Oat-based Symbiotic Beverage Fermented by Lactobacillus plantarum, Lactobacillus paracasei ssp. casei, and Lactobacillus acidophilus

ABSTRACT: Oats and probiotics have long been recognized for their health benefits. The objectives of this study were (1) to study the ability of Lactobacillus plantarum (B-28), Lactobacillus paracasei ssp. casei (B-29) isolated from a traditional Bulgarian cereal-based fermented beverage, and Lactobacillus acidophilus from Chr. Hansen, Milwaukee, Wis., U.S.A., to remove cholesterol from the media and to adhere to the Caco-2 cell line, (2) to optimize the fermentation conditions to develop a beverage using these probiotics and oats with acceptable sensory and nutritional qualities, and (3) to assess the quality and shelf-life of this beverage and survivability of probiotics in the beverage. Lactobacillus acidophilus , B-28, and B-29 were found to remove 70.67%± 2.35%, 20.26%± 2.63%, and 16.75%± 3.83% of cholesterol from media and the percentage of adhesion was 4.69%± 0.78%, 1.92%± 0.78%, and 8.36%± 0.78%, respectively. The blend of oat flour, sugar, inulin, and whey protein concentrate was cooked in water and fermented for 12 h at 37°C by 2 ± 10⁶ colony-forming units (CFU) /mL each of B-28 and B-29 and 2 ± 10⁸ CFU/mL of L. acidophilus . The beverage had 0.87%± 0.03% of dietary fiber and had better sensory qualities compared with the commercially available similar product. The probiotics survived for 10 wk of storage at 4°C, except for L. acidophilus , which survived for about 4 wk. The population of B-28 was 1.77 ± 10⁶ to 1.29 ± 10⁷ CFU/mL and that of B-29 was 7.39 ± 10⁷ to 4.49 ± 10⁸ CFU/mL throughout the storage period. The oat-based symbiotic beverage is a functional drink providing both probiotics and prebiotics at the same time.     

Prevention of gastrointestinal infection using immunobiological methods with milk fermented with Lactobacillus casei and Lactobacillus acidophilus

The protective effect of feeding milk fermented with a mixture of Lactobacillus casei sp. and Lb. acidophilus sp. against Salmonella typhimurium infection in mice was compared with that obtained feeding milks fermented with these microorganisms individually. The survival rate obtained after oral infection with Sal. typhimurium was 100% in mice pretreated by feeding during 8 d with the mixture of Lb. casei and Lb. acidophilus fermented milks. Similar treatments with the individual milks were ineffective. Moreover, mice became more susceptible to infection with Sal. typhimurium after such treatment. The colonization of liver and spleen with the pathogen was markedly inhibited by the pretreatment with the mixture of fermented milk, while such inhibition was not observed using the Lb. casei and Lb. acidophilus milks. The highest levels of anti-salmonellae antibodies in serum and in intestinal fluid were found in the group of mice fed with the mixture and with Lb. casei fermented milk respectively. However, this latter milk was not effective in protecting against Sal. typhimurium. When the mice were first infected with Sal. typhimurium and then fed with the mixture of fermented milks, pathogen colonization was not prevented. The results suggest that the augmentation of resistance to salmonellae caused by the treatment with Lb. casei- + Lb. acidophilus-fermented milk was due to the anti-salmonellae protective immunity mainly mediated by the mucosal tissue. Milk fermented with this mixture could be used as an immunobiological method to prevent gastrointestinal infection.     

温度对嗜酸乳杆菌和干酪乳杆菌发酵羊奶的影响

以鲜羊奶为原料,保加利亚乳杆菌和嗜热乳链球菌为基础发酵剂,在此基础上分别添加嗜酸乳杆菌和干酪乳杆菌,研究温度对两菌株发酵羊奶过程中酸度、pH、活菌数及总活茵数的影响.结果表明:嗜酸乳杆菌酸羊奶的最佳发酵温度为37℃,此时凝乳时间为4h,酸度、pH、嗜酸乳杆菌数和总活菌数分别为97.8 0T、3.88、1.8×107 CFU/mL和1.38×109 CFU/mI,感官评价总分可达到8.35分;干酪乳杆菌羊奶的最佳发酵温度为39℃,此时酸羊奶凝固时间为4.5h,酸度、pH、干酪乳杆菌数和总茵数分别为79.2 °T、4.48、1.56×108 CFU/mL和1.81×109 CFU/mL,感官评价总分可达到7.00分.     

The increased viability of probiotic Lactobacillus salivarius NRRL B-30514 encapsulated in emulsions with multiple lipid-protein-pectin layers

Probiotics have demonstrated various health benefits but have poor stability to sustain food processing and storage conditions, as well as after ingestion. Biopolymer beads are commonly studied to encapsulate probiotic cells to improve their stability, but the millimeter-dimension of these beads may not meet the quality requirement of food products. The aim of this study was to enhance the viability of Lactobacillus salivarius NRRL B-30514 by encapsulation in emulsion droplets with multiple lipid-protein-pectin layers. Spray-dried L. salivarius was suspended in melted anhydrous milk fat that was then emulsified in a neutral aqueous phase with whey protein isolate or sodium caseinate to prepare primary solid/oil/water (S/O/W) emulsions. Subsequently, pectin was electrostatically deposited onto the droplet surface at pH 3.0 to form secondary emulsions. The encapsulation efficiency was up to 90%. After 20-day storage at 4 °C, the viable cell counts of bacteria in secondary emulsions at pH 3.0 and primary emulsions at 7.0 were 3 log higher than the respective free cell controls. After heating at 63 °C for 30 min, free L. salivarius was inactivated to be undetectable, while about 2.0 log CFU/mL was observed for primary (at pH 7.0) and secondary (at pH 3.0) emulsion treatments. Additionally, a 5 log-CFU/g-powder reduction was observed after spray drying free L. salivarius, while a 2 log CFU/g reduction was observed for emulsion treatments with capsules smaller than 20 μm. Furthermore, cross-linking the secondary emulsion with calcium enhanced the viability of L. salivarius after the simulated gastric and intestinal digestions. Therefore, the studied S/O/W emulsion systems may be used to improve the viability of probiotics during processing, storage, and gastrointestinal digestion.     

Effect of nutrient supplements on growth and viability of Lactobacillus johnsonii NRRL B-2178 in whey

The main objectives of the paper were to study the effects of nutrient supplementation of cows' whey on the growth and viability of the probiotic microorganism Lactobacillus johnsonii NRRL B-2178. Because of the short lifespan of probiotics, this study was aimed at the evaluation of the contribution of nutrients in the improvement of growth and viability of the microorganism for the purpose of its further application in the formulation of whey-based beverages. A maximal cell growth of 8.70 log₁₀ cfu mL⁻¹ and viability of less than 5 days were achieved during the fermentation of whey supplemented with yeast extract (3.0%) and inulin (1.0%), including the synergistic effect of temperature 39 °C. Elimination of inulin from the fermentation process reduced the viable cell count by 0.2 log₁₀ cfu mL⁻¹, but the addition of 1.0% inulin after fermentation extended the viability of Lb. johnsonii NRRL B-2178 by 10 days.     

Production of free conjugated linoleic acid by Lactobacillus acidophilus and Lactobacillus casei of human intestinal origin

A gas chromatographic procedure was used for analysis of conjugated linoleic acid (CLA) isomers cis-9, trans-11-octadecadienoic; trans-10, cis-12 octadecadienoic; and trans-9, trans-11-octadecadienoic (c9t11, t10c12, t9t11) produced by lactobacilli. Four different cultures, two strains each of Lactobacillus acidophilus and Lactobacillus casei were tested for their ability to produce CLA from free linoleic acid in MRS broth supplemented with linoleic acid. Different concentrations of linoleic acid (0, 0.05, 0.1, 0.2 and 0.5 mg/ml) were added to MRS broth, inoculated with the lactobacilli, and incubated at 37 degrees C. Viable counts and amounts of individual isomers of CLA (c9t11, t10c12, t9t11) were measured at 0, 24, 48, and 72 h. All the cultures were able to produce free CLA in media supplemented with linoleic acid. Maximum production of CLA (80.14 to 131.63 microg/ml) was observed at 24 h of incubation in broth containing 0.02% of free linoleic acid. No significant (P > 0.05) increases in total CLA levels were observed after 24 h of incubation. The ability of the cultures to produce CLA in skim milk supplemented with 0.02% free linoleic acid also was studied. In this medium, the total amounts of free CLA after 24 h of incubation ranged from 54.31 to 116.53 microg/ml. The use of lactic acid bacteria able to form free CLA in cultured dairy products may have potential health or nutritional benefits. Free CLA in the products likely would be more readily available for absorption from the digestive tract than if it were incorporated into the cells of the starter culture.     

Effect of pasteurisation on ascorbic acid,dehydroascorbic acid,tocopherols and fatty acids in pooled mature human milk

Ascorbic and dehydroascorbic acids (vitamin C), tocopherols (vitamin E) and unsaturated fatty acids are heat-sensitive and therefore, their concentrations in human milk could be affected by pasteurisation. Here we determined the concentrations of ascorbic acid plus dehydroascorbic acid, ascorbic acid alone, and α- and γ-tocopherols, and the percentages of fatty acids in samples of human milk after pasteurisation by a slow (62.5 °C, 30 min) or fast heating (100 °C, 5 min) procedure. Both methods led to a significant decrease in the concentrations of ascorbic acid plus dehydroascorbic acid (12% and 29%), ascorbic acid (26% and 41%), α-tocopherol (17% and 34%) and γ-tocopherol (13% and 32%), respectively. However, milk fatty acids, including the polyunsaturated long-chain fatty acids, were unaffected by the two methods. On the basis of these observations, we recommend that human milk be treated using a slow pasteurisation. In addition, we propose ascorbic acid as a marker of the degree of heat treatment.     

酸奶中嗜酸乳杆菌和植物乳杆菌检测方法的研究

根据嗜酸乳杆菌、植物乳杆菌、嗜热链球菌和保加利亚乳杆菌对不同碳水化合物的选择性利用，设计了适合上述4种乳酸菌的选择性计数培养基，并对其效果进行了验证。结果表明，利用MRS培养基、麦芽糖-MRS培养基和山梨醇-MRS培养基(两种培养基共同使用)能满足选择性计数的要求。以所设计的选择性培养基，研究了含有4种乳酸菌的酸奶中不同乳酸菌在产品货架期内活菌数的变化规律。     

响应面法提高植物乳酸菌和嗜酸乳杆菌发酵椰奶的活菌数

为进一步提高椰子饮料营养价值,以新鲜椰子为原料,嗜酸乳杆菌和植物乳杆菌为菌种,辅以蔗糖和葡萄糖,经发酵研制出一种具独特风味、富含益生菌的发酵椰奶。通过单因素实验研究发酵温度、发酵时间、接种量及菌比值对发酵椰奶中活菌数的影响。采用Box-Behnken实验设计和响应面法,以活菌数为指标,优化发酵椰奶的发酵工艺条件。结果表明,当发酵温度为41℃、发酵时间为18 h、接种量为9%、菌比例(植物乳杆菌/嗜酸乳杆菌)(Z/S)为1时,发酵椰奶活菌数含量达2.011×108 CFU/mL,与模型预测值较为吻合,远高于市场标准。发酵椰奶富含的乳酸菌均可定殖于人体胃肠道,发挥护肠保健作用,保留了纯正的椰香,市场前景广阔。     

Rennet paste from lambs fed a milk substitute supplemented with Lactobacillus acidophilus: effects on lipolysis in ovine cheese

The present work was undertaken to evaluate the effects of Lactobacillus acidophilus supplementation of a milk substitute on the features of lamb rennet paste used for cheese making. Lipolysis in cheese manufactured with rennet paste from lambs receiving supplemented milk was also evaluated. Lambs were subjected to 3 different feeding regimens (mother suckling, MS; artificial rearing, AR; and artificial rearing with 7 log₁₀ cfu/mL of Lb. acidophilus supplementation of the milk substitute, ARLb) and slaughtered at 20 and 40 d of age for each feeding treatment. Abomasa of the lambs were processed to rennet paste. Microbial loads, enzymatic activities (chymosin, pepsin, and lipases), and renneting characteristics of the lamb rennet paste were determined. Free fatty acids and conjugated linoleic acids were detected in cheese at 60 d of ripening. Addition of 7 log₁₀ cfu/mL of Lb. acidophilus to the milk substitute was carried out successfully. Total recovery of viable cells was recorded in milk supplied daily to the lambs in the ARLb group. The ARLb rennet had greater amounts of lactobacilli than did the MS or AR rennet, irrespective of the slaughter age of the lambs, and the ARLb rennet had higher concentrations of lactococci when lambs were slaughtered at 40 d of age. Chymosin and lipase activities were also higher in ARLb rennet than in MS or AR rennet from lambs slaughtered at an older age. Milk supplementation of ARLb lambs resulted in improved coagulating ability of the rennet and enhanced cheese lipolysis after 60 d of ripening. A reduction of all free fatty acids was observed in all cheeses when passing from 20 to 40 d of slaughter of the lambs. Conjugated linoleic acids were more abundant in ARLb cheeses at both 20 and 40 d. Therefore, supplementation of the milk substitute with Lb. acidophilus improved the enzymatic features of rennet and the healthful and nutritional characteristics of it the ovine cheese. Moreover, the addition of lactobacilli to the milk substitute made it possible to increase the slaughter age of lambs without detrimental effects on rennet characteristics.     

Production of beta-glucosidase and hydrolysis of isoflavone phytoestrogens by Lactobacillus acidophilus, Bifidobacterium lactis, and Lactobacillus casei in soymilk

ABSTRACT:  The study determined β-glucosidase activity of commercial probiotic organisms for hydrolysis of isoflavone to aglycones in fermenting soymilk. Soymilk made with soy protein isolate (SPI) was fermented with Lactobacillus acidophilus LAFTI^® L10, Bifidobacterium lactis LAFTI^® B94, and Lactobacillus casei LAFTI^® L26 at 37 °C for 48 h and the fermented soymilk was stored for 28 d at 4 °C. β-Glucosidase activity of organisms was determined using ρ-nitrophenyl β-D-glucopyranoside as a substrate and the hydrolysis of isoflavone glycosides to aglycones by these organisms was carried out. The highest level of growth occurred at 12 h for L. casei L26, 24 h for B. lactis B94, and 36 h for L. acidophilus L10 during fermentation in soymilk. Survival after storage at 4 °C for 28 d was 20%, 15%, and 11% greater ( P < 0.05) than initial cell counts, respectively. All the bacteria produced β-glucosidase, which hydrolyzed isoflavone β-glycosides to isoflavone aglycones. The decrease in the concentration of β-glycosides and the increase in the concentration of aglycones were significant ( P < 0.05) in the fermented soymilk. Increased isoflavone aglycone content in fermented soymilk is likely to improve the biological functionality of soymilk.     

Selective enumeration of Lactobacillus delbrueckii ssp. bulgaricus,Streptococcus thermophilus,Lactobacillus acidophilus,bifidobacteria, Lactobacillus casei,Lactobacillus rhamnosus,and propionibacteria

Nineteen bacteriological media were evaluated to assess their suitability to selectively enumerate Lactobacillus delbrueckii ssp. bulgaricus, Streptococcus thermophilus, Lactobacillus casei, Lactobacillus rhamnosus, Lactobacillus acidophilus, bifidobacteria, and propionibacteria. Bacteriological media evaluated included Streptococcus thermophilus agar, pH modified MRS agar, MRS-vancomycine agar, MRS-bile agar, MRS-NaCl agar, MRS-lithium chloride agar, MRS-NNLP (nalidixic acid, neomycin sulfate, lithium chloride and paramomycine sulfate) agar, reinforced clostridial agar, sugar-based (such as maltose, galactose, sorbitol, manitol, esculin) media, sodium lactate agar, arabinose agar, raffinose agar, xylose agar, and L. casei agar. Incubations were carried out under aerobic and anaerobic conditions at 27, 30, 37, 43, and 45 degrees C for 24, 72 h, and 7 to 9 d. S. thermophilus agar and aerobic incubation at 37 degrees C for 24 h were suitable for S. thermophilus. L. delbrueckii ssp. bulgaricus could be enumerated using MRS agar (pH 4.58 or pH 5.20) and under anaerobic incubation at 45 degrees C for 72 h. MRS-vancomycine agar and anaerobic incubation at 43 degrees C for 72 h were suitable to enumerate L. rhamnosus. MRS-vancomycine agar and anaerobic incubation at 37 degrees C for 72 h were selective for L. casei. To estimate the counts of L. casei by subtraction method, counts of L. rhamnosus on MRS-vancomycine agar at 43 degrees C for 72 h under anaerobic incubation could be subtracted from total counts of L. casei and L. rhamnosus enumerated on MRS-vancomycine agar at 37 degrees C for 72 h under anaerobic incubation. L. acidophilus could be enumerated using MRS-agar at 43 degrees C for 72 h or Basal agar-maltose agar at 43 degrees C for 72 h or BA-sorbitol agar at 37 degrees C for 72 h, under anaerobic incubation. Bifidobacteria could be enumerated on MRS-NNLP agar under anaerobic incubation at 37 degrees C for 72 h. Propionibacteria could be enumerated on sodium lactate agar under anaerobic incubation at 30 degrees C for 7 to 9 d. A subtraction method was most suitable for counting propionibacteria in the presence of other lactic acid bacteria from a product. For this method, counts of lactic bacteria at d 3 on sodium lactate agar under anaerobic incubation at 30 degrees C were subtracted from counts at d 7 of lactic bacteria and propionibacteria.     

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.     

乳酸菌发酵荠菜－番茄复合蔬菜汁的动力学研究

以荠菜和番茄为基料,用嗜酸乳杆菌和植物乳杆菌为试验菌种进行乳酸发酵,在发酵工艺优化的基础上,对5L 罐乳酸菌分批发酵过程进行研究,建立了菌体生长、产物生成和底物消耗动力学模型,经验证,模型的理论值和实验值拟和较好。所建动力学模型对乳酸菌发酵荠菜- 番茄复合蔬菜汁的反应器设计、工艺原理以及有效的控制发酵过程具有指导意义。     

Conjugated linoleic acid conversion by six Lactobacillus plantarum strains cultured in MRS broth supplemented with sunflower oil and soymilk

Six strains of Lactobacillus plantarum, isolated from traditional dairy products of minority nationalities, were evaluated for their ability to produce conjugated linoleic acid (CLA) from free linoleic acid in vitro. All the 6 strains were found to be capable of converting linoleic acid to CLA when using sunflower oil as substrate or during soymilk fermentation. The inhibitory effect of linoleic acid on the growth of the L. plantarum was also discussed. The production of CLA was increased with adding high concentration of substrate in sunflower oil and IMAU60042 produced the highest CLA both in sunflower oil and soymilk. The CLA was composted by 2 isomers: cis9, trans11-CLA and tran10, cis12-CLA, and cis9, tran s11-CLA covered the most part of the total CLA formed except for L. plantarum P8. The production of CLA was decreased during the storage of fermented soymilk. The CLA contents decreased significantly in the first week, also more quickly in 2 wk. Especially, tran10, cis12-CLA decreased more rapidly than cis9, tran11-CLA. No dramatic change was observed among other 8 fatty acids in soymilk. The proportion of unsaturated fatty acids varied after fermentation with different L. plantrum strains, but all decreased the during storage. The research on the ability of converting CLA of L. plantrum strains could be basis for the future research and development of fermented soymilk products. PRACTICAL APPLICATION: Desirable probiotic traits, such as acid and bile tolerance, aggregation activity, and antibacterial activity, have been proved for the 6 Lactobacillus plantarum strains. The 6 L. plantarum strains might be used in the fermentation of soymilk to produce multifunctional probiotic soymilk products, especially the rich CLA contents.     

Evaluation of enzymic potential for biotransformation of isoflavone phytoestrogen in soymilk by Bifidobacterium animalis,Lactobacillus acidophilus and Lactobacillus casei

Three strains of Lactobacillus acidophilus, two of Lactobacillus casei and one of Bifidobacterium were screened for β-glucosidase activity using ρ-nitrophenyl-β-d-glucopyranoside as a substrate and their potential for the breakdown of isoflavone glucosides to the biologically active aglycones in soymilk. Isoflavones quantification with HPLC and β-glucosidase activity were performed after 0, 12, 24, 36, and 48 h of incubation in soymilk at 37 °C. All six micro-organisms produced β-glucosidase, which hydrolysed the predominant isoflavone β-glucosides. There was a significant increase and decrease (P < 0.05) in the concentration of isoflavone aglycones and glucosides, respectively, in fermented soymilk. Based on the concentration of isoflavones during peak β-glucosidase activity, the hydrolytic potential was calculated. L. acidophilus 4461 had the highest aglycone concentration of 76.9% after 24 h of incubation, up from 8% in unfermented soymilk (at 0 h). It also had the best isoflavone hydrolytic index of 2.01, signifying its importance in altering the biological activity of soymilk.     

Phylogenetic study of Lactobacillus acidophilus group, L. casei group and L. plantarum group based on partial hsp60, pheS and tuf gene sequences

The partial nucleotide sequences encoding elongation factor Tu (tuf gene), 60-kDa heat shock protein (hsp60 gene) and phenylalanyl-tRNA synthase (pheS gene) were determined to assess the suitability as phylogenetic markers for discriminating the closely related species in Lactobacillus acidophilus group, L. casei group and L. plantarum group. A total of 234 lactobacilli were chosen from traditional fermented dairy products that were not exactly assigned to species based on biochemical tests and 16S rRNA gene sequences. The sequencing of partial tuf, hsp60 and pheS gene of all strains was performed, and then, the phylogenetic trees were constructed by neighbor-joining method. Phylogenetic tree revealed three genes provided better resolution of each Lactobacillus species than 16S rDNA, and all of strains were clearly identified as L. casei (63 strains), L. plantarum (58 strains) and L. helveticus (113 strains) by comparison of sequences with the type strains. From our results, the partial sequences of three genes had a higher discriminatory power than 16S rRNA gene sequences and were an alternative molecular tool for the taxonomical analysis of L. casei group, L. plantarum group and L. acidophilus group.     

大豆多肽对乳酸菌增殖作用的影响研究

研究大豆多肽对乳酸菌增殖作用.用大豆多肽按比例替代脱脂乳粉制作复原乳,接种乳酸菌于41 ℃恒温静态培养,测定乳酸菌培养过程中培养液的pH值、滴定酸度、乳酸菌生长量以及贮藏期活菌数.试验结果表明,大豆多肽添加量为3%时,乳酸菌活菌数增菌数达到最高,为1.2×107 CFU/mL.发酵终点乳酸菌活菌数是空白对照组菌数的1.3倍.