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A Ti4+-functionalized magnetic covalent organic framework material with flexible branched polymers (mCOF@ε-PL@THBA-Ti4+) built via an immobilized metal ion affinity chromatography (IMAC) enrichment strategy was proposed through post-synthesis modification. Hydrophilic ε-poly- L -lysine (ε-PL) rich in amino active groups was first introduced in the fabrication of the phosphopeptide enrichment material to increase the hydrophilicity while providing more functional modification pathways of the material. 2,3,4-Trihydroxy-benzaldehyde (THBA) provides abundant binding sites for the immobilization of numerous Ti4+, which is advantageous for the subsequent efficient phosphopeptide enrichment. The magnetic nanocomposite exhibited outstanding performance of phosphopeptide enrichment with good selectivity (1 : 5000), a low detection limit (2 fmol), and relatively high loading capacity (66.7 mg g−1). What's more, after treatment with mCOF@ε-PL@THBA-Ti4+, 16 endogenous phosphopeptides from fresh saliva of healthy people were recognized by MALDI-TOF MS, and 50 phosphopeptides belonging to 35 phosphoproteins from the serum of uremia patients were detected by nano-LC-MS/MS. Proteomics data analysis for the differential protein selection between uremia and normal controls was conducted using R software, and four down-regulated and three up-regulated proteins were obtained. The results suggested that the prepared material has potential applications in biomarker discovery. [ABSTRACT FROM AUTHOR]

Protein glycosylation is of great significance in various physiological processes. Nevertheless, it remains a huge challenge to identify glycopeptides in complex biosamples by the direct mass spectrometry analysis due to the low ion efficiency and low abundance of glycopeptides. In this study, a novel hydrogel (denoted as ZIF-8/SAP) with a stable three-dimensional (3D) network structure and excellent hydrophilicity was successfully fabricated to capture glycopeptides with high efficiency. Owing to the unique characteristics, ZIF-8/SAP exhibited great selectivity (1 : 1000), great sensitivity (1 fmol μL−1), large binding capacity (300 mg g−1) and satisfactory reusability (seven cycles). Inspired by the great enrichment performance of the as-prepared material toward glycopeptides, ZIF-8/SAP was further applied to capture glycopeptides from a real human serum sample. The experimental results demonstrated that 217 N-glycosylation sites were identified in 283 N-glycopeptides, corresponding to 95 glycoproteins identified from 10 μL human serum by the nano-LC-MS/MS analysis, revealing the great potential of the novel ZIF-8/SAP hydrogel for glycopeptide enrichment and glycoproteomic research. [ABSTRACT FROM AUTHOR]