Quantitation of bivalirudin, a novel anticoagulant peptide, in human plasma by LC-MS/MS: Method development, validation andapplication to pharmacokinetics

A rapid and sensitive method based on liquid chromatography tandem mass spectrometry (LC-MS/MS) for the determination of a novel anticoagulant peptide bivalirudin in human plasma has been developed and validated. Plasma samples were precipitated protein with acetonitrile and re extracted with dichloromethane, after which the analyte and triptorelin as an internal standard (IS) were separated on a 300SB-C18 column (150 mm x 4.6 mm i.d., 5 mu m particle size) using 0.1% formic acid:methanol (45:55, v/v) as mobile phase. The triple-quadrupole mass spectrometer, equipped with electrospray ionization (ESI) interface, was operated in the positive ion mode, and the multiple reaction monitoring (MRM) transitions of bivalirudin and IS were at m/z 1091.0 -> 650.4 and m/z 656.5 -> 249.3, respectively. The lower limit of quantification (LLOQ) was 1 ng/mL for 100 mu L plasma sample and the assay was linear over the concentration range 1-1000 ng/mL. The accuracy was within a range from -0.4% to 0.5% in terms of relative error (RE) and the intra- and inter-day precisions in terms of relative standard deviation (RSD) were <= 2.92 and <= 3.36, respectively. The method was successfully applied to a pharmacokinetic study involving intravenous administration of bivalirudin (0.5 mg/kg) to Chinese volunteers. (C) 2013 Xi'an Jiaotong University. Production and hosting by Elsevier B.V. All rights reserved.