Recently, focus has been placed on renewable sources, as they can be provided in large quantities at the lowest possible cost, in order to create nanoparticles. One of these sources is Zygnema moss which used in the present investigation to create Copper oxide nanoparticles (CuONPs). Several phenols and flavonoids were identified the extract of Zygnema sp. via analysis of High performance liquid chromatography. These constituents served as reducing and stabilizing agents for CuONPs. Characterization of CuONPs was performed via UV-visible spectrum that demonstrated peak at 252 nm, Transmission electron microscopy that showed spherical CuONPs with mean diameter of 30.06 nm, Fourier transform infrared spectroscopy that confirm that presence of several functional groups aided to formation of CuONPs. The crystallographic pattern of CuONPs was recorded via X-ray diffraction analysis. Antimicrobial potential of CuONPs was compared to copper acetate and antibiotic/antifungal drug. CuONPs exhibited more inhibition zones against S. aureus (32 ± 0.1 mm), E. coli (36 ± 0.1 mm), S. typhi (27 ± 0.2 mm), E. faecalis (37 ± 0.1 mm), C. albicans (34 ± 0.3 mm) than copper acetate and antibiotic/antifungal drug. Promising MIC values of were recorded against S. aureus, E. coli, and S. typhi. CuONPs at 200 ppm inhibited the growth of C. lunata, F. oxysporium, A. flavus, and Mucor circinelloid with inhibtion of 76.92, 73.33, 63.63, and 53.84%, respectively regarded the control 100% growth. The photocatalytic role of CuONPs was recorded for degradation of reactive red (RR195) and reactive blue (RB) dyes with maximum degradation of 84.66% and 90.82%, respectively at 75 min. Moreover, the optimal dyes degradation was 84.66 and 90.82%, respectively at 40 °C.