Nanotechnology has grown in importance in modern material science research in recent decades. Green nanoparticles have sparked widespread interest due to their inherent characteristics of rapidity, environmental friendliness, and cost-effectiveness. In this study, zinc oxide nanoparticles were successfully synthesized for the first time using Citrullus colocynthis leaf extract. SEM with EDAX was used to investigate the morphology of zinc oxide nanoparticles. The crystalline nature of the nanoparticles was exposed through X-ray diffraction (XRD) experiments, which was responsible for reduction, stabilization, and capping agents. The disc diffusion method was used to test the antibacterial activity of zinc oxide against Pseudomonas aeruginosa, E. coli, Klebsiella pneumoniae, and Staphylococcus aureus. The ZnO nanoparticles had a strong antibacterial activity against the tested bacterial strains.