CHARACTERIZATION OF AMINO-ACID ON GLUTATHIONE ADDUCTS OF CIS-2-BUTENE-1,4-DIAL, A REACTIVE METABOLITE OF FURAN

Metabolic activation of the hepatocarcinogen furan yields metabolites that react covalently with proteins, cis-2-Butene-1,4-dial is a micros omal metabolite of furan, This reactive aldehyde is thought to be the toxic metabolite that is responsible for the carcinogenic activity of furan. In order to characterize the chemistry by which this unsaturate d dialdehyde could alkylate proteins, the products formed upon reactio n of cis-2-butene-1,4-dial with model nucleophiles in PH 7.4 buffer we re investigated, N-alpha-Acetyl-L-cysteine (AcCys) reacts with cis-2-b utene-1,4-dial to form N-substituted pyrrolin-2-one adducts. N-Acetyl- L-cysteine (AcCys) reacts rapidly with cis-2-butene-1,4-dial to form m ultiple uncharacterized products. The inclusion of AcLys in this react ion mixture yielded an N-substituted 3-(S-acetylcysteinyl)pyrrole addu ct which Links the two amino acid residues. Related compounds were iso lated when cis-2-butene-1,4-dial and glutathione (GSH) were combined. In this case, cis-2-butene-1,4-dial cross-linked two molecules of GSH resulting in either cyclic or acyclic adducts depending on the relativ e GSH concentration. Incubation of furan with rat liver microsomes in the presence of [glycine-2-H-3]GSH led to the formation of radioactive peaks that coeluted with synthetic standards for the bisgluthathione conjugates. These studies demonstrate that the reactive cis-2-butene-1 ,4-dial formed during the microsomal oxidation of furan reacts rapidly and completely with amino acid residues to farm pyrrole and pyrrolin- 2-one derivatives. Therefore, this metabolite is a likely candidate fo r the activated furan derivative that binds to proteins. The ease with which cis-2-butene-1,4-dial cross-links amino acids suggests that pyr role-thiol cross-links may be involved in the toxicity observed follow ing furan exposure.