Azadirachta indica -Mediated Synthesis of Iron Oxide Nanozymes with Superior Peroxidase-Like Catalytic Efficiency

Background: Metal-based nanomaterials with natural enzyme-like activity, nanozymes are gaining popularity as alternatives to biological enzymes due to their superior stability and cost-effectiveness. The catalytic efficacy of iron oxide nanozymes may be diminished by aggregation during chemical production. Objective: This study aimed to synthesize iron oxide nanozymes using Azadirachta indica leaf extract and compare their structural composition and OPD–H₂O₂ peroxidase-like catalytic activity with chemically synthesized nanozymes. Methods: Iron oxide nanozymes were synthesized using chemical co-precipitation and Azadirachta indica-mediated green synthesis. The synthesized materials were characterized by scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDS). Peroxidase-like activity was evaluated using oxidation of o-phenylenediamine (OPD) in the presence of hydrogen peroxide(H2O2). Kinetic parameters were estimated using Michaelis–Menten analysis. Results: Green-synthesized nanozymes showed comparatively reduced aggregation and lower residual chlorine signal than chemically synthesized nanozymes. Azadirachta indica-mediated nanozymes generated the maximum absorbance at 451 nm in the OPD–H2O₂ assay, suggesting more peroxidase-like activity. Kinetic analysis revealed that green-synthesized nanozymes had a higher Vmax and a roughly 5.4-fold higher Vmax/Km-based catalytic effectiveness than chemically synthesized nanozymes. Conclusion: Iron oxide nanozymes with enhanced catalytic performance were generated by Azadirachta indica-mediated synthesis, most likely as a result of phytochemical-assisted stabilization and surface modification. Before biosensing or environmental usage, additional phase characterization, replicate-based validation, and application-specific testing are needed