Abstract for TR-471

Cobalt sulfate is used in the electroplating and electro chemical 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 B6C3F1 mice were exposed to cobalt sulfate 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).

Two-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 3.0 mg/m3 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/m3, the 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 1.0 mg/m3 and one 3.0 mg/m3 female.

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.

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 were increased in 3.0 mg/m3 males and females.

Two-year study in mice

Groups of 50 male and 50 female mice 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 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 tration in 3.0 mg/m3 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 in 1.0 and 3.0 mg/m3 males and females and hyper plasia of the olfactory epithelium in 3.0 mg/m3 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 Helico bacter hepaticus. In NTP studies with H. hepaticus- associated hepatitis, increased incidences of hemangiosarcoma 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 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 neo-plasms 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.

Studies

Summary of the Two-year Carcinogenesis and Genetic Toxicology Studies of Cobalt Sulfate Heptahydrate
	Male F344/N Rats	Female F344/N Rats	Male B6C3F1 Mice	Female B6C3F1 Mice
Doses	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
Body weights	Exposed groups similar to chamber controls	Exposed groups similar to chamber controls	3.0 mg/m3 group slightly lower than chamber controls	Exposed groups slightly higher than controls
Survival rates	17/50, 15/50, 21/50,15/50	28/50, 25/49, 26/50, 30/50	22/50, 31/50, 24/50,20/50	34/50, 37/50, 32/50, 28/50
Nonneoplastic effects	Lung: proteinosis (0/50, 16/50, 40/48, 47/50); alveolar epithelial metaplasia (0/50, 50/50, 48/48, 49/50); granulomatous alveolar inflammation (2/50, 50/50, 48/48, 50/50); interstitial fibrosis (1/50, 50/50, 48/48, 49/50); alveolar epithelial hyperplasia (9/50, 20/50, 20/48, 23/50) Nose: lateral wall hyperplasia (2/50, 14/50, 21/49, 20/50); olfactory epithelial atrophy (8/50, 24/50, 42/49, 48/50); lateral wall squamous metaplasia (1/50, 3/50, 5/49, 8/50); olfactory epithelial metaplasia (5/50, 1/50, 5/49, 30/50) Larynx: epiglottis squamous metaplasia (0/50, 10/49, 37/48, 50/50)	Lung: proteinosis (0/50, 36/49, 49/50, 49/50); alveolar epithelial metaplasia (2/50, 47/49, 50/50, 49/50); granulomatous alveolar inflammation (9/50, 47/49, 50/50, 49/50); interstitial fibrosis (7/50, 47/49, 50/50, 49/50); alveolar epithelial hyperplasia (15/50, 7/49, 20/50, 33/50); squamous metaplasia (0/50, 1/49, 8/50, 3/50); atypical alveolar epithelial hyperplasia (0/50, 0/49, 3/50, 5/50) Nose: lateral wall hyperplasia (1/50, 8/49, 26/50, 38/50); olfactory epithelial atrophy (5/50, 29/49, 46/50, 47/50); lateral wall squamous metaplasia (1/50, 1/49, 4/50, 10/50); olfactory epithelial metaplasia (2/50, 2/49, 3/50, 40/50) Larynx: epiglottis squamous metaplasia (1/50, 22/49, 39/50, 48/50)	Lung: diffuse histiocytic cell infiltrate (1/50, 2/50, 4/50, 10/50) Nose: olfactory epithelial atrophy (0/50, 0/50, 29/48, 48/49); olfactory epithelial hyperplasia (0/50, 0/50, 0/48, 10/49) Larynx: squamous metaplasia (0/48, 37/49, 48/48, 44/49)	Lung: focal histiocytic cell infiltrate (2/50, 5/50, 7/50, 10/50) Nose: olfactory epithelial atrophy (0/50, 2/50, 12/49, 46/48); olfactory epithelial hyperplasia (0/50, 0/50, 0/49, 30/48) Larynx: squamous metaplasia (0/50, 45/49, 40/47, 50/50)
Neoplastic effects	Lung: alveolar/bronchiolar adenoma (1/50, 4/50, 1/48, 6/50); alveolar/bronchiolar carcinoma (0/50, 0/50, 3/48, 1/50); alveolar/bronchiolar adenoma or carcinoma (1/50, 4/50, 4/48, 7/50)	Lung: alveolar/bronchiolar adenoma (0/50, 1/49, 10/50, 9/50); alveolar/bronchiolar carcinoma (0/50, 2/49, 6/50, 6/50); alveolar/bronchiolar adenoma, alveolar/bronchiolar carcinoma, or 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)	Lung: alveolar/bronchiolar adenoma (9/50, 12/50, 13/50, 18/50); alveolar/bronchiolar carcinoma (4/50, 5/50, 7/50, 11/50); alveolar/bronchiolar adenoma or carcinoma (11/50, 14/50, 19/50, 28/50)	Lung: alveolar/bronchiolar adenoma (3/50, 6/50, 9/50, 10/50); alveolar/bronchiolar carcinoma (1/50, 1/50, 4/50, 9/50); alveolar/bronchiolar adenoma or carcinoma (4/50, 7/50, 13/50, 18/50)
Uncertain findings	Adrenal medulla: benign, complex, or malignant pheochromocytoma (15/50, 19/50, 25/49, 20/50)	None	None	None
Levels of evidence of carcinogenic activity	Some evidence	Clear evidence	Clear evidence	Clear evidence

Two-year Genetic Toxicology Studies of Cobalt Sulfate Heptahydrate
Assay	Results
Salmonella typhimurium gene mutations:	Positive in strain TA100 with and without S9 Negative in strains TA98 and TA1535 with and without S9