A Simple and Rapid Method for the Determination of Pennogenin Diglycoside in Rat Plasma by HPLC-MS: Application to the Pharmacokinetics of the Extract in Gongxuening Capsules

A sensitive, selective, and reproducible reversed-phase high-performance liquid chromatography method coupled with electrospray ionization-ion trap mass spectrometry (LC-ESI-ITMS) was developed for the simultaneous quantification of pennogenin diglycoside (PD) in a small volume (100 µL) of rat plasma. PD was extracted from rat plasma samples using liquid-liquid extraction with methanol, digoxin as internal standard. The chromatographic separation was performed on a reversed-phase Agilent C18 (250 mm × 4.6 mm i.d., 5 µm particle size) analytical column using a mobile phase of 0.1% formic acid solution-acetonitrile (75:25, v/v) at a flow-rate of 1.0 mL/min. The mass spectrometer was operated in the negative ion mode at the deprotonated-molecular ions [M-H]- of parent drug. Calibration curve in spiked plasma was linear (correlation coefficient r = 0.999) from 0.5 to 50.0 mg/mL. For the different samples with concentration of 0.50, 5.00, and 50.0 µg/mL, recoveries of PD were (86.45 ± 4.39)%, (91.40 ± 4.40)%, and (93.79 ± 3.29)%, respectively (n = 3). The intra-day assay relative standard deviation at 0.50, 5.00, and 50.0 mg/mL of PD were 4.29%, 5.66%, and 4.03% (n = 3), respectively. The interday assay %RSD at the previously mentioned concentrations were 5.53%, 4.99%, and 4.31% (n = 3), respectively. The method was successfully applied to the pharmacokinetic study of PD in rats following either intravenous administration of PD solution or oral administration of the extract in Gongxuening capsules, a famous patent Chinese botanic drug.