Liquid chromatographic-tandem mass spectrometric determination of amprenavir (agenerase) in serum/plasma of human immunodeficiency virus type-1 infected patients receiving combination antiretroviral therapy.

A selective assay method for quantitation of amprenavir (agenerase) in human immunodeficiency virus type-1 infected patient serum or plasma using liquid chromatography-tandem mass spectrometry (LC-MS-MS) is described. Amprenavir and an internal standard (reserpine) are extracted by liquid-liquid extraction and chromatographically separated by a reversed-phase C18-analytical column. The triple quadrupole LC-MS-MS system is operated in the positive-ion mode and multiple reaction monitoring is used for drug quantitation. The method has been validated over the range of 0.05-10.0 microg/ml. The RSDs for the intra-day and inter-day determinations ranged from 5.3 to 6.1% and from 4.7 to 6.2%, respectively. The average assay accuracy at two different concentrations ranged from 96.0 to 103.0% and the extraction recovery of amprenavir was 90.8%. The lower limit of quantitation was 0.05 microg/ml. Using a short microbore column, the analysis was completed in less than 5 min.