This study outlines the synthesis of biogenic copper oxide nanoparticles (CuONPs) using an extract derived from Cassia fstula Linn (Cf) leaves through a green synthesis approach. Characterization of the synthesized CfBio-CuONPs was carried out using UV- VIS, FTIR, DLS, XRD, and TEM studies. The CfBio-CuONPs exhibited a prominent peak at 272 nm in UV–VIS spectroscopy, and XRD measurements confrmed their crystalline nature. The FTIR spectrum of CfBio-CuONPs revealed the presence of functional groups such as O–H and aromatic groups. TEM analysis confrmed that the CfBio-CuONPs were predominantly spherical with diameters ranging from 15 to 25 nm. Subsequently, the antibacterial potential of CfBio-CuONPs was evaluated against four pathogenic bacteria, including Escherichia coli, Pseudomonas aeruginosa, Staphylococcus epidermidis, and Bacillus subtilis. Among these, B. subtilis exhibited the highest zone of inhibition (26.93±2.01 mm), followed by E. coli (24.25±1.04 mm), P. aeruginosa (23.98±0.97 mm), and S. epidermidis (22.97±1.20 mm). CfBio-CuONPs demonstrated maximum antioxidant activity (78±1.54%) at a dose-dependent concentration of 2000 µg/ml. Furthermore, in vitro toxicity assessment using the toxtrak test indicated that CfBio-CuONPs exhibited a signifcantly stronger toxic efect value/PI against E. coli (93.52%) compared to P. aeruginosa (92.65%), B. subtilis (91.25%), and S. epidermidis (82.89%). These results underscore the notable toxicity of CfBio-CuONPs against E. coli, surpassing that against other bacteria and conventional antibiotics. This study highlights the potential utility of CfBio-CuONPs for eradicating pathogenic microorganisms and suggests potential implications for ecotoxicology
