Acetic acid-and furfural-based adaptive evolution of Saccharomyces cerevisiae strains for improving stress tolerance and lignocellulosic ethanol production

Acetic acid and furfural are known as prevalent inhibitors deriving from pretreatment during lignocellulosic ethanol production.They negatively impact cell growth,glucose uptake and ethanol biosynthesis of Saccharomyces cerevisiae strains.Development of industrial S.cerevisiae strains with high tolerance towards these inhibitors is thus critical for efficient lignocellulosic ethanol production.In this study,the acetic acid or furfural tolerance of different S.cerevisiae strains could be significantly enhanced after adaptive evolution via serial cultivation for 40 generations under stress conditions.The acetic acid-based adaptive strain SPSC01-TA9 produced 30.5 g·L^(-1)ethanol with a yield of 0.46 g·g^(-1)in the presence of 9 g·L^(-1)acetic acid,while the acetic acid/furfural-based adaptive strain SPSC01-TAF94 produced more ethanol of 36.2 g·L^(-1)with increased yield up to 0.49 g·g^(-1)in the presence of both 9 g·L^(-1)acetic acid and 4 g·L^(-1)furfural.Significant improvements were also observed during non-detoxified corn stover hydrolysate culture by SPSC01-TAF94,which achieved ethanol production and yield of 29.1 g·L^(-1)and 0.49 g·g^(-1),respectively,the growth and fermentation efficiency of acetic acid/furfural-based adaptive strain in hydrolysate was 95%higher than those of wildtype strains,indicating the acetic acid-and furfural-based adaptive evolution strategy could be an effective approach for improving lignocellulosic ethanol production.The adapted strains developed in this study with enhanced tolerance against acetic acid and furfural could be potentially contribute to economically feasible and sustainable lignocellulosic biorefinery.

Postbiotics from Saccharomyces cerevisiae fermentation stabilize microbiota in rumen liquid digesta during grain-based subacute ruminal acidosis(SARA) in lactating dairy cows

Background Subacute ruminal acidosis(SARA)is a common metabolic disorder of high yielding dairy cows,and it is associated with dysbiosis of the rumen and gut microbiome and host inflammation.This study evaluated the impact of two postbiotics from Saccharomyces cerevisiae fermentation products(SCFP)on rumen liquid associated microbiota of lactating dairy cows subjected to repeated grain-based SARA challenges.A total of 32 rumen cannulated cows were randomly assigned to 4 treatments from 4 weeks before until 12 weeks after parturition.Treatment groups included a Control diet or diets supplemented with postbiotics(SCFPa,14 g/d Original XPC;SCFPb-1X,19 g/d Nutri Tek;SCFPb-2X,38 g/d Nutri Tek,Diamond V,Cedar Rapids,IA,USA).Grain-based SARA challenges were conducted during week 5(SARA1)and week 8(SARA2)after parturition by replacing 20%DM of the base total mixed ration(TMR)with pellets containing 50%ground barley and 50%ground wheat.Total DNA from rumen liquid samples was subjected to V3–V416S r RNA gene amplicon sequencing.Characteristics of rumen microbiota were compared among treatments and SARA stages.Results Both SARA challenges reduced the diversity and richness of rumen liquid microbiota,altered the overall composition(β-diversity),and its predicted functionality including carbohydrates and amino acids metabolic pathways.The SARA challenges also reduced the number of significant associations among different taxa,number of hub taxa and their composition in the microbial co-occurrence networks.Supplementation with SCFP postbiotics,in particular SCFPb-2X,enhanced the robustness of the rumen microbiota.The SCFP supplemented cows had less fluctuation in relative abundances of community members when exposed to SARA challenges.The SCFP supplementation promoted the populations of lactate utilizing and fibrolytic bacteria,including members of Ruminococcaceae and Lachnospiraceae,and also increased the numbers of hub taxa during non-SARA and SARA stages.Supplementation with SCFPb-2X prevented the fluctuations in the abundances of hub taxa that were positively correlated with the acetate concentration,andα-andβ-diversity metrics in rumen liquid digesta.Conclusions Induction of SARA challenges reduced microbiota richness and diversity and caused fluctuations in major bacterial phyla in rumen liquid microbiota in lactating dairy cows.Supplementation of SCFP postbiotics could attenuate adverse effects of SARA on rumen liquid microbiota.

基于Lachancea thermotolerans-Saccharomyces cerevisiae双酵母混合发酵酸啤酒的工艺研究

酸啤酒正在被越来越多的消费者接受,也正在成为啤酒企业多元化布局的方向之一。该研究在不添加外源风味物质的前提下,基于啤酒双酵母混合发酵体系,将耐热拉钱斯氏酵母(Lachancea thermotolerans)与酿酒酵母(Saccharomyces cerevisiae)组合按照10∶1同时接种进行啤酒发酵,耐热拉钱斯氏酵母的发酵过程可产生一定量的乳酸(1.5~1.8 g/L),降低酒液pH值(3.3~3.4),得到酸味柔和的新型酸啤酒。相比于单一菌株发酵,双酵母混合发酵酸啤酒体系的风味物质产量都有所提高,产生了更多乙酸异丁酯和乙酸异戊酯等酯类物质,增强了酸啤酒的水果香气,果香馥郁、爽口协调,对啤酒的质量产生了积极影响,并且减少了糖化锅酸化等工艺操作步骤,实现更快速的发酵和稳定的过程控制,促进了酸啤酒的大规模生产应用。

Saccharomyces cerevisiae prevents postoperative recurrence of Crohn's disease modeled by ileocecal resection in HLA-B27 transgenic rats

BACKGROUND Postoperative recurrence(POR)after ileocecal resection(ICR)affects most Crohn's disease patients within 3-5 years after surgery.Adherent-invasive Escherichia coli(AIEC)typified by the LF82 strain are pathobionts that are frequently detected in POR of Crohn's disease and have a potential role in the early stages of the disease pathogenesis.Saccharomyces cerevisiae CNCM I-3856 is a probiotic yeast reported to inhibit AIEC adhesion to intestinal epithelial cells and to favor their elimination from the gut.AIM To evaluate the efficacy of CNCM I-3856 in preventing POR induced by LF82 in an HLA-B27 transgenic(TgB27)rat model.METHODS Sixty-four rats[strain F344,38 TgB27,26 control non-Tg(nTg)]underwent an ICR at the 12th wk(W12)of life and were sacrificed at the 18th wk(W18)of life.TgB27 rats were challenged daily with oral administration of LF82(109 colony forming units(CFUs)/day(d),n=8),PBS(n=5),CNCM I-3856(109 CFUs/d,n=7)or a combination of LF82 and CNCM I-3856(n=18).nTg rats receiving LF82(n=5),PBS(n=5),CNCM I-3856(n=7)or CNCM I-3856 and LF82(n=9)under the same conditions were used as controls.POR was analyzed using macroscopic(from 0 to 4)and histologic(from 0 to 6)scores.Luminal LF82 quantifications were performed weekly for each animal.Adherent LF82 and inflammatory/regulatory cytokines were quantified in biopsies at W12 and W18.Data are expressed as the median with the interquartile range.RESULTS nTg animals did not develop POR.A total of 7/8(87%)of the TgB27 rats receiving LF82 alone had POR(macroscopic score≥2),which was significantly prevented by CNCM I-3856 administration[6/18(33%)TgB27 rats,P=0.01].Macroscopic lesions were located 2 cm above the anastomosis in the TgB27 rats receiving LF82 alone and consisted of ulcerations with a score of 3.5(2-4).Seven out of 18 TgB27 rats(39%)receiving CNCM I-3856 and LF82 had no macroscopic lesions.Compared to untreated TgB27 animals receiving LF82 alone,coadministration of CNCM I-3856 and LF82 significantly reduced the macroscopic[3.5(2-4)vs 1(0-3),P=0.002]and histological lesions by more than 50%[4.5(3.3-5.8)vs 2(1.3-3),P=0.003].The levels of adherent LF82 were correlated with anastomotic macroscopic scores in TgB27 rats(r=0.49,P=0.006),with a higher risk of POR in animals having high levels of luminal LF82(71.4%vs 25%,P=0.02).Administration of CNCM I-3856 significantly reduced the levels of luminal and adherent LF82,increased the production of interleukin(IL)-10 and decreased the production of IL-23 and IL-17 in TgB27 rats.CONCLUSION In a reliable model of POR induced by LF82 in TgB27 rats,CNCM I-3856 prevents macroscopic POR by decreasing LF82 infection and gut inflammation.

Inhibition effect of expression of Cu/Zn superoxide dismutase from rice on synthesis of Glutathione in Saccharomyces cerevisiae

The expression of a rice Cu/Zn superoxide dismutase （Cu/Zn-SOD） in Saccharomyces cerevisiae regulated by GAPDH promoter, involved in the inhibition of endogenous Glutathione （GSH） synthesis, and the competitive expression was detected by constructing the expression vector transferred Cu/Zn-SOD gene into wild-type S. cerevisiae. Transcription and expression of the Cu/Zn-SOD gene in S. cerevisiawere were confirmed by northern blot and SDS-PAGE, respectively, and activity of the Cu/Zn-SOD from crude extracts was enzymatically detected based on the effect of nitroblue tetrazolium （NBT） after running a native polyacrylamide gel. The GSH synthesis was also tested by DTNB （5, 5＇-Dithiobis （2-nitrobenzoic acid）） method. Results showed that GSH synthesis was evidently suppressed by the expression of Cu/Zn-SOD gene in both control and heat shock strains. It implied that the expression of the Cu/Zn-SOD gene in S. cerevisiae has more potential facility in response to oxidative exposure than that of endogenous GSH, although Cu/Zn-SOD and GSH were both contributed to the function of oxygen radical oxidoreduction.

Reducing Yield of Fusel Alcohols by Saccharomyces cerevisiae of Compound Mutagenesis Through UV-MPMS Method

Excessive fusel alcohol contents will cause the beer to produce off-flavors and cause dizziness and headaches.Reducing the contents of fusel alcohols in beer is very important to people's health.The excessive fusel alcohol contents in beer is a common problem in the industry.How to control the contents of fusel alcohols in a reasonable range is of great significance for improving beer quality.After one round of ultraviolet(UV)and one round of multifunctional plasma mutagenesis system(MPMS)mutagenesis,the yeast strains with lower fusel oil yield and more stablility could be screened.According to the relationship between the fusel alcohol Harris metabolic pathway of brewer's yeast and lactic acid metabolism,excellent strains were obtained by triple screening with lactic acid medium,calcium carbonate medium and 2,3,5-triphenyl tetrazolium chloride upper medium.The content of fusel alcohol in the finished beer fermentation test of screened strain Z43 was 52.1±0.142 mg•L^(-1),which was 43%lower than that of the starting strain,and other fermentation properties remained unchanged.After eight passages,it was verified that the strain was stable and heritable.These results showed that strain Z43 presented promising characteristics for use in the production of beer with a potentially low contents of fusel alcohols.

啤酒酵母（Saccharomyces cerevisiae）M－05变株合成谷胱甘肽的研究Ⅰ．菌种的选育

从１２种５５株酵母菌株中筛选出１株相对富含谷胱甘肽（ＧＳＨ）的菌株──啤酒酵母Ｓ─１２，经紫外线及硫酸二乙酯等复合诱变处理，以甲硫氨酸缺陷型（Ｍｅｔ－）为筛选模型，获得１株高产谷胱甘肽变株Ｍ—０５，其干细胞每ｇ含有ＧＳＨ１４．４３ｍｇ，较出发菌株提高了６８．４％．

伴有辅酶再生过程的Saccharomyces cerevisiae B5不对称还原2’-氯-苯乙酮的催化反应动力学

对以5%(体积比)乙醇为辅助底物的伴有辅酶再生过程的SaccharomycescerevisiaeB5不对称还原2’-氯-苯乙酮制备手性药物中间体R-2’-氯-1-苯乙醇的生物催化反应建立了描述底物消耗和产物合成的动力学模型。考察了反应过程中辅酶的种类和含量,以及底物和产物随时间的变化量。研究表明,参加反应的还原剂是辅酶Ⅰ。当底物初始浓度≤8.09mmol?L?1,可不考虑底物对微生物的毒害作用,反应可看作两个均符合顺序反应机制的氧化还原反应的耦联。通过实验数据对动力学方程式的拟合,得到动力学参数:Vm1=8.0×10?4mol?L?1?h?1,KmB1=9.0×10-4mol?L?1,KiA1=2.0×10-6mol?L?1。动力学模型模拟计算结果与实验值能较好地吻合。

Screening of Saccharomyces Strains Highly Producing Glutathione and Breeding of Its Ethionine-resistant Mutants

[Objective] The aim of this study was to screen Saccharomyces for glutathione over-production. [Method] Ethionine-resistant mutants were obtained through UV mutagenesis and rational screening. [Result] A high GSH-producing strain HSJB1 was isolated from soil, and the biomass for this strain by flask shaking fermentation was 3.87 g/L while the GSH yield was 91.87 mg/L. According to the morphological, physiological and biochemical characteristics of cells, this strain was primarily identified as Saccharomyces cerevisiae. An ethionine-resistant mutant YBS77 was obtained through UV mutagenesis of the original strain HSJB1, and the biomass for this strain by flask shaking fermentation was 7.60 g dry cell weight/L while the GSH yield was 211.96 mg/L. [Conclusion] The biomass of the mutant obtained by breeding is increased by 96.38% than that of the original strain, and the GSH yield of the mutant obtained by breeding is increased by 130.72% than that from the original strain, which indicates that the breeding method is feasible.

酵母(Saccharomyces cerevisiae)及酵母提取物对肉鸡肉质的影响(英文)

本文旨在研究酵母(Saccharomyces cerevisiae)及其提取物对肉鸡肉质的影响。试验采用单因子随机分组,选取1日龄雄性肉鸡180只,随机分为3组,每组6个重复,每个重复10只。分别在基础饲粮中添加0.5%酵母和0.3%酵母提取物作为试验组饲粮。试验结果表明,饲粮添加酵母提取物可显著增加肉鸡小腿肉的最终pH(P<0.05),同时饲粮添加酵母有增加肉中水分含量及降低水煮失重率的趋势,但未达到统计学显著水平(P>0.05)。饲粮添加酵母及酵母提取物与对照组相比均可显著降低生肉和熟肉的剪切力(P<0.05)。饲粮添加酵母与对照组相比可显著降低TBARS值(P<0.05)。由此可知,饲粮添加酵母及酵母提取物可改善肉鸡肉质和嫩度,同时酵母及酵母提取物均具有降低氧化的作用。

酿酒酵母Saccharomyces cerevisiae重组菌株木糖醇发酵的初步研究

木糖还原酶催化木糖为木糖醇的反应 ,是木糖代谢的第一步。将木糖还原酶的基因XYL1引入酿酒酵母中 ,构建得到高效表达XYL1基因的重组酿酒酵母菌株HYEX2 ,该重组菌株的木糖还原酶比活力为 7.47U/mg。研究表明 ,该菌株获得转化木糖产生木糖醇的能力 ,当辅助碳源葡萄糖的浓度为 2 % ,并在发酵 30h左右添加木糖 ,木糖醇的转化率可达到 0 .94g/ g。

酒精酵母Saccharomyces cerevisiae 4-S的高密度培养及其酒精发酵性能

实验比较了分批培养、分批补料及连续流加培养对酒精酵母细胞生长的影响。结果显示,分批补料流加或连续流加(流加速率0.133 g.L-1.m-1)有利于酒精酵母菌Saccharomyces cerevisiae4-S的快速生长和得到较高的细胞浓度,分批补料流加或连续流加培养44 h,发酵液中酒精酵母细胞浓度分别达到71.82(细胞干重-DCW)g.L-1和82.01(DCW)g.L-1,比分批培养分别提高了155.68%和191.95%。同时,对高密度酒精酵母的酒精发酵性能和循环利用研究结果显示,在细胞浓度为82.01(DCW)g.L-1,糖浓度为350 g.L-1,发酵36h发酵液中乙醇浓度达到16.23%,比普通酵母浓度28.09(DCW)g.L-1批发酵时间缩短12 h,乙醇浓度提高了24.4%,酵母的有效重复利用次数达4批次。

Saccharomyces cerevisiae对浓香型白酒发酵的影响

为了探究酿酒酵母对浓香型白酒发酵的影响,在混菌固态发酵糟醅中强化接种酿酒酵母,发酵50 d后分析糟醅主要理化指标、风味物质含量及酿造微生物区系构成,发现糟醅中浓香型白酒主要风味物质含量均有所下降,特别是酯类物质生成量减少,乙酸乙酯显著下降,下降幅度达71.0%;同时,糟醅中细菌和真菌多样性降低,优势细菌仍为Lactobacillus和Pseudomonas,而真菌区系中,曲霉属的丰度从1.8%上升到60.4%,取代接合酵母成为新的优势真菌。结果表明,本实验条件下酿酒酵母对浓香型白酒发酵有一定不利影响。

利用Saccharomyces cerevisiae KD控制苹果汁中棒曲霉素的污染

研究一株食品生产用酿酒酵母Saccharomyces cerevisiae KD在培养基及市售100%苹果汁中对棒曲霉素污染的控制作用。通过高效液相色谱法对棒曲霉素进行定量,分析起始棒曲霉素浓度、菌体接种量和培养基p H对S.cerevisiae KD去除棒曲霉素活力的影响;利用酶标仪监测S.cerevisiae KD的生长状况,且通过检测可溶性固形物、酸度、总酚、黄酮含量对菌体发酵后苹果汁的品质进行了评估。结果表明:有氧条件下S.cerevisiae KD能够在28 h内完全去除培养基中的棒曲霉素,其去除机理包括物理吸附和酶解;在较低的起始棒曲霉素浓度和较高的菌体接种量条件下,S.cerevisiae KD对棒曲霉素的去除率较高,但在培养后期,不同菌体接种量下棒曲霉素的去除率接近一致;实验还发现酸性条件有利于S.cerevisiae KD去除棒曲霉素。此外,S.cerevisiae KD对棒曲霉素的耐受性较强,甚至在棒曲霉素浓度高达100 mg/L的环境中依然能较好生长。在市售100%苹果汁中,S.cerevisiae KD也能高效控制棒曲霉素的污染,且与Lactococcus lactis MG1363联合发酵2 d后,果汁中已无棒曲霉素检出,总酚含量显著高于发酵前苹果汁(p<0.05),发酵果汁的品质较好。结论:S.cerevisiae KD可有效控制食品中棒曲霉素的污染,具有潜在的应用前景。

响应面法优化Saccharomyces cerevisiae FL-1培养基

以筛选得到的Saccharomycescerevisiae(面包酵母)FL-1菌株作为研究对象,考察了培养基中碳、氮源和酵母浸出物对该酵母生物量的影响,通过响应面法建立了生物量和蔗糖、硫酸铵及酵母浸出物浓度之间的关系,实现培养基的优化。摇瓶发酵结果表明,由蔗糖、硫酸铵和酵母浸出物为主要成分的培养基能满足S.cerevisiaeFL-1生长繁殖对营养物质的需求,拟合得到的模型较好地符合实际,在考察范围内,酵母浸出物对生物量收率影响程度最高,蔗糖次之,硫酸铵最弱。最后,次氯酸钠氧化S.cerevisiaeFL-1的自溶体可制备目标产物(1→3)-β-D-葡聚糖。

去氢木香内酯特异性抑制Saccharomyces cerevisiae细胞壁中β-葡聚糖的合成

基于酿酒酵母(Saccharomyces cerevisiae)及其细胞壁合成酶缺陷突变株组成的靶标定向全细胞筛选模型,对去氢木香内酯抗真菌的作用机制进行研究,发现去氢木香内酯特异性抑制β-葡聚糖合酶基因缺陷突变株Δfks1,其最低抑制浓度MIC值为16μg/mL,比野生型WHU2a和几丁质合酶基因缺陷突变株均低2倍;其对WHU2a生长的抑制能被山梨醇等渗溶液部分挽救;IC20浓度的去氢木香内酯处理后WHU2a的β-葡聚糖含量降低了16.83%,512μg/mL去氢木香内酯抑制了26.66%的β-葡聚糖合酶活性。研究结果表明,去氢木香内酯能特异性抑制真菌细胞壁中葡聚糖的合成,从而抑制真菌细胞生长。

Debaryomyces vanriji和Saccharomyces cerevisiae的混合培养及其对葡萄酒风味的影响

葡萄酒的风味是衡量葡萄酒品质的一个重要指标。研究了Debaryomycesvanriji和Saccharomycescerevisiae(酿酒酵母)混合培养的生长情况及其对葡萄酒风味的影响。结果表明,混合培养得到的β-葡萄糖苷酶的活性更高,发酵后的葡萄酒中脂肪酸、酯、萜烯醇等的含量更丰富,从而增强或改善了葡萄酒的风味。

吩噻嗪类钙调素抑制剂对芽殖酵母(Saccharomyces cerevisiae)和粟酒裂殖酵母(Schizosaccharomyces pombe)细胞增殖的不同效应

低浓度的TFP、CPZ及FPZ能明显促进S.pombe细胞的增殖,培养液中Ca^(2+)浓度越低,TFP等的促进作用越显著,Ca^(2+)与TFP具有协同促进S.pombe增殖的功能;但20μmol/L TFP能终止S.cerevisiae细胞周期的运转。TFP的细胞膜通透性研究表明,20μmol/L TFP就能进入到S.cerevisiae细胞中,但不易穿过S.pombe细胞膜却能明显促进Ca^(2+)的内流,引起S.pombe胞内Ca^(2+)总量的迅速增加。因此认为低浓度的TFP促进S.pombe细胞的增殖,不是因为TFP进入到胞内作为CaM的抑制剂而起的作用,而是通过促进胞外钙的内流从而促进S.pombe细胞的增殖。

乙醇对酿酒酵母Saccharomyces cerevisiae应激性的研究进展

在酿酒发酵工业中,虽然酿酒酵母Saccharomyces cerevisiae具有较高的乙醇耐受能力和乙醇产率,但也有一个乙醇的耐受能力极限。乙醇应激对酿酒酵母所产生的影响,包括乙醇应激对细胞质膜、HSP热激蛋白、抗氧化酶、海藻糖的影响以及高温高压等因素如何影响酿酒酵母的乙醇耐受力;以及当前研究中存在的问题。

原位预处理甘蔗糖蜜对耐高温酿酒酵母突变株Saccharomyces cerevisiae AQ生产乙醇的影响

【目的】探讨和分析原位预处理糖蜜促进酿酒酵母生长和乙醇生产的原因,开发一条绿色、低成本糖蜜乙醇生产途径。【方法】先在不同温度和初始糖浓度条件下,考察酿酒酵母原始菌株Saccharomyces cerevisiae A及其突变株Saccharomyces cerevisiae AQ的生长和乙醇生产性能差异,再以原位预处理前后糖蜜为发酵底物,测定突变株Saccharomyces cerevisiae AQ在不同培养基中的生长量、出芽率、乙醇产量、胞内超氧化物歧化酶(SOD)、过氧化物酶、细胞质内ATP酶和线粒体内ATP酶活力,研究原位预处理糖蜜对其生理特性的影响。【结果】在高温发酵糖蜜过程中,突变株Saccharomyces cerevisiae AQ较原始菌株Saccharomyces cerevisiae A表现出更好的生长和乙醇发酵稳定性。当以原位预处理糖蜜作为Saccharomyces cerevisiae AQ唯一碳源时,其胞内SOD酶、过氧化物酶、细胞质内ATP酶和线粒体内ATP酶活力较以糖蜜原料为唯一碳源时分别提高2.51倍,0.92倍,1.80倍和1.45倍,乙醇收率为31.07%,较以糖蜜原料为唯一碳源时提高36.26%。【结论】突变株Saccharomyces cerevisiae AQ较原始菌株Saccharomyces cerevisiae A更适用于糖蜜发酵生产乙醇体系,且新型的原位预处理方法能通过增强Saccharomyces cerevisiae AQ在糖蜜培养基中的呼吸作用,提高菌株活力,从而进一步提高乙醇收率。