The Biocatalytic Degradation of Organic Dyes Using Laccase Immobilized Magnetic Nanoparticles

Free laccase has limitations for its use in industrial applications that require laccase immobilization on proper support, to improve its catalytic activity. Herein, the nanoparticles of magnetic iron oxide (Fe3O4) and copper ferrite (CuFe2O4) were successfully used as support for the immobilization of free laccase, using glutaraldehyde as a cross-linker. The immobilization conditions of laccase on the surface of nanoparticles were optimized to reach the maximum activity of the immobilized enzyme. The synthesized free nanoparticles and the nanoparticle-immobilized laccase were characterized using different techniques, including X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR), scanning electron microscope (SEM), vibrating sample magnetometer (VSM), and thermogravimetric analysis (TGA). CuFe2O4 nanoparticles, as support, enhanced laccase activity compared to free laccase and Fe3O4 nanoparticle-immobilized laccase that appeared during the study of pH, temperature, and storage stability on free and immobilized laccase. The CuFe2O4 and Fe3O4 nanoparticle-immobilized laccase showed superior activity in a wide pH range, temperature range, and storage period, up to 20 days at 4.0 °C, when compared to free laccase. Additionally, the synthesized nanobiocatalysts were examined and optimized for the biodegradation of the anionic dye Direct Red 23 (DR23). HPLC analysis was used to confirm the dye degradation. The reusability of immobilized laccases for the biodegradation of DR23 dye was investigated for up to six successive cycles, with a decolorization efficiency over 70.0%, which indicated good reusability and excellent stability.

Preparation and characterization of oxidized Multi-walled Carbon Nanotubes-Immobilized Aspergillus sp. Laccase Hybrid Materials

This work deals with preparation and characterization of immobilized laccase (Aspergillus sp.) over oxidized multi-walled carbon nanotubes (ox-MWCNTs) via simple mixing technique. The resulting materials were characterized by Fourier transform infrared spectroscopy (FTIR), thermogravimetric analysis (TGA), transmission electron microscope (TEM) and particle size distribution analysis using dynamic light scattering technique (DLS). The results showed that the TEM images exhibited more separate individual carbon bundles with particle size around of 396 nm after enzyme immobilization rather than the spaghetti-like tubes with size about 180 nm in the case of ox-MWCNTs. Also, the lowering in the zeta potential negative value (-5 mv) proved that the free carboxyl groups at ox-MWCNT surface were decreased after enzyme immobilization. Moreover, the thermal stability was decreased after enzyme immobilization using TGA. These results confirmed that the laccase could be reacted at the side walls of the ox-MWCNTs without structure damage. The biocatalytic effect of the immobilized laccase was investigated after its incubation with silver nitrate solution for 1 and 24 h. It can be concluded that the biocatalytic efficiency of the immobilized laccase could be enhanced after its incubation with silver nitrate solution for 24 h at room temperature relative to the free form. On the other hand, the enzyme stability was improved after immobilization up to 50ºC and at pH 3.0, while no remarkable differences on the activity values were observed for immobilized and free laccases at acidic pH range (4-6).

Degradation and Detoxification of Congo Red azo dye by Immobilized Laccase of Streptomyces sviceus

The discharge of textile effluents enriched with reactive azo dyes is of critical importance owing to inability of the dyes to degrade in waste water and their carcinogenic, mutagenic effects to various organisms. This study initiated based on the need to gaze into molecular mechanism of marine bacterial bioremediation process to develop strategies for the decolorization and detoxification of the synthetic azo dyes. The experimental work carried out to explore decolorization and degradation efficacy of laccase derived from marine actinobacteria, Streptomyces sviceus by choosing Congo red-21 as model azo dye. The extracellular production of laccase was confirmed with plate assay in medium supplemented with ABTS as substrate. Laccase was purified to homogeneity from 72hrs culture of Streptomyces sviceus by Fast performance liquid chromatography and the molecular size of laccase was noticed as 60 kDa. The purified laccase was immobilized with an efficiency of 82% by Calcium alginate method. The crude, purified and immobilized forms of the laccase enzyme was used to decolorize the Congo red-21. Crude laccase enzyme showed 69% of decolorization of Congo red-21 after 48h where as purified and immobilized laccase represented 78% and 92% of colour removal after 24 h respectively. Fourier-transform infrared spectroscopy, High Performance Liquid Chromatography and Gas chromatography–mass spectrometry were used to unravel the molecular mechanism of dye detoxification and also identify nontoxic products released from Congo Red-21 upon administration with immobilized laccase. Based on GC-MS data, it may deduce that immobilized laccase of Streptomyces sviceus cleaves the Congo red-21 dye followed by oxidative cleavage, desulfonation, deamination, demethylation process.

Optimization and efficacy studies of Laccase immobilized on Zein-Polyvinyl pyrrolidone nano fibrous membrane in decolorization of Acid Red 1

Azo dyes are widely used in textile industries. A significant portion of these recalcitrant dyes are being discharged to the natural waters. Due to their low biodegradability they pose serious pollution problems if untreated. In this work, decolourization studies of Acid Red 1 (AR1) by laccase enzyme immobilized onto Zein – Polyvinyl pyrrolidone composite nanofiber is done. The nanofibers were characterized by SEM and FTIR analysis. pH and temperature profiles of immobilized enzyme were found to be broader than its free counterpart. The Km value was found to be 0.243 mM for free laccase and 0.958 mM for immobilized laccase. Similarly Vmax for the free enzyme was 3.572 U/mg compared to 2.48 U/mg of immobilized laccase. The relative activity of immobilized laccase was 64.91% after storage for 30 days at room temperature while it was 28.64% for free laccase. The temperature and pH for AR 1 decolorization were optimized and was found to be 60 °C and 5 respectively. Also decolorization percentage was found to be 91.67% for immobilized laccase and 72.03% of free laccase in the presence of natural mediators like vanillin. From phytotoxicity studies it was found that the germination rate, shoot and root length was increased compared to untreated dye. Therefore, Zein-PVP nanofiber immobilized laccase could be an ideal candidate for the textile dye decolorization.