Simultaneous determination of thirteen flavonoids from Xiaobuxin-Tang extract using high-performance liquid chromatography coupled with electrospray ionization mass spectrometry.

A simple and reliable high performance liquid chromatography coupled with electrospray ionization mass spectrometry (HPLC-ESI-MS) analysis method was established to simultaneously determine thirteen flavonoids of Xiaobuxing-Tang in intestine perfusate, namely onpordin, 3'-O-methylorobol, glycitein, patuletin, genistein, luteolin, quercetin, nepitrin, quercimeritrin, daidzin, patulitrin, quercetagitrin and 3-glucosylisorhamnetin. Detection was performed on a quadrupole mass spectrometer equipped with an electrospray ionization (ESI) source operating in negative ionization mode. Negative ion ESI was used to form deprotonated molecules at m/z 315 for onpordin, m/z 299 for 3'-O-methylorobol, m/z 283 for glycitein, m/z 331 for patuletin, m/z 269 for genistein, m/z 285 for luteolin, m/z 301 for quercetin, m/z 477 for nepitrin, m/z 463 for quercimeritrin, m/z 461 for daidzin, m/z 493 for patulitrin, m/z 479 for quercetagitrin, m/z 477 for 3-glucosylisorhamnetin and m/z 609.2 for rutin. The linearity, sensitivity, selectivity, repeatability, accuracy, precision, recovery and matrix effect of the assay were evaluated. The proposed method was successfully applied to simultaneous determination of these thirteen flavonoids, and using this method, the intestinal absorption profiles of thirteen flavonoids were preliminarily predicted.