Real-time monitoring of peptide grafting onto chitosan films using capillary electrophoresis

Chitosan being antimicrobial and biocompatible is attractive as a cell growth substrate. To improve cell attachment RGDS peptides (arginine-glycine-aspartic acid-serine) were covalently grafted to chitosan films, through the widely used coupling agents EDC-HCl (1ethyl-3-(3-dimethylaminopropyl)carbodiimide) and NHS (N-hydroxysuccinimide), via the carboxylic acid function of the RGDS molecule. The grating reaction was monitored, for the first time, in real time using free-solution capillary electrophoresis (CE). This enabled fast separation and determination of the peptide and all other reactants in one separation with no sample preparation. Covalent RGDS peptide grafting onto the chitosan film surface was demonstrated using solid-state NMR of swollen films. CE indicated that oligomers of RGDS, not simply RGDS, were grafted on the film, with a likely hyperbranched structure. To assess the functional properties of the grafted films cell growth was compared on control and peptide-grafted chitosan films. Light microscopy and polymerase chain reaction (PCR) analysis demonstrated greatly improved cell attachment to RGDS-grafted chitosan films.