Highlights:Graphical/Tabular Abstract  Preparation of chitosangelatin based wound dressing film  Incoporation of St.John's Worth to wound dressing film  Antibacterial effect of St.John's Worth Figure A. Preparation scheme for St.john's wort supported wound dressing filmsPurpose: The present study aimed to investigate the potential use of St.John's Wort (Hypericum perforatum) incorporated films based on chitosan and gelatin as wound dressing. Theory and Methods:Wound infections due to various organisms often lead to challenging complications for public health during wound care management and have been conducted many studies to develop new wound dressing materials which plays critical role in the healing process . A new, innovative wound dressing material still needs to be developed to overcome the current limitations by accompanying the unique features, such as promoted moist environment, rapid wound healing, mechanical protection, non-cytotoxic to healthy tissue, antimicrobial/antifungal effects. In this work, St. John's Wort (Hypericum perforatum) supported wound dressing films based on chitosan and gelatin were prepared by using solvent casting technique. As wound dressing film structures were confirmed by FTIR and optical microscope its physical properties such as thickness, mass, water vapor permission rate (WVPR), water absorption capacity, water content and dressing pH were determined. Also, antibacterial activities were determined by the agar disc diffusion method for gram positive (S. Aureus, B. Subtilis) and gram negative (S. Typhi, E. Coli) microorganisms. Results:St.John's Wort (Hypericum perforatum) incorporated films that are investigated for their potential as wound dressing films have water absorption capacities of 107-646%, water contents of 6.4-23.1% and water vapor permeability rate (WVPR) of 356.25-984.38 g/m2day respectively. Also, pH values of films were range of 5.26-7.66 and these values are suitable for wound dressing application. It has been observed that all films exhibit antibacterial activity by forming inhibition diameters against four different microorganisms. Conclusion:Material selection is an important step to design an ideal wound dressing. St.John's Wort (Hypericum perforatum) incorporated films based on chitosan and gelatin were successfully prepared using a sequential solvent casting method. The results obtained in this study showed that St.John's Wort incorporated films could be used as potential candidates for wound healing application