The purpose of this study was to assess the phytochemical screening and antibacterial activity of Borassus flabellifer L. leaf extracts collected in the Thoothukudi district's Inland and Coastal areas against human pathogens. The antibacterial activity of B. flabellifer L. leaves was investigated using the agar well diffusion technique against a variety of test microorganisms, including bacteria and fungi. In the study on photochemistry, compounds such as flavonoids, alkaloids, phenol, protein, saponins, tannins, carbohydrates, and glycosides were investigated. Inland areas had the most (21 mm) and least (8 mm) zones of inhibition for E.coli and Staphylococcus albus, respectively. In coastal areas E.coli (21 mm) and Staphylococcus albus (9 mm) had the largest and smallest inhibition zones, respectively. B. flabellifer L. was found to have a higher rate of growth inhibition against some human pathogens. Ethanol and acetone extracts of B.flabellifer L. leaves have potent antibacterial properties and it could be used in the agro-food and pharmaceutical industries to create natural antioxidants.