The purpose of the study was to develop new antimicrobial hydrogels from natural resources that may promote wound healing and prevent bacterial skin infection. The new hydrogels were synthesized by crosslinking chitosan with a vanillin isomer, 5-methoxysalicylaldehyde, by a friendly and easy method. To characterize these hydrogels, their structural and morphological properties were explored by FTIR, 1H NMR, SEM, POM, and TGA. In view of the targeted application, swelling behavior, biodegradability, antimicrobial activity and biocompatibility were investigated in vitro. Structural and morphological studies confirmed the formation of new hydrogels via the imination reaction concomitant with the supramolecular organization. The hydrogels were highly porous with the average pore diameter around 80 μm, and a swelling rate controlled by the crosslinking density and medium pH. The hydrogels showed a progressive weight loss in the presence of lysozyme up to 35%, during 21 days of testing. They proved non-cytotoxic effect on Normal Human Dermal Fibroblasts using MTS test and powerful antifungal activity against Candida Albicans, as determined by disk diffusion assay. All these properties indicate the new hydrogels as a promising option for the treatment of various skin lesions.