In this study, green synthesis, characterization and antibacterial activity of zinc oxide nanoparticles was studied. The zinc oxide nanoparticles were successfully synthesized via a fast, convenient, cost-effective, and environmentally friendly method by biologically reducing Zinc Nitrate hexahydrate Zn (NO3)2.6H2O solution with Khat (Catha Edulis) leaf extract under optimum condition. The presence of active flavonoids, phenolic groups, alkaloids, steroids, and tannins, which were in the biomass of the leaf extract before and after reduction was identified using qualitative screening methods (observing the color changes) and FT-IR Spectroscopy. The formation of ZnO NPs was visually indicated by the color changes from colorless to light yellow color. Biosynthesized nanoparticles were also characterized by UV-Visible, FT-IR, and XRD spectroscopies. The reduction process was simple and convenient to handle and was monitored by UV-Visible spectroscopy that showed surface Plasmon resonance (SPR) of the Zinc Oxide nanoparticles at 322 nm. This result clearly revealed the formation of Zinc Oxide nanoparticles. X-ray diffraction was used to investigate the crystal structure and the average particle size of Zinc Oxide powder around 44.6 nm using the line width of the plane, refraction peak using the Scherer's equation. The nanoparticles were evaluated for antimicrobial activities against Gram-positive and Gram-negative bacteria, which exhibited maximum zone of inhibition against E.coli (19 mm) while least activity was seen against S. aureus. Keywords: Green synthesis, Zinc Oxide nanoparticles, Khat, UV-Visible spectroscopy and