NATIONAL TOXICOLOGY PROGRAM Technical Report Series No. 259 \,~ s:UlVIC/E,r. ­ ('~ CARCINOGENESIS STUDIES OF ETHYL ACRYLATE (CAS NO. 140-88-5) IN F344/N RATS AND B6C3Ft 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 ofdisease. By bringing to­ gether the relevant programs, staff, and resources from the U.S. Public Health Service, DHHS, the National Toxicology Program has centralized and strengthened activities relating to toxicology research, testing and test development/validation 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 ofHealth; 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. Special Note: A draft of this Technical Report was peer reviewed in public session and approved by the NTP Board of Scientific Counselors' Technical Reports Review Subcommittee in February 1983, before the NTP adopted use of levels ofevidence ofcarcinogenicity for its carcinogenesis studies. In October 1983, the NTP adopted the policy that the experimental data and laboratory records from all NTP Toxicology and Carcinogenesis Studies not yet printed and distributed would be audited. [A summary of the data audit is presented in Appendix N.] Consequently, printing and distribution of this Technical Report have been delayed and the format differs from that of Technical Reports peer reviewed more recently. This final Technical Report supercedes all previous versions of this report that have been distributed. Ethyl Acrylate NTP TECHNICAL REPORT ON THE CARCINOGENESIS STUDIES OF ETHYL ACRYLATE (CAS NO. 140-88-5) IN F344/N RATS AND B6C3F1 MICE (GAVAGE STUDIES) NATIONAL TOXICOLOGY PROGRAM P.O. Box 12233 Research Triangle Park, NC 27709 December 1986 NTP TR 259 NIH Publication No. 87-2515 U.S. DEPARTMENT OF HEALTH AND HUMAN SERVICES Public Health Service National Institutes of Health NOTE TO THE READER This is one in a series of experiments designed to determine whether selected chemicals produce cancer in animals. Chemicals selected for testing in the NTP carcinogenesis program are chosen primarily on the bases of human exposure, level of production, and chemical structure. Selection per se is not an indicator of a chemical's carcinogenic potential. Negative results, in which the test animals do not have a greater incidence of cancer than control animals, do not necessarily mean that a test chemical is not a carcinogen, inasmuch as the experiments are conducted under a limited set of conditions. Positive results demonstrate that a test chemical is carcinogenic for animals under the conditions of the test and indicate that exposure to the chemical has the potential for hazard to humans. The determination of the risk to humans from chemicals found to be carcinogenic in animals requires a wider analysis which extends beyond the purview of this study. This study was initiated by the National Cancer Institute's Carcinogenesis Testing Program, now part of the National Institute of Environmental Health Sciences, National Toxicology Program. Comments and questions about the National Toxicology Program Technical Reports on Carcino­ genesis Studies should be directed to the National Toxicology Program, located at Research Triangle Park, NC 27709 (919-541-3991) or at Room 835B, Westwood Towers, 5401 Westbard Ave., Bethesda, MD 20205 (301-496-1152). Although every effort is made to prepare the Technical Reports as accurately as possible, mistakes may occur. Readers are requested to communicate any mistakes to the Deputy Director, NTP (P.O. Box 12233, Research Triangle Park, NC 27709), so that corrective action may be taken. Further, anyone who is a ware of related ongoing or published studies not mentioned in this report is encouraged to make this information known to the NTP. These NTP Technical Reports are available for sale from the National Technical Information Service, U.S. Department of Commerce, 5285 Port Royal Road, Springfield, VA 22161 (703-487­ 4650). Single copies of this carcinogenesis studies 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. Ethyl Acrylate 2 TABLE OF CONTENTS Page Abstract.......................................................................... 7 Contributors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8 Reviewers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10 Summary of Peer Review Comments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . 11 I. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13 Chemical Identification.. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14 Production and Use ........................................................... 14 Metabolism . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14 Animal Toxicity and Teratogenicity ............................................. 15 Human Toxicity and Exposure ................................................. 15 Mutagenicity and Carcinogenicity ............................................... 16 Rationale for Testing .......................................................... 17 II. Materials and Methods ........................................................ 19 Chemical Analyses . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20 Dose Preparation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20 Testing Chronology ........................................................... 21 Single-Dose Studies ........................................................... 21 Fourteen-Day Studies ......................................................... 22 Thirteen-Week Studies ........................................................ 22 Two-Year Studies. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23 Study Design .............................................................. 23 Source and Specifications of Test Animals ..................................... 23 Animal Maintenance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23 Clinical Examinations and Pathology ......................................... 24 Data Recording and Statistical Methods. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24 III. Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 31 Rats ........................................................................ 32 Single-Dose Studies ........................................................... 32 Fourteen-Day Studies ......................................................... 32 Thirteen-Week Studies ........................................................ 34 Two-Year Studies. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35 Body Weights and Clinical Signs ............................................. 35 Survival . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 36 Pathology and Statistical Analyses of Results .................................. 38 Mice ........................................................................ 41 Single-Dose Studies ........................................................... 41 Fourteen-Day Studies ......................................................... 41 Thirteen-Week Studies ........................................................ 43 Two-Year Studies. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 45 Body Weights and Clinical Signs ............................................. 45 Survival ................................................................... 47 Pathology and Statistical Analyses of Results .................................. 48 IV. Discussion and Conclusions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 51 V. References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 57 TABLES Table I Testing Chronology for Ethyl Acrylate .................................. 21 Table 2 Experimental Design and Materials and Methods ......................... 26 Table 3 Survival and Mean Body Weights of Rats Administered Ethyl Acrylate by Gavage for 14 Days (First Study) ............................ 33 Table 4 Survival and Mean Body Weights of Rats Administered Ethyl Acrylate by Gavage for 14 Days (Third Study) ........................... 33 3 Ethyl Acrylate Table 5 Survival and Mean Body Weights of Rats Administered Ethyl Acrylate by Gavage for 13 Weeks ....................................... 34 Table 6 Mean Body Weights (Relative to Controls) of Rats Administered Ethyl Acrylate by Gavage in the Two-Year Studies ....................... 36 Table 7 Numbers of Rats with Forestomach Lesions ............................. 38 Table 8 Analysis of Forestomach Tumors in Rats ................................ 39 Table 9 Numbers of Rats with Retinopathy or Cataracts .......................... 40 Table 10 Survival and Mean Body Weights of Mice Administered Ethyl Acrylate by Gavage for 14 Days (First Study) ............................ 41 Table ll Survival and Mean Body Weights of Mice Administered Ethyl Acrylate by Gavage for 14 Days (Third Study) ........................... 42 Table 12 Survival and Mean Body Weights of Mice Administered Ethyl Acrylate by Gavage for 13 Weeks (First Study) ........................... 43 Table 13 Survival and Mean Body Weights of Mice Administered Ethyl Acrylate by Gavage for 13 Weeks (Second Study) ........................ 44 Table 14 Mean Body Weights (Relative to Controls) of Mice Administered Ethyl Acrylate by Gavage in the Two-Year Studies ....................... 46 Table 15 Numbers of Mice with Forestomach Lesions ............................. 48 Table 16 Analysis of Forestomach Tumors in Mice ............................... 49 Table 17 Negative Trends in Overall Tumor Incidences in Mice. . . . . . . . . . . . . . . . . . . . . 50 Table 18 Comparison of Forestomach Tumors in Rats and Mice Based on Ethyl Acrylate Concentration in the Corn Oil Gavage Solution . . . . . . . . . . . . . 53 FIGURES Figure l Growth Curves for Rats Administered Ethyl Acrylate in Corn Oil by Gavage . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35 Figure 2 Survival Curves for Rats Administered Ethyl Acrylate in Corn Oil by Gavage ........................................................... 37 Figure 3 Growth Curves for Mice Administered Ethyl Acrylate in Corn Oil by Gavage ........................................................... 45 Figure 4 Survival Curves for Mice Administered Ethyl Acrylate in Corn Oil by Gavage ........................................................... 47 Figure 5 Infrared Absorption Spectrum of Ethyl Acrylate (Lot No. 37201) ........... 177 Figure 6 Infrared Absorption Spectrum of Ethyl Acrylate (Lot No. 343029) .......... 178 Figure 7 Nuclear Magnetic Resonance Spectrum of Ethyl Acrylate (Lot No. 37201) ...................................................... 180 Figure 8 Nuclear Magnetic Resonance Spectrum of Ethyl Acrylate (Lot No. 343029) ..................................................... I8l Ethyl Acrylate 4 APPENDIXES Appendix A Summary of the Incidence of Neoplasms in Rats Administered Ethyl Acrylate in Corn by Gavage ...................................... 61 Table AI Summary of the Incidence of Neoplasms in Male Rats Administered Ethyl Acrylate in Corn Oil by Gavage ................................... 62 Table A2 Summary of the Incidence of Neoplasms in Female Rats Administered Ethyl Acrylate in Corn Oil by .Gavage ................................... 68 Table A3 Individual Animal Tumor Pathology of Male Rats in the Two-Year Study of Ethyl Acryla'te . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 72 Table A4 Individual Animal Tumor Pathology of Female Rats in the Two-Year Study of Ethyl Acrylate ...................................... 78 Appendix B Summary of the Incidence of Neoplasms in Mice Administered Ethyl Acrylate in Corn Oil by Gavage ................................... 85 Table Bl Summary of the Incidence of Neoplasms in Male Mice Administered Ethyl Acrylate in Corn Oil by Gavage . . . . . . . . . . . . . . . . . . . . . . 86 Table B2 Summary of the Incidence of Neoplasms in Female Mice Administered Ethyl Acrylate in Corn Oil by Gavage ...................... 90 Table B3 Individual Animal Tumor Pathology of Male Mice in the Two-Year Study of Ethyl Acrylate ...................................... 96 Table B4 Individual Animal Tumor Pathology of Female Mice in the Two-Year Study of Ethyl Acrylate ...................................... 102 Appendix C Summary of the Incidence of Nonneoplastic Lesions in Rats Administered Ethyl Acrylate in Corn Oil by Gavage ...................... 109 Table Cl Summary of the Incidence of Nonneoplastic Lesions in Male Rats Administered Ethyl Acrylate in Corn Oil by Gavage .................. 110 Table C2 Summary of the Incidence of Nonneoplastic Lesions in Female Rats Administered Ethyl Acrylate in Corn Oil by Gavage .................. 116 Appendix D Summary of the Incidence of Nonneoplastic Lesions in Mice Administered Ethyl Acrylate in Corn Oil by Gavage ...................... 121 Table 01 Summary of the Incidence of Nonneoplastic Lesions in Male Mice Administered Ethyl Acrylate in Corn Oil by Gavage ................. 122 Table 02 Summary of the Incidence of Nonneoplastic Lesions in Female Mice Administered Ethyl Acrylate in Corn Oil by Gavage .................128 Appendix E Historical Incidences of Tumors in F344/ N Rats and B6C3Fl Mice Receiving Corn Oil by Gavage ............................ 135 Table El Historical Incidence of Stomach Tumors in Male F344/N Rats Receiving Corn Oil by Gavage ............................. 136 Table E2 Historical Incidence of Stomach Tumors in Female F344/N Rats Receiving Corn Oil by Gavage ............................. 136 Table E3 Historical Incidence of Pancreatic Acinar Cell Adenomas in Male F344/N Rats Receiving Corn Oil by Gavage ...................... 137 Table E4 Historical Incidence of Liver Tumors in Male B6C3F Mice 1 Receiving Corn Oil by Gavage .........................................137 Table E5 Historical Incidence of Stomach Tumors in Male B6C3Fl Mice Receiving Corn Oil by Gavage .........................................138 Table E6 Historical Incidence of Stomach Tumors in Female B6C3F1 Mice Receiving Corn Oil by Gavage ......................................... 138 5 Ethyl Aqrylate Table E7 Historical Incidence oflntegumentary Basal Cell Tumors in Male F344/N Rats Receiving Corn Oil by Gavage ••••••••••••••••••••••••••••••••••••••••••••••••••.•••••••• 139 Table E8 Historical Incidence of Adrenal Tumors in Male F344/N Rats Receiving Corn Oil by Gavage .•••••• 139 Table E9 Historical Incidence ofThyroid Tumors in Male B6C3F Mice Receiving Corn Oil by Gavage •••••• 140 1 Table ElO Historical Incidence of Hematopoietic Tumors in Male B6C3F Mice Receiving Corn Oil 1 by Gavage ••••.•••••.••••••••••••••••••••••••••••••••••••••.....••..•••••.•• 140 Appendix F Analysis of Primary Tumors in Rats and Mice •• , •• , •• , •• , • , , ••• , •••••••••• , •••••••••• 141 Table Fl Analysis of Primary Tumors in Male Rats • , •• , , , •• , , , •• , , • , • , , , ••••••••••• 142 o ••••••••• Table F2 Analysis of Primary Tumors in Female Rats , •••••••••••••• , • , •••••••••••••••••••••••• 148 Table F3 Analysis of Primary Tumors in Male Mice •••••••••••••••• , , ••••••••••••••••••••••••• 151 Table F2 Analysis of Primary Tumors in Female Mice ••••••••••••••••••••••••••••••••••••••••• 156 Appendix G Mutagenesis Results for Ethyl Acrylate in Salmonella Typhimurium ••••• , , ••••••••••••••••• 159 Table Gl Results of Mutagenicity Tests of Ethyl Acrylate In Salmonella Typhimurium TA98 Performed at SRI International , •••••••••••••••••••••••••••••••••••••••••••••••••••••••••• 161 Table G2 Results of Mutagenicity Tests of Ethyl Acrylate in Salmonella Typhimurium TA98 Performed at Case Western Reserve University ••••••••••••••••••••••••••••••••••••••..••••••• 162 Table G3 Results of Mutagenicity Tests of Ethyl Acrylate in Salmonella Typhimurium TAtOO Performed at SRI International , •••••.•••••••••••••••••••••••••••••••••••••••••••••.•••••• 163 Table G4 Results of Mutagenicity Tests of Ethyl Acrylate in Salmonella Typhimurfum TAtOO Performed at Case Western Reserve University ••••••••••••••••••••••••••••••••••••••••••••••• 164 Table G5 Results ofMutagenicity Tests of Ethyl Acrylate in Salmonella Typhimurium TA1535 Performed at SRI International .•••••••••••••••••••••••••••••••••••••••••••••••••••••••••• 165 Table G6 Results of Mutagenicity Tests of Ethyl Acrylate in Salmonella Typhimurium TA1535 Performed at Case Western Reserve University ••••••••••••••••••••••••••••••••••••••••••••••• 166 Table G7 Results of Mutagenicity Tests of Ethyl Acrylate in Salmonella Ty/)himurium TA1537 Performed at SRI International .•••••••••••••••••••••••••••••••••••••••••••••••••••••••••• 167 Table G8 Results of Mutagenicity Tests of Ethyl Acrylate in Salmonella Typhimurium TA1537 Performed at Case Western Reserve University •••••••••••••••••••••••••••••••••••.••••••••••• 168 Appendix H Sentinel Animal Serology Data for the Ethyl Acrylate Bioassay •••• , •••••••••••••••••••••• 169 Table H 1 Murine Virus Antibody Determinations for Rats in the Two-Year Study .••••••••••••••••••••• 171 Table H2 Murine Virus Antibody Determinations for Mice in the Two-Year Study ••••••••••••••••••••• 172 Appendix I Analysis of Ethyl Acrylate •••••••••••••••••••••••••••••••••••••••••••••••••••••• 173 Appendix J Analysis of Ethyl Acrylate in Waterfor Stability of Ethyl Acrylate •• , •••••••••••••••••••••• 183 Appendix K Analysis of Ethyl Acrylate in Corn Oil for Stability of Ethyl Acrylate--Midwest Research Institute o. 185 Appendix L Analysis of Ethyl Acrylate in Corn Oil for Concentrations of Ethyl Acrylate ••••••••••••••••••• 187 Table Ll Analysis of Ethyl Acrylate in Corn Oil 188 o ••••••••••••••••••••••••••••••••••••••••••••• Appendix M Metabolism of Ethyl Acrylate in the Stomachs of Male and Female F344.1N Rats 189 o ••••••••••••••• Appendix N Data Audit Summary .••••••••••••••••••••••••••••••••••••••••••••••••••••••••• 223 Ethyl Acrylate 6 CARCINOGENESIS STUDIES OF ETHYL ACRYLATE H....._ ,.,.H c=c H' ' COCH2CH3 II 0 ETHYL ACRYLATE CAS NO. 140-88-5 Mol. Wt. 100.12 ABSTRACT Carcinogenesis studies of ethyl acrylate were conducted by administering this test chemical in corn oil by gavage to groups of 50 male and 50 female F344/ N rats and B6C3F1 mice at doses of 100 or 200 mgj kg. Ethyl acrylate was administered five times per week for 103 weeks. Groups of 50 rats and 50 mice of each sex received corn oil by gavage on the same schedule and served as vehicle controls. Survival of dosed male and female rats and mice was comparable with that of the corresponding vehicle controls. There was no evidence of systemic toxicity in the prechronic or in the 2-year studies. Compound-related increased incidences of hyperkeratosis, inflammation, and hyperplasia of the forestomach were observed in rats and mice in the prechronic as well as 2-year studies. In the 2-year studies, squamous cell papillomas and squamous cell carcinomas of the forestomach occurred at the site of chemical deposition with significant positive trends and increased incidences in dosed groups versus vehicle controls for both sexes of rats and mice. Nonneoplastic and neoplastic forestomach lesion frequencies were related to the concentration of ethyl acrylate in dosing solutions used. Significant negative trends for several common rodent tumors were found in treated animals in the 2-year studies. Under the conditions ofthese studies, ethyl acrylate was carcinogenic for the forestomach of F344/ N rats and B6C3F1 mice, causing squamous cell carcinomas in male rats and male mice, squamous cell papillomas in male and female rats and male mice, and squamous cell papillomas or carcinomas (combined) in male and female rats and mice. Evidence for carcinogenicity was greater in males than in females. Ethyl acrylate also caused irritation of the forestomach mucosa in male and female rats and mice. 7 Ethyl Acrylate Ethyl Acrylate 8