Assay of uridine diphosphate glucuronosyltransferase by high-pressure liquid chromatography

A new and simple method for the determination of rat liver microsomal uridine diphosphate glucuronosyltransferase activity toward 4-nitrophenol, phenolphthalein, and testosterone has been developed with the use of high-pressure liquid chromatography (hplc). After incubation of the substrate with microsomal fractions in the presence of uridine diphosphate glucuronic acid, the reaction is stopped by heating, mixed with methanol, and centrifuged to give the supernatant, which is analyzed directly by hplc. The unreacted substrate as well as its glucuronide can be quantitated from the peak-height method. When the pure glucuronide is not available, the standard curve for the glucuronide is obtained from the enzymatically produced glucuronide.