西北黄土地区石油污染土壤生物修复的菌种筛选与影响因素
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摘要
西北黄土地区的石油开发利用已有90多年的历史,该地区油田面积大,油井多,在石油的生产、贮运、炼制、加工及使用过程中,都会有石油烃类的溢出和排放,造成大面积的石油污染,使原本脆弱的自然环境更加恶化。本文在此背景下,筛选、分离得到高效降解石油的优势菌,对其进行了初步鉴定;并对影响其降解性能的营养条件和环境因素进行实验研究,主要研究成果如下:
1.通过筛选,得到五株在好氧条件下,对石油烃有较强降解能力的优势菌株GX1、GX2、GX3、GX4和GX5,经鉴定分别为假单胞菌属(Pseudomonas sp.)、邻单胞菌属(Plesionmonas sp.)、黄单胞菌属(Xanthomonas sp.)、假单胞菌属(Pseudomonas sp.)和动胶菌属(Zoogloea sp.)。
2.研究了pH值、营养物质、油质量浓度对菌体生长和除油率的影响,结果表明,这五株菌在初始pH值为7～9,最佳氮源为NH_4NO_3,氮磷比大约控制在4:1左右时,降解效果最好。在原油初始浓度较低时,除油率随石油烃浓度的升高而有微小的提高,当达到一定的浓度时除油率下降,不同菌株耐油程度不同。
3.添加氮磷营养盐对土壤中石油污染物的降解是必要的,激化了油污土壤中的土著菌,提高其微生物活性,使得最能适应环境的优势菌群开始急剧增加;接种了驯化培养的高效石油降解菌并添加其生长所需的营养盐,能极大地促进了石油烃的降解。
4.在降解初期,上层土壤降解效果较好,这是因为在初始湿度相差不大的情况下,上层土壤具有良好的透气性,进而增强微生物的活性,促进石油烃的生物降解。而在降解的中、后期,中下层土壤降解效果明显好于上层。这是由于中下层土壤含水量较高,微生物得到充分足的水分供应,促进细胞活性、加快代谢速率,从而增强降解效果。
5.微生物数量和脱氢酶活性与石油污染物生物降解之间存在良好的相关性,表明了
The exploitation and utilization of petroleum in northwest loess area has a long history of more than 90 years. Due to large aera of oil field and numerous oil wells, there are large quantity of waste petroleum hydrocarbon in the process of petroleum production, storage, transportation, refinement, processing and use. All of this can create oil pollution in large area, which causes the frail natural environment to be originally wicked. Under this background highly efficient petroleum-degrading bacteria were screened, isolated from oil-polluted soil, and they have been identified primarily. The nutritional conditions and the environmental factors which affect its degeneration performance were studied.The main conclusions of the study were as follows.1. Five strains of highly efficient petroleum degrading bacteria, GX1, GX2, GX3, GX4andGX5, were isolated from oil-polluted soil in aerobic condition. They have been identified respectively as Pseudomonas sp. Plesionmonas sp. Xanthomonas sp. Pseudomonas sp. and Zoogloea sp. .2. the pH value, nutrients and oil concentration that affect the bacteria growth and oil degradiation were studied. The experimental results indicated that the five bacterial strains was efficient under initial pH of 7~9, the best nitrogen source of ammonium nitrate (NH_4NO_3), the ratio of nitrogen/phosphorus of 4 :1. With the low initial concentration of petroleum, the degrading rate had a tiny increase along with increasing oil concentration;it decreased when the oil concentration reached a certain value. Different bacterial strains had different capability for petroleum tolerance.3. Regarding to petroleum hydrocarbon biodegradation, it is necessary to add nutrients to contaminated soil, which intensifed indigenous fungus in oiled soil and improved
its microorganism activity, at the same time increased predominant bacterial strains quantity;It promoted petroleum hydrocarbon biodegradation process by the addition of highly efficient petroleum degrading bacteria and its nutrients.4. In the initial period of the degradation, petroleum hydrocarbon biodegradation effect was better in the upper layer soil, because the upper layer of soil had the better permeability when the initial humidity was almost same. Then enhanced microorganism activy and promoted petroleum hydrocarbon biodegradation. But in the middle, last period, petroleum hydrocarbon biodegradation effect was better in the center, lower layer soil. This is because of the high moisture content in the lower layer soil, it supplied the full moisture content for the microorganism that promoted cell activeness and speeded up metabolism speed, thus enhanced degradation effect.5. The total number of microorganisms and the dehydrogenase activity were closely related to the petroleum hydrocarbon biodegradation, which indicated that they were both good indexes of the contaminated soil bioremediation. However the dehydrogenase activity which compared with the microorganism quantity can reflect microorganism's active condition in contaminated soil's bioremediation.
1.通过筛选,得到五株在好氧条件下,对石油烃有较强降解能力的优势菌株GX1、GX2、GX3、GX4和GX5,经鉴定分别为假单胞菌属(Pseudomonas sp.)、邻单胞菌属(Plesionmonas sp.)、黄单胞菌属(Xanthomonas sp.)、假单胞菌属(Pseudomonas sp.)和动胶菌属(Zoogloea sp.)。
2.研究了pH值、营养物质、油质量浓度对菌体生长和除油率的影响,结果表明,这五株菌在初始pH值为7～9,最佳氮源为NH_4NO_3,氮磷比大约控制在4:1左右时,降解效果最好。在原油初始浓度较低时,除油率随石油烃浓度的升高而有微小的提高,当达到一定的浓度时除油率下降,不同菌株耐油程度不同。
3.添加氮磷营养盐对土壤中石油污染物的降解是必要的,激化了油污土壤中的土著菌,提高其微生物活性,使得最能适应环境的优势菌群开始急剧增加;接种了驯化培养的高效石油降解菌并添加其生长所需的营养盐,能极大地促进了石油烃的降解。
4.在降解初期,上层土壤降解效果较好,这是因为在初始湿度相差不大的情况下,上层土壤具有良好的透气性,进而增强微生物的活性,促进石油烃的生物降解。而在降解的中、后期,中下层土壤降解效果明显好于上层。这是由于中下层土壤含水量较高,微生物得到充分足的水分供应,促进细胞活性、加快代谢速率,从而增强降解效果。
5.微生物数量和脱氢酶活性与石油污染物生物降解之间存在良好的相关性,表明了
The exploitation and utilization of petroleum in northwest loess area has a long history of more than 90 years. Due to large aera of oil field and numerous oil wells, there are large quantity of waste petroleum hydrocarbon in the process of petroleum production, storage, transportation, refinement, processing and use. All of this can create oil pollution in large area, which causes the frail natural environment to be originally wicked. Under this background highly efficient petroleum-degrading bacteria were screened, isolated from oil-polluted soil, and they have been identified primarily. The nutritional conditions and the environmental factors which affect its degeneration performance were studied.The main conclusions of the study were as follows.1. Five strains of highly efficient petroleum degrading bacteria, GX1, GX2, GX3, GX4andGX5, were isolated from oil-polluted soil in aerobic condition. They have been identified respectively as Pseudomonas sp. Plesionmonas sp. Xanthomonas sp. Pseudomonas sp. and Zoogloea sp. .2. the pH value, nutrients and oil concentration that affect the bacteria growth and oil degradiation were studied. The experimental results indicated that the five bacterial strains was efficient under initial pH of 7~9, the best nitrogen source of ammonium nitrate (NH_4NO_3), the ratio of nitrogen/phosphorus of 4 :1. With the low initial concentration of petroleum, the degrading rate had a tiny increase along with increasing oil concentration;it decreased when the oil concentration reached a certain value. Different bacterial strains had different capability for petroleum tolerance.3. Regarding to petroleum hydrocarbon biodegradation, it is necessary to add nutrients to contaminated soil, which intensifed indigenous fungus in oiled soil and improved
its microorganism activity, at the same time increased predominant bacterial strains quantity;It promoted petroleum hydrocarbon biodegradation process by the addition of highly efficient petroleum degrading bacteria and its nutrients.4. In the initial period of the degradation, petroleum hydrocarbon biodegradation effect was better in the upper layer soil, because the upper layer of soil had the better permeability when the initial humidity was almost same. Then enhanced microorganism activy and promoted petroleum hydrocarbon biodegradation. But in the middle, last period, petroleum hydrocarbon biodegradation effect was better in the center, lower layer soil. This is because of the high moisture content in the lower layer soil, it supplied the full moisture content for the microorganism that promoted cell activeness and speeded up metabolism speed, thus enhanced degradation effect.5. The total number of microorganisms and the dehydrogenase activity were closely related to the petroleum hydrocarbon biodegradation, which indicated that they were both good indexes of the contaminated soil bioremediation. However the dehydrogenase activity which compared with the microorganism quantity can reflect microorganism's active condition in contaminated soil's bioremediation.
引文
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[3] 中央电视台焦点访谈节目.“非法采油何时了”.2000.12.23
[4] 张旭,李广贺,黄巍.石油烃污染土层生物修复模拟实验研究[J].清华大学学报(自然科学版),2000,40(11):106～108
[5] 李晔,陈新才,王焰新.石油污染土壤生物修复的最佳生态条件研究[J].环境科学与技术,2004,27(4):17～19
[6] 夏北成.环境污染物生物降解[M].北京:化学工业出版社,2002.3
[7] J.H. Vandermelmen, A. Foda and C. Stuttard. Toxicity VS Mutagen city of some crude oils, Distillates and their water soluble fractions [J]. Water Res. 1985, 19(10) : 1283～1289
[8] Susan, C. et al. Bioremediation of soil contaminated with poly nuclear aromatic hydrocarbons (PAHs): A review environment [J]. Pollution, 1993, 81:229～249
[9] Hilberts, B. et al. In Site Techniques in Contaminated Soil. In: K. Wilf et al. (Eds.) [J]. Kluwer Aeddemic Publishers, 1985, 675～693
[10] Morgan, P. and Watkinson, R. J. Hydrocarbon degradation in soil and methods for soil biotreatment [J]. Biotechnology, 1989, 8:305～333
[11] America Petroleum Institute, the Land Treatability of Appendix Ⅶ Weshington, D.C. 1984
[12] 张久根等.国内土壤污染研究状况[J].环境科学研究,1990,3:56～60
[13] 沈德中.石油污染生化治理进展[J].国外科技,1992,9:40～43
[14] Zitrides, T.G. Effect of bioremediation on PAHs residues in soil [J]. Pollution Engineering, 1990, 22(5):57～62
[15] 丁克强,骆永明.生物修复石油污染土壤[J].土壤,2001,4:179～184
[16] Audrew R. et al. Environment Progress, 1992, 11 (40): 318～322
[17] 焦瑞身等编著,生物工程概论.化学工业出版社,1991,85～87
[18] 李慧容,白腐真菌的研究进展.环境科学进展,1996,4(6):69～77
[19] 齐永强,王红旗.微生物处理土壤石油污染的研究进展[J].上海环境科学,2002,21(3):177～180
[20] Salanitro JP. Crude oil hydrocarbon bioremediation and soil ecotoxicity assessment [J]. Environ SciTech, 1997, 31(6): 1769～1776
[21] JM亨特著.胡伯良译.石油地球化学和地质学[M1.北京:石油工业出版社,1986
[22] Robert M Garrett. Photo oxidation of crude oils [J]. Environ Sci Technal, 1998, 32:3719～3723
[23] angehrn D. Movement and fate of residual internal oil contaminants in bioremediated soil [J]. Environ Toxicol Chem, 1999, 18(10): 2225～2231
[24] Row land A P. Effects of beach sand properties, temperature and rainfall on the degradation rates of oil in buried beach sand mixtures [J]. Environ Pollut, 2000, 109(1): 109～118
[25] Rhykerd RL. Influence of Salinity on bioremediation of oil in soil [J]. Environ Pollut, 1995, 90(1): 127～130
[26] A Haiim. Growth of hydrocarbon-utilizing isolated in chemically defined media [J]. International Biodeterioration & Biodegradation, 1996, 37(12): 61～68
[27] Lin Q. Effects of bioremediation agents on oil degradation in mineral and sandy salt marsh sediments [J]. Environ Technal, 1999, 18(10): 825～837
[28] Brink R H. Biodegradation of organic chemicals in the enviornment [A]. In: McKinney JD, ed. Environmengt Health Chemistry [C]. Lancaster, PA:Technomic Publishing Company, 1981, 245～264
[29] Gibson D T.Microbial metabolism [A]. In: Hutzinger O, ed.The Handbook of Enviornment chemistry, Volume 2.Part A, Reactions and Processes[C]. New York, NY: Springer-Verlag, 1980, 344～356
[30] Benka-Coker M O. Applicability of evaluating the ability of michobes isloted from and oil spill site to degrade oil [J]. Environ Monit Assess, 1997, 45(3): 259～272
[31] Carriere PEP. Enhanced biodegradation of creosote-contaminated soil [J]. Waste Management, 1995, 579～583
[32] 魏德洲,秦煜民.H_2O_2在石油污染土壤微生物治理过程中的作用[J].中国环境科学,1997,17(5):429～431
[33] Chaineau CH. Land treatment of oil-based drill cuttings in an agricultural soil [J]. Environ Qual, 1996, 25(4): 858～867
[34] Geerdink M J. Microbial decontamination of polluted soil in a slurry process [J]. Environ Eng, 1996, 122(11): 975～982
[35] 全国政协人口资源环境委员会.西部大开发与水资源文集[C].北京:中国水利电力出版社,2000:239
[36] Cahaway, J.W. and Doyle, J.R. Innovative remedial action at awood-treating superfund site [J]. Tappi Journal, 1991, 74:113～118
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