Highly Efficient Enrichment Method for Glycopeptide Analyses: Using Specific and Nonspecific Nanoparticles Synergistically

Abstract

We invented a new method for highly efficient and specific enrichment of glycopeptides using two different nanomaterials synergistically. One is boronic-acid-functionalized Fe₃O₄ nanoparticles, enriching glycopeptides through formation of cyclic boronate esters between the boronic acid groups and the cis–diol groups on glycopeptides. The other nanomaterial is conventional poly­(methyl methacrylate) nanobeads, which have strong adsorption toward nonglycopeptides. By optimizing the proportion of these two materials, extremely high sensitivity and selectivity are achieved in analyzing the standard glycopeptides/nonglycopeptides mixture solutions. Since the washing step is not necessary for these conditions, the enrichment process is simplified and the recovery efficiency of target glycopeptides reaches 90%. Finally, this approach is successfully applied to analyze human serum with the sample volume as little as 1 μL, in which 147 different N-glycosylation peptides within 66 unique glycoproteins are identified. All these performances by the synergistic enrichment are much better than employing one specific enrichment agent alone