导　　师： 冯贵颖;廖银章;刘晓风

学科专业： 083002

授予学位： 硕士

作　　者： ;

机构地区： 西北农林科技大学

摘　　要： 本文以探索高效的木质素降解为宗旨,对木质素降解菌株的分离、筛选、产酶特性及对天然木质素的降解进行了较为系统的研究和分析,结果如下: 1.从本实验室选育的12株菌株中筛选得到一株高效木质素降解菌,编号为m-6。通过液体产酶培养,发现该菌株主要分泌胞外漆酶,漆酶的最高活力达到127.78 U//L;对碱溶木质素的降解可达到61.25 /%;固态产酶培养后,漆酶的最高活力达到143.06U//L,第60天时,稻草粉的失重可达72.13 /%。该菌产酶最适pH为4,最适温度32℃。 2.研究W//O十六烷基三甲基溴化铵/(CTAB/)微乳液体系和HAc-NaAc缓冲液体系中, m-6菌株固态发酵后产生的胞外木质素降解酶漆酶/(Lac/)、木质素过氧化物酶/(Lip/)、锰过氧化物酶/(Mnp/)酶促反应的最佳反应条件。通过对Lip/(以2, 2’-连氮-二/(3-乙苯基并噻唑-磺酸/)为底物/)、Mnp/(以MnSO/_4为底物/)和Lac/(以藜芦醇为底物/) 3种木质素降解酶在W//O型CTAB微乳液体系和HAc-NaAc缓冲液体系中的酶活分别进行测定,以研究不同温度,pH和底物浓度对酶促反应进行的影响,找到最佳反应条件,并比较两种反应体系下的酶活。在W//O型CTAB微乳液体系中,Lip、Mnp和Lac 3种酶的酶促反应的最佳反应条件为:温度37℃,pH分别为4.5,4.5和3.5;在HAc-NaAc缓冲液体系中,3种酶的酶促反应的最佳反应条件为:温度37℃,pH分别为5.0,5.0和3.0。在两种反应体系中最佳反应底物的浓度相同,即藜芦醇、MnSO/_4、2, 2’-连氮-二/(3-乙苯基并噻唑-磺酸/)浓度分别为0.053,0.116,0.492 mmol//L。在W//O型CTAB微乳液中Lip和Mnp酶活比其在HAc-NaAc缓冲液体系中分别提高了81.45 /%和36.75/%,但Lac酶活却减少了2.914倍。3种木质素降解酶在W//O型CTAB微乳液体系和HAc-NaAc缓冲液体系的最佳反应条件有所差别,并且W//O型CTAB微乳液体系有利于Lip和Mnp酶促反应的进行,但该体系不利于Lac酶促反应的进行。 1. An efficient lignin-degrading fungus designated as m-6 was screened from 12 strains of fungus preserved in our laboratory. In liquid culture medium, m-6 mainly produced extracellular laccase, the peak value of laccase activity was reached to 127.78U//L; its degradation rate of alkaline soluble lignin was 61.25/%. After solid substrate fermentation, the peak value of laccase activity was reached to 143.06U//L, on the 60th day, the weight-loss rate of rice straw was achieved to 72.13/%. The optimum ligninase producing conditions of m-6 were pH 4 and 32℃. 2. The reaction conditions of ligninolytic enzyme system, which contain lignin peroxidase /(Lip/), Manganese /(Mnp/) and Laccase /(Lac/), were investigated in W//O microemulsion of centyltrimethy ammonium bromide /(CTAB/) /(W//O CTAB ME/) and HAc-NaAc buffer solution /(HAc-NaAc BS/). The enzymes were excreted from white-rot fungi m-6 which cultured from solid fermentation. Through measured the Lip /(3,4-Dimethoxybenzyl alcohol as its substrate/), Mnp /(MnSO/_4 as its substrate/) and Lac /(2,2'-azino- acid/(ABTS/) as its substrate/) activities in W//O CTAB ME and HAc-NaAc BS, the factors which affect ligninolytic enzyme activity could be found to study the optimum reactive conditions of ligninolytic enzyme system, and the enzymes’activities in these two different systems were compared. In W//O CTAB ME, the optimal temperature was 37℃and the optimal pH are 4.5, 4.5 and 3.5 for Lip, Mnp and Lac respectively. In HAc-NaAc BS, the optimal temperature was also 37℃and the optimal pH was 5.0, 5.0 and 3.0 for Lip, Mnp and Lac respectively; The optimal substrate concentration was 0.053,0.116 and 0.492 mmol//L for Lip, Mnp and Lac respectively.The reasults showed that the Lip and Mnp activitives are improved 81.45 /% and 36.75 /%, while the Lac activity decreased 2.914 times. There were great difference about ligninolytic enzyme activity of m-6 in W//O CTAB ME and In HAc-NaAc BS. The Lip and Mnp activity could be improved in W//O CTAB ME, while the W//O sy

关 键 词： 筛选 木质素降解 型 微乳液 酶活 组合菌

领　　域： [环境科学与工程]