Proteolytic profiles of yogurt and probiotic bacteria

Nine strains of Streptococcus thermophilus, 6 strains of Lactobacillus delbrueckii ssp. bulgaricus, 14 strains of Lactobacillus acidophilus and 13 strains of Bifidobacterium spp. were screened for proteolytic, amino-, di-, tri- and endopeptidase activity by using the o-pthaldialdehyde-based spectrophotometric assay. Strains showing the highest and lowest proteolytic activity were further studied for their peptidase activities at the extracellular and intracellular levels. The amounts of free amino groups released by S. thermophilus, L. delbrueckii ssp. bulgaricus and L. acidophilus strains were higher than that by Bifidobacterium strains. Aminopeptidase activity was detected for all bacterial strains both at the extracellular and intracellular levels. The specific activity towards the six substrates studied was higher at the intracellular level for all strains. High dipeptidase activity was demonstrated by all bacterial strains for L. delbrueckii ssp. bulgaricus, L. acidophilus, and Bifidobacterium spp. whereas S. thermophilus had greater dipeptidase activity at the extracellular level. All bacterial cultures tested were able to hydrolyse large biologically active peptides, bradykinin, Ala–Ala–Ala–Ala–Ala and the tripeptide substrate Gly–Ala–Tyr at both the extracellular and intracellular levels. However, with the substrates ending with a C-terminal of phenylalanine, the hydrolysis only occurred at the intracellular level.     

Production of oligosaccharides in yogurt containing bifidobacteria and yogurt cultures

Yogurts were prepared by using yogurt cultures combined to mixed cultures of bifidobacteria (Bifidobacterium animalis, Bifidobacterium bifidum, Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium longum) and by adding a preincubation step (1.5 h at 50 degrees C) with bifidobacteria to the conventional method of manufacture in order to produce oligosaccharides. The survival of bifidobacteria was drastically affected during storage of yogurts, except for products containing B. animalis, in which viable counts remained at >10(6) cfu/g after 28 d of storage at 4 degrees C. Oligosaccharides with a degree of polymerization of 3 were produced during the preincubation step (0.31 to 0.68%), and the amount in the final products varied according to the species of bifidobacteria inoculated during the preincubation step or the concentration of bifidobacteria used as second inoculum during the fermentation process. In fact, the higher concentration of oligosaccharides measured at the end of the fermentation process (0.72%) and the 28 d-storage period (0.67%) was obtained for yogurts containing B. infantis. However, yogurts containing B. breve showed higher beta-galactosidase activities and had lower lactose concentrations after the fermentation process and the storage period than the other yogurts. The use of a mixed cultures of bifidobacteria (B. animalis, B. infantis, or B. breve) thus allows the production of yogurts in which bifidobacteria can survive in relatively high cell numbers and contain appreciable amount of oligosaccharides.     

黑豆双歧酸奶的研制

以黑豆为主要原料,通过加入双歧杆菌等发酵剂,研制出一种营养丰富、色泽美观的发酵酸豆奶。对其产品配方、发酵工艺条件进行了初步的分析探讨。     

Screening of lactic acid bacteria and bifidobacteria for potential probiotic use in Iberian dry fermented sausages

The purpose of this study was to select lactic acid bacteria and bifibobacteria strains as potential probiotic cultures during the processing of Iberian dry fermented sausages. A total of 1000 strains were isolated from Iberian dry fermented sausages (363), and human (337) and pig faeces (300) in different culture media. Around 30% of these strains, mainly isolated from Iberian dry fermented sausages in LAMVAB agar, were pre-selected for testing as potential probiotics by their ability to grow adequately at the pH values and NaCl concentrations of these meat products during the ripening process. Of the in vitro investigations used to predict the survival of a strain in conditions present in the gastro intestinal tract, exposure to pH 2.5 showed itself to be a highly discriminating factor with only 51 out of 312 pre-selected strains resisting adequately after 1.5h of exposure. All acid-resistant isolates identified as lactobacilli originated from human faeces (Lactobacillus casei and Lactobacillus fermentum) and pig faeces (Lactobacillus reuteri, Lactobacillus animalis, Lactobacillus murinus, and Lactobacillus vaginalis). Pediococcus acidilactici strains were isolated from Iberian dry fermented sausages and pig faeces, whereas the greatest number of Enterococcus strains were identified as Enterococcus faecium, with this species being isolated from Iberian dry fermented sausages, and human and pig faeces. Most of these strains are promising probiotic meat culture candidates suitable for Iberian dry fermented sausages.     

Metabolic activities and probiotic potential of bifidobacteria

It has been shown that the gut microbiota regulates fat storage in the body and that disturbances in its composition can lead to the development of certain metabolic disease states. Bifidobacteria are found among the resident microbiota in the gastrointestinal tract (GIT) and their metabolic activities have been shown to beneficially influence the human host. It has been reported that they inhibit intestinal colonisation by pathogenic microorganisms and have anti-carcinogenic, immunostimulatory, and anti-diarrhoeal properties, as well as aiding in the alleviation of lactose intolerance and ability to lower serum cholesterol levels in humans. One particular health promoting property of bifidobacteria is bioactive fatty acid production, which when ingested, may confer health benefits on the host. A bioactive fatty acid produced by bifidobacteria is conjugated linoleic acid (CLA), of which cis-9, trans-11 (c9, t11) and trans-10, cis-12 (t10, c12) CLA are the main biologically active CLA isomers. The production of CLA by Bifidobacterium can also have a positive effect on the immune system of the human host leading to numerous health benefits. This is an example of the metabolic activities of an ingested bacterium being beneficial to the host, rather than the direct interaction of the bacterium with the host.     

螺旋藻益生菌酸乳的研制

探讨了在纯鲜牛乳中添加螺旋藻制剂,经益生乳酸菌发酵,制得一种浅绿色、风味独特的新型营养保健酸奶.通过四因素三水平L9(34)正交试验确定了最佳配方工艺为:合格鲜牛乳,白砂糖7.0%,果胶0.08%(均为质量分数),益生菌L.casei Zhang 4×106g-1,直投式发酵剂CHR HANSEN YC-X11 0.006%,螺旋藻提取液6.0%,发酵温度42℃,发酵时间4 h.     

Development and characterization of probiotic (co)encapsulates in biopolymeric matrices and evaluation of survival in a millet yogurt formulation

The formulation of probiotics-enriched products still remains a challenge for the food industry due to the loss of viability, mainly occurring upon consumption and during storage. To tackle this challenge, the current study investigated the potential of using sodium alginate and inulin (SIN) in combination with various encapsulating materials such as skim milk (SKIM), whey protein concentrate (WPC), soy protein concentrate (SPC), and flaxseed oil (FS) to increase the viability of Lactobacillus casei upon freeze-drying, under simulated gastrointestinal conditions, during 28 days of storage at 4°C, and in a formulation of millet yogurt. Microstructural properties of microcapsules and co-microcapsules by SEM, oxidative stability of flaxseed oil in co-microcapsules, and physicochemical and sensory analysis of the product were performed. The produced microcapsules (SIN-PRO-SKIM, SIN-PRO-WP, and SIN-PRO-SP) and co-microcapsules (SIN-PRO-FS-SKIM, SIN-PRO-FS-WP, and SIN-PRO-FS-SP) had a high encapsulation rate >90%. Moreover, encapsulated and co-encapsulated strains exhibited a high in vitro viability accounting for 9.24 log10 CFU/g (SIN-PRO-SKIM), 8.96 log10 CFU/g (SIN-PRO-WP), and 8.74 log10 CFU/g (SIN-PRO-SP) for encapsulated and 10.08 log10 CFU/g (SIN-PRO-FS-SKIM), 10.03 log10 CFU/g (SIN-PRO-FS-WP), and 10.14 log10 CFU/g (SIN-PRO-FS-SP) for co-encapsulated. Moreover, encapsulated and co-encapsulated cells showed higher survival upon storage than free cells. Also, the SEM analysis showed spherical particles of 77.92–230.13 µm in size. The physicochemical and sensory analysis revealed an interesting nutritional content in the millet yogurt. The results indicate that the SIN matrix has significant promise as probiotic encapsulating material as it may provide efficient cell protection while also providing considerable physicochemical and nutritional benefits in functional foods.     

Selection of probiotic bacteria for the fermentation of a soy beverage in combination with Streptococcus thermophilus

Lactobacillus delbrueckii subsp. lactis R0187, Lactobacillus helveticus R0052, Lactobacillus rhamnosus R0011 and Bifidobacterium longum R0175 were examined for their ability to grow in combination with Streptococcus thermophilus cultures in milk and a laboratory soy beverage (LSB; both standardized to 4.5% protein and 2.3% fat). Strains R0011 and R0187 did not rapidly acidify the soy beverage despite good growth rates on soy carbohydrates. The S. thermophilus populations in the LSB were similar to that of milk even though milk had 30% more buffering capacity. In milk but not in soy, symbiosis with respect to acidification rate was observed between S. thermophilus and L. helveticus or B. longum. The populations of L. helveticus in the fermented products were similar in pure cultures or in the presence of the streptococci. However B. longum did not compete well in the mixed culture. Fermentation conditions varied as a function of the ability of S. thermophilus strains to acidify media to a pH of 4.65 (between 8 and 24 h). The probiotic populations in the mixed culture were influenced by the S. thermophilus strain and by the time of fermentation. Variations in growth rates of the bacteria did not appear to be linked to differences in initial redox or α-amino nitrogen levels. Strain selection enabled the preparation of a mixed starter, probiotic-fermented soy beverage containing 1.1 × 10⁸ CFU/mL of L. helveticus R0052, which represented approximately 13% of the total final population.     

Effects of inulin and sugar levels on the viability of yogurt and probiotic bacteria and the physical and sensory characteristics in probiotic ice-cream

Ice-cream containing probiotic bacteria was produced by mixing fortified milk fermented with probiotic strains with ice-cream mixes with different sugar concentrations (15, 18, 21% (w/w)). Cultures were grown (37 °C, 12 h) in UHT skimmed milk supplemented with or without inulin addition (1% and 2%). The fermented milk was added to the ice-cream mix to a level of 10% w/w.     

Growth and metabolism of selected strains of probiotic bacteria in milk

Growth and metabolism of five probiotic strains with well-documented health effects were studied in ultra-high temperature (UHT) treated milk, supplemented with 0.5% (w/v) tryptone or 0.75% (w/v) fructose. The probiotic strains were Lactobacillus acidophilus La5, Lb. acidophilus 1748, Lactobacillus rhamnosus GG, Lactobacillus reuteri SD 2112 and Bifidobacterium animalis BB12. Fermentation was followed for 72 h at 37 degrees C and the samples were analysed for pH, log cfu ml(-1), volatile compounds, organic acids and carbon dioxide. The strains reduced pH from 6.7 to between 3.9 and 4.4 after 24 h of incubation. All strains attained viable cell counts above 8.7-9.18 log cfu ml(-1) after 6-16 h of incubation. The two Lb. acidophilus strains showed a stable level of viable cells during 12-72 h of incubation but the three other strains showed a reduction of 0.4-1.1 log cfu ml(-1) from 24 to 72 h of incubation. However, all strains showed cell levels between 7.8 and 8.7 log cfu ml(-1) after 72 h of incubation. After 48 h of incubation, the amount of lactic acid produced varied according to strain from 6949 to 14,000 mg kg(-1) and acetic acid produced varied from 0 to 6901 mg kg(-1). Three of the strains metabolised citrate but only low amounts of diacetyl and acetoin were detected within strains, 0.2-0.8 and 6.5-10 mg kg(-1), respectively. Carbon dioxide produced varied from 221 to 3942 mg kg(-1) and was connected to the citrate-fermenting ability of the strain used and their carbohydrate fermentation pathway. Three of the strains produced detectable levels of acetaldehyde and the concentration varied from 9.4 to 12.6 mg kg(-1) after 24 h of incubation. All five probiotic strains showed very different profiles of metabolites during fermentation; however, the two Lb. acidophilus strains were the most alike. Our findings show the importance of controlling the fermentation time since the probiotic strains produced different amounts of metabolic products according to fermentation time.     

Effects of beta-glucan addition to a probiotic containing yogurt

ABSTRACT:  This study investigated the effects of addition of β-glucan from 2 different cereal sources (oat and barley) on growth and metabolic activity of Bifidobacterium animalis ssp. lactis (Bb-12™) as determined by plating on a selective medium in yogurt during prolonged cold storage. These yogurt batches were compared to unsupplemented and inulin supplemented controls. All batches were also assessed for syneresis. Oat β-glucan addition resulted in improved probiotic viability and stability comparable to that of inulin. It also enhanced lactic and propionic acid production. The barley β-glucan addition suppressed proteolytic activity more than that from oat. These improvements were hindered by greater syneresis caused likely by thermodynamic incompatibility. Small amplitude oscillatory measurements of acidified model mixture of β-glucan/skim milk solids showed formation of casein gel within the β-glucan network. Binary mixtures of β-glucan and skim milk solids had apparent pseudoplastic and non-Newtonian behavior governed mainly by β-glucan contribution. Above critical concentrations, the mixtures underwent phase separation with the lower phase rich in protein. The phase diagram also showed that the addition of β-glucan may be possible at or below 0.24 w/w%.     

双麦麦精对酸奶益生菌及其感官性质的影响

双麦麦精是以大麦,其他麦类为原料通过酶解提取的一种天然营养物质。在酸奶制作过程中添加不同比例的双麦麦精,采用平板涂布的方法检测不同储存时间下,酸奶中的保加利亚乳杆菌和嗜热链球菌的活菌数。并且通过感官实验评定双麦麦精对酸奶口感、口味、色泽以及香味的影响。结果表明,当双麦麦精添加量为3%,蔗糖4%时,对酸奶中益生菌的促生长作用最好。同时验证了3%双麦麦精对双歧杆菌也有明显的益生作用。而且,3%添加量的双麦麦精在益生作用以及感官性质方面都比同等添加量的低聚果糖效果好。     

Viability of microencapsulated bifidobacteria in set yogurt during refrigerated storage

Bifidobacteria are probiotic organisms that improve the microbial balance in the human gut. They can be incorporated as live cultures in fermented dairy foods, including yogurt, for transmission to humans. Because bifidobacteria are sensitive to high acidity, their viability in yogurt is limited. The objective of the present study was to investigate the effect of microencapsulation on the viability of bifidobacteria in yogurt during refrigerated storage for 30 d. Live bifidobacterial cells were encapsulated in kappa-carrageenan. Cell enumeration, determination of titratable acidity and pH, quantitation of lactic and acetic acids, and sensory evaluation (consumer test) were carried out on the yogurt samples. Microbiological results showed a decline of 78 and 70.5% in the population of Bifidobacterium longum B6 and B. longum ATCC 15708, respectively, for the treatments containing nonencapsulated cells. No difference in bifidobacterial population was observed in the encapsulated treatments. The acetic acid content in the yogurt with nonencapsulated bifidobacteria was higher than that in the plain yogurt (control) and encapsulated treatments. The increase in lactic acid content during storage was not different among the various treatments for B. longum B6, but was greater for nonencapsulated than encapsulated B. longum 15708 and the control. Consumers judged the nonencapsulated treatment as the most sour, which was likely due to the higher acetic acid content. Consumers preferred the control and nonencapsulated treatments over the encapsulated treatment. Microencapsulation appears to increase the viability of bifidobacteria in yogurt. This technique can be used to transmit bifidobacteria via fermented products provided that sensory characteristics of the product are improved or maintained.     

Some low homogenization pressures improve certain probiotic characteristics of yogurt culture bacteria and Lactobacillus acidophilus LA-K

Lactobacillus delbrueckii ssp. bulgaricus, Streptococcus salivarius ssp. thermophilus, and Lactobacillus acidophilus are dairy cultures widely used in the manufacture of cultured dairy products. Commonly used homogenization pressures in the dairy industry are 13.80 MPa or less. It is not known whether low homogenization pressures can stimulate bacteria to improve their probiotic characteristics. Objectives were to determine the effect of homogenization at 0, 3.45, 6.90, 10.34, and 13.80 MPa on acid tolerance, bile tolerance, protease activity, and growth of L. delbrueckii ssp. bulgaricus LB-12, S. salivarius ssp. thermophilus ST-M5, and L. acidophilus LA-K. The cultures were individually inoculated in cool autoclaved skim milk (4°C) and homogenized for 5 continuous passes. Growth and bile tolerance of samples were determined hourly for 10 h of incubation. Acid tolerance was determined every 20 min for 120 min of incubation. Protease activity was determined at 0, 12, and 24 h of incubation. All homogenization pressures studied improved acid tolerance of L. delbrueckii ssp. bulgaricus LB-12 but had no beneficial effect on protease activity and had negative effects on growth and bile tolerance. A pressure of 6.90 MPa improved acid tolerance, bile tolerance, and protease activity of S. salivarius ssp. thermophilus ST-M5, but none of the homogenization pressures studied had an effect on its growth. Homogenization pressures of 13.80 and 6.90 MPa improved acid tolerance and bile tolerance, respectively, of L. acidophilus LA-K but had no effect on protease activity and its growth. Some low homogenization pressures positively influenced some characteristics of yogurt culture bacteria and L. acidophilus LA-K. Culture pretreatment with some low homogenization pressures can be recommended for improvement of certain probiotic characteristics.     

决明子酸豆奶的研制

以大豆、牛奶为主要原料,添加适量决明子,研制出一种健眼明目的功能性保健乳品。对决明子酸豆奶配方组合和复合稳定剂的配比、发酵工艺进行了探讨。结果表明:以酸奶稳定剂29%、双叉奶稳定剂57%、酪蛋白酸钠14%的配比组合成一种复合稳定剂,对控制产品乳清析出具有很好的效果;以决明子浓缩液∶豆奶牛奶混合液=1∶20、发酵剂量3%、蔗糖量8%、稳定剂量0.3%组成产品配方,可很好地解决产品口感问题。     

传统发酵牦牛酸乳中益生性乳酸菌的体外筛选

为获得具有潜在益生特性的乳酸菌,实验以抗人工胃液、胆盐中生长效率和细胞表面疏水性为筛选指标,对羊八井地区传统发酵牦牛酸乳中分离出的38株乳酸菌进行体外筛选。最终通过测定结果综合比较得出菌株编号为YBJ-3、YBJ-11、YBJ-15、YBJ-17、YBJ-30和YBJ-33乳酸菌的益生特性较好,其中4株乳酸菌抗人工胃液能力大于90%,在胆盐中良好生长。通过16S r DNA序列分析鉴定,菌株编号YBJ-3为屎肠球菌(Enterococcus faecium)、YBJ-11为发酵乳杆菌(Lactobacillus fermentum)、YBJ-15为耐久肠球菌(Enterococcus durans)、YBJ-17为植物乳杆菌(Lactobacillus plantarum)、YBJ-30为植物乳杆菌(Lactobacillus plantarum)、YBJ-33为干酪乳杆菌(Lactobacillus casei)。     

Effects of Chlorella vulgaris and Arthrospira platensis addition on viability of probiotic bacteria in yogurt and its biochemical properties

It is a practice to add microalgae into plain and probiotic fermented milks in order to promote the functionality of these products via their direct health effects as well as the enhancing impact on viability of probiotic microorganisms in product and in gastrointestinal tract. In this study, the effects of addition of two species of microalgae including Arthrospira platensis and Chlorella vulgaris (seven yogurt treatments containing three concentrations for each microalgae—0.25, 0.50, and 1.00?%—and a control without microalgae) on pH, titrable acidity, and redox potential changes as well as on the viability of probiotic bacteria during fermentation and during a 28-day refrigerated storage period (5?°C) were investigated in yogurt. Also, the amounts of lactic and acetic acids at the end of fermentation were assessed. The culture composition of yogurt was ABY type, containing Lactobacillus acidophilus LA-5, Bifidobacterium lactis BB-12, Lactobacillus delbrueckii ssp. bulgaricus, and Stresptococcus themophilus. The addition of microalgae significantly (p?<?0.05) increased the viability of L. acidophilus and bifdobacteria at the end of fermentation and during the storage period. Treatments containing A. platensis had slower pH decline, faster acidity increase, longer incubation time, and greater final titrable acidity than those containing C. vulgaris and control. In treatments containing 0.5 or 1?% microalgae, the viability was almost higher than 10⁷ cfu/mL until the end of refrigerated storage.     

Comparison of four methods to enumerate probiotic bifidobacteria in a fermented food product

Four methods of enumeration were compared by monitoring levels of probiotic bifidobacteria in fermented oat drink during storage. Strains of Bifidobacterium longum and B. lactis were quantified by plate counts, fluorescent in situ hybridization (FISH), quantitative real-time PCR and commercial LIVE/DEAD BacLight bacterial viability kit, and the methods were further developed to suit the enumeration of bifidobacteria in fermented foods. Plate counts of both B. lactis and B. longum were lower than the PCR and FISH counts. The LIVE/DEAD counts of B. lactis were comparable to PCR and FISH counts. The plate counts of B. lactis were slightly but significantly lower than LIVE/DEAD counts, suggesting that the cells that were not able to grow on plates may have become dormant. The plate counts of B. longum were several log units lower than LIVE/DEAD counts, suggesting that a remarkable part of the cells were dormant. Real-time PCR and FISH were shown to suit the quantification of the total amount of probiotic bifidobacteria in foods. Plate counts and LIVE/DEAD counts provided conflicting information on viability especially in the case of B. longum. We conclude that the choice of enumeration method for probiotic bacteria may have significant effect on the results of the analysis. The strain-specific properties and the objects of the analysis should be taken into account when enumeration methods for different probiotic strains are chosen.     

Production of soy yogurt enriched with glyceollins

Soy milk was prepared from regular soybean (M1), soybean germinated for 3 days at 25°C (M2), and soybean germinated under fungal infection (M3). Rhizopus microsporus var. oligosporus ATCC 22959 was used as the elicitor for glyceollin production. Each soy milk was fermented with Streptococcus infantarius 12 and Weissella sp. 4 (1:5, v/v) for 12 h at 37°C. Significant induction of glyceollins was confirmed only in M3 soy milk and glyceollins maintained stably during 12 h fermentation period. The concentration of glyceollins in M3 yogurt was 2,400.4±83 and 2,525.2±158 μg/g dry matter (d.m.) at 0 and 12 h, respectively. The amount of daidzein was significantly higher in M3 yogurt (635.1±21) than that of M1 (417±11) and M2 (545±17 μg/g d.m.) yogurt in 12 h (p<0.05). M2 yogurt had the highest amount of genistein (695.3±17) followed by M3 (634.5±26) and M1 (612.5±14 μg/g d.m.) yogurt. M3 soy yogurt also showed the highest content of total phenolic compounds (5.37 mg/g) and antioxidant activity. The results indicated that functional soy yogurt can be prepared from soybean enriched with glyceollins.