Simultaneous Determination of Formononetin, Calycosin and Rhamnocitrin from Astragalus Complanatus by UHPLC-MS-MS in Rat Plasma: Application to a Pharmacokinetic Study

This assay provided a novel and generally applicable method to simultaneously determine formononetin, calycosin and rhamnocitrin in rat plasma based on ultra-high performance liquid chromatography with tandem mass spectrometry. A single step of protein precipitation procedure with methanol was utilized, and luteolin was chosen as an internal standard. Chromatographic separation was achieved using a Waters Symmetry-C18 column, and the applied isocratic elution program allowed for the simultaneous determination of the three Flavones in a one-step chromatographic separation with a total run time of 3.5 min. The fully validated methodology for the analytes demonstrated high sensitivity, good accuracy and precision. The average recoveries of the analytes and internal standard were all above 91.0% and no obvious matrix effect was observed. This method was successfully applied to the preclinical pharmacokinetic studies of formononetin, calycosin and rhamnocitrin in rats. The results would be helpful to provide some references to clinical application of this herb.