Dendritic cross-linked MXene@Fe3O4 for co-immobilization of HRP/GOX bienzyme system and its application in 2,4-dichlorophenol degradation

To address the challenges of quantitative immobilization and unavoidable spatial hindrance of multiple enzymes in traditional multi-enzyme cascade immobilization, we developed a novel bienzyme immobilization system based on a hierarchically structured MXene@LDHs@Fe₃O₄ (MLF) carrier. The MLF composite, with a high theoretical surface area (∼300 m 2 /g), adjustable interlayer spacing (10–50 nm), and strong magnetization (20 emu/g), provides an ideal support for high-capacity enzyme loading. The system was fabricated through the assembly of MXene nanosheets with layered double hydroxides (LDHs) and magnetic Fe₃O₄ nanoparticles, integrating horseradish peroxidase (HRP) on the MLF core, a polydopamine (PDA) interfacial barrier, and surface-immobilized glucose oxidase (GOx) through melamine-genipin mediated dendritic cross-linking. This spatially organized catalytic architecture enables quantitative enzyme loading while minimizing spatial hindrance, thereby enhancing catalytic efficiency. The co-immobilized bienzyme reactor (GOX@PDA@HRP@MLF) achieved loading capacities of 272.46 mg/g for HRP and 414.33 mg/g for GOx, with an activity recovery of 85.3 % during 2,4-dichlorophenol degradation. The immobilized enzymes retained ∼80 % of their initial activity after five operational cycles. Under optimized conditions, the GOX@PDA@HRP@MLF material achieved a degradation rate of 4.6 for 2,4-dichlorophenol in a continuous flow system, representing a ∼21-fold increase in space-time yield compared to random multi-enzyme immobilization on single-layer resin materials. These results demonstrate that the dendritic cross-linking strategy employed in this study can serve as a universal method for multi-enzyme immobilization, with significant potential for industrial applications.