Formation of Glutathione Conjugates by Reactive Metabolites of Vinylidene Chloride in Microsomes and Isolated Hepatocytes1

Oxidation of the vinyl halide carcinogen and hepatotoxin vinylidene chloride (VDC) by microsomal cytochrome P-450 yields 2,2-dichloroacetaldehyde, 2-chloroacetyl chloride, 2-chloroacetic acid, and 1,1-dichloroethylene oxide. The roles of these metab olites in covalent modification of proteins and reduced glutathione (GSH) were examined. 2-Chloroacetyl chloride reacted with model thiols at least 103-fold faster than did 1,1-dichloroethylene oxide and at least 105-fold faster than did 2,2-dichloroacetalde hyde or 2-chloroacetic acid. Microsomal covalent binding of [14C]VDC was inhibited by GSH but not by lysine, suggesting that protein thiols, rather than amino groups, are major targets. Liver microsomes catalyzed the formation of three GSH:VDC metabolite conjugates, identified as S-(2,2-dichloro-1-hydroxy)ethylglutathione, 2-(S-glutathionyl)acetate, and S-(2-glutathionyl)acetylglutathione, a novel conjugate containing both sta ble (thioether) and labile (thioester) linkages. The latter two conjugates also were formed in isolated rat hepatocytes and measurable amounts of 2-{S-glutathionyl)acetate were released into the incubation medium. Both 2-(S-g!utathionyl)acetate and S-(2-glutathionyl)acetylglutathione were formed with [^SjGSH added to the hepatic medium, indicating that reactive VDC metabolites are capable of crossing the plasma membrane to react with extracellular targets. Unlabeled S-(2-glutathionyl)acetylglutathione underwent carbonyl substitution with added [35S]GSH, suggesting that this conjugate may participate in modification of protein thiols. This conjugate also underwent hydrolysis with a half-life of approximately 3 hr. GSH:VDC me tabolite conjugates may serve as accessible models for labile covalent adducts formed between VDC metabolites and protein thiols.