DNA-encoded copper nanozymes with tunable peroxidase -mimicking activity for colorimetric detection of glyphosate

Glyphosate (Gly), a widely used organophosphorus pesticide, has raised concern due to its environmental persistence and adverse health effects. Conventional detection methods are often limited by high expenses, complex operations, and insufficient sensitivity, making them unsuitable for on-site application. Herein, we report a colorimetric biosensor based on DNA-encoded copper (DNA/Cu) nanozymes for the sensitive detection of Gly. The DNA/Cu nanozymes were synthesized via a one-step coordination-driven self-assembly at room temperature without additional reagents, and their peroxidase-like activity was modulated by altering the DNA sequence. The nanozyme efficiently catalyzes the H₂O₂-mediated oxidation of 3,3',5,5'-tetramethylbenzidine (TMB), producing a blue coloration. Mechanistic studies confirm that hydroxyl radicals (·OH) generated from H₂O₂ decomposition are the dominant reactive intermediates. Gly suppresses this activity by chelating Cu²⁺ and disrupting the nanozyme structure, thereby blocking the generation of ·OH. Based on this inhibition mechanism, the biosensor exhibits a good linear response to Gly from 0.1 to 5 μg/mL, with a detection limit of 13 ng/mL and excellent specificity. To facilitate on-site monitoring, the biosensor was integrated with a smartphone-based colorimetric reader, enabling quantitative analysis via RGB ratio measurement and achieving a practical LOD of 35 ng/mL. This DNA/Cu nanozyme-based colorimetric platform represents a promising tool for Gly detection in food safety and environmental monitoring.