Spectroscopic characterisation and identification of ecdysteroids using high-performance liquid chromatography combined with on-line UV-diode array, FT-infrared and H-1-nuclear magnetic resonance spectroscopy and time of flight mass spectrometry

A prototype multiply hyphenated reversed-phase HPLC system has been applied to the analysis of a mixture of pure ecdysteroids and an ecdysteroid-conta ining plant extract. Characterisation was achieved via a combination of dio de array UV, H-1 NMR, FT-IR spectroscopy and time of flight (TOF) mass spec trometry. This combination of spectrometers allowed the collection of UV, H -1 NMR, IR and mass spectra for a mixture of pure standards enabling almost complete structural characterisation to be performed. The technique was th en applied to a partially purified plant extract in which 20-hydroxyecdyson e and polypodine B were identified despite incomplete chromatographic resol ution and the presence of co-chromatographing interferents. The experimenta l difficulties in the use of such a systems for these analytes are describe d. (C) 2001 Elsevier Science B.V. All rights reserved.