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。     

耐高渗酵母产赤藓糖醇的影响因素

球拟酵母OS-194是一株单产赤藓糖醇的耐高渗酵母,该菌株高产赤藓糖醇的最佳培养基配方为葡萄糖10g/dL,酵母膏0.5g/dL,尿素0.1g/dL.最适培养条件是在摇瓶转速150r/min的条件下于35℃培养4d.在上述培养条件下,该菌株赤藓糖醇的耗糖转化率高达29.6%.磷是限制OS-194菌株高产赤藓糖醇的主要因素,当培养液中的磷质量浓度低于31.5mg/L时,赤藓糖醇的产量最高;随着磷质量浓度的升高,该菌株赤藓糖醇的产量降低,而酒精的产量和生物量却有明显升高.同时,OS-194菌株还能利用果糖、蔗糖和D-甘露糖产赤藓糖醇.     

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%。     

耐高渗酵母B845产赤藓糖醇的研究

以自然界中筛选分离到的耐高渗酵母B8为出发菌株,经过紫外诱变,得到了赤藓糖醇产量较高的突变株B845。对B845进了一系列的发酵工艺条件试验。经初步研究,B845在葡萄糖20%,酵母膏0.5%,脲0.1%、pH6.0的发酵液中经35℃摇瓶5天,可产赤藓糖醇75mg/ml,对糖转化率41.2%。     

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。     

解脂亚罗酵母赤藓糖醇发酵过程的研究

从自然界分离得到1株可发酵葡萄糖为赤藓糖醇的酵母菌Y-22。经鉴定,属于解脂亚罗酵母。该酵母菌在高糖条件下转化赤藓糖醇的能力比众多从国内保藏机构获得和自然界分离得到的同种、异种酵母及未鉴定耐渗酵母高很多。在5 m3和50 m3容量的发酵罐上进行发酵试验,对葡萄糖的转化率可以达到47%,赤藓糖醇在96 h内可达150 g/L以上。文中还对赤藓糖醇的分批发酵和流加发酵过程进行了讨论。     

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。     

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。     

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。     

甘氨酸、脯氨酸促进高渗环境下Yarrowia lipolytica发酵甘油产赤藓糖醇

解脂耶氏酵母(Yarrowia lipolytica)可很好地发酵甘油生产赤藓糖醇。NaCl作为渗透压调节剂提升发酵体系渗透压有利于提高赤藓糖醇产量,但高渗会抑制酵母生长,延长发酵周期,降低生产强度。该研究以甘氨酸、脯氨酸为渗透压保护剂,在高渗环境下,研究其如何提升酵母细胞耐高渗能力。结果发现,Y.lipolytica可吸收胞外甘氨酸和脯氨酸并在胞内积累以抵御高渗胁迫,并显著提升高渗环境下的菌体量,促进赤藓糖醇的高效合成。在7 L发酵罐水平,初始渗透压为4.17±0.17 osmol/kg时,发酵初始添加30 mg/L甘氨酸和40 mg/L脯氨酸,发酵时间由108 h减少到90 h,赤藓糖醇最终产量达到了93.6±4.2 g/L,生产强度为1.04±0.05 g/(L·h),产物得率为0.49±0.03 g/g,分别比未添加保护剂时增加了4.12%,25.3%和4.26%。     

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.     

产多元醇酵母的耐高渗生长特征

应用低水活度选择性增殖的方法从花卉、果园土壤、水果和蜂产品等样品中分离得到２６７株耐高渗酵母,其中２０６株生产阿拉伯醇、赤藓糖醇和甘油等各种多元醇．对随机挑选的６２株产多元醇的耐高渗酵母在不同质量浓度葡萄糖培养基中的生长特征进行了研究,结果显示酵母的耐高渗生长能力与其产多元醇的类别之间存在关联．在所调查的酵母中,产赤藓糖醇的酵母耐高渗生长能力最强、产阿拉伯醇的酵母耐高渗生长能力次之,产甘油的酵母耐高渗生长能力较弱．     

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。     

Production of L-erythrose via L-erythrulose from erythritol using microbial and enzymatic reactions

A rare aldotetrose, L-erythrose, was produced from erythritol via a two-step reaction. In the first step, complete oxidation of erythritol to L-erythrulose was achieved by using Gluconobacter frateurii IFO 3254. Washed cell suspension of the strain grown on tryptic soy broth (TSB) supplemented with 1% d-sorbitol was used to carry out the transformation reaction at 30 degrees C with shaking at 170 rpm. At 10% substrate concentration, 98% erythritol was converted to L-erythrulose within 48 h. The produced L-erythrulose was then used as a substrate for the production of L-erythrose. The isomerization of L-erythrulose to L-erythrose was carried out using constitutively produced L-ribose isomerase (l-RI) from the mutant strain Acinetobacter sp. DL-28 grown on D-lyxose mineral salt medium. At equilibrium, the yield of L-erythrose from L-erythrulose was 18% and finally 1.7 g L-erythrose was obtained from 10 g erythritol. After a number of simple purification steps, the product was isolated from the reaction mixture by ion-exchange column chromatography (Dowex 50W-X2, Ca2+). The structure of the product was determined after NaBH4 reduction from Infrared (IR) and 13C nuclear magnetic resonance (NMR) spectra.     

从酸菜液中筛选产γ-氨基丁酸的菌株

从腌制的酸菜液中,采用乳酸菌分离纯化法,经初筛、复筛得到一株产γ-氨基丁酸的菌株,编号为LpL-0212,对菌株进行形态学观察和生理生化实验及16S rDNA、atpA基因序列分析鉴定,鉴定该菌株为Enterococcus faecium。在含2%谷氨酸钠的TYG发酵培养基中静置培养24h,经薄层层析定性、高效液相色谱法测定,发酵液中的γ-氨基丁酸含量可达到102.37μmol/L。     

PRODUCTION of FLAVOR BYAMUTANT of YEAST

ABSTRACT. Strain CCU-N₁₆-18143 was derived with N-methyl-N'-nitro-N-nitrosoguanidine (NTG) treatment three times from a wild strain of yeast CCU-16 as a mutant to produce more delicious and intense flavors for nutrient beverage from glucose medium. This mutant was identified by computer system as Saccharomyces sp. the optimal culture medium for the production of flavor is one liter of medium containing 100 g glucose, 12 g ammonium nitrate, 3 g yeast extract, 1 g magnesium sulfate, 1 g ammonium sulfate, 2 g potassium dihydrogen phosphate, pH: 3.5. the optimal culture conditions are: temperature: 30C; agitation: 150 rpm., 50 ml medium in 500-ml Hinton flask; incubation time: 96 h. the volatile flavor compounds in the culture medium were analyzed by capillary gas chromatography and capillary gas chromatography-mass spectrometry. It was found that more volatile compounds were produced by mutant strain than wild strain. the content of isoamyl alcohol increased from 7.76 to 61.64%, whereas ethyl alcohol decreased from 85.25 to 5.77%.     

Production of L-sorbitol from L-fructose by Aureobasidium pullulans LP23 isolated from soy sauce mash

A strain LP23 that can convert L-fructose to L-sorbitol was isolated from soy sauce mash and identified as Aureobasidium pullulans. The cells grown on L-arabinose were found to have relatively high L-fructose to L-sorbitol conversion potential. Addition of erythritol to the reaction mixture considerably accelerated the conversion rate of L-fructose to L-sorbitol. During the conversion reaction, erythritol was added to the reaction mixture at 8-h intervals to maintain the concentration of erythritol at 1.0%. The final conversion ratios were 82.8%, 95.3%, 92.4%, and 42.6% using washed cells when the concentrations of L-fructose were 1.0%, 2.0%, 5.0% and 10.0%, respectively. The product from L-fructose was identified as L-sorbitol by HPLC analysis, infrared spectroscopy, optical rotation and melting point measurements.     

Effect of Medium Composition on Glucoamylase Production During Batch Fermentation of Recombinant Saccharomyces cerevisiae

Plasmid stability of the recombinant Saccharomyces cerevisiae C468/pGAC9 (ATCC 20690) strain harboring a pGAC9 plasmid with glucoamylase genes has been investigated in shake flasks and in a bioreactor system using various compositions of media containing glucose or starch as the main carbon and energy source. The medium composition affected both the growth characteristics of S. cerevisiae and stability of the plasmid. Superior plasmid stability was obtained in yeast minimal medium and in complex medium with 0.5 to 2% D‐glucose. Plasmid stability of 92% was obtained in complex medium with 2% D‐glucose yielding 48 units of glucoamylase/g of cells compared to 54% plasmid stability achieved with 2% soluble starch, which yielded 23 units of glucoamylase/g of cells. The plasmid stability increased at high growth rates and decreased with increasing starch concentration in the complex media as compared to glucose medium. The kinetic characteristics of biomass and glucoamylase production were investigated, and a growth kinetic model was used to interpret the experimental results.