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摘要：作为一种潜在的可再生资源，纤文素和木质素具有很大的利用价值。但是因为纤文素经常会与半纤文素以及木质素等一些较难分解物质结合在一起，因此在利用秸秆等含纤文素丰富的有机质之前，就要将其与木质素和半纤文素分开，从而使纤文素暴露出来，以便更好地与降解酶接触；相对于木质素而言，半纤文素降解较为容易，因此秸秆中纤文素利用的核心就是木质素降解问题，通过微生物来完成木质素的去处是具有潜在价值的热点之一。本实验目的就是筛选木质素酶产生菌株。从学校腐木以及柳树、松树、葡萄树等树皮中分离纯化出能够降解秸秆中木质素的真菌，并测定漆酶和木质素过氧化物酶的活性及菌种进行鉴定，从而筛选出高产漆酶和木质素过氧化物酶的真菌。为筛选能高效降解木质素的菌株，从11份采集的朽木与树皮样品中分离纯化获得36株菌，将纯化获得的菌株分别接入到PDA-愈创木酚和PDA-苯胺蓝培养基上进行初筛，观察其颜色变化，根据变色圈的大小和深浅筛选漆酶产生菌，根据褪色圈的大小筛选木质素过氧化物酶产生菌，共筛选出11株产漆酶或木质素过氧化物酶的菌株，将这11株菌接入液体产酶培养基中进行液体静止培养并测定木质素酶活力，最终获得1株高产木质素酶的真菌菌株L7。其所产木质素过氧化物酶活力达到207.1U/L；在液体培养基中，漆酶测定值较小，受误差影响较大，特采用固体培养基测漆酶，将板子上显色明显的菌株接种到固体培养基上，测定漆酶活性，最终筛选一株高产漆酶菌株1-1，其漆酶活力达到1033.1U/g。34365
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毕业论文关键字：木质素; 漆酶; 木质素过氧化物酶; 愈创木酚; 苯胺蓝
Separation Purification and Identification of Lignin Enzyme Producing Fungi
Abstract: As a potentially renewable resource, cellulose and lignin have great value. But the cellulose is easy to combine with hemicellulose, lignin and other matters which are hard to break down, so it must be released from lignocellulose and hemicellulose in the parcel before cellulose was used. And hemicellulose can be degraded easily, therefore, the key of cellulose degradation is the degradation of lignin, so the use of microbial degradation of lignin and cellulose has been one of the attention. The objective of this experiment is screening strains which can produce lignin enzyme. In order to separate and purify lignin-degradation fungi, fungi were screened from the rotten wood and the barks of willow, pine and grape trees. The activity of laccase and lignin peroxidase were determined to screen strains which can produce high yield laccase and lignin peroxidase. 36 strains was purified from the rotten wood and barks, with the purpose of screening high efficient lignin-degradation fungi, these purified strains were inoculated in the PDA-guaiacol plate and PDA-aniline blue plate to observe the color change, screening laccase producing fungi according to the size and depth of color ring and screening lignin peroxidase producing fungi according to the size of fade circle. 11 strains were screened and then inoculated in the liquid culture medium with static culture to determine the lignin enzyme activity, results showed that L7 was the most excellent strain with the higher lignin-degrading enzymes activities for its lignin peroxidase were up to 207.1U/L. In the liquid culture medium, the activity of laccase is smaller, and the numerical value is influenced bigger by error, so using the solid medium measure laccase activity, results showed that 1-1 was the most excellent strain with the higher laccase enzymes activities for its laccase activity were up to 1033.1U/g.
Key words: lignin; laccase; lignin peroxidase; guaiacol; aniline blue
目录
引言 1
1实验材料与方法 3
1.1样本来源 3