Pharmacokinetics and bioavailability of hapepunine in mice by ultra-performance liquid chromatography–tandem mass spectrometry

An ultra-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) method was established to determine the hapepunine in mouse blood, and the pharmacokinetics of hapepunine after intravenous (1.0 mg/kg) and intragastric (2.5, 5, and 10 mg/kg) administrations was studied. Delavinone was used as an internal standard. The UPLC ethylene bridged hybrid (BEH) C18 column was used for chromatographic separation. The mobile phase consisted of acetonitrile and 0.1% formic acid with a gradient elution flow rate of 0.4 mL/min. Multiple reaction monitoring (MRM) mode was used for quantitative analysis of hapepunine in electrospray ionization (ESI) positive interface. Proteins from mouse blood were removed by acetonitrile precipitation. The verification method was established in accordance with the US Food and Drug Administration (FDA) bioanalytical method validation guidelines. In the concentration range of 1–1000 ng/mL, the hapepunine in the mouse blood was linear ( r 2 > 0.995), and the lower limit of quantification was 1.0 ng/mL. In the mouse blood, the intra-day precision coefficient of variation (CV) was less than 12%, the inter-day precision CV was less than 14%. The accuracy ranged from 91.7% to 109.3%. The average recovery was higher than 76.7%, and the matrix effect was between 86.0% and 106.4%. The UPLC–MS/MS method was sensitive, rapid, and selective and was successfully applied to the pharmacokinetic study of hapepunine in mice. The absolute bioavailability of hapepunine was 22.0%.