Determination of paeoniflorin, calycosin‐7‐O‐β‐d‐glucoside, ononin, calycosin and formononetin in rat plasma after oral administration of Buyang Huanwu decoction for their pharmacokinetic study by liquid chromatography–mass spectrometry

The purpose of this study was to simultaneously investigate the pharmacokinetics of five bioactive compounds in rat plasma after oral administration of Buyang Huanwu decoction (BYHWD) using high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS). The separations were performed on a Thermo Hypersil Gold C(18) analytical column (50 × 2.1 mm, 3 µm) with the column temperature kept at 30°C. The quantitative analysis was performed using a quadrupole mass spectrometer detector operated under selected ion monitoring mode. A linear gradient elution of A (0.1% formic acid solution) and B (100% acetonitrile) was used at a flow rate of 0.2 mL/min. The method was validated within the concentration ranges 1.8-450, 6.0-1500, 2.0-500, 1.2-300 and 1.2-150 ng/mL for paeoniflorin, calycosin-7-O-β-d-glucoside, ononin, calycosin and formononetin, respectively. The calibration curves were linear with correlation coefficients > 0.99. The lower limits of quantitations were < 6.0 ng/mL. The method was further applied to assess the pharmacokinetics of the five bioactive constituents of BYHWD in rat plasma.