石林喀斯特碳酸钙高效沉积菌的筛选及其沉积条件优化
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摘要
"微生物水泥"为重要建筑和珍贵石质文物的保护提供了更优越的解决方案,但目前我国缺乏具有自主知识产权的高效沉积碳酸钙的菌株.研究从云南石林喀斯特的土壤和风化物样品中分离到细菌856株,其中185株具有脲酶活性,29株具有明显的碳酸钙沉积能力,6株能高效沉积碳酸钙.根据菌株生理特性,选取SL313进行沉积条件的响应面优化.根据16S rRNA基因相似性,SL313初步鉴定为Staphylococcus xylosus,目前未见该种菌株的碳酸钙沉积研究.响应面优化后的条件为p H=6.00,尿素126 mg/m L,硝酸钙344.4 mg/m L,胰蛋白胨25 g/L,碳酸钙沉积量为0.165 g/m L,比优化前提高了17.9%.对沉积物的X射线衍射分析表明其为球霰石晶体,扫描电镜显示沉积物呈球状堆积,表面偶见菌体.本研究为"微生物水泥"的开发提供了大量备选菌株,也为喀斯特微生物资源利用提供了新的思路.
Microbe cement provides superior solutions for the protection of important buildings and precious stone relics,few independent intellectual property rights of the efficient calcium carbonate mineralization strain in China.In this study,856 strains of bacteria were isolated from soil and weathered products in Shilin Karsts of Yunnan,185 of them had urease activity,29 of them could induce the formation of calcium carbonate,6 of them could induce the formation of calcium carbonate efficiently. According to the physiological characteristics of the strain,SL313 was selected to optimize the medium composition by using response surface methodology.SL313 was identified as Staphylococcus xylosus,which has never been studied about the calcium carbonate deposition before.The results showed that the optimal condition was p H = 6. 00,urea 126 mg/m L,calcium nitrate 344. 4 mg/m L,tryptone 25 g/L,and the Ca CO3 production reached 0.165 g/m L,which was 17.9% higher than that under the initial culture conditions.Scanning electron microscopy and X-ray diffraction analysis was used to observe and study the crystal structure and chemical composition of the formed calcium carbonate. The results showed that SL313 could accelerate the formation of vaterite crystal. This study provides a large number of candidate strains for the development of microbial cement and new ideas for the utilization of microbial resources in Karsts.
Microbe cement provides superior solutions for the protection of important buildings and precious stone relics,few independent intellectual property rights of the efficient calcium carbonate mineralization strain in China.In this study,856 strains of bacteria were isolated from soil and weathered products in Shilin Karsts of Yunnan,185 of them had urease activity,29 of them could induce the formation of calcium carbonate,6 of them could induce the formation of calcium carbonate efficiently. According to the physiological characteristics of the strain,SL313 was selected to optimize the medium composition by using response surface methodology.SL313 was identified as Staphylococcus xylosus,which has never been studied about the calcium carbonate deposition before.The results showed that the optimal condition was p H = 6. 00,urea 126 mg/m L,calcium nitrate 344. 4 mg/m L,tryptone 25 g/L,and the Ca CO3 production reached 0.165 g/m L,which was 17.9% higher than that under the initial culture conditions.Scanning electron microscopy and X-ray diffraction analysis was used to observe and study the crystal structure and chemical composition of the formed calcium carbonate. The results showed that SL313 could accelerate the formation of vaterite crystal. This study provides a large number of candidate strains for the development of microbial cement and new ideas for the utilization of microbial resources in Karsts.
引文
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[2]KANTZAS A,STEHMEIER L,MARENTETTE D F,et al.A novel method of sand consolidation through bacteriogenic mineral plugging[C].Proc of CIM Annual Technical Conference Calgary Canada:Petroleum Society of Canada,1992:1-15.
[3]钱春香,王欣,於孝牛.微生物水泥研究与应用进展[J].材料工程,2015,43(8):92-103.QIAN C X,WANG X,YU X N.Research and application development of microbe cement[J].Journal of Materials Engineering,2015,43(8):92-103.
[4]陶犁.徐霞客与云南喀斯特旅游资源[J].云南教育学院学报,1995,11(2):77-80.TAO L.Xu xiake and Yunnan karst tourism resources[J].Journal of Yunnan College of Education,1995,11(2):77-80.
[5]竹文坤,牟涛,段涛,等.响应面法优化微生物诱导碳酸钙沉积培养基[J].化工进展,2014,33(6):1 533-1 538.ZHU W K,MU T,DUAN T,et al.Optimization for microbiological-induced Ca CO3precipitation medium using response surface methodology[J].Chemical Industry and Engineering Progress,2014,33(6):1 533-1 538.
[6]竹文坤,罗学刚.碳酸盐矿化茵诱导碳酸钙沉淀条件的优化[J].非金属矿,2012,35(3):1-4.ZHU W K,LUO X G.Optimization of calcium carbonate precipitation for carbonate mineralization bacteria[J].Non-Metallic Mines,20l2,35(3):1-4.
[7]贾强,陈晓瀚,孙增斌,等.微生物诱导沉积碳酸钙提高产量的试验研究[J],山东建筑大学学报,2015,30(5):423-428.JIA Q,CHEN X H,SUN Z B,et al.The raising calcite yield experiment for bacteria induced precipitation[J].Journal of Shandong Jianzhu University,2015,30(5):423-428.
[8]成亮,钱春香,王瑞兴,等.碳酸岩矿化菌诱导碳酸钙晶体形成机理研究[J].化学学报,2007,65(19):2 133-2 138.CHENG L,QIAN C X,WANG R X,et al.Study on the mechanism of calcium carbonate formation induced by carbonate-mineralization microbe[J].Acta Chimica Sinica,2007,65(19):2 133-2 138.
[9]成亮,钱春香,王瑞兴,等.碳酸盐矿化菌调控碳酸钙结晶动力学、形态学的研究[J].功能材料,2007,38(9):1 511-1 515.CHENG L,QIAN C X,WANG R X,et al.Study on kinetics and morphology of formation of Ca CO3crystal induced by carbonate-mineralization microbe[J].Journal of Functional Materials,2007,38(9):1 511-1 515.
[10]李辉,连宾,龚国洪,等.碳酸钙颗粒的细菌诱导形成[J].高校地质学报,2011,17(1):112-117.LI H,LIAN B,GONG G H,et al.The formation of calcium carbonate particles induced by bacteria[J].Geological Journal of China Universities,2011,17(1):112-117.
[11]李沛豪,屈文俊.细菌诱导矿化保护历史建筑遗产的机理及效果[J].硅酸盐学报,2009,37(4):497-505.LI P H,QU W J.Mechanism and performance of remediation for historic buildings by bacterially induced mineralization[J].Journal of the Chinese Ceramic Society,2009,37(4):497-505.
[12]尹晓爽,张慧,杨文忠,等.一种假单胞菌诱导Ca CO3结晶[J].应用化学,2010,27(8):911-915.YIN X S,ZHANG H,YANG W Z.et al.Calcium carbonate formation induced by Pseudomonas[J].Chinese Journal of Applied Chemistry,2010,27(8):911-915.
[13]李沛豪,屈文俊.细菌诱导碳酸钙矿化材料及其应用前景[J].建筑材料学报,2009,12(4):482-486.LI P H,QU W J.Applications of calcium carbonate mineralization material induced by bacterium[J].Journal of Building Materials,2009,12(4):482-486.
[14]李沛豪,屈文俊.细菌诱导碳酸钙沉积修复混凝土裂缝[J].土木工程学报,2010,43(11):64-70.LI P H,QU W J.Remediation of concrete cracks by bacterially induced calcium carbonate deposition[J].China Civil Engineer Journal,2010,43(11):64-70.
[15]LI W,YU L J,YUAN D X,et al.A study of the activity and ecological significance of carbonic anhydrase from soil and its microbes from different karst ecosystems of Southwest China[J].Plant and Soil,2005,272:133-141.
[16]贾强,张鑫,侯宏涛,等.微生物沉积碳酸钙修复混凝土裂缝的现场试验[J].建筑材料学报,2013,16(4):667-672.JIA Q,ZHANG X,HOU H T,et al.Field experiment of crack Repair by microbiological precipitation of Ca CO3[J].Journal of Building Materials,2013,16(4):667-672.
[17]王瑞兴,钱春香,王剑云,等.水泥石表面微生物沉积碳酸钙覆膜的不同工艺[J].硅酸盐学报,2008,36(10):1 378-1 384.WANG R X,QIAN C X,WANG J Y,et al.Different treated methods of microbiologically deposited Ca CO3layer on hardened cement paste surface[J].Journal of the Chinese Ceramic Society,2008,36(10):1 378-1 384.
[18]蒋建建,刘子琦,贺秋芳,等.岩溶洞穴中碳酸钙沉积菌种的筛选及初步研究[J].西南师范大学学报:自然科学版,2012,37(12):109-114.JIANG J J,LIU Z Q,HE Q F,et al.A preliminary study and filtration on bacteria species depositing Ca CO3in Karst caves[J].Journal of Southwest China Normal University:Natural Science Edition,2012,37(12):109-114.
[19]孙莺.中国不同地区尿素分解菌的分离筛选及其诱导碳酸钙沉积能力的比较研究[D].兰州:兰州大学,2017.SUN Y.Separation screening of urea decomposing bacteria in different regions of China and its comparative study on its induction of calcium carbonate deposition[D].Lanzhou:Lanzhou University,2017.
[20]肖炜,杨亚玲,刘宏伟,等.昆明盐矿古老岩盐沉积中可培养细菌多样性研究[J].微生物学报,2006,46(6):967-972.XIAO W,YANG Y L,LIU H W,et al.Culturable bacterial diversity of the ancient salt deposits in the Kunming Salt Mine,China[J].Acta Microbiologica Sinica,2006,46(6):967-972.
[21]XIAO W,WANG Z G,WANG Y X,et al.Comparative molecular analysis of the prokaryotic diversity of two salt mine soils in southwest China[J].Journal of Basic Microbiology,2013,53:1-11.
[22]李为,刘丽萍,曹龙,等.碳酸盐生物沉积作用的研究现状与展望[J].地球科学进展,2009,24(6):598-605.LI W,LIU L P,CAO L,et al.Research status and prospect of biological precipitation of carbonate[J].Advances in Earth Science,2009,24(6):598-605.
[23]STOCKS-FISCHER S,GALINAT J K,BANG S S.Microbiological precipitation of Ca CO3[J].Soil Biology and Biochemistry,1999,31(11):1 563-1 571.
[24]FERRIS F G,PHOENIX V,FUJITA Y,et al.Kinetics of calcite precipitation induced by ureolytic bacteria at10 to 20℃in Artificial groundwater[J].Geochimica Et Cosmochimica Acta,2003,67(8):1 701-1 722.
[25]BASKAR S,BASKAR R,MAUCLAIRE L,et al.Microbially induced calcite precipitation in culture experiments:Possible origin for stalactites in Sahastradhara caves,Dehradun,India[J].Current Science,2006,90(1):58-64.
[26]LIAN B,HU Q N,CHEN J,et al.Carbonate biomineralization induced by soil bacterium Bacillus megaterium[J].Geochimica et Cosmochimica Acta,2006,70(12):5 522-5 535.
[27]MUYNCK W D,BELIE N D,VERSTRAETE W,et al.Microbial carbonate precipitation in construction materials:A review[J].Ecological Engineering,2010,36(2):118-136.
[28]ACHAL V,PAN X L.Influence of calcium sources on microbially induced calcium carbonate precipitation by Bacillus sp.CR2[J].Applied Biochemistry Biotechnology,2014,173:307-317.
[2]KANTZAS A,STEHMEIER L,MARENTETTE D F,et al.A novel method of sand consolidation through bacteriogenic mineral plugging[C].Proc of CIM Annual Technical Conference Calgary Canada:Petroleum Society of Canada,1992:1-15.
[3]钱春香,王欣,於孝牛.微生物水泥研究与应用进展[J].材料工程,2015,43(8):92-103.QIAN C X,WANG X,YU X N.Research and application development of microbe cement[J].Journal of Materials Engineering,2015,43(8):92-103.
[4]陶犁.徐霞客与云南喀斯特旅游资源[J].云南教育学院学报,1995,11(2):77-80.TAO L.Xu xiake and Yunnan karst tourism resources[J].Journal of Yunnan College of Education,1995,11(2):77-80.
[5]竹文坤,牟涛,段涛,等.响应面法优化微生物诱导碳酸钙沉积培养基[J].化工进展,2014,33(6):1 533-1 538.ZHU W K,MU T,DUAN T,et al.Optimization for microbiological-induced Ca CO3precipitation medium using response surface methodology[J].Chemical Industry and Engineering Progress,2014,33(6):1 533-1 538.
[6]竹文坤,罗学刚.碳酸盐矿化茵诱导碳酸钙沉淀条件的优化[J].非金属矿,2012,35(3):1-4.ZHU W K,LUO X G.Optimization of calcium carbonate precipitation for carbonate mineralization bacteria[J].Non-Metallic Mines,20l2,35(3):1-4.
[7]贾强,陈晓瀚,孙增斌,等.微生物诱导沉积碳酸钙提高产量的试验研究[J],山东建筑大学学报,2015,30(5):423-428.JIA Q,CHEN X H,SUN Z B,et al.The raising calcite yield experiment for bacteria induced precipitation[J].Journal of Shandong Jianzhu University,2015,30(5):423-428.
[8]成亮,钱春香,王瑞兴,等.碳酸岩矿化菌诱导碳酸钙晶体形成机理研究[J].化学学报,2007,65(19):2 133-2 138.CHENG L,QIAN C X,WANG R X,et al.Study on the mechanism of calcium carbonate formation induced by carbonate-mineralization microbe[J].Acta Chimica Sinica,2007,65(19):2 133-2 138.
[9]成亮,钱春香,王瑞兴,等.碳酸盐矿化菌调控碳酸钙结晶动力学、形态学的研究[J].功能材料,2007,38(9):1 511-1 515.CHENG L,QIAN C X,WANG R X,et al.Study on kinetics and morphology of formation of Ca CO3crystal induced by carbonate-mineralization microbe[J].Journal of Functional Materials,2007,38(9):1 511-1 515.
[10]李辉,连宾,龚国洪,等.碳酸钙颗粒的细菌诱导形成[J].高校地质学报,2011,17(1):112-117.LI H,LIAN B,GONG G H,et al.The formation of calcium carbonate particles induced by bacteria[J].Geological Journal of China Universities,2011,17(1):112-117.
[11]李沛豪,屈文俊.细菌诱导矿化保护历史建筑遗产的机理及效果[J].硅酸盐学报,2009,37(4):497-505.LI P H,QU W J.Mechanism and performance of remediation for historic buildings by bacterially induced mineralization[J].Journal of the Chinese Ceramic Society,2009,37(4):497-505.
[12]尹晓爽,张慧,杨文忠,等.一种假单胞菌诱导Ca CO3结晶[J].应用化学,2010,27(8):911-915.YIN X S,ZHANG H,YANG W Z.et al.Calcium carbonate formation induced by Pseudomonas[J].Chinese Journal of Applied Chemistry,2010,27(8):911-915.
[13]李沛豪,屈文俊.细菌诱导碳酸钙矿化材料及其应用前景[J].建筑材料学报,2009,12(4):482-486.LI P H,QU W J.Applications of calcium carbonate mineralization material induced by bacterium[J].Journal of Building Materials,2009,12(4):482-486.
[14]李沛豪,屈文俊.细菌诱导碳酸钙沉积修复混凝土裂缝[J].土木工程学报,2010,43(11):64-70.LI P H,QU W J.Remediation of concrete cracks by bacterially induced calcium carbonate deposition[J].China Civil Engineer Journal,2010,43(11):64-70.
[15]LI W,YU L J,YUAN D X,et al.A study of the activity and ecological significance of carbonic anhydrase from soil and its microbes from different karst ecosystems of Southwest China[J].Plant and Soil,2005,272:133-141.
[16]贾强,张鑫,侯宏涛,等.微生物沉积碳酸钙修复混凝土裂缝的现场试验[J].建筑材料学报,2013,16(4):667-672.JIA Q,ZHANG X,HOU H T,et al.Field experiment of crack Repair by microbiological precipitation of Ca CO3[J].Journal of Building Materials,2013,16(4):667-672.
[17]王瑞兴,钱春香,王剑云,等.水泥石表面微生物沉积碳酸钙覆膜的不同工艺[J].硅酸盐学报,2008,36(10):1 378-1 384.WANG R X,QIAN C X,WANG J Y,et al.Different treated methods of microbiologically deposited Ca CO3layer on hardened cement paste surface[J].Journal of the Chinese Ceramic Society,2008,36(10):1 378-1 384.
[18]蒋建建,刘子琦,贺秋芳,等.岩溶洞穴中碳酸钙沉积菌种的筛选及初步研究[J].西南师范大学学报:自然科学版,2012,37(12):109-114.JIANG J J,LIU Z Q,HE Q F,et al.A preliminary study and filtration on bacteria species depositing Ca CO3in Karst caves[J].Journal of Southwest China Normal University:Natural Science Edition,2012,37(12):109-114.
[19]孙莺.中国不同地区尿素分解菌的分离筛选及其诱导碳酸钙沉积能力的比较研究[D].兰州:兰州大学,2017.SUN Y.Separation screening of urea decomposing bacteria in different regions of China and its comparative study on its induction of calcium carbonate deposition[D].Lanzhou:Lanzhou University,2017.
[20]肖炜,杨亚玲,刘宏伟,等.昆明盐矿古老岩盐沉积中可培养细菌多样性研究[J].微生物学报,2006,46(6):967-972.XIAO W,YANG Y L,LIU H W,et al.Culturable bacterial diversity of the ancient salt deposits in the Kunming Salt Mine,China[J].Acta Microbiologica Sinica,2006,46(6):967-972.
[21]XIAO W,WANG Z G,WANG Y X,et al.Comparative molecular analysis of the prokaryotic diversity of two salt mine soils in southwest China[J].Journal of Basic Microbiology,2013,53:1-11.
[22]李为,刘丽萍,曹龙,等.碳酸盐生物沉积作用的研究现状与展望[J].地球科学进展,2009,24(6):598-605.LI W,LIU L P,CAO L,et al.Research status and prospect of biological precipitation of carbonate[J].Advances in Earth Science,2009,24(6):598-605.
[23]STOCKS-FISCHER S,GALINAT J K,BANG S S.Microbiological precipitation of Ca CO3[J].Soil Biology and Biochemistry,1999,31(11):1 563-1 571.
[24]FERRIS F G,PHOENIX V,FUJITA Y,et al.Kinetics of calcite precipitation induced by ureolytic bacteria at10 to 20℃in Artificial groundwater[J].Geochimica Et Cosmochimica Acta,2003,67(8):1 701-1 722.
[25]BASKAR S,BASKAR R,MAUCLAIRE L,et al.Microbially induced calcite precipitation in culture experiments:Possible origin for stalactites in Sahastradhara caves,Dehradun,India[J].Current Science,2006,90(1):58-64.
[26]LIAN B,HU Q N,CHEN J,et al.Carbonate biomineralization induced by soil bacterium Bacillus megaterium[J].Geochimica et Cosmochimica Acta,2006,70(12):5 522-5 535.
[27]MUYNCK W D,BELIE N D,VERSTRAETE W,et al.Microbial carbonate precipitation in construction materials:A review[J].Ecological Engineering,2010,36(2):118-136.
[28]ACHAL V,PAN X L.Influence of calcium sources on microbially induced calcium carbonate precipitation by Bacillus sp.CR2[J].Applied Biochemistry Biotechnology,2014,173:307-317.
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