The silver nanoparticles of 1-100 nm size have the potential to be used in many medical fields, such as medical devices, therapies and molecular diagnostics, due to their antimicrobial properties. The biosynthesis of nanoparticles with numerous important advantages over chemical and physical methods has attracted attention today. In this study, the biosynthesis of the silver nanoparticles was performed using the Hypericum perforatum leaf extract to enhance the antibacterial effect of the silver nanoparticles. The morphological characteristic of the obtained mixture was determined using Transmission Electron Microscopy (TEM). Also the formation of the mixture was identified using UV-Vis spectrophotometer. The antibacterial activity of the synthesized Ag NPs using H. perforatum leaf extract was carried out in Gram (+) bacteria (Staphylococcus aureus ATCC 29213, Staphylococcus aureus ATCC 95923, Staphylococcus epidermidis ATCC 12228) and Gram (-) bacteria (Klebsiella pneumonia ATCC 700603, Escherichia coli ATCC 25922, Proteus mirabilis ATCC 27853, Shigella, Acinetobacter baumannii ATCC BAA 747) by disc diffusion method on solid media. The biosynthesized silver nanoparticles using the H. perforatum leaf showed higher antibacterial activity than the antibiotics.