Simultaneous determination of Guanfu base G and its active metabolites by UPLC–MS/MS in rat plasma and its application to a pharmacokinetic study

Abstract

To establish a rapid and sensitive ultra performance liquid chromatography mass spectrometry (UPLC-MS/MS) method for the determination of concentration of guanfu base G (GFG) and its active metabolites in rat plasma. The GFG and its active metabolites and the internal standard (phenacetin) were separated on an Acquity UPLC(®) BEH C18 chromatography column (2.1mm×50mm I.D., 1.7μm) using gradient elution with a mobile phase of methanol and ultrapure water at a flow rate of 0.4mL/min. The detection was performed on a Xevo triple quadrupole tandem mass spectrometer by multiple reaction monitoring (MRM) mode to monitor the precursor-to-product ion transitions of m/z 472.26→m/z 310.03 for GFG and m/z 180.00→m/z 109.99 for phenacetin (IS) using a positive electrospray ionization interface. The lower limit of quantification (LLOQ) was 1ng/mL, the limit of detection (LOD) was 0.3ng/mL, and the linear calibration curve was obtained over the concentration range of 1-200ng/mL. The intra-day and inter-day assay variations were measured to be below 10.97%, and the accuracy values (relative error) ranged from 95.4% to 103.6%. After validation, this method was successfully applied to a pharmacokinetic study where rats were intravenous administration of 5mg/kg GFG. A simple, rapid, sensitive, and accurate method for the determination of the concentration of GFG and its metabolites in rat plasma was developed and validated.