In vitro glucuronidation of peroxisomal proliferators: 2-ethylhexanoic acid enantiomers and their structural analogs.

In order to investigate the glucuronidation of 2-ethylhexanoic acid (2-EHA), a metabolite of the plasticizer di-(2-ethylhexyl) adipate, by liver microsomes of several mammalian species including man, a gas chromatography method for the quantification of the corresponding glucuronides was developed. The variation coefficients for intra- and interassay repeatability were less than 3 and 7%, respectively. The rat liver UDP-glucuronosyl-transferase (UGT) presented similar Km and Vmax toward the two enantiomers. The glucuronidation of the racemate 2-EHA or its enantiomers was strongly increased up to six times by treatment of the rats with phenobarbital and, to a lesser extent, by 3-methylcholanthrene. In contrast, the treatment of the rats clofibrate did not modify the activity. The induction was not stereoselective. The Gunn rats, which present a genetic defect in the bilirubin UGT isoforms, were able to glucuronidate the drug as well as the congenic strain. Moreover, the UGT-2B1 isoform, stably expressed in V79 cells, glucuronidated 2-EHA in an appreciable amount. Interspecies comparison indicated that the most active glucuronidation of 2-EHA occurred in the dog and the rat. The lowest activities were observed in the man and the rabbit. In all species considered, except rabbit and guinea pig which glucuronidated the R isomer faster, the R and S enantiomers were glucuronidated to a similar extent. The glucuronidation activity toward compounds chemically related to 2-EHA increased as a function of molecular weight, but was not affected by the position of the methyl or the ethyl moiety on the hydrocarbon chain. A correlation between the glucuronidation rate of 2-EHA and analogs and the activity of PCoA oxidase was observed.

Characterization of the in vitro glucuronidation of flurbiprofen enantiomers.

To investigate the glucuronidation of the R- and S-enantiomers of the nonsteroidal antiinflammatory drug, flurbiprofen, by liver microsomes of several mammals, including humans, a new and reliable HPLC method for the separation and quantification of the corresponding diastereoisomeric glucuronides has been developed. Interspecies comparison revealed that the glucuronidation of flurbiprofen was highly efficient with liver microsomes of humans, monkeys, rats, and guinea pigs (in decreasing ranking order). Gunn rats, which present a genetic defect in the bilirubin UDP-glucuronosyltransferase (UGT) isoforms, were still able to glucuronidate the drug. The R-enantiomer was glucuronidated faster than the S-form by liver microsomes of rats and humans. Although the KM of glucuronidation of R- and S-enantiomers by rat liver UGT were in same order of magnitude (apparent KM 0.52 and 0.57 mM, respectively), the apparent Vmax's were significantly different (9.34 and 5.48 nmol/min.mg of protein). Regardless of the enantiomer considered, the glucuronidation of flurbiprofen was strongly increased up to 5-fold by treatment of rats with phenobarbital and, at a lower extent, by 3-methylcholanthrene. In contrast, the treatment of rats with ciprofibrate markedly decreased the activity. Glucuronidation of R-flurbiprofen was more enhanced by phenobarbital than that of the S-antipode. Each flurbiprofen enantiomer could weakly inhibit the glucuronidation of its antipode in a noncompetitive way. The apparent Ki was 0.51 mM with R-flurbiprofen as a substrate, and 0.37 mM with S-enantiomer. On the other hand, the rat liver UGT2B1 isoform, stably expressed in V79 cells, could glucuronidate flurbiprofen in an appreciable amount.(ABSTRACT TRUNCATED AT 250 WORDS)