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.     

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.     

Turkish kefir and kefir grains: microbial enumeration and electron microscobic observation†

Microbial populations in kefir and kefir grains were enumerated by plating. Total lactic acid bacteria, lactoccocci, lactobacilli and yeast populations increased during fermentation and increased slightly during cold storage. Kefir grains had a lactic acid bacteria : yeast ratio of 10 ⁹  : 10 ⁶ . In further studies, kefir grains were observed using scanning electron microscopy (SEM) methods, which indicated yeast colonization on the surface and middle part of the kefir grain. Three types of lactobacilli (short, long and curved) were noted throughout the grain. Lactococci were not observed under SEM; preparation of kefir grains for SEM may have caused removal of lactococci from the grains.     

Distribution of microorganisms with particular reference to encapsulated bacteria in kefir grains

Propagable and non-propagable kefir grains in a form resembling cauliflower florets were observed by scanning electron microscopy (SEM) and light microscopy. In propagable grains short and long rod-shaped bacteria and yeasts formed separate colonies on the outside surface and inside. Internally, filaments which derived from capsules around the cells extended radially from a population of long rod-shaped bacteria. In non-propagable grains long rod-shaped bacteria with filamentous appendages were not observed, but only short rod-shaped bacteria and yeasts. Indian ink preparations showed presence of encapsulated bacteria in propagable grains and absence of these in non-propagable ones. The above results suggest that encapsulated bacteria are responsible for propagation of kefir grains.     

Assessment of the microbial diversity of Brazilian kefir grains by PCR-DGGE and pyrosequencing analysis

The microbial diversity and community structure of three different kefir grains from different parts of Brazil were examined via the combination of two culture-independent methods: PCR-denaturing gradient gel electrophoresis (PCR-DGGE) and pyrosequencing. PCR-DGGE showed Lactobacillus kefiranofaciens and Lactobacillus kefiri to be the major bacterial populations in all three grains. The yeast community was dominated by Saccharomyces cerevisiae. Pyrosequencing produced a total of 14,314 partial 16S rDNA sequence reads from the three grains. Sequence analysis grouped the reads into three phyla, of which Firmicutes was dominant. Members of the genus Lactobacillus were the most abundant operational taxonomic units (OTUs) in all samples, accounting for up to 96% of the sequences. OTUs belonging to other lactic and acetic acid bacteria genera, such as Lactococcus, Leuconostoc, Streptococcus and Acetobacter, were also identified at low levels. Two of the grains showed identical DGGE profiles and a similar number of OTUs, while the third sample showed the highest diversity by both techniques. Pyrosequencing allowed the identification of bacteria that were present in small numbers and rarely associated with the microbial community of this complex ecosystem.     

Diversity and functional characteristics of lactic acid bacteria from traditional kefir grains

Traditional kefir grains were collected from distinct parts of Turkey, and their microbial profile was determined. A wide bacterial biota was observed formed by distinct lactic acid bacteria (LAB) in which Lactococcus lactis strains appeared to be dominant. Yeast species were also identified in kefir grains. Significant levels of antifungal and antibacterial activities were monitored in kefir isolates. All tested LAB produced an exopolysaccharide (EPS) containing glucose and galactose, and some strains formed a fructan‐type EPS. Importantly, low levels of antibiotic resistance were observed among the kefir isolates.     

开菲尔粒中乳酸菌的分离方法研究

本文分别对开菲尔滤液和洗液中乳酸菌的分离方法进行了研究。结果表明:使用添加了0.5%山梨酸钾的乳清改良培养基,倾注法(划线法)厌氧培养乳酸菌分离效果较好,并且滤液中的菌株比洗液中的长势更好,活力更高。     

Effect of different growth conditions on biomass increase in kefir grains

Kefir is a functional dairy product and the effects of kefir consumption on health have been well documented. Kefir grains have naturally high numbers of lactic acid bacteria and yeasts and are used in manufacturing kefir. The biomass of kefir grains slowly increases after successive fermentations. The effects of adding whey protein isolate, modified whey protein (MWP, fat replacer; Carbery Inc., Cork, Ireland), or inulin to milk and different atmospheric conditions (ambient or 6% CO₂) during fermentation on the increase in biomass of kefir grains were investigated. Reconstituted milks (10% milk powder) enriched with whey protein isolate (2%), MWP (2%), and inulin (2%) were inoculated with kefir grains and fermented in ambient and 6% CO₂ incubators at 25°C until a final pH of 4.6 was reached. Biomass increments of kefir grains were determined weekly over 30 d. Lactic acid bacteria and yeast contents of kefir grains were also determined. The highest biomass increase (392%) was found in kefir grains grown in milk supplemented with whey protein isolate under ambient atmospheric conditions. Application of CO₂ did not provide a significant supporting effect on the biomass of kefir grains. Addition of MWP significantly accelerated the formation of kefir grain biomass (223%). The use of whey protein isolate, MWP, or inulin in milk did not cause any adverse effects on the microbial flora of kefir grains.     

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.     

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.     

Investigation on culturable microflora in tibetan kefir grains from different areas of china

Four samples of Tibetan kefir grains (TK-ZJUJ 01-04) from Tibet and surrounding areas were investigated via phenotypic and genotypic methods to compare and analyze the diversity of culturable microflora among different origins. As a result, 4 genera of microorganisms from TK-ZJUJ01: Bacillus subtilis (2.9 × 10(7) cfu/mL), Lactococcus lactis (8.2 × 10(7) cfu/mL), Kluyveromyces marxianus (3.0 × 10(6) cfu/mL), Saccharomyces cerevisiae (9.0 × 10(6) cfu/mL); 4 genera from TK-ZJUJ02: Lactobacillus kefiri (1.0 × 10(8) cfu/mL), Pichia kudriavzevii (5.0 × 10(6) cfu/mL), K. marxianus (1.9 × 10(7) cfu/mL), Kazachstania unispora (6.2 × 10(7) cfu/mL); 6 genera from TK-ZJUJ03: Leuconostoc lactis (4.6 × 10(7) cfu/mL), L. lactis (3.0 × 10(7) cfu/mL), Lactobacillus plantarum (3.0 × 10(7) cfu/mL), K. unispora (3.0 × 10(6) cfu/mL), K. marxianus (2.0 × 10(6) cfu/mL), (1.7 × 10(7) cfu/mL); and 4 genera from TK-ZJUJ04: L. plantarum (1.8 × 10(7) cfu/mL), Acetobacter fabarum (5.0 × 10(6) cfu/mL), K. unispora (6.2 × 10(7) cfu/mL), Pichia guilliermondii (6.2 × 10(7) cfu/mL) were identified. Yeasts like P. kudriavzevii and P. guilliermondii isolated in this study were the first time reported in Tibetan kefir grains. For TK-ZJUJ 01-03, lactic acid bacteria were the major microorganisms, which accounted for more than 50% of all the microbial population, while for TK-ZJUJ04, the largest microbial group was yeasts which accounted for more than 50%. In a word, study of diversity and composition of microflora provided us theoretical foundation for further investigation and application of Tibetan kefir grains. Practical Application: This is the basic research in order to develop and industrialize a new kind of yogurt starter which is naturally formed microbiota with both lactic acid bacteria and yeasts in it.     

Gelling properties of kefiran, a food-grade polysaccharide obtained from kefir grain

The physicochemical and gelling properties of kefiran, a water-soluble glucogalactan with probed health-promoting properties, were investigated. Gel permeation chromatograms revealed a single distribution of molecular weight corresponding to 10⁷ Da. Intrinsic viscosity of kefiran determined using Huggins extrapolations was 6.0 dl/g and using Kramer approximations was 5.95 dl/g.Kefiran has a Newtonian behaviour in diluted solutions, which becomes pseudoplastic at higher concentrations. Rheological behaviour of the solution before and after freeze drying was evaluated by small deformation oscillatory rheological measurements. The mechanical spectrum of solution corresponded to an entangled network behaviour. After freeze–thaw treatment of the solution, a rheological behaviour transition from a liquid-like system to a gel was observed. The storage modulus (G′) in cryogels was 35 times higher than the value obtained for the solution. Rheological characteristics of the cryogel were influenced by kefiran concentration. As the polymer concentration increased, higher number of interactions was evident for the increment in both moduli (G′ and G″). The behaviour of kefiran cryogels about 37 °C determines its ability to melt at mouth temperature. These results suggest that kefiran cryogels could be an interesting alternative for its application in food formulations.     

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.     

开菲尔粒中乳酸菌的分离与鉴定

从4个不同来源的开菲尔粒中分离得到40株乳酸菌，并对这40株乳酸菌进行了鉴定．结果为德氏乳杆菌8株，德氏乳杆菌保加力亚种6株，高加索奶乳杆菌2株，嗜酸乳杆菌1株，粪肠球菌7株，屎肠球菌2株，嗜热链球菌1株，乳脂链球菌3株。     

开菲尔粒中酵母菌的分离及其发酵性能的研究

本文分别对开菲尔滤液和洗液中酵母菌的分离方法及其在牛乳中的发酵性能进行了研究。结果表明：从滤液和洗液中分离筛选出不同类型的乳糖发酵性酵母。它们在牛乳中发酵产酒精能力较强，确定了其在牛乳中的最佳发酵条件是：发酵温度为28℃，发酵时间为60h，接种量为3％。     

开菲尔粒及其发酵液中菌相分析

为了研制开菲尔发酵剂，本试验测定了开菲尔粒及其发酵液中几种主要微生物的数量。两者结果均表明其中乳酸菌多于醋酸菌，醋酸菌多于酵母菌，而且乳酸菌类中大多数为兼性厌氧菌。     

Utilization of African grains for sourdough bread making

Abstract: Acha and Iburu flours were singly subjected to sourdough fermentation with previously selected autochthonous starters. Sourdoughs were used (30%, wt/wt) as aroma carriers and acidifiers during short time fermentation with the addition of baker's yeast. Acha and Iburu sourdough breads were compared to wheat sourdough bread started with the same strains and to breads made with the same formula but using baker's yeast alone. During Acha and Iburu sourdough fermentations, starter lactic acid bacteria reached almost the same cell density found in wheat sourdoughs. Acidification was more intense. Iburu sourdough bread had the highest total titratable acidity, the lowest pH, and contained the highest levels of free amino acids and phytase activity. The values of in vitro protein digestibility did not differ between Acha sourdough and wheat sourdough breads, while Iburu sourdough bread showed a slightly lower value. Acha and Iburu sourdough breads showed lower specific volume and higher density with respect to wheat sourdough breads. Nevertheless, Acha and Iburu sourdough breads were preferred for hardness and resilience. As shown by sensory analysis, Acha and especially Iburu sourdough breads were appreciated for color, acid taste and flavor, and overall acceptability. Practical Application: This study was aimed at evaluating the technological and nutritional properties of the African cereals Acha and Iburu. Sourdough fermentation and the use of selected starters increased the nutritional and sensory qualities and the potential application for bakery industry.     

Comparative study of the biochemical changes and volatile compound formations during the production of novel whey-based kefir beverages and traditional milk kefir

Cheese whey (CW) and deproteinised cheese whey (DCW) were investigated for their suitability as novel substrates for the production of kefir-like beverages. Lactose consumption, ethanol production, as well as organic acids and volatile compounds formation, were determined during CW and DCW fermentation by kefir grains and compared with values obtained during the production of traditional milk kefir. The results showed that kefir grains were able to utilise lactose from CW and DCW and produce similar amounts of ethanol (7.8–8.3 g/l), lactic acid (5.0 g/l) and acetic acid (0.7 g/l) to those obtained during milk fermentation. In addition, the concentration of higher alcohols (2-methyl-1-butanol, 3-methyl-1-butanol, 1-hexanol, 2-methyl-1-propanol, and 1-propanol), ester (ethyl acetate) and aldehyde (acetaldehyde) in cheese whey-based kefir and milk kefir beverages were also produced in similar amounts. Cheese whey and deproteinised cheese whey may therefore serve as substrates for the production of kefir-like beverages similar to milk kefir.