～（60）Co-γ射线对白腐真菌和黑曲霉的诱变效应以及对木质素降解的研究
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摘要
白腐真菌(White rot fungi)和黑曲霉(Aspergillus Niger)为丝状真菌，白腐真菌属于担子菌门、多孔菌科、层孔菌属、木质拟层孔菌，分泌漆酶(Laccase)、锰过氧化物酶(Mn-peroxidase，Mnp)和木质素过氧化物酶(Lingin-peroxidase，Lip)；黑曲霉属于半知菌门、丝孢菌目、丛梗菌科、黑曲霉属、黑曲霉，是羧甲基纤维素酶(exoglucanase，EC)和β-葡萄糖苷酶(β-glucosidase，βG)和木聚糖酶(xylanase．EC3.2.1.8)的高产菌株，这两个菌株对纤维素和木质素有独特的降解能力。本论文以这两菌株为研究对象，利用~(60)C_0—γ射线(~(60)C_0 gamma rays)，在不同辐照时间、不同辐照剂量条件下对其孢子悬浮液辐射处理，通过测酶活力，与原菌株对照，选出酶活力提高幅度较大的诱变菌株，并对诱变菌株传代，研究酶活力的稳定性及不同碳源、氮源对菌丝干重和产酶量的影响。论文采用蔗渣固体发酵试验研究对照菌株和诱变菌株对甘蔗渣纤维素和木质素的降解效果，以及甘蔗渣的失重和粗蛋白含量变化情况，进一步考察诱变菌株的生长速度、产酶量较对照组是否有所提高。最后研究固体发酵的产酶条件，试验结果表明：
(1) ~(60)C_0—γ射线对白腐真菌和黑曲霉辐射效果明显，在0.6kgy剂量下，白腐真菌致死率为94.6％，黑曲霉致死率为99.49％；1.2kgy剂量下，白腐真菌存活率为0.0045％，黑曲霉存活率为0.0026％；但对黑曲霉的筛选及酶活测定结果表明其正突变率为0。从筛选到的6株白腐真菌正突变菌中，漆酶活力提高幅度最大一株达到115.28％，锰过氧化物酶活力提高108.09％，
张变英60co--丫射线对白腐真菌和黑曲霉的诱变效应以及对木质素降解的研究2
经四次传代稳定性较好。
(2)诱变菌株和原菌株对碳源及氮源的要求上不存在差异。
(3)白腐真菌诱变菌株和原菌株发酵甘蔗渣，经30天培养后，原菌株
NDF降解率为1 6.87%，K 1 ason木素降解率为26.63%;诱变株NDF降解率为
28.01%，K!ason木素降解率40.75%;经20天培养后，原菌株NO「降解率为
1 2.60%，K!ason木素降解率为2.99%;诱变株NDF降解率为1 0.89%，K 1 ason
木素降解率10.69%o
(4)固体发酵的最佳氮源为蛋白膝，其次为硫酸铁，最佳含水量为44%，
氮源最佳浓度为3%。
White-rot fungi (5.132 coriolus versicolor fome) and Aspergillus Niger (3.316) are all filar fungus, white rot fungi (5.132) excretes laccase and Mn-peroxidase(Mnp); Aspergillus niger(3.316) excretes endoglucanase (EG), xylanase(EC) and B-glucosidase ( BG), Two species have high degradability to cellulose ,hemi-cellulose and lignin.The degradability of White-rot fungi (5.132) and Aspergillus Niger (3.316)were investigated to bagasse by solid-state fermentation for thirty days, the result shown :
(1) The degradability of white-rot fungi to neutral detergent fibre (NDF) is 16.87% and to Klason lignin is 26.63%.
(2) After Aspergillus Niger (3.316) degraded, neutral detergent fibre (NDF) and Klason lignin is higher than before.
In this study , Sporule suspension of white-rot fungi (5.132) and Aspergillus Niger (3.316) are radiated with 60Co gamma rays under the different radiate dose and different radiate time. Enzyme vitalities of laccase and Mn-peroxidase, endoglucanase (EG) and B-glucosidase (BG)were mensurated. compared with original strains ,their Enzyme vitalities are higher than original strains. 60Co gamma rays have physiological and biochemical effect on fungus sporules which is radiated. Enzyme vitality enhancing attributed two effects or single effect, biochemical effect include gene mutation .By
several multiply and repeated isolated, getting six high produced strains of white rot fungi, study shown, six isolated strains of white rot fungi have higher enzyme vitality and mycelial weightiness, and that enzyme vitality have good stabilization, mutation strains have no difference in the demand of carbon source and nitrogen source from original strains. The research of solid-state fermentation of bagasse indicated, six high produced strains of white-rot fungi have higher degradability to cellulose and lignin than original strains ,the difference is evident. The result shown :
(1) The effect of 60Co gamma rays on white-rot fungi and Aspergillus Niger is evident. Under the 0.6kgy dose, white-rot fungi deadly rate is 94.6%, Aspergillus Niger deadly rate is 99.49%; under 1.2kgy dose, livability rate of white-rot fungi is 0.0045%, livability rate of Aspergillus Niger is 0.0026%.Result of isolation for positive mutation of Aspergillus Niger is zero.
(2) white-rot fungi laccase vitality enhance 157.7%, Mnp vitality enhance 137.24%. mutiply four times, laccase and Mn-peroxidase vitality have good stabilization.
(3) After solid state fermentation, original strains degradability to bagasse NDF is 16.87%, to Klason lignin is 26.63%, mutation strains degradability of NDF to bagasse is 28.01%, to Klason lignin is 40.75%.
(1) ~(60)C_0—γ射线对白腐真菌和黑曲霉辐射效果明显，在0.6kgy剂量下，白腐真菌致死率为94.6％，黑曲霉致死率为99.49％；1.2kgy剂量下，白腐真菌存活率为0.0045％，黑曲霉存活率为0.0026％；但对黑曲霉的筛选及酶活测定结果表明其正突变率为0。从筛选到的6株白腐真菌正突变菌中，漆酶活力提高幅度最大一株达到115.28％，锰过氧化物酶活力提高108.09％，
张变英60co--丫射线对白腐真菌和黑曲霉的诱变效应以及对木质素降解的研究2
经四次传代稳定性较好。
(2)诱变菌株和原菌株对碳源及氮源的要求上不存在差异。
(3)白腐真菌诱变菌株和原菌株发酵甘蔗渣，经30天培养后，原菌株
NDF降解率为1 6.87%，K 1 ason木素降解率为26.63%;诱变株NDF降解率为
28.01%，K!ason木素降解率40.75%;经20天培养后，原菌株NO「降解率为
1 2.60%，K!ason木素降解率为2.99%;诱变株NDF降解率为1 0.89%，K 1 ason
木素降解率10.69%o
(4)固体发酵的最佳氮源为蛋白膝，其次为硫酸铁，最佳含水量为44%，
氮源最佳浓度为3%。
White-rot fungi (5.132 coriolus versicolor fome) and Aspergillus Niger (3.316) are all filar fungus, white rot fungi (5.132) excretes laccase and Mn-peroxidase(Mnp); Aspergillus niger(3.316) excretes endoglucanase (EG), xylanase(EC) and B-glucosidase ( BG), Two species have high degradability to cellulose ,hemi-cellulose and lignin.The degradability of White-rot fungi (5.132) and Aspergillus Niger (3.316)were investigated to bagasse by solid-state fermentation for thirty days, the result shown :
(1) The degradability of white-rot fungi to neutral detergent fibre (NDF) is 16.87% and to Klason lignin is 26.63%.
(2) After Aspergillus Niger (3.316) degraded, neutral detergent fibre (NDF) and Klason lignin is higher than before.
In this study , Sporule suspension of white-rot fungi (5.132) and Aspergillus Niger (3.316) are radiated with 60Co gamma rays under the different radiate dose and different radiate time. Enzyme vitalities of laccase and Mn-peroxidase, endoglucanase (EG) and B-glucosidase (BG)were mensurated. compared with original strains ,their Enzyme vitalities are higher than original strains. 60Co gamma rays have physiological and biochemical effect on fungus sporules which is radiated. Enzyme vitality enhancing attributed two effects or single effect, biochemical effect include gene mutation .By
several multiply and repeated isolated, getting six high produced strains of white rot fungi, study shown, six isolated strains of white rot fungi have higher enzyme vitality and mycelial weightiness, and that enzyme vitality have good stabilization, mutation strains have no difference in the demand of carbon source and nitrogen source from original strains. The research of solid-state fermentation of bagasse indicated, six high produced strains of white-rot fungi have higher degradability to cellulose and lignin than original strains ,the difference is evident. The result shown :
(1) The effect of 60Co gamma rays on white-rot fungi and Aspergillus Niger is evident. Under the 0.6kgy dose, white-rot fungi deadly rate is 94.6%, Aspergillus Niger deadly rate is 99.49%; under 1.2kgy dose, livability rate of white-rot fungi is 0.0045%, livability rate of Aspergillus Niger is 0.0026%.Result of isolation for positive mutation of Aspergillus Niger is zero.
(2) white-rot fungi laccase vitality enhance 157.7%, Mnp vitality enhance 137.24%. mutiply four times, laccase and Mn-peroxidase vitality have good stabilization.
(3) After solid state fermentation, original strains degradability to bagasse NDF is 16.87%, to Klason lignin is 26.63%, mutation strains degradability of NDF to bagasse is 28.01%, to Klason lignin is 40.75%.
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