In present investigation, copper oxide (CuO) nanostructures have been prepared via green chemistry. Olea europaea leaf extract act as strong chelating agent for tailoring physical as well as bio-medical characteristics of CuO at the nano-size. Physical characterisation such as scanning electron microscope analysis depicts the formation of homogenised spherical shape nanoparticles (NPs) with average size of 42 nm. X-ray diffraction and Fourier transform infrared spectroscopy further confirmed the crystalline pure phase and monoclinic structure. High performance liquid chromatography (HPLC) testing is performed to evaluate the relative concentration of bioactive molecules in the O. europaea leaf extract. From HPLC results capping action of organic molecules around CuO-NPs is hypothesised. The antimicrobial potency of biosynthesised CuO-NPs have been evaluated using colony forming unit (CFU) counting assay and disc diffusion method which shows a significant zone of inhibition against bacterial and fungal strains may be highly potential for future antimicrobial pharmaceutics. Furthermore, reduction of various precursors by plant extract will reduce environmental impact over chemical synthesis.