Simultaneous quantification of zidovudine and its glucuronide in serum by high-performance liquid chromatography

A sensitive high-performance liquid chromatography (HPLC) assay has been developed to simultaneously determine levels of the anti human immunodeficiency virus agent, zidovudine (AZT), and its major metabolite (the 5'-O-glucuronide) in serum. Samples were first mixed with an internal standard (a stereoisomer of AZT), then prepared for analysis using solid-phase extraction columns and chromatographed using a reversed-phase analytical column. Isocratic elution with a mobile phase of 15% acetonitrile, buffered to pH 2.70 with ammonium phosphate, gave good resolution of the three analytes and endogenous serum components. The HPLC analysis time required per sample was 34 min and analyte recoveries were reproducibly high (greater than 93%). Replicate analyses of prepared standards gave satisfactory precision and accuracy, with coefficients of variation less than 15% and deviations from expected concentrations less than 10%.