Fabrication of imine-based COFs with different linker lengths for HRP and GOx immobilization and colorimetric detection of glucose and H2O2

Abstract

Covalent organic frameworks (COFs) have emerged as excellent candidates for enzyme immobilization due to their high surface tunability, diverse structures, inherent porosity, and metal-free characteristics. However, reports on optimizing the efficiency of immobilized enzymes in COFs by controlling linker length are scarce. In this work, horseradish peroxidase (HRP) and glucose oxidase (GOx), are co-immobilized on three imine-based COFs with different linker lengths using a one-pot method, resulting in HRP&GOX@COF (COF = LZU1, TbBD, TbDI). The immobilization of HRP and GOX using these three COF materials led to a slight decrease in enzyme activity compared to the free enzymes; however, a significant improvement in the stability of the immobilized enzymes was achieved. Besides, the experimental results demonstrate that all three materials HRP&GOX@COF (COF = LZU1, TbBD, TbDI) exhibit good colorimetric detection, stability, and recyclability for glucose and hydrogen peroxide (H₂O₂), using 2,2′-biazobis (3-ethylbenzothiazole-6-sulfonic acid) diammonium salt (ABTS) as a chromogenic agent. Notably, HRP&GOX@LZU1, which has the shortest linker, shows the highest enzyme loading efficiency (45.3 %) and the most effective colorimetric detection, with detection limits of 0.265 mM for glucose and 0.100 mM for H₂O₂. This work highlights the importance of linker length in designing COFs for enzyme immobilization and colorimetric recognition of H₂O₂ and glucose.