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PubMed1994

NTP Toxicology and Carcinogenesis Studies of Hexachlorocyclopentadiene (CAS No. 77-47-4) in F344/N Rats and B6C3F1 Mice (Inhalation Studies).

Hexachlorocyclopentadiene is an intermediate used in the manufacture of flame retardants, resins, and chlorinated cyclodiene pesticides. Toxicology and carcinogenesis studies were conducted by exposing male and female F344/N rats and B6C3F1 mice to atmospheres containing hexachlorocyclopentadiene (approximately 98% pure) for 6 hours per day, 5 days per week, for 13 weeks or 2 years. A stop-exposure evaluation was conducted in male B6C3F1 mice to determine the influence of exposure level and exposure duration on the development of nonneoplastic lesions of the respiratory tract and on their regression or progression after exposure was stopped. Genetic toxicology studies were conducted in Salmonella typhimunum, cultured Chinese hamster ovary cells, Drosophila melanogaster, and mouse peripheral blood samples were analyzed for frequency of micronucleated normochromatic erythrocytes. 13-WEEK STUDY IN RATS: Groups of 10 male and 10 female rats were exposed to atmospheres containing 0, 0.04, 0.15, 0.4, 1, or 2 ppm (equivalent to 0, 0.45, 1.67, 4.46, 11.14, and 22.28 mg/m(3)) hexachlorocyclopentadiene. Additional rats were exposed to 0, 0.04, 0.4, or 2 ppm hexachlorocyclopentadiene and evaluated for differences in clinical pathology parameters. All rats in the 1 and 2 ppm groups died during the first 4 weeks of the study. The final mean body weight and mean body weight gain of males exposed to 0.4 ppm were significantly lower than those of the controls. Listlessness was observed in 2 ppm rats from week 1, in 1 ppm rats from week 2, and in 0.4 ppm rats during week 3. Rats exposed to 1 or 2 ppm also experienced respiratory distress. No chemical-related differences in hematology, clinical chemistry, or urinalysis parameters were observed in male or female rats. Absolute and relative lung weights of 0.4 ppm males were significantly greater than those of the controls. Inflammation (necrotizing, chronic, or suppurative) of the nose, larynx, trachea, and lung was observed in 0.4, 1, and 2 ppm males and females. Squamous metaplasia of the epithelial lining of the nose of 0.4 ppm males and 1 and 2 ppm males and females was also observed. 13-WEEK STUDY IN MICE: Groups of 10 male and 10 female mice were exposed to atmospheres containing 0, 0.04, 0.15, 0.4, 1, or 2 ppm (equivalent to 0, 0.45, 1.67, 4.46, 11.14, and 22.28 mg/m(3)) hexachlorocyclopentadiene. Additional mice were exposed to 0, 0.04, 0.4, or 2 ppm and evaluated for differences in clinical pathology parameters. All 2 ppm mice died during the first week of exposure. All 1 ppm mice died during the first 5 weeks of exposure. Five males and two females in the 0.4 ppm group died during the first 2 weeks of exposure. Deaths in the other groups were not related to hexachlorocyclopentadiene exposure. Final mean body weights of males exposed to 0.15 and 0.4 ppm and the body weight gain of 0.4 ppm males were significantly lower than those of the controls. Treatment-related clinical findings included listlessness in 0.4 and 1 ppm males and females. No chemical-related differences in hematology, clinical chemistry, or urinalysis parameters were observed in male or female mice. Necrosis or inflammation of the nose, larynx, trachea, or lung occurred in mice exposed to 0.4,1, and 2 ppm hexachlorocyclopentadiene. Squamous metaplasia of the larynx or trachea was observed in 0.15, 0.4, and 1 ppm males and in 0.4 and 1 ppm females. 2-YEAR STUDY IN RATS: Survival, Body Weights, Clinical Findings, and Urinalysis Groups of 60 male and 60 female rats were exposed to atmospheres containing 0, 0.01, 0.05, or 0.2 ppm (equivalent to 0, 0.11, 0.56, and 2.28 mg/m(3)) hexachlorocyclopentadiene. Survival rates and mean body weights of exposed rats were similar to those of the controls. No chemical-related clinical findings were observed in male or female rats during the 2-year study. No differences in urinalysis parameters at the 15-month interim evaluation could be attributed to exposure to hexachlorocyclopentadiene. Pathology Findings: No increases in neoplasm incidences could be attributed to hexachlorocyclorocyclopentadiene. Toxicity was limited to the respiratory tract and included an increase in the incidence of pigmentation of the respiratory epithelium of the nose, trachea, and the bronchi and bronchioles of the lung in both males and females. Exposure to hexachlorocyclopentadiene also caused an increase in the incidence of squamous metaplasia of the laryngeal epithelium of exposed females; the incidences in 0.01 and 0.2 ppm females were significantly greater than that of the controls. The severity of squamous metaplasia was minimal in all exposed and control females. 2-YEAR STUDY IN MICE: Survival, Body Weights, Clinical Findings, and Urinalysis Groups of 60 male and 60 female mice were exposed to atmospheres containing 0, 0.01, 0.05, or 0.2 ppm (equivalent to 0, 0.11, 0.56, and 2.28 mg/m(3)) hexachlorocyclopentadiene. The 2-year survival rate of female mice in the 0.2 ppm group was marginally lower than that of the controls due to a higher incidence of ovarian inflammation in 0.2 ppm females. Mean body weights of 0.2 ppm males (weeks 62 to 103) and females (throughout the study) were lower than those of the controls. No clinical findings in male or female mice were attributed to chemical exposure during the 2-year study. There were no chemical-related differences in urinalysis parameters at the 15-month interim evaluation. Pathology Findings: The site of toxicity of hexachlorocyclopentadiene exposure in mice in the 2-year study was the respiratory tract. Chemical-related pigmentation of the respiratory epithelium of the nose, trachea, and lung and suppurative inflammation of the nose were observed. No increased neoplasm incidences in males or females could be attributed to hexachlorocyclopentadiene exposure. STOP-EXPOSURE EVALUATION: Survival, Body Weights, and Clinical Findings Groups of male mice were exposed to atmospheres containing 0.2 ppm hexachlorocyclopentadiene for 33 or 66 weeks or 0.5 ppm for 26 or 42 weeks followed by exposure to air until the end of the study. Fifty male mice from each stop-exposure group were evaluated at 2 years. Two-year survival rates of stop exposure groups were similar to that of the controls. Final mean body weights of stop-exposure groups were similar to that of the controls. No chemical related clinical findings were observed. Pathology Findings: Nonneoplastic respiratory tract lesions similar to those observed in the core study were observed in males in the stop-exposure groups. Chemical-related pigmentation and inflammation of the respiratory epithelium were persistent as indicated by their presence in many male mice after recovery periods of 62 to 78 weeks, and the incidence and severity of the lesions were related to exposure concentration and duration. GENETIC TOXICOLOGY: Hexachlorocyclopentadiene was not mutagenic in Salmonella typhimurium strains TA98, TA100, TA1535, and TA1537 when tested with and without S9. Hexachlorocyclopentadiene did induce sister chromatid exchanges and chromosomal aberrations in cultured Chinese hamster ovary cells, with and without S9. No induction of sex-linked recessive lethal mutations was observed in male Drosophila melanogaster treated with hexachlorocyclopentadiene by feeding or injection, and no increase in the frequency of micronucleated erythrocytes was seen in male or female B6C3F1 mice exposed to hexachlorocyclopentadiene by inhalation for 13 weeks. CONCLUSIONS: Under the conditions of these 2-year studies, there was no evidence of carcinogenic activity of hexachlorocyclopentadiene in male or female F344/N rats or B6C3F1 mice exposed to 0.01, 0.05, or 0.2 ppm. Exposure of rats to hexachlorocyclopentadiene produced pigmentation of the respiratory epithelium of the nose, trachea (males), and bronchi and bronchioles of the lung. Squamous metaplasia of the laryngeal epithelium occurred in female rats exposed to hexachlorocyclopentadiene. Suppurative inflammation of the nose as well as pigmentation of the respiratory mucosal epithelium occurred in mice exposed to hexachlorocyclopentadiene. Synonyms: Perchlorocyclopentadiene, hexachloro-1,3-cyclopentadiene, HEX, HCPD, HCCP, HCCPD Trade Name: C-56-Graphlox

Journal Article↗
PubMed1994

NTP Toxicology and Carcinogenesis Studies of 4,4'-Thiobis(6- t -butyl- m -cresol) (CAS No. 96-69-5) in F344/N Rats and B6C3F1 Mice (Feed Studies).

4,4'-Thiobis(6- t -butyl- m -cresol) (TBBC) is used in the rubber and plastics industries as an antioxidant. TBBC is also used as a stabilizer in polyethylene and polyolefin packaging materials for foodstuffs. Toxicology and carcinogenesis studies were conducted by administering TBBC (99% pure) in feed to groups of male and female F344/N rats and B6C3F1 mice for 15 days, 13 weeks, and 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium and cultured Chinese hamster ovary cells. 15-DAY STUDY IN RATS: Groups of 10 male and 10 female F344/N rats were fed diets containing 0, 1,000, 2,500, 5,000, 10,000 or 25,000 ppm TBBC for 15 days. Rats given to 1,000, 2,500, 5,000, or 10,000 ppm received approximate doses of 95, 235, 335, or 365 mg TBBC per kilogram body weight per day (males) or 85, 220, 325, or 270 mg/kg per day (females). Approximate doses for rats receiving 25,000 ppm could not be calculated due to early deaths. All 25,000 ppm rats and three male and four female 10,000 ppm rats died. Surviving rats in the 10,000 ppm groups had a significant weight loss and the final mean body weights of 5,000 and 10,000 ppm male and female rats were significantly lower than those of the controls. Male and female rats exposed to 5,000, 10,000, or 25,000 ppm TBBC consumed markedly less feed than the controls. Diarrhea occurred in 5,000, 10,000, and 25,000 ppm males and females. The principal lesions attributed to the administration of TBBC were renal papillary and tubule necroses which occurred in 10,000 ppm rats. Focal necrosis or erosions of the glandular stomach also occurred in some 10,000 ppm rats. Changes observed in the thymus and spleen were attributed to debilitation or stress; bone marrow depletion was attributed to nutrient deficiency accompanying weight loss. 15-DAY STUDY IN MICE: Groups of 10 male and 10 female B6C3F1, mice were fed diets containing 0, 1,000, 2,500, 5,000, 10,000, or 25,000 ppm TBBC for 15 days. Mice given 1,000, 2,500, or 5,000 ppm received approximate doses of 285, 585, or 475 mg TBBC per kilogram body weight per day (males) or 360, 950, or 1,030 mg/kg per day (females). Approximate doses for mice given 10,000 or 25,000 ppm could not be calculated due to early deaths. All 10,000 and 25,000 ppm mice died, as did eight males and eight females given 5,000 ppm. A significant weight loss occurred in surviving 5,000 ppm males and females and the final mean body weights of 2,500 ppm females and 5,000 ppm males and females were significantly lower than those of the controls. Feed consumption by mice given 5,000, 10,000, or 25,000 ppm was markedly reduced. Diarrhea occurred in all 25,000 ppm mice and in most male and female mice given 5,000 or 10,000 ppm. Renal tubule necrosis occurred in eight males and three females in the 5,000 ppm groups. Lymphocytic depletion of Iymphoid tissues in many 5,000 ppm males and females was attributed to debilitation and stress or to nutrient deficiency accompanying weight loss. 13-WEEK STUDY IN RATS: Groups of 10 male and 10 female F344/N rats were fed diets containing 0, 250, 500, 1,000, 2,500, or 5,000 ppm TBBC for 13 weeks. These exposure levels delivered approximate doses of 15, 30, 60, 165, or 315 mg TBBC per kilogram body weight per day (males) or 15, 35, 70, 170, or 325 mg/kg per day (females). All rats survived to the end of the study. The final mean body weight of 5,000 ppm males was 40% lower than that of the controls; the final mean body weight of 5,000 ppm females was 27% lower than that of the controls. Feed consumption by male and female rats exposed to 5,000 ppm TBBC was markedly lower than that by the controls throughout the study. The absolute and relative liver weights of 5,000 ppm females were significantly greater than those of the controls. Serum alkaline phosphatase (ALP) levels were significantly higher in 2,500 and 5,000 ppm males and slightly higher in 5,000 ppm females. Serum alanine aminotransferase levels were significantly higher in 2,500 and 5,000 ppm males and females. Hematocrit and hemoglobin concentrations and mean erythroions and mean erythrocyte volume (MCV) values were significantly lower in 1,000, 2,500, and 5,000 ppm males than in controls; MCV values were also significantly lower in 5,000 ppm females. A dose-related significant increase in forelimb and hindlimb grip strength was observed in exposed male and female rats. Histopathologic findings in the liver of 2,500 and 5,000 ppm males and females included hypertrophy of Kupffer cells, bile duct hyperplasia, and individual cell necrosis of hepatocytes; centrilobular hepatocyte hypertrophy also occurred in males and females exposed to 5,000 ppm TBBC. Macrophages were increased in size and number in the mesenteric Iymph nodes of males and females exposed to 5,000 ppm, and to a lesser extent in 2,500 ppm male and female rats. Pigmentation and degeneration of the renal cortical tubule epithelial cells was also present in males and females in the 2,500 and 5,000 ppm groups; cortical tubule necrosis occurred in 5,000 ppm males and females. 13-WEEK STUDY IN MICE: Groups of up to 10 male and 10 female B6C3F1 mice were fed diets containing 0, 100, 250, 500, 1,000, or 2,500 ppm TBBC for 13 weeks. These exposure levels delivered approximate doses of 15, 30, 65, 145, or 345 mg TBBC per kilogram body weight per day (males) or 10, 35, 60, 165, or 340 mg/kg per day (females). All mice survived to the end of the study. The final mean body weights of 2,500 ppm males and of 500,1,000, or 2,500 ppm females were significantly lower than those of the controls. Feed consumption by 2,500 ppm males averaged 24% lower than that by controls through week 3 and was similar to that by controls for the remainder of the study. Feed consumption by females receiving 2,500 ppm averaged 27% less than that by the controls during most of the study. The absolute and relative liver weights of males and females exposed to 2,500 ppm TBBC were slightly but significantly greater than those of the controls. Males exposed to 500, 1,000, or 2,500 ppm and females exposed to 2,500 ppm had significantly increased absolute and relative spleen weights. No clinical findings in mice were considered chemical related. Hematocrit concentrations and erythrocyte counts of males receiving 1,000 or 2,500 ppm were significantly less than those of the controls; hemoglobin concentration in males receiving 2,500 ppm was significantly less and mean erythrocyte volume was significantly less in males receiving 2,500 ppm. Females in the 1,000 and 2,500 ppm groups had significantly decreased hematocrit concentrations and erythrocyte counts; 2,500 ppm females also had significantly decreased hemoglobin concentrations and mean erythrocyte volumes. Kupffer cell hypertrophy, bile duct hyperplasia, and an increase in size and number of macrophages in mesenteric Iymph nodes were present in 2,500 ppm male and female mice. 2-YEAR STUDY IN RATS: Doses selected for the 2-year study of TBBC were based on the lower body weights and liver and kidney toxicity observed at 5,000 ppm in the 13-week study. Groups of 115 male and 75 female F344/N rats were fed diets containing 0, 500, 1,000, or 2,500 ppm TBBC for 2 years. Based on average daily feed consumption, these exposure levels resulted in a daily ingestion of TBBC of approximately 20, 40, or 100 mg/kg body weight for males and 20, 45, or 120 mg/kg body weight for females. Hematology, clinical chemistry, and urinalysis evaluations were performed on 15 male and 15 female rats from each group at 3, 9, and 15 months. Also at 15 months, an additional 10 male and 10 female rats from each group were evaluated for histopathology, hematology, and clinical chemistry. Forty male rats per group were evaluated for neurotoxic effects. Survival, Body Weights, Feed Consumption, and Clinical Findings: Two-year survival rates and mean body weights of exposed male and female rats were generally similar to those of the controls. The mean body weights of 2,500 ppm male rats were slightly lower than those of the controls throughout the study. At week 65, the mean body weight of 2,500 ppm females was 14% lower than that of the controls, but the final mean body weight of this group was 6% lower than that of the control group. Feed consumption, behavior, and general health and appearance of exposed male and female rats were similar to those of the controls. Hematology and Clinical Chemistry: Results of the hematology evaluation were not uniformly consistent at 3, 9, and 15 months in one set of rats, nor were they consistent between the two sets of rats evaluated at 15 months. Slight but significant decreases in hematocrit levels, hemoglobin concentrations, and erythrocyte counts were observed in the 1,000 and 2,500 ppm groups in one set of males at 15 months. Similar significant decreases in hematocrit level and hemoglobin concentration occurred in 2,500 ppm females at 9 months. Mean erythrocyte hemoglobin and mean erythrocyte hemoglobin concentration of 2,500 ppm females were also significantly lower than those of controls at 9 months and in both sets of female rats evaluated at 15 months. Platelet counts of 2,500 ppm male and female rats were slightly but significantly higher than those of controls at 3 and 9 months. Platelet counts were also slightly but significantly increased in 2,500 ppm males of one set evaluated at 15 months, and in 2,500 ppm females of the second set evaluated at 15 months. Serum activities of alkaline phosphatase, alanine aminotransferase, and sorbitol dehydrogenase in 2,500 ppm males were significantly greater than those in the controls at 3, 9, and 15 months. Alkaline phosphatase activities in both sets of 1,000 ppm males evaluated at 15 months were also significantly greater than those of controls. Serum activities of alanine aminotransferase and sorbitol dehydrogenase in 2,500 ppm females were also significantly greater than those in controls at 3, 9, and 15 months. Neurotoxicity Findings: There were no significant inhibitory effects of TBBC on motor nerve excitability or conduction, neuromuscular transmission, or muscle contractility. There were no microscopic lesions in the sciatic nerve, quadriceps muscle, or teased nerve preparations of sciatic nerve that could be attributed to TBBC administration. Pathology Findings: At the 15-month interim evaluation, the absolute and relative liver weights of 2,500 ppm female rats were significantly greater than those of controls; at 15 months and at the end of the study, the incidences of Kupffer cell hypertrophy, hepatocyte cytoplasmic vacuolization, and mixed cell foci were also significantly increased. At the end of the study, the incidence of hepatocellular fatty change was significantly increased in 2,500 ppm females. The incidence of Kupffer cell hypertrophy was significantly increased in 2,500 ppm males at 15 months and at 2 years; the incidence of cytoplasmic vacuolization was significantly increased in all exposed males at 15 months but only moderately increased in 1,000 and 2,500 ppm males at 2 years; the incidence of basophilic foci was significantly increased in 2,500 ppm males at 15 months and the incidence of mixed cell foci was significantly increased in 1,000 and 2,500 ppm male rats at 2 years. The incidences of hepatocellular adenoma or carcinoma (combined) in exposed male rats were not significantly greater than that in the controls (0 ppm, 1/50; 500 ppm, 3/50; 1,000 ppm, 3/50; 2,500 ppm, 5/49), were within the historical control range, and were not considered chemical related. The severity of nephropathy was significantly increased in 2,500 ppm female rats. There was a significant negative trend in the incidence of mammary gland fibroadenoma, adenoma, or carcinoma (combined) in female rats (32/50, 24/50, 11/50, 16/50), and the incidences of fibroadenoma in 1,000 and 2,500 ppm females were significantly less than that of the controls. 2-YEAR STUDY IN MICE: Because of the reduction in body weights, the increase in liver and spleen weights, and the accompanying histopathologic changes in the liver of 2,500 ppm male and female mice in the 13-week study, the doses selected for the 2-year study were 250, 500, and 1,000 ppm. Groups of 80 male and 80 female mice were fed diets containing 0, 250, 500, or 1,000 ppm TBBC for 2 years. Based on average daily feed consumption, these exposure levels resulted in the daily ingestion of approximately 30, 60, or 145 mg TBBC/kg body weight for males and 45, 110, or 255 mg TBBC/kg body weight for females. Nine or 10 animals from each exposure group were evaluated at 3, 9, and 15 months. Survival, Body Weights, Feed Consumption, and Clinical Findings: Two-year survival rates of exposed male and female mice were similar to those of the controls. The final mean body weights of male and female mice exposed to 1,000 ppm were 8% and 18% lower than those of the controls, respectively. The final mean body weights of females exposed to 250 or 500 ppm were 8% to 9% lower than that of the controls. Feed consumption by exposed males was similar to that by controls, and there were no clinical findings attributed to TBBC administration. Hematology and Clinical Chemistry: Hematocrit level, hemoglobin concentration, and erythrocyte count in 1,000 ppm male mice were significantly lower than those in controls at the 15-month interim evaluation. Serum alkaline phosphatase activities in 1,000 ppm males were slightly but significantly greater than those in controls at 3 and 9 months, as was the serum alkaline phosphatase activity in 1,000 ppm females at 9 months. Serum levels of total bilirubin in all exposed groups of males were significantly greater than those in controls at 9 and 15 months. Pathology Findings: In the liver of male mice, negative trends in the incidences of fatty change, clear cell foci, and adenoma or carcinoma combined occurred at the end of the 2-year study. There were no compound-related increased incidences of neoplasms or nonneoplastic lesions in mice receiving TBBC for 2 years. A negative trend in the incidence of fatty change in the liver of male mice also occurred at 15 months. GENETIC TOXICOLOGY: 4,4'-Thiobis(6- t -butyl- m -cresol) was not mutagenic in Salmonella typhimurium strains TA98, TA100, TA1535, or TA1537 with or without exogenous metabolic activation (S9). Sister chromatid exchanges were induced in cultured Chinese hamster ovary cells treated with TBBC, with and without S9, but no increases in chromosomal aberrations were noted in cultured Chinese hamster ovary cells after treatment with TBBC. CONCLUSIONS: Under the conditions of these 2-year feed studies, there was no evidence of carcinogenic activity of 4,4'-thiobis(6- t -butyl- m -cresol) in male or female F344/N rats administered 500, 1,000, or 2,500 ppm or in male or female B6C3F1, mice administered 250, 500, or 1,000 ppm. Nonneoplastic lesions associated with exposure to TBBC included: Kupffer cell hypertrophy, cytoplasmic vacuolization, and mixed cell foci in the liver of male and female rats, fatty change in the liver of female, rats, and an increase in the severity of nephropathy in the kidney of female rats. In addition, decreased incidences of fibroadenoma, adenoma, or carcinoma (combined) were observed in the mammary gland of female rats. Decreases also occurred in the incidences of fatty change, clear cell foci, and adenoma or carcinoma (combined) in the liver of male mice. Synonyms: 4,4'-Thiobis(6- t -butyl-3-cresol); bis(3- t -butyl-4-hydroxy-6-methylphenyl)sulfide

Journal Article↗
PubMed1993

NTP Toxicology and Carcinogenesis Studies of Pentachloroanisole (CAS No. 1825-21-4) in F344 Rats and B6C3F1 Mice (Feed Studies).

Pentachloroanisole is a chlorinated aromatic compound which is widely distributed at low levels in the environment and in food products. Formation of pentachloroanisole in the environment may result from the degradation of structurally related, commercially important, ubiquitous chlorinated aromatic compounds such as pentachlorophenol and pentachloronitrobenzene which are known rodent toxins or carcinogens. Toxicology and carcinogenesis studies were conducted by administering pentachloroanisole (>99% pure) in corn oil by gavage to groups of male and female F344/N rats and B6C3F1 mice for 16 days, 13 weeks, or 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium strains, mouse lymphoma cells, and Chinese hamster ovary cells. 16-DAY STUDIES IN RATS: Groups of five male and five female F344/N rats were administered pentachloroanisole in corn oil by gavage once per day, 5 days per week, for 16 days at doses of 0, 100, 125, 150, 175, or 200 mg/kg body weight. Deaths occurred during days 2 and 3 in rats receiving doses of 125 mg/kg or greater; these deaths were considered directly related to pentachloroanisole administration. No biologically significant changes in mean body weight gains or final body weights were noted in the 100 mg/kg groups of rats. Because of the high early mortality rate, valid comparisons of body weight differences in other dose groups could not be made. Inactivity was noted in all dose groups. Rats administered doses of 125 mg/kg or greater also exhibited dyspnea. 16-DAY STUDIES IN MICE: Groups of five male and five female B6C3F1 mice were administered pentachloroanisole in corn oil by gavage once per day, 5 days per week, for 16 days at doses of 0, 100, 175, 250, 325, or 400 mg/kg. Deaths occurred during days 2 and 3 in mice receiving doses of 175 mg/kg or greater; these deaths were considered directly related to chemical administration. No biologically significant changes in mean body weight gains or final body weights were noted in 100 mg/kg males or 100 or 175 mg/kg females. Because of the high early mortality rate, valid comparisons of body weight differences in other dose groups could not be made. Inactivity was noted in dosed mice. 13-WEEK STUDIES IN RATS: Groups of 10 male and 10 female rats were administered pentachloroanisole in corn oil by gavage once per day, 5 days per week, for 13 weeks at doses of 0, 40, 80, 120, 140, or 180 mg/kg body weight. Most rats receiving doses of 120 mg/kg or greater died during the first week of the study as a direct result of pentachloroanisole administration. Mean body weight gains of males administered 40 or 80 mg/kg and of females administered 40, 80, or 120 mg/kg pentachloroanisole were significantly lower than those of the controls. Most dosed rats exhibited temporary inactivity for several hours after dosing. Relative liver and kidney weights of males administered 40 or 80 mg/kg and absolute and/or relative liver and kidney weights of females administered 40 to 120 mg/kg were significantly greater than those of the controls. Lesions observed in males administered 80 mg/kg or more and in females administered 120 mg/kg or more included pulmonary congestion, hemorrhage, and/or edema, meningeal congestion, and hepatocellular necrosis, glycogen depletion, and degeneration of biliary epithelium in the liver. 13-WEEK STUDIES IN MICE: Groups of 10 male and 10 female mice were administered pentachloroanisole in corn oil by gavage once per day, 5 days per week, for 13 weeks at doses of 0, 40, 80, 120, 140, or 180 mg/kg body weight. Most mice administered doses of 120 mg/kg or higher died during the first week of the study as a direct result of pentachloroanisole administration. Mean body weight gains of females administered 40 to 140 mg/kg were significantly greater than that of the controls, but those of dosed males were similar to that of the controls. Most dosed mice exhibited temporary inactivity for several hours after dosing. Absolute and relative liver weights of males administered 80 mg/kg, absolute and relative liver weights of femats of females administered 40 to 180 mg/kg, and absolute and relative kidney weights of females administered 80 to 180 mg/kg pentachloroanisole were also significantly greater than those of the controls. Lesions observed in males administered 40 mg/kg or more and in females administered 80 mg/kg or more included pulmonary congestion and/or edema, adrenal congestion, lymphoid depletion of lymph nodes and thymus, hepatocellular cytomegaly and karyomegaly, and pigment accumulation in hepatocytes and Kupffer cells. 2-YEAR STUDIES IN RATS: Based on the chemical-related mortality and liver lesions seen in the 16-day and 13-week studies, doses selected for the 2-year studies were 0, 10, 20, and 40 mg/kg for males and 0, 20, and 40 mg/kg for females. Groups of 70 male and 70 female rats were administered pentachloroanisole in corn oil by gavage 5 days per week for up to 2 years. At 9 and 15 months, up to 10 animals per group were selected for interim evaluations. Survival, Body Weights, and Clinical Findings: The survival of high-dose males was significantly decreased (vehicle control, 24/50; low-dose, 20/50; mid-dose, 24/50; high-dose, 14/50); most deaths in the high-dose group occurred at or before week 16. The majority of deaths in the mid- and high-dose groups may have been due to pentachloroanisole-related hyperthermia. The survival of dosed females was greater than that of the controls (29/50, 35/50, 44/50). Final mean body weights of mid- and high-dose males were 7% and 10% lower than that of the controls; final mean body weight of high-dose females was 11% lower than that of the controls. Final mean body weights of other dose groups were similar to those of the vehicle controls. At the 9-month interim evaluation, mean rectal temperature of males administered 40 mg/kg was significantly greater than that of the controls. Relative liver and kidney weights of males and females administered 20 or 40 mg/kg were significantly greater than those of controls. At the 15-month interim evaluation, relative liver weights of dosed females and absolute liver weights of 40 mg/kg females were significantly greater than those of the controls, as were relative liver and kidney weights of 40 mg/kg males. Pathology Findings: In the 2-year studies, administration of pentachloroanisole to males was associated with significant increases in the incidences of benign adrenal medulla pheochromocytomas. The incidence of benign adrenal medulla pheochromocytomas was marginally increased in high-dose females and slightly exceeded the range of the historical controls. Incidences of adrenal medulla hyperplasia were increased in dosed female rats, but not in dosed males. The incidences of pancreatic adenomas and focal hyperplasia were decreased in dosed males. The incidences of mammary gland fibroadenomas and uterine stromal polyps and sarcomas (combined) were decreased in high-dose females. Treatment-related increased incidences of intracytoplasmic pigmentation occurred in renal tubule epithelium, olfactory epithelium, and hepatocytes of males and females. Congestion and hemorrhage of the lungs, lymph nodes, thymus, adrenal cortex, and meninges, as well as hepatocellular centrilobular necrosis occurred almost exclusively in mid- and high-dose males that died or were killed moribund before the end of the studies. 2-YEAR STUDIES IN MICE: Based on the chemical-related mortality and liver lesions seen in the 16-day and 13-week studies, doses selected for the 2-year studies were 0, 20, and 40 mg/kg. Groups of 70 male and 70 female mice were administered pentachloroanisole in corn oil by gavage 5 days per week for up to 2 years. At 9 and 15 months, up to 10 animals per group were selected for interim evaluations. Survival, Body Weights, and Clinical Findings: The survival of dosed males was similar to that of the controls; survival of high-dose females was lower than that of the controls (24/50, 25/50, 16/50). The decreased survival of the high-dose females was attributed primarily to ovarian abscesses which were observed after moribund sacrifice. At the 9-month interim evaluation, the mean body weight of males administered 40 mg/kg was significantly lower than that of the vehicle controls. Absolute and relative liver weights of females and the relative liver weight of males administered 40 mg/kg were significantly greater than those of the controls. Final mean body weights of low- and high-dose males were 11% and 17% lower than that of the controls. Final mean body weights of dosed females were similar to that of the controls. There were no clinical findings attributed to pentachloroanisole administration. Pathology Findings: Centrilobular hepatocyte cytomegaly and pigment accumulation in hepatocytes and Kupffer cells were seen in dosed mice, but not in controls at the 9- and 15-month interim evaluations. In the 2-year studies, the incidence of benign pheochromocytomas was significantly increased in high-dose males. Dosed males also exhibited increased incidences of adrenal medulla hyperplasia and hypertrophy. The incidences of hemangiosarcomas of the liver were significantly increased in dosed males. Increased incidences of hepatocellular cytologic alteration, biliary tract hyperplasia, and Kupffer cell pigmentation occurred in dosed males and females; the incidences of mixed cell foci were also increased in dosed males. Cytologic alteration encompassed hepatocellular cytomegaly, karyomegaly, hepatocyte degeneration and necrosis, and multinucleated giant cell formation, and was considered an advanced stage of the pathologic process observed at 13 weeks. GENETIC TOXICOLOGY: Pentachloroanisole was mutagenic in Salmonella typhimurium strains TA98 and TA1537 in the absence but not in the presence of exogenous metabolic activation (S9). No clear mutagenic activity was observed in TA100 with hamster S9, without S9, or in TA1535 with or without S9. An equivocal response was observed in TA100 with rat S9. Pentachloroanisole was positive for induction of trifluorothymidine resistance in mouse lymphoma L5178Y cells with S9; the response observed without S9 was weak and inconsistent. In cytogenetic tests with Chinese hamster ovary cells, pentachloroanisole induced sister chromatid exchanges, but not chromosomal aberrations, with and without S9. TOXICOKINETICS: Male and female F344/N rats and B6C3F1 mice were administered 10, 20, or 40 mg/kg pentachloroanisole by gavage or 10 mg/kg pentachloroanisole intravenously (Appendix H). A rapid elimination of pentachloroanisole and a rapid formation of its main metabolite, pentachlorophenol, were seen in both species after an intravenous or an oral dose of pentachloroanisole. The area under the concentration-versus-time curve of pentachloroanisole increased with dosage in each species but the dose proportionality was lost above 20 mg/kg. No sex-related differences were found in the rate of absorption of pentachloroanisole from the GI tract, in the area under the concentration-versus-time curve, or in the overall rate elimination of pentachloroanisole. However, in female rats the area under the concentration-versus-time curve of pentachlorophenol was significantly larger than in male rats. No such difference was observed in mice. CONCLUSIONS: Under the conditions of these 2-year gavage studies, there was some evidence of carcinogenic activity of pentachloroanisole in male F344/N rats based on increased incidences of benign pheochromocytomas of the adrenal medulla. There was equivocal evidence of carcinogenic activity of pentachloroanisole in female F344/N rats based on marginally increased incidences of benign pheochromocytomas of the adrenal medulla. There was some evidence of carcinogenic activity of pentachloroanisole in male B6C3F1 mice based on increased incidences of benign pheochromocytomas of the adrenal medulla and hemangiosarcomas of the liver. There was no evidence of carcinogenic activity of pentachloroanisole in female B6C3F1 mice given doses of 20 or 40 mg/kg. Pentachloroanisole administration was associated with increased incidences of adrenal medulla hyperplasia in female rats and increased incidences of pigmentation in the renal tubule epithelium, olfactory epithelium, and hepatocytes of male and female rats. In addition, decreased incidences of pancreatic adenomas and focal hyperplasia in male rats and decreased incidences of mammary gland fibroadenomas and uterine stromal polyps and sarcomas (combined) in female rats were observed. Hyperthermia-related lesions in male rats receiving 20 or 40 mg/kg were considered indirectly related to pentachloroanisole administration. Pentachloroanisole administration was associated with increased incidences of adrenal medulla hyperplasia and hypertrophy and hepatocellular mixed cell foci in male mice. In male and female mice, nonneoplastic liver lesions associated with pentachloroanisole administration included hepatocellular cytologic alteration, Kupffer cell pigmentation, biliary tract hyperplasia, and subacute inflammation. Synonyms: 2,3,4,5,6-pentachloroanisole; methyl pentachlorophenate; methyl pentachlorophenyl ether; o-methylpentachlorophenol; pentachloromethoxybenzene; pentachlorophenyl methyl ether

Journal Article↗
PubMed1993

NTP Toxicology and Carcinogenesis Studies of Promethazine Hydrochloride (CAS No. 58-33-3) in F344/N Rats and B6C3F1 Mice (Gavage Studies).

Promethazine hydrochloride is a drug used for the management of allergic conditions, motion sickness and nausea, and as a sedative to (treat psychiatric disorders. This drug was nominated for testing by the Food and Drug Administration because of its widespread use in human medicine and because of lack of data on its potential carcinogenicity. Oral administration is the most common route of human exposure. Toxicology and carcinogenicity studies were conducted by administering promethazine hydrochloride (>99% pure) in distilled water by gavage to groups of male and female F344/N rats and B6C3F1 mice for 16 days, 13 weeks, or 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, in cultured Chinese hamster ovary cells, and in Drosophila melanogaster. 16-DAY STUDY IN RATS: Groups of five male and five female rats received 0, 18.5, 55.5, 166.5, 500, or 1,500 mg promethazine hydrochloride/kg body weight once daily, 5 days per week for a total of 12 doses in a 16-day period. All rats receiving 1,500 mg/kg, four males and four females receiving 500 mg/kg, and one male and one female receiving 166.5 mg/kg died during the study. No deaths occurred in the remaining dose groups. Final mean body weights of rats receiving 166.5 mg/kg were significantly lower (12% to 25%) than those of the controls. Clinical findings included decreased activity, ocular discharge, and labored breathing in males and females receiving 166.5, 500, and 1,500 mg/kg as well as tremors in females receiving 166.5 and 500 mg/kg. There were dose-related increases in the absolute and relative liver weights of rats. Focal suppurative inflammation occurred in the nose of some male and female rats receiving 55 or 166.5 mg/kg and in the trachea of some male and female rats receiving 166.5 mg/kg. 16-DAY STUDY IN MICE: Groups of five male and five female mice received 0, 18.8, 37.5, 75, 150, or 300 mg promethazine hydrochloride/kg body weight once daily, 5 days per week for a total of 12 doses in a 16-day period. Two females receiving 75 mg/kg, one male and one female receiving 150 mg/kg, and four females receiving 300 mg/kg died during the study. No deaths occurred in the remaining dose groups. Final mean body weights of mice receiving promethazine hydrochloride were similar to those of the controls. However, in male and female controls, the final mean body weights were 11% to 12% lower than the initial mean body weights. Clinical findings occurred as early as the first day of the study and included decreased activity in male and female mice receiving 150 and 300 mg/kg. Tremors occurred in one male and five females in the 300 mg/kg group on day 1 and in one male in the 150 mg/kg group and five males and one female in the 300 mg/kg group on day 2. Absolute and relative liver weights of male mice receiving 75, 150, or 300 mg/kg were significantly greater than those of the controls. No chemical related lesions were present in male or female mice. 13-WEEK STUDY IN RATS: Groups of 10 male and 10 female rats received 0, 3.7, 11.1, 33.3, 100, or 300 mg promethazine hydrochloride/kg body weight once daily, 5 days per week for 13 weeks. One female receiving 100 mg/kg and six males and nine females receiving 300 mg/kg died during the study. No deaths occurred in the remaining dose groups. Final mean body weights of male rats receiving 100 or 300 mg/kg were significantly lower (19% to 22%) than those of the controls. Mean body weight gain of females receiving 100 mg/kg was significantly lower (14%) than that of the controls. Clinical findings in rats included hunched posture and labored breathing. Absolute and relative liver weights of males receiving 11.1, 33.3, 100, or 300 mg/kg and females receiving 33.3 or 100 mg/kg were significantly greater than those of the controls. Focal suppurative inflammation of the nose and trachea occurred with an increased incidence in rats receiving 100 and 300 mg/kg. A dose-related increased incidence of vacuolar degeneration of the nasal olfactory epithelium occurred in male and female rats that received 11.1, 33.3, or urred in male and female rats that received 11.1, 33.3, or 100 mg/kg. 13-WEEK STUDY IN MICE: Groups of 10 male and 10 female mice received 0, 5, 15, 45, 135, or 405 mg promethazine hydrochloride/kg body weight once daily, 5 days per week for 13 weeks. One control female, one female receiving 5 mg/kg, two females receiving 45 mg/kg, four females receiving 135 mg/kg, and all mice receiving 405 mg/kg died during the study. No deaths occurred in the remaining dose group. Final mean body weights of mice receiving 135 mg/kg were significantly lower (8% to 9%) than those of the controls. Clinical findings of toxicity included labored breathing and decreased activity in one 135 mg/kg female. Absolute and relative liver weights increased in a dose-related trend in both sexes. No chemical-related lesions were observed in mice. 2-YEAR STUDY IN RATS: Based on mortality and body weight differences observed at higher levels, doses of promethazine hydrochloride selected for the 2-year study in rats were 0, 8.3, 16.6, and 33.3 mg/kg. Groups of 60 male or 60 female rats were administered promethazine hydrochloride in deionized water by gavage once daily, 5 days per week for up to 103 weeks. Up to ten male and ten female rats per dose group were evaluated at 15 months. Survival, Body Weights, and Clinical Findings: There was a significant dose-related decrease in survival of rats. The survival rates in the 16.6 and 33.3 mg/kg male groups and in the 33.3 mg/kg female group were significantly lower than those of the controls. The final mean body weight of male rats receiving 33.3 mg/kg promethazine hydrochloride was 10% lower than that of the controls. Final mean body weights of female rats in the 16.6 and 33.3 mg/kg groups were 9% and 11% lower than that of the controls, respectively. No chemical-related clinical findings were noted in any dose group. Significant increases in the absolute and relative liver weights of mid- and high-dose female rats and the relative liver weights of mid- and high-dose male rats were observed at the 15-month interim evaluation. There were no biologically significant differences in the hematology or clinical chemistry parameters measured at 15 months. Pathology Findings: No neoplasms that could be attributed to promethazine hydrochloride administration were found in male or female rats. Several neoplasms occurred with a significantly decreased incidence in rats receiving promethazine hydrochloride. These included adrenal medullary pheochromocytoma (benign or malignant) and pituitary gland adenoma in the 33.3 mg/kg males and uterine stromal polyp in the 33.3 mg/kg females. The decreased incidences of adrenal medullary pheochromocytoma were chemical related. The decreased incidences of pituitary gland adenoma and uterine stromal polyp may have been related to chemical administration. Diffuse fatty change of the liver of male rats increased with dose and was attributed to chemical administration. 2-YEAR STUDY IN MICE: Based on mortality and body weight differences observed at higher levels, the doses of promethazine hydrochloride selected for the 2-year study were 0, 11.25, 22.5, and 45 mg/kg for male mice and 0, 3.75, 7.5, and 15 mg/kg for female mice. Groups of 60 male or 60 female mice were administered promethazine hydrochloride in deionized water by gavage once daily, 5 days per week for up to 103 weeks. Up to 10 male and 10 female mice per dose group were evaluated at 15 months. Survival, Body Weights, and Clinical Findings: Survival of mice receiving promethazine hydrochloride was similar to that of the controls. Mean body weights of mice were within 7% of those of the controls throughout the study. There were no chemical-related clinical findings in male or female mice. There were no differences in hematology or clinical chemistry parameters measured at 15 months that were attributed to the administration of promethazine hydrochloride. Pathology Findings: There were no neoplasms or nonneoplastic lesions that were attributed to the administration of promethazine hydrochloride. GENETIC TOXICOLOGY: Promethazine hydrochloride did not induce gene mutations in Salmonella typhimurium strains TA97, TA98, TA100, TA1535, or TA1537, or a significant increase in chromosomal aberrations in cultured Chinese hamster ovary cells; both of these tests were conducted with and without exogenous metabolic activation (S9). A small dose-related increase in sister chromatid exchanges was observed in cultured Chinese hamster ovary cells in the presence of S9; this response was considered to be equivocal. No increase in sister chromatid exchanges was observed in the absence of S9. Promethazine hydrochloride did not induce sex-linked recessive lethal mutations in germ cells of male Drosophila melanogaster administered the chemical by feeding or injection. CONCLUSIONS: Under the conditions of these 2-year gavage studies, there was no evidence of carcinogenic activity of promethazine hydrochloride in male or female F344/N rats receiving 8.3, 16.6, or 33.3 mg/kg. There was no evidence of carcinogenic activity of promethazine hydrochloride in male B6C3F1 mice receiving 11.25, 22.5, or 45 mg/kg. There was no evidence of carcinogenic activity of promethazine hydrochloride in female B6C3F1 mice receiving 3.75, 7.5, or 15 mg/kg. The decrease in the incidences of adrenal medullary pheochromocytoma in male rats was considered to be related to promethazine hydrochloride administration. The decrease in the incidences of pituitary gland adenoma in male rats and uterine stromal polyp in female rats may have been related to promethazine administration. Synonyms: Phenothiazine,10-(2-(dimethylamino)propyl)-,monochlorohydrate; 10H-phenothiazine-10-ethanamine;10-(2-dimethylamino-2-methylethyl)phenothiazine hydrochloride; N-(2 -dimethylamino-2 -methyl)ethylphenothiazine hydrochloride Trade names: Diprazi; Kinetosin; Phenergan; Phenergan hydrochloride; Promine; Pipolfen; Plletia; Prorex; Promantine; Pyrethia; Romergan hydrochlonde

Journal Article↗
PubMed1993

NTP Toxicology and Carcinogenesis Studies of 3,4-Dihydrocoumarin (CAS No. 119-84-6) in F344/N Rats and B6C3F1 Mice (Gavage Studies).

3,4-Dihydrocoumarin was nominated by the Food and Drug Administration and the National Cancer Institute for study because of its widespread use as a flavoring agent in beverages, gelatins, puddings, candy, and other food items; as a fragrance in perfumes, creams, and cosmetics; and because of interest in the structure-activity relationships of the coumarin derivatives. Toxicity and carcinogenicity studies were conducted by administering 3,4-dihydrocoumarin (99% pure) in corn oil by gavage to groups of male and female F344/N rats and B6C3F1 mice for 16 days, 13 weeks, and 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, cultured Chinese hamster ovary cells, and peripheral blood cells of mice. 16-DAY STUDY IN RATS: Groups of five male and five female rats received 3,4-dihydrocoumarin in corn oil by gavage at doses of 0, 190, 375, 750, 1,500, or 3,000 mg/kg body weight 5 days per week for a total of 12 doses in a 16-day period. All male and female rats given 3,000 mg/kg, and four male rats and five female rats given 1,500 mg/kg died. Body weight gains and final mean body weights of rats receiving 190, 375, or 750 mg/kg were similar to those of the controls. There were no clinical findings of organ-specific toxicity or evidence of impaired blood coagulation. 16-DAY STUDY IN MICE: Groups of five male and five female mice received 3,4-dihydrocoumarin in corn oil by gavage at doses of 0, 140, 280, 560, 1,125, or 2,250 mg/kg body weight 5 days per week for a total of 12 doses in a 16-day period. All mice given 2,250 mg/kg died. Body weight gains and final mean body weights of mice receiving 140, 280, 560, and 1,125 mg/kg were similar to those of the controls. There were no clinical findings of organ-specific toxicity or evidence of impaired blood coagulation. 13-WEEK STUDY IN RATS: Groups of 10 male and 10 female rats received 3,4-dihydrocoumarin in corn oil by gavage at doses of 0, 75, 150, 300, 600, or 1,200 mg/kg body weight 5 days per week for 13 weeks. Two male rats and five female rats given 1,200 mg/kg died. The body weight gain and final mean body weight of male rats that received 1,200 mg/kg were significantly lower than those of the controls, but the final mean body weights of other dosed groups of male rats and all dosed groups of female rats were similar to or slightly greater than those of the controls. Platelet counts were significantly lower in males and females receiving 600 and 1,200 mg/kg and in females receiving 300 mg/kg. Hemoglobin and hematocrit values and erythrocyte counts were significantly lower in males that received 300 mg/kg or more. The absolute and relative liver and kidney weights of males and females receiving 600 and 1,200 mg/kg were significantly greater than those of the controls. Hepatocellular hypertrophy was observed in rats given 300, 600, and 1,200 mg/kg. The high dose selected for the 2-year study was 600 mg/kg, which was below the level at which mortality, lower final mean body weights, and treatment-related liver lesions were observed. 13-WEEK STUDY IN MICE: Groups of 10 male and 10 female mice received 3,4-dihydrocoumarin in corn oil by gavage at doses of 0, 100, 200, 400, 800, or 1,600 mg/kg body weight 5 days per week for 13 weeks. Eight male and five female mice receiving 1,600 mg/kg died. Deaths in other groups were attributed to dosing accidents. Final mean body weights of dosed male and female mice were similar to those of the controls, and there were no treatment-related changes in any hematologic parameters. The absolute and relative liver weights of males and females that received 1,600 mg/kg and the relative kidney weight of males that received 1,600 mg/kg were significantly greater than those of the controls. No treatment-related lesions were noted. The high dose selected for the 2-year study was 600 mg/kg, which was below the level at which mortality, lower final mean body weights, and treatment-related liver lesions were observed. 2-YEAR STUDY IN RATS: Groups of 60 male and 60 female rats received 3,4-dihydrocoumarin in corn oil by gavage at age at doses of 0, 150, 300, or 600 mg/kg body weight. After 15 months, up to 10 animals from each group were evaluated. Survival, Body Weights, and Clinical Findings: Survival rates of dosed male rats were lower than that of the controls (O mg/kg, 28/51; 150 mg/kg, 12/50; 300 mg/kg, 8/50; 600 mg/kg, 2/50) but survival rates of dosed female rats were similar to that of the controls (31/50, 21/51, 26/50, 23/51). The decreased survival in dosed male rats was attributed to a chemical-related increase in the severity of nephropathy. The final mean body weight of male rats receiving 600 mg/kg was lower than that of the controls, but the final mean body weights of other dosed groups of male rats and all dosed groups of female rats were similar to those of the controls. No clinical findings related to chemical administration were observed. Hematology and Clinical Chemistry: At the 15-month interim evaluation, the hemoglobin concentrations, mean erythrocyte volumes, or mean erythrocyte hemoglobin concentrations in the 300 and 600 mg/kg female rats were slightly, but significantly, lower than those of the controls. In males, only the hemoglobin concentration in the 600 mg/kg group was significantly lower. Serum levels of alkaline phosphatase, alanine aminotransferase, sorbitol dehydrogenase, or g-glutamyltransferase in the 300 and 600 mg/kg male rats were significantly higher than those in the controls. In females, alkaline phosphatase and g-glutamyltransferase levels were significantly higher in the 600 mg/kg group. Pathology Findings: The principal lesions associated with the administration of 3,4-dihydrocoumarin to rats occurred in the kidney and forestomach. There was a chemical related increase in the severity of nephropathy in all dosed male rats and in 300 and 600 mg/kg female rats. There was a corresponding increased incidence of parathyroid gland hyperplasia, probably as a result of compromised renal function. In the standard evaluation of single kidney sections, renal tubule adenomas were observed in one 150 and two 600 mg/kg males and one each in the control, 150, and 300 mg/kg females. Transitional cell carcinomas were also observed in two 600 mg/kg male rats. However, an extended evaluation of step sections identified significantly higher incidences of focal hyperplasia and adenoma in the 600 mg/kg males than in controls (hyperplasia: 0/50, 5/48, 6/47, 8/50; adenoma: 1/50,1/48, 3/47, 6/50). The incidence of forestomach ulcers in all groups of dosed male rats was significantly greater than that of the controls (4/47, 14/48, 20/50, 16/46). STOP-EXPOSURE EVALUATION: A group of 40 male rats received 600 mg/kg 3,4-dihydrocoumarin in corn oil by gavage for 9 months, when 20 of the animals were necropsied and evaluated. The remainder of the male rats received only the corn oil vehicle until they died or until the end of the study. Similarly, a group of 30 male rats received 600 mg/kg 3,4-dihydrocoumarin in corn oil by gavage for 15 months, when 10 of the rats were necropsied and evaluated. The remaining 20 rats received only corn oil until the end of the study. A group of 20 vehicle control male rats was necropsied at 9 months, and another 10 vehicle control male rats were necropsied at 15 months. The severity of nephropathy in male rats of the stop-exposure groups was significantly greater than that of males examined at the 9- and 15-month interim evaluations. This was expected because nephropathy is a progressive degenerative disease that naturally increases in severity with age. 2-YEAR STUDY IN MICE: Groups of 70 male and 70 female mice received 3,4-dihydrocoumarin in corn oil by gavage at doses of 0, 200, 400, or 800 mg/kg body weight. After 15 months, five to 10 animals from each group were evaluated. Additional groups of 8 to 10 animals were evaluated for clinical pathology after 15 months. Survival, Body Weights, and Clinical Findings Survival rates of dosed male and female mice were similar to those of the controls (males: O mg/kg, 42/50; 200 mg/kg, 39/51; 400 mg/kg, 34/51; 800 mg/kg, 38/50; females: 36/51, 39/50, 41/50, 28/52). Final mean body weights of dosed male and female mice were similar to those of the controls. No clinical findings were noted that were related to chemical administration. Hematology and Clinical Chemistry: There were no differences in hematology or clinical chemistry parameters that were considered to be chemical related. Pathology Findings: The principal neoplasms associated with the administration of 3,4-dihydrocoumarin to mice occurred in the liver. There were significantly increased incidences of hepatocellular adenomas in all groups of dosed female mice. Further, the incidences of multiple hepatocellular adenomas in dosed female mice were greater than that of the controls (control, 0/51; 200 mg/kg, 6/50; 400 mg/kg, 9/50; 800 mg/kg, 9/52). However, there was no corresponding increased incidence of hepatocellular carcinoma in dosed female mice (3/51, 2/50, 4/50, 6/52), and the incidences of hepatocellular adenoma or carcinoma were similar between dosed and control male groups (adenoma: 29/50, 23/51, 36/51, 31/50; carcinoma: 11/50, 11/51, 11/51, 6/50). The incidence of alveolar/bronchiolar adenoma in the 200 and 400 mg/kg male mice was marginally greater than that of the controls (8/50,15/50,15/51,10/50). However, these neoplasms were not considered chemical related because the increased incidence was slight and there was no corresponding increased incidence in the 800 mg/kg group. The incidence of alveolar/bronchiolar neoplasms in female mice was similar between the dosed and control groups (adenoma: 2/51, 5/50, 1/48, 3/51; carcinoma: 0/51, 1/50, 0/48, 0/51). In the standard evaluation of single sections of kidney, focal hyperplasia and adenoma or carcinoma of the renal tubule were identified in several dosed male mice, but not in controls [adenoma or carcinoma (combined): 0/50,1/51, 2/51,1/49; hyperplasia: 2/50, 2/51, 5/51, 2/49]. In an extended evaluation of step sections, a few additional males with focal hyperplasia or renal tubule adenomas were identified in the dosed groups. However, the incidences of these lesions in dosed groups of male mice were not significantly greater than those of the controls, and did not increase with dose (hyperplasia: 0/50,1/51, 3/51, 1/49; renal tubule adenoma: 0/50, 0/51, 2/51, 1/49). Therefore, the low number of renal tubule neoplasms in male mice was not considered to be chemical related. GENETIC TOXICOLOGY: 3,4-Dihydrocoumarin did not induce gene mutations in Salmonella typhimurium strains TA98, TA100, TA1535, or TA1537 with or without exogenous metabolic activation (S9). It induced sister chromatid exchanges but not chromosomal aberrations in cultured Chinese hamster ovary cells, with and without S9. No induction of micronuclei was noted in peripheral blood erythrocyte samples obtained from male and female B6C3F1 mice at the end of the 13-week toxicology study. CONCLUSIONS: Under the conditions of these 2-year gavage studies, there was some evidence of carcinogenic activity of 3,4-dihydrocoumarin in male F344/N rats based on increased incidences of renal tubule adenomas and focal hyperplasia. The transitional cell carcinomas in two 600 mg/kg males may also have been chemical related. There was no evidence of carcinogenic activity of 3,4-dihydrocoumarin in female F344/N rats receiving 150, 300, or 600 mg/kg. There was no evidence of carcinogenic activity of 3,4-dihydrocoumarin in male B6C3F1 mice receiving 200, 400, or 800 mg/kg. There was some evidence of carcinogenic activity in female B6C3F1 mice based on increased incidences of hepatocellular adenoma and hepatocellular adenoma or carcinoma (combined). 3,4-Dihydrocoumarin caused ulcers, hyperplasia, and inflammation of the forestomach, parathyroid gland hyperplasia, and increased severity of nephropathy in male rats. Synonyms: 1,2-benzodihydropyrone, 2H-1-benzopyran-2-one, 2-chromanone, 3,4-dihydro-2H-1-benzopyran-2-one, dihydrocoumarin, hydrocoumarin, o-hydroycinnamic acid, delta-lactone-hydrocinnamic acid, melilotin, melilotine, melilotol, 2-oxochroman

Journal Article↗
PubMed1992

NTP Toxicology and Carcinogenesis Studies of HC Yellow 4 (CAS No. 59820-43-8) in F344 Rats and B6C3F1 Mice (Feed Studies).

HC Yellow 4 is used in semipermanent hair dyes. Toxicology and carcinogenesis studies were conducted by administering HC Yellow 4 (greater than 93% pure) in feed to groups of F344/N rats and B6C3F1 mice of each sex for 14 days, 13 weeks, and 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, cultured Chinese hamster ovary cells, and Drosophila melanogaster. 14-Day Studies: Groups of five rats of each sex were given 0, 5,000, 10,000, 20,000, 40,000, or 80,000 ppm and groups of five mice of each sex were given 0, 1,250, 2,500, 5,000, 10,000, or 20,000 ppm HC Yellow 4 in feed for 14 days. All animals survived to the end of the studies. Final mean body weights of male rats that received 20,000 ppm or more, female rats that received 10,000 ppm or more, and female mice that received 20,000 ppm were significantly lower than those of the controls. The mean body weights of exposed and control groups of male mice were similar. No chemical-related decrease in feed consumption was observed. No chemical-related clinical findings or changes in absolute or relative organ weights occurred in rats or mice. No gross or microscopic changes were related to HC Yellow 4 administration in rats or mice. 13-Week Studies: Groups of 10 rats of each sex were fed diets containing 0, 2,500, 5,000, 10,000, 20,000, or 40,000 ppm and groups of 10 mice of each sex were fed diets containing 0, 5,000, 10,000, 20,000, 40,000, or 80,000 ppm HC Yellow 4 for 13 weeks. All rats survived to study termination. Chemical-related deaths occurred at the two highest dose levels in male and female mice. Final mean body weights of male rats that received 10,000 ppm or greater, female rats that received 20,000 or 40,000 ppm, and mice that received 10,000 ppm or greater were significantly lower than those of the controls. There were no biologically significant changes in absolute or relative organ weights. Mineralization of the renal papilla occurred in all male rats in the 40,000 ppm group. Thyroid pigmentation occurred in rats receiving 40,000 ppm and in mice at all dose levels. Uterine atrophy occurred in female rats in the 20,000 and 40,000 ppm groups and female mice in the 40,000 and 80,000 ppm groups. Lymphoid depletion and atrophy of the spleen occurred in male mice that received 40,000 or 80,000 ppm and female mice that received 80,000 ppm. Atrophy of the thymus occurred in male and female mice that received 40,000 or 80,000 ppm. 2-Year Studies: Groups of 70 male rats were fed diets containing 0, 2,500, or 5,000 ppm and groups of 70 female rats and 70 mice of each sex were fed diets containing 0, 5,000, or 10,000 ppm HC Yellow 4 for up to 2 years. Interim evaluations were performed on 10 rats and 10 mice from each dose group at 6 and 15 months. No biologically significant changes in absolute or relative organ weight or hematology or clinical chemistry values were found in these rats or mice. No compound-related lesions were seen in exposed rats. In exposed mice, pigmentation of the thyroid gland was observed at the 6-month interim evaluations; pigmentation and hyperplasia of the thyroid gland were seen at the 15-month interim evaluations. Body Weight, Survival, and Feed Consumption in the 2-Year Studies: The mean body weight of female rats that received 10,000 ppm was significantly lower than that of the controls. The mean body weights of mice receiving 10,000 ppm were 20% to 30% lower than those of the controls during the second year of the studies. The survival of exposed rats and mice was similar to that of the controls. Neoplasms and Nonneoplastic Lesions in the 2-Year Studies: Pituitary gland pars distalis adenomas were marginally increased in exposed male rats (0 ppm, 17/45; 2,500 ppm, 20/49; 5,000 ppm, 28/49), and there was a concomitant dose-related increase in the incidence of hyperplasia (8/45, 13/49, 18/49). There was no increase in the incidence of pituitary gland adenomas or carcinomas in female rats (34/49, 35/48, 30/49). In mice, no neoplasms were considered related to chemical administration. However, a dose-related incr, a dose-related increased incidence of thyroid gland pigmentation and follicular cell hyperplasia occurred in both sexes of mice. Genetic Toxicology: HC Yellow 4 was mutagenic in Salmonella typhimurium strains TA100, TA1537, and TA98 with and without exogenous metabolic activation (S9); the response in strain TA1535 without S9 was equivocal. HC Yellow 4 induced sister chromatid exchanges in Chinese hamster ovary cells in the absence but not the presence of S9 activation; no induction of chromosomal aberrations occurred in Chinese hamster ovary cells, with or without S9. HC Yellow 4 induced sex-linked recessive lethal mutations in germ cells of adult male Drosophila melanogaster when administered by injection; results of a reciprocal translocation test in D. melanogaster were negative. Conclusions: Under the conditions of these 2-year feed studies, there was equivocal evidence of carcinogenic activity of HC Yellow 4 in male F344/N rats based on the increased incidence of pituitary gland adenomas and hyperplasia. The male rats may have been able to tolerate a slightly higher dose of the chemical. There was no evidence of carcinogenic activity of HC Yellow 4 in female F344/N rats given 5,000 or 10,000 ppm. There was no evidence of carcinogenic activity of HC Yellow 4 in male or female B6C3F1 mice given 5,000 or 10,000 ppm. There was a chemical-related increase in the incidence of thyroid gland pigmentation and follicular cell hyperplasia in mice. Synonym: N,O-di(2-hydroxyethyl)-2-amino-5-nitrophenol

Journal Article↗
PubMed1993

NTP Toxicology and Carcinogenesis Studies of p-Nitroaniline (CAS No. 100-01-6) in B6C3F1 Mice (Gavage Studies).

p-Nitroaniline is an intermediate in the preparation of several azo dyes used for coloring consumer products. Toxicology and carcinogenicity studies were conducted by administering p-nitroaniline (>99% pure) in corn oil by gavage to groups of male and female B6C3F1 mice for 14 days, 13 weeks, and 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, Chinese hamster ovary cells, mouse Iymphoma cells, and Drosophila melanogaster. 14-DAY STUDIES: Groups of five male and five female B6C3F1 mice received p-nitroaniline in corn oil by gavage at doses of 0, 10, 30, 100, 300, or 1,000 mg/kg body weight 5 days per week for 2 weeks. All mice that received 1,000 mg/kg died from chemical-related toxicity by day 4 of the studies. Final mean body weights of mice receiving 300 mg/kg or less were similar to those of the controls. Hematology results were consistent with chemical-related methemoglobinemia and regenerative anemia. Met hemoglobin concentrations in all groups of dosed mice were significantly higher than those in controls. Hematocrit values in mice that received 300 mg/kg and total erythrocyte counts in mice that received 100 or 300 mg/kg were significantly lower than those in controls. Reticulocyte counts in 300 mg/kg male mice and in 100 or 300 mg/kg females were significantly higher than controls. Heinz bodies were observed in erythrocytes of all 300 mg/kg mice and in two 100 mg/kg male mice. The absolute and relative spleen weights of 100 and 300 mg/kg mice were significantly greater than those of the controls. Hematopoiesis and pigment (hemosiderin) accumulation were observed in the splenic red pulp of males and females receiving 300 mg/kg; pigment (hemosiderin) accumulation in Kupffer cells of the liver was also seen in male mice at this dose level. 13-WEEK STUDIES: Groups of 20 male and 20 female B6C3F1 mice received p-nitroaniline in corn oil by gavage at doses of 0, 1, 3, 10, 30, or 100 mg/kg body weight 5 days per week for up to 13 weeks. Eight or nine mice in each group were evaluated at 7 weeks. There were no deaths associated with exposure to p-nitroaniline, and final mean body weights of dosed mice were similar to those of the controls. Hematologic and pathologic findings at 7 and 13 weeks were similar to those seen in the 14-day studies and occurred primarily in the 30 and 100 mg/kg groups. Met hemoglobin concentrations were increased and hematocrit levels and erythrocyte counts were decreased relative to those of the controls. Heinz bodies were observed in erythrocytes and nucleated erythrocytes and reticulocytes were increased in number. Absolute and relative spleen weights of male and female mice receiving 30 and 100 mg/kg were significantly greater than those of controls at 7 and 13 weeks. Absolute and relative liver weights of female mice necropsied at 7 weeks were significantly greater in the 30 and 100 mg/kg groups; by 13 weeks, both absolute and relative liver weights were similar to control values. The incidence or severity of splenic hematopoiesis and pigmentation (hemosiderin) increased with dose at the 7-week interim evaluations and at the end of the studies. Pigment (hemosiderin) was also present in Kupffer cells of the liver in dosed male mice. 2-YEAR STUDIES: Groups of 70 male and 70 female B6C3F1 mice received p-nitroaniline in corn oil by gavage at doses of 0, 3, 30, or 100 mg/kg body weight for 5 days per week for up to 103 weeks. The dose selection was based on the hematologic and pathologic findings of the 13-week studies. Nine or ten mice from each group were evaluated at 9 and 15 months for the presence of chemical-related lesions. Body Weights, Clinical Findings, Survival, and Hematology: Mean body weights of male and female mice that received p-nitroaniline were similar to those of control mice throughout the 2-year studies. There were no clinical findings associated with chemical exposure, and survival of dosed mice was similar to that of controls. The hematology findings at the 9 and 15-month interim evaluations were similar to those in the 14-day and 13-week s 13-week studies. The methemoglobin concentrations were significantly higher in all 30 or 100 mg/kg mice; sulfhemoglobin concentrations were significantly higher at 9 months in all 30 or 100 mg/kg female mice and at 15 months in 100 mg/kg females. Hematocrit and erythrocyte counts in 100 mg/kg mice were significantly lower than those in controls. By 9 months, reticulocyte counts were significantly higher in all 30 or 100 mg/kg mice. At 15 months, only the 100 mg/kg mice exhibited significantly higher reticulocyte counts. Neoplasms and Nonneoplastic Lesions: Lesions related to the administration of p-nitroaniline occurred in the spleen, liver, and bone marrow, primarily in mice receiving 30 or 100 mg/kg; these were observed at the 9- and 15-month interim evaluations and at the end of the studies. There were increases in the incidence or severity of splenic congestion, hematopoiesis, pigment (hemosiderin) accumulation, Kupffer cell pigmentation in the liver, and bone marrow hypercellularity (hyperplasia). The incidences of hemangiosarcoma of the liver (0 ppm, 0/50; 3 ppm, 1/50; 30 ppm, 2/50; 100 ppm, 4/50) and hemangioma or hemangiosarcoma (combined) at all sites (5/50, 3/50, 4/50, 10/50) were marginally increased in 100 mg/kg male mice. The incidence of hepatocellular adenoma or carcinoma (combined) was significantly decreased (25/50, 26/50, 25/50, 13/50) in 100 mg/kg male mice. GENETIC TOXICOLOGY: p-Nitroaniline is mutagenic in vitro. It was tested in two laboratories for induction of gene mutations in several strains of Salmonella typhimurium. Both studies showed positive results in strain TA98, with and without S9 activation; results were negative for all other strains. p-Nitroaniline was tested in two laboratories for induction of sister cremated exchanges and chromosomal aberrations in Chinese hamster ovary cells. In the sister cremated exchange study, one laboratory reported negative results without S9 and positive results with S9; the second laboratory reported equivocal results without S9 and negative results with S9. In the chromosomal aberrations study, both laboratories found positive results with S9. Without S9, one laboratory reported weakly positive results while the other reported negative results. p-Nitroaniline induced trifluorothymidine resistance in L5178Y mouse Iymphoma cells in the absence of S9; no induction of trifluorothymidine resistance was noted with S9. In contrast to the positive results in the previous tests, p-nitroaniline did not induce sex-linked recessive lethal mutations in germ cells of male Drosophila melanogaster when administered by feeding or injection to adult males or by feeding to larvae. CONCLUSIONS: Under the conditions of these 2-year gavage studies there was equivocal evidence of carcinogenic activity of p-nitroaniline in male B6C3F1 mice based on the increased incidences of hemangiosarcoma of the liver and hemangioma or hemangiosarcoma (combined) at all sites. There was no evidence of carcinogenic activity of p-nitroaniline in female B6C3F1 mice receiving doses of 3, 30, or 100 mg/kg.

Journal Article↗
PubMed1993

NTP Toxicology and Carcinogenesis Studies of p-Nitrophenol (CAS No. 100-02-7) in Swiss Webster Mice (Dermal Studies).

p-Nitrophenol is used in the production of acetaminophen, methyl and ethyl parathion insecticides, fungicides, and dyestuffs. Toxicology and carcinogenesis studies of p-nitrophenol (greater than 97% pure) were conducted by dermal application to male and female Swiss-Webster mice for 18 months. Dermal application was selected as the route of chemical administration because of possible skin absorption from p-nitrophenol-treated leather footwear. Genetic toxicology studies were conducted in Salmonella typhimurium, Chinese hamster ovary cells, and Drosophila melanogaster. 18-MONTH STUDIES: Groups of 60 Swiss-Webster mice of each sex received p-nitrophenol in acetone applied to the interscapular skin. Doses of 0, 40, 80, or 160 mg/kg p-nitrophenol were administered to mice 3 days per week for 78 weeks. At the end of the study, survival rates of mice receiving 0, 40, 80, or 160 mg/kg p-nitrophenol were 29/60, 17/60, 26/60, and 24/60 for males and 35/60, 26/60, 33/60, and 27/60 for females. Deaths after 60 weeks were caused by generalized amyloidosis and secondary kidney failure. The severity of amyloidosis was similar among dosed and control animals. At the end of the study, the final mean body weights of the dosed groups of each sex were similar to those of the controls. No biologically significant lesions were observed that were related to the dermal administration of p-nitrophenol. GENETIC TOXICOLOGY: p-Nitrophenol was not mutagenic in Salmonella typhimurium (strains TA100, TA1535, TA1537, and TA98) with or without exogenous metabolic (S9) activation, or in germ cells of male Drosophila melanogaster administered p-nitrophenol in feed or by injection. In Chinese hamster ovary cells, no induction of sister chromatid exchanges was observed with or without S9, but a significant increase in chromosomal aberrations occurred in trials conducted with S9. CONCLUSIONS: Under the conditions of these 18-month dermal studies there was no evidence of carcinogenic activity in male or female Swiss-Webster mice receiving 40, 80, or 160 mg/kg p-nitrophenol. Synonyms: 4-hydroxynitrobenzene, p-hydroxynitrobenzene, 4-nitrophenol, paranitrophenol, PNP, Niphen

Journal Article↗
PubMed1993

NTP Toxicology and Carcinogenesis Studies of o-Nitroanisole (CAS No. 91-23-6) in F344 Rats and B6C3F1 Mice (Feed Studies).

o-Nitroanisole is used as an intermediate for the preparation of o-anisidine and in the manufacture of azo dyes. Toxicology and carcinogenesis studies were conducted by administering o-nitroanisole (>99% pure) in the diet to groups of male and female F344 rats and B6C3F1 mice for 14 days, 13 weeks, and 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, Chinese hamster ovary cells, and mouse lymphoma cells. 14-DAY STUDIES: Groups of five male and five female F344 rats received diets containing 0, 583, 1,166, 2,332, 4,665, or 9,330 ppm o-nitroanisole. Mean body weight gains and final mean body weights of males in the 4,665 and 9,330 ppm groups were lower than those of the controls. Absolute liver weights were significantly increased in males receiving 1,166 ppm or more and in females receiving 583 ppm or more. Groups of five male and five female B6C3F1 mice received diets containing 0, 250, 500, 1,000, 2,000, or 4,000 ppm o-nitroanisole. Mean body weight gains and final mean body weights of males that received 250 ppm and females that received 4,000 ppm were significantly lower than those of the controls. No other chemical-associated effects were observed. 13-WEEK STUDIES: Groups of 10 male and 10 female F344 rats received diets containing 0, 200, 600, 2,000, 6,000, or 18,000 ppm o-nitroanisole. Final mean body weights and feed consumption by male and female rats receiving 6,000 and 18,000 ppm were lower than those of the controls. Hemoglobin and hematocrit values were significantly lower and methemoglobin levels significantly higher in males in the 6,000 and 18,000 ppm groups than in controls. Absolute liver weights were significantly increased in females that received 200, 600, 2,000, and 6,000 ppm, absolute kidney weights were significantly increased in males that received 600, 2,000, and 6,000 ppm, and absolute spleen weights were significantly increased in males and females that received 6,000 and 18,000 ppm. Groups of 10 male and 10 female B6C3F1 mice received diets containing 0, 60, 200, 600, 2,000, or 6,000 ppm o-nitroanisole. Final mean body weight gains, final mean body weights, and feed consumption by male and female mice receiving 6,000 ppm were lower than those of the controls. Hemoglobin and hematocrit values in males and females that received 2,000 or 6,000 ppm were significantly lower than those in the controls. The absolute and relative liver weights of females in the 600 ppm group and relative liver weights of males and females in the 2,000 and 6,000 ppm groups were significantly greater than those of controls. Lesions associated with exposure to o-nitroanisole were present in the urinary bladder, spleen, kidney, liver, testis, and uterus of rats. Diffuse hyperplasia of the transitional epithelium of the urinary bladder occurred in all male and female rats that received 6,000 and 18,000 ppm. A transitional cell papilloma occurred in one male and transitional cell carcinomas occurred in two males and three females receiving 18,000 ppm. Congestion of the red pulp and capsular hyperplasia of the spleen and hepatocellular hypertrophy of the liver were present in males and females from the 18,000 ppm groups. Multifocal degeneration and necrosis of the renal tubule epithelium with infiltration of mononuclear inflammatory cells were present in male rats that received 600, 2,000, and 6,000 ppm. At the 18,000 ppm level, degeneration of the seminiferous epithelium accompanied by loss of spermatogenic cells and decreased numbers of spermatozoa were observed in the testes of male rats, while uterine atrophy was observed in female rats. Hepatocyte hypertrophy of the centrilobular and midzonal regions of liver lobules was present in mice that received 200 ppm and increased in severity at higher exposure levels. 2-YEAR STUDIES: The doses selected for the 2-year study of o-nitroanisole in rats were based on lower mean body weights, reduced feed consumption, and increased severity of regenerative anemia in male and female rats receiving 6,000 and 18,000 ppm during the 13-week study. Groups of 6roups of 60 male and 60 female F344 rats received diets containing 0, 222, 666, or 2,000 ppm o-nitroanisole. Groups of 60 male and 60 female B6C3F1 mice received diets containing 0, 666, 2,000, or 6,000 ppm o-nitroanisole. After 15 months, up to 10 animals from each group were evaluated for chemical-related lesions. Survival, Body Weights, Feed Consumption, and Clinical Findings: Survival of male rats receiving 2,000 ppm was significantly lower than that of the controls due to increased severity of nephropathy. Survival of 222 and 666 ppm male rats and all exposed female rats was similar to that of the controls. Survival of groups of exposed male and female mice was similar to that of the controls. The final mean body weight of male rats receiving 2,000 ppm was lower than that of the controls. Final mean body weights of male and female mice that received 2,000 and 6,000 ppm were lower than those of the controls. Feed consumption by male and female rats was similar to that by the controls. The only clinical finding in male or female mice attributable to chemical administration was discolored urine. Neoplasms and Nonneoplastic Lesions: The incidence of mononuclear cell leukemia was significantly increased in male rats that received 666 and 2,000 ppm and in female rats that received 2,000 ppm (males: 0 ppm, 26/50; 222 ppm, 25/50; 666 ppm, 42/50; 2,000 ppm, 34/50; females: 14/50, 11/50, 14/50, 26/50). Nephropathy occurred in all male rats; the severity increased with exposure level. Focal hyperplasia of the renal tubule epithelium was present in three males receiving 222 ppm and two males receiving 2,000 ppm. Renal tubule adenomas occurred in one male from each of the 222, 666, and 2,000 ppm groups, and renal tubule carcinomas occurred in two males from the 2,000 ppm group. Focal hyperplasia of the transitional epithelium of the urinary bladder was present in one female rat that received 222 ppm and two male rats and six female rats that received 2,000 ppm. A transitional cell papilloma occurred in the urinary bladder of one female rat from the 2,000 ppm group, and a transitional cell carcinoma occurred in another female from the 2,000 ppm group. The incidence of forestomach ulcers increased in male rats that received 2,000 ppm, and the incidence of focal hyperplasia of the forestomach increased with exposure level in male and female rats. In addition, squamous cell papillomas of the forestomach were present in one female receiving 222 ppm, one male receiving 666 ppm, and one male and one female receiving 2,000 ppm, while squamous cell carcinomas were present in one male receiving 666 ppm and one male and one female receiving 2,000 ppm. The incidences of pituitary gland adenomas in male rats and mammary gland fibroadenomas in female rats decreased with exposure level. The incidence of cellular alteration in the liver was significantly increased in exposed groups of male and female mice. The incidences of hepatocellular adenoma, hepatocellular adenoma or carcinoma (combined), and hepatocellular carcinoma or hepatoblastoma (combined) were significantly increased in male mice receiving 2,000 and 6,000 ppm. The incidences of hepatocellular adenoma or carcinoma were significantly increased in female mice that received 2,000 ppm. STOP-EXPOSURE STUDY: Groups of 60 male and 60 female F344 rats received diets containing 0, 6,000, or 18,000 ppm o-nitroanisole for 27 weeks and were then maintained on control feed without further chemical exposure for up to an additional 77 weeks. Up to 10 rats from each group were evaluated for the presence of chemical-related lesions at 3, 6, 9, and 15 months. Survival and Body Weights: Survival of exposed male and female rats was significantly lower than that of the controls as a result of moribund deaths associated with significantly increased incidences of urinary bladder neoplasms, primarily transitional cell carcinomas. All male rats that received 18,000 ppm were dead by week 48 and all females that received 18,000 ppm were dead by week 61. Mean body weights of exposed male and female rats were lower than those of the controls throughout the study. Neoplasms and Nonneoplastic Lesions: Hyperplasia of the transitional epithelium of the urinary bladder was present in nearly all exposed male and female rats examined at the interim evaluations. A transitional cell carcinoma was first observed at the 3-month interim evaluation in a male rat that received 18,000 ppm. At the 6- and 9-month interim evaluations, transitional cell papillomas or carcinomas were observed in both exposed groups of male rats. Transitional cell carcinomas were observed at the 6-month interim evaluation in females receiving 18,000 ppm and at the 9-month interim evaluation in females receiving 6,000 and 18,000 ppm. Adenomatous polyps of the large intestine were observed in a small number of exposed rats at the 6-, 9-, and 15-month interim evaluations. At the end of the study, the incidence of adenomatous polyps of the large intestine was significantly increased in all exposed groups and carcinomas of the large intestine were present in four males and two females from the 18,000 ppm groups. The incidence of hyperplasia of the transitional epithelium of the kidney pelvis was significantly increased in exposed male and female rats and transitional cell papillomas were present in three males and one female that received 18,000 ppm. Transitional cell carcinomas of the kidney were present in one male receiving 6,000 ppm and six males and one female receiving 18,000 ppm. Transitional cell carcinomas of the urinary bladder were seen in nearly all exposed male and female rats. Of the males and females receiving 6,000 ppm which were without carcinomas, three males and one female had transitional cell papillomas. Generalized centrilobular hypertrophy, focal hepatocellular necrosis, multifocal hepatocellular cytoplasmic vacuolation, and Kupffer cell pigmentation were observed in the livers of male and female rats at the 3- and 6-month interim evaluations; however, only Kupffer cell pigmentation was observed at the end of the study. Congestion of the red pulp of the spleen was observed in nearly all exposed male and female rats at the 3-, 6-, and 9-month interim evaluations but the incidence was only slightly increased in the 18,000 ppm groups at the end of the study. Degeneration and atrophy of the seminiferous tubule epithelium of the testes were observed at the 3- and 6-month interim evaluations in all male rats receiving 18,000 ppm. GENETIC TOXICOLOGY: o-Nitroanisole was tested in two laboratories for mutagenicity in Salmonella typhimurium strains TA97, TA98, TA100, TA1535, and TA1537 with and without exogenous metabolic activation (S9). Positive responses were observed at both laboratories in TA100 with and without S9 activation. One laboratory found no increase in mutations, while the second laboratory detected a weakly positive response in TA1535 without S9. No mutagenic activity was observed in the other tester strains. o-Nitroanisole was positive in the mouse lymphoma assay for induction of trifluorothymidine resistance in L5178Y cells without S9 activation. In cytogenetic tests with Chinese hamster ovary cells, o-nitroanisole induced a significant increase in chromosomal aberrations at the highest dose tested in the presence of S9 activation; sister chromatid exchanges were induced both with and without S9. CONCLUSIONS: Under the conditions of these feed studies there was clear evidence of carcinogenic activity of o-nitroanisole in male and female F344 rats that received diets containing 6,000 or 18,000 ppm for 6 months based on overall increased incidences of benign and malignant neoplasms of the urinary bladder, transitional cell neoplasms of the kidney, and benign and malignant neoplasms of the large intestine. There was a chemical-related increased incidence of mononuclear cell leukemia in male and female rats receiving diets containing 222, 666, or 2,000 ppm o-nitroanisole for 2 years. Marginally increased incidences of uncommon renal tubule neoplasms in male rats and forestomach neoplasms in male and female rats were considered uncertain findings. There was clear evidence of carcinogenic activity of o-nitroanisole in male B6C3F1 mice based on increased incidences of benign and malignant hepatocellular neoplasms. There was some evidence of carcinogenic activity of o-nitroanisole in female B6C3F1 mice based on increased incidences of hepatocellular adenomas. Increased severity of nephropathy in male rats, and increased incidences of focal hyperplasia of the renal tubule epithelium and forestomach ulcers in male rats, and of transitional cell hyperplasia of the urinary bladder, focal hyperplasia of the forestomach, and hyperplasia of transitional epithelium of the kidney pelvis in male and female rats were associated with exposure to o-nitroanisole. Synonyms: Methoxynitrobenzene, nitrophenyl methyl ether

Journal Article↗
PubMed1993

NTP Toxicology and Carcinogenesis Studies of 1,3-Butadiene (CAS No. 106-99-0) in B6C3F1 Mice (Inhalation Studies).

1,3-Butadiene is produced in large volumes for use in the manufacture of synthetic rubber and of thermoplastic resins. In previous inhalation studies conducted by the NTP (NTP, 1984) there was clear evidence of multiple organ carcinogenicity in male and female mice exposed to 625 or 1,250 ppm 1,3-butadiene for 60 or 61 weeks. To better characterize exposure-response relationships for neoplasms and nonneoplastic lesions, toxicology and carcinogenesis studies were conducted by exposing groups of male and female B6C3F1 mice to air containing 1,3-butadiene (greater than 99% pure) for up to 2 years. An additional study in male B6C3F1 mice, in which exposure to 1,3-butadiene was stopped after limited exposure periods (13, 26, 40, or 52 weeks), was performed to assess the effects of varying concentration and duration of exposure on the incidences of 1,3-butadiene-induced neoplasms. In vitro genetic toxicology studies were conducted in Salmonella typhimurium and mouse lymphoma cells. In vivo genetic effects were assayed in germ cells of male Drosophila melanogaster and in bone marrow and peripheral blood cells of B6C3F1 mice. 2-Year Studies: Groups of 70 male and 70 female mice were exposed to air containing 0, 6.25, 20, 62.5, or 200 ppm 1,3-butadiene for 6 hours per day, 5 days per week for up to 2 years; groups of 90 male and 90 female mice were exposed to 625 ppm 1,3-butadiene on the same schedule. Up to 10 animals from each group were examined after 9 and 15 months of exposure. Survival and Body Weight in the 2-Year Studies: Two-year survival was decreased for males and females exposed to concentrations of 20 ppm or above, primarily due to the development of chemical-related malignant neoplasms. No female mice exposed to 200 or 625 ppm or males exposed to 625 ppm survived to the end of the studies (males: 35/50, 39/50, 24/50, 22/50, 4/50, 0/70; females: 37/50, 33/50, 24/50, 11/50, 0/50, 0/70). Mean body weights of exposed male and female mice were similar to those of the controls. Hematologic Effects in the 2-Year Studies: Hematologic parameters were evaluated after 9 and 15 months of exposure. At 9 months, decreases in erythrocyte counts, hemoglobin concentration, and packed red cell volume were observed in male mice exposed to 62.5 ppm or above and in female mice exposed to 200 or 625 ppm. Mean erythrocyte volume was increased in male mice exposed to 625 ppm and in females exposed to 200 or 625 ppm. At 15 months, decreases in erythrocyte counts, hemoglobin concentration, and packed red cell volume and increases in mean erythrocyte volume were observed in male and female mice exposed to 625 ppm. Neoplasms and Nonneoplastic Lesions in the 2-Year Studies: Exposure of mice to 1,3-butadiene induced benign and malignant neoplasms at multiple sites. Statistically significant increases in the incidences of neoplasms at one or more sites were seen at concentrations of 20 ppm and higher in males and 6.25 ppm and higher in females. There was no exposure level in this study at which a significant carcinogenic response was not observed. Statistically significant increases occurred in the incidences of malignant lymphoma; histiocytic sarcoma; cardiac hemangiosarcoma; harderian gland adenoma; hepatocellular adenoma and carcinoma; alveolar/bronchiolar adenoma and carcinoma; mammary gland carcinoma, adenoacanthoma, and malignant mixed tumor (females only); benign and malignant ovarian granulosa cell tumor; and forestomach squamous cell papilloma and carcinoma. Low incidences of uncommon neoplasms also occurred in exposed male and female mice, including intestinal carcinomas in males, renal tubule adenomas in males and females, skin sarcomas (all types combined) in females, and Zymbal's gland adenomas and carcinomas in females. Lymphocytic lymphomas appeared as early as week 23 and were the principal cause of death of male and female mice exposed to 625 ppm 1,3-butadiene. The early and extensive development of lethal lymphocytic lymphomas in mice exposed to 625 ppm resulted in a reduced number of mice at risk for neoplasms developing laterg later at other sites. Exposure-response relationships for 1,3-butadiene-induced neoplasms were more clearly characterized at concentrations below 625 ppm and after adjustment for intercurrent mortality. Increased incidences of nonneoplastic lesions in exposed mice included bone marrow atrophy; testicular atrophy; ovarian atrophy, angiectasis, germinal epithelial hyperplasia, and granulosa cell hyperplasia; uterine atrophy; cardiac endothelial hyperplasia and mineralization; alveolar epithelial hyperplasia; forestomach epithelial hyperplasia; and harderian gland hyperplasia. Stop-Exposure Study: The stop-exposure study consisted of groups of 50 male mice exposed to 1,3-butadiene at concentrations of 200 ppm for 40 weeks, 625 ppm for 13 weeks, 312 ppm for 52 weeks, or 625 ppm for 26 weeks. After the exposures were completed, these groups were placed in control chambers for the remainder of the 2-year study. The total exposure of 1,3-butadiene (concentration times duration of exposure) of the 13- and 40-week stop-exposure groups was approximately 8,000 ppm-weeks, while that of the 26- and 52-week stop-exposure groups was approximately 16,000 ppm-weeks. The survival of all stop-exposure groups was markedly lower than that of the controls. The incidences of lymphocytic lymphoma, histiocytic sarcoma, cardiac hemangiosarcoma, alveolar/bronchiolar adenoma and carcinoma, forestomach squamous cell papilloma and carcinoma, hepatocellular adenoma, harderian gland adenoma and adenocarcinoma, and preputial gland carcinoma were significantly increased. Neoplasms were induced at most of these sites after only 13 weeks of exposure to 1,3-butadiene. Additionally, low numbers of malignant gliomas and neuroblastomas of the brain and Zymbal's gland carcinomas occurred in one or more stop-exposure groups. At similar total exposures, the incidence of lymphocytic lymphoma was greater with exposure to a higher concentration of 1,3-butadiene for a short time compared with exposure to a lower concentration for an extended period (34% at 625 ppm for 13 weeks versus 12% at 200 ppm for 40 weeks; 60% at 625 ppm for 26 weeks versus 8% at 312 ppm for 52 weeks). Genetic Toxicology: 1,3-Butadiene has been tested both in vitro and in vivo for mutagenic activity. In vitro, positive results were obtained in the Salmonella typhimurium gene mutation assay with strain TA1535; mutagenic activity was not observed in other S. typhimurium strains (TA100, TA97, and TA98). 1,3-Butadiene was negative in the mouse lymphoma assay for induction of trifluorothymidine resistance in L5178Y cells with and without S9. In vivo, 1,3-butadiene did not induce sex-linked recessive lethal mutations in germ cells of male Drosophila melanogaster; however, it did induce significant increases in chromosomal aberrations and sister chromatid exchanges in bone marrow cells of mice exposed for 2 weeks by inhalation. In addition, significant increases in micronucleated erythrocytes were observed in peripheral blood samples obtained from male and female mice exposed to 1,3-butadiene for 2 or 13 weeks or 15 months by inhalation. Conclusions: The previous inhalation studies of 1,3-butadiene (TR-288) in male and female B6C3F1 mice provided clear evidence of carcinogenicity at exposure concentrations of 625 or 1,250 ppm. The present inhalation studies - 2-year exposures of 6.25, 20, 62.5, 200, or 625 ppm or shorter duration exposures of 200, 312, or 625 ppm - provide a better characterization of the concentration-dependent responses for 1,3-butadiene-induced neoplasms and nonneoplastic lesions. The present studies confirmed the clear evidence of carcinogenicity of 1,3-butadiene in male B6C3F1 mice based on increased incidences of neoplasms in the hematopoietic system, heart, lung, forestomach, liver, harderian gland, preputial gland, brain, and kidney. There was clear evidence of carcinogenicity of 1,3-butadiene in female B6C3F1 mice based on increased incidences of neoplasms in the hematopoietic system, heart, lung, forestomach, liver, harderian gland, ovary, and mammary gland. Low incidences of intestinal carcinomas in male mice, Zymbal's gland carcinomas in male and female mice, and renal tubule adenomas and skin sarcomas in female mice may also have been related to administration of 1,3-butadiene. Synonyms: alpha,gamma-Butadiene; bivinyl; divinyl; erythrene; vinylethylene; biethylene; pyrrolylene

Journal Article↗
PubMed1994

NTP Toxicology and Carcinogenesis Studies of Tricresyl Phosphate (CAS No. 1330-78-5) in F344/N Rats and B6C3F1 Mice (Gavage and Feed Studies).

Tricresyl phosphate is an organophosphate plasticizer widely used in vinyl plastics and as a fire retardant additive for hydraulic fluids. Toxicology and carcinogenesis studies were conducted by administering a mixed isomer preparation of 79% tricresyl phosphate esters (consisting of 21% tri- m-cresyl phosphate, 4% tri- p-cresyl phosphate, less than 1% tri- o-cresyl phosphate, and other unidentified tricresyl phosphate esters) by gavage to groups of F344/N rats and B6C3F1 mice for 16 days and 13 weeks, and in feed to groups of F344/N rats and B6C3F1 mice for 13 weeks and 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium and cultured Chinese hamster ovary cells. 16-DAY GAVAGE STUDY IN RATS: Groups of 10 male and 10 female rats received tricresyl phosphate in corn oil by gavage at doses of 0, 360, 730, 1,450, 2,900, or 5,800 mg/kg body weight, 5 days per week, for a total of 13 or 14 doses in a 16-day period. One female receiving 1,450 mg/kg and five males and eight females receiving 2,900 mg/kg died before the end of the study. Final mean body weights of male and female rats that received 1,450, 2,900, or 5,800 mg/kg were significantly lower than those of the controls. Necrosis of the mandibular lymph node, spleen, and thymus occurred primarily in rats receiving 2,900 and 5,800 mg/kg. Diffuse aspermatogenesis occurred in the testes of male rats that received 2,900 and 5,800 mg/kg. Changes in neurobehavioral parameters in groups that received 1,450, 2,900, or 5,800 mg/kg were confounded by mortality and reduced body weights and were not attributed to a direct neurotoxic response. 16-DAY GAVAGE STUDY IN MICE: Groups of 10 male and 10 female mice received tricresyl phosphate in corn oil by gavage at doses of 0, 360, 730, 1,450, 2,900, or 5,800 mg/kg body weight, 5 days per week, for a total of 13 or 14 doses in a 16-day period. Five males and all females that received 1,450 mg/kg, all mice that received 2,900 mg/kg, and four males and one female that received 5,800 mg/kg died before the end of the study. Final mean body weights of male mice that received 1,450 and 5,800 mg/kg were significantly lower than that of the controls. Final mean body weights of female mice that received 360, 730, or 5,800 mg/kg were significantly greater than that of the controls. Necrosis of the mandibular lymph node, thymus, and spleen occurred primarily in mice receiving 2,900 and 5,800 mg/kg. Hindlimb grip strengths of male mice that received 360 and 1,450 mg/kg and male and female mice that received 730 and 5,800 mg/kg were significantly lower than those of the controls at the end of the study. 13-WEEK GAVAGE STUDY IN RATS: Groups of 10 male and 10 female rats received tricresyl phosphate in corn oil by gavage at doses of 0, 50, 100, 200, 400, or 800 mg/kg body weight. All rats survived to the end of the study. Final mean body weights of male rats receiving 200, 400, and 800 mg/kg were significantly lower than that of the controls. Cytoplasmic vacuolization of the adrenal cortex occurred in all dosed groups and the severity increased with dose. Ovarian interstitial cell hypertrophy occurred in all dosed groups of females. Atrophy of the seminiferous tubules occurred in male rats that received 400 and 800 mg/kg. There were no biologically significant changes in neurobehavioral parameters in rats. 13-WEEK GAVAGE STUDY IN MICE: Groups of 10 male and 10 female mice received tricresyl phosphate in corn oil by gavage at doses of 0, 50, 100, 200, 400, or 800 mg/kg body weight. All mice survived to the end of the study. Final mean body weights of male mice receiving 200 mg/kg and of male and female mice receiving 400 and 800 mg/kg were significantly lower than those of the controls. Cytoplasmic vacuolization of the adrenal cortex occurred in all dosed groups of mice and the severity increased with dose. Ovarian interstitial cell hypertrophy was present in all dosed groups of female mice. Multifocal degeneration of the spinal cord occurred in males and females that received 100, 200, 400, and 800 mg/kg, and multifocal degenerg/kg, and multifocal degeneration of the sciatic nerve occurred in males that received 200, 400, and 800 mg/kg and females that received 100, 200, 400, and 800 mg/kg. Hindlimb grip strengths of male mice that received 200, 400, or 800 mg/kg were significantly lower than that of the controls at the end of the study. 13-WEEK FEED STUDY IN RATS: Groups of 10 male and 10 female rats were fed diets containing 0, 900, 1,700, 3,300, 6,600, or 13,000 ppm of tricresyl phosphate. All rats survived to the end of the study. Final mean body weights of males and females exposed to 6,600 and 13,000 ppm and females exposed to 3,300 ppm were significantly lower than those of controls. Feed consumption by male and female rats exposed to 13,000 ppm was lower than that by controls during the first week of the study. Dietary levels of 900, 1,700, 3,300, 6,600 or 13,000 ppm tricresyl phosphate were estimated to deliver daily doses of 55, 120, 220, 430, or 750 mg/kg body weight (males) and 65, 120, 230, 430, or 770 mg/kg (females). There were no biologically significant changes in neurobehavioral parameters in rats. Cytoplasmic vacuolization of the adrenal cortex occurred in all exposed groups of rats. Hyperplasia of ovarian interstitial cells and inflammation of the ovarian interstitium occurred in all exposed groups of females. Renal papillary edema and renal papillary necrosis occurred in 13,000 ppm males and females and in 6,600 ppm females. Basophilic hypertrophy of the pituitary gland pars distalis and atrophy of the seminiferous tubules occurred in 6,600 and 13,000 ppm males. Dose selection for the 2-year study in rats was based on lower mean body weights; toxic responses observed in the kidney, pituitary gland, and testis of males and the kidney of females exposed to 6,600 and 13,000 ppm; the presence of cytoplasmic vacuolization of the adrenal cortex in exposed males and females; and the occurrence of ovarian interstitial cell hyperplasia in females exposed to 900 and 1,700 ppm. 13-WEEK FEED STUDY IN MICE: Groups of 10 male and 10 female mice were fed diets containing 0, 250, 500, 1,000, 2,100, or 4,200 ppm of tricresyl phosphate. All mice survived to the end of the study. Mean body weights of 4,200 ppm males and of females exposed to 2,100 and 4,200 ppm were lower than those of controls throughout the study. Feed consumption by females exposed to 1,000, 2,100, or 4,200 ppm was lower than that by controls during week 12. Dietary levels of 250, 500, 1,000, 2,100, or 4,200 ppm tricresyl phosphate were estimated to deliver average daily doses of 45, 110, 180, 380, or 900 mg/kg body weight (males) and 65, 130, 230, 530, or 1,050 mg/kg (females). Interpretation of grip strength changes observed in groups receiving 2,100 or 4,200 ppm were confounded by the reduced body weights of these groups. Cytoplasmic vacuolization of the adrenal cortex occurred in all exposed groups of male and female mice with the exception of 250 ppm males. Papillary hyperplasia of the gallbladder mucosa occurred in male mice exposed to 500 ppm or more and in female mice exposed to 1,000 ppm or more. Axonal degeneration occurred in males and females exposed to 2,100 and 4,200 ppm and females exposed to 1,000 ppm. Renal tubule regeneration occurred in all 4,200 ppm male mice. Dose selection for the 2-year study in mice was based on the presence of axonal degeneration at concentrations of 1,000 ppm or more and cytoplasmic vacuolization of the adrenal cortex at concentrations of 500 ppm or more in males and in all exposed groups of females. 2-YEAR FEED STUDY IN RATS: Groups of 95 male and 95 female rats were fed diets containing 0, 75, 150, or 300 ppm of tricresyl phosphate. An additional group of 95 male and 95 female rats were fed diets containing 600 ppm of tricresyl phosphate for 22 weeks and then received only control feed. After 3, 9, and 15 months of chemical exposure, up to 15 males and 15 females per group were evaluated for forelimb and hindlimb grip strength, then necropsied and evaluated for histopathologic lesions. Survival, Mean Body Weights, and Feed Consumption: Survival of exposed rats was similar to that of controls. The final mean body weights of all exposed groups of males and females were similar to those of the controls. Feed consumption by exposed groups of male and female rats was similar to that by the controls. Dietary levels of 75, 150, or 300 ppm tricresyl phosphate were estimated to deliver average daily doses of 3, 6, or 13 mg/kg body weight (males) and 4, 7, or 15 mg/kg (females). Pathology Findings: There were no chemical-related increased incidences of neoplasms in rats. Cytoplasmic vacuolization of the adrenal cortex occurred in 600 ppm males and 150, 300, and 600 ppm females at the 3-month interim evaluation. At 9 and 15 months, cytoplasmic vacuolization occurred only in female rats, primarily in the 300 ppm group. Cytoplasmic vacuolization of the adrenal cortex and ovarian interstitial cell hyperplasia occurred in female rats exposed to 300 ppm throughout the 2-year study and the incidence and severity were significantly increased at the end of the study. 2-YEAR FEED STUDY IN MICE: Groups of 95 male and 95 female mice were fed diets containing 0, 60, 125, or 250 ppm of tricresyl phosphate. After 3, 9, and 15 months of chemical exposure, up to 15 males and 15 females per group were evaluated for forelimb and hindlimb grip strength, then necropsied and evaluated for histopathologic lesions. Survival, Mean Body Weights, and Feed Consumption: Survival of exposed groups of male and female mice was similar to that of the controls. The final mean body weights of males and females receiving tricresyl phosphate were similar to those of controls. Feed consumption by exposed groups of male and female mice was similar to that by the controls. Dietary levels of 60, 125, or 250 ppm tricresyl phosphate were estimated to deliver average daily doses of 7, 13, or 27 mg/kg body weight (males) and 8, 18, or 37 mg/kg (females). Pathology Findings: There were no chemical-related increased incidences of neoplasms in mice. Ceroid pigmentation of the adrenal cortex occurred in all groups of mice throughout most of the 2-year study, with the exception of 60 and 125 ppm females at the 3-month interim evaluation; however, the severity was markedly increased in female mice receiving 250 ppm. Incidences of clear cell foci, fatty change, and ceroid pigmentation of the liver were significantly increased in male mice that received 125 or 250 ppm. GENETIC TOXICOLOGY: Tricresyl phosphate was not mutagenic in Salmonella typhimurium strains TA98, TA100, TA1535, or TA1537, nor did it induce chromosomal aberrations or sister chromatid exchanges in cultured Chinese hamster ovary cells. These in vitro assays were all conducted with and without exogenous metabolic activation (S9). CONCLUSIONS: Under the conditions of these 2-year feed studies, there was no evidence of carcinogenic activity of tricresyl phosphate in male or female F344/N rats that received 75, 150, or 300 ppm. There was no evidence of carcinogenic activity of tricresyl phosphate in male or female B6C3F1 mice that received 60, 125, or 250 ppm. Nonneoplastic lesions associated with exposure to tricresyl phosphate included cytoplasmic vacuolization of the adrenal cortex and ovarian interstitial cell hyperplasia in female rats, increased incidences of clear cell focus, fatty change, and ceroid pigmentation of the liver in male mice, and increased severity of ceroid pigmentation of the adrenal cortex in female mice.

Journal Article↗
PubMed1994

NTP Toxicology and Carcinogenesis Studies of Barium Chloride Dihydrate (CAS No. 10326-27-9) in F344/N Rats and B6C3F1 Mice (Drinking Water Studies).

Barium chloride dihydrate, a white crystalline granule or powder, is used in pigments, aluminum refining, leather tanning and coloring, the manufacture of magnesium metal, ceramics, glass, and paper products, as a pesticide, and in medicine as a cardiac stimulant. Toxicology and carcinogenicity studies were conducted by administering barium chloride dihydrate (99% pure) in drinking water to F344/N rats and B6C3F1 mice for 15 days, 13 weeks, and 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, cultured Chinese hamster ovary cells, and mouse lymphoma cells. 15-DAY STUDY IN RATS: Groups of five males and five females received barium chloride dihydrate in the drinking water at concentrations of 0, 125, 250, 500, 1,000, or 2,000 ppm for 15 days, corresponding to average daily doses of 10, 15, 35, 60, or 110 mg barium/kg body weight to males and females. No chemical-related deaths, differences in final mean body weights, or clinical findings of toxicity were observed. Water consumption by male and female rats exposed to 2,000 ppm was slightly less (S16%) than controls during week 2. There were no significant differences in absolute or relative organ weights between exposed and control rats. No biologically significant differences in hematology, clinical chemistry, or neurobehavioral parameters occurred in rats. 15-DAY STUDY IN MICE: Groups of five males and five females received barium chloride dihydrate in the drinking water at concentrations of 0, 40, 80,173, 346, or 692 ppm for 15 days, corresponding to average daily doses of 5,10, 20, 40, or 70 mg barium/kg body weight to males and 5, 10, 15, 40, or 85 mg barium/kg body weight to females. No chemical-related deaths, differences in mean body weights or in water consumption, or clinical findings of toxicity were observed in mice. The relative liver weight of males receiving 692 ppm was significantly greater than that of the controls. The absolute and relative liver weights of females that received 692 ppm were significantly greater than those of the controls. No histopathologic evidence of toxicity was observed in mice. 13-WEEK STUDY IN RATS: Groups of 10 males and 10 females received barium chloride dihydrate in the drinking water at concentrations of 0, 125, 500, 1,000, 2,000, or 4,000 ppm for 13 weeks, corresponding to average daily doses of 10, 30, 65, 110, or 200 mg barium/kg body weight to males and 10, 35, 65, 115, or 180 mg barium/kg body weight to females. Three males and one female in the 4,000 ppm groups died during the last week of the study. The final mean body weights of male and female rats receiving 4,000 ppm were significantly lower (13% and 8%) than those of the controls. Water consumption by male and female rats in the 4,000 ppm groups was approximately 30% lower than that by the controls. No clearly chemical-related clinical findings of toxicity or neurobehavioral or cardiovascular effects were noted. Serum phosphorus levels in 2,000 and 4,000 ppm male and female rats were significantly higher than those in controls, but there were no biologically significant differences in hematology parameters or in serum sodium, potassium, or calcium levels. Renal tubule dilatation in the outer stripe of the outer medulla and cortex occurred in male and female rats receiving 4,000 ppm. 13-WEEK STUDY IN MICE: Groups of 10 males and 10 females received barium chloride dihydrate in the drinking water at concentrations of 0, 125, 500, 1,000, 2,000, or 4,000 ppm for 13 weeks, corresponding to average daily doses of 15, 55, 100, 205, or 450 mg barium/kg body weight to males and 15, 60, 110, 200, or 495 mg barium/kg body weight to females. Six males and seven females that received 4,000 ppm and one male that received 125 ppm died during the study. Final mean body weights of male and female mice receiving 4,000 ppm were significantly lower (>30%) than those of controls. Water consumption by male mice in the 4,000 ppm group was 18% lower than that by the controls; water consumption by other exposed groups of male and female mice was similar to thatd groups of male and female mice was similar to that by the controls. Clinical findings of toxicity were limited to debilitation in the surviving male and female mice receiving 4,000 ppm. The absolute and/or relative liver weights of mice receiving 1,000, 2,000, and 4,000 ppm were significantly lower than those of the controls. Multifocal to diffuse nephropathy characterized by tubule dilatation, regeneration, and atrophy occurred in 4,000 ppm male and female mice. 2-YEAR STUDY IN RATS: Groups of 60 males and 60 females received barium chloride dihydrate in the drinking water at concentrations of 0, 500, 1,250, or 2,500 ppm for 104 (males) or 105 weeks (females), corresponding to average daily doses of 15, 30, or 60 mg barium/kg body weight for males and 15, 45, or 75 mg barium/kg body weight for females. The high dose of 2,500 ppm was selected based on decreased final mean body weights, mortality, decreased water consumption, and chemical-related kidney lesions observed in the 4,000 ppm groups in the 13-week study. Survival, Body Weights, Water Consumption, and Clinical Findings: Two-year survival of exposed male and female rats was similar to that of the controls. The final mean body weights of male and female rats that received 2,500 ppm were (5% and 11%) lower than those of controls. Beginning as early as week 5, water consumption by male and female rats receiving 2,500 ppm was substantially lower than that by controls (male: 11% to 30%; female: 19% to 33%). There were no chemical-related clinical findings. Hematology and Clinical Chemistry: There were no chemical-related differences in hematology or clinical chemistry parameters in male or female rats. Special Studies: At the 15-month interim evaluation, the plasma barium concentrations (mg/ml) were significantly increased in males receiving 1,250 and 2,500 ppm and in all exposed groups of females (male: 0 ppm, 0.98; 500 ppm, 1.00; 1,250 ppm, 1.23; 2,500 ppm, 1.68; female: 0 ppm, 0.74; 500 ppm, 0.99; 1,250 ppm, 0.97; 2,500 ppm, 1.43). Barium levels in bone in rats from the 2,500 ppm groups were about 400 times greater than those in the controls. Pathology Findings: At the end of 2 years, there were no increased incidences of neoplasms or nonneoplastic lesions that could be attributed to barium chloride dihydrate. However, there were dose-related decreased incidences of adrenal medulla pheochromocytomas and mononuclear cell leukemia in male rats. 2-YEAR STUDY IN MICE: Groups of 60 males and 60 females received barium chloride dihydrate in the drinking water at concentrations of 0, 500, 1,250, or 2,500 ppm for 103 (males) or 104 weeks (females), corresponding to average daily doses of 30, 75, or 160 mg barium/kg body weight for males and 40, 90, or 200 mg barium/kg body weight for females. The high dose of 2,500 ppm was selected based on decreased final mean body weights, mortality, decreased water consumption, and chemical-related kidney lesions observed in the 4,000 ppm groups in the 13-week study. Survival, Body Weights, Water Consumption, and Clinical Findings: Two-year survival of male and female mice receiving 2,500 ppm was significantly lower than that of the controls due to renal toxicity. Final mean body weights of 2,500 ppm males and females were 9% and 12% lower than those of controls. Water consumption by male and female mice receiving barium chloride was similar to that by the controls. There were no chemical-related clinical findings. Hematology and Clinical Chemistry: There were no differences in hematology or clinical chemistry parameters measured at the 15-month interim evaluation. Special Studies: At the 15-month interim evaluation, plasma barium concentrations (mg/mL) were significantly increased in all exposed groups of mice (male: 0 ppm, 0.62; 500 ppm, 0.77; 1,250 ppm, 0.89; 2,500 ppm, 1.49; female: 0 ppm, 0.52; 500 ppm, 0.74; 1,250 ppm, 1.01; 2,500 ppm, 1.35). Pathology Findings: At the end of the 2-year study, there were increased incidences of nephropathy in male and female mice (male: 1/50, 0/50, 2/48, 19/50; female: 0/50, 2/53, 1/50, 37/54). There were no chemical-related increased incidences of neoplasms in male or female mice. The incidence of hepatocellular adenoma was significantly decreased in male mice receiving 2,500 ppm. GENETIC TOXICOLOGY: Barium chloride dihydrate was not mutagenic in Salmonella typhimurium strains TA97, TA98, TA100, TA1535, or TA1537, with or without exogenous metabolic activation (S9). It was mutagenic in L5178Y mouse lymphoma cells in the presence of S9, but it did not induce sister chromatid exchanges or chromosomal aberrations in cultured Chinese hamster ovary cells, with or without S9. CONCLUSIONS: Under the conditions of these 2-year drinking water studies, there was no evidence of carcinogenic activity of barium chloride dihydrate in male or female F344/N rats that received 500, 1,250, or 2,500 ppm. There was no evidence of carcinogenic activity of barium chloride dihydrate in male or female B6C3F1 mice that received 500, 1,250, or 2,500 ppm. There were chemical-related increased incidences of nephropathy in male and female mice.

Journal Article↗
PubMed1993

NTP Toxicology and Carcinogenesis Studies of Benzyl Acetate (CAS No. 140-11-4) in F344/N Rats and B6C3F1 Mice Feed Studies).

Benzyl acetate is used as a flavoring agent in foods, as a fragrance in soaps and perfumes, as a solvent for cellulose acetate and nitrate, and as a component of printing inks and varnish removers. The NTP previously studied the toxicology and carcinogenicity of this chemical in F344/N rats and B6C3F1 mice using the gavage route of administration and corn oil as a vehicle. Benzyl acetate increased the incidences of pancreatic acinar cell adenomas in male rats and the incidences of hepatocellular adenomas and forestomach neoplasms in male and female mice. Because of the confounding effect of corn oil on the incidences of pancreatic neoplasms and because of controversy over the use of the gavage route of administration, the NTP decided to restudy benzyl acetate using the dosed feed route of administration. In these repeat studies, male and female F344/N rats and B6C3F1 mice received benzyl acetate (at least 98% pure) in feed for 13 weeks and 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium nunnery, cultured Chinese hamster ovary cells, LS178Y mouse lymphoma cells, Drosophila melanogaster, and mouse bone marrow and peripheral blood cells. 13-WEEK STUDY IN RATS: Groups of 10 male and 10 female F344/N rats were fed diets containing 0, 3,130, 6,250,12,500, 25,000, or 50,000 ppm (0, 230, 460, 900,1,750, or 3,900 mg/kg body weight for males and 0, 240, 480, 930,1,870, or 4,500 mg/kg for females) benzyl acetate for 13 weeks. Nine male and nine female rats receiving 50,000 ppm benzyl acetate died or were killed moribund between weeks 2 and 8 of the study. The mean body weight gain and the final mean body weight of 25,000 ppm males were significantly lower (P</=0.01) than those of the control group. Feed consumption by exposed rats, except the 25,000 and S0,000 ppm males and 50,000 ppm females, was similar to that by the controls. The reduced feed consumption by 25,000 and 50,000 ppm males and 50,000 ppm females may have been due to toxicity or decreased palatability. Tremors and ataxia occurred only in the 50,000 ppm rats. These findings were first observed on day 15 in nine males and six females and continued until the end of the study. Cholesterol levels in 12,500 and 25,000 ppm females and triglyceride levels in 25,000 ppm females were lower than those in the controls. Chemical-related lesions occurred in the brain, kidney, tongue, and skeletal muscles of the thigh. Necrosis of the brain involving the cerebellum and/or hippocampus, degeneration and regeneration of the renal tubule epithelium, and degeneration and sarcolemma nuclear hyperplasia of the tongue and skeletal muscles occurred in most male and female 50,000 ppm rats. This effect was observed in the 1,000 mg/kg group in the previous gavage study (NTP, 1986). 13-WEEK STUDY IN MICE: Groups of 10 male and 10 female B6C3F1 mice were fed diets containing 0, 3, 130, 6,250, 12,500, 25,000, or 50,000 ppm (0, 425, 1,000, 2,000, 3,700, or 7,900 mg/kg body weight for males and 0, 650, 1,280, 2,980, 4,300, or 9,400 mg/kg for females) benzyl acetate. One 50,000 ppm male mouse died and one 50,000 ppm female mouse was killed moribund before the end of the study. Mean body weight gains and final mean body weights of all exposed male and female mice were significantly lower than those of the controls and the mean body weight gains decreased with increased exposure level. Feed consumption by 3,130 ppm males and all exposed females was lower than that by the controls. Tremors occurred only in females and were first observed on day 16 in three females receiving 50,000 ppm, day 94 in one female receiving 25,000 ppm, and day 93 in one female receiving 12,500 ppm. The tremors continued until the end of the study. Necrosis of the brain involving the hippocampus occurred in four 50,000 ppm mice, one male and three females. Hepatocellular necrosis also occurred in the male with brain lesions. On reexamination of the previous 13-week gavage study (NTP, 1986), a similar lesion was seen in the brain of one 1,000 mg/kg female mouse; none were seen in 1,000 mg/kg male mi male mice. The lesion was less severe than that described in the present dosed feed study. The highest dose used in the gavage study was 1,000 mg/kg compared to an estimated high dose of 7,200 mg/kg for the feed study. 2-YEAR STUDY IN RATS: The doses selected for the 2-year feed study of benzyl acetate in F344/N rats were based on lower survival, mean body weights, and feed consumption, and on increased incidences of histopathologic brain lesions in 50,000 ppm male and female rats in the 13-week study. Groups of 60 male and 60 female F344/N rats were fed diets containing 0, 3,000, 6,000, or 12,000 ppm benzyl acetate for 2 years. Survival, Body Weights, Feed and Compound Consumption, and Clinical Pathology: Survival of exposed rats was similar to that of the controls. The mean body weights of the 12,000 ppm males and exposed females were approximately 5&percnt; lower than those of the controls throughout most of the study. The feed consumption by 12,000 ppm males was slightly lower than that by the controls. Dietary levels of 3,000, 6,000, and 12,000 ppm benzyl acetate were estimated to result in average daily consumption levels of 130, 260, and 510 mg/kg body weight (males) and 145, 290, and 575 mg/kg (females). No biologically significant changes in hematology or clinical chemistry parameters were found that could be attributed to benzyl acetate administration. Pathology Findings: No compound-related increased incidences of neoplasms or nonneoplastic lesions occurred in male or female F344/N rats receiving benzyl acetate for as long as 2 years. 2-YEAR STUDY IN MICE: The doses selected for the 2-year feed study of benzyl acetate in B6C3F1 mice were based primarily on lower body weight gains and lower final mean body weights of exposed mice in the 13-week study. Groups of 60 male and 60 female B6C3F1 mice were fed diets containing 0, 330, 1,000, or 3,000 ppm benzyl acetate for 2 years. Survival, Body Weights, Feed and Compound Consumption, and Clinical Pathology: Survival of all exposed mice, except the 3,000 ppm females, was similar to that of the control groups. Survival of 3,000 ppm females was significantly higher than that of the control group. Throughout the 2-year study, the mean body weights of 1,000 and 3,000 ppm males and females were 2&percnt; to 14&percnt; lower than those of the control groups. Dietary levels of 330, 1,000, and 3,000 ppm benzyl acetate were estimated to result in average daily consumption levels of 35, 110, and 345 mg/kg (males) and 40, 130, and 375 mg/kg (females). No biologically significant changes in hematology or clinical chemistry parameters were observed in mice receiving 330,1,000, or 3,000 ppm benzyl acetate. Pathology Findings: No increase in neoplasm incidence in mice could be attributed to benzyl acetate administration in feed. This contrasts with the previous finding that administration of benzyl acetate in corn oil by gavage once daily 5 days a week for as long as 2 years was carcinogenic to mice, causing increased incidences of hepatocellular neoplasms and forestomach neoplasms. The contrast in results between the two studies may be due to differences in the dose levels used (highest dose: gavage, 1,000 mg/kg a day; feed, 360 mg/kg a day). Dose-related increased incidences or severities of nonneoplastic nasal lesions occurred in the most posterior portions of the nasal cavity in all exposed groups. The lesions occurred in the majority of the exposed mice and consisted of atrophy and degeneration, primarily of the olfactory epithelium, cystic hyperplasia of the nasal submucosal glands, pigmentation of the mucosal epithelium, and exudate accumulation. GENETIC TOXICOLOGY: Benzyl acetate was not mutagenic in Salmonella typhimurium strains TA98, TA100, TA1535, or TA1537, with or without exogenous metabolic activation (S9). However, a positive response was observed for benzyl acetate, with and without S9, in the mouse lymphoma assay for induction of trifluorothymidine resistance in L5178Y cells. No significant increases in the frequencies of sister chromatid exchanges or chromosomal aberrations occurred in cultured Chinese hamster ovary cells treated with benzyl acetate in vitro, with or without S9, and no increases in either sister chromatid exchanges or chromosomal aberrations occurred in bone marrow cells of male mice treated in vivo by intraperitoneal injection. No increase in sex-linked recessive lethal germ cell mutations occurred in male Drosophila melanogaster administered benzyl acetate in feed or by injection. Tests of benzyl acetate for induction of micronucleated erythrocytes in bone marrow and peripheral blood of mice were also negative. CONCLUSIONS: Under the conditions of these 2-year feed studies, there was no evidence of carcinogenic activity of benzyl acetate in male or female F344/N rats receiving 3,000, 6,000, or 12,000 ppm; however, rats may have tolerated higher doses. There was no evidence of carcinogenic activity of benzyl acetate in male or female B6C3F1 mice receiving 330, 1,000, or 3,000 ppm. Nasal lesions associated with benzyl acetate exposure in male and female mice included nasal mucosa atrophy and degeneration (primarily of the olfactory epithelium), cystic hyperplasia of the nasal submucosal gland, and luminal exudate and pigmentation of the nasal mucosal epithelium. In previous 2-year gavage studies (TR-250), benzyl acetate increased the incidence of acinar cell adenomas of the exocrine pancreas in male F344/N rats; the gavage vehicle may have been a contributing factor. There was no evidence of carcinogenic activity in female F344/N rats receiving 250 or 500 mg/kg a day. There was some evidence of carcinogenic activity in male and female B6C3F1 mice, indicated by the increased incidences of hepatocellular adenomas and squamous cell neoplasms of the forestomach. Synonyms: acetic acid benzyl ester, acetic acid phenyl methyl ester, (acetoxymethyl)benzene, acetoxytoluene, benzyl ethanoate, phenylmethyl acetate

Journal Article↗
PubMed1993

NTP Toxicology and Carcinogenesis Studies of Manganese (II) Sulfate Monohydrate (CAS No. 10034-96-5) in F344/N Rats and B6C3F1 Mice (Feed Studies).

Manganese is the 12th most abundant element in the earth's crust. The base metal does not occur naturally, but is a component of more than 100 minerals, including sulfides, oxides, carbonates, silicates, phosphates, and borates. In addition to occurring in foods and drinking water, manganese occurs in the atmosphere from dust, volcanic activity, forest fires, and industrial emissions. Manganese (II) sulfate monohydrate was chosen for study because of its stability, solubility, and availability. Toxicology and carcinogenesis studies were conducted by administering manganese (II) sulfate monohydrate (97% pure) in feed to groups of male and female F344/N rats and B6C3F1 mice for 14 days, 13 weeks, and 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, germ cells of Drosophila melanogaster, and cultured Chinese hamster ovary cells. 14-DAY STUDY IN RATS: Groups of five male and five female rats received diets containing 0, 3,130, 6,250, 12,500, 25,000, or 50,000 ppm manganese (II) sulfate monohydrate. All rats survived to the end of the study. Male rats exposed to 50,000 ppm had a mean body weight gain 57% lower and a final mean body weight 13% lower than those of the controls. The mean body weight gain of 50,000 ppm females was 20% lower and the final mean body weight was 7% lower than those of the controls. During the second week, 50,000 ppm males and females exhibited diarrhea. 14-DAY STUDY IN MICE: Groups of five male and five female mice received diets containing 0, 3,130, 6,250, 12,500, 25,000, or 50,000 ppm manganese (II) sulfate monohydrate. One female mouse in the 25,000 ppm group died on day 1 of unknown causes; all other mice survived to the end of the study. Differences in body weights between exposed and control mice could not be attributed to chemical administration. 13-WEEK STUDY IN RATS: Groups of 10 male and 10 female rats received diets containing 0, 1,600, 3,130, 6,250, 12,500, or 25,000 ppm manganese (II) sulfate monohydrate. Mean daily ingestion of manganese (II) sulfate monohydrate ranged from 110 to 1,700 mg/kg body weight in males and 115 to 2,000 mg/kg in females. All rats survived to the end of the study. Mean body weight gains were marginally lower than that of controls in males exposed to 3,130 ppm or more; mean body weight gains were significantly lower than that of the controls in females exposed to 6,250,12,500, or 25,000 ppm. At the end of the study, absolute and relative liver weights of all exposed male rats and of 25,000 ppm female rats were significantly lower than those of controls. The total leukocyte count in males was similar between exposed and control rats; however, neutrophil counts of all exposed groups were greater than those of the controls, whereas lymphocyte counts of the 6,250, 12,500, and 25,000 ppm groups were significantly lower than those of the controls. Total leukocyte counts in 6,250,12,500, and 25,000 ppm females were significantly decreased because of a decrease in lymphocytes. Male rats also demonstrated marginal but significant increases in percent hematocrit and erythrocyte count in the 6,250,12,500, and 25,000 ppm groups. No clinical or histopathologic findings in rats were chemical related. 13-WEEK STUDY IN MICE: Groups of 10 male and 10 female mice received diets containing 0, 3,130, 6,250, 12,500, 25,000, or 50,000 ppm manganese (II) sulfate monohydrate. Mean daily ingestion of manganese (II) sulfate monohydrate ranged from 330 to 7,400 mg/kg body weight in males and 390 to 6,900 mg/kg body weight in females. No deaths were chemical related. The mean body weight gains of exposed male mice and of 50,000 ppm female mice were significantly lower than those of controls. The absolute and relative liver weights of 50,000 ppm males were significantly lower than those of controls. The percent hematocrit and hemoglobin concentration of males and females exposed to 50,000 ppm were lower than those of the controls, and the mean erythrocyte volumes were significantly lower than those of the controls. The total leukocyte counts of males in the 25,eukocyte counts of males in the 25,000 and 50,000 ppm groups were significantly lower than that of the controls. No clinical findings were attributed to manganese (II) sulfate monohydrate ingestion. Epithelial hyperplasia and hyperkeratosis of the forestomach occurred in three 50,000 ppm males. 2-YEAR STUDY IN RATS: Groups of 70 male and 70 female rats were fed diets containing 0, 1,500, 5,000, or 15,000 ppm manganese (II) sulfate monohydrate. Based on average daily feed consumption, these doses resulted in the daily ingestion of 60, 200, or 615 mg/kg body weight (males) or 70, 230, or 715 mg/kg (females). Eight to 10 rats from each group were evaluated at 9 and 15 months. Survival, Body Weights, Feed Consumption, and Clinical Findings: Survival of 15,000 ppm male rats in the 2-year study was significantly lower than that of the control group. The deaths of males in the control and exposure groups were attributed to a variety of spontaneous neoplastic and nonneoplastic lesions; however, the greater number of deaths in the 15,000 ppm group resulted from increased incidences of advanced renal disease related to ingestion of manganese (II) sulfate monohydrate. The decreased survival of the 15,000 ppm males did not occur until approximately week 93 of the study; before week 93, survival was similar in all groups. Survival of exposed females was similar to that of the controls. The mean body weight of 15,000 ppm male rats was within 5&percnt; of the control group until week 89, by week 104, the mean body weight of 15,000 ppm males was 10&percnt; lower than that of the control group. The mean body weights of 1,500 and 5,000 ppm male rats and all exposed female groups were similar to those of the controls throughout the study. Feed consumption by all exposure groups was similar to that by the control groups. No clinical findings were attributed to manganese (II) sulfate monohydrate ingestion. Hematology, Clinical Chemistry, and Tissue Metal Concentration Analyses No differences in hematology and clinical chemistry parameters attributable to the ingestion of manganese (II) sulfate monohydrate occurred between exposed and control groups. At both the 9- and 15-month interim evaluations, tissue concentrations of manganese were significantly elevated in the livers of 5,000 and 15,000 ppm male and female rats, with an accompanying depression of hepatic iron. Pathology Findings: The ingestion of diets containing 15,000 ppm manganese (II) sulfate monohydrate was associated with a marginal increase in the average severity of nephropathy in male rats (0 ppm, 2.9; 1,500 ppm, 3.0; 5,000 ppm, 3.0; 15,000 ppm, 3.2). The increased severity of nephropathy in the 15,000 ppm male rats was accompanied by significantly increased incidences of mineralization of the blood vessels (4/52, 10/51, 6/51,17/52) and glandular stomach (8/52,13/51, 9/51, 23/52), parathyroid gland hyperplasia (14/51, 14/46, 12/49, 23/50), and fibrous osteodystrophy of the femur (12/52,14/51,12/51, 24/52). These lesions are manifestations of renal failure, uremia, and secondary hyperparathyroidism. The increased incidence of advanced renal disease caused reduced survival of the high-dose male rats. No increase in the incidence of neoplasms in male or female rats was attributed to the ingestion of diets containing manganese (II) sulfate monohydrate. 2-YEAR STUDY IN MICE: Groups of 70 male and 70 female mice received diets containing 0, 1,500, 5,000, or 15,000 ppm manganese (II) sulfate monohydrate. These levels resulted in an average daily ingestion of 160, 540, or 1,800 mg/kg body weight (males) or 200, 700, or 2,250 mg/kg (females). Nine or 10 mice from each group were evaluated at the 9-month and 15-month interim evaluations. Survival, Body Weights, Feed Consumption, and Clinical Findings: Survival rates of exposed male and female mice in the 2-year study were similar to those of the control groups. The mean body weights of exposed male mice were similar to that of the control group. Compared to controls, female mice had exposure related lower mean body weights after week 37, and the final mean body weights for the 1,500, 5,000, and 15,000 ppm groups were 6&percnt;, 9&percnt;, and 13&percnt; lower than that of the control group. Feed consumption by all exposure groups was similar to that by the control groups. No clinical findings were attributed to the administration of manganese (II) sulfate monohydrate. Hematology, Clinical Chemistry, and Tissue Metal Concentration Analyses No chemical-related differences between exposed and control groups occurred in hematology or clinical chemistry parameters. At the 9- and 15-month interim evaluations, tissue concentrations of manganese were significantly elevated in the livers of the 5,000 and 15,000 ppm groups. Hepatic iron levels were significantly lower in exposed females at the 9-month interim evaluation and in 5,000 and 15,000 males and all exposed females at the 15-month interim evaluation. Pathology Findings: Incidences of thyroid follicular dilatation and hyperplasia were significantly greater in 15,000 ppm male and female mice than in controls. Follicular cell adenomas occurred in one 15,000 ppm male at the 15-month interim evaluation and in three 15,000 ppm males at the end of the study but not in the lower exposure groups or the control group. Follicular cell adenomas also occurred in two control, one 1,500, and five 15,000 ppm female mice at the end of the study. It is uncertain if the slightly increased incidence of follicular cell adenoma is related to the ingestion of manganese (II) sulfate monohydrate. The incidences of focal hyperplasia of the forestomach epithelium were significantly greater in the 15,000 ppm male and exposed female groups. The hyperplasia was associated with ulcers and inflammation in some mice, particularly males. GENETIC TOXICOLOGY: Manganese (II) sulfate monohydrate was not mutagenic in Salmonella typhimurium strains TA97, TA98, TA100, TA1535, or TA1537, with or without exogenous metabolic activation (S9), and did not induce sex-linked recessive lethal mutations in germ cells of male Drosophila melanogaster. Tests for induction of sister chromatid exchanges and chromosomal aberrations in cultured Chinese hamster ovary cells treated without S9 were positive; with S9, only the sister chromatid exchange test with manganese (11) sulfate monohydrate was positive. CONCLUSIONS: Under the conditions of these 2-year feed studies, there was no evidence of carcinogenic activity of manganese (II) sulfate monohydrate in male or female F344/N rats receiving 1,500, 5,000, or 15,000 ppm. There was equivocal evidence of carcinogenic activity of manganese (II) sulfate monohydrate in male and female B6C3F1 mice, based on the marginally increased incidences of thyroid gland follicular cell adenoma and the significantly increased incidences of follicular cell hyperplasia. The ingestion of diets containing manganese (II) sulfate monohydrate was associated with an increased severity of nephropathy in male rats, focal squamous hyperplasia of the forestomach in male and female mice, and ulcers and inflammation of the forestomach in male mice. These studies were not designed to assess any neurotoxicity that might have been expected with chronic exposure to sufficiently high doses of manganese. Synonyms: Manganese sulfate; manganous sulfate; sulfuric acid. manganese2+ salt (1:1), monohydrate

Journal Article↗
PubMed1992

Toxicology and Carcinogenesis Studies of g-Butyrolactone (CAS No. 96-48-0) in F344/N Rats and B6C3F1 Mice (Gavage Studies).

g-Butyrolactone is an intermediate in the synthesis of polymers used as film formers in hair sprays, blood plasma extenders, and clarifying agents in beer and wine. Toxicology and carcinogenesis studies were conducted by administering g-butyrolactone (greater than 97% pure) in corn oil by gavage to groups of F344/N rats and B6C3F1 mice of each sex, 5 days per week for 16 days, 13 weeks, and 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, Drosophila melanogaster, and Chinese hamster ovary cells. 16-Day Studies: Groups of five rats of each sex received doses of 0, 75, 150, 300, 600, or 1,200 mg of g-butyrolactone per kg of body weight and groups of five mice of each sex received doses of 0, 87, 175, 350, 700, or 1,400 mg/kg. All male and female rats given 1,200 mg/kg and one male rat given 600 mg/kg died within 3 days. The mean body weight gain of female rats given 600 mg/kg was significantly lower than that of the controls. Mean body weight gains of the other female dose groups and all male dose groups were similar to those of the controls. All of the male and four female mice receiving 1,400 mg/kg died during the studies. Mean body weight gains of dosed mice were generally similar to those of the controls. Rats receiving 600 or 1,200 mg/kg and mice receiving 350 mg/kg or more became inactive or recumbent with irregular respiration following dosing. 13-Week Studies: Groups of 10 rats of each sex received doses of 0, 56, 112, 225, 450, or 900 mg of g-butyrolactone per kg of body weight and groups of 10 mice of each sex received doses of 0, 65, 131, 262, 525, or 1,050 mg/kg. One female and all male rats given 900 mg/kg died during the studies. The final mean body weight and mean body weight gain of male rats receiving 450 mg/kg were significantly lower than those of the controls; final mean body weights and body weight gains of all female rat dose groups were similar to those of the controls. There was an increased incidence of focal inflammation of the nasal mucosa in rats administered g-butyrolactone. Three male mice and one female receiving 1,050 mg/kg died from g-butyrolactone toxicity during the studies. The mean body weight gain and final mean body weight of high-dose male mice were lower than those of the controls; the mean body weight gains and final mean body weights of dosed female mice were similar to those of the controls. No lesions related to the administration of g-butyrolactone occurred in mice of either sex. 2-Year Studies: The doses administered to groups of 50 animals per sex were 0, 112, and 225 mg ofg-butyrolactone per kg of body weight for male rats; 0, 225, and 450 mg/kg for female rats; and 0, 262, and 525 mg/kg for male and female mice. Body Weight and Survival in the 2-Year Studies: The mean body weights of male rats administered g-butyrolactone were similar to those of the controls throughout the study. The mean body weight of high-dose females was lower than that of the controls after week 5 and was 10% to 20% lower than that of the controls throughout the second year. The survival of high-dose male rats was slightly higher than that of the controls (control, 24/50; low-dose, 27/50, high-dose, 32/50) due primarily to a lower incidence of mononuclear cell leukemia in the high-dose group (16/50, 15/50, 9/50). The survival of dosed females was similar to that of the controls (28/50, 27/50, 28/50). The mean body weights of dosed male mice were lower than those of the controls throughout the study, but the differences in mean body weights decreased when male mice were housed individually at week 67. The final mean body weights of dosed male mice were 6% lower than that of the controls. Mean body weights of dosed female mice were also lower than those of the controls throughout the study, and the final mean body weights were from 14% to 17% lower than that of the controls. The survival in high-dose male mice was significantly lower than that of the controls (35/50, 30/50, 12/50) due to bite wounds and fighting in high-dose males recovering from the sedative effects of g-becovering from the sedative effects of g-butyrolactone. The survival of female dosed mice was similar to that of the controls (38/50, 34/50, 38/50). Neoplasms and Nonneoplastic Lesions in the 2-Year Studies: No increased incidences of neoplasms or nonneoplastic lesions in male rats were related to the administration of g-butyrolactone for 2 years. In female rats, negative trends were observed in the incidences of cysts (42/50, 35/50, 23/50) and fibroadenomas of the mammary gland (22/50, 14/50, 6/50) and in cysts of the pituitary pars distalis (25/49, 13/37, 11/48). These decreases were considered to be related to g-butyrolactone administration. Increased incidences of proliferative lesions, primarily hyperplasia, of the adrenal medulla in low-dose male mice were associated with g-butyrolactone administration (pheochromocytoma, benign or malignant: 2/48, 6/50, 1/50; hyperplasia: 2/48, 9/50, 4/50). The incidence of hepatocellular neoplasms in both dose groups of male mice was lower than the incidence in the controls (hepatocellular adenoma or carcinoma: 24/50, 8/50, 9/50). Genetic Toxicology: g-Butyrolactone was not mutagenic, with or without exogenous metabolic activation (S9), in Salmonella typhimurium strains TA98, TA100, TA1535, and TA1537, nor did it induce sex-linked recessive lethal mutations in germ cells of male Drosophila melanogaster when administered in feed or by injection. Positive results were obtained, however, in cytogenetic tests with Chinese hamster ovary cells; g-butyrolactone induced sister chromatid exchanges and chromosomal aberrations in trials conducted in the presence of S9 activation. Conclusions: Under the conditions of these 2-year gavage studies, there was no evidence of carcinogenic activity of g-butyrolactone in male F344/N rats given 112 or 225 mg/kg or in female F344/N rats given 225 or 450 mg/kg in corn oil. There was equivocal evidence of carcinogenic activity of g-butyrolactone in male B6C3F1 mice based on marginally increased incidences of adrenal medulla pheochromocytomas and hyperplasia in the low-dose group. The sensitivity of the study in male mice to detect a carcinogenic effect was reduced by the low survival of the high-dose group associated with fighting. There was no evidence of carcinogenic activity of g-butyrolactone in female B6C3F1 mice given 262 or 525 mg/kg in corn oil. A decreased incidence of hepatocellular neoplasms in dosed male mice and decreased incidences of mammary gland fibroadenomas and cysts and pituitary cysts in female rats were associated with the administration of g-butyrolactone. Synonyms: Dihydro-2(3H)-furanone (8CI) (9CI), 1,2-butanolide, butyrolactone, 1,4-butanolide, 4-butyrolactone, 4-hydroxybutanoic acid lactone, g-hydroxybutyric acid cyclic ester, g-hydroxybutyric acid lactone, g-lactone 4-hydroxy-butanoic acid, butyric acid lactone, butyryl lactone, 4-hydroxybutyric acid lactone, tetrahydro-2-furanone, 4-butanolide, 4-deoxytetronic acid, g-hydroxybutyrolactone

Journal Article↗
PubMed1993

Toxicology and Carcinogenesis Studies of Furan (CAS No. 110-00-9) in F344 Rats and B6C3F1 Mice(Gavage Studies).

Furan serves as an intermediate in the synthesis and preparation of numerous linear polymers used to prepare temperature-resistant structural laminates and to prepare copolymers used in machine dishwashing products as alternatives to phosphorus- and nitrogen-containing detergents. Toxicology and carcinogenesis studies were conducted by administering furan (purity > 99%) in corn oil by gavage to groups of F344/N rats and B6C3F1 mice of each sex for 16 days, 13 weeks, and 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, Drosophila melanogaster, mouse bone marrow cells, mouse L5178Y lymphoma cells, and Chinese hamster ovary cells. 16-Day Studies: Groups of five male rats received doses of 0, 5, 10, 20, 40, or 80 mg of furan per kg of body weight and groups of five female rats and five mice of each sex received doses of 0, 10, 20, 40, 80, and 160 mg/kg in corn oil by gavage. All male and female mice and female rats that received 160 mg/kg, all male and female rats and all male and four female mice that received 80 mg/kg, and three male mice that received 40 mg/kg died by day 8. Final mean body weights of male rats that received 20 mg/kg and of male and female rats that received 40 mg/kg were significantly lower than controls. Final mean body weights of male mice that received 10 or 20 mg/kg were significantly greater than controls. Mottled and enlarged livers were observed at necropsy in male rats that received 20, 40, or 80 mg/kg and in females that received 40, 80, or 160 mg/kg. No lesions were observed at necropsy that were considered related to furan administration in mice. 13-Week Studies: Groups of 10 rats of each sex and groups of 10 female mice received doses of 0, 4, 8, 15, 30, or 60 mg of furan per kg of body weight, and groups of 10 male mice received doses of 0, 2, 4, 8, 15, or 30 mg/kg in corn oil by gavage. Nine male and four female rats that received 60 mg/kg died before the end of the studies. There were no chemical-related deaths in mice. Final mean body weights of male rats that received 15 or 30 mg/kg and female rats that received 60 mg/kg were significantly lower than controls. Final mean body weights of male mice that received 60 mg/kg were significantly lower than controls. Relative and absolute liver weights in both sexes of rats and mice were increased in groups that received furan, as were relative and absolute kidney weights in female rats that received furan. Thymus weights were decreased in all groups of rats that received furan. Toxic lesions of the liver (bile duct hyperplasia, cholangiofibrosis, cytomegaly and degeneration of hepatocytes, and nodular hyperplasia of hepatocytes) were associated with furan administration in all dose groups of rats; the severity of the lesions increased with dose. Kidney lesions (tubule dilatation and necrosis of tubule epithelium) were present in rats that received 30 or 60 mg/kg. Thymic atrophy and testicular or ovarian atrophy were also observed in rats exposed to 60 mg/kg furan. Toxic liver lesions (cytomegaly, degeneration, and necrosis of hepatocytes) were also present in all groups of furan-exposed mice. Bile duct hyperplasia and cholangiofibrosis were observed in groups of mice receiving 30 or 60 mg/kg. Doses selected for the 2-year studies of rats and mice were based on the hepatotoxicity associated with exposure to furan. 2-Year Studies: Groups of 70 rats of each sex were administered 2, 4, or 8 mg furan per kg body weight in corn oil by gavage 5 days per week for 2 years. After 9 and 15 months of chemical exposure, 10 rats per group were evaluated for the presence of treatment-associated lesions. Groups of 50 mice of each sex received doses of 8 or 15 mg/kg furan 5 days per week for 2 years. Body Weight and Survival: Mean body weights of male rats that received 8 mg/kg furan were lower than controls from approximately week 73 to the end of the study. Survival of male and female rats that received 8 mg/kg was lower than controls from approximately week 85 to the end of the studies as a result of moribund condition associatedassociated with liver and biliary tract neoplasms and mononuclear cell leukemia. Mean body weights of male and female mice that received 15 mg/kg furan were lower than controls during the studies. Survival of low- and high-dose male and high-dose female mice was lower than controls from approximately week 80 to the end of the studies as a result of moribund condition associated with liver neoplasms. Neoplastic and Nonneoplastic Lesions: Cholangiocarcinoma of the liver occurred in all groups of dosed rats (males: control, 0/50; low dose, 43/50; mid dose, 48/50; high dose, 49/50; females: 0/50; 49/50; 50/50; 48/50) and was present in many rats of each sex at the 9- and 15-month interim evaluations (9-month: males - 0/10, 5/10, 7/10, 10/10; females - 0/10, 4/10, 9/10, 10/10; 15-month: males - 0/10, 7/10, 9/10, 6/10; females - 0/10, 9/10, 9/10, 7/10). Hepatocellular adenomas or carcinomas (combined) were significantly increased in male rats after 2 years of chemical administration (1/50, 5/50, 22/50, 35/50) and hepatocellular adenomas were significantly increased in female rats (0/50, 2/50, 4/50, 7/50); hepatocellular neoplasms were not observed at the 9- or 15-month interim evaluations. Increased incidences of numerous nonneoplastic liver lesions were present in rats administered furan. These lesions included biliary tract fibrosis, hyperplasia, chronic inflammation, and proliferation and hepatocyte cytomegaly, cytoplasmic vacuolization, degeneration, nodular hyperplasia, and necrosis. The incidence of mononuclear cell leukemia was increased in male and female rats that received 4 or 8 mg/kg furan (males: 8/50, 11/50, 17/50, 25/50; females: 8/50, 9/50, 17/50, 21/50); the incidence in the 8 mg/kg groups of each sex exceeded the historical control ranges for corn oil gavage studies. The severity of nephropathy increased with dose and the incidence was significantly increased in all groups of dosed rats; this increased severity was accompanied by an associated increased incidence of parathyroid hyperplasia (renal secondary hyperparathyroidism). The incidence of forestomach hyperplasia was increased in male and female rats (males: 1/50, 4/49, 7/50, 6/50; females: 0/50, 2/50, 5/50, 5/50) and the incidence of subacute inflammation of the forestomach was increased in female rats (0/50, 1/50, 5/50, 6/50). No forestomach neoplasms were observed in males; a squamous papilloma was present in one low-dose female. The incidences of hepatocellular adenomas and carcinomas were significantly increased in mice receiving furan (males: adenoma - 20/50, 33/50, 42/50; carcinoma - 7/50, 32/50, 34/50; females: adenoma - 5/50, 31/50, 48/50; carcinoma - 2/50, 7/50, 27/50). The incidences of numerous nonneoplastic hepatocellular lesions were increased in dosed mice. These lesions included hepatocyte cytomegaly, degeneration, necrosis, multifocal hyperplasia, and cytoplasmic vacuolization and biliary tract dilatation, fibrosis, hyperplasia, and inflammation. The incidences of benign pheochromocytoma and focal hyperplasia of the adrenal medulla were increased in low- and high-dose male and in high-dose female mice (benign pheochromocytoma: males - 1/49, 6/50, 10/50; females - 2/50, 1/50, 6/50). The incidences of squamous papilloma, focal inflammation, and papillary hyperplasia of the forestomach were increased in male mice (squamous papilloma: 0/49, 1/50, 3/50; focal inflammation: 9/49, 13/50, 21/50; papillary hyperplasia: 7/49, 14/50, 22/50). Stop-Exposure Study: A separate 2-year study was conducted in which 50 male rats were administered 30 mg/kg furan in corn oil by gavage 5 days per week for 13 weeks and then maintained for the remainder of the 2 years without additional furan administration. Groups of 10 animals were evaluated for the presence of treatment-related lesions at the end of the 13-week period of furan administration and at 9 and 15 months. Neoplastic and Nonneoplastic Lesions: Cholangiocarcinoma of the liver occurred with an overall incidence of 100&percnt; (40/40) and hepatocellular carcinoma occurred with an overall incidence of 15&percnt; (6/40) in stop-exposure male rats that survived at least 9 months. Cholangiocarcinoma was observed in all 10 males at both the 9-month and 15-month interim evaluations. Hepatocellular carcinoma was first observed in 2 males at the 15-month interim evaluation. Genetic Toxicology: Furan was negative for induction of gene mutations in Salmonella typhimurium strains TA100, TA1535, TA1537, and TA98 in the presence and the absence of exogenous metabolic activation (S9). Furan was negative for the induction of sex-linked recessive lethal mutations in germ cells of male Drosophila melanogaster when administered either by feeding or by injection. In vitro tests for genotoxicity in mammalian cells, however, were positive. Furan induced trifluorothymidine resistance in mouse L5178Y lymphoma cells in the absence of S9, and sister chromatid exchanges and chromosomal aberrations in Chinese hamster ovary cells, with and without S9. Furan administered to male B6C3F1 mice by intraperitoneal injection induced chromosomal aberrations but not sister chromatid exchanges in bone marrow cells. Conclusions: Under the conditions of these 2-year gavage studies there was clear evidence of carcinogenic activity of furan in male and female F344/N rats based on increased incidences of cholangiocarcinoma and hepatocellular neoplasms of the liver and on increased incidences of mononuclear cell leukemia. There was clear evidence of carcinogenic activity of furan in male and female B6C3F1 mice based on increased incidences of hepatocellular neoplasms of the liver and benign pheochromocytomas of the adrenal gland. Nonneoplastic liver lesions associated with furan administration in rats and mice included biliary tract fibrosis, hyperplasia, inflammation, and proliferation, as well as hepatocellular cytomegaly, degeneration, hyperplasia, necrosis, and vacuolization. In rats, increased severity of nephropathy with an associated increased incidence of parathyroid hyperplasia was associated with exposure to furan. Synonyms: Divinylene oxide, tetrole, furfuran, oxole, 1,4-epoxy-1,3-butadiene, axole, oxacyclopentadiene

Journal Article↗
PubMed1992

NTP Technical Report on the Toxicology and Carcinogenesis Studies of 2,4-Diaminophenol Dihydrochloride (CAS NO. 137-09-7) in F344/N Rats and B6C3F1 Mice (Gavage Studies).

2,4-Diaminophenol dihydrochloride is used in the manufacture of dyes and as a color accelerator in photographic developers. Toxicology and carcinogenesis studies were conducted by administering 2,4-diaminophenol dihydrochloride (greater than 97% pure) in corn oil by gavage to F344/N rats and B6C3F1 mice for 16 days, 13 weeks, and 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium, mouse lymphoma cells, Drosophila melanogaster, and Chinese hamster ovary cells. 16-Day Studies: Groups of five rats of each sex received doses of 0, 6, 13, 25, 50, or 100 mg/kg and groups of five mice of each sex received doses of 0, 13, 25, 50, 100, or 200 mg/kg of 2,4-diaminophenol dihydrochloride in corn oil by gavage. There were no deaths among rats during the 16-day studies and chemical exposure had no effect on final mean body weights. Organ weight differences were unrelated to chemical exposure. Renal tubule necrosis was present in male and female rats in the 25, 50, and 100 mg/kg dose groups. All male and four female mice that received 200 mg/kg, all males and three females that received 100 mg/kg, and two female mice that received 50 mg/kg died before the end of the studies. Final mean body weights of surviving dosed mice were similar to controls. Absolute liver weights were increased in female mice that received 50 and 100 mg/kg and relative liver weights were increased in all female dose groups. Renal tubule necrosis was present in the 100 mg/kg male and female dose groups. 13-Week Studies: Groups of ten male and female rats received doses of 0, 12, 25, 50, 100, or 200 mg/kg and groups of ten male and female mice received doses of 0, 5, 9, 19, 38, or 75 mg/kg of 2,4-diaminophenol dihydrochloride in corn oil by gavage. All female and nine male rats that received 200 mg/kg and four males and one female in the 100 mg/kg rat groups died before the end of the studies. Final mean body weights of male rats that received 50 or 100 mg/kg were significantly lower than controls. Relative kidney weights were significantly increased in all male dose groups; absolute and relative kidney and liver weights were significantly increased in females that received 50 and 100 mg/kg. Lesions in rats associated with chemical exposure included renal tubule necrosis in males that received 25 mg/kg and above and in females that received 100 or 200 mg/kg. Forestomach ulcers with acanthosis and hyperkeratosis were present at 50 mg/kg and above in both sexes. Pigment, presumably 2,4-diaminophenol dihydrochloride or a metabolite, was present in the duodenum and within the renal tubule epithelium of all dose groups. Splenic lymphoid depletion was present in both sexes and bone marrow hyperplasia was present in groups of males that received 100 or 200 mg/kg. Hemosiderin was present in Kupffer cells of males that received 100 or 200 mg/kg. The severity of splenic extramedullary hematopoiesis increased with dose in males. There were no deaths among mice attributed to chemical exposure. Final mean body weight of male mice that received 75 mg/kg was 8% lower than for the controls. Absolute and relative liver weights were increased in males that received 9 mg/kg or above and in all female dose groups. Absolute and relative kidney weights of 38 and 75 mg/kg male mice and all dosed female mice were significantly increased. Absolute heart weights were increased in the 19, 38, and 75 mg/kg female groups. Lesions in 75 mg/kg mice included increased incidences of renal tubule regeneration; forestomach acanthosis and hyperkeratosis occurred in 38 and 75 mg/kg groups. Hemosiderin was present in the spleen of mice from all dose groups and in hepatic Kupffer cells of female mice that received 38 or 75 mg/kg. Pigment was present in the duodenum of males and females that received 9 mg/kg and above and in the renal tubule epithelium of both sexes that received 75 mg/kg. 2-Year Studies: The 2-year studies were conducted by administering 2,4-diaminophenol dihydrochloride in corn oil by gavage to groups of 60 male and 60 female F344/N rats and B6C3F1 mice. Tence. Ten animals from each dose group were evaluated after 15 months. Rats received doses of 0, 12.5, or 25 mg/kg; mice received doses of 0, 19, or 38 mg/kg. Body Weight and Survival in the 2-Year Studies: Mean body weights of high-dose male and female rats and of high-dose male mice were lower than those of the respective controls. Survival of dosed rats and mice was similar to controls throughout the 2-year studies. Neoplastic and Nonneoplastic Effects in the 2-Year Studies: The severity of nephropathy increased in high-dose male rats. The incidence of renal tubule cell hyperplasia was increased in rats that received 2,4-diaminophenol dihydrochloride; however, there were no renal tubule cell neoplasms attributed to chemical exposure. Renal tubule cell hyperplasia was present in three high-dose male mice and a renal tubule adenoma was present in one of the high-dose males with hyperplasia; two additional high-dose males also had renal tubule adenomas. These are uncommon neoplasms in male mice and were considered related to chemical exposure. In a supplemental review of step-sectioned kidneys, six additional hyperplasia were seen in high-dose male mice and adenomas in three other high-dose males. A tubule cell carcinoma was present in one low-dose female mouse. The incidences of several nonneoplastic kidney lesions, including necrosis and renal tubule regeneration, were also increased in male and female mice. Pigment was observed in the lamina propria of the duodenum, the submucosa of the forestomach, and in pancreatic and mesenteric lymph nodes in dosed rats. Hemosiderin was present in the renal tubule epithelial cells of low- and high-dose rats. The incidence of forestomach acanthosis was increased in dosed male mice and pigment was present in liver Kupffer cells, in the lamina propria of the duodenum, and in the mesenteric lymph nodes of male and female mice. Pigment and hemosiderin were also present in the renal tubule epithelium of dosed mice. Genetic Toxicology: 2,4-Diaminophenol dihydrochloride was mutagenic in the Salmonella typhimurium strain TA98 in the presence of exogenous metabolic activation (S9); it was not mutagenic in strain TA98 in the absence of S9, nor was it mutagenic in strains TA100, TA1535, or TA1537 with or without S9. 2,4-Diaminophenol dihydrochloride was positive in the mouse lymphoma assay for induction of trifluorothymidine resistance in L5178Y cells without S9; it was not tested with S9. No induction of sister chromatid exchanges or chromosomal aberrations was observed in Chinese hamster ovary cells treated with 2,4-diaminophenol dihydrochloride with and without S9. In the Drosophila melanogaster sex-linked recessive lethal assay, 2,4-diaminophenol dihydrochloride gave equivocal results when administered in the feed and negative results when administered by injection. Conclusions: Under the conditions of these 2-year gavage studies, there was no evidence of carcinogenic activity of 2,4-diaminophenol dihydrochloride in male or female F344/N rats that received 12.5 or 25 mg/kg. There was some evidence of carcinogenic activity of 2,4-diaminophenol dihydrochloride in male B6C3F1 mice based on increased incidences of renal tubule adenomas; there was no evidence of carcinogenic activity of 2,4-diaminophenol dihydrochloride in female B6C3F1 mice that received 19 or 38 mg/kg. Administration of 2,4-diaminophenol dihydrochloride to rats was associated with increased severity of nephropathy in males and females, increased incidence of nephropathy in females, and focal renal tubule hyperplasia in males and females. In mice, chemical exposure was associated with renal tubule necrosis and regeneration in males and females and forestomach acanthosis in males. Synonyms: Acrol, amidol, dianol

Journal Article↗
PubMed1992

Toxicology and Carcinogenesis Studies of 4,4'-Diamino-2,2'-Stilbenedisulfonic Acid Disodium Salt (CAS No. 7336-20-1) in F344 Rats and B6C3F1 Mice (Feed Studies).

4,4'-Diamino-2,2'-stilbenedisulfonic acid, disodium salt, is used in the synthesis of dyes and optical brighteners or fluorescent whitening agents. Toxicology and carcinogenesis studies were conducted by administering the chemical (approximately 14% water, 6% sodium chloride, 4% impurities, and 76% 4,4'-diamino-2,2'-stilbenedisulfonic acid) in feed to groups of F344/N rats and B6C3F1 mice of each sex for 14 days, 13 weeks, and 2 years. Genetic toxicology studies were conducted in Salmonella typhimurium and Chinese hamster ovary cells. 14-Day Studies: Groups of five rats and five mice of each sex were given 0, 6,250, 12,500, 25,000, 50,000, or 100,000 ppm 4,4'-diamino-2,2'-stilbenedisulfonic acid, disodium salt, in feed for 14 days. All rats and mice survived to the end of the studies. The mean body weight gain of male rats receiving 50,000 or 100,000 ppm and of female rats and male and female mice receiving 100,000 ppm was significantly lower than those of the respective controls. Clinical findings included diarrhea in the rats and mice receiving 100,000 ppm. There were no chemical-related changes in absolute or relative organ weights in rats or mice. There were no gross or microscopic lesions related to chemical administration in rats or mice. 13-Week Studies: Groups of 10 rats and 10 mice of each sex were given 0, 6,250, 12,500, 25,000, 50,000, or 100,000 ppm 4,4'-diamino-2,2'-stilbenedisulfonic acid, disodium salt, in feed for 13 weeks. One female rat, six male mice, and one female mouse in the 100,000 ppm dose groups died during the studies. Mean body weight gain was significantly decreased in male rats and female mice receiving 50,000 or 100,000 ppm, in male mice receiving 25,000, 50,000, or 100,000 ppm, and in female rats receiving 100,000 ppm. Clinical findings in rats that received 50,000 or 100,000 ppm and in mice that received 100,000 ppm included diarrhea, emaciation, and hyperemia of the perineum. There were no biologically significant changes in absolute or relative organ weights or clinical pathology results in rats or mice. Histopathologic lesions present in rats receiving 100,000 ppm were bone marrow hypercellularity and chronic inflammation of the anus and rectum. Ulcerative inflammation of the anus and rectum was observed in mice receiving 25,000 ppm and above. Female mice in the 6,250, 12,500, 25,000, and 50,000 ppm dose groups had increased incidences of cystic endometrial hyperplasia. 2-Year Studies: Doses selected for the 2-year studies were based on mortality, decreased body weight gains, and the presence of diarrhea and chronic inflammation of the anus/rectum in rats and mice during the 13-week studies. Groups of 60 rats of each sex were given 0, 12,500 or 25,000 ppm and groups of 60 mice of each sex were given 0, 6,250, or 12,500 ppm 4,4'-diamino-2,2'-stilbenedisulfonic acid, disodium salt, in feed for up to 103 weeks. Interim evaluations were performed on 10 rats and 10 mice from each dose group at 15 months. There were no biologically significant absolute or relative organ weight, clinical pathology, or histopathology findings in rats or mice administered 4,4'-diamino-2,2'-stilbenedisulfonic acid, disodium salt, in feed for 15 months. Body Weight, Feed Consumption, Survival, and Clinical Findings in the 2-Year Studies: Mean body weights were marginally decreased for high-dose male and female rats and female mice. Feed consumption by dosed rats and mice was similar to feed consumption by the controls throughout the studies. Survival was similar among control and treated groups of rats and mice. No clinical findings related to chemical administration were observed in rats or mice. Nonneoplastic and Neoplastic Effects in the 2-Year Studies: There were no chemical-related increased incidences of neoplasms at any site in rats. Ulcers of the forestomach or glandular stomach occurred in dosed rats (males: 1/50, 5/50, 4/50; females: 0/50, 1/50, 4/50), and may have been related to the administration of 4,4'-diamino-2,2'-stilbenedisulfonic acid, disodium salt. There were no chemical-related incidences ohemical-related incidences of neoplasms, nonneoplastic lesions, or other toxic effects in mice in the 2-year studies. Although the animals might have been able to tolerate slightly higher doses, results of the 13-week studies indicate that a doubling of the highest doses could not have been tolerated. Genetic Toxicology: 4,4'-Diamino-2,2'-stilbenedisulfonic acid was not mutagenic in Salmonella typhimurium strains TA100, TA1535, TA1537, or TA98 with or without S9 metabolic activation. 4,4'-Diamino-2,2'-stilbenedisulfonic acid did not induce sister chromatid exchanges or chromosomal aberrations in Chinese hamster ovary cells in the presence or absence of S9. Conclusions: Under the conditions of these 2-year feed studies, there was no evidence of carcinogenic activity of 4,4'-diamino-2,2'-stilbenedisulfonic acid, disodium salt, in male or female F344/N rats receiving 12,500 or 25,000 ppm. There was no evidence of carcinogenic activity of 4,4'-diamino-2,2'-stilbenedisulfonic acid, disodium salt, in male or female B6C3F1 mice receiving 6,250 or 12,500 ppm. Synonyms: Amsonic acid; diaminostilbene disulphonate (DASD); 2,2'-(1,2-ethenediyl)bis[5-amino-benzenesulfonic acid]; 2,2'-disulfo-4,4'-stilbenediamine; 2,2'-stilbenedisulfonic acid; 4,4'-diamino-2,2'-benzenesulfonic acid; 2,2'-(1,2-ethenediyl)bis(5-amino-) diaminostilbenedisulfonic acid; flavonic acid; p,p'-diaminostilbene-o,o'-disulfonic acid; 4,4'-diaminostilbene-2,2'-disulfonic acid

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