Detection, characterization and identification of major constituents in Zhimu‐Baihe herb‐pair extract by fast high‐performance liquid chromatography and time‐of‐flight mass spectrometry through dynamic adjustment of fragmentor voltage

This work describes a novel methodology for unequivocal identification of chemical constituents in Zhimu-Baihe herb-pair (ZMBHHP). Compounds were removed from ZMBHHP by ultrasonic extraction with 70% ethanol, and then analyzed by fast high-performance liquid chromatography (HPLC) coupled with time-of-flight mass spectrometry (TOFMS). The accurate-mass capability of the TOF analyzer allowed reliable confirmation of the identity of the detected compounds, normally with mass errors below 3 ppm in routine analysis. This mass accuracy was sufficient to verify the elemental compositions of the chemical constituents in ZMBHHP. With dynamic adjustment of fragmentor voltage in TOFMS, an efficient ion transmission was achieved to obtain the best sensitivity and abundant fragmentation. By accurate mass measurements for each molecular ion and subsequent fragment ions, a reliable identification and differentiation of 24 saponins, 3 xanthones, 1 anthraquinone and 2 alkaloids was described here, including four groups of isomers. It is concluded that this fast and sensitive HPLC/ESI-TOFMS technique is powerful in qualitative analysis of complex herbal medicines in terms of sensitivity, selectivity, resolving power, time savings and lower solvent consumption. Furthermore, the data gathered in this study may be helpful for understanding the synergistic nature of this herb pair in traditional Chinese medicine (TCM) and further pharmacokinetic studies of ZMBHHP.