Simultaneous quantitative determination of the HIV protease inhibitors amprenavir, indinavir, nelfinavir, ritonavir and saquinavir in human plasma by ion-pair high-performance liquid chromatography with ultraviolet detection.

A reversed-phase high-performance liquid chromatographic assay for the simultaneous quantitative determination of five HIV protease inhibitors (i.e. amprenavir, indinavir, nelfinavir, ritonavir, and saquinavir) in human plasma is described. Sample pretreatment consisted of solid-phase extraction prior to ion-pair, reversed-phase high-performance liquid chromatography with ultraviolet detection at 210 nm (amprenavir, indinavir and nelfinavir) and 239 nm (saquinavir and ritonavir). For amprenavir, indinavir and saquinavir the method has been validated over the range of 25 ng/ml to 25 microg/ml using a 0.6 ml sample volume. For nelfinavir and ritonavir the method has been validated over the range of 50 ng/ml to 25 microg/ml. The method proved to be accurate, with an average accuracy at four concentrations ranging from 90.6 to 109.2%, and precise, with the within-day and between-day precision ranging from 1.8 to 6.7%, and 0.7 to 7.6%, respectively. The protease inhibitors which can be quantified by using this assay proved to be stable under various conditions. This assay can readily be used in a hospital laboratory for the routine monitoring of plasma concentrations of these protease inhibitors.