阿魏菇与胶红酵母共培养产漆酶对活性艳蓝W-RV的脱色

利用阿魏菇与胶红酵母共培养所产漆酶对染料活性艳蓝W-RV进行脱色,同时考察不同p H、温度、染料浓度和漆酶酶活等条件对脱色的影响。结果显示,酸性范围内的p H有利于活性艳蓝W-RV的脱色,较高温度并不适于脱色反应。在研究不同染料浓度的影响时发现,不同浓度染料的脱色率在7 h反应后都达到了稳定,而漆酶酶活在达到200 U/L后脱色率不再变化。因此,最终得到的最适反应条件为p H 4.5、30℃、染料质量浓度100 mg/L、漆酶酶活200 U/L,在此条件下,活性艳蓝W-RV的最高脱色率为89.91%。     

红平菇产漆酶条件优化及对孔雀绿染料脱色的研究

采用LNAS(低氮天冬酰胺-琥珀酸)培养基添加方式,对红平菇Pleurotus djamor HP1进行培养,检测不同时间培养液对不同底物的氧化作用,进而得到光密度值的变化情况,作为漆酶的产生及活性测定的主要依据。结果表明:在含Cu2+的培养液中漆酶最大酶活为235.4 U/L。含Cu2+的培养液添加底物木屑后漆酶最大酶活为458.8 U/L。提取经优化筛选后的培养基培养出的漆酶粗酶液,对4种具有不同化学结构的染料进行了脱色试验。结果表明:三苯基甲烷类的孔雀绿在6 h时脱色率为87.5%,蒽醌类的SN4R在24 h时脱色率为49.4%,偶氮类的甲基橙在24 h时脱色率为45%,杂环类的中性红在24 h时脱色率为23.6%。因此,显示出红平菇漆酶对孔雀绿染料脱色具有较大的应用潜力,进而对废水处理具有更好的应用前景。     

灵芝漆酶催化阳离子红2GL脱色的研究

真菌漆酶在纺织物染料脱色及其废水净化等领域有着巨大的应用潜力。阳离子红2GL是使用广泛又难以处理的一种染料,现有的方法治理效果差。本研究优化了灵芝漆酶催化阳离子红2GL脱色的主要工艺参数:最适pH、温度、ABTS用量、漆酶用量和染料浓度分别为4.5、20℃、0.083mmol/L、10U/mL和50mg/L。在所得的最优条件下反应30min,阳离子红2GL的脱色率可达90.3%;反应24h,脱色率达100.0%。     

短小芽孢杆菌LC01漆酶基因在毕赤酵母中的胞外表达及重组漆酶酶学性质研究

为了获得表达量高、稳定性好及染料脱色效率高的细菌漆酶,通过PCR扩增出短小芽孢杆菌LC01的漆酶基因并构建重组表达载体pPICZαA-lac,转化毕赤酵母菌株SMD1168H后利用甲醇诱导培养重组菌获得重组漆酶,纯化并分析了重组漆酶的性质。重组菌株产漆酶活性在第7天达到最高,为1 390 U/L。纯化的重组漆酶分子量为65 kD,以丁香醛连氮为底物的最适反应温度和pH分别为70℃和6.8。在pH 9.0放置10 d活性没有下降,在70℃保温10 h后仍保留36%的酶活。Al~(3+)、Fe~(3+)和Mn~(2+)完全抑制漆酶活性。在介体乙酰丁香酮参与下该漆酶能够有效脱色RB亮蓝、活性黑5和靛红,在pH 9.0时6 h的脱色率达到了90%以上,表明该重组漆酶能有效应用于染料废水的脱色处理。     

白囊耙齿菌对茜素红脱色条件优化及其毒性变化检测

白囊耙齿菌Irpex lacteus是分离自倒木上的一株可以分泌漆酶和锰过氧化物酶的白腐真菌。利用I. lacteus对固体条件下的活性黑、活性红、结晶紫、茜素红和孔雀石绿进行脱色能力的检测,通过单因素和正交试验优化I. lacteus对茜素红的脱色条件,并以3种作物发芽率为指标测定茜素红被I. lacteus脱色前后的毒性变化。结果显示,I. lacteus对5种染料均可脱色,其中对茜素红染料的脱色更为彻底;单因素和正交试验优化I. lacteus对茜素红的脱色条件为：pH 7.0、葡萄糖10.0g/L、硫酸铵0.66g/L、接种量2片（Φ=8.0mm）、100.0mL三角瓶装液20.0mL,优化条件下I. lacteus对茜素红脱色10d时的脱色率为88.26%,与未优化前的脱色率相比提高了60.50%;茜素红染料被I. lacteus脱色前后毒性大小排序为：染料原液>染料脱色后>PDB培养基处理,表明茜素红染料存在一定的毒性,I. lacteus脱色茜素红后可以使其毒性减弱。通过本研究,为I. lacteus在茜素红等染料废水脱色以及降低染料废水毒性方面的应用奠定基础。     

灵芝漆酶催化直接耐晒翠蓝GL脱色条件的优化

采用灵芝菌株发酵所得的漆酶, 对酞菁类染料直接耐晒翠蓝GL进行了催化脱色实验, 确定了脱色反应的最适条件。结果表明: 单独使用灵芝漆酶粗酶液对直接耐晒翠蓝GL具有很好的脱色效果。其最适脱色pH为3.0, 最适脱色温度为40°C, 最适漆酶用量是40 U/mL, 最适染料浓度为60 mg/L。以上述最适脱色条件对直接耐晒翠蓝GL进行脱色实验, 反应70 min, 脱色率可达94.3%。研究结果显示, 所试灵芝漆酶在印染废水治理方面具有良好的应用前景。     

刺芹侧耳产漆酶条件优化及对偶氮染料甲基橙的脱色

漆酶是一种含铜的单电子多酚氧化酶,能够催化氧化各种酚类及多种染料,在处理染料废水方面具有巨大的潜力。刺芹侧耳Pleurotus eryngii具有较强的产漆酶能力,但漆酶产量在较大程度上受环境条件限制。本文研究了氮源含量、pH、温度、金属离子等环境条件对刺芹侧耳产漆酶能力的影响,优化了其产漆酶条件,并用其粗酶液对典型偶氮类染料甲基橙进行脱色,结果表明,在氮源0.5%（W/W）、pH 5.5、温度28℃、添加5.0mmol/L Mg ²⁺的培养条件下,刺芹侧耳产漆酶能力最强,培养6d时,漆酶酶活可达78.0U/L。用优化培养的刺芹侧耳粗酶液对偶氮染料甲基橙进行脱色,28h后脱色率可达90%,脱色反应为准一级动力学反应,甲基橙并未完全矿化,而是生成小分子中间产物。     

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。     

烟管孔菌G11对活性染料的脱色

本研究从未成熟的有机蓝莓表皮分离、纯化得到一株白腐真菌G11,通过对菌株G11的形态特征、ITS序列同源性比对以及系统发育分析,鉴定菌株G11为一株烟管孔菌Bjerkandera adusta。菌株G11可以产生木质素过氧化物酶、漆酶和锰过氧化物酶等木质素降解酶。菌株G11对8种不同染料的脱色效果显示其对活性染料的脱色效果最好,脱色率达到90%所需时间最短。以菌株G11为研究对象,研究其对不同浓度的活性黑和活性红的脱色能力,结果表明：菌株G11对活性红和活性黑具有显著的脱色能力。在脱色15d时,菌株G11对浓度为10、50、100、250、500mg/L活性红的脱色率分别为99%、98%、95%、94%和92%;对浓度为10、50、100、250、500mg/L活性黑的脱色率分别为98%、97%、95%、93%和90%。     

重组海洋细菌漆酶Lac15脱色人工合成纺织染料的潜力

以重组海洋细菌漆酶Lac15(rLac15)为生物催化剂,对蒽醌类和偶氮类染料进行脱色,考察了rLac15对人工合成纺织染料的脱色潜能。通过研究催化的介体、酶量、pH、染料浓度以及温度对脱色效率的影响,进一步优化了rLac15对部分蒽醌类和偶氮类人工合成纺织染料的脱色条件。以丁香酸甲酯为介体,在pH 8.5、45℃条件下反应1 h,20 U/L rLac15对100μmol/L偶氮染料类Acid Red 6B(AR-6B),Reactive Blue 194(M-2GE),Reactive Brilliant Orange(K-7R)和Reactive Blue 171(KE-R)具有较好的脱色效果,脱色率分别达到95%、93%、76%和61%。随着染料浓度的增加,脱色率呈下降趋势,但当染料浓度达到200μmol/L时,M-2GE和AR-6B仍可保持80%以上脱色率。在常温下,rLac15对AR-6B、M-2GE、K-7R和KE-R显示较高脱色率,25℃反应24 h,分别达到96%、86%、66%和66%。rLac15是具有常温以及偏碱性环境脱色能力的细菌漆酶,具有潜在工业应用价值。     

Production of Laccase by Trametes hirsuta Grown in an Immersion Bioreactor and its Application in the Docolorization of Dyes from a Leather Factory

An alternative system for producing laccase on a bioreactor scale by the white‐rot fungus Trametes hirsuta is proposed. The experiments were performed in an immersion bioreactor (employing cuttings of stainless steel sponges as a support) and the culture medium was supplemented with copper sulfate (1 mM). Operating under these conditions, it was possible to obtain a maximum laccase activity of nearly 5,000 U/L within 9 days. In addition, the ability of the crude laccase produced to decolorize two synthetic acid dyes utilized in the leather industry (Luganil Green and Sella Solid Red) was investigated. The effect of the pH and the enzyme activity on decolorization was analyzed. It was found that a pH of 4.0 and a laccase activity of 300 U/L were optimal for Luganil dye decolorization (16.2 % in 2 hours). Sella Solid Red showed its highest decolorization (around 40 % in 2 hours) when used at pH 5.0 and at a laccase activity of 1,000 U/L.     

毛栓菌产漆酶条件优化及该酶对合成染料脱色的特性

【目的】提高菌株Trametes hirsuta SYBC-L19漆酶产量,并研究该酶对合成染料脱色的性质。【方法】通过单因素和响应面设计,对产漆酶培养基进行优化。【结果】最优培养基为:玉米粉20.0 g/L、马铃薯淀粉32.4 g/L、酒石酸铵2.9 g/L、吐温80 0.5 g/L、CuSO4.5H2O 2.0 mmol/L、香兰素0.54 mmol/L、NaH2PO4.2H2O 2.0 g/L、MgSO4.7H2O0.5 g/L、MnSO4.H2O 0.1 g/L;最佳培养条件为:培养温度30°C,初始pH 6.0,装液量40 mL/250 mL,接种量8%。【结论】培养8 d酶活达35 U/mL,是优化前的39倍。对漆酶催化合成染料脱色进行了考察,发现该酶在60°C下对偶氮类染料AR1和RB5能迅速脱色,5 min内即可完成。     

云芝Trametes versicolor1126所产漆酶对靛蓝废水脱色的初步研究

考察了云芝Trametes versicolor 1126发酵培养中漆酶酶活和pH的变化,同时研究了羧甲基纤维素钠及苯酚添加量对漆酶活力的影响。结果表明当培养基中加入0.8%羧甲基纤维素钠、100mg/L苯酚时,均能明显提高漆酶的活力。以漆酶/HBT介质体系对靛蓝废水进行脱色,反应100min后,脱色率达90.8%。使用漆酶处理靛蓝废水具有广阔的应用前景     

云芝Trametes versicolor1126所产漆酶对靛蓝废水脱色的初步研究

考察了云芝Trametes versicolor1126发酵培养中漆酶酶活和pH的变化,同时研究了羧甲基纤维素钠及苯酚添加量对漆酶活力的影响。结果表明当培养基中加入0.8%羧甲基纤维素钠、100mg/L苯酚时,均能明显提高漆酶的活力。以漆酶/HBT介质体系对靛蓝废水进行脱色,反应100min后,脱色率达90.8%。使用漆酶处理靛蓝废水具有广阔的应用前景。     

The use of a radial basis neural network and genetic algorithm for improving the efficiency of laccase-mediated dye decolourization

A radial basis function neural network (RBF) and genetic algorithm (GA) were applied to improve the efficiency of the oxidative decolourization of the recalcitrant dye Reactive Black 5 (RB 5) by a technical laccase (Trametes spp.) and the natural mediator acetosyringone (ACS). The decolourization of RB 5 in aqueous solution was studied with a 3(4) factorial design including different levels of laccase (2, 100, 200UL(-1)), acetosyringone (5, 50, 100μM), pH value (3, 4.5, 6) and incubation time (10, 20, 30min). The generated RBF network was mathematically evaluated by several statistical indices and revealed better results than a classical quadratic response surface (RS) model. The experimental data showed that within 10min of incubation time a complete decolourization (>90%) was achieved by using the highest amount of laccase (200UL(-1)) and acetosyringone (100μM) at pH 6. By applying the RBF-GA methodology, the efficiency of the laccase-mediated decolourization was improved by minimising the required amount of laccase and acetosyringone by 25% and 21.7% respectively. Complete decolourization (>90%) was obtained within 10min at the GA-optimised process conditions of laccase (150UL(-1)) and acetosyringone (78.3μM) at pH 5.67. These results illustrate that the RBF-GA methodology could be a powerful technique during scale-up studies.     

Contribution of manganese peroxidase and laccase to dye decoloration by Trametes versicolor

During dye decoloration by Trametes versicolor ATCC 20869 in modified Kirk’s medium, manganese peroxidase (MnP) and laccase were produced, but not lignin peroxidase, cellobiose dehydrogenase or manganese-independent peroxidase. Purified MnP decolorized azo dyes [amaranth, reactive black 5 (RB5) and Cibacron brilliant yellow] in Mn²⁺-dependent reactions but did not decolorize an anthraquinone dye [Remazol brilliant blue R (RBBR)]. However, the purified laccase decolorized RBBR five to ten times faster than the azo dyes and the addition of a redox mediator, 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid), did not alter decoloration rates. Amaranth and RB5 were decolorized the most rapidly by MnP since they have a hydroxyl group in an ortho position and a sulfonate group in the meta position relative to the azo bond. During a typical batch decoloration with the fungal culture, the ratio of laccase:MnP was 10:1 to 20:1 (based on enzyme activity) and increased to greater than 30:1 after decoloration was complete. Since MnP decolorized amaranth about 30 times more rapidly than laccase per unit of enzyme activity, MnP should have contributed more to decoloration than laccase in batch cultures.     

Extracellular ligninolytic enzymes by Lentinus polychrous Lév. under solid-state fermentation of potential agro-industrial wastes and their effectiveness in decolorization of synthetic dyes

Six agro-industrial wastes were evaluated as a support for ligninolytic enzyme production by the white-rot fungus Lentinus polychrous Lév. under solid-state fermentation. Enzyme production was markedly different according to the substrate used. Rice bran (RB) yielded the highest laccase activity of 1,449 U/L (after 21 days of culture) with specific activity of 4.4 U/g substrate. Rice bran supplemented with rice husk (RH) (2:1 by wt) showed high laccase activity of 1,425 U/L with specific activity of 10.0 U/g substrate (after 17 days of culture). The crude enzyme of the RH-RB culture also contained manganese peroxidase (MnP) and manganese-independent peroxidase (MIP) activities in relative proportions of 1.9:1.4:1 of laccase:MnP:MIP, respectively. Zymogram studies showed the same isoenzyme pattern with these ligninolytic enzymes. The high enzyme production level and low substrate cost of SSF-L. polychrous Lév. suggest that it has potential for industrial applications. Our studies showed that the crude enzyme from this culture exhibited in vitro decolorization of Indigo Carmine. The highest efficiency of dye decolorization was observed under alkaline conditions (pH 9.0) at an initial dye concentration of 10 mg/L. The rather high pH conditions and high efficiency in Indigo Carmine decolorization make the enzyme further interest for the applications in treatment of waste water from the textile industry, which contains synthetic dyes.     

Overlap of laccases/cellobiose dehydrogenase activities during the decolourisation of anthraquinonic dyes with close chemical structures by Pycnoporus strains

Pycnoporus strains were used as model to understand the role of laccases in the in vivo decolourisation of three anthraquinonic dyes. The decolourisation capability of Pycnoporus sanguineus MUCL 41582 (PS7), which produces laccases as the main oxidative enzyme, was assayed and compared with the decolourisation capability of a control strain, Pycnoporus cinnabarinus MUCL 39533 (PC330) described as laccase-deficient strain. In absence of dye, laccase activity was observed during the trophophase and the idiophase with PS7, while no laccase activity was observed with PC330. Acid Blue 62 (ABu62), Acid Blue 281 (ABu281) and Reactive Blue 19 (RBu19) caused an increase in laccase activity and surprisingly laccase activity was detected with PC330. In vitro, oxidation of all three anthraquinones by a laccase preparation was obtained to a lesser extent than the whole cell process; suggesting that other factor(s) could be required for a complete decolourisation. As the time space of laccase production in the tested fungi was not perfectly coincidental with the decolourisation process, the activity of cellobiose dehydrogenase (CDH) was monitored. Present early in the broth during the growth of the fungi, CDH displayed in vitro a synergism with laccases in the decolourisation of ABu62, and an antagonism with laccases in the decolourisation of ABu281 and RBu19.