摘要
酚类化合物是废水中常见的一类高毒性和难于降解的有机物。由于酚的毒性大,而且涉及水生生物的生长和繁殖,污染饮用水水源,因此如何能够有效安全地降解酚类化合物即成为人们一直关心的问题。在众多的治理方法中,生物法具有成本低、无二次污染等特点而成为含酚废水处理的主流。但酚具有强毒性,因此需要降解菌种在具有较强的降酚能力的同时,还具有较好的耐受高浓度酚的能力,从而进一步提高降酚效率。
基于以上的一些问题,本文以获得诱变高效菌株为主要研究目的,以粘质沙雷氏菌(S. marcescens AB 90027)为出发菌株,采用等离子体诱变结合高浓度酚筛选的方法,获得了遗传性状较稳定的优势降酚菌株S. marcescens AB 90027 P15b-HQ3,降酚能力较原菌株有了很大的提高。
微生物共代谢是处理难降解有机物的重要方式,论文探讨了S. marcescens AB 90027 P15b-HQ3共代谢处理对苯二酚的降解特性。文中比较了在以对苯二酚作为唯一碳源及共代谢基质存在下S. marcescens AB 90027 P15b-HQ3的降解过程,并研究了不同碳源、氮源对该菌株共代谢降解对苯二酚的影响。结果表明:共代谢基质的添加显著提高对苯二酚的降解率及缩短处理时间。2 g/L的甘油作为碳源时,14天对苯二酚的降解率达80.4%且半衰期仅为3.5 d,优于其它碳源的作用效果;蛋白胨的加入虽然稍影响了对苯二酚的降解但同时进一步缩短细菌的共代谢处理时间,当蛋白胨含量为1.5 g/L时能有效调节两者间的平衡并提高对苯二酚的降解效率。论文对S. marcescens AB 90027 P15b-HQ3共代谢降解对苯二酚的研究发现,接种量为5%(v/v)、Mn2+离子浓度为0.1 mg/L,Fe2+离子浓度为0.1 mg/L,Cu2+离子浓度为0.1 mg/L, Ca2+离子浓度为10 mg/L时,对苯二酚去除最为理想,9 d后降解率达80.5%。通过对降解条件的优化,提高了对苯二酚的降解率及缩短其降解周期,为共代谢降解酚类化合物提供了可靠的理论基础。共代谢-酶催化双氧水联合有利于对苯二酚的进一步降解,且在一定条件下双氧水的添加量存在一个最佳值:3 mL 10%H2O2处理100 mL 2000 mg/L对苯二酚的降解率达97.6%;一次性滴加双氧水与分批滴加双氧水的方式对降解效果影响不大,两者的降解率在96.9%-97.4%之间。
Phenolic compounds are a class of strong toxic and refractory organic compounds frequently found in waste water. Since the phenolic compounds affect the growth and reproduction of aquatic organism and contaminates the drinking water sources, how to degradate such compounds in utility and safe has been an important problem regarded by human. Of all the methods of degradating phenolic compounds, biodegradation has been a main trend because of its low cost and no secondary pollution. For the strong toxicity of phenolic waste water, it is necessary to get one strain with fine degradation ability and high tolerance to phenolic compounds, which could elevate the efficiency of degradation.
Based on the problem discussed above, this thesis studied the mutation of efficient strains of hydroquinone degradation. The bacteria source is S. marcescens AB 90027, which was mutated to obtain S. marcescens AB 90027 P15b-HQ3 by means of plasma induced mutation combining with the selection of hydroquinone tolerance. The results showed that the hydroquinone degradation ratio of the mutant above was higher than that of the parent strain.
Co-metabolism is one of the most important methods for refractory organic compounds degradation.The study dealed with the degradation of hydroquinone by S. marcescens AB 90027 P15b- HQ3 cultured with different sources of carbon, nitrogen. The results showed that addition of carbon source accelerated the degradation of hydroquinone under co-metabolism process. Compared to other carbon sources studied, glycerine was the best carbon source for the degradation of hydroquinone. After 14 days’hydroquinone degradation, the degradation ratio of hydroquinone was 80.4% and the half-life of hydroquinone was 3.5 d while the concentration of glycerine was 2 g/L. Supplement of peptone hindered the degradation of hydroquinone , however, peptone further decurtated degradation of hydroquinone. It is available to accommodate the balance of both as the concentration of peptone was 1.5 g/L. The optimum degradation mediums were glycerine 2 g/L, peptone 1.5 g/L, Mn2+ 0.1 mg/L, Fe2+ 0.1 mg/L, Cu2+ 0.1 mg/L, Ca2+ 10 mg/L while the degradation ratio of hydroquinone reached to 80.5% 9 days’degradation. The yield was enhanced and the degradation time was shortened by optimization of culture media and degradation conditions in flasks, which provided dependable rationale for co-metabolism of phenolic compounds.
The addition of H2O2 profited for hydroquinone further degradation, and there was an optimum value at the condition of degradation: 3 mL 10%H2O2 could removal most 100 mL 2000 mg/L hydroquinone and the degradation ratio was 97.6%. Besides, compared to the method of dropwising H2O2 in batch, dropwising H2O2 at one time had the same effect in hydroquinone degradation, and the degradation ratios of both were between 96.9%-97.4%.
引文
[1]史长林,李俊,姚宏伟等.含酚废水处理方法的研究[J].河北煤炭建筑工程学院学报, 1995, 8(4):4~9.
[2]王莉莉,杨孙楷.我国高浓度含酚废水的治理技术近况[J].环境污染与防治, 1995, 17(5):77~79.
[3]孔繁翔,严国安,尹大强.环境生物学[M].高等教育出版社, 2000
[4]于萍,姚琳,罗运柏.高浓度含酚废水处理的新工艺[J].工业水处理, 2002, 22(9):5~8.
[5]王红娟,奚红霞,夏启斌等.含酚废水处理技术的现状与开发前景[J].工业水处理, 2002, 22(6):6~9.
[6]刘相伟.工业含酚废水处理技术的现状与进展[J].工业水处理, 1998, 18(2): 4~6.
[7]庞晓坤.离子束诱变选育高效降酚微生物的研究[D].安徽:安徽农业大学出版社,2006.
[8]李湛江.硝基苯厌氧降解菌的选育及含硝基苯废水降解工艺研究[D].广州:华南理工大学出版社,1999.
[9]杨彦希,尹萍,杨惠芳.一株高效脱酚菌麦芽糖假丝酵母10-4的研究[J].微生物学通报, 1995, 22(4):208~211.
[10]罗国维,杨丹青,林世光.投菌生物接触氧化法处理洁霉素废水的机理研究[J].环境科学, 1994, 15(6):20~22.
[11] S. Buffet-Bataillon, V. Rabier, et al. Outbreak of Serratia marcescens in a neonatal intensive care unit: contaminated unmedicated liquid soap and risk factors[J]. Journal of Hospital Infection, 2009, 5(16):1~6.
[12]朱建国.临床常见细菌鉴定手册[M].北京:医科大学&中国协和医科大学联合出版社, 1993.
[13] Melissa M. Statham, Amit Vohra, Deepak K. Mehta, et al. Serratia marcescens causing cervical necrotizing oropharyngitis[J]. International Journal of Pediatric Otorhinolaryngology, 2009, 73:467~473.
[14] Gales A. C.,Biedenbach D. J.,Winokur P., et al. Carbapenem-resistant Serratia marcescens isolates producing Bush group 2fβ-lactamase (SME-1) in the United States: results from the MYSTIC ProgrammeDiagnostic[J]. Microbiology and Infectious Disease,2001, 39:125~127.
[15] Bidlan R., Manomani H. K.. Aerobic degradation of dichlorodiphenyltrichlor- oethane (DDT) by Serratia marcescens DT-1P[J]. Process Biochemistry, 2002, 38: 49~56.
[16] Ijah U. J. J.. Studies on relative capabilities of bacterial and yeast isolates from tropical soil in degrading crude oil[J]. Waste Management, 1998, 18:293~299
[17]周梅先,董钧锋.抗生素菌种选育的研究进展[J].生物学通报, 2006, 41(8): 3~4.
[18]赵化侨.等离子体化学和工艺[J],合肥:中国科学技术出版社, 1993.
[19]薛开先.国外医学:遗传学分册[M], 1998, 21(3):157~161.
[20]柴之芳.环境研究中的核科学技术战略研讨会纪要──核技术发展和科学基金[J].核技术, 2000, 23(6):431~432.
[21] Laoussi Mounir. Sterilization of contaminated matter with an atmospheric pressure plasma[J]. IEEE Transations on Plasma Science, 1996, 24:1188~1191.
[22] Garate Eusebio, Evans Kirk, Gornostaeva Olga, et al. The 1998 IEEE Internati- onal Conference on Plasma Science[J]. Raleigh, NC, USA, 1998.
[23] Georgiev I.V.,Bulgaranova Z.H.,Kumanova B..INT J Mater Prod Technol, 1995,10(3):522~529.
[24]凌代俊,曹金祥,鲁润龙等.一种新型物理诱变剂—低温空气等离子体的遗传毒性[J].核技术, 2001, 24(8): 668~673.
[25]凌代俊,曹金祥.低温等离子体诱导产生的一种“空洞”形细胞[J].核技术, 2003, 26(4): 284~286.
[26] Fisher J., Reeves E.A.,Isaac G.H.,et al. J Mater Sci Mater Meal Med,1997,8:375~378.
[27]余增亮,邵春林,杨剑波.离子刻蚀生物样品的初步研究[J].安徽农业大学学报, 1994, 26(3): 260~264.
[28]卫增泉,韩光武,周光明等.超低能离子注入作物育种的一种重要机制[J].生物物理学报, 1996, 12(3): 315~320.
[29]宋道军,余汛,韩建伟等.离子注入不同辐射敏感性微生物自由基与存活关系的研究[J].激光生物学学报, 1998, 7(4): 245~248.
[30]宋道军,姚建铭,邵春林等.离子注入微生物产生“马鞍型”存活曲线的可能作用机制[J].核技术. 1999, 22(3): 129~132.
[31] E. Garate, K. Evans, O. Gornostaeva, et al. Atmospheric plasma induced sterilization and chemical neutralization[J]. IEEE Int. Conf. Plasma Science, 1998:183.
[32] M. Rader, I. Alexeff, P. P. Tsai, et al. Electrostatic charging apparatus and method[P], U. S. Patent 5592357, 1997.
[33] M. Laroussi. Sterilization of tools and infectious waste by plasmas[J]. Bull. Amer. Phys. Soc. Div. Plasma Phys., 1995, 40(11):1685~1686.
[34] K. Kelly-Wintenberg, T.C. Montie, C. Brickman, et al. Room temperature sterilization of surfaces and fabrics with a one atmosphere uniform glowdischarge plasma[J]. J. Ind. Microbiol & Biotech. 1998, 20:69~74.
[35] H.W. Herrmann, I. Henins, J. Park, et al. Discharge physics and chemistry of a novel atmospheric pressure plasma source[J]. Phys. Plasmas, 1999, 6(5): 2284~2289.
[36] A. Scutze, Y. Jeong, S. E. Babyan, et al. The atmospheric-pressure plasma jet: a review and comparison to other plasma sources[J]. IEEE Trans. Plasma Sci., 1998, 26:1685~1694.
[37] J. P. Richardson, F.F. Dyer, F.C. Dobbs, et al. On the use of the resistive barrier discharge to kill bacteria: recent results[J]. IEEE Int. Conf. Plasma Science, 2000, 109.
[38] M. Laroussi, J. P. Richardson, and F. C. Dobbs. Biochemical pathways in the interaction of non-equilibrium plasmas with bacteria[J]. Proc. Electromed., Portsmouth, VA, 2001:33~34.
[39] A. I. Kuzmichev, I. A. Soloshenko, V. V. Tsiolko, et al. Feature of sterilization by different type of atmospheric pressure discharges[J]. High Pressure Low Temperatuer Plasma Chemisrty, Greifswald. Germany, 2001:402~406.
[40] Thomas Schultz, Elena Samoylova, Wolfgang Radloff, et al. Efficient Deactivation of a Model Base Pair via Excited-State Hydrogen Transfer[J]. Science, 2004, 306(3):1765~1768.
[41] Rami Ben Gadri, J. Reece Roth, et al. Sterilization and plasma processing of room temperature surfaces with a one atmosphere uniform glow discharge plasma (OAUGDP) [J]. Elsevier Surface and Coarings Technology, 2000, 131:528~542.
[42] M. Yamamoto, M. Nishioka, M. Sadakata. Proc. Sterilization using a corona discharge with H2O2 droplets and examination of effective species[J]. 15th Int. Symp. Plasma Chemistry, Vol.Ⅱ, Orleans, France, 2001:743~751.
[43] Alexandra Mess, Tobias Klar, Petra Gnau, et al. Crystal Structure of a Photolyase Bound to a CPD-Like DNA Lesion After in Situ Repair[J]. Science. 2004, 306(3): 1789~1793.
[44] M. Laroussi, F.Leipold. Evaluation of the roles of reactive species, heat, and UV radiation in the inactivation of bacterial cells by air plasmas at atmospheric pressure[J]. Intenrational Journal of Mass Spectrometry, 2004, 233:81~86.
[45] M. Laroussi, Senior Member. Nonthermal decontamination of biological media by atmospheric-pressure plasmas: review, analysis, and prospects[J]. IEEE Trans. Plasma Sci., 2002, 30( 4):1409~1415.
[46] M. Laroussi, D. A. Mendis, M. Rosenberg. Plasma interaction with microbes[J]. New Journal of Physics, 2003, 5:41.1~41.10.
[47]石兴民,郝炳华,袁育康等.气体等离子体对细菌损伤作用的电镜观察[J].中国消毒学杂志, 2003, 20(4): 284~286.
[48]王锡录,水野彰.一种新灭菌方法的研究[J].东北师大学报自然科学版, 1999, 6(2): 27~30.
[49]董春娟,吕丙南,陈志强等.处理生物难降解物质的有效方式——共代谢[J].化工环保, 2003, 23(2): 82~85.
[50]李萍,刘俊新.废水中难降解性有机污染物的共代谢降解[J].环境污染治理技术与设备, 2002, 3(11): 43~46.
[51]沈同,王镜岩.生物化学[M].北京:人民教育出版社, 1981.
[52]瞿福平,张晓健,何苗等.易降解有机物对氯苯好氧生物降解性能影响[J].中国环境科学, 1998, 18(5): 407~409.
[53]全向春,王建龙,韩力平等.喹啉与葡萄糖共基质条件下生物降解的动力学分析[J].环境科学学报, 2001, 21(4): 416~419.
[54] Gerald E., Speital Jr, Robert L., et al. Cometabolism in biofilm reactors[J]. Water Sci. and Technology, 1995, 31(l): 215~225.
[55] K. Miserez, S. Philips, W. Verstraete. New Biology for Advanced Wastewater Treatment[J]. Water Sci. and Technology, 1999, 40(4): 137~144.
[56]李华钟,章燕芳.白腐菌与染料废水的处理[J].工业水处理, 2001, 21(5): 1~5.
[57] Sandra L.. Polychlorinated Biphenyl Reductive Dechlorination by Vitamin B12s: Thermodynamics and Regiospecificity[J]. Environ. Sci. Technol, 1999, 33(6):857~863.
[58]张锡辉, R. Bajpai.以关键酶为基础共代谢模型的建立—以甲烷细菌共代谢三氯乙烯为例[J].环境科学学报, 2000, 20(5): 558~562.
[59]王永杰,李顺鹏,沈标等.有机磷农药广谱活性降解菌的分离及其生理特性研究[J].南京农业大学学报,1999, 22(2): 42~45.
[60]隋红,李鑫钥,段云霞等.三氯乙烯共代谢生物降解研究[J].农业环境科学学报,2004, 23(1):170~173.
[61]朱柱,李和平.固定化细胞技术处理含酚废水的研究[J].重庆环境科学, 2000, 22(6): 64~67.
[62]赵玲.焦化废水生化处理工艺的改进[J].工业水处理, 1999, 19(1): 44.
[63]吴立波,王建龙,黄霞等.自固定化高效菌种强化处理焦化废水研究[J].中国给水排水, 1999, 15(5): 1~4.
[64]罗国维,物丹青,林世光.投菌生物接触氧化法处理洁霉素废水的机理研究[J].环境科学, 1994, 15(6): 20~22.
[65] Ling Dai-jun, Cao Jin-xiang, Lu Run-long, et al. Genotoxicity of a new type ofnon-pollutant mutagen,low-temperature air plasma[J]. Nuclear Technique, 2001, 24(8):668~673
[66] Laroussi M., Richardson J. P., Dobbs F. C., et al. Effects of nonequilibrium atmospheric pressure plasmas on the heterotrompic pathways of bacteria and on their cell morphology[J]. Applied Physics Letters, 2002, 81(4): 772~776.
[67] Laroussi M., Leipold F. Evaluation of the roles of reactive species, heat, and UV radiation in the inactivation of bacterial cells by air plasmas at atmospheric pressure[J]. Intenrational Journal of Mass Specrtometry, 2004, 233: 81~86.
[68] Hou Ying-min. Induced Mutation of Strain Producing 1,3-Propanediol by Plasma[D]. Dalian: Dalian University of Technology, 2006.
[69]宋道军,余汛,余增亮.低能离子束对微生物细胞的直接作用和间接作用研究[J].高技术通讯, 1999, 1: 47~50.
[70] Xia Le-xian, Zeng Jia, Ding Jian-lan, et al. Comparison of three induced mutation methods for Acidiothiobacillus caldus in processing sphalerite[J]. 2007, Minerals Engineering, 20: 1323~1326.
[71] DeCaprio A.P.. The Toxicology of Hydroquinone—Relevance to Occupational and Environmental Exposure. Critical Reviews in Toxicology[J]. 1999, 29: 283~330.
[72] J.A. Daniel, M.Y. Chris, R.H. Michael. Molecular and cellular fundamentals of aerobic cometabolism of trichloroethylene[J]. Biodegradation , 2001, 12: 81~103.
[73] H.K. Michael, J.H. Oliver. Cometabolic degradation of chlorophenols by Acinetobacter species[J]. Water Res., 1999, 33: 562~574.
[74] M. Ziagova, K.M. Liakopoulou. Comparison of cometabolic degradation of 1,2-dichlorobenzene by Pseudomonas sp. and Staphylococcus xylosus.Enzyme Microb[J]. Technol., 2007, 40 (5): 1244~1250.
[75]姜燕,马毅,吴桂峰等.添加不同共代谢基质处理苯胺废水的研究[J].上海化工, 2007, 32(11): 1~3.
[76]李烨,刘菲,史敬华等.以醋酸为共代谢基质时四氯乙烯的的生物降解初步研究[J].水文地质工程地质, 2006, 3: 7~10.
[77]孙剑辉,梁志伟,孙靖宇.不同共代谢基质下厌氧生物降解间苯二酚的研究[J].环境科学与技术, 2007, 30(5): 24~26.
[78]姚珺,赵野,何苗.共代谢对难降解有机物生物降解性能的影响[J].环境科学与技术, 2006, 29 (3): 11~12.
[79]程丽华,黄君利,王丽等. Fenton试剂的特性及其在废水处理中的应用[J].化学工程师, 2001, 84(3): 24~25.