Preparation of cotton wool modified with boric acid functionalized titania for selective enrichment of glycopeptides.

An effective enrichment technique is a prerequisite to enable comprehensive analysis of protein glycosylation. In this work, a method was developed for modifying cotton wool with boric acid-functionized titania nanomaterial. The modified cotton wool was used for the preparation of solid phase extraction pipette tips for glycopeptide enrichment. Owing to the unique properties of this novel solid phase material, an enrichment protocol was proposed by using a mixed-mode of hydrophilic interaction liquid chromatography, titania affinity chromatography and boronate affinity chromatography. The performance of the tips was assessed by capturing glycopeptides from tryptic digests of horse radish peroxidase (HRP) and human immunoglobulin G. Modified cotton wool tips allowed the remove of most nonglycosylated peptides. 26 glycopeptides were detected from the HRP digest. The detection limit for glycopeptide determined by MALDI-TOF MS was below 5 fmol. The developed tips are particularly convenient tools for sample pretreatment and show a great potential in glycosylation analysis.