Quantitative determination of rupestonic acid in rat plasma by high-performance liquid chromatography-tandem mass spectrometry and its application in a pharmacokinetic study.

A sensitive and specific high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (HPLC-ESI-MS/MS) method was developed and validated for determination of rupestonic acid in rat plasma. Protein precipitation method was used to extract rupestonic acid and the internal standard (IS) warfarin sodium from rats plasma. The chromatographic separation was performed on an Agela Venusil XBP Phenyl column with an isocratic mobile phase consisting of methanol-0.1% formic acid in water (40:60, v/v), pumped at 0.4 mL/min. Rupestonic acid and the internal standard (IS) warfarin sodium were detected at m/z 247.2 → 203.1 and 307.1 → 161.3 in positive ion and multiple reaction monitoring mode respectively. The standard curves were linear over the concentration range of 2.5-5000 ng/mL (r(2) > 0.99). The within-day and between-day precision values for rupestonic acid at four concentrations were 4.7-5.7 and 4.4-8.7%, respectively. The method described herein was fully validated and successfully applied to the pharmacokinetic study after an intravenous administration of rupestonic acid in rats.