Electrospray Mass Spectrometry with Consecutive Fragmentation Steps (ESI‐MSn) as a Tool for Rapid and Sensitive Analysis of Ginsenosides and Their Galactosyl Derivatives

The ginsenosides Rb₁ ( 3 ) and Rg₁ ( 4 ) isolated from Panax ginseng were enzymatically modified with galactosyltransferase to furnish new derivatives carrying galactose units in one or both sugar chains at position C(20) and/or C(3) or C(6) of the protopanaxadiol and protopanaxatriol aglycones 1 and 2 , respectively. To determine the linkage position(s) of the introduced galactose unit(s), an electrospray‐ionization MS analysis with consecutive fragmentation steps (ESI‐MSⁿ) was carried out using an ion‐trap mass spectrometer (Figs. 2 and 3). It was shown that both sugar moieties, located at different positions of the protopanaxadiol and protopanaxatriol aglycone, can be easily differentiated and analyzed in the subsequent fragmentation steps. Collision‐induced dissociation (CID) of the Na⁺‐ionized molecule (MS²) leads to cleavage of the most labile O?C(20) glycosidic bond, liberating the C(20) oligosaccharide fragment ion that can be analyzed in a subsequent fragmentation step (MS³). MS³ of the C(20) monodeglycosylated ginsenoside leads to cleavage of the second sugar moiety, allowing structure analysis of this fragment ion (MS⁴). By this method, the linkages of the monosaccharides and branching positions can be rapidly determined using only a few μl of a 10^?5 M sample solution.