Simultaneous determination of two flavonoids and six pentacyclic triterpenes by UPLC-MS/MS: application to a comparative pharmacokinetic study in normal and arthritic rats after oral administration of Huo Luo Xiao Ling Dan or its single herb extract

Abstract

A sensitive and reliable UPLC-MS/MS method was developed and validated for the simultaneous determination of two flavonoids (liquiritigenin and isoliquiritigenin) and six pentacyclic triterpenes (glycyrrhetinic acid, 11-keto-β-boswellic acid, α-boswellic acid, β-boswellic acid, acetyl-α-boswellic acid and acetyl-β-boswellic acid) in rat plasma after oral administration of Huo Luo Xiao Ling Dan or its single herb. The plasma samples were extracted by liquid–liquid extraction with emodin as the internal standard. Good chromatographic separation was achieved on a Shim-pack XR-ODS II column with a mobile phase of methanol and 5 mM ammonium acetate water. Detection was performed on a UPLC-MS/MS system with an electrospray ionization source in negative ionization and multiple reaction monitoring mode. The lower limits of quantification were 0.05–4 ng mL⁻¹ for all the analytes. The intra- and inter-day precision and accuracy of the analytes were well within acceptance criteria (15%), and the mean extraction recoveries of the analytes and emodin from rat plasma were all more than 70.6%. The validated method was successfully applied to a comparative pharmacokinetic study of eight active ingredients in normal and arthritic rats. The results showed that there were remarkable differences in the pharmacokinetic properties of the analytes between herbal formula and single herb groups, in normal and arthritic groups.