Mono- and dimethylation of arsenic in rat liver cytosol in vitro

Abstract

Production of methylarsonate and dimethylarsinate from radiolabelled [73 As]arsenite and [73 As]arsenate was examined in an assay system that contained cytosol prepared from a 20% homogenate (w/v) of livers from 8- 10-week-old male Fischer 344 rats. After a 60-min incubation at 37 degrees C with added S-adenosylmethionine and glutathione, up to 50% of carrier-free [73As]arsenite and about 15% of carrier-free [73As]arsenate were methylated. Incubation of cytosol at 100% degrees C for 1 min before addition to the assay system completely abolished methylation of arsenite. Production of methylarsonate increased in proportion to the arsenite concentration in the assay system; however, 50 microM arsenite inhibited production of dimethylarsinate. Methylarsonate production from carrier-free [73-As]arsenite was not dependent on addition of exogenous S-adenosylmethionine to the assay system. Addition of 0.1 mM S-adenosylmethionine maximized dimethylarsinate production. Addition of 0.1 or 1.0 mM S-adenosylhomocysteine decreased methylation of arsenite, especially dimethylarsinate production. Omission of glutathione from the assay system nearly abolished the methylation of arsenite. Addition of exogenous glutathione to the assay system (up to 20 mM) decreased protein binding of arsenic and increased the production of methylarsonate and dimethylarsinate. The effects of sodium selenite, mercuric chloride, EDTA, p-anisic acid and 2,3-dichloro-alpha-methylbenzylamine on the methylation of arsenite were determined. Addition of 10 microM selenite to the assay system nearly abolished the formation of either methylated species. Addition of 1 or 10 microM mercuric chloride inhibited dimethylarsinate production in a concentration-dependent manner but had little effect on methylarsonate yield. Addition of 10 mM EDTA to the assay system inhibited formation of both methylated metabolites, suggesting that an endogenous divalent cation might be involved in enzymatic methylation of arsenic. Neither p-anisic acid, an inhibitor of cytosolic methyltransferases, nor 2,3-dichloro-alpha-methylbenzylamine, an inhibitor of microsomal methyltransferases, inhibited the conversion of inorganic arsenic to mono- or dimethylated metabolites.