Abstract: In this work, the thiolated chitosan was prepared by modification of chitosan. The composite film was prepared with thiolated chitosan as the film-forming matrix, adding gellan gum, glycerin, calcium chloride and natamycin as modifier. Based on the tensile strength, elongation at break, water vapor transmittance and transmittance of the film, the optimum addition amount of these four substances were determined by single factor experiments. By the Plackett-Burman experiment and the steepest climb experiment, the factors that had significant influence on the tensile strength of the film and the optimal experiment range were determined. Finally, the response surface experiment was carried out, and the tensile strength was taken as the evaluation index. The secondary response prediction model was obtained, and the optimal ratio of composite film was optimized. The results showed that the tensile strength of the single chitosan film was 0.928 MPa, but the elongation at break was only 5.91%. The tensile strength of the single thiolated chitosan film was only 0.350 MPa, and the elongation at break was 14.47%. When the basic film solution contained 0.20g of thiolated chitosan, the modified conditions were 0.18 g gellan gum, 1.00 g glycerol, 0.17 g calcium chloride and 0.01 g natamycin, the tensile strength of the composite film reached the maximum of 4.986±0.087 MPa. Under these, the tensile strength at break of the modified chitosan composite film was obviously (P<0.05) improved. The results of this study would provide theoretical support for the preparation of modified chitosan composite film, the improvement of composite properties and its application in the field of food preservation.