A simple and sensitive LC-MS/MS approach for simultaneous quantification of six bioactive compounds in rats following oral administration of aqueous extract and ultrafine powder of Astragalus propinquus: Application to a comparative pharmacokinetic study

A rapid, simple and selective approach based on ultra-high performance liquid chromatography tandem with mass spectrometry was established and validated for simultaneous quantification of six compounds in rat plasma, including astragaloside I, astragaloside II, astragaloside IV, calycosin‑7‑O‑β‑d‑glucoside, ononin and formononetin. The approach was applied to a comparative pharmacokinetic investigation of routine aqueous extract and ultrafine powder of Astragalus propinquus. Six compounds were extracted from plasma by using one-step protein precipitation. Chromatographic separation was conducted with a Waters BEH C18 UHPLC column using mobile phases of acetonitrile/0.1% formic acid-water. Multiple reaction monitoring (MRM) was optimized for mass quantification in a triple quadrupole mass spectrometer. The approach elicited good linearity for the six compounds (r2 > 0.9991) in the concentration ranges. The lower limits of quantification (LLOQ) of astragaloside IV, astragaloside II, astragaloside I, calycosin‑7‑O‑β‑d‑glucoside, ononin and formononetin were determined as 6.0, 9.6, 11.8, 5.8, 6.9, and 9.4 ng/mL, respectively. Intra- and inter-day precision, accuracy, selectivity and stability of the method met the required limits. The extraction recoveries of the six compounds were from 91.1% to 107.5% and the matrix effects ranged from 92.1% to 106.7%. The normalized Cmax and AUC0-t values of calycosin‑7‑O‑β‑d‑glucoside, and formononetin of the ultrafine powder group was much higher than that of the routine aqueous extract. Administration of Astragalus propinquus in the form of ultrafine powder showed enhanced bioavailability than that of routine aqueous extract.