Department of Chemistry and Biochemistry, Mendel University in Brno, Zemedelska 1, 61300 Brno, Czech Republic.
Central European Institute of Technology, Brno University of Technology, Technicka 3058/10, 61600 Brno, Czech Republic.
Int J Environ Res Public Health. 2017 Dec 23;15(1):23. doi: 10.3390/ijerph15010023.
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 and/or , 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 and 30 °C for . The CLEAs retained 80% of their initial activity for and 74% for 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 showed 95% decolourisation potential and CLEA from demonstrated 90% decolourisation efficiency within 10 h for all dyes used. These results suggest that these CLEAs have promising potential in dye decolourisation.
获得酶最佳性能的关键通常是设计合适的酶，并优化其固定化条件。在这项研究中，使用了来自捷克森林的天然白腐真菌和/或的漆酶。从这些真菌中，制备了交联酶聚集体（CLEA），并在优化实验条件时对其进行了表征。基于优化步骤，使用饱和硫酸铵溶液（75wt%）作为沉淀剂，并研究了不同浓度的戊二醛作为交联剂。与游离漆酶相比，在所优化条件下形成的 CLEA 聚集体显示出更高的催化效率和稳定性（热稳定性、pH 值稳定性和储存稳定性，防止变性）以及更高的可重复使用性，对于两种真菌菌株都是如此。似乎最佳戊二醛浓度为 50mM，并且在低温 4°C 下观察到更高的交联效率。与游离漆酶相比，两种真菌的 CLEA 漆酶的最佳 pH 值略有增加。结果表明，对于 和 ，游离漆酶和 CLEA 漆酶的最佳温度分别为 35°C 和 30°C。经过 70 天的培养，固定化 CLEA 酶对 和 的初始活性保留了 80%。还研究了制备的交联酶聚集体对孔雀石绿、溴百里酚蓝和甲基红染料的脱色活性。来自 的固定化 CLEA 漆酶对所有使用的染料表现出 95%的脱色潜力，而来自 的 CLEA 漆酶在 10 小时内对所有染料的脱色效率达到 90%。这些结果表明，这些 CLEA 在染料脱色方面具有很大的应用潜力。
