Rapid determination of ruscogenin in rat plasma with application to pharmacokinetic study

A sensitive and rapid ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method was developed to determine ruscogenin in rat plasma using midazolam as the internal standard (IS). Sample preparation was accomplished through a liquid-liquid extraction procedure with ethyl acetate to 0.2mL plasma sample. The analyte and IS were separated on an Acquity UPLC BEH C18 column (2.1mm×50mm, 1.7μm) with the mobile phase of acetonitrile and 1% formic acid in water with gradient elution at a flow rate of 0.40mL/min. Ruscogenin and IS were eluted at 1.74 and 1.11min, respectively. The detection was performed on a triple quadrupole tandem mass spectrometer equipped with positive-ion electrospray ionization (ESI) by multiple reactions monitoring (MRM) of the transitions at m/z 431.2→287.0 for ruscogenin and m/z 326.2→291.1 for IS. The linearity of this method was found to be within the concentration range of 2-1000ng/mL with a lower limit of quantification of 2ng/mL. Only 2.0min was needed for an analytical run. The matrix effect was 92.4-107.3% for ruscogenin. The intra- and inter-day precision (RSD%) were less than 11.2% and accuracy (RE%) was within ±9.8%. The recovery ranged from 75.4% to 86.3%. Ruscogenin was sufficiently stable under all relevant analytical conditions. The method was also successfully applied to the pharmacokinetic study of ruscogenin in rats.