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A liquid chromatography-tandem mass spectrometry method was developed and validated for simultaneous determination of six bioactive constituents including vitexin, orientin, isoorientin, 2″-O-β- d -xylopyranosyl isoorientin, 2″-O-β- d -xylopyranosyl isovitexin, and 6-C- l -α-arabipyranosyl vitexin in rats’ various tissues using isoquercitrin as the internal standard. Biological samples were pretreated by protein precipitation with acetonitrile. Chromatographic separation was carried out on a C18 column with a gradient mobile phase consisting of acetonitrile and 0.1% aqueous formic acid. All analytes and internal standard were quantitated through electrospray ionization in negative ion selected reaction monitoring mode. The mass transitions were as follows: m/z 431 → 311 for vitexin, m/z 447 → 327 for orientin or isoorientin, m/z 579 → 459 for 2″-O-β- d -xylopyranosyl isoorientin, m/z 563 → 293 for 2″-O-β- d -xylopyranosyl isovitexin, m/z 563 → 353 for 6-C- l -α-arabipyranosyl vitexin, and m/z 463 → 300 for the internal standard, respectively. The lower limits of quantification for the six analytes in different tissue homogenates were 0.8–2.2 ng/ml. The validated assay was successfully applied to a tissue distribution study of the six components in rats after intravenous administration of total flavonoids of Scorzonera austriaca Willd; Asteraceae. The results of the tissue distribution study showed that the high concentrations of six components were mainly in the kidney.

revista brasileira de farmacognosia

Simultaneous determination of six bioactive components of total flavonoids of Scorzonera austriaca in rat tissues by LC-MS/MS: application to a tissue distribution study