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The use of biomaterials in the synthesis of nanoparticles is one of the most up-to-date focuses in modern nanotechnologies and nanosciences. More and more research on green methods of producing metal oxide nanoparticles (NP) is taking place, with the goal to overcome the possible dangers of toxic chemicals for a safe and innocuous environment. In this study, we synthesized copper nanoparticles (CuNPs) using Fortunella margarita leaves’ extract, which reflects its novelty in the field of nanosciences. The visual observation of a color change from dark green to bluish green clearly shows the instant and spontaneous formation of CuNPs when the phytochemicals of F. margarita come in contact with Cu⁺² ions. The synthesis of CuNPs was carried out at different conditions, including pH, temperature, concentration ratio and time, and were characterized with UV-Vis absorption spectra, scanning electron microscope (SEM) and X-ray diffraction (XRD). The UV-Vis analysis reveals the surface plasmon resonance property (SPR) of CuNPs, showing a characteristic absorption peak at 679 nm, while SEM reveals the spherical but agglomerated shape of CuNPs of the size within the range of 51.26–56.66 nm.