Rapid determination of 3′, 4′-dimethoxy flavonol-3-β-d-glucopyranoside in rat plasma by LC-MS/MS method followed by protein precipitation

A reliable liquid chromatography/tandem mass (LC-MS/MS) method was developed for quantitation of 3′, 4′-dimethoxy flavonol-3-β-d-glucopyranoside (DF3G) in rat plasma using pantoprazole as the internal standard. An Agilent C18 column (100 mm × 3 mm, 3.5 μm) was performed for chromatographic separation with the mobile phase composed of methanol-water (containing formic acid) at a flow rate of 0.4 mL/min. A triple-quadrupole mass spectrometry equipped with an electrospray ionization (ESI) source in positive ion mode was applied to quantitative analysis by multiple reacting monitoring (MRM). The MRM precursor-product ion transition of DF3G and Pantoprazole (IS) were m/z 461.1 → 299.2 and 383.9 → 200.2, respectively. Calibration curves were recovered in a concentration range of 1-500 ng/mL for plasma with a limit of lower quantification (LLOQ) of 1 ng/mL. The intra-day and inter-day precision were not >10%. The accuracy of DF3G ranged from −10% to 0.53% in quality control (QC) samples at three concentrations. DF3G was not degraded during the analysis and the storage period. All the data were validated in accordance with the FDA bioanalytical method validation guideline. The LC-MS/MS method was successfully employed in the pharmacokinetic study of the DF3G after oral administration and intravenous injection in rats.