Efficient separation of phosphopeptides employing a Ti/Nb-functionalized core-shell structure solid-phase extraction nanosphere

A strategy for effectively enriching global phosphopeptides was successfully developed by using ammonia methyl phosphate (APA) as a novel chelating ligand and Ti 4+ and Nb 5+ as double functional ions (referred to as Fe 3 O 4 @mSiO 2 @APA@Ti 4+ /Nb 5+ ). With the advantage of large specific surface area (151.1 m 2 /g), preeminent immobilized ability for metal ions (about 8% of total atoms), and unbiased enrichment towards phosphopeptides, Fe 3 O 4 @mSiO 2 @APA@Ti 4+ /Nb 5+ displays high selectivity (maximum mass ratio β-casein to BSA is 1:1500), low limit of detection (LOD, as low as 0.05 fmol), good relative standard deviation (RSD, lower than 7%), recovery rate of 87% ( 18 O isotope labeling method), outstanding phosphopeptide loading capacity (330 μg/mg), and at least five times re-use abilities. In the examination of the actual sample, 24 phosphopeptides were successfully detected in saliva and 4 phosphopeptides were also selectively extracted from human serum. All experiments have shown that Fe 3 O 4 @mSiO 2 @APA@Ti 4+ /Nb 5+ exhibits exciting potential in view of the challenge of low abundance of phosphopeptides. Graphical abstract