红芝LYL263所产漆酶在溶胶-凝胶体系中的固定化研究

Immobilization of Laccase from Ganoderma lucidum LYL 263 in Sol-gel System

[目的]对红芝LYL263漆酶进行不同溶胶-凝胶体系的固定化研究,以期找到适宜该漆酶的固定化材料。[方法]将海藻酸钠-明胶、海藻酸钠-壳聚糖和海藻酸钠-明胶-壳聚糖3种溶胶-凝胶体系用于红芝LYL263所产漆酶的固定化研究,并初步分析固定化酶的酶学性质。[结果]海藻酸钠-明胶-壳聚糖体系固定后的漆酶酶活分别是海藻酸钠-明胶和海藻酸钠-壳聚糖的2.14倍和2.75倍,该体系最佳材料组成为:海藻酸钠浓度2.0%、明胶浓度1.0%、壳聚糖浓度0.3%、氯化钙浓度36.0%。该试验首次将苯甲酸用于包埋法固定化漆酶,单因素试验结果显示,适宜的苯甲酸浓度能有效提高固定化漆酶酶活;该体系固定化漆酶最佳条件为:苯甲酸浓度2mmol/L、戊二醛浓度0.32%、交联时间50 min、酶浓度10.0%,此时固定化酶活高达635.7 U/g。对固定化漆酶酶学性质的研究表明,固定化酶的热稳定性比游离酶高,冻干的固定化酶活是未冻干固定化酶的1.59倍,但冻干的固定化酶操作稳定性较差。[结论]在酶固定化材料中,各个物质的组成对酶固定化有非常大的影响,该研究对红芝LYL263漆酶的固定化材料选择有重要意义。

[Objective] The aim was to study the immobilization of laccase from Canoderma lucidum in three different sol-gel systems to find out the optimum materials for laccase immobilization.[Method] Laccase from G.lucidum LYL 263 was immobilized in sodium alginate-gelatin,sodium alginate-chitosan and sodium alginate-gelatin-chitosan sol-gel systems,respectively;and the laccase activity was preliminarily analyzed.[Result] The activity of laccase immobilized in sodium alginate-gelatin-chitosan was 2.14 folds and 2.75 folds respectively,compared to sodium alginate-gelatin and sodium alginate-chitosan;and this optimum system contained sodium alginate 2.0%,gelatin 1.0%,chitosan 0.3% and calcium chloride 36%.Benzoic acid was firstly used to embed immobilized laccase in this study,and single-factor experiments revealed that the activity of laccase could be effectively improved by suitable content of benzoic acid.The optimal conditions for laccase immobilization in the system were benzoic acid 2 mmol/L,glutaraldehyde 0.32%,cross-linking reaction time 50 min and enzyme concentration 10.0%;and then the activity of the immobilized enzyme reached 635.7 U/g with these conditions.The thermal stability of the immobilized laccase was higher than free laccase,and the activity of immobilized enzyme treated by freeze-drying was 1.59 folds compared to the non-freeze-dried,but the freeze-dried immobilized laccase had poor operational stability.[Conclusion] The compositions of the material had significant influence on laccase immobilization and this study will provide reference to select materials for immobilization of laccase from G.lucidum LYL 263.