Amine functionalization of collagen matrices with multifunctional polyethylene glycol systems

A method to functionalize collagen-based biomaterials with free amine groups was established in an attempt to improve their potential for tethering of bioactive molecules. Collagen sponges were incorporated with amine-terminated multifunctional polyethylene glycol (PEG) derivatives after N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide and N-hydroxysuccinimide (EDC/NHS) cross-linking. The extent of the incorporation of different amounts and different numbers of active moieties of amine-terminated PEG systems into the collagen scaffolds was evaluated using ninhydrin assay, Fourier transform infrared spectrophotometry (FTIR), collagenase degradation assay, denaturation temperature measurements, and in vitro cell studies. A 3% 8-arm amine-terminated PEG was found to be the minimum required effective concentration to functionalize EDC/NHS stabilized collagen scaffolds. EDC/NHS stabilized scaffolds treated with 3% 8-arm amine-terminated PEG exhibited significantly improved denaturation temperature and resistance to collagenase degradation over non-cross-linked scaffolds (p < 0.002). Biological evaluation using 3T3 cells demonstrated that the produced scaffolds facilitated maintenance of the cells' morphology, metabolic activity, and ability to proliferate in vitro. Overall, our results indicate that amine-terminated PEG systems can be used as means to enhance the functionality of collagenous structures.