An analytical strategy for accurate, rapid and sensitive quantitative analysis of isoflavones in traditional Chinese medicines using ultra-high performance supercritical fluid chromatography: Take Radix Puerariae as an example

Isoflavones are phenolic phytoestrogens due to their structural similarity to estradiol, so they usually serve as active component for quality control of traditional Chinese medicines (TCMs) rich in isoflavones. However, TCMs contains various kinds of similar isoflavones, especially isomers, which to a significant extent hinders accurate analysis of isoflavones in TCMs. Here, we present a novel analytical strategy for quality control of TCMs rich in isoflavones using ultra-high performance supercritical fluid chromatography coupled to photodiode array detection (UHPSFC–PDA) and tandem mass spectrometry (UHPSFC–MS/MS). Both chromatography and mass spectrometry parameters were optimized in order to develop an accurate, rapid, sensitive method for quantification of isoflavones. The reproducibility of quantitative analysis of multi-components by single marker (QAMS) using UHPSFC–PDA was discussed in terms of mobile phase gradient, temperature and backpressure for the first time. An analytical method for the analysis of isoflavones using UHPSFC–MS/MS was developed for the first time, and the established method was successfully applied to quantify isoflavones in three species of Radix Puerariae. The study showed Radix Pueraria Peduncularis contained higher amounts of isoflavones than Radix Puerariae Thomsonii, and it is worth noting that Radix Pueraria Peduncularis was often overlooked by researchers. It took less than 8 min with the current method and the limit of detection was not more than 0.05 ng/ml, which was definitely sufficient for anlysis of various samples from TCMs without enrichment.