A robust glycan labeling strategy using a new cationic hydrazide tag for MALDI-MS-based rapid and sensitive glycomics analysis.

Chemical derivatization of glycans is a common strategy to increase the analytical performance of MALDI-MS-based glycan profiling techniques. Hydrazide, one of the most popular tags, offers important advantages including allowing purification-free procedures. Several hydrazides have thus been used for glycomics combined with an on-target strategy to further simplify the analytical procedures. Usually, gentle heating and mildly acidic conditions with somewhat long reaction times are needed for these hydrazide derivatizations to reach a high reaction efficiency, which makes the current hydrazide tags not yet perfectly conducive to high-throughput analysis. To further optimize these hydrazide tags for high-throughput analysis, based on the structure of a reported hydrazide and the theoretical calculations, a new cationic hydrazide tag, 4-(hydrazinecarbonyl)-N,N,N-trimethylbenzenaminium (HTMBA), was designed, synthesized and tested in this work. HTMBA could completely derivatize glycans at room temperature in several seconds under very mildly acidic conditions (<3% acetic acid). A 19-fold enhancement in the signal intensity was obtained without interference from alkali adduct ions in the MALDI-MS detection of HTMBA-labeled maltoheptaose. To broaden the applicability of HTMBA, an HTMBA on-target derivatization (HOD) strategy was developed and fully validated with maltoheptaose and RNase B, and the method showed a good repeatability and stability. Finally, the HOD strategy was successfully applied to serum samples, 44 glycans in human serum were detected, and the O-acetylation information of sialic acid in horse serum was preserved. These results showed that the HOD strategy was suitable for the MS-based rapid analysis of all glycoforms in complex biological samples.