DIASTEREOISOMERIC GLUCURONIDES OF OXAZEPAM - ISOLATION AND STEREOSELECTIVE ENZYMIC-HYDROLYSIS

Abstract

Oxazepam glucuronide isolated from swine urine by previously published methods was separated into its diastereoisomers by ion-exchange chromatography on a preparative scale. Quantitative high-performance liquid chromatography was used to monitor the separation. The 2 isomers were obtained in analytically pure form and then characterized by elemental analysis, oxazepam content, mass spectrometry, UV spectroscopy, optical rotation and optical rotatory dispersion-circular dichroism. The latter permitted the assignment of the dextrorotatory and the levorotatory isomers to the (S)- and (R)- configurations, respectively. Rates of enzymic hydrolysis depend on the configuration of the substrate as well as on the enzyme preparation used. Rate of cleavage was highest with the (S)-(+)-glucuronide and .beta.-glucuronidase from Escherichia coli. This enzyme possesses the highest degree of stereoselectivity; it hydrolyzes the (S)-(+)-isomer more than 400 times faster than the (R)-(-)-form. Bovine liver glucuronidase is less stereoselective, whereas glucuronidase preparations of molluscan [Helix pomatia] origin exhibit little stereoselectivity. The ready hydrolysis of one of the glucuronides by an enzyme from an intestinal microorganism may play a role in the enterohepatic circulation of oxazepam.