One‐Pot and Gram‐Scale Synthesis of Fe‐Based Nanozymes with Tunable O2 Activation Pathway and Specificity Between Associated Enzymatic Reactions

Abstract Nanozymes have recently gained attention for their low cost and high stability. However, unlike natural enzymes, they often exhibit multiple enzyme‐like activities, complicating their use in selective bioassays. Since H 2 O 2 and O 2 are common substrates in these reactions, controlling their activation—and thus reaction specificity—is crucial. Recent advances in tuning the chemical state of cerium have enabled control over H 2 O 2 activation pathways for tunable peroxidase/haloperoxidase‐like activities. In contrast, the control of O 2 activation on an element in oxidase/laccase nanozymes and the impact of its chemical state on these activities remains unexplored. Herein, a facile one‐pot method is presented for the gram‐scale synthesis of Fe‐based nanozymes with tunable compositions of Fe 3 O 4 and Fe 3 C by adjusting preparation temperatures. The Fe 3 O 4 ‐containing samples exhibit superior laccase‐like activity, while the Fe 3 C‐containing counterparts demonstrate better oxidase‐like activity. This divergent O 2 activation behavior is linked to their surface Fe species: the abundant reactive Fe 2+ in Fe 3 O 4 promotes laccase‐like activity via Fe 3+ ‐superoxo formation, whereas metallic Fe in Fe 3 C facilitates OH radical generation for oxidase‐like activity. Controlled O 2 activation pathways in these Fe‐based nanozymes demonstrate improved sensitivity in the corresponding biomolecule detection, which should inform the design of nanozymes with enhanced activity and specificity.