Simultaneous determination of nine antiretroviral compounds in human plasma using liquid chromatography

A rapid, sensitive and specific high-performance liquid chromatographic (HPLC) method using UV detection was developed for the determination of nine antiretroviral compounds commonly found in plasma from patients receiving antiretroviral therapy. Analytes include indinavir, saquinavir, ritonavir, amprenavir, lopinavir, delavirdine, efavirenz, nelfinavir and its M8 metabolite. Analytes were isolated from plasma using tert.-butyl methyl ether and separation achieved via reversed-phase liquid chromatography on a C(8) column with a gradient mobile phase. Detection at 210 nm provided adequate sensitivity. Limit of quantification is 50 ng/ml and all analytes demonstrated linearity across 50-10000 ng/ml from a single 200-microliter plasma sample. Recovery from plasma was consistently high (>80%). This novel HPLC methodology allows us to simultaneously determine plasma concentrations of nine antiretrovirals, including lopinavir, in HIV-infected patients on a single HPLC system.