MONOMETHYLATION AND DIMETHYLATION OF ARSENIC IN RAT-LIVER CYTOSOL IN-VITRO

Production of methylarsonate and dimethylarsinate from radiolabelled [ As-73]arsenite and [As-73]arsenate was examined in an assay system tha t contained cytosol prepared from a 20% homogenate (w/v) of livers fro m 1-10-week-old male Fischer 344 rats. After a 60-min incubation at 37 degrees C with added S-adenosyhnethionine and glutathione, up to 50% of carrier-free [As-73]arsenite and about 15% of carrier-free [As-73]a rsenate were methylated. Incubation of cytosol at 100 degrees C for 1 min before addition to the assay system completely abolished methylati on of arsenite, Production of methylarsonate increased in proportion t o the arsenite concentration in the assay system; however, 50 mu M ars enite inhibited production of dimethylarsinate. Methylarsonate product ion from carrier-free [As-73]arsenite was not dependent on addition of exogenous S-adenosylmethionine to the assay system. Addition of 0.1 m M S-adenosylmethionine maximized dimethylarsinate production. Addition of 0.1 or 1.0 mM S-adenosylhomocysteine decreased methylation of arse nite, especially dimethylarsinate production. Omission of glutathione from the assay system nearly abolished the methylation of arsenite. Ad dition of exogenous glutathione to the assay system (up to 20 mM) decr eased protein binding of arsenic and increased the production of methy larsonate and dimethylarsinate. The effects of sodium selenite, mercur ic chloride, EDTA, p-anisic acid and 2,3-dichloro-alpha-methylbenzylam ine on the methylation of arsenite were determined. Addition of 10 mu M selenite to the assay system nearly abolished the formation of eithe r methylated species. Addition of 1 or 10 mu M mercuric chloride inhib ited dimethylarsinate production in a concentration-dependent manner b ut had little effect on methylarsonate yield. Addition of 10 mM EDTA t o the assay system inhibited formation of both methylated metabolites, suggesting that an endogenous divalent cation might be involved in en zymatic methylation of arsenic. Neither p-anisic acid, an inhibitor of cytosolic methyltransferases, nor 2,3-dichloro-alpha-methylbenzylamin e, an inhibitor of microsomal methyltransferases, inhibited the conver sion of inorganic arsenic to mono- or dimethylated metabolites.