Simultaneous determination of ten active constituents of Yankening Capsule in rat plasma by ultra high performance liquid chromatography with tandem mass spectrometry.

Abstract

An ultra high performance liquid chromatography with tandem mass spectrometry (U-HPLC-MS/MS) method was developed for simultaneous determination and pharmacokinetic study of ten active constituents, phellodendrine, coptisine, jatrorrhizine, berberine, palmatine, baicalin, wogonoside, baicalein, wogonin and emodin in rat plasma after oral administration of Yankening Capsule. After mixing with two internal standards tetrahydropalmatine and rutin, plasma samples were pretreated by protein precipitation with anhydrous ethanol-acetonitrile (9:1, v/v). The U-HPLC separation was carried on a ZORBAX RRHD Eclipse Plus C18 column (2.1 mm × 50 mm, 1.8 μm) with gradient elution using a mobile phase composed of methanol and water (containing 0.3% formic acid) at a flow rate of 0.3 mL min(-1). The detection was performed on a triple quadrupole tandem mass spectrometer by multiple reaction monitoring via electrospray ionization source with positive-negative ionization mode. The calibration curves of ten analytes showed good linearity (r>0.9979). The lower limits of quantification of phellodendrine, coptisine, jatrorrhizine, berberine, palmatine, baicalin, wogonoside, baicalein, wogonin and emodin were 0.50, 0.50, 0.30, 0.30, 0.30, 10, 3.0, 8.0, 1.0, 8.0 μg L(-1), respectively. The relative standard deviation of intra-day precision and inter-day precision were in the range from 1.13% to 5.96% and from 0.65% to 8.85%, respectively. The matrix effects of all analytes were found to be within the acceptable range with a range of 89.99-109.3%. The method is reliable and rapid and has been applied successfully to pharmacokinetic study of the ten active constituents in rat plasma after oral administration of Yankening Capsule.