Simultaneous Determination of Typhaneoside and Isorhamnetin-3-O-Neohesperidoside in Rats After Oral Administration of Pollen Typhae Extract by UPLC-MS/MS.

For the first time, a selective and rapid ultra-performance liquid chromatography method with tandem mass spectrometric (UPLC-MS/MS) detection for simultaneous determination of typhaneoside and isorhamnetin-3-O-neohesperidoside in rat plasma was developed and validated, which was applied to the pharmacokinetic study of Pollen Typhae extract. The separation was carried out on an ACQUITY UPLC(TM) BEH C18 column with gradient elution using mobile phase including acetonitrile and water (containing 0.1% formic acid). The flow rate was 0.4 mL/min. The detection was conducted by means of electrospray ionization mass spectrometry in negative ion mode with multiple reaction monitoring. The assays were linear over the concentration range of 0.5-100 ng/mL, and the lower limit of quantification was 0.5 ng/mL for typhaneoside and isorhamnetin-3-O-neohesperidoside. The method was validated in terms of intra- and interday precision (<9.37%), accuracy (within ±10.91%), linearity, specificity and stability, and has been successfully applied to a pharmacokinetic study of Pollen Typhae extract in rats after oral administration.