Nickel oxide nanoparticles synthesized at room temperature (RT NiO NPs) and 60°C temperature treated nickel oxide nanoparticles (RT60 NiO NPs) were synthesized by the co-precipitation methods. The antibacterial studies performed against set of (gram positive) S.aureus and (gram negative) E. coli bacterial strains showed that the RT60 NiO NPs possessed a greater antibacterial effect as compared to the RT NiO NPs. The cytotoxic effect of the RT NiO and RT60 NiO NPs were examined in cultured (MCF-7) human breast cancer cells.