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High-performance liquid chromatographic quantification of 4-methylumbelliferyl-beta-D-glucuronide as a probe for
B Sperker1, M Schick, H K Kroemer
1Dr. Margarete Fischer-Bosch Institute of Clinical Pharmacology, Stuttgart, Germany.
Abstract:
An internally standardized HPLC method to determine the concentration of 4-methylumbelliferone liberated from 4-methylumbelliferyl-beta-D-glucuronide by human beta-glucuronidase was developed. The assay allows the precise and rapid measurement of specific enzyme activity in human tissue homogenates. Without prior extraction the incubation mixture can be separated using a C8 column followed by fluorescence detection. The assay showed good accuracy and precision with a detection limit of 20 nM and a limit of quantification of 167 nM. The suitability of the method was shown in enzyme kinetic experiments with human liver homogenates.