Characterisation of Homoflavonoids from Three Ophioglossum Species Using Liquid Chromatography with Diode Array Detection and Electrospray Ionisation Tandem Mass Spectrometry

Introduction Homoflavonoids, characterised by one more carbon atom directly added to C6–C3–C6 backbone of flavonoids, are rich in the species of genus Ophioglossum. Up to now we have little knowledge about their MS fragmentation patterns. It is therefore necessary to investigate their MS fragmentation pathways so as to distinguish them from other types of flavonoids. Objective To develop a rapid method for identifying homoflavonoids from Ophioglossum based on their characteristic MS fragmentation. Methods Mass spectrometry fragmentation pathways and qualitative analysis of homoflavonoids in three ferns of Ophioglosssum were investigated by using high-performance liquid chromatography coupled with diode-array detection and electrospray ionisation tandem mass spectrometry (HPLC–DAD–ESI/MSn). Results The analyses of the MSn spectra of the homoflavonoids allowed us to classify them into two types according to their fragmentation characteristics. The type I homoflavonoids, with an attached additional carbon atom to the C-3 position of the C-ring, presented the initial competing loss of H2O and CH2O from their aglycone ions, compared to the initial removal of H2O or CO in the case of the type II homoflavonoids, which bear the additional carbon atom at the C-2’ site of the B-ring and forming ring D. The above characteristic fragmentations of homoflavonoids were quite different from those of other flavonoids, and were successfully applied to identify homoflavonoids in the crude extracts of three Ophioglossum species. Conclusion The HPLC–DAD–ESI/MSn method obtained in the present study provided a powerful tool for identifying homoflavonoids from ferns in the genus Ophioglossum. Copyright © 2013 John Wiley & Sons, Ltd.