DETERMINATION OF PRANLUKAST AND ITS METABOLITES IN HUMAN PLASMA BY LCMS/MS WITH PROSPEKT(TM) ONLINE SOLID-PHASE EXTRACTION/

A highly sensitive and selective liquid chromatography/ionspray tandem mass spectrometry (LC/MS/MS) method was developed and validated for t he determination of Pranlukast and its oxidative metabolites (SB 24010 3, SE 241484 and SE 218663) in human plasma in order to support pharma cokinetic studies. The method employed direct injection of human plasm a into an on-line solid phase extraction (SPE) PROSPEKT(TM) instrument for isolation of the analytes followed by column switching to the LC/ MS/MS. The use of on-line SPE resulted in reduced sample preparation t ime and cleaner extracts, therefore minimizing ion suppression and HPL C back-pressures issues. The use of a 20 mM ammonium acetate-methanol system and a step gradient yielded intense ion species, excellent sepa ration between the polar metabolites and the parent drug and sufficien t selectivity for baseline resolution of the two positional isomers, S E 240103 and SE 218663. Pranlukast, its metabolites and the internal s tandard (SK&F 108566) were quantified using a turbo-ionspray interface by negative ion selected reaction monitoring (SRM). The lower limit o f quantification (LLQ) for the assay was 10.0 ng ml(-1) for Pranlukast and 1.00 ng ml(-1) for its metabolites based on a 100 mu l plasma ali quot. The calibration curves were linear for analyte concentrations ra nging from 10.0 to 2000 ng ml(-1) for Pranlukast and 1.00 to 200 ng ml (-1) for the metabolites. The calculated intra- and inter-assay precis ion from quality control (QC) samples resulted in mean variability val ues of less than 12% for all analytes Pranlukast and its metabolites w ere shown to be stable under routine analysis conditions for clinical trial samples. The method provides automated sample analysis in a tota l cycle time of 5 min with improved robustness, sensitivity, selectivi ty, accuracy and reproducibility compared to the existing methodology. (C) 1998 John Wiley & Sons, Ltd.