UPLC-MS/MS Determination of Chlorogenic Acid, Hyperoside and Astragalin in Plasma and its Pharmacokinetic Application in Liver Injury Rats

Background: Cuscutae Semen (CS) is reported to show a hepatoprotective effect. Chlorogenic acid, hyperoside and astragalin are three major biologically active components from CS. Objective: A sensitive method based on ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was developed and validated to quantify the three components in rat plasma and was successfully used to study pharmacokinetics in liver injury rats. Methods: Plasma samples were prepared with protein precipitation by acetonitrile. Chromatographic separation was achieved on ACQUITY-XBridge BEH C18 column with gradient elution using the mobile phase containing 0.05% formic acid in water (A) and acetonitrile (B). The three components were quantified using Electrospray Ionization (ESI) source in the negative multiple Reaction Monitoring (MRM) mode. Results: Calibration curves of each analyte showed good linearity with correlation coefficients over 0.99. Accuracies (RE%) and precisions (RSD%) were within 15%. The method was stable. Recovery of the target compounds in plasma samples ranged from 87.00% to 102.29%. No matrix effect was found to influence the quantitative method. Conclusion: The UPLC-MS/MS method met the acceptance criteria and was successfully applied to the simultaneous determination of chlorogenic acid, hyperoside and astragalin in rat plasma for the first time. It is suitable for pharmacokinetic application in liver injury rats. It provides the foundation for further development and utilization of the hepatoprotective effect of cuscutae semen.