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Rapid profiling of cantharidin analogues in Mylabris phalerata Pallas by ultra-performance liquid chromatography-quadrupole time-of-flight-tandem mass spectrometry.

We evaluated the protective effect and toxicity of extracts from Mylabris phalerata Pallas by measuring the activated partial thromboplastin time (APTT), prothrombin time (PT), venous thrombosis and acute toxicity in rats. Results showed the petroleum ether (PE) and water fractions of M. phalerata inhibited thrombosis but hardly prolonged the APTT and PT in rats. The trichloromethane (TCM) fraction had obvious toxicity with a LD50 of 0.2 g/kg in vivo, and contained many cantharidin analogues (CAs) by ultra-performance liquid chromatography-quadrupole ion trap-tandem mass spectrometry (UPLC-QTRAP-MS/MS). CAs are the major potential bioactivity constituent in M. phalerata. An effective and reliable UPLC-QTRAP-MS/MS method was successfully developed to separate and identify CAs. The fragmentation patterns of five purified compounds were applied for elucidate the structure of their analogues. Thirty four CAs were characterized or tentatively identified, eight of which are proposed to be novel compounds (13-17, 20, 21, 23), and their fragmentation patterns were investigated for the first time. Most importantly, a rapid and reliable UPLC-MS method was developed to identify CAs of M. phalerata. As shown, this method contributed to the discovery of most of these unknown analogues or their metabolites in M. phalerata effectively and quickly, and did not rely on limited chemical structural diversity libraries.