Determination of acacetin in rat plasma by UPLC-MS/MS and its application to a pharmacokinetic study.

A rapid, sensitive and selective ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) was developed and validated for the determination and pharmacokinetic investigation of acacetin in rat plasma. Sample preparation was accomplished through a simple one-step deproteinization procedure with 0.2 mL of acetonitrile to a 0.1 mL plasma sample. Plasma samples were separated by UPLC on an Acquity UPLC BEH C18 column using a mobile phase consisting of acetonitrile-0.1% formic acid in water with gradient elution. The total run time was 2.0 min and the elution of acacetin was at 0.83 min. The detection was performed on a triple quadrupole tandem mass spectrometer equipped with positive-ion electrospray ionization (ESI) by multiple reaction monitoring (MRM) of the transitions at m/z 285.3→242.2 for acacetin and m/z 237.2→194.3 for carbamazepine (internal standard). The calibration curve was linear over the range of 1-1600 ng/mL with a lower limit of quantitation (LLOQ) of 1.0 ng/mL. Mean recovery of acacetin in plasma was in the range of 78.4-85.2%. Intra-day and inter-day precision were both <10.5%. This method was successfully applied in pharmacokinetic study after intravenous administration of 5.0mg/kg acacetin in rats.