Simultaneous saccharification and fermentation of wheat bran flour into ethanol using coculture of amylotic Aspergillus niger and thermotolerant Kluyveromyces marxianus

Studies on simultaneous saccharification and fermentation (SSF) of wheat bran flour, a grain milling residue as the substrate using coculture method were carried out with strains of starch digesting Aspergillus niger and nonstarch digesting and sugar fermenting Kluyveromyces marxianus in batch fermentation. Experiments based on central composite design (CCD) were conducted to maximize the glucose yield and to study the effects of substrate concentration, pH, temperature, and enzyme concentration on percentage conversion of wheat bran flour starch to glucose by treatment with fungal α-amylase and the above parameters were optimized using response surface methodology (RSM). The optimum values of substrate concentration, pH, temperature, and enzyme concentration were found to be 200 g/L, 5.5, 65°C and 7.5 IU, respectively, in the starch saccharification step. The effects of pH, temperature and substrate concentration on ethanol concentration, biomass and reducing sugar concentration were also investigated. The optimum temperature and pH were found to be 30°C and 5.5, respectively. The wheat bran flour solution equivalent to 6% (w/V) initial starch concentration gave the highest ethanol concentration of 23.1 g/L after 48 h of fermentation at optimum conditions of pH and temperature. The growth kinetics was modeled using Monod model and Logistic model and product formation kinetics using Leudeking-Piret model. Simultaneous saccharificiation and fermentation of liquefied wheat bran starch to bioethanol was studied using coculture of amylolytic fungus A. niger and nonamylolytic sugar fermenting K. marxianus.     

Effect of different fermentation parameters on bioethanol production from corn meal hydrolyzates by free and immobilized cells of Saccharomyces cerevisiae var. ellipsoideus

BACKGROUND: Bioethanol produced from renewable biomass, such as corn meal, is a biofuel that is both renewable and environmentally friendly. Significant scientific and technological investments will be needed to achieve substitution of conventional fossil fuels with alternative fuels. The ethanol fermentation of enzymatically obtained corn meal hydrolyzates by free and immobilized cells of Saccharomyces cerevisiae var. ellipsoideus yeast in a batch system was studied. The initial glucose and inoculum concentration and the time required for the efficient ethanol production were optimized taking into account parameters such as ethanol concentration, ethanol yield, percentage of the theoretical yield of ethanol and volumetric productivity in both immobilized and free cell systems. RESULTS: The yeast cells were immobilized in Ca–alginate by an electrostatic droplet generation method. An optimal initial inoculum concentration of 2% (v/v) and optimal fermentation time of 38 h for both immobilized and free yeasts were determined. An optimal initial glucose concentration of 150 g L^?1 for free system was achieved. At the initial glucose concentration of 176 g L no substrate or product inhibition were achieved with immobilized yeast. CONCLUSION: By immobilization of the yeast into Ca–alginate using the method of electrostatic droplet generation a superior system was realized, which exhibited lower substrate inhibition and higher tolerance to ethanol. The cells of S. cerevisiae var. ellipsoideus yeast entrapped in Ca–alginate showed good physical and chemical stability, and no substrate and product diffusion restrictions were noticed. Copyright © 2008 Society of Chemical Industry     

Production of bioethanol from corn meal hydrolyzates

The two-step enzymatic hydrolysis of corn meal by commercially available α-amylase and glucoamylase and further ethanol fermentation of the obtained hydrolyzates by Saccharomyces cerevisiae yeast was studied. The conditions of starch hydrolysis such as substrate and enzyme concentration and the time required for enzymatic action were optimized taking into account both the effects of hydrolysis and ethanol fermentation. The corn meal hydrolyzates obtained were good substrates for ethanol fermentation by S. cerevisiae. The yield of ethanol of more than 80% (w/w) of the theoretical was achieved with a satisfactory volumetric productivity P (g/l h). No shortage of fermentable sugars was observed during simultaneous hydrolysis and fermentation. In this process, the savings in energy by carrying out the saccharification step at lower temperature (32 °C) could be realized, as well as a reduction of the process time for 4 h.     

Modulated gluconic acid production from immobilized cells of Aspergillus niger ORS‐4.410 utilizing grape must

BACKGROUND: Gluconic acid (GA) production by immobilized cells of mutant Aspergillus niger ORS‐4.410 on polyurethane sponge (PUS) and calcium‐alginate (Ca‐alginate) was evaluated in repeated batches of solid state surface fermentation (SSF) and submerged fermentation (SmF) conditions, respectively, utilizing rectified grape must as carbon source. RESULTS: The passive immobilization of cells in fermentation medium solid support of having 0.4 cm³ cube size, 4% spore suspension, 0.6 g inoculum of PUS immobilized cells at 32 °C and 2.0 L min^?1 resulted in the maximum GA production (88.16 g L^?1) with a 92.8% yield, while the Ca‐alginate matrix with a 0.5 cm diameter bead size, 2–3% spore suspension, 15 g inoculum at 34 °C and 150 rpm agitation speed revealed 67.19 g L^?1 GA with a 85.2% yield. Repeated use of PUS showed higher levels of GA (110.94 g L^?1) in the third–fourth fermentation cycles with 95–98% yield and 22.50 g L^?1 d^?1 productivity under SSF that was 2.5‐fold higher than the productivity obtained from a typical fermentation cycle, and 54% greater than the productivity obtained with repetitive use of Ca‐alginate immobilized cells of A. niger under SmF. CONCLUSION: Using immobilized cells of A. niger in PUS, the rectified form of grape must can be utilized for GA production as an alternative source of carbohydrate by replacing the conventional fermentation conditions. Copyright © 2008 Society of Chemical Industry     

Enzymatic Hydrolysis of Corn Stover after Pretreatment with Dilute Sulfuric Acid

Although many previous studies have been carried on the enzymatic hydrolysis of corn stover after pretreatment with dilute sulfuric acid, this paper emphasizes the use of different conditions to attain the highest yields of two sugars, xylose and glucose, from both stages. The pretreatment was performed at a range of sulfuric acid concentrations of 2, 4 and 6 % at 80, 100 and 120 °C. Up to 77 % xylose yield was obtained while the glucose yield was only 8.4 %. The corresponding solid phase was hydrolyzed by cellulase and the influences of five factors and their interactions on enzyme hydrolysis were evaluated by response surface methodology based on one‐factor‐at‐a‐time experiments. The optimal levels for each variable to obtain the highest reducing sugar yield were as follows: enzyme concentration of 22 FPU/g substrate, substrate concentration of 77 g/L, temperature of 49 °C, pH 4.8 and reaction time of 38 h. A reducing sugar yield of 42.11 g/100 g substrate was achieved, which was consistent with the predicted value of 42.13 g/100 g substrate. Compared with the one‐factor‐at‐a‐time experiments, there was a 9.4 % increase in reducing sugar yield when the enzyme concentration was decreased to 3 FPU/g substrate, the substrate concentration increased to 17 g/L and the reaction time dropped to 22 h. The total sugar released from the two stages was 62.81 g/100 g substrate.     

Production of L-aminoacylase by fermentation of Pseudomonas sp. BA2

The growth and enzymatic production of Pseudomonas sp. BA2 a new L -aminoacylase-producing microorganism, were studied in a bench-top fermenter. Multiple fermentations were carried out in order to determine the optical pH and temperature values. The influence of the substrate concentration on both growth and L aminoacylase activity was also investigated. The maximum growth rate and the greatest yield of enzyme were obtained when the fermentation was carried out at pH 7·5, 25°C and DOT ≥ 50%. N-Acetyl-DL -alanine, at a concentration 20 g dm^?3, was used as the sole carbon and nitrogen source. The fermentation process provided a maximum biomass concentration of 3·36 g dry weight dm^?3. The highest L -aminoacylase production (11429 U g^?1 dry weight) was obtained after 39 h of cultivation. The results were a significant improvement over those previously reported.     

OPTIMIZATION OF PROCESS CONDITIONS USING RESPONSE SURFACE METHODOLOGY (RSM) FOR ETHANOL PRODUCTION FROM WASTE CASHEW APPLE JUICE BY ZYMOMONAS MOBILIS

Ethanol has been known for a long time, being perhaps the oldest product obtained through traditional biotechnology. It is an attractive, sustainable energy source for fuel additives. Based on a four-level central composite design (CCD) involving the variables substrate composition (20–100%) X₁, pH (4.5–6.5) X₂, incubation temperature (28°–36°C) X₃, and fermentation time (12–60 h) X₄, a response surface methodology (RSM) for the production of ethanol using waste cashew apple juice as substrate by Zymomonas mobilis MTCC 090 was standardized. The design contains a total of 31 experimental trials with the first 16 organized in a fractional factorial design and 25 to 31 involving the replications of the central points. Data obtained from RSM on ethanol production were subjected to the analysis of variance (ANOVA) and analyzed using a second-order polynomial equation, which resulted in the optimized process conditions of 62% (v/v) as substrate concentration, pH of 5.5, temperature of 32°C, and fermentation time of 37 h. Maximum ethanol concentration (12.64 g/L) was obtained at the optimized conditions in an anaerobic batch fermentation.     

Enzymatic hydrolysis and fermentation of lime pretreated wheat straw to ethanol

BACKGROUND: The objective of this work is to develop an efficient pretreatment method that can help enzymes break down the complex carbohydrates present in wheat straw to sugars, and to then ferment of all these sugars to ethanol. RESULTS: The yield of sugars from wheat straw (8.6%, w/v) by lime pretreatment (100 mg g^?1 straw, 121 °C, 1 h) and enzymatic hydrolysis (45 °C, pH 5.0, 120 h) using a cocktail of three commercial enzyme preparations (cellulase, β‐glucosidase, and xylanase) at the dose level of 0.15 mL of each enzyme preparation g^?1 straw was 568 ± 13 mg g^?1 (82% yield). The concentration of ethanol from lime pretreated enzyme saccharified wheat straw (78 g) hydrolyzate by recombinant Escherichia coli strain FBR5 at pH 6.5 and 35 °C in 24 h was 22.5 ± 0.6 g L^?1 with a yield of 0.50 g g^?1 available sugars (0.29 g g^?1 straw). The ethanol concentration was 20.6 ± 0.4 g L^?1 with a yield of 0.26 g g^?1 straw in the case of simultaneous saccharification and fermentation by the E. coli strain at pH 6.0 and 35 °C in 72 h. CONCLUSION: The results are important in choosing a suitable pretreatment option for developing bioprocess technologies for conversion of wheat straw to fuel ethanol. Copyright © 2007 Society of Chemical Industry     

Preparation of D-lysine by chemical reaction and microbial asymmetric transformation

The DL-lysine crystals from the racemization of L-lysine was treated as substrate with Hafnia alvei AS1.1009 intact cells as biocatalysts to produce crystalline D-lysine with a yield of 56.6% from the reaction mixture after simple purification. In the presence of 0.10 molar equivalent of salicylaldehyde, L-lysine racemization can be completed within 4 h in 1.0 mol/L of NaOH at 100°C. The activation energy of the processes was 62187.86 J/mol. The characteristics of Hafnia alvei AS1.1009 decarboxylase were studied. Under the conditions of pH 8.0, temperature 37°C, cell concentration 10 g/L, tween-80 0.5 g/L, substrate concentration 30 g/L, and the specific activity of up to 3840 U, L-lysine can be completely degraded by the decarboxylase for 12 h under the optimal conditions. __________ Translated from Modern Chemical Industry, 2007, 27(5): 32–34 [译自: 现代化工]     

FED-BATCH SIMULTANEOUS SACCHARIFICATION AND FERMENTATION OF MICROWAVE/ACID/ALKALI/H2O2 PRETREATED RICE STRAW FOR PRODUCTION OF ETHANOL

The batch simultaneous saccharification and fermentation (SSF) of microwave/acid/alkali/H₂O₂ pretreated rice straw to ethanol was optimized using cellulase from Trichoderma reesei and Saccharomyces cerevisiae YC-097 cells prior to the fed-batch SSF studies. The batch SSF optima were 10% w/v substrate, 40°C, 15 mg cellulase/g substrate, initial pH 5.3, and 72 hours. Under the optimum conditions the ethanol concentration and its yield were 29.1 g/L and 61.3% respectively. Based on the optimal batch SSF, the fed-batch SSF was investigated and its operation parameters were optimized. Under its optimal conditions the ethanol concentration reached 57.3 g/L, while its productivity and yield were only slightly less than those in the batch SSF. This suggests that fed-batch SSF is a potential operation mode for effective ethanol production from microwave/acid/alkali/H₂O₂ pretreated rice straw.     

利用Clostridium saccharobutylicum DSM 13864 连续发酵生产丁醇

为使用廉价糖质原料进行连续发酵生产丁醇提供理论依据和实验指导,以Clostridium saccharobutylicum DSM 13864为发酵菌种,考察了稀释率和温度等因素对以葡萄糖为原料的四级连续发酵生产丁醇的影响。结果表明：高稀释率有利于酸的积累和菌体的生长,低稀释率有利于溶剂的生产。当稀释率为0.05 h?1,4个发酵罐温度控制在37 ℃时,总溶剂产量为11.57 g/L,其中丁醇7.29 g/L,生产率为0.145 g/(L?h)。在0.05 h?1稀释率的条件下进行变温连续发酵证实低温有利于溶剂的积累,当一级和二级罐温度控制在37 ℃,三级和四级罐温度控制在33 ℃时总溶剂产量最高为13.69 g/L,其中丁醇为8.36 g/L,生产率为0.171 g/(L?h)。     

Production of 2,3-butanediol from diverse saccharides via fermentation

The present study aimed to evaluate the potential use of whey to produce 2,3-BD via the fermentation of lactose and its monosaccharides, glucose and galactose, in a synthetic culture medium (medium 9, M9) using a modified strain of Escherichia coli K12 MG1655 (E. coli JFR12) at a 0.1 L/L (10 vol%) inoculum ratio, 37 °C, atmospheric pressure, an initial pH 7.4, and 100 rpm for 72 h varying the saccharide concentration from 12.5, 25, and 50 g/L. The 2,3-BD yield was ∼80 % of the theoretical yield using 25 g/L of glucose and lactose, corresponding to 0.38 g/g saccharides at a fermentation time of 48 h (glucose) and 72 h (lactose). However, the 2,3-BD yield was halved (0.19 g/g galactose), fermenting 25 g/L of galactose at 48 h. Taking into account these results, two important conclusions were determined: i) E. coli JFR12 could transform galactose into 2,3-BD although its yield was half of the yield observed with glucose at 48 h; and ii) E. coli JFR12 was as efficient as other natural 2,3-BD producers such as Klebsiella species fermenting lactose. However, the E. coli strain has the advantage of being an innocuous strain. To the best of our knowledge, there is no other study presenting the production of 2,3-BD from galactose and lactose with a genetically modified E. coli strain.     

Preparation of D-lysine by chemical reaction and microbial asymmetric transformation

The DL-lysine crystals from the racemization of L-lysine was treated as substrate with Hafnia alvei AS1.1009 intact cells as biocatalysts to produce crystalline D-lysine with a yield of 56.6% from the reaction mixture after simple purification. In the presence of 0.10 molar equivalent of salicylaldehyde, L-lysine racemization can be completed within 4 h in 1.0 mol/L of NaOH at 100°C. The activation energy of the processes was 62187.86 J/mol. The characteristics of Hafnia alvei AS1.1009 decarboxylase were studied. Under the conditions of pH 8.0, temperature 37°C, cell concentration 10 g/L, tween-80 0.5 g/L, substrate concentration 30 g/L, and the specific activity of up to 3840 U, L-lysine can be completely degraded by the decarboxylase for 12 h under the optimal conditions.     

Continuous ethanol production from carob pod extract by immobilized Saccharomyces cerevisiae in a packed-bed reactor

The continuous production of ethanol from carob pod extract by immobilized Saccharomyces cerevisiae in a packed-bed reactor has been investigated. At a substrate concentration of 150 g dm^?3, maximum ethanol productivity of 16 g dm^?3 h^?1 was obtained at D = 0·4 h^?1 with 62·3% of theoretical yield and 83·6% sugars′ utilization. At a dilution rate of 0·1 h^?1, optimal ethanol productivity was achieved in the pH range 3·5–5·5, temperature range 30–35·C and initial sugar concentration of 200 g dm^?3. Maximum ethanol productivity of 24·5 g dm^?3 h^?1 was obtained at D = 0·5 h^?1 with 58·8% of theoretical yield and 85% sugars′ utilization when non-sterilized carob pod extract containing 200 g dm^?3 total sugars was used as feed material. The bioreactor system was operated at a constant dilution rate of 0·5 h^?1 for 30 days without loss of the original immobilized yeast activity. In this case, the average ethanol productivity, ethanol yield (% of theoretical) and sugars′ utilization were 25 g dm^?3 h^?1, 58·8% and 85·5%, respectively.     

Simultaneous saccharification and fermentation of wheat bran flour into ethanol using coculture of amylotic Aspergillus niger and thermotolerant Kluyveromyces marxianus

Studies on simultaneous saccharification and fermentation (SSF) of wheat bran flour, a grain milling residue as the substrate using coculture method were carried out with strains of starch digesting Aspergillus niger and nonstarch digesting and sugar fermenting Kluyveromyces marxianus in batch fermentation. Experiments based on central composite design (CCD) were conducted to maximize the glucose yield and to study the effects of substrate concentration, pH, temperature, and enzyme concentration on percentage conversion of wheat bran flour starch to glucose by treatment with fungal α-amylase and the above parameters were optimized using response surface methodology (RSM). The optimum values of substrate concentration, pH, temperature, and enzyme concentration were found to be 200 g/L, 5.5, 65°C and 7.5 IU, respectively, in the starch saccharification step. The effects of pH, temperature and substrate concentration on ethanol concentration, biomass and reducing sugar concentration were also investigated. The optimum temperature and pH were found to be 30°C and 5.5, respectively. The wheat bran flour solution equivalent to 6% (w/V) initial starch concentration gave the highest ethanol concentration of 23.1 g/L after 48 h of fermentation at optimum conditions of pH and temperature. The growth kinetics was modeled using Monod model and Logistic model and product formation kinetics using Leudeking-Piret model. Simultaneous saccharificiation and fermentation of liquefied wheat bran starch to bioethanol was studied using coculture of amylolytic fungus A. niger and nonamylolytic sugar fermenting K. marxianus.     

Batch kinetics and modelling of propionic acid fermentation

Batch propionic acid fermentation kinetics was studied using five different initial concentrations of lactose (i.e., 37 g/L, 45g/L, 50g/L, 57 g/L and 73 g/L) at constant temperature (30°C) and pH (6.5) under anaerobic conditions using Propionibacterium acidipropionici (ATCC 4875). When the initial substrate concentration was 37 g/L, 45 g/L, 50 g/L, 57 g/L and 73 g/L, then, correspondingly, 16 g/L, 19 g/L, 22.25 g/L, 25.3 g/L and 26.3 g/L of propionic acid was accumulated in the fermentation broth. Increasing the supply of lactose in the fermentation medium led to the accumulation of by products succinate, acetate and pyruvate. Maximum propionate yield (0.44 g/g) and comparatively lesser impurities (byproducts) were achieved with 57 g/L initial lactose concentration. The batch growth kinetics was eventually used to develop and test a mathematical model for propionic acid fermentation by P. acidipropionici at pH 6.5 and S_o = 57 g/L. Y_X/S^max was found to be the most sensitive parameter of the model. The same model also successfully simulated the batch kinetics observed at S_o = 37 g/L. However the model failed to simulate the fermentation kinetics observed at S_o = 73 g/L. The developed model can be used for process optimization studies.     

Enhanced production of l(+)-lactic acid by floc-form culture of Rhizopus oryzae

A process to optimize l-lactic acid production from glucose by Rhizopus oryzae, based on sustaining floc morphology throughout the fermentation process, is herein performed. During the fermentation, supplementary ammonium sulfate was added intermittently to maintain the ammonia level of the culture medium always higher than 0.1 g/L. With replenish of nitrogen source, mycelia flocs did not aggregate, and the lactic acid production was optimized upon the fermentation being controlled at pH 4.3–4.5 by adding calcium carbonate slurry. In contrast, without supplementary addition of nitrogen source, mycelial clumps formed, resulting in a poor production of lactic acid. In the initial batch fermentation process, the final concentration of lactic acid produced was 109 g/L, with a yield (g lactic acid/g glucose consumed) of 0.87 and a productivity of 2.73 g/L h, using 125 g/L of glucose as substrate. For the first four cycles of repeated-batch fermentation, the average final concentration, the productivity and the yield of lactic acid were 113 g/L, 4.03 g/L h and 0.90, respectively.     

The effects of mixing,temperature, and nutrient concentration on the fermentation of a mixed sugar solution simulating the hexose content of waste sulfite liquor

The results of changes in temperature, mixing and nutrient concentration on the conversion by yeast of D-mannose, D-galactose, and D-glucose using Saccharomyces cerevisiae strain were studied. These sugars are present in waste sulfite liquor. From the operating conditions used in this study, the most rapid removal of sugars found at a pH of 5 were temperature 36°C, mixing 700 rpm, and nutrient concentration 13.3 g/1. The fermentation was carried in a three-litre batch reactor and the main variables measured were dissolved oxygen, yeast growth, alcohol production, total sugar consumption, and D-mannose, D-galactose and D-glucose utilization. It was found that the fermentation could be fitted to the published Kono-Assai model.     

Sago starch saccharification and simultaneous ethanol fermentation by amyloglucosidase and Zymomonas mobilis

Simultaneous saccharification and ethanol fermentation (SSF) of sago starch was studied using amyloglucosidase (AMG) and Zymomonas mobilis. The optimal concentration of AMG and operating temperature for the SSF process were found to be 0.5% (v/w) and 35°C, respectively. Under these conditions with 150 g dm^?3 sago starch as a substrate, the final ethanol concentration obtained was 69.2 g dm^?3 and ethanol yield, Y_P/S, 0.50 g g^?1 (97% of theoretical yield). Sago starch in the concentration range of 100–200 g dm^?3 was efficiently converted into ethanol. When compared to a two-step process involving separate saccharification and fermentation stages, the SSF reduced the total process time by half.     

Optimized reuse and bioconversion from retentate of pre‐filtered palm oil mill effluent (POME) into microbial protease by Aspergillus terreus using response surface methodology

BACKGROUND: The membrane filtration process enables the treatment of wastewater, producing permeate which is less polluted. However, disposal is usually required for the retentate, which is produced as a concentrated constituent along with the permeate. In this study, the authors explored the possibility of reusing, rather than disposing of, the retentate of pre‐filtered palm oil mill effluent (POME) as a fermentation substrate in protease production by a wild type strain of Aspergillus terreus IMI 282743. In addition, the quantitative and interactive effects of the concentration factor for retentate, temperature, inoculum concentration, and fermentation time on the optimization of protease production were investigated using response surface methodology (RSM). RESULTS: Using RSM, the optimum conditions were found to be a concentration factor of 7.27, temperature of 37.95 °C, inoculum concentration of 1.30% (v/v) and fermentation time of 3.83 days. The protease production was increased 4.37‐fold in comparison with the results obtained under non‐optimized conditions. CONCLUSION: To a certain extent, protease production could be enhanced with an increase in concentration factor and temperature, and a decrease of inoculum concentration and fermentation time. Also, POME retentate was found to be a good substrate for protease production with high product activity and without nutrient supplementation. Copyright © 2009 Society of Chemical Industry