High-performance liquid chromatographic determination of thiopurine metabolites of azathioprine in biological fluids.

A selective and sensitive reversed-phase liquid chromatographic method for the analysis of thiopurine bases, nucleosides and nucleotides in biological samples was developed. A simple and rapid sample treatment procedure using perchloric acid deproteinization with dithiothreitol for the analysis of thiopurine bases and nucleosides is presented. The addition of dithiothreitol during sample collection and treatment improves recoveries. This procedure also allows the determination of thiopurine nucleotides by hydrolysis to their free bases after heating of the perchloric acid extract. The method was applied to the analysis of thiopurine metabolites in plasma and erythrocytes from lung-transplant patients under azathioprine therapy.