A Method for the Determination of UDP-Glucuronosyltransferase Activity toward Arylcarboxylic Acids

✍ Scribed by M. Pritchard; S. Fournelgigleux; G. Siest; J. Magdalou

• Publisher: Elsevier Science
• Year: 1993
• Tongue: English
• Weight: 969 KB
• Volume: 212
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1006/abio.1993.1358

✦ Synopsis

A fast and reliable method for the assay of UDP-glucuronosyltransferase (UGT) activity toward aglycones containing a carboxylic acid function is described. The procedure involves incubation with UDP-[U- (\left.{ }^{14} \mathrm{C}\right]-) glucuronic acid, the common substrate for the reaction, solid-phase separation of the radiolabeled acylglucuronides and unreacted cofactor, and quantification by liquid scintillation counting. The technique was validated for each of the seven substrates tested by reversedphase HPLC, and was then applied successfully to the determination of optimal conditions for the activation of the carboxylic acid-UGT, and the estimation of kinetic constants for the glucuronidation of clofibric acid, 2-naphthylacetic acid, naproxen, and 4,4,4-triphenylbutanoic acid in rat liver microsomes. From the results obtained, we believe that this is an assay which, with only minor modification, could be applied to a wide range of carboxylic acid substrates for which, until now, specific and sensitive assays have been largely unavailable. ({1993) Academic Press, Inc.