Highly selective SiO₂-NH₂@TiO₂ hollow microspheres for simultaneous enrichment of phosphopeptides and glycopeptides

Glycosylation and phosphorylation as the commonest and most important posttranslational modifications of proteins play critical roles in biological processes. Specific and sensitive strategies have been developed to identify the glycosylation and phosphorylation of proteins by mass spectrometry but most of these methods have mainly focused on enriching glycopeptides or phosphopeptides separately, and few of them can enrich both of them. In this work, highly selective SiO₂-NH₂@TiO₂ hollow microspheres were designed and synthesized for the simultaneous selective enrichment of both phosphopeptides and glycopeptides. Because of the bifunctionalized property of the titanium dioxide and the amino group, the hollow microspheres were successfully applied for the simultaneous enrichment of phosphopeptides and glycopeptides. We evaluated the enrichment selectivity of the SiO₂-NH₂@TiO₂ hollow microspheres by capturing phosphopeptides and glycopeptides from a peptide mixture of bovine β-casein, horseradish peroxidase, and bovine serum albumin in a molar ratio of 1:1:500 (1.7 × 10^-12 mol β-casein and horseradish peroxidase, 8.5 × 10^-10 mol bovine serum albumin in 100 μL). Graphical Abstract Workflow of the simultaneous enrichment strategy for phosphopeptides and glycopeptides by SiO₂-NH₂@TiO₂ hollow microspheres.