Degradation of azo dyes using laccase immobilized on mechanically ground silk fibroin nanofibers.

The extensive use of azo dyes in the textile industry poses serious environmental and health hazards due to their carcinogenic and mutagenic properties, largely stemming from the stability of their azo bonds (-NN-) which resist natural degradation. Enzymatic azo dye degradation using laccases offers an eco-friendly solution. However, the limited operational stability and reusability of free (non-immobilized) laccases in continuous degradation systems hinders their industrial application. This study developed a continuous azo dye degradation system based on membranes composed of silk fibroin nanofibers functionalized with covalently immobilized laccase (SFNF-laccase). The system was evaluated for catalytic performance, operational stability, and suitability for treating industrial effluents. SFNF-laccase, prepared by covalently immobilizing laccase onto mechanically ground SFNFs, successfully degraded the azo dyes Direct Red 23, Acid Blue 92, and Trypan Blue in the presence of azinobis(3-ethylbenzothiazoline-6-sulfonic acid ammonium salt) as a redox mediator through azo bond cleavage. The immobilized laccase exhibited higher activity across a broader pH and temperature range than free laccase. Notably, it retained 70 % of its initial activity after 9 days of storage (vs. 26 % for free laccase) and maintained 42 % activity after 8 days of continuous operation in a membrane flow-through reactor. It also exhibited moderate resistance to freeze-drying and freeze-thawing, indicating its potential for industrial storage and transportation. These results demonstrate that SFNF-laccase membranes are robust and sustainable biocatalytic platforms for the continuous treatment of azo dye-contaminated industrial effluents.