西部传统发酵乳品中乳酸菌筛分及其亚硝酸盐降解能力

为探究西部牧区传统发酵乳品中乳酸菌多样性及其优势菌种的亚硝酸盐降解能力,为工业化利用提供参考,应用16S rDNA技术鉴定分离菌株,并对分离菌株亚硝酸盐降解能力采用比色法进行分析,比较不同地区、不同发酵乳品中不同种类乳酸菌亚硝酸盐降解能力的差异。结果表明：1）104?份样品中共分离得到275?株乳酸菌,鉴定出6?个属23?个种,其中瑞士乳杆菌（Lactobacillus helveticus）为青海、甘肃、新疆、内蒙古、西藏传统发酵乳品中共有菌株,L. helveticus和植物乳杆菌（L. plantarum）为酸牦牛奶、酸马奶、酸驼奶、奶渣发酵乳品中共有菌株。2）275?株乳酸菌亚硝酸盐降解率在4.8%～99.9%之间波动,其中50%的菌株表现出较好的亚硝酸盐降解能力,平均在91.3%以上。3）降解能力因菌株来源和乳酸菌种类不同存在差异性,来源于西藏的菌株显著高于青海、甘肃、新疆的菌株（P＜0.05）;来源于奶渣中的菌株显著高于酸牦牛奶和酸马奶中的菌株（P＜0.05）;在分离出的8?种优势菌株中L. plantarum、副干酪乳杆菌（L. paracasei）、短乳杆菌（L. breris）显示了稳定高效的亚硝酸盐降解能力,其中L. plantarum降解能力最强,且部分菌种间降解能力存在显著性差异（P＜0.05）。     

Isolation and identification of microorganisms in Kazakhstan koumiss and their application in preparing cow-milk koumiss

Koumiss is a type of famous fermented mare milk and considered an important nutritious beverage in central Asian countries. However, the production of koumiss cannot meet public demand in the market due to availability of mare milk. In the present study, 52 lactic acid bacteria and 20 yeast strains from traditional homemade Kazakhstan koumiss were isolated and identified. The isolates were used in a trial that included fermented cow milk, and the flavor profiles, color, and taste to determine their contribution in the co-fermentation of cow milk. Based on the sensory evaluation, KZLAB13 and KZY10 strains were selected as the best cofermentation combinations. The optimal fermentation conditions were confirmed as the ratio of the starter culture 2.4:1.6 % (vol/vol) KZLAB13 strain to KZY10 strain and a temperature of 36°C for 16 h using response surface methodology. After evaluating the quality of the optimized cow-milk koumiss compared with the Kazakhstan koumiss, results suggested that cow milk fermented by these 2 strains possessed a promising taste, flavor, and physicochemical and rheological properties. Altogether, our results showed that cow milk fermented with a combination of KZLAB13 and KZY10 strains can simulate the taste, flavor, and quality of traditional koumiss. Our study provided a novel alternative to mare-milk koumiss and could be used in dairy programs to fulfill the needs of people.     

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。     

传统乳制品中乳酸菌的筛选及发酵性能研究

为获得优良发酵性能的乳酸菌,采用传统培养方法从传统发酵乳样品中分离乳酸菌,并将分离菌株制备发酵乳。通过考察发酵乳的感官品质筛选优良菌株,采用形态学观察及16S rDNA序列分析对筛选菌株进行鉴定;并对其生长性能、产酸性能及其制备发酵乳贮藏期内品质变化进行测定。结果显示,共分离得到67株乳酸菌,筛选出5株性能较佳的菌株（编号为C-1、Q164、Q165、Q166和Q167）,均被鉴定为嗜热链球菌（Streptococcus thermophilus）。其中,菌株C-1生长性能和产酸性能最佳,该菌株制备的发酵乳奶香明显,口感细腻顺滑,感官评分最高（93.5分）,30 ℃贮藏21 d后,酸度为86.0 °T,活菌数为7.2×103 CFU/g。综上,嗜热链球菌C-1具备优良发酵性能菌株的应用潜力。     

甘肃牧区传统发酵乳制品中优良乳酸菌的分离筛选

从甘肃牧区传统发酵乳制品中分离筛选适合发酵乳生产的乳酸菌,对分离纯化的乳酸菌进行了发酵性能检测,筛选出产酸快、发酵活力高、凝乳时间短、后酸化能力弱、遗传性状稳定的菌株。筛选的6株菌株单菌发酵牛乳,均在6.5 h内凝乳,发酵乳酸度＞70 °T,乳酸菌活菌数＞1×108 CFU/mL,发酵乳组织状态、滋味、气味等感官指标良好,可作为生产发酵乳的优良乳酸菌菌种进行开发利用。     

酸马乳发酵过程中乳酸菌与酵母菌生长的相互影响

研究新疆酸马乳中乳酸乳球菌WLB5、干酪乳杆菌MLS5与马克思克鲁维酵母菌WWMJ1间的相互作用。结果表明：在酸马乳发酵过程中,马克思克鲁维酵母菌WWMJ1可以促进干酪乳杆菌MLS5的生长,干酪乳杆菌MLS5对马克思克鲁维酵母菌WWMJ1的生长有抑制作用,乳酸乳球菌WLB5能促进马克思克鲁维酵母菌WWMJ1的生长。乳酸乳球菌WLB5和干酪乳杆菌MLS5混合发酵有助于提高酸马乳中乳酸菌总活菌数。本研究可为酵母菌在发酵乳制品中的应用及开发新型乳制品提供一定参考。     

Identification of the bacterial biodiversity in koumiss by denaturing gradient gel electrophoresis and species-specific polymerase chain reaction

Bacterial biodiversity in traditional koumiss fermented milk was studied by denaturing gradient gel electrophoresis (DGGE). Target DNA bands were identified according to the reference species ladder, constructed in this study. Comigrating bands present in the DGGE profiles were resolved by species-specific PCR. The results revealed a novel bacterial profile and extensive bacterial biodiversity in koumiss. The dominant lactic acid bacteria included Lactobacillus acidophilus, Lactobacillus helveticus, Lactobacillus fermentum, and Lactobacillus kefiranofaciens. Frequently encountered bacterial species were Enterococcus faecalis, Lactococcus lactis, Lactobacillus paracasei, Lactobacillus kitasatonis, and Lactobacillus kefiri. Leuconostoc mesenteroides, Streptococcus thermophilus, Lactobacillus buchneri, and Lactobacillus jensenii were occasionally found in this product. In addition, L. buchneri, L. jensenii, and L. kitasatonis, which were never previously isolated by culture-dependent methods, were identified for the first time in the Xinjiang koumiss. Furthermore, conventional cultivation was performed by plating samples on M17, de Man-Rogosa-Sharpe, Halligan-Pearce, and Kenner fecal media. The results revealed that lactobacilli were the dominant species in the koumiss ecosystem, which was consistent with the results obtained by the DGGE analysis. This is the first systematic study of the microbial composition in koumiss, and our findings will be helpful in selecting appropriate strains for the manufacture of this product at the industrial level.     

核桃粕发酵乳菌种筛选及发酵条件优化研究

该试验对乳酸菌发酵核桃粕乳的不同菌株进行筛选,通过pH值、酸度及感官评定分析,从9株乳酸菌中筛选出嗜酸乳杆菌(Lactobacillus acidophilus)和植物乳杆菌(Lactobacillus plantarum)两株优良菌种。研究两株乳酸菌的复配比例,并与传统发酵剂发酵的核桃粕乳进行品质对比。结果表明,以嗜酸乳杆菌:植物乳杆菌(1∶1)接种发酵后得到的发酵核桃粕发酵乳综合品质最佳,其感官评分90分,氨基酸态氮含量57.0 mg/L,活菌总数7.35×107 CFU/mL,经发酵后的营养价值明显优于传统发酵剂发酵的核桃粕乳。     

Technological characterization of lactococci isolated from traditional Chinese fermented milks

To screen lactic acid bacteria for starter cultures in cheese production, 21 Lactococcus strains previously isolated from natural fermented milk and koumiss made by herdsman families in the Xinjiang, Gansu, and Qinghai provinces of China were evaluated for optimal growth temperature, acidification activity, proteolytic activity, aminopeptidase activity, and autolytic activity. All isolates presented low acidification rates, and the pH value did not reach 5.3 after 6 h of inoculation in sterile reconstituted skim milk at 30°C. Strains X9C2 and T7C showed the highest proteolytic activity of 24.67 and 23.58 mg of glycine/L of milk, respectively. For aminopeptidase activity, strains X9C2 and T1C2 displayed the highest activities of 30.56 and 27.70 U/mg of protein using l-leucine-p-nitroanilide as substrate, respectively. Autolytic activity in simulated cheese-like buffer ranged from 7.45 to 34.76%, and strains Q14C2 and Q16C showed the highest values of 34.76 and 34.20%, respectively. Collectively, one main finding is that some technological characteristics of Lactococcus isolates from Chinese traditional fermented products varied greatly. Some isolates with potentially important properties could be valuable for application as starter cultures of cheese or could constitute a mixed culture.     

功能性开菲尔牛奶酒最佳发酵条件的研究

本文利用开菲尔粒中分离的酵母菌和乳球菌采用两步发酵法研制开菲尔牛奶酒。针对影响牛奶酒制品最终品质的三个因素,采用三因素三水平L9(34)正交试验确定了功能性开菲尔牛奶酒的最佳发酵条件。成品开菲尔牛奶酒口感细腻、酸度适中,风味柔和,有浓郁的醇香和酯香味。     

马奶酒中抗单增李斯特氏菌细菌素产生菌的筛选及鉴定

从内蒙古传统乳制品酸马奶酒中分离出144株乳酸菌,通过双层琼脂平板扩散法筛选出具有明显抑菌作用的菌株.在排除有机酸、H2O2等的干扰后,其中1株乳酸菌的发酵上清液对受试的单核细胞增生李斯特氏菌表现出明显抑制作用;进一步硫酸铵沉淀、透析及浓缩处理后,其抑菌活性显著增强,说明其代谢产物中含有蛋白质类抑菌物质,可能是细菌素.经生理、生化试验和形态学观察,将该菌株鉴定为屎肠球菌(Enterococcus faecium).     

青海省海西地区传统发酵山羊乳中乳酸菌的分离及鉴定

从采自青海省海西地区牧民家庭的11份以山羊奶为主要原料的传统发酵乳中分离得到33株乳酸菌，经过形态特征．生理生化特性，糖发酵试验和乳酸旋光性的测定，将这33株乳酸茵初步鉴定为7株Lldelbrueckiisubsp．Lactis，4株L．delbrueckii subsp．bulgaricus.3株L．acidophilus，1株L，coryniformis subsp，corynformis，12株S．thermophilus，Ped．日cidilaaici 3株，Ped．Unnaeequi 2株，Leu．mesenteriodes subsp.mesentemiodes 1株，其中，杆菌占所分离菌株的45．45％，球菌占所分离菌株的54．55％。     

Identification of lactic acid bacteria in traditional fermented yak milk and evaluation of their application in fermented milk products

In this study, 53 strains of lactic acid bacteria (LAB) isolated from Xueo, a traditional fermented yak milk in the western Sichuan Plateau of China, were identified and their use in fermented milk was evaluated. All gram-positive and catalase-negative strains were divided into 6 groups at the level of 87% similarity using amplified ribosomal DNA restriction analysis. These groups were identified as 6 species using 16S rDNA sequence analysis and atpA gene analysis. The dominant LAB strains in Xueo were Enterococcus durans, Lactobacillus fermentum, and Lactobacillus paracasei, accounting for 45.3, 22.6, and 17.0% of isolates, respectively. Milk fermented with most of the representative strains was high in quality, exhibiting relatively high viscosity, moderate acidity, good sensory quality, and high counts of viable LAB. Fermented milk of E. durans SCA16 and L. fermentum SCA52 achieved the highest scores for overall sensory quality. Most strains displayed antimicrobial activity against at least 1 of 9 spoilage microorganisms. Lactic acid was the main factor inhibiting the growth of spoilage bacteria, and H(2)O(2) was also inhibitory to some extent. Excluding the influence of acid and H(2)O(2), strains SCA52 (L. fermentum) and SCA7 (Lactobacillus plantarum) were antagonistic against some of the indicators, suggesting that the 2 strains may produce a bacteriocin-like substance. Therefore, the development of superior strains isolated from Xueo to ferment milk with similar flavor and texture to Xueo is expected.     

发酵酸菜来源乳酸菌的益生特性及其在发酵乳中的应用

通过pH值、胆盐耐受性实验从发酵酸菜中筛选性能优良的益生乳酸菌株,经16S rRNA序列分析鉴定得4 株植物乳杆菌A44、B51、B54、C53和2 株戊糖乳杆菌A16、B72。经疏水、黏附、自凝聚和溶血能力实验评价6 株乳酸菌的益生特性,其中植物乳杆菌A44对氯仿和二甲苯的疏水性均大于80%,对Caco-2细胞的黏附率为13.57%,放置5 h的自凝聚率超过60%,与其他菌株相比具有更好的益生特性且无溶血活性。因此选用植物乳杆菌A44进一步研究其在发酵乳中的功能特性,结果表明：植物乳杆菌A44作为辅助发酵剂添加后对4 ℃贮藏7 d期间发酵乳pH值、滴定酸度和持水性均无显著影响（P＞0.05）,但是可以显著提高发酵乳的活菌数和黏度（P＜0.05）,活菌数达到8.45（lg（CFU/mL））。本研究筛选得到的植物乳杆菌A44是一株性能优良的益生乳酸菌,具有作为发酵乳益生菌辅助发酵剂的潜在应用前景。     

Ribotyping and biotyping of Lactobacillus helveticus from the koumiss

The lactic acid bacteria are very important components involved in the milk products processing. Lactobacillus helveticus, a homofermentative thermophilic lactobacillus commonly occurring in cheeses, has been isolated as the prevailing species in natural koumiss in this work. The species identification of six studied L. helveticus strains was based on rep-PCR fingerprinting with (GTG)₅ primer. Biotyping (API 50CH kit, conventional tests) showed phenotypic heterogeneity among isolates and enabled the identification to the genus Lactobacillus only. Ribotyping with restriction enzyme EcoRI yielded a strain-specific restriction patterns and allowed good strain differentiation. Obtained ribotypes of the isolates originating from the koumiss gave unique band patterns with low similarity to the selected reference cultures of L. helveticus. Despite low number of strains analyzed, the ribotype data showed certain heterogeneity that seemed to be not only strain dependent, but also related to the source of isolates. We concluded that analyzed isolates from koumiss represent a new ecovar, L. helveticus ecovar Koumiss.     

Characterization of the lactic acid bacteria in ewe's milk and cheese from northwest Argentina

Indigenous lactic acid bacteria in ewe's milk and artisanal cheese were studied in four samples of fresh raw milk and four 1-month-old cheeses from the provinces of northwest Argentina. Mean growth counts on M17, MRS, and MSE agar media did not show significant differences (P < 0.05) in raw milk and cheeses. Isolates of lactic acid bacteria from milk were identified as Enterococcus (48%), lactococci (14%), leuconostocs (8%), and lactobacilli (30%). All lactococci were identified as Lactococcus lactis (subsp. lactis and subsp. cremoris). Lactobacilli were identified as Lactobacillus plantarum (92%) and Lactobacillus acidophilus (8%). Enterococci (59%) and lactobacilli (41%) were isolated from cheeses. L. plantarum (93%), L. acidophilus (5%), and Lactobacillus casei (2%) were most frequently isolated. L. lactis subsp. lactis biovar diacetylactis strains were considered as fast acid producers. L. lactis subsp. cremoris strains were slow acid producers. L. plantarum and L. casei strains identified from the cheeses showed slow acid production. The majority of the lactobacilli and Lactococcus lactis strains utilized citrate and produced diacetyl and acetoin in milk. Enzyme activities (API-ZYM tests) of lactococci were low, but activities of L. plantarum strains were considerably higher. The predominance of L. plantarum in artisanal cheese is probably important in the ripening of these cheeses due to their physiological and biochemical characteristics.     

阿尤恩地区自然发酵牛乳中乳酸菌分离鉴定及多样性研究

为了分离、保藏自然发酵乳中乳酸菌菌种,丰富自然发酵乳中乳酸菌多样性信息.本文采用传统的纯培养分离方法和宏基因组16S rRNA基因测序技术对阿尤恩地区自然发酵牛乳的乳酸菌多样性进行研究.纯培养结果表明:5份自然发酵牛乳中共分离出111株乳酸菌,鉴定为5个属10个种,其中Lactococcus lactis,占总分离株的...     

发酵型核桃乳生产专用乳杆菌的选育

从发酵面包、白芥丝、泡菜、酸菜中分离到14株乳杆菌,经核桃乳发酵产酸特性初筛,确定菌株L5、L7、L8及L12适合核桃乳发酵。通过菌落形态、菌体形态和16S rRNA基因序列分析,确定4株菌分别为鼠李糖乳杆菌（Lactobacillus rhamnosus）、副干酪乳杆菌（Lactobacillus paracasei）、食窦魏斯氏菌（Weissella cibaria）干酪乳杆菌（Lactobacillus casei）。将上述菌株与实验室保存的嗜热链球菌SA进行核桃乳发酵实验,感官评分分别为78分、95分、83分和82分;4个发酵核桃乳中9种氨基酸含量均有不同程度提高,其中菌株L7发酵产品的氨基酸含量增幅最大,由0.67%增至1.60%。因此副干酪乳杆菌L7更适合发酵型核桃乳的生产。     

Characterisation of lactic acid bacteria isolated from fermented milk "laban"

The technological properties of 96 lactic acid bacteria isolated from Lebanese traditional fermented milk "laban" were characterised. They were classified by phenotypic and biochemical analyses as Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus, thus indicating that laban is a fermented milk similar to yogurt. Most strains of L. bulgaricus (87.5%) exhibited a high acidification activity, whereas strains of streptococci showed low acidification ability. 33.3% of streptococci strains and 25% of lactobacilli strains displayed similar acidification performances as European strains. Results obtained for syneresis, texture and rheological parameters led us to consider that isolated strains were not low polymer-producing strains. Some of them displayed interesting characteristics such as low syneresis and high values for rheological parameters. The major flavour compounds found in pure cultures were acetaldehyde, acetone, 2-butanone, dimethyl disulfide, acetoin, 2,3-butanedione, 2,3-pentanedione, and acetic, hexanoic and butanoic acids. Acetaldehyde (7.4%) and organic acids (48.3%) were mainly produced by L. bulgaricus strains, whereas streptococci cultures contained high relative levels of 2,3-butanedione and acetoin, which represented around 82% of the total flavour compounds. Finally, strains isolated from laban samples exhibited different technological properties than those used in yogurt production, thus conferring specific characteristics to this product.     

Identification and first characterization of lactic acid bacteria isolated from the artisanal ovine cheese Pecorino del Poro

A total of 225 lactic acid bacteria, isolated from the artisanal raw ovine milk cheese Pecorino del Poro, were identified and given an initial characterization; the best 18 strains were tested for their antagonistic activity against autochthonous Escherichia coli . An intense antagonistic activity against E. coli growth, induced by strains of Lactobacillus paracasei ssp. paracasei or L. curvatus , was observed; the E. coli inhibition can be ascribed to the production of bacteriocin-like compounds. The lactobacilli seem able to improve the hygienic quality of other artisanal raw milk cheeses.