Simultaneous determination of clevidipine and its primary metabolite in dog plasma by liquid chromatography–tandem mass spectrometry: Application to pharmacokinetic study

Abstract A simple and rapid liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed and validated for the simultaneous determination of clevidipine and its primary metabolite H152/81 in dog plasma after protein precipitation with acetonitrile using felodipine as the internal standard (IS). Chromatographic separation was performed on a XB C18 column (2.1 mm × 50 mm, 3.5 μm) under isocratic conditions with the mobile phase consisting of acetonitrile and 20 mM ammonium acetate buffer (pH 7.0) (40:60, v/v) at the flow rate of 0.3 ml/min. The run time was 5.5 min. Mass spectrometric analysis was performed on a triple quadrupole mass spectrometer operated in the multiple reaction monitoring (MRM) mode with the transitions of m / z 473.0→338.2 for clevidipine, m / z 356.1→324.0 for H152/81 and m / z 383.9→338.2 for the IS. The method was fully validated in terms of selectivity, linearity, lower limit of quantification (LLOQ), accuracy, precision, stability, matrix effect and recovery over a concentration range of 0.15–200 ng/ml for clevidipine and 10–2000 ng/ml for H152/81, respectively. The analytical method was applied to support a pharmacokinetic study of simultaneous determination of clevidipine and H152/81 in ten healthy beagle dogs.