Hydrogen peroxide (H2O2) is a key reactive oxygen species involved in physiological regulation and pathological pro‑

cesses, and its accurate quantification is essential for bioanalysis and disease-related studies. Herein, we report the synthe‑

sis of a series of Cu-Fe oxide nanozymes via a simple antisolvent crystallization strategy and systematically investigate 

their peroxidase-like catalytic activity toward colorimetric H2O2 detection. By tuning the Cu/Fe molar ratio, the catalytic 

activity of the nanozymes can be effectively regulated, with Cu2FeOx exhibiting the highest peroxidase-like activity toward 

3,3′,5,5′-tetramethylbenzidine (TMB) oxidation in the presence of H2O2 under acidic conditions and enabling sensitive 

colorimetric detection of H2O2 over a linear range of 10–200 µM with a detection limit of 0.572 µM. Furthermore, the 

proposed sensing platform demonstrates satisfactory selectivity and reproducibility, and is successfully applied to the 

determination of H2O2 released into cell culture supernatants after chemical stimulation. This work highlights the potential 

of composition‑regulated Cu-Fe oxide nanozymes as robust artificial enzymes for colorimetric H2O2 sensing and provides 

insights into their application in bioanalytical systems.