热压结合处理对低温火腿中耐压腐败菌的抑制作用
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摘要
本文主要探究热压结合处理(50℃,350 MPa保压10 min)对低温火腿中绿色魏斯菌和肠膜明串珠菌的抑制作用。通过薄层平板计数法对热压结合处理的杀菌效果进行判断,使用流式细胞技术判断细胞的生理状态,通过扫描电镜(scanning electron microscope,SEM)和透射电镜(transmission electron microscope,TEM)观察细胞形态结构的变化,利用紫外分光光度法测定热压结合处理对细菌细胞膜通透性的影响。结果表明:经热压结合处理后,绿色魏斯菌和肠膜明串珠菌的存活率分别为-4.74和-6.61lg(CFU/m L),其生长变得迟缓,对数期生长速率降低。扫描电镜结果显示热压结合处理对两种菌的形态造成明显的变形,透射电镜结果显示细胞内部出现团状聚集和透电子区。热压结合处理对细胞膜产生损伤,细胞膜通透性变大,导致细胞外紫外吸收物质也极显著增加(p <0.001),细胞处于亚致死状态。热压结合处理会对细胞膜产生一定的形变及损坏,使得细胞内核酸和蛋白质流出,同时细胞内部也会发生蛋白变性和细胞质皱缩,达到抑制耐压菌生长繁殖的目的。
The inhibition of high pressure thermal sterilization treatment(350 MPa and 50 ℃ for 10 min) on Weissella viridescens and Leuconostoc mesenteroides of low temperature cooked ham were investigated in the study.The germicidal efficacy of high pressure thermal sterilization was evaluated by thin layer plate counting method. The flow cytometry was used to determine the physiological state of the cell,SEM and TEM were used to observe cell morphological changes and ultrastructure of bacteria.The influence of cell membrane permeability was evaluated by UV spectrophotometry.The results showed that,after high pressure thermal sterilization,the survival rate of Weissella viridescens and Leuconostoc mesenteroides was-4.74 and-6.61lg(CFU/m L),respectively. These two baroduric bacteria appeared growth retardation and lower growth rate in exponential phase under high-pressure thermal sterilization. SEM and TEM results showed high pressure thermal sterilization induced obvious morphologic changes of cells and intracellular cytoplasm aggregate with large electron transmission area. The combination of heat and pressure treatment damaged the cell membrane,and made the cell membrane permeability larger,which caused a significant increase in extracellular ultraviolet absorption content(p < 0.001),and the cells would be in a sublethal state.The combination of heat and pressure would cause certain deformation and damage to the cell membrane,which would lead to the flow out of nucleic acid and protein in cell,and the protein denaturation and cytoplasm shrinkage would occur inside the cell,thereby inhibiting the growth and reproduction of the pressure-resistant bacteria.
The inhibition of high pressure thermal sterilization treatment(350 MPa and 50 ℃ for 10 min) on Weissella viridescens and Leuconostoc mesenteroides of low temperature cooked ham were investigated in the study.The germicidal efficacy of high pressure thermal sterilization was evaluated by thin layer plate counting method. The flow cytometry was used to determine the physiological state of the cell,SEM and TEM were used to observe cell morphological changes and ultrastructure of bacteria.The influence of cell membrane permeability was evaluated by UV spectrophotometry.The results showed that,after high pressure thermal sterilization,the survival rate of Weissella viridescens and Leuconostoc mesenteroides was-4.74 and-6.61lg(CFU/m L),respectively. These two baroduric bacteria appeared growth retardation and lower growth rate in exponential phase under high-pressure thermal sterilization. SEM and TEM results showed high pressure thermal sterilization induced obvious morphologic changes of cells and intracellular cytoplasm aggregate with large electron transmission area. The combination of heat and pressure treatment damaged the cell membrane,and made the cell membrane permeability larger,which caused a significant increase in extracellular ultraviolet absorption content(p < 0.001),and the cells would be in a sublethal state.The combination of heat and pressure would cause certain deformation and damage to the cell membrane,which would lead to the flow out of nucleic acid and protein in cell,and the protein denaturation and cytoplasm shrinkage would occur inside the cell,thereby inhibiting the growth and reproduction of the pressure-resistant bacteria.
引文
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[28]蒋娜.番茄溃疡病菌VBNC状态的诱导、复苏及机制研究[D].北京:中国农业大学,2016.
[29]陆海霞,黄小鸣,朱军莉.超高压对单增李斯特菌细胞膜的损伤和致死机理[J].微生物学报,2014,54(7):746-753.
[30]王平平.镍对玉米幼苗生长及DNA损伤效应的影响研究[D].长春:吉林农业大学,2015.
[31]张东晨,张明旭,陈清如,等.脱硫微生物氧化亚铁硫杆菌遗传特性的分子生物学试验[J].煤炭学报,2006,31(1):104-107.
[2]徐雅梦,姜晓冰,于涛,等.真空包装低温熟牛肉中腐败菌的分离与鉴定[J].肉类研究,2017,31(8):23-27.
[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.
[4]韩衍青.应用超高压技术延长低温火腿的货架期[D].南京:南京农业大学,2011.
[5]全拓.肉制品中主要微生物的检测与研究[D].重庆:西南大学,2012.
[6]Naderi N,House J D,Pouliot Y,et al.Effects of high hydrostatic pressure processing on hen egg compounds and egg products[J].Comprehensive Reviews in Food Science and Food Safety,2017,16(4):707-720.
[7]费楠,李芳菲,党苗苗,等.非热杀菌技术特点及在肉制品加工中的应用[J].食品安全质量检测学报,2015(2):540-544.
[8]刘蓄瑾,胡志和,刘军军,等.超高压和热杀菌对黄金贝可消化性和食用品质的影响[J].食品科学,2016,37(7):72-77.
[9]Clérybarraud C,Gaubert A,Masson P,et al.Combined effects of high hydrostatic pressure and temperature for inactivation of Bacillus anthracis spores[J].Applied and Environmental Microbiology,2004,70(1):635-637.
[10]曾庆梅,谢慧明,潘见,等.超高压处理对枯草芽孢杆菌超微结构的影响[J].高压物理学报,2006,20:83-87
[11]高芳.热压结合处理对肉品中耐压腐败菌的失活动力学研究[D].北京:中国农业大学,2017.
[12]Balasubramaniam V M,Martínezmonteagudo S I,Gupta R.Principles and application of high pressure-based technologies in the food industry[J].Annual Review of Food Science and Technology,2015,6(435-462):435.
[13]王莉.二氧化碳协同食品超高压杀菌研究[D].合肥:合肥工业大学,2012.
[14]陈婵娟,宁喜斌.亚致死热损伤和复苏状态的副溶血性弧菌特性[J].食品工业科技,2013,34(14):160-165.
[15]莘似韵,孙晓红,徐忆宁,等.超高压对单增李斯特菌生物被膜形成的影响[J].上海海洋大学学报,2017,26(2):294-300.
[16]钱天乐,周逸卿,邹珍友,等.微生物扫描电镜样品清洗方法的改进与固定干燥方法比较[J].安徽农业科学,2009,37(23):10886-10888.
[17]Chiang M L,Ho W L,Chou C C.Response of Vibrio parahaemolyticus to ethanol shock[J].Food Microbiology,2006,23(5):461-467.
[18]Müller S,Nebe-von-Caron G.Functional single-cell analyses:low cytometry and cell sorting of microbial populations and communities[J].FEMS Microbiology Reviews,2010,34(4):554-587.
[19]Soejima T,Iida K,Qin T,et al.Discrimination of live,antituberculosis agent-injured,and dead Mycobacterium tuberculosis using flow cytometry[J].FEMS Microbiology Letters,2009,294(1):74-81.
[20]Klotz B,Ma1as P,Mackey B M.The relationship between membrane damage,release of protein and loss of viability in Escherichia coli exposed to high hydrostatic pressure[J].International Journal of Food Microbiology,2010,137(2-3):214-220.
[21]姚春艳,刘美凤,付瑞东,等.加压CO2对大肠杆菌细胞膜的损伤作用[J].微生物学通报,2013,40(2):256-265.
[22]孙新生,徐幸莲,韩衍青,等.超高压协同温度处理对烟熏火腿中单增李斯特菌生长预测模型的建立[J].食品工业科技,2012,33(13):118-121.
[23]季宇彬,王胜惠,高世勇,等.龙葵碱对H_(22)荷瘤小鼠细胞膜流动性和膜蛋白水平的影响[J].中草药,2005,36:239-241.
[24]郭红莲,曹勤红,任东涛,等.激光光钳法细胞膜弹性测量技术的建立和白细胞膜弹性的测量[J].科学通报,2003,48:246-251.
[25]杭瑜瑜,陆海霞,励建荣.超高压处理对副溶血性弧菌的影响[J].微生物学报,2009,49(11):1489-1493.
[26]VerthéK,Verstraete W.Use of flow cytometry for analysis of phage-mediated killing of Enterobacter aerogenes[J].Research in Microbiology,2006,157(7):613-618.
[27]张梦露,党志,伍凤姬,等.利用流式细胞术研究鞘氨醇单胞菌GY2B降解菲过程中细菌表面特性的变化[J].环境科学,2014,35(4):1449-1456.
[28]蒋娜.番茄溃疡病菌VBNC状态的诱导、复苏及机制研究[D].北京:中国农业大学,2016.
[29]陆海霞,黄小鸣,朱军莉.超高压对单增李斯特菌细胞膜的损伤和致死机理[J].微生物学报,2014,54(7):746-753.
[30]王平平.镍对玉米幼苗生长及DNA损伤效应的影响研究[D].长春:吉林农业大学,2015.
[31]张东晨,张明旭,陈清如,等.脱硫微生物氧化亚铁硫杆菌遗传特性的分子生物学试验[J].煤炭学报,2006,31(1):104-107.
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