Films based on kefiran,an exopolysaccharide obtained from kefir grain: Development and characterization

Kefiran, an exopolysaccharide produced by microorganisms present in the kefir grains, is a glucogalactan that has several health promoting properties. In the present work, the ability of kefiran to form films and the effect of glycerol addition at different concentrations on film properties was evaluated. Kefiran was able to form films at concentrations ranging from 5 to 10 g/kg. The concentration 10 g/kg was selected because the films were easily removed from the plate. All film-forming solutions exhibited a pseudoplastic behavior; glycerol addition did not modify the solution rheological properties.     

Development and characterisation of a new biodegradable edible film made from kefiran,an exopolysaccharide obtained from kefir grains

This study examined the feasibility of using kefiran, an exopolysaccharide obtained from kefir grains, as a new film-forming material. Kefiran-based films, with and without glycerol as plasticizer, were prepared by a casting and solvent-evaporation method. To study the impact of the incorporation of glycerol into the film matrix, physical, mechanical, and thermal properties of the films were investigated. As expected, the increase of glycerol concentration from 15% to 35% w/w increased extensibility but decreased tensile strength, implying higher mobility of polymer chains by the plasticizing effect of glycerol. Water vapour permeability of films was found to increase as the plasticizer content increased. Glass transition temperatures decreased as a result of plasticization as glycerol content increased. The properties of the films were related to their microstructure, which was observed by scanning electron microscopy. Thus, it was observed that plasticizer is a significant factor in the properties of these films and their food technology applications.     

Development of a new method for determination of exopolysaccharide quantity in fermented milk products and its application in technology of kefir production

An easy-to-use method for quantitative analysis of bacterial exopolysaccharides (EPS) was developed. The acid treatment was resultant in removing proteins and lactose and allowed to avoid long-term operation as dialysis. It was shown that, as opposed to anthron and phenol–sulphuric acid methods, reducing sugars method after EPS hydrolysis with glucose as standard provided reliable results, no matter to the EPS monosaccharide composition.     

Microbiological,physicochemical and rheological properties of fermented soymilk produced with exopolysaccharide (EPS) producing lactic acid bacteria strains

Growth of Lactobacillus plantarum 70810, Lactobacillus rhamnosus 6005 and a commercial yogurt starter culture in soymilk was investigated in the present study. It was found that the fermented soymilk using L. plantarum 70810 had significantly higher viable cell counts, water holding capacity (WHC, 88.27%), apparent viscosity (1840.35 mPa s) and exopolysaccharide (EPS) amount (832.15 mg/L) than the other two starter cultures in soymilk. Direct observation of microstructure in fermented soymilk indicated that the network structures of EPS-protein could improve the texture of fermented soymilk. Considering that the beneficial effects of L. plantarum 70810 in fermented soymilk, volatile compounds in fermented soymilk were further identified. Then the growth and fermentation characteristics of L. plantarum 70810 including changes in viable cell counts, pH, titratable acidity, apparent viscosity and EPS production during storage were investigated. In comparison to original soymilk base, the concentrations of the characteristic flavor compounds for fermented soymilk using L. plantarum 70810 increased, whereas hexanal, 2-pentylfuran and 2-pentanone in relation to beany flavor of soymilk decreased. In addition, fermented soymilk using L. plantarum 70810 maintained high viable cell count (>10⁸ cfu/mL), apparent viscosity (966.43 mPa s) and amounts of EPS (635.49 mg/mL) during storage at 4 °C for 21 days.     

Characterisation of the exopolysaccharide kefiran produced by lactic acid bacteria entrapped within natural kefir grains

The main goal of this work was to characterise and quantify the exopolysaccharide kefiran and to discover an effective procedure for its isolation from kefir grains, originating from the Caucasian Mountains. Capillary electrophoresis was used for the characterisation and quantification of the d ‐glucose and d ‐galactose in our samples at a mass ratio of 1:0.7. The effect of fermentation time on growth of kefir grains and the content of kefiran within the grains were determined. The pH profiles were monitored dynamically. In addition, the influence of fermentation temperature on kefir grains mass concentration (γ_KG) and the content of kefiran within the grains (w_KEF/KG) were studied. The highest values for both were obtained at 30 °C.     

Restoration of kefir grains subjected to different treatments

The aim of the study was to find a way to recover the quality of kefir grains that had been subjected to the following treatments: homogenisation, rinsing the grains with water, freeze‐drying and milling, freezing in liquid nitrogen and then frozen storage, and cool storage. The grains were studied in respect of their later replication in milk, their size and their microbiota composition. The daily transfer of treated kefir grains, except freeze‐dried ones, into fresh milk was effective in respect of the recovery of their growth dynamics, size and microbiota balance. The growth dynamics of grains in milk seems to be a very good indicator of their vital and technological functions.     

Application of culture-dependent and culture-independent methods for the identification of Lactobacillus kefiranofaciens in microbial consortia present in kefir grains

The biological and technological characteristics of kefiran as well as its importance in grain integrity led us to analyze the microbial kefir grain consortium with focus on Lactobacillus kefiranofaciens. The presence of L. kefiranofaciens in the nine kefir grains studied was demonstrated by denaturing gradient gel electrophoresis. By culture dependent methods applying a methodology focused on the search of this species, 22 isolates with typical morphology were obtained and identified applying a combination of SDS-PAGE of whole cell proteins, (GTG)₅-PCR and sequence analysis of the housekeeping gene encoding the α-subunit of bacterial phenylalanyl-tRNA synthase (pheS). This polyphasic approach allowed the reliable identification of 11 L. kefiranofaciens, 5 Lactobacillus paracasei, 4 Lactobacillus kefiri and 2 Lactobacillus parakefiri isolates. Isolated L. kefiranofaciens strains produced polysaccharide in strain-dependent concentrations and EPS produced by them also differed in the degree of polymerization. The isolation and accurate identification of L. kefiranofaciens is relevant taking into account the important role of this microorganism in the grain ecosystem as well as its potential application as starter in food fermentations.     

Isolation and characterization of the microbial population of different South African kefir grains

Kefir, a slightly acidic fermented milk, is produced by adding lactic acid bacteria and yeasts, in the form of grains, to milk. The bacteria and yeasts present in the kefir grains are known to vary widely. Selective growth media and morphological and biochemical characteristics were used for the isolation and identification of the microbes present in the grains from eight different sources in South Africa. The kefir grains were activated in milk for only 24 h to prevent any changes in the microbial population of the grains. The microbial numbers varied between 6.4 × 10⁴ and 8.5 × 10 ⁸  cfu/g on the media selective for the bacterial species and between 1.5 × 10 ⁵ and 3.7 × 10 ⁸   cfu/g on the media selective for the yeast species. The bacterial genera that were identified included Lactobacillus , Leuconostoc and Lactococcus and the yeast genera included Zygosaccharomyces , Candida and Saccharomyces . The distribution frequencies of the microbes in the different grains were determined and most of the grains were dominated by two microbial species. No pediococci, acetic acid bacteria or propionibacteria were detected.     

Microbiological study of lactic acid bacteria in kefir grains by culture-dependent and culture-independent methods

Lactic acid bacteria (LAB) in different original kefir grains were first assessed using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) by a culture-dependent way, and were further confirmed by DNA sequencing techniques. Results indicated that a combined method of cultivation with PCR-DGGE and subsequent DNA sequencing could successfully identify four LAB strains from three kefir grains from Taiwan (named Hsinchu, Mongolia and Ilan). Lactobacillus kefiri accounted, in the three kefir grains, for at least half of the isolated colonies while Lb. kefiranofaciens was the second most frequently isolated species. Leuconostoc mesenteroides was less frequently found but still in the three kefir grains conversely to Lactococcus lactis which based on culture-dependent isolation was only found in two of the kefir grains. It was interesting to find that all three kefir grains contain similar LAB species. Furthermore, the DGGE as a culture-independent method was also applied to detect the LAB strains. Results indicated that Lb. kefiranofaciens was found in all three kefir grains, whereas Lb. kefiri was only observed in Hsinchu kefir grain and Lc. lactis was found in both Mongolia and Ilan samples. Two additional strains, Pseudomonas spp. and E. coli, were also detected in kefir grains.     

Identification of microbial flora in kefir grains produced in Turkey using PCR

Kefir grains might have different ratios and/or content of microflora according to their origin. The purpose of this study was to determine microbial flora of kefir grains produced in three regional universities in Turkey using polymerase chain reaction (PCR) and consensus sequence primers. According to the results of PCR products with the specific primers, the following were identified as the natural inhabitants of the kefir grains: Lactobacillus acidophilus, Lactobacillus helveticus, Streptococcus thermophilus, Bifidobacterium bifidum and Kluyveromyces marxianus kefir. The results of this study revealed that traditional kefir produced by using kefir grains as natural starter cultures contains lactic acid bacteria especially Lactobacillus spp. One of the sources also contained B. bifidum. This is the first record on the presence of B. bifidum in kefir grains.     

Production of microbial exopolysaccharides in the sourdough and its effects on the rheological properties of dough

Exopolysaccharides (EPS) are exogenous microbial metabolites which are secreted mainly by bacteria and microalgae during growth. In addition to natural polysaccharides present in cereal grains flour and dough, microbial flora is usually involved in production of polysaccharide on sourdough fermentation. Total polysaccharides (microbial and flour) were extracted from sourdough and dough samples dehydrated and were added at the rate of 0%, 0.25%, 0.5%, 1%, 1.5%, 2% and 2.5% (w/w flour based) on the dough to investigate its effects on the rheological properties of the dough. Addition of polysaccharides to the dough increased the water absorption and decreased the dough softening after 20 min. Resistance to extension after 45, 90 and 135 min resting time was decreased by increasing the percentage of the added polysaccharides. Longer fermentation time for each level of polysaccharides led to greater stability. No significant differences were observed in the extensibility of dough. The overall effects of different levels of added polysaccharides resulted in a decrease in resistance to extension ratio of the samples. Energy input decreased in all cases. It seems therefore that addition of polysaccharides may be useful when bread is to be made with stronger flour and longer fermentation time is needed.     

Structural investigation and response surface optimisation for improvement of kefiran production yield from a low-cost culture medium

Kefiran, a water-soluble heteropolysaccharide with molecular weight of 1.35 × 10⁶ Da and a specific optical rotation of +64° (c 1.0, H₂O), was isolated from kefir grains grown in cheese whey and further purified through DEAE-Sepharose XK26. Response surface methodology was employed to optimise the culture conditions for kefiran production from kefir grains to be lactose concentration 67 g/l, yeast extract 13 g/l, pH 5.7 and temperature 24 °C. Intrinsic viscosity was 5.84 dl/g using the Huggins extrapolation and 5.53 dl/g using the Kramer extrapolation. Monosaccharide analysis revealed that kefiran is composed of glucose (Glc) and galactose (Gal) in a relative molar ratio of 1.0:1.1. Its structural features were elucidated by a combination of FT-IR, methylation and GC–MS analysis, periodate oxidation–Smith degradation, partial acid hydrolysis and NMR spectroscopy (¹H, ¹³C and HMBC). The data obtained indicated that kefiran possessed a backbone of (1 → 6)-linked Glc, (1 → 3)-linked Gal, (1 → 4)-linked Gal, (1 → 4)-linked Glc and (1 → 2,6)-linked Gal, with a branch attached to O-2 of Gal residues and terminated with Glc residues.     

Cheese whey fermented with kefir micro‐organisms: Antagonism against Salmonella and immunomodulatory capacity

The antagonistic effect against Salmonella spp. and the immunomodulatory capacity of whey fermented with kefir grains and with three strains isolated from them – Lactobacillus plantarum CIDCA 8327, Lactobacillus kefiri CIDCA 8348 and Kluyveromyces marxianus var. marxianus CIDCA 8154 – were evaluated. Both fermented products interfered with the capacity of Salmonella spp. to associate and invade Caco‐2/TC7 cells and reduced up to the basal level the expression of the CCL20 in response to a pro‐inflammatory stimulus on Caco‐2 CCL20:luc cells. The products with potential application as probiotics obtained by fermentation of whey with kefir micro‐organisms represent an alternative to increase whey economic value.     

In vitro comparison of probiotic properties of Lactobacillus casei Zhang, a potential new probiotic, with selected probiotic strains

Since 2004, our research group has isolated 240 Lactobacillus strains from Koumiss, a traditional fermented alcoholic beverage prepared from mare's milk in Inner Mongolia, Xinjiang of China and Mongolia. Among these Lactobacillus strains a novel strain with potential probiotic properties, Lactobacillus casei Zhang, was screened out and studied extensively for its probiotic properties, health-promoting effects and fermentation characteristics. In vitro tests indicated that L. casei Zhang had high tolerance to simulated gastric, intestine juices and bile salts, similar to commercial probiotic strains such as Lactobacillus acidophilus NCFM, Lactobacillus rhamnosus GG, L. casei Shirota and Bifidobacterium animalis Bb12. Higher acid-production activity and proteolytic activity was observed in the fermented milk inoculated with L. casei Zhang during the refrigerated storage than in the samples inoculated with the selected commercial probiotics. The yogurt samples fermented with L. casei Zhang exhibited similarer viable count (1.0 × 10⁹ cfu/mL) as the other samples after 28 d of refrigerated storage. The results suggested that L. casei Zhang showed good potential for application in functional foods and health-related products.     

Physicochemical properties of starches obtained from Polish potato cultivars

Selected physicochemical, thermal, and rheological properties of starches isolated from new Polish potato varieties were determined. The starches contained 25.7–30.0 g/100 g d.m. of amylose and 59.5–90.2 mg/100 g d.m. of phosphorus. Gelatinization temperatures were 62.6–64.0, 68.9–69.9, and 73.6–77.0°C for T_O, T_P, and T_E, respectively, whilst enthalpy of gelatinization amounted to 11.1–15.3 J/g. The retrogradation degree of starch was from 52.90 to 78.53%. Pasting curves showed significant differences between the starches. Peak viscosity and final viscosity ranges were 2035–4458 and 1931–2985 mPa · s, respectively. Starch pastes exhibited non‐Newtonian, shear thinning, and thixotropic behavior. After cooling they demonstrated diversified viscoelastic properties, however, all of them were classified as weak gels. Significant linear correlations among selected rheological parameters and amylose and phosphorus content were found. Results of principal component analysis demonstrated an ability to differentiate the starches isolated from different potato varieties.     

传统发酵牦牛酸乳中高产胞外多糖乳酸菌的筛选与鉴定

目的:以四川红原、西藏羊八井地区传统牦牛酸乳中分离出的57株乳酸菌作为实验菌种,筛选出高产胞外多糖乳酸菌。方法:通过菌落拉丝法、硫酸-苯酚法的测定。结果:实验菌株胞外多糖产量在19.95⁹1.08μg/m L之间,其中菌株代号为32-2、67-1、41-1和27的胞外多糖产量较高,分别为91.08、89.76、87.22、87.40μg/m L。通过16S r DNA序列同源性鉴定表明代号32-2菌株鉴定为屎肠球菌（Enterococcus faecium）、代号67-1菌株为肠膜明串株菌肠膜亚种（Leuconostoc mesenteroides subsp.mesenteroides）、代号41-1菌株为干酪乳杆菌（Lactobacillus casei）、代号27菌株为肠膜明串珠菌（Leuconostoc mesenteroides）。结论:为研究具有提高酸乳品质能力的发酵菌株提供依据。      

分离自藏灵菇的乳酸菌的益生特性

从藏灵菇中分离纯化5株乳酸菌,初步鉴定2株为嗜酸乳杆菌,3株为乳酸乳球菌。选取其中2株菌研究其益生特性。结果表明,从藏灵菇中分离出的乳酸菌具有良好的益生特性,2株乳酸菌在pH值为4～6可生长良好;耐热范围为30～60℃;胆盐耐受性为0.1%～0.5%;发酵液对金黄色葡萄球菌、沙门氏菌、大肠杆菌等肠道病原菌有抑制作用;对抗生素有不同程度的耐药性。     

一株西藏灵菇嗜热链球菌产胞外多糖的流变学特性

以一株西藏灵菇嗜热链球菌为研究对象,系统地研究质量浓度、剪切速率、热处理、pH及盐离子对其所产胞外多糖流变学特性的影响。结果表明,该胞外多糖溶液具有良好的增稠性,质量浓度为8.6 mg/mL时其黏度可达到85.63mPa.s;多糖溶液呈高度假塑性,剪切稀释现象明显;热稳定性差,当温度由20℃升至100℃时,其黏度下降了74.4%,分别于60,80℃热处理150min,对多糖溶液黏度有不同程度的影响;多糖溶液黏度在酸性条件下相对稳定,但在中性和碱性条件下明显升高;并且随Ca2+浓度升高,多糖溶液黏度显著下降。     

Gelation,thermal and pasting properties of pigeon pea (Cajanus cajan L.), dolichos bean (Dolichos lablab L.) and jack bean (Canavalia ensiformis) flours

This study evaluated the rheological, thermal and pasting properties of pigeon pea (PP), dolichos bean (DB) and jack bean (JB) legume flours and gels. Starch and protein contents were also measured and its molecular weight distribution was determined by electrophoresis. PP and DB showed the highest viscosities while JB had the highest pasting temperature. The minimum flour concentrations for gel formation were estimated at 6–8% for DB and PP and 10% for JB. Above these concentrations all flour suspensions heated to 95 °C led to gels with a solid-like behavior. Differential scanning calorimetry showed two endothermic peaks in all flours at 80–89 °C and 96–100 °C. Avrami model was successfully fitted to the hardening kinetics of PP and DB gels stored at 4 °C. The half-life times were 22 and 6 h for PP and DB respectively. PP and DB flours were able to form self-supporting gels and could be applied in the formulation of gel-like foods.