Compartmentalized multi-enzyme immobilization in hierarchical shell-shell structure for glucose detection

Recent years, incorporation of multiple enzymes into matrix is considered as promising candidates for constructing highly sensitive glucose-responsive systems. However, the stochastic nature of traditional multi-enzyme immobilization techniques makes it challenging to precisely control the spatial arrangement of enzyme molecules. Here, a new hierarchical shell-shell (HSS) multi-enzyme structure was constructed using interfacial growth method and colloidal hydrated zinc sulphate to be used as a soft template (Cyt C@HSS@GOx). In this process, glutathione (GSH) hybrid ZIF-67 was used as the guest MOF and hollow ZIF-8 as the host MOF. The resultant Cyt C@HSS@GOx exhibited high activity by separating cytochrome C (Cyt C) and glucose oxidase (GOx) in two independent compartments. This method can accurately locate the spatial position of multiple enzymes, weaken enzyme-to-enzyme inhibition, and enhance the channeling effect of substrates, thus improving the catalytic performance of multiple enzymes. The resulting Cyt C@HSS@GOx showed 230% higher activity than free enzymes, and maintained good stability under extreme pH and temperature conditions. Furthermore, Cyt C@HSS@GOx demonstrated 71.25% retention of its initial activity following seven cycles, providing good reusability. Particularly, Cyt C@HSS@GOx has been demonstrated to possess a glucose detection limit as low as 0.38 μmol/L, with a reaction time of only three minutes. This strategy provides a rapid and accurate glucose detection method.