丛梗孢酵母发酵产赤藓糖醇条件的优化

丛梗孢酵母BH010是从蜂蜜样品中分离得到的产赤藓糖醇菌株。该实验研究了发酵培养基及发酵条件对丛梗孢酵母赤藓糖醇产量的影响。单因素实验及正交实验的结果表明,最佳发酵培养基及发酵条件为:葡萄糖含量(质量浓度)35%、酵母膏含量(质量浓度)1%、CaCl2.2H2O(质量浓度)0.2%,初始pH6.0,接种量1%,30℃摇瓶培养9d。最终赤藓糖醇产量为110.61g/L发酵液,比普通发酵条件下提高85.56%。     

高产赤藓糖醇菌株RH-UV-L4-F9发酵条件的优化

以高产赤藓糖醇菌株RH-UV-L4-F9为研究对象,采用生物统计方法分别对该菌株的发酵培养基和培养条件进行优化。发酵培养基最佳配比为葡萄糖30%,酵母膏0.5%,脲1%,MgSO_4 0.05%;最适发酵条件为34℃,初始pH值6.0.摇床转数180r/min。最适条件下赤藓糖醇产量为157.4mg/mL。     

响应面法优化产赤藓糖醇菌株RH-UV-L4-F9的发酵条件

为进一步提高糖蜜玉米浆发酵生产赤藓糖醇的产量,以高产赤藓糖醇耐高渗酵母RH-UV-L4-F9为出发菌株,采用响应面法优化发酵工艺参数。根据Box-Benhnken设计原理,在单因素试验基础上,采用三因素三水平响应面分析,确定最佳发酵工艺参数为：温度32.3℃、pH5.1、摇床转速176r/min。在此优化条件下赤藓糖醇产量达到166.89mg/mL,是初始条件下的1.2倍。     

Torulopsis sp.ERY237产赤藓糖醇工艺条件的研究

以Torulopsis sp.ERY237作为出发菌株,考察了不同碳源、氮源、无机盐类以及温度等因素对菌种产赤藓糖醇的影响,建立和优化了赤藓糖醇摇瓶发酵培养基配方、发酵工艺条件,同时研究了发酵过程中菌体生物量、pH值、产物浓度的动态变化。结果表明,菌株的最适培养基配方为(g/L):葡萄糖300,玉米浆3.5,C_([Cu~(2+)])1.5,C_([Mn~(2+)])10;适宜的培养条件为初始pH值自然,温度30℃,装液量50 mL/500 mL,转速200 r/min,在此条件下培养132 h赤藓糖醇产量达87.8 g/L,是优化前产量的1.9倍,发酵时间缩短了12 h。     

圆酵母B84512单倍体的制备及其产赤藓糖醇特性分析

圆酵母B84512为工业用赤藓糖醇生产菌株,为获得其遗传育种单倍体亲本,以Mcclary产孢培养基进行该菌株子囊孢子萌发,萌发条件为:30℃,培养7 d,菌株产孢率为45%;以蜗牛酶裂解子囊孢子细胞壁150 min,共筛选出10株单倍体菌株,其中3株在发酵培养基中合成赤藓糖醇的能力相对较高,分别命名为Torula sp.B84512-7,Torula sp.B84512-8及Torula sp.B84512-9。经PCR验证,前两者为α型,后者为a型。将3株单倍体两两杂合,发现杂合子Torula sp.B84512-79的发酵性能最佳,赤藓糖醇产量高达97 g/L,约为出发菌株的56%。赤藓糖醇产量较高的单倍体亲本Torula sp.B84512-7及Torula sp.B84512-9可用于基因工程育种。     

碳源对圆酵母(Torula sp.)B84512发酵合成赤藓糖醇及副产物甘油的影响

研究了圆酵母(Torula sp.)B84512以不同碳源发酵产赤藓糖醇过程中副产物甘油的生成与消耗情况。发现该菌株在以任何碳源为底物发酵过程中均会产生甘油,且在发酵中后期甘油逐渐被消耗。以甘油为唯一碳源时该菌株合成赤藓糖醇的速率及产率均低于葡萄糖。葡萄糖为圆酵母B84512发酵产赤藓糖醇的最佳碳源。采用分批补料的方式提高赤藓糖醇的产率并期望能抑制甘油的生成,实验结果表明补料至总糖浓度为50%时赤藓糖醇产量最高为253 g/L,产率为1.03 g/(L.h)。但甘油产量与葡萄糖的浓度呈正相关,分批补料并不能有效抑制甘油的生成,反而导致发酵周期大大延长,对于工业化生产极其不利。通过对甘油的生成及消耗过程中关键酶胞浆3-磷酸甘油脱氢酶(ctGPD)、3-磷酸甘油酯酶(GPP)、线粒体3-磷酸甘油脱氢酶(mtGPD)酶活测定,确定胞浆3-磷酸甘油脱氢酶为甘油合成途径的关键酶,为以后对圆酵母B84512中甘油代谢途径的基因工程改造选育奠定了基础。     

丛梗孢酵母产赤藓糖醇的诱变选育

利用微波-硫酸二乙酯复合诱变对产赤藓糖醇丛梗孢酵母E54进行处理,以高渗平板和摇瓶发酵为筛选方法,得到遗传稳定的诱变高产株EW29;再采用氮离子注入对EW29进行诱变处理,摇瓶发酵筛选得到诱变株EN59,其90h发酵液中赤藓糖醇产量达到55.13g/L,较EW29提高20.3%,较E54提高36.9%,遗传稳定性较好。对突变株EN59的发酵培养基进行了优化,在优化培养基葡萄糖250g/L、酵母膏5g/L、KH2PO4 0.3g/L、MnSO4 ·4H2O 0.04g/L、CuSO4 ·5H2O 0.03g/L,初始pH4的条件下,90h发酵液中赤藓糖醇平均产量达到69.00g/L以上。在优化培养基的基础上进行5L罐发酵放大实验,发酵126h赤藓糖醇产量达到71.14g/L。     

无机盐渗透压对假丝酵母SK25.001发酵生产赤藓糖醇的影响

以假丝酵母SK25.001为生产菌,通过研究其发酵产赤藓糖醇的碳源、氮源、碳氮比以及NaCl、KCl对其发酵产赤藓糖醇的影响,来探索无机盐(NaCl,KCl)渗透压对赤藓糖醇发酵的影响。结果发现,葡萄糖、酵母粉分别是其最佳碳源和氮源,最佳碳氮比为20∶1,转化率达到了14.2%;向发酵培养基中添加不同浓度的KCl或NaCl后发现,菌体生长速度随着KCl或NaCl浓度增大而降低,在KCl浓度为0.4 mol/L或NaCl浓度为0.3 mol/L时赤藓糖醇产量达到最大,达到了18.4 g/L和17.4 g/L;将NaCl和KCl的浓度用渗透压表示发现赤藓糖醇的转化率随着渗透压的增大而升高,高渗透压抑制菌体的生长。     

两阶段调控葡萄糖质量浓度强化赤藓糖醇发酵的研究

该研究在5 L发酵罐水平上研究了不同初始葡萄糖质量浓度对解脂耶氏酵母（Yarrowia lipolytica）JZ-204生长和发酵产赤藓糖醇的影响。结果表明,100 g/L初始葡萄糖质量浓度有利于菌体生长,高初始葡萄糖质量浓度（300 g/L和400 g/L）有利于赤藓糖醇合成。基于此,提出两阶段葡萄糖质量浓度调控策略,即0 h时以初始葡萄糖质量浓度为100 g/L,22 h后通过补加葡萄糖使总糖量达300 g/L进行发酵。结果表明,与分批发酵相比（100 g/L、200 g/L、300 g/L、400 g/L）,采用该调控策略,赤藓糖醇产量达到最高水平92.66 g/L,分别比分批发酵提高了1 347.81%、84.54%、14.66%、7.57%;生产强度达到最高的0.48 g/（L·h）,分别比分批发酵提高了300%、37.14%、29.73%、20.00%。该调控策略为赤藓糖醇的高效发酵合成提供参考。     

产赤藓糖醇菌株的筛选与鉴定

利用含有300g/L 葡萄糖的高渗培养基从蜂蜜、花粉、土壤等样品中筛选耐高渗酵母菌,经薄层层析和高效液相色谱分析得到两株产赤藓糖醇且不产甘油的酵母菌,通过高碘酸氧化法筛选出其中赤藓糖醇产量较高的一株菌株E54。菌株E54 在含葡萄糖200g/L、酵母膏5g/L 的发酵培养基中发酵90h,赤藓糖醇产量为41.1g/L,转化率为22.8%。通过形态观察、生理生化实验、5.8S rDNA 序列分析并构建系统进化树,初步鉴定E54 为Moniliellaacetoabutans(丛梗孢酵母)。     

High-level production of erythritol by strain 618A-01 isolated from pollen

We have isolated a microorganism (strain 618A-01) from pollen which has the ability to produce erythritol when grown in the presence of glucose as the carbon source. When cultivated in a medium consisting of 20% glucose and 1% dried bouillon in a shake flask, 75 g/l erythritol was produced after 950 h, corresponding to a 37.5% yield against glucose consumption. No other polyols, including glycerol, were detected in the medium. Positive-ion fast atom bombardment mass spectrometry and 1H- and 13C-NMR analyses confirmed that the fermentation product was erythritol. Scanning electron microscopic analysis clearly demonstrated that the cells grown on YPD medium at 30 degrees C showed yeast-like morphology, while they appeared like hyphae at 37 degrees C. The complete 18S rRNA sequence of the isolate was determined, which showed high identity (99.5%) with the genus Ustilago of the phylum Basidiomycota. The data strongly suggest that strain 618A-01 belongs to the class Ustilaginomycetes. The culture conditions for the production of erythritol by the isolate were examined. The use of medium containing 1% tryptone, 0.5% yeast extract and 0.5% NaCl yielded the highest cell growth and erythritol productivity among the media tested. Continuous glucose feeding at 6-7% to the fermentor further increased the production of erythritol, and we obtained a maximal 100 g/l erythritol after 530 h, with a 39.3% yield.     

Ethanol yield and volatile compound content in fermentation of agave must by Kluyveromyces marxianus UMPe-1 comparing with Saccharomyces cerevisiae baker's yeast used in tequila production

In tequila production, fermentation is an important step. Fermentation determines the ethanol productivity and organoleptic properties of the beverage. In this study, a yeast isolated from native residual agave must was identified as Kluyveromyces marxianus UMPe-1 by 26S rRNA sequencing. This yeast was compared with the baker's yeast Saccharomyces cerevisiae Pan1. Our findings demonstrate that the UMPe-1 yeast was able to support the sugar content of agave must and glucose up to 22% (w/v) and tolerated 10% (v/v) ethanol concentration in the medium with 50% cells survival. Pilot and industrial fermentation of agave must tests showed that the K. marxianus UMPe-1 yeast produced ethanol with yields of 94% and 96% with respect to fermentable sugar content (glucose and fructose, constituting 98%). The S. cerevisiae Pan1 baker's yeast, however, which is commonly used in some tequila factories, showed 76% and 70% yield. At the industrial level, UMPe-1 yeast shows a maximum velocity of fermentable sugar consumption of 2.27g·L(-1)·h(-1) and ethanol production of 1.38g·L(-1)·h(-1), providing 58.78g ethanol·L(-1) at 72h fermentation, which corresponds to 96% yield. In addition, the major and minor volatile compounds in the tequila beverage obtained from UMPe-1 yeast were increased. Importantly, 29 volatile compounds were identified, while the beverage obtained from Pan1-yeast contained fewer compounds and in lower concentrations. The results suggest that the K. marxianus UMPe-1 is a suitable yeast for agave must fermentation, showing high ethanol productivity and increased volatile compound content comparing with a S. cerevisiae baker's yeast used in tequila production.     

Optimization of carotenoids by <Emphasis Type="Italic">Rhodotorula glutinis</Emphasis>

Response surface methodology was applied to maximize the yield and productivity of carotenoids by Rhodotorula glutinis strain 1151 using supplemented tomato waste based medium. Higher concentration of tomato waste extract and yeast extract favored the production of carotenoids. In contrast to carotenogenesis higher concentration of yeast extract negatively affected the formation of biomass whereas higher amount of glucose in the medium favored biomass indicating that carotenogenesis is not correlated to biomass. The optimal concentration of medium components for maximum total carotenoids and corresponding biomass production as obtained from model were calculated to be as 660 mL/L, 1.5, 4.5, 7.4, and 10 g/L for tomato extract, malt extract, yeast extract, peptone, and glucose, respectively.     

Sporidiobolus pararoseus WZ012产类胡萝卜素发酵条件的优化

以响应面方法对Sporidiobolus pararoseus WZ012产类胡萝卜素的摇瓶发酵条件进行了优化。在Plackett-Burman实验的基础上,利用中心组合设计研究了4个主要因素(葡萄糖、酵母膏、KH2PO4、接种量)对产类胡萝卜素的影响。结果表明,较高浓度的酵母膏和较大的接种量有助于类胡萝卜素的生产,而较高浓度的葡萄糖和KH2PO4有助于生物量的提高,生物量与类胡萝卜素产量之间没有必然的联系。优化得到的发酵培养基组成(g/L)为:葡萄糖60.23、酵母膏12.21、KH2PO41.68、接种量为8.38%。在此条件下红酵母产类胡萝卜素的最大产量为29.3mg/L,较优化前提高了54.1%。类胡萝卜素成分分析表明主要含有β-胡萝卜素、圆酵母素和红酵母红素这3种类胡萝卜素物质,其中β-胡萝卜素占62.5%。     

Sophorolipid production by Candida bombicola: medium composition and culture methods

Candida bombicola is able to produce sophorolipid molecules with surfactant properties when grown in a medium composed of two different carbon sources (usually sugar and oil) and a nitrogen source (frequently yeast extract). In this work, the composition of the medium and the culture method employed have been studied. The influences of glucose concentration, properties of oil and yeast extract concentration have been taken into account. Accordingly, a production medium composition is proposed (100 g/l glucose, 100 g/l sunflower oil and 1 g/l yeast extract). The most frequent culture methods reported in literature, i.e. batch, medium pulse mode and resting-cell methods, have been tested. The resting-cell method was found to produce the highest final concentration of sophorolipid, obtaining a good yield of carbon sources in a relatively short time. Under the best operational conditions and using the resting-cell method, 120 g/l sophorolipid was obtained in 8 d, with a carbon source yield of 0.60. Product distribution has also been investigated and the sophorolipid molecular structure of opened or cycled molecules has been determined under different operational conditions. Low yeast extract concentration and long fermentation time enhance the production of cycled structures by all the production methods studied.     

高产酵母菌株(Rhodotorula bacarum)产普鲁兰多糖过程中搅拌和通气条件的优化

在过去的研究中发现普鲁兰多糖高产酵母菌株摇瓶发酵的最佳条件是 180r/min ,2 8℃ ,6 0h ,在此条件下该菌株可以产生质量体积分数为 5 9%的普鲁兰多糖。本文研究发现在 5L发酵罐中通气量和搅拌速度对该酵母菌株的普鲁兰多糖产量有很明显的影响。研究结果表明产普鲁兰多糖的最适通气量和搅拌速度分别是6 . 5L/min和 30 .0r/min。相应的 ,用每升含 80g的葡萄糖做发酵培养基 ,在 2 8℃条件下培养 72h ,其普鲁兰多糖的产量质量体积分数为达到 7. 5 % ,这是迄今为止所报道的产普鲁兰多糖酵母中产量最高的菌株。作者发现这株酵母菌产普鲁兰多糖高效合成过程的原因是其有很高的葡萄糖转化率。