NATIONAL TOXICOLOGY PROGRAM Technical Report Series No. 332 TOXICOLOGY AND CARCINOGENESIS STUDIES OF 2-MERCAPTOBENZOTHIAZOLE (CAS NO. 149-30-4) IN F344/N RATS AND B6C3F1 MICE (GAVAGE STUDIES) U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES Public Health Service National Institutes of Health NATIONAL TOXICOLOGY PROGRAM The National Toxicology Program (NTP), established in 1978,develops and evaluates scientific information about potentially toxic and hazardous chemicals. This knowledge can be used for protecting the health of the American people and for the primary prevention of disease. By bringing to- gether the relevant programs, staff, and resources from the US. Public Health Service, DHHS, the National Toxicology Program has centralized and strengthened activities relating to toxicology research, testing and test developmenthalidation efforts, and the dissemination of toxicological in- formation to the public and scientific communities and to the research and regulatory agencies. The NTP is made up of four charter DHHS agencies: the National Cancer Institute (NCI), National Institutes of Health; the National Institute of En- vironmental Health Sciences (NIEHS), National Institutes of Health; the National Center for Toxicological Research (NCTR), Food and Drug Ad- ministration; and the National Institute for Occupational Safety and Health (NIOSH), Centers for Disease Control. In July 1981, the Carcino- genesis Bioassay Testing Program, NCI, was transferred to the NIEHS. 2-Mercaptobenzothiazole,NTP TR 332 NTP TECHNICAL REPORT ON THE TOXICOLOGY AND CARCINOGENESIS STUDIES OF 2-MERCAPTOBENZOTHIAZOLE (CAS NO. 149-30-4) IN F344/N RATS AND B6C3F1 MICE (GAVAGE STUDIES) Michael P. Dieter, Ph.D., Chemical Manager NATIONAL TOXICOLOGY PROGRAM P.O. Box S2233 Research Triangle Park, NC 27709 May 1988 NTP TR 332 NIH Publication No. 88-2588 U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES Public Health Service National Institutes of Health NOTETOTHEREADER This study was performed under the direction of the National Institute of Environmental Health Sci- ences as a function of the National Toxicology Program. The studies described in this Technical Re- port have been conducted in compliance with NTP chemical health and safety requirements and must meet or exceed all applicable Federal, state, and local health and safety regulations. Animal care and use were in accordance with the U.S.Public Health Service Policy on Humane Care and Use of Ani- mals. All NTP toxicology and carcinogenesis studies are subjected to a data audit before being pre- sented for public peer review. Although every effort is made to prepare the Technical Reports as accurately as possible, mistakes may occur. Readers are requested to identify any mistakes so that corrective action may be taken. Further, anyone who is aware of related ongoing or published studies not mentioned in this report is encouraged to make this information known to the NTP. Comments and questions about the National Toxicology Program Technical Reports on Toxicology and Carcinogenesis Studies should be directed to Dr. J.E. Huff, National Toxicology Program, P.O. Box 12233,Research Triangle Park, NC 27709 (919-541-3780). These NTP Technical Reports are available for sale from the National Technical Information Service, U.S.Department of Commerce, 5285 Port Royal Road, Sprin~ield,VA 22161 (703-487-4650). Single copies of this Technical Report are available without charge (and while supplies last) from the NTP Public Information Office, National Toxicology Program, P.O.Box 12233, Research Triangle Park, NC 27709. 2-Mercaptobenzothiazole, NTP TR 332 2 CONTENTS PAGE NOTE TO THE READER ................................................................... 2 ABSTRACT ............................................................................... 5 EXPLANATION OF LEVELS OF EVIDENCE OF CARCINOGENIC ACTIVITY ....................... 8 CONTRIBUTORS .......................................................................... 9 PEER REVIEW PANEL ................................................................... io SUMMARY OF PEER REVIEW COMMENTS .................................................. ii I. INTRODUCTION ................................................................... 13 II. MATERIALS AND METHODS ........................................................ 19 PROCUREMENT AND CHARACTERIZATION OF 2-MERCAPTOBENZOTHIAZOLE ........20 PREPARATION AND CHARACTERIZATION OF DOSE MIXTURES ...................... 25 FIRST SIXTEEN-DAY STUDIES .................................................... 27 SECOND SIXTEEN-DAY STUDIES .................................................. 27 THIRTEEN-WEEK STUDIES ....................................................... 27 TWO-YEAR STUDIES ............................................................. 27 STUDY DESIGN ............................................................... 27 SOURCE AND SPECIFICATIONS OF ANIMALS .................................... 27 ANIMAL MAINTENANCE ....................................................... 30 CLINICAL EXAMINATIONS AND PATHOLOGY .................................... 30 STATISTICAL METHODS ....................................................... 31 III. RESULTS ......................................................................... 33 RATS .......................................................................... 34 SIXTEEN-DAY STUDIES ........................................................ 34 THIRTEEN-WEEK SI"NJIES ..................................................... 34 TWO-YEAR STUDIES .......................................................... 35 BODY WEIGHTS AND CLINICAL SIGNS ........................................ 35 SURVIVAL ................................................................. 38 PATHOLOGY AND STATISTICAL ANALYSES OF RESULTS ....................... 38 MICE .......................................................................... 46 SIXTEEN-DAY STUDIES ....................................................... -46 THIRTEEN-WEEK STUDIES .................................................... -47 TWO-YEAR STUDIES ........................................................... 48 BODY WEIGHTS AND CLINICAL SIGNS ........................................ 48 3 2.Mercaptobenzothiazole. NTP TR 332 CONTENTS (Continued) PAGE SURVIVAL . e. 51 PATHOLOGY AND STATISTICAL ANALYSES OF RESULTS , ... , .,.,... ..,.. .51 IV. DISCUSSION AND CONCLUSIONS .. ... ,... ... ..... .. ..... ..... .55 V. REFERENCES ................................................................... 0.59 APPENDIXES APPENDIX A SUMMARY OF LESIONS IN MALE RATS IN THE TWO-YEAR GAVAGE STUDY OF 2-MERCAPTOBENZOTHIAZOLE .. .. ...... ... ..65 APPENDIX B SUMMARY OF LESIONS IN FEMALE RATS IN THE TWO-YEAR GAVAGE STUDY OF 2-MERCAPTOBENZOTHIAZOLE .. , .. ... .. ... .89 APPENDIX C SUMMARY OF LESIONS IN MALE MICE IN THE TWO-YEAR GAVAGE STUDY OF 2-MERCAPTOBENZOTHIAZOLE . ..... .. .I11 APPENDIX D SUMMARY OF LESIONS IN FEMALE MICE IN THE TWO-YEAR GAVAGE STUDY OF 2-MERCAPTOBENZOTHIAZOLE .... .. ... .. ..I31 APPENDIX E GENETIC TOXICOLOGY OF 2-MERCAPTOBENZOTHIAZOLE ,.. , ....,.I@ APPENDIX F SENTINEL ANIMAL PROGRAM ,.... .. ..... .... ... ... ,161 APPENDIX G INGREDIENTS, NUTRIENT COMPOSITION, AND CONTAMINANT LEVELS IN NIH 07 RAT AND MOUSE RAT'ION ... .... .... .... ..165 APPENDIX H AUDIT SUMMARY ... .. .. ....... ..... ...... ,171 2-Mercaptobenzothiazole,NTP TR 332 4 2-MERCAPTOBENZOTHIAZOLE CAS NO.149-30-4 C7H5NSZ Molecular weight 167.25 Synonyms and trade names: Captax, Dermacid, Mertax, Thiotax, 2(3H)-Benzothiazolethione, 2-Benzothiazolyl mercaptan ABSTRACT Toxicology and carcinogenesis studies of technical-grade 2-mercaptobenzothiazole (96%-97% pure), a rubber accelerant and preservative, were conducted by administering the chemical by gavage in a corn oil vehicle to groups of F344/N rats and B6C3F1 mice of each sex for 16 days, 13 weeks, or 2 years. 2-Mercaptobenzothiazole was nominated for study by the National Institute of Environmental Health Sciences and the National Institute for Occupational Safety and Health. Sixteen-Day and Thirteen-Week Studies: In 16-day studies, mean body weight gains of rats receiving 2,500 mgkg were 6-7 g lower than those of vehicle controls; 415 male and 515 female mice dosed with 3,000 mg/kg and 415 female mice dosed with 1,500 mg/kg died; lethargy and prostration occurred in most of these animals after gavage. Based on these results, doses selected for both species in the 13- week studies were 0,94 (mice only), 188,375,750, and 1,500 mg/kg. In 13-week studies, no chemical-related deaths occurred in rats, but body weight gains in males dosed with 1,500 mg/kg and in females dosed with 750 or 1,500 mg/kg were lower than those in the vehicle control groups. Hepatomegaly occurred at the two highest doses in males and at all doses in females; however, no microscopic pathologic changes were noted in any tissue. More than half the mice dosed with 1,500 mg/kg died, but no compound-related body weight changes occurred. Clinical signs in mice were dose related and included lethargy in animals dosed with 375 mg/kg and lacrimation, sali- vation, and clonic seizure in some dosed with 750 or 1,500 mg/kg. No association between these clini- cal signs of toxicity and gross or microscopic pathologic effects was observed. Doses selected for the 2- year studies were 0, 375, and 750 mg/kg for male rats and for mice of each sex and 0, 188, or 375 mg/kg for female rats. Body Weight and Survival in the Two-year Studies: Fifty animals of each species and sex were ad- ministered 2-mercaptobenzothiazole