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.