Abstract: BACKGROUND: The increase in multi-drug resistant bacterial infections has become a major problem in modern healthcare due to the development of bacterial resistance to antibiotics and the development of new antibacterial alternative drug materials is of great importance.
OBJECTIVE: To synthesize and perform a series of characterization of a CeO2 nanoenzyme to investigate its biocompatibility and antibacterial properties against Escherichia coli.
METHODS: CeO2 nanoenzymes were synthesized using a hydrothermal method. The morphology, product composition, and chemical composition were analyzed using characterization methods such as X-ray diffraction, X-ray photoelectron spectroscopy, Fourier infrared analysis, Raman spectroscopy, scanning electron microscopy, and transmission electron microscopy. The peroxide-mimetic enzyme activity of CeO2 nanoenzymes was characterized using TMB color development assay. The toxic effect of CeO2 nanoenzymes at different concentrations (10, 25, and 50 μg/mL) on mouse fibroblast L929 cells was evaluated using the CCK-8 assay. The antibacterial properties of CeO2 nanoenzymes against Escherichia coli under different conditions were evaluated using the plate coating method. Changes in intra-bacterial reactive oxygen species after treatment with different conditions were detected using a reactive oxygen species detection kit.
RESULTS AND CONCLUSION: (1) The morphology of the synthesized CeO2 nanoparticles was rod-shaped, with Ce3+ accounting for 29.87% of the total Ce3+/Ce4+ and an average grain size of 7.4 nm. In a slightly acidic environment containing TMB and pH=5.5, CeO2 nanoenzymes mixed with H2O2 showed excellent peroxidase activity, but did not show peroxidase simulated activity at pH=7.4. (2) There was no statistically significant difference in the toxic effects of CeO2 nanoparticles at various mass concentrations on mouse fibroblast L929 cells. (3) In a slightly acidic environment at pH 5.5, Escherichia coli was inhibited to a certain extent in the presence of CeO2 nanoenzyme alone at a concentration of 10 μg/mL, with a decrease in CFU results of about 0.5 log (P < 0.01); in a slightly acidic environment containing 50 μmol/L H2O2, CeO2 nanoenzyme showed excellent antibacterial effects against Escherichia coli, with a decrease in Escherichia coli CFU results of by about 1.5 log (P < 0.001). After CeO2 nanoenzymes interacted with Escherichia coli, the level of reactive oxygen species in Escherichia coli increased (P < 0.05); after CeO2 nanoenzymes interacted with Escherichia coli together with H2O2, the level of reactive oxygen species in Escherichia coli increased significantly (P < 0.001). (4) The results show that the CeO2 nanoenzymes have good biocompatibility, are inherently antibacterial, and can exhibit peroxidase activity in a slightly acidic environment containing low concentrations of H2O2, and generate reactive oxygen species to kill bacteria, thus showing excellent antibacterial effects.

Key words: cerium oxide, nanoenzyme, nanomaterial, antibacterial property, biocompatibility, Escherichia coli