COMPARISON OF PROCEDURES FOR MEASURING THE QUATERNARY N-GLUCURONIDES OF AMITRIPTYLINE AND DIPHENHYDRAMINE IN HUMAN URINE WITH AND WITHOUT HYDROLYSIS

The activities of beta-glucuronidases from Helix pomatia, Escherichia coli and rat towards the N-glucuronides of amitriptyline and diphenhyd ramine were considerably lower than those towards standard substrates. The two N-glucuronides were analysed in urine samples by the followin g procedures: HPLC of the intact conjugate after solid-phase extractio n on a cation exchanger cartridge or after direct injection of urine; HPLC of the aglycone after hydrolysis with beta-glucuronidase from H. pomatia or E. coli or after alkaline hydrolysis. Solid-phase extractio n led to the highest recovery and precision, and sensitivity can be im proved by extracting a larger volume of urine. On application to sampl es from patients under treatment with amitriptyline, the results of al l procedures except alkaline hydrolysis were in good agreement. When d iphenhydramine N-glucuronide was analysed in urine samples of voluntee rs, solid-phase extraction, hydrolysis by E. coli glucuronidase and al kaline hydrolysis resulted in similar values.