LC–MS/MS determination of naringin, hesperidin and neohesperidin in rat serum after orally administrating the decoction of Bulpleurum falcatum L. and Fractus aurantii

To identify and quantify biologically active components in rat serum after orally administrating the decoction of Bulpleurum falcatum L. and Fractus aurantii, one of traditional Chinese medicines (TCMs), high-performance liquid chromatography (HPLC)-tandem mass spectrometry method was developed and validated. The HPLC separation was carried out on a Waters Nova Pak C(18) column using acetonitrile and water as mobile phase after the sample of rat serum was cleaned up with solid-phase extraction. Atmospheric pressure chemical ionization in the negative ion mode and selected reaction monitoring (SRM) method was developed to determine the active components. Three flavonoids of hesperidin, neohesperidin and naringin were identified in the serum by comparing their retention times and three independent SRM precursor/product ion transitions with those of corresponding reference standards. The concentrations of naringin, hesperidin and neohesperidin in rat serum determined by SRM measurement were 16.3, 11.9 and 14.3 ng/ml, respectively, after orally administrating the decoction of B. falcatum L. and F. aurantii. This method was validated in terms of recovery, linearity, accuracy and precision (intra- and interday variation). The recoveries from spiked control samples were 93.0, 89.3 and 91.2% for hesperidin, neohesperidin and naringin, respectively. Linearity in rat serum was observed over the range of 2.0-50.0 ng/ml. Percent bias (accuracy) and precision were well within the acceptable range and the relative standard deviation (R.S.D.) of the measured rat serum samples was less than 10% (n=5).