Differentiation of Oligosaccharide Linkage Isomers by Electron Activated Dissociation

The electron activated dissociation (ExD) behavior of oligosaccharides was determined by the electron energies and the type of charge carriers. Depending on the electron energy, metal-adducted oligosaccharides can undergo several fragmentation processes, including electron capture dissociation (ECD) at low energies, hot-ECD at intermediate energies, and electronic excitation dissociation (EED) at high energies. In this study, these ExD techniques were applied to the differentiation of oligosaccharide linkage isomers. Three pairs of permethylated oligosaccharide linkage isomers were investigated, including milk oligosaccharide isomers LNT and LNnT, LNFP I and LNFP III, and blood group antigens Sialyl Lewis^a (SLe^a) and Sialyl Lewis^x (SLe^x). Although ECD produced extensive fragmentations, the cross-ring fragments generated were insufficient for determination of the linkage differences and were often accompanied by neutral losses. In the case of SLe^a and SLe^x, owing to the preferred binding of metal cation to the sialic acid residue, ECD fragmentation was dominated by the loss of the sialic acid, and the cross-ring cleavages at the reducing end were suppressed. By contrast, EED produced abundant cross-ring fragments for definitive differentiation of all three pairs of oligosaccharide linkage isomers

Because the metal cation is usually the preferred electron capture site of metal-adducted oligosaccharides, the sites of ECD fragmentation are often limited by the binding positions of the metal cation. Such limitation is not as significant in EED, as it is initiated by ionization of a ring or glycosidic oxygen. Thus, EED generally produces a broader range of fragment ions than ECD. EED also generates cross-ring fragments with fewer neutral losses and therefore with less ambiguity. These results suggest that EED is a valuable tool for oligosaccharide linkage determination This research is supported by NIH-NCRR Grant Nos. P41 RR010888 and S10 RR025082.