Identification and Characterization of Steroidal Saponins from Paris Polyphylla var. Yunnanensis by UPLC/oaTOF MS~(E)

摘要

Rhizoma Paridis ("ChongLou" in Chinese) is an herb used in many famous Traditional Chinese Medicine (TCM) remedies such as Yunnan Baiyao and Gongxuening amongst others. It is included as Paris polyphylla Smith var. yunnanensis (Franch) Hand-Mazz. (PPY) in Chinese Pharmacopeia. ChongLou is used to treat fractures, parotitis, homeostasis, snake bite, and abscesses. Rhizoma paridis saponins (RPS) are reported to be the main component in ChongLou. Among them, major aglycones are diosgenin, pennnogenin etc. Ingredient identification from ChonLou is important in elucidating its working mechanism and enabling enhanced QC methods for evaluating domestic ChongLou. The main purpose of this study is to perform ingredient analysis of ChongLou by using the UPLC/oaTOF MS~(E) data acquisition strategy (Figure 1). This strategy takes the advantages of the UPLC for high resolution, high sensitivity and high speed separation, as well as the oaTOF exact mass measurement capability. The UPLC/MS~(E) experiment was performed such that the mass spectrometer acquired data in two separate functions simultaneously. The first scan function uses low collision energy so that precursor ion information can be obtained; the second scan function uses elevated collision energy so that fragmentation data can be acquired. Our work is mainly focused on saponin identifications. We have 21 saponin standards in-house. These 21 saponins were separated using the traditional large pore resin column (home made) and were positively identified and confirmed by NMR. In this work, by using the UPLC/oaTOF MS~(E) strategy, we first studied the LC/TOF MS behavior of these 21 known saponins, this include understand the fragmentation behaviors of these 21 saponins. Then, we perform UPLC/oaTOF MS~(E) analysis on ChongLou extract. We use MS full scan data to screen possible new saponins, then, we use the MSMS fragment information that was obtained from the same LC injection to perform structural analysis and to compare with what we have already learn from the 21 known saponins. In the end, new saponins were positively identified.