Biogenic Synthesis and Characterization of Chitosan-CuO Nanocomposite and Evaluation of Antibacterial Activity against Gram-Positive and-Negative Bacteria

Chitosan-copper oxide (CHT-CuO) nanocomposite was synthesized using olive leaf extract (OLE) as reducing agent and CuSO₄·5H₂ O as precursor. CHT-CuO nanocomposite was prepared using an in situ method in which OLE was added to a solution of chitosan and CuSO₄·5H₂ O mixture in the ratio of 1:5 (v/v) and heated at a temperature of 90^◦ C. The obtained CHT-CuO nanocomposite was characterized using field emission scanning electron microscopy (FE-SEM), X-ray diffraction (XRD), ultraviolet-visible (UV-Vis) spectrophotometry, energy-dispersive X-ray spectroscopy (EDAX), Fourier transform infrared spectroscopy (FTIR), and high-resolution transmission electron microscopy (TEM). TEM results indicated that CHT-CuO nanocomposite are spherical in shape with size ranging from 3.5 to 6.0 nm. Antibacterial activity of the synthesized nanocomposites was evaluated against Gram-positive (Bacillus cereus, Staphyloccous haemolytica and Micrococcus Luteus) and Gram-negative (Escherichia coli, Pseudomonas citronellolis, Pseudomonas aeruginosa, kliebisella sp., Bradyrhizobium japon-icum and Ralstonia pickettii) species by cup platting or disc diffusion method. Overall, against all tested bacterial strains, the diameters of the inhibition zone of the three nanocomposites fell between 6 and 24 mm, and the order of the antimicrobial activity was as follows: CuO-1.0 > CuO-0.5 > CuO-2.0. The reference antibiotic amoxicillin and ciprofloxacin showed greater activity based on the diame-ter of zones of inhibition (between 15–32 mm) except for S. heamolytica and P. citronellolis bacteria strains. The nanocomposites MIC/MBC were between 0.1 and 0.01% against all tested bacteria, except S. heamolityca (>0.1%). Based on MIC/MBC values, CuO-0.5 and CuO-1.0 were more active than CuO-2.0, in line with the observations from the disc diffusion experiment. The findings indicate that these nanocomposites are efficacious against bacteria; however, Gram-positive bacteria were less susceptible. The synthesized CHT-CuO nanocomposite shows promising antimicrobial activities and could be utilized as an antibacterial agent in packaging and medical applications.