Herein, zinc oxide nanoparticles (ZnO NPs) were synthesized using Parthenium hysterophorus whole plant aqueous extract as reducing and capping agents. The synthesized ZnO NPs were characterized via UV-Vis spectroscopy, Fourier-transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), transmission electron microscopy (TEM), X-ray diffraction (XRD), and dynamic light scattering (DLS). An intrinsic optical absorbance of ZnO NPs occurred at 337 nm in the UV-Vis spectrum. The FTIR analysis revealed the presence of secondary metabolites responsible for reducing and stabilizing the nanoparticles. Furthermore, SEM and TEM images revealed that ZnO NPs were spherical with an average particle size of 38 nm. The XRD analysis revealed that ZnO NPs had a hexagonal wurtzite crystal structure with a crystallite size of 42.6 nm. The synthesized nanoparticles were investigated for degradation ability against methylene blue dye at varying conditions of ZnO NPs’ dosage, methylene blue concentrations, pH, temperature, and interaction time. Degradation efficiency of 55.69% was obtained at optimal conditions using 50 mg of ZnO NPs, 5 mg/L of MB dye concentration, and pH 12 and at 65°C within 32 minutes. Due to their novel green synthesis route, Parthenium hysterophorus-mediated ZnO NPs are promising candidates for removing persistent organic dyes from aquatic environments.