New hydrophilic material based on hydrogel polymer for the selective enrichment of intact glycopeptides from serum protein digests

The paper describes the preparation and characterization of a new HILIC material for the enrichment of N-linked glycopeptides. The material was prepared using 2-acrylamido-2-methyl-1-propanesulfonic acid as the monomer and ethylene glycol dimethacrylate as the cross-linker. The material was developed by a Box-Behnken experimental design, taking into consideration the amount of monomer-to-crosslinker ratio, the composition, and the amount of porogen mixture. By this approach, the property of the resulting polymer could be fine-tuned to modulate the hydrophilicity and porosity. As HILIC enrichment is mostly dependent on hydrophilic interactions, including H-bonding, the amount of swelling was expected to have an important function, therefore the optimization considered a monomer percent in the range of 20–80%, which implied very different water swelling capacities. After assessing the potential of this new polymer family on fetuin digests, the 17 materials resulting from the Box-Behnken experimental design were used for the enrichment of glycopeptides from serum protein digests. The materials displayed a superior performance over cotton HILIC enrichment, both in terms of the number of enriched N-linked glycopeptides and selectivity, providing up to 762 N-linked glycopeptides with 77% selectivity. The optimization indicated that a high amount of monomer significantly affected the number of enriched glycopeptides, which is also closely connected with the hydrogel nature of the resulting polymers. The results not only provide one additional HILIC material for the enrichment of glycopeptides but also pave the way for the use and development of hydrogel materials for the enrichment of N-linked glycopeptides.