自絮凝颗粒酵母高浓度高强度乙醇发酵的研究
|
摘要
高浓度乙醇发酵对燃料乙醇工业具有重要意义。但高浓度产物对酵母生长和乙醇发酵产生强烈的抑制作用,使目前国内外大部分乙醇连续发酵装置的发酵终点乙醇浓度限制在13%(v/v)以下。本文实验利用SPSC01自絮凝颗粒酵母为研究对象,首先考察了乙醇冲击方式对酵母细胞存活率的影响。发现经过诱导的酵母细胞与未经诱导而直接进入高浓度乙醇环境的酵母细胞相比,存活时间延长了2.5倍。但同时还发现,虽然乙醇耐受性经过诱导,但经受长时间乙醇冲击,会造成细胞活性的快速丧失。细胞存活率在乙醇浓度达到120 g·L~(-1)后10 h,45 h和90 h分别丧失了24.6%,79.1%和100%。
为避免剧烈的长时间的乙醇冲击,本实验设计了重复间歇发酵工艺,利用SPSC01自絮凝颗粒酵母絮凝沉降的优势,在几乎不增加成本的情况下实现细胞与发酵液分离,以便及时移走高浓度乙醇,即在间歇发酵终点使酵母自然沉降以获得高浓度菌体,移走含高浓度乙醇的发酵液并补充新鲜培养基进行下一批次间歇发酵。本实验共进行了10个批次,实际操作总时间为142.73 h,共获得高浓度乙醇发酵液13.44 L,平均乙醇浓度122 g·L~(-1),平均残糖浓度0.46 g·L~(-1),平均乙醇得率为0.46,平均生产强度为6.62g·L~(-1).h~(-1)。与使用游离酵母的批次和连续发酵相比,生产强度分别提高了1.4倍和1.7倍;而与使用SPSC01自絮凝颗粒酵母进行的乙醇连续发酵工艺相比,生产强度提高了12.8%。同时,在发酵结束时,细胞存活率仍保持在88%左右,细胞活性得到很好的保持。
为了进一步降低成本,本实验对此工艺的条件进行了初步优化,主要集中在对接种量和培养基营养条件的优化方面。从乙醇得率,生产强度及成本等角度综合考虑,初步确定最佳种子培养基体积为2L,培养基最佳营养条件为酵母粉12g·L~(-1),蛋白胨8g·L~(-1)。
Very high gravity(VHG) ethanol production technologies have garnered great attention. However,as a toxic metabolite,ethanol exerts a strong inhibition on yeast cell growth and ethanol production,which limits ethanol concentrations in the broth to no more than 13%(v) for most ethanol production plants at the present time.In this study,the effect of ethanol inhibition process on the viabilities of yeast cells was investigated by using a self-flocculating yeast strain SPSC01.Compared with those suddenly subjected to a high ethanol environment, the survival time of cells was prolonged by a factor of 2.5 when the ethanol concentration was increased gradually.Rapid loss of cell viability was also observed after a long-time inhibition of high ethanol.These experimental results indicated that about 24.6%,79.1%and 100% cell viability were lost after being subjected to 120 g·L~(-1) ethanol for 10 h,45 h and 90 h, respectively,even the ethanol tolerance of yeast cells had already been induced.
A repeated batch operation strategy was then proposed for high gravity ethanol fermentation.Yeast cells were separated by self-sedimentation after each turn of fermentation to hold a high level of biomass concentration,while ethanol was removed selectively before the next turn of fermentation to maintain a high level of cell viability.Ten batches have been run for 142.73 h during the whole fermentation process.Totally,13.44 L fermentation broth with high ethanol was obtained.The final ethanol concentration,residual glucose and ethanol yield were 122 g·L~(-1),0.46 g·L~(-1) and 0.46,respectively.The ethanol productivity was calculated to be 6.62 g·L~(-1)·h~(-1),which increased by factors of 1.4 and 1.7 compared with those achieved by batch and continuous,fermentations using free yeast cells,and the ethanol productivity was increased 12.8%compared with continuous fermentation using the same self-flocculating yeast strain.Furthermore,about 88%cell viability was maintained at the end of the fermentation.
The inoculation volume and amount of nutrients in the medium were further optimized to reduce the whole production cost.Considering the ethanol yield,ethanol productivity and cost,the optimal volume of broth for seed cultivation was determined to be 2 L,and the concentrations of yeast extract and peptone in the medium were further optimized to be 12 g·L~(-1)and 8 g·L~(-1),respectively.
为避免剧烈的长时间的乙醇冲击,本实验设计了重复间歇发酵工艺,利用SPSC01自絮凝颗粒酵母絮凝沉降的优势,在几乎不增加成本的情况下实现细胞与发酵液分离,以便及时移走高浓度乙醇,即在间歇发酵终点使酵母自然沉降以获得高浓度菌体,移走含高浓度乙醇的发酵液并补充新鲜培养基进行下一批次间歇发酵。本实验共进行了10个批次,实际操作总时间为142.73 h,共获得高浓度乙醇发酵液13.44 L,平均乙醇浓度122 g·L~(-1),平均残糖浓度0.46 g·L~(-1),平均乙醇得率为0.46,平均生产强度为6.62g·L~(-1).h~(-1)。与使用游离酵母的批次和连续发酵相比,生产强度分别提高了1.4倍和1.7倍;而与使用SPSC01自絮凝颗粒酵母进行的乙醇连续发酵工艺相比,生产强度提高了12.8%。同时,在发酵结束时,细胞存活率仍保持在88%左右,细胞活性得到很好的保持。
为了进一步降低成本,本实验对此工艺的条件进行了初步优化,主要集中在对接种量和培养基营养条件的优化方面。从乙醇得率,生产强度及成本等角度综合考虑,初步确定最佳种子培养基体积为2L,培养基最佳营养条件为酵母粉12g·L~(-1),蛋白胨8g·L~(-1)。
Very high gravity(VHG) ethanol production technologies have garnered great attention. However,as a toxic metabolite,ethanol exerts a strong inhibition on yeast cell growth and ethanol production,which limits ethanol concentrations in the broth to no more than 13%(v) for most ethanol production plants at the present time.In this study,the effect of ethanol inhibition process on the viabilities of yeast cells was investigated by using a self-flocculating yeast strain SPSC01.Compared with those suddenly subjected to a high ethanol environment, the survival time of cells was prolonged by a factor of 2.5 when the ethanol concentration was increased gradually.Rapid loss of cell viability was also observed after a long-time inhibition of high ethanol.These experimental results indicated that about 24.6%,79.1%and 100% cell viability were lost after being subjected to 120 g·L~(-1) ethanol for 10 h,45 h and 90 h, respectively,even the ethanol tolerance of yeast cells had already been induced.
A repeated batch operation strategy was then proposed for high gravity ethanol fermentation.Yeast cells were separated by self-sedimentation after each turn of fermentation to hold a high level of biomass concentration,while ethanol was removed selectively before the next turn of fermentation to maintain a high level of cell viability.Ten batches have been run for 142.73 h during the whole fermentation process.Totally,13.44 L fermentation broth with high ethanol was obtained.The final ethanol concentration,residual glucose and ethanol yield were 122 g·L~(-1),0.46 g·L~(-1) and 0.46,respectively.The ethanol productivity was calculated to be 6.62 g·L~(-1)·h~(-1),which increased by factors of 1.4 and 1.7 compared with those achieved by batch and continuous,fermentations using free yeast cells,and the ethanol productivity was increased 12.8%compared with continuous fermentation using the same self-flocculating yeast strain.Furthermore,about 88%cell viability was maintained at the end of the fermentation.
The inoculation volume and amount of nutrients in the medium were further optimized to reduce the whole production cost.Considering the ethanol yield,ethanol productivity and cost,the optimal volume of broth for seed cultivation was determined to be 2 L,and the concentrations of yeast extract and peptone in the medium were further optimized to be 12 g·L~(-1)and 8 g·L~(-1),respectively.
引文
[1]白凤武.无载体固定化细胞的研究进展.生物工程进展,2000,20(2):32-36.
[2]苏小军,熊兴耀,谭兴和等.燃料乙醇发酵技术研究进展.湖南农业大学学报(自然科学版),2007,33(4):480-485.
[3]Yan Lin,Shuzo Tanaka.Ethanol fermentation from biomass resources:Current state and prospects.Appled Microbiology and Biotechnology,2006,69(6):627-642.
[4]黄宇彤,杜连祥,赵继湘.世界燃料酒精工业形势.酿酒,2001,28(5):24-26.
[5]高寿清.燃料酒精发展的国际情况与分析.食品与发酵,2001,27(12):59-62.
[6]王正友.美国生物燃料乙醇生产近况.粮油食品科技,2006,14(1):1-62.
[7]张绪霞,董海洲,侯汉学等.燃料酒精制备及其开发前景.粮食与油脂,2006,(2):7-9.
[8]廖兴华,夏延斌,周传云.燃料酒精的发展现状和研究趋势.酿酒,2007,34(5):18-20.
[9]黄宇彤,杜连祥,赵继湘.世界燃料酒精生产形势.酿酒,2001,28(5):24-26.
[10]李盛贤,贾树彪,顾立文.利用纤维素原料生产燃料酒精的研究进展.酿酒,2005,32(2):13-16.
[11]侯保朝,杜风光,郭永豪等.高浓度酒精发酵.酿酒科技,2005,(4):93-96.
[12]陈丽杰,白凤武,WA Anderson等.超高浓度培养基条件下酵母细胞生长及酒精生成的准稳态动力学研究.生物加工工程,2004,(2):25-29.
[13]Pim van Hoek,Erik de Hulster,Johannes P.van Dijken,et al.Fermentative Capacity in High-Cell-Density Fed-Batch Cultures of Baker's Yeast.Biotechnology and Bioengineering,2000,68(5):517-523.
[14]吕欣,毛忠贵.高浓度酒精发酵研究进展.酿酒科技,2003,(5):58-59.
[15]Rose A H.Recent research on industrially important strains of Saccharomyces cerevisiae.London:Academic Press,1980,(3):103-109.
[16]Thomas S D,Hossack J A,Rose A H.Plasma membrane lipid composition and ethanol tolerance in Saccharomyces cerevisiae.Archives of Microbiology,1978,117(3):239-245.
[17]Beavan M J,Charpentier C,Rose A H.Production and tolerance of ethanol in relation to phospholipid fatty-acid composition in Saccharomyces cerevisiae NCYC 431.Journal of Gen Microbiol,1982,128(7):1447-1455.
[18]K A Jacques,T P Lyons,D R Kelsall.The Alcohol Textbook.Nottingham:Nottinghum University Press,1999:55-67.
[19]T D' Amore,G G Stewart.Ethanol tolerance of yeast.Enzyme and Microbial Technology,1987,9(6):322-330.
[20]陈思纭,萧熙佩.酵母生物化学.济南:山东科技出版社,1990:41-49.
[21]Rosslyn M,Bireh,et al.Influence of magnesium ions on heat shock and ethanol stress responses of Saccharomyces cerevislae.Enzyme and Microbial Technology,2000,26(9):678-687.
[22]上原悌次郎.酵母菌呼吸与发酵调节.发酵工业,1977,(5):6-15.
[23]黄亚东.应用膜分离技术提高酒精发酵的研究.酿酒,1998,(6):26-28.
[24]Correia S I,Uden V N.Temperature profiles of ethanol tolerance:effects of ethanol on the minimum and the maximum temperatures for growth of the yeasts Saccharomyces cerevisiae and Kluyveromyces fragilis.Biotechnology and Bioengineering,1983,25(6):1665-1667.
[25]Jones A M,Ingledew W M.Fuel Alcohol Production:Appraisal of Nitrogeneous Yeast Foods for Very High Gravity Wheat Mash Fermentation.Process Biochemistry,1994,29(6):483-488.
[26]池振明,高峻.酵母菌耐酒精机制的研究进展.微生物学通报,1999,26(5):373-376.
[27]池振明.高浓度酒精发酵技术的研究进展.食品与发酵工业,1995,(4):80-85.
[28]毛志群,张伟,檀建新等.燃料乙醇用高产酒精酵母的筛选及鉴定.酿酒,2003,30(3):35-37.
[29]刘建军,姜鲁燕,赵祥颖等.高产酒精酵母菌种的选育.酿酒,2003,30(3):57-59.
[30]林秋叶,赵长新,金凤燮等.两株耐高渗酿酒酵母的生理特性比较.酿酒,2003,30(2):27-28.
[31]Bayrock D P,Ingledew W M.Application of multistage continuous fermentation for Production of fuel alcohol by very-high-gravity fermentation technology.Journal of Industrial Microbiology and Biotechnology,2001,27(2):87-93.
[32]F W.Bai,W A.Anderson,M Moo-Young.Ethanol fermentation technologies from sugar and starch feedstocks.Biotechnology Advances,2008,26(1):89-105.
[33]F W Bai,L J Chen,W A Anderson,et al.Parameter oscillation in high gravity medium continuous ethanol fermentation and their attenuation on multi-stage pack column bioreactor system.Biotechnology and Bioengineering,2004,88(5):558-566.
[34]吕欣,段作营,毛忠贵等.氮源与无机盐对高浓度酒精发酵的影响.西北农林科技大学学报(自然科学版),2003,31(4):159-162.
[35]S Alfenore,C Molina-Jouve,S E Guillouet,et al.Improving ethanol production and viability of Saccharomyces cerevisiae by a vitamin feeding strategy during fed-batch process.Appled Microbiology and Biotechnology,2002,60(1):67-72.
[36]Baily J E,Ollis D F.Biochemical Engineering Fundamentals.NewYork:McGraw-Hill,1986:99-113.
[37]肖美燕,徐尔尼,陈志文.包埋法固定化细胞技术的研究进展.食品科学,2003,(4):36-38.
[38]王健.固定化酵母酒精萃取发酵技术研究.成都:西华大学,2007:12-15.
[39]Atkinson B,Mavituna F.Immobilized Cell Systems.Biochem.Eng.& Biotech.Handbook.2nd.NewYork:Stoekton Press,1991:768.
[40]Sree N K,Sridhar M,Suresh K,et al.Isolation of thermotolerant,osmotolerant,flocculating Saccharomyces cerevisiae for ethanol Production.Bioresource Technology,2000,72(1):43-46.
[41]Nahvi I,Emtiazi G,Alkabi L.Isolation of a flocculating Saccharomyces cerevisiae and investigation of its performance in the fermentation of beet molasses to ethanol.Biomass Bioenergy,2002,23(6):481-486.
[42]靳艳,白凤武,冯朴荪等.融合株SPCS发酵生产酒精的工艺研究Ⅰ.絮凝速率、发酵过程的最适温度和pH.微生物学报,1996,36(2):115-120.
[43]白凤武,云战友,谢健等.以玉米为原料采用絮凝颗粒酵母连续发酵新技术生产乙醇并副产SCP 饲料工艺的探讨.食品与发酵工业,1994,(3):60-65.
[44]白凤武,谢健,李宁等.融合株SPSC发酵生产乙醇的工艺研究—自絮凝细胞颗粒粒径分布、细胞生长和产物乙醇生成动力学.生物工程学报,1999,15(4):455-461.
[45]严正,孜力汗,白凤武等.多级串联悬浮床反应器系统中自絮凝颗粒酵母乙醇连续发酵耦合废糟液直接全循环使用的研究.生物工程学报,2005,21(4):628-632.
[46]徐铁军,赵心清,白凤武.在组合生物反应器系统中进行自絮凝酵母SPSC01乙醇连续发酵的研究.生物工程学报,2005,21(1):113-117.
[47]S Alfenore,X Cameleyre,L Benbadis,et al.Aeration strategy:a need for very high ethanol performance in Saccharomyces cerevisiae fed-batch process.Appled Microbiology and Biotechnology,2004,63(5):537-542.
[48]Bu'lock J D,Comberbach D M,Ghommidh C.A study of continuous ethanol production using a highly flocculent yeast in the gas lift tower fermentor.Chem.Eng.J,1984,29(1):9-24.
[49]Jones S T,Korus R A,Admassu W,et al.Ethanol fermentation in a continuous tower fermentor.Biotechnology and Bioengineering,1984,26(7):742-747.
[50]Admussu W,Korus R A.Ethanol fermentation with a flocculating yeast.Chemical Engineering Journal,1985,31(1):B1-BB.
[51]Kuriyama H,Ishibashi H,Miyagawa H,et al.Optimization of two-stage continuous ethanol fermentation using flocculating yeast.Biotechnology Letters,1993,15(4):415-420.
[521Kida K,Asano S,Yamadaki M,et al.Continuous high-ethanol fermentation from cane molasses by a flocculating yeast.Journal of Fermentation and Bioengineering,1990,69(1):39-45.
[53]Limtong S,Nakata M,Funahashi H,et al.Continuous ethanol production by a concentrated culture of flocculating yeast.Journal of fermentation technology,1984,62(1):55-62.
[54]白凤武,秦金来,谢健等.絮凝酵母颗粒悬浮床反应器中试及酒精连续发酵工艺的研究.生物工程学报,1995,11(3):255-259.
[55]葛旭萌,俞路,赵心清.自絮凝颗粒酵母在线检测与定量表征.化工学报,2005,56(2):306-311.
[56]Ge X M,Zhao X Q,Bai F W.Online monitoring and Characterization of Flocculating Yeast Cell Flocs During Continuous Ethanol Fermentation.Biotechnology and Bioengineering,2005,90(5):523-531.
[57]雷娟娟,赵心清,薛闯等.絮凝颗粒粒度分布对自絮凝酵母SPSC01乙醇耐受能力的影响.生物工程学报,2008,24(2):1-6
[58]王博,葛旭萌,李宁等.自絮凝颗粒酵母乙醇连续发酵耦合酵母回用工艺的研究.生物工程学报,2006,22(5):816-819
[59]G Dasari,M A Worth,M A Connor,et.al.Reasons for the apparent difference in the effects of produced and added ethanol on culture viability during rapid fermentation by Saccharomyces cerevisiae.Biotechnology and Bioengineering,1990,35(2):109-122
[60]Saibidor J,Grego J.Fatty acid alternations in Saccharomyces cerevisiae exposed to ethanol stress.FEMS Microbiol Lett,1992,93:13-16.
[2]苏小军,熊兴耀,谭兴和等.燃料乙醇发酵技术研究进展.湖南农业大学学报(自然科学版),2007,33(4):480-485.
[3]Yan Lin,Shuzo Tanaka.Ethanol fermentation from biomass resources:Current state and prospects.Appled Microbiology and Biotechnology,2006,69(6):627-642.
[4]黄宇彤,杜连祥,赵继湘.世界燃料酒精工业形势.酿酒,2001,28(5):24-26.
[5]高寿清.燃料酒精发展的国际情况与分析.食品与发酵,2001,27(12):59-62.
[6]王正友.美国生物燃料乙醇生产近况.粮油食品科技,2006,14(1):1-62.
[7]张绪霞,董海洲,侯汉学等.燃料酒精制备及其开发前景.粮食与油脂,2006,(2):7-9.
[8]廖兴华,夏延斌,周传云.燃料酒精的发展现状和研究趋势.酿酒,2007,34(5):18-20.
[9]黄宇彤,杜连祥,赵继湘.世界燃料酒精生产形势.酿酒,2001,28(5):24-26.
[10]李盛贤,贾树彪,顾立文.利用纤维素原料生产燃料酒精的研究进展.酿酒,2005,32(2):13-16.
[11]侯保朝,杜风光,郭永豪等.高浓度酒精发酵.酿酒科技,2005,(4):93-96.
[12]陈丽杰,白凤武,WA Anderson等.超高浓度培养基条件下酵母细胞生长及酒精生成的准稳态动力学研究.生物加工工程,2004,(2):25-29.
[13]Pim van Hoek,Erik de Hulster,Johannes P.van Dijken,et al.Fermentative Capacity in High-Cell-Density Fed-Batch Cultures of Baker's Yeast.Biotechnology and Bioengineering,2000,68(5):517-523.
[14]吕欣,毛忠贵.高浓度酒精发酵研究进展.酿酒科技,2003,(5):58-59.
[15]Rose A H.Recent research on industrially important strains of Saccharomyces cerevisiae.London:Academic Press,1980,(3):103-109.
[16]Thomas S D,Hossack J A,Rose A H.Plasma membrane lipid composition and ethanol tolerance in Saccharomyces cerevisiae.Archives of Microbiology,1978,117(3):239-245.
[17]Beavan M J,Charpentier C,Rose A H.Production and tolerance of ethanol in relation to phospholipid fatty-acid composition in Saccharomyces cerevisiae NCYC 431.Journal of Gen Microbiol,1982,128(7):1447-1455.
[18]K A Jacques,T P Lyons,D R Kelsall.The Alcohol Textbook.Nottingham:Nottinghum University Press,1999:55-67.
[19]T D' Amore,G G Stewart.Ethanol tolerance of yeast.Enzyme and Microbial Technology,1987,9(6):322-330.
[20]陈思纭,萧熙佩.酵母生物化学.济南:山东科技出版社,1990:41-49.
[21]Rosslyn M,Bireh,et al.Influence of magnesium ions on heat shock and ethanol stress responses of Saccharomyces cerevislae.Enzyme and Microbial Technology,2000,26(9):678-687.
[22]上原悌次郎.酵母菌呼吸与发酵调节.发酵工业,1977,(5):6-15.
[23]黄亚东.应用膜分离技术提高酒精发酵的研究.酿酒,1998,(6):26-28.
[24]Correia S I,Uden V N.Temperature profiles of ethanol tolerance:effects of ethanol on the minimum and the maximum temperatures for growth of the yeasts Saccharomyces cerevisiae and Kluyveromyces fragilis.Biotechnology and Bioengineering,1983,25(6):1665-1667.
[25]Jones A M,Ingledew W M.Fuel Alcohol Production:Appraisal of Nitrogeneous Yeast Foods for Very High Gravity Wheat Mash Fermentation.Process Biochemistry,1994,29(6):483-488.
[26]池振明,高峻.酵母菌耐酒精机制的研究进展.微生物学通报,1999,26(5):373-376.
[27]池振明.高浓度酒精发酵技术的研究进展.食品与发酵工业,1995,(4):80-85.
[28]毛志群,张伟,檀建新等.燃料乙醇用高产酒精酵母的筛选及鉴定.酿酒,2003,30(3):35-37.
[29]刘建军,姜鲁燕,赵祥颖等.高产酒精酵母菌种的选育.酿酒,2003,30(3):57-59.
[30]林秋叶,赵长新,金凤燮等.两株耐高渗酿酒酵母的生理特性比较.酿酒,2003,30(2):27-28.
[31]Bayrock D P,Ingledew W M.Application of multistage continuous fermentation for Production of fuel alcohol by very-high-gravity fermentation technology.Journal of Industrial Microbiology and Biotechnology,2001,27(2):87-93.
[32]F W.Bai,W A.Anderson,M Moo-Young.Ethanol fermentation technologies from sugar and starch feedstocks.Biotechnology Advances,2008,26(1):89-105.
[33]F W Bai,L J Chen,W A Anderson,et al.Parameter oscillation in high gravity medium continuous ethanol fermentation and their attenuation on multi-stage pack column bioreactor system.Biotechnology and Bioengineering,2004,88(5):558-566.
[34]吕欣,段作营,毛忠贵等.氮源与无机盐对高浓度酒精发酵的影响.西北农林科技大学学报(自然科学版),2003,31(4):159-162.
[35]S Alfenore,C Molina-Jouve,S E Guillouet,et al.Improving ethanol production and viability of Saccharomyces cerevisiae by a vitamin feeding strategy during fed-batch process.Appled Microbiology and Biotechnology,2002,60(1):67-72.
[36]Baily J E,Ollis D F.Biochemical Engineering Fundamentals.NewYork:McGraw-Hill,1986:99-113.
[37]肖美燕,徐尔尼,陈志文.包埋法固定化细胞技术的研究进展.食品科学,2003,(4):36-38.
[38]王健.固定化酵母酒精萃取发酵技术研究.成都:西华大学,2007:12-15.
[39]Atkinson B,Mavituna F.Immobilized Cell Systems.Biochem.Eng.& Biotech.Handbook.2nd.NewYork:Stoekton Press,1991:768.
[40]Sree N K,Sridhar M,Suresh K,et al.Isolation of thermotolerant,osmotolerant,flocculating Saccharomyces cerevisiae for ethanol Production.Bioresource Technology,2000,72(1):43-46.
[41]Nahvi I,Emtiazi G,Alkabi L.Isolation of a flocculating Saccharomyces cerevisiae and investigation of its performance in the fermentation of beet molasses to ethanol.Biomass Bioenergy,2002,23(6):481-486.
[42]靳艳,白凤武,冯朴荪等.融合株SPCS发酵生产酒精的工艺研究Ⅰ.絮凝速率、发酵过程的最适温度和pH.微生物学报,1996,36(2):115-120.
[43]白凤武,云战友,谢健等.以玉米为原料采用絮凝颗粒酵母连续发酵新技术生产乙醇并副产SCP 饲料工艺的探讨.食品与发酵工业,1994,(3):60-65.
[44]白凤武,谢健,李宁等.融合株SPSC发酵生产乙醇的工艺研究—自絮凝细胞颗粒粒径分布、细胞生长和产物乙醇生成动力学.生物工程学报,1999,15(4):455-461.
[45]严正,孜力汗,白凤武等.多级串联悬浮床反应器系统中自絮凝颗粒酵母乙醇连续发酵耦合废糟液直接全循环使用的研究.生物工程学报,2005,21(4):628-632.
[46]徐铁军,赵心清,白凤武.在组合生物反应器系统中进行自絮凝酵母SPSC01乙醇连续发酵的研究.生物工程学报,2005,21(1):113-117.
[47]S Alfenore,X Cameleyre,L Benbadis,et al.Aeration strategy:a need for very high ethanol performance in Saccharomyces cerevisiae fed-batch process.Appled Microbiology and Biotechnology,2004,63(5):537-542.
[48]Bu'lock J D,Comberbach D M,Ghommidh C.A study of continuous ethanol production using a highly flocculent yeast in the gas lift tower fermentor.Chem.Eng.J,1984,29(1):9-24.
[49]Jones S T,Korus R A,Admassu W,et al.Ethanol fermentation in a continuous tower fermentor.Biotechnology and Bioengineering,1984,26(7):742-747.
[50]Admussu W,Korus R A.Ethanol fermentation with a flocculating yeast.Chemical Engineering Journal,1985,31(1):B1-BB.
[51]Kuriyama H,Ishibashi H,Miyagawa H,et al.Optimization of two-stage continuous ethanol fermentation using flocculating yeast.Biotechnology Letters,1993,15(4):415-420.
[521Kida K,Asano S,Yamadaki M,et al.Continuous high-ethanol fermentation from cane molasses by a flocculating yeast.Journal of Fermentation and Bioengineering,1990,69(1):39-45.
[53]Limtong S,Nakata M,Funahashi H,et al.Continuous ethanol production by a concentrated culture of flocculating yeast.Journal of fermentation technology,1984,62(1):55-62.
[54]白凤武,秦金来,谢健等.絮凝酵母颗粒悬浮床反应器中试及酒精连续发酵工艺的研究.生物工程学报,1995,11(3):255-259.
[55]葛旭萌,俞路,赵心清.自絮凝颗粒酵母在线检测与定量表征.化工学报,2005,56(2):306-311.
[56]Ge X M,Zhao X Q,Bai F W.Online monitoring and Characterization of Flocculating Yeast Cell Flocs During Continuous Ethanol Fermentation.Biotechnology and Bioengineering,2005,90(5):523-531.
[57]雷娟娟,赵心清,薛闯等.絮凝颗粒粒度分布对自絮凝酵母SPSC01乙醇耐受能力的影响.生物工程学报,2008,24(2):1-6
[58]王博,葛旭萌,李宁等.自絮凝颗粒酵母乙醇连续发酵耦合酵母回用工艺的研究.生物工程学报,2006,22(5):816-819
[59]G Dasari,M A Worth,M A Connor,et.al.Reasons for the apparent difference in the effects of produced and added ethanol on culture viability during rapid fermentation by Saccharomyces cerevisiae.Biotechnology and Bioengineering,1990,35(2):109-122
[60]Saibidor J,Grego J.Fatty acid alternations in Saccharomyces cerevisiae exposed to ethanol stress.FEMS Microbiol Lett,1992,93:13-16.