Double-crystallized GOx-entrapped copper nanoflowers enabling interfacial cascade catalysis for electrochemical glucose sensing.

Artificial enzyme-nanozyme cascade systems that enable interfacial coupling between biocatalytic and inorganic redox processes are of significant interest for biointerface engineering. In this study, a double-crystallized Cu nanoflower-based enzyme-nanozyme cascade platform was introduced by immobilizing GOx-integrated Cu nanoflowers (D-Cu@GOx NFs) onto an electrode surface. The double-crystallization process generates a hierarchical Cu nanoflower architecture that spatially confines enzyme and nanozyme components, enabling proximity-driven cascade catalysis through a redox-active Cu framework. Electrochemical characterization revealed that incorporation of GOx slightly modified the interfacial redox behavior while maintaining quasi-reversible electron-transfer characteristics. Chronoamperometric measurements demonstrated concentration-dependent responses toward both H2O2 and glucose, confirming that the Cu nanoflower structure retains intrinsic peroxidase-like activity through reversible Cu2+/Cu+ redox cycling. The D-Cu@GOx NF modified electrode exhibited a linear glucose response up to 31200 µM, with analytical parameters calculated from the linear region below 5200 µM. These results indicate that the integrated D-Cu@GOx NF architecture retains both enzymatic glucose-responsive and Cu-associated electrochemical functionality, supporting its application as a hybrid enzyme-nanozyme electrochemical interface. This work demonstrates a double-crystallized nanoflower platform as a biointerface for integrating enzyme-nanozyme cascade systems.