Chitosan as a nontoxic, biodegradable, and biocompatible biopolymer with film-forming properties can also be modified to improve its parameters. Modification of polymer films by the addition of nanoparticles is an increasingly common solution due to the higher efficiency of products at the nanoscale compared to the macroscale. In this work, thin chitosan films enriched with biogenic zinc oxide nanoparticles (ZnONPs) from Fusarium solani IOR 825 were obtained by the solvent evaporation method. The influence of nanoadditive on the physicochemical, mechanical, and antimicrobial properties of the polymeric matrix was evaluated. Two different concentrations of ZnONPs were added to the chitosan solution. Spectrometric measurements, mechanical tests, microscopic imaging, and microbiological tests were performed for nanoparticles-modified and control samples. Analysis revealed that ZnONPs influence the properties of chitosan films. FTIR spectroscopy showed changes that are the result of interactions between polymer matrix and the additive. Modified samples were characterized by increased values of Young’s modulus and tensile strength. SEM analysis combined with energy-dispersive X-ray spectrometry confirmed the presence of zinc in the modified films. The addition of nanoparticles slightly affected the surface morphology of the tested samples, and an increase in roughness was observed. Microbiological tests showed the biostatic activity of the films containing ZnONPs. The obtained films based on chitosan with the addition of ZnONPs can be considered easy-to-obtain biomaterials with potential use as cosmetic and biomedical products.