Cape gooseberry plant leaf extract mediated iron oxide nanoparticles (CG-IONPs) were prepared for the antibacterial study. The optimum precursor salt concentration, pH of the reaction mixture, the ratio between reducing agent and precursor salt, and time for the synthesis of iron nanoparticles were found to be 5 mM, 9.0, 3:7, and 0th, respectively. The synthesized iron nanoparticles were characterized by UV/Vis absorption spectroscopy, Photoluminescence Spectroscopy (PL), Transmission Electron Microscopy (TEM), X-ray photoelectron spectroscopy (XPS), Thermal Analysis (TG/DTA), and Fourier transform infrared spectroscopy (FTIR). A FTIR measurement was carried out to identify the possible molecules like carbonyl, CH, and OH bands which were responsible for the reduction and formation of the nanoparticles. From the X-ray diffraction (XRD) method, it was found that the average particle size of magnetite nanoparticles was found to be 24.18 nm. TEM analysis has been carried out to determine the size and shape of the synthesized CG-IONPs. The thermal stability of the synthesized CG-IONPs was tested by performing TG/DTA analysis, using a nitrogen atmosphere. XPS analysis was carried out to measure the elemental composition and oxidation states of the elements present in CG-IONPs. The PL analysis has been employed in the present study to explore the luminescent character of CG-IONPs. The synthesized CG-IONPs had antibacterial activity against pathogenic bacteria like Staphylococcus aureus, Streptococcus agalactiae, Escherichia coli, and Salmonella enterica by the well diffusion method. This biosynthesis method has been found to be cost-effective, good for the environment, and likely to be useful in many fields.