The use of mild conditions to perform the entrapment of biomolecules in polymeric matrices is a crucial step in a broad range of applications as biosensors, biocarrier-mediated facilitated transport membranes, and drug-controlled release devices. In this study, we investigated the crosslinking of poly(vinyl alcohol) (PVA) by glutaraldehyde in the absence of an acid catalyst and organic solvents to improve the water resistance of the hydrophilic biocompatible polymer. Glutaraldehyde was chosen as the crosslinking agent because it favors the intermolecular reaction with PVA and is able to bind nonspecifically to proteins. The effects of the temperature and glutaraldehyde content on the thermal and structural properties of the PVA films were examined. Membranes prepared at 40C showed a maximum crosslinking density for low glutaraldehyde content namely, 0.04 wt % in the spreading solution. Higher amounts of the crosslinker led to the branching of PVA. The increase in membrane thermal properties and reduction in crystallinity were ascribed to the crosslinking treatment, which was confirmed by Fourier transform infrared analysis. The oxygen permeability of the films was reduced up to 2.7 times, which indicated that the crosslinking of the polymer was successfully accomplished. V

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