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〖JP2〗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.versicolor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。将粗酶液用于染料的脱色降解试验,在酶活力为3.3IU/mL〖JP〗,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%。     

三相鼓泡塔生物反应器培养云芝菌合成漆酶

为了提高云芝菌发酵生产漆酶的效率,提出了一种利用自絮凝菌丝球在三相鼓泡塔生物反应器中重复分批发酵产漆酶的新方法。在优化后的产酶条件下,考察维生素C的添加浓度对漆酶活力的影响,并通过在培养基中添加维生素C进行漆酶多批次培养。研究结果表明,维生素C的添加浓度为1.50mmol/L时,可使漆酶活力提高2.6倍。连续进行了10批培养,每批最大漆酶的活力均在1000 U/mL以上,最高酶活出现在第五批为1919.6 U/mL,总培养时间达25 d。此方法所得漆酶对染料靛蓝具有明显的脱色降解作用,在介体1-羟基苯并三唑（HBT）用量0.10%,漆酶用量100 U/L条件下作用40 min时,靛蓝脱色率达到96.7%。该方法采用的三相鼓泡塔生物反应器性能稳定、菌丝球可重复使用,该方法有利于漆酶的高效、规模化生产。     

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。     

栓孔菌属漆酶高产菌株的初步筛选及其产酶条件的优化

利用显色反应对栓孔菌属(Trametes)进行了漆酶高产菌株的筛选,并对目标菌株的产酶条件进行了优化,在添加愈创木酚的固体培养基中,通过显色反应初步筛选出漆酶高产菌株东方栓孔菌Trametes orientalis Cui 6300;进一步通过单因子分析、正交试验和ABTS法确定了菌株Cui 6300的最适产酶条件:麦芽糖15 g/L,蛋白胨3 g/L,pH 4.8,Cu2+2.0 mmol/L,培养温度28°C,接种饼直径1.5 cm,此时酶活最高可达19.923 U/mL;同时探索了Cu2+浓度及添加时间对其菌丝生物量和漆酶活力的影响。研究表明,Cu2+最适添加浓度为2.0 mmol/L,添加时间为接种后第3天。     

Ganoderma lucidum U-281漆酶催化偶氮染料活性黑5脱色

漆酶在纺织染料脱色及印染废水处理领域有着广阔的应用前景。活性黑5是纺织印染中应用广泛的偶氮类活性染料,结构复杂,生物降解性低。以灵芝菌Ganoderma lucidum U-281所产漆酶对活性黑5进行氧化脱色,采用单因素逐一优化方法得到了U-281漆酶催化活性黑5脱色的工艺参数：染料初始浓度25mg/L、漆酶用量2.0U/mL、铜离子添加量40mmol/L、pH 6.0、40℃。在优化条件下,4h可使RB5脱色62.34%,24h可完成90%以上的脱色效果。     

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,对其兴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。     

粗毛栓菌菌丝球非灭菌条件下对12种染料的脱色研究

以新型白腐真菌——粗毛栓菌Trametes gallica为材料,研究了优化后的该菌菌丝球在非灭菌条件下对直接染料、中性染料、三苯甲烷类染料以及蒽醌类染料共12种染料的脱色能力、脱色机制,以及pH、温度、染料初始浓度等参数对该菌菌丝球脱色效果的影响。结果表明,优化条件下制备的粗毛栓菌菌丝球脱色活力良好,4℃下保存20d后仍保持有原脱色活力的95%;活菌丝球比死菌丝球对染料具有更强的耐受性和更好的脱色效果;非灭菌条件下活菌丝球对12种染料的适宜脱色条件为pH 3.0–5.0、25℃、染料浓度为50mg/L、处理36–60h,该条件下粗毛栓菌菌丝球在60h内脱色率均在55%以上,其中粗毛栓菌菌丝球对亚甲基蓝脱色率最高可达到96.40%。紫外可见光谱分析和显微观察结果表明,48h内粗毛栓菌菌丝球在非灭菌条件下对12种染料的脱色是由吸附引起,无二次污染物的产生。粗毛栓菌的这些优良特性显示了其在工业染料废水处理中的广阔应用前景。     

适于罗布麻生物脱胶的果胶酶产生菌分离筛选与表征

在青海柴达木盆地沤制的罗布麻表皮中分离得到两株适于罗布麻脱胶的高产果胶酶菌株,经形态、生理生化指标鉴证和16SrDNA菌种鉴定,确定一株为新的果胶酶产生菌琼氏不动杆菌（Acinetobacter junii）,一株为枯草芽孢杆菌（Bacillus subtilis）。水解圈实验得出前者H／C值（H／C为水解圈直径H与菌落直径C之比）为5,后者为3;经酶活力测定,在37℃下,琼氏不动杆菌（Acinetobacter junii）在11h达到产酶高峰（酶活力为103．2IU／mL）,枯草芽孢杆菌（Bacillus subtilis）培养至9h达到产酶高峰（酶活力为91．6IU／mL）,但前者最高酶活力比后者高12．5％;经脱胶实验得出两菌残胶率分别为18．47％和17．31％,对罗布麻生物脱胶有较好的适用性。     

端梗霉Z45产漆酶培养基的优化及其对染料的脱色

菌株Acrophialophora sp.Z45是一株产漆酶的端梗霉属真菌,本文依据不同培养基中菌株Z45对愈创木酚的氧化圈大小探究了影响该菌株产漆酶的因素并设计正交试验优化其产漆酶培养基,进而比较了菌株Z45在土豆葡萄糖培养基和优化后的产漆酶培养基中的漆酶活力差异,最后基于漆酶与染料脱色的相关性评价了菌株Z45对8种合成染料的脱色能力。结果表明,单因素及正交试验确定了菌株Z45优化后的产漆酶培养基为：基础产酶培养基中以蔗糖为碳源、硝酸钠为氮源、C/N比为45∶1、pH为5.0;在优化后的产漆酶培养基中菌株Z45的漆酶活性显著提高;菌株Z45对溴酚蓝、中性红、亚甲基蓝、甲基蓝和结晶紫等5种染料均有一定的脱色能力,其中对三苯甲烷染料甲基蓝的脱色能力最强,菌株Z45的粗酶液能使甲基蓝快速脱色。在较低甲基蓝浓度的固体培养基上,甲基蓝完全脱色的时间不受染料浓度的影响;而较高甲基蓝浓度下,甲基蓝完全脱色的时间随染料浓度升高逐渐延长。     

血红密孔菌高产漆酶菌株的筛选及其对烟梗的生物降解

白腐菌是目前已知的唯一能将木质素彻底降解的微生物,而漆酶在木质素分解过程中起着重要的作用,被广泛应用于农作物秸秆或甘蔗渣等多种类型生物质的生物预处理和生物降解。本研究利用白腐菌产漆酶发酵培养基对30株血红密孔菌Pycnoporus sanguineus菌株进行筛选,得到了多株漆酶高产菌株,并研究了血红密孔菌发酵粗酶液和菌丝对烟梗的生物降解条件。研究结果表明:血红密孔菌及其产生的漆酶表现出了对烟梗木质素较强的生物降解能力。在漆酶浓度为40U/mL、温度30℃、pH4.5的条件下处理24h,烟梗中木质素的降解率可达到50.4%,纤维素和半纤维素的降解率分别为17.5%和17.3%;漆酶浓度为5U/mL、温度30℃、pH4.5的条件下处理48h,木质素降解率可达到65.1%。血红密孔菌菌丝也表现出对烟梗较好的生物降解效果,接种培养7d后烟梗中木质素降解率可达30%以上,21d后木质素的降解率可达79.1%,而纤维素和半纤维素的降解率仅为20%和12%左右。本研究不但为生物质材料的生物预处理和生物降解提供了优质的白腐菌及漆酶资源,还为通过烟梗的生物预处理提高烟草梗丝和卷烟品质提供了重要参数,具有一定的应用前景。     

The evaluation of pre-grown mycelial pellets in decolorization of textile dyes during repeated batch process

This study was undertaken for the possibility of application of pre-grown pellets for biotechnological treatment of dyes and textile industry waste waters. Mycelial pellets of five different white rot fungi were tested for their dye decolorization activity. The pellets of Funalia trogii, Phanerochaete chrysosporium and Trametes versicolor were determined as the most effective ones. The decolorization ability of viable pellets was compared with the decolorization (adsorption) ability of dead pellets during repeated batch studies. Astrazon Black dye was decolorized effectively, about 90%, by viable pellets of all fungi during the first use. Viable F. trogii pellets were found as the most effective pellets. Upon pellet treatment not only a high decolorization but also reduced toxicity (antimicrobial activity) of the Astrazon Black dye was recorded. This type of decolorization activity with commercial or crude laccase was partially observed. Growing cells of F. trogii in batch system showed lower efficiency in color removal of mixed dyes compared to the pre-grown pellets in repeated batch system. The results in this study showed that mycelial pellets could effectively be used as an alternative to traditional physicochemical processes.     

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。     

Activity and elution profile of laccase during biological decolorization and dephenolization of olive mill wastewater

The performance and enzymatic strategy exhibited by basidiomycete Euc-1, a laccase producing strain, was investigated during the biodegradation of olive mill wastewater (OMW). This strain yielded better decolorization of solidified OMW than Phanerochaete chrysosporium and removed 90% of phenols (initial concentration=800 mg l(-1)), 73% of color (initial A465=4.4), and 45% of chemical oxygen demand in batch cultures containing OMW. Since partial phenol removal occurred before the detection of enzymatic activity, no plausible correlation could be established between them. In contrast, decolorization occurred only after the detection of laccase activity and coincided with its production over time. Two laccase fractions (Lac1 and Lac2) were separated by chromatography. OMW strongly induced Lac2 that was almost absent in defined liquid medium. Furthermore, Lac2 was the main laccase fraction in the presence of OMW. This study pointed out that basidiomycete Euc-1 and its ligninolytic system could be a useful tool for the bioremediation of wastewater generated in the process of olive oil extraction.     

Rapid decolorization of azo dyes by crude manganese peroxidase from Schizophyllum sp. F17 in solid-state fermentation

The production of ligninolytic enzymes by the fungus Schizophyllum sp. F17 using a cost-effective medium comprised of agro-industrial residues in solid-state fermentation (SSF) was optimized. The maximum activities of the enzymes manganese peroxidase (MnP), laccase (Lac), and lignin peroxidases (LiP) were 1,200, 586, and 109 U/L, respectively, on day 5 of SSF. In vitro decolorization of three structurally different azo dyes by the extracellular enzymes was monitored to determine its decolorization capability. The results indicated that crude MnP, but not LiP and Lac, played a crucial role in the decolorization of azo dyes. After optimization of the dye decolorization system with crude MnP, the decolorization rates of Orange IV and Orange G, at an initial dye concentration of 50 mg/L, were enhanced to 76 and 57%, respectively, after 20 min of reaction at pH 4 and 35°C. However, only 8% decolorization of Congo red was observed. This enzymatic reaction system revealed a rapid decolorization of azo dyes with a low MnP activity of 24 U/L. Thus, this study could be the basis for the production and application of MnP on a larger scale using a low-cost substrate.     

Decolorization of reactive dyes by a thermostable laccase produced by Ganoderma lucidum in solid state culture

Dye decolorizing potential of the white rot fungus Ganoderma lucidum KMK2 was demonstrated for recalcitrant textile dyes. G. lucidum produced laccase as the dominant lignolytic enzyme during solid state fermentation (SSF) of wheat bran (WB), a natural lignocellulosic substrate. Crude enzyme shows excellent decolorization activity to anthraquinone dye Remazol Brilliant Blue R (RBBR) without redox mediator whereas diazo dye Remazol Black-5 (RB-5) requires a redox mediator. Polyacrylamide gel electrophoresis (PAGE) of crude enzyme confirms that the laccase enzyme was the major enzyme involved in decolorization of either dyes. Native and SDS-PAGE indicates that the presence of single laccase with molecular weight of 43 kDa. N-Hydroxybenzotriazole (HBT) at a concentration of 1 mM was found as the best redox mediator. RB-5 (50 mg l^−l) was decolorized by 62% and 77.4% within 1 and 2 h, respectively by the crude laccase (25 U ml⁻¹). RBBR (50 mg l^−l) was decolorized by 90% within 20 h, however, it was more efficient in presence of HBT showing 92% decolorization within 2 h. Crude laccase showed high thermostability and maximum decolorization activity at 60 °C and pH 4.0. The decolorization was completely inhibited by the laccase inhibitor sodium azide (0.5 mM). Enzyme inactivation method is a good method which averts the undesirable color formation in the reaction mixture after decolorization. High thermostability and efficient decolorization suggest that this crude enzyme could be effectively used to decolorize the synthetic dyes from effluents.     

Continuous decolorization of bleached kraft effluents byCoriolus versicolor in the form of pellets

Summary Continuous decolorization of kraft black liquor by mycelial pellets ofCoriolus versicolor in the presence of glucose as co-substrate is discussed. A linear relationship was developed between the rate of decolorization and the liquor concentration. The rate constant was equal to 0.00961 gmyc^–1 h^–` at 22°C. During the continuous experiments the pellets exhibited no apparent loss of activity when the liquor concentration was in the range of 400 CU l^–1 to 5000 CU l^–1. However, in the repeated batch experiments a loss of activity was observed which was dependent on the initial liquor concentration. The half-life of pellets was equal to 4.7, 9.4 and 20.2 days for the initial liquor concentration of 1380, 31 780 and 6990 CU l^–1, respectively. The production of the extracellular enzyme, laccase, was followed but could not be used as an indicator of the ligninolytic activity. The involvement of the intracellular enzymes ofC. versicolor in the decolorization process is described.     

Dephenolization and decolorization of olive mill wastewater through sequential batch and co-culture applications

In a previous work it was reported adapted Trametes versicolor FPRL 28A INI culture was used to treat undiluted olive mill wastewater (OMW) without addition of any nutrients with significant amount of total phenolics were removed. However, decolorization was not so pronounced. Therefore, the aim of this study is to enhance the efficiency of dephenolization and decolorization of the primary treatment with adapted Trametes versicolor, incorporating a secondary biological treatment step using different microorganisms with sequential batch and co-culture applications. Through sequential batch applications Funalia trogii ATCC 20080 was found to have a higher potential in terms of total phenolics removal and decolorization amongst the tested organisms and better results were obtained from sequential batch applications as compared to co-culture experiments. In sequential batch applications, up to 91% total phenolics were removed and 64% decolorization was achieved after 24 days with 20% (v/v) inoculation rate of F. trogii when malt extract broth was used in inoculum preparation. In addition, significant accumulation of laccase (2019 ± 121.13Ul⁻¹) and manganese peroxidase (463 ± 33.89 Ul⁻¹) activities were attained. In co-culture applications highest total phenolics removal and decolorization were 78 and 39%, respectively, with non-adapted T. versicolor, whereas highest laccase and manganese peroxidase acitivities were obtained with F. trogii as 2219 Ul⁻¹  ± 176.14^. and 513 ± 4.12 Ul⁻¹, respectively.     

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。     

Dye decolorization by <Emphasis Type="Italic">Trametes hirsuta</Emphasis> immobilized into alginate beads

Summary The present paper studies the production of laccase by Trametes hirsuta immobilized into alginate beads in an airlift bioreactor. In order to enhance laccase production fresh ammonium chloride was added, which led to the production, of high laccase activities (around 1000 U l⁻¹). The bioreactor operated for 40 days without operational problems and the bioparticles maintained their shape throughout fermentation. Dye decolorization was performed at bioreactor scale operating in the batch mode. High decolorization percentages were obtained in a short time (96% for indigo carmine and 69% for phenol red in 24 h), indicating the suitability of this process for application to synthetic dye decolorization. On the other hand, in vitro decolorization of several industrial azo dyes by crude laccase produced in the above bioreactor was also performed. It was found that some of the dyes needed the addition of 1-hydroxybenzotriazole for their decolorization.