Preparation and Optimisation of Cross-Linked Enzyme Aggregates Using Native Isolate White Rot Fungi Trametes versicolor and Fomes fomentarius for the Decolourisation of Synthetic Dyes

Abstract

The key to obtaining an optimum performance of an enzyme is often a question of devising a suitable enzyme and optimisation of conditions for its immobilization. In this study, laccases from the native isolates of white rot fungi Fomes fomentarius and/or Trametes versicolor, obtained from Czech forests, were used. From these, cross-linked enzyme aggregates (CLEA) were prepared and characterised when the experimental conditions were optimized. Based on the optimization steps, saturated ammonium sulphate solution (75 wt.%) was used as the precipitating agent, and different concentrations of glutaraldehyde as a cross-linking agent were investigated. CLEA aggregates formed under the optimal conditions showed higher catalytic efficiency and stabilities (thermal, pH, and storage, against denaturation) as well as high reusability compared to free laccase for both fungal strains. The best concentration of glutaraldehyde seemed to be 50 mM and higher efficiency of cross-linking was observed at a low temperature 4 °C. An insignificant increase in optimum pH for CLEA laccases with respect to free laccases for both fungi was observed. The results show that the optimum temperature for both free laccase and CLEA laccase was 35 °C for T. versicolor and 30 °C for F. fomentarius. The CLEAs retained 80% of their initial activity for Trametes and 74% for Fomes after 70 days of cultivation. Prepared cross-linked enzyme aggregates were also investigated for their decolourisation activity on malachite green, bromothymol blue, and methyl red dyes. Immobilised CLEA laccase from Trametes versicolor showed 95% decolourisation potential and CLEA from Fomes fomentarius demonstrated 90% decolourisation efficiency within 10 h for all dyes used. These results suggest that these CLEAs have promising potential in dye decolourisation.

Crosslinked Enzyme Aggregates (CLEAs) of Laccases from Pleurotus citrinopileatus Induced in Olive Oil Mill Wastewater (OOMW)

“…Some of the most recent efforts include the preparation of laccase CLEAs and their application to the decolorization of triphenyl methane and reactive dyes [ 29 ], the development of laccase-containing combi-CLEAs for the degradation of pharmaceuticals [ 30 ], and the immobilization of laccase in combi-CLEAs and their application in acetaminophen degradation [ 19 ]. For dye decolorizations, other CLEAs prepared from various laccases have also been employed, such as from Pleurotus ostreatus [ 29 ], Trametes versicolor [ 31 , 32 ], Fomes fomentarius [ 31 ], and Cerrena spp. [ 33 ].…”

Influence of Glutaraldehyde Cross-Linking Modes on the Recyclability of Immobilized Lipase B from Candida antarctica for Transesterification of Soy Bean Oil

“…However, it was described that high concentrations of cross-linking agent increased the enzyme rigidity, which might have influenced in the active site mobility, leading to a decrease in the reaction yield [ 27 ]. Moreover, excessive glutaraldehyde concentrations might affect the tridimensional structure of the enzyme [ 28 ].…”

Microbial lipases and their industrial applications: a comprehensive review

“…Enzymes obtained by crystallization, atomization, and aggregation leads by crosslinking approaches [ 174 ]. The immobilization of the enzyme arises subsequent in the development of Cross-Linked (soluble) Enzyme (CLE), Cross-Linked Enzyme Crystals (CLEC), Crosslinked Enzyme Aggregates (CLEA), and Cross-Linked Spraydrying Enzyme (CSDE) when these enzymes are placed in a medium containing a cross-linking agent [ 175 , 176 ]. Due to the elimination of solid support; besides being an adaptable process are highly catalyzed enzymatic activities, high stability, and low cost of production is the main advantage of cross-linking process, it is possible to get more healthy and stable enzymes for industrial uses [ 177 , 178 ].…”