鼠李糖脂产生菌M7-6在模拟油藏条件下的培养基优化
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摘要
基于已筛选出的鼠李糖脂产生菌M7-6,在模拟油藏条件(温度、pH、矿化度及缺氧)下,对该菌株的激活剂配方进行了碳源、氮源、碳氮比(C/N)、无机盐等因素的优化,并考察了该菌株在模拟油藏条件下的最佳接种量;利用厌氧发酵罐对菌株M7-6进行了扩大培养,评价菌M7-6的原位代谢活性及与其他微生物类群的竞争作用。结果表明:以甘油为碳源、硝酸盐为氮源、C/N为14.4∶1时,最利于菌株M7-6在模拟油藏条件下生产鼠李糖脂,最小接种量为1%(体积分数)。在厌氧发酵罐中,菌株M7-6可以将培养体系的表面张力降至38.4 mN/m;并且体系中烃降解菌和产酸菌数量有所增加,而硫酸盐还原菌数逐渐减少。
Based on a rhamnolipid producing bacterial strain Pseudomonas aeruginosa M7-6,medium( carbon source,nitrogen source,C/N ratio,and mineral factors) was optimized for efficiently producing rhamnolipid under simulated oil reservoir conditions( temperature,p H and absence of oxygen). Using the optimized medium,effect of different inoculums amount on anaerobic production of rhamnolipid by M7-6was studied. Strain M7-6 was cultured in a 6-L anaerobic fermentor using oilfield production water to prepare medium,to evaluate the rhamnolipid production by M7-6 under simulated conditions and study the competitive activity of M7-6 with other native microorganisms. M7-6 could efficiently produce rhamnolipid under simulated conditions with glycerol as carbon source and nitrate as nitrogen source. Theoptimum C/N ratio is 14. 4 ∶ 1,and the least inoculums amount is 1%( V/V). Strain M7-6 can reduce surface tension of culture system to 38. 4 m N/m in the anaerobic fermenter. Moreover,the numbers of oil degrading bacteria and acid producing bacteria were increased,and the number of sulfate reducing bacteria was decreased.
Based on a rhamnolipid producing bacterial strain Pseudomonas aeruginosa M7-6,medium( carbon source,nitrogen source,C/N ratio,and mineral factors) was optimized for efficiently producing rhamnolipid under simulated oil reservoir conditions( temperature,p H and absence of oxygen). Using the optimized medium,effect of different inoculums amount on anaerobic production of rhamnolipid by M7-6was studied. Strain M7-6 was cultured in a 6-L anaerobic fermentor using oilfield production water to prepare medium,to evaluate the rhamnolipid production by M7-6 under simulated conditions and study the competitive activity of M7-6 with other native microorganisms. M7-6 could efficiently produce rhamnolipid under simulated conditions with glycerol as carbon source and nitrate as nitrogen source. Theoptimum C/N ratio is 14. 4 ∶ 1,and the least inoculums amount is 1%( V/V). Strain M7-6 can reduce surface tension of culture system to 38. 4 m N/m in the anaerobic fermenter. Moreover,the numbers of oil degrading bacteria and acid producing bacteria were increased,and the number of sulfate reducing bacteria was decreased.
引文
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[30]王靖,章厚名,安明泉,等.高效产表面活性剂菌株(Lz2-1)的筛选及其特性研究[J].石油化工高等学校学报,2009,23(3):33-36.
[2]赵国文,张丽萍,白利涛.生物表面活性剂及其应用[J].日用化学工业,2010,40(4):293-295.
[3]杜瑾,郝建安,张晓青,等.微生物合成鼠李糖脂生物表面活性剂的研究进展[J].化学与生物工程,2015,32(4):5-11.
[4]李俊峰,刘丽.脂肽类生物表面活性剂的研究进展[J].化学与生物工程,2015,32(1):12-15.
[5]吴柏志,李宜强.PBS菌的趋化性与提高原油采收率理解[J].环境科学,2005,26(5):132-136.
[6]赵玲莉,高雁,张涛,等.微生物驱油技术在克拉玛依油田的应用[J].石油化工应用,2015,34(9):40-42.
[7]王冠,崔延杰,张凯,等.弱凝胶驱后高含水稠油油藏微生物提高采收率技术研究及应用[J].长江大学学报(自科版),2015,12(11):74-79.
[8]班允赫,张潆月,史荣久,等.一株快速产脂肽解淀粉芽孢杆菌的筛选及其产物特性[J].生态学杂志,2015,34(6):1682-1688.
[9]张清军,张敏,王京秀,等.稠油污水原位升级达标现场试验研究[J].化学与生物工程,2013,30(8):63-69.
[10]冯海柱,程武刚,陈刚,等.生物表面活性剂提高采收率技术室内研究[J].当代化工,2015,44(2):243-244.
[11]汪卫东,汪竹,耿雪莉,等.美国微生物采油技术现场应用效果分析[J].油气地质与采收率,2002,9(6):75-76.
[12]乐建君,于盛鸿,崔长海,等.低渗透油田微生物采油现场试验研究[J].精细石油化工进展,2004,5(2):13-15.
[13]伍晓林,赵玲侠,乐建君,等.大庆油田聚驱后油藏内源微生物激活剂的筛选和效果评价[J].南开大学学报(自然科学版),2012,45(4):105-110.
[14]梁小龙,赵峰,史荣久,等.原生质体融合构建高效产脂肽工程菌[J].中国生物工程杂志,2014,34(11):76-84.
[15]赵峰.厌氧产鼠李糖脂S2-氧化工程菌的构建及微生物驱油潜力评价[D].北京:中国科学院大学,2013.
[16]ZHAO F,SHI R,ZHAO J,et al.Heterologous production of Pseudomonas aeruginosa rhamnolipid under anaerobic conditions for microbial enhanced oil recovery[J].J Appl Microbiol,2015,118(2):379-389.
[17]ZHAO F,MANDLAA M,HAO J,et al.Optimization of culture medium for anaerobic production of rhamnolipid byrecombinant Pseudomonas stutzeri Rhl for microbial enhanced oil recovery[J].Lett Appl Microbiol,2014,59(2):231-237.
[18]李万峰,颜学军.大肠菌群最大可能数(MPN)的数学推导与计算机程序的应用[J].中国现代医学杂志,2009,19(6):923-925.
[19]刘淑艳,马惠蕊,蒋丹,等.最可能数的精确计算及其在食品微生物检验中的应用[J].中国卫生统计,201l,28(10):516-519.
[20]YOUSSEF N H,DUNCAN K E,NAGLE D P,et al.Comparison of methods to detect biosurfactant production by diverse microorganisms[J].J Microbiol Methods,2004,56:339-347.
[21]孙琳,蒲万芬,吴雅丽.表面活性剂高温乳化性能研究[J].油田化学,2011,28(3):275-279.
[22]ZHAO F,ZHANG J,SHI R,et al.Production of biosurfactant by a Pseudomonas aeruginosa isolate and its applicability to in situ microbial enhanced oil recovery under anoxic conditions[J].RSC Adv,2015,45(5):36044-36050.
[23]顾生辉,朱莉,詹晓北,等.以甘油为底物鼠李糖脂高产菌株的诱变选育[J].生物加工过程,2015,13(1):54-59.
[24]孙全莲.铜绿假单胞菌的筛选及产鼠李糖脂发酵条件的研究[D].西安:西北大学,2014.
[25]夏文杰,董汉平,俞理.鼠李糖脂发酵条件优化和采油应用研究[J].深圳大学学报(理工版),2010,27(4):482-489.
[26]梁生康,王修林,单宝田.假单胞O-2-2利用油脂废水生产鼠李糖脂研究[J].现代化工,2005,25(增刊1):192-196.
[27]WU J Y,YEH K L,LU W B,et al.Rhamnolipid production with indigenous Pseudomonas aeruginosa EM1 isolated from oilcontaminated site[J].Bioresour Technol,2008,99(5):1157-1164.
[28]SOBERON-CHAVEZ G,LEPINE F,DEZIEL E.Production of rhamnolipids by Pseudomonas aeruginosa[J].Appl Microbiol Biotechnol,2005,68(6):718-725.
[29]GHOJAVAND H,VAHABZADEH F,AZIZMOHSENI F.A halotolerant,thermotolerant,and facultative biosurfactant producer:identification and molecular characterization of a bacterium and evolution of emulsifier stability of a lipopeptide biosurfactant[J].Biotechnol Bioproc Eng,2011,16:72-80.
[30]王靖,章厚名,安明泉,等.高效产表面活性剂菌株(Lz2-1)的筛选及其特性研究[J].石油化工高等学校学报,2009,23(3):33-36.