原油降解菌株HC13的降解特性研究及其条件优化
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摘要
为减少原油污染的危害,各国学者都在研究利用微生物降解环境中的原油污染物.该实验以原油为唯一碳源制备培养基,采用平板菌落分离法,分离获得11株能够利用原油生长的菌株,重量法测得其中HC13菌株的原油降解率最高,为57%.16S rRNA基因序列初步鉴定其为假单胞菌(Pseudomonas sp.).气相与质谱联用(GC-MS)分析发现菌株HC13对短直链烷烃的降解率都在60%以上.结果表明:控制单一变量可得到其生长最适条件为pH值为7,温度为30℃,原油质量分数为1.5%,接种量为2%,转速为140 rpm.
In order to reduce the harm of the crude oil pollution, scholars around the world are studying the use of microbial degradation of crude oil pollutants in the environment.In this experiment,crude oil was used as the sole carbon source to prepare the culture medium.11 strains which are capable of growing from crude oil were isolated from crude oil by plate colony separation method.Results show that the HC13 strain enjoy the highest crude oil degradation rate of 57%.The 16 S rRNA gene sequence was initially identified as Pseudomonas sp.Gas phase and mass spectrometry(GC-MS) analysis shows that the degradation rate of strain HC13 on short linear paraffins is above 60%.The results show that optimum conditions for growth was obtained by controlling the single variable are as follows:pH 7,temperature 30 ℃,crude oil mass concentration 1.5%,inoculum volume 2%,and rotation speed 140 rpm.
In order to reduce the harm of the crude oil pollution, scholars around the world are studying the use of microbial degradation of crude oil pollutants in the environment.In this experiment,crude oil was used as the sole carbon source to prepare the culture medium.11 strains which are capable of growing from crude oil were isolated from crude oil by plate colony separation method.Results show that the HC13 strain enjoy the highest crude oil degradation rate of 57%.The 16 S rRNA gene sequence was initially identified as Pseudomonas sp.Gas phase and mass spectrometry(GC-MS) analysis shows that the degradation rate of strain HC13 on short linear paraffins is above 60%.The results show that optimum conditions for growth was obtained by controlling the single variable are as follows:pH 7,temperature 30 ℃,crude oil mass concentration 1.5%,inoculum volume 2%,and rotation speed 140 rpm.
引文
[1] SALANITRO J P,DORN P B,HUESEMANN M H ,et al.Crude oil hydrocarbon bioremediation and soil ecotoxicity assessment[J].Environmental Science & Technology,1997,31(6):1769-1776.
[2] PERRY J J.Microbial metabolism of cyclic alkanes[J].Petroleum Microbiology,1984:61-98.
[3] 李丽,张利平,张元亮.石油烃类化合物降解菌的研究概况[J].微生物学通报,2001,28(5):89-92.
[4] KVENVOLDEN K A,COOPER C K.Natural seepage of crude oil into the marine environment[J].Geo-Marine Letters,2003,23(3/4):140-146.
[5] 郭静仪.含油废水的生物处理[D].广州:暨南大学,2005.
[6] 刘明,孙鹏,张祥胜.原油降解菌YSL28的分离鉴定及降解特性研究[J].东北农业大学学报,2013,44(5):47-50.
[7] 谢鲲鹏,宫正,赵慧,等.高效原油降解菌的筛选及其降解能力的研究[J].辽宁师范大学学报(自然科学版),2005,28(2):228-231.
[8] 杨蕊琪,薛林贵,常思静,等.祁连山不同海拔低温原油降解菌群的分布特性研究[J].冰川冻土,2016,38(3):785-793.
[9] MEDINA-BELLVER J I,MARíN P,DELGADO A,et al.Evidence for in situ crude oil biodegradation after the Prestige oil spill[J].Environmental Microbiology,2010,7(6):773-779.
[10] ZOBELL C E.Action of microorganisms on hydrocarbons[J].Bacteriological Reviews,1946,10(1/2):1.
[11] DAS K,MUKHERJEE A K.Crude petroleum-oil biodegradation efficiency of Bacillus subtilis and Pseudomonas aeruginosa strains isolated from a petroleum-oil contaminated soil from north-east India[J].Bioresour Technol,2007,98(7):1339-1345.
[12] 陈宇.石油烃降解菌降解性能、机理与应用研究[D].天津:天津大学,2014.
[13] THRONE-HOLST M,WENTZEL A,ELLINGSEN T E,et al.Identification of novel genes involved in Long-Chain n-Alkane degradation by Acinetobacter sp.Strain DSM 17874[J].Applied and Environmental Microbiology,2007,73(10):3327-3332.
[14] 贾燕,尹华,彭辉,等.石油降解菌株的筛选、初步鉴定及其特性[J].暨南大学学报(自然科学版),2007,28(3):296-301.
[15] WANG W,WANG L,SHAO Z.Diversity and abundance of oil-degrading bacteria and alkane hydroxylase (alkB) genes in the subtropical seawater of Xiamen island[J].Microbial Ecology,2010,60(2):429-439.
[16] CUI Z,TAN T,LAI Q,et al.A pyrene-degrading consortium from deep-sea sediment of the West Pacific and its key member Cycloclasticus sp.P1[J].Environmental Microbiology,2010,10(8):1948-1963.
[17] 杨蕊琪.青藏高原土壤中原油降解细菌的多样性及降解特性研究[D].兰州:兰州交通大学,2016.
[18] KUSKE C R,BARNS S M,BUSCH J D.Diverse uncultivated bacterial groups from soils of the arid Southwestern United States that are present in many geographic regions[J].Applied and Environmental Microbiology,1997,63(9):3614-3621.
[19] ALAALEH E,DROBIOVA H,OBUEKWE C.Predominant culturable crude oil-degrading bacteria in the coast of Kuwait[J].International Biodeterioration & Biodegradation,2009,63(4):400-406.
[20] GENNARO P D,MORENO B,ANNONI E,et al.Dynamic changes in bacterial community structure and in naphthalene dioxygenase expression in vermicompost-amended PAH-contaminated soils[J].Journal of Hazardous Materials,2009,172(2/3):1464-1469.
[21] 王博.土壤石油降解菌的筛选与微生物生态修复研究[D].南京:南京农业大学,2012.
[22] WANG X B,CHI C Q,NIE Y,et al.Degradation of petroleum hydrocarbons (C6-C40) and crude oil by a novel Dietzia strain[J].Bioresour Technology,2011,102(17):7755-7762.
[23] AISLABIE J,SAUL D J,FOGHT J M.Bioremediation of hydrocarbon-contaminated polar soils[J].Extremophiles,2006,10(3):171-179.
[24] LIU Q,TANG J,BAI Z,et al.Distribution of petroleum degrading genes and factor analysis of petroleum contaminated soil from the Dagang Oilfield,China[J].Scientific Reports,2015,5:11068.
[25] BRAKSTAD O G,LODENG A G G.Microbial diversity during biodegradation of crude oil in seawater from the North Sea[J].Microbial Ecology,2005,49(1):94-103.
[26] WEISBURG W G,BARNS S M,PELLETIER D A,et al.16S ribosomal DNA amplification for phylogenetic study[J].Journal of Bacteriology,1991,173(2):697-703.
[27] AYOTAMUNO M J,KOGBARA R B,OGAJI S O T,et al.Bioremediation of a crude-oil polluted agricultural-soil at Port Harcourt,Nigeria[J].Applied Energy,2006,83(11):1249-1257.
[28] 王艺霖.青藏高原土壤中低温高效原油降解菌的筛选、鉴定及其降解特性的研究[D].兰州:兰州交通大学,2015.
[29] 刘庆龙,唐景春,万晓彤.污染土壤中主要石油降解基因AlkB和Nah定量检测方法的建立和应用[J].分析化学,2014,42(9):1348-1353.
[30] KNIEMEYER O,HEIDER J.Ethylbenzene dehydrogenase,a novel hydrocarbon-oxidizing molybdenum/ iron-sulfur/heme enzyme[J].Journal of Biological Chemistry,2001,276(24):21381-6.
[31] GODINI K,SAMARGHANDI M R,ZAFARI D,et al.Isolation and identification of new strains of crude oil degrading bacteria from Kharg Island,Iran[J].Petroleum Science and Technology,2018:1-6.
[32] PARTHIPAN P,ELUMALAI P,TING Y P,et al.Characterization of hydrocarbon degrading bacteria isolated from Indian crude oil reservoir and their influence on biocorrosion of carbon steel API 5LX[J].International Biodeterioration & Biodegradation,2018,129:67-80.
[33] 赵姣,屈撑囤,鱼涛,等.高效原油降解菌的分离鉴定及降解特性分析[J].油田化学,2017,34(3):532-537.
[34] 信艳娟,吴佩春,曹旭鹏,等.大连湾原油降解菌的分离和多样性分析[J].微生物学通报,2013,40(6):979-987.
[2] PERRY J J.Microbial metabolism of cyclic alkanes[J].Petroleum Microbiology,1984:61-98.
[3] 李丽,张利平,张元亮.石油烃类化合物降解菌的研究概况[J].微生物学通报,2001,28(5):89-92.
[4] KVENVOLDEN K A,COOPER C K.Natural seepage of crude oil into the marine environment[J].Geo-Marine Letters,2003,23(3/4):140-146.
[5] 郭静仪.含油废水的生物处理[D].广州:暨南大学,2005.
[6] 刘明,孙鹏,张祥胜.原油降解菌YSL28的分离鉴定及降解特性研究[J].东北农业大学学报,2013,44(5):47-50.
[7] 谢鲲鹏,宫正,赵慧,等.高效原油降解菌的筛选及其降解能力的研究[J].辽宁师范大学学报(自然科学版),2005,28(2):228-231.
[8] 杨蕊琪,薛林贵,常思静,等.祁连山不同海拔低温原油降解菌群的分布特性研究[J].冰川冻土,2016,38(3):785-793.
[9] MEDINA-BELLVER J I,MARíN P,DELGADO A,et al.Evidence for in situ crude oil biodegradation after the Prestige oil spill[J].Environmental Microbiology,2010,7(6):773-779.
[10] ZOBELL C E.Action of microorganisms on hydrocarbons[J].Bacteriological Reviews,1946,10(1/2):1.
[11] DAS K,MUKHERJEE A K.Crude petroleum-oil biodegradation efficiency of Bacillus subtilis and Pseudomonas aeruginosa strains isolated from a petroleum-oil contaminated soil from north-east India[J].Bioresour Technol,2007,98(7):1339-1345.
[12] 陈宇.石油烃降解菌降解性能、机理与应用研究[D].天津:天津大学,2014.
[13] THRONE-HOLST M,WENTZEL A,ELLINGSEN T E,et al.Identification of novel genes involved in Long-Chain n-Alkane degradation by Acinetobacter sp.Strain DSM 17874[J].Applied and Environmental Microbiology,2007,73(10):3327-3332.
[14] 贾燕,尹华,彭辉,等.石油降解菌株的筛选、初步鉴定及其特性[J].暨南大学学报(自然科学版),2007,28(3):296-301.
[15] WANG W,WANG L,SHAO Z.Diversity and abundance of oil-degrading bacteria and alkane hydroxylase (alkB) genes in the subtropical seawater of Xiamen island[J].Microbial Ecology,2010,60(2):429-439.
[16] CUI Z,TAN T,LAI Q,et al.A pyrene-degrading consortium from deep-sea sediment of the West Pacific and its key member Cycloclasticus sp.P1[J].Environmental Microbiology,2010,10(8):1948-1963.
[17] 杨蕊琪.青藏高原土壤中原油降解细菌的多样性及降解特性研究[D].兰州:兰州交通大学,2016.
[18] KUSKE C R,BARNS S M,BUSCH J D.Diverse uncultivated bacterial groups from soils of the arid Southwestern United States that are present in many geographic regions[J].Applied and Environmental Microbiology,1997,63(9):3614-3621.
[19] ALAALEH E,DROBIOVA H,OBUEKWE C.Predominant culturable crude oil-degrading bacteria in the coast of Kuwait[J].International Biodeterioration & Biodegradation,2009,63(4):400-406.
[20] GENNARO P D,MORENO B,ANNONI E,et al.Dynamic changes in bacterial community structure and in naphthalene dioxygenase expression in vermicompost-amended PAH-contaminated soils[J].Journal of Hazardous Materials,2009,172(2/3):1464-1469.
[21] 王博.土壤石油降解菌的筛选与微生物生态修复研究[D].南京:南京农业大学,2012.
[22] WANG X B,CHI C Q,NIE Y,et al.Degradation of petroleum hydrocarbons (C6-C40) and crude oil by a novel Dietzia strain[J].Bioresour Technology,2011,102(17):7755-7762.
[23] AISLABIE J,SAUL D J,FOGHT J M.Bioremediation of hydrocarbon-contaminated polar soils[J].Extremophiles,2006,10(3):171-179.
[24] LIU Q,TANG J,BAI Z,et al.Distribution of petroleum degrading genes and factor analysis of petroleum contaminated soil from the Dagang Oilfield,China[J].Scientific Reports,2015,5:11068.
[25] BRAKSTAD O G,LODENG A G G.Microbial diversity during biodegradation of crude oil in seawater from the North Sea[J].Microbial Ecology,2005,49(1):94-103.
[26] WEISBURG W G,BARNS S M,PELLETIER D A,et al.16S ribosomal DNA amplification for phylogenetic study[J].Journal of Bacteriology,1991,173(2):697-703.
[27] AYOTAMUNO M J,KOGBARA R B,OGAJI S O T,et al.Bioremediation of a crude-oil polluted agricultural-soil at Port Harcourt,Nigeria[J].Applied Energy,2006,83(11):1249-1257.
[28] 王艺霖.青藏高原土壤中低温高效原油降解菌的筛选、鉴定及其降解特性的研究[D].兰州:兰州交通大学,2015.
[29] 刘庆龙,唐景春,万晓彤.污染土壤中主要石油降解基因AlkB和Nah定量检测方法的建立和应用[J].分析化学,2014,42(9):1348-1353.
[30] KNIEMEYER O,HEIDER J.Ethylbenzene dehydrogenase,a novel hydrocarbon-oxidizing molybdenum/ iron-sulfur/heme enzyme[J].Journal of Biological Chemistry,2001,276(24):21381-6.
[31] GODINI K,SAMARGHANDI M R,ZAFARI D,et al.Isolation and identification of new strains of crude oil degrading bacteria from Kharg Island,Iran[J].Petroleum Science and Technology,2018:1-6.
[32] PARTHIPAN P,ELUMALAI P,TING Y P,et al.Characterization of hydrocarbon degrading bacteria isolated from Indian crude oil reservoir and their influence on biocorrosion of carbon steel API 5LX[J].International Biodeterioration & Biodegradation,2018,129:67-80.
[33] 赵姣,屈撑囤,鱼涛,等.高效原油降解菌的分离鉴定及降解特性分析[J].油田化学,2017,34(3):532-537.
[34] 信艳娟,吴佩春,曹旭鹏,等.大连湾原油降解菌的分离和多样性分析[J].微生物学通报,2013,40(6):979-987.