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A rapid, sensitive and convenient method based on ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) was developed and validated for the simultaneous quantification of calycosin-7-O-β- d -glucoside (CCSG), ononin, calycosin, (6aR,11aR)-9,10-dimethoxypterocarpan-3-O-β- d -glucopyanoside (DPPG), and 7,2′-dihydroxy-3′,4′-dimethoxyisoflavan-7-O-β- d -glucopyanoside (DIFG) in rat plasma after oral administration of the methanol extraction from  Radix Astragali . Theophylline played the role of internal standard (IS). Preparation of plasma samples by liquid-liquid extraction method with ethyl acetate after precipitation of protein with methanol. The analytes were detected with a triple quadrupole tandem mass spectrometery (MS) in multiple reaction monitoring (MRM) mode and a positive ion electrospray ionization (ESI). The method was validated with the concentration ranges of 1.96–62.69 ng/mL for CCSG, 1.70–54.5 ng/mL for ononin, 1.85–59.06 ng/mL for calycosin, 2.14–137.24 ng/mL for DPPG and1.96–125.25 ng/mL for DIFG, respectively. The method had the lower limit of quantification (LLOQ) with 0.49, 0.21, 0.92, 1.07, and 0.98 ng/mL for CCSG, ononin, calycosin, DPPG and DIFG respectively, and the precision less than 10%. The RSD of the accuracy was in the range of −4.35–8.91%. The results may be helpful to provide more accurate references to clinical application of this herb.

acta chromatographica

Simultaneous determination and pharmacokinetics studies of five isoflavones in rat plasma by UHPLC-MS/MS after oral administration of Radix Astragali extract