海藻糖对面包酵母高糖耐性的影响

通过高温刺激使面包酵母积累较高的胞内海藻糖,考察胞内海藻糖含量对面包酵母高糖发酵力和高渗存活率的影响。研究结果表明,发酵结束前2h培养温度提高至40℃后,菌株BY-6胞内海藻糖含量由4.21%提高到9.76%,高糖发酵力提高了17.2%,高渗存活率也有一定提高。另外对3株面包酵母的海藻糖积累能力和面团发酵力的比较表明,海藻糖积累能力强的酵母菌株高糖耐性较好。     

面包酵母耐冷冻性影响因子的研究进展

论述了影响面包酵母耐冷冻性的各种因子。耐冷冻面包酵母是冷冻面团技术的关键,许多因素与酵母冷冻耐受性有关,如酵母胞内海藻糖含量、甘油含量、酒精含量、热休克蛋白含量、不同脂肪成分的比例、某些带电荷的氨基酸含量、呼吸能力、控制质膜的水运送的基因的表达等等,而海藻糖在酵母胞内起到的耐冷冻作用被广泛证明。     

高糖胁迫对酿酒酵母抗氧化活性及代谢的影响

本文以酿酒酵母(Saccharomy cescerevisiae)为模式生物,研究了高糖培养条件对酿酒酵母的生长、抗氧化酶活性及海藻糖、甘油代谢的影响。结果表明:当葡萄糖浓度达到40%时,酿酒酵母生长的对数期延长,对酿酒酵母的生长产生了抑制作用。酿酒酵母在培养4~10 h范围内四组酿酒酵母细胞(20 g/L葡萄糖组、40 g/L葡萄糖组、60 g/L葡萄糖组和80 g/L葡萄糖组)内海藻糖的积累随着胁迫时间的增加发生显著变化(P<0.05),海藻糖的积累量呈先升高后下降的趋势,在8 h时高糖组海藻糖积累量均达到一个最高点,胞内海藻糖的浓度最高达到0.0955 mg/mL。在不同葡萄糖浓度胁迫下,酵母细胞胞内外甘油的积累随着时间增加呈现出先上升后下降的趋势,但是胞内甘油的积累量在80 g/L葡萄糖浓度时达到最高,而胞外甘油的积累量在60 g/L葡萄糖浓度时达到最高。这些结果说明在高糖胁迫下甘油和海藻糖是酿酒酵母的主要相容性溶质。另外,高糖处理后,与对照组相比,高糖组酿酒酵母胞内超氧化物歧化酶(Superoxide Dismutase,SOD)、过氧化氢酶(Catalase,CAT)和谷胱甘肽过氧化物酶(GSH-Px)活性均显著升高(P<0.05),说明这些抗氧化物酶活性物质对维持有机体胞内正常渗透压起到关键作用。该研究结果将为今后研究酿酒酵母耐高糖渗透压方面提供理论依据。     

中性海藻糖酶基因缺失对面包酵母耐冷冻性的影响

研究了中性海藻糖酶基因(NTH1和NTH2)对面包酵母耐冷冻性的影响。通过分析中性海藻糖酶基因缺失突变株和亲本菌株(BY6-9α)的生长曲线、生物量、比生长速率、中性海藻糖酶酶活力、胞内海藻糖含量、胞内海藻糖降解速率、冷冻存活率、冷冻前产气量、比发酵力、冷冻后产气量和相对发酵力,结果表明,中性海藻糖酶基因对酵母生物量、比生长速率、胞内海藻糖含量和比发酵力均无显著影响,而相对于亲本菌株(BY6-9α),TL-101(nth1)和TL-201(nth1 nth2)的胞内海藻糖降解速率分别减慢了9.22%和15.60%,冷冻后CO2产生量分别提高了63.04%和65.22%,-20℃冷冻28 d后冷冻存活率分别提高了91.06%和103.01%,冷冻后相对发酵力分别提高了95.95%和116.04%,这充分说明敲除NTH1基因能明显改善酵母菌株的耐冷冻特性,而且酵母的耐冷冻特性与胞内海藻糖降解速率呈负相关。TL-102(nth2)的耐冷冻特性与亲本菌株无显著差异,说明单敲NTH2基因对酵母耐冷冻性无显著影响。     

面包酵母海藻糖含量与酵母耐性之间的关系

以存活率为指标,探讨了不同时期不同面包酵母的耐盐性能、耐酒精性能、耐高温性能以及耐冷冻性能,并且测定了不同时期菌体胞内海藻糖含量,研究结果表明胞内海藻糖含量与酵母的耐受性之间存在一定的相关性,海藻糖含量越高,酵母的耐性越好。     

游离高表达Mal62基因对面包酵母耐冷冻性的影响

通过测定胞内海藻糖积累量、冷冻前后相对发酵力以及存活率的变化,对比游离高表达麦芽糖酶基因（Md62）的突变株BYCPM与亲本BY14的海藻糖合成能力,研究Ma／62基因游离高表达与酵母耐冷冻性之间的关系。结果表明．Ma／62基因游离高表达与酵母耐冷冻性有一定的相关性,突变株耐冷冻性改善,其在烘焙产业中具有潜在商业价值。     

高糖胁迫下蜂蜜接合酵母碳代谢流的研究

研究蜂蜜接合酵母LGL-1在高糖胁迫下的代谢特性及代谢流变化。采用高效液相色谱等方法测定蜂蜜接合酵母LGL-1在葡萄糖质量浓度100 g/L和400 g/L条件下积累的代谢产物(酒精、海藻糖、甘油、苹果酸、α-酮戊二酸、柠檬酸及乳酸),通过构建代谢矩阵方程组,采用Matlab软件和归一法分析关键节点的代谢流分布情况。结果表明,蜂蜜接合酵母LGL-1高糖胁迫时的碳流量偏向流入海藻糖、甘油、乙醇和TCA循环的代谢支路,与常规培养相比,海藻糖、甘油和TCA循环的通量分别增加了50.00%,17.73%和4.79%,高糖培养与常规培养的乙醇代谢通量接近。在高糖胁迫条件下蜂蜜接合酵母LGL-1积累的海藻糖、甘油以及TCA循环的通量变化也反映海藻糖、甘油以及TCA循环对酵母细胞的重要保护功能。     

蜂蜜接合酵母对营养物质利用及其高糖胁迫应激代谢特性分析

研究6 株蜂蜜接合酵母利用碳源、氮源和维生素的能力,发现蜂蜜接合酵母6-7431对营养物质的代谢能力强于其他菌株,具有较好应用潜力。进一步利用扫描电镜、高效液相色谱检测等方法对该菌株在高糖胁迫下的细胞形态、繁殖方式和应激代谢产物进行探究,结果发现在300～550 g/L的高糖环境中,菌株6-7431能正常出芽生殖,其代谢产生的乙醇、甘油、海藻糖、有机酸（苹果酸、α-酮戊二酸和柠檬酸）等物质含量都远高于100 g/L含糖环境;在550～700 g/L高糖环境中,菌株6-7431的繁殖方式由出芽生殖逐渐变为孢子生殖,代谢产生的乙醇含量低于100 g/L含糖环境,但苹果酸和α-酮戊二酸质量浓度仍有所增加;在750 g/L高糖环境中,菌株6-7431严重失水,形态干瘪,其产生的乙醇体积分数为0.6%,基本停止发酵活动。研究结果揭示了高糖环境对蜂蜜接合酵母细胞形态、繁殖方式以及应激代谢产物的影响,对分析酵母的高糖耐受机制具有重要的参考价值。     

提高酱油酵母海藻糖方法研究

生物体处于恶劣环境中时,海藻糖能起到保护作用,在生产干酵母前应尽量提高胞内海藻糖的含量。文章研究了耐盐酵母中海藻糖的提高方法,即盐度和温度刺激。研究发现:S酵母和T酵母在9%盐度刺激下海藻糖分别由7.0%和7.5%增加到8.4%和9.6%;在40℃,30min的热刺激下海藻糖含量由8.4%和9.6%增加到10.1%和10.9%。变异菌株S3-2和T3-5在适宜的盐度和温度刺激下,细胞内海藻糖含量也有所提高,但总体低于原始菌株。     

高适应性面包酵母菌株的杂交选育

为了改善耐高糖酵母在不同含糖量面团中的发酵力,首先制备了2 4 0株耐高糖酵母单倍体,通过高麦芽糖发酵筛选培养基筛选出了2 8株麦芽糖发酵性能良好的单倍体菌株,通过测定这些单倍体在无糖面团中的发酵力,发现9株单倍体菌株的发酵力优于或等于其亲本,其中4株为a型,5株为α型。通过它们之间的杂交得到2 0 0株杂交株,在无糖、中糖、高糖面团中测定这些杂交株的发酵力,获得4株在不同含糖量(0～30 % )面团中都具有高发酵力的高适应性面包酵母杂交株。研究表明,来自同一亲本单倍体之间的杂交有可能改善面包酵母的某些特征。     

基于代谢组学和转录组学分析工业面包酵母（Saccharomyces cerevisiae）ABY3冷冻胁迫应答机制

为研究面包酵母（Saccharomyces cerevisiae）响应冷冻胁迫的机理，对面包酵母－20 ℃处理7 d前后的发酵菌液进行胞内的代谢组学和转录组学分析。在－20 ℃、7 d的环境胁迫下，面包酵母不加糖模拟面团发酵后的存活率为43%，发酵力下降42%。冷冻胁迫下，面包酵母胞内24 种代谢物的变化与494 种基因的表达差异与应答机制相关。通过差异代谢通路分析得出：冷冻胁迫下，胞内氨基酸的匮乏与质膜僵硬化可能是影响细胞生长和发酵性能的主要原因，而胞内不饱和脂肪酸相对含量的增加和海藻糖的积累并不能消除低温对细胞的损伤。研究结果可完善酵母冷冻胁迫应答机理，为耐性调节机制的研究提供思路，对冷冻面团的优化和技术发展具有重要意义。     

Proline accumulation in baker's yeast enhances high-sucrose stress tolerance and fermentation ability in sweet dough

During bread-making processes, yeast cells are exposed to various baking-associated stresses. High-sucrose concentrations exert severe osmotic stress that seriously damages cellular components by generation of reactive oxygen species (ROS). Previously, we found that the accumulation of proline conferred freeze-thaw stress tolerance and the baker's yeast strain that accumulated proline retained higher-level fermentation abilities in frozen doughs than the wild-type strain. In this study, we constructed self-cloning diploid baker's yeast strains that accumulate proline. These resultant strains showed higher cell viability and lower intracellular oxidation levels than that observed in the wild-type strain under high-sucrose stress condition. Proline accumulation also enhanced the fermentation ability in high-sucrose-containing dough. These results demonstrate the usefulness of proline-accumulating baker's yeast for sweet dough baking.     

Expression of the GPD1 and GPP2 orthologues and glycerol retention during growth of Debaryomyces hansenii at high NaCl concentrations

The highly NaCl-tolerant yeast Debaryomyces hansenii produces and obtains high levels of intracellular glycerol as a compatible solute when grown at high NaCl concentrations. The effect of high NaCl concentrations (4%, 8% and 12% w/v) on the glycerol production and the levels of intra- and extracellular glycerol was determined for two D. hansenii strains with different NaCl tolerance and compared to one strain of the moderately NaCl-tolerant yeast Saccharomyces cerevisiae. Initially, high NaCl tolerance seems to be determined by enhanced glycerol production, due to an increased expression of DhGPD1 and DhGPP2 (AL436338) in D. hansenii and GPD1 and GPP2 in S. cerevisiae; however, the ability to obtain high levels of intracellular glycerol seems to be more important. The two D. hansenii strains had higher levels of intracellular glycerol than the S. cerevisiae strain and were able to obtain high levels of intracellular glycerol, even at very high NaCl concentrations, indicating the presence of, for example, a type of closing channel, as previously described for other yeast species.     

空气等离子体技术对面包酵母的诱变选育研究

采用空气等离子体诱变技术对面包酵母进行诱变,以高发酵力为筛选依据,筛选出一株生产性能优良的酵母菌株。结果表明,采用空气等离子体技术对面包酵母BY-3进行诱变最佳时间为10 s。通过诱变,筛选得到一株高发酵力的酵母菌株3G-28。与原始菌株BY-3相比,菌株3G-28的发酵力提高了48.57%;具有更好的耐高糖性能,其蔗糖酶活力较初始菌株BY-3降低了71.89%,抗葡萄糖阻遏现象的能力提高了34.84%;具有更好的传代稳定性和低温适应性:传代15次,其发酵力仍可保持87.50%;在-20℃低温下进行保藏,两周后,其菌株的相对发酵力可达到76.10%,酵母泥的相对发酵力为83.91%。研究表明,空气等离子体技术能够有效的对面包酵母BY-3进行诱变,为面包酵母的改良育种提供了一定的依据。     

Identification of genes whose expressions are enhanced or reduced in baker's yeast during fed-batch culture process using molasses medium by DNA microarray analysis

Genes whose expression levels are enhanced or reduced during the cultivation process that uses cane molasses in baker's yeast production were identified in this study. The results showed that baker's yeast grown in molasses medium had higher fermentation ability and stress tolerance compared with baker's yeast grown in synthetic medium. Molasses apparently provided not only sugar as a carbon source but also provided functional components that enhanced or reduced expression of genes involved in fermentation ability and stress tolerance. To identify the genes whose expression is enhanced or reduced during cultivation in molasses medium, DNA microarray analysis was then used to compare the gene expression profile of cells grown in molasses with that of cells grown in synthetic medium. To simulate the commercial baker's yeast production process, cells were cultivated using a fed-batch culture system. In molasses medium, genes involved in the synthesis or uptake of vitamins (e.g., biotin, pyridoxine and thiamine) showed enhanced expression, suggesting that vitamin concentrations in molasses medium were lower than those in synthetic medium. Genes involved in formate dehydrogenase and maltose assimilation showed enhanced expression in molasses medium. In contrast, genes involved in iron utilization (e.g., siderophore, iron transporter and ferroxidase) showed enhanced expression in synthetic medium, suggesting that iron starvation occurred. The genes involved in the metabolism of amino acids also showed enhanced expression in synthetic medium. This identification of genes provides information that will help improve the baker's yeast production process.     

面包酵母自交提升耐高糖发酵力的研究

用工业耐高糖面包酵母BH3制备了102株单倍体,通过四分体分析发现,控制生物量和耐高糖发酵力的主效基因在此菌株中是杂合的,并且控制生物量的主效基因和控制接合型的基因MAT极大可能是连锁的。结果表明,通过自交和人工选择相结合的方式,筛选出耐高糖且发酵活力优良的18株单倍体,构建27个交配组合,其中有11个组合的自交菌株高于亲本的耐高糖发酵力水平,最高能达到亲本水平的116.73%,体现出菌株BH3在自交系选育上有进步空间。     

TOR1基因缺失对酿酒酵母耐受性的影响

酿酒酵母是最常见且应用最为广泛的酵母菌种,是以糖质和淀粉质为原料的乙醇发酵最经典的菌株。在发酵过程中,有很多不可避免的胁迫环境如高温条件、高渗条件等出现,这些胁迫会阻碍细胞生长并降低细胞的发酵能力,给发酵行业带来一定的经济损失。因此,为改善菌种的耐受性,该研究主要以实验室现有菌株AY12a为亲本菌株,URA3基因作筛选标记,通过胞内同源重组,实现TOR1基因的敲除,最终成功构建突变株AY12a-tor1Δ。对酵母进行耐受性的测定,发现AY12a-tor 1Δ具有一定的耐高温性能,在高渗条件下也有一定的耐受性,同时具有一定的氧化环境耐受性。同时将突变株与AY12a进行模拟白酒发酵(玉米浓醪发酵),并对发酵完成后的酒度、残糖、48 h细胞存活率、CO 2失重及发酵时间进行测定。发酵数据显示突变株AY12a-tor 1Δ乙醇产量有所上升,残糖含量下降,48 h细胞存活率没有下降,发酵时间有所延长。     

Development of a novel microbial sensor with baker's yeast cells for monitoring temperature control during cold food chain

A novel microbial sensor containing a commercial baker's yeast with a high freeze tolerance was developed for visibly detecting inappropriate temperature control of food. When the yeast cells fermented glucose, the resulting gas production triggered the microbial sensor. The biosensor was a simple, small bag containing a solution of yeast cells, yeast extract, glucose, and glycerol sealed up with multilayer transparent film with barriers against oxygen and humidity. Fine adjustment of gas productivity in the biosensor at low temperatures was achieved by changing either or both concentrations of glucose and yeast cells. Moreover, the amount of time that food was exposed to inappropriate temperatures could be deduced by the amount of gas produced in the biosensor. The biosensor was stable without any functional loss for up to 1 week in frozen storage. The biosensor could offer a useful tool for securing food safety by maintaining low-temperature control in every stage from farm to fork, including during transportation, in the store, and at home.