Characterization and identification of triterpenoid saponins in crude extracts from Clematis spp. by high-performance liquid chromatography/electrospray ionization with multi-stage tandem mass spectrometry

High-performance liquid chromatography coupled with electrospray ionization multi-stage tandem mass spectrometry (HPLC/ESI-MS(n)) was applied to characterize and identify triterpenoid saponins in crude extracts from nine species of Clematis L. After separation on a Zorbax SB-C(18) column, negative ESI-MS experiments were performed. The quasi-molecular ions [M-H]-, [M+Cl]- and [M+HCOO]- were observed in the full-scan MS spectra of all compounds. The MS(n) (n = 2-4) data of the [M-H]- ions provided structural information on the sugar sequence of the oligosaccharide chains and on the aglycone of the saponins. In addition, the fragmentation mechanisms could be deduced from the fragment ions. As a result, eight saponins were unambiguously identified in C. ganpiniana by comparison with reference compounds. In addition, a new compound was tentatively identified as 3-O-ribopyranosyl --> rhamnopyranosyl --> (glucopyranosyl) --> arabinopyranosylhederagenin 28-O-rhamnopyranosyl --> glucopyranosyl --> glucopyranosyl ester (peak 1), and another one was tentatively deduced to be 3-O-glucopyranosyl --> ribopyranosyl --> rhamnopyranosyl --> arabinopyranosylhederagenin 28-O-rhamnopyranosyl --> glucopyranosyl --> glucopyranosyl ester (peak 5) from the genus Clematis L. for the first time. By ESI-MS(n), non-isomeric saponins could be discriminated rapidly. It is of interest that cleavage preferentially occurrs at the ester bond at C-28 and the charge is easy to transfer onto the oligosaccharide chain when the ester bond of a monodesmosidic saponin like HNH cleaves.