The aim of this research is to use red onion extract (Allium cepa) in the green synthesis of zinc oxide nanoparticles (ZnO NPs). With an average crystal size of 8.13 nm, X-ray diffraction (XRD) scanning was used to determine the hexagonal wurtzite structure of the ZnO NPs. Infrared spectra (FT-IR) provided evidence of capping and biogenesis stability of ZnO NPs. The absorption peak at 374 nm with an energy gap of 3.32 eV was determined by surface plasmon resonance via UV-visible analysis. Morphological analysis was performed, and the results demonstrated the formation of spherical ZnO NPs with sizes ranging from 2.83 to 15.35 nm. The great purity, intensity, and crystal width of zinc and oxygen was determined by EDS. When compared to a common antibiotic such as ciprofloxacin, which was used as a positive control in the inhibitory zone, ZnO NPs showed more antibacterial effect against E. coli than S. aureus bacteria. Onion extract and DMSO were added to the control wells, but these additions did not produce an inhibitory zone that could be seen.