corn 24.50 

Ground hard winter wheat 23.00 

Soybean meal (49% protein) 12.00 

Fish meal (60% protein) 10.00 

Wheat middlings 10.00 

Dried skim milk 5.00 

Alfalfa meal (dehydrated, 17% protein) 4.00 

Corn gluten meal (60% protein) 3.00 

Soy oil 2.50 

Dried brewer’s yeast 2.00 

Dry molasses 1.50 

Dicalcium phosphate 1.25 

Ground limestone 0.50 

Salt 0.50 

Premixes (vitamin and mineral) 0.25 

a NCI, 1976; NIH, 1978 b Ingredients were ground to pass through a U.S. Standard Screen No. 16 before being mixed. 

TABLE G2 

Vitamins and Minerals in NIH-07 Rat and Mouse Rationa 

Amount Source 

Vitamins 

A 5,500,000 IU Stabilized vitamin A palmitate or acetate 

D3 4,600,000 IU D-activated animal sterol 

K3 

d-"-Tocopheryl acetate 

2.8 g 

20,000 IU 

Menadione 

Choline 560.0 g Choline chloride 

Folic acid 2.2 g 

Niacin 30.0 g 

d-Pantothenic acid 18.0 g d-Calcium pantothenate 

Riboflavin 3.4 g 

Thiamine 10.0 g Thiamine mononitrate 

B12 4,000 μg 

Pyridoxine 1.7 g Pyridoxine hydrochloride 

Biotin 140.0 mg d-Biotin 

Minerals 

Iron 120.0 g Iron sulfate 

Manganese 60.0 g Manganous oxide 

Zinc 16.0 g Zinc oxide 

Copper 4.0 g Copper sulfate 

Iodine 1.4 g Calcium iodate 

Cobalt 0.4 g Cobalt carbonate 

a Per ton (2,000 lb) of finished product 
Cobalt Sulfate Heptahydrate, NTP TR 471 227 

TABLE G3 

Nutrient Composition of NIH-07 Rat and Mouse Ration 

Mean ± Standard 

Nutrient Deviation Range Number of Samples 

Protein (% by weight) 23.40 ± 0.56 22.2 – 24.3 25 

Crude fat (% by weight) 5.31 ± 0.19 5.00 – 5.90 25 

Crude fiber (% by weight) 3.36 ± 0.33 2.60 – 4.30 25 

Ash (% by weight) 6.43 ± 0.20 6.12 – 6.81 25 

Amino Acids (% of total diet) 

Arginine 1.280 ± 0.083 1.110 – 1.390 11 

Cystine 0.308 ± 0.071 0.181 – 0.400 11 

Glycine 1.158 ± 0.048 1.060 – 1.220 11 

Histidine 0.584 ± 0.027 0.531 – 0.630 11 

Isoleucine 0.917 ± 0.033 0.867 – 0.965 11 

Leucine 1.975 ± 0.051 1.850 – 2.040 11 

Lysine 1.274 ± 0.049 1.200 – 1.370 11 

Methionine 0.437 ± 0.109 0.306 – 0.699 11 

Phenylalanine 0.999 ± 0.120 0.665 – 1.110 11 

Threonine 0.904 ± 0.058 0.824 – 0.985 11 

Tryptophan 0.218 ± 0.153 0.107 – 0.671 11 

Tyrosine 0.685 ± 0.094 0.564 – 0.794 11 

Valine 1.086 ± 0.055 0.962 – 1.170 11 

Essential Fatty Acids (% of total diet) 

Linoleic 2.407 ± 0.227 1.830 – 2.570 10 

Linolenic 0.259 ± 0.065 0.100 – 0.320 10 

Vitamins 

Vitamin A (IU/kg) 6,738 ± 1,318 5,730 – 11,450 25 

Vitamin D (IU/kg) 4,450 ± 1,382 3,000 – 6,300 4 

"-Tocopherol (ppm) 35.43 ± 8.98 22.5 – 48.9 11 

Thiamine (ppm) 17.48 ± 2.10 14.0 – 22.0 25 

Riboflavin (ppm) 7.83 ± 0.923 6.10 – 9.00 11 

Niacin (ppm) 99.22 ± 24.27 65.0 – 150.0 11 

Pantothenic acid (ppm) 30.55 ± 3.52 23.0 – 34.6 11 

Pyridoxine (ppm) 9.11 ± 2.53 5.60 – 14.0 11 

Folic acid (ppm) 2.46 ± 0.63 1.80 – 3.70 11 

Biotin (ppm) 0.268 ± 0.047 0.190 – 0.354 11 

Vitamin B12 (ppb) 40.5 ± 19.1 10.6 – 65.0 11 

Choline (ppm) 2,991 ± 382 2,300 – 3,430 10 

Minerals 

Calcium (%) 1.16 ± 0.10 1.00 – 1.49 25 

Phosphorus (%) 0.92 ± 0.05 0.76 – 1.00 25 

Potassium (%) 0.886 ± 0.063 0.772 – 0.971 9 

Chloride (%) 0.529 ± 0.087 0.380 – 0.635 9 

Sodium (%) 0.316 ± 0.033 0.258 – 0.371 11 

Magnesium (%) 0.166 ± 0.010 0.148 – 0.181 11 

Sulfur (%) 0.272 ± 0.059 0.208 – 0.420 10 

Iron (ppm) 350.5 ± 87.3 255.0 – 523.0 11 

Manganese (ppm) 92.48 ± 5.14 81.7 – 99.4 11 

Zinc (ppm) 59.33 ± 10.2 46.1 – 81.6 11 

Copper (ppm) 11.81 ± 2.50 9.09 – 15.4 11 

Iodine (ppm) 3.54 ± 1.19 1.52 – 5.83 10 

Chromium (ppm) 1.66 ± 0.46 0.85 – 2.09 11 

Cobalt (ppm) 0.76 ± 0.23 0.49 – 1.15 7 
228 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE G4 

Contaminant Levels in NIH-07 Rat and Mouse Rationa 

Mean ± Standard

 Deviationb Range Number of Samples 

Contaminants 

Arsenic (ppm) 0.42 ± 0.20 0.10 – 0.70 25 

Cadmium (ppm) 0.13 ± 0.07 0.04 – 0.20 25 

Lead (ppm) 0.35 ± 0.24 0.10 – 1.00 25 c Mercury (ppm) 0.02 0.02 – 0.03 25 

Selenium (ppm) 0.33 ± 0.11 0.05 – 0.40 25 

Aflatoxins (ppm) <5.0 25 

Nitrate nitrogen (ppm)d 8.99 ± 4.49 2.90 – 17.0 25 

Nitrite nitrogen (ppm)d 0.15 ± 0.08 0.10 – 0.40 25 e BHA (ppm)	 1.80 ± 1.94 1.00 – 10.0 25 e BHT (ppm) 1.56 ± 1.58 1.00 – 8.00 25 

Aerobic plate count (CFU/g) 95,908 ± 162,569 4,100 – 710,000 25 

Coliform (MPN/g) 3 ± 0.3 3 – 4 25 

Escherichia coli (MPN/g) <3 25 

Salmonella (MPN/g) Negative 25 

Total nitrosoamines (ppb)f 7.36 ± 1.75 4.70 – 11.40 25 

N-Nitrosodimethylamine (ppb)f 5.40 ± 1.18 2.90 – 8.20 25 

N-Nitrosopyrrolidine (ppb)f 1.96 ± 1.05 1.00 – 4.30 25 

Pesticides (ppm) 

"-BHC <0.01 25 

$-BHC <0.02 25 

(-BHC <0.01 25 

*-BHC <0.01 25 

Heptachlor <0.01 25 

Aldrin <0.01 25 

Heptachlor epoxide <0.01 25 

DDE <0.01 25 

DDD <0.01 25 

DDT <0.01 25 

HCB <0.01 25 

Mirex <0.01 25 

Methoxychlor <0.05 25 

Dieldrin <0.01 25 

Endrin <0.01 25 

Telodrin <0.01 25 

Chlordane <0.05 25 

Toxaphene <0.10 25 

Estimated PCBs <0.20 25 

Ronnel <0.01 25 

Ethion <0.02 25 

Trithion <0.05 25 

Diazinon <0.10 25 

Methyl parathion <0.02 25 

Ethyl parathion <0.02 25 

Malathion 0.23 ± 0.23 0.05 – 0.97 25 

Endosulfan I <0.01 25 

Endosulfan II <0.01 25 

Endosulfan sulfate <0.03 25 

a CFU ù colony forming units; MPN ù most probable number; BHC ù hexachlorocyclohexane or benzene hexachloride b	 For values less than the limit of detection, the detection limit is given as the mean. 

All values except for the September, November, and December 1991 milling dates (0.03 ppm) were less than the detection limit. The 

detection limit is given as the mean. d	 Sources of contamination: alfalfa, grains, and fish meal e Sources of contamination: soy oil and fish meal f All values were corrected for percent recovery. 

c 
229 

APPENDIX H
 

SENTINEL ANIMAL PROGRAM
 

METHODS ............................................................... 230 

TABLE H1 Murine Virus Antibody Determinations for Rats and Mice 

in the 2-Year Studies of Cobalt Sulfate Heptahydrate ........................ 232 
230 Cobalt Sulfate Hepathydrate, NTP TR 471 

SENTINEL ANIMAL PROGRAM
 

METHODS 

Rodents used in the Carcinogenesis Program of the National Toxicology Program are produced in optimally 

clean facilities to eliminate potential pathogens that may affect study results. The Sentinel Animal Program is 

part of the periodic monitoring of animal health that occurs during the toxicologic evaluation of chemical 

compounds. Under this program, the disease state of the rodents is monitored via serology on sera from 

extra (sentinel) animals in the study rooms. These animals and the study animals are subject to identical 

environmental conditions. The sentinel animals come from the same production source and weanling groups 

as the animals used for the studies of chemical compounds. 

Serum samples were collected from randomly selected rats and mice during the 2-year studies of cobalt 

sulfate heptahydrate. Blood from each animal was collected and allowed to clot, and the serum was 

separated. The samples were processed appropriately and sent to Microbiological Associates, Inc. (Bethesda, 

MD), for determination of antibody titers. The laboratory serology methods and viral agents for which 

testing was performed are tabulated below; the times at which blood was collected during the studies are also 

listed. 

Method and Test Time of Analysis 

RATS 

ELISA 

Mycoplasma arthritidis 

Mycoplasma pulmonis 

PVM (pneumonia virus of mice) 

RCV/SDA (rat coronavirus/

 sialodacryoadenitis virus) 

Sendai 

Study termination 

Study termination 

6, 12, and 18 months, study termination 

6, 12, and 18 months, study termination 

6, 12, and 18 months, study termination 

Immunofluorescence Assay 

RCV/SDA 6 months 

Hemagglutination Inhibition 

H-1 (Toolan's H-1 virus) 

KRV (Kilham rat virus) 

6, 12, and 18 months, study termination 

6, 12, and 18 months, study termination 
Cobalt Sulfate Heptahydrate, NTP TR 471 231 

MICE 

ELISA 

Ectromelia virus 

EDIM (epizootic diarrhea of infant mice) 

GDVII (mouse encephalomyelitis virus) 

LCM (lymphocytic choriomeningitis virus) 

Mouse adenoma virus-FL 

MHV (mouse hepatitis virus) 

M. arthritidis 

M. pulmonis 

PVM
 

Reovirus 3
 

Sendai
 

Immunofluorescence Assay 

EDIM 

LCM 

MHV 

Reovirus 3 

Hemagglutination Inhibition 

K (papovavirus) 

MVM (minute virus of mice) 

Polyoma virus 

Results of serology tests are presented in Table H1. 

6, 12, and 18 months, study termination 

6, 12, and 18 months, study termination 

6, 12, and 18 months, study termination 

6, 12, and 18 months, study termination 

6, 12, and 18 months, study termination 

6, 12, and 18 months, study termination 

Study termination 

Study termination 

6, 12, and 18 months, study termination 

6, 12, and 18 months, study termination 

6, 12, and 18 months, study termination 

18 months and study termination 

6 months 

Study termination 

18 months and study termination 

6, 12, and 18 months, study termination 

6, 12, and 18 months, study termination 

6, 12, and 18 months, study termination 
232 Cobalt Sulfate Hepathydrate, NTP TR 471 

TABLE H1 

Murine Virus Antibody Determinations for Rats and Mice in the 2-Year Studies 

of Cobalt Sulfate Heptahydrate 

Interval Incidence of Antibody 

in Sentinel Animals 

Positive Serologic 

Reaction for 

Rats 

6 Months 

12 Months 

18 Months 

Study termination 

0/16 

0/16 

0/16 

6/10 

None positive 

None positive 

None positive 

M. arthriditisa 

Mice 

6 Months 

12 Months 

18 Months 

Study termination 

0/10 

0/9 

0/10 

3/10 

None positive 

None positive 

None positive 

M. arthriditis 

a Further evaluation of samples positive for M. arthritidis by immunoblot and Western blot procedures indicated that the positive titers may 

have been due to cross reaction with antibodies of nonpathogenic Mycoplasma or other agents. There were no clinical findings or 

histopathologic changes of M. arthritidis infection in animals with positive titers. Accordingly, M. arthritidis-positive titers were 

considered false positives. 
233 

APPENDIX I
 

K-RAS MUTATION FREQUENCY AND SPECTRA
 

IN LUNG NEOPLASMS FROM B6C3F1 MICE 
 

EXPOSED TO COBALT SULFATE HEPTAHYDRATE
 

FOR 2 YEARS
 

Robert C. Sills, Jennifer Neil, H. Lily Hong, John R. Bucher,
 

Gary A. Boorman, and Theodora R. Devereux
 

National Institute of Environmental Health Sciences
 

Research Triangle Park, North Carolina
 

INTRODUCTION ............................................................ 234
 

MATERIALS AND METHODS .................................................... 234
 

RESULTS ................................................................ 235
 

DISCUSSION .............................................................. 235
 

REFERENCES 237
 

TABLE I1 K-ras Mutations in Lung Neoplasms of B6C3F1
 

 Mice ......................... 239

TABLE I2 K-ras Mutation Profile in Lung Neoplasms of B6C3F1
  Mice .................... 239
234 Cobalt Sulfate Heptahydrate, NTP TR 471 

K-RAS MUTATION FREQUENCY AND SPECTRA 

IN LUNG NEOPLASMS FROM B6C3F MICE 1 

EXPOSED TO COBALT SULFATE HEPTAHYDRATE 

FOR 2 YEARS 

INTRODUCTION 

Lung neoplasms occur in B6C3F mice with a typical incidence of 20% in control males and 10% in control 1 

females by 2 years of age. Molecular analysis of lung neoplasms for genetic alterations in cancer genes such 

as the ras proto-oncogene provides additional mechanistic information to help distinguish spontaneous 

neoplasms from chemical-induced neoplasms. For example, chemical-induced neoplasms in mice may have a 

higher frequency of proto-oncogene activation, particularly by point mutations in codon 12, 13, or 61 of 

K-ras genes (Sills et al., 1995). The frequency of ras activation in these neoplasms is often greater than that 

detected in neoplasms occurring in control animals (Devereux et al., 1991), and there is evidence for 

chemical specificity in the pattern of mutations. The specific types of oncogene-activating mutations induced 

by a chemical carcinogen often agree with what is expected based on the DNA adducts formed by the agent 

(Devereux et al., 1993a). Even for “nongenotoxic carcinogens,” patterns of ras gene mutations in 

neoplasms can give clues about the mechanism of tumorigenesis (Devereux et al., 1993b). 

MATERIALS AND METHODS 

3 Lung neoplasms: Male and female B6C3F mice were exposed to 0, 0.3, 1.0, or 3.0 mg/m cobalt sulfate 1 

heptahydrate by inhalation for 6 hours per day, 5 days per week for 2 years. At necropsy, lung neoplasms 

were fixed in 10% neutral buffered formalin, routinely processed, embedded in paraffin, sectioned to a 

thickness of 5 μm, and stained with hematoxylin and eosin. Subsequently, six unstained serial sections 

(10 μm thick) were prepared from paraffin blocks containing alveolar/bronchiolar adenomas or carcinomas 

for isolation of DNA for polymerase chain reaction (PCR)-based assays. In order to isolate adequate 

amounts of DNA, lung neoplasms greater than 1 mm in diameter were identified for analysis. A total of 

32 paraffin-embedded neoplasms were examined for genetic alterations in the K-ras gene. This included 

26 neoplasms from cobalt sulfate heptahydrate-exposed mice and six neoplasms from control mice. 

DNA isolation: The DNA isolation procedure is described in Marmur (1961) and Sills et al. (1995). The 

paraffin-embedded tissue was deparaffinized and rehydrated before digesting with proteinase K (Wright and 

Manos, 1990). The neoplasm tissue was digested with 10 mg/mL pronase in 1% sodium dodecyl sulfate in 

TNE buffer (10 mM Tris; 150 mM NaCl; and 2 mM EDTA disodium salt, pH 7.5). DNA was extracted 

with phenol and chloroform and precipitated with ethanol. DNA was quantified by optical density at 

260 nm, and 200 ng/μL was used for amplification. 

DNA amplification: DNA was amplified by PCR (Saiki et al., 1988; Sills et al., 1995); details of the use of 

nested primers are described in Devereux et al. (1991, 1993b). 

Restriction fragment length polymorphic identification: For identification of K-ras mutations at codon 61, 

restriction fragment length polymorphism (RFLP) was used, and most of exon 2 surrounding codon 61 was 

amplified (Sukumuar and Barbacid, 1990). The sense primer used for amplification of exon 2 was 

5N-GACATCTTAGACACAGCAGTT-3N. A restriction site for XbaI or TaqI enzyme (New England 

Biolaboratory, Beverly, MA) is created by the presence of an A to T or A to G mutation in the second base 

of codon 61. By using this technique, codon 61 CTA and CGA mutations were detected by XbaI and TaqI 

digestion, respectively; the normal sequence (CAA) of codon 61 is not cut by these enzymes. The reaction 
Cobalt Sulfate Heptahydrate, NTP TR 471 235 

was incubated at 37E C (XbaI) or 60E C (TaqI) for 2 hours. Fifteen μL of the mixture with bromophenol 

blue dye was loaded onto the 6% acrylamide tris-borate-EDTA (TBE) gel (8 × 8 cm × 1 mm; 15 wells) 

(Novex, San Diego, CA). The gel was run at 100 volts for 1 hour on the Novex gel electrophoresis unit. 

Gels were stained with a 5 μg/mL solution of ethidium bromide for 20 minutes and then destained in distilled 

water. Ethidium bromide-stained bands were visualized using a 312 nm ultraviolet viewing box and 

photographed. 

“Cold” single-strand conformation polymorphism analysis (SSCP): A mixture consisting of 5 μL of PCR 

product (double-stranded DNA), 0.6 μL of 1M methylmercury hydroxide, 1 μL of 15% W/V Ficoll 

(molecular weight 400,000) loading buffer containing 0.25% bromophenol blue and 0.25% xylene cyanol, 

and 13.4 μL of 1X TBE buffer (Novex, San Diego, CA) was prepared to yield a total volume of 20 μL. 

This nonradioactive mixture was heated to 85E C for 5 minutes and then plunged into ice prior to loading the 

entire 20 μL into the gel. A 20% polyacrylamide TBE gel was used for K-ras with the matching gel 

electrophoresis unit (Novex, San Diego, CA). The buffer chamber was filled with 1.5X TBE buffer. The 

gel was run at 300 volts in a 5E C cold room until a light blue marker reached the bottom of the gel. A 

positive control for K-ras mutations and one undenatured DNA control (without methylmercury hydroxide 

and no heat) were run with unknown samples. Gels were stained with a 0.5 μg/mL solution of ethidium 

bromide for 20 minutes and destained in distilled water for 5 minutes. The stained bands were visualized 

under a UV viewing box and photographed. For identification of K-ras mutation at codon 61, RFLP was 

used with XbaI enzyme digestion and “cold” SSCP analysis was performed on the same 20% gel. 

Direct sequencing: Direct sequencing of the amplified first and second exon of the K-ras gene was 

performed as described by Tindall and Stankowski (1989) using previously described sequencing primers 

(Devereux et al., 1991). 

RESULTS 

In order to determine if the cobalt sulfate heptahydrate-induced neoplasms contained a K-ras mutation profile 

similar to that observed with “spontaneous” neoplasms, sample groups of six neoplasms consisting of 3 adenomas and carcinomas from the chamber control, seven, eight, and 11 neoplasms from the 0.3 mg/m , 3 1.0, and 3.0 mg/m dose groups, respectively, were evaluated by PCR amplification of K-ras exon 1 or 

K-ras exon 2 followed by RFLP for the two codon 61 mutations CTA and CGA in the B6C3F1 mouse 

(Table I1). SSCP was used as an alternative screening method for detection of K-ras mutations in DNA, and 

mutations were confirmed by direct sequencing. Mutation spectra in codons 12, 13, and 61 of the K-ras 

gene had some similarity to those identified in spontaneous lung neoplasms. 

Of the K-ras mutations detected, a higher frequency (5/9, 55%) of G to T transversions was detected at the 

second base of codon 12 compared to 0/1 (0%) for chamber controls or 1/24 (4%) for NTP historical 

controls. K-ras codon 61 CTA or CGA mutations were not present in the cobalt sulfate heptahydrateinduced lung neoplasms. A trend toward a dose-response relationship in the frequency of K-ras mutations 

was observed in cobalt sulfate heptahydrate-induced lung neoplasms: 14% versus 38% versus 45% for the 3 0.3, 1.0, and 3.0 mg/m doses, respectively (Table I2). There were generally no differences in the mutation 

frequency or spectra between benign and malignant lung neoplasms (data not shown). 

DISCUSSION 

In examining the K-ras mutations detected in the cobalt sulfate heptahydrate study, a higher frequency (5/9, 

55%) of G to T transversions was detected at codon 12 compared to 0/1 (0%) for chamber controls or 1/24 

(4%) for historical controls. These findings are consistent with the work of Zeiger et al. (1992), in which 

cobalt sulfate heptahydrate showed a weakly positive response in Salmonella typhimurium strain TA100 in 

the absence of exogenous metabolic activation as well as with hamster or rat liver S9. 
236 Cobalt Sulfate Heptahydrate, NTP TR 471 

The higher number of G to T transversions at codon 12 is supportive evidence that cobalt sulfate 

heptahydrate may indirectly damage DNA by oxidative stress. GGT to GTT mutations appear to be 

infrequent in spontaneous lung neoplasms from B6C3F1 mice (Table I1). However, G to T transversions are

commonly detected DNA base changes associated with active oxygen species and are consistent with 8-OH-G 

adducts produced during oxidative damage to DNA (Tchou et al., 1991; Shigenaga and Ames, 1991; Janssen 

et al., 1993). 8-OH-G is a suspect lesion in the formation of both spontaneous cancers and those induced by 

various agents such as 4-nitroquinoline oxide, ionizing radiation, KBrO , and 2-nitropropane (Floyd, 1990; 3 

Foley et al., 1993). Thus, cobalt sulfate heptahydrate exposure in B6C3F mice may have resulted in the 1 

generation of hydroxyl radicals that could have enhanced G to T transversions at the second base of codon 

12. Consistent with these findings is the work of Shi et al. (1993), in which cobalt sulfate heptahydrate was 

shown to catalyze the production of oxygen-based free radicals. 

The observation of similar frequencies and spectra of mutations in cobalt sulfate heptahydrate-induced 

alveolar/bronchiolar adenomas and carcinomas is consistent with other studies showing that K-ras activation 

occurs early and may be an initiating event in murine lung carcinogenesis (Sills et al., 1995). If mutations in 

the K-ras gene occurred later, an increased frequency of K-ras mutations would be expected in carcinomas. 

In B6C3F mice exposed to cobalt sulfate heptahydrate, specific K-ras mutations did not correlate with 1 

specific morphological patterns or sizes of lung neoplasms, a finding supported by Ohmori (1992). 
Cobalt Sulfate Heptahydrate, NTP TR 471 237 

REFERENCES 

Devereux, T.R., Anderson, M.W., and Belinsky, S.A. (1991). Role of ras protooncogene activation in the 

formation of spontaneous and nitrosamine-induced lung tumors in the resistant C3H mouse. Carcinogenesis 

12, 299-303. 

Devereux, T.R., Belinsky, S.A., Maronpot, R.R., White, C., Hegi, M., Patel, A.C., Foley, J.F., 

Greenwell, A., and Anderson, M.W. (1993a). Comparison of pulmonary O6-methylguanine DNA adduct 

levels and K-ras activation in lung tumors from resistant and susceptible mouse strains. Mol. Carcinog. 8, 

177-185. 

Devereux, T.R., Foley, J.F., Maronpot, R.R., Kari, F., and Anderson, M.W. (1993b). Ras proto-oncogene 

activation in liver and lung tumors from B6C3F mice exposed chronically to methylene chloride. 1 

Carcinogenesis 14, 795-801. 

Floyd, R.A. (1990). Role of oxygen free radicals in carcinogenesis and brain ischemia. FASEB J. 4, 

2587-2597. 

Foley, J., Ton, T., Maronpot, R., Butterworth, B., and Goldsworthy, T.L. (1993). Comparison of 

proliferating cell nuclear antigen to tritiated thymidine as a marker of proliferating hepatocytes in rats. 

Environ. Health Perspect. 101, 199-206. 

Janssen, Y.M.W., Van Houten, B., Borm, P.J.A., and Mossman, B.T. (1993). Cell and tissue responses to 

oxidative damage. Lab. Invest. 69, 261-274. 

Marmur, J. (1961). A procedure for the isolation of deoxyribonucleic acid from micro-organisms. J. Mol. 

Biol. 3, 208-218. 

Ohmori, H. (1992). Comparison of K-ras gene mutation among simultaneously occurring multiple urethaneinduced lung tumors in individual mice. Carcinogenesis 13, 851-855. 

Saiki, R.K., Gelfand, D.H., Stoffel, S., Scharf, S.J., Higuchi, R., Horn, G.T., Mullis, K.B., and 

Erlich, H.A. (1988). Primer-directed enzymatic amplification of DNA with a thermostable DNA 

polymerase. Science 239, 487-491. 

Shi, X., Dalal, N.S., and Kasprzak, K.S. (1993). Generation of free radicals from model lipid 

hydroperoxides and H2O2 by Co(II) in the presence of cysteinyl and histidyl chelators. Chem. Res. Toxicol. 

6, 277-283. 

Shigenaga, M.K., and Ames, B.N. (1991). Assays for 8-hydroxy-2N-deoxyguanosine: A biomarker of 

in vivo oxidative DNA damage. Free Radic. Biol. Med. 10, 211-216. 

Sills, R.C., Hong, H.L., Greenwell, A., Herbert, R.A., Boorman, G.A., and Devereux, T.R. (1995). 

Increased frequency of K-ras mutations in lung neoplasms from female B6C3F mice exposed to ozone for 24 1 

or 30 months. Carcinogenesis 16, 1623-1628. 

Sukumuar, S., and Barbacid, M. (1990). Specific patterns of oncogene activation in transplacentally induced 

tumors. Proc. Natl. Acad. Sci. USA 87, 718-722. 
238 Cobalt Sulfate Heptahydrate, NTP TR 471 

Tchou, J., Kasai, H., Shibutani, S., Chung, M.H., Laval, J., Grollamm, A.P., and Nishimura, S. (1991). 

8-Oxogyanine (8-hydroxyguanine) DNA glycosylase and its substrate specificity. Proc. Natl. Acad. Sci. 

USA 88, 4690-4694. 

Tindall, K.R., and Stankowski, L.F., Jr. (1989). Molecular analysis of spontaneous mutations at the gpt 

locus in Chinese hamster ovary (AS52) cells. Mutat. Res. 220, 241-253. 

Wright, D.K., and Manos, M.M. (1990). Sample preparation from paraffin-embedded tissues. In 

PCR Protocols: A Guide to Methods and Applications (M.A. Innis, D.H. Gelfand, J.J. Sninsky, and 

T.J. White, Eds.), pp. 153-158. Academic Press, San Diego, CA. 

Zeiger, E., Anderson, B., Haworth, S., Lawlor, T., and Mortelmans, K. (1992). Salmonella mutagenicity 

tests: V. Results from the testing of 311 chemicals. Environ. Mol. Mutagen. 19 (Suppl. 21), 2-141. 
Cobalt Sulfate Heptahydrate, NTP TR 471 239 

TABLE I1 

K-ras Mutations in Lung Neoplasms of B6C3F Mice 1 

Activated 

Codon 12 

NùGGT 

Codon 13 

NùGGC 

Codon 61 

NùCAA 

Treatment K-ras (%) GTT GAT TGT CGT CTT ATT CGC GAC CTA CAT CAC CGA 

Control,

 Historical 24/75 (33%) 1 9 4 0 0 0 3 0 0 4 1 2 

Control,

 Chamber 1/6 (17%) 0 0 1 0 0 0 0 0 0 0 0 0 

Cobalt Sulfate Heptahydratea 

9/26 (35%) 5 2 1 0 0 0 1 0 0 0 0 0 

Ozone 19/27 (70%) 5 3 2 0 0 0 0 1 8 0 0 0 

1,3-Butadiene 6/9 (67%) 0 0 0 0 0 0 6 0 0 0 0 0 

Tetranitromethane 

10/10 (100%) 0 10 0 0 0 0 0 0 0 0 0 0 

Methylene

 Chloride 11/54 (20%) 1 1 1 0 0 0 1 0 1 1 4 1 

a One animal had two neoplasms of the same type and mutation (GTT) which were counted as one neoplasm. If counted as two neoplasms, 

the activated K-ras would be 10/27 (37%), and there would be 6 codon 12 GTT mutations. 

TABLE I2 

K-ras Mutation Profile in Lung Neoplasms of B6C3F Mice 1 

Treatment 

Concentration 

(mg/m )

3 

Activated 

K-ras (%) GTT GAT 

Codon 12 

NùGGT 

TGT CGT CTT ATT

Codon 13 

NùGGC 

CGC CTA 

Codon 61 

NùCAA 

CAT CAC CGA 

Chamber and Historical Control 

25/81 (31%) 1 

(1%) 

9 

(11%) 

5 

(6%) 

0 0 0 3 

(4%) 

0 4 

(5%) 

1 

(1%) 

2

(2%) 

Cobalt Sulfate Heptahydrate

 0.3 1/7 (14%) 1 

(14%)

0 0 0 0 0 0 0 0 0 0 

1.0 3/8 (38%) 2 

(25%) 

0 0 0 0 0 1 

(13%)

0 0 0 0 

3.0 5/11 (45%) 2 2 

(18%) (18%) 

1 

(9%)

0 0 0 0 0 0 0 0 

Totala 9/26 (35%) 5 

(19%) 

2 

(8%) 

1 

(4%) 

0 0 0 1 

(4%) 

0 0 0 0 

a One animal had two neoplasms of the same type and mutation (GTT) which were counted as one neoplasm. If counted as two neoplasms, 

the activated K-ras would be 10/27 (37%), and there would be 6 codon 12 GTT mutations. 
 

 

241 

APPENDIX J
 

IMPACT OF HELICOBACTER HEPATICUS
 

INFECTION IN B6C3F1
  MICE

FROM 12 NTP 2-YEAR CARCINOGENESIS STUDIES
 

1	 1 1 1 James R. Hailey , Joseph K. Haseman , John R. Bucher , Ann E. Radovsky , 2 2 1	 1 David E. Malarkey , Richard T. Miller , Abraham Nyska , and Robert R. Maronpot

1

National Institute of Environmental Health Sciences
 

Research Triangle Park, North Carolina
 

2

Department of Microbiology, Pathology, and Parasitology
 

College of Veterinary Medicine
 

North Carolina State University
 

Raleigh, North Carolina
 

ABSTRACT ............................................................... 242
 

INTRODUCTION ............................................................ 242
 

MATERIALS AND METHODS .................................................... 243
 

RESULTS AND DISCUSSION ..................................................... 245
 

REFERENCES .............................................................. 252
 

TABLE J1 Incidence of Helicobacter hepaticus-Associated Hepatitis in Control B6C3F1 Mice 
 

from Nine NTP 2-Year Studies ....................................... 256
 

TABLE J2 Identification of Helicobacter hepaticus with PCR-RFLP-Based Assays
 

in Control B6C3F Mice from 32 NTP 2-Year Studies 1
 

and Three NTP 13-Week Studies ...................................... 256
 

TABLE J3 Comparison of Neoplasm Incidences in Control B6C3F1 Mice 
 

from Helicobacter hepaticus-Affected and Unaffected NTP 2-Year Studies .......... 257
 

TABLE J4 Liver Neoplasm Incidences and Body Weights of Control B6C3F1 Mice 
 

in Relation to Study Start Dates of Helicobacter hepaticus-Affected 

and Unaffected NTP 2-Year Studies .................................... 258
 

TABLE J5 Association of Liver Neoplasm Incidence
 

and Severity of Helicobacter hepaticus-Associated Hepatitis 

in Control B6C3F1
  Mice from Nine Affected NTP 2-Year Studies ................ 259

TABLE J6	 H-ras Codon 61 AAA Mutations in Spontaneous Liver Neoplasms 

in Control B6C3F1
  Mice from Helicobacter hepaticus-Affected

and Unaffected NTP 2-Year Studies .................................... 259
 

TABLE J7 Proliferating Cell Nuclear Antigen Labeling Indices in the Liver 

of Control B6C3F1 Mice 
 ............................................ 260

TABLE J8	 Summary of Target Sites of Carcinogenicity in B6C3F Mice 1
 

from NTP 2-Year Studies with Helicobacter hepaticus-Associated Hepatitis ......... 261
 
242 Cobalt Sulfate Heptahydrate, NTP TR 471 

IMPACT OF HELICOBACTER HEPATICUS 

INFECTION IN B6C3F MICE 1 

FROM 12 NTP 2-YEAR CARCINOGENESIS STUDIES 

ABSTRACT 

Male and female B6C3F1 mice from 12 NTP 2-year carcinogenesis studies were found to be infected with

Helicobacter hepaticus. Many of the male mice from nine of these studies (“affected” studies) had an 

associated hepatitis. The current evaluations were performed in an attempt to determine if the data from the 

H. hepaticus-affected NTP B6C3F1 mouse studies were compromised and unsuitable for cancer hazard

identification. The incidences of neoplasms of the liver (both hepatocellular neoplasms and 

hemangiosarcoma), but not of other organs in control male B6C3F1 mice, were found to be increased in

affected studies compared to control males from unaffected studies. The increased incidence of 

hepatocellular neoplasms was observed in those males exhibiting H. hepaticus-associated hepatitis. Other 

observations further differentiated control male mice from affected and unaffected studies. H-ras codon 61 

CAA-to-AAA mutations were less common in liver neoplasms in males from affected studies compared to 

historical and unaffected study controls. In addition, increases in cell proliferation rates and apoptosis were 

observed in the livers of male mice with H. hepaticus-associated hepatitis. These data support the hypothesis 

that the increased incidence of liver neoplasms is associated with H. hepaticus and that hepatitis may be 

important in the pathogenesis. Therefore, interpretation of carcinogenic effects in the liver of B6C3F1 mice

may be confounded if there is H. hepaticus-associated hepatitis. 

INTRODUCTION 

Helicobacter-Induced Diseases 

Since the bacterium H. pylori was isolated from humans in 1983, numerous Helicobacter species have been 

identified in several laboratory and domestic animal species. Their pathogenicity varies, with some species 

inducing significant disease while others appear merely to colonize the gastrointestinal tract. H. pylori is 

known to cause chronic gastritis and peptic ulcers in humans (Marshall and Warren, 1984; Graham, 1989; 

Lee et al., 1993) and, more recently, has been linked to adenocarcinoma and mucosa-associated lymphoma 

of the stomach (Fox et al., 1989; Nomura et al., 1991; Parsonnet et al., 1991; Wotherspoon et al., 1993). 

Based on epidemiological and pathology findings, the International Agency for Research on Cancer (1994) 

has classified H. pylori as a group 1 carcinogen in humans. H. hepaticus is associated with an increase in 

liver neoplasm incidences in A/JCr mice (Ward et al., 1994a; Fox et al., 1996). 

H. hepaticus commonly colonizes the gastrointestinal tract of many strains of mice from many sources (Fox 

et al., 1994; Ward et al., 1994b; Shames et al., 1995). It has been shown to be pathogenic, with hepatitis 

highly prevalent in some strains of mice (A/JCr, BALB/cAnNCr, C3H/HeNCr, SJL/NCr, and SCID/NCr) 

(Ward et al., 1994b). Intestinal colonization does not necessarily result in subsequent hepatitis, and the 

conditions that lead to migration of the organism from the intestine to the liver have not been determined. 

H. hepaticus appears to reside primarily within the bile canaliculi. Male mice were reported to have a 

greater incidence and severity of hepatitis than female mice, and this finding occurred in NTP studies as 

well. The recently identified H. bilis, like H. hepaticus, colonizes the biliary tract, liver, and intestine of 

mice. While H. bilis has been identified in animals with chronic hepatitis, whether it caused the hepatitis is 

not known (Fox et al., 1995). 

The pathogenesis of H. hepaticus-induced disease has not been fully characterized. In susceptible strains of 

mice, H. hepaticus can cause acute, focal, nonsuppurative, necrotizing hepatitis, which progresses to 

chronic, active hepatitis characterized by minimal necrosis, hepatocytomegaly, oval cell hyperplasia, and 
Cobalt Sulfate Heptahydrate, NTP TR 471 243 

cholangitis. H. hepaticus has been found to possess high levels of urease (Fox et al., 1994). H. hepaticus is 

often isolated from the cecum and colon but is not necessarily isolated from the liver of A/JCr mice, even 

though these animals develop severe hepatitis. Culture supernatants from several strains of H. hepaticus and 

several other Helicobacter species were shown to cause cytopathic effects in a rodent hepatocyte cell line 

(Taylor et al., 1995). Ward et al. (1996) suggested that autoimmunity may play a role in the progressive 

hepatitis and carcinogenesis in livers infected with H. hepaticus. 

NTP Infectious Disease Surveillance 

In 1993, during the histological evaluation of an NTP 2-year study, pathologists identified a constellation of 

liver lesions (hepatitis) in control and treated male mice that was consistent with what would later be 

described in mice infected with H. hepaticus (Ward et al., 1993, 1994a; Fox et al., 1994). Subsequently, 

pathology results from all mouse studies begun since 1984 (67 two-year studies) were reviewed for diagnoses 

of the characteristic hepatitis; the lesions were identified in nine studies (NTP, 1998a,b,c,d,e,f). Silver stains 

revealed helical bacteria consistent with Helicobacter present in the liver of male mice in the nine studies. 

Every reasonable measure is taken to prevent the occurrence of infectious diseases during NTP 2-year 

carcinogenicity studies. When infections occasionally occur, care is taken to identify the causal agent and its 

source, measures are taken to ensure that animals in later studies will not be infected, and the potential 

impact on biological parameters (primarily neoplastic endpoints) important in interpretation of the study is 

determined. To date, animals (control and treated) from a few studies have had a mild pulmonary 

inflammatory response presumed to be caused by an infectious agent. In other studies, there have been 

utero-ovarian infections with Klebsiella sp. (Rao et al., 1987) and fungal infections of the nasal cavity. For 

scientifically valid reasons, interpretation of chemical-related effects was not considered significantly 

compromised in any of these studies. Unlike the previous infections, H. hepaticus involves the liver, the 

major metabolic organ, and has been associated with an increase in incidences of liver neoplasms in the 

A/JCr mouse (Ward et al., 1994a). Therefore, when the contemporary epizootic of H. hepaticus infection in 

the United States affected several NTP studies, use of the data for hazard identification was questioned. The 

first step was to determine the extent of the infection within NTP studies and then evaluate the impact the 

infection had on biological parameters important in interpretation of the carcinogenic potential of test 

chemicals. 

MATERIALS AND METHODS 

Histologic Examination 

Studies in which mice were potentially infected with H. hepaticus were identified by reviewing the summary 

pathology tables for characteristic diagnoses: oval and/or biliary epithelial hyperplasia, hepatocyte 

enlargement (often diagnosed as karyomegaly), chronic inflammation, and regenerative hyperplasia. All 

13-week and 2-year studies begun by the NTP since 1984 and for which complete pathology data were 

available (67 two-year studies) were examined. Eight contemporary studies in which the characteristic 

lesions were not identified from pathology tables were randomly selected for histologic reevaluation. Slides 

containing sections of hematoxylin- and eosin-stained livers from 20 to 25 control and 20 to 25 high-dose 

male mice from each of seven 2-year studies and one 13-week study (10 animals from each group) were 

reexamined microscopically for the presence of hepatitis potentially related to H. hepaticus infection. 

Hepatitis consistent with that observed with H. hepaticus infection was not observed in any of these studies. 

Liver sections from five or more animals from each of nine 2-year studies in which hepatitis was observed 

were prepared using the Warthin-Starry silver stain or Steiner’s modification to identify silver-positive helical 

bacteria. 
244 Cobalt Sulfate Heptahydrate, NTP TR 471 

PCR-RFLP Detection of Helicobacter DNA 

Assays based on polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) were 

conducted at the NIEHS (Malarkey et al., 1997) and the University of Missouri Research Animal Diagnostic 

and Investigative Laboratory (MU-RADIL) (Riley et al., 1996) on liver tissue from approximately 

20 animals from each of 32 NTP 2-year studies (including the nine affected studies) and three NTP 13-week 

studies. The majority of these studies were selected because they were begun at approximately the same time 

(1988-1990) as the nine affected studies. Also, two earlier studies (1984-1985; mouse life-span and 

p-nitroaniline studies) and one later study (1993; methyleugenol) were selected. The mouse life-span study 

was designed to evaluate the incidences of spontaneous changes associated with age; therefore, there is no 

NTP Technical Report. Pathology peer review is not complete for the methyleugenol study, and the NTP 

Technical Report (NTP, 1998g) has not been completed. Frozen tissue was available from 22 of these 

studies, while only formalin-fixed tissue was available for the remaining ten 2-year studies and the three 

13-week studies. Most of the assays were conducted by MU-RADIL, which used Helicobacter genusspecific primers; MU-RADIL used restriction endonucleases on a subset of positives to determine if the 

species was H. hepaticus. DNA was isolated from frozen liver samples with a QIAamp Tissue Kit (Qiagen 

Inc., Chatsworth, CA) according to the manufacturer’s recommendations or routine phenol/chloroform 

extraction (Malarkey et al., 1997). DNA content and purity were determined spectrophotometrically by 

measuring the A260/A280 optical density ratio. To isolate DNA from paraffin-embedded samples, five 10-μm 

sections were washed twice with 1 mL xylene and twice with 500 μL ethanol. Tissues were then dried 

within a vacuum centrifuge prior to DNA isolation as described above. Routine measures were taken to 

avoid contamination at every step from tissue collection to PCR amplification, and concurrently run controls 

without DNA were consistently negative. 

Statistical Analyses 

Multiple regression procedures were used to compare control neoplasm rates in the nine affected studies with 

the 26 unaffected contemporary studies which had no histologic evidence of H. hepaticus-associated liver 

disease. While frozen liver tissue was unavailable from 13 of these 26 studies, none showed the hepatitis 

indicative of H. hepaticus and thus were assumed to be unaffected. Potential confounding factors such as 

body weight, date study was begun, route of administration, and animal supplier were included as 

covariables in the statistical analysis. 

Analysis for H-ras Codon 61 CAA-to-AAA Mutations 

For analyses of formalin-fixed tissue, three to five unstained serial sections (10 μm thick) were cut from 

paraffin blocks containing hepatocellular adenomas or carcinomas. Paraffin-embedded tissues were 

deparaffinized and rehydrated prior to being digested with proteinase k overnight at 55E C to isolate DNA. 

Frozen tissues were digested with 10 mg/mL pronase in 1% sodium dodecyl sulfate in TNE buffer (10 mM 

TRIS, 150 mM NaCl, and 2 mM EDTA; pH 7.5) overnight at 37E C; DNA was isolated by phenol 

chloroform extraction and precipitated with ethanol (Marmur, 1961; Sills et al., 1995). 

Nested primers were used for amplification of exon 2 of H-ras by PCR. The outer primers were 5'-CCA 

CTA AGC CTG TTG TGT TTT GCA G-3' (forward primer) and 5'-CTG TAC TGA TGG ATG TCC TCG 

AAG GA-3' (reverse primer). The inner primers (second round of amplification) were 5'-GAC ATC TTA 

GAC ACA GCA GTT-3' (forward primer) and 5'-GGT GTT GTT GAT GGC AAA TAC-3' (reverse 

primer). Although the normal sequence of codon 60 is GCT, the forward PCR primer is made with a T at 

the penultimate 3' base to create the restriction site for Mse1. 

A nonradioactive RFLP method was employed to identify CAA-to-AAA mutations in the H-ras gene at 

codon 61 in liver neoplasms (Lee and Drinkwater, 1995). This was based on Mse1 enzyme restriction 

cutting only the sequence 5'-TTAA-3'. Thus, Mse1 will detect CÿA conversion mutation at the first 

position of codon 61. 
Cobalt Sulfate Heptahydrate, NTP TR 471 245 

Analysis of PCNA and Apoptosis 

Detailed methods are included in a report by Nyska et al. (1997). Cell proliferation was assessed in 

nonneoplastic areas of the liver, kidney, and lung by determining a PCNA S-phase labeling index (the 

percentage of cells in S phase). The identification of apoptotic cells was based on morphologic criteria 

(Garewal et al., 1996; Goldsworthy et al., 1996) and confirmed immunohistochemically by the terminal 

deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) procedure (Gavrieli et al., 

1992). 

RESULTS AND DISCUSSION 

Identification of H. hepaticus Infection in NTP Studies 

Determining the extent of H. hepaticus infection involved a three-pronged approach of histologic evaluation, 

silver stains, and PCR-RFLP based assays; all were necessary because of the limitations identified for each. 

In NTP studies, and as reported in other studies (Ward et al., 1994b), there were no obvious clinical signs of 

infection, and the only significant histologic lesion (hepatitis) was observed in the liver, primarily in males. 

Therefore, summary pathology tables were reviewed to identify studies that may have been affected by 

H. hepaticus-associated hepatitis. Male mice from nine studies were identified (Table J1) as having the 

hepatitis. Eight of the nine studies were begun during a time span of about 6 months (July 1990 to 

January 1991), while the other study was begun much earlier (October 1988). The hepatitis was not 

observed in any 13-week studies. Use of histologic evaluation for identification of infected animals has 

limitations, however. It is somewhat insensitive, as H. hepaticus has been cultured and identified by PCRRFLP methods within livers of animals with no histological evidence of infection (Fox et al., 1998). This 

may be explained in part by the limited sampling (two liver sections) and the sometimes focal nature of 

H. hepaticus-associated hepatitis. Also, while in the more severely affected animals the hepatitis appears 

somewhat characteristic, component lesions of the hepatitis are not pathognomonic, and, when the hepatitis is 

subtle in 2-year old animals, it is more difficult to recognize or attribute to H. hepaticus. 

Within affected studies, the incidences of the hepatitis in male mice varied from 16% to 78% (Table J1). 

While generally mild to moderate, the hepatitis varied in severity from barely detectable in some animals to 

extensive liver involvement and regeneration in others. Only a few females were identified as having the 

characteristic hepatitis (Table J1). In general, the incidences and severities of H. hepaticus-associated 

hepatitis were similar between control and treated groups. This constellation of nonneoplastic liver lesions, 

while not pathognomonic, was certainly suggestive of an H. hepaticus infection, particularly when observed 

in control animals. Characteristic lesions included proliferation of oval and/or biliary epithelial cells, 

hepatocyte enlargement (diagnosed as karyomegaly), and chronic inflammation. In many instances, areas of 

regenerative hyperplasia were identified within diseased liver. 

Helicobacter spp. are not usually observed on routine histologic examination of hematoxylin and eosinstained sections of liver. The methods for confirmation of infection with Helicobacter include WarthinStarry silver stain or Steiner’s modification (Garvey et al., 1985) of this stain for direct microscopic 

observation of the organisms in tissue; however, this can be a relatively insensitive technique when few 

organisms are present. In most instances, histologic differentiation between Helicobacter species is not 

possible. Speciation can usually be accomplished with electron microscopy, but this technique is both time 

consuming and labor intensive. Microbiologic culture of feces, cecal smears, and fresh or frozen liver is 

also possible. Currently, assays involving amplification of the DNA of the organism using PCR are the most 

rapid and perhaps the most sensitive methods of detection, and the use of restriction endonucleases has 

allowed a determination of the species present. PCR-based methods also can be used on feces, cecal 

contents, or liver homogenates and are most sensitive when using fresh or frozen tissue (Riley et al., 1996; 

Malarkey et al., 1997). 
246 Cobalt Sulfate Heptahydrate, NTP TR 471 

Using Warthin-Starry silver stains or Steiner’s modification on the livers of five or more animals per study, 

helical bacteria (Helicobacter) were identified in animals from the nine affected studies. In some animals, 

helical bacteria were numerous, suggesting a heavy bacterial burden in these infected animals. However, 

even in these animals with abundant organisms, few to none were observed in proliferative hepatic lesions 

such as foci and neoplasms. Helical bacteria were not identified in approximately 25% of males with 

moderate hepatitis and were rarely identified in males without hepatitis or in females. The absence of 

identification of helical organisms by silver stains does not preclude infection, nor does the presence of 

organisms confirm H. hepaticus. Based upon current knowledge, however, the characteristic liver lesions in 

B6C3F mice, coupled with the presence of silver-positive helical organisms, are highly suggestive of 1 

H. hepaticus infection. 

As the NTP evaluation evolved, PCR-based assays were developed that appeared more sensitive than 

histologic evaluation and silver stains for identification and speciation of Helicobacter. Therefore, 

PCR-RFLP-based assays were used to confirm the presence of pathogenic Helicobacter (primarily 

H. hepaticus) within the nine affected studies and to determine whether there was H. hepaticus infection in 

other NTP studies. Unfortunately, none of the PCR-based assays had been specifically developed for, or 

proven reliable for use with, formalin-fixed tissue. Frozen tissue was available from a limited number of 

animals from a limited number of NTP studies, including only three of the nine affected studies. 

Furthermore, available frozen liver was almost always limited to tissue from a neoplasm, and, based upon 

results obtained with silver stains, organisms are generally not readily observed within proliferative hepatic 

lesions, even when organisms are abundant in adjacent liver tissue. Because the availability of frozen tissue 

was limited, a PCR-RFLP-based assay was developed and evaluated (Malarkey et al., 1997) for use with 

frozen or formalin-fixed tissue. 

The NIEHS and MU-RADIL laboratories conducted PCR-RFLP-based assays on 32 NTP 2-year studies and 

three NTP 13-week studies (data not shown); frozen tissues from 22 of the 2-year studies were available. All 

three bioassays in which hepatitis was identified and for which frozen tissue was available were positive for 

H. hepaticus by the PCR-RFLP-based assays (Table J2). At a third laboratory, H. hepaticus was also 

cultured from the liver tissue of animals in one of these studies (Fox et al., 1998). Formalin-fixed tissues 

from two of the three studies were evaluated and were also positive; these tissues had been fixed in formalin 

for less than 48 hours. In the other six affected studies, for which only formalin-fixed tissue was available, 

H. hepaticus was identified in only 1 of 120 animals (Table J2). This decreased sensitivity was considered to 

be related to the prolonged formalin fixation (Malarkey et al., 1997) rather than proof of an absence of 

H. hepaticus. The presence or absence of H. hepaticus apparently cannot be confirmed with current 

PCR-RFLP-based assays in liver that has been fixed in formalin for long periods (weeks or months). In the 

three 13-week studies with formalin-fixed tissue, only 1 of 30 animals was positive for H. hepaticus. 

Within the three affected, PCR-RFLP-positive 2-year studies, H. hepaticus was often identified by PCR in 

frozen livers of mice that had no overt hepatitis. In fact, based upon the combined data from two studies 

(including PCR results from three laboratories), of 57 animals without characteristic liver lesions, 13 of 

24 male mice (54%) and 17 of 33 female mice (52%) were positive for H. hepaticus. Furthermore, 

H. hepaticus was identified by PCR in frozen liver of several animals from three “unaffected” studies in 

which hepatitis typical of that associated with H. hepaticus was not observed (Table J2). Apparent 

variability occurs between various strains of mice and between individual mice from affected studies in 

developing hepatitis in response to H. hepaticus infection. One would assume that, within affected studies, 

most or all animals have been exposed to the organism, and even animals resistant to developing hepatitis 

may have organisms within the liver. This assumption is supported by the fact that animals without hepatitis 

are often positive with PCR-RFLP-based assays. Therefore, although alternative explanations are possible, 

the three PCR-RFLP-positive studies in which liver lesions are absent are assumed to be true positives. In 

fact, helical organisms were identified with a silver stain in one animal from one of these studies (Malarkey 

et al., 1997). Therefore, in addition to assessing the affect of H. hepaticus in the nine affected 2-year 
Cobalt Sulfate Heptahydrate, NTP TR 471 247 

studies, the significance of a positive PCR-RFLP assay for H. hepaticus in the absence of liver lesions is also 

an important question. 

Inconsistent Results with PCR-Based Methods 

As with any technique, the PCR-RFLP-based assays have limitations even when used to assay fresh and 

frozen tissue. One assessment of the variability in results of PCR and serologic analyses for Helicobacter 

among three commercial laboratories revealed significant inconsistencies (Dew et al., 1997). Others 

(J.M. Ward and J. Thigpen, personal communications) have obtained similarly inconsistent results when 

sending replicate samples to different laboratories. Though the number of samples evaluated by both the 

NIEHS and MU-RADIL laboratories was limited, there was good, but not complete, correlation of 

PCR-RFLP results. Also, within the affected studies, the PCR assays were not positive in some animals with 

liver disease. This result may be explained, in part, by the fact that the only frozen tissues available were 

neoplasms; as described above, neoplasms are expected to have fewer organisms. 

Analysis of H. hepaticus-Affected and Unaffected Studies 

for Incidence of Common Neoplasms 

To determine whether the incidences of various neoplasms were different between control groups from 

affected and unaffected studies, the nine affected studies were compared to 26 unaffected studies begun at 

relatively similar times (Table J3). There were no statistically significant differences in body weight or 

survival among the affected and unaffected studies. The neoplasms evaluated represent those that occurred at 

high enough incidences in various organs for statistically significant differences to be detected. Using 

multiple regression procedures, male mice in the nine affected studies were demonstrated to have a 

significantly (P<0.05) increased incidence of only two neoplasm types, both of which were in the liver 

(hepatocellular neoplasms and hemangiosarcoma), when compared to the unaffected studies. Because of 

these differences, there was also a corresponding significant difference in the overall incidence of malignant 

neoplasms (all sites) as well as in the overall proportion of neoplasm-bearing animals. No other tissue site 

showed a significant difference in the incidence of neoplasms. For female mice, the slightly increased 

incidence of hepatocellular neoplasms observed in the affected studies was not statistically significant. 

This seemingly simple analysis is complicated by several potential confounding variables. There have been 

coordinate, time-related increases in body weight and in the incidence of liver neoplasms in mice in NTP 

studies (Haseman, 1992). Table J4 presents the liver neoplasm incidences in relation to the dates the studies 

began and clearly shows the increases in liver neoplasm incidences and body weights (Seilkop, 1995). In 

assessing differences in neoplasm incidences between H. hepaticus-affected and unaffected studies, the most 

relevant comparison would be between studies begun at approximately the same time. The starts of 20 of the 

26 unaffected studies were clustered near the early part of the time frame (April 1988 to June 1990), while 

the starts of the affected studies were clustered toward the later end, with eight of the nine studies begun 

between July 1990 and January 1991; incidences of liver neoplasms in these later studies are expected to be 

higher based on trends in body weight alone. While the slightly increased incidences of liver neoplasms 

observed in female control mice in the nine affected studies is likely due to clustering in time, clearly, this 

alone cannot account for the increased liver neoplasm incidences observed in control male mice in the 

affected studies (Table J3). 

Ideally, unaffected studies used in the above comparison should not only be free of histologic evidence of 

infection with H. hepaticus but should be confirmed as negative by PCR assays. Thirteen of these 26 studies 

could not be confirmed as negative by PCR because frozen tissue was not available; however, 

H. hepaticus-associated hepatitis was not present in any of the 26 studies. Because these and other data 

reported to date suggest that hepatitis is associated with neoplasm development in the liver, it seems 

reasonable to include those 13 studies, unconfirmed by PCR, in this analysis. The majority of the 13 studies 

confirmed as negative by PCR were begun much earlier than the clearly affected studies, and, therefore, 

comparing them alone to the nine affected studies is not reasonable. Although not presented here, a number 
248 Cobalt Sulfate Heptahydrate, NTP TR 471 

of comparisons were made with various groupings of studies based on the degree of confidence in their 

infection status. Although the outcomes of the various comparisons varied somewhat, incidences of 

hepatocellular neoplasms and hemangiosarcomas of the liver were consistently increased in control male mice 

from affected studies compared to control males from unaffected studies. Significantly increased liver 

neoplasm incidences generally were not observed in females. Importantly, the following data corroborate the 

findings and association with H. hepaticus identified in these analyses. 

Analysis of Hepatitis-Positive and Hepatitis-Negative Mice 

for Liver Neoplasm Incidence 

Several infectious agents known to be associated with increased incidences of neoplasms cause chronic 

inflammation in the target tissue or organ. It is commonly hypothesized that this inflammatory process may 

cause or contribute to the development of neoplasms. One approach to address this was to stratify the mice 

from the affected studies according to the severity of hepatitis and examine liver neoplasm incidences in 

relation to these groupings. Thus, animals within the nine affected studies were placed into three groups: 

1) animals with mild to moderate hepatitis considered related to H. hepaticus infection (+), 2) animals with 

minimal to mild hepatitis that may have been associated with H. hepaticus (±), and 3) animals with no 

hepatitis that was considered to be associated with H. hepaticus (S). Within these groupings, the incidence 

of liver neoplasms was significantly increased (P<0.05) in males with mild to moderate H. hepaticusassociated hepatitis (+) when compared to animals without such hepatitis (Table J5). The neoplasm 

incidence in animals with minimal lesions (±) was also increased. The liver neoplasm incidence in males 

without hepatitis (58%) was similar to the incidence (54.8%) in males from the 26 unaffected studies 

(Table J3). This analysis clearly suggests an association of H. hepaticus-associated hepatitis with increased 

liver neoplasm incidences. Females showed a similar trend, albeit not significant; however, these 

comparisons are weak because of the low numbers of females with hepatitis. 

Analysis of H-ras Oncogene Mutations in Liver Neoplasms in Mice 

from Affected and Unaffected Studies 

Liver neoplasms commonly occur in control B6C3F mice in 2-year studies. In the historical database of 1 

333 male and female mice with liver neoplasms, 106 (32%) had H-ras codon 61 CAA-to-AAA mutations 

(Maronpot et al., 1995). This historical control database is composed primarily of male data; however, 

adequate numbers of females have been assayed, and there was no significant difference in the incidences of 

CAA-to-AAA mutations between males and females. 

In an attempt to examine further whether H. hepaticus infection had an effect on the development of 

hepatocellular neoplasms, neoplasms from control male mice from selected affected (NTP, 1998a,b,c) and 

unaffected (NTP, 1993, 1998h) studies were evaluated for H-ras codon 61 CAA-to-AAA mutations 

(Table J6). Only 6% (2/33) of the hepatocellular neoplasms from control males with hepatitis from three 

affected studies had this mutation. This percentage is significantly (P<0.01) less than the 32% (11/34) 

observed in males from the two unaffected studies and less than the 32% (106/333) that occurred in 

historical control animals. In addition, neoplasms from males without hepatitis from the affected, PCRpositive triethanolamine study (NTP, 1998a) and the unaffected, PCR-positive methyleugenol study (NTP, 

1998g) were evaluated; the incidences of mutations in those groups were 3/14 (21%) and 2/17 (12%), 

respectively. 

Neoplasms from control female mice (none had hepatitis) from affected and unaffected studies were evaluated 

for the CAA-to-AAA mutation (Table J6). The mutation rate was low in both the affected studies (1/25; 4%) 

and the unaffected study (1/11; 9%) when compared to the 32% observed in the historical control groups. 

The finding of a different H-ras mutation profile in neoplasms of male mice from affected studies tends to 

support the association of increased neoplasm incidences with H. hepaticus, although there is no mechanistic 

understanding behind this observation. In a study of H. hepaticus-infected A/JCr mice, ras mutations were 
Cobalt Sulfate Heptahydrate, NTP TR 471 249 

not detected in the 25 hepatocellular neoplasms analyzed using a PCR/single-strand conformation 

polymorphism assay (Sipowicz et al., 1997). Because of the low spontaneous rate of liver neoplasms in the 

A/JCr mouse, there are few or no conclusive data on ras mutations in uninfected animals, however. Point 

mutations at codons 12, 13, and 61 of the Ki-, Ha- and N-ras genes were not identified in 45 early gastric 

carcinomas in humans, whether or not H. pylori was present (Craanen et al., 1995). If the increased 

incidence of hepatocellular neoplasms is associated with hepatitis, as many suspect, then one would expect 

the neoplasms from animals without hepatitis to have a similar mutational profile as that of the historical 

controls. The data do not provide a clear answer, because the hepatitis-free males from the affected 

triethanolamine study (NTP, 1998a) and the males from the methyleugenol study (NTP, 1998g), which were 

positive by PCR but lacked hepatitis, had mutation frequencies between those of the unaffected controls and 

the hepatitis-positive mice. Furthermore, mutations in neoplasms from females, none of which had hepatitis, 

from two affected and one unaffected study were very low compared to the historical controls. These 

findings were unexpected, and their significance is not understood. 

H. hepaticus-Associated Alterations in Cell Kinetics 

Studies evaluating cell kinetics were completed to explore further the link between hepatitis and the increased 

incidence of liver neoplasms (Table J7; Nyska et al., 1997). One of the major objectives was to determine 

whether there were differences between PCNA labeling indices in the livers of animals with hepatitis from 

three affected studies, cobalt sulfate heptahydrate, chloroprene, and triethanolamine (NTP, 1998a,b,c), 

compared to animals without hepatitis, whether from the same three affected studies or from an unaffected 9 study, 1-trans-delta -tetrahydrocannabinol (NTP, 1996). Male mice with hepatitis from the three affected 

studies had a significantly increased (P<0.001) labeling index, with a 24-fold increase over males from the 

unaffected study and a sixfold increase over males without hepatitis from the same three affected studies 

(Table J7). The labeling index increase in these mice was substantial and was considered biologically 

significant. Male mice without hepatitis from the three affected studies had a significantly greater labeling 

index (increased fourfold) than male mice from the unaffected study (Table J7). The significance of this 

finding is uncertain, as differences of a similar magnitude were observed in other comparisons. For 9 example, the labeling index of females from the unaffected 1-trans-delta -tetrahydrocannabinol study

(Table J7; NTP, 1996) was increased fivefold over females from the PCR-positive, hepatitis-negative 

scopolamine hydrobromide trihydrate study (NTP, 1997). Such differences may be within the limits of 

normal variability for 2-year-old animals. 

A second objective of the cell proliferation studies of the liver was to determine if labeling indices were 

increased in animals from the PCR-positive, hepatitis-negative methyleugenol (NTP, 1998g), scopolamine 

hydrobromide trihydrate (NTP, 1997), and mouse life-span studies compared to an unaffected PCR-negative 9 and hepatitis-negative 1-trans-delta -tetrahydrocannabinol study (NTP, 1996). The scopolamine 

hydrobromide trihydrate study was evaluated and included in the study by Nyska et al. (1997), while the 

methyleugenol and mouse life-span studies were completed later and are included in Table J7. The labeling 

indices of males from two of these three studies were almost identical to those of males from the unaffected 

study. However, the labeling index of males from the mouse life-span study is increased approximately 

fivefold over that of males from the unaffected study as well as fivefold over the labeling indices of males 

from the two like studies of scopolamine hydrobromide trihydrate and methyleugenol. This finding suggests 

that the increase observed in the mouse life-span study is not attributable to the presence of H. hepaticus, as 

two other studies also positive for H. hepaticus did not show a similar increase. 

The cell proliferation data for the liver from NTP studies are consistent with data from a study by Fox et al. 

(1996) in which cell proliferation indices were evaluated at 8, 10, and 13 months in the A/JCr mouse, which 

is generally believed to be more susceptible to H. hepaticus-associated hepatitis than the B6C3F1 mouse. In 

the study by Fox et al. (1996), cell proliferation rates were significantly increased at all time points in males. 

Some increases were observed in females in that study but did not reach statistical significance. An increased 
250 Cobalt Sulfate Heptahydrate, NTP TR 471 

incidence of hepatocellular neoplasms was observed only in the males. Though liver lesions were observed 

in females in that study, they were less severe than those in males. 

In addition to the liver, cell proliferation indices (PCNA) were evaluated in the kidneys and lungs of male 

and female mice in affected studies versus those in unaffected studies (Nyska et al., 1997). No apparent 

effect of H. hepaticus infection or the presence of hepatitis on PCNA indices was observed for the kidneys or 

lungs. 

Apoptosis (programmed cell death) is another important parameter in evaluations of cell kinetics. The 

apoptotic index in the liver of male mice with hepatitis from an affected study, cobalt sulfate heptahydrate 

(NTP, 1998b), was significantly (P<0.01) greater than that observed in males from the unaffected 1-trans­ 9 delta -tetrahydrocannabinol study and the PCR-positive, hepatitis-negative scopolamine hydrobromide

trihydrate study (Nyska et al., 1997). For females, there were no significant differences among the three 

studies. 

Two 13-week studies which were begun during the same time as the nine affected studies were randomly 

selected for evaluation of PCNA indices. H. hepaticus was not identified in either of the studies by 

PCR-RFLP; however, as with all NTP 13-week studies, only tissue fixed in formalin for an unspecified 

period was available. Because of this, no true negative control group was available; therefore, the labeling 

index of these 19- to 20-week-old animals was compared to values cited in the literature (Eldridge and 

Goldsworthy, 1996) for 20-week-old B6C3F mice. The labeling index in the NTP studies clearly was not 1 

increased (data not shown). 

The Impact of H. hepaticus on the Interpretation of 2-Year Carcinogenesis Studies 

Increases in the incidences of neoplasms are associated with a number of infectious agents. The chronic 

inflammation caused by these agents has been hypothesized to be important in the pathogenesis of the 

increased neoplasm incidences (e.g., gastric cancer associated with H. pylori). The increased incidences of 

liver neoplasms in male mice from the nine affected NTP studies were observed in the animals with 

H. hepaticus-associated hepatitis. Neoplasms from males with hepatitis tended to have an H-ras mutation 

profile different from that of animals from unaffected studies. Further, cell replication rates at 2 years were 

significantly higher in males with hepatitis compared to those in males without hepatitis. The data suggest 

that H. hepaticus-associated hepatitis is associated with the increased incidences of liver neoplasms in the 

male B6C3F1 mouse. Therefore, the most important consideration in evaluating the impact of H. hepaticus 

infection on the interpretation of study results appears to be the presence or absence of significant hepatitis. 

For any carcinogenicity study, data within and specific to the individual study provide the greatest basis for 

an accurate interpretation. However, it is prudent to consider and evaluate all data or information which may 

affect the interpretation. Based upon the data presented in this and other reports, general guidelines emerge 

that may be useful in interpreting potential chemical-associated carcinogenic effects in H. hepaticus-infected 

B6C3F mice. In a study with sufficient evidence of H. hepaticus-associated hepatitis (>10% of the animals 1 

having the characteristic hepatitis may be a reasonable guideline), interpretation of increased incidences of 

liver neoplasms (hepatocellular neoplasms and hemangiosarcoma) of male mice is considered to be 

potentially confounded. 

Altered chemical uptake and metabolism, due to the intestinal load of H. hepaticus and to H. hepaticusassociated liver disease, respectively, are possible reasons for considering that the male mouse response to 

chemical administration at sites other than the liver should also be considered confounded. Data do not 

currently exist that definitively answer this question. In this group of nine studies, however, there is no 

evidence to suggest that affected mice responded to chemical treatment in organs other than the liver in a 

manner different from mice in nonaffected studies. Within each study, there was excellent concordance in 

chemical-associated neoplasms between the male mice and the females, which had little or no hepatitis 
Cobalt Sulfate Heptahydrate, NTP TR 471 251 

(Table J8). Furthermore, analyses indicate that H. hepaticus is not associated with neoplastic responses 

outside the liver; incidences of neoplasms at sites other than the liver were not different between control 

groups from affected and unaffected studies (Table J3). Cell replication rates in two major organs (lung and 

kidney) also were not increased in control groups from affected studies compared to those from unaffected 

studies. 

One of the more difficult issues to address is whether interpretation of a treatment-related increase in liver 

neoplasm incidences in the female mouse is confounded when H. hepaticus-associated hepatitis is present 

within the male mice in the study. Most evidence to date links hepatitis with the increased liver neoplasm 

incidences observed in males, and female B6C3F mice in affected studies do not have significant hepatitis at 1 

2 years. The lack of hepatitis in females, however, is based on an analysis in which only late time points 

were evaluated histologically. Therefore, it is conceivable that hepatitis along with increased cell 

proliferation could have occurred earlier and resolved by 18 months to 2 years. Data collected to date, 

however, suggest that H. hepaticus-associated hepatitis is a late-developing and persistent disease in the 

B6C3F1 mouse. H. hepaticus-associated hepatitis has never been observed in any NTP 13-week studies, 

including five begun during the same 6-month time span as eight of the nine affected 2-year studies. Also, 

within affected 2-year studies, more males (51%) that were 18 to 24 months of age had hepatitis than those 

(34%) that were 12 to 18 months of age. This is consistent with a report by Ward et al. (1994b) that 

H. hepaticus-associated liver lesions are not observed at early time points in the B6C3F1 mouse. 

Nonetheless, within affected studies, female control mice did have a slightly elevated incidence of liver 

neoplasms when compared to control mice from unaffected studies, and the data derived from the H-ras 

mutation frequency analysis were inconclusive. The possibility that H. hepaticus-infected female mice from 

affected studies may respond differently to a liver carcinogen than mice from unaffected studies cannot be 

eliminated at this time. However, because within an affected study hepatitis is observed only rarely in 

females, until definitive data suggest otherwise, it is concluded that the interpretation of an apparent 

chemical-induced neoplastic effect in the liver of female mice is not confounded. To censor the few females 

with H. hepaticus-associated hepatitis from any statistical analyses of hepatocellular neoplasms would be 

prudent. Studies in the ostensibly more sensitive A/JCr mouse (Fox et al., 1996) also showed significant 

increases in neoplasm incidences and cell proliferation rates in the liver of H. hepaticus-infected males, but 

not females. 

Another concern is how to interpret possible chemical-related effects in a study in which the status of 

H. hepaticus infection cannot be determined by PCR-RFLP because only tissues fixed in formalin for more 

than 48 hours are available. While histologic evaluation is inadequate to identify infection, it appears 

adequate for identifying hepatitis severe enough to alter the outcome of the study. Therefore, in the absence 

of significant histologic evidence of H. hepaticus-associated hepatitis, the outcome of a 2-year study should 

not be considered potentially compromised. 

The causality between H. hepaticus infection and neoplasia has not been proven in the B6C3F1 mouse in

these studies, nor has the mechanism of this association been determined; further studies are needed. 

However, sufficient information exists to make reasonable scientific judgments relative to the interpretation 

of data from the nine 2-year carcinogenicity studies in the B6C3F1 mouse. Refinements to the above 

interpretive positions may occur if warranted by future information. 

REFERENCES 

Craanen, M.E., Blok, P., Top, B., Boerrigter, L., Dekker, W., Offerhaus, G.J.A., Tytgat, G.N.J., and 

Rodenhuis, S. (1995). Absence of ras gene mutations in early gastric carcinomas. Gut 37, 758-762. 
252 Cobalt Sulfate Heptahydrate, NTP TR 471 

Dew, J.A., Clifton, L.G., Sanders, B.L., and Reynolds, R.P. (1997). Comparison of results of Helicobacter 

tests performed by commercial laboratories. Contemp. Top. (AALAS) 36, 60. (Abstr.) 

Eldridge, S.R., and Goldsworthy, S.M. (1996). Cell proliferation rates in common cancer target tissues of 

B6C3F1 mice and F344 rats: Effects of age, gender, and choice of marker. Fundam. Appl. Toxicol. 32, 

159-167. 

Fox, J.G., Correa, P., Taylor, N.S., Zavala, D., Fontham, E., Janney, F., Rodriquez, E., Hunter, F., and 

Diavolitsis, S. (1989). Campylobacter pylori-associated gastritis and immune response in a population at 

increased risk of gastric carcinoma. Am. J. Gastroenterol. 84, 775-781. 

Fox, J.G., Dewhirst, F.E., Tully, J.G., Paster, B.J., Yan, L., Taylor, N.S., Collins, M.J., Jr., 

Gorelick, P.L., and Ward, J.M. (1994). Helicobacter hepaticus sp. nov., a microaerophilic bacterium 

isolated from livers and intestinal mucosal scrapings from mice. J. Clin. Microbiol. 32, 1238-1245. 

Fox, J.G., Yan, L.L., Dewhirst, F.E., Paster, B.J., Shames, B., Murphy, J.C., Hayward, A., Belcher, J.C., 

and Mendes, E.N. (1995). Helicobacter bilis sp. nov., a novel Helicobacter species isolated from bile, 

livers, and intestines of aged, inbred mice. J. Clin. Microbiol. 33, 445-454. 

Fox, J.G., Li, X., Yan, L., Cahill, R.J., Hurley, R., Lewis, R., and Murphy, J.C. (1996). Chronic 

proliferative hepatitis in A/JCr mice associated with persistent Helicobacter hepaticus infection: A model of 

Helicobacter-induced carcinogenesis. Infect. Immun. 64, 1548-1558. 

Fox, J.G., MacGregor, J., Shen, Z., Li, X., Lewis, R., and Dangler, C.A. (1998). Role of Helicobacter 

hepaticus in confounding results of a triethanolamine carcinogenesis study in mice. J. Clin. Microbiol. 

(in press) 

Garewal, H., Bernstein, H., Bernstein, C., Sampliner, R., and Payne, C. (1996). Reduced bile acid-induced 

apoptosis in “normal” colorectal mucosa: A potential biological marker for cancer risk. Cancer Res. 56, 

1480-1483. 

Garvey, W., Fathi, A., and Bigelow, F. (1985). Modified Steiner for the demonstration of spirochetes. 

J. Histotechnol. 8, 15-17. 

Gavrieli, Y., Sherman, Y., and Ben-Sasson, S.A. (1992). Identification of programmed cell death in situ via 

specific labeling of nuclear DNA fragmentation. J. Cell Biol. 119, 493-501. 

Goldsworthy, T.L., Fransson-Steen, R., and Maronpot, R.R. (1996). Importance of and approaches to 

quantification of hepatocyte apoptosis. Toxicol. Pathol. 24, 24-35. 

Graham, D.Y. (1989). Campylobacter pylori and peptic ulcer disease. Gastroenterology 96, 615-625. 

Haseman, J.K. (1992). Value of historical controls in the interpretation of rodent tumor data. Drug Inf. J. 

26, 191-200. 

International Agency for Research on Cancer (IARC) (1994). IARC Monographs on the Evaluation of 

Carcinogenic Risks to Humans: Schistosomes, Liver Flukes and Helicobacter pylori, Vol. 61. IARC, Lyon, 

France. 

Lee, A., Fox, J., and Hazell, S. (1993). Pathogenicity of Helicobacter pylori: A perspective. Infect. 

Immun. 61, 1601-1610. 
Cobalt Sulfate Heptahydrate, NTP TR 471 253 

Lee, G.-H., and Drinkwater, N.R. (1995). Hepatocarcinogenesis in BXH recombinant inbred strains of 

mice: Analysis of diverse phenotypic effects of the hepatocarcinogen sensitivity loci. Mol. Carcinog. 14, 

190-197. 

Malarkey, D.E., Ton, T.-V., Hailey, J.R., and Devereaux, T.R. (1997). A PCR-RFLP method for the 

detection of Helicobacter hepaticus in frozen or fixed liver from B6C3F mice. Toxicol. Pathol. 25, 1 

606-612.
 

Marmur, J. (1961). A procedure for the isolation of deoxyribonucleic acid from micro-organisms. J. Mol.
 

Biol. 3, 208-218.
 

Maronpot, R.R., Fox, T., Malarkey, D.E., and Goldsworthy, T.L. (1995). Mutations in the ras protooncogene: Clues to etiology and molecular pathogenesis of mouse liver tumors. Toxicology 101, 125-156.
 

Marshall, B.J., and Warren, J.R. (1984). Unidentified curved bacilli in the stomach of patients with gastritis
 

and peptic ulceration. Lancet 1, 1311-1314.
 

National Toxicology Program (NTP) (1993). Toxicology and Carcinogenesis Studies of Oxazepam
 

(CAS No. 604-75-1) in Swiss Webster and B6C3F Mice (Feed Studies). Technical Report Series No. 443. 1 

NIH Publication No. 93-3359. U.S. Department of Health and Human Services, Public Health Service, 

National Institutes of Health, Research Triangle Park, NC. 

9 National Toxicology Program (NTP) (1996). Toxicology and Carcinogenesis Studies of 1-Trans-delta ­

tetrahydrocannabinol (CAS No. 1972-08-3) in F344/N Rats and B6C3F Mice (Gavage Studies). Technical 1 

Report Series No. 446. NIH Publication No. 97-3362. U.S. Department of Health and Human Services, 

Public Health Service, National Institutes of Health, Research Triangle Park, NC. 

National Toxicology Program (NTP) (1997). Toxicology and Carcinogenesis Studies of Scopolamine 

Hydrobromide Trihydrate (CAS No. 6533-68-2) in F344/N Rats and B6C3F Mice (Gavage Studies). 1 

Technical Report Series No. 445. NIH Publication No. 97-3361. U.S. Department of Health and Human 

Services, Public Health Service, National Institutes of Health, Research Triangle Park, NC. 

National Toxicology Program (NTP) (1998a). Toxicology and Carcinogenesis Studies of Triethanolamine 

(CAS No. 102-71-6) in F344/N Rats and B6C3F Mice (Dermal Studies). Technical Report Series No. 449. 1 

NIH Publication No. 98-3365. U.S. Department of Health and Human Services, Public Health Service, 

National Institutes of Health, Research Triangle Park, NC. (in preparation) 

National Toxicology Program (NTP) (1998b). Toxicology and Carcinogenesis Studies of Cobalt Sulfate 

Heptahydrate (CAS No. 10026-24-1) in F344/N Rats and B6C3F Mice (Inhalation Studies). Technical 1 

Report Series No. 471. NIH Publication No. 98-3961. U.S. Department of Health and Human Services, 

Public Health Service, National Institutes of Health, Research Triangle Park, NC. (in press) 

National Toxicology Program (NTP) (1998c). Toxicology and Carcinogenesis Studies of Chloroprene 

(CAS No. 126-99-8) in F344/N Rats and B6C3F Mice (Inhalation Studies). Technical Report Series 1 

No. 467. NIH Publication No. 98-3957. U.S. Department of Health and Human Services, Public Health 

Service, National Institutes of Health, Research Triangle Park, NC. (in press) 

National Toxicology Program (NTP) (1998d). Toxicology and Carcinogenesis Studies of Technical Grade 

Sodium Xylenesulfonate (CAS No. 1300-72-7) in F344/N Rats and B6C3F Mice (Dermal Studies). 1 

Technical Report Series No. 464. NIH Publication No. 98-3380. U.S. Department of Health and Human 

Services, Public Health Service, National Institutes of Health, Research Triangle Park, NC. (in press) 
254 Cobalt Sulfate Heptahydrate, NTP TR 471 

National Toxicology Program (NTP) (1998e). Toxicology and Carcinogenesis Studies of AZT 

(CAS No. 30516-87-1) and AZT/"-Interferon A/D in B6C3F Mice (Gavage Studies). Technical Report 1 

Series No. 469. NIH Publication No. 98-3959. U.S. Department of Health and Human Services, Public 

Health Service, National Institutes of Health, Research Triangle Park, NC. (in press) 

National Toxicology Program (NTP) (1998f). Toxicology and Carcinogenesis Studies of Theophylline 

(CAS No. 58-55-9) in F344/N Rats and B6C3F Mice (Feed and Gavage Studies). Technical Report Series 1 

No. 473. NIH Publication No. 98-3963. U.S. Department of Health and Human Services, Public Health 

Service, National Institutes of Health, Research Triangle Park, NC. (in press) 

National Toxicology Program (NTP) (1998g). Toxicology and Carcinogenesis Studies of Methyleugenol 

(CAS No. 93-15-2) in F344/N Rats and B6C3F Mice (Gavage Studies). Technical Report Series No. 491. 1 

NIH Publication No. 98-3950. U.S. Department of Health and Human Services, Public Health Service, 

National Institutes of Health, Research Triangle Park, NC. (in preparation) 

National Toxicology Program (NTP) (1998h). Toxicology and Carcinogenesis Studies of Diethanolamine 

(CAS No. 111-42-2) in F344/N Rats and B6C3F Mice (Dermal Studies). Technical Report Series No. 478. 1 

NIH Publication No. 98-3968. U.S. Department of Health and Human Services, Public Health Service, 

National Institutes of Health, Research Triangle Park, NC. (in press) 

Nomura, A., Stemmermann, G.N., Chyou, P., Kato, I., Perez-Perez, G.I., and Blaser, M.J. (1991). 

Helicobacter pylori infection and gastric carcinoma among Japanese Americans in Hawaii. N. Engl. J. Med. 

325, 1132-1136. 

Nyska, A., Maronpot, R.R., Eldridge, S.R., Haseman, J.K., and Hailey, J.R. (1997). Alteration in cell 

kinetics in control B6C3F mice infected with Helicobacter hepaticus. Toxicol. Pathol. 25, 591-596. 1 

Parsonnet, J., Friedman, G.D., Vandersteen, D.P., Chang, Y., Vogelman, J.H., Orentreich, N., and Sibley, 

R.K. (1991). Helicobacter pylori infection and the risk of gastric carcinoma. N. Engl. J. Med. 325, 

1127-1131. 

Rao, G.N., Hickman, R.L., Seilkop, S.K., and Boorman, G.A. (1987). Utero-ovarian infection in aged
 

B6C3F1 mice. Lab. Animal Sci. 37, 153-158.
 

Riley, L.K., Franklin, C.L., Hook, R.R., Jr., and Besch-Williford, C. (1996). Identification of murine
 

Helicobacters by PCR and restriction enzyme analyses. J. Clin. Microbiol. 34, 942-946.
 

Seilkop, S.K. (1995). The effect of body weight on tumor incidence and carcinogenicity testing in B6C3F1
 

mice and F344 rats. Fundam. Appl. Toxicol. 24, 247-259.
 

Shames, B., Fox, J.G., Dewhirst, F., Yan, L., Shen, Z., and Taylor, N.S. (1995). Identification of
 

widespread Helicobacter hepaticus infection in feces in commercial mouse colonies by culture and PCR
 

assay. J. Clin. Microbiol. 33, 2968-2972.
 

Sills, R.C., Hong, H.L., Greenwell, A., Herbert, R.A., Boorman, G.A., and Devereux, T.R. (1995). 

Increased frequency of K-ras mutations in lung neoplasms from female B6C3F1 mice exposed to ozone for
 

24 or 30 months. Carcinogenesis 16, 1623-1628.
 

Sipowicz, M.A., Weghorst, C.M., Shio, Y.-H., Buzard, G.S., Calvert, R.J., Anver, M.R.,
 

Anderson, L.M., and Rice, J.M. (1997). Lack of p53 and ras mutations in Helicobacter hepaticus-induced
 

liver tumors in A/JCr mice. Carcinogenesis 18, 233-236.
 
Cobalt Sulfate Heptahydrate, NTP TR 471 255 

Taylor, N.S., Fox, J.G., and Yan, L. (1995). In-vitro hepatotoxic factor in Helicobacter hepaticus, 

H. pylori and other Helicobacter species. J. Med. Microbiol. 42, 48-52. 

Ward, J.M., Anver, M.R., Haines, D.C., Tully, J.G., Jr., Collins, M.J., Jr., Gorelick, P.L., Anderson, L., 

Rice, J.M., and Russell, R.J. (1993). A unique hepatitis in mice associated with a helical bacterium. 

Toxicol. Pathol. 21, 591. (Abstr.) 

Ward, J.M., Fox, J.G., Anver, M.R., Haines, D.C., George, C.V., Collins, M.J., Jr., Gorelick, P.L., 

Nagashima, K., Gonda, M.A., Gilden, R.V., Tully, J.G., Russell, R.J., Benveniste, R.E., Paster, B.J., 

Dewhirst, F.E., Donovan, J.C., Anderson, L.M., and Rice, J.M. (1994a). Chronic active hepatitis and 

associated liver tumors in mice caused by a persistent bacterial infection with a novel Helicobacter species. 

J. Natl. Cancer Inst. 86, 1222-1227. 

Ward, J.M., Anver, M.R., Haines, D.C., and Benveniste, R.E. (1994b). Chronic active hepatitis in mice 

caused by Helicobacter hepaticus. Am. J. Pathol. 145, 959-968. 

Ward, J.M., Benveniste, R.E., Fox, C.H., Battles, J.K., Gonda, M.A., and Tully, J.G. (1996). 

Autoimmunity in chronic active Helicobacter hepatitis of mice. Am. J. Pathol. 148, 509-517. 

Wotherspoon, A.C., Doglioni, C., Diss, T.C., Pan, L., Moschini, A., de Boni, M., and Isaacson, P.G. 

(1993). Regression of primary low-grade B-cell gastric lymphoma of mucosa-associated lymphoid tissue 

type after eradication of Helicobacter pylori. Lancet 342, 575-577. 
256 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE J1 

Incidence of Helicobacter hepaticus-Associated Hepatitis in Control B6C3F Mice 1 

from Nine NTP 2-Year Studiesa 

Incidence of Hepatitis (%) 

Study Males Females 

Sodium xylenesulfonate 78 4 

AZT/5,000 U "-interferon A/D 76 4 

Cobalt sulfate heptahydrate 72 8 

AZT/500 U "-interferon A/D 66 0 

Chloroprene 54 0 

Theophylline 32 0 

"-Interferon A/D 22 4 

Triethanolamine 20 0 

AZT 16 2 

Average 48 2 

a Includes regeneration and mild to marked (excludes minimal) chronic inflammation, karyomegaly, oval cell hyperplasia, and bile duct 

hyperplasia. AZTù3N-azido-3N-deoxythymidine 

TABLE J2 

Identification of Helicobacter hepaticus with PCR-RFLP-Based Assays in Control B6C3F Mice 1 

from 32 NTP 2-Year Studies and Three NTP 13-Week Studiesa 

H. hepaticus-Positive Studiesb 

Type of Sample Total Affected Unaffected 

Studies Studies Studies 

13-Week Studies 

Formalin-fixed liver 3 — 1/3c 

2-Year Studies 

Frozen liver 

Formalin-fixed liver 

22 

10 

3/3 

1/6c 3/19 0/4 

a PCR-RFLPùpolymerase chain reaction-restriction fragment length polymorphism b	 Number of H. hepaticus-positive studies/number of affected or unaffected studies. Affected studies are those in which hepatitis typical of 

that associated with H. hepaticus infection occurred in many male mice. 

Only one animal in the positive study was positive for H. hepaticus. c 
Cobalt Sulfate Heptahydrate, NTP TR 471 257 

TABLE J3 

Comparison of Neoplasm Incidences in Control B6C3F1 Mice

from Helicobacter hepaticus-Affected and Unaffected NTP 2-Year Studies 

Males Females 

Affected Unaffected Affected Unaffected 

Studiesa Studies Studies Studies 

Number of studies 9 26 9 26 

Survival (%) 64 71 68 68 

12-Month body wt (g) 48.0 48.3 48.1 47.0 

Neoplasm incidence (%) 

Liver 71.3* 54.8 50.3 40.5 

Lung 26.6 23.2 7.6 10.3 

Pituitary gland 0.4 0.8 14.7 14.3 

Harderian gland 5.6 6.1 6.0 4.9 

Lymphoma 6.9 6.3 16.2 15.5 

Circulatory system 9.8 6.0 5.3 4.7

 liver only 7.1* 2.5 — — 

All benign 61.8 57.2 59.1 54.6 

All malignant 61.3* 40.9 50.0 44.2 

All neoplasms 88.0* 77.4 82.7 75.4 

* Significantly different (P#0.05) from the unaffected studies a Affected studies are those in which hepatitis typical of that associated with H. hepaticus infection occurred in many male mice. 
258 Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE J4 

Liver Neoplasm Incidences and Body Weights of Control B6C3F Mice 1 

in Relation to Study Start Dates of Helicobacter hepaticus-Affected and Unaffected NTP 2-Year Studiesa 

Liver Neoplasm Incidence (%) Mean Body Weight (g) 

Study Start Date Affected Unaffected Affected Unaffected a Studies Studies Studies Studies 

Male 

April to September 1988 — 43.8 (8)b — 46.2 (8) 

October 1988 62.0 (1) — 48.3 (1) — 

November 1988 to September 1989 — 52.6 (7) — 48.7 (7) 

October 1989 to June 1990 — 61.2 (5) — 48.9 (5) 

July 1990 to January 1991 72.5 (8) 66.2 (4) 48.0 (8) 49.0 (4) 

February 1991 to April 1992 — 68.0 (2) — 52.8 (2) 

Average 71.3 54.8 48.0 48.3 

Female 

April to September 1988 — 31.1 (8) — 44.8 (8) 

October 1988 46.0 (1) — 46.4 (1) — 

November 1988 to September 1989 — 39.9 (7) — 47.2 (7) 

October 1989 to June 1990 — 38.6 (5) — 45.9 (5) 

July 1990 to January 1991 50.9 (8) 54.2 (4) 48.3 (8) 48.0 (4) 

February 1991 to April 1992 — 58.0 (2) — 55.6 (2) 

Average 50.3 40.5 48.1 47.0 

a Includes nine affected studies (those in which hepatitis typical of that associated with H. hepaticus infection occurred in many male mice) 

and 26 unaffected studies 

b Number of studies is given in parentheses. 
Cobalt Sulfate Heptahydrate, NTP TR 471 259 

TABLE J5 

Association of Liver Neoplasm Incidence and Severity of Helicobacter hepaticus-Associated Hepatitis a in Control B6C3F Mice from Nine Affected NTP 2-Year Studies 1 

Liver Neoplasm Incidence 

Severity of Hepatitis Males Females 

Absent 

Minimal 

Mild/moderate 

Significance of association 

101/175 (58%) 

44/57 (77%) 

176/218 (81%) 

P<0.05 

196/396 (49%) 

23/42 (55%) 

7/11 (64%) 

NSb 

a Affected studies are those in which hepatitis typical of that associated with H. hepaticus infection occurred in many male mice. b NSùnot significant 

TABLE J6 

H-ras Codon 61 AAA Mutations in Spontaneous Liver Neoplasms in Control B6C3F Mice 1 

from Helicobacter hepaticus-Affected and Unaffected NTP 2-Year Studies 

a Study Affected H-ras AAA Mutations 

Male 

Cobalt sulfate heptahydrate + 0/10 (0%) 

Chloroprene + 1/13 (8%) 

Triethanolamine + 1/10 (10%) 

Oxazepam — 7/18 (39%) 

Diethanolamine — 4/16 (25%) 

Historical control database 106/333 (32%) 

Female 

Chloroprene + 0/10 (0%) 

Triethanolamine + 1/15 (7%) 

Diethanolamine — 1/11 (9%) 

Historical control database 106/333 (32%) 

a +ùaffected; — ùnot affected. Affected studies are those in which hepatitis typical of that associated with H. hepaticus infection occurred 

in many male mice. 
260 

c 

Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE J7 a Proliferating Cell Nuclear Antigen Labeling Indices in the Liver of Control B6C3F1 Mice

Hepatitis No. of PCNA Labeling Average PCNA 

Animals Indexb Labeling Indexc 

Male 

Cobalt sulfate heptahydrated 

Chloroprened 

Triethanolamined 

+ 

+ 

+ 

15 

12 

9 

0.535 ± 0.129 

1.452 ± 0.386 

1.215 ± 0.374 1.011 

Cobalt sulfate heptahydrate 

Chloroprene 

Triethanolamine 

— 

— 

— 

7 

10 

12 

0.175 ± 0.117 

0.296 ± 0.124 

0.100 ± 0.042 0.186 

1-Trans-delta -tetrahydrocannabinol 9 e 

Scopolamine hydrobromide trihydratef 

Methyleugenolf 

Mouse life-span studyf 

— 

— 

— 

— 

15 

14 

14 

15 

0.042 ± 0.011 

0.043 ± 0.012 

0.077 ± 0.020 

0.217 ± 0.880 

Female 

Cobalt sulfate heptahydrate + 5 0.161 ± 0.062 

Cobalt sulfate heptahydrate 

Chloroprene 

Triethanolamine 

— 

— 

— 

17 

12 

12 

0.055 ± 0.015 

0.154 ± 0.050 

0.138 ± 0.053 0.108 

1-Trans-delta -tetrahydrocannabinol 9 — 13 0.156 ± 0.047 

Scopolamine hydrobromide trihydrate — 15 0.032 ± 0.009 

a A portion of these data are presented in Nyska et al. (1997). +ùhepatitis present; — ùno hepatitis present b Mean ± standard error; PCNAùproliferating cell nuclear antigen
 

Average of the mean labeling indices for animals from all three studies d	 Affected study (one in which hepatitis typical of that associated with H. hepaticus occurred in many male mice) e Unaffected study (one in which the typical hepatitis did not occur in mice) f	 Unaffected study with no typical hepatitis, but positive for H. hepaticus by polymerase chain reaction-restriction fragment length 

polymorphism-based assay 
261 

c 

Cobalt Sulfate Heptahydrate, NTP TR 471 

TABLE J8 

Summary of Target Sites of Carcinogenicity in B6C3F Mice from NTP 2-Year Studies 1 

with Helicobacter hepaticus-Associated Hepatitis 

Males	 Females 

Chloroprene Lung 

Circulatory systema Lung Circulatory system 

Harderian gland Harderian gland 

Forestomach Forestomach 

Kidney Liver 

Skin 

Mesentery 

Zymbal’s gland 

Mammary gland 

Cobalt sulfate heptahydrateb Lung Lung 

Triethanolamine Liver Liver 

AZTc None Vagina 

Sodium xylenesulfonate None None 

Theophylline None None 

a Hemangioma and hemangiosarcoma of the liver were excluded from the analysis in males. b	 An apparent treatment-related increase in the incidence of hemangiosarcoma of the liver was discounted in male mice because of the 

presence of H. hepaticus. 

AZTù3N-azido-3N-deoxythymidine. Includes four studies: AZT; "-interferon A/D; AZT/500 U "-interferon A/D; and 

AZT/5,000 U "-interferon A/D 
262 Cobalt Sulfate Heptahydrate, NTP TR 471