Covalent immobilization of catalase on periodate-oxidized chitin microspheres for efficient hydrogen peroxide decomposition

Hierarchical CNMs enable covalent catalase immobilization with robust structure. • CAT-ACNMs retain high activity across various pH, solvents, and trypsin conditions. • CAT-ACNMs show excellent reusability and storage stability for real long-term use. A persistent bottleneck in industrial biocatalysis is the engineering of immobilized enzymes that possess both structural resilience and high recyclability under rigorous conditions. Herein, hierarchical chitin-based nanofibrous microspheres (CNMs) were directly constructed via thermally induced self-assembly in a NaOH/urea system, followed by controlled sodium periodate oxidation to introduce aldehyde functionalities for covalent enzyme anchoring. Catalase (CAT) was subsequently anchored through Schiff-base formation, resulting in the CAT-ACNMs complex characterized by a 3D interconnected porous framework. While the immobilization process led to a decrease in the apparent rate constant from 0.42 to 0.032 s −1 , the resulting biocatalyst demonstrated significantly improved environmental tolerance. CAT-ACNMs maintained over 80.0% of their peak activity across a pH range of 5.0–8.0 and exhibited notable stability in various organic solvents (40.7–83.5% retention). Furthermore, the biocatalyst displayed exceptional longevity, preserving ∼90.8% of its initial performance after 8 consecutive cycles and three weeks of storage. The synergistic integration of hierarchical porosity and covalent spatial confinement grants CAT-ACNMs superior operational robustness, underscoring their suitability for sustained biocatalytic processes in complex media. Download: Download high-res image (266KB) Download: Download full-size image Hierarchical CNMs were fabricated via thermally induced self-assembly and aldehyde-functionalized to enable covalent immobilization of catalase, forming CAT-ACNMs with a stable interconnected porous structure. The immobilized enzyme exhibits high activity and enhanced durability, maintaining excellent tolerance to pH, solvents, and trypsin, along with superior reusability and storage stability for long-term applications.