Received for publication February 23, 2005.
Revised June 28, 2005.
Accepted for publication June 29, 2005.

Glutathione trapping to measure microsomal oxidation of furan to cis-2-butene-1,4-dial

Abstract

Furan is a liver carcinogen and toxicant. Furan is oxidized to the reactive dialdehyde, cis-2-butene-1,4-dial, by microsomal enzymes. This reactive metabolite readily reacts with glutathione nonenzymatically to form conjugates. An HPLC-electrochemical method for the detection of cis-2-butene-1,4-dial-GSH conjugates in microsomal preparations was developed in order to measure the extent of furan metabolism to cis-2-butene-1,4-dial in vitro. Previously unobserved mono-GSH reaction products of cis-2-butene-1,4-dial were detected in addition to the already characterized bis-GSH conjugates. Chemical characterization of these compounds indicated that the -amino group of glutathione had reacted with cis-2-butene-1,4-dial to form a thiol substituted pyrrole adduct. The analytical method was employed to estimate the extent of furan oxidation in rat liver microsomes from untreated or acetone-pretreated F344 rats as well as in human P450 2E1 supersomes. Our results confirm that cytochrome P450 2E1 can catalyze the oxidation of furan to cis-2-butene-1,4-dial. However, the data is also consistent with the involvement of other P450 enzymes in the oxidation of furan in untreated animals. This assay will be a valuable tool to explore tissue and species differences in rates of furan oxidation.

Key words: chemical carcinogenesis, metabolite identification, metabolite kinetics, microsomes, reactive metabolites