采用玉米秸秆水解糖和玉米浆发酵生产丁二酸

研究了以玉米秸秆水解糖为碳源,不同氮源条件下琥珀酸放线杆菌Actinobacillus succinogenesSF-9的丁二酸发酵产酸能力。结果表明玉米浆可以替代酵母膏作为丁二酸发酵的廉价氮源。厌氧摇瓶丁二酸发酵单因素试验,得到在初糖浓度50 g/L时,玉米浆的较佳用量为20 g/L。在5 L搅拌罐上,考察了不同初始玉米秸秆水解糖浓度对A.succinogenes SF-9发酵生产丁二酸的影响,结果显示高初始秸秆糖浓度对琥珀酸放线杆菌的生长有抑制作用。采用补料分批发酵,发酵60 h丁二酸的产量达到42.7g/L,丁二酸产率82.7%,生产强度0.81 g/(L·h)。丁二酸的产量和生产强度较分批发酵有明显提高。     

Fermentation of Detoxified Acid-Hydrolyzed Pyrolytic Anhydrosugars into Bioethanol with <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> 2.399

Pyrolysate obtained from the pyrolysis of waste cotton is a source of fermentable sugars that could be fermented into bioethanol fuel and other chemicals via microbial fermentation. However, pyrolysate is a complex mixture of fermentable and non-fermentable substrates causing inhibition of the microbial growth. The aim of this study was to detoxify the hydrolysate and then ferment it into bio-ethanol fuel in shake flasks and fermenter applying yeast strain Saccharomyces cerevisiae 2.399. Pyrolysate was hydrolyzed to glucose with 0.2 M sulfuric acid, neutralized with Ba(OH)₂ followed by treatment with ethyl acetate and activated carbon to remove fermentation inhibitors. The effect of various fermentation parameters such as inoculum concentration, pH and hydrolysate glucose was evaluated in shake flasks for optimum ethanol fermentation. With respect to inoculum concentration, 20% v/v inoculum i.e. 8.0 × 10⁸–1.2 × 10⁹ cells/mL was the optimum level for producing 8.62 ± 0.33 g/L ethanol at 9 h of fermentation with a maximum yield of 0.46 g ethanol/g glucose. The optimum pH for hydrolysate glucose fermentation was found to be 6.0 that produced 8.57 ± 0.66 g/L ethanol. Maximum ethanol concentration, 14.78 g/L was obtained for 4% hydrolysate glucose concentration after 16 h of fermentation. Scale-up studies in stirred fermenter produced much higher productivity (1.32 g/L/h^–1) compared to shake flask fermentation (0.92 g/L/h^–1). The yield of ethanol reached a maximum of 91% and 89% of the theoretical yield of ethanol in shake flasks and fermenter, respectively. The complex of integrated models of development was applied, that has been successfully tested previously for the mathematical analysis of the fermentation processes.     

Strain development and medium optimization for fumaric acid production

Rhizopus oryzae RUR709 mutant was isolated based on halo size from selection medium via mutagenesis with UV and γ-rays, and the production of fumaric acid in the submerged fermentation was assessed. The maximum concentration of fumaric acid was obtained using 0.5% corn steep liquor (CSL) as the nitrogen source. Organic nitrogen sources were shown to be more effective in fumaric acid production than inorganic nitrogen sources. Using optimum medium obtained by response surface methodology (RSM), the maximum concentration of fumaric acid achieved in flask culture was 26.2 g/L, which is fairly close to the 27.4 g/L predicted by the model. The highest concentration of fumaric acid in the stirred-tank reactor generated by the R. oryzae RUR709 mutant was 32.1 g/L and yield (0.45 g/g) and productivity (0.32 g/L/h) were highest at 4 days.     

玉米浆在产甘油假丝酵母甘油发酵中的作用机理

以复合培养基和合成培养基进行比较发酵,研究了玉米浆在产甘油假丝酵母甘油发酵过程中的作用机理。结果表明:玉米浆中的磷、氮和微量元素是影响产甘油假丝酵母甘油发酵的3个关键因素。当玉米浆磷浓度为121·75mg/L(玉米浆浓度为14g/L),最大甘油转化率达到53·44%。玉米浆磷可以调节EMP途径与HMP途径之间碳架代谢流的分布,随着玉米浆浓度进一步增加,过量磷能抑制HMP途径而激活EMP途径,因而复合培养基各项发酵参数的变化非常显著。玉米浆氮对磷的调节功能有协同作用,但并不是产甘油假丝酵母甘油发酵的理想氮源。玉米浆中的微量元素能够显著提高葡萄糖的消耗速率、促进菌体的生长和增加甘油的产量。     

树干毕赤酵母和酿酒酵母混合糖发酵产乙醇

以树干毕赤酵母和酿酒酵母为发酵菌株,酸性蒸汽爆破玉米秸秆预水解液和纯糖模拟液为C源,采用固定化酵母细胞的方法,研究了酸爆玉米秸秆预水解液初始pH、N源种类及其浓度、3种发酵模式对树干毕赤酵母戊糖发酵的影响。结果表明:玉米秸秆预水解液适合发酵的初始pH范围为6.0～7.0;1.0 g/L的(NH4)2SO4作为N源,在40 g/L葡萄糖和25 g/L木糖培养基中发酵24 h,糖利用率达到99.47%,乙醇质量浓度为24.72 g/L,优于尿素和蛋白胨作为N源;3种模式的发酵体系中,以游离树干毕赤酵母和固定化酿酒酵母发酵性能最好,糖利用率和乙醇得率分别为99.43%和96.39%。     

Ethanol production from syngas by Clostridium strain P11 using corn steep liquor as a nutrient replacement to yeast extract

The feasibility of replacing yeast extract (YE) by corn steep liquor (CSL), a low cost nutrient source, for syngas fermentation to produce ethanol using Clostridium strain P11 was investigated. About 32% more ethanol (1.7 g L⁻¹) was produced with 20 g L⁻¹ CSL media in 250-mL bottle fermentations compared to media with 1 g L⁻¹ YE after 360 h. Maximum ethanol concentrations after 360 h of fermentation in a 7.5-L fermentor with 10 and 20 g L⁻¹ CSL media were 8.6 and 9.6 g L⁻¹, respectively, which represent 57% and 60% of the theoretical ethanol yields from CO. Only about 6.1 g L⁻¹ of ethanol was obtained in the medium with 1 g L⁻¹ YE after 360 h, which represents 53% of the theoretical ethanol yield from CO. The use of CSL also enhanced butanol production by sevenfold compared to YE in bottle fermentations. These results demonstrate that CSL can replace YE as the primary medium component and significantly enhance ethanol production by Clostridium strain P11.     

Production of ethanol and xylitol from corn cobs by yeasts

Saccharomyces cerevisiae and Candida tropicalis were used separately and as co-culture for simultaneous saccharification and fermentation (SSF) of 5-20% (w/v) dry corn cobs. A maximal ethanol concentration of 27, 23, 21 g/l (w/v) from 200 g/l (w/v) dry corn cobs was obtained by S. cerevisiae, C. tropicalis and the co-culture, respectively, after 96 h of fermentation. However, theoretical yields of 82%, 71% and 63% were observed from 50 g/l dry corn cobs for the above cultures, respectively. Maximal xylitol concentration of 21, 20 and 15 g/l from 200 g/l (w/v) dry corn cobs was obtained by C. tropicalis, co-culture, and S. cerevisiae, respectively. Maximum theoretical yields of 79.0%, 77.0% and 58% were observed from 50 g/l of corn cobs, respectively. The volumetric productivities for ethanol and xylitol increased with the increase in substrate concentration, whereas, yield decreased. Glycerol and acetic acid were formed as minor by-products. S. cerevisiae and C. tropicalis resulted in better product yields (0.42 and 0.36 g/g) for ethanol and (0.52 and 0.71 g/g) for xylitol, respectively, whereas, the co-culture showed moderate level of ethanol (0.32 g/g) and almost maximal levels of xylitol (0.69 g/g).     

Fermentative production of succinic acid from glucose and corn steep liquor byAnaerobiospirillum succiniciproducens

Anaerobiospirillum succiniciproducens requires expensive complex nitrogen sources such as yeast extract and polypeptone for its growth and succinic acid production. It was found thatA. succiniciproducens was able to grow in a minimal medium containing glucose when supplemented with corn steep liquor (CSL) as the sole complex nitrogen source. The concentration of CSL had a significant effect on the glucose consumption byA. succiniciproducens. When 10–15 g/L of CSL was supplemented, cells were grown to an OD₆₆₀ of 3.5 and produced 17.8 g/L succinic acid with 20 g/L glucose. These results are similar to those obtained by supplementing yeast extract and polypeptone, thereby suggesting that succinic acid can be produced more economically using glucose and CSL.     

Optimization of pH,Temperature and Inoculum Ratio for the Production of β‐D‐Galactosidase by Kluyveromyces marxianus Using a Lactose‐free Medium

The pH, temperature and inoculum ratio for the production of β‐galactosidase by Kluyveromyces marxianus CDB 002 were optimized using sugar‐cane molasses (100 g/l) in a lactose‐free medium. The temperature optimum was evaluated in the range from 28–37 °C. Lactase production was initiated after substrate consumption indicating a reversible enzyme inhibition or catabolic repression. The specific enzyme activity after 45 h was between 456.3 U/g cell mass (37 °C) and 733.3 U/g (34 °C), whereas the highest volumetric activity was obtained at 30 °C: 21.8 U/ml. This is generally consistent with results from other authors that used whey as a carbon source. Ethanol as a by‐product reached its maximum concentration after 10–14 h (31.1–40.5 g/l), but was completely consumed afterwards. A pH of 5.5 without further control gave the best production rate for lactase (484.4 U/l × h). In this process, the pH was stable during cell growth at 5.5 and then went up to pH 7.2 after 45 h. At a fixed pH of 5.5 or 6.5, the production rates achieved 313.3 U/l × h and 233.3 U/ l × h, respectively. These results differed from those of other authors, who suggested a fixed pH at 7.0 using whey as a carbon source. There were no significant differences between inoculum ratios of 1% [v/v] and 10% [v/v] so that 1% is the preferable ratio as it is cheaper. Yeast extract (10 g/l) and peptone (20 g/l) were used as the vitamin and nitrogen source, respectively, for the studies of temperature and pH. These were substituted by corn steep liquor (100 g/l) for inoculum ratio experiments. Production of lactase using sugar cane molasses in a lactose‐free medium gave better enzyme productivity rates than obtained by other authors using whey. The optimum conditions for β‐galactosidase synthesis were a temperature of 30–34 °C and an inoculum ratio of 1% [v/v], an initial pH of 5.5 without any further control or a control of 5.5 during cell growth. Then the pH was raised up to 7.     

L-lactic acid production by Lactobacillus casei fermentation with corn steep liquor-supplemented acid-hydrolysate of soybean meal

Batch and fed-batch fermentation studies were performed to evaluate the potential of corn steep liquor (CSL)-supplemented acid-hydrolysate of soybean meal (AHSM) as an alternative to yeast extract (YE) for the production of L-lactic acid by Lactobacillus casei LA-04-1. The CSL-supplemented AHSM gave an outstanding result in supporting L-lactic acid production from glucose. In the exponential fed-batch fermentation, the concentration, yield and productivity of L-lactic acid were 162.5 g/L, 89.7% and 1.69 g/L per h, respectively, which were lower than those with 20 g/L YE (180 g/L, 90.3%, 2.14 g/L per h) after 96 h of fermentation. However, the raw material cost of the nitrogen resource was estimated as only 25% of that using the YE.     

Characterization and evaluation of corn steep liquid in acetone-butanol-ethanol production by Clostridium acetobutylicum

Corn steep liquor (CSL) obtained from a commercial starch manufacturing process was analyzed and tested as a complex nutrient source for ABE (acetone, butanol, and ethanol) production by Clostridium acetobutylicum PJC4BK_AdhE1. CSL contained a wealth of trace elements and nitrogenous components, proteins and amino acids. ABE production increased the content of CSL was raised up to 6% (v/v) in medium and then decreased at higher contents. In 6% CSL-containing medium, C. acetobutylicum PJC4BK_AdhE1 produced 21.4 g/L of ABE with a yield of 0.41 g/g in 18 h of fermentation. Although the final titer of ABE was similar in CSL containing media and Clostridial Growth Medium (CGM, containing yeast extract and asparagines as complex nutrients), the yield and productivity of ABE in 6% CSL-containing medium were found to be higher than 32 and 26%, respectively.     

Optimization of nutritional requirements for gentamicin production by Micromonospora echinospora

Effect of various fermentation media, carbon sources, nitrogen sources, phosphate concentration and culture requirements includes inoculum levels and age were determined on gentamicin production and biomass dry weight production for Micromonospora echinospora, a gentamicin producing strain. Of the substrates tested, starch as a sole carbon source promoted maximal gentamicin production, while maltose promoted maximal growth. Yeast extract as a sole nitrogen source promoted maximal growth, while soyabean meal for gentamicin production. Increasing phosphate concentration enhanced gentamicin production and observed optimum production at 1.2 g/1 (6% v/v) of phosphate having 72 h old inoculum in the medium. Highest gentamicin production was obtained after cultivation with shaking for 120 h in a medium containing starch 0.75% (w/v), soyabean meal 0.5%, K2HPO4 0.12%, CaCO3 0.4%, FeSO4 0.003% and CoCl2 0.0001%. The gentamicin production was 1.2-fold in this medium as compared to basal medium.     

A high-throughput transient gene expression system for switchgrass (Panicum virgatum L.) seedlings

Background

Fermentations using Escherichia coli KO11, Saccharomyces cerevisiae 424A(LNH-ST), and Zymomonas mobilis AX101 are compared side-by-side on corn steep liquor (CSL) media and the water extract and enzymatic hydrolysate from ammonia fiber expansion (AFEX)-pretreated corn stover.

Results

The three ethanologens are able produce ethanol from a CSL-supplemented co-fermentation at a metabolic yield, final concentration and rate greater than 0.42 g/g consumed sugars, 40 g/L and 0.7 g/L/h (0-48 h), respectively. Xylose-only fermentation of the tested ethanologenic bacteria are five to eight times faster than 424A(LNH-ST) in the CSL fermentation. All tested strains grow and co-ferment sugars at 15% w/v solids loading equivalent of ammonia fiber explosion (AFEX)-pretreated corn stover water extract. However, both KO11 and 424A(LNH-ST) exhibit higher growth robustness than AX101. In 18% w/w solids loading lignocellulosic hydrolysate from AFEX pretreatment, complete glucose fermentations can be achieved at a rate greater than 0.77 g/L/h. In contrast to results from fermentation in CSL, S. cerevisiae 424A(LNH-ST) consumed xylose at the greatest extent and rate in the hydrolysate compared to the bacteria tested.

Conclusions

Our results confirm that glucose fermentations among the tested strains are effective even at high solids loading (18% by weight). However, xylose consumption in the lignocellulosic hydrolysate is the major bottleneck affecting overall yield, titer or rate of the process. In comparison, Saccharomyces cerevisiae 424A(LNH-ST) is the most relevant strains for industrial production for its ability to ferment both glucose and xylose from undetoxified and unsupplemented hydrolysate from AFEX-pretreated corn stover at high yield.     

以玉米浆和木薯为原料机械搅拌发酵制备细菌纤维素的研究

本文以玉米浆和木薯为原料,用机械搅拌式发酵罐制备细菌纤维素（BC）,对发酵过程的纤维素产量、还原糖消耗、溶氧变化和茵浓变化进行了监测,并以葡萄糖一蛋白胨-酵母粉培养基为对照进行了比较。实验得出玉米浆作氮源时不溶BC的产量为9．2g／L,而氮源成本只是对照组的15％;木薯水解液作碳源时的不溶BC产量达到11．7g／L,比对照组（10．8g／L）高8％;而用玉米浆搭配木薯水解液发酵生产BC,产量也达到10．1g／L,验证了这两种天然原料的廉价高效性,用于工业生产细菌纤维素具有良好的前景。     

A media design program for lactic acid production coupled with extraction by electrodialysis

The aim of this study was to investigate industrial media for lactic acid fermentation to reduce the cost of nitrogen sources. Corn steep liquor (CSL) was successfully used at 5% (v/v) in batch fermentations. Use of soluble CSL improved the productivity approximately 20% with an advantage of clearer fermentation broth. Yeast extract (YE)-complemented CSL media further increased the productivity. It was found that 3.1 g L(-1) yeast extract and 5% CSL could be an effective substitute for 15 g L(-1) yeast extract in 10% glucose medium. Spent brewery yeast was also used as a sole nitrogen source equivalent to 5% CSL. Lactic acid was recovered by electrodialysis from the cell free broth. Depleted cell free broth supplemented with 5 g L(-1) of yeast extract performed reasonably in batch cultures. Reuse of the fermentation broth may reduce the cost of raw materials as well as minimize the fermentation wastes.     

Direct and efficient ethanol production from high-yielding rice using a Saccharomyces cerevisiae strain that express amylases

Efficient ethanol producing yeast Saccharomyces cerevisiae cannot produce ethanol from raw starch directly. Thus the conventional ethanol production required expensive and complex process. In this study, we developed a direct and efficient ethanol production process from high-yielding rice harvested in Japan by using amylase expressing yeast without any pretreatment or addition of enzymes or nutrients. Ethanol productivity from high-yielding brown rice (1.1g/L/h) was about 5-fold higher than that obtained from purified raw corn starch (0.2g/L/h) when nutrients were added. Using an inoculum volume equivalent to 10% of the fermentation volume without any nutrient supplementation resulted in ethanol productivity and yield reaching 1.2g/L/h and 101%, respectively, in a 24-h period. High-yielding rice was demonstrated to be a suitable feedstock for bioethanol production. In addition, our polyploid amylase-expressing yeast was sufficiently robust to produce ethanol efficiently from real biomass. This is first report of direct ethanol production on real biomass using an amylase-expressing yeast strain without any pretreatment or commercial enzyme addition.     

产酸丙酸杆菌生长特性及5L罐发酵动力学研究

论文在摇瓶水平对产酸丙酸杆菌基本生长特性（温度、pH、摇床转速、接种量、种龄等）、碳源、氮源利用情况、产物抑制及5 L罐发酵动力学进行了研究。结果表明,该菌在32℃,初始pH 6．5,摇床转速150 r/min,接种24 h的种子液,接种量为5％条件下,产酸丙酸杆菌生长及产酸水平达最高值;该菌可利用碳源十分广泛,但对氮源要求比较高,只可利用有机氮源;在不同初始葡萄糖浓度下,产酸丙酸杆菌生长及产酸水平差异不大,无明显底物抑制现象;在2g/L的初始丙酸盐浓度下,该菌生长受到明显抑制;在5L发酵罐中,初始葡萄糖浓度为58．8 g/L,发酵72 h,葡萄糖消耗完全,丙酸终浓度达22．4 g/L,丙酸得率和产率分别达0．381 g/g和0．295 g/（L·h）,丙酸占总酸比例达72．10％。     

Improvement of ethanol yield from raw corn flour by <Emphasis Type="Italic">Rhizopus</Emphasis> sp.

A novel raw-starch-digesting glucoamylase producer, Rhizopus sp. W-08, and Saccharomyces cerevisiae Z-06 were used in a fed batch process for simultaneous saccharification and fermentation of raw corn flour. Ethanol concentration of 21% (v/v) was obtained after 48 h. The conversion efficiency of raw corn flour to ethanol was 94.5% of the theoretical ethanol yield.     

Improved production of curdlan with concentrated cells ofAgrobacterium sp.

The addition of a limited concentration of yeast extract to a minimal salt medium (MSM) enhanced cell growth and increased the production of curdlan whereas nitrogenlimitation was found to be essential for the higher production of curdlan byAgrobacterium sp. ATCC 31749. As the amount of the inoculum increased, the cell growth as well as the production of curdlan also increased in the MSM without a nitrogen source. The cell growth and production of curdlan increased as the initial pH of the medium decreased as low as 5.0. The conversion rate and concentration of curdlan from 2% (w/v) glucose in the MSM with concentrated cells under nitrogen deletion was 67% and 13.4 g/L, respectively. The highest conversion rate of curdlan under the conditions optimized in this study was 71% when the glucose concentration was 1% (w/v).     

Optimization of fermentation conditions for ethanol production from whey

Summary Optimal conditions for ethanol production in 7% whey solutions by the yeast Candida pseudotropicalis ATCC 8619 included initial pH of 4.57 and 30°C. Complete fermentation of the available lactose took place without supplementary nutrients; additions of nitrogen or phosphorus salts, yeast extract or corn steep liquor resulted in increased yeast production and lower ethanol yields. A positive correlation was observed between increases in yeast inocula and lactose utilization and ethanol production rates; 8.35 g/l of ethanol was obtained within 22 h by using yeast inoculum of 13.9 g/l. No differences in fermentation rates or ethanol yields were observed when whole or deproteinized whey solutions were used. Concentrated whey permeates, obtained after removal of the valuable proteins from whey, can be effectively fermented for ethanol production.