Hydrophilic Nb5+-immobilized magnetic core–shell microsphere – A novel immobilized metal ion affinity chromatography material for highly selective enrichment of phosphopeptides

Rapid and selective enrichment of phosphopeptides from complex biological samples is essential and challenging in phosphorylated proteomics. In this work, for the first time, niobium ions were directly immobilized on the surface of polydopamine-coated magnetic microspheres through a facile and effective synthetic route. The Fe 3 O 4 @polydopamine-Nb 5+ (denoted as Fe 3 O 4 @PD-Nb 5+ ) microspheres possess merits of high hydrophilicity and good biological compatibility, and demonstrated low limit of detection (2   fmol). The selectivity was also basically satisfactory (β-casein:BSA   =   1:500) to capture phosphopeptides. They were also successfully applied for enrichment of phosphopeptides from real biological samples such as human serum and nonfat milk. Compared with Fe 3 O 4 @PD-Ti 4+ microspheres, the Fe 3 O 4 @PD-Nb 5+ microspheres exhibit superior selectivity to multi-phosphorylated peptides, and thus may be complementary to the conventional IMAC materials.