Determination of 3'-amino-3'-deoxythymidine, a cytotoxic metabolite of 3'-azido-3'-deoxythymidine, in human plasma by ion-pair high-performance liquid chromatography.

ABSTRACT

A sensitive high-performance liquid chromatographic assay has been developed to determine the levels of 3'-amino-3'-deoxythymidine (AMT), a cytotoxic metabolite of 3'-azido-3'-deoxy-thymidine (AZT, zidovudine), in human plasma. The sample pretreatment involved solid-phase extraction using cation-exchange extraction columns. Chromatography was carried out on a C8 column, using a mobile phase of methanol-0.01 M ammonium acetate (pH 5)-0.25 M sodium dioctylsulfosuccinate (60404, v/v/v) and ultraviolet detection at 265 nm. The method has been validated, and stability tests under various conditions have been performed. The lower limit of quantitation is 5 ng/ml (using 500-microliters human plasma samples). The bioanalytical assay has been used for the determination of AMT in patients with AIDS who used AZT.