乳酸对阴沟肠杆菌的抑菌作用及其机制分析

以自主分离的一株冰鲜鸡肉产品的优势腐败菌——阴沟肠杆菌为指示菌,研究乳酸对该菌的抑菌机制。结果发现各质量分数乳酸（1.0%、0.5%、0.25%）均对阴沟肠杆菌具有一定的杀菌效果;乳酸处理后菌体的细胞电势增加,ATP和紫外吸收物质有明显的泄漏;透射电镜观察发现,乳酸处理后的菌株形态变得不规则,细胞壁损伤明显,细胞内容物渗出;通过激光扫描共聚焦显微镜和流式细胞仪检测也发现,乳酸处理后的菌株细胞通透性增加。因此,乳酸抑菌效应主要通过作用于细胞壁,导致细胞壁损伤和细胞通透性增加,改变细胞内外电势,造成内容物的泄漏,发挥杀菌作用。     

α-乙酰乳酸脱羧酶基因在酿酒酵母中稳定表达的策略

综述了宿主细胞的表达特点，外源基因的来源，表达载体和转化方法对α-乙酰乳酸脱羧酶基因在酿酒酵母中稳定表达的影响，并在此基础上提出了相应的策略。     

奶粉中阴沟肠杆菌的检测方法研究

本研究发现对奶粉中低含量阴沟肠杆菌的检测,可以采用灭菌蒸馏水36℃增菌24h后,选用自主设计的阴沟肠杆菌分离琼脂进行选择性分离,再用赖氨酸脱羧酶、鸟氨酸脱羧酶、精氨酸双水解酶这一组生化实验进行进一步筛选,最后选择ID 32E或VITEK GNI+进行生化鉴定.     

含α-乙酰乳酸脱羧酶基因的啤酒酵母工程菌的构建

摘要：利用基因工程手段将食品级醋化醋杆菌的α-乙酰乳酸脱羧酶基因引入啤酒酵母中，使啤酒酵母获得编码此酶的基因并表达，检测到α-乙酰乳酸脱羧酶的活性，获得一株性能优良的啤酒酵母工程菌。经测试，构建的工程菌与出发菌在生理、生化、发酵特性等方面无差异，但发酵液中双乙酰的峰值及还原时间较出发菌降低1／3左右。     

α-乙酰乳酸脱羧酶应用的研究

使用α-乙酰乳酸脱羧酶，可减少双乙酰还原时间，使发酵周期缩短4d，降低双乙酰峰值，成品啤酒双乙酰反弹幅度小，延长啤酒保存期，提升啤酒质量和稳定性。     

香芹酚对阴沟肠杆菌的抑菌作用及机理

香芹酚具有较好的抑菌效果,符合人们对食品防腐剂安全无毒、无副作用的要求。本研究通过最小抑菌浓度和杀菌实验分析了香芹酚对阴沟肠杆菌（Enterobacter cloacae C4）的抑菌活性,并检测其对阴沟肠杆菌细胞膜电势、胞外ATP化学发光值的影响,使用激光共聚焦显微镜观察细胞渗透性,并通过扫描电子显微镜观察香芹酚对阴沟肠杆菌细胞形态的影响,从而探究香芹酚对阴沟肠杆菌的抑菌机理。结果表明：香芹酚对阴沟肠杆菌的最小抑菌浓度为0.5 μL/mL;阴沟肠杆菌细胞膜通透性随着香芹酚含量（0.25～2 μL/mL）的增加而逐渐增加。经香芹酚处理的阴沟肠杆菌细胞膜内外电势改变,ATP大量漏出。通过扫描电子显微镜观察,发现经香芹酚处理的菌体细胞形态干瘪且明显被破坏,含量高于1 μL/mL的香芹酚几乎能够溶解菌体。推测香芹酚通过作用于细胞膜改变膜内外电势,造成细胞膜通透性改变,引起细胞ATP等内容物泄露,从而影响细胞功能,达到抑菌的作用。     

国产α-乙酰乳酸脱羧酶在啤酒生产中的应用

双乙酰的前驱物质是α-乙酰乳酸,在生产过程中添加α-乙酰乳酸脱羧酶可迅速降解α-乙酰乳酸,从而降低双乙酰含量,缩短发酵周期.通过两年来对进口和国产α-乙酰乳酸脱羧酶的试用比较,认为国产α-乙酰乳酸脱羧酶的使用效果完全可靠,能够替代进口同类酶制剂.     

百里香酚对食源阴沟肠杆菌生物膜形成的抑制作用

本文研究了百里香酚在亚抑菌浓度条件下对阴沟肠杆菌(Enterobacter cloacae) C4生物膜形成的抑制作用。首先确定百里香酚对E.cloacae C4的最小抑菌浓度,然后测定亚抑菌浓度的百里香酚对生物膜内菌落总数、菌体泳动能力和胞外多糖含量的影响,观察生物膜内的细菌形态以及三维结构的变化,并进一步测定与生物膜形成相关基因的相对表达量。结果表明:百里香酚对该菌的最小抑菌浓度(MIC)为256 μg/mL。1/4 MIC百里香酚在不影响浮游细菌生长和生物膜内活菌总数的情况下,显著(P<0.05)抑制菌体的泳动能力和生物膜内胞外多糖的合成,与生物膜形成相关(细胞粘附和胞外多糖合成)基因相对表达量显著下调。百里香酚处理后生物膜内细菌菌体变长、生物膜厚度降低,膜结构松散。因此百里香酚在亚抑菌浓度下对阴沟肠杆菌C4生物膜形成有抑制作用。     

树脂固定化α-乙酰乳酸脱羧酶的初步研究

采用7种树脂对α-乙酰乳酸脱羧酶进行固定化,并将固定化酶应用于啤酒发酵实验,从中筛选出降低发酵液中双乙酰能力较强的大孔弱碱性丙烯酸系阴离子交换树脂D314FD作为固定化的载体。通过单因素和正交实验确定固定化ALDC的最优条件为:每克树脂加入1.25mL原酶液,吸附时间10h,吸附pH7.5,吸附温度15℃,在此条件下固定化效率可达44%。发酵实验亦表明,固定化ALDC可以显著降低发酵液中双乙酰的含量,降低率达40%以上。      

树脂固定化α-乙酰乳酸脱羧酶的初步研究

采用7种树脂对α-乙酰乳酸脱羧酶进行固定化,并将固定化酶应用于啤酒发酵实验,从中筛选出降低发酵液中双乙酰能力较强的大孔弱碱性丙烯酸系阴离子交换树脂D314FD作为固定化的载体。通过单因素和正交实验确定固定化ALDC的最优条件为:每克树脂加入1.25mL原酶液,吸附时间10h,吸附pH7.5,吸附温度15℃,在此条件下固定化效率可达44%。发酵实验亦表明,固定化ALDC可以显著降低发酵液中双乙酰的含量,降低率达40%以上。     

婴幼儿奶粉中阪崎肠杆菌双重荧光PCR快速检测方法的建立

旨在建立针对阪崎肠杆菌的双重荧光PCR快速检测方法。以阪崎肠杆菌局部大分子合成(MMS)操纵子和外膜蛋白A(ompA)为靶基因,建立双重荧光PCR反应体系,探讨该体系的特异性、灵敏度和抗干扰能力。结果表明,双重荧光PCR体系对阪崎肠杆菌的灵敏度为4.3×10³ CFU/mL,人工污染初始菌量为2 CFU/100 g奶粉样品增菌24 h即可检出;39株实验菌中的15株阪崎肠杆菌出现特异性扩增,24株非阪崎肠杆菌未出现特异性扩增。本研究所建立的双重荧光PCR体系特异好、灵敏度较高及抗干扰能力强,可用于婴幼儿奶粉中阪崎肠杆菌的快速检测。     

Biochemical characteristics of Enterobacter agglomerans and related strains found in buckwheat seeds

Thirty strains of bacteria were randomly isolated and identified from buckwheat seeds. The phenotypic characteristics of these strains agree well with those of the Enterobacter agglomerans-Erwinia herbicola complex. On the basis of the difference in indole production and gas production from D-glucose, the isolates were divided into 3 phenotypic groups, viz. I, II and III. Twenty two strains were in phenotypic group I, which is negative for indole production and gas production from D-glucose, and resembles Pantoea agglomerons. All six strains in phenotypic group II, which is positive for indole production and negative for gas production from D-glucose, were identified as Erwinia ananas. Two strains in phenotypic group III, which is negative for indole production and positive for gas production from D-glucose, were identified as Rahnella aquatilis.     

Inactivation of Enterobacter sakazakii by pulsed electric field in buffered peptone water and infant formula milk

The effect of high-intensity pulsed electric field (PEF) treatment on the survival of Enterobacter sakazakii suspended in buffered peptone water (BPW) and powdered infant formula milk (IFM) was evaluated. Reference medium and IFM samples were treated with PEF. Electric field intensity and treatment time were varied from 10 to 40 kV cm⁻¹ and from 60 to 3895 μs, respectively. Samples of buffered peptone water (3 g L⁻¹) and IFM were inoculated with E. sakazakii (CECT 858) (10⁹ cfu mL⁻¹) and then treated with PEF. The inactivation data were adjusted to the Weibull frequency distribution function and Bigelow model, and constants were calculated for both substrates. A maximum 2.7 log (cfu mL⁻¹) reduction was achieved in BPW after exposure of E. sakazakii to PEF for 360 μs (2.5 μs pulse width) at 40 kV cm⁻¹. In IFM, exposure of E. sakazakii to PEF, with the same conditions, led to a 1.2 log (cfu mL⁻¹) reduction. The greater the field strength and treatment time, the greater the inactivation achieved in both substrates. Even though further research will be necessary, according to the results, there are good prospects for the use of PEF in hospitals to achieve safe reconstituted infant formula before storage at refrigerated temperatures.     

Isolation of Enterobacter sakazakii and other Enterobacteriaceae from powdered infant formula milk and related products

The presence of Enterobacter sakazakii and other Enterobacteriaceae was surveyed in 82 powdered infant formula milk (IFM) and 404 other food products. The presence of Ent. sakazakii was detected using the conventional method (growth on violet red bile glucose agar plus yellow pigment production on TSA) and a new chromogenic medium (Druggan–Forsythe–Iversen agar, DFI) which enables results to be obtained 2 days earlier than the conventional method. Ent. sakazakii was isolated from 2/82 powdered IFM, 5/49 dried infant foods, 3/72 milk powder, 2/62 cheese products and various dry food ingredients, especially herbs and spices (40/122). Ent. sakazakii was isolated from 67 samples using the DFI medium, however only 19 of the samples were positive following the conventional method. The largest difference in isolation between the two methods was with dry food ingredients.Although Enterobacteriaceae were enumerated from one powdered IFM sample (Klebsiella ozaenae, 200 cfu/g), 7/82 had detectable Enterobacteriaceae after enrichment in EE broth. Using the ISO 6579 2002 method and immuno-magnetic separation technique no Salmonella serovars were isolated from powdered IFM, dried infant foods or milk samples. Therefore hygienic production of powdered IFM and milk production as monitored by control of Salmonella and enumeration of Enterobacteriaceae did not control Ent. sakazakii.     

新金色分枝杆菌CRISPR基因编辑技术的构建及其初步应用

新金色分枝杆菌Mycobacterium neoaurum JC-12是一株用于转化植物甾醇合成甾体激素类药物中间体的重要菌株,传统的基因编辑方法效率低、周期长、操作复杂并且需带入抗性标签.近期 CRISPR(clustered regularly interspaced short palindromic repea...     

带鱼纯鱼肉重组工艺以及制品蛋白质体外消化的研究

研究了带鱼纯鱼肉的重组工艺。在单因素试验的基础上,在5℃反应温度下,以谷氨酰胺转胺酶用量、作用时间和NaCl用量3个因素为工艺参数,以鱼肉重组制品的凝胶强度为响应值,通过响应面分析对工艺条件进行优化,优化后的工艺条件为:谷氨酰胺转胺酶用量1.25%,作用时间为2.97 h,NaCl用量为1.01%。在优化工艺条件下,鱼肉重组制品的凝胶强度为2 437.41 g.mm,其蛋白质体外消化率为89.14%,稍高于其他实验组。     

红枣、胡萝卜复合饮料制作技术研究

本文以红枣、胡萝卜为主要原料进行可保存6个月以上的清亮型饮料加工技术研究,运用不同澄清方法进行澄清试验,并对饮料 所含固体物质,总糖含量,透光率VC含量进行了测定。     

超临界流体CO2萃取枸杞子红色素的工艺条件研究

本文采用超临界CO2 流体萃取方法 ,从枸杞子中萃取红色素。讨论样品含水量、样品粉碎度、萃取温度、压力、时间和流速对萃取率的影响 ,从而确定最佳萃取工艺条件 ,为批量生产提供依据     

红曲菌Mn-SOD基因克隆、表达及序列分析

为得到产Mn-SOD工程菌及表达产物,通过设计出红曲菌H4000 Mn-SOD简并引物得到目的基因片段,再设计全长引物利用PCR技术扩增红曲霉Mn-SOD基因的全长序列。经EcoR Ⅰ和Hind Ⅲ酶切后连接至相同酶切的表达载体pET28a,并转化至E.coli BL21进行诱导表达。克隆得到的基因预测编码152个氨基酸,预测相对分子量为17 kDa。同时,将克隆得到的Mn-SOD基因与NCBI数据库进行比对,发现该序列与橙色红曲霉超氧化物歧化酶(SOD)基因相似度达到99%,与炭疽菌、米曲霉、黄曲霉的Mn-SOD基因也有较高的相似度。通过SDS-PAGE检测蛋白表达情况,目标蛋白相对分子质量约为19 kDa,与预测分子量基本相符。该表达蛋白在pH2.0保温处理1 h,仍具有较高的Mn-SOD酶活。