Comprehensive two-dimensional Liquid Chromatography (LCxLC)

Traditional high-performance liquid chromatography (HPLC) uses a single column, which is not sufficient for analysis of complex samples. If the number of components exceeds 37% of the peak capacity, peak resolution is statistically reduced. And if several ingredients of a sample are eluted and ionized at the same time in the ion source, this can lead to ion suppression and/or formation of artefacts. Therefore, a high-performance chromatographic platform in front of the MS is always the gold standard. Comprehensive two-dimensional liquid chromatography (LCxLC) is characterized by greatly increased resolving power and selectivity as compared with one-dimensional separation. The general setup of an LCxLC system comprises of two separation dimensions, which are coupled online via an interface, as shown in Fig. 1. The interface is usually a ten or eight-port switching valve with two equivalent sampling loops or trapping columns, operating in alternating cycles. The eluents from first dimensional elution were collected in the loop and subsequently transferred into the second dimensional separation column.

The great advantage of increased peak capacity of LCxLC enables the characterization of very complex samples, in present case, Hedyotis diffusa (Rubiaceae). Therefore, an LCxLC-QTOF-MS system was used to analyse flavonoids glycosides (FGs) and iridoid glycosides (IGs) in aqueous extraction of H. diffusa, as shown in Fig. 2. Tentative identification of compounds was done by accurate mass interpretation and validation by UV spectrum. A clear classification of FGs (Green solid line), acylated FGs (White dot line) and iridoid glycosides (IGs) was shown in different regions of the LCxLC contour plot. Totally, five FGs, four acylated FGs and three IGs were tentative identified. In addition, several novel flavonoids were found, which demonstrate that LCxLC-QTOF-MS detection has also a great potential in herbal medicine analysis.

Work was done by Dr. Duxin Li