干酪乳杆菌LC-15冻干发酵剂制备条件研究

在益生菌发酵乳制品的开发与应用研究中,益生菌发酵剂的制备具有重要的研究价值。对一株具有降低胆固醇作用的益生菌干酪乳杆菌LC-15（Lactobadllus casei LC-15）进行冻干发酵剂制备研究。通过正交优化得到最佳冻干保护剂组合为：脱脂奶粉100g／L,甘油30mL／L,麦芽糊精100g／L,海藻糖150g／L,L-谷氨酸钠10g／L。将此冻干保护剂应用于干酪乳杆菌LC-15冷冻干燥,其存活率达到82．2％,活菌数达4．4×10^10g^-1。在此基础上,对冻干过程中干酪乳杆菌LC-15的预培养条件和预冻条件进行考察,得到的优化结果为：在37℃预培养60min后以-40℃预冻3h,存活率最高可达到84．8％,活菌数达4．9×10^10g^-1,凝乳时间为13h。     

水分对干酪乳杆菌LC-134冻干粉的贮存稳定性的影响

本文研究了水分含量和水分活度(Aw)对干酪乳杆菌LC-134(简称LC-134)冻干粉在贮存过程中的稳定性影响。将不同水分含量和Aw的LC-134冻干粉分别在-18℃和25℃条件下贮存12个月和6个月,定期测定LC-134冻干粉中的活菌数的变化趋势,并且探讨保护剂中的聚葡萄糖含量对冻干粉的水分含量和Aw的影响;研究表明,贮存于-18℃条件下、LC-134冻干粉水分含量≤(5.14±0.06)×10-2 g/g或者Aw≤0.31±0.01时,在贮存的12个月中,活菌数没有明显的下降;然而,贮存于25℃条件下达到活菌数稳定,则要求LC-134冻干粉的水分含量低于(3.06±0.05)×10-2 g/g或者Aw低于0.25±0.01。聚葡萄糖的添加,可降低水分含量相同的LC-134冻干粉的Aw,而且水分含量越高,这种作用越明显。     

干酪乳杆菌LC-15的增菌培养基设计及生长动力学研究

对1株具有降胆固醇作用的干酪乳杆菌(Lactobacillus casei)LC-15的增殖培养基进行了筛选和优化,同时对其在分批培养时的生长动力学进行了研究。所得增菌培养基配方为0.1mol/L pH6.8的Na2HPO4-柠檬酸配制经蛋白酶预水解的10%脱脂乳冻干粉,再添加0.5%的葡萄糖,1.5%的胰蛋白胨,0.3%酵母浸膏,在此培养基中的最大菌数可达8.8×109cfu/mL,分别比MRS及脱脂复原乳中提高了1.35倍和0.75倍。得到的干酪乳杆菌在优化培养基中分批培养的生长动力学模型为:N(t)=3.4088/(1+e3.3560-0.3254t)。     

干酪乳杆菌胞外多糖LCP1的流变性研究

通过对干酪乳杆菌LC2W胞外多糖LCP1溶液流变性质研究发现,高浓度LCP1溶液呈剪切变稀,低浓度时,剪切速率对黏度没有影响,溶液黏度随温度升高下降明显,pH对LCP1溶液黏度影响很小,盐可以降低LCP1溶液黏度,但黏度降低与盐浓度有关,通过黏弹性研究发现LCP1水溶液不能形成胶体,乌氏黏度计测得LCP1水溶液固有黏度[η]为1.930 dL/g。     

两株干酪乳杆菌的安全性评价

干酪乳杆菌X-29和X-15是两株分离于黄酒浸米浆水中的乳酸菌,这两株菌均具有高的水解淀粉活性。本实验主要是评价这两株菌的安全性,为其应用于发酵米制品提供安全性保障。通过急性毒性实验、耐药性实验、有害代谢产物和溶血实验,确定这两株菌株是否具备安全性。结果表明该两株菌株不会引起急性毒性和易位现象;不产生有害代谢产物;对大部分常见抗生素没有耐药性;不会产生溶血现象。通过对这两株菌的安全性评价可知干酪乳杆菌X-29和X-15是安全的。     

益生乳酸菌Lactobacillus casei Zhang从基础研究到产业化开发

2001年-2009年,内蒙古农业大学乳品生物技术与工程教育部重点实验室从采集自中国西部7个省市及蒙古国14个省市不同少数民族地区951份自然发酵乳制品等样品中分离鉴定出3 388株乳酸菌,建立了中国首个原创性乳酸菌菌种资源库;从菌种库中分离自传统发酵酸马奶(koumiss)的243株乳杆菌中筛选获得1株性能优良的乳酸菌—Lactobacillus casei Zhang(L.casei Zhang)。采用5种动物模型和人体试验进行了功能评价,并利用基因组学和蛋白质组学研究技术对L.casei Zhang益生分子机制进行了深入剖析。经过多年系统的基础研究,自主解决了其产业化的关键技术问题,包括其直投式发酵剂、发酵乳制品和发酵豆乳益生菌饮料的研发并实现了产业化。     

Viability of probiotic (Bifidobacterium, Lactobacillus acidophilus and Lactobacillus casei) and nonprobiotic microflora in Argentinian Fresco cheese

We evaluated the suitability of Argentinian Fresco cheese as a food carrier of probiotic cultures. We used cultures of Bifidobacterium bifidum (two strains), Bifidobacterium longum (two strains), Bifidobacterium sp. (one strain), Lactobacillus acidophilus (two strains), and Lactobacillus casei (two strains) in different combinations, as probiotic adjuncts. Probiotic, lactic starter (Lactococcus lactis and Streptococcus thermophilus), and contaminant (coliforms, yeasts, and molds) organisms were counted at 0, 30, and 60 d of refrigerated storage. Furthermore, the acid resistance of probiotic and starter bacteria was determined from hydrochloric solutions (pH 2 and 3) of Fresco cheese. The results showed that nine different combinations of bifidobacteria and L. acidophilus had a satisfactory viability (count decreases in 60 d <1 log order) in the cheese. Both combinations of bifidobacteria and L. casei cultures assayed also showed a satisfactory survival (counts decreased <1 log order for bifidobacteria but no decrease was detected for L. casei). On the other hand, the three combinations of bifidobacteria, L. acidophilus, and L. casei tested adapted well to the Fresco cheese environment. When a cheese homogenate at pH 3 was used to partially simulate the acidic conditions in the stomach, the probiotic cultures had an excellent ability to remain viable up to 3 h. At pH 2, the cell viability was more affected; B. bifidum was the most resistant organism. This study showed that the Argentinian Fresco cheese could be used as an adequate carrier of probiotic bacteria.     

The physicochemical surface characteristics of Lactobacillus casei

The surface and interfacial properties of five strains of Lactobacillus casei, commonly found in cheese, were investigated in an attempt to identify surface properties that might be exploited to effect adsorption of the cells onto milk fat globules. All measures of surface hydrophobicity revealed these strains to be moderately hydrophobic compared to a reference strain of Mycobacterium smegmatis which was very hydrophobic. In chromatography experiments, L. casei had no affinity for either hydrophobic (octyl-sepharose) or control (sepharose) columns in conditions of physiological pH and low ionic strength, despite a favorable free energy of interfacial interaction (ΔG^IF_IW2) as derived from contact angle measurements. Adhesion of L. casei strains to these materials was effected by manipulation of eluent pH and ionic strength. In contrast, adhesion of L. casei strains to n-hexadecane concurred with ΔG^IF_IW2. Surfaces of L. casei strains subjected to X-ray photoelectron spectroscopy exhibited similar amounts of carbon and oxygen but displayed considerable diversity in surface nitrogen and phosphorous contents. Carbon existed predominantly as C-(O, N) and oxygen as -OH on L. casei surfaces but carbon on M. smegmatis surfaces was present predominantly as C-(C,H).     

Survival of Lactobacillus casei and functional characteristics of reduced sugar red beetroot yoghurt with natural sugar substitutes

In this study, probiotic gel fermentation and functional characteristics of reduced sugar yoghurt with red beetroot bioactive components and steviol glycosides from Stevia rebaudiana extracts as natural source of noncaloric sweeteners were investigated. A significant increase in Lactobacillus casei viability and fermented gel network and a decrease in syneresis were determined in yoghurt sweetened with steviol glycosides and high content of red beetroot bioactive components such as total phenolics, anthocyanins and total antioxidant capacity (DPPH and FRAP). The survival of L. casei was maintained within probiotic and therapeutic levels (>9 log₁₀ CFU/g), due to the possible prebiotic effect of steviol glycosides and fermentable fibres and phenolic compounds of red beetroot during 28 days of cold storage of the probiotic yoghurts. Colour properties (L*, a*, b*, ΔE*, C*, h*) and such sensory properties as smoothness, sweetness astringency were significantly affected. The highest organic acid content was found in red beetroot enriched yoghurt with stevia, while the most dominant organic acids were butyric, lactic, malic and citric. As a result, it was determined that fermented milk products, which are supported by phytochemical and steviol glycoside interactions, have adequate nutritional effects, high probiotic viability and acceptable sensory properties     

Survival of Lactobacillus casei,Lactobacillus plantarum and Bifidobacterium bifidum in free and microencapsulated forms on Iranian white cheese produced by ultrafiltration

Survival of probiotic strains Lactobacillus casei ( ATCC 39392 ), Lactobacillus plantarum ( ATCC 8014 ) and Bifidobacterium bifidum ( ATCC 29521 ) was investigated either in microencapsulated or in free form in the Iranian white cheese produced by ultrafiltration technique. The results indicated that the survival of encapsulated probiotic bacteria was higher than free cells. Both free and microencapsulated forms were successful in keeping counts of L. casei, L. plantarum and B. bifidum in the cheese high enough for the therapeutic minimum (10⁶–10⁷ cfu/g) after 60 days. Addition of probiotic adjunct also did not alter the chemical composition, but pH was lower in probiotic cheeses.     

Probiotic effects of feeding heat-killed Lactobacillus acidophilus and Lactobacillus casei to Candida albicans-colonized immunodeficient mice

Probiotic bacteria can protect immunodeficient mice from orogastric candidiasis but cause some pathology of their own. Severely immunodeficient patients may be at risk if fed viable probiotics, so this study evaluated the probiotic potential of nonviable probiotic bacteria to protect immunodeficient mice from Candida albicans infections. Heat-killed probiotic bacteria were fed to gnotobiotic bg/bg-nu/nu and bg/bg-nu/+ mice to ascertain if they could protect the mice from mucosal and systemic candidiasis. Both heat-killed Lactobacillus acidophilus (HKLA) and heat-killed Lactobacillus casei (HKLC), in comparison to control mice not fed the probiotic bacteria but challenged (oral) with C. albicans, suppressed the severity of orogastric candidiasis in bg/bg-nu/nu mice at 2 weeks after colonization with C. albicans, inhibited disseminated candidiasis in C. albicans-colonized bg/bg-nu/+ mice at 4 weeks after colonization, and suppressed the number of viable C. albicans in the alimentary tract. HKLA, but not HKLC, treatment inhibited disseminated candidiasis in bg/bg-nu/nu mice at 2 weeks after oral challenge and enhanced the proliferative responses of splenocytes from C. albicans-colonized bg/bg-nu/+ mice to C. albicans antigens. Neither HKLA nor HKLC were able to prolong the survival of gnotobiotic bg/bg-nu/nu mice after oral challenge with C. albicans. These results demonstrate that heat-killed lactobacilli can induce some (limited) protection (probiotic effect) against candidiasis in mice.     

副干酪乳杆菌海藻酸钠微胶囊包埋工艺

研究副干酪乳杆菌海藻酸钠微胶囊的包埋效果,副干酪乳杆菌在冻干、贮藏及模拟胃胀道环境中的存活情况,并优化包埋工艺参数。结果表明,大豆蛋白分离物(SPI)是适宜的内层壁材,低聚异麦芽糖对副干酪乳杆菌的冷冻保护效果最佳。当SPI用量为3%,低聚异麦芽糖添加量5%,海藻酸钠浓度2%、氯化钙浓度为0.2 mo L/L,副干酪乳杆菌的包埋率可达93.31%,冷冻干燥后微胶囊4℃储存28 d的副干酪乳杆菌存活率达58.97%,微胶囊副干酪乳杆菌在模拟胃液中3 h存活率达67.52%,模拟肠液中45 min基本得到释放。由于挤压法制备的副干酪乳杆菌微胶囊较高的存活率,操作简便、经济,因此有较为广阔的工业化前景。     

干酪乳杆菌细菌素的抗菌机制分析

目的：分析和预测干酪乳杆菌细菌素基因及其编码蛋白的分子结构和理化性质,明确细菌素的抗菌机制。方法：利用生物信息学软件进行细菌素编码蛋白的结构预测和功能分析,采用氨基酸定点突变技术对预测的氨基酸位点进行点突变,检测突变体的抗菌活性。结果：干酪乳杆菌细菌素（LacA和LacB）属于热稳定、分泌信号肽的跨膜蛋白,LacA蛋白存在典型的抗菌结构域GxxxG,构建LacA蛋白中第40位V和第57位I的突变载体,两个位点中任何一个发生突变都明显影响细菌素的抗菌活性。结论：推测V40和I57是干酪乳杆菌细菌素发挥抑菌活性的关键氨基酸位点,为今后探索干酪乳杆菌细菌素（class IIb）抗菌机制提供理论依据。     

干酪乳杆菌对农家干酪抗氧化活性及在其模拟胃肠道中存活能力的研究

本研究利用向农家干酪加入干酪乳杆菌的方式,研究农家干酪的抗氧化活性及在模拟胃肠道中乳酸菌的存活能力。通过对DPPH自由基、羟自由基的清除的能力以及对铁的还原能力指标的测定,利用PH 4.6-SN及12%TCA-SN进行抗氧化活性分析。结果表明:在第30 d时,DPPH自由基的清除能力达到最高值,为316μM Trolox,还原能力为0.647;羟自由基的清除率在第25 d时最高达到54.38%,在第30 d时有所下降。在对第20 d的农家干酪的水溶性提取液的冷冻干燥粉进行抗氧化活性的评价时,抗氧化活性得到提高;综上所述,添加干酪乳杆菌的农家干酪在储藏期内抗氧化活性显著增强。选择第20 d的农家干酪进行模拟胃肠道的存活能力研究,发现活菌数降低了5 log_(10) cfu/g,最终为7 log_(10) cfu/g,可起到益生作用。因此,加入干酪乳杆菌显著提高农家干酪抗氧化活性,农家干酪可以起到益生作用。     

Factors affecting the caseinolytic activity of Lactobacillus casei and Lactobacillus plantarum

Whole cell suspensions of some strains of each Lactobacillus casei and Lactobacillus plantarum were assayed for their caseinolytic activity in 0.1 M NaH₂PO₄ buffer, pH 6.5, at 30 °C, using different assay methods. Azocasein was not as sensitive as casein (Hammarsten) as a substrate. Inclusion of glucose in the assay mixture reduced the released α-amino groups as evidenced by fluorescent labelling, but generally increased the amounts of excreted amino acids. Divalent cations, including calcium ions, played only a minor role in the activation of the cell-bound proteinase, whereas NaCl inhibited it markedly. Inhibitor studies suggest that the enzyme is a serine proteinase. The different assay methods used did not give identical results. Fluorescent labelling of the free α-amino groups at pH 6.0 appears, on the contrary, to be a more reliable method.     

干酪乳杆菌纯种发酵大豆乳培养基的优化调配

从益生菌乳制品中自行分离选育出的生长繁殖力强、发酵活力高的干酪乳杆菌(05-20)为试验菌株,研究了牛乳和蔗糖在大豆乳中的添加量对干酪乳杆菌在大豆乳中发酵的凝乳时间、凝乳时活菌数、pH值、滴定酸度及产品感官风味的影响,通过方差分析确定了发酵培养基中牛乳和蔗糖的最适添加量分别为20%和7%,完成了以大豆乳作为干酪乳杆菌最佳载体的初步探索,为进一步研制开发益生菌发酵大豆乳制品奠定了基础.     

氧气对干酪乳杆菌生长和活力的影响

以静置培养为对照,通过振荡培养制造有氧环境,对干酪乳杆菌的生长曲线、菌体活力进行分析。研究发现在有氧条件下菌体生长受到抑制,菌体密度小于静置发酵,在4℃保存时的活力迅速下降,另外,有氧生长菌体对热、胆汁等条件的耐受性较差,在50℃处理4h导致菌体迅速死亡,在含有胆汁的培养基中,菌体生长几乎停滞。通过透射电镜观察发现有氧条件下菌体细胞壁发生破裂,菌体细胞质成分溶出,导致菌体死亡。因此,氧气对干酪乳杆菌的生长具有抑制作用,造成细胞损伤,导致菌体活力降低。