中温乳化肠中凝结芽孢杆菌芽孢萌发及热致死规律
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摘要
从中温乳化香肠中筛选出一株凝结芽孢杆菌芽孢,以20种氨基酸、D-果糖、D-葡萄糖、氯化钾为萌发剂,通过相差显微镜及生长曲线测定仪测定600 nm波长下光密度值,研究该芽孢萌发规律,同时研究7种防腐剂对芽孢萌发效果影响及不同温度热胁迫处理后的芽孢致死率。结果表明:葡萄糖可以促进凝结芽孢杆菌芽孢萌发,增大其浓度对芽孢萌发率无影响;绘制了芽孢萌发率标准曲线,相关系数为0.977 8;亚硝酸钠、山梨酸钾、乳酸链球菌素(nisin)和脱氢乙酸钠对芽孢萌发无影响,乳酸钠、葡萄糖酸-δ-内酯和双乙酸钠可以抑制芽孢萌发;采用90℃及95℃对芽孢热胁迫致死效果较差,100℃热处理最高致死率可达92.09%,105~120℃热胁迫处理对该芽孢致死率在15 min内可以接近100%。
The spore germination of a Bacillus coagulans stain isolated from medium-temperature sterilized emulsified sausage was investigated using 20 amino acids,D-fructose,D-glucose and potassium chloride asgermination inducers by phase contrast microscopic morphology observation and measurement of optical density(OD) values at 600 nm with a Bioscreen growth analyzer.Also,we examined the effect of seven preservatives on spore germination and we evaluated spore inactivation after heat treatment at different temperatures.Results showed that glucose could promote the germination of Bacillus coagulans spore,and the spore germination percentage did not changed with increasing glucose concentration.A standard curve with a correlation coefficient of 0.977 8 was established to determine the spore germination percentage.Sodium nitrite,potassium sorbate,nisin and dehydrogenation sodium acetate had no effects on spore germination,whereas sodium lactate,glucose acid-δ-lactone and sodium acetate could inhibit spore germination.When treated at 100℃,the inactivation percentage was up to 92.09%,and at 105-120℃,the value was almost 100%within 15 min;however,the spores were not effectively inactivated at 90 and 95℃.
The spore germination of a Bacillus coagulans stain isolated from medium-temperature sterilized emulsified sausage was investigated using 20 amino acids,D-fructose,D-glucose and potassium chloride asgermination inducers by phase contrast microscopic morphology observation and measurement of optical density(OD) values at 600 nm with a Bioscreen growth analyzer.Also,we examined the effect of seven preservatives on spore germination and we evaluated spore inactivation after heat treatment at different temperatures.Results showed that glucose could promote the germination of Bacillus coagulans spore,and the spore germination percentage did not changed with increasing glucose concentration.A standard curve with a correlation coefficient of 0.977 8 was established to determine the spore germination percentage.Sodium nitrite,potassium sorbate,nisin and dehydrogenation sodium acetate had no effects on spore germination,whereas sodium lactate,glucose acid-δ-lactone and sodium acetate could inhibit spore germination.When treated at 100℃,the inactivation percentage was up to 92.09%,and at 105-120℃,the value was almost 100%within 15 min;however,the spores were not effectively inactivated at 90 and 95℃.
引文
[1]余岳,王春维,祝爱侠,等.一株凝结芽孢杆菌芽孢抗逆性研究及其产孢条件优化[J].饲料工业,2013(7):43-47.DOI:10.3969/j.issn.1001-991X.2013.07.012.
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[13]AVILA M,G MEZ-TORRES N,HERN NDEZ M,et al.Inhibitory activity of reuterin,nisin,lysozyme and nitrite against vegetative cells and spores of dairy-related Clostridium species[J].International Journal of Food Microbiology,2014,172(7):70-75.DOI:10.1016/j.ijfoodmicro.2013.12.002.
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[15]ARAN N.The effect of calcium and sodium lactates on growth from spores of Bacillus cereus and Clostridium perfringens in a'sousvide'beef goulash under temperature abuse[J].International Journal of Food Microbiology,2001,63(1/2):117-23.DOI:10.1016/S0168-1605(00)00412-8.
[16]VELUGOTI P R,RAJAGOPAL L,JUNEJA V,et al.Use of calcium,potassium,and sodium lactates to control germination and outgrowth of Clostridium perfringens spores during chilling of injected pork[J].Food Microbiology,2007,24(7/8):687-694.DOI:10.1016/j.fm.2007.04.004.
[17]王涛.食品组分与抑菌剂对嗜热脂肪芽孢杆菌芽孢耐热性的影响[D].无锡:江南大学,2011.
[18]WESCHE A M,GURTLERJ B,MARKS B P,et al.Stress,sublethal injury,resuscitation,and virulence of bacterial food borne pathogens[J].Journal of Food Protection,2009,72(5):1121-1138.
[19]NORIEGA E,VELLIOU E,DERLINDEN E V,et al.Effect of cell immobilization on heat-induced sublethal injury of Escherichia coli,Salmonella,Typhimurium and Listeria innocua[J].Food Microbiology,2013,36(36):355-64.DOI:10.1016/j.fm.2013.06.015.
[20]宣晓婷,丁甜,刘东红.食品中亚致死损伤单增李斯特菌的研究进展[J].食品科学,2015,36(3):280-284.DOI:10.7506/spkx1002-6630-201503052.
[21]林丽花,柯芙容,詹湉湉,等.凝结芽孢杆菌对黄羽肉鸡生产性能、血清生化指标及抗氧化功能的影响[J].动物营养学报,2014,26(12):3806-3813.DOI:10.3969/j.issn.1006-267x.2014.12.034.
[22]董惠钧,姜俊云,郑立军,等.新型微生态益生菌凝结芽孢杆菌研究进展[J].食品科学,2010,31(1):292-294.
[23]VERCAMMEN A,VIVIJ S B,LURQUIN I,et al.Germination and inactivation of Bacillus coagulans and Alicyclobacillus acidoterrestris spores by high hydrostatic pressure treatment in buffer and tomato sauce[J].International Journal of Food Microbiology,2012,152(3):162-167.DOI:10.1016/j.ijfoodmicro.2011.02.019.
[24]DARYAEI H,BALASUBRAMANIAM V M.Kinetics of Bacillus coagulans spore inactivation in tomato juice by combined pressureheat treatment[J].Food Control,2013,30(1):168-175.DOI:10.1016/j.foodcont.2012.06.031.
[25]ZIMMERMANN M,SCHAFFNER D W,ARAG O G M F.Modeling the inactivation kinetics of Bacillus coagulans spores in tomato pulp from the combined effect of high pressure and moderate temperature[J].LWT-Food Science and Technology,2013,53(1):107-112.DOI:10.1016/j.lwt.2013.01.026.
[26]PENGJ,MAHJ H,SOMAVAT R,et al.Thermal inactivation kinetics of Bacillus coagulans spores in tomato juice[J].Joumal of Food Protection,2012,75(7):1236-1242.DOI:10.4315/0362-028X.J FP-11-490.
[27]潘晓倩,赵燕,张顺亮,等.中温乳化香肠中一株优势腐败菌的分离鉴定与生物学特性[J].食品科学,2016,37(7):93-98.DOI:10.7506/spkx1002-6630-201607018.
[28]ICIEK J,PAPIEWSKA A,MOLSKA M.Inactivation of Bacillus stearothermophilus spores during thermal processing[J].Journal of Food Engineering,2006,77(3):406-410.DOI:10.1016/j.jfoodeng.2005.07.005
[29]中华人民共和国卫生部.GB/T 4789.2—2010食品微生物学检验菌落总数测定[S].北京:中国标准出版社,2010.
[30]中华人民共和国国家卫生和计划生育委员会.GB 2760—2011食品添加剂使用标准[S].北京:中国标准出版社,2014.
[31]KORZA G,SETLOW P.Topology and accessibility of germination proteins in the Bacillus subtilis spore inner membrane[J].Journal of Bacteriology,2013,195(7):1484-1491.DOI:10.1128/J B.02262-12.
[32]GHOSH S,SETLOW P.The preparation,germination properties and stability of superdormant spores of Bacillus cereus[J].Journal of Applied Microbiology,2010,108(2):582-590.DOI:10.1111/j.1365-2672.2009.04442.x.
[33]MORRIS S L,WALSH R C,HANSEN J N.Identification and characterization of some bacterial membrane sulfhydryl groups which are targets of bacteriostatic and antibiotic action[J].Journal of Biological Chemistry,1984,259(21):13590-13594.
[34]GUT I M,BLANKE S R,DONK W A V D.Mechanism of inhibition of Bacillus anthracis spore outgrowth by the lantibiotic nisin[J].Acs Chemical Biology,2011,6(7):744-752.DOI:10.1021/cb1004178.
[35]AOUADHI C,ROUISSI Z,MEJRI S,et al.Inactivation of Bacillus sporothermodurans spores by nisin and temperature studied by design of experiments in water and milk[J].Food Microbiology,2014,38(2):270-275.DOI:10.1016/j.fm.2013.10.005.
[2]艾婷,张顺亮,王守伟,等.中温杀菌对乳化肠挥发性风味物质变化的影响[J].肉类研究,2014,28(12):6-9.
[3]韩衍青,徐宝才,徐幸莲,等.真空包装熟肉制品中的特定腐败微生物及其控制[J].中国食品学报,2011,11(7):148-156.DOI:10.16429/j.1009-7848.2011.07.028.
[4]DOULGERAKI A I,ERCOLINI D,VILLANI F,et al.Spoilage microbiota associated to the storage of raw meat in different conditions[J].International Journal of Food Microbiology,2012,157(2):130-141.DOI:10.1016/j.ijfoodmicro.2012.05.020.
[5]YOST C K,NATTRESS F M.The use of multiplex PCR reactions to characterize populations of lactic acid bacteria associated with meat spoilage[J].Letters in Applied Microbiology,2000,31:129-133.DOI:10.1046/j.1365-2672.2000.00776.x.
[6]CHENOLL E,MACIAN M C,ELIZAQUIVEL P,et al.Lactic acid bacteria associated with vacuum-packed cooked meatproduct spoilage:population analysis by rDNA-based methods[J].Journal of Applied Microbiology,2007,102:498-508.DOI:10.1111/j.1365-2672.2006.03081.x.
[7]MATAMOROS S,ANDR S,HUE I,et al.Identification of lactic acid bacteria involved in the spoilage of pasteurized"foie gras"products[J].Meat Science,2010,85(3):467-471.DOI:10.1016/j.meatsci.2010.02.017.
[8]ALFARO B,HERNANDEZ I,le MARC Y,et al.Modelling the effect of the temperature and carbon dioxide on the growth of spoilage bacteria in packed fish products[J].Food Control,2013,29(2):429-437.DOI:10.1016/j.foodcont.2012.05.046.
[9]GHOSH S,SETLOW P.The preparation,germination properties and stability of superdormant spores of Bacillus cereus[J].Journal of Applied Microbiology,2010,108(2):582-590.DOI:10.1111/j.1365-2672.2009.04442.x.
[10]GEORGET E,KUSHMAN A,CALLANAN M,et al.Geobacillus stearothermophilus,ATCC 7953 spore chemical germination mechanisms in model systems[J].Food Control,2014,50:141-149.DOI:10.1016/j.foodcont.2014.08.044.
[11]BLACK E P,KOZIOLDUBE K,GUAN D,et al.Factors influencing germination of Bacillus subtilis spores via activation of nutrient receptors by high pressure[J].Applied and Environmental Microbiology,2005,71(10):5879-5887.DOI:10.1128/AEM.71.10.5879-5887.2005.
[12]NANGUY S P M,PERRIER-CORNET J M,BENSOUSSAN M,et al.Impact of water activity of diverse media on spore germination of Aspergillus,and Penicillium species[J].International Journal of Food Microbiology,2010,142(1/2):273-276.DOI:10.1016/j.ijfoodmicro.2010.06.031.
[13]AVILA M,G MEZ-TORRES N,HERN NDEZ M,et al.Inhibitory activity of reuterin,nisin,lysozyme and nitrite against vegetative cells and spores of dairy-related Clostridium species[J].International Journal of Food Microbiology,2014,172(7):70-75.DOI:10.1016/j.ijfoodmicro.2013.12.002.
[14]AOUADHI C,MEJRI S,MAAROUFI A.Inhibitory effects of nisin and potassium sorbate alone orin combination on vegetative cells growth and spore germination of Bacillus sporothermodurans in milk[J].Food Microbiology,2015,46(101):40-45.DOI:10.1016/j.fm.2014.07.004.
[15]ARAN N.The effect of calcium and sodium lactates on growth from spores of Bacillus cereus and Clostridium perfringens in a'sousvide'beef goulash under temperature abuse[J].International Journal of Food Microbiology,2001,63(1/2):117-23.DOI:10.1016/S0168-1605(00)00412-8.
[16]VELUGOTI P R,RAJAGOPAL L,JUNEJA V,et al.Use of calcium,potassium,and sodium lactates to control germination and outgrowth of Clostridium perfringens spores during chilling of injected pork[J].Food Microbiology,2007,24(7/8):687-694.DOI:10.1016/j.fm.2007.04.004.
[17]王涛.食品组分与抑菌剂对嗜热脂肪芽孢杆菌芽孢耐热性的影响[D].无锡:江南大学,2011.
[18]WESCHE A M,GURTLERJ B,MARKS B P,et al.Stress,sublethal injury,resuscitation,and virulence of bacterial food borne pathogens[J].Journal of Food Protection,2009,72(5):1121-1138.
[19]NORIEGA E,VELLIOU E,DERLINDEN E V,et al.Effect of cell immobilization on heat-induced sublethal injury of Escherichia coli,Salmonella,Typhimurium and Listeria innocua[J].Food Microbiology,2013,36(36):355-64.DOI:10.1016/j.fm.2013.06.015.
[20]宣晓婷,丁甜,刘东红.食品中亚致死损伤单增李斯特菌的研究进展[J].食品科学,2015,36(3):280-284.DOI:10.7506/spkx1002-6630-201503052.
[21]林丽花,柯芙容,詹湉湉,等.凝结芽孢杆菌对黄羽肉鸡生产性能、血清生化指标及抗氧化功能的影响[J].动物营养学报,2014,26(12):3806-3813.DOI:10.3969/j.issn.1006-267x.2014.12.034.
[22]董惠钧,姜俊云,郑立军,等.新型微生态益生菌凝结芽孢杆菌研究进展[J].食品科学,2010,31(1):292-294.
[23]VERCAMMEN A,VIVIJ S B,LURQUIN I,et al.Germination and inactivation of Bacillus coagulans and Alicyclobacillus acidoterrestris spores by high hydrostatic pressure treatment in buffer and tomato sauce[J].International Journal of Food Microbiology,2012,152(3):162-167.DOI:10.1016/j.ijfoodmicro.2011.02.019.
[24]DARYAEI H,BALASUBRAMANIAM V M.Kinetics of Bacillus coagulans spore inactivation in tomato juice by combined pressureheat treatment[J].Food Control,2013,30(1):168-175.DOI:10.1016/j.foodcont.2012.06.031.
[25]ZIMMERMANN M,SCHAFFNER D W,ARAG O G M F.Modeling the inactivation kinetics of Bacillus coagulans spores in tomato pulp from the combined effect of high pressure and moderate temperature[J].LWT-Food Science and Technology,2013,53(1):107-112.DOI:10.1016/j.lwt.2013.01.026.
[26]PENGJ,MAHJ H,SOMAVAT R,et al.Thermal inactivation kinetics of Bacillus coagulans spores in tomato juice[J].Joumal of Food Protection,2012,75(7):1236-1242.DOI:10.4315/0362-028X.J FP-11-490.
[27]潘晓倩,赵燕,张顺亮,等.中温乳化香肠中一株优势腐败菌的分离鉴定与生物学特性[J].食品科学,2016,37(7):93-98.DOI:10.7506/spkx1002-6630-201607018.
[28]ICIEK J,PAPIEWSKA A,MOLSKA M.Inactivation of Bacillus stearothermophilus spores during thermal processing[J].Journal of Food Engineering,2006,77(3):406-410.DOI:10.1016/j.jfoodeng.2005.07.005
[29]中华人民共和国卫生部.GB/T 4789.2—2010食品微生物学检验菌落总数测定[S].北京:中国标准出版社,2010.
[30]中华人民共和国国家卫生和计划生育委员会.GB 2760—2011食品添加剂使用标准[S].北京:中国标准出版社,2014.
[31]KORZA G,SETLOW P.Topology and accessibility of germination proteins in the Bacillus subtilis spore inner membrane[J].Journal of Bacteriology,2013,195(7):1484-1491.DOI:10.1128/J B.02262-12.
[32]GHOSH S,SETLOW P.The preparation,germination properties and stability of superdormant spores of Bacillus cereus[J].Journal of Applied Microbiology,2010,108(2):582-590.DOI:10.1111/j.1365-2672.2009.04442.x.
[33]MORRIS S L,WALSH R C,HANSEN J N.Identification and characterization of some bacterial membrane sulfhydryl groups which are targets of bacteriostatic and antibiotic action[J].Journal of Biological Chemistry,1984,259(21):13590-13594.
[34]GUT I M,BLANKE S R,DONK W A V D.Mechanism of inhibition of Bacillus anthracis spore outgrowth by the lantibiotic nisin[J].Acs Chemical Biology,2011,6(7):744-752.DOI:10.1021/cb1004178.
[35]AOUADHI C,ROUISSI Z,MEJRI S,et al.Inactivation of Bacillus sporothermodurans spores by nisin and temperature studied by design of experiments in water and milk[J].Food Microbiology,2014,38(2):270-275.DOI:10.1016/j.fm.2013.10.005.