白酒酿造中产正丙醇的微生物溯源研究
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摘要
利用生物信息数据资源,对白酒酿造过程中产正丙醇微生物进行定向补充筛查,并对筛查后的菌株进行了发酵验证。结果表明,基于KEGG数据库明确了正丙醇的产生途径为丙酸代谢途径,其催化的关键酶为丙二醇脱水酶、1,3-丙二醇脱氢酶和1-丙醇脱氢酶;通过NCBI数据库确定了表达该3种关键酶的微生物种类信息,结合白酒发酵过程中的微生物群落构成,确定乳杆菌属(Lactobacillus)具有代谢产生正丙醇的潜在特性;发酵结果显示,面包乳杆菌(Lactobacillus panis)代谢生成正丙醇的能力相对较强,产量为263.7 mg/L。该研究结果表明,乳酸杆菌是白酒酿造中产正丙醇的重要微生物来源之一。该研究为白酒酿造过程正丙醇的调控提供新的研究方向,同时该方法也可以为其他食品发酵过程中目标风味物质的微生物溯源提供新思路。
In this study, 1-propanol-producing microorganisms were directional screened from Baijiu(Chinese liquor) brewing process by bioinformatics databases. And then the screened strains were subjected to fermentation validation. The results showed that the generation pathway of 1-propanol was involved in propanoate metabolism based on KEGG database, and then it confirmed that three key enzymes involved for catalyzation of 1-propanol synthesis were propanediol dehydratase, 1,3-propanediol dehydrogenase, and 1-propanol dehydrogenase. The Microbial species information of three key enzymes was confirmed by NCBI database. Combined with the microbial community in Baijiu fermentation process, the results showed that Lactobacillus had the potential capability for producing 1-propanol. The fermentation results showed that Lactobacillus panis had higher 1-propanol-producing capability, and the production was 263.7 mg/L. These results indicated that Lactobacillus was one of the most important microbial sources for producing 1-propanol during Baijiu fermentation process. The method provided a new research direction for regulation of 1-propanol. Meanwhile, the method could also provide a new idea for microbial traceability of target flavor substances in other food fermentation process.
In this study, 1-propanol-producing microorganisms were directional screened from Baijiu(Chinese liquor) brewing process by bioinformatics databases. And then the screened strains were subjected to fermentation validation. The results showed that the generation pathway of 1-propanol was involved in propanoate metabolism based on KEGG database, and then it confirmed that three key enzymes involved for catalyzation of 1-propanol synthesis were propanediol dehydratase, 1,3-propanediol dehydrogenase, and 1-propanol dehydrogenase. The Microbial species information of three key enzymes was confirmed by NCBI database. Combined with the microbial community in Baijiu fermentation process, the results showed that Lactobacillus had the potential capability for producing 1-propanol. The fermentation results showed that Lactobacillus panis had higher 1-propanol-producing capability, and the production was 263.7 mg/L. These results indicated that Lactobacillus was one of the most important microbial sources for producing 1-propanol during Baijiu fermentation process. The method provided a new research direction for regulation of 1-propanol. Meanwhile, the method could also provide a new idea for microbial traceability of target flavor substances in other food fermentation process.
引文
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[6]WU Q,ZHU W A,WANG W,et al.Effect of yeast species on the terpenoids profile of Chinese light-style liquor[J].Food Chem,2015,168:390-395.
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[14]柏永昊,张明春,缪礼鸿.芽孢杆菌对白酒发酵过程中正丙醇含量的影响[J].酿酒科技,2013(11):26-29.
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[16]王霜.产酱香功能芽孢杆菌的筛选及浓酱兼香型白酒复合功能强化曲的研究[D].武汉:武汉轻工大学,2017.
[17]游玲,任羽,王涛,等.酵母对浓香型白酒糟醅中高级醇生成的影响[J].食品与发酵工业,2016,42(2):23-28.
[18]赵长青,徐莎,杨阳,等.浓香型白酒酿造大曲及糟醅中功能芽孢杆菌的筛选[J].食品工业科技,2017,38(7):151-155.
[19]MASTUBARA M,URANO N,YAMADA S,et al.Fermentative production of 1-propanol from D-glucose,l-rhamnose and glycerol using recombinant Escherichia coli[J].J Biosci Bioeng,2016,122(4):421-426.
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[21]ZHANG L Q,WU C D,DING X F,et al.Characterisation of microbial communities in Chinese liquor fermentation starters Daqu using nested PCR-DGGE[J].World J Microb Biotechn,2014,30(12):3055-3063.
[22]ZHENG X W,YAN Z,HAN B Z,et al.Complex microbiota of a Chinese"Fen"liquor fermentation starter(Fen-Daqu),revealed by culturedependent and culture-independent methods[J].Food Microbiol,2012,31(2):293-300.
[23]HU X L,DU H,XU Y.Identification and quantification of the caproic acid-producing bacterium Clostridium kluyveri in the fermentation of pit mud used for Chinese strong-aroma type liquor production[J].Int JFood Microbiol,2015,214:116-122.
[24]LIU X,GUO K L,ZHANG H X.Determination of microbial diversity in Daqu,a fermentation starter culture of Maotai liquor,using nested PCR-denaturing gradient gel electrophoresis[J].World J Microb Biotechn,2012,28(6):2375-2381.
[25]LI K,ZHANG Q,ZHONG X T,et al.Microbial diversity and succession in the Chinese Luzhou-flavor liquor fermenting cover lees as evaluated by SSU r RNA Profiles[J].Ind J Microbiol,2013,53(4):425-431.
[26]郭敏,黄永光,邱树毅,等.高通量测序在酱香白酒微生态多样性研究中的应用[J].中国酿造,2017,36(5):146-151.
[27]王鹏,吴群,徐岩.中国白酒发酵过程中的核心微生物群及其与环境因子的关系[J].微生物学报,2018,58(1):142-153.
[28]刘凡,周新虎,陈翔,等.洋河浓香型白酒发酵过程酒醅微生物群落结构解析及其与有机酸合成的相关性[J].微生物学报,2018,58(12):2087-2099.
[29]WANG L,WANG Y Y,WANG D Q,et al.Dynamic changes in the bacterial community in Moutai liquor fermentation process characterized by deep sequencing[J].J Inst Brew,2015,121(4):603-608.
[30]FAN G S,SUN B G,FU Z L,et al.Analysis of physicochemical indices,volatile flavor components,and microbial community of a light-flavor Daqu[J].J Am Soc Brew Chem,2018,76(3):209-218.
[31]YANG F,CHEN L Q,LIU Y F,et al.Identification of microorganisms producing lactic acid during solid-state fermentation of Maotai flavour liquor[J].J Inst Brew,2018.
[32]HONG X T,CHEN J,LIU L,et al.Metagenomic sequencing reveals the relationship between microbiota composition and quality of Chinese rice wine[J].Sci Report,2016,6:26621.
[33]WANG X S,DU H,ZHANG Y,et al.Environmental microbiota drives microbial succession and metabolic profiles during Chinese liquor fermentation[J].Appl Environ Microbiol,2017,84(4):02369-17.
[34]WANG J,ZHONG Q P,YANG Y Y,et al.Comparison of bacterial diversity between two traditional starters and the round-koji-maker starter for traditional cantonese chi-flavor liquor brewing[J].Front Microbiol,2018,9:1053.
[35]CHEN Q P,KANG G T,ZHOU T N,et al.Significant difference of the culturable anaerobic microbial species in GujingTribute pitmud with different ages[J].Mat Sci Eng,2018,394:22-25.
[36]LI X R,MA E B,YAN L Z,et al.Bacterial and fungal diversity in the traditional Chinese liquor fermentation process[J].Int J Food Microbiol,2011,146(1):31-37.
[37]ZHANG W X,QIAO Z W,TORU S,et al.Analysis of the bacterial community in Zaopei during production of Chinese Luzhou-flavor liquor[J].J Inst Brew,2005,111(2):215-222.
[2]YOU L,WANG T,YANG Z R,et al.Performance of indigenous yeasts in the processing of Chinese strong-flavoured liquor during spontaneous mixed solid-state or submerged fermentation[J].J Inst Brew,2015,121(2):295-303.
[3]FU Z L,SUN B G,LI X T,et al.Isolation and characterization of a high ethyl acetate-producing yeast from Laobaigan Daqu and its fermentation conditions for producing high-quality Baijiu[J].Biotechnol Biotechnolog Equip,2018,32(5):1218-1227.
[4]ZOU W,YE G B,ZHANGK Z.Diversity,function,and application of Clostridium in Chinese strong flavor Baijiu ecosystem:a review[J].J Food Sci,2018,83(5):1193-1199.
[5]DU H,LIU B,WANG X S,et al.Exploring the microbial origins of,p-cresol and its co-occurrence pattern in the Chinese liquor-making process[J].Int J Food Microbiol,2017,260:27-35.
[6]WU Q,ZHU W A,WANG W,et al.Effect of yeast species on the terpenoids profile of Chinese light-style liquor[J].Food Chem,2015,168:390-395.
[7]ZHAO C Q,YAN X L,YANG S T,et al.Screening of Bacillus strains from Luzhou-flavor liquor making for high-yield ethyl hexanoate and low-yield propanol[J].LWT-Food Sci Technol,2017,77:60-66.
[8]陈向田.芝麻香型白酒关键细菌的分离与风味功能研究[J].酿酒,2018,45(5):95-96.
[9]AlEJANDRA E Z,ARTURO V P D L,ALEJANDRO S F.The road to metagenomics:from microbiology to DNA sequencing technologies and bioinformatics[J].Front Genet,2015,6(104):348.
[10]贾栋,贾小云,马瑞燕.生物信息学数据库及查询[J].山西农业大学学报(自然科学版),2013,33(6):520-525.
[11]PALLEN M J.Microbial bioinformatics 2020[J].Microbial Biotechnol,2016,9(5):681-686.
[12]ORTIZ-ESTRADA A M,GOLLAS-GALV魣N T,MART魱NEZ-C魷RDOVAL R,et al.Predictive functional profiles using metagenomic 16S r RNAdata:a novel approach to understanding the microbial ecology of aquaculture systems[J].Rev Aquacult,2018:1-12.
[13]谢方安.谈白酒香气成分和作用[J].酿酒,2006,33(5):52-55.
[14]柏永昊,张明春,缪礼鸿.芽孢杆菌对白酒发酵过程中正丙醇含量的影响[J].酿酒科技,2013(11):26-29.
[15]孙神英.不同酵母菌对白酒风味成分的影响[D].武汉:武汉轻工大学,2014.
[16]王霜.产酱香功能芽孢杆菌的筛选及浓酱兼香型白酒复合功能强化曲的研究[D].武汉:武汉轻工大学,2017.
[17]游玲,任羽,王涛,等.酵母对浓香型白酒糟醅中高级醇生成的影响[J].食品与发酵工业,2016,42(2):23-28.
[18]赵长青,徐莎,杨阳,等.浓香型白酒酿造大曲及糟醅中功能芽孢杆菌的筛选[J].食品工业科技,2017,38(7):151-155.
[19]MASTUBARA M,URANO N,YAMADA S,et al.Fermentative production of 1-propanol from D-glucose,l-rhamnose and glycerol using recombinant Escherichia coli[J].J Biosci Bioeng,2016,122(4):421-426.
[20]SIEBERT D,WENDISCH V F.Metabolic pathway engineering for production of 1,2-propanediol and 1-propanol by Corynebacterium glutamicum[J].Biotechnol Biofuel,2015,8(1):91-103.
[21]ZHANG L Q,WU C D,DING X F,et al.Characterisation of microbial communities in Chinese liquor fermentation starters Daqu using nested PCR-DGGE[J].World J Microb Biotechn,2014,30(12):3055-3063.
[22]ZHENG X W,YAN Z,HAN B Z,et al.Complex microbiota of a Chinese"Fen"liquor fermentation starter(Fen-Daqu),revealed by culturedependent and culture-independent methods[J].Food Microbiol,2012,31(2):293-300.
[23]HU X L,DU H,XU Y.Identification and quantification of the caproic acid-producing bacterium Clostridium kluyveri in the fermentation of pit mud used for Chinese strong-aroma type liquor production[J].Int JFood Microbiol,2015,214:116-122.
[24]LIU X,GUO K L,ZHANG H X.Determination of microbial diversity in Daqu,a fermentation starter culture of Maotai liquor,using nested PCR-denaturing gradient gel electrophoresis[J].World J Microb Biotechn,2012,28(6):2375-2381.
[25]LI K,ZHANG Q,ZHONG X T,et al.Microbial diversity and succession in the Chinese Luzhou-flavor liquor fermenting cover lees as evaluated by SSU r RNA Profiles[J].Ind J Microbiol,2013,53(4):425-431.
[26]郭敏,黄永光,邱树毅,等.高通量测序在酱香白酒微生态多样性研究中的应用[J].中国酿造,2017,36(5):146-151.
[27]王鹏,吴群,徐岩.中国白酒发酵过程中的核心微生物群及其与环境因子的关系[J].微生物学报,2018,58(1):142-153.
[28]刘凡,周新虎,陈翔,等.洋河浓香型白酒发酵过程酒醅微生物群落结构解析及其与有机酸合成的相关性[J].微生物学报,2018,58(12):2087-2099.
[29]WANG L,WANG Y Y,WANG D Q,et al.Dynamic changes in the bacterial community in Moutai liquor fermentation process characterized by deep sequencing[J].J Inst Brew,2015,121(4):603-608.
[30]FAN G S,SUN B G,FU Z L,et al.Analysis of physicochemical indices,volatile flavor components,and microbial community of a light-flavor Daqu[J].J Am Soc Brew Chem,2018,76(3):209-218.
[31]YANG F,CHEN L Q,LIU Y F,et al.Identification of microorganisms producing lactic acid during solid-state fermentation of Maotai flavour liquor[J].J Inst Brew,2018.
[32]HONG X T,CHEN J,LIU L,et al.Metagenomic sequencing reveals the relationship between microbiota composition and quality of Chinese rice wine[J].Sci Report,2016,6:26621.
[33]WANG X S,DU H,ZHANG Y,et al.Environmental microbiota drives microbial succession and metabolic profiles during Chinese liquor fermentation[J].Appl Environ Microbiol,2017,84(4):02369-17.
[34]WANG J,ZHONG Q P,YANG Y Y,et al.Comparison of bacterial diversity between two traditional starters and the round-koji-maker starter for traditional cantonese chi-flavor liquor brewing[J].Front Microbiol,2018,9:1053.
[35]CHEN Q P,KANG G T,ZHOU T N,et al.Significant difference of the culturable anaerobic microbial species in GujingTribute pitmud with different ages[J].Mat Sci Eng,2018,394:22-25.
[36]LI X R,MA E B,YAN L Z,et al.Bacterial and fungal diversity in the traditional Chinese liquor fermentation process[J].Int J Food Microbiol,2011,146(1):31-37.
[37]ZHANG W X,QIAO Z W,TORU S,et al.Analysis of the bacterial community in Zaopei during production of Chinese Luzhou-flavor liquor[J].J Inst Brew,2005,111(2):215-222.