One of the most recent areas of interest in current nanotechnologies and nanosciences is the use of biomaterials in the manufacturing of nanoparticles. More and more research is being carried out on environmentally friendly methods to create metal oxide nanoparticles (NP), with the intention of preventing any potential risks associated with harmful substances for a safe and healthy environment. In this study, Copper Oxide (CuO) is synthesized utilizing Opuntia ficus‐indica as the plant extract using a Microwave Combustion Technique (MCM) and its comparison against the Conventional Combustion Method (CCM) are investigated. The synthesized CuO nanoparticles were characterized using X‐ray Diffraction (XRD), Fourier Transform Infrared Spectroscopy (FT‐IR), Scanning Electron Microscopy (SEM), and Energy Dispersive X‐ray Analysis (EDX) analysis. Photoluminescence spectroscopy was undertaken to acquire emission and absorption spectra and determine defects in the structures of all synthesized nanopowder samples. The antibacterial activity of the CuO nanoparticles was evaluated in‐vitro using gram‐negative and gram‐positive bacteria (Bacillus subtilis, Staphylococcus aureus). Enhanced anti‐bactericidal activity was shown against Gram‐negative bacteria compared to Gram‐positive bacteria. Through these findings, the use of CuO using Opuntia ficus indica extracts is hereby shown to be a cost‐effective and environmentally friendly alternative and that can be used in a variety of applications.