Phosphate-functionalized magnetic microspheres for immobilization of Zr4+ ions for selective enrichment of the phosphopeptides

In this work, we developed phosphate functionalized magnetic Fe3O4@C microspheres to immobilize Zr4+ ions for selective extraction and concentration of phosphopeptides for mass spectrometry analysis. Firstly, we synthesized Fe3O4@C magnetic microspheres as our previous work reported. Then, the microspheres were functionalized with phosphate groups through a simple hydrolysis reaction using 3-(trihydroxysilyl)propyl methylphosphate. And the Zr4+ ions were immobilized on phosphate-functionalized magnetic microspheres by using phosphate chelator. Finally, we successfully employed Zr4+-phosphate functionalized magnetic microspheres to selectively isolate the phosphopeptides from tryptic digests of standard protein and real samples including rat brain. All the experimental results demonstrate the enrichment efficiency and selectivity of the method we reported here.