Dual metal cations coated magnetic mesoporous silica probe for highly selective capture of endogenous phosphopeptides...

For the first time, dual metal ions (Ti⁴⁺-Zr⁴⁺) were successfully modified into the channel of magnetic mesoporous silica to obtain an affinity probe for highly selective capture of endogenous phosphopeptides in biological samples. The newly prepared Fe₃O₄@mSiO₂@Ti⁴⁺-Zr⁴⁺ composites possessed the advantages of ordered mesoporous channels, superparamagnetism, and enhanced affinity properties of dual metal ions of Ti⁴⁺ and Zr⁴⁺. The phosphopeptide enrichment efficiency of the Fe₃O₄@mSiO₂@Ti⁴⁺-Zr⁴⁺ composite was investigated, and the result indicated an ultrahigh size exclusive ability (weight ratio of β-casein tryptic digests, BSA, and α-casein protein reached up to 1:1000:1000). Compared to magnetic affinity probes with single metal ions (Fe₃O₄@mSiO₂@Ti⁴⁺, Fe₃O₄@mSiO₂@Zr⁴⁺), the composite possessed stronger specificity, higher sensitivity, and better efficiency; and more importantly, it showed much enhanced enrichment ability towards both mono- and multi-phosphorylated peptides. Additionally, by utilizing the Fe₃O₄@mSiO₂@Ti⁴⁺-Zr⁴⁺ affinity probe, a total number of 104 endogenous phosphopeptides including 95 mono-phosphopeptides and 9 multi-phosphopeptides were captured and identified from human saliva, indicating the great potential for the application of the novel probe for the peptidome analysis in the future. Graphic abstract.