Rapid identification of C21 steroidal saponins in Cynanchum versicolor Bunge by electrospray ionization multi-stage tandem mass spectrometry and liquid chromatography/tandem mass spectrometry

Electrospray ionization multi-stage tandem mass spectrometry (ESI-MSn) and liquid chromatography coupled with on-line electrospray ionization tandem mass spectrometry (LC/ESI-MSn) were performed to elucidate the clearage rule of nine investigated C-21 steroidal saponins and identify them in the saponin fraction of 90% ethanolic extracts from the root and rhizome of Cynanchum versicolor Bunge. The fragments Of C-21 steroidal saponins in positive and negative ESI-MSn were used to deduce their mass spectral fragmentation mechanisms, and their structures were further confirmed by ESI-MSn in positive mode. The MSn spectra of the [M+Na](+) ions for saponins provided a wealth of structural information on glycosidic bond cleavage, which allowed a straightforward interpretation of spectra, with respect to the identifications of features such as the sequences of sugars attached to saponins and sugar type. By using LC/ESI-MSn, nine C-21 steroidal saponins were detected in the saponin fraction of C. versicolor, and an isomer of atratoglaucoside A was elucidated simultaneously. All nine compounds showed an abundant ion for the loss of 46 Da (HCOOH) from [M+Na](+). The losses of monosaccharide sequences and aglycone as neutral fragmentation from [M+Na-HCOOH](+) were also acquired as the characteristic ions of these C-21 steroidal saponins. It provided important information on monosaccharide sequences and in particular on sugar types and could be used to identify and elucidate other C-21 steroidal saponins. These studies allowed us to rapidly identify C-21 steroidal saponins from Radix cynanchi atrati. It is indicated that the described method had wide applicability to rapidly screen and provide structural confirmation On C-21 steroidal saponins in crude materials. Copyright (c) 2007 John Wiley & Sons, Ltd.