Development of an ethanol/sorbitol process using a ceramic membrane cell recycle bioreactor

The properties of a ceramic microfiltration membrane unit have been investigated as a component of a cell recycle bioreactor used for the production of ethanol/sorbitol. The conversion is carried out by a glucose/fructose oxidoreductase (and bound co-factor NADP) using cells of Zymomonas mobilis and an equimolar mixture of glucose and fructose. With 100 g/l each of glucose and fructose, a product stream containing 40 g/l sorbitol and 75 g/l ethanol was obtained. The potential economic benefits of such a process are discussed.     

Wood blocks as a carrier for Saccharomyces Cerevisiae used in the production of ethanol and fructose

Saccharomyces cerevisiae ATCC 39859 was immobilized onto small cubes of wood in order to produce very enriched fructose syrup from synthetic glucose-fructose mixtures, through the selective fermentation of glucose. The kinetics of growth and ethanol production rates were studied. Several tests to assess the influence of substrate and product concentration on the production rates were carried out and appropriate rate equations were proposed as a design basis for continuous immobilized reactors. The ethanol production rate and cell growth rate were found to be inhibited linearly by both substrate and product concentrations. A maximum ethanol productivity of 21.9 g 1⁻¹ h⁻¹ was attained from a feed containing 10% (by weight) glucose and 10% (by weight) fructose. The ethanol concentration was 29.6 g 1⁻¹, the glucose conversion was 78% and a fructose yield of 99% was obtained. This resulted in a final fructose:glucose ratio of 2.7. At lower ethanol productivity levels the fructose:glucose ratio increased, as did the ethanol concentration in the effluent. The ethanol productivities obtained in this study were 33%–132% higher than those obtained in a previous study using the same system, under similar conditions, with the cells immobilized in alginate beads.     

Pure fructose syrup and ethanol production from high fructose corn syrup supplemented with Jerusalem artichoke juice

Saccharomyces cerevisiae ATCC 36859 preferentially consumes glucose from glucose–fructose mixtures. Synthetic media and complex media containing high fructose corn syrup supplemented with Jerusalem artichoke juice were used for the production of pure fructose syrup by the conversion of glucose to ethanol. Fructose was not converted in these processes. Increasing the concentration of Jerusalem artichoke juice increased the yields of ethanol and biomass and decreased the process time. A similar effect was obtained at a low juice concentration when a larger amount of biomass was used for the inoculum. The product from this process contained only fructose and ethanol. Use of food-grade materials results in a pure fructose syrup that is suitable for human consumption.     

Utilization of raffinose and melibiose by a mutant of Saccharomyces cerevisiae

The mutant Saccharomyces cerevisiae ATCC 36858 was used in the production of ethanol and/or fructose from synthetic media in batch processes with raffinose, melibiose or sucrose. The mutant was able to hydrolyze all the sugars used and to selectively ferment glucose and galactose to ethanol while fructose accumulated in the fermentation medium. The fructose yield was above 89% of the theoretical value in the media with either raffinose or sucrose, when initial concentrations were between 131.5 g dm^?3 and 242.0 g dm^?3. The ethanol yields were 82% and 77% of the theoretical values in the media with melibiose and sucrose, respectively, and about 72% of the theoretical value when raffinose was used. The fructose fraction in the carbohydrate content of the produced syrups was more than 96% when raffinose concentration was below 189.1 g dm^?3. However, even at a sucrose concentration of 187.9 g dm^?3, the produced syrup contained 100% fructose. Some oligosaccharides were also produced in all tested media. The produced oligosaccharides were consumed by the end of the fermentation process. These findings can be useful in the production of ethanol and high fructose syrups using raw materials based on sucrose and raffinose such as molasses. Copyright © 2003 Society of Chemical Industry     

Xylitol production by different yeasts: Kinetic study and biosynthesis from cashew apple bagasse hydrolysate

Xylitol is a polyol with sweetening and anti-cariogenic properties and is highly relevant to the food and pharmaceutical industries. This work evaluates xylitol production by three yeasts (Candida tropicalis, Kluyveromyces marxianus CCA510, and Kluyveromyces marxianus ATCC 36907) from different carbon sources: D-xylose (medium xylose [MX]) and D-xylose plus D-glucose (medium xylose and glucose [MXG]). The potential of xylitol production from hemicellulose hydrolysate of cashew bagasse was evaluated. In MX medium, K. marxianus CCA510 was the strain that produced higher xylitol concentration (17.04 g · L⁻¹). However, K. marxianus ATCC 36907 and C. tropicalis produced 13.22 and 9.54 g · L⁻¹, respectively. On the other hand, in MXG medium, probably due to the presence of glucose as a carbon source, lower xylitol production was observed for all microorganisms: the CCA510 strain produced 13.30 g · L⁻¹ of xylitol, while C. tropicalis and ATCC 36907 produced 11.42 and 0.64 g · L⁻¹, respectively. Additionally, the production of ethanol as a secondary product was also noted. According to the results of the kinetic study, xylitol formation is associated with the growth and consumption of substrate (xylose), which is a typical behaviour of a primary metabolite for the three yeasts. Furthermore, the strain K. marxianus CCA510 was able to produce xylitol from cashew apple bagasse hydrolysate (CABH), evidencing its potential for use in bioprocesses related to biorefinery. In view of the results reported here, it was possible to clarify in detail the kinetics of xylitol production by the yeast strains evaluated, which had the ability to produce xylitol from CABH, providing added value to this agro-industrial residue.     

The Role of Hexokinase and Hexose Transporters in Preferential Use of Glucose over Fructose and Downstream Metabolic Pathways in the Yeast Yarrowia lipolytica

The development of efficient bioprocesses requires inexpensive and renewable substrates. Molasses, a by-product of the sugar industry, contains mostly sucrose, a disaccharide composed of glucose and fructose, both easily absorbed by microorganisms. Yarrowia lipolytica, a platform for the production of various chemicals, can be engineered for sucrose utilization by heterologous invertase expression, yet the problem of preferential use of glucose over fructose remains, as fructose consumption begins only after glucose depletion what significantly extends the bioprocesses. We investigated the role of hexose transporters and hexokinase (native and fructophilic) in this preference. Analysis of growth profiles and kinetics of monosaccharide utilization has proven that the glucose preference in Y. lipolytica depends primarily on the affinity of native hexokinase for glucose. Interestingly, combined overexpression of either hexokinase with hexose transporters significantly accelerated citric acid biosynthesis and enhanced pentose phosphate pathway leading to secretion of polyols (31.5 g/L vs. no polyols in the control strain). So far, polyol biosynthesis was efficient in glycerol-containing media. Moreover, overexpression of fructophilic hexokinase in combination with hexose transporters not only shortened this process to 48 h (84 h for the medium with glycerol) but also allowed to obtain 23% more polyols (40 g/L) compared to the glycerol medium (32.5 g/L).     

Production of fructose and ethanol from media with high sucrose concentrations by a mutant of Saccharomyces cerevisiae

The production of enriched fructose syrups and ethanol from a synthetic medium with high sucrose concentrations was studied in a batch process using Saccharomyces cerevisiae ATCC 36858. The results showed that the fructose yield was above 92% of theoretical values in synthetic media with sucrose concentrations between 180 g dm^?3 and 726 g dm^?3. Ethanol yield was about 82% in media with sucrose concentrations up to 451 g dm^?3. A product containing 178 g dm^?3 fructose, which represents 97% of the total sugar content, and 79 g dm^?3 ethanol was obtained using a medium with 360 g dm^?3 sucrose. The fructose fraction in the carbohydrates content in the produced syrups decreased with increases in the initial sucrose concentration. In a medium with initial sucrose concentration of 574 g dm^?3, the fructose content in the produced broth was 59% of the total carbohydrates. Glycerol and fructo‐oligosaccharides were also produced in this process. The produced fructo‐oligosaccharides started to be consumed when the concentration of sucrose in the media was less than 30% of its initial value. Complete hydrolysis of these sugars was noticed in media with sucrose concentrations below 451 g dm^?3. These findings will be useful in the production of ethanol and high fructose syrups using sucrose‐based raw materials with high concentrations of this carbohydrate. © 2001 Society of Chemical Industry     

Effect of extraction and enzymatic esterification of ethanol on glucose consumption by two Saccharomyces cerevisiae strains: a comparative study

Extractive alcoholic fermentations of high glucose concentrations (300 and 400 g dm^?3) using a flocculent (saké) and a non‐flocculent (DER24) Saccharomyces cerevisiae strain were compared. The introduction of a Rhizomucor miehei lipase, in the extractive fermentations of 300 g dm^?3 of glucose, increased the ethanol extraction due to its esterification with oleic acid, allowing complete glucose consumption at an organic solvent/fermentation medium phase ratio of 1. In these conditions, an increase of ethanol yield was observed. Total glucose consumption was also obtained in enzymatic extractive fermentations of 400 g dm^?3 of glucose, but only when oleic acid was added at the exponential growth phase. From the comparison of the extractive fermentation performances obtained using the two yeast strains it was observed that the flocculent strain led to a lower glucose metabolisation rate. This behaviour was related to the highest diffusional limitations that occur in the presence of flocs. The developed processes showed that the association of alcoholic fermentation with enzymatic extraction led to the reduction of inhibitory effects as well as to the simultaneous production of fatty esters which are compounds with several commercial applications. © 2001 Society of Chemical Industry     

Biosynthesis of lipids and organic acids by Yarrowia lipolytica strains cultivated on glucose

The biochemical behavior of wild‐type or genetically modified (presenting decreased expression of intracellular acyl‐CoA oxidases) Yarrowia lipolytica strains cultivated on commercial glucose was studied. Flask nitrogen‐limited cultures were performed favoring the production of organic acids (and potentially the accumulation of lipid). Nitrogen depletion induced secretion of citric acid, while intracellular lipid was not produced in high quantities. Maximum total citric acid up to 49 g/L (yield 0.85 g/g glucose) was produced. In some of the wild‐type strains, an increase of glucose in the medium also induced noticeable production of acetic acid. Increasing the amount of added glucose led to an increase in the total lipid quantity (%) produced, although in the stationary growth phase the concentration of lipid declined, indicating lipid degradation even for the genetically modified strains. Total lipid amount did not exceed the value of 14%, while neutral fractions increased with increase in glucose concentration. In all cases, the total microbial lipids and major lipid fractions were composed of C₁₆ and C₁₈ (principally unsaturated) fatty acids. Finally, in several of the strains cultured in media containing a low glucose concentration, unicellular morphology was observed, while at high glucose concentrations mycelia were predominant.     

Production,thermal stability and immobilisation of inulinase from Fmarium oxysporum

Fusarium oxysporum produced maximum extracellular inulinase after 9 days of its growth at 25°C on a medium (pH 5.5) containing 3% fructan and 0.2% sodium nitrate. The level of this enzyme decreased on the addition of either glucose, fructose, galactose or sucrose to F. oxysporum already growing on a fructan-containing medium. A significant increase in invertase production which resulted in an increase of the invertase/inulinase (S/I) ratio, was observed on addition of inulin to this fungus growing on other carbon sources. Glycerol (10%) gave better protection to inulinase against thermal denaturation at 50°C compared to ethylene glycol and sorbitol. Inulinase immobilised in polyacrylamide gel retained 45% of its original activity. The immobilised enzyme showed a higher optimum temperature (45°C) compared to free enzyme (37°C). The immobilised enzyme after storage at 25°C for 96 h showed 58% activity. Thermal stability of entrapped inulinase increased in the presence of inulin.     

Mutation Breeding of Extracellular Polysaccharide-Producing Microalga Crypthecodinium cohnii by a Novel Mutagenesis with Atmospheric and Room Temperature Plasma

Extracellular polysaccharides (EPS) produced by marine microalgae have the potential to be used as antioxidants, antiviral agents, immunomodulators, and anti-inflammatory agents. Although the marine microalga Crypthecodinium cohnii releases EPS during the process of docosahexaenoic acid (DHA) production, the yield of EPS remains relatively low. To improve the EPS production, a novel mutagenesis of C. cohnii was conducted by atmospheric and room temperature plasma (ARTP). Of the 12 mutants obtained, 10 mutants exhibited significantly enhanced EPS yield on biomass as compared with the wild type strain. Among them, mutant M7 was the best as it could produce an EPS volumetric yield of 1.02 g/L, EPS yield on biomass of 0.39 g/g and EPS yield on glucose of 94 mg/g, which were 33.85%, 85.35% and 57.17% higher than that of the wild type strain, respectively. Results of the present study indicated that mutagenesis of the marine microalga C. cohnii by ARTP was highly effective leading to the high-yield production of EPS.     

Kinetic Study and Mathematical Modelling of the Growth of S. cerevisiae 522D in Presence of K2 Killer Protein

This paper deals with the study of the kinetics of batch fermentations for a sensitive strain of S. cerevisiae growing in a culture medium containing K2 killer protein. The inhibition due to the killer toxin was measured by the reduction of the viable biomass, ethanol production and glucose consumption compared with a reference fermentation. The reference culture was run under the same conditions, but using a heat-denatured solution of killer protein. Results showed a decrease of the viable population of 67·8% after 15 h incubation. The fermentation time for the total consumption of the glucose was significantly affected by the presence of the killer toxin. The specific rate of ethanol production was also affected during the fermentation. However, the yields of ethanol and biomass were very similar for both fermentations. A kinetic mathematical model was proposed to explain the dynamics of the S. cerevisiae 522D growth in the presence of the K2 killer protein. The results of the suggested simulation were in agreement with the experimental data. © 1997 SCI.     

Production of L‐lactic acid with very high gravity distillery wastewater from ethanol fermentation by a newly isolated Enterococcus hawaiiensis

BACKGROUND: A great amount of wastewater with high contents of chemical oxygen demand (COD) are produced by ethanol production. It would be useful to utilize distillery wastewater to produce L‐lactic acid, which could be a high additional value byproduct of ethanol production. The fermentation process of L‐lactic acid production by a newly isolated Enterococcus hawaiiensis CICIM‐CU B0114 is reported for the first time. RESULTS: The strain produced 56 g L^?1 of L‐lactic acid after cultivation for 48 h in optimized medium consisting of (g L^?1) 80 glucose, 10 peptone, 10 yeast extract, 1.5 Na₂HPO₄ and 0.2 MgSO₄. E. hawaiiensis CICIM‐CU B0114 was isolated and purified by subculture for growing and producing L‐lactic acid in distillery wastewater of very high gravity (VHG) from ethanol fermentation. L‐lactic acid fermentation was further studied with distillery wastewater substrate in 7 L and 15 L fermentors. The results showed that L‐lactic acid concentrations of 52 g L^?1 and 68 g L^?1 was achieved in 7 L and 15 L fermentors with the initial sugar concentrations of 67 g L^?1 and 87 g L^?1, respectively. CONCLUSION: The production of L‐lactic acid by the newly isolated E. hawaiiensis CICIM‐CU B0114 was carried out and the fermentation medium was optimized by orthogonal experimental design. This new strain holds the promise of L‐lactic acid production utilizing distillery wastewater from VHG ethanol fermentation. Copyright © 2010 Society of Chemical Industry     

碳氮源及pH调控对假肠膜明串珠菌产甘露醇的影响

为了降低生物法制备甘露醇的成本,以假肠膜明串珠菌Leuconostoc pseudomesenteroides G123为研究对象,对培养基中的氮源、葡萄糖与果糖比例和pH调控过程进行了优化,提高了果糖转化率和甘露醇产量。5L发酵罐中结果显示:采用2g/L的酵母粉作为单一氮源,葡萄糖和果糖的比例为0.35:1,初始pH值7.5,发酵过程中保持pH值不低于4.5,甘露醇产量可达57.24g/L,甘露醇对果糖的转化率为83.2%。该过程副产D-乳酸20.32g/L,其光学纯度达99.9%,具有回收价值,甘露醇与D-乳酸对糖总转化率为89.38%,有助于降低生物法制备甘露醇的成本。     

Direct ethanol production from rice straw by coculture with two high-performing fungi

To develop efficient and economical direct ethanol production from fine rice straw crashed mechanically, two high-performing fungi, which can secret hyperactive cellulases and/or ferment effectively various sugars, were selected from some strains belong to Mucor circinelloides preserved in our laboratory. The simultaneous saccharification and fermentation (SSF) by coculture with these fungi was investigated. The screening of high-performing fungi resulted in the selection of NBRC 4572 as an ethanol-producing fungus and NBRC 5398 as a cellulase-secreting fungus. The strain 4572 produced ethanol aerobically from glucose and xylose in high yields of 0.420 g/g at 36 h and 0.478 g/g at 60 h, respectively, but secreted fairly low cellulases. On the other hand, the strain 5398 also produced ethanol from glucose in yield of 0.340 g/g though it had a little growth in xylose culture. However, it secreted hyperactive cellulases that are essential for hydrolysis of rice straw in culture and the maximum activities of endo-β-glucanase and β-glucosidase were 2.11 U/L and 1.47 U/L, respectively. In SSF of rice straw by coculture with two fungi selected, the ethanol production reached 1.28 g/L after 96 h when the inoculation ratio of the strain 5398 to the strain 4572 was 9.     

成对电合成葡萄糖酸及山梨醇的研究

采用成对合成新工艺电解葡萄糖制取葡萄糖酸及山梨醇 ,研究了电流、温度、介质浓度、葡萄糖浓度及电解时间对葡萄糖电解过程的影响。结果表明 ,在电流密度为 1mA/cm2 ,温度为 2 5℃ ,c(硫酸 ) =0 .30mol/L介质中 ,浓度为 3.6× 10 -2 mol/L的葡萄糖电解 90min后 ,其转化率可达 90 %。     

Genetic Variation in Primary Metabolites of Pastinaca sativa; Can Herbivores Act as Selective Agents?

Although insect herbivory has been shown to act as a selective agent on plant secondary metabolism, whether primary metabolites contribute to resistance and can respond to selection by herbivores remains untested. In the wild parsnip (Pastinaca sativa), its principal herbivore, Depressaria pastinacella, acts as a selective agent on furanocoumarin resistance factors. In this study, we determined whether webworms can, by causing differential reductions in fitness, act as selective agents on parsnip primary metabolites. Estimates of narrow-sense heritabilities were significantly different from zero for C18 fatty acids in buds and developing fruits, fructose and sorbitol in buds, fructose, myo-inositol, bergapten, and psoralen in fruits. Wild parsnips protected from webworms by insecticide produced 2.5 times as much seed biomass as unsprayed plants; that webworms accounted for this difference in plant fitness was indicated by a significant negative relationship between reproductive effort and an index of webworm damage. Only a handful of metabolites influenced resistance to webworms; these included osthol, sorbitol, and protein in developing fruits as well as previously documented furanocoumarins. Osthol, a coumarinic compound, enhanced resistance, as did protein content, while sorbitol lowered resistance. Other primary metabolites may affect resistance to webworms, but their effect was context-dependent, that is, their effect depended on concentrations of other metabolites (epistasis). Susceptible plant phenotypes were found to have average chemical compositions. Although there was genetic variation in some of the primary metabolites in parsnips, the epistatic nature of their involvement in resistance and the lack of genetic variation in some suggest that selection on them from webworms will be either inconsistent or ineffective.     

过表达谷氧还蛋白基因GRX5提高酿酒酵母乙酸耐性

利用可再生的纤维素原料生产燃料乙醇是国内外研究的热点。但纤维素原料一些预处理过程产生的乙酸对酿酒酵母细胞生长和乙醇发酵产生强烈抑制,因此,提高酿酒酵母细胞的乙酸耐受性是提高纤维素乙醇发酵效率的重要手段。本文研究了谷氧还蛋白家族中GRX5p的编码基因的过表达对酿酒酵母在乙酸胁迫条件下细胞生长和发酵性能的影响。结果表明,过表达GRX5的重组菌株在含有5 g·L^-1乙酸的平板中生长优于对照菌株;在含有5 g·L^-1乙酸的培养基中进行乙醇发酵,过表达GRX5的重组菌株可在48 h基本消耗培养基中所有的葡萄糖,发酵周期比对照菌株缩短了12 h。过表达GRX5菌株的乙醇生产强度为0.897 g·L^-1·h^-1,比对照提高了28.5%。代谢物分析结果表明,过表达GRX5的重组菌株可产生更多的保护性物质海藻糖和甘油,有利于增强菌株胁迫耐受性。     

成对电解同时合成甘露醇、山梨醇和葡萄糖酸盐

采用成对电解合成技术 ,在阴极电还原葡萄糖为甘露醇和山梨醇 ,葡萄糖的转化率达74 6 % ;在阳极电氧化葡萄糖为葡萄糖酸盐 ,其中葡萄糖酸钠的收率为 90 3 % ,电解槽的总收率为 16 4 9%。     

Mathematical model for ethanol production from mixed sugars by Pichia stipitis

A mathematical model for estimating the dynamic behavior of ethanol production from mixed sugars such as glucose and xylose is presented. This model was constructed by introducing the term for ethanol production into the diauxic growth model proposed previously by the authors. It was assumed that the first substrate, glucose, is metabolized by a constitutive enzyme and the second substrate, xylose, is utilized by an inducible enzyme. The synthesis of the inducible enzyme is controlled by the catabolite repression caused by glucose and with the induction caused by xylose as an inducer, and the xylose is then converted into cell mass and ethanol by the inducible enzyme. The parameters of the model were estimated from the experimental data in the medium containing glucose and xylose, singly or in combination in a batch culture. The application of the model was examined for experiments in both batch and continuous culture, with glucose and xylose as carbon sources. The calculated values, according to the model, corresponded satisfactorily with experimental data, such as cell growth, substrate consumption, and ethanol production, especially in the estimation of the lag times between the first log phase and the second log phase in cell growth and ethanol production. © 2001 Society of Chemical Industry