Dual-Enzyme Cascade Composed of Chitosan Coated FeS₂ Nanozyme and Glucose Oxidase for Sensitive Glucose Detection

Immobilizing enzymes with nanozymes to catalyze cascade reactions overcomes many of the shortcomings of biological enzymes in industrial manufacturing. In the study, glucose oxidases were covalently bound to FeS₂ nanozymes as immobilization carriers while chitosan encapsulation increased the activity and stability of the immobilized enzymes. The immobilized enzymes exhibited a 10% greater increase in catalytic efficiency than the free enzymes while also being more stable and catalytically active in environments with an alkaline pH of 9.0 and a high temperature of 100 °C. Additionally, the FeS₂ nanozyme-driven double-enzyme cascade reaction showed high glucose selectivity, even in the presence of lactose, dopamine, and uric acid, with a limit of detection (LOD) (S/N = 3) as low as 1.9 × 10⁻⁶ M. This research demonstrates that nanozymes may be employed as ideal carriers for biological enzymes and that the nanozymes can catalyze cascade reactions together with natural enzymes, offering new insights into interactions between natural and synthetic biosystems.