Stoichiometric Regulation of Gallic Acid-Copper Nanozymes for Optimized Coordination Structure, Peroxidase-Like...

Gallic acid (GA)-copper nanozymes were prepared at GA:Cu²⁺ stoichiometric ratios ranging from 1:1 to 5:1 to elucidate how stoichiometry governs coordination assembly, colloidal stability, electronic structure, peroxidase-like catalysis, and antibacterial activity. Among all formulations, the 2:1 sample exhibited the smallest hydrodynamic diameter (95.65 ± 4.20 nm), the lowest polydispersity index (0.19 ± 0.01), and the most negative zeta potential (-27.7 ± 2.80 mV), indicating superior dispersion stability. Morphological and XRD analyses revealed a relatively uniform, weakly aggregated, and predominantly amorphous metal-phenolic network. FT-IR, 2D-COS, UV-Vis, and XPS collectively confirmed Cu─O coordination and stoichiometry-dependent modulation of the local electronic environment; the appearance of a new absorption band near 320 nm was attributed to ligand-to-metal charge transfer. The 2:1 nanozyme also showed the highest peroxidase-like activity, with Km values of 0.034 mM for TMB and 4.39 mM for H₂O₂. EPR analysis verified ·OH generation during H₂O₂ activation. DFT, BCP, and IGMH analyses further demonstrated that the optimal 2:1 configuration adopted the most stable coordination mode, with the lowest bond energy (-69.8834 kcal/mol) and a HOMO-LUMO gap of 5.7082 eV. In antibacterial assays, the optimized nanozyme exhibited concentration-dependent killing against both Escherichia coli and Staphylococcus aureus, which was markedly enhanced in the presence of H₂O₂. These results establish a stoichiometry-guided structure-performance relationship for GA-Cu nanozymes and demonstrate that stoichiometric regulation is an important design parameter for optimizing their coordination structure, colloidal stability, and catalytic activity. Overall, the optimized GA-Cu_2:1 formulation provides a rational basis for further development and evaluation of metal-phenolic catalytic antibacterial materials for potential food-safety applications. PRACTICAL APPLICATIONS: Utilizes naturally abundant gallic acid and copper ions as sustainable, low-cost raw materials for nanozyme construction. Stoichiometric regulation (GA:Cu² = 2:1) achieves an optimal coordination structure, colloidal stability, and peroxidase-like activity, enabling enhanced ROS-mediated antibacterial efficacy. Offers a biocompatible, catalytic antibacterial system for active food packaging or antimicrobial coatings to extend the shelf life of chilled meat and other perishable foods.