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UPLC-MS/MS Quantification of Fenspiride in Human Plasma
Yuriy V Pidpruzhnykov, Valerii E Sabko, Volodymyr V Iurchenko and Igor A Zupanets
A new simple and rapid ultra-performance liquid chromatography - tandem mass spectrometry (UPLC-MS/ MS) method has been developed and validated for the quantification of fenspiride in human plasma. Following a simple protein precipitation with acetonitrile, the analyte and internal standard (bupivacaine) were separated on an ACQUITY UPLC ® BEH С18 column (50 mm×2.1 mm, 1.7 μm) using a gradient mixture of acetonitrile and water (both containing 0.2% formic acid) as a mobile phase at flow rate 0.4 mL/min. The retention time of fenspiride and bupivacaine was 0.64 and 0.87 min, respectively. Detection was operated in positive mode by monitoring the transitions of m/z 261→105 for fenspiride and m/z 289→140 for bupivacaine. The lower limit of quantification was 2 ng/mL by using 200 μL aliquots of plasma. The range of quantification of fenspiride was 2-500 ng/mL. Within-run and between-run precision of analysis was <9.5%, accuracy ranged from 91.5% to 112.4%, average recovery from 99.3% to 101.9%. The analyte was stable in human plasma following three freeze/thaw cycles and for 5 months following storage at -70°C. The method was successfully applied to the bioequivalence study of generic and brand coated tablets of fenspiride