Negative ion mass spectrometry of sialylated carbohydrates: Discriminationof N-acetylneuraminic acid linkages by MALDI-TOF and ESI-TOF mass spectrometry

Negative ion MALDI and electrospray fragmentation spectra were recorded fro m 12 sialylated carbohydrates ranging from trisaccharides to biantennary N- linked glycans, D-Arabinosazone was found to be the most satisfactory MALDI matrix for these compounds. Fragmentation mechanisms were investigated wit h the aid of several synthesized analogues of the sugars labeled with C-13 and H-2. The substitution position of the sialic acid (alpha 2-->3 or alpha 2-->6) was found to have a dramatic effect on the overall fragmentation pa ttern of these compounds, and several features of the spectra were identifi ed that allowed the substitution pattern to be determined. In particular, t he appearance of an ion at m/z 306 appeared to be diagnostic of the presenc e of an alpha 2-->6-linked sialic acid. Selection and further fragmentation of the in-source (cone-voltage) fragment ion corresponding to the trisacch aride Neu5A alpha t2-->3(or 6)Gal beta 1-->4GlcNAc from larger, N-linked gl ycans, ionized by electrospray, gave fragmentation patterns identical to th ose of the reference trisaccharides, thus providing a method for confirming the sialic acid linkage.