In-situ encapsulated enzymes in covalent-organic frameworks for glucose biosensor design with high-performance cascade catalysis

• A bio-friendly strategy to encapsulate multienzymes in COF is developed. • This process is simple and can be used for the glucose biosensor fabrication. • This biosensor detects glucose in serum with high sensitivity and good selectivity. • The biosensor shows promising for the diagnosis of diabetic patients. Covalent organic frameworks (COFs) are desired capsule for enzyme immobilization. However, the synthetic conditions of COFs usually require harsh conditions, such as high temperatures, acid/ base catalysts, or organic solvents. Many reported approaches for enzyme immobilization are physical adsorption, post-modification of COF, or MOF as a sacrificial template, which are low loading efficiency, and require multistep with complex processes. Here, we used a bio-friendly, one-step strategy to encapsulate HPR and glucose oxidase (GOx) in COF (GOx-HRP@COF) in aqueous solution at room temperature. Compared to two-step or mixture encapsulation approaches, GOx-HRP@COF biocomposites show superior enzymatic bioactivity and improved stability under a variety of harsh environments. Moreover, they can be used for biosensor fabrication for glucose detection based on a cascade catalytic system. After optimizing the experimental parameters, the limit of detection (LOD) of glucose was 28 μM with a wide detection range of 0.05–8 mM. Moreover, the glucose in serum can be determined with this system and the results fit the concentration window of glucose in clinical test. This biocomposite is easy to operate and has a remarkable 90 % enzymatic activity preservation over 7 days, showing significant potential value in point-of-care testing (POCT). Moreover, the biosensor can be extended to other substrates by altering the immobilized enzymes, showing great prospective values to fabricate another sensing platform.