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Copper oxide nanoparticles are the most common metal oxides. (CuONPs) The good capability of copper-tolerant probiotic of Lactobacillus ssp tolerating, Because of its unique properties and applications, these bacteria have gained more interest as a natural microbial cell nano-factory for a more effective and environmentally friendly method of biosynthesis of these nanoparticles. We examined the antibacterial activity of the Copper Oxide Nanoparticles on some pathogenic bacteria challenge of multidrug resistance (MDR) .It is clear in this the study that Copper Oxide has stronger inhibitory effect on pathogenic bacteriathe largest inhibition region of Cuo NPs from L.plantarium in E coli was 28 mm at 500 g/ml MIC followed by E. faecalis 25mm in the same concentration. The antibiofilm activity of the Copper Oxide Nanoparticles on some pathogenic bacteria formation biofilm was quantified in well plate assay , congo red method and Tube method ,The efect of copper oxide nanoparticles on strong ,modrate biofilm to weak or non formation. Antioxidant activity was determined using 2,2-diphenyl-2-picrylhydrazyl (DPPH), the Copper Oxide Nanoparticles from Lactobacillus plantarum had the highest antioxidant activity (73.60 %,67.11%,58.97 at 250,200,150 μg/ml) in concentrations (250,200,150,100,50) μg/ml.