A B S T R A C TBackground and Objectives: Studies on packaged food by activated biopolymer starch films containing antimicrobial material is interesting field of food. Science This study investigates the effect of chitosan and rosemary essential oil on properties of starch films and peanut were packaged in them. Materials and Methods:In this study, composite films based on starch containing 0, 2, 4, 6 and 8% chitosan and values of 0, 0.5, 1, 1.5 and 2 % rosemary essential oil produced by casting method, peanut packaging in them and the growth of Aspergillus flavus and aflatoxin B 1 production were investigated up to 12 days; as well as, the optimal point and regression model with RSM, Minitab 17. Results:Results of films showed chitosan and rosemary had decreasing effect on water vapor permeability, solubility and moisture absorption and increasing effect on tensile strength. The optimal point for physical properties of film was proposed 2% rosemary and 7.27% chitosan. In addition, the film reduced number of Aspergillus flavus about 4700 cfu/g, both chitosan and rosemary had a significant effect in this respect, although rosemary effect was more. The decreasing effect in the production of aflatoxin was found. The microbial optimal condition for film with 2% rosemary and 4% chitosan was created after 12 days, which showed the antimicrobial effect of film over time.Conclusions: It seems, rosemary has a major variable to improve physical and microbial properties of the films, while, chitosan has significant effect on physical properties.