固定化漆酶反应器的构建及应用  

Construction and application of immobilized laccase reactor

作　　者：林沚葳 叶秀云[1] 林娟[1] 杨捷[1] LIN Zhiwei;YE Xiuyun;LIN Juan;YANG Jie(Fujian Key Laboratory of Marine Enzyme Engineering,Fuzhou University,Fuzhou 350116,China)

机构地区：[1]福州大学福建省海洋酶工程重点实验室,福州350116

出　　处：《应用与环境生物学报》2021年第5期1297-1303,共7页Chinese Journal of Applied and Environmental Biology

基　　金：国家自然科学基金项目(31671795)资助。

摘　　要：漆酶具备较强的工业应用潜力,但经济性和稳定性的不足限制了其推广应用,酶的固定化被认为是提高漆酶稳定性的一种有效方法.以来自Cerrena sp.HYB07菌株的漆酶作为研究对象,以壳聚糖为载体制备固定化漆酶,构建实验室规模的酶反应器并应用于染料脱色,探讨固定化漆酶的应用效果.结果显示,以愈创木酚为底物,优化固定化漆酶的制备条件,所得固定化漆酶的最高酶活力为43.5 U/g.固定化后漆酶的热稳定性增强,65℃条件下固定化酶半衰期是游离酶的5.3倍.将固定化漆酶填充于酶反应器中处理remazol brilliant blue reactive(RBBR)和孔雀石绿,酶反应器由进样池、反应池和出样池构成,设置为批次和循环两种模式.批次模式下,重复使用10次,固定化漆酶仍可保留60%以上的酶活力及脱色率.与批次模式相比,循环模式下固定化漆酶对染料的脱色效率更高,固定化酶表现出了比游离酶更好的操作稳定性,可实现对孔雀石绿的完全脱色.经漆酶处理后,孔雀石绿的微生物毒性、植物毒性和细胞毒性都显著降低.本研究表明固定化可以提高漆酶的稳定性和使用效率,该结果可为漆酶在染料脱色脱毒方面的应用提供理论依据.Laccases are valuable for a variety of industrial applications.Nonetheless,large-scale applications of laccases are limited by the economy and efficiency of the enzymes.Enzyme stability can be improved by immobilization.In the present study,the laccase from Cerrena sp.HYB07 was covalently immobilized on chitosan,and continuous decolorization of dyes was achieved with a lab-scale bioreactor,with guaiacol as the substrate;the immobilized enzyme activity was 43.5 U/g after optimization of immobilization conditions.Immobilized laccase displayed higher thermostability,with a half-life at 65℃,5.3 times that of the free enzyme.The reactor was composed of an injection cell,reaction vessel,and collector,set to batch and cycle modes.The immobilized enzyme was used in an enzyme reactor to decolorize Remazol Brilliant Blue Reactive(RBBR)and malachite green(MG).After ten decolorization cycles,the activity of immobilized laccase and the decolorization rates of the dyes remained at 60%.Compared with the batch mode,the decolorization efficiency of the immobilized enzyme was higher in the circulating mode.Decolorization of MG by the immobilized enzyme was more thorough than that of the free enzyme.After laccase treatment,the microbial toxicity,phytotoxicity,and cytotoxicity of MG were significantly reduced.The stability and decolorization efficiency of laccase improved after immobilization.Furthermore,immobilized laccase has better operational stability in a bioreactor.The results of this study can prove the application value of laccase in dye decolorization and detoxification.

关 键 词：漆酶 壳聚糖 固定化 酶反应器 孔雀石绿 

分 类 号：Q814.2[生物学—生物工程]