Characterization and formation of the glutathione conjugate of clofibric acid.

The incubation of 1-O-clofibryl glucuronide (1-O-CAG), a metabolite of clofibrate, with glutathione (GSH) resulted in the appearance of a new peak when analyzed by HPLC. The use of HPLC coupled to electrospray-MS permitted the identification of the peak as S-(p-chlorophenoxy-2-methylpropanoyl)glutathione (CA-SG), formed by nucleophilic displacement of the glucuronide moiety from 1-O-CAG. Conjugate formation was enhanced 8-fold by rat liver glutathione S-transferases (GSTs). GSH was unreactive with isomers of 1-O-CAG formed by acyl migration, indicating that 1-O-CAG itself was the preferred substrate. Rearrangement of 1-O-CAG to its isomers in vitro, was found to be decreased in the presence of GSH. In vivo studies indicated that, following an intravenous infusion of clofibric acid to rats (75 mg/kg), the concentration of CA-SG excreted in bile over 4 hr, was approximately 0.1% of the concurrent CAG concentrations. Although these results indicate a minor role for GST-catalyzed reactions in clofibrate metabolism in vivo, they do define 1-O-acyl-linked glucuronides as a new class of substrates for GSTs.