Liquid chromatography/electrospray tandem mass spectrometry method for thequantitation of fosinoprilat in human serum using automated 96-well solid-phase extraction for sample preparation

A sensitive, specific, accurate and reproducible liquid chromatography/elec trospray tandem mass spectrometry method was developed and validated for th e quantitation of fosinoprilat in 0.2 mt of human serum. The method employe d acidification (with pH 4.0 sodium acetate buffer) of the serum samples to minimize the hydrolysis of the prodrug fosinopril to fosinoprilat prior to purification by automated 96-well solid-phase extraction. The required chr omatographic separation of fosinoprilat and fosinopril was achieved isocrat ically on a Luna C8 analytical column (2 x 50 mm, 3 mu m). The total run ti me was 2 min. The mobile phase contained methanol and water with 10 mM ammo nium acetate, Detection was by positive ion electrospray tandem mass spectr ometry, The standard curve, which ranged from 2.00 to 500 ng/mL, was fitted to a 1/x(2) weighted linear regression model. Fosinoprilat quality control (QC) samples used to determine the accuracy and precision of the method we re prepared in human serum at concentrations of 5,00, 200, 400 and 1000 ng/ mL, The assay accuracy was within 8% (dev), The intra- and inter-assay prec isions were within 6 and 3% (RSD), respectively. Fosinopril QC samples used to gauge the rate of hydrolysis of Fosinopril to fosinopritat during the a ssay procedure were prepared in human serum at 500 ng/mL, The hydrolysis of fosinopril to fosinoprilat was less than or equal to 1%. This degree of co nversion would cause little error in the analysis of post-dose serum sample s since such samples are known to contain low levels of the prodrug compare d with the drug. Copyright (C) 2000 John Wiley & Sons, Ltd.