摘要
偶氮染料和三苯甲烷染料广泛应用于印染、食品、造纸等各个领域,由于很多染料都具有生物毒性且难以自然降解,因此,对环境水体造成了严重的污染。传统的理化处理方法处理成本昂贵且易造成二次污染,严重阻碍了其广泛应用,而生物处理方法作为一种环境友好型处理手段受到环境工作者的青睐。某些细菌具有生长快、处理效率高等优势,在染料废水的生物治理中具有广阔的应用前景,通过对这类细菌脱色特性和脱色机理详细深入的研究,可以为染料废水的生物治理奠定理论基础。
本论文分别从腐木覆盖的土壤和曝气池的污泥中筛选到了两株染料高效脱色菌株Pseudomonas sp菌株DY1和Micrococcus sp菌株BD15,详细研究了两种菌株对偶氮染料或三苯甲烷染料的脱色特性,并采用统计学手段对其脱色条件进行优化；采用现代分析手段对两种菌株对偶氮染料或三苯甲烷染料的脱色机理进行了深入研究；并最终通过菌株固定化和毒理学研究探讨了这两株菌的实际应用潜能。本论文的主要研究结果如下：
(1)染料高效降解菌株的分离和鉴定：从自然环境中分离到两株偶氮染料酸性黑172或三苯甲烷染料孔雀石绿高效脱色菌株,通过16S rDNA和生理生化鉴定确定这两株菌株分别为Pseudomonas sp菌株DY1和Micrococcus sp菌株BD15,其中菌株DY1对酸性黑172和孔雀石绿均有较好的脱色效果,而菌株BD15则仅对孔雀石绿脱色效果较好。
(2)菌株DY1对酸性黑172的脱色特性和条件优化：实验结果表明大多数氮源能够促进脱色,而碳源则对脱色没有影响或抑制脱色。高浓度的Fe2+、Fe3+和Ca2+(10 mM)存在时,仍观察到较高的脱色率。在此基础上,采用PB设计筛选出酸性黑172脱色的主效应因子：pH、温度、Fe3+浓度和NaH2PO4浓度。随后采用响应面设计对主效应因子进行优化,结果表明该过程可以用二次模型进行解释,脱色的最佳操作条件为：pH 6.23、30℃、8.0 mM的Fe3+和10.0 g/L的NaH2PO4,验证实验结果在实验模型的置信区间内。此外,在最优条件下,该菌株对酸性黑172的脱色动力学符合对数动力学模型(R2=0.964)。
(3)菌株DY1加热致死细胞和活细胞对酸性黑172脱色特性的比较研究：菌株DY1对酸性黑172的吸附动力学数据与假二次模型的拟合度较好。起始染料浓度能够显著促进热致死细胞的吸附量,其最大吸附量可达2.98 mmol/g biomass;而活细胞对染料的吸附量则不受染料浓度的影响。随着菌体处理温度的升高,该菌株对染料的吸附能力也随之增加。电位滴定和FTIR扫描结果表明氨基基团在吸附的过程中起到关键作用。SEM、AFM和TEM分析说明对菌体进行热处理后细胞壁的通透性增加且胞内蛋白变性。不同细胞成分的吸附实验结果肯定了胞内蛋白是导致热致死细胞吸附量增大的主要原因。
(4)菌株DY1对孔雀石绿的降解特性、降解酶系和降解途径研究：实验结果表明该菌株在振荡条件下、24 h内对高浓度的孔雀石绿(100-1000 mg/L)显示出较高的脱色能力(脱色率为90.3-97.2%),而在静止条件下,该菌株的脱色率比振荡条件下低,但仍高达78.9%以上。脱色的最佳pH和温度分别为pH 6.6和28-30℃,Mg2+和Mn2+(1 mM)能够显著促进脱色。通过UV-Visible、GC-MS和LC-MS分析获得降解中间产物主要为孔雀石绿甲醇、二甲氨基二苯甲酮、甲氨基二苯甲酮、氨基二苯甲酮、二苯甲酰甲烷和N,N-二甲基苯胺。酶分析实验结果表明锰过氧化物酶、NADH-DCIP还原酶和MG还原酶可能与该菌株降解孔雀石绿相关。
(5)菌株BD15对孔雀石绿的脱色特性和脱色条件优化：采用光学显微分析和紫外分光光度分析确定脱色是由生物吸附还是生物降解引起的,并采用PB设计研究操作因素对脱色的影响,随后通过响应面设计优化操作条件。实验结果说明该菌株对孔雀石绿的脱色主要是由生物降解引起。孔雀石绿、尿素和酵母粉浓度以及接种量对脱色有显著的促进作用,而CuCl2和MgCl2浓度以及温度则对脱色有显著抑制作用。脱色的最佳操作条件为：1.0 g/L的尿素、0.9 g/L的酵母粉、100 mg/L的孔雀石绿、0.1 g/L的接种量(干重)以及温度为25.2℃。在最优条件下,孔雀石绿在1h内的脱色率高达96.9%。此外,该菌株对孔雀石绿的脱色动力学数据与二次动力学模型拟合度较好。
(6)菌株BD15对孔雀石绿的降解酶系和降解途径研究：采用UV-Visible、FTIR、GC-MS和LC-MS分析,获得菌株BD15对孔雀石绿的降解中间产物主要包括：二甲氨基二苯甲酮、米氏酮、甲氨基二苯甲酮、氨基二苯甲酮、4-甲氨基苯甲酸、4-羟基-N,N-二甲基苯铵、N,N-二甲基苯铵、羟基二甲氨基二苯甲酮和对氨基苯酚。根据降解中间产物推测了菌株BD15降解孔雀石绿的可能途径。酶分析实验结果表明,漆酶和NADH-DCIP还原酶可能与该菌株降解孔雀石绿相关。
(7)菌株固定化和毒理学实验：实验结果表明菌株DY1和BD15固定化后,均可反复多次使用,且脱色效果较好,经处理后的染料生物毒性显著降低。
Azo dyes and triphenylmethane dyes are extensively used in textile, food and paper industries, but considered to be toxic and recalcitrant. The pollution caused by the dyes is serious nowadays. The conventional physicochemical treatments are not only expensive but also may cause secondary pollution; as an eco-friendly method, the biological treatments are received more and more attentions. Some bacteria maybe have a great potential in application because these strains could grow quickly and may decolorize the dye efficiently. Studying the characteristics and mechanism of dye decolorization by these strains could be helpful for the bioremediation of the dye.
In this research, Pseudomonas sp. strain DY1 and Micrococcus sp. strain BD15 were isolated from the soil with rotten woods and sludge in aeration tank; the characteristics of the decolorization was studied systematically and the optimal conditions of the decolorization was obtain by the statistical method; the mechanism of the decolorization was investigated by different analysis; and finally, the practical application potential of these two strains was confirmed by the immobilization and toxicology tests. The results of the reseach can be devided into the following parts:
(1) The isolation and identification of dye-degrading bacteria:Pseudomonas sp. strain DY1 and Micrococcus sp. strain BD15 were isolated from the polluted soil and identified by the 16S rDNA and biochemical tests. The strain DY1 could decolorize both Acid Black 172 and Malachite Green efficiently, whereas strain BD15 could only decolorize Malachite Green efficiently.
(2) The characteristics and optimization of Acid Black 172 decolorization by the strain DY1:It was observed that most of the tested nitrogen source enhanced decolorization, while carbon source inhibited or had no influence on decolorization. Efficient decolorization was still observed when high concentrations of Fe2+, Fe3+ and Ca2+ existed. Based on the above results, a Plackett-Burman design was used to select the operational parameters for decolorization of Acid Black 172. Four significant parameters, including pH, temperature, concentrations of Fe3+ and NaH2PO4, were optimized. A quadratic model obtained from the response surface design was constructed on experimental data. The optimal condition for decolorization was found to be pH 6.23,30℃,8.0 mM of Fe3+, and 10.0 g/L of NaH2PO4. The confirmatory experiments (86.0% decolorization percentage within the confidence interval) subsequently verified the accuracy of the experimental model. Moreover, the decolorization under the optimal condition fitted the logarithmic model well (R2= 0.964).
(3) Biosorption mechanisms of the metal-complex dye Acid Black 172 by live and heat-treated biomass of strain DY1:Kinetic data for Acid Black 172 adsorption by strain DY1 fit a pseudo-second-order model. Increased initial dye concentration could significantly enhance the amount of dye adsorbed by heat-treated biomass in which the maximum amount of dye adsorbed was as high as 2.98 mmol/g biomass, whereas it had no significant influence on dye sorption by live biomass. As treated temperature increased, the biomass showed gradual increase of dye sorption ability. Experiments using potentiometric titration and FTIR indicated that amine groups (NH2) played a prominent role in biosorption of Acid Black 172. SEM, AFM and TEM analysis indicated that heat treatment of the biomass increased the permeability of the cell walls and denatured the intracellular proteins. The results of biosorption experiments by different cell components confirmed that intracellular proteins contributed to the increased biosorption of Acid Black 172 by heat-treated biomass.
(4) The characteristics, degradation products and enzyme analysis of Malachite Green by strain DY1:The results showed that this strain demonstrated high decolorizing capability (90.3-97.2%) at high concentrations of MG (100-1000 mg/L) under shaking condition within 24 h. In static conditions, lower but still effective decolorization (78.9-84.3%) was achieved. The optimal pH and temperature for the decolorization was pH 6.6 and 28-30℃, respectively. Mg2+ and Mn2+(1 mM) were observed to significantly enhance the decolorization. The intermediates of the MG degradation under aerobic condition identified by UV-Visible, GC-MS and LC-MS analysis included malachite green carbinol, (dimethyl amino-phenyl)-phenyl-methanone, (methyl amino-phenyl)-phenyl-methanone, (amino phenyl)-phenyl methanone, di-benzyl methane and N, N-dimethylaniline. The enzyme analysis indicated that Mn-peroxidase, NADH-DCIP and MG reductase were involved in the biodegradation of MG.
(5) The characteristics and optimization of Malachite Green by strain BD15: Optical microscope and UV-visible analysis were carried out to determine whether the decolorization was due to biosorption or biodegradation. Plackett-Burman design was employed to investigate the effect of various parameters on decolorization, and response surface method was then used to explore the optimal decolorization conditions. The results indicated that the decolorization by the strain was mainly due to biodegradation. Concentration of MG, urea and yeast extract, and inoculums size had significantly positive effect on decolorization, while concentration of CuCl2 and MgCl2, and temperature had significantly negative effect. The optimal conditions for decolorization were 1.0 g/L urea,0.9 g/L yeast extract,100 mg/L MG,0.1 g/L inoculums (dry weight), and at 25.2℃. Under the optimal conditions,96.9% MG was removed by the strain within 1 h, which was highly efficient microbial decolorization according to our knowledge. Moreover, the kinetic data for decolorization fit a second-order model well.
(6) The degradation products of Malachite Green by strain BD15 and enzymes analysis:The results of UV-Visible, FTIR, GC-MS and LC-MS analysis indicated that the intermediates of the MG degradation by strain BD15 included:(dimethyl amino-phenyl)-phenyl-methanone, Michler's ketone, (methyl amino-phenyl)-phenyl-methanone, (amino phenyl)-phenyl methanone, 4-methylaminobenzoic acid,4-hydroxyl-N, N-dimethylaniline, N, N-dimethylaniline, hydroxyl-(dimethyl amino-phenyl)-phenyl-methanone and 4- hydroxyl-aniline. Based on the above results, the possible degradation pathway of MG was proposed. The enzymes analysis indicated that laccase and NADH-DCIP reductase were involved in the degradation of MG.
(7) The cell immobilization and toxicology tests:The results indicated that the immobilized cells of strain DY1 and BD15 could be used repeatedly, and the decolorization efficiency was effective. The toxicity of the dye solution after decolorization was significantly decreased compared to before decolorization.
引文
[1]G. McMullan, C. Meehan, A. Conneely, N. Kirby, T. Robinson, P. Nigam, I. M. Banat, R. Marchant, W. F. Smyth, Microbial decolourisation and degradation of textile dyes, Applied Microbiology and Biotechnology,2001,56:81-87.
[2]何瑾馨,染料化学,中国纺织工业出版社。2004,30-100.
[3]O.J. Hao, H. Kim, P.C. Chiang, Decolorization of Wastewater, Critical Reviews in Environmental Science and Technology,1999,30:449-505.
[4]S. Srivastava, R. Sinha, D. Roy, Toxicological effects of malachite green, Aquatic Toxicology, 2004,66:319-329.
[5]蒋志国,王连,于文敦,赵成芳,刘越,染料工业废水的治理方法,污染防冶技术,2000,13：107-109.
[6]刘友勋,染料的环境污染及其处理方法,中南论坛(综合版),2006,2：98-107.
[7]洪颖,陈国松,张红漫,张之翼,蒋皎梅,蒽醌类染料废水处理的研究进展,工业水处理,2004,10：5-8.
[8]戴日成,张统,印染废水水质特征及处理技术综述.给水排水,2000,26：33-37.
[9]汤心虎,黄秀微,无机／有机复合絮凝剂对印染废水脱色的研究,水处理技术,2001,27：267-270.
[10]F. Rafii, J.D. Hall, C.E. Cerniglia, Mutagenicity of Azo Dyes Used in Foods, Drugs and Cosmetics Before and After Reduction by Clostridium Species from the Human Intestinal Tract, Food and Chemical Toxicology,1997,35:897-901.
[11]D. Dillon, R. Combes, E. Zeiger, Activation by caecal reduction of the azo dye D&C Red No. 9 to a bacterial mutagen, Mutagenesis,1994,9:295-299.
[12]A. Kaur, R.S. Sandhu, I.S. Grover, Screening of azo dyes for mutagenicity with Ames-Salmonella assay, Environmental and Molecular Mutagenesis,1993,22:188-190.
[13]K.T. Chung, C.E. Cerniglia, Mutagenicity of azo dyes:Structure-activity relationships, Mutation Research,1992,277:201-220.
[14]G.A.R. Oliveira, E.R.A. Ferraz, F.M.D. Chequer, M.D. Grando, J.P.F. Angeli, M.S. Tsuboy, J.C. Marcarini, M.S. Mantovani, M.E. Osugi, T.M. Lizier, M.V.B. Zanoni, D.P. Oliveira, Chlorination treatment of aqueous samples reduces, but does not eliminate, the mutagenic effect of the azo dyes Disperse Red 1, Disperse Red 13 and Disperse Orange 1, Mutation Research,2010, 703:200-208.
[15]G. Paramaguru, A. Kathiravan, S. Selvaraj, P. Venuvanalingam, R. Renganathan, Interaction of anthraquinone dyes with lysozyme:Evidences from spectroscopic and docking studies, Journal of Hazardous Materials,2010,175:985-991.
[16]T.C. Marrs, H.F. Colgrave, P. Rice, J.A.G. Edginton, B. Morris, The repeated dose toxicity of a smoke containing Disperse Blue 180, an anthraquinone dye mixture, Journal of Hazardous Materials,1989,21:73-88.
[17]R.H. Jaskot, D.L. Costa, Toxicity of an Anthraquinone Violet Dye mixture following inhalation exposure, intratracheal instillation, or gavage, Fundamental and Applied Toxicology, 1994,22:103-112.
[18]C. Novotny, N. Dias, A. Kapanen, K. Malachova, M. Vandrovcova, M. Itavaara, N. Lima, Comparative use of bacterial, algal and protozoan tests to study toxicity of azo-and anthraquinone dyes, Chemosphere,2006,63:1436-1442.
[19]J.M. Fanchiang, D.H. Tseng, Degradation of anthraquinone dye C.I. Reactive Blue 19 in aqueous solution by ozonation, Chemosphere,2009,77:214-221.
[20]S.J. Srivastava, N.D. Singh, A.K. Srivastava, R. Sinha, Acute toxicity of malachite green and its effects on certain blood parameters of a catfish, Heteropneustes fossilis, Aquatic Toxicology, 1995,31:241-247.
[21]L.D. Wright, Effect of malachite green and formaline on the survival of large mouth bass eggs and fry. The Progressive Fish-Culturist,1976,38:155-157.
[22]刘友勋,耐碱真菌Myrothecium sp. IMER1对染料脱色的机理研究,博士论文。
[23]M. Ghaedi, A. Hekmati Jah, S. Khodadoust, R. Sahraei, A. Daneshfar, A. Mihandoost, M.K. Purkait, Cadmium telluride nanoparticles loaded on activated carbon as adsorbent for removal of sunset yellow, Spectrochimica Acta Part A,2012,90:22-27.
[24]C. Soma, M. Rumeau, C. Sergent, Use ofmineral membranes in the treatment of texiles effluents pore intl continorganic membrane, France, Montpeller,1989,10:523-526.
[25]刘艳,解立平,费学宁,姜远光,一体式光催化-膜分离反应器处理酸性红B染料废水,天津工业大学学报,2011,30：60-64.
[26]E. Kusvuran, O. Gulnaz, A. Samil, O. Yildirim, Decolorization of malachite green, decolorization kinetics and stoichiometry of ozone-malachite green and removal of antibacterial activity with ozonation processes, Journal of Hazardous Materials,2011,186:133-143.
[27]M.A. Oturan, E. Guivarch, N. Oturan, I. Sires, Oxidation pathways of malachite green by Fe3+-catalyzed electro-Fenton process, Applied Catalysis B:Environmental,2008,82:244-254.
[28]周光元,卢海青,CQS1.2/0.6型采气废水处理装置,石油机械,2004,32：35-39.
[29]陈跃,染料废水处理技术及研究趋势,黄石理工学院学报,2011,27：8-14.
[30]赵雪,何瑾馨,展义臻,印染废水处理技术的研究进展,化学工业与工程技术,2009,30：38-42.
[31]T. Robinson, G. McMullan, R. Marchant, P. Nigam, Remediation of dyes in textile effluent, a critical review on current treatment technologies with a proposed alternative. Bioresource Technology,2001,77:247-255.
[32]徐文东,文湘华,微生物在含染料废水处理中的应用,环境污染治理技术与设备,2000,1：9-16.
[33]Y. Fu, T. Viraraghavan, Fungal decolorization of dye wastewaters:a review, Bioresource Technology,2001,79:251-262.
[34]D. Eaton, H.M. Chang, T.K. Kirk, Fungal decolorization of Kraft bleach plant effluent. Tappi Journal,1980,63:103-109.
[35]T. Fukuzumi, H. Chang, (Eds.), Microbiology, Chemistry and Potential Applications, vol.1. CRC press, Boca Raton FL, USA, pp.215-230.
[36]J.K. Glenn, M.H. Gold, Decolorization of several polymeric dyes by the lignin-degrading basidiomycete Phanerochaete chrysosporium, Applied and Environmental Microbiology,1983,45: 1741-1747.
[37]M.A.M. Martins, I.C. Ferreira, I.M. Santos, M.J. Queiroz, N. Lima, Biodegradation of bioaccessible textile azo dyes by Phanerochaete chrysosporium, Journal of Biotechnology,2001, 89:91-98.
[38]K.V. Radha, I. Regupathi, A. Arunagiri, T. Murugesan, Decolorization studies of synthetic dyes using Phanerochaete chrysosporium and their kinetics, Process Biochemistry,2005,40: 3337-3345.
[39]S. Senthilkumar, M. Perumalsamy, H.J. Prabhu, Decolourization potential of white-rot fungus Phanerochaete chrysosporium on synthetic dye bath effluent containing Amido black 10B, Journal of Saudi Chemical Society, doi:10.1016/j.jscs.2011.10.010
[40]Y. Fu, T. Viraraghavan, Removal of a dye from an aqueous solution by the fungus Aspergillus niger, Water Quality Research Journal of Canada,2000,35:95-111
[41]H. Horitsu, M. Takada, E. ldaka, M. Tomoyeda, T. Ogawa, Degradation of p-aminoazobenzene by Bacillus subtilis, European Journal of Applied Microbiology,1977,4: 217-224.
[42]杨清香,贾振杰,杨敏,微生物染料脱色研究进展,微生物学通报,2006,33：144-148.
[43]M.Y. Xu, J. Guo, G.Q. Zeng, X.Y. Zhong, G.P. Sun, Decolorization of anthraquinone dye by Shewanella decolorationis S12, Applied Microbiology and Biotechnology,2006,71:246-251.
[44]A. Khalid, M. Arshad, D.E. Crowley, Accelerated decolorization of structurally different azo dyes by newly isolated bacterial strains, Applied Microbiology and Biotechnology,2008,78: 361-369.
[45]C.I. Pearce, J.R. Lloyd, J.T. Guthrie, The removal of colour from textile wastewater using whole bacterial cells:a review, Dyes and Pigments,2003,58:179-196.
[46]N.J. Willmott, The use of bacteria-polymer composites for the removal of colour from reactive dye effluents. PhD thesis, UK:University of Leeds; 1997.
[47]E.S. Yoo, J. Libra, L. Adrian, Mechanism of decolorization of azo dyes in anaerobic mixed culture, Journal of Environmental Engineering,2001,127:844-849.
[48]K. Vijayaraghavan, Y.S. Yun, Utilization of fermentation waste (Corynebacterium glutamicum) for biosorption of Reactive Black 5 from aqueous solution, Journal of Hazardous Materials,2007,141:45-52.
[49]G. Bayramoglu, M.Y. Arica, Biosorption of benzidine based textile dyes "Direct Blue 1 and Direct Red 128" using native and heat-treated biomass of Trametes versicolor, Journal of Hazardous Materials,2007,143:135-143.
[50]L. Velasquez, J. Dussan, Biosorption and bioaccumulation of heavy metals on dead and living biomass of Bacillus sphaericus, Journal of Hazardous Materials,2009,167:713-716.
[51]A. Pandey, P. Singh, L. Iyengar, Bacterial decolorization and degradation of azo dyes, International Biodeterioration & Biodegradation,2007,59:73-84.
[52]M. Kudlich, A. Keck, J. Klein, A. Stolz, Localization of the enzyme system involved in the anaerobic reduction of azo dyes by Sphingomonas sp. strain BN6 and effect of artificial redox mediators on the rate of azo dye reduction, Applied and Environmental Microoiology,1997,63: 3691-3694.
[53]B. Yan, J.T. Zhou, J. Wang, C.H. Du, H.M. Hou, Z.Y.Song, Y.M. Bao, Expression and characterization of the gene encoding azoreductase from Rhodobacter sphaeroides AS1.1737, FEMS Microbiology Letters,2004,236:129-136.
[54]U. Shedbalkar, R. Dhanve, J. Jadhav, Biodegradation of triphenylmethane dye cotton blue by Penicillium ochrochloron MTCC 517, Journal of Hazardous Materials,2008,157:472-479.
[55]C.Y. Chen, Y.T. Huang, I.H. Ho, Y.C. Chung, Biological decolorization of dye solution containing malachite green by Pandoraea pulmonicola YC32 using a batch and continuous system, Journal of Hazardous Materials,2009,172:1439-1445.
[56]F.P. Van der Zee, S. Villaverde, Combined anaerobic-aerobic treatment of azo dyes--A short review of bioreactor studies, Water Research,2005,39:1425-1440.
[57]A. Stolz, Basic and applied aspects in the microbial degradation of azo dyes, Applied Microbiology and Biotechnology,2001,56:69-80.
[58]Y. Fu, T. Viraraghavan, Dye biosorption sites in Aspergillus niger, Bioresource Technology, 2002,82:139-145.
[59]S.M. Ghoreishi, R. Haghighi, Chemical catalytic reaction and biological oxidation for treatment of non-biodegradable textile effluent, Chemical Engineering Journal,2003,95: 163-169.
[60]B. Xin, G. Chen, W. Zheng, Bioaccumulation of Cu-complex reactive dye by growing pellets of Penicillium oxalicum and its mechanism, Water Research,2010,44:3565-3572.
[61]E.D.A. Mohey, J.A. Libra, U. Wiesmann, Kinetics of decolorization and mineralization of the azo dye Reactive Black 5 by hydrogen peroxide and UV light, Water Science and Technology, 2001,44:295-301.
[62]J. Wu, T. Wang, Ozonation of aqueous azo dye in a semi-batch reactor, Water Research,2001, 35:1093-1099.
[63]K. Chojnacka, Biosorption and bioaccumulation-the prospects for practical applications, Environment International,2010,36:299-307.
[64]J.G. Holt, N.R. Krieg, P.H. Sneath, J.T. Staley, S.T. Williams, Bergey's manual of detrerminative bacteriology. In. Lippincott Williams and Wilkins, Baltimore.9th ed.1994, pp 93-168.
[65]东秀珠,蔡妙英等,常见细菌系统鉴定手册,北京：学出版社,2001.2.
[66]周德庆,微生物实验手册,上海：上海科学技术出版社,1983.
[67]J.P. Chen, Y.S. Lin, Decolorization of azo dye by immobilized Pseudomonas luteola entrapped in alginate-silicate sol-gel beads, Process Biochemistry,2007,42:934-942.
[68]H.M. Pinheiro, E. Touraud, O. Thomas, Aromatic amines from azo dye reduction:status review with emphasis on direct UV spectrophotometric detection in textile industry wastewaters, Dyes and Pigments,2004,61:121-139.
[69]T. Li, J.T. Guthrie, Colour removal from aqueous solution of metal-complex azo dyes using bacterial cells of Shewanella strain J18 143, Bioresource Technology,2010,101:4291-4295.
[70]K. Ravikumar, B. Deebika, K. Balu, Decolourization of aqueous dye solutions by a novel adsorbent:Application of statistical designs and surface plots for the optimization and regression analysis, Journal of Hazardous Materials,2005,122:75-83.
[71]I.M. Banat, P. Nigam, D. Singh, R. Marchant, Microbial decolorization of textile-dye containing effluents:A review, Bioresource Technology,1996,58:217-227.
[72]E. Forgacs, T. Cserhati, G. Oros, Removal of synthetic dyes from wastewaters:a review, Environment International,2004,30:953-971.
[73]J. Yu, X.W. Wang, P.L. Yue, Optimal Decolorization and Kinetic Modeling of Synthetic Dyes by Pseudomonas Strains, Water Research,2001,35:3579-3586.
[74]N.K. Kilic, J.L. Nielsen, M. Yuce, G. Donmez, Characterization of a simple bacterial consortium for effective treatment of wastewaters with reactive dyes and Cr(VI), Chemosphere, 2007,67:826-831.
[75]M. Sukumar, A. Sivasamy, G Swaminathan, Decolorization of textile dye effluent by genetically improved bacterial strains, Applied Biochemistry and Biotechnology,2007,136:53-62.
[76]M. Chhabra, S. Mishra, T.P. Sreekrishnan, Laccase/mediator assisted degradation of triarylmethane dyes in a continuous membrane reactor, Journal of Biotechnology,2009,143: 69-78.
[77]N. Casas, T. Parella, T. Vicent, G. Caminal, M. Sarra, Metabolites from the biodegradation of triphenylmethane dyes by trametes versicolor or laccase, Chemosphere,2009,75:1344-1349.
[78]Z.W. Wang, X.L. Liu, Medium optimization for antifungal active substances production from a newly isolated Paenibacillus sp. using response surface methodology, Bioresource Technology, 2008,99:8245-8251.
[79]A.R. Cockshott, G.R. Sullivan, Improving the fermentation medium for Echinocandin B production. Part I:sequential statistical experimental design, Process Biochemistry,2001,36: 647-660.
[80]L. Levin, F. Forchiassin, A. Viale, Ligninolytic enzyme production and dye decolorization by Trametes trogii:application of the Plackett-Burman experimental design to evaluate nutritional requirements, Process Biochemistry,2005,40:1381-1387.
[81]M Amini, H. Younesi, N. Bahramifar, A.A.Z. Lorestani, F. Ghorbani, A. Daneshi, M. Sharifzadeh, Application of response surface methodology for optimization of lead biosorption in an aqueous solution by Aspergillus niger, Journal of Hazardous Materials,2008,154:694-702.
[82]S. Mohana, S. Shrivastava, J. Divecha, D. Madamwar, Response surface methodology for optimization of medium for decolorization of textile dye Direct Black 22 by a novel bacterial consortium, Bioresource Technology,2008,99:562-569.
[83]S.L.C. Ferreira, R.E. Bruns, H.S. Ferreira, GD. Matos, J.M. David, G.C. Brandao, E.G.P. da Silva, L.A. Portugal, P.S. dos Reis, A.S. Souza, W.N.L. dos Santos, Box-Behnken design:An alternative for the optimization of analytical methods, Analytical Chimica Acta,2007,597: 179-186.
[84]R.G. Saratale, G.D. Saratale, J.S. Chang, S.P. Govindwar, Decolorization and biodegradation of textile dye Navy blue HER by Trichosporon beigelii NCIM-3326, Journal of Hazardous Materials,2009a,166:1421-1428.
[85]R.G. Saratale, G.D. Saratale, J.S. Chang, S.P. Govindwar, Ecofriendly degradation of sulfonated diazo dye C.I. Reactive Green 19A using Micrococcus glutamicus NCIM-2168, Bioresource Technology,2009b,100:3897-3905.
[86]C.V. Nachiyar, G.S. Rajakumar, Purification and characterization of an oxygen insensitive azoreductase from Pseudomonas aeruginosa, Enzyme and Microbial Technology,2005,36: 503-509.
[87]M.Y. Xu, J. Guo, X.Y. Kong, X.J. Chen, G.P. Sun, Fe (Ⅲ)-enhanced azo reduction by Shewanella decolorationis S12, Applied Microbiology and Biotechnology,2007,74:1342-1349.
[88]Z. Aksu, Application of biosorption for the removal of organic pollutants:a review, Process Biochemistry,2005,40:997-1026.
[89]S.H. Chang, K.S. Wang, S.J. Chao, T.H. Peng, L.C. Huang, Degradation of azo and anthraquinone dyes by a low-cost Fe0/air process, Journal of Hazardous Materials,2009,166: 1127-1133.
[90]B.E. Tastan, S. Ertugrul, G. Donmez, Effective bioremoval of reactive dye and heavy metals by Aspergillus versicolor, Bioresource Technology,2010,101:870-876.
[91]G. Wei, L. Fan, W. Zhu, Y. Fu, J. Yu, M. Tang, Isolation and characterization of the heavy metal resistant bacteria CCNWRS33-2 isolated from root nodule of Lespedeza cuneata in gold mine tailings in China, Journal of Hazardous Material,2009,162:50-56.
[92]Y.G. Hong, X.J. Chen, J. Guo, Z.C. Xu, M.Y. Xu, G.P. Sun, Effects of electron donors and acceptors on anaerobic reduction of azo dyes by Shewanella decolorationis S12, Applied Microbiology and Biotechnology,2007,74:230-238
[93]P. Kaushik, A. Malik, Fungal dye decolourization:Recent advances and future potential, Environment International,2009,35,127-141.
[94]A.R. Khataee, G. Dehghan, A. Ebadi, M. Zarei, M. Pourhassan, Biological treatment of a dye solution by Macroalgae Chara sp.:Effect of operational parameters, intermediates identification and artificial neural network modeling, Bioresource Technology,2010,101:2252-2258.
[95]G.K. Parshetti, A.A. Telke, D.C. Kalyani, S.P. Gocindwar, Decolorization and detoxification of sulfonated azo dye methyl orange by Kocuria rosea MTCC 1532, Journal of Hazardous Materials,2010,176:503-509.
[96]K. Vijayaraghavan, Y.S. Yun, Bacterial biosorbents and biosorption, Biotechnology Advances, 2008,26:266-291.
[97]Z. Aksu, E. Balibek, Effect of salinity on metal-complex dye biosorption by Rhizopus arrhizus, Journal of Environmental Management,2010,91:1546-1555.
[98]Z. Aksu, G. Donmez, A comparative study on the biosorption characteristics some yeasts for Remazol Blue reactive dye, Chemosphere,2003,50:1075-1083.
[99]T.V.N. Padmesh, K. Vijayaraghavan, G. Sekaran, M. Velan, Batch and column studies on biosorption of acid dyes on fresh water macro alga Azolla filiculoides, Journal of Hazardous Materials,2005, B125:121-129.
[100]G. Bayramoglu, G. Celik, M.Y. Arica, Biosorption of Reactive Blue 4 dye by native and treated fungus Phanerocheate chrysosporium:Batch and continuous flow system studies, Journal of Hazardous Materials,2006, B137:1689-1697.
[101]T. Akar, I. Tosun, Z. Kaynak, E. Kavas, G. Incirkus, S.T. Akar, Assessment of the biosorption characteristics of a macro-fungus for the decolorization of Acid Red 44 (AR44) dye, Journal of Hazardous Materials,2009a,171:865-871.
[102]S.T. Akar, A.S. Ozcan, T. Akar, A. Ozcan, Z. Kaynak, Biosorption of a reactive textile dye from aqueous solutions utilizing an agro-waste, Desalination,2009b,249:757-761.
[103]F. Colaka, N. Atarb, A. Olgun, Biosorption of acidic dyes from aqueous solution by Paenibacillus macerans:Kinetic, thermodynamic and equilibrium studies, Chemical Engineering Journal,2009,150:122-130.
[104]S.T. Akar, A. Gorgulu, Z. Kaynak, B. Anilan, T. Akar, Biosorption of Reactive Blue 49 dye under batch and continuous mode using a mixed biosorbent of macro-fungus Agaricus bisporus and Thuja orientalis cones, Chemical Engineering Journal,2009c,148:26-34.
[105]M.C. Ncibi, B. Mahjoub, A.M. Ben Hamissa, R. Ben Mansour, M. Seffen, Biosorption of textile metal-complexed dye from aqueous medium using Posidonia oceanica (L.) leaf sheaths: Mathematical modeling, Desalination,2009,243:109-121.
[106]L.D. Fiorentin, D.E.G. Trigueros, A.N. Modenes, F.R. Espinoza-Quinones, N.C. Pereira, S.T.D. Barros, O.A.A. Santos, Biosorption of reactive blue 5G dye onto drying orange bagasse in batch system:Kinetic and equilibrium modeling, Chemical Engineering Journal,2010,163: 68-77.
[107]T.L. Hu, Sorption of reactive dyes by Aeromonas biomass, Water Science and Technology, 1992,26:357-66.
[108]T. Akar, I. Tosun, Z. Kaynak, E. Ozkara, O. Yeni, E.N. Sahin, S.T. Akar, An attractive agro-industrial by-product in environmental cleanup:Dye biosorption potential of untreated olive pomace, Journal of Hazardous Materials,2009d,166:1217-1225.
[109]K. Vijayaraghavan, J. Mao, Y.S. Yun, Biosorption of methylene blue from aqueous solution using free and polysulfone-immobilized Corynebacterium glutamicum:Batch and column studies, Bioresource Technology,2008,99:2864-2871.
[110]L.C. Fang, P. Cai, W.L. Chen, W. Liang, Z.N. Hong, Q.Y. Huang, Impact of cell wall structure on the behavior of bacterial cells in the binding of copper and cadmium, Colloids and Surfaces A:Physicochem. Eng. Aspects,2009,347:50-55.
[111]J.S. Cox, D.S. Smith, L.A. Warren, F.G. Ferris, Characterizing heterogeneous bacterial surface functional groups using discrete affinity spectra for proton binding, Environmental Science & Technology,1999,33:4514-4521.
[112]E.K. Putra, R. Pranowo, J. Sunarso, N. Indraswati, S. Ismadji, Performance of activated carbon and bentonite for adsorption of amoxicillin from wastewater:Mechanisms, isotherms and kinetics, Water Research,2009,43:2419-2430.
[113]S.Garip, A.C. Gozen, F. Severcan, Use of Fourier transform infrared spectroscopy for rapid comparative analysis of Bacillus and Micrococcus isolates, Food Chemistry,2009,113: 1301-1307.
[114]C.Y. Jia, D.Z. Wei, W.G. Liu, C. Han, S.L. Gao, Y.J. Wang, Selective adsorption of bacteria on sulfide minerals surface, Transactions of Nonferrous Metals Society of China,2008,18: 1247-1252.
[115]Z. Filip, S. Hermann, An attempt to differentiate Pseudomonas spp. and other soil bacteria by FT-IR spectroscopy, European Journal of Soil Biology,2001,37:137-143.
[116]S.W. Won, M.H. Han, Y.S. Yun, Different binding mechanisms in biosorption of reactive dyes according to their reactivity, Water Research,2008,42:4847-4855.
[117]Y. Zhang, D. Wei, R. Huang, M. Yang, S. Zhang, X. Dou, D. Wang, V. Vimonses, Binding mechanisms and QSAR modeling of aromatic pollutant biosorption on Penicillium oxalicum biomass, Chemical Engineering Journal.2011,166:624-630.
[118]J. Wang, S.Y. He, L.N. Xu, N. Gu, Transmission electron microscopy and atomic forcemicroscopy characterization of nickel deposition on bacterial cells, Chinese Science Bulletin, 2007,52:2919-2924.
[119]S.K. Kazya, S.F. D'Souzab, P. Sar, Uranium and thorium sequestration by a Pseudomonas sp.:Mechanism and chemical characterization, Journal of Hazardous Materials,2009,163:65-72.
[120]N.S. Maurya, A.K. Mittal, P. Cornel, E. Rother, Biosorption of dyes using dead macro fungi: Effect of dye structure, ionic strength and pH, Bioresource Technology,2006,97:512-521.
[121]A. Srinivasan, T. Viraraghavan, Decolorization of dye wastewaters by biosorbents:A review, Journal of Environmental Management,2010,91:1915-1929.
[122]Y.Z. Zhang, B. Zhou, X.P. Zhang, P. Huang, C.H. Li, Y. Liu, Interaction of malachite green with bovine serum albumin:Determination of the binding mechanism and binding site by spectroscopic methods, Journal of Hazardous Materials,2009,163:1345-1352.
[123]V.T. Fessard, T. Godard, S. Huet, A. Mourot, J.M. Poul, Mutagenicity of malachite green and leucomalachite green in vitro tests, Journal of Applied Toxicology,1999,19:421-430.
[124]I.A. Rahman, B. Saad, S. Shaidan, E.S. Sya Rizal, Adsorption characteristics of malachite green on activated carbon derived from rice husks produced by chemical-thermal process, Bioresource Technology,2005,96:1578-1583.
[125]L. Papinutti, N. Mouso, F. Forchiassin, Removal and degradation of the fungicide dye malachite green from aqueous solution using the system wheat bran-Fomes sclerodermeus, Enzyme and Microbial Technology,2006,39:848-853.
[126]B.H. Hameed, T.W. Lee, Degradation of malachite green in aqueous solution by Fenton process, Journal of Hazardous Materials,2009,164:468-472.
[127]R. Han, Y. Wang, Q. Sun, L. Wang, J. Song, X. He, C. Dou, Malachite green adsorption onto natural zeolite and reuse by microwave irradiation. Journal of Hazardous Materials,2010,175: 1056-1061.
[128]W. Jing, J.B. Gil, KIM Kyoung-Sook, LEE Young-Choon, SUNG Nak-Chang, Isolation and characterization of Pseudomonas otitidis WL-13 and its capacity to decolorize triphenylmethane dyes, Journal of Environmental Science,2009,21:960-964.
[129]G. Parshetti, S. Kalme, G. Saratale, S. Govindwar, Biodegradation of Malachite Green by Kocuria rosea MTCC 1532, Acta Chimica Slovenica,2006,53:492-498.
[130]N. Daneshvar, A.R. Khataee, M.H. Rasoulifard, M. Pourhassan, Biodegradation of dye solution containing Malachite Green:Optimization of effective parameters using Taguchi method, Journal of Hazardous Materials,2007,143:214-219.
[131]L. Ayed, K. Chaieb, A. Cheref, A. Bakhrouf, Biodegradation of triphenylmethane dye Malachite Green by Sphingomonas paucimobilis, World Journal of Microbiology and Biotechnology,2009,25:705-711.
[132]M. Tien, T.K. Kirk, Lignin peroxidase of Phanerochaete chrysosporium, Methods in Enzymology,1988,161:238-249.
[133]H. Bermek, I. Gulseren, K. Li, H. Jung, C. Tamerler, The effect of fungal morphology on ligninolytic enzyme production by a recently isolated wood-degrading fungus Trichophyton rubrum LSK-27, World Journal of Microbiology and Biotechnology,2004,20:345-349.
[134]J.P. Jadhav, S.P. Govindwar, Biotransformation of malachite green by Saccharomyces cerevisiae MTCC 463. Yeast,2006,23:315-323.
[135]D.C. Kalyani, P.S. Patil, J.P. Jadhav, S.P. Govindwar, Biodegradation of reactive textile dye Red BLI by an isolated bacterium Pseudomonas sp. SUK1, Bioresource Technology,2008,99: 4635-4641.
[136]R.K. Sani, U.C. Banerjee, Decolorization of triphenylmethane dyes and textile and dye-stuff effluent by Kurthia sp., Enzyme and Microbial Technolgy,1999,24:433-437.
[137]D. Deng, J. Guo, G. Zeng, G. Sun, Decolorization of anthraquinone, triphenylmethane and azo dyes by a new isolated Bacillus cereus strain DC11, International Biodeteriation and Biodegradation,2008,62:263-269.
[138]M.M. Kammoun, H.Z. Mechichi, L. Belbahri, S. Woodward, T. Mechichi, Malachite green decolourization and detoxification by the laccase from a newly isolated strain of Trametes sp., International Biodeteriation and Biodegradation,2009,63:600-606.
[139]L.T. Li, Q. Hong, X. Yan, G.H. Fang, S.W. Ali, S.P. Li, Isolation of a malachite green-degrading Pseudomonas sp. MDB-1 strain and cloning of the tmr2 gene, Biodegradation, 2009,20:769-776.
[140]C.H. Chen, C.F. Chang, S.M. Liu, Partial degradation mechanisms of malachite green and methyl violet B by Shewanella decolorationis NTOU1 under anaerobic conditions, Journal of Hazardous Materials,2010,177:281-289.
[141]U. Shedbalkar, J.P. Jadhav, Detoxification of Malachite Green and Textile Industrial Effluent by Penicillium ochrochloron, Biotechnology and Bioprocess Engineering,2011,16: 196-204.
[142]L.N. Du, S. Wang, G. Li, B. Wang, X.M. Jia, Y.H. Zhao, Y.L. Chen, Biodegradation of malachite green by Pseudomonas sp. strain DY1 under aerobic condition:characteristics, degradation products, enzyme analysis and phytotoxicity, Ecotoxicology,2011,20:438-446.
[143]S.I. Mulla, R.S. Hoskeri, Y.S. Shouche, N.H.Z. innekar, Biodegradation of 2-Nitrotoluene by Micrococcus sp. strain SMN-1, Biodegradation,2011,22:95-102.
[144]W.S. El-Sayed, A.F. El-Baz, A.M. Othman, Biodegradation of melamine formaldehyde by Micrococcus sp. strain MF-1 isolated from aminoplastic wastewater effluent, International Biodeteriation and Biodegradation,2006,57:75-81.
[145]C.L. Zheng, B.C. Qu, J. Wang, J.T. Zhou, J. Wang, H. Lu, Isolation and characterization of a novel nitrobenzene-degrading bacterium with high salinity tolerance:Micrococcus luteus, Journal of Hazardous Materials,2009,165:1152-1158.
[146]S.S. Singh, A.K. Dikshit, Optimization of the parameters for decolourization by Aspergillus niger of anaerobically digested distillery spentwash pretreated with polyaluminium chloride, Journal of Hazardous Materials,2010,176:864-869.
[147]L. Ayed, E. Khelifi, H.B. Jannet, H. Miladi, A. Cheref, S. Achour, A. Bakhrouf, Response surface methodology for decolorization of azo dye Methyl Orange by bacterial consortium: Produced enzymes and metabolites characterization, Chemical Engineering Journal,2010a,165: 200-208.
[148]D. Das, D. Charumathi, N. Das, Combined effects of sugarcane bagasse extract and synthetic dyes on the growth and bioaccumulation properties of Pichia fermentans MTCC 189, Journal of Hazardous Materials,2010,183:497-505.
[149]X.Y. Lin, X.H. Xu, C.H. Yang, Y.H. Zhao, Z.H. Feng, Y.Y. Dong, Activities of antioxidant enzymes in three bacteria exposed to bensulfuron-methyl, Ecotoxicology and Environmental Safety,2009,72:1899-1904.
[150]杨雪芬, 赵旭德,固定化微生物技术在印染废水处理方面的研究进展,污染防治技术,2011,24：47-49.
[151]K.C. Chen, J.Y. Wu, C.C. Huang, Y.M. Liang, S.C.J. Hwang, Decolorization of azo dye using PVA-immobilized microorganisms, Journal of Biotechnology,2003,101:241-252.[152]闫鹏,固定化生物技术在废水处理中的研究,绿色科技,2011,3：92-93.
[153]U.U. Jadhav, V.V. Dawkar, G.S. Ghodake, S.P. Govindwar, Biodegradation of Direct Red 5B, a textile dye by newly isolated Comamonas sp. UVS, Journal of Hazardous Materials,2008, 158:507-516.
[154]L. Lyubenova, E. Nehnevajova, R. Herzig, P. Schroder, Response of antioxidant enzymes in Nicotiana tabacum clones during phytoextraction of heavy metals, Environmental Science and Pollution Research,2009,16:573-581.