Introduction In gas chromatographic-mass spectroscopic analysis of buprenorphine metabolites, the urine specimen must be first hydrolyzed to release buprenorphine and norbuprenorphine from their glucuronide conjugates. For evaluation of existing hydrolysis methods and to find out the optimal hydrolysis conditions, buprenorphine-3-1~-D-glucuronide (B3G) and norbuprenorphine-3-II-D-glucuronide (NB3G) were synthesized. In a previous communication we reported on the optimum conditions for

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... of B3G. Because we have previously shown that no hydrolysis was achieved under basic conditions and that acid hydrolysis resulted in extensive degradation, only enzymatic hydrolysis was examined for NB3G. This study therefore reports on the optimum enzymatic conditions for the hydrolysis of NB3G. Urine fortified with synthetic NB3G was hydrolyzed with I~-glucuronidases from different source species, including Helix pomatia, Escherichia coli, and Patella vulgata. Glusulase, a preparation containing both the 13-glucuronidase (H. pomatia) and