漆酶固定化及其在小型酶反应器中的应用

以尼龙网为载体,戊二醛为交联剂,固定化真菌漆酶。用固定化酶在自行设计的小反应器中降解低浓度造纸废水,试验结果如下：当废水COD的浓度控制在3000 mg/L左右,降解时间为26h,废水的COD去除率达到35%。     

动物细胞在鼓泡式生物反应器中的死亡速率

通过实验测定,证明生物反应器中细胞死亡速率与气体鼓泡速率成正比而与反应器体积成反比。实验发现气泡大小对细胞死亡速率具有两种作用,一种作用在于影响气泡表面积生成速率;另一种作用则在于影响细胞在气泡表面的吸附程度,其最佳直径为5mm左右。血清和Pluronic F68能显著降低细胞死亡速率,当Pluronic F68浓度达到0．1％时,kd趋于零。所有这些实验结果均与前文提出的生物反应器设计模型具有很好的一致性。     

粗毛栓菌产漆酶对考马斯亮蓝的脱色降解

粗毛栓菌粗酶液经聚丙烯酰胺凝胶电泳,在CBBG－250染色后,漆酶带处有一透明圈,经纯化漆酶和CBBG－250溶液直接作用以及菌体的培养结果证实,漆酶对CBBG－250具有一定的脱色降解作用,在漆酶活力达118u/ml时,CBBG－250溶液在595nm波长的光吸收60min内下降了32．4％。该粗毛栓菌产漆酶对工业染料废水也具有一定的降解脱色作用。     

毛云芝菌利用糖蜜酒精废水产漆酶培养基优化

为提高毛云芝菌的漆酶产量,降低生产成本,同时使糖蜜废水中的营养成分得到充分利用,变废为宝,以糖蜜废水作为碳源,对毛云芝菌发酵产漆酶进行研究。在单因素试验基础上,以不同浓度糖蜜废水、尿素含量为自变量,漆酶活力为因变量,采用Box-Behnken设计方法得到最佳的培养基配方为糖蜜废水浓度47%,尿素添加量0.5%,漆酶活力可达1 810.0 U/mL,约为优化前的6倍。说明毛云芝菌利用糖蜜废水作为碳源生产漆酶是可行的,为糖蜜废水资源化利用高效生产有价值产品奠定了理论基础。     

培养条件对毛栓菌漆酶分泌的影响

研究了碳源、氮源、愈创木酚、香兰素及培养条件对漆酶分泌的影响;结果表明,麦草粉作碳源、(NH_4)_2SO_4作氮源有利于漆酶的分泌,适宜浓度的愈创木酚和香兰素等对漆酶的产生有一定的作用;pH在3.0～8.0的范围内对漆酶的分泌影响差别不大,培养温度、接种量、通气量对漆酶的分泌有较大影响。     

毛栓菌(Tramets trogii)液体培养产漆酶的研究

实验研究了碳源、氮源和综合因素对毛栓菌(Tramets trogii)液体培养产漆酶和生长情况的影响。碳源为玉米面、氮源为豆饼粉时,菌株的产漆酶酶活最高;碳源为可溶性淀粉,氮源为麦麸时,对菌株的生长有利;在液体条件下,菌株产漆酶的最佳培养基组成为玉米面3.5%,豆饼粉4.0%,KH2PO40.3%,MgSO40.15%,VB10.04%。     

胶质射脉革菌漆酶的复合诱导及其脱色性能

[目的]研究复合诱导对胶质射脉革菌BBEL0901产漆酶的影响及其粗酶液对染料的脱色性能。[方法]采用分光光度法测定漆酶活性及其染料脱色性能。[结果]甘草渣和Cu2+对BBEL0901产漆酶均有促进作用,于发酵的第6 d向含甘草渣的低氮培养基中添加Cu2+(0.5 mmol/L)对漆酶的协同诱导作用最显著,活性达325.1 U/m L,分别为Cu2+单独诱导、甘草渣单独诱导和未诱导的4倍、14.4倍和19.8倍。粗酶液对不同结构的染料均有较强的脱色能力,对三苯甲烷类染料的脱色效果最好,脱色率达80%以上。[结论]甘草渣与Cu2+对BBEL0901漆酶具有协同诱导作用,酶活最高可达325.1 U/m L,产业化潜力大,所得粗酶液在染料脱色方面具有良好的应用前景。     

白腐菌对培养环境pH的调节及其产漆酶的相关性

研究了不同种属白腐菌多孔菌属C1(Polyporussp.)、侧耳属B2(Pleurotussp.)、香菇属A3(Lentinusedodes)对培养环境pH的调节,及其与菌株产漆酶状况的相关性。结果表明:3株白腐菌均可调节培养液酸化,培养环境的pH可分别由初始的4.5~5.0持续下降至培养结束时3.0~3.5水平,相应C1、B2、A3三者的漆酶合成与分泌在pH值较高(pH 4.5以上)时均表现较弱(酶活分别处于约1 500、200、50 U/mL以下的水平),在pH 3.2~4.5范围内的不同偏酸性条件下则得到较好表达(最高酶活分别达5 000、340、144 U/mL)。统计学分析指出,白腐菌调节环境pH状况与其产漆酶状况之间存在显著或极显著相关性,但不同白腐菌调节培养环境pH的能力之间并不具有显著性差异。     

层孔菌产漆酶的摇瓶最适培养条件研究*

筛选到一株高产漆酶的层孔菌W-1 ,对其液体产漆酶的培养条件进行了优化 ,确定了最适的碳源、氮源 ,并用正交试验确定了培养基中碳源和氮源的浓度。在最优化的培养条件下 ,培养 7d后 ,W-1产生的漆酶活力可以达到 7.1U/mL。采用补料的方法 ,可以得到大量的漆酶粗酶液。     

碳源和氮源对彩绒革盖菌液体发酵合成漆酶的影响

研究了碳源、氮源对彩绒革盖菌液体发酵合成漆酶的影响。结果表明,在所选的几种碳、氮源中,麸皮为试验菌株合成漆酶的较好碳源;酵母浸膏、酒石酸铵、蛋白胨均是比较理想的氮源。不同的碳氮比对彩绒革盖菌漆酶的产量有着显著的影响,高碳高氮是其生产漆酶的最佳条件。在适宜的培养条件下该菌株能合成高活力的漆酶,其酶活力可达298 U/mL以上,产酶周期为6~7 d。     

彩绒革盖菌漆酶产酶条件研究

本文研究了碳源、氮源、愈创木酚、香兰素及培养条件对漆酶分泌的影响;结果表明,淀粉作碳源、干酷素作氮源有利于漆酶的分泌,适宜浓度的愈创木酚和香兰素等对漆酶的产生有一定的作用;pH在3.0－8.0的范围内对漆酶的分泌影响差别不大,培养温度,接种量、通气量对漆酶的分泌有较大影响。     

彩绒革盖菌漆酶产酶条件研究

本文研究了碳源、氮源、愈创木酚、香兰素及培养条件对漆酶分泌的影响;结果表明,淀粉作碳源、干酪素作氮源有利于漆酶的分泌,适宜浓度的愈创木酚和香兰素等对漆酶的产生有一定的作用;pH在3.0～8.0的范围内对漆酶的分泌影响差别不大,培养温度、接种量、通气量对漆酶的分泌有较大影响。     

Laccase activity in lignin degradation by Coriolus versicolor in vivo and in vitro studies

Abstract Laccase activity in ligninolytic cultures of Coriolus versicolor was inhibited by the addition of antibodies. The degree and rate of lignin degradation was unaffected by this inhibition in comparison with cultures which had normal laccase activity. Although experiments in vitro showed that milled wood lignin was depolymerised by laccase in the presence of hydrogen peroxide, and that this reaction was inhibited by antibody, it is concluded that this degradative reaction was not the function of laccase in vivo.     

应用Coriolus vericolor 菌丝球脱色染料及印染废水的研究

对白腐真菌（Coriolus vericolor）产生漆酶进行了研究。发现该菌产漆酶的最适初始pH值为4．5。提高微量元素浓度或添加藜芦醇都可使C.versiclor的产酶能力增加,添加Tween80会有一定的抑制作用。采用C.versicolor菌丝球进行重复分批产酶试验,结果表明菌丝球的稳定性很好,同一批菌丝球可连续利用14次,平均每批酶活力可保持在6．72U／mL,产酶能力优于聚氨酯泡沫固定化菌丝。将粗酶液用要料的脱色降解试验,在酶活力为3．3IU／mL,酸性橙浓度为500mg/L条件下,经过24h反应,脱色率达到98．5％;对含弱酸大红和卡布龙红的印染废水进行脱色试验,脱色率也达到了93％。     

Production of Decolorizing Activity for Molasses Pigment by Coriolus versicolor Ps4a

The distribution of molasses pigment (melanoidin) decolorizing activity (MDA) was investigated in various Basidiomycetes. MDA was only found in some genera of the white-rot-fungi group of which Coriolus versicolor Ps4a showed high activity, a decolorization yield of approximately 80% under the optimal conditions. Production of MDA by C. versicolor was almost completely coincident with the growth of mycelia. The main MDA was due to intracellular enzymes and induced by the molasses pigment. The induced enzyme consisted of two types, namely a sugar dependent enzyme and a sugar independent enzyme. The decolorization by C. versicolor was due to the decomposition of the molasses pigment.     

Relationship between Fungal Biomass Production and the Brightening of Hardwood Kraft Pulp by Coriolus versicolor

The white-rot fungus Coriolus versicolor increased the brightness of hardwood kraft pulp by two mechanisms depending on the concentration of available nitrogen. In low-nitrogen conditions, the brightening process was a chemical effect mediated by the fungus, associated with the removal of residual lignin in the pulp; kappa number was used as an indicator of lignin concentration. A five-day treatment in low-nitrogen conditions increased the brightness of hardwood kraft pulp from 36.2 to 54.5%, with a corresponding decrease in kappa number from 12.0 to 8.5, equivalent to a reduction in the lignin concentration from ca. 2.0% (wt/wt) to ca. 1.4% (wt/wt). Under these conditions, we concluded that the brightening of the pulp was a secondary metabolic event initiated after the depletion of available nitrogen. This method of brightening has been described as bleaching or biobleaching. By contrast, in high-nitrogen conditions, the brightening was a physical effect associated with the dilution of the dark pulp fibers by the relatively high levels of brighter fungal mycelium produced. Since this method of brightening was not evidently associated with lignin removal, it cannot be described as bleaching. In pulp samples brightened in high-nitrogen conditions, as brightness increased, there was a corresponding increase in kappa number. This observation was explained by the consumption of potassium permanganate by the fungal mycelium, which interfered with kappa number determinations at high fungal biomass levels.     

云芝发酵产胃蛋白酶抑制剂的反应机理探讨

目的:对一种从云芝发酵液中提取得到的胃蛋白酶抑制剂的动力学性质和反应机理进行研究.方法:采用Lineweaver-Bunk双倒数法和UltroGelACA-54凝胶过滤法:结果:实验证明抑制剂为胞内产物.其成分是蛋白质,分子量为23 100Da,K值为0.996 μmol/L.结论:抑制剂对胃酶的抑制类型属于非竞争和反竞争混合型抑制模式,它与胃酶结合作用的方式可能是在酶的活性基同形成氢键并封锁酶与底物的结合部位.     

彩绒革盖菌固体发酵生产木质素酶工艺优化研究*

采用均匀设计U15(5^8)和双温度培养法进行彩绒革盖菌固体发酵生产木质素酶,对漆酶、愈创木酚酶、多酚氧化酶的活力进行回归分析。结果表明：应用双温度培养法进行彩绒革盖菌固体发酵生产木质素酶时,在自然补给氧气,培养基pH自然(约6．5),并保持环境湿度约60％的条件下,20d是适宜的发酵周期;玉米浆、麸皮、(NH4)2SO4、水分适宜作为固体培养基的成分;少量的Tween-80有利于木质素酶的生产。     

Coriolus versicolor Laccase Catalyzes the Decarboxylation of 2-(4-Hydroxyphenyl)-glycine and 4-Hydroxymandelic Acid

Cyclodextrin glucanotransferase [1,4-α-D-glucan 4-α-D-(l,4-α-D-glucano)-transferase (cyclizing), EC 2.4.1.19] from an alkalophilic Bacillus species A2–5a had a wider acceptor specificity than that from B. macerans, which was similar to those from B. stearo-thermophilus and B. circulans.

Glucosyl rhamnose produced by the CGTase was identified as glucopyranosyl-α-l,4-rhamnopyranose by α- and β-glucosidase treatments, and ¹H- and ^{1 3}C-NMR analyses.