This study aimed to determine the effects of the ozone treatment of film-forming solutions (FFSs) containing whey protein concentrate (WPC) and gelatine on biopolymer films’ physical, mechanical, and thermal properties. Film samples were produced from a FFS that was ozone-treated at 0 (control), 5, 10, and 30 min. Ozone treatment caused an increase in the pH values of the FFS, whereas the zeta potential remained negative. The films became lighter in colour, slightly greenish, and more opaque with longer ozonation times. The control sample showed the highest thermogravimetric weight loss (92.15%). The weight loss of the samples decreased with increasing ozone treatment time. The application of ozone treatment on the FFS enhanced the films’ mechanical properties. Increased ozone treatment time improved the film samples’ tensile strength, elongation at break, and toughness values. In conclusion, the results of this study demonstrate that the ozone treatment of FFS containing whey protein concentrate and gelatine can significantly enhance the physical, mechanical, and thermal properties of biopolymer films. These results highlight the potential of ozone treatment as a viable method for improving the performance and quality of biopolymer films used in food packaging, offering promising advantages for sustainable and environmentally friendly packaging solutions.