The composition of a sample solvent has a crucial impact on separations in hydrophilic interaction liquid chromatography (HILIC). In this short communication, we studied the effect of an organic modifier in the sample solvent on the solubility of different tryptic glycopeptides of hemopexin and haptoglobin proteins. The results showed that the solubility of glycopeptides in solvents with a high acetonitrile content depends on the type of attached N-glycan. We observed lower solubility in larger glycans attached to the same peptide backbone, and we demonstrated that glycopeptides containing sialic acids precipitate more readily than those without sialic acid. Therefore, the sample solvent composition in HILIC must be carefully optimized for accurate quantitative data collection and for adequate separation.

Department of Analytical Chemistry Faculty of Science Charles University Hlavova 8 128 43 Prague 2 Czech Republic

Department of Biochemistry Faculty of Science Charles University Hlavova 8 128 43 Prague 2 Czech Republic

Department of Oncology Lombardi Comprehensive Cancer Center PSB GF9 Georgetown University 3800 Reservoir Road NW Washington DC 20057 United States; Department of Biochemistry and Molecular and Cellular Biology Georgetown University 3800 Reservoir Road NW Washington DC 20057 United States; Clinical and Translational Glycoscience Research Center Georgetown University Washington DC 20057 United States

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