Application of mixed polymer brushes based on poly(2-methyl-2-oxazoline) and poly(acrylic acid) to on-line preconcentration of lysozyme by capillary electrophoresis

Abstract

A capillary coated with mixed polymer brushes that shows switchability toward lysozyme adsorption was developed. This capillary was applied for the on-line preconcentration of lysozyme by capillary electrophoresis (CE) in order to enhance the detection sensitivity. First, poly(2-methyl-2-oxazoline) (PMOXA) and poly(acrylic acid) (PAA) were synthesized by cationic ring-opening polymerization and reversible addition-fragmentation chain transfer (RAFT) polymerization, respectively. Then, glycidyl methacrylate (GMA) and PMOXA were used to prepare poly(2-methyl-2-oxazoline)-random-glycidyl methacrylate (PMOXA-r-GMA) via radical copolymerization, and poly(acrylic acid)-block-poly(glycidyl methacrylate) (PAA-b-PGMA) was obtained by the RAFT polymerization of GMA and PAA. A mixed solution of PMOXA-r-GMA and PAA-b-PGMA at a certain mass ratio was then injected into the capillary. Subsequent annealing provided capillary materials coated with mixed polymer brushes based on PMOXA and PAA. X-ray photoelectron spectroscopy (XPS) analysis was performed to determine the surface composition of the capillary raw materials. When the mass concentration of the mixed solution was 20 g/L and the mass ratio of PMOXA-r-GMA and PAA-b-PGMA was 1:1, the carboxyl content in the coating increased with increasing chain length of PAA. Fluorescein isothiocyanate-labeled lysozyme (FITC-lysozyme) adsorption assay demonstrated that the coated capillary had switchable properties for lysozyme adsorption upon pH and ionic strength (I) trigger. At pH 7 (I=10-5mol/L), the capillary could adsorb a large amount of lysozyme, which could be released at pH 3 (I=10-1mol/L). Subsequently, this coated capillary was applied to the on-line preconcentration of lysozyme by CE. The sensitivity enhancement factor was 17.69 when the chain length of PAA was 2.2 times that of PMOXA. The limit of detection could reach 8.7×10-5g/L. On-line preconcentration of lysozyme was performed for five successive times on the same day and on five consecutive days. The intraday and interday relative standard deviations (RSDs) for the peak areas were 2.9% and 4.1%, respectively. The intraday and interday RSDs for the migration times were 0.9% and 2.1%, respectively. The developed method for the preparation of the coated capillary with good stability only needs one step in this work, and this research will supply a simple and effective way to analyze trace protein by CE.