传统自然发酵酸肉中细菌群落多样性与风味品质分析
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摘要
为探讨传统自然发酵酸肉中细菌群落多样性与风味品质的关系,采用Ion S5 XL测序平台,对4种传统自然发酵酸肉中细菌16S rDNA V3～V4区进行高通量测序,并结合电子鼻和固相微萃取-气相色谱-质谱(solid phase microextraction-gaschromatography-massspectrometry,SPME-GC-MS)分析等手段揭示酸肉细菌群落结构及其风味品质的相关性。结果表明,样品获得序列数平均为110 395条。4种酸肉样品中总体细菌群落包含11个门,其中厚壁菌门占绝对优势,约占总细菌群落的83.73%～98.92%,其次是变形菌门和放线菌门。主要优势菌属为乳杆菌属、魏斯氏菌属和乳球菌属。利用SPME-GC-MS从酸肉样品中检测到的挥发性物质包括酸类、醇类、醛类、酯类、酮类、萜烯类等化合物共126种。各样品之间的菌属丰度存在一定的差异性,其菌群组成与工艺密切相关,而菌群种类和数量影响产品风味。与传统方法相比,高通量测序得到的细菌多样性信息更接近于样品微生态,能够全面解析自然发酵肉制品酸肉的细菌多样性,为传统食品的现代化改造和质量安全控制提供科学支撑。
The present study was conducted in order to investigate the relationship between the diversity of the bacterial community in four naturally fermented sour meat products and their flavor qualities. The bacterial community composition was investigated by 16S rDNA V3-V4 region high throughput sequencing using Ion S5 XL sequencing platform. Meanwhile, these samples were subjected to electronic nose evaluation and solid phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS) analysis. The results revealed that the average number of sequences per sample was 110 395. The overall bacterial community compositions of the four samples were quite complicated, including 11 phyla, among which, Firmicutes was absolutely dominant, accounting for 83.73%–98.92% of the total number, followed by Proteobacteria and Actinobacteria. The dominant genera were Lactobacillus, Weissella and Lactococcus. A total of 126 volatile compounds were identified from four naturally fermented sour meats, including acids, alcohols, aldehydes, esters, ketones and terpenes. There were differences in the abundance of bacterial species among the four samples and the microfloral community was closely to the production process. The bacterial diversity involved in the fermentation process was one of the most important factors shaping the flavor characteristics of traditional sour meat products. Compared to traditional methods, the information about bacterial community diversity provided by high-throughput sequencing method was closer to the microecology of the samples, which were helpful to better understand the microfloral composition of sour meat and might provide a scientific support for the modernization of the traditional food industry and food quality and safety control.
The present study was conducted in order to investigate the relationship between the diversity of the bacterial community in four naturally fermented sour meat products and their flavor qualities. The bacterial community composition was investigated by 16S rDNA V3-V4 region high throughput sequencing using Ion S5 XL sequencing platform. Meanwhile, these samples were subjected to electronic nose evaluation and solid phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS) analysis. The results revealed that the average number of sequences per sample was 110 395. The overall bacterial community compositions of the four samples were quite complicated, including 11 phyla, among which, Firmicutes was absolutely dominant, accounting for 83.73%–98.92% of the total number, followed by Proteobacteria and Actinobacteria. The dominant genera were Lactobacillus, Weissella and Lactococcus. A total of 126 volatile compounds were identified from four naturally fermented sour meats, including acids, alcohols, aldehydes, esters, ketones and terpenes. There were differences in the abundance of bacterial species among the four samples and the microfloral community was closely to the production process. The bacterial diversity involved in the fermentation process was one of the most important factors shaping the flavor characteristics of traditional sour meat products. Compared to traditional methods, the information about bacterial community diversity provided by high-throughput sequencing method was closer to the microecology of the samples, which were helpful to better understand the microfloral composition of sour meat and might provide a scientific support for the modernization of the traditional food industry and food quality and safety control.
引文
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[17]李迎楠,刘文营,成晓瑜.GC-MS结合电子鼻分析温度对肉味香精风味品质的影响[J].食品科学,2016,37(14):104-109.DOI:10.7506/spkx1002-6630-201614018.
[18]米瑞芳,陈曦,戚彪,等.乳杆菌发酵剂对酸肉挥发性风味成分的影响[J].肉类研究,2018,32(4):48-55.DOI:10.7506/rlyj1001-8123-201804009.
[19]GUTSCHE K A,TRAN T B T,VOGEL R F.Production of volatile compounds by Lactobacillus sakei from branched chainα-keto acids[J].Food Microbiology,2012,29(2):224-228.DOI:10.1016/j.fm.2011.06.010.
[20]周才琼,李艳芳,杜木英,等.渝黔地区传统酸肉发酵过程中微生物区系研究[J].食品工业科技,2010,31(4):171-175.
[21]FONTANA C,BASSI D,LóPEZ C,et al.Microbial ecology involved in the ripening of naturally fermented llama meat sausages.A focus on lactobacilli diversity[J].International Journal of Food Microbiology,2016,236:17-25.DOI:10.1016/j.ijfoodmicro.2016.07.002.
[22]CHEN C,CHEN X H,JIANG M,et al.A newly discovered bacteriocin from Weissella hellenica D1501 associated with Chinese Dong fermented meat (Nanx Wudl)[J].Food Control,2014,42(2):116-124.DOI:10.1016/j.foodcont.2014.01.031.
[23]余冰,周红丽,李宗军.固相微萃取分析发酵肉制品中的挥发性风味组分[J].湖南农业大学学报(自然科学版),2007,33(2):232-234.DOI:10.13331/j.cnki.jhau.2007.02.026.
[24]周才琼,代小容,杜木英.酸肉发酵过程中挥发性风味物质形成的研究[J].食品科学,2010,31(7):98-104.
[25]黄群,滕慧,宋洪波,等.固相微萃取结合GC-MS分析湘西酸肉挥发性成分[J].食品工业,2016,37(11):277-280.
[26]SIDIRA M,KANDYLIS P,KANELLAKI M,et al.Effect of curing salts and probiotic cultures on the evolution of flavor compounds in dry-fermented sausages during ripening[J].Food Chemistry,2016,201:334-338.DOI:10.1016/j.foodchem.2016.01.084.
[27]朱健辉.不同等级金华火腿蛋白质和脂肪水解产物与挥发性风味物质分析[D].南京:南京农业大学,2005:50.
[28]JELE?H,MAJCHER M,GINJA A,et al.Determination of compounds responsible for tempeh aroma[J].Food Chemistry,2013,141(1):459-465.DOI:10.1016/j.foodchem.2013.03.047.
[29]LEROY F,VERLUYTEN J,DE V L.Functional meat starter cultures for improved sausage fermentation[J].International Journal of Food Microbiology,2006,106(3):270-285.DOI:10.1016/j.ijfoodmicro.2005.06.027.
[30]CHEN Q,LIU Q,SUN Q X,et al.Flavour formation from hydrolysis of pork sarcoplasmic protein extract by a unique LAB culture isolated from Harbin dry sausage[J].Meat Science,2015,100:110-117.DOI:10.1016/j.meatsci.2014.10.001.
[31]褚福娟,孔保华,黄永.发酵肉制品常见微生物及其对风味的影响[J].肉类研究,2008,22(1):3-6.DOI:10.3969/j.issn.1001-8123.2008.01.004.
[32]KARGOZARI M,MOINI S,AKHONDZADEH B A,et al.Effect of autochthonous starter cultures isolated from Siahmazgi cheese on physicochemical,microbiological and volatile compound profiles and sensorial attributes of sucuk,a Turkish dry-fermented sausage[J].Meat Science,2014,97(1):104-114.DOI:10.1016/j.meatsci.2014.01.013.
[33]SIDIRA M,KANDYLIS P,KANELLAKI M,et al.Effect of immobilized Lactobacillus casei on the evolution of flavor compounds in probiotic dry-fermented sausages during ripening[J].Meat Science,2015,100:41-51.DOI:10.1016/j.meatsci.2014.09.011.
[34]黄业传,李婷婷,龚道楷,等.高压结合热处理对猪肉风味的影响[J].食品科学,2018,39(2):227-232.DOI:10.7506/spkx1002-6630-201802036.
[35]赵健,王蒙,谢建春,等.黑猪肉关键香气物质分析鉴定[J].食品科学,2018,39(2):203-209.DOI:10.7506/spkx1002-6630-201802032.
[36]MONTANARI C,BARGOSSI E,GARDINI A,et al.Correlation between volatile profiles of Italian fermented sausages and their size and starter culture[J].Food Chemistry,2016,192:736-744.DOI:10.1016/j.foodchem.2015.07.062.
[37]KOT W,NEVE H,HELLER K J,et al.Bacteriophages of Leuconostoc,Oenococcus,and Weissella[J].Front Microbiol,2014,5(2):186-195.DOI:10.3389/fmicb.2014.00186.
[2]张倩,郭晓芸,张海松,等.贵州荔波传统酸肉发酵期间理化成分与发酵风味物质分析[J].食品科学,2013,34(4):173-177.
[3]孙文静,卫飞,袁军,等.酸肉发酵中脂肪的变化及对小鼠血脂的影响[J].食品科学,2013,34(11):263-267.DOI:10.7506/spkx1002-6630-201311057.
[4]叶春,马顺强,张倩,等.酸肉肽抗氧化活性的研究[J].食品科学,2010,31(7):94-97.
[5]丁苗,刘洋,葛平珍,等.发酵酸肉中降胆固醇乳酸菌的筛选、鉴定及降胆固醇作用[J].食品科学,2014,35(19):203-207.DOI:10.7506/spkx1002-6630-201419041.
[6]张倩,郭晓芸,范丽平,等.贵州荔波传统酸肉优势菌筛选、鉴定与发酵性能研究[J].食品工业科技,2015,36(13):189-194.DOI:10.13386/j.issn1002-0306.2015.13.031.
[7]湛剑龙,陈韵,黄林波,等.贵州少数民族酸肉、酸鱼中乳酸菌的分离鉴定[J].肉类研究,2013,27(7):40-43.
[8]李宗军,江汉湖.中国传统酸肉发酵过程中微生物的消长变化[J].微生物学通报,2004,31(4):9-13.DOI:10.3969/j.issn.0253-2654.2004.04.003.
[9]陈韵,胡萍,湛剑龙,等.运用PCR-DGGE技术分析贵州侗族酸肉中细菌多样性[J].食品与机械,2014(2):188-191.DOI:10.3969/j.issn.1003-5788.2014.02.047.
[10]QUIJADA N M,FILIPPIS F D,SANZ J J,et al.Different Lactobacillus populations dominate in“Chorizo de León”manufacturing performed in different production plants[J].Food Microbiology,2018,70:94-102.DOI:10.1016/j.fm.2017.09.009.
[11]PARENTE E,COCOLIN L,FILIPPIS F D,et al.Food microbionet:a database for the visualisation and exploration of food bacterial communities based on network analysis[J].International Journal of Food Microbiology,2016,219:28-37.DOI:10.1016/j.ijfoodmicro.2015.12.001.
[12]PO?KA J,REBECCHI A,PISACANE V,et al.Bacterial diversity in typical Italian salami at different ripening stages as revealed by highthroughput sequencing of 16S rRNA amplicons[J].Food Microbiology,2015,46:342-356.DOI:10.1016/j.fm.2014.08.023.
[13]于美娟,谭欢,马美湖,等.传统固态发酵鱼中细菌群落多样性与品质特征分析[J].食品科学,2017,38(8):86-95.DOI:10.7506/spkx1002-6630-201708015.
[14]MARTIN M.Cutadapt removes adapter sequences from highthroughput sequencing reads[J].Embnet Journal,2011,17(1):10-12.DOI:10.14806/ej.17.1.200.
[15]HAAS B J,GEVERS D,EARL A M,et al.Chimeric 16S rRNAsequence formation and detection in Sanger and 454-pyrosequenced PCR amplicons[J].Genome Research,2011,21(3):494-504.DOI:10.1101/gr.112730.110.
[16]QUAST C,PRUESSE E,YILMAZ P,et al.The SILVA ribosomal RNA gene database project:improved data processing and web-based tools[J].Nucleic Acids Research,2013,41(Database issue):590-596.DOI:10.1093/nar/gks1219.
[17]李迎楠,刘文营,成晓瑜.GC-MS结合电子鼻分析温度对肉味香精风味品质的影响[J].食品科学,2016,37(14):104-109.DOI:10.7506/spkx1002-6630-201614018.
[18]米瑞芳,陈曦,戚彪,等.乳杆菌发酵剂对酸肉挥发性风味成分的影响[J].肉类研究,2018,32(4):48-55.DOI:10.7506/rlyj1001-8123-201804009.
[19]GUTSCHE K A,TRAN T B T,VOGEL R F.Production of volatile compounds by Lactobacillus sakei from branched chainα-keto acids[J].Food Microbiology,2012,29(2):224-228.DOI:10.1016/j.fm.2011.06.010.
[20]周才琼,李艳芳,杜木英,等.渝黔地区传统酸肉发酵过程中微生物区系研究[J].食品工业科技,2010,31(4):171-175.
[21]FONTANA C,BASSI D,LóPEZ C,et al.Microbial ecology involved in the ripening of naturally fermented llama meat sausages.A focus on lactobacilli diversity[J].International Journal of Food Microbiology,2016,236:17-25.DOI:10.1016/j.ijfoodmicro.2016.07.002.
[22]CHEN C,CHEN X H,JIANG M,et al.A newly discovered bacteriocin from Weissella hellenica D1501 associated with Chinese Dong fermented meat (Nanx Wudl)[J].Food Control,2014,42(2):116-124.DOI:10.1016/j.foodcont.2014.01.031.
[23]余冰,周红丽,李宗军.固相微萃取分析发酵肉制品中的挥发性风味组分[J].湖南农业大学学报(自然科学版),2007,33(2):232-234.DOI:10.13331/j.cnki.jhau.2007.02.026.
[24]周才琼,代小容,杜木英.酸肉发酵过程中挥发性风味物质形成的研究[J].食品科学,2010,31(7):98-104.
[25]黄群,滕慧,宋洪波,等.固相微萃取结合GC-MS分析湘西酸肉挥发性成分[J].食品工业,2016,37(11):277-280.
[26]SIDIRA M,KANDYLIS P,KANELLAKI M,et al.Effect of curing salts and probiotic cultures on the evolution of flavor compounds in dry-fermented sausages during ripening[J].Food Chemistry,2016,201:334-338.DOI:10.1016/j.foodchem.2016.01.084.
[27]朱健辉.不同等级金华火腿蛋白质和脂肪水解产物与挥发性风味物质分析[D].南京:南京农业大学,2005:50.
[28]JELE?H,MAJCHER M,GINJA A,et al.Determination of compounds responsible for tempeh aroma[J].Food Chemistry,2013,141(1):459-465.DOI:10.1016/j.foodchem.2013.03.047.
[29]LEROY F,VERLUYTEN J,DE V L.Functional meat starter cultures for improved sausage fermentation[J].International Journal of Food Microbiology,2006,106(3):270-285.DOI:10.1016/j.ijfoodmicro.2005.06.027.
[30]CHEN Q,LIU Q,SUN Q X,et al.Flavour formation from hydrolysis of pork sarcoplasmic protein extract by a unique LAB culture isolated from Harbin dry sausage[J].Meat Science,2015,100:110-117.DOI:10.1016/j.meatsci.2014.10.001.
[31]褚福娟,孔保华,黄永.发酵肉制品常见微生物及其对风味的影响[J].肉类研究,2008,22(1):3-6.DOI:10.3969/j.issn.1001-8123.2008.01.004.
[32]KARGOZARI M,MOINI S,AKHONDZADEH B A,et al.Effect of autochthonous starter cultures isolated from Siahmazgi cheese on physicochemical,microbiological and volatile compound profiles and sensorial attributes of sucuk,a Turkish dry-fermented sausage[J].Meat Science,2014,97(1):104-114.DOI:10.1016/j.meatsci.2014.01.013.
[33]SIDIRA M,KANDYLIS P,KANELLAKI M,et al.Effect of immobilized Lactobacillus casei on the evolution of flavor compounds in probiotic dry-fermented sausages during ripening[J].Meat Science,2015,100:41-51.DOI:10.1016/j.meatsci.2014.09.011.
[34]黄业传,李婷婷,龚道楷,等.高压结合热处理对猪肉风味的影响[J].食品科学,2018,39(2):227-232.DOI:10.7506/spkx1002-6630-201802036.
[35]赵健,王蒙,谢建春,等.黑猪肉关键香气物质分析鉴定[J].食品科学,2018,39(2):203-209.DOI:10.7506/spkx1002-6630-201802032.
[36]MONTANARI C,BARGOSSI E,GARDINI A,et al.Correlation between volatile profiles of Italian fermented sausages and their size and starter culture[J].Food Chemistry,2016,192:736-744.DOI:10.1016/j.foodchem.2015.07.062.
[37]KOT W,NEVE H,HELLER K J,et al.Bacteriophages of Leuconostoc,Oenococcus,and Weissella[J].Front Microbiol,2014,5(2):186-195.DOI:10.3389/fmicb.2014.00186.