Magnetic microspheres modified with Ti(IV) and Nb(V) for enrichment of phosphopeptides.

Magnetic microspheres (FeO) were coated with polydopamine (PDA) and loaded with the metal ions Ti(IV) and Nb(V) to give a material of type FeO@PDA-Ti/Nb. It is shown to be useful for affinity chromatography and for enrichment of phosphopeptides from both standard protein solutions and real samples. For comparison, such microspheres loaded with single metal ions only (FeO@PDA-Ti and FeO@PDA-Nb) and their physical mixtures were also investigated under identical conditions. The binary metal ion-loaded magnetic microspheres display better enrichment efficiency than the single metal ion-loaded microspheres and their physical mixture. Both multiphosphopeptides and monophosphopeptides can be extracted. The FeO@PDA-Ti/Nb microspheres exhibit ultra-high sensitivity (the lowest detection amount being 2 fmol) and selectivity at a low mass ratio such as in case of β-casein/BSA (1:1000). Graphical abstract Magnetic microspheres (FeO) were coated with polydopamine (PDA) and loaded with the metal ions Ti(IV) and Nb(V) to give a material of type FeO@PDA-Ti/Nb. Results showed its great potential as an affinity probe in phosphoproteome research due to rapid magnetic separation of phosphopeptides, ultrahigh sensitivity and selectivity, and remarkable reusability.