Development and validation of an UHPLC–MS/MS approach for simultaneous quantification of five bioactive saponins in rat plasma: Application to a comparative pharmacokinetic study of aqueous extracts of raw and salt-processed Achyranthes bidentata

Abstract

A simple, accurate and sensitive ultra high-performance liquid chromatography-tandem mass spectrometry approach was established for the simultaneous determination of β-ecdysterone, 25S-inokosterone, ginsenoside Ro, chikusetsusaponin IV and chikusetsusaponin IVa in rat plasma after oral administration of raw and salt-processed Achyranthes bidentata extract. The saponins were completely separated on a Waters BEH C18 UHPLC column by using acetonitrile/0.1% formic acid-water as mobile phases. The mass analysis was performed in a triple quadrupole mass spectrometer using multiple reaction monitoring (MRM) with negative scan mode. The sample preparations for protein removal were accomplished using a simple acetonitrile precipitation method. The calibration curves displayed good linearity (r2 > 0.9998) with the concentration ranges of 24.4–6100 ng mL−1, 25.6–6400 ng mL−1, 20.4–8500 ng mL−1, 21.6–5400 ng mL−1, 21.6–6100 ng mL−1 for the five saponins, respectively. The intra-day and inter-day precisions (RSD) of the five saponins were less than 3.95% and the bias of the accuracies ranged from −4.50% to 4.84%. The extraction recoveries of the five saponins ranged from 95.2% to 104.8% and the matrix effects were satisfactory. In comparison with the raw group, the parameters of Cmax and AUC0-t of β-ecdysterone, 25S-inokosterone, ginsenoside Ro, and chikusetsusaponin IVa elevated remarkably (p < 0.05) after oral delivery of the extract of salt-processed Achyranthes bidentata, which revealed that salt-processing could increase bioavailability of β-ecdysterone, 25S-inokosterone, ginsenoside Ro and chikusetsusaponin IVa.