Testing of collagenase covalently linked to chitosan nanofibers for biomedical applications

Abstract

Nanofibers biofunctionalized with enzymes are materials of interest in the biomedical field due to their potential application in wound healing. This work describes the production and characterisation of electrospun chitosan nanofibers biofunctionalized with a microbial collagenase. The morphology and microstructure of the chitosan nanofibers prepared with Nanospider technology were examined using a scanning electron microscope (SEM). Amino groups of a chitosan nanofiber were connected with the carboxylic acid groups of the collagenase using EDAC and sulfo-NHS methodology. Collagenase enzymatic activity was evaluated with a peptidic substrate (Pz-peptide, 4-phenylazobenzyloxycarbonyl-Pro-Leu-Gly-Pro-D-Arg) in order to estimate the ability of this biomaterial to be used as an enzymatic debriding wound dressing. The characterisation of collagenase chitosan nanofibers in terms of measuring apparent Michaelis-Menten constants (KM(app) and vmax) revealed a higher enzyme to substrate affinity than with the soluble form of the collagenase. The resulting novel collagenase chitosan nanofibers exhibited excellent operational stability and long shelf life for at least 6 weeks. Therefore, the novel collagenase chitosan nanofibers are expected to be a potential scaffold for wound healing applications, such as e.g. enzymatic debridement.