The in situ formation of hydrogels is utilized in numerous biomedical applications, notably for surgical adhesives and sealants. Unfortunately, the deleterious attributes of contemporary materials and methods, with serious concerns over their poor physical properties and in vivo cytotoxicity, requires the development of new approaches to the in situ generation of biocompatible hydrogels. Our approach utilizes oxygen as a ubiquitous, environmentally-borne initiation stimulus to generate free radicals capable of initiating polymerization reactions. This is accomplished by utilizing glucose oxidase, an oxidoreductase enzyme that reduces oxygen and concomitantly oxidizes glucose into hydrogen peroxide and gluconolactone, respectively. The hydrogen peroxide generated in situ is subsequently converted into polymerization-initiating radicals by Fe2+, via the Fenton reaction, or by the action of a second oxidoreductase, horseradish peroxidase. These radicals are utilized to initiate polymerization via the thiol–ene reaction, a radical-mediated, step-growth polymerization mechanism. Unlike other radical-mediated polymerization reactions, such as the chain-growth polymerization of (meth)acrylates, thiol–ene polymerizations exhibit extraordinary resistance to oxygen inhibition, a crucial trait for systems intended for utilization under aerobic conditions. The generation of hydrogels via aerobic, enzyme-mediated thiol–ene polymerization was demonstrated with a model monomer system as well as with functionalized chitosan, a polysaccharide well-known for its biocompatibility, mucoadhesion, anti-bacterial properties, and facile modification via its primary amines. Reaction kinetics experiments, performed with real-time FTIR spectroscopy and rheometry, reveal the limitations of the glucose/glucose oxidase/Fe2+ ternary initiating system; excessive Fe2+ concentrations lead to inhibitory and terminating reactions that consume radicals and limit attainable reaction rates. To circumvent these limitations, horseradish peroxidase, a second oxidoreductase, is utilized as an alternative means to generate radicals from hydrogen peroxide.

Fabrication of chitosan-based hydrogels via enzyme-mediated thiol-ene polymerization

251st National Meeting (2016)

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