模拟人体胃肠道环境筛选益生乳杆菌

【目的】筛选具有益生特性的乳杆菌作为保健型酸奶的候选菌株。【方法】从健康人肠道和奶豆腐中分离筛选出耐受人工胃液的乳杆菌,对其进行体外益生特性(人工胃肠液耐受性、胆盐耐受性、抑菌活性及胆固醇降解能力)研究。【结果】从在乳杆菌分离培养基上有溶钙圈的41株菌株中筛选出5株耐酸、耐人工胃液较强的菌株,经16S rR NA基因测序鉴定,其中3株为乳杆菌,分别命名为植物乳杆菌Lp MT-3、植物乳杆菌Lp MT-5和唾液乳杆菌LsA F-7。在人工胃液中3株菌的耐受力均强于商品化的对照菌株LGG(鼠李糖乳杆菌GG);转入肠液4 h后直至26 h,Lp MT-5存活率基本稳定在45%左右,仅次于LGG。胆盐浓度为0.10%时,3株乳杆菌的耐胆盐能力均强于LGG;胆盐浓度为0.20%时,Lp MT-3和LsA F-7仍能存活。3株乳杆菌均具有抑菌活性,对粪肠球菌的抑制最明显,其次是金黄色葡萄球菌,对大肠杆菌、沙门氏菌的抑制作用较差。3株乳杆菌对胆固醇的清除效力依次为Lp MT-3LpM T-5Ls AF-7;清除率依次为Ls AF-7Lp MT-3LpM T-5。【结论】筛选出3株适应人体胃肠液环境、耐胆盐、抑菌及降胆固醇活力强的乳杆菌,可作为进一步开发新的益生菌产品和保健型酸奶的菌株。     

一株具降胆固醇功能乳酸菌的机制研究

目的对动物双歧杆菌SQ-0001的体外环境耐受性及其降胆固醇作用进行研究。方法用邻苯二甲醛法对pH 3.0和0.3%牛胆酸钠有一定耐受性的动物双歧杆菌SQ-0001的体外降胆固醇机制进行了研究。结果动物双歧杆菌SQ-0001对pH 3.0及0.3%胆盐环境具有一定的耐受性;该菌在胆盐浓度为0.2%,培养时间为24 h时,胆固醇脱除率最高,达到91.59%;该菌降胆固醇的作用主要以吸收为主,吸收率达到63.87%,其次是降解率,为23.81%。结论动物双歧杆菌SQ-0001的环境耐受能力良好,具有很好体外降胆固醇作用,该菌株可以作为日后降胆固醇的潜在益生菌。     

产淀粉酶益生乳杆菌的筛选及鉴定

目的以健康仔猪肠道及粪便样品为基础,从中筛选产淀粉酶的乳杆菌菌株,并评价其作为益生菌候选菌株潜力。方法用MRS培养基分别从仔猪新鲜粪便和小肠黏膜上分离到乳酸菌,采用改良的产淀粉酶选择性培养基初筛得到能降解淀粉活性的菌株,并研究菌株的淀粉水解活性、抗逆能力、粘附特性及对抗生素的敏感性。结果从备选的485株乳酸菌中筛选得到具有初步淀粉酶活性的菌株25株（占总筛选数量的5.2%）,复筛选育得到具有较强淀粉酶活性的乳杆菌3株。进一步研究了这3株乳杆菌的抗逆能力、粘附特性以及对抗生素的耐药性,并对最终选育得到的菌株进行生理生化及16S rRNA分子鉴定。经选育鉴定的罗伊乳杆菌G8-5淀粉降解能力最强,并能耐受pH 3.0的酸度、1.0%的胆盐浓度,在小肠上皮细胞上的粘附效率超过15个以上,并对常用的抗生素具有较高的敏感率。结论罗伊乳杆菌G8-5符合安全益生菌的要求,可以作为产淀粉酶的益生乳杆菌优良的候选菌株。     

具有血管紧张素转化酶抑制活性益生菌筛选的研究

经体外血管紧张素转换酶（ACE）抑制活性实验,从保存的40株菌中筛选到7株具有降压活性的益生菌,其中菌株DM8319粗提样品经初步凝胶层析纯化后其IC50值从0．038mg／ml降到0．016mg／ml,达到了初步分离效果。后依据益生菌筛选标准,对上述菌株抗氧化性、耐酸、耐胆盐、抑菌和耐药性能进行考察。结果显示,菌株DM84013最适用于益生菌降压产品的研究,经初步鉴定该菌种为长双歧杆菌。     

益生菌降胆固醇功能研究进展

本文对益生菌在动物及人体上的降胆固醇功能及可能机理进行了综述。益生菌是一类能够对人体健康起到促进作用的活体微生物。现已发现某些乳杆菌、双歧杆菌和肠球菌属的一些菌株具有降低血清胆固醇水平的能力。由于实验动物和人体在生理学上存在差异,因此同一菌株在实验动物和人体上获得的结论可能不同。关于益生菌降胆固醇功能的机理,研究者提出了不同的假说。这些假说包括:(1)益生菌将胆固醇吸收至细胞膜或细胞质中;(2)益生菌将胆固醇吸附到细胞表面;(3)胆固醇和游离胆盐在酸性环境下发生共沉淀;(4)结合胆盐被益生菌的胆盐水解酶水解成了游离胆盐,后者具有较低的溶解度,不易被肠道回收;(5)胆酸被益生菌的荚膜胞外多糖黏附到了细胞表面;(6)益生菌发酵肠道食源性未消化的碳水化合物产生丙酸,后者能够抑制肝脏胆固醇的生物合成,从而导致血清胆固醇水平降低;(7)益生菌通过下调NPC1L1蛋白基因表达来降低小肠细胞对胆固醇的吸收;(8)益生菌抑制胆固醇乳化胶束的形成。这些假说将为我们认识益生菌的降胆固醇机制及潜在降胆固醇功能菌株的筛选提供了依据。     

解脂亚罗酵母菌体外降甘油三酯降胆固醇效果研究

筛选出高效降三酰甘油降胆固醇菌株,为新型药物、食品的制备提供优良菌株。从样品中筛选分离出酵母菌株,并测定菌株降三酰甘油能力,采用磷硫铁比色法测定菌株降胆固醇能力,通过耐酸、耐胆盐及毒性实验研究菌株各项性能,通过生理生化及26S rDNA法对菌株进行鉴定。结果显示,试验筛选出1株高效降脂降胆固醇酵母菌EAM-ZT003T,降三酰甘油能力高达80.2%,菌株具有很好的耐酸、耐胆盐能力,为期42 d的小鼠毒性试验发现,小鼠未出现异常体征,未发现异常死亡,且解剖观察各器官一切正常,菌株经26S rDNA鉴定为解脂亚罗酵母菌(Yarrowia lipolytica)。     

具有降胆固醇功能益生菌的筛选研究

目的通过体外实验从发酵制品及人体肠道内分离筛选出安全高效的降胆固醇乳酸菌,并研究其生理生化特性,为今后功能性乳制品的开发提供优良菌种。方法应用MRS．胆固醇培养基液体发酵法进行目的菌株筛选,并测定其耐酸及耐胆盐等能力。结果筛选出4株具有降胆固醇能力的乳酸菌,其降解率分别为DM8403163．1％、DM8403470．3％、DM850381．1％、DM8605663．1％;同时4株乳酸菌均表现出不同程度的耐酸及耐胆盐能力,其中DM86056能力最强,其他3株乳酸菌次之;经鉴定,DM84031为保加利亚乳杆菌、DM84034为嗜热链球菌、DM8503为詹氏乳杆菌、DN86056为屎肠球菌。结论经上述实验综合比较,DM86056降胆固醇能力比较强且耐酸及耐胆盐等能力也较强,因此有待于将其应用于动物体内进行深入研究。     

两株乳酸杆菌益生菌特性的研究

对两株乳酸杆菌作为益生菌的特性进行分析。采用人工胃液、人工肠液,及体外培养人结肠癌细胞株HT-29,模拟 上消化道环境,检测候选菌株L.acidophilus1.1878和L.rhamnosus1.120对上消化道环境的耐受性和粘附性;以琼脂扩散打孔 法,检测候选菌株SCS对4种常见肠道致病菌的抑菌能力。结果表明,两株候选菌株可耐受pH>3.0的人工胃液,及0.2%的 胆盐浓度;光镜观察L.acidophilus1.1878、L.rhamnosus1.120粘附于HT-29细胞边缘,平均4.5～7个/细胞,扫描电镜观察 L.acidophilus1.1878、L.rhamnosus1.120粘附于HT-29细胞表面的刷状缘,且细胞表面结构完整;对4种肠道致病菌的抑菌 圈直径均在12mm以上。证明实验观察的两株乳酸杆菌具有较好的耐酸、耐胆盐及粘附、抑菌特性,符合益生菌的标准。     

金针菇源降胆固醇土著乳酸菌的筛选与鉴定

以金针菇子实体为材料,通过添加溴甲酚紫的MRS培养基分离得到98株土著乳酸菌。以降胆固醇的效率和模拟肠道环境的耐酸和耐胆盐的耐受力为指标进行复筛,获得了2株降胆固醇以及耐酸耐胆盐综合能力都较强的菌株4J1和4J17,经16S r DNA鉴定及生理生化试验鉴定,结果表明该两株乳酸菌均为假肠膜明串珠菌(Leuconostoc pseudomesenteroides)。本研究结果扩宽了降胆固醇的益生乳酸菌的种质资源,同时为金针菇的发酵加工产品提供良好的益生发酵菌株。     

发酵柚子皮戊糖片球菌的分离及生物学特性分析

目的从发酵柚子皮中分离具有优良性能的戊糖片球菌。方法对5株戊糖片球菌分离株(WPPE01、WPPE02、WPPE03、WPPE04和WPPE05)以及标准株CGMCC 1.2695的生物学特性和安全性进行系统分析。结果 WPPE02和WPPE04的耐酸性优于其他菌株,WPPE01和WPPE05的耐胆盐能力优于其他菌株,WPPE01和WPPE03同时具有较强的自聚及粘附能力,WPPE02具有最强的盐耐受能力,WPPE03的耐热性能最强。与其他受试细菌相比,5株戊糖片球菌对单增李斯特菌具有最强的抑制效果,WPPE01和WPPE03对大肠埃希菌的抑制效果显著优于标准株。分离株主要通过置换的方式抑制大肠埃希菌O157:H7对Caco-2细胞的粘附且WPPE02和WPPE03的抑制效果最强。全部戊糖片球菌均对部分抗生素敏感且不具有溶血性。结论 5株从发酵柚子皮中分离的戊糖片球菌具有作为益生菌应用的潜能。     

3株罗伊氏乳杆菌生物学特性的分析比较

【目的】对3株罗伊氏乳杆菌的生物学特性进行分析比较,为后期生产应用提供一定的参考。【方法】对实验室保藏的3株罗伊氏乳杆菌的生长曲线、pH曲线、耐受人工胃液能力、耐受猪胆盐能力、黏附能力、抑菌能力和对抗生素的耐药性等特性进行了分析比较。【结果】3株菌生长趋势大致相同;3株菌对人工胃液均具有良好的耐受性,且可以有效地抑制大肠杆菌和金黄色葡萄球菌的生长;菌株L0和L2对高胆盐的环境耐受性较差,菌株L1则对高胆盐环境具有极强的耐受性;菌株L1和L2具有很强的黏附能力;3株菌对20种抗生素表现出不同的耐受性。【结论】菌株L1的生物学特性明显优于其他两株菌株,有利于后期的生产应用。     

The effect of bile salts on survival and morphology of a potential probiotic strain Lactobacillus acidophilus M92

Bile tolerance is an important criterion in the selection of microbial strains for probiotic use. The survival and morphological changes of a potential probiotic strain, Lactobacillus acidophilus M92, in the presence of bile salts were examined. Lactobacillus acidophilus M92 has shown a satisfactory degree of tolerance against oxgall and individual bile salts tested, especially to taurocholate. The higher resistance of L. acidophilus M92 against taurine-conjugated bile salts relative to deconjugated and glycine-conjugated bile salts was attributed to its reaction to the stronger acidity of the former. Furthermore, bile salt hydrolase (BSH) was active when L. acidophilus M92 was grown in the presence of sodium taurocholate. The rate of BSH activity was highest at the exponential growth phase. It was hypothesised that BSH activity may be important for the bile salt resistance of this strain. The colonial and cellular morphology may also be a valuable parameter in the selection of bile salt-resistant Lactobacillus strains for probiotic use. Smooth (S) and rough (R) colonies, appeared in the original L. acidophilus M92 bacterial culture and demonstrated a different degree of bile tolerance. Rough colonies were more sensitive to bile salts than smooth ones. The R colony cells assumed a round form, probably induced by gaps in the cell wall caused by the cytotoxicity of glycodeoxycholate. This revised version was published online in July 2006 with corrections to the Cover Date.     

益生菌降解胆固醇的作用及机理研究进展

益生菌是来源于宿主并对宿主健康有一定促进作用的微生物。降胆固醇功能是某些益生菌的主要益生功能之一。近年来国际上对益生菌降胆固醇的体外和体内研究进展,主要包括菌体生长与降胆固醇的关系、pH的影响、胆酸盐的作用、胆酸盐水解酶活性、益生元的使用、人和动物喂养实验等方面。目前人们提出的益生菌降胆固醇作用机理的假说主要有共沉淀作用、酶对胆酸盐的分解作用、胆固醇掺入细胞膜、细菌对胆固醇的吸收等,这些机理假说尚有待进一步研究证实。降胆固醇益生菌制品研发的前景十分广阔。     

Screening of lactic acid bacteria with cholesterol-lowering and triglyceride-lowering activity in vitro and evaluation of probiotic function

To screen the lactic acid bacteria with cholesterol-lowering and triglyceride-lowering activity in vitro and evaluate their probiotic function. By plate separating, cholesterol-lowering and triglyceride-lowering activity in vitro were determined; and by evaluating the probiotic functions, including tolerances to simulated gastric and intestinal juice, the antibacterial spectrum, and the adhesion ability to Caco-2 cells, the probiotic strains with cholesterol-lowering and triglyceride-lowering activity in vitro were screened, and then were identified by phenotypical and physiological tests and 16Sr DNA. Finally, the cholesterol-lowering and triglyceride-lowering activity in vivo of the strains were evaluated using male Sprague-Dawley rats. Two strains L2-16 and L2-73 with stronger cholesterol-lowering and triglyceride-lowering activity in vitro, stronger tolerance to simulated gastric and intestinal juice and adhesion ability to Caco-2 cells, and wider antibacterial spectrum were screened from traditional Chinese fermented cucumber and were identified as Lactobacillus acidophilus and Enterococcus faecalis, respectively. Compared with a hyperlipidemia diet without lactic acid bacteria, the diet supplemented with Lactobacillus acidophilus L2-16 and Enterococcus faecalis L2-73 significantly reduced serum total cholesterol, triglycerides, and low-density lipoprotein cholesterol levels, and liver total cholesterol and triglyceride levels in rats (P?<?0.05). Moreover, the diet supplemented with Lactobacillus acidophilus L2-16 and Enterococcus faecalis L2-73 significantly increased the fecal elimination of bile acids (P?<?0.05). Lactobacillus acidophilus L2-16 and Enterococcus faecalis L2-73 may have application prospect in the production of some fermented foods such as fermented vegetables, milk, or meat, and probiotic preparations with the function to lower the serum lipid and liver lipid levels.     

Bile Salt Hydrolase (Bsh) Activity Screening of Lactobacilli: In Vitro Selection of Indigenous Lactobacillus Strains with Potential Bile Salt Hydrolysing and Cholesterol-Lowering Ability

The bile salt hydrolase (Bsh) activity of probiotic bacterium residing in gastrointestinal tract has often being associated with its cholesterol-lowering effects. Hence, Bsh activity was explored in this study as the criterion for the selection of most potential Bsh-active and cholesterol-lowering indigenous Lactobacillus strains. Forty lactobacilli were adjudged Bsh active after a preliminary screening of 102 lactobacilli and occurrence of Bsh activity correlated well with their natural habitats. Of the 40 shortlisted lactobacilli, fifteen putative Lactobacillus strains were selected and further tested for their comparative Bsh activity. In the end, indigenous Lactobacillus plantarum strains Lp91 and Lp21 were emerged as the promising Bsh-active lactobacilli with their substrate preference inclined more towards glycocholate than other bile acid amino conjugates. In addition, strains Lp91 and Lp21 also exhibited significantly high bile salt deconjugation, cholesterol assimilation and cholesterol co-precipitation ability in vitro. In conclusion, indigenous L. plantarum strains Lp91 and Lp21 may be the promising candidate probiotics to elucidate the ecological significance of probiotic Bsh activity in vivo.     

商品鸡盲肠内容物乳酸菌的分离与鉴定

实验以健康商品鸡盲肠为菌源,对附着其上的乳酸菌进行分离与鉴定,就所获得的菌株的耐酸耐胆汁、黏附特性、抑菌性进行初步研究。实验结果表明,从商品鸡盲肠中分离出6株乳酸菌,根据耐酸耐胆汁试验,病原菌生长抑制试验以及肠道黏膜粘附试验,筛选出2株优势菌株CX001和CX005,经乳杆菌属的生化鉴定,初步确定为短乳杆菌。     

乳杆菌耐胆汁、降解结合胆盐和同化胆固醇能力的研究

对8株植物乳杆菌的胆汁耐受力、降解结合胆盐能力以及同化胆固醇能力进行了研究。不同的菌株在添加了牛胆汁的MRS中生长速度具有明显差异,同化胆固醇能力也明显不同,而降解结合胆盐的能力没有明显区别。分析发现,菌株的胆汁耐受力和降解结合胆盐能力,胆汁耐受力和同化胆固醇能力,以及降解结合胆盐能力和同化胆固醇能力之间都没有明显的相关性。