Sensitive Analyses of Neutral N-Glycans using Anion-Doped Liquid Matrix G3CA by Negative-Ion Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry

Negative-ion fragmentation of N-glycans has been proven to be more informative than that of positive-ion. In particular, it defines structural features such as the specific composition of the two antennae and the location of fucose. However, negative-ion formation of neutral N-glycans by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) remains a challenging task, and the detection limit of N-glycans in negative-ion mode is merely at the subpicomole level. Thus, practical applications are limited. In this study, combinations of five liquid matrices and nine anions were used to ionize N-glycans as anionic adducts, and their performances for sensitive analyses were evaluated. The best results were obtained with anion doped liquid matrix G(3)CA, which consists of p-coumaric acid and 1,1,3,3-tetramethylguanidine; the detection limits of anion adducted N-glycans were 1 fmol/well for NO3-, and 100 amol/well for BF4-. Negative ion MS2 spectra of 1 fmol N-glycans were successfully acquired with a sufficient signal-to-noise ratio and were quite useful for MS based structural determination. The anion doped G(3)CA matrix opens the way for sensitive and rapid analysis of neutral N-glycans in negative-ion MALDI at a low femtomole level.