Influence of isomalto-oligosaccharides on intestinal microbiota in rats

Aims: Isomalto‐oligosaccharides (IMO) with α(1→6) and α(1→4) glucosidic linkages are produced by enzymatic conversion of starch. IMO are only partially digestible but data on their influence on intestinal microbiota are limited. It was the aim of this study to investigate the effect of IMO diet on intestinal microbiota and short‐chain fatty acids production (SCFA) in rats. Methods and results: Three groups of F344 rats, each consisting of six animals, were fed IMO, inulin or a control diets for six weeks. A qualitative assessment of the intestinal microbiota was achieved by PCR‐denaturing gradient gel electrophoresis (DGGE). Major bacterial taxa were quantified by quantitative PCR (qPCR), and SCFA were measured using gas chromatography. Quantitative PCR demonstrated that lactobacilli were one of the dominant bacterial taxa in faecal samples from rats. IMO increased the number of lactobacilli and the total number of intestinal bacteria in rats fed IMO compared with animals receiving control and inulin diets. Furthermore, PCR‐DGGE with lactobacilli‐specific primers showed an altered biodiversity of lactobacilli in rats fed IMO compared with control diet. Conclusions: IMO selectively stimulates lactobacilli and increases their diversity in rats. Significance and impact of study: Isomalto‐oligosaccharides specifically stimulate growth of intestinal lactobacilli in a rat model system.     

Co-immobilization of dextransucrase and dextranase for the facilitated synthesis of isomalto-oligosaccharides: Preparation, characterization and modeling

Co-Immobilization of dextransucrase (DS) and dextranase (DN) into calcium alginate includes the co-entrapment of soluble DS and adsorbed DN. DS converts sucrose into dextran, which is the substrate for DN, so that isomalto-oligosaccharides (IMOs) are follow-up products of dextran hydrolysis. The boundary conditions for the successful preparation are investigated with respect to choice of DN adsorbate, surface modifications using blotting agents and optimal enzyme activity ratios. Further, repetitive batch experiments suggest the selection of medium activity ratios for continuous use (0.3 U(DN)U(-1) (DS), e.g.). Product formation at various cosubstrate:substrate concentrations as well as at different DN:DS ratios are discussed. Moreover, the complexity of the bi-enzymatic system can be reduced considering the molar ratios of cosubstrate:substrate (glucose:sucrose). Based on these factors, a mechanistic kinetic model is developed, which distinguishes the corresponding contributions of the two enzymes upon overall product formation. In general, at low glucose:sucrose ratios isomaltose synthesis is featured primarily by DN action. Yet with increasing amounts of glucose both the quantity and quality of DN substrate changes, so that its contribution to product formation decreases in an exponential manner; still the overall product yield continuously increases due to enhanced DS contribution.     

Very high gravity wort fermentation by immobilised yeast

Using calcium alginate-entrapped yeast, 24% (w/w) wort was successfully fermented within 8 days. This is half the time needed for fermentation by free yeast. The highest ethanol concentration obtained was 10.5% (v/v). When the original wort gravity was increased, the specific rate of ethanol production remained constant 0.16 g gh^–1 and the viability did not fall bellow 95% of living cells. Protection of cell against osmotic stress by gel matrix was also confirmed by trehalose measurement. The maximum intracellular trehalose content in calcium alginate-entrapped yeast was 3 times lower compared to free yeast at 30% (w/w) wort fermentation.     

土霉素菌渣源酵母菌的筛选及其培养条件的优化

【背景】随着制药行业的蓬勃发展,中国每年产生大量的抗生素菌渣,已造成了不可忽视的环境污染和资源浪费等问题。由于抗生素菌渣中含有丰富的蛋白质等有机物质,利用其蛋白质等进行二次生产或将成为解决抗生素菌渣处理问题的一种有效方法。【目的】从土霉素菌渣中筛选出天然酵母菌菌株,并利用土霉素菌渣作为酵母菌生长的主要培养基成分,通过其培养条件的优化,实现土霉素菌渣的资源化利用。【方法】以土霉素菌渣为样品,运用稀释涂布法进行酵母菌的筛选,通过其形态学观察和18S rRNA基因序列分析等方法对菌株进行鉴定。通过单因素试验与Box-Behnken Design试验结合,对培养条件进行优化,确定筛选菌株在以土霉素菌渣为主要成分培养基中的最佳生长条件。【结果】筛选的土霉素菌渣源酵母菌为一株希腊接合囊酵母菌(Zygoascus hellenicus)Y1,其最佳培养条件为:土霉素菌渣添加量为5%,葡萄糖添加量为0.5%,接菌量为2%,在pH 5.0、32℃、160r/min条件下培养24h。【结论】筛选到一株希腊接合囊酵母菌Y1,该菌能够很好地利用土霉素菌渣进行生长,实现了土霉素菌渣的资源化利用,大大减少了菌渣的...     

Batch ethanol fermentation of molasses: a correlation between the time necessary to complete the fermentation and the initial concentrations of sugar and yeast cells

Batch fermentations of sugar-cane blackstrap molasses to ethanol, using pressed yeast as inoculum, demonstrated an exponential relationship between the time necessary to complete the fermentation and the initial concentrations of sugar and yeast cells. The parameters of the derived exponential equations depended on the experimental conditions.     

Xylose and cellobiose fermentation to ethanol by the thermotolerant methylotrophic yeast Hansenula polymorpha

Wild-type strains of the thermotolerant methylotrophic yeast Hansenula polymorpha are able to ferment glucose, cellobiose and xylose to ethanol. H. polymorpha most actively fermented sugars to ethanol at 37 degrees C, whereas the well-known xylose-fermenting yeast Pichia stipitis could not effectively ferment carbon substrates at this temperature. H. polymorpha even could ferment both glucose and xylose up to 45 degrees C. This species appeared to be more ethanol tolerant than P. stipitis but more susceptible than Saccharomyces cerevisiae. A riboflavin-deficient mutant of H. polymorpha increased its ethanol productivity from glucose and xylose under suboptimal supply with riboflavin. Mutants of H. polymorpha defective in alcohol dehydrogenase activity produced lower amounts of ethanol from glucose, whereas levels of ethanol production from xylose were identical for the wild-type strain and the alcohol dehydrogenase-defective mutant.     

Discrepancy in glucose and fructose utilisation during fermentation by Saccharomyces cerevisiae wine yeast strains

While unfermented grape must contains approximately equal amounts of the two hexoses glucose and fructose, wine producers worldwide often have to contend with high residual fructose levels (>2 gl(-1)) that may account for undesirable sweetness in finished dry wine. Here, we investigate the fermentation kinetics of glucose and fructose and the influence of certain environmental parameters on hexose utilisation by wine yeast. Seventeen Saccharomyces cerevisiae strains, including commercial wine yeast strains, were evaluated in laboratory-scale wine fermentations using natural Colombard grape must that contained similar amounts of glucose and fructose (approximately 110 gl(-1) each). All strains showed preference for glucose, but to varying degrees. The discrepancy between glucose and fructose utilisation increased during the course of fermentation in a strain-dependent manner. We ranked the S. cerevisiae strains according to their rate of increase in GF discrepancy and we showed that this rate of increase is not correlated with the fermentation capacity of the strains. We also investigated the effect of ethanol and nitrogen addition on hexose utilisation during wine fermentation in both natural and synthetic grape must. Addition of ethanol had a stronger inhibitory effect on fructose than on glucose utilisation. Supplementation of must with assimilable nitrogen stimulated fructose utilisation more than glucose utilisation. These results show that the discrepancy between glucose and fructose utilisation during fermentation is not a fixed parameter but is dependent on the inherent properties of the yeast strain and on the external conditions.     

Production of ethanol from guava pulp by yeast strains

Guava pulp used for ethanol production by three yeast strains contained 10% (w/v) total sugars and was pH 4.1. Ethanol production at the optimum sugar concentration of 10%, at pH 4.1 and 30°C was 1.5%, 3.6% and 3.9% (w/v) by Saccharomyces cerevisiae MTCC 1972, Isolate-1 and Isolate-2, respectively, at 60 h fermentation. Higher sugar concentrations at 15 and 20% were inhibitory for ethanol production by all test cultures. The maximum production of ethanol at optimum natural sugar concentration (10%) of guava pulp, was 5.8% (w/v) at pH 5.0 by Isolate-2 over 36 h fermentation, which was only slightly more than the quantity of ethanol produced by Saccharomyces cerevisiae (5.0%) and Isolate-1 (5.3%) over 36 and 60h fermentation, respectively.     

Treatment of thin stillage by Aspergillus oryzae and its effect on alcoholic fermentation in syrup liquid

The microbial degradation of thin stillage for environment-friendly treatment has been studied extensively in recent years, and useful compounds in the treated-thin stillage are expected to be utilized in the subsequent fermentation. In this study, an Aspergillus oryzae H18, suitable for growing in thin stillage, was isolated from soil and served to degrade the organic matter in thin stillage, with the increase in pH (from 3·75 to 4·8) and decrease in chemical oxygen demand (COD, 81·3% removal rate). The effect of thin stillage as backset water after degradation of the strain H18 on alcohol production in syrup liquid was investigated. Compared with zero addition of thin stillage, the alcohol yield in mixed syrup liquid increased by 8·6% when the concentration of treated-thin stillage was 20%. After the addition of nutrients at proper concentration (0·5% urea, 1% molasses, 0·25% NaCl, 0·2% NaH₂PO₄, 0·3% MgSO₄ and 0·25% CaCl₂) in thin stillage, the alcohol yield in yeast fermentation was increased by 32·7% when mixed syrup liquid (with 40% thin stillage treated by H18) was employed, in comparison to control group without thin stillage addition. Meanwhile, the fermentation time was shortened, and alcohol production rate was enhanced.     

Lumping kinetics of ABE fermentation wastewater treatment by oleaginous yeast Trichosporon cutaneum

Lumping kinetics models were built for the biological treatment of acetone–butanol–ethanol (ABE) fermentation wastewater by oleaginous yeast Trichosporon cutaneum with different fermentation temperatures. Compared with high temperature (33°C, 306?K) and low temperature (23°C, 296?K), medium temperature (28°C, 301?K) was beneficial for the cell growth and chemical oxygen demand (COD) degradation during the early stage of fermentation but the final yeast biomass and COD removal were influenced little. By lumping method, the materials in the bioconversion network were divided into five lumps (COD, lipid, polysaccharide, other intracellular products, other extracellular products), and the nine rate constants (k₁–k₉) for the models can well explain the bioconversion laws. The Gibbs free energy (G) for this bioconversion was positive, showing that it cannot happen spontaneous, but the existence of yeast can after the chemical equilibrium and make the bioconversion to be possible. Overall, the possibility of using lumping kinetics for elucidating the laws of materials conversion in the biological treatment of ABE fermentation wastewater by T. cutaneum has been initially proved and this method has great potential for further application.     

Influence of alginate properties and gel reinforcement on fermentation characteristics of immobilized yeast cells

The effect of alginate composition, gel concentration, gelation method, cell loading and surface area on fermentation characteristics of immobilized yeast cells have been investigated. Molecular weight and G/M ratio had only little effect on fermentation velocity and gel strength, while increasing the alginate concentration caused a sizeable decrease in fermentation velocity and an increase in gel strength. The internally gelled immobilizates generally showed a higher fermentation velocity for the same gel strength and no decrease in gel strength was seen during fermentation. With high initial cell loadings, the fermentation velocity per g of immobilizate was higher, but the productivity per cell was lower than with low initial cell loadings. The difference decreased with time. Specific surface area (surface/volume) was shown to be an important factor for the observed productivity per gram of immobilizate, with high S/V ratios giving the highest productivity. Gel shape had no influence on fermentation velocity for a given S/V ratio. Gelation behaviour of externally gelled beads was determined by estimating the amount of cells liberated during gel formation through measurement of invertase activity (yeast-bound) in the gelling solution. A method for reinforcement of internally gelled alginate slabs with a nylon mesh was developed and utilized for production of a continuous fermentation reactor with reinforced gels.     

Effect of carbon dioxide on yeast growth and fermentation

Inhibition of yeast function by ethanol and by high substrate concentrations is well recognized and, to a limited extent, quantified. The role of carbon dioxide in affecting yeast metabolism (particularly growth processes) is not clear although inhibition is generally found at moderate to high concentrations of the dissolved gas. A similar situation exists with other microorganisms and with other fermentation systems. An understanding of the role of carbon dioxide, and particularly of its inhibitory effects on enzyme action and membrane function, is required if the observed global inhibition of yeasts and other fermentation systems is to be partitioned to its appropriate causes.     

Effect of oils and fatty acids on molasses fermentation by distillers' yeast

Some vegetable oils, and the mixed fatty acids derived from them, stimulate sugar utilization and both the rate of alcohol production and yield of alcohol during molasses fermentation by yeast. The effect is particularly prominent during fermentation at a higher temperature of 40°C.     

Genetic and fermentation properties of the K2 killer yeast, Saccharomyces cerevisiae T206

Saccharomyces cerevisiae T206 K⁺R⁺, a K₂ killer yeast, was differentiated from other NCYC killer strains of S. cerevisiae on the basis of CHEF-karyotyping and mycoviral RNA separations. Genomic DNA of strain T206 was resolved into 13 chromosome bands, ranging from approximately 0.2 to 2.2 Mb. The resident virus in strain T206 yielded L and M RNA species of approximately 5.1 kb and 2.0 kb, respectively. In micro-scale vinifications, strain T206 showed a lethal effect on a K^-R^- mesophilic wine yeast. Metabolite accumulation and toxin activity were measured over a narrow pH range of 3.2 to 3.5. Contrary to known fermentation trends, the challenged fermentations were neither stuck nor protracted although over 70% of the cell population was killed. Toxin-sensitive cells showed cytosolic efflux.     

Xylitol conversion by fermentation using five yeast strains and polyelectrolyte-assisted ultrafiltration

Batch fermentations for xylitol production were conducted using Candida boidinii (BCRC 21432), C. guilliermondii (BCRC 21549), C. tropicalis (BCRC 20520), C. utilis (BCRC 20334), and P. anomala (BCRC 21359) together with a mixture of sugars simulating lignocellulosic hydrolysates as the carbon source. C. tropicalis had the highest bioconversion yield (Y_P/S) of 0.79 g g⁻¹ (g xylitol·g xylose⁻¹) over 48 h. Additional fermentations with C. tropicalis achieved Y_P/S values of 0.6 and 0.39 g g⁻¹ after 96 and 72 h using urea and soybean meal as the nitrogen sources, respectively. Ethanol and arabitol were also produced in all fermentation. Xylitol in the fermentation broth was recovered by cross-flow ultrafiltration. With prior application of 2 mg polydiallyl dimethylammonium chloride l⁻¹ on the membrane surface, protein in the permeate was reduced from 7.1 to 1.5 mg l⁻¹ after 2 h.     

废糟液全循环条件下絮凝酵母乙醇连续发酵

丙酸是以玉米为原料自絮凝酵母乙醇连续发酵系统废糟液全循环过程中积累的主要抑制物。基于丙酸对酵母细胞抑制机理,开发了3种废糟液全循环条件下乙醇连续发酵工艺策略。首先根据高温导致丙酸生成的现象,去除了物料灭菌环节,使发酵液丙酸浓度显著降低,生物量和乙醇浓度分别提高了59.3%和7.4%。其次,以丙酸浓度达到半数抑制浓度(IC50)40 mmol/L为目标,通过拟合丙酸积累数据预测废糟液全循环的最长运行时间,发酵装置运行应控制在此时间范围内。再次,较低的环境pH值提高了丙酸毒性,而实验证明发酵液pH为5.5时,丙酸对细胞生长的抑制影响最小,因此控制发酵过程中的pH有利于弱化丙酸毒性。     

Surface display of transglucosidase on Escherichia coli by using the ice nucleation protein of Xanthomonas campestris and its application in glucosylation of hydroquinone

A surface anchoring motif using the ice nucleation protein (INP) of Xanthomonas campestris pv. campestris BCRC 12,846 for display of transglucosidase has been developed. The transglucosidase gene from Xanthomonas campestris pv. campestris BCRC 12,608 was fused to the truncated ina gene. This truncated INP consisting of N- and C-terminal domains (INPNC) was able to direct the expressed transglucosidase fusion protein to the cell surface of E. coli with apparent high enzymatic activity. The localization of the truncated INPNC-transglucosidase fusion protein was examined by Western blot analysis and immunofluorescence labeling, and by whole-cell enzyme activity in the glucosylation of hydroquinone. The glucosylation reaction was carried out at 40 degrees C for 1 h, which gave 23 g/L of alpha-arbutin, and the molar conversion based on the amount of hydroquinone reached 83%. The use of whole-cells of the wild type strain resulted in an alpha-arbutin concentration of 4 g/L and a molar conversion of 16% only under the same conditions. The results suggested that E. coli displaying transglucosidase using truncated INPNC as an anchoring motif can be employed as a whole-cell biocatalyst in glucosylation.     

自絮凝颗粒酵母乙醇连续发酵耦合废液全循环工艺的研究

在以CO2为驱动力的单级悬浮床生物反应器中,进行了自絮凝颗粒酵母乙醇连续发酵耦合废液全循环实验研究。以双酶法制备的玉米粉糖化液为底物,系统连续运行了28 d,每隔5 d将收集到的发酵液集中精馏处理,得到的废糟液直接用于玉米粉调浆制糖。实验数据表明,在稀释率为0.05 h-1条件下,发酵液中乙醇、残还原糖、残总糖质量浓度基本稳定,其平均值为82.97,30.02和40.87 g.L-1。对废液循环工艺过程中,发酵液中的8种高沸点有机酸进行了定量分析,发现发酵液中仅丙酮酸有明显积累,并最终在0.1~0.3 mol.L-1范围内波动。实验结果表明:系统中存在的高沸点副产物不对乙醇发酵产生明显抑制作用,乙醇连续发酵能够在废液全循环条件下良好运行。     

高生物量富铁酵母菌的选育及其发酵条件的研究

对402株不同种属的酵母菌株进行初筛、复筛,筛选到一株生物量较高的二倍体菌株ZY-46(Saccharomyces cerevisiae)和一株铁富集量较高的二倍体菌株ZY-173(Saccharomyces kluyveri)。然后以它们为出发菌,分别进行单倍体分离、硫酸二乙酯(DES)诱变,并通过原生质体融合,得到一株高生物量富铁酵母融合菌株ZYF-15。在优化的发酵条件下,该融合菌株生物量可达11．2g／L,细胞铁含量达24．5mg／g干细胞,细胞总铁含量分别比原始亲株ZY-46和ZY-173提高了2．6倍和1．9倍。