On-line identification of phenanthroindolizidine alkaloids in a crude extract from Tylophora atrofolliculata by liquid chromatography combined with tandem mass spectrometry.

Electrospray ionization multi-stage tandem mass spectrometry (ESI-MS(n)) and liquid chromatography coupled with sequential mass spectrometry (LC/MS(n)) were applied to identify trace-level phenanthroindolizidine alkaloids in crude extracts from Tylophora atrofolliculata. Based on the relationship between the characteristic fragmentation reactions and the structural features of related compounds of known structure from this plant, the bioactive crude extract was analyzed in detail by positive and negative ion ESI-MS(n), LC/UV-MS and LC/MS(n) techniques. A total of nine constituents in the crude extract were identified rapidly, including several isomers; seven of these constituents are new and two are known compounds. The structures of four of these constituents were subsequently confirmed by nuclear magnetic resonance (NMR) and accurate mass measurements using high-resolution fast-atom bombardment mass spectrometry (FAB-HRMS).