Development and validation of a sensitive UHPLC-MS/MS method for quantitation of prucalopride in rat plasma and its application to pharmacokinetics study

•Developing and verifying of a high precision and accuracy UHPLC-MS/MS method for the determination of prucalopride in rat plasma.•The comparative pharmacokinetic study of prucalopride following oral administration of different dosages to female and male rats is for the first time reported.•This selective and sensitive method (LLOQ = 100 pg/mL) requires quite low plasma sample volume (100 μL).•Extraction performed with a simple and effective protein precipitation preparation procedure for prucalopride.

A rapid, sensitive, selective and accurate ultra high performance liquid chromatography with tandem mass spectrometry (UHPLC-MS/MS) method was developed and validated for the quantitation of prucalopride in rat plasma using carbamazepine as an internal standard (IS). Separation was achieved on a Waters ACQUITY UHPLC® HSS C18 column (2.1 mm × 50 mm, 1.8 μm) column with a gradient mobile phase consisting of acetonitrile-water (containing 0.1% formic acid) as mobile phase at a flow rate of 0.2 mL/min. Prucalopride and IS were monitored using positive electrospray triple quadrupole mass spectrometer (Waters Xevo TQD) via multiple reaction monitoring (MRM) mode. The monitored transitions were set at m/z 367.99 → 195.89 and m/z 236.97 → 194.04 for prucalopride and IS, respectively. The achieved lower limit of quantitation was 0.1 ng/mL. The validated method had an excellent linearity in the range of 0.1-100 ng/mL (r > 0.996). The intra- and inter-day precisions were both ≤ 7.8% for prucalopride and IS, and the average intra- and inter-day accuracies ranged from −3.0% to 8.5%. Extraction recoveries at three levels QC concentrations were in the range of 90.0-110.0% for prucalopride and 99.6% for IS. Matrix effects were found to be acceptable. The validated assay was successfully applied to a pharmacokinetic study of prucalopride following oral administration of 0.25, 0.5, 1.0 mg/kg to female and male rats respectively.