Determination of cephalomannine in rat plasma by gradient elution UPLC–MS/MS method

• First report of the determination of cephalomannine in rat plasma using UPLC–MS/MS.
• Sample preparation with a simple one-step protein precipitation by perchloric acid–methanol (1:9, v/v).
• A relatively larger number of samples can be analyzed for 2.0 min per sample.

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 cephalomannine in rat plasma. Sample preparation was accomplished through a simple one-step deproteinization procedure with 0.2 mL of perchloric acid–methanol (1:9, v/v) 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 cephalomannine was at 1.60 min. The detection was performed on a triple quadrupole tandem mass spectrometer in the multiple reaction-monitoring (MRM) mode using the respective transitions m/z 832.8 → 264.1 for cephalomannine and m/z 812.6 → 286.0 for 10-DAT (internal standard), respectively. The calibration curve was linear over the range of 10–2000 ng/mL with a lower limit of quantitation (LLOQ) of 10 ng/mL. Mean recovery of cephalomannine in plasma was in the range of 80.9–85.3%. Intra-day and inter-day precision were both <11.2%. This method was successfully applied in pharmacokinetic study after intravenous administration of 5.0 mg/kg cephalomannine in rats.