Sample records for repeated fed-batch fermentation

  1. Improving cellulase productivity of Penicillium oxalicum RE-10 by repeated fed-batch fermentation strategy. (United States)

    Han, Xiaolong; Song, Wenxia; Liu, Guodong; Li, Zhonghai; Yang, Piao; Qu, Yinbo


    Medium optimization and repeated fed-batch fermentation were performed to improve the cellulase productivity by P. oxalicum RE-10 in submerged fermentation. First, Plackett-Burman design (PBD) and central composite design (CCD) were used to optimize the medium for cellulase production. PBD demonstrated wheat bran and NaNO3 had significant influences on cellulase production. The CCD results showed the maximum filter paper activity (FPA) production of 8.61U/mL could be achieved in Erlenmeyer flasks. The maximal FPA reached 12.69U/mL by submerged batch fermentation in a 7.5-L stirred tank, 1.76-fold higher than that on the original medium. Then, the repeated fed-batch fermentation strategy was performed successfully for increasing the cellulase productivity from 105.75U/L/h in batch fermentation to 158.38U/L/h. The cellulase activity and the glucan conversion of delignined corn cob residue hydrolysis had no significant difference between the enzymes sampled from different cycles of the repeated fed-batch fermentation and that from batch culture.

  2. One Approach for Dynamic L-lysine Modelling of Repeated Fed-batch Fermentation

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    Kalin Todorov


    Full Text Available This article deals with establishment of dynamic unstructured model of variable volume fed-batch fermentation process with intensive droppings for L-lysine production. The presented approach of the investigation includes the following main procedures: description of the process by generalized stoichiometric equations; preliminary data processing and calculation of specific rates for main kinetic variables; identification of the specific rates as a second-order non-linear dynamic models; establishment and optimisation of dynamic model of the process; simulation researches. MATLAB is used as a research environment.

  3. Butanol production by immobilised Clostridium acetobutylicum in repeated batch, fed-batch, and continuous modes of fermentation. (United States)

    Dolejš, Igor; Krasňan, Vladimír; Stloukal, Radek; Rosenberg, Michal; Rebroš, Martin


    Clostridium acetobutylicum immobilised in polyvinylalcohol, lens-shaped hydrogel capsules (LentiKats(®)) was studied for production of butanol and other products of acetone-butanol-ethanol fermentation. After optimising the immobilisation protocol for anaerobic bacteria, continuous, repeated batch, and fed-batch fermentations in repeated batch mode were performed. Using glucose as a substrate, butanol productivity of 0.41 g/L/h and solvent productivity of 0.63 g/L/h were observed at a dilution rate of 0.05 h(-1) during continuous fermentation with a concentrated substrate (60 g/L). Through the process of repeated batch fermentation, the duration of fermentation was reduced from 27.8h (free-cell fermentation) to 3.3h (immobilised cells) with a solvent productivity of 0.77 g/L/h (butanol 0.57 g/L/h). The highest butanol and solvent productivities of 1.21 and 1.91 g/L/h were observed during fed-batch fermentation operated in repeated batch mode with yields of butanol (0.15 g/g) and solvents (0.24 g/g), respectively, produced per gram of glucose.

  4. Supervision of Fed-Batch Fermentations

    DEFF Research Database (Denmark)

    Gregersen, Lars; Jørgensen, Sten Bay


    Process faults may be detected on-line using existing measurements based upon modelling that is entirely data driven. A multivariate statistical model is developed and used for fault diagnosis of an industrial fed-batch fermentation process. Data from several (25) batches are used to develop a mo...

  5. Modelling of L-valine Repeated Fed-batch Fermentation Process Taking into Account the Dissolved Oxygen Tension

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    Tzanko Georgiev


    Full Text Available This article deals with synthesis of dynamic unstructured model of variable volume fed-batch fermentation process with intensive droppings for L-valine production. The presented approach of the investigation includes the following main procedures: description of the process by generalized stoichiometric equations; preliminary data processing and calculation of specific rates for main kinetic variables; identification of the specific rates takes into account the dissolved oxygen tension; establishment and optimisation of dynamic model of the process; simulation researches. MATLAB is used as a research environment.

  6. Optimal control of a fed-batch fermentation

    Energy Technology Data Exchange (ETDEWEB)

    Dekkers, R.M.


    The common cultivation of bakers' yeast is an aerobic fed-batch fermentation under sugar-limited growth. The ultimate objective of on-line computer control is to optimize the process through maximizing the productivity of biomass formation while minimizing the consumption of raw materials for the product. Results obtained on the optimal control of a fed-batch fermentation are given. The aspects to be considered are instrumentation, state estimation, optimization and process control.

  7. Continuous Cellulosic Bioethanol Fermentation by Cyclic Fed-Batch Cocultivation (United States)

    Jiang, He-Long; He, Qiang; He, Zhili; Hemme, Christopher L.; Wu, Liyou


    Cocultivation of cellulolytic and saccharolytic microbial populations is a promising strategy to improve bioethanol production from the fermentation of recalcitrant cellulosic materials. Earlier studies have demonstrated the effectiveness of cocultivation in enhancing ethanolic fermentation of cellulose in batch fermentation. To further enhance process efficiency, a semicontinuous cyclic fed-batch fermentor configuration was evaluated for its potential in enhancing the efficiency of cellulose fermentation using cocultivation. Cocultures of cellulolytic Clostridium thermocellum LQRI and saccharolytic Thermoanaerobacter pseudethanolicus strain X514 were tested in the semicontinuous fermentor as a model system. Initial cellulose concentration and pH were identified as the key process parameters controlling cellulose fermentation performance in the fixed-volume cyclic fed-batch coculture system. At an initial cellulose concentration of 40 g liter−1, the concentration of ethanol produced with pH control was 4.5-fold higher than that without pH control. It was also found that efficient cellulosic bioethanol production by cocultivation was sustained in the semicontinuous configuration, with bioethanol production reaching 474 mM in 96 h with an initial cellulose concentration of 80 g liter−1 and pH controlled at 6.5 to 6.8. These results suggested the advantages of the cyclic fed-batch process for cellulosic bioethanol fermentation by the cocultures. PMID:23275517

  8. Pilot scale repeated fed-batch fermentation processes of the wine yeast Dekkera bruxellensis for mass production of resveratrol from Polygonum cuspidatum. (United States)

    Kuo, Hsiao-Ping; Wang, Reuben; Lin, Yi-Sheng; Lai, Jinn-Tsyy; Lo, Yi-Chen; Huang, Shyue-Tsong


    Resveratrol has long been used as an ingredient in functional foods. Currently, Polygonum cuspidatum extract is the greatest natural source for resveratrol because of high concentrations of glycosidic-linked resveratrol. Thus, developing a cost-effective procedure to hydrolyze glucoside could substantially enhance resveratrol production from P. cuspidatum. This study selected Dekkera bruxellensis from several microorganisms based on its bioconversion and enzyme-specific activities. We demonstrated that the cells could be reused at least nine times while maintaining an average of 180.67U/L β-glucosidase activity. The average resveratrol bioconversion efficiency within five rounds of repeated usage was 108.77±0.88%. This process worked effectively when the volume was increased to 1200L, a volume at which approximately 35mgL(-1)h(-1) resveratrol per round was produced. This repeated fed-batch bioconversion process for resveratrol production is comparable to enzyme or cell immobilization strategies in terms of reusing cycles, but without incurring additional costs for immobilization. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Enhanced phenylpyruvic acid production with Proteus vulgaris in fed-batch and continuous fermentation. (United States)

    Coban, Hasan B; Demirci, Ali; Patterson, Paul H; Elias, Ryan J


    Phenylpyruvic acid is a deaminated form of phenylalanine and is used in various areas such as development of cheese and wine flavors, diagnosis of phenylketonuria, and to decrease excessive nitrogen accumulation in the manure of farm animals. However, reported phenylpyruvic acid fermentation studies in the literature have been usually performed at shake-flask scale with low production. In this study, phenylpyruvic acid production was evaluated in bench-top bioreactors by conducting fed-batch and continuous fermentation for the first time. As a result, maximum phenylpyruvic acid concentrations increased from 1350 mg/L (batch fermentation) to 2958 mg/L utilizing fed-batch fermentation. Furthermore, phenylpyruvic acid productivity was increased from 48 mg/L/hr (batch fermentation) to 104 and 259 mg/L/hr by conducting fed-batch and continuous fermentation, respectively. Overall, this study demonstrated that fed-batch and continuous fermentation significantly improved phenylpyruvic acid production in bench-scale bioreactor production.

  10. Batch and fed-batch fermentation of Bacillus thuringiensis using starch industry wastewater as fermentation substrate. (United States)

    Vu, Khanh Dang; Tyagi, Rajeshwar Dayal; Valéro, José R; Surampalli, Rao Y


    Bacillus thuringiensis var. kurstaki biopesticide was produced in batch and fed-batch fermentation modes using starch industry wastewater as sole substrate. Fed-batch fermentation with two intermittent feeds (at 10 and 20 h) during the fermentation of 72 h gave the maximum delta-endotoxin concentration (1,672.6 mg/L) and entomotoxicity (Tx) (18.5 x 10(6) SBU/mL) in fermented broth which were significantly higher than maximum delta-endotoxin concentration (511.0 mg/L) and Tx (15.8 x 10(6) SBU/mL) obtained in batch process. However, fed-batch fermentation with three intermittent feeds (at 10, 20 and 34 h) of the fermentation resulted in the formation of asporogenous variant (Spo-) from 36 h to the end of fermentation (72 h) which resulted in a significant decrease in spore and delta-endotoxin concentration and finally the Tx value. Tx of suspended pellets (27.4 x 10(6) SBU/mL) obtained in fed-batch fermentation with two feeds was the highest value as compared to other cases.

  11. On-line Scheduling Algorithm for Penicillin Fed-batch Fermentation

    Institute of Scientific and Technical Information of China (English)

    XUE Yao-feng; YUAN Jing-qi


    An on-line scheduling algorithm to maximize gross profit of penicillin fed-batch fermentation is proposed. According to the on-line classification method, fed-batch fermentation batches are classified into three categories. Using the scheduling strategy, the optimal termination sequence of batches is obtained. Pseudo on-line simulations for testing the proposed algorithm with the data from industrial scale penicillin fermentation are carried out.

  12. Batch and Fed-Batch Fermentation System on Ethanol Production from Whey using Kluyveromyces marxianus

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    H Hadiyanto


    Full Text Available Nowadays reserve of fossil fuel has gradually depleted. This condition forces many researchers to  find energy alternatives which is renewable and sustainable in the future. Ethanol derived from cheese industrial waste (whey using fermentation process can be a new perspective in order to secure both energy and environment. The aim of this study was  to compare the operation modes (batch and fed-batch of fermentation system on ethanol production from whey using Kluyveromyces marxianus. The result showed that the fermentation process for ethanol production by fed-batch system was higher at some point of parameters compared with batch system. Growth rate and ethanol yield (YP/S of fed-batch fermentation were 0.122/h and 0.21 gP/gS respectively; growth rate and ethanol yield (YP/S of batch fermentation were 0.107/h, and 0.12 g ethanol/g substrate, respectively. Based on the data of biomass and ethanol concentrations, the fermentation process for ethanol production by fed-batch system were higher at some point of parameters compared to batch system. Periodic substrate addition performed on fed-batch system leads the yeast growth in low substrate concentrations and consequently  increasing their activity and ethanol productivity. Keywords: batch; ethanol; fed-batch; fermentation;Kluyveromyces marxianus, whey


    Institute of Scientific and Technical Information of China (English)


    1 IntroductionGlycerol fed-batch fermentation is attractive tocommercial application since it can control theglucose concentration by changing the feed rate andget a high glycerol yield, therefore it is essential todevelop an optimal glucose feed strategy. For mostof fed-batch fermentation, optimization of feed ratewas based on Pontryagin's maximum principle [if.Since the term of feed rate appears linearly in theHamiltonian, the optimal feed rate profile usuallyconsists of ba,lg-bang intervals and singular ...

  14. Enhancement of thermoalkaliphilic xylanase production by Pichia pastoris through novel fed-batch strategy in high cell-density fermentation

    National Research Council Canada - National Science Library

    Shang, Tingting; Si, Dayong; Zhang, Dongyan; Liu, Xuhui; Zhao, Longmei; Hu, Cong; Fu, Yu; Zhang, Rijun


    .... This study aims to enhance thermoalkaliphilic xylanase production in Pichia pastoris through fermentation parameters optimization and novel efficient fed-batch strategy in high cell-density fermentation...

  15. Enhanced incorporation yield of cysteine for glutathione overproduction by fed-batch fermentation of Saccharomyces cerevisiae. (United States)

    Lorenz, Eric; Schmacht, Maximilian; Stahl, Ulf; Senz, Martin


    In the following work a high cell density fed-batch process with Saccharomyces cerevisiae coupled with a high efficient incorporation of cysteine for glutathione (GSH) overproduction was developed. Therefore, a feeding strategy based on the respiratory quotient (RQ) was applied to ensure high biomass (96.1g/l). Furthermore, the optimal cysteine concentration and time of cysteine addition were investigated. Low concentrations of cysteine at late fermentation phases resulted in relatively high incorporation yields of about 0.40mol/mol and maintained the physiology of cultivated yeast. By changing the cysteine feeding from standard single shot to continuous addition, an often observed cell specific toxicity, triggered by high cysteine concentrations, could be prevented and the cysteine incorporation yield (0.54±0.01mol/mol) and GSH content (1650.7±42.8mg/l; 1.76±0.08%) were maximized, respectively. The developed process was transferred from laboratory into pilot plant scale. Further, the reduced cell specific toxicity enabled the development of a repeated fed-batch procedure with a suitable performance concerning cysteine incorporation yield (0.40±0.1mol/mol), biomass (84.2±1.2g/l) and GSH content (1304.7±61.4mg/l).

  16. Fed-batch fermentation dealing with nitrogen limitation in microbial transglutaminase production by Streptoverticillium mobaraense

    NARCIS (Netherlands)

    Rinzema, A; Tramper, J; de Bruin, E; Bol, J

    In the later stages of a batch fermentation for microbial transglutaminase production by Streptoverticillium mobaraense the availability of a nitrogen source accessible to the microorganism becomes critical. Fed-batch fermentation is investigated with the aim of avoiding this substrate limitation.

  17. Fed-batch fermentation dealing with nitrogen limitation in microbial transglutaminase production by Streptoverticillium mobaraense

    NARCIS (Netherlands)

    Zhu, Y.; Rinzema, A.; Tramper, J.; Bruin, E. de; Bol, J.


    In the later stages of a batch fermentation for microbial transglutaminase production by Streptoverticillium mobaraense the availability of a nitrogen source accessible to the microorganism becomes critical. Fed-batch fermentation is investigated with the aim of avoiding this substrate limitation. W

  18. Production of Ammonium Lactate by Fed-batch Fermentation of Rhizopus oryzae from Corncob Hydrolysate

    Institute of Scientific and Technical Information of China (English)

    BAI Dong-mei; LI Shi-zhong; LIN Fang-qian


    L- (+)-Lactic acid production from corncob hydrolysate as a cheap carbohydrate source by fed-batch fermentation of Rhizopus oryzae HZS6 was studied. After 96 h of fermentation in a 5 L fermentor, the final concentration of ammonium L-(+)-lactate, average productivity(based on initial xylose concentration) and max(+)-lactate was 98.8%.

  19. High cell density fed-batch fermentations for lipase production: feeding strategies and oxygen transfer. (United States)

    Salehmin, M N I; Annuar, M S M; Chisti, Y


    This review is focused on the production of microbial lipases by high cell density fermentation. Lipases are among the most widely used of the enzyme catalysts. Although lipases are produced by animals and plants, industrial lipases are sourced almost exclusively from microorganisms. Many of the commercial lipases are produced using recombinant species. Microbial lipases are mostly produced by batch and fed-batch fermentation. Lipases are generally secreted by the cell into the extracellular environment. Thus, a crude preparation of lipases can be obtained by removing the microbial cells from the fermentation broth. This crude cell-free broth may be further concentrated and used as is, or lipases may be purified from it to various levels. For many large volume applications, lipases must be produced at extremely low cost. High cell density fermentation is a promising method for low-cost production: it allows a high concentration of the biomass and the enzyme to be attained rapidly and this eases the downstream recovery of the enzyme. High density fermentation enhances enzyme productivity compared with the traditional submerged culture batch fermentation. In production of enzymes, a high cell density is generally achieved through fed-batch operation, not through perfusion culture which is cumbersome. The feeding strategies used in fed-batch fermentations for producing lipases and the implications of these strategies are discussed. Most lipase-producing microbial fermentations require oxygen. Oxygen transfer in such fermentations is discussed.

  20. A cellular automata model for simulating fed-batch penicillin fermentation process

    Institute of Scientific and Technical Information of China (English)

    Yu Naigong; Ruan Xiaogang


    A cellular automata model to simulate penicillin fed-batch fermentation process(CAPFM)was established in this study,based on a morphologically structured dynamic penicillin production model,that is in turn based on the growth mechanism of penicillin producing microorganisms and the characteristics of penicillin fed-batch fermentation.CAPFM uses the three-dimensional cellular automata as a growth space,and a Moore-type neighborhood as the cellular neighborhood.The transition roles of CAPFM are designed based on mechanical and structural kinetic models of penicillin batch-fed fermentation processes.Every cell of CAPFM represents a single or specific number of penicillin producing microorganisms,and has various state.The simulation experimental results show that CAPFM replicates the evolutionary behavior of penicillin batch-fed fermentation processes described by the structured penicillin production kinetic model accordingly.

  1. Optimal substrate feeding policy for a fed batch fermentation with substrate and product inhibition kinetics. (United States)

    Hong, J


    The optimal substrate feeding policy for the fed batch fermentation which is governed by product and substrate inhibited kinetics is presented. The conjunction point between nonsingular and singular arcs and the feeding policy along the singular arc are derived analytically in terms of the concentrations of substrate and product and the liquid volume. Thus, it is possible to determine the feeding rate by monitoring the state variables (i.e., closed loop control). As a specific example, an optimization study of the fed batch fermentation for ethanol production by Saccharomyces cerevisiae is presented. It is shown that the optimal feeding patterns are heavily dependent upon the initial conditions. The point selectivity provides the guideline for predicting the optimal feeding patterns and explaining the results of rigorous mathematical analysis.

  2. Oxygen Control for an Industrial Pilot-Scale Fed-Batch Filamentous Fungal Fermentation


    Bodizs, Levente; Titica, Mariana; Faria, Nuno; Srinivasan, Bala; Dochain, Denis; Bonvin, Dominique


    Industrial filamentous fungal fermentations are typically operated in fed- batch mode. Oxygen control represents an important operational challenge due to the varying biomass concentration. In this study, oxygen control is implemented by manipulating the substrate feed rate, i.e. the rate of oxygen consumption. It turns out that the setpoint for dissolved oxygen represents a trade-off since a low dissolved oxygen value favors productivity but can also induce oxygen limitation. This pape...

  3. Modelling of Fed-batch Fermentation Process with Droppings for L-lysine Production

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    Velitchka Ivanova


    Full Text Available The aim of the article is the development of dynamic unstructured model of L-lysine fed-batch fermentation process with droppings. This approach includes the following procedures: description of the process by generalized stoichiometric equations; preliminary data processing; identification of the specific rates (growth rate (mu , substrate utilization rate (nu, production rate (rho; establishment and optimization of the dynamic model of the process; simulation researches.

  4. Scale-up of dextransucrase production by Leuconostoc mesenteroides in fed batch fermentation

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    Michelena Georgina L.


    Full Text Available Fed batch fermentation was carried out for the dextransucrase enzyme production from Leuconostoc mesenteroides and the production was scale-up using oxygen transfer criteriuom. It was found that in 5 L vessel fermentation capacity, the best agitation speed was 225 min-1 and aeration rate was 0.15 vvm, obtaining dextransucrase activity of 127 DSU/mL.. The maximum enzyme production velocity coincide with the maximum growth velocity between 6 and 7 h of fermentation, which confirmed that dextransucrase production was associated with microbial growth. High enzyme yields were achieved during scale up based on oxygen transfer rate.

  5. Enhancing Production of Alkaline Polygalacturonate Lyase from Bacillus subtilis by Fed-Batch Fermentation


    Mouyong Zou; Fenfen Guo; Xuezhi Li; Jian Zhao; Yinbo Qu


    Alkaline polygalacturonate lyase (PGL, EC is an enzyme used in many industries. We developed a fed-batch fermentation process that combines the enzymatic pretreatment of the carbon source with controlling the pH of the fermentative broth to enhance the PGL production from Bacillus subtilis 7-3-3 to decrease the production cost. Maintaining the fermentation broth at pH 6.5 prior to feeding with ammonia and at pH 6.0 after feeding significantly improved PGL activity (743.5 U mL-1) comp...

  6. Enhanced submerged Aspergillus ficuum phytase production by implementation of fed-batch fermentation. (United States)

    Coban, Hasan B; Demirci, Ali


    Phytase is an important feed and food additive, which is both used in animal and human diets. Phytase has been used to increase the absorption of several divalent ions, amino acids, and proteins in the bodies and to decrease the excessive phosphorus release in the manure to prevent negative effects on the environment. To date, microbial phytase has been mostly produced in solid-state fermentations with insignificant production volumes. There are only a few studies in the literature that phytase productions were performed in submerged bench-top reactor scale. In our previous studies, growth parameters (temperature, pH, and aeration) and important fermentation medium ingredients (glucose, Na-phytate, and CaSO4) were optimized. This study was undertaken for further enhancement of phytase production with Aspergillus ficuum in bench-top bioreactors by conducting fed-batch fermentations. The results showed that addition of 60 g of glucose and 10 g of Na-phytate at 96 h of fermentation increased phytase activity to 3.84 and 4.82 U/ml, respectively. Therefore, the maximum phytase activity was further enhanced with addition of glucose and Na-phytate by 11 and 40 %, respectively, as compared to batch phytase fermentations. It was also reported that phytase activity increased higher in early log stage additions than late log stage additions because of higher microbial activity. In addition, the phytase activity in fed-batch fermentation did not drop significantly as compared to the batch fermentation. Overall, this study shows that fungal phytase can be successfully produced in submerged fed-batch fermentations.

  7. Omega-3 production by fermentation of Yarrowia lipolytica: From fed-batch to continuous. (United States)

    Xie, Dongming; Miller, Edward; Sharpe, Pamela; Jackson, Ethel; Zhu, Quinn


    The omega-3 fatty acid, cis-5,8,11,14,17-eicosapentaenoic acid (C20:5; EPA) has wide-ranging benefits in improving heart health, immune function, and mental health. A sustainable source of EPA production through fermentation of metabolically engineered Yarrowia lipolytica has been developed. In this paper, key fed-batch fermentation conditions were identified to achieve 25% EPA in the yeast biomass, which is so far the highest EPA titer reported in the literature. Dynamic models of the EPA fermentation process were established for analyzing, optimizing, and scaling up the fermentation process. In addition, model simulations were used to develop a two-stage continuous process and compare to single-stage continuous and fed- batch processes. The two stage continuous process, which is equipped with a smaller growth fermentor (Stage 1) and a larger production fermentor (Stage 2), was found to be a superior process to achieve high titer, rate, and yield of EPA. A two-stage continuous fermentation experiment with Y. lipolytica strain Z7334 was designed using the model simulation and then tested in a 2 L and 5 L fermentation system for 1,008 h. Compared with the standard 2 L fed-batch process, the two-stage continuous fermentation process improved the overall EPA productivity by 80% and EPA concentration in the fermenter by 40% while achieving comparable EPA titer in biomass and similar conversion yield from glucose. During the long-term experiment it was also found that the Y. lipolytica strain evolved to reduce byproduct and increase lipid production. This is one of the few continuous fermentation examples that demonstrated improved productivity and concentration of a final product with similar conversion yield compared with a fed-batch process. This paper suggests the two-stage continuous fermentation could be an effective process to achieve improved production of omega-3 and other fermentation products where non-growth or partially growth associated kinetics

  8. A novel model-based control strategy for aerobic filamentous fungal fed-batch fermentation processes

    DEFF Research Database (Denmark)

    Mears, Lisa; Stocks, Stuart M.; Albaek, Mads O.


    A novel model-based control strategy has been developed for filamentous fungal fed-batch fermentation processes. The system of interest is a pilot scale (550 L) filamentous fungus process operating at Novozymes A/S. In such processes, it is desirable to maximize the total product achieved...... in a batch in a defined process time. In order to achieve this goal, it is important to maximize both the product concentration, and also the total final mass in the fed-batch system. To this end, we describe the development of a control strategy which aims to achieve maximum tank fill, while avoiding oxygen...... limited conditions. This requires a two stage approach: (i) calculation of the tank start fill; and (ii) on-line control in order to maximize fill subject to oxygen transfer limitations. First, a mechanistic model was applied off-line in order to determine the appropriate start fill for processes...

  9. Optimization of cyclosporin A production by Beauveria nivea in continuous fed-batch fermentation

    Directory of Open Access Journals (Sweden)

    Dong Huijun


    Full Text Available To develop the effective control method for fed-batch culture of cyclosporin A production, we chose fructose, L-valine and (NH42HPO4 as feeding nutrients and compared their productivities in relation to different concentrations. The feeding rate of three kinds of feeding materials was controlled to maintain the suitable residual concentration. The fed-batch fermentation results indicated that the optimal concentrations of fructose, L-valine and (NH42HPO4 were about 20 g/L, 0.5 g/L and 0.6 g/L for cyclosporin A production, respectively. The cultivation of Beauveria nivea could produce cyclosporin A up to 6.2 g/L for 240 hrs through a continuous feeding-rate-controlled-batch process under the optimal feeding conditions.

  10. Fed-batch simultaneous saccharification and ethanol fermentation of native corn starch

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    Włodzimierz Grajek


    Full Text Available Background. The most important innovations in boethanol production in the last decade were: simultaneous saccharification and fermentation processes (SSF, high gravity fermentation, the use of new enzyme preparation able to hydrolyse native granular starch and construction of genetically modified strains of microorganisms able to carry out simultaneous production of hydrolytic enzymes and fermentation of C6 and C5 sugars. The aim of this study was to assess the efficiency of ethanol fermentation using new type of amylolytic enzymes able to hydrolyse native corn starch in a SSF process. Material and methods. The simultaneous saccharification and fermentation of raw corn flour by fed-batch processes using Saccharomyces cerevisiae strain Red Star Ethanol Red and Stargen 001 enzyme preparation was performed. As experimental variable were investigated: fermentation temperature (35-37-40°C, rate of mash stirring (100 and 200 rpm, fermentation time (0-92 h and dosage of corn flour (different portion and different time. Results. It was found that optimal temperature for fed-batch SSF process was 37°C at initial pH of 5.0. However, the yeast intensively fermented the saccharides also at 40°C. The fermentation stirring rate has significant effect on starch utilization and fermentation production. The prolongation of fermentation time over 72 h has no substantiation in additional ethanol production. In all experimental fermentations the level of produced organic acids was very low, significantly below toxic concentration for the yeast. Conclusions. It was stated that the use of new method of starch raw material preparation resulted in satisfied fermentation yield and allowed to reduce energy requirements for starch liquefaction.  

  11. Proteome profiling illustrated by a large-scale fed-batch fermentation of Penicillium chrysogenum

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    Michaela Helmel


    Full Text Available Filamentous fungi are employed for the large-scale production of value-added products, including organic acids, enzymes, and antibiotics and bioprocess characterization is essential for production optimization but relies on empiricism-based strategies. Protein expression profiles in an industrial scale, 180 h fed-batch fermentation of Penicillium chrysogenum are presented. The biomass of P. chrysogenum, as well as the specific penicillin V production rate and fungal morphology were monitored during fermentation to be compared with obtained protein profiles. Our results demonstrate a correlation between proteomics data and biomass concentration, morphological changes, and penicillin production.

  12. Enhancing Production of Alkaline Polygalacturonate Lyase from Bacillus subtilis by Fed-Batch Fermentation (United States)

    Zou, Mouyong; Guo, Fenfen; Li, Xuezhi; Zhao, Jian; Qu, Yinbo


    Alkaline polygalacturonate lyase (PGL, EC is an enzyme used in many industries. We developed a fed-batch fermentation process that combines the enzymatic pretreatment of the carbon source with controlling the pH of the fermentative broth to enhance the PGL production from Bacillus subtilis 7-3-3 to decrease the production cost. Maintaining the fermentation broth at pH 6.5 prior to feeding with ammonia and at pH 6.0 after feeding significantly improved PGL activity (743.5 U mL−1) compared with the control (202.5 U mL−1). The average PGL productivity reached 19.6 U mL−1 h−1 after 38 h of fermentation. The crude PGL was suitable for environmentally friendly ramie enzymatic degumming. PMID:24603713

  13. Production of gamma-aminobutyric acid by Lactobacillus brevis NCL912 using fed-batch fermentation

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    Huang Guidong


    Full Text Available Abstract Background Gamma-aminobutyric acid is a major inhibitory neurotransmitter in mammalian brains, and has several well-known physiological functions. Lactic acid bacteria possess special physiological activities and are generally regarded as safe. Therefore, using lactic acid bacteria as cell factories for gamma-aminobutyric acid production is a fascinating project and opens up a vast range of prospects for making use of GABA and LAB. We previously screened a high GABA-producer Lactobacillus brevis NCL912 and optimized its fermentation medium composition. The results indicated that the strain showed potential in large-scale fermentation for the production of gamma-aminobutyric acid. To increase the yielding of GABA, further study on the fermentation process is needed before the industrial application in the future. In this article we investigated the impacts of pyridoxal-5'-phosphate, pH, temperature and initial glutamate concentration on gamma-aminobutyric acid production by Lactobacillus brevis NCL912 in flask cultures. According to the data obtained in the above, a simple and effective fed-batch fermentation method was developed to highly efficiently convert glutamate to gamma-aminobutyric acid. Results Pyridoxal-5'-phosphate did not affect the cell growth and gamma-aminobutyric acid production of Lb. brevis NCL912. Temperature, pH and initial glutamate concentration had significant effects on the cell growth and gamma-aminobutyric acid production of Lb. brevis NCL912. The optimal temperature, pH and initial glutamate concentration were 30-35°C, 5.0 and 250-500 mM. In the following fed-batch fermentations, temperature, pH and initial glutamate concentration were fixed as 32°C, 5.0 and 400 mM. 280.70 g (1.5 mol and 224.56 g (1.2 mol glutamate were supplemented into the bioreactor at 12 h and 24 h, respectively. Under the selected fermentation conditions, gamma-aminobutyric acid was rapidly produced at the first 36 h and almost not

  14. Nisin production of Lactococcus lactis N8 with hemin-stimulated cell respiration in fed-batch fermentation system. (United States)

    Kördikanlıoğlu, Burcu; Şimşek, Ömer; Saris, Per E J


    In this study, nisin production of Lactococcus lactis N8 was optimized by independent variables of glucose, hemin and oxygen concentrations in fed-batch fermentation in which respiration of cells was stimulated with hemin. Response surface model was able to explain the changes of the nisin production of L. lactis N8 in fed-batch fermentation system with high fidelity (R(2) 98%) and insignificant lack of fit. Accordingly, the equation developed indicated the optimum parameters for glucose, hemin, and dissolved oxygen were 8 g L(-1) h(-1) , 3 μg mL(-1) and 40%, respectively. While 1711 IU mL(-1) nisin was produced by L. lactis N8 in control fed-batch fermentation, 5410 IU mL(-1) nisin production was achieved within the relevant optimum parameters where the respiration of cell was stimulated with hemin. Accordingly, nisin production was enhanced 3.1 fold in fed-batch fermentation using hemin. In conclusion the nisin production of L. lactis N8 was enhanced extensively as a result of increasing the biomass by stimulating the cell respiration with adding the hemin in the fed-batch fermentation.

  15. Alcoholic fermentation with flocculant Saccharomyces cerevisiae in fed-batch process. (United States)

    Guidini, Carla Zanella; Marquez, Líbia Diniz Santos; de Almeida Silva, Helisângela; de Resende, Miriam Maria; Cardoso, Vicelma Luiz; Ribeiro, Eloízio Júlio


    Studies have been conducted on selecting yeast strains for use in fermentation for ethanol production to improve the performance of industrial plants and decrease production costs. In this paper, we study alcoholic fermentation in a fed-batch process using a Saccharomyces cerevisiae yeast strain with flocculant characteristics. Central composite design (CCD) was used to determine the optimal combination of the variables involved, with the sucrose concentration of 170 g/L, a cellular concentration in the inoculum of 40% (v/v), and a filling time of 6 h, which resulted in a 92.20% yield relative to the theoretical maximum yield, a productivity of 6.01 g/L h and a residual sucrose concentration of 44.33 g/L. With some changes in the process such as recirculation of medium during the fermentation process and increase in cellular concentration in the inoculum after use of the CCD was possible to reduce the residual sucrose concentration to 2.8 g/L in 9 h of fermentation and increase yield and productivity for 92.75% and 9.26 g/L h, respectively. A model was developed to describe the inhibition of alcoholic fermentation kinetics by the substrate and the product. The maximum specific growth rate was 0.103 h(-1), with K(I) and K(s) values of 109.86 and 30.24 g/L, respectively. The experimental results from the fed-batch reactor show a good fit with the proposed model, resulting in a maximum growth rate of 0.080 h(-1).

  16. Dark fermentation of ground wheat starch for bio-hydrogen production by fed-batch operation

    Energy Technology Data Exchange (ETDEWEB)

    Kargi, Fikret; Pamukoglu, M. Yunus [Department of Environmental Engineering, Dokuz Eylul University, 35160 Buca, Izmir (Turkey)


    Ground wheat solution was used for bio-hydrogen production by dark fermentation using heat-treated anaerobic sludge in a completely mixed fermenter operating in fed-batch mode. The feed wheat powder (WP) solution was fed to the anaerobic fermenter with a constant flow rate of 8.33 mL h{sup -1} (200 mL d{sup -1}). Cumulative hydrogen production, starch utilization and hydrogen yields were determined at three different WP loading rates corresponding to the feed WP concentrations of 10, 20 and 30 g L{sup -1}. The residual starch (substrate) concentration in the fermenter decreased with operation time while starch consumption was increasing. The highest cumulative hydrogen production (3600 mL), hydrogen yield (465 mL H{sub 2} g{sup -1} starch or 3.1 mol H{sub 2} mol{sup -1} glucose) and hydrogen production rate (864 mL H{sub 2} d{sup -1}) were obtained after 4 days of fed-batch operation with the 20 g L{sup -1} feed WP concentration corresponding to a WP loading rate of 4 g WP d{sup -1}. Low feed WP concentrations (10 g L{sup -1}) resulted in low hydrogen yields and rates due to substrate limitations. High feed WP concentrations (30 g L{sup -1}) resulted in the formation of volatile fatty acids (VFAs) in high concentrations causing inhibition on the rate and yield of hydrogen production. (author)

  17. User-friendly optimization approach of fed-batch fermentation conditions for the production of iturin A using artificial neural networks and support vector machine

    Directory of Open Access Journals (Sweden)

    Fudi Chen


    Conclusion: According to the modeling results, the GRNN is considered as the most suitable ANN model for the design of the fed-batch fermentation conditions for the production of iturin A because of its high robustness and precision, and the SVM is also considered as a very suitable alternative model. Under the tolerance of 30%, the prediction accuracies of the GRNN and SVM are both 100% respectively in repeated experiments.

  18. A Genetic Algorithm for Feeding Trajectory Optimisation of Fed-batch Fermentation Processes

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    Stoyan Tzonkov


    Full Text Available In this work a genetic algorithm is proposed with the purpose of the feeding trajectory optimization during a fed-batch fermentation of E. coli. The feed rate profiles are evaluated based on a number of objective functions. Optimization results obtained for different feeding trajectories demonstrate that the genetic algorithm works well and shows good computational performance. Developed optimal feed profiles meet the defined criteria. The ration of the substrate concentration and the difference between actual cell concentration and theoretical maximum cell concentration is defined as the most appropriate objective function. In this case the final cell concentration of 43 g·l-1 and final product concentration of 125 g·l-1 are achieved and there is not significant excess of substrate.

  19. A dynamic metabolic flux balance based model of fed-batch fermentation of Bordetella pertussis. (United States)

    Budman, Hector; Patel, Nilesh; Tamer, Melih; Al-Gherwi, Walid


    A mathematical model based on a dynamic metabolic flux balance (DMFB) is developed for a process of fed-batch fermentation of Bordetella pertussis. The model is based on the maximization of growth rate at each time interval subject to stoichiometric constraints. The model is calibrated and verified with experimental data obtained in two different bioreactor experimental systems. It was found that the model calibration was mostly sensitive to the consumption or production rates of tyrosine and, for high supplementation rates, to the consumption rate of glutamate. Following this calibration the model correctly predicts biomass and by-products concentrations for different supplementation rates. Comparisons of model predictions to oxygen uptake and carbon emission rates measurements indicate that the TCA cycle is fully functional.

  20. Time Series Analysis of Fed-batch Fermentation Process for L-valine Production

    Directory of Open Access Journals (Sweden)

    Tzanko Georgiev


    Full Text Available Fed-batch fermentation processes are some of the most efficient and wildly applied types of cultivation for industrial production of most amino acids including L-valine. Time series analysis is an important tool for description of the experimental data. This article deals with statistical inference from the time series analysis of generalised stoichiometric equations as a hypothesis for modelling and optimisation. The aim of the article is to develop some time series models of generalized stoichiometric equations. The identification procedure includes the following steps: description of the process by generalized stoichiometric equations; preliminary data processing; model structure selection for each stoichiometric equation; estimation of the model's parameters; verification of the derived models.

  1. The development of an industrial-scale fed-batch fermentation simulation. (United States)

    Goldrick, Stephen; Ştefan, Andrei; Lovett, David; Montague, Gary; Lennox, Barry


    This paper describes a simulation of an industrial-scale fed-batch fermentation that can be used as a benchmark in process systems analysis and control studies. The simulation was developed using a mechanistic model and validated using historical data collected from an industrial-scale penicillin fermentation process. Each batch was carried out in a 100,000 L bioreactor that used an industrial strain of Penicillium chrysogenum. The manipulated variables recorded during each batch were used as inputs to the simulator and the predicted outputs were then compared with the on-line and off-line measurements recorded in the real process. The simulator adapted a previously published structured model to describe the penicillin fermentation and extended it to include the main environmental effects of dissolved oxygen, viscosity, temperature, pH and dissolved carbon dioxide. In addition the effects of nitrogen and phenylacetic acid concentrations on the biomass and penicillin production rates were also included. The simulated model predictions of all the on-line and off-line process measurements, including the off-gas analysis, were in good agreement with the batch records. The simulator and industrial process data are available to download at and can be used to evaluate, study and improve on the current control strategy implemented on this facility. Crown Copyright © 2014. Published by Elsevier B.V. All rights reserved.

  2. Kinetic study of batch and fed-batch enzymatic saccharification of pretreated substrate and subsequent fermentation to ethanol

    Directory of Open Access Journals (Sweden)

    Gupta Rishi


    Full Text Available Abstract Background Enzymatic hydrolysis, the rate limiting step in the process development for biofuel, is always hampered by its low sugar concentration. High solid enzymatic saccharification could solve this problem but has several other drawbacks such as low rate of reaction. In the present study we have attempted to enhance the concentration of sugars in enzymatic hydrolysate of delignified Prosopis juliflora, using a fed-batch enzymatic hydrolysis approach. Results The enzymatic hydrolysis was carried out at elevated solid loading up to 20% (w/v and a comparison kinetics of batch and fed-batch enzymatic hydrolysis was carried out using kinetic regimes. Under batch mode, the actual sugar concentration values at 20% initial substrate consistency were found deviated from the predicted values and the maximum sugar concentration obtained was 80.78 g/L. Fed-batch strategy was implemented to enhance the final sugar concentration to 127 g/L. The batch and fed-batch enzymatic hydrolysates were fermented with Saccharomyces cerevisiae and ethanol production of 34.78 g/L and 52.83 g/L, respectively, were achieved. Furthermore, model simulations showed that higher insoluble solids in the feed resulted in both smaller reactor volume and shorter residence time. Conclusion Fed-batch enzymatic hydrolysis is an efficient procedure for enhancing the sugar concentration in the hydrolysate. Restricting the process to suitable kinetic regimes could result in higher conversion rates.

  3. Improvement in the bioreactor specific productivity by coupling continuous reactor with repeated fed-batch reactor for acetone-butanol-ethanol production. (United States)

    Setlhaku, Mpho; Brunberg, Sina; Villa, Eva Del Amor; Wichmann, Rolf


    In comparison to the different fermentation modes for the production of acetone, butanol and ethanol (ABE) researched to date, the continuous fermentation is the most economically favored. Continuous fermentation with two or more reactor cascade is reported to be the most efficient as it results in a more stable solvent production process. In this work, it is shown that a continuous (first-stage) reactor coupled to a repeated fed-batch (second stage) is superior to batch and fed-batch fermentations, including two-stage continuous fermentation. This is due to the efficient catalyst use, reported through the specific product rate and rapid glucose consumption rate. High solvents are produced at 19.4 g(ABE) l⁻¹, with volumetric productivities of 0.92 g(butanol) l⁻¹ h⁻¹ and 1.47 g(ABE) l ⁻¹ h⁻¹. The bioreactor specific productivities of 0.62 and 0.39 g g⁻¹(cdw) h⁻¹ obtained show a high catalyst activity. This new process mode has not been reported before in the development of ABE fermentation and it shows great potential and superiority to the existing fermentation methods. Copyright © 2012 Elsevier B.V. All rights reserved.

  4. Optimization of ectoine synthesis through fed-batch fermentation of Brevibacterium epidermis. (United States)

    Onraedt, Annelies E; Walcarius, Bart A; Soetaert, Wim K; Vandamme, Erick J


    A production process for ectoine has been developed, using Brevibacterium epidermis DSM20659 as the producer strain. First, the optimal conditions for intracellular synthesis of ectoine were determined. The size of the intracellular ectoine pool is shown to be dependent on the external salt concentration, type of carbon source, and yeast extract concentration. Under the optimized conditions of 1 M NaCl, 50 g/L monosodium glutamate, and 2.5 g/L yeast extract, a maximum concentration of intracellular ectoine of 0.9 g/L was obtained in shake flask cultures. After optimizing the batch fermentation parameters of temperature, pH, agitation, and aeration, the yield could be further increased by applying the fed-batch fermentation principle in 1.5- to 2-L fermentors. Glutamate and yeast extract were fed to the bacterial cells such that the total glutamate concentration in the broth remained constant. A total yield of 8 g ectoine/L fermentation broth was obtained with a productivity of 2 g ectoine/L/day. After the bacterial cells were harvested from the culture broth, the ectoine was recovered from them by a two-step extraction with water and ethanol. Crystallization of the product was obtained after concentration of the extract via evaporation under reduced pressure. After this downstream process, 55% of the ectoine produced in the fermentor could be crystallized in four fractions. The first fractions were of very high purity (98%). This production process can compete with other described production processes for ectoine in productivity and simplicity. Further advantages are the relatively low amounts of NaCl needed and the absence of hydroxyectoine, often a byproduct, in the final product.

  5. Kinetic characterization and fed-batch fermentation for maximal simultaneous production of esterase and protease from Lysinibacillus fusiformis AU01. (United States)

    Divakar, K; Suryia Prabha, M; Nandhinidevi, G; Gautam, P


    The simultaneous production of intracellular esterase and extracellular protease from the strain Lysinibacillus fusiformis AU01 was studied in detail. The production was performed both under batch and fed-batch modes. The maximum yield of intracellular esterase and protease was obtained under full oxygen saturation at the beginning of the fermentation. The data were fitted to the Luedeking-Piret model and it was shown that the enzyme (both esterase and protease) production was growth associated. A decrease in intracellular esterase and increase in the extracellular esterase were observed during late stationary phase. The appearance of intracellular proteins in extracellular media and decrease in viable cell count and biomass during late stationary phase confirmed that the presence of extracellular esterase is due to cell lysis. Even though the fed-batch fermentation with different feeding strategies showed improved productivity, feeding yeast extract under DO-stat fermentation conditions showed highest intracellular esterase and protease production. Under DO-stat fed-batch cultivation, maximum intracellular esterase activity of 820 × 10(3) U/L and extracellular protease activity of 172 × 10(3) U/L were obtained at the 16th hr. Intracellular esterase and extracellular protease production were increased fivefold and fourfold, respectively, when compared to batch fermentation performed under shake flask conditions.

  6. Experimental optimization of a real time fed-batch fermentation process using Markov decision process. (United States)

    Saucedo, V M; Karim, M N


    This article describes a methodology that implements a Markov decision process (MDP) optimization technique in a real time fed-batch experiment. Biological systems can be better modeled under the stochastic framework and MDP is shown to be a suitable technique for their optimization. A nonlinear input/output model is used to calculate the probability transitions. All elements of the MDP are identified according to physical parameters. Finally, this study compares the results obtained when optimizing ethanol production using the infinite horizon problem, with total expected discount policy, to previous experimental results aimed at optimizing ethanol production using a recombinant Escherichia coli fed-batch cultivation. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 55: 317-327, 1997.

  7. Actinoplanes utahensis ZJB-08196 fed-batch fermentation at elevated osmolality for enhancing acarbose production. (United States)

    Wang, Ya-Jun; Liu, Li-Ling; Wang, Yuan-Shan; Xue, Ya-Ping; Zheng, Yu-Guo; Shen, Yin-Chu


    Acarbose, a potent α-glucosidase inhibitor, is as an oral anti-diabetic drug for treatment of the type two, noninsulin-dependent diabetes. Actinoplanes utahensis ZJB-08196, an osmosis-resistant actinomycete, had a broad osmolality optimum between 309 mOsm kg(-1) and 719 mOsm kg(-1). Utilizing this unique feature, an fed-batch culture process under preferential osmolality was constructed through intermittently feeding broths with feed medium consisting of 14.0 g l(-1) maltose, 6.0 g l(-1) glucose and 9.0 g l(-1) soybean meal, at 48 h, 72 h, 96 h and 120 h. This intermittent fed-batch culture produced a peak acarbose titer of 4878 mg l(-1), increased by 15.9% over the batch culture.

  8. High bioethanol titre from Manihot glaziovii through fed-batch simultaneous saccharification and fermentation in Automatic Gas Potential Test System. (United States)

    Moshi, Anselm P; Crespo, Carla F; Badshah, Malik; Hosea, Kenneth M M; Mshandete, Anthony Manoni; Mattiasson, Bo


    A process for the production of high bioethanol titre was established through fed-batch and simultaneous saccharification and fermentation (FB-SSF) of wild, non-edible cassava Manihot glaziovii. FB-SSF allowed fermentation of up to 390g/L of starch-derived glucose achieving high bioethanol concentration of up to 190g/L (24% v/v) with yields of around 94% of the theoretical value. The wild cassava M. glaziovii starch is hydrolysable with a low dosage of amylolytic enzymes (0.1-0.15% v/w, Termamyl® and AMG®). The Automatic Gas Potential Test System (AMPTS) was adapted to yeast ethanol fermentation and demonstrated to be an accurate, reliable and flexible device for studying the kinetics of yeast in SSF and FB-SSF. The bioethanol derived stoichiometrically from the CO2 registered in the AMPTS software correlated positively with samples analysed by HPLC (R(2)=0.99).


    Directory of Open Access Journals (Sweden)

    Anahita Dehkhoda


    Full Text Available With the aim of increasing the sugars concentration in dilute-acid ligno-cellulosic hydrolyzate to more than 100 g/l for industrial applications, the hydrolyzate from spruce was concentrated about threefold by high-pressure or vacuum evaporations. It was then fermented by repeated fed-batch cultivation using flocculating Saccharomyces cerevisiae with no prior detoxification. The sugars and inhibitors concentrations in the hydrolyzates were compared after the evaporations and also fermenta-tion. The evaporations were carried out either under vacuum (VEH at 0.5 bar and 80°C or with 1.3 bar pressure (HPEH at 107.5°C, which resulted in 153.3 and 164.6 g/l total sugars, respectively. No sugar decomposition occurred during either of the evaporations, while more than 96% of furfural and to a lesser extent formic and acetic acids disappeared from the hydrolyzates. However, HMF and levulinic acid remained in the hydrolyzates and were concentrated proportionally. The concentrated hydrolyzates were then fermented in a 4 l bioreactor with 12-22 g/l yeast and 0.14-0.22 h-1 initial dilute rates (ID. More than 84% of the fermentable sugars present in the VEH were fermented by fed-batch cultivation using 12 g/l yeast and initial dilution rate (ID of 0.22 h-1, and resulted in 0.40±0.01 g/g ethanol from the fermentable sugars in one cycle of fermentation. Fermentation of HPEH was as successful as VEH and resulted in more than 86% of the sugar consumption under the corresponding conditions. By lowering the initial dilution rate to 0.14 h-1, more than 97% of the total fermentable sugars were consumed, and ethanol yield was 0.44±0.01 g/g in one cycle of fermentation. The yeast was able to convert or assimilate HMF, levulinic, acetic, and formic acids by 96, 30, 43, and 74%, respectively.

  10. Intact cell mass spectrometry as a progress tracking tool for batch and fed-batch fermentation processes. (United States)

    Helmel, Michaela; Marchetti-Deschmann, Martina; Raus, Martin; Posch, Andreas E; Herwig, Christoph; Šebela, Marek; Allmaier, Günter


    Penicillin production during a fermentation process using industrial strains of Penicillium chrysogenum is a research topic permanently discussed since the accidental discovery of the antibiotic. Intact cell mass spectrometry (ICMS) can be a fast and novel monitoring tool for the fermentation progress during penicillin V production in a nearly real-time fashion. This method is already used for the characterization of microorganisms and the differentiation of fungal strains; therefore, the application of ICMS to samples directly harvested from a fermenter is a promising possibility to get fast information about the progress of fungal growth. After the optimization of the ICMS method to penicillin V fermentation broth samples, the obtained ICMS data were evaluated by hierarchical cluster analysis or an in-house software solution written especially for ICMS data comparison. Growth stages of a batch and fed-batch fermentation of Penicillium chrysogenum are differentiated by one of those statistical approaches. The application of two matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) instruments in the linear positive ion mode from different vendors demonstrated the universal applicability of the developed ICMS method. The base for a fast and easy-to-use method for monitoring the fermentation progress of P. chrysogenum is created with this ICMS method developed especially for fermentation broth samples.

  11. Batch and fed-batch simultaneous saccharification and fermentation of primary sludge from pulp and paper mills. (United States)

    Mendes, Cátia Vanessa Teixeira; Rocha, Jorge Manuel Dos Santos; de Menezes, Fabrícia Farias; Carvalho, Maria da Graça Videira Sousa


    Primary sludge from a Portuguese pulp and paper mill, containing 60% of carbohydrates, and unbleached pulp (as reference material), with 93% of carbohydrates, were used to produce ethanol by simultaneous saccharification and fermentation (SSF). SSF was performed in batch or fed-batch conditions without the need of a pretreatment. Cellic(®) CTec2 was the cellulolytic enzymatic complex used and Saccharomyces cerevisiae (baker's yeast or ATCC 26602 strain) or the thermotolerant yeast Kluyveromyces marxianus NCYC 1426 were employed. Primary sludge was successfully converted to ethanol and the best results in SSF efficiency were obtained with S. cerevisiae. An ethanol concentration of 22.7 g L(-1) was produced using a content of 50 g L(-1) of carbohydrates from primary sludge, in batch conditions, with a global conversion yield of 81% and a production rate of 0.94 g L(-1) h(-1). Fed-batch operation enabled higher solids content (total carbohydrate concentration of 200 g L(-1), equivalent to a consistency of 33%) and a reduction of three-quarters of cellulolytic enzyme load, leading to an ethanol concentration of 40.7 g L(-1), although with lower yield and productivity. Xylitol with a concentration up to 7 g L(-1) was also identified as by-product in the primary sludge bioconversion process.

  12. Sucrose fed-batch strategy enhanced biomass, polysaccharide, and ganoderic acids production in fermentation of Ganoderma lucidum 5.26. (United States)

    Wei, Zhen-hua; Liu, Lianliang; Guo, Xiao-feng; Li, Yan-jun; Hou, Bao-chao; Fan, Qiu-ling; Wang, Kai-xiang; Luo, Yingdi; Zhong, Jian-jiang


    Ganoderma, as a Chinese traditional medicine, has multiple bioactivities. However, industrial production was limited due to low yield during Ganoderma fermentation. In this work, sucrose was found to greatly enhance intracellular polysaccharide (IPS) content and specific extracellular polysaccharide (EPS) production rate. The mechanism was studied by analyzing the activities of enzymes related to polysaccharide biosynthesis. The results revealed that sucrose regulated the activities of phosphoglucomutase and phosphoglucose isomerase. When glucose and sucrose mixture was used as carbon source, biomass, polysaccharide and ganoderic acids (GAs) production was greatly enhanced. A sucrose fed-batch strategy was developed in 10-L bioreactor, and was scaled up to 300-L bioreactor. The biomass, EPS and IPS production was 25.5, 2.9 and 4.8 g/L, respectively, which was the highest biomass and IPS production in pilot scale. This study provides information for further understanding the regulation mechanism of Ganoderma polysaccharide biosynthesis. It demonstrates that sucrose fed-batch is a useful strategy for enhancing Ganoderma biomass, polysaccharide and GAs production.

  13. Global expression profiling of Bacillus subtilis cells during industrial-close fed-batch fermentations with different nitrogen sources. (United States)

    Jürgen, Britta; Tobisch, Steffen; Wümpelmann, Mogens; Gördes, Dirk; Koch, Andreas; Thurow, Kerstin; Albrecht, Dirk; Hecker, Michael; Schweder, Thomas


    A detailed gene expression analysis of industrial-close Bacillus subtilis fed-batch fermentation processes with casamino acids as the only nitrogen source and with a reduced casamino acid concentration but supplemented by ammonia was carried out. Although glutamine and arginine are supposed to be the preferred nitrogen sources of B. subtilis, we demonstrate that a combined feeding of ammonia and casamino acids supports cell growth under fed-batch fermentation conditions. The transcriptome and proteome analyses revealed that the additional feeding of ammonia in combination with a reduced amino acid concentration results in a significantly lower expression level of the glnAR or tnrA genes, coding for proteins, which are mainly involved in the nitrogen metabolism of B. subtilis. However, the mRNA levels of the genes of the ilvBHC-leuABD and hom-thrCB operons were significantly increased, indicating a valine, leucine, isoleucine, and threonine limitation under these fermentation conditions. In contrast, during the fermentation with casamino acids as the only nitrogen source, several genes, which play a crucial role in nitrogen metabolism of B. subtilis (e.g., glnAR, nasCDE, nrgAB, and ureABC), were up-regulated, indicating a nitrogen limitation under these conditions. Furthermore, increased expression of genes, which are involved in motility and chemotaxis (e.g., hag, fliT) and in acetoin metabolism (e.g., acoABCL), was determined during the fermentation with the mixed nitrogen source of casamino acids and ammonia, indicating a carbon limitation under these fermentation conditions. Under high cell density and slow growth rate conditions a weak up-regulation of autolysis genes could be observed as well as the induction of a number of genes involved in motility, chemotaxis and general stress response. Results of this study allowed the selection of marker genes, which could be used for the monitoring of B. subtilis fermentation processes. The data suggest for example aco

  14. Fed-batch alcoholic fermentation of sugar cane blackstrap molasses: Influence of the feeding rate on yeast yield and productivity

    Energy Technology Data Exchange (ETDEWEB)

    Carvalho, J.C.M. de; Aquarone, E.; Sato, S.; Brazzach, M.L.; Moraes, D.A. (Faculdade de Ciencias Farmaceuticas, Sao Paulo Univ., SP (Brazil)); Borzani, W. (Centro de Desenvolvimento Biotecnologico, Joinville, SC (Brazil))


    Fed-batch ethanol fermentation tests of sugar cane blackstrap molasses were carried out at 32deg C and pH 4.5-5.0, using pressed yeast as inoculum, and with no air supply. Two values of the fermentor filling-up time were adopted: 5 h and 7 h. The feeding rates obeyed equation F=F[sub 0].e[sup K.t], with K equal to 0.0, 0.2, 0.4, 0.6 and 0.8 h[sup -1]. The average yeast yields and the average yeast productivities increased up to 33% and 45%, respectively, while the ethanol yield (average=76%; standard deviation=4%) was practically unaffected when K increased from 0 to 0.8 h[sup -1]. (orig.).

  15. Improvement of Photorhabdus temperata strain K122 bioinsecticide production by batch and fed-batch fermentations optimization. (United States)

    Jallouli, Wafa; Jaoua, Samir; Zouari, Nabil


    Optimization of a fermentation process for bioinsecticides production by Photorhabdus temperata strain K122 was investigated into fully controlled 3-L fermenter using an optimized medium (OM). Development of large-scale inocula showed that the composition of the growth medium greatly influenced the physiological state of P. temperata cells. The effect of pH, agitation and dissolved oxygen concentration (DO) on the growth, culturability and oral toxicity of P. temperata cells were also investigated. Indeed, maintaining the pH at 7 and controlling DO concentration at 50 % saturation throughout the fermentation process, improved biomass production, CFU counts and oral toxicity by 41.1, 35 and 32.1 %, respectively, as compared to cultures carried out in 500 mL shake flasks. At such conditions, 8 g/L glucose fed-batch fermentation, enhanced cell lysis and variants small colony (Vsm) polymorphism appearance. To overcome such limitations, glucose concentration should be maintained at 4 g/L. In this case, P. temperata cells were produced at high cell density and culturability reaching 4.5 and 1.2 × 10(9) cells/mL, respectively. In addition, the stability of the primary form was maintained for a long period in the stationary growth phase and Vsm polymorphism was completely avoided that can be crucial for scale-up the bioprocess of P. temperata bioinsecticide.

  16. Feeding strategies for the enhanced production of α-arbutin in the fed-batch fermentation of Xanthomonas maltophilia BT-112. (United States)

    Liu, Chunqiao; Zhang, Peng; Zhang, Shurong; Xu, Tao; Wang, Fang; Deng, Li


    To develop a cost-effective method for the enhanced production of α-arbutin using Xanthomonas maltophilia BT-112 as a biocatalyst, different fed-batch strategies such as constant feed rate fed-batch, constant hydroquinone (HQ) concentration fed-batch, exponential fed-batch and DO-control pulse fed-batch (DPFB) on α-arbutin production were investigated. The research results indicated that DPFB was an effective method for α-arbutin production. When fermentation with DO-control pulse feeding strategy to feed HQ and yeast extract was applied, the maximum concentrations of α-arbutin and cell dry weight were 61.7 and 4.21 g/L, respectively. The α-arbutin production was 394% higher than that of the control (batch culture) and the molar conversion yield of α-arbutin reached 94.5% based on the amount of HQ supplied (240 mM). Therefore, the results in this work provide an efficient and easily controlled method for industrial-scale production of α-arbutin.

  17. Biosynthesis of Citric Acid from Glycerol by Acetate Mutants of Yarrowia lipolytica in Fed-Batch Fermentation

    Directory of Open Access Journals (Sweden)

    Anita Rywińska


    Full Text Available Pure and crude glycerol from biodiesel production have been used as substrates for citric acid production by acetate-negative mutants of Yarrowia lipolytica in fed-batch fermentation. Both the final concentration and the yield of the product were the highest when Y. lipolytica Wratislavia AWG7 strain was used in the culture with pure or crude glycerol. With a medium containing 200 g/L of glycerol, production reached a maximum of citric acid of 139 g/L after 120 h. This high yield of the product (up to 0.69 g of citric acid per gram of glycerol consumed was achieved with both pure and crude glycerol. Lower yield of citric acid in the culture with Y. lipolytica Wratislavia K1 strain (about 0.45 g/g resulted from increased erythritol concentrations (up to 40 g/L, accumulated simultaneously with the citric acid. The concentration of isocitric acid, a by-product in this fermentation, was very low, in the range from 2.6 to 4.6 g/L.

  18. Differential expression of small RNAs under chemical stress and fed-batch fermentation in E. coli

    DEFF Research Database (Denmark)

    Rau, Martin Holm; Nielsen, Alex Toftgaard; Long, Katherine


    applications, the involvement of sRNAs in this process is not well understood. We have used RNA sequencing to map sRNA expression in E. coli under chemical stress and high cell density fermentation conditions with the aim of identifying sRNAs involved in the transcriptional response and those with potential...

  19. Hydrogen production from starch by co-culture of Clostridium acetobutylicum and Rhodobacter sphaeroides in one step hybrid dark- and photofermentation in repeated fed-batch reactor. (United States)

    Zagrodnik, R; Łaniecki, M


    Hydrogen production from starch by a co-culture hybrid dark and photofermentation under repeated fed-batch conditions at different organic loading rates (OLR) was studied. Effective cooperation between bacteria in co-culture during initial days was observed at controlled pH 7.0. However, at pH above 6.5 dark fermentation phase was redirected from H2 formation towards production of formic acid, lactic acid and ethanol (which are not coupled with hydrogen production) with simultaneous lower starch removal efficiency. This resulted in decrease in the hydrogen production rate. The highest H2 production in co-culture process (3.23LH2/Lmedium - after 11days) was achieved at OLR of 1.5gstarch/L/day, and it was twofold higher than for dark fermentation process (1.59LH2/Lmedium). The highest H2 yield in the co-culture (2.62molH2/molhexose) was obtained at the OLR of 0.375gstarch/L/day. Different pH requirements of bacteria were proven to be a key limitation in co-culture system. Copyright © 2016 Elsevier Ltd. All rights reserved.

  20. Optimal Control of a Fed-batch Fermentation Process by Neuro-Dynamic Programming

    Directory of Open Access Journals (Sweden)

    Tatiana Ilkova


    Full Text Available In this paper the method for optimal control of a fermentation process is presented, that is based on an approach for optimal control - Neuro-Dynamic programming. For this aim the approximation neural network is developed and the decision of the optimization problem is improved by an iteration mode founded on the Bellman equation. With this approach computing time and procedure are decreased and quality of the biomass at the end of the process is increased.

  1. High concentrations of cellulosic ethanol achieved by fed batch semi simultaneous saccharification and fermentation of waste-paper. (United States)

    Elliston, Adam; Collins, Samuel R A; Wilson, David R; Roberts, Ian N; Waldron, Keith W


    A fundamental goal of second generation ethanol production is to increase the ethanol concentration to 10% (v/v) or more to optimise distillation costs. Semi simultaneous saccharification and fermentations (SSSF) were conducted at small pilot scale (5L) utilising fed-batch additions of solid shredded copier paper substrate. Early addition of Accellerase® 1500 at 16 FPU/g substrate and 30 U/g β-glucosidase followed by substrate only batch addition allowed low final equivalent enzyme concentrations to be achieved (3.7 FPU/g substrate) whilst maintaining digestion. Batch addition resulted in a cumulative substrate concentration equivalent to 65% (w/v). This in turn resulted in the production of high concentrations of ethanol (11.6% v/v). The success of this strategy relied on the capacity of the bioreactor to perform high shear mixing as required. Further research into the timing and number of substrate additions could lead to further improvement in overall yields from the 65.5% attained. Copyright © 2013 Elsevier Ltd. All rights reserved.

  2. Multivariate Curve Resolution and Carbon Balance Constraint to Unravel FTIR Spectra from Fed-Batch Fermentation Samples

    Directory of Open Access Journals (Sweden)

    Dennis Vier


    Full Text Available The current work investigates the capability of a tailored multivariate curve resolution–alternating least squares (MCR-ALS algorithm to analyse glucose, phosphate, ammonium and acetate dynamics simultaneously in an E. coli BL21 fed-batch fermentation. The high-cell-density (HCDC process is monitored by ex situ online attenuated total reflection (ATR Fourier transform infrared (FTIR spectroscopy and several in situ online process sensors. This approach efficiently utilises automatically generated process data to reduce the time and cost consuming reference measurement effort for multivariate calibration. To determine metabolite concentrations with accuracies between ±0.19 and ±0.96·gL−l, the presented utilisation needs primarily—besides online sensor measurements—single FTIR measurements for each of the components of interest. The ambiguities in alternating least squares solutions for concentration estimation are reduced by the insertion of analytical process knowledge primarily in the form of elementary carbon mass balances. Thus, in this way, the established idea of mass balance constraints in MCR combines with the consistency check of measured data by carbon balances, as commonly applied in bioprocess engineering. The constraints are calculated based on online process data and theoretical assumptions. This increased calculation effort is able to replace, to a large extent, the need for manually conducted quantitative chemical analysis, leads to good estimations of concentration profiles and a better process understanding.

  3. Development of an on-line state estimator for fed-batch filamentous fungal fermentations

    DEFF Research Database (Denmark)

    Mears, Lisa; Stocks, Stuart M.; Albæk, Mads O.

    Bioprocesses can be challenging to model due to complex and non-linear process dynamics [1]. In addition there is a lack of robust, on-line sensors for key parameters of interest in the field, such as substrate, product and biomass concentration [2]. These factors lead to limitations in the ability...... to monitor and control bioprocess systems. There is therefore an interest in state estimation, in order to model these key process states based on available on-line measurements [1]. This work discusses the application of a first principle model to pilot scale filamentous fungal fermentation systems operated...... pressure [4], [5]. This stoichiometric-based coupled process model is successfully applied on-line as a state estimator in order to predict the biomass and product concentration, from robust, available on-line measurements. Such state estimators will be valuable as part of control strategy development...

  4. Fed batch fermentation and purification strategy for high yield production of Brucella melitensis recombinant Omp 28 kDa protein and its application in disease diagnosis. (United States)

    Karothia, B S; Athmaram, T N; D, Thavaselvam; Ashu, Kumar; Tiwari, Sapna; Singh, Anil K; Sathyaseelan, K; Gopalan, N


    Brucellosis is a disease caused by bacteria belonging to the genus Brucella. It affects cattle, goat, sheep, dog and humans. The serodiagnosis of brucellosis involves detection of antibodies generated against the LPS or whole cell bacterial extracts, however these tests lack sensitivity and specificity. The present study was performed to optimize the culture condition for the production of recombinant Brucella melitensis outer membrane protein 28 kDa protein in E.coli via fed batch fermentation. Expression was induced with 1.5mM isopropyl β thiogalactoside and the expressed recombinant protein was purified using Ni-NTA affinity chromatography. After fed-batch fermentation the dry cell weight of 17.81 g/L and a purified protein yield of 210.10 mg/L was obtained. The purified Brucella melitensis recombinant Omp 28 kDa protein was analyzed through SDS- poly acrylamide gel electrophoresis and western blotting. The obtained recombinant protein was evaluated for its diagnostic application through Indirect ELISA using brucellosis suspected human sera samples. Our results clearly indicate that recombinant Omp28 produced via fed batch fermentation has immense potential as a diagnostic reagent that could be employed in sero monitoring of brucellosis.

  5. Fed-batch cultivation of baker's yeast followed by nitrogen or carbon starvation: effects on fermentative capacity and content of trehalose and glycogen

    DEFF Research Database (Denmark)

    Jørgensen, Henning; Olsson, Lisbeth; Rønnow, B.


    An industrial strain of Saccharomyces cerevisiae (DGI 342) was cultivated in fed-batch cultivations at a specific growth rate of 0.2 h(-1). The yeast was then exposed to carbon or nitrogen starvation for up to 8 h, to study the effect of starvation on fermentative capacity and content of protein...... of the yeast cells, and the fermentative capacity per gram dry-weight decreased by 40%. The protein content in the carbon-starved yeast increased as a result of starvation due to the fact that the content of glycogen was reduced. The fermentative capacity per gram dry-weight was, however, unaltered....

  6. Effects of starch loading rate on performance of combined fed-batch fermentation of ground wheat for bio-hydrogen production

    Energy Technology Data Exchange (ETDEWEB)

    Ozmihci, Serpil; Kargi, Fikret [Department of Environmental Engineering, Dokuz Eylul University, 35160 Buca, Izmir (Turkey)


    Ground wheat powder solution (10 g L{sup -1}) was subjected to combined dark and light fermentations for bio-hydrogen production by fed-batch operation. A mixture of heat treated anaerobic sludge (AN) and Rhodobacter sphaeroides-NRRL (RS-NRRL) were used as the mixed culture of dark and light fermentation bacteria with an initial dark/light biomass ratio of 1/2. Effects of wheat starch loading rate on the rate and yield of bio-hydrogen formation were investigated. The highest cumulative hydrogen formation (CHF = 3460 ml), hydrogen yield (201 ml H{sub 2} g{sup -1} starch) and formation rate (18.1 ml h{sup -1}) were obtained with a starch loading rate of 80.4 mg S h{sup -1}. Complete starch hydrolysis and glucose fermentation were achieved within 96 h of fed-batch operation producing volatile fatty acids (VFA) and H{sub 2}. Fermentation of VFAs by photo-fermentation for bio-hydrogen production was most effective at starch loading rate of 80.4 mg S h{sup -1}. Hydrogen formation by combined fermentation took place by a fast dark fermentation followed by a rather slow light fermentation after a lag period. (author)

  7. Control of agitation and aeration rates in the production of surfactin in foam overflowing fed-batch culture with industrial fermentation. (United States)

    Yao, Shulin; Zhao, Shengming; Lu, Zhaoxin; Gao, Yuqi; Lv, Fengxia; Bie, Xiaomei


    Bacillus amyloliquefaciens fmb50 produces a high yield of surfactin, a lipopeptide-type biosurfactant that has been widely studied and has potential applications in many fields. A foam overflowing culture has been successfully used in the combined production-enrichment fermentation of surfactin. In this study, the agitation and aeration rates were found to have relationships with foam formation and surfactin enrichment. A maximum surfactin concentration of 4.7g/l of foam was obtained after 21h of culture with an agitation rate of 150rpm and an aeration rate of 1vvm in fed-batch culture. By controlling the foam overflow rate (fout) of a fed-batch culture, surfactin concentration in the foam was continuously maintained above 4g/l. Copyright © 2015. Publicado por Elsevier España, S.L.U.

  8. Efficient production of ethanol from empty palm fruit bunch fibers by fed-batch simultaneous saccharification and fermentation using Saccharomyces cerevisiae. (United States)

    Park, Jang Min; Oh, Baek-Rock; Seo, Jeong-Woo; Hong, Won-Kyung; Yu, Anna; Sohn, Jung-Hoon; Kim, Chul Ho


    The concentration of ethanol produced from lignocellulosic biomass should be at least 40 g l(-1) [about 5 % (v/v)] to minimize the cost of distillation process. In this study, the conditions for the simultaneous saccharification and fermentation (SSF) at fed-batch mode for the production of ethanol from alkali-pretreated empty palm fruit bunch fibers (EFB) were investigated. Optimal conditions for the production of ethanol were identified as temperature, 30 °C; enzyme loading, 15 filter paper unit g(-1) biomass; and yeast (Saccharomyces cerevisiae) loading, 5 g l(-1) of dry cell weight. Under these conditions, an economical ethanol concentration was achieved within 17 h, which further increased up to 62.5 g l(-1) after 95 h with 70.6 % of the theoretical yield. To our knowledge, this is the first report to evaluate the economic ethanol production from alkali-pretreated EFB in fed-batch SSF using S. cerevisiae.

  9. Kinetics of sugars consumption and ethanol inhibition in carob pulp fermentation by Saccharomyces cerevisiae in batch and fed-batch cultures. (United States)

    Lima-Costa, Maria Emília; Tavares, Catarina; Raposo, Sara; Rodrigues, Brígida; Peinado, José M


    The waste materials from the carob processing industry are a potential resource for second-generation bioethanol production. These by-products are small carob kibbles with a high content of soluble sugars (45-50%). Batch and fed-batch Saccharomyces cerevisiae fermentations of high density sugar from carob pods were analyzed in terms of the kinetics of sugars consumption and ethanol inhibition. In all the batch runs, 90-95% of the total sugar was consumed and transformed into ethanol with a yield close to the theoretical maximum (0.47-0.50 g/g), and a final ethanol concentration of 100-110 g/l. In fed-batch runs, fresh carob extract was added when glucose had been consumed. This addition and the subsequent decrease of ethanol concentrations by dilution increased the final ethanol production up to 130 g/l. It seems that invertase activity and yeast tolerance to ethanol are the main factors to be controlled in carob fermentations. The efficiency of highly concentrated carob fermentation makes it a very promising process for use in a second-generation ethanol biorefinery.

  10. Reduction of N-terminal methionylation while increasing titer by lowering metabolic and protein production rates in E. coli auto-induced fed-batch fermentation. (United States)

    Xu, Jianlin; Qian, Yueming; Skonezny, Paul M; You, Li; Xing, Zizhuo; Meyers, David S; Stankavage, Robert J; Pan, Shih-Hsie; Li, Zheng Jian


    A standard fed-batch fermentation process using 1 mM isopropyl-β-D: -thiogalactopyranoside (IPTG) induction at 37 °C in complex batch and feed media had been developed for manufacturing of a therapeutic protein (TP) expressed in inclusion bodies (IBs) by E. coli BL21 (DE3) driven by T7 promoter. Six unauthentic TP N-terminal variants were identified, of which methionylated TP (Met-TP) ratio was predominant. We hypothesized that lowering metabolic and protein production rates would reduce the Met-TP ratio while improving TP titer. The standard process was surprisingly auto-induced without added IPTG due to galactose in the complex media. Without changing either the clone or the batch medium, a new process was developed using lower feed rates and auto-induction at 29 °C after glucose depletion while increasing induction duration. In comparison to the standard process, the new process reduced the unauthentic Met-TP ratio from 23.6 to 9.6 %, increased the TP titer by 85 %, and the specific production yield from 210 to 330 mg TP per gram of dry cell weight. Furthermore, the TP recovery yield in the purified IBs was improved by ~20 %. Adding together, ~105 % more TP recovered in the purified IBs from per liter of fermentation broth for the new process than the standard process. The basic principles of lowering metabolic and production rates should be applicable to other recombinant protein production in IBs by fed-batch fermentations.

  11. Advances in fed-batch ethanol fermentation technologies%乙醇补料发酵技术研究进展

    Institute of Scientific and Technical Information of China (English)

    郭加明; 杨功勋; 胡纯铿; 詹美蓉; 张新华


    As a renewable and clean energy,development and utilization of fuel ethanol has attracted much attention,and consequently there is increasingly in-depth research of the fermentation processes. In recent years,various approaches of fed-batch fermentation have been attempted to investigate their suitability for ethanol production and eventually demonstrated their good potential with the advantages,such as decreased substrate inhibition,and alleviated effects of the toxic compounds released in the dilute-acid pretreatment of lignocellulose. However,due to the existing problems,such as complexity of ethanol fermentation process and lack of in-depth investigations into feeding control strategies,the application of this technology to large-scale production of fuel ethanol has been restricted. This paper reviews major progress of ethanol fed-batch fermentation technologies , particularly focusing on the application of fed-batch approaches to ethanol production using lignocellulose as feedstock and high concentration ethanol fermentation,as well as specific fed-batch control strategies. Finally,more efforts should be made to understand fed-batch ethanol fermentation kinetics and feeding control theory,and to develop new types of sensors and online monitoring technologies.%作为一种可再生的清洁能源,燃料乙醇的开发利用备受关注,对其发酵工艺的研究也日益深入。近年来,补料发酵工艺逐渐应用于燃料乙醇的生产研究中,并以其降低基质抑制和减轻纤维素稀酸水解液中有毒成分的影响等优点而显示了良好的发展潜力,但由于发酵过程的复杂性和对补料控制策略的研究尚不深入等存在的问题,使该技术在燃料乙醇规模化生产中的应用受到制约。本文介绍了国内外乙醇补料发酵研究的主要进展,着重概述补料发酵技术在乙醇两大重要发酵工艺--纤维素乙醇工艺和超高浓度乙醇发酵工艺中的应用以及补料调

  12. Fed batch enzymatic saccharification of food waste improves the sugar concentration in the hydrolysates and eventually the ethanol fermentation by Saccharomyces cerevisiae H058

    Directory of Open Access Journals (Sweden)

    Shoubao Yan


    Full Text Available The enzymatic hydrolysis of food waste by commercially available enzymes and the subsequent ethanol fermentation of the hydrolysates by Saccharomyces cerecisiae H058 were studied in this work. The optimum batch enzymatic conditions were found to be saccharification pH of 4.5, temperature of 55!, glucoamylase concentration of 120 u/g, α-amylase concentration of 10 u/g, solid-liquid ratio of 1: 0.75 (w/w. Fed batch hydrolysis process was started with a solid-liquid ratio of 1: 1 (w/w, with solid food waste added at time lapse of 2 h to get a final solid-liquid ratio of 1: 0.5 (w/w. After 4 h of reaction, the reducing sugar concentration reached 194.43 g/L with a enzymatic digestibility of 93.12%. Further fermentation of the batch and fed batch enzymatic hydrolysates, which contained reducing sugar concentration of 131.41 and 194.43 g/L respectively, was performed using Saccharomyces cerevisiae H058, 62.93 and 90.72 g/L ethanol was obtained within 48 h.

  13. An investigation into the preservation of microbial cell banks for α-amylase production during 5 l fed-batch Bacillus licheniformis fermentations. (United States)

    Hancocks, Nichola H; Thomas, Colin R; Stocks, Stuart M; Hewitt, Christopher J


    Fluorescent staining techniques were used for a systematic examination of methods used to cryopreserve microbial cell banks. The aim of cryopreservation here is to ensure subsequent reproducible fermentation performance rather than just post thaw viability. Bacillus licheniformis cell physiology post-thaw is dependent on the cryopreservant (either Tween 80, glycerol or dimethyl sulphoxide) and whilst this had a profound effect on the length of the lag phase, during subsequent 5 l fed-batch fermentations, it had little effect on maximum specific growth rate, final biomass concentration or α-amylase activity. Tween 80 not only protected the cells during freezing but also helped them recover post-thaw resulting in shorter process times.

  14. Characteristics of corn stover pretreated with liquid hot water and fed-batch semi-simultaneous saccharification and fermentation for bioethanol production. (United States)

    Li, Xuezhi; Lu, Jie; Zhao, Jian; Qu, Yinbo


    Corn stover is a promising feedstock for bioethanol production because of its abundant availability in China. To obtain higher ethanol concentration and higher ethanol yield, liquid hot water (LHW) pretreatment and fed-batch semi-simultaneous saccharification and fermentation (S-SSF) were used to enhance the enzymatic digestibility of corn stover and improve bioconversion of cellulose to ethanol. The results show that solid residues from LHW pretreatment of corn stover can be effectively converted into ethanol at severity factors ranging from 3.95 to 4.54, and the highest amount of xylan removed was approximately 89%. The ethanol concentrations of 38.4 g/L and 39.4 g/L as well as ethanol yields of 78.6% and 79.7% at severity factors of 3.95 and 4.54, respectively, were obtained by fed-batch S-SSF in an optimum conditions (initial substrate consistency of 10%, and 6.1% solid residues added into system at the prehydrolysis time of 6 h). The changes in surface morphological structure, specific surface area, pore volume and diameter of corn stover subjected to LHW process were also analyzed for interpreting the possible improvement mechanism.

  15. Characteristics of corn stover pretreated with liquid hot water and fed-batch semi-simultaneous saccharification and fermentation for bioethanol production.

    Directory of Open Access Journals (Sweden)

    Xuezhi Li

    Full Text Available Corn stover is a promising feedstock for bioethanol production because of its abundant availability in China. To obtain higher ethanol concentration and higher ethanol yield, liquid hot water (LHW pretreatment and fed-batch semi-simultaneous saccharification and fermentation (S-SSF were used to enhance the enzymatic digestibility of corn stover and improve bioconversion of cellulose to ethanol. The results show that solid residues from LHW pretreatment of corn stover can be effectively converted into ethanol at severity factors ranging from 3.95 to 4.54, and the highest amount of xylan removed was approximately 89%. The ethanol concentrations of 38.4 g/L and 39.4 g/L as well as ethanol yields of 78.6% and 79.7% at severity factors of 3.95 and 4.54, respectively, were obtained by fed-batch S-SSF in an optimum conditions (initial substrate consistency of 10%, and 6.1% solid residues added into system at the prehydrolysis time of 6 h. The changes in surface morphological structure, specific surface area, pore volume and diameter of corn stover subjected to LHW process were also analyzed for interpreting the possible improvement mechanism.

  16. Improved 5-Aminolevulinic Acid Production with Recombinant Escherichia coli by a Short-term Dissolved Oxygen Shock in Fed-batch Fermentation

    Institute of Scientific and Technical Information of China (English)

    杨俊; 朱力; 傅维琦; 林逸君; 林建平; 岑沛霖


    5-Aminolevulinic acid (ALA) is a common precursor for tetrapyrrole compounds in all kinds of organ-isms and has wide applications in agriculture and medicines. In this study, a new strategy, i.e. short-term dissolved oxygen (DO) shock during aerobic fermentation, was introduced to produce 5-aminolevulinic acid with a recombi-nant E. coli. Effects of duration time of DO shock operation on plasmid concentration, intracellular ALA synthase (ALAS) activity and ALA production were investigated in Erlenmeyer shake flasks. The results indicated that both ALAS activity and ALA yield were enhanced in an anaerobic operation of 45 min in the early exponential phase during fermentation, while they decreased when the anaerobic operation time was further increased to 60 min. The DO shock protocol was confirmed with the fed-batch fermentation in a 15 L fermenter and the ALA production achieved 9.4 g·L-1 (72 mmol·L-1), which is the highest yield in the fermentation broth reported up to now.

  17. An integral term adaptive neural control of fed-batch fermentation biotechnological process; Control neuronal adaptable con termino integral para un proceso biotecnologico de fermentacion por lote alimentado

    Energy Technology Data Exchange (ETDEWEB)

    Baruch, Ieroham; Hernandez, Luis Alberto; Barrera Cortes, Josefina [Centro de Investigacion y de Estudios Avanzados, Instituto Politecnico Nacional, Mexico D.F. (Mexico)


    A nonlinear mathematical model of aerobic biotechnological process of a fed-batch fermentation system is derived using ordinary differential equations. A neurocontrol is applied using Recurrent Trainable Neural Network (RTNN) plus integral term; the first network performs an approximation of the plant's output; the second network generates the control signal so that the biomass concentration could be regulated by the nutrient influent flow rate into the bioreactor. [Spanish] Un modelo matematico no lineal de un proceso biotecnologico aerobio de un sistema de fermentacion por lote alimentado es presentado mediante ecuaciones diferenciales ordinarias. Es propuesto un control utilizando dos redes neuronales recurrentes entrenables (RNRE) con la adicion de un termino integral; la primera red representa un aproximador de la salida de la planta y la segunda genera la senal de control tal que la concentracion de la biomasa pueda ser regulada mediante la alimentacion de un flujo con nutrientes al biorreactor.

  18. High-titer n-butanol production by clostridium acetobutylicum JB200 in fed-batch fermentation with intermittent gas stripping. (United States)

    Xue, Chuang; Zhao, Jingbo; Lu, Congcong; Yang, Shang-Tian; Bai, Fengwu; Tang, I-Ching


    Acetone-butanol-ethanol (ABE) fermentation with a hyper-butanol producing Clostridium acetobutylicum JB200 was studied for its potential to produce a high titer of butanol that can be readily recovered with gas stripping. In batch fermentation without gas stripping, a final butanol concentration of 19.1 g/L was produced from 86.4 g/L glucose consumed in 78 h, and butanol productivity and yield were 0.24 g/L h and 0.21 g/g, respectively. In contrast, when gas stripping was applied intermittently in fed-batch fermentation, 172 g/L ABE (113.3 g/L butanol, 49.2 g/L acetone, 9.7 g/L ethanol) were produced from 474.9 g/L glucose in six feeding cycles over 326 h. The overall productivity and yield were 0.53 g/L h and 0.36 g/g for ABE and 0.35 g/L h and 0.24 g/g for butanol, respectively. The higher productivity was attributed to the reduced butanol concentration in the fermentation broth by gas stripping that alleviated butanol inhibition, whereas the increased butanol yield could be attributed to the reduced acids accumulation as most acids produced in acidogenesis were reassimilated by cells for ABE production. The intermittent gas stripping produced a highly concentrated condensate containing 195.9 g/L ABE or 150.5 g/L butanol that far exceeded butanol solubility in water. After liquid-liquid demixing or phase separation, a final product containing ~610 g/L butanol, ~40 g/L acetone, ~10 g/L ethanol, and no acids was obtained. Compared to conventional ABE fermentation, the fed-batch fermentation with intermittent gas stripping has the potential to reduce at least 90% of energy consumption and water usage in n-butanol production from glucose. Copyright © 2012 Wiley Periodicals, Inc.

  19. 法夫酵母产虾青素的反复分批及反复分批补料发酵%Repeated batch and fed-batch process for astaxanthin production by Phaffia rhodozyma

    Institute of Scientific and Technical Information of China (English)

    肖安风; 倪辉; 李利君; 蔡慧农


    A comparative study of batch and repeated batch process was carried out for astaxanthin fermentation of Phaffia rhodozyma to develop a more economical method for astaxanthin industrial production. In shaking flask fermentation, the change of biomass and astaxanthin production was studied to compare the five-day cycle with four-day cycle of repeated batch culture of P. thodozyma. Astaxanthin production increased at first and then decreased subsequently in seven cycles, yet the yield of astaxanthin in the next six cycles remains higher than that of the first cycle. Comparing the average production of astaxanthin in the seven cycles, four-day cycle performed even better than five-day cycle. Subsequently, a repeated fed-batch process was used in a 5-1 bioreactor. The experimental data showed that biomass and astaxanthin production of the second batch could reach the level of the first batch, no matter that the carbon source was glucose or hydrolysis sugar of starch. This result showed that this strain had good stability, and thus repeated batch and fed-batch process could be applied in astaxanthin fermentation for economical purpose.%以生物量和虾青素产量为指标,考察法夫酵母多批次半连续培养产虾青素的稳定性.实验结果显示,在摇瓶上分别以4 d和5 d为周期反复分批培养法夫酵母,虾青素产量呈现先增加再下降的趋势,但第2代至第7代虾青素产量仍高于第 1代,并且4 d为周期的虾青素平均产量略高于5 d的.在5 L罐法夫酵母进行反复分批补料发酵中,不管是补加30%的葡萄糖还是补加30%的淀粉水解糖,第2个批次发酵的生物量和虾青素产量均达到第1个批次的水平,表明菌种稳定性较好.

  20. Enhanced production of glycyrrhetic acid 3-O-mono-β-D-glucuronide by fed-batch fermentation using pH and dissolved oxygen as feedback parameters☆

    Institute of Scientific and Technical Information of China (English)

    Bo Lü; Xiaogang Yang; Xudong Feng; Chun Li


    Glycyrrhetic acid 3-O-mono-β-D-glucuronide (GAMG), the major functional ingredient in licorice, has widespread applications in food, pharmacy and cosmetics industry. The production of GAMG through Penicil ium purpurogenum Li-3 cultivation was for the first time performed through both batch and fed-batch processes in bioreactors. In batch process, under optimal conditions (pH 5.0, temperature 32 °C, agitation speed 100 r·min−1), 3.55 g·L−1 GAMG was obtained in a 2.5 L fermentor. To further enhance GAMG production, a fine fed-batch process was developed by using pH and DO as feedback parameters. Starting from 48 h, 100 ml 90 g·L−1 substrate Glycyrrhizin (GL) was fed each time when pH increased to above 5.0 and DO was increased to above 80%. This strategy can significantly enhance GAMG production:the achieved GL conversion was 95.34%with GAMG yield of 95.15%, and GAMG concentration was 16.62 g·L−1 which was 5 times higher than that of batch. Then, a two-step separation strat-egy was established to separate GAMG from fermentation broth by crude extraction of 15 ml column packed with D101 resin followed by fine purification with preparative C18 chromatography. The obtained GAMG purity was 95.79%. This study provides a new insight into the industrial bioprocess of high-level GAMG production.

  1. Fed-batch fermentation for n-butanol production from cassava bagasse hydrolysate in a fibrous bed bioreactor with continuous gas stripping. (United States)

    Lu, Congcong; Zhao, Jingbo; Yang, Shang-Tian; Wei, Dong


    Concentrated cassava bagasse hydrolysate (CBH) containing 584.4 g/L glucose was studied for acetone-butanol-ethanol (ABE) fermentation with a hyper-butanol-producing Clostridium acetobutylicum strain in a fibrous bed bioreactor with gas stripping for continuous butanol recovery. With periodical nutrient supplementation, stable production of n-butanol from glucose in the CBH was maintained in the fed-batch fermentation over 263 h with an average sugar consumption rate of 1.28 g/L h and butanol productivity of 0.32±0.03 g/L h. A total of 108.5 g/L ABE (butanol: 76.4 g/L, acetone: 27.0 g/L, ethanol: 5.1 g/L) was produced, with an overall yield of 0.32±0.03 g/g glucose for ABE and 0.23±0.01 g/g glucose for butanol. The gas stripping process generated a product containing 10-16% (w/v) of butanol, ~4% (w/v) of acetone, a small amount of ethanol (butanol solution of ~64% (w/v) after phase separation. Copyright © 2011 Elsevier Ltd. All rights reserved.

  2. Expression of a mutated SPT15 gene in Saccharomyces cerevisiae enhances both cell growth and ethanol production in microaerobic batch, fed-batch, and simultaneous saccharification and fermentations. (United States)

    Seong, Yeong-Je; Park, Haeseong; Yang, Jungwoo; Kim, Soo-Jung; Choi, Wonja; Kim, Kyoung Heon; Park, Yong-Cheol


    The SPT15 gene encodes a Saccharomyces cerevisiae TATA-binding protein, which is able to globally control the transcription levels of various metabolic and regulatory genes. In this study, a SPT15 gene mutant (S42N, S78R, S163P, and I212N) was expressed in S. cerevisiae BY4741 (BSPT15-M3), of which effects on fermentative yeast properties were evaluated in a series of culture types. By applying different nitrogen sources and air supply conditions in batch culture, organic nitrogen sources and microaerobic condition were decided to be more favorable for both cell growth and ethanol production of the BSPT15-M3 strain than the control S. cerevisiae BY4741 strain expressing the SPT15 gene (BSPT15wt). Microaerobic fed-batch cultures of BSPT15-M3 with glucose shock in the presence of high ethanol content resulted in a 9.5-13.4% higher glucose consumption rate and ethanol productivity than those for the BSPT15wt strain. In addition, BSPT15-M3 showed 4.5 and 3.9% increases in ethanol productivity from cassava hydrolysates and corn starch in simultaneous saccharification and fermentation processes, respectively. It was concluded that overexpression of the mutated SPT15 gene would be a potent strategy to develop robust S. cerevisiae strains with enhanced cell growth and ethanol production abilities.

  3. L-缬氨酸高产菌XQ-8补料分批发酵的研究%Study on the fed-batch fermentation of L-valine hyper-producer XQ-8

    Institute of Scientific and Technical Information of China (English)

    张伟国’; 钱和; 乎守涛; 刘康乐; 程国平; 张苏龙; 聂晓东


    在分批发酵优化条件基础上,通过对补料分批发酵方式发酵过程的各种参数,包括产酸率、转化率和发酵周期等进行了研究,确定了L-缬氨酸高产菌XQ-8补料分批发酵的最优条件。在最优补料分批发酵奈件下发酵72h左右,L-缬氨酸产量达72g/L,糖酸转化率达38%以上,其结果明显优于分批培养。%On the basis of optimization of batch fermentation conditions, by way of the various parameters of the fed -batch fermentation process,including valine production,glucose/valine conversion rate and fermentation period were studied to determine optimal fed-batch fermentation conditions of the L-valine producer XQ-8.1n the optimal fed-batch fermentation conditions for about 72h,L-valine production was up to 72g/L,glucose/valine conversion rate was more than 38% ,the results were Petter than batch fermentation.

  4. In situ phenol removal from fed-batch fermentations of solvent tolerant Pseudomonas putida S12 by pertraction

    NARCIS (Netherlands)

    Heerema, L.; Wierckx, N.; Roelands, C.P.M.; Hanemaaijer, J.H.; Goetheer, E.L.V.; Verdoes, D.; Keurentjes, J.


    In situ phenol pertraction with 1-octanol has been experimentally studied to improve the production of the model component phenol by a recombinant strain of Pseudomonas putida S12. When the phenol concentration in the reactor reaches 2mM, the cells in fermentations without phenol removal are inhibit

  5. Ethanol production from biomass by repetitive solid-state fed-batch fermentation with continuous recovery of ethanol

    Energy Technology Data Exchange (ETDEWEB)

    Moukamnerd, Churairat; Kino-oka, Masahiro; Sugiyama, Minetaka; Kaneko, Yoshinobu; Harashima, Satoshi; Katakura, Yoshio [Osaka Univ. (Japan). Dept. of Biotechnology; Boonchird, Chuenchit [Mahidol Univ., Bangkok (Thailand). Dept. of Biotechnology; Noda, Hideo [Kansai Chemical Engineering Co., Ltd., Amagasaki (Japan); Ninomiya, Kazuaki [Kanazawa Univ. (Japan). Inst. of International Environment Technology; Shioya, Suteaki [Sojo Univ., Kumamoto (Japan). Dept. of Applied Life Science


    To save cost and input energy for bioethanol production, a consolidated continuous solid-state fermentation system composed of a rotating drum reactor, a humidifier, and a condenser was developed. Biomass, saccharifying enzymes, yeast, and a minimum amount of water are introduced into the system. Ethanol produced by simultaneous saccharification and fermentation is continuously recovered as vapor from the headspace of the reactor, while the humidifier compensates for the water loss. From raw corn starch as a biomass model, 95 {+-} 3, 226 {+-} 9, 458 {+-} 26, and 509 {+-} 64 g l{sup -1} of ethanol solutions were recovered continuously when the ethanol content in reactor was controlled at 10-20, 30-50, 50-70 and 75-85 g kg-mixture{sup -1}, respectively. The residue showed a lesser volume and higher solid content than that obtained by conventional liquid fermentation. The cost and energy for intensive waste water treatment are decreased, and the continuous fermentation enabled the sustainability of enzyme activity and yeast in the system. (orig.)

  6. Ethanol production from Sorghum bicolor using both separate and simultaneous saccharification and fermentation in batch and fed batch systems

    DEFF Research Database (Denmark)

    Mehmood, Sajid; Gulfraz, M.; Rana, N. F.;


    The objective of this work was to find the best combination of different experimental conditions during pre-treatment, enzymatic saccharification, detoxification of inhibitors and fermentation of Sorghum bicolor straw for ethanol production. The optimization of pre-treatment using different...

  7. Enhancement of biomass and fermentation activity of surplus brewers' yeast in a fed-batch process

    Energy Technology Data Exchange (ETDEWEB)

    Strel, B. (Ministry of Science and Technology, Ljubljana (Slovenia). Industrial Property Office); Grba, S.; Maric, V. (Zagreb Univ. (Croatia). Faculty of Food Technology and Biotechnology)


    The growth of surplus brewers' yeast in a fedbatch process was studied with the aim of increasing the fermentation activity of the yeast cells and of optimizing the growth conditions: 20 h cultivation at 30deg C and pH 5.0-5.5 using beet molasses as substrate, with a regulated feeding rate, showed satisfactory results. Under the chosen conditions, the final amount of biomass increased more than fivefold, achieving a specific growth rate of 0.1 h[sup -1] and substrate yield coefficient of 0.54 g.g[sup -1]. The increase in fermentation activity of yeast cells during cultivation correlated very well with the concentration of reduced glutathione, which increased from 1.2 to 2.7 mg.g[sup -1] (dry matter). At the same time the fermentation activity increased fivefold, which related to the nitrogen content of the yeast cells. Ethanol formation throughout the cultivation did not exceed 0.5 g.l[sup -1]. (orig.).

  8. Bioconversion of sawdust into ethanol using dilute sulfuric acid-assisted continuous twin screw-driven reactor pretreatment and fed-batch simultaneous saccharification and fermentation. (United States)

    Kim, Tae Hyun; Choi, Chang Ho; Oh, Kyeong Keun


    Ethanol production from poplar sawdust using sulfuric acid-assisted continuous twin screw-driven reactor (CTSR) pretreatment followed by simultaneous saccharification and fermentation (SSF) was investigated. Pretreatment with high acid concentration increased the cellulose content in the pretreated solid (74.9-76.9% in the range of 4.0-5.5wt.% H(2)SO(4)). The sugar content (XMG; xylan+mannan+galactan) in the treated-solid was 11.1-15.2% and 0.9-5.7% with 0.5wt.% and 7.0wt.%, respectively. The XMG recovery yield of the sample treated with 4.0wt.% H(2)SO(4) at 185°C was maximized at 88.6%. Enzymatic hydrolysis test showed a cellulose digestibility of 67.1%, 70.1%, and 73.6% with 15, 30, and 45FPU/g-cellulose, respectively. In the fed-batch SSF tests with initial enzyme addition, the ethanol yield of each stage almost reached a maximum at 28h, 48h, and 56h, respectively, with yields of 63.9% (16.5g/L), 78.4% (30.1g/L), and 81.7% (39.9g/L), respectively.

  9. High-titer lactic acid production by Lactobacillus pentosus FL0421 from corn stover using fed-batch simultaneous saccharification and fermentation. (United States)

    Hu, Jinlong; Lin, Yanxu; Zhang, Zhenting; Xiang, Ting; Mei, Yuxia; Zhao, Shumiao; Liang, Yunxiang; Peng, Nan


    Because the cost of refined sugar substrate and limit of worldwide food availability, lignocellulosic materials are attractive for use in lactic acid (LA) production. In this study, we found Lactobacillus pentosus strain FL0421 produced LA with high yields (0.52-0.82g/g stover) from five NaOH-pretreated and washed agro stovers through simultaneous saccharification and fermentation (SSF). We developed a fed-batch SSF process at 37°C and pH 6.0 using the cellulase of 30FPU/g stover and 10g/L yeast extract in a 5-L bioreactor to produce LA from 14% (w/w) NaOH-pretreated and washed corn stover under non-sterile condition. The LA-titer, yield and productivity reached 92.30g/L, 0.66g/g stover and 1.92g/L/h, respectively; and acetic acid titer and yield reached 34.27g/L and 0.24g/g stover. This study presented a feasible process for LA production from agro stovers and provided a candidate strain for genetic engineering for high-titer and -yield lignocellulosic LA production. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Fed-batch Fermentation Kinetics of Cyclosporine A by Beauveria nivea%雪白白僵菌产环孢菌素A分批补料发酵动力学

    Institute of Scientific and Technical Information of China (English)

    姜俊云; 董惠钧; 闫同顺


    对在30 L发酵罐中雪白白僵菌发酵环孢菌素A的分批补料发酵过程进行了动力学研究。通过对环孢菌素A分批发酵数据进行分析,基于Logistic模型和Luedeking-Piret方程,建立了环孢菌素A发酵动力学模型,包括细胞生长、底物消耗和产物合成回归方程。应用Origin7.5软件对模型进行非线性拟合计算,建立的动力学模型与实验值拟合良好,能较准确反映环孢菌素A分批补料发酵动力学过程。%The kinetic models of the fed-batch fermentation of cyclosporin A in 301 bioreactor were studied. Based on the fed-batch fermentation data and Logistic model, the kinetics models of cyclosporin A fermentation for cell growth, cyclosporin production and substrate consumpsion were built up. The program of origin 7.5 version was used to fit the model. The analysis results showed that the good agreement of predicted values with the experimental values, and that the kinetic models could provide reasonable descriptions for the process of cyclosporin A fed-batch fermenta- tion. The development of fed-batch fermentation kinetics of cyclosporine A would help to regular the fermentation of cyclosporine A and increase the fermentation titer.

  11. The Influence on Fermentation Level of Bacillus coagulans by Adopting Segmented Fed-batch Fermentation Technology%分段式补料批次发酵对凝结芽孢杆菌发酵水平的影响

    Institute of Scientific and Technical Information of China (English)

    高书锋; 孔利华; 周映华; 胡新旭; 吴胜莲; 缪东; 周小玲; 刘惠知


    试验采用分段式补料批次发酵技术对1株畜禽用凝结芽孢杆菌的发酵水平进行了研究,对数期补料促使菌体量大量积累,稳定期补料促进芽孢大量形成,从而达到高菌体量和高芽孢率的目的.试验结果表明,对数期补加淀粉量为总淀粉量的10%,豆粉和鱼粉(质量比为2∶1)补加量为总豆粉和鱼粉量(质量比为2∶1)的5%,补加方式为2次等量补加(间歇10~12 h),发酵水平由分批发酵6.80×109 CFU/mL提高到8.30×109 CFU/mL;稳定期最佳补料浓度为0.10 g/L碳酸钙、0.156 g/L磷酸二氢钠、0.30 g/L蛋氨酸,最佳补料方式为1次性补加,经稳定期补料优化,芽孢率由分批发酵的75.78%提高到85.63%.因此,采用分段式补料批次发酵技术能够进一步提升发酵液的菌体数和芽孢率.%The experiment was aimed to research on fermentation level of Bacillus coagulans strain applied for animals by adopting segmented fed-batch fermentation technology. The fed-batch in exponential and stationary phases respectively increased the number of bacterium and promoted spore formation. The results showed in exponential phase that the quantity of starch additives was 10% of the total,the quantity of soya bean flour and fish meal additives was 5% of the total,the best fed-batch mode was fed twice equally (interval time for 10 to 12 h) ,on this conditions the living bacteria amount could reach 8. 30 × 109 CFU/mL from 6. 80×109 CFU/mL. The results showed in stationary phase that the best fed-batch concentration was calcium carbonate 0. 10 g/L,sodium dihydrogen phosphate 0. 156 g/L and methionine 0. 30 g/L,the best fed-batch mode was one-off feeding,on this conditions the ratio of spore-forming could reach 85. 63% from 75. 78%. Therefore, the research through the experiment on Bacillus coagulans suggested that both the living bacteria and the ratio of spore-forming of fermentation broth further increased by adopting segmented fed-batch

  12. Phenol degradation by Ralstonia eutropha: colorimetric determination of 2-hydroxymuconate semialdehyde accumulation to control feed strategy in fed-batch fermentations. (United States)

    Léonard, D; Youssef, C B; Destruhaut, C; Lindley, N D; Queinnec, I


    Phenol biodegradation by Ralstonia eutropha was modeled in different culture modes to assess phenol feeding in biotechnological depollution processes. The substrate-inhibited growth of R. eutropha was described by the Haldane equation with a Ks of 2 mg/L, a Ki of 350 mg/L and a mumax of 0.41 h(-1). Furthermore, growth in several culture modes was characterized by the appearance of a yellow color, due to production of a metabolic intermediate of the phenol catabolic pathway, 2-hydroxymuconic semialdehyde (2-hms) which was directly correlated to the growth rate and/or the phenol-degradation rate, because these two parameters are coupled (as seen by the constant growth yield of 0.68 g biomass/g phenol whatever the phenol concentration). This correlation between color appearance and metabolic activity was used to develop a control procedure for optimal phenol degradation. A mass-balance equation modeling approach combined with a filtering step using an extended Kalman filter enabled state variables of the biological system to be simulated. A PI controller, using the estimation of the phenol concentration provided by the modeling step, was then built to maintain the phenol concentration at a constant set-point of 0.1 g/L which corresponded to a constant specific growth rate of 0.3 h(-1), close to the maximal specific growth value of the strain. This monitoring strategy, validated for two fed-batch cultures, could lead, in self-cycling fermentation systems, to a productivity of more than 19 kg of phenol consumed/m(3)/d which is the highest value reported to date in the literature. This system of monitoring metabolic activity also protected the bacterial culture against toxicity problems due to the transient accumulation of phenol. Copyright 1999 John Wiley & Sons, Inc.

  13. Biogas Production from Protein-Rich Biomass: Fed-Batch Anaerobic Fermentation of Casein and of Pig Blood and Associated Changes in Microbial Community Composition (United States)

    Kovács, Etelka; Wirth, Roland; Maróti, Gergely; Bagi, Zoltán; Rákhely, Gábor; Kovács, Kornél L.


    It is generally accepted as a fact in the biogas technology that protein-rich biomass substrates should be avoided due to inevitable process inhibition. Substrate compositions with a low C/N ratio are considered difficult to handle and may lead to process failure, though protein-rich industrial waste products have outstanding biogas generation potential. This common belief has been challenged by using protein-rich substrates, i.e. casein and precipitated pig blood protein in laboratory scale continuously stirred mesophilic fed-batch biogas fermenters. Both substrates proved suitable for sustained biogas production (0.447 L CH4/g protein oDM, i.e. organic total solids) in high yield without any additives, following a period of adaptation of the microbial community. The apparent key limiting factors in the anaerobic degradation of these proteinaceous materials were the accumulation of ammonia and hydrogen sulfide. Changes in time in the composition of the microbiological community were determined by next-generation sequencing-based metagenomic analyses. Characteristic rearrangements of the biogas-producing community upon protein feeding and specific differences due to the individual protein substrates were recognized. The results clearly demonstrate that sustained biogas production is readily achievable, provided the system is well-characterized, understood and controlled. Biogas yields (0.45 L CH4/g oDM) significantly exceeding those of the commonly used agricultural substrates (0.25-0.28 L CH4/g oDM) were routinely obtained. The results amply reveal that these high-energy-content waste products can be converted to biogas, a renewable energy carrier with flexible uses that can replace fossil natural gas in its applications. Process control, with appropriate acclimation of the microbial community to the unusual substrate, is necessary. Metagenomic analysis of the microbial community by next-generation sequencing allows a precise determination of the alterations in

  14. Biogas production from protein-rich biomass: fed-batch anaerobic fermentation of casein and of pig blood and associated changes in microbial community composition.

    Directory of Open Access Journals (Sweden)

    Etelka Kovács

    Full Text Available It is generally accepted as a fact in the biogas technology that protein-rich biomass substrates should be avoided due to inevitable process inhibition. Substrate compositions with a low C/N ratio are considered difficult to handle and may lead to process failure, though protein-rich industrial waste products have outstanding biogas generation potential. This common belief has been challenged by using protein-rich substrates, i.e. casein and precipitated pig blood protein in laboratory scale continuously stirred mesophilic fed-batch biogas fermenters. Both substrates proved suitable for sustained biogas production (0.447 L CH4/g protein oDM, i.e. organic total solids in high yield without any additives, following a period of adaptation of the microbial community. The apparent key limiting factors in the anaerobic degradation of these proteinaceous materials were the accumulation of ammonia and hydrogen sulfide. Changes in time in the composition of the microbiological community were determined by next-generation sequencing-based metagenomic analyses. Characteristic rearrangements of the biogas-producing community upon protein feeding and specific differences due to the individual protein substrates were recognized. The results clearly demonstrate that sustained biogas production is readily achievable, provided the system is well-characterized, understood and controlled. Biogas yields (0.45 L CH4/g oDM significantly exceeding those of the commonly used agricultural substrates (0.25-0.28 L CH4/g oDM were routinely obtained. The results amply reveal that these high-energy-content waste products can be converted to biogas, a renewable energy carrier with flexible uses that can replace fossil natural gas in its applications. Process control, with appropriate acclimation of the microbial community to the unusual substrate, is necessary. Metagenomic analysis of the microbial community by next-generation sequencing allows a precise determination of the

  15. Biogas production from protein-rich biomass: fed-batch anaerobic fermentation of casein and of pig blood and associated changes in microbial community composition. (United States)

    Kovács, Etelka; Wirth, Roland; Maróti, Gergely; Bagi, Zoltán; Rákhely, Gábor; Kovács, Kornél L


    It is generally accepted as a fact in the biogas technology that protein-rich biomass substrates should be avoided due to inevitable process inhibition. Substrate compositions with a low C/N ratio are considered difficult to handle and may lead to process failure, though protein-rich industrial waste products have outstanding biogas generation potential. This common belief has been challenged by using protein-rich substrates, i.e. casein and precipitated pig blood protein in laboratory scale continuously stirred mesophilic fed-batch biogas fermenters. Both substrates proved suitable for sustained biogas production (0.447 L CH4/g protein oDM, i.e. organic total solids) in high yield without any additives, following a period of adaptation of the microbial community. The apparent key limiting factors in the anaerobic degradation of these proteinaceous materials were the accumulation of ammonia and hydrogen sulfide. Changes in time in the composition of the microbiological community were determined by next-generation sequencing-based metagenomic analyses. Characteristic rearrangements of the biogas-producing community upon protein feeding and specific differences due to the individual protein substrates were recognized. The results clearly demonstrate that sustained biogas production is readily achievable, provided the system is well-characterized, understood and controlled. Biogas yields (0.45 L CH4/g oDM) significantly exceeding those of the commonly used agricultural substrates (0.25-0.28 L CH4/g oDM) were routinely obtained. The results amply reveal that these high-energy-content waste products can be converted to biogas, a renewable energy carrier with flexible uses that can replace fossil natural gas in its applications. Process control, with appropriate acclimation of the microbial community to the unusual substrate, is necessary. Metagenomic analysis of the microbial community by next-generation sequencing allows a precise determination of the alterations in

  16. 木醋杆菌分批补料发酵法生产广式米醋%Fed-batch fermentation by Gluconacetobacter xylinus to produce Guangdong rice vinegar

    Institute of Scientific and Technical Information of China (English)

    傅亮; 易九龙; 陈思谦; 吴炳鸿


    The feasibility of fed-batch fermentation to improve the total acidity of Guangdong rice vinegar was investigated. The main contents include the distribution of Gluconacetobacter xylinus in fermented liquid and bacterial cellulose membrane, the variation of total acidity by single-batch fermentation and fed-batch fermentation, the optimal alcohol content of raw and the effect on total acidity by fed-batch fermentation. The result shows that the cell number of Bacterial cellulose membrane is 300 times than in the fermented liquid and the optimal alcohol content of raw is 5% (V/V). And fed-batch fermentation is advantageous in improving total acidity. The result of orthogonal test shows that the optimal conditions were: the sixth day began to add, add every 2 days one time, every time add 2%(volume of alcohol / volume of fermented liquid) and 3 times. Under this con-dition, the total acidity was at 7. 29 g/100 mL, 73. 6% higher than 4. 2 g/100 mL by single-batch fermentation.%研究分批补料发酵法提高广式米醋总酸度的可行性.主要内容包括发酵过程中木醋杆菌RF4在发酵液及细菌纤维素膜内的菌体数分布比较、单批和分批补料发酵法总酸的变化规律、原料最适酒精度及分批补料发酵法对总酸的影响.结果表明:细菌纤维素膜内的菌体数是发酵液中的300倍左右,最适原料初始酒精度为5% (V/V),分批补料发酵法有利于显著提高总酸度.通过正交优化,分批补料发酵最优工艺为发酵第6天每隔2d补加2%(酒精体积/发酵液体积)的酒精,补加3次,最终酸度可达7.29 g/l00 mL,较单批发酵的4.2 g/l00mL提高73.6%.

  17. Microbial fed-batch production of 1,3-propanediol using raw glycerol with suspended and immobilized Klebsiella pneumoniae. (United States)

    Jun, Sun-Ae; Moon, Chuloo; Kang, Cheol-Hee; Kong, Sean W; Sang, Byoung-In; Um, Youngsoon


    The production of 1,3-propanediol (1,3-PD) was investigated with Klebsiella pneumoniae DSM 4799 using raw glycerol without purification obtained from a biodiesel production process. Fed-batch cultures with suspended cells revealed that 1,3-PD production was more effective when utilizing raw glycerol than pure glycerol (productivity after 47 h of fermentation, 0.84 g L(-1) 1 h(-1) versus 1.51 g L(-1) h(-1) with pure and raw glycerol,respectively). In addition, more than 80 g/L of 1,3-PD was produced using raw glycerol;this is the highest 1,3-PD concentration reported thus far for K. pneumoniae using raw glycerol. Repeated fed-batch fermentation with cell immobilization in a fixed-bed reactor was performed to enhance 1,3-PD production. Production of 1,3-PD increased with the cycle number (1.06 g L(-1) h(-1) versus 1.61 g L(-1) h(-1) at the first and fourth cycle, respectively)due to successful cell immobilization. During 46 cycles of fed-batch fermentation taking place over 1,460 h, a stable and reproducible 1,3-PD production performance was observed with both pure and raw glycerol. Based on our results, repeated fed batch with immobilized cells is an efficient fermentor configuration, and raw glycerol can be utilized to produce 1,3-PD without inhibitory effects caused by accumulated impurities.

  18. Optimization of the fed-batch fermentation process for raspberry wine by response surface methodology%响应面法优化树莓酒流加发酵工艺

    Institute of Scientific and Technical Information of China (English)

    马荣山; 王艳平; 穆晶


    以红树莓为原料,利用响应面法对树莓酒的流加发酵工艺条件进行优化,在单因素基础上,选取酵母接种量、流加糖时间、流加糖量为影响因子,以树莓酒酒精体积分数为响应值进行响应面分析.结果表明,经优化后树莓酒的最佳流加发酵工艺条件为酵母接种量1.0‰,分别在发酵5d、6d共流加糖30mL,发酵10d得到酒精度为10.5%vol的树莓酒.树莓酒酒色玫红,果香柔和,酒质柔顺,所得产品是一种符合现代人健康理念的低度发酵营养酒.%In this study, the process conditions of raspberry wine fed-batch fermentation were optimized using response surface methodology. Based on single factor experiment, the inoculum of yeast, feeding time and quantity of caramel were chosen as influencing factors, and the alcoholicity was selected as response value. It was found that the optimum fed-batch fermentation conditions were as followed: inoculum of active dry yeast 1.0%>, feeding 30ml caramel at 5d and 6d of the fermentation and fermentation time 10d. Under these conditions, alcohol concentration in obtained raspberry wine was 10.5%vol, and displayed a rose pink color, and had a fruity, soft and supple taste. This product is a low-alcohol nutritional wine that caters to modern people' s health concept

  19. The potential of random forest and neural networks for biomass and recombinant protein modeling in Escherichia coli fed-batch fermentations. (United States)

    Melcher, Michael; Scharl, Theresa; Spangl, Bernhard; Luchner, Markus; Cserjan, Monika; Bayer, Karl; Leisch, Friedrich; Striedner, Gerald


    Product quality assurance strategies in production of biopharmaceuticals currently undergo a transformation from empirical "quality by testing" to rational, knowledge-based "quality by design" approaches. The major challenges in this context are the fragmentary understanding of bioprocesses and the severely limited real-time access to process variables related to product quality and quantity. Data driven modeling of process variables in combination with model predictive process control concepts represent a potential solution to these problems. The selection of statistical techniques best qualified for bioprocess data analysis and modeling is a key criterion. In this work a series of recombinant Escherichia coli fed-batch production processes with varying cultivation conditions employing a comprehensive on- and offline process monitoring platform was conducted. The applicability of two machine learning methods, random forest and neural networks, for the prediction of cell dry mass and recombinant protein based on online available process parameters and two-dimensional multi-wavelength fluorescence spectroscopy is investigated. Models solely based on routinely measured process variables give a satisfying prediction accuracy of about ± 4% for the cell dry mass, while additional spectroscopic information allows for an estimation of the protein concentration within ± 12%. The results clearly argue for a combined approach: neural networks as modeling technique and random forest as variable selection tool.

  20. A special reactor design for investigations of mixing time effects in a scaled-down industrial L-lysine fed-batch fermentation process (United States)

    Schilling; Pfefferle; Bachmann; Leuchtenberger; Deckwer


    A specially designed model reactor based on a 42-L laboratory fermentor was equipped with six stirrers (Rushton turbines) and five cylindrical disks. In this model reactor, the mixing time, Theta(90), turned out to be 13 times longer compared with the 42-L standard laboratory fermentor fitted with two Rushton turbines and four wall-fixed longitudinal baffles. To prove the suitability of the model reactor for scaledown studies of mixing-time-dependent processes, parallel exponential fed-batch cultivations were carried out with the leucine-auxotrophic strain, Corynebacterium glutamicum DSM 5715, serving as a microbial test system. L‐Leucine, the process-limiting substrate, was fed onto the liquid surface of both reactors. Cultivations were conducted using the same inoculum material and equal oxygen supply. The model reactor showed reduced sugar consumption (-14%), reduced ammonium consumption (-19%), and reduced biomass formation (-7%), which resulted in a decrease in L-lysine formation (-12%). These findings were reflected in less specific enzyme activity, which was determined for citrate synthase (CS), phosphoenolpyruvate carboxylase (PEP-C), and aspartate kinase (AK). The reduced specific activity of CS correlated with lower CO(2) evolution (-36%) during cultivation. The model reactor represents a valuable tool to simulate the conditions of poor mixing and inhomogeneous substrate distribution in bioreactors of industrial scale. Copyright 1999 John Wiley & Sons, Inc.

  1. High-titer lactic acid production from NaOH-pretreated corn stover by Bacillus coagulans LA204 using fed-batch simultaneous saccharification and fermentation under non-sterile condition. (United States)

    Hu, Jinlong; Zhang, Zhenting; Lin, Yanxu; Zhao, Shumiao; Mei, Yuxia; Liang, Yunxiang; Peng, Nan


    Lactic acid (LA) is an important chemical with various industrial applications. Non-food feedstock is commercially attractive for use in LA production; however, efficient LA fermentation from lignocellulosic biomass resulting in both high yield and titer faces technical obstacles. In this study, the thermophilic bacterium Bacillus coagulans LA204 demonstrated considerable ability to ferment glucose, xylose, and cellobiose to LA. Importantly, LA204 produces LA from several NaOH-pretreated agro stovers, with remarkably high yields through simultaneous saccharification and fermentation (SSF). A fed-batch SSF process conducted at 50°C and pH 6.0, using a cellulase concentration of 30 FPU (filter paper unit)/g stover and 10 g/L yeast extract in a 5-L bioreactor, was developed to produce LA from 14.4% (w/w) NaOH-pretreated non-sterile corn stover. LA titer, yield, and average productivity reached 97.59 g/L, 0.68 g/g stover, and 1.63 g/L/h, respectively. This study presents a feasible process for lignocellulosic LA production from abundant agro stovers.

  2. Use of a stress-minimisation paradigm in high cell density fed-batch Escherichia coli fermentations to optimise recombinant protein production. (United States)

    Wyre, Chris; Overton, Tim W


    Production of recombinant proteins is an industrially important technique in the biopharmaceutical sector. Many recombinant proteins are problematic to generate in a soluble form in bacteria as they readily form insoluble inclusion bodies. Recombinant protein solubility can be enhanced by minimising stress imposed on bacteria through decreasing growth temperature and the rate of recombinant protein production. In this study, we determined whether these stress-minimisation techniques can be successfully applied to industrially relevant high cell density Escherichia coli fermentations generating a recombinant protein prone to forming inclusion bodies, CheY-GFP. Flow cytometry was used as a routine technique to rapidly determine bacterial productivity and physiology at the single cell level, enabling determination of culture heterogeneity. We show that stress minimisation can be applied to high cell density fermentations (up to a dry cell weight of >70 g L(-1)) using semi-defined media and glucose or glycerol as carbon sources, and using early or late induction of recombinant protein production, to produce high yields (up to 6 g L(-1)) of aggregation-prone recombinant protein in a soluble form. These results clearly demonstrate that stress minimisation is a viable option for the optimisation of high cell density industrial fermentations for the production of high yields of difficult-to-produce recombinant proteins, and present a workflow for the application of stress-minimisation techniques in a variety of fermentation protocols.

  3. Fed-batch production of citric acid by Candida lipolytica grown on n-paraffins. (United States)

    Crolla, A; Kennedy, K J


    This study reports on the effects of fermentor agitation and fed-batch mode of operation on citric acid production from Candida lipolytica using n-paraffin as the carbon source. An optimum range of agitation speeds in the 800-1000 rpm range corresponding to Reynolds numbers of 50000-63000 (based on initial batch conditions) seemed to give the best balance between substrate utilization for biomass growth and citric acid production. Application of multiple fed-batch feedings can be used to extend the batch fermentation and increase final citric acid concentrations and product yield. The three-cycle fed-batch system increased overall citric acid yields to 0.8-1.0 g citricacid/g n-paraffin, approximately a 100% improvement in product yield from those observed in the single cycle fed-batch system and a 200% improvement over normal batch operation. The three-cycle fed-batch mode of operation also increased the final citric acid concentration to 42 g/l from about 12 and 6g/l for single fed-batch cycle and normal batch modes of operation, respectively. Increased citric acid concentrations in three-cycle fed-batch mode was achieved at longer fermentation times.

  4. Butanol production employing fed-batch fermentation by Clostridium acetobutylicum GX01 using alkali-pretreated sugarcane bagasse hydrolysed by enzymes from Thermoascus aurantiacus QS 7-2-4. (United States)

    Pang, Zong-Wen; Lu, Wei; Zhang, Hui; Liang, Zheng-Wu; Liang, Jing-Juan; Du, Liang-Wei; Duan, Cheng-Jie; Feng, Jia-Xun


    Sugarcane bagasse (SB) is a potential feedstock for butanol production. However, biological production of butanol from SB is less economically viable. In this study, evaluation of eight pretreatments on SB showed that alkali pretreatment efficiently removed lignin from SB while retaining the intact native structure of the released microfibrils. In total, 99% of cellulose and 100% of hemicellulose in alkali-pretreated SB were hydrolysed by enzymes from Thermoascus aurantiacus. The hydrolysate was used to produce butanol in a fed-batch fermentation by Clostridium acetobutylicum. At 60h, 14.17 and 21.11gL(-1) of butanol and acetone-butanol-ethanol (ABE) were produced from 68.89gL(-1) of total sugars, respectively, yielding 0.22 and 0.33gg(-1) of sugars. The maximum yield of butanol and ABE reached 15.4g and 22.9g per 100g raw SB, respectively. This established process may have potential application for butanol production from SB. Copyright © 2016 Elsevier Ltd. All rights reserved.

  5. Scaling up a virginiamycin production by a high-yield Streptomyces virginiae VKM Ac-2738D strain using adsorbing resin addition and fed-batch fermentation under controlled conditions. (United States)

    Dzhavakhiya, Vakhtang; Savushkin, Vyacheslav; Ovchinnikov, Alexander; Glagolev, Vladislav; Savelyeva, Veronika; Popova, Evgeniya; Novak, Nikita; Glagoleva, Elena


    Virginiamycin produced by Streptomyces virginiae as a natural mix of macrocyclic peptidolactones M and S is widely used in the industrial production of ethanol fuel and as an antibiotic feed additive for cattle and poultry. Its main antimicrobial components, M1 and S1 factors, act synergistically if the M1:S1 ratio in the final product is 70-75:25-30. This fact significantly complicates the development of stable high-yield strains suitable for industrial application. In the previous work, authors obtained a mutant S. virginiae VKM Ac-2738D strain, characterized by a high productivity in flasks and the optimum M1:S1 ratio (75:25) in the final product. In this study, the scale-up of the virginiamycin production by VKM AC-2738D from shake flasks to a pilot-scale (100 L) stirred fermentor was carried out and the possibility of the in situ use of synthetic adsorbing resins to remove virginiamycin from culture broth was assessed. After the optimization of pH and dissolved oxygen concentration (6.8-7.0 and 50%, respectively), the fed-batch fermentation of VKM Ac-2738D with continuous addition of 50% sucrose solution (5 g/L/day starting from 48 h of fermentation) resulted in a final virginiamycin titer of 4.9 g/L. Among four tested resins, Diaion(®) HP21 added to fermentation medium prior to sterilization absorbed 98.5% of the total virginiamycin that simplifies its further recovery procedure and increased its total titer to 5.6 g/L at the M1:S1 ratio of 74:26. The developed technology has several important advantages, which include (1) the optimum M1:S1 ratio in the final product, (2) the possibility to use sucrose as a carbon source instead of traditionally used and more expensive glucose or D-maltose, and (3) selective binding of up to 98.5% of produced virginiamycin on the adsorbing resin.

  6. Study on Cysteine Addition in Fed-Batch Fermentation of S-Adenosylmethionine Production by Saccharomyces Cerevisiae%产腺苷甲硫氨酸酿酒酵母半胱氨酸补料工艺的研究

    Institute of Scientific and Technical Information of China (English)

    夏毅; 杨依顺; 周长林; 汪维云


    /L which was equivalent to reducing sugar feeding as 0. 8 g/L-h in fed-batch,after 16 h then adding 2 mmol/L cysteine,the maximum DCW reached 15. 50 g/L at 26 h and the concentration of SAM was improved to 5. 02 g/L at 34 h in 36 h fermentation. After optimization of fermentation process,concentration of SAM in fermented liquid increased by 43. 8% than using glucose as carbon source without addition of cysteine. In this study, both molasses as carbon source and cysteine addition were helpful to improve the SAM accumulation.


    Institute of Scientific and Technical Information of China (English)

    贾钧辉; 郑宇; 杨青娟; 张艳春; 申雁冰; 王敏


    本论文对生防枯草芽孢杆菌B579补料分批发酵工艺进行了优化,提高了菌体浓度和最终芽孢浓度,为生防菌剂的大规模生产奠定了基础.为获得较高的最终芽孢浓度,分别优化了葡萄糖补加时机、浓度控制范围以及发酵过程pH.利用7L发酵罐,当葡萄糖浓度降至3.0g/L时开始连续补加葡萄糖至3.0-6.0 g/L,发酵过程控制pH为7.0,培养24h菌体浓度达到3.9×1010CFU/mL,继续培养至40h芽孢浓度达到2.8×1010 CFU/mL,分别是分批发酵的7.5倍和7倍.发酵过程葡萄糖浓度对菌体生长和芽孢形成有较大影响,过高的葡萄糖浓度会抑制芽孢的生成,发酵过程控制合适的葡萄糖浓度有利于菌体浓度和芽孢浓度的提高.%In this research, the cell concentration and the final spore concentration were improved by optimizing the fed - batch fermentation process of biocontrol Bacillus subtilis B579, and this study provide the basis for biopesticide production in large scale. For obtaining a higher concentration of spores, the glucose - feed time, controlled glucose concentration and pH of medium was optimized, respectively. In a 7 L fermenter, the glucose was supplemented when its concentration was below 3. 0 g/L and was controlled between 3.0 - 6.0 g/L. The fermentation pH was controlled as 7.0. In this condition, the cell concentration was 3.9 1010CFU/mL after 24 h cultivation and the spore concentration of 2. 8 1010CFU/mL was achieved after 40 h cultivation, respectively, which represent a 7.5 -fold and 7 —fold increasement compared with those of batch fermentation. Glucose concentration showed a significant effect on cell growth and sporulation. It is important to control glucose -concentration for improving cell growth and the final spore concentration, because high concentration of glucose will inhibit the sporulation B. subtilis.

  8. Study on the Effects of Fed-batch Culture Method of Escherichia coli XD-12 Fermentation%流加培养方式对大肠杆菌XD-12发酵的影响研究

    Institute of Scientific and Technical Information of China (English)

    潘自皓; 顾薇; 潘扬


    [Objective] The research aimed to study the fed-batch culture method of Escherichia coli and enhance the fermentation concentration of transaminase donor.E. coli XD-12. [Method] The effects of carbon source feeding, nitrogen source feeding,feeding with pH control on E. coli fermentation were studied to obtain the optimal culture conditions. [ Result] The optimal cultural conditions lot transaminase-produeing E.coli were as Mows; temperature of 37 ℃ .agitation speed of 400 r/min,aeration quantity of 1.5 L/min,initial pH of medium of 7.0,for controlling fermentation process pH of 7.5,initial glucose concentration of 5 g/L,initial nitrogen source of 5 g/L peptone + 1.5 g/L beef extract, 120 g/L glucose solution was intermittently fed at an interval of 2 h after glucose concentration declined to 2 g/L And 15 g/L peptone and 4. 5 g/L beef extract were intermittently fed at an interval of 2 h after 8 hours. After culture 24 h under these conditions, the cell dry weight concentration of E.coli reached 9.66 g/L,104.7% higher than that in batch culture. [ Conclusion] This research was of an important realistic significance for reducing the production cost of preparing L-phenylalanine by enzyme method,enhancing the production efficiency of L-phenylala-nine,sufficing the increasing market requirements of L-phenylalanine.%[目的]研究大肠杆菌流加培养方式,提高转氨醇供体——大肠杆菌XD-12的发酵浓度.[方法]通过研究碳源流加、氮源流加、pH控制流加对发酵的影响,获得了优化的培养条件.[结果]产转氨酶大肠杆菌的最佳培养条件为:温度37℃,搅拌转速500 r/min,通气量1.5 L/min,培养基初始pH为7.0,控制发酵过程pH为7.5,初始葡萄糖浓度5 g/L,初始氮源为5g/L蛋白胨+1.5 g/L牛肉膏,从葡萄糖浓度下降为2 g/L开始每隔2h问歇流加120 g/L的糖,从8h起每隔2h间歇流加15 9/L蛋白胨+4.5 g/L牛肉膏.在此条件下培养24h,大肠杆菌的茵体干重浓度达9.66 g

  9. Fed-batch pediocin production by Pediococcus acidilactici NRRL B-5627 on whey. (United States)

    Pérez Guerra, Nelson; Bernárdez, Paula Fajardo; Agrasar, Ana Torrado; López Macías, Cristina; Castro, Lorenzo Pastrana


    Cell growth and pediocin production by Pediococcus acidilactici NRRL B-5627 on whey were compared by using batch fermentation and re-alkalized fed-batch fermentation. The batch fermentations were performed on DWG [DW (diluted whey) supplemented with 1% (w/v) glucose], DWYE [DW supplemented with 2% (w/v) yeast extract] and DWGYE (DW supplemented with 1% glucose plus 2% yeast extract) media. The fed-batch culture on DWYE medium was fed with a mixture of concentrated whey (48 g of total sugars/l) supplemented with 2% yeast extract and 400 g/l concentrated glucose. The re-alkalized fed-batch culture was characterized by higher biomass (6.57 g/l) and pediocin [517.6 BU (bacteriocin activity units)/ml] concentrations compared with the batch processes on MRS (de Man, Rogosa and Sharpe) broth (1.76 g/l and 493.2 BU/ml), DW (0.17 g/l and 57.7 BU/ml), DWG (0.14 g/l and 53.6 BU/ml), DWYE (1.43 g/l and 187.6 BU/ml) and DWGYE (1.28 g/l and 167.3 BU/ml) media. A mixed acid fermentation was observed during the growth of P. acidilactici NRRL B-5627 in the fed-batch culture on DWYE medium, and other products (acetic acid and ethanol) in addition to lactic acid accumulated in the medium. Mathematical models were set up to describe fed-batch production of biomass and pediocin by P. acidilactici. The models developed offer a better fit and a more realistic description of the experimental biomass and pediocin production data when compared with the logistic and Luedeking and Piret model.

  10. Batch and fed-batch production of butyric acid by Clostridium butyricum ZJUCB

    Institute of Scientific and Technical Information of China (English)

    HE Guo-qing; KONG Qing; CHEN Qi-he; RUAN Hui


    The production of butyric acid by Clostridium butyricum ZJUCB at various pH values was investigated. In order to study the effect of pH on cell growth, butyric acid biosynthesis and reducing sugar consumption, different cultivation pH values ranging from 6.0 to 7.5 were evaluated in 5-L bioreactor. In controlled pH batch fermentation, the optimum pH for cell growth and butyric acid production was 6.5 with a cell yield of 3.65 g/L and butyric acid yield of 12.25 g/L. Based on these results, this study then compared batch and fed-batch fermentation of butyric acid production at pH 6.5. Maximum value (16.74 g/L) of butyric acid concentration was obtained in fed-batch fermentation compared to 12.25 g/L in batch fermentation. It was concluded that cultivation under fed-batch fermentation mode could enhance butyric acid production significantly (P<0.01) by C. butyricum ZJUCB.

  11. Improved production of human type II procollagen in the yeast Pichia pastoris in shake flasks by a wireless-controlled fed-batch system

    Directory of Open Access Journals (Sweden)

    Myllyharju Johanna


    Full Text Available Abstract Background Here we describe a new technical solution for optimization of Pichia pastoris shake flask cultures with the example of production of stable human type II collagen. Production of recombinant proteins in P. pastoris is usually performed by controlling gene expression with the strong AOX1 promoter, which is induced by addition of methanol. Optimization of processes using the AOX1 promoter in P. pastoris is generally done in bioreactors by fed-batch fermentation with a controlled continuous addition of methanol for avoiding methanol toxification and carbon/energy starvation. The development of feeding protocols and the study of AOX1-controlled recombinant protein production have been largely made in shake flasks, although shake flasks have very limited possibilities for measurement and control. Results By applying on-line pO2 monitoring we demonstrate that the widely used pulse feeding of methanol results in long phases of methanol exhaustion and consequently low expression of AOX1 controlled genes. Furthermore, we provide a solution to apply the fed-batch strategy in shake flasks. The presented solution applies a wireless feeding unit which can be flexibly positioned and allows the use of computer-controlled feeding profiles. By using the human collagen II as an example we show that a quasi-continuous feeding profile, being the simplest way of a fed-batch fermentation, results in a higher production level of human collagen II. Moreover, the product has a higher proteolytic stability compared to control cultures due to the increased expression of human collagen prolyl 4-hydroxylase as monitored by mRNA and protein levels. Conclusion The recommended standard protocol for methanol addition in shake flasks using pulse feeding is non-optimal and leads to repeated long phases of methanol starvation. The problem can be solved by applying the fed-batch technology. The presented wireless feeding unit, together with an on-line monitoring

  12. Modified Multi-Population Genetic Algorithm for Yeast Fed-batch Cultivation Parameter Identification

    Directory of Open Access Journals (Sweden)

    Angelova M.


    Full Text Available In this work, a modified multi-population genetic algorithm is developed for the purpose of parameter identification of fermentation process model. Modified multi-population genetic algorithm is similar to the multi-population one and its development is instigated by modified genetic algorithm, similar to simple one. A comparison of four types of genetic algorithms, namely simple, modified, multipopulation and modified multi-population is presented for parameter identification of a fed-batch cultivation of Saccharomyces cerevisiae

  13. L-lysine Production by Phased pH Feedback Fed-batch Fermentation Based on High Throughput Bioreactor%基于高通量生物反应器和分段式pH反馈补料技术发酵生产L-赖氨酸

    Institute of Scientific and Technical Information of China (English)

    梁恒宇; 林海龙; 孙际宾; 卢宗梅; 陈博; 孙村民


    利用高通量生物反应器,以在线监测的pH为直接反馈补料信号,以葡萄糖、氨水和硫酸铵混合溶液为流加液进行补料发酵生产L-赖氨酸。当流加液中葡萄糖含量均为360 g/L时,对流加液中硫酸铵添加量、氨水添加量和发酵培养基接种量进行了单因素优化,确定一段式流加培养最佳条件为氨水添加量180 mL/L、硫酸铵添加量40 g/L、接种量为5 mL/45 mL培养基。分段式补料培养研究结果表明,在赖氨酸发酵的不同阶段采用不同配比的流加液进行分段式培养可以进一步提高赖氨酸的产酸浓度,同时降低残糖和残铵氮含量。三段式pH反馈补料发酵可以将赖氨酸产酸浓度提高到(56.85±0.98) g/L,与二段式和一段式相比分别提高8.65%和23.64%。%L-lysine fed-batch fermentations were carried out in high throughput bioreactors, and pH was used as a direct feedback signal to feed the solution mixed with glucose, ammonia water and ammonium sulfate. The single factor optimization experiments on addition amount of ammonia water, ammonium sulfate and inoculation quantity, when glucose level in feeding liquid was 360 g/L, were developed. The best composition of one section fermentation feeding liquid was ammonia water 180 mL/L, ammonium sulfate 40 g/L and inoculation quantity 5 mL/45 mL. The results of phased fed-batch showed that the yield of L-lysine was increased meanwhile the level residual sugar and ammonium was decreased, when feeding liquid with different composition was used at different fermentation stages. The production was enhanced by (56.85 ±0.98) g/L at the end of tri-section pH feedback fed-batch fermentation, which was increased by 8.65%and 23.64%respectively compared with bi-section and one section styles.

  14. Development of fed-batch profiles for efficient biosynthesis of catechol-O-methyltransferase

    Directory of Open Access Journals (Sweden)

    G.M. Espírito Santo


    Full Text Available Catechol-O-methyltransferase (COMT, EC plays a crucial role in dopamine metabolism which has intimately linked this enzyme to some neurodegenerative diseases, such as Parkinson's disease. In recent years, in the attempt of developing new therapeutic strategies for Parkinson's disease, there has been a growing interest in the search for effective COMT inhibitors. In order to do so, large amounts of COMT in an active form are needed, and the best way to achieve this is by up-scaling its production through biotechnological processes. In this work, a fed-batch process for the biosynthesis of the soluble isoform of COMT in Escherichia coli is proposed. This final process was selected through the evaluation of the effect of different dissolved oxygen concentrations, carbon and nitrogen source concentrations and feeding profiles on enzymatic production and cell viability, while controlling various parameters (pH, temperature, starting time of the feeding and induction phases and carbon source concentration during the process. After several batch and fed-batch experiments, a final specific COMT activity of 442.34 nmol/h/mg with approximately 80% of viable cells at the end of the fermentation were achieved. Overall, the results described herein provide a great improvement on hSCOMT production in recombinant bacteria and provide a new and viable option for the use of a fed-batch fermentation with a constant feeding profile to the large scale production of this enzyme.

  15. Monitoring and robust adaptive control of fed-batch cultures of microorganisms exhibiting overflow metabolism [abstract

    Directory of Open Access Journals (Sweden)

    Vande Wouwer, A.


    Full Text Available Overflow metabolism characterizes cells strains that are likely to produce inhibiting by-products resulting from an excess of substrate feeding and a saturated respiratory capacity. The critical substrate level separating the two different metabolic pathways is generally not well defined. Monitoring of this kind of cultures, going from model identification to state estimation, is first discussed. Then, a review of control techniques which all aim at maximizing the cell productivity of fed-batch fermentations is presented. Two main adaptive control strategies, one using an estimation of the critical substrate level as set-point and another regulating the by-product concentration, are proposed. Finally, experimental investigations of an adaptive RST control scheme using the observer polynomial for the regulation of the ethanol concentration in Saccharomyces cerevisiae fed-batch cultures ranging from laboratory to industrial scales, are also presented.

  16. Fed-Batch Biomolecule Production by Bacillus subtilis: A State of the Art Review. (United States)

    Öztürk, Sibel; Çalık, Pınar; Özdamar, Tunçer H


    Bacillus subtilis is a highly promising production system for various biomolecules. This review begins with the algorithm of fed-batch operations (FBOs) and then illustrates the approaches to design the initial production medium and/or feed stream. Additionally, the feeding strategies developed with or without feedback control for fed-batch B. subtilis fermentations were compiled with a special emphasis on recombinant protein (r-protein) production. For biomolecule production by wild-type B. subtilis, due to the different intracellular production patterns, no consensus exists on the FBO strategy that gives the maximum productivity, whereas for r-protein production appropriate feeding strategies vary depending on the promoter used. Thus, we conclude that the B. subtilis community is still seeking an approved strong promoter and generalized FBO strategies.

  17. Optimization of a feed medium for fed-batch culture of insect cells using a genetic algorithm

    NARCIS (Netherlands)

    Marteijn, R.C.L.; Jurrius, O.; Dhont, J.; Gooijer, de C.D.; Tramper, J.; Martens, D.E.


    Insect cells have been cultured for over 30 years, but their application is still hampered by low cell densities in batch fermentations and expensive culture media. With respect to the culture method, the fed-batch culture mode is often found to give the best yields. However, optimization of the

  18. Ethanol production by anaerobic thermophilic bacteria: kinetics in fed-batch cultures of Clostridium thermohydrosulfuricum

    Energy Technology Data Exchange (ETDEWEB)

    Toukourou, F.; Donaduzzi, L.; Miclo, A.; Germain, P. (Lab. of Industrial Microbiology, ENSAIA-INPL, Vandoeuvre les Nancy (FR))


    Fed-batch fermentations of Clostridium thermohydrosulfuricum are carried out using a medium rich in nitrogen source and with glucose as growth limiting factor. The ethanol/lactate yield increases as the specific growth rate and specific rate of consumption of glucose diminish. Under the experimental conditions chosen here this yield attained 3.66 moles. mole/sup -1/ with a maximal ethanol concentration of 12 g.1/sup -1/. In batch fermentation, the maximum concentration of ethanol did not exceed 8 g.1/sup -1/ independent of the concentration in glucose or nitrogen source applied. (author).

  19. Fed-batch process for the psychrotolerant marine bacterium Pseudoalteromonas haloplanktis

    Directory of Open Access Journals (Sweden)

    Lalk Michael


    Full Text Available Abstract Background Pseudoalteromonas haloplanktis is a cold-adapted γ-proteobacterium isolated from Antarctic sea ice. It is characterized by remarkably high growth rates at low temperatures. P. haloplanktis is one of the model organisms of cold-adapted bacteria and has been suggested as an alternative host for the soluble overproduction of heterologous proteins which tend to form inclusion bodies in established expression hosts. Despite the progress in establishing P. haloplanktis as an alternative expression host the cell densities obtained with this organism, which is unable to use glucose as a carbon source, are still low. Here we present the first fed-batch cultivation strategy for this auspicious alternative expression host. Results The key for the fed-batch cultivation of P. haloplanktis was the replacement of peptone by casamino acids, which have a much higher solubility and allow a better growth control. In contrast to the peptone medium, on which P. haloplanktis showed different growth phases, on a casamino acids-containing, phosphate-buffered medium P. haloplanktis grew exponentially with a constant growth rate until the stationary phase. A fed-batch process was established by feeding of casamino acids with a constant rate resulting in a cell dry weight of about 11 g l-1 (OD540 = 28 which is a twofold increase of the highest densities which have been obtained with P. haloplanktis so far and an eightfold increase of the density obtained in standard shake flask cultures. The cell density was limited in the fed-batch cultivation by the relatively low solubility of casamino acids (about 100 g l-1, which was proven by pulse addition of casamino acid powder which increased the cell density to about 20 g l-1 (OD540 = 55. Conclusion The growth of P. haloplanktis to higher cell densities on complex medium is possible. A first fed-batch fermentation strategy could be established which is feasible to be used in lab-scale or for industrial

  20. Modeling of the pyruvate production with Escherichia coli in a fed-batch bioreactor. (United States)

    Zelić, B; Vasić-Racki, D; Wandrey, C; Takors, R


    A family of 10 competing, unstructured models has been developed to model cell growth, substrate consumption, and product formation of the pyruvate producing strain Escherichia coli YYC202 ldhA::Kan strain used in fed-batch processes. The strain is completely blocked in its ability to convert pyruvate into acetyl-CoA or acetate (using glucose as the carbon source) resulting in an acetate auxotrophy during growth in glucose minimal medium. Parameter estimation was carried out using data from fed-batch fermentation performed at constant glucose feed rates of q(VG)=10 mL h(-1). Acetate was fed according to the previously developed feeding strategy. While the model identification was realized by least-square fit, the model discrimination was based on the model selection criterion (MSC). The validation of model parameters was performed applying data from two different fed-batch experiments with glucose feed rate q(VG)=20 and 30 mL h(-1), respectively. Consequently, the most suitable model was identified that reflected the pyruvate and biomass curves adequately by considering a pyruvate inhibited growth (Jerusalimsky approach) and pyruvate inhibited product formation (described by modified Luedeking-Piret/Levenspiel term).

  1. From Fed-batch to Continuous Enzymatic Biodiesel Production

    DEFF Research Database (Denmark)


    In this this paper, we use mechanistic modelling to guide the development of acontinuous enzymatic process that is performed as a fed-batch operation. In this workwe use the enzymatic biodiesel process as a case study. A mechanistic model developedin our previous work was used to determine...... measured components (triglycerides, diglycerides, monoglycerides, free fatty acid and fatty acid methyl esters(biodiesel)) much better than using fed-batch data alone given the smaller residuals. We also observe a reduction in the correlation between the parameters.The model was then used to predict that 5...... reactors are required (with a combined residence time of 30 hours) to reach a final biodiesel concentration within 2 % of the95.6 mass % achieved in a fed-batch operation, for 24 hours....

  2. An Advisory System for On-line Control of Fed-batch Cultivation of Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Ljakova K.


    Full Text Available Free software for entering and documenting data EpiData is here used for design of an advisory system for on-line control of a fermentation process. Based on the preliminary developed system for functional state recognition, presented here system will advise the user which new functional state can be reached and what kind of control actions have to be taken. New-designed system appears as an expert system and comprises knowledge of well-trained operators of cultivation processes. Developed advisory system is further applied for a fed-batch cultivation of Saccharomyces cerevisiae.

  3. Optimal parametric sensitivity control for a fed-batch reactor

    NARCIS (Netherlands)

    Stigter, J.D.; Keesman, K.J.


    The paper presents a method to derive an optimal parametric sensitivity controller for optimal estimation of a set of parameters in an experiment. The method is demonstrated for a fed batch bio-reactor case study for optimal estimation of the saturation constant Ks and, albeit intuitively, the param

  4. Optimal parametric sensitivity control of a fed-batch reactor

    NARCIS (Netherlands)

    Stigter, J.D.; Keesman, K.J.


    The paper presents an optimal parametric sensitivity controller for estimation of a set of parameters in an experiment. The method is demonstrated for a fed-batch bioreactor case study for optimal estimation of the half-saturation constant KS and the parameter combination µmaxX/Y in which µmax is th

  5. Nisin production in realkalized fed-batch cultures in whey with feeding with lactose- or glucose-containing substrates. (United States)

    Costas Malvido, Mónica; Alonso González, Elisa; Pérez Guerra, Nelson


    Nisin production by Lactococcus lactis CECT 539 was followed in batch cultures in whey supplemented with different concentrations of glucose and in two realkalized fed-batch fermentations in unsupplemented whey, which were fed, respectively, with concentrated solutions of lactose and glucose. In the batch fermentations, supplementation of whey with glucose inhibited both the growth and bacteriocin production. However, fed-batch cultures were characterized with high productions of biomass (1.34 and 1.51 g l(-1)) and nisin (50.6 and 60.3 BU ml(-1)) in comparison to the batch fermentations in unsupplemented whey (0.48 g l(-1) and 22.5 BU ml(-1)) and MRS broth (1.59 g l(-1) and 50.0 BU ml(-1)). In the two realkalized fed-batch fermentations, the increase in bacteriocin production parallels both the biomass production and pH drop generated in each realkalization and feeding cycle, suggesting that nisin was synthesized as a pH-dependent primary metabolite. A shift from homolactic to heterolactic fermentation was observed at the 108 h of incubation, and other metabolites (acetic acid and butane-2,3-diol) in addition to lactic acid accumulated in the medium. On the other hand, the feeding with glucose improved the efficiencies in glucose, nitrogen, and phosphorus consumption as compared to the batch cultures. The realkalized fed-batch fermentations showed to be an effective strategy to enhance nisin production in whey by using an appropriate feeding strategy to avoid the substrate inhibition.

  6. Maximizing yellow pigment production in fed-batch culture of Monascus sp. (United States)

    Krairak, S; Yamamura, K; Irie, R; Nakajima, M; Shimizu, H; Chim-Anage, P; Yongsmith, B; Shioya, S


    Yellow pigment production in exponential fed-batch cultivation of Monascus sp. was studied. Due to the difficulty of measuring the optical density for accurate determination of the cell concentration, a capacitance probe was employed on-line. The feed rate needed to keep the specific growth rate, mu, constant in fed-batch culture was determined on the basis of the cell concentration measured by the capacitance probe. Control of mu was improved by using updated information on the cell concentration compared with the simple feed-forward determination method using the initial cell concentration only. The highest specific pigment production rate was achieved with a mu of 0.02 h(-1) in the feeding phase. However, among several fermentation examined, the largest pigment production in the final step was obtained at a mu of 0.01 h(-1); in each case the same amount of substrates was used. An investigation of the optimal initial glucose concentration revealed that pigment production was maximum when the initial glucose concentration in the batch mode was 10 g/l and mu was 0.01 h(-1) in the fed-batch mode. It was also found that the pellet weight in the fermentation could be accurately estimated by image analysis. The ratio of the mycelium weight to the total cell weight estimated from information on the total cell weight and the estimated pellet weight was found to be more than 80%. However, no clear quantitative relationship could be discerned between the specific pigment production rate, rho, and the ratio of mycelium in the cell population.

  7. Optimal feed rate profiles for fed-batch culture in penicillin production

    Directory of Open Access Journals (Sweden)

    Murray Moo-Young


    Full Text Available The fed-batch optimization of penicillin productivity was applied as an example of optimization algorithm verification. The objective function of this problem was to optimize penicillin productivity by determination of feed rate trajectory. This study compared the optimized results derived from the proposed algorithm and from the iterative dynamic programming. Three decision variables for the proposed algorithm comprised ts (switching time from exponential to linear feeding schedules, K (constant in feed rate equation, and ε (a multiplier on substrate requirement. Estimation of this set of decision variables employed Markov chain Monte Carlo procedures (the Gibbs parameter sampling and the Metropolis-Hasting algorithm using an originally given set of initial values. The optimization procedure was divided into two time periods as follows: i the time period of exponential feeding policy, t ts. The calculation procedure of the first period of fermentation time had been proposed by integrating Pontryagin’s optimum principle and Luedeking-Piret equation. The feed rate profile during the later period was obtained from the direct substitution of desired substrate requirement derived from Monod equation. The optimal feed-rate profile corresponded to the values of decision variables as follows [ts K ε] = [35.9370.096 2.087]. The proposed algorithm was appropriate for determination of optimal feed-rate trajectories in any fed-batch problems provided that the product formation rate agrees with a Luedecking-Piret model.

  8. Fed-batch versus batch cultures of Yarrowia lipolytica for γ-decalactone production from methyl ricinoleate. (United States)

    Gomes, Nelma; Teixeira, José A; Belo, Isabel


    Constant medium feeding rate and intermittent fed-batch fermentation strategies were investigated aiming to increase the yields of γ-decalactone production by Yarrowia lipolytica, using methyl ricinoleate as substrate and ricinoleic acid source. The accumulation of another compound, 3-hydroxy-γ-decalactone, was also analyzed since it derives from the direct precursor of γ-decalactone thereby providing information about the enzymatic activities of the pathway. Both strategies were compared with the traditional batch mode in terms of overall productivity and yield in respect to the substrate. Although the productivity of γ-decalactone was considerably higher in the batch mode (168 mg l(-1) h(-1)), substrate conversion to lactone (73 mg γ-decalactone g(-1)) was greater in the intermittent fed-batch giving 6.8 g γ-decalactone l(-1). This last strategy therefore has potential for γ-decalactone production at an industrial level.

  9. Effects of the feeding ratio of food waste on fed-batch aerobic composting and its microbial community. (United States)

    Wang, Xiaojun; Pan, Songqing; Zhang, Zhaoji; Lin, Xiangyu; Zhang, Yuzhen; Chen, Shaohua


    To determine the suitable feeding ratio for fed-batch aerobic composting, four fermenters were operated by adding 0%, 5%, 10% or 15% of food waste every day. The results showed that the 5% and 10% treatments were able to maintain continuous thermophilic conditions, while the 15% treatment performed badly in regard to composting temperature, which was probably due to the negative effects of excessive moisture on microbial activity. As composting proceeded, both the 5% and the 10% treatments reached maturity and achieved weight losses of approximately 65%. High-throughput sequencing results indicated that Firmicutes, Proteobacteria, Bacteroidetes and Actinobacteria were the dominant phyla of the community structure. The communities sampled at the thermophilic phases had high similarity and relatively low diversity, while species diversity increased in the maturity phase. This study was devoted to optimizing the fed-batch composting process and assessing bacterial communities, both of which were supplied as a reference for practical application.

  10. Improved production of medium-chain-length Polyhydroxyalkanotes in glucose-based fed-batch cultivations of metabolically engineered Pseudomonas putida strains

    NARCIS (Netherlands)

    Poblete-Castro, I.; Rodriguez, A.L.; Lam, M.C.; Kessler, W.


    One of the major challenges in metabolic engineering for enhanced synthesis of value-added chemicals is to design and develop new strains which can be translated into well-controlled fermentation processes using bioreactors. The aim of this study was to assess the influence of various fed-batch stra

  11. Estimation of optimal feeding strategies for fed-batch bioprocesses. (United States)

    Franco-Lara, Ezequiel; Weuster-Botz, Dirk


    A generic methodology for feeding strategy optimization is presented. This approach uses a genetic algorithm to search for optimal feeding profiles represented by means of artificial neural networks (ANN). Exemplified on a fed-batch hybridoma cell cultivation, the approach has proven to be able to cope with complex optimization tasks handling intricate constraints and objective functions. Furthermore, the performance of the method is compared with other previously reported standard techniques like: (1) optimal control theory, (2) first order conjugate gradient, (3) dynamical programming, (4) extended evolutionary strategies. The methodology presents no restrictions concerning the number or complexity of the state variables and therefore constitutes a remarkable alternative for process development and optimization.

  12. Fed-batch production of tetanus toxin by Clostridium tetani. (United States)

    Fratelli, Fernando; Siquini, Tatiana Joly; de Abreu, Marcelo Estima; Higashi, Hisako Gondo; Converti, Attilio; de Carvalho, João Carlos Monteiro


    This study deals with the effects of the initial nitrogen source (NZ Case TT) level and the protocol of glucose addition during the fed-batch production of tetanus toxin by Clostridium tetani. An increase in the initial concentration of NZ Case TT (NZ(0)) accelerated cell growth, increased the consumption of the nitrogen source as well as the final yield of tetanus toxin, which achieved the highest values (50-60 L(f)/mL) for NZ(0) > or = 50 g/L. The addition of glucose at fixed times (16, 56, and 88 h) ensured a toxin yield ( approximately 60 L(f)/mL) about 33% higher than those of fed-batch runs with addition at fixed concentration ( approximately 45 L(f)/mL) and about 300% higher than those obtained in reference batch runs nowadays used at industrial scale. The results of this work promise to substantially improve the present production of tetanus toxin and may be adopted for human vaccine production after detoxification and purification.

  13. Batch and High Cell Density Fed-Batch Culture Productions of an Organophosphorus Hydrolase (United States)


    0.02 g H3BO3, 0.01 g NaMoO4@ 2H2O , and 0.01 g CuSO4 . Fed-Batch Fermentations were carried out in the same Bio-Flow 3000 unit fitted with 10 L...per L): 3.0 g nitrilotriacetic acid, 6.0 MgSO4@7H2O, 1.0 g NaCl, 1.0 g MnSO4@H2O, 0.5 g FeSO4@7H20, 0.1 CaCl2@ 2H2O , 0.1 CoCl2@6H2O, 0.1 g ZnSO4@7H2O

  14. Exponential fed-batch strategy for enhancing biosurfactant production by Bacillus subtilis. (United States)

    Amin, G A


    Surfactin produced by Bacillus subtilis BDCC-TUSA-3 from Maldex-15 was used as a growth-associated product in a conventional batch process. Maldex-15 is a cheap industrial by-product recovered during manufacturing of high fructose syrup from corn starch. Surfactin production was greatly improved in exponential fed-batch fermentation. Maldex-15 and other nutrients were exponentially fed into the culture based on the specific growth rate of the bacterium. In order to maximize surfactin yield and productivity, conversion of different quantities of Maldex-15 into surfactin was investigated in five different fermentation runs. In all runs, most of the Maldex-15 was consumed and converted into surfactin and cell biomass with appreciable efficiencies. The best results were obtained with the fermentation run supplied with 204 g Maldex-15. Up to 36.1 g l(-1) of surfactin and cell biomass of 31.8 g l(-1) were achieved in 12 h. Also, a marked substrate yield of 0.272 g g(-1) and volumetric reactor productivity of 2.58 g 1(-1) h(-1) were obtained, confirming the establishment of a cost-effective commercial surfactin production.

  15. Implementation of Sliding Mode Controller with Boundary Layer for Saccharomyces cerevisiae Fed-batch Cultivation

    Directory of Open Access Journals (Sweden)

    Stoyan Tzonkov


    Full Text Available An implementation of sliding mode control for yeast fed-batch cultivation is presented in this paper. Developed controller has been implemented on two real fed-batch cultivations of Saccharomyces cerevisiae. The controller successfully stabilizes the process and shows a very good performance at high input disturbances.


    Directory of Open Access Journals (Sweden)

    Leonard D. Holmes


    Full Text Available The present study deals with the batch and fed-batch mass production of Steinernema carpocapsae. S. carpocapsae is an entomoparasitic nematode that is used as a biological control agent of soil-borne crop insect pests. The ability and efficiency of fed-batch culture process was successful through the utilization of the nematode’s bacterial symbiont Xenorhabdus nematophila. Results from the fed-batch process were compared to those obtain from the standard batch process. The fed-batch process successively improved the mass production process of S. carpocapsae employing liquid medium technology. Within the first week of the fed-batch process (day six, the nematode density obtained was 202,000 nematodes mL−1; whereas on day six, batch culture mode resulted in a nematode density of 23,000 nematodes mL−1. The fed-batch process was superior to that of batch production with a yield approximately 8.8-fold higher. In fed-batch process, the nematode yield was improved 88.6 % higher within a short amount of time compared to the batch process. Fed-batch seems to make the process more efficient and possibly economically viable.

  17. A fast approach to determine a fed batch feeding profile for recombinant Pichia pastoris strains

    Directory of Open Access Journals (Sweden)

    Herwig Christoph


    Full Text Available Abstract Background The microorganism Pichia pastoris is a commonly used microbial host for the expression of recombinant proteins in biotechnology and biopharmaceutical industry. To speed up process development, a fast methodology to determine strain characteristic parameters, which are needed to subsequently set up fed batch feeding profiles, is required. Results Here, we show the general applicability of a novel approach to quantify a certain minimal set of bioprocess-relevant parameters, i.e. the adaptation time of the culture to methanol, the specific substrate uptake rate during the adaptation phase and the maximum specific substrate uptake rate, based on fast and easy-to-do batch cultivations with repeated methanol pulses in a batch culture. A detailed analysis of the adaptation of different P. pastoris strains to methanol was conducted and revealed that each strain showed very different characteristics during adaptation, illustrating the need of individual screenings for an optimal parameter definition during this phase. Based on the results obtained in batch cultivations, dynamic feeding profiles based on the specific substrate uptake rate were employed for different P. pastoris strains. In these experiments the maximum specific substrate uptake rate, which had been defined in batch experiments, also represented the upper limit of methanol uptake, underlining the validity of the determined process-relevant parameters and the overall experimental strategy. Conclusion In this study, we show that a fast approach to determine a minimal set of strain characteristic parameters based on easy-to-do batch cultivations with methanol pulses is generally applicable for different P. pastoris strains and that dynamic fed batch strategies can be designed on the specific substrate uptake rate without running the risk of methanol accumulation.

  18. A fast approach to determine a fed batch feeding profile for recombinant Pichia pastoris strains. (United States)

    Dietzsch, Christian; Spadiut, Oliver; Herwig, Christoph


    The microorganism Pichia pastoris is a commonly used microbial host for the expression of recombinant proteins in biotechnology and biopharmaceutical industry. To speed up process development, a fast methodology to determine strain characteristic parameters, which are needed to subsequently set up fed batch feeding profiles, is required. Here, we show the general applicability of a novel approach to quantify a certain minimal set of bioprocess-relevant parameters, i.e. the adaptation time of the culture to methanol, the specific substrate uptake rate during the adaptation phase and the maximum specific substrate uptake rate, based on fast and easy-to-do batch cultivations with repeated methanol pulses in a batch culture. A detailed analysis of the adaptation of different P. pastoris strains to methanol was conducted and revealed that each strain showed very different characteristics during adaptation, illustrating the need of individual screenings for an optimal parameter definition during this phase. Based on the results obtained in batch cultivations, dynamic feeding profiles based on the specific substrate uptake rate were employed for different P. pastoris strains. In these experiments the maximum specific substrate uptake rate, which had been defined in batch experiments, also represented the upper limit of methanol uptake, underlining the validity of the determined process-relevant parameters and the overall experimental strategy. In this study, we show that a fast approach to determine a minimal set of strain characteristic parameters based on easy-to-do batch cultivations with methanol pulses is generally applicable for different P. pastoris strains and that dynamic fed batch strategies can be designed on the specific substrate uptake rate without running the risk of methanol accumulation.

  19. Application of the Lyapunov Exponent to Evaluate Noise Filtering Methods for a Fed-batch Bioreactor for PHB Production

    Directory of Open Access Journals (Sweden)

    Pratap R. Patnaik


    Full Text Available Large-scale fed-batch fermentations are often subject to noise carried by the feed streams. This noise corrupts the process data and may destabilize the fermentation. So it is important to retrieve clear signals from noisy data. This is done by noise filters. The performances of some commonly used filters have been studied for poly-β-hydroxybutyrate production by Ralstonia eutropha. In simulated experiments, Gaussian noise was added to the flow rates of the carbon and nitrogen substrates. The filters were compared by means of the Lyapunov exponents of the outputs and their closeness to the noise-free performance. Negative exponents indicate a stable fermentation. An auto-associative neural filter performed the best, followed by a combination of a cusum filter and an extended Kalman filter. Butterworth filters were inferior and inadequate.

  20. Aeration strategy: a need for very high ethanol performance in Saccharomyces cerevisiae fed-batch process. (United States)

    Alfenore, S; Cameleyre, X; Benbadis, L; Bideaux, C; Uribelarrea, J-L; Goma, G; Molina-Jouve, C; Guillouet, S E


    In order to identify an optimal aeration strategy for intensifying bio-fuel ethanol production in fermentation processes where growth and production have to be managed simultaneously, we quantified the effect of aeration conditions--oxygen limited vs non limited culture (micro-aerobic vs aerobic culture)--on the dynamic behaviour of Saccharomyces cerevisiae cultivated in very high ethanol performance fed-batch cultures. Fermentation parameters and kinetics were established within a range of ethanol concentrations (up to 147 g l(-1)), which very few studies have addressed. Higher ethanol titres (147 vs 131 g l(-1) in 45 h) and average productivity (3.3 vs 2.6 g l(-1) h(-1)) were obtained in cultures without oxygen limitation. Compared to micro-aerobic culture, full aeration led to a 23% increase in the viable cell mass as a result of the concomitant increase in growth rate and yield, with lower ethanol inhibition. The second beneficial effect of aeration was better management of by-product production, with production of glycerol, the main by-product, being strongly reduced from 12 to 4 g l(-1). We demonstrate that aeration strategy is as much a determining factor as vitamin feeding (Alfenore et al. 2002) in very high ethanol performance (147 g l(-1) in 45 h) in order to achieve a highly competitive dynamic process.

  1. Production of biomethane from palm oil mill effluent (POME) with fed batch system in beam-shaped digester (United States)

    Aznury, Martha; Amin, Jaksen M.; Hasan, Abu; Himmatuliza, Astinesia


    Palm oil mill effluent (POME) is the biggest liquid waste which is produced from palm oil production. POME are containing organic matter, high levels of biological oxygen demand (BOD) and chemical oxygen demand (COD) were 28000 mg/L and 48000 mg/L. To reduce the levels of pollution caused by POME, is necessary to do stages of processing using a biological process that involves aerobic and anaerobic bacteria so that it can be utilized as a new product that has economic value, one is biogas. The processing into biogas in anaerobic performed by fed batch system. In the ratio between POME and activated microorganismes are 70:30%. The process of anaerobic fermentation in fed batch is done by time variation of the addition of the substrate. The mixture of POME and activated microorganismes were fermented for a month and then after one month substrates were added gradually as much as 1 liter into the digester with a variety of additional time are 1, 2, and 5 days. The interval of addition of the substrate give effect to the pH and the quantity of biogas produced. The highest increasing of the quantity of biomethane was 25.14 mol% at the time the addition of substrate every fifth day.

  2. Soft sensor control of metabolic fluxes in a recombinant Escherichia coli fed-batch cultivation producing green fluorescence protein. (United States)

    Gustavsson, Robert; Mandenius, Carl-Fredrik


    A soft sensor approach is described for controlling metabolic overflow from mixed-acid fermentation and glucose overflow metabolism in a fed-batch cultivation for production of recombinant green fluorescence protein (GFP) in Escherichia coli. The hardware part of the sensor consisted of a near-infrared in situ probe that monitored the E. coli biomass and an HPLC analyzer equipped with a filtration unit that measured the overflow metabolites. The computational part of the soft sensor used basic kinetic equations and summations for estimation of specific rates and total metabolite concentrations. Two control strategies for media feeding of the fed-batch cultivation were evaluated: (1) controlling the specific rates of overflow metabolism and mixed-acid fermentation metabolites at a fixed pre-set target values, and (2) controlling the concentration of the sum of these metabolites at a set level. The results indicate that the latter strategy was more efficient for maintaining a high titer and low variability of the produced recombinant GFP protein.

  3. 红曲霉色素流加培养的初步研究%Preliminary Study on Fed-batch Culture of Monascus Pigments

    Institute of Scientific and Technical Information of China (English)

    杨旭; 曹岚; 李旭


    Objective:The optimization methods of monascus pigments with submerged fermentation are studied.The level of fermentation is improved and the products contain a high concentration of monascus pigments.Methods:L8 (27) orthogonal experiment and fed-batch fermentation method of monaseus purpureus liquid are designed to select the most efficient fermentation medium for monaseus purpureus,reduce inhibition caused by over-rich nutrients and improve the fermentation concentration and level through different batch fermentation modes.Conclusion:Optimized shake flask fermentation medium with maltose of 8°Bx,soluble starch of 3%,soya bean protein powder of 4%,NaCl of 0.5%,magnesium sulfate of 0.05% and dipotassium hydrogen phosphate of 0.1%.The optimum fed-batch mode is obtained by fed-batch fermentation for 2 L batch tank,the best feeding submerged fermentation starts feeding after 60 h,filling every 20 min,completes after 12 h.Compared with batch fermentation,the valence of fed-batch fermented monascus pigment increases by 57%.%目的:研究红曲霉液体深层发酵的优化方法,提高其发酵水平,生产出含有较高红曲霉色素的产品.方法:采用L8(27)正交实验法和红曲霉液体流加发酵方法,通过不同的补料发酵方式,降低营养物质过浓而产生的阻碍作用,提高发酵浓度和水平.结果:优化后的培养基为饴糖8°Bx,可溶淀粉3%,大豆蛋白粉4%,氯化钠0.5%,硫酸镁0.05%,磷酸氢二钾0.1%.通过对2L发酵罐的补料发酵得出最佳的补料方式:液体深层发酵60 h以后开始补料,每20 min补1次,12 h补完,同分批发酵相比,流加发酵红曲色素的效价提高57%.

  4. On the optimal control of fed-batch reactors with substrate-inhibited kinetics. (United States)

    Cazzador, L


    The optimal feed rate profiles, for fed-batch fermentation that maximizes the biomass production and accounts for time, are analyzed. The solution can be found only if the final arc of the optimal control is a batch arc, since in this case the final concentrations of substrate and biomass can be determined by ulterior conditions on the mass balance and on the final growth rate of biomass and thus it is possible to solve the resulting time optimal problem by using Green's theorem. This evidences the "turnpike property" of the solution, which tries to spend the maximum time on or at least near the singular arc along which the substrate concentration is maintained constant. The optimality of the final batch arc is related to the time operational cost in the performance index. The sequence of the control depends on the initial conditions for which six different regions, with the respective patterns, have been identified, in case the performance index allows the control sequence to have a final batch.

  5. De novo biosynthesis of biodiesel by Escherichia coli in optimized fed-batch cultivation.

    Directory of Open Access Journals (Sweden)

    Yangkai Duan

    Full Text Available Biodiesel is a renewable alternative to petroleum diesel fuel that can contribute to carbon dioxide emission reduction and energy supply. Biodiesel is composed of fatty acid alkyl esters, including fatty acid methyl esters (FAMEs and fatty acid ethyl esters (FAEEs, and is currently produced through the transesterification reaction of methanol (or ethanol and triacylglycerols (TAGs. TAGs are mainly obtained from oilseed plants and microalgae. A sustainable supply of TAGs is a major bottleneck for current biodiesel production. Here we report the de novo biosynthesis of FAEEs from glucose, which can be derived from lignocellulosic biomass, in genetically engineered Escherichia coli by introduction of the ethanol-producing pathway from Zymomonas mobilis, genetic manipulation to increase the pool of fatty acyl-CoA, and heterologous expression of acyl-coenzyme A: diacylglycerol acyltransferase from Acinetobacter baylyi. An optimized fed-batch microbial fermentation of the modified E. coli strain yielded a titer of 922 mg L(-1 FAEEs that consisted primarily of ethyl palmitate, -oleate, -myristate and -palmitoleate.

  6. Fed-batch CHO cell culture for lab-scale antibody production

    DEFF Research Database (Denmark)

    Fan, Yuzhou; Ley, Daniel; Andersen, Mikael Rørdam


    Fed-batch culture is the most commonly used upstream process in industry today for recombinant monoclonal antibody production using Chinese hamster ovary cells. Developing and optimizing this process in the lab is crucial for establishing process knowledge, which enable rapid and predictable tech......-transfer to manufacturing scale. In this chapter, we will describe stepwise how to carry out fed-batch CHO cell culture for lab-scale antibody production....

  7. Fed-batch production of the hydrophobins RodA and RodB from Aspergillus fumigatus in host Pichia pastoris

    DEFF Research Database (Denmark)

    Pedersen, Mona Højgaard; Borodina, Irina; Frisvad, Jens Christian;

    . The expression of the RodA and RodB genes was first studied in culture flasks in buffered complex methanol medium as protein production was dependent on the methanol-induced AOX1 promoter. Later production was scaled up to a 2 L fed-batch fermentor. Hydrophobins were purified using His-select Nickel Affinity gel....... The emulsifying properties of recombinant hydrophobins were investigated using oil-water emulsions studied by light microscopy. Results: Protein bands of expected size were detected by SDS-PAGE and western blotting in the fermentation broth. Fed-batch production yielded approximately 300 mg/L. rRodB showed good...... emulsifying properties. Conclusion: RodA and RodB from A. fumigatus were successfully produced by yeast host Pichia pastoris with good yields....

  8. Production of Medium Chain Length Polyhydroxyalkanoates From Oleic Acid Using Pseudomonas putida PGA1 by Fed Batch Culture

    Directory of Open Access Journals (Sweden)

    Sidik Marsudi


    Full Text Available Bacterial polyhydroxyalkanoates (PHAs are a class of p0lymers currently receiving much attention because of their potential as renewable and biodegradable plastics. A wide variety of bacteria has been reported to produce PHAs including Pseudomonas strains. These strains are known as versatile medium chain length PHAs (PHAs-mcl producers using fatty acids as carbon source. Oleic acid was used to produce PHAs-mcl using Pseudomonas putida PGA 1 by continuous feeding of both nitrogen and carbon source, in a fed batch culture. During cell growth, PHAs also accumulated, indicating that PHA production in this organism is growth associated. Residual cell increased until the nitrogen source was depleted. At the end of fermentation, final cell concentration, PHA content, and roductivity were 30.2 g/L, 44.8 % of cell dry weight, and 0.188 g/l/h, respectively.

  9. Growth kinetics of Saccharomyces cerevisiae in batch and fed-batch cultivation using sugarcane molasses and glucose syrup from cassava starch. (United States)

    Win, S S; Impoolsup, A; Noomhorm, A


    Growth kinetics of Saccharomyces cerevisiae in glucose syrup from cassava starch and sugarcane molasses were studied using batch and fed-batch cultivation. The optimum temperature and pH required for growth were 30 degrees C and pH 5.5, respectively. In batch culture the productivity and overall cell yield were 0.31 g L-1 h-1 and 0.23 g cells g-1 sugar, respectively, on glucose syrup and 0.22 g L-1 h-1 and 0.18 g cells g-1 sugar, respectively, on molasses. In fed-batch cultivation, a productivity of 3.12 g L-1 h-1 and an overall cell yield of 0.52 g cells g-1 sugar were achieved in glucose syrup cultivation and a productivity of 2.33 g L-1 h-1 and an overall cell yield of 0.46 g cells g-1 sugar were achieved in molasses cultivation by controlling the reducing sugar concentration at its optimum level obtained from the fermentation model. By using an on-line ethanol sensor combined with a porous Teflon tubing method in automating the feeding of substrate in the fed-batch culture, a productivity of 2.15 g L-1 h-1 with a yield of 0.47 g cells g-1 sugar was achieved using glucose syrup as substrate when ethanol concentration was kept at a constant level by automatic control.

  10. Simulation of kefiran production of Lactobacillus kefiranofaciens JCM6985 in fed-batch reactor

    Directory of Open Access Journals (Sweden)

    Benjamas Cheirsilp


    Full Text Available Kinetics of kefiran production by Lactobacillus kefiranofaciens JCM6985 has been investigated. A mathematical model taking into account the mechanism of exopolysaccharides production has been developed. Experiments were carried out in batch mode in order to obtain kinetic model parameters that were further applied to simulate fed-batch processes. A simplification of parameter fitting was also introduced for complicated model. The fed-batch mode allows more flexibility in the control of the substrate concentration as well as product concentration in the culture medium. Based on the batch mathematical model, a fed-batch model was developed and simulations were done. Simulation study in fed-batch reactor resulted that substrate concentration should be controlled at 20 g L-1 to soften the product inhibition and also to stimulate utilization of substrate and its hydrolysate. From simulation results of different feeding techniques, it was found that constant feeding at 0.01 L h-1 was most practically effective feeding profile for exopolysaccharides production in fed-batch mode.

  11. Fed-batch production of gluconic acid by terpene-treated Aspergillus niger spores. (United States)

    Ramachandran, Sumitra; Fontanille, Pierre; Pandey, Ashok; Larroche, Christian


    Aspergillus niger spores were used as catalyst in the bioconversion of glucose to gluconic acid. Spores produced by solid-state fermentation were treated with 15 different terpenes including monoterpenes and monoterpenoids to permeabilize and inhibit spore germination. It was found that spore membrane permeability is significantly increased by treatment with terpenoids when compared to monoterpenes. Best results were obtained with citral and isonovalal. Studies were carried out to optimize spores concentration (10(7)-10(10) spores/mL), terpene concentrations in the bioconversion medium and time of exposure (1-18 h) needed for permeabilization of spores. Fed-batch production of gluconate was done in a bioreactor with the best conditions [10(9) spores/mL of freeze-thawed spores treated with citral (3% v/v) for 5 h] followed by sequential additions of glucose powder and pH-regulated with a solution containing 2 mol/L of either NaOH or KOH. Bioconversion performance of the spore enzyme was compared with the commercial glucose oxidase at 50, 60, and 70 degrees C. Results showed that the spore enzyme was comparatively stable at 60 degrees C. It was also found that the spores could be reutilized for more than 14 cycles with almost similar reaction rate. Similar biocatalytic activity was rendered by spores even after its storage of 1 year at -20 degrees C. This study provided an experimental evidence of the significant catalytic role played by A. niger spore in bioconversion of glucose to gluconic acid with high yield and stability, giving protection to glucose oxidase.

  12. Scale-up bioprocess development for production of the antibiotic valinomycin in Escherichia coli based on consistent fed-batch cultivations. (United States)

    Li, Jian; Jaitzig, Jennifer; Lu, Ping; Süssmuth, Roderich D; Neubauer, Peter


    Heterologous production of natural products in Escherichia coli has emerged as an attractive strategy to obtain molecules of interest. Although technically feasible most of them are still constrained to laboratory scale production. Therefore, it is necessary to develop reasonable scale-up strategies for bioprocesses aiming at the overproduction of targeted natural products under industrial scale conditions. To this end, we used the production of the antibiotic valinomycin in E. coli as a model system for scalable bioprocess development based on consistent fed-batch cultivations. In this work, the glucose limited fed-batch strategy based on pure mineral salt medium was used throughout all scales for valinomycin production. The optimal glucose feed rate was initially detected by the use of a biocatalytically controlled glucose release (EnBase® technology) in parallel cultivations in 24-well plates with continuous monitoring of pH and dissolved oxygen. These results were confirmed in shake flasks, where the accumulation of valinomycin was highest when the specific growth rate decreased below 0.1 h(-1). This correlation was also observed for high cell density fed-batch cultivations in a lab-scale bioreactor. The bioreactor fermentation produced valinomycin with titers of more than 2 mg L(-1) based on the feeding of a concentrated glucose solution. Valinomycin production was not affected by oscillating conditions (i.e. glucose and oxygen) in a scale-down two-compartment reactor, which could mimic similar situations in industrial bioreactors, suggesting that the process is very robust and a scaling of the process to a larger industrial scale appears a realistic scenario. Valinomycin production was scaled up from mL volumes to 10 L with consistent use of the fed-batch technology. This work presents a robust and reliable approach for scalable bioprocess development and represents an example for the consistent development of a process for a heterologously expressed natural

  13. A high-yielding, generic fed-batch process for recombinant antibody production of GS-engineered cell lines

    DEFF Research Database (Denmark)

    Fan, Li; Zhao, Liang; Sun, Yating;


    An animal component-free and chemically defined fed-batch process for GS-engineered cell lines producing recombinant antibodies has been developed. The fed-batch process relied on supplying sufficient nutrients to match their consumption, simultaneously minimizing the accumulation of byproducts....... This generic and high-yielding fed-batch process would shorten development time, and ensure process stability, thereby facilitating the manufacture of therapeutic antibodies by GS-engineered cell lines....

  14. Fed-batch and perfusion culture processes: economic, environmental, and operational feasibility under uncertainty. (United States)

    Pollock, James; Ho, Sa V; Farid, Suzanne S


    This article evaluates the current and future potential of batch and continuous cell culture technologies via a case study based on the commercial manufacture of monoclonal antibodies. The case study compares fed-batch culture to two perfusion technologies: spin-filter perfusion and an emerging perfusion technology utilizing alternating tangential flow (ATF) perfusion. The operational, economic, and environmental feasibility of whole bioprocesses based on these systems was evaluated using a prototype dynamic decision-support tool built at UCL encompassing process economics, discrete-event simulation and uncertainty analysis, and combined with a multi-attribute decision-making technique so as to enable a holistic assessment. The strategies were compared across a range of scales and titres so as to visualize how their ranking changes in different industry scenarios. The deterministic analysis indicated that the ATF perfusion strategy has the potential to offer cost of goods savings of 20% when compared to conventional fed-batch manufacturing processes when a fivefold increase in maximum viable cell densities was assumed. Savings were also seen when the ATF cell density dropped to a threefold increase over the fed-batch strategy for most combinations of titres and production scales. In contrast, the fed-batch strategy performed better in terms of environmental sustainability with a lower water and consumable usage profile. The impact of uncertainty and failure rates on the feasibility of the strategies was explored using Monte Carlo simulation. The risk analysis results demonstrated the enhanced robustness of the fed-batch process but also highlighted that the ATF process was still the most cost-effective option even under uncertainty. The multi-attribute decision-making analysis provided insight into the limited use of spin-filter perfusion strategies in industry. The resulting sensitivity spider plots enabled identification of the critical ratio of weightings of

  15. Change in hyphal morphology of Aspergillus Oryzae during fed-batch cultivation

    DEFF Research Database (Denmark)

    Haack, Martin Brian; Olsson, Lisbeth; Hansen, K


    Industrial enzymes are often produced by filamentous fungi in fed-batch cultivations. During cultivation, the different morphological forms displayed by the fungi have an impact on the overall production. The morphology of a recombinant lipase producing Aspergillus oryzae strain was investigated...

  16. Automatic tuning and adaptation for specific growth rate control of fed-batch cultivation

    NARCIS (Netherlands)

    Soons, Z.I.T.A.; Straten, van G.; Boxtel, van A.J.B.


    To ensure consistency between fed-batch cultivations for the production of vaccines or other bio-pharmaceuticals it is desirable to control the specific growth rate to a pre-set constant value. This is a challenge because the dynamics of the process is considerably changing due to the increase in

  17. Combined age and segregated kinetic model for industrial-scale penicillin fed-batch cultivation

    NARCIS (Netherlands)

    Wang, Z.F.; Lauwerijssen, M.J.C.; Yuan, J.Q.


    This paper proposes a cell age model forPenicillium chrysogenum fed-batch cultivation to supply a qualitative insight into morphology-associated dynamics. The average ages of the segregated cell populations, such as growing cells, non-growing cells and intact productive cells, were estimated by this

  18. Selection of chemically defined media for CHO cell fed-batch culture processes

    NARCIS (Netherlands)

    Pan, X.; Streefland, M.; Dalm, C.; Wijffels, R.H.; Martens, D.E.


    Two CHO cell clones derived from the same parental CHOBC cell line and producing the same monoclonal antibody (BC-G, a low producing clone; BC-P, a high producing clone) were tested in four basal media in all possible combinations with three feeds (=12 conditions) in fed-batch cultures.
    Higher a

  19. Sensor combination and chemometric variable selection for online monitoring of Streptomyces coelicolor fed-batch cultivations

    DEFF Research Database (Denmark)

    Ödman, Peter; Johansen, C.L.; Olsson, L.


    Fed-batch cultivations of Streptomyces coelicolor, producing the antibiotic actinorhodin, were monitored online by multiwavelength fluorescence spectroscopy and off-gas analysis. Partial least squares (PLS), locally weighted regression, and multilinear PLS (N-PLS) models were built for prediction...

  20. Production of farnesene and santalene by Saccharomyces cerevisiae using fed-batch cultivations with RQ-controlled feed. (United States)

    Tippmann, Stefan; Scalcinati, Gionata; Siewers, Verena; Nielsen, Jens


    Terpenes have various applications as fragrances, cosmetics and fuels. One of the most prominent examples is the sesquiterpene farnesene, which can be used as diesel substitute in its hydrogenated form farnesane. Recent metabolic engineering efforts have enabled efficient production of several terpenes in Saccharomyces cerevisiae and Escherichia coli. Plant terpene synthases take on an essential function for sesquiterpene production as they catalyze the specific conversion of the universal precursor farnesyl diphosphate (FPP) to the sesquiterpene of interest and thereby impose limitations on the overall productivity. Using farnesene as a case study, we chose three terpene synthases with distinct plant origins and compared their applicability for farnesene production in the yeast S. cerevisiae. Differences regarding the efficiency of these enzymes were observed in shake flask cultivation with maximal final titers of 4 mg/L using α-farnesene synthase from Malus domestica. By employing two existing platform strains optimized for sesquiterpene production, final titers could be raised up 170 mg/L in fed-batch fermentations with RQ-controlled exponential feeding. Based on these experiments, the difference between the selected synthases was not significant. Lastly, the same fermentation setup was used to compare these results to production of the fragrance sesquiterpene santalene, and almost equivalent titers were obtained with 163 mg/L, using the highest producing strain expressing a santalene synthase from Clausena lansium. However, a reduction of the product yield on biomass by 50% could indicate a higher catalytic efficiency of the farnesene synthase. © 2015 Wiley Periodicals, Inc.

  1. Fuzzy logic feedback control for fed-batch enzymatic hydrolysis of lignocellulosic biomass. (United States)

    Tai, Chao; Voltan, Diego S; Keshwani, Deepak R; Meyer, George E; Kuhar, Pankaj S


    A fuzzy logic feedback control system was developed for process monitoring and feeding control in fed-batch enzymatic hydrolysis of a lignocellulosic biomass, dilute acid-pretreated corn stover. Digested glucose from hydrolysis reaction was assigned as input while doser feeding time and speed of pretreated biomass were responses from fuzzy logic control system. Membership functions for these three variables and rule-base were created based on batch hydrolysis data. The system response was first tested in LabVIEW environment then the performance was evaluated through real-time hydrolysis reaction. The feeding operations were determined timely by fuzzy logic control system and efficient responses were shown to plateau phases during hydrolysis. Feeding of proper amount of cellulose and maintaining solids content was well balanced. Fuzzy logic proved to be a robust and effective online feeding control tool for fed-batch enzymatic hydrolysis.

  2. Fed-batch bioreactor process with recombinant Saccharomyces cerevisiae growing on cheese whey

    Directory of Open Access Journals (Sweden)

    R. Rech


    Full Text Available Saccharomyces cerevisiae strain W303 was transformed with two yeast integrative plasmids containing Kluyveromyces lactis LAC4 and LAC12 genes that codify beta-galactosidase and lactose permease respectively. The BLR030 recombinant strain was selected due to its growth and beta-galactosidase production capacity. Different culture media based on deproteinized cheese whey (DCW were tested and the best composition (containing DCW, supplemented with yeast extract 1 %, and peptone 3 % (w/v was chosen for bioreactor experiments. Batch, and fed-batch cultures with linear ascending feeding for 25 (FB25, 35 (FB35, and 50 (FB50 hours, were performed. FB35 and FB50 produced the highest beta-galactosidase specific activities (around 1,800 U/g cells, and also the best productivities (180 U/L.h. Results show the potential use of fed-batch cultures of recombinant S. cerevisiae on industrial applications using supplemented whey as substrate.


    Directory of Open Access Journals (Sweden)

    C.C.F. da Cunha


    Full Text Available In this work, the growth of Bacillus thuringiensis var. israelensis, a bioinsecticide producer, is investigated. Experiments were carried out in batch mode in order to obtain kinetic model parameters that were further applied to simulate fed-batch processes. The fed-batch mode allows more flexibility in the control of the substrate concentration in the culture medium. Different techniques, such as constant feeding, "bang-bang" control and model based control (exponential feeding and singular control, were compared. For the techniques based on a model, combinations of models with and without a substrate inhibition parameter were used to represent the simulated process and the internal model of the feeding controller. Singular control based on the model with an inhibition parameter proved to be the most robust controller.

  4. Feed development for fed-batch CHO production process by semisteady state analysis. (United States)

    Khattak, Sarwat F; Xing, Zizhuo; Kenty, Brian; Koyrakh, Inna; Li, Zheng Jian


    Semisteady state cultures are useful for studying cell physiology and facilitating media development. Two semisteady states with a viable cell density of 5.5 million cells/mL were obtained in CHO cell cultures and compared with a fed-batch mode control. In the first semisteady state, the culture was maintained at 5 mM glucose and 0.5 mM glutamine. The second condition had threefold higher concentrations of both nutrients, which led to a 10% increase in lactate production, a 78% increase in ammonia production, and a 30% reduction in cell growth rate. The differences between the two semisteady states indicate that maintaining relatively low levels of glucose and glutamine can reduce the production of lactate and ammonia. Specific amino acid production and consumption indicated further metabolic differences between the two semisteady states and fed-batch mode. The results from this experiment shed light in the feeding strategy for a fed-batch process and feed medium enhancement. The fed-batch process utilizes a feeding strategy whereby the feed added was based on glucose levels in the bioreactor. To evaluate if a fixed feed strategy would improve robustness and process consistency, two alternative feeding strategies were implemented. A constant volume feed of 30% or 40% of the initial culture volume fed over the course of cell culture was evaluated. The results indicate that a constant volumetric-based feed can be more beneficial than a glucose-based feeding strategy. This study demonstrated the applicability of analyzing CHO cultures in semisteady state for feed enhancement and continuous process improvement. Copyright 2009 American Institute of Chemical Engineers

  5. Effect of feeding methods on the astaxanthin production by Phaffia rhodozyma in fed-batch process

    Directory of Open Access Journals (Sweden)

    Danilo Gomes Moriel


    Full Text Available The effect of feeding methods on the production of astaxanthin by the yeast Phaffia rhodozyma ATCC 24202 was studied, using continuous and pulsed fed-batch processes and low cost materials as substrates (sugar cane juice and urea. In continuous fed-batch processes, a cellular astaxanthin concentration of 383.73 µg/g biomass was obtained. But in pulsed fed-batch processes a reduction in the cellular astaxanthin concentration (303.34 µg/g biomass was observed. Thus the continuous fed-batch processes could be an alternative to industrial production of astaxanthin, allowing an increase in the biomass productivity without losses on astaxanthin production by the yeast.O efeito da alimentação na produção de astaxantina pela levedura Phaffia rhodozyma ATCC 24202 foi estudado, utilizando processos descontínuo alimentado com alimentação contínua e intermitente, e matérias-primas de baixo custo como substratos (caldo de cana de açúcar e uréia. Em processos descontínuo alimentado com alimentação contínua, uma concentração celular de astaxantina de 383,73 µg/g biomassa foi obtida. Entretanto, em processos descontínuo alimentado com alimentação intermitente, uma redução na concentração celular de astaxantina (303,34 µg/g biomassa foi observada. Desta forma, processos descontínuo alimentado com alimentação contínua poderiam ser uma alternativa na produção industrial de astaxantina, permitindo um aumento na produtividade de biomassa sem perdas na produção de astaxantina pela levedura.

  6. Preferences based Control Design of Complex Fed-batch Cultivation Process

    Directory of Open Access Journals (Sweden)

    Yuri Pavlov


    Full Text Available In the paper is presented preferences based control design and stabilization of the growth rate of fed-batch cultivation processes. The control is based on an enlarged Wang-Monod-Yerusalimsky kinetic model. Expected utility theory is one of the approaches for utilization of conceptual information (expert preferences. In the article is discussed utilization of stochastic machine learning procedures for evaluation of expert utilities as criteria for optimization.

  7. Perfusion seed cultures improve biopharmaceutical fed-batch production capacity and product quality. (United States)

    Yang, William C; Lu, Jiuyi; Kwiatkowski, Chris; Yuan, Hang; Kshirsagar, Rashmi; Ryll, Thomas; Huang, Yao-Ming


    Volumetric productivity and product quality are two key performance indicators for any biopharmaceutical cell culture process. In this work, we showed proof-of-concept for improving both through the use of alternating tangential flow perfusion seed cultures coupled with high-seed fed-batch production cultures. First, we optimized the perfusion N-1 stage, the seed train bioreactor stage immediately prior to the production bioreactor stage, to minimize the consumption of perfusion media for one CHO cell line and then successfully applied the optimized perfusion process to a different CHO cell line. Exponential growth was observed throughout the N-1 duration, reaching >40 × 10(6) vc/mL at the end of the perfusion N-1 stage. The cultures were subsequently split into high-seed (10 × 10(6) vc/mL) fed-batch production cultures. This strategy significantly shortened the culture duration. The high-seed fed-batch production processes for cell lines A and B reached 5 g/L titer in 12 days, while their respective low-seed processes reached the same titer in 17 days. The shortened production culture duration potentially generates a 30% increase in manufacturing capacity while yielding comparable product quality. When perfusion N-1 and high-seed fed-batch production were applied to cell line C, higher levels of the active protein were obtained, compared to the low-seed process. This, combined with correspondingly lower levels of the inactive species, can enhance the overall process yield for the active species. Using three different CHO cell lines, we showed that perfusion seed cultures can optimize capacity utilization and improve process efficiency by increasing volumetric productivity while maintaining or improving product quality. © 2014 American Institute of Chemical Engineers.

  8. Developing high cell density fed-batch cultivation strategies for heterologous protein production in Pichia pastoris using the nitrogen source-regulated FLD1 Promoter. (United States)

    Resina, David; Cos, Oriol; Ferrer, Pau; Valero, Francisco


    A Pichia pastoris strain expressing a Rhizopus oryzae lipase gene under the transcriptional control of the promoter from the P. pastoris formaldehyde dehydrogenase 1 gene (PFLD) was utilized to study the feasibility of this expression system for recombinant protein production using methanol-free fed-batch high cell density cultivations. We have developed a simple and reliable fed-batch strategy using the PFLD system based on the use of methylamine and sorbitol as nitrogen and carbon sources, respectively, for the induction phase. Three different fed-batch fermentations were performed at three different constant growth rates, i.e., at a low growth rate (0.005/h), at an intermediate growth rate of (0.01/h), and at a constant residual sorbitol concentration of 8 g/L, i.e., allowing cells to grow at high (near micro(max)) growth rate (0.02/h). Important differences were observed between the lower and higher growth rate cultivation phases in terms of specific production rate (q(p)) profiles. In all three cases, maximum q(p) were reached soon after the start of the induction phase; after that maximum, an exponential decrease reaching final values close to zero were observed, except for the cells growing at near micro(max). The best results in terms of Y(P/X), productivity and specific productivity were obtained when the microorganism was growing at the highest growth rate. Furthermore, such results were significantly better in relation to those obtained with the PAOX-based system expressing the same protein.

  9. Optimization of glutathione production in batch and fed-batch cultures by the wild-type and recombinant strains of the methylotrophic yeast Hansenula polymorpha DL-1

    Directory of Open Access Journals (Sweden)

    Malyshev Alexander Y


    Full Text Available Abstract Background Tripeptide glutathione (gamma-glutamyl-L-cysteinyl-glycine is the most abundant non-protein thiol that protects cells from metabolic and oxidative stresses and is widely used as medicine, food additives and in cosmetic industry. The methylotrophic yeast Hansenula polymorpha is regarded as a rich source of glutathione due to the role of this thiol in detoxifications of key intermediates of methanol metabolism. Cellular and extracellular glutathione production of H. polymorpha DL-1 in the wild type and recombinant strains which overexpress genes of glutathione biosynthesis (GSH2 and its precursor cysteine (MET4 was studied. Results Glutathione producing capacity of H. polymorpha DL-1 depending on parameters of cultivation (dissolved oxygen tension, pH, stirrer speed, carbon substrate (glucose, methanol and type of overexpressed genes of glutathione and its precursor biosynthesis during batch and fed-batch fermentations were studied. Under optimized conditions of glucose fed-batch cultivation, the glutathione productivity of the engineered strains was increased from ~900 up to ~ 2300 mg of Total Intracellular Glutathione (TIG or GSH+GSSGin, per liter of culture medium. Meantime, methanol fed-batch cultivation of one of the recombinant strains allowed achieving the extracellular glutathione productivity up to 250 mg of Total Extracellular Glutathione (TEG or GSH+GSSGex, per liter of the culture medium. Conclusions H. polymorpha is an competitive glutathione producer as compared to other known yeast and bacteria strains (Saccharomyces cerevisiae, Candida utilis, Escherichia coli, Lactococcus lactis etc. with good perspectives for further improvement especially for production of extracellular form of glutathione.

  10. Self-tuning GMV control of glucose concentration in fed-batch baker's yeast production. (United States)

    Hitit, Zeynep Yilmazer; Boyacioglu, Havva; Ozyurt, Baran; Ertunc, Suna; Hapoglu, Hale; Akay, Bulent


    A detailed system identification procedure and self-tuning generalized minimum variance (STGMV) control of glucose concentration during the aerobic fed-batch yeast growth were realized. In order to determine the best values of the forgetting factor (λ), initial value of the covariance matrix (α), and order of the Auto-Regressive Moving Average with eXogenous (ARMAX) model (n a, n b), transient response data obtained from the real process wereutilized. Glucose flow rate was adjusted according to the STGMV control algorithm coded in Visual Basic in an online computer connected to the system. Conventional PID algorithm was also implemented for the control of the glucose concentration in aerobic fed-batch yeast cultivation. Controller performances were examined by evaluating the integrals of squared errors (ISEs) at constant and random set point profiles. Also, batch cultivation was performed, and microorganism concentration at the end of the batch run was compared with the fed-batch cultivation case. From the system identification step, the best parameter estimation was accomplished with the values λ = 0.9, α = 1,000 and n a = 3, n b = 2. Theoretical control studies show that the STGMV control system was successful at both constant and random glucose concentration set profiles. In addition, random effects given to the set point, STGMV control algorithm were performed successfully in experimental study.

  11. Production of clavulanic acid and cephamycin C by Streptomyces clavuligerus under different fed-batch conditions

    Directory of Open Access Journals (Sweden)

    C. Bellão


    Full Text Available The effect of carbon source and feeding conditions on the production of clavulanic acid (CA and cephamycin C (CephC by Streptomyces clavuligerus was investigated. In fed-batch experiments performed with glycerol feeding, production of CA exceeded that of CephC, and reached 1022 mg.L-1. Highest CephC production (566.5 mg.L-1 was obtained in fed-batch cultivation with glycerol feeding. In fed-batch experiments performed with starch feeding, the production of CephC was in general higher than that of CA. A dissociation index (DI was used to identify feeding conditions that favored production of CephC relative to CA. In all cultures with glycerol, DI values were less than unity, indicating higher production of CA compared to CephC. Conversely, in cultures fed with starch, the DI values obtained were greater than unity. However, no carbon source or feeding condition was able to completely dissociate the production of CA from that of CephC.

  12. Effect of auxotrophies on yeast performance in aerated fed-batch reactor

    Energy Technology Data Exchange (ETDEWEB)

    Landi, Carmine; Paciello, Lucia [Dept. Ingegneria Industriale, Universita di Salerno, Via Ponte Don Melillo, 84084 Fisciano, Salerno (Italy); Alteriis, Elisabetta de [Dept. Biologia Strutturale e Funzionale, Universita degli Studi di Napoli ' Federico II' , Via Cinthia, 80100 Napoli (Italy); Brambilla, Luca [Dept. Biotecnologie e Bioscienze, Universita Milano-Bicocca, Piazza della Scienza, 20126 Milano (Italy); Parascandola, Palma, E-mail: [Dept. Ingegneria Industriale, Universita di Salerno, Via Ponte Don Melillo, 84084 Fisciano, Salerno (Italy)


    Highlights: Black-Right-Pointing-Pointer The paper contributes to fill the gap existing between the basic and applied research. Black-Right-Pointing-Pointer Mathematical model sheds light on the physiology of auxotrophic yeast strains. Black-Right-Pointing-Pointer Yeast behavior in fed-batch is influenced by biological and environmental determinants. Black-Right-Pointing-Pointer Process optimization would make possible the production of heterologous proteins which are not yet on the market. -- Abstract: A systematic investigation on the effects of auxotrophies on the performance of yeast in aerated fed-batch reactor was carried out. Six isogenic strains from the CEN.PK family of Saccharomyces cerevisiae, one prototroph and five auxotrophs, were grown in aerated fed-batch reactor using the same operative conditions and a proper nutritional supplementation. The performance of the strains, in terms of final biomass decreased with increasing the number of auxotrophies. Auxotrophy for leucine exerted a profound negative effect on the performance of the strains. Accumulation of reactive oxygen species (ROS) in the cells of the strain carrying four auxotrophies and its significant viability loss, were indicative of an oxidative stress response induced by exposure of cells to the environmental conditions. The mathematical model was fundamental to highlight how the carbon flux, depending on the number and type of auxotrophies, was diverted towards the production of increasingly large quantities of energy for maintenance.

  13. Biological treatment of high strength waste water by fed-batch operation

    Energy Technology Data Exchange (ETDEWEB)

    Kargi, F. [Dept. of Environmental Engineering, Dokuz Eyluel Univ. Bornova, Izmir (Turkey)


    Biological treatment systems for high strength wastewaters are usually operated in continuous mode such as activated sludge systems. When operated at steady-state, continuous systems result in constant effluent standards. However, in the presence of shock loadings and/or toxic compounds in feed wastewater, system performance drops quite significantly as a result of partial loss of microbial activity. In fed-batch operation, wastewater is fed to the aeration tank with a flow rate determined by effluent standards. In this type of operation, wastewater can be fed to biological oxidation unit intermittently or continuously with a low flow rate without any effluent removal. Feed flow rate is adjusted by measuring COD concentration in the effluent. As a result of intermittent addition of wastewater high COD concentrations and toxic compounds are diluted in large volume of aeration tank and inhibition effects of those compounds are reduced. As a result, biological oxidation of these compounds take place at a much higher rate. In order to show the aforementioned advantage of fed-batch operation, a high strength synthetic wastewater consisting of diluted molasses, urea, KH{sub 2}PO{sub 4} and MgSO{sub 4} was treated in an biological aeration tank by fed-batch operation. Organisms used were an active and dominant culture of Zooglea ramigera commonly encountered in activated sludge operations. COD removal kinetics was found to be first order and the rate constant was determined. (orig.). With 6 figs., 1 tab.

  14. Nonlinear GPC with In-place Trained RLS-SVM Model for DOC Control in a Fed-batch Bloreactor

    Institute of Scientific and Technical Information of China (English)

    冯絮影; 于涛; 王建林


    In this study, Saccharomyces cerevisiae (baker's yeast) was produced in a fed-batch bioreactor at the optimal dissolved oxygen concentration (DOC) and growth medium temperature. However, it is very difficult to control the DOC using conventional controllers because of the poorly understood and constantly changing dynamics of the bioprocess. A generalized predictive controller (GPC) based on a nonlinear autoregressive integrated moving average exogenous (NARIMAX) model is presented to stabilize the DOC by manipulation of air flow rate. The NARIMAX model is built by an improved recursive least-squares support vector machine, which is trained by an in-place computation scheme and avoids the computation of the inverse of a large matrix and memory reallocation. The proposed nonlinear GPC algorithm requires little preliminary knowledge of the fermentation process, and directly obtains the nonlinear model in matrix form by using iterative multiple modeling instead of linearization at each sampling period. By application of an on-line bioreactor control, experimental results demonstrate the robustness, effectiveness and advantages of the new controller.

  15. Modeling of Xanthophyllomyces dendrorhous growth on glucose and overflow metabolism in batch and fed-batch cultures for astaxanthin production. (United States)

    Liu, Yuan-Shuai; Wu, Jian-Yong


    An astaxanthin-producing yeast Xanthophyllomyces dendrorhous ENM5 was cultivated in a liquid medium containing 50 g/L glucose as the major carbon source in stirred fermentors (1.5-L working volume) in fully aerobic conditions. Ethanol was produced during the exponential growth phase as a result of overflow metabolism or fermentative catabolism of glucose by yeast cells. After accumulating to a peak of 3.5 g/L, the ethanol was consumed by yeast cells as a carbon source when glucose in the culture was nearly exhausted. High initial glucose concentrations and ethanol accumulation in the culture had inhibitory effects on cell growth. Astaxanthin production was partially associated with cell growth. Based on these culture characteristics, we constructed a modified Monod kinetic model incorporating substrate (glucose) and product (ethanol) inhibition to describe the relationship of cell growth rate with glucose and ethanol concentrations. This kinetic model, coupled with the Luedeking-Piret equation for the astaxanthin production, gave satisfactory prediction of the biomass production, glucose consumption, ethanol formation and consumption, and astaxanthin production in batch cultures over 25-75 g/L glucose concentration ranges. The model was also applied to fed-batch cultures to predict the optimum feeding scheme (feeding glucose and corn steep liquor) for astaxanthin production, leading to a high volumetric yield (28.6 mg/L) and a high productivity (5.36 mg/L/day).

  16. Genetic Algorithmic Optimization of PHB Production by a Mixed Culture in an Optimally Dispersed Fed-batch Bioreactor

    Directory of Open Access Journals (Sweden)

    Pratap R. Patnaik


    Full Text Available Poly-β-hydroxybutyrate (PHB is an energy-storage polymer whose properties are similar to those of chemical polymers such as polyethylene and polypropylene. Moreover, PHB is biodegradable, absorbed by human tissues and less energy-consuming than synthetic polymers. Although Ralstonia eutropha is widely used to synthesize PHB, it is inefficient in utilizing glucose and similar sugars. Therefore a co-culture of R. eutropha and Lactobacillus delbrueckii is preferred since the latter can convert glucose to lactate, which R. eutropha can metabolize easily. Tohyama et al. [24] maximized PHB production in a well-mixed fed-batch bioreactor with glucose and (NH42SO4 as the primary substrates. Since production-scale bioreactors often deviate from ideal laboratory-scale reactors, a large bioreactor was simulated by means of a dispersion model with the kinetics determined by Tohyama et al. [24] and dispersion set at an optimum Peclet number of 20 [32]. The time-dependent feed rates of the two substrates were determined through a genetic algorithm (GA to maximize PHB production. This bioreactor produced 22.2% more PHB per liter and 12.8% more cell mass than achieved by Tohyama et al. [24]. These results, and similar observations with other fermentations, indicate the feasibility of enhancing the efficiency of large nonideal bioreactors through GA optimizations.

  17. Effect of fed-batch on calcium biotransformation of mussel shell by Acetobacter sp.%分批补料对醋酸菌发酵转化贻贝壳钙源的影响研究

    Institute of Scientific and Technical Information of China (English)

    李晓娇; 刘书来; 丁玉庭


    研究了不同初始酒精浓度对醋酸发酵的影响及分批补料对贝壳钙源发酵的影响.醋酸茵在初始酒精浓度为6%vol时的产酸速率、菌体生长速率都较快,且其发酵周期适中.在此基础上,研究了分批补料发酵过程中菌体生长、产物及副产物的合成规律.结果表明:分批补料发酵通过改善发酵的环境条件,进而提高钙离子的转化率.与分批发酵相比,发酵中钙离子的转化率由18.08%提高到了37.33%,钙离子的总浓度由16.96mg/mL提高到了33.99mg/mL.因此,分批补料发酵可显著提高代谢产物的产量,促进贝壳钙源的生物转化率.%The effects of initial alcohol concentrations on acetic acid fermentation and the influence of fed-batch on calcium biotransformation of mussel shell were investigated. When the initial alcohol concentration was 6%vol, the acetic acid production and growth rate of f Acetobacter sp. were faster, and the fermentation time was proper. Base on this study, the cell growth rate and synthetic rates of metabolites in fed-batch fermentation were investigated. The results showed that fed-batch fermentation can improve the calcium conversion rate through changing the fermentation environment. Comparing with batch fermentation, the calcium conversion rate increased from 18.08% to 37.33%, and the total concentration of calcium increased from 16.96mg/ml to 33.99mg/ml in fed-bath fermentation. The fed-batch fermentation could significantly enhance the yield of acetic acid and promote the efficiency of calcium conversion.

  18. A discretized model for enzymatic hydrolysis of cellulose in a fed-batch process. (United States)

    Tervasmäki, Petri; Sotaniemi, Ville; Kangas, Jani; Taskila, Sanna; Ojamo, Heikki; Tanskanen, Juha


    In the enzymatic hydrolysis of cellulose, several phenomena have been proposed to cause a decrease in the reaction rate with increasing conversion. The importance of each phenomenon is difficult to distinguish from batch hydrolysis data. Thus, kinetic models for the enzymatic hydrolysis of cellulose often suffer from poor parameter identifiability. This work presents a model that is applicable to fed-batch hydrolysis by discretizing the substrate based on the feeding time. Different scenarios are tested to explain the observed decrease in reaction rate with increasing conversion, and comprehensive assessment of the parameter sensitivities is carried out. The proposed model performed well in the broad range of experimental conditions used in this study and when compared to literature data. Furthermore, the use of data from fed-batch experiments and discretization of the model substrate to populations was found to be very informative when assessing the importance of the rate-decreasing phenomena in the model. Copyright © 2016 Elsevier Ltd. All rights reserved.

  19. Cell mass energetic yields of fed-batch culture by Lipomyces starkeyi. (United States)

    Anschau, Andréia; Franco, Telma Teixeira


    Estimation of the energy capacity of a microbial cell mass on the basis of its lipid content and elemental composition can be used for the comparative evaluation of different microbial sources of biodiesel. Lipomyces starkeyi cell mass concentration reached 94.6 g/L with 37.4% of lipids in a fed-batch process using xylose and urea as substrates. The fatty acid composition of the yeast oil was quite similar to that of palm oil. L. starkeyi converted more than 80% of the energy contained in xylose into cell mass energy yield. The approach used in this study makes it possible to determine the energy of a cell mass by its elemental composition. A heat of combustion (Q c) of 25.7 (kJ/g) was obtained for the cell mass after 142 h of fed-batch cultivation, which represents approximately 56% of the energy content of diesel oil (45.4 kJ/g). The Q c of the triacylglycerols produced was 48.9 (kJ/g), indicating the potential of this oleaginous yeast for biodiesel production. Our work developed here provides a simple and efficient tool for characterization of this cell mass to further our understanding of its use as a feedstock for bioenergy production.

  20. Evaluation of selected control strategies for fed-batch cultures of a hybridoma cell line. (United States)

    Pörtner, Ralf; Schwabe, Jan-Oliver; Frahm, Björn


    While fed-batch suspension culture of animal cells continues to be of industrial importance for the large-scale production of pharmaceutical products, existing control concepts are still insufficient. The present paper illustrates the advantages and disadvantages of different fed-batch strategies, including fixed-feed trajectories, control via OUR (oxygen uptake rate) (stoichiometric feeding), a priori determination of feed trajectories based on a kinetic model and the model-based adaptive OLFO (open-loop-feedback-optimal) control strategy. A recommendation as to which control strategy should be used for a specific process has to consider the respective process. For an established process with a well characterized and stable production cell line, probably the application of a fixed feed trajectory should be recommended. An adaptive, model-based control strategy could be the method of choice during cell-line development or for rapid production of small amounts of product for clinical trials, owing to its universal character and because it does not require intensive process development.

  1. Fed-batch microbioreactor platform for scale down and analysis of a plasmid DNA production process. (United States)

    Bower, Diana M; Lee, Kevin S; Ram, Rajeev J; Prather, Kristala L J


    The rising costs of bioprocess research and development emphasize the need for high-throughput, low-cost alternatives to bench-scale bioreactors for process development. In particular, there is a need for platforms that can go beyond simple batch growth of the organism of interest to include more advanced monitoring, control, and operation schemes such as fed-batch or continuous. We have developed a 1-mL microbioreactor capable of monitoring and control of dissolved oxygen, pH, and temperature. Optical density can also be measured online for continuous monitoring of cell growth. To test our microbioreactor platform, we used production of a plasmid DNA vaccine vector (pVAX1-GFP) in Escherichia coli via a fed-batch temperature-inducible process as a model system. We demonstrated that our platform can accurately predict growth, glycerol and acetate concentrations, as well as plasmid copy number and quality obtained in a bench-scale bioreactor. The predictive abilities of the micro-scale system were robust over a range of feed rates as long as key process parameters, such as dissolved oxygen, were kept constant across scales. We have highlighted plasmid DNA production as a potential application for our microbioreactor, but the device has broad utility for microbial process development in other industries as well. Copyright © 2012 Wiley Periodicals, Inc.

  2. A fully defined, fed-batch, recombinant NS0 culture process for monoclonal antibody production. (United States)

    Hermes, Paul A; Castro, Chris D


    To manufacture a glycoprotein, mammalian cells expressing the desired protein are often grown in fed-batch mode. Feeding an undefined, nonanimal hydrolysate helps the cells receive sufficient nutrition, but makes systems difficult to optimize. Even different lots of the same hydrolysate may have significant variability; furthermore, individual components may actually be detrimental to the cells. Switching to fully defined feeds could eliminate these issues. For monoclonal antibody (mAb) production by fed-batch NS0 cells, this article describes the replacement of a hydrolysate-based feed with a fully defined, animal-component-free feed system. The defined feed initially had 67 components, but additional experiments allowed a reduction to 25 components. The mAb titer is approximately 20% higher than in the undefined system, and the feed volume is circa 20% lower. The two systems generated antibodies with similar glycosylation profiles. Other benefits of the defined feed system include lower raw material costs, the ability to optimize key nutrient concentrations, greater confidence in raw material quality, and the elimination of potential, hydrolysate-associated endotoxin issues.

  3. Concentrated fed-batch cell culture increases manufacturing capacity without additional volumetric capacity. (United States)

    Yang, William C; Minkler, Daniel F; Kshirsagar, Rashmi; Ryll, Thomas; Huang, Yao-Ming


    Biomanufacturing factories of the future are transitioning from large, single-product facilities toward smaller, multi-product, flexible facilities. Flexible capacity allows companies to adapt to ever-changing pipeline and market demands. Concentrated fed-batch (CFB) cell culture enables flexible manufacturing capacity with limited volumetric capacity; it intensifies cell culture titers such that the output of a smaller facility can rival that of a larger facility. We tested this hypothesis at bench scale by developing a feeding strategy for CFB and applying it to two cell lines. CFB improved cell line A output by 105% and cell line B output by 70% compared to traditional fed-batch (TFB) processes. CFB did not greatly change cell line A product quality, but it improved cell line B charge heterogeneity, suggesting that CFB has both process and product quality benefits. We projected CFB output gains in the context of a 2000-L small-scale facility, but the output was lower than that of a 15,000-L large-scale TFB facility. CFB's high cell mass also complicated operations, eroded volumetric productivity, and showed our current processes require significant improvements in specific productivity in order to realize their full potential and savings in manufacturing. Thus, improving specific productivity can resolve CFB's cost, scale-up, and operability challenges.

  4. A review of control strategies for manipulating the feed rate in fed-batch fermentation processes

    DEFF Research Database (Denmark)

    Mears, Lisa; Stocks, Stuart M.; Sin, Gürkan


    . This review covers a range of strategies which have been employed to use the feed rate as a manipulated variable in a control strategy. The feed rate is chosen as the focus for this review, as it is seen that this variable may be used towards many different objectives depending on the process of interest......, the characteristics of the strain, or the product being produced, which leads to different drivers for process optimisation. This review summarises the methods, as well as focusing on the different objectives for the controllers, and the choice of measured variables involved in the strategy. The discussion includes...... a summary of considerations for control strategy development....

  5. Mechanistic Models for Process Development and Optimization of Fed-batch Fermentation Systems

    DEFF Research Database (Denmark)

    Mears, Lisa; Stocks, Stuart M.; Albæk, Mads O.


    into account the oxygen transfer conditions, as well as the evaporation rates of the system. Mechanistic models are valuable tools which are applicable for both process development and optimization. The state estimator described will be a valuable tool for future work as part of control strategy development...... for on-line process control and optimization....

  6. Optimization of fed-batch fermentation for a staphylokinase-hirudin ...

    African Journals Online (AJOL)



    Aug 9, 2010 ... similar in the three feeding mediums, but the yield of total protein and STH were significantly ... The efficacy of STH production was 37 times more than that ... sion recombinant plasmid pBV220-STH was described in our.

  7. Fed-batch cultivation of Desmodesmus sp. in anaerobic digestion wastewater for improved nutrient removal and biodiesel production. (United States)

    Ji, Fang; Zhou, Yuguang; Pang, Aiping; Ning, Li; Rodgers, Kibet; Liu, Ying; Dong, Renjie


    Desmodesmus sp. was used in anaerobically digested wastewater (ADW) for nutrients removal and the biodiesel production was measured and compared using fed-batch cultivation was investigated and compared with batch cultivation. The Desmodesmus sp. was able to remove 236.143, 268.238 and 6.427 mg/L of TN, NH4-N and PO4-P respectively after 40 d of fed-batch cultivation, while in batch cultivation the quantities of TN, NH4-N and PO4-P removed were 33.331, 37.227 and 1.323 mg/L. Biomass production of Desmodesmus sp. was also enhanced in fed-batch cultivation, when ADW loading was carried out every 2 days; the biomass concentration peaked at 1.039 g/L, which was three times higher than that obtained in batch cultivation (0.385 g/L). The highest lipid production (261.8 mg/L) was also recorded in fed-batch cultivation as compared to batch cultivation (83.3 mg/L). Fed-batch cultivation of Desmodesmus sp. could provide effective control of nutrients limitation and/or ammonia inhibition on microalgae cultivation.

  8. Fuzzy control of ethanol concentration its application to maximum glutathione production in yeast fed-batch culture. (United States)

    Alfafara, C G; Miura, K; Shimizu, H; Shioya, S; Suga, K; Suzuki, K


    A fuzzy logic controller (FLC) for the control of ethanol concentration was developed and utilized to realize the maximum production of glutathione (GSH) in yeast fedbatch culture. A conventional fuzzy controller, which uses the control error and its rate of change in the premise part of the linguistic rules, worked well when the initial error of ethanol concentration was small. However, when the initial error was large, controller overreaction resulted in an overshoot.An improved fuzzy controller was obtained to avoid controller overreaction by diagnostic determination of "glucose emergency states" (i.e., glucose accumulation or deficiency), and then appropriate emergency control action was obtained by the use of weight coefficients and modification of linguistic rules to decrease the overreaction of the controller when the fermentation was in the emergency state. The improved fuzzy controller was able to control a constant ethanol concentration under conditions of large initial error.The improved fuzzy control system was used in the GSH production phase of the optimal operation to indirectly control the specific growth rate mu to its critical value micro(c). In the GSH production phase of the fed-batch culture, the optimal solution was to control micro to micro(c) in order to maintain a maximum specific GSH production rate. The value of micro(c) also coincided with the critical specific growth rate at which no ethanol formation occurs. Therefore, the control of micro to micro(c) could be done indirectly by maintaining a constant ethanol concentration, that is, zero net ethanol formation, through proper manipulation of the glucose feed rate. Maximum production of GSH was realized using the developed FLC; maximum production was a consequence of the substrate feeding strategy and cysteine addition, and the FLC was a simple way to realize the strategy.

  9. Modelling of Escherichia coli Cultivations: Acetate Inhibition in a Fed-batch Culture

    Directory of Open Access Journals (Sweden)

    Stoyan Tzonkov


    Full Text Available A set of three competing, unstructured models has been proposed to model biomass growth, glucose utilization, acetate formation, dissolved oxygen consumption and carbon dioxide accumulation of a fed-batch cultivation process of Escherichia coli. The inhibiting effect of acetate on growth of E. coli cultures is included in the considered models. The model identification is carried out using experimental data from the cultivation process. Genetic algorithms are used for parameter estimation. The model discrimination is based on the four criteria, namely sum of square errors, Fisher criterion, Akaike information criterion and minimum description length criterion. The most suitable model is identified that reflects the state variables curves adequately by considering acetate inhibited growth according to the Jerusalimsky approach.

  10. Mucor miehei's microbial rennin production characteristics in a fed-batch proccess

    Directory of Open Access Journals (Sweden)

    C. P. Sánchez Henao


    Full Text Available The Mucor miehei zygomycete produces an acid protease (EC: resembling calf rennet chymosin characteristics. It has been suggested that low glucose concentration levels could be why enzyme synthesis, co-mes to an end even though enzyme production is still great (Escobar and Barnett, 1993, 1995. To overcome this possible limitation, a two stage research process was designed; the relationship between protease production and sugar consumption was studied initially to determine the periods of time when enzyme production is still high and glucose concentration close to zero. The following stage concentrated on developing a glucose fed-batch process during the afore mentioned time periods to observe any increase or decrease in enzyme production. During the batch studies, it was found that maximum enzyme activity (EA was 165 UC/ml for an average glucose consumption rate of 0.1813 g/1 h. Based on the previous.

  11. Comparison of biomass estimation techniques for a Bacillus thuringiensis fed-batch culture

    Energy Technology Data Exchange (ETDEWEB)

    Cunha, C.C.F. [University of Newcastle upon Tyne (United Kingdom). Dept. of Chemical and Process Engineering]. E-mail:; Souza Junior, M.B. [Universidade Federal, Rio de Janeiro, RJ (Brazil). Escola de Quimica]. E-mail:


    In this work, the ability of artificial neural nets was investigated for the on-line biomass prediction of the simulated growth of a strain of Bacillus thuringiensis in fed-batch mode. For this purpose, multilayered backpropagation nets with sigmoid nodes were trained. The patterns were composed of input data on current values of biomass concentration, limiting substrate concentration and dilution rate, and output data on prediction of biomass concentration for the following step. The dilution rate was disturbed by a PRBS input, and simulations were conducted using a phenomenological experimentally validated model. The nets were able to predict the biomass concentration for different feeding techniques, and they were also compared with the variable estimation technique using the extended Kalman filter. (author)

  12. Improved methane production from brown algae under high salinity by fed-batch acclimation. (United States)

    Miura, Toyokazu; Kita, Akihisa; Okamura, Yoshiko; Aki, Tsunehiro; Matsumura, Yukihiko; Tajima, Takahisa; Kato, Junichi; Nakashimada, Yutaka


    Here, a methanogenic microbial community was developed from marine sediments to have improved methane productivity from brown algae under high salinity. Fed-batch cultivation was conducted by adding dry seaweed at 1wt% total solid (TS) based on the liquid weight of the NaCl-containing sediment per round of cultivation. The methane production rate and level of salinity increased 8-fold and 1.6-fold, respectively, at the 10th round of cultivation. Moreover, the rate of methane production remained high, even at the 10th round of cultivation, with accumulation of salts derived from 10wt% TS of seaweed. The salinity of the 10th-round culture was equivalent to 5% NaCl. The improved methane production was attributed to enhanced acetoclastic methanogenesis because acetate became rapidly converted to methane during cultivation. The family Fusobacteriaceae and the genus Methanosaeta, the acetoclastic methanogen, predominated in bacteria and archaea, respectively, after the cultivation.

  13. Statistical methods in media optimization for batch and fed-batch animal cell culture. (United States)

    De Alwis, Diliny M; Dutton, Roshni L; Scharer, Jeno; Moo-Young, Murray


    Hybridoma 130-8F producing anti-F monoclonal antibodies (MAb) were grown in batch and fed-batch mode with glutamine as the limiting substrate. The initial concentration of glucose varied between 10 and 25 mM but was not growth limiting. Monoclonal antibody production was identified as being partially growth associated. Employing the cumulative cell hour concept, external metabolic flux estimates were calculated during the exponential growth phase for MAb, glucose, amino acids, ammonia and lactate. Through nutritional profiling using principal component analysis (PCA) followed by partial least squares regression (PLS), key metabolites were identified and grouped for significant positive, significant negative, low level, and negligible correlation to MAb production, cellular growth, glucose consumption, and ammonia and lactate production. Significant relationships peculiar to Hybridoma 130-8F were identified, such as demand for two normally non-essential amino acids (asparagine and aspartic acid), and the positive correlation between MAb and ammonia production.

  14. Simulation and prediction of protein production in fed-batch E. coli cultures: An engineering approach. (United States)

    Calleja, Daniel; Kavanagh, John; de Mas, Carles; López-Santín, Josep


    An overall model describing the dynamic behavior of fed-batch E. coli processes for protein production has been built, calibrated and validated. Using a macroscopic approach, the model consists of three interconnected blocks allowing simulation of biomass, inducer and protein concentration profiles with time. The model incorporates calculation of the extra and intracellular inducer concentration, as well as repressor-inducer dynamics leading to a successful prediction of the product concentration. The parameters of the model were estimated using experimental data of a rhamnulose-1-phosphate aldolase-producer strain, grown under a wide range of experimental conditions. After validation, the model has successfully predicted the behavior of different strains producing two different proteins: fructose-6-phosphate aldolase and ω-transaminase. In summary, the presented approach represents a powerful tool for E. coli production process simulation and control.

  15. On-line optimal control for fed-batch culture of baker's yeast production

    Energy Technology Data Exchange (ETDEWEB)

    Wu, W.T.; Chen, K.C.; Chiou, H.W.


    A method of on-line optimal control for fed-batch culture of bakers yeast production is proposed. The feed rate is taken as the control variable. The specific growth rate of the yeast is the output variable and is determined from the balance equation of oxygen. A moving model is obtained by using the data from the feed rate and the specific growth rate. Based on the moving model, an optimal feed rate for fed-batch culture is then achieved. 11 references.

  16. Brunovsky Normal Form of Monod Kinetics Models and Growth Rate Control of a Fed-batch Cultivation Process

    Directory of Open Access Journals (Sweden)

    Pavlov Y.


    Full Text Available A mathematical methodology that gives assistance to design of fed-batch stabilization and control is presented. The methodology is based both on Utility theory and optimal Control theory. The Utility theory deals with the expressed subjective preferences and allows for the expert preferences to be taken in consideration in complex biotechnological systems as criteria for control and optimization. The Control theory is used for parameters stabilization of a fed-batch cultivation process. The control is written based on information of the growth rate. The simulations show good efficiency of the control laws.

  17. Segmented linear modeling of CHO fed-batch culture and its application to large scale production. (United States)

    Ben Yahia, Bassem; Gourevitch, Boris; Malphettes, Laetitia; Heinzle, Elmar


    We describe a systematic approach to model CHO metabolism during biopharmaceutical production across a wide range of cell culture conditions. To this end, we applied the metabolic steady state concept. We analyzed and modeled the production rates of metabolites as a function of the specific growth rate. First, the total number of metabolic steady state phases and the location of the breakpoints were determined by recursive partitioning. For this, the smoothed derivative of the metabolic rates with respect to the growth rate were used followed by hierarchical clustering of the obtained partition. We then applied a piecewise regression to the metabolic rates with the previously determined number of phases. This allowed identifying the growth rates at which the cells underwent a metabolic shift. The resulting model with piecewise linear relationships between metabolic rates and the growth rate did well describe cellular metabolism in the fed-batch cultures. Using the model structure and parameter values from a small-scale cell culture (2 L) training dataset, it was possible to predict metabolic rates of new fed-batch cultures just using the experimental specific growth rates. Such prediction was successful both at the laboratory scale with 2 L bioreactors but also at the production scale of 2000 L. This type of modeling provides a flexible framework to set a solid foundation for metabolic flux analysis and mechanistic type of modeling. Biotechnol. Bioeng. 2017;114: 785-797. © 2016 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc. © 2016 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc.

  18. Temperature limited fed-batch technique for control of proteolysis in Pichia pastoris bioreactor cultures

    Directory of Open Access Journals (Sweden)

    Garcia Percival


    Full Text Available Abstract Background A temperature limited fed-batch (TLFB technique is described and used for Pichia pastoris Mut+ strain cultures and compared with the traditional methanol limited fed-batch (MLFB technique. A recombinant fusion protein composed of a cellulose-binding module (CBM from Neocallimastix patriciarum cellulase 6A and lipase B from Candida antarctica (CALB, was produced and secreted by this strain. Results A protein concentration of about 1 g L-1 was produced in the MLFB process. However, this product was considerably degraded by protease(s. By applying the TLFB process, the yield was increased to 2 g L-1 full-length product and no proteolytic degradation was observed. Flow cytometry analysis showed that the percentage of dead cells increased rapidly during the initial methanol feed phase in the MLFB process and reached a maximum of about 12% after about 40–70 hours of methanol feeding. In the TLFB process, cell death rate was low and constant and reached 4% dead cells at the end of cultivation (about 150 hours methanol feeding time. The lower cell death rate in the TLFB correlated with a lower protease activity in the culture supernatant. The specific alcohol oxidase (AOX activity in the TLFB process was 3.5 times higher than in the MLFB process. Conclusion Three mechanisms that may contribute to the much higher accumulation of product in the TLFB process are: 1 reduced proteolysis due to lower temperature, 2 reduced proteolysis due to lower cell death and protease release to the medium, 3 increased synthesis rate due to higher AOX activity.

  19. Quantitative evaluation of yeast's requirement for glycerol formation in very high ethanol performance fed-batch process

    Directory of Open Access Journals (Sweden)

    Nevoigt Elke


    Full Text Available Abstract Background Glycerol is the major by-product accounting for up to 5% of the carbon in Saccharomyces cerevisiae ethanolic fermentation. Decreasing glycerol formation may redirect part of the carbon toward ethanol production. However, abolishment of glycerol formation strongly affects yeast's robustness towards different types of stress occurring in an industrial process. In order to assess whether glycerol production can be reduced to a certain extent without jeopardising growth and stress tolerance, the yeast's capacity to synthesize glycerol was adjusted by fine-tuning the activity of the rate-controlling enzyme glycerol 3-phosphate dehydrogenase (GPDH. Two engineered strains whose specific GPDH activity was significantly reduced by two different degrees were comprehensively characterized in a previously developed Very High Ethanol Performance (VHEP fed-batch process. Results The prototrophic strain CEN.PK113-7D was chosen for decreasing glycerol formation capacity. The fine-tuned reduction of specific GPDH activity was achieved by replacing the native GPD1 promoter in the yeast genome by previously generated well-characterized TEF promoter mutant versions in a gpd2Δ background. Two TEF promoter mutant versions were selected for this study, resulting in a residual GPDH activity of 55 and 6%, respectively. The corresponding strains were referred to here as TEFmut7 and TEFmut2. The genetic modifications were accompanied to a strong reduction in glycerol yield on glucose; the level of reduction compared to the wild-type was 61% in TEFmut7 and 88% in TEFmut2. The overall ethanol production yield on glucose was improved from 0.43 g g-1 in the wild type to 0.44 g g-1 measured in TEFmut7 and 0.45 g g-1 in TEFmut2. Although maximal growth rate in the engineered strains was reduced by 20 and 30%, for TEFmut7 and TEFmut2 respectively, strains' ethanol stress robustness was hardly affected; i.e. values for final ethanol concentration (117 ± 4 g

  20. Glucoamylase production in batch, chemostat and fed-batch cultivations by an industrial strain of Aspergillus niger

    DEFF Research Database (Denmark)

    Pedersen, Henrik; Beyer, Michael; Nielsen, Jens


    The Aspergillus niger strain BO-1 was grown in batch, continuous (chemostat) and fed-batch cultivations in order to study the production of the extracellular enzyme glucoamylase under different growth conditions. In the pH range 2.5-6.0, the specific glucoamylase productivity and the specific gro...

  1. High-cell-density fed-batch cultivation of the docosahexaenoic acid producing marine alga Crypthecodinium cohnii

    NARCIS (Netherlands)

    Swaaf, de M.E.; Sijtsma, L.; Pronk, J.T.


    The heterotrophic marine alga Crypthecodinium cohnii is known to produce docosahexaenoic acid (DHA), a polyunsaturated fatty acid with food and pharmaceutical applications, during batch cultivation on complex media containing sea salt, yeast extract, and glucose. In the present study, fed-batch cult

  2. Glucoamylase production in batch, chemostat and fed-batch cultivations by an industrial strain of Aspergillus niger

    DEFF Research Database (Denmark)

    Pedersen, Henrik; Beyer, Michael; Nielsen, Jens


    The Aspergillus niger strain BO-1 was grown in batch, continuous (chemostat) and fed-batch cultivations in order to study the production of the extracellular enzyme glucoamylase under different growth conditions. In the pH range 2.5-6.0, the specific glucoamylase productivity and the specific...

  3. High-cell-density fed-batch cultivation of the docosahexaenoic acid producing marine alga Crypthecodinium cohnii

    NARCIS (Netherlands)

    Swaaf, de M.E.; Sijtsma, L.; Pronk, J.T.


    The heterotrophic marine alga Crypthecodinium cohnii is known to produce docosahexaenoic acid (DHA), a polyunsaturated fatty acid with food and pharmaceutical applications, during batch cultivation on complex media containing sea salt, yeast extract, and glucose. In the present study, fed-batch cult

  4. Degradation of toluene and trichloroethylene by Burkholderia cepacia G4 in growth-limited fed-batch culture

    NARCIS (Netherlands)

    Mars, Astrid E.; Houwing, Joukje; Dolfing, Jan; Janssen, Dick B.


    Burkholderia (Pseudomonas) cepacia G4 was cultivated in a fed-batch bioreactor on either toluene or toluene plus trichloroethylene (TCE), The culture was allowed to reach a constant cell density under conditions in which the amount of toluene supplied equals the maintenance energy demand of the cult

  5. Determination of model parameters for zinc (II) ion biosorption onto powdered waste sludge (PWS) in a fed-batch system. (United States)

    Kargi, Fikret; Cikla, Sinem


    Biosorption of zinc (II) ions onto pre-treated powdered waste sludge (PWS) was investigated using a completely mixed tank operating in fed-batch mode instead of an adsorption column. Experiments with variable feed flow rate (0.05-0.5 L h(-1)), feed Zn(II) ion concentrations (37.5-275 mg L(-1)) and amount of adsorbent (1-6 g PWS) were performed using fed-batch operation at pH 5 and room temperature (20-25 degrees C). Break-through curves describing variations of aqueous (effluent) zinc ion concentrations with time were determined for different operating conditions. Percent zinc removal from the aqueous phase decreased, but the biosorbed (solid phase) zinc ion concentration increased with increasing feed flow rate and zinc concentration. A modified Bohart-Adams equation was used to determine the biosorption capacity of PWS (q'(s)) and the rate constant (K) for zinc ion biosorption. Biosorption capacity (q'(s)=57.7 g Zn kg(-1) PWS) of PWS in fed-batch operation was found to be comparable with powdered activated carbon (PAC) in column operations. However, the adsorption rate constant (K=9.17 m(3) kg(-1) h(-1)) in fed-batch operation was an order of magnitude larger than those obtained in adsorption columns because of elimination of mass transfer limitations encountered in the column operations. Therefore, a completely mixed tank operated in fed-batch mode was proven to be more advantageous as compared to adsorption columns due to better contact between the phases yielding faster adsorption rates.

  6. In Vitro Growth of Curcuma longa L. in Response to Five Mineral Elements and Plant Density in Fed-Batch Culture Systems (United States)

    El-Hawaz, Rabia F.; Bridges, William C.; Adelberg, Jeffrey W.


    Plant density was varied with P, Ca, Mg, and KNO3 in a multifactor experiment to improve Curcuma longa L. micropropagation, biomass and microrhizome development in fed-batch liquid culture. The experiment had two paired D-optimal designs, testing sucrose fed-batch and nutrient sucrose fed-batch techniques. When sucrose became depleted, volume was restored to 5% m/v sucrose in 200 ml of modified liquid MS medium by adding sucrose solutions. Similarly, nutrient sucrose fed-batch was restored to set points with double concentration of treatments’ macronutrient and MS micronutrient solutions, along with sucrose solutions. Changes in the amounts of water and sucrose supplementations were driven by the interaction of P and KNO3 concentrations. Increasing P from 1.25 to 6.25 mM increased both multiplication and biomass. The multiplication ratio was greatest in the nutrient sucrose fed-batch technique with the highest level of P, 6 buds/vessel, and the lowest level of Ca and KNO3. The highest density (18 buds/vessel) produced the highest fresh biomass at the highest concentrations of KNO3 and P with nutrient sucrose fed-batch, and moderate Ca and Mg concentrations. However, maximal rhizome dry biomass required highest P, sucrose fed-batch, and a moderate plant density. Different media formulations and fed-batch techniques were identified to maximize the propagation and storage organ responses. A single experimental design was used to optimize these dual purposes. PMID:25830292

  7. In vitro growth of Curcuma longa L. in response to five mineral elements and plant density in fed-batch culture systems. (United States)

    El-Hawaz, Rabia F; Bridges, William C; Adelberg, Jeffrey W


    Plant density was varied with P, Ca, Mg, and KNO3 in a multifactor experiment to improve Curcuma longa L. micropropagation, biomass and microrhizome development in fed-batch liquid culture. The experiment had two paired D-optimal designs, testing sucrose fed-batch and nutrient sucrose fed-batch techniques. When sucrose became depleted, volume was restored to 5% m/v sucrose in 200 ml of modified liquid MS medium by adding sucrose solutions. Similarly, nutrient sucrose fed-batch was restored to set points with double concentration of treatments' macronutrient and MS micronutrient solutions, along with sucrose solutions. Changes in the amounts of water and sucrose supplementations were driven by the interaction of P and KNO3 concentrations. Increasing P from 1.25 to 6.25 mM increased both multiplication and biomass. The multiplication ratio was greatest in the nutrient sucrose fed-batch technique with the highest level of P, 6 buds/vessel, and the lowest level of Ca and KNO3. The highest density (18 buds/vessel) produced the highest fresh biomass at the highest concentrations of KNO3 and P with nutrient sucrose fed-batch, and moderate Ca and Mg concentrations. However, maximal rhizome dry biomass required highest P, sucrose fed-batch, and a moderate plant density. Different media formulations and fed-batch techniques were identified to maximize the propagation and storage organ responses. A single experimental design was used to optimize these dual purposes.

  8. Fed-Batch Production of Bacterial Ghosts Using Dielectric Spectroscopy for Dynamic Process Control. (United States)

    Meitz, Andrea; Sagmeister, Patrick; Lubitz, Werner; Herwig, Christoph; Langemann, Timo


    The Bacterial Ghost (BG) platform technology evolved from a microbiological expression system incorporating the ϕX174 lysis gene E. E-lysis generates empty but structurally intact cell envelopes (BGs) from Gram-negative bacteria which have been suggested as candidate vaccines, immunotherapeutic agents or drug delivery vehicles. E-lysis is a highly dynamic and complex biological process that puts exceptional demands towards process understanding and control. The development of a both economic and robust fed-batch production process for BGs required a toolset capable of dealing with rapidly changing concentrations of viable biomass during the E-lysis phase. This challenge was addressed using a transfer function combining dielectric spectroscopy and soft-sensor based biomass estimation for monitoring the rapid decline of viable biomass during the E-lysis phase. The transfer function was implemented to a feed-controller, which followed the permittivity signal closely and was capable of maintaining a constant specific substrate uptake rate during lysis phase. With the described toolset, we were able to increase the yield of BG production processes by a factor of 8-10 when compared to currently used batch procedures reaching lysis efficiencies >98%. This provides elevated potentials for commercial application of the Bacterial Ghost platform technology.

  9. Metabolic characterization of a CHO cell size increase phase in fed-batch cultures. (United States)

    Pan, Xiao; Dalm, Ciska; Wijffels, René H; Martens, Dirk E


    Normally, the growth profile of a CHO cell fed-batch process can be divided into two main phases based on changes in cell concentration, being an exponential growth phase and a stationary (non-growth) phase. In this study, an additional phase is observed during which the cell division comes to a halt but the cell growth continues in the form of an increase in cell size. The cell size increase (SI) phase occurs between the exponential proliferation phase (also called the number increase or NI phase) and the stationary phase. During the SI phase, the average volume and dry weight per cell increase threefold linearly with time. The average mAb specific productivity per cell increases linearly with the cell volume and therefore is on average two times higher in the SI phase than in the NI phase. The specific essential amino acids consumption rates per cell remain fairly constant between the NI and the SI phase, which agrees with the similar biomass production rate per cell between these two phases. Accumulation of fatty acids and formation of lipid droplets in the cells are observed during the SI phase, indicating that the fatty acids synthesis rate exceeds the demand for the synthesis of membrane lipids. A metabolic comparison between NI and SI phase shows that the cells with a larger size produce more mAb per unit of O2 and nutrient consumed, which can be used for further process optimization.

  10. Optimization and robustness analysis of hybridoma cell fed-batch cultures using the overflow metabolism model. (United States)

    Amribt, Z; Dewasme, L; Vande Wouwer, A; Bogaerts, Ph


    The maximization of biomass productivity in fed-batch cultures of hybridoma cells is analyzed based on the overflow metabolism model. Due to overflow metabolism, often attributed to limited oxygen capacity, lactate and ammonia are formed when the substrate concentrations (glucose and glutamine) are above a critical value, which results in a decrease in biomass productivity. Optimal feeding rate, on the one hand, for a single feed stream containing both glucose and glutamine and, on the other hand, for two separate feed streams of glucose and glutamine are determined using a Nelder-Mead simplex optimization algorithm. The optimal multi exponential feed rate trajectory improves the biomass productivity by 10 % as compared to the optimal single exponential feed rate. Moreover, this result is validated by the one obtained with the analytical approach in which glucose and glutamine are fed to the culture so as to control the hybridoma cells at the critical metabolic state, which allows maximizing the biomass productivity. The robustness analysis of optimal feeding profiles obtained with different optimization strategies is considered, first, with respect to parameter uncertainties and, finally, to model structure errors.

  11. Fed-Batch Production of Bacterial Ghosts Using Dielectric Spectroscopy for Dynamic Process Control

    Directory of Open Access Journals (Sweden)

    Andrea Meitz


    Full Text Available The Bacterial Ghost (BG platform technology evolved from a microbiological expression system incorporating the ϕX174 lysis gene E. E-lysis generates empty but structurally intact cell envelopes (BGs from Gram-negative bacteria which have been suggested as candidate vaccines, immunotherapeutic agents or drug delivery vehicles. E-lysis is a highly dynamic and complex biological process that puts exceptional demands towards process understanding and control. The development of a both economic and robust fed-batch production process for BGs required a toolset capable of dealing with rapidly changing concentrations of viable biomass during the E-lysis phase. This challenge was addressed using a transfer function combining dielectric spectroscopy and soft-sensor based biomass estimation for monitoring the rapid decline of viable biomass during the E-lysis phase. The transfer function was implemented to a feed-controller, which followed the permittivity signal closely and was capable of maintaining a constant specific substrate uptake rate during lysis phase. With the described toolset, we were able to increase the yield of BG production processes by a factor of 8–10 when compared to currently used batch procedures reaching lysis efficiencies >98%. This provides elevated potentials for commercial application of the Bacterial Ghost platform technology.

  12. Modulation of mAb quality attributes using microliter scale fed-batch cultures. (United States)

    Rouiller, Yolande; Périlleux, Arnaud; Vesin, Marie-Noëlle; Stettler, Matthieu; Jordan, Martin; Broly, Hervé


    A high-throughput DoE approach performed in a 96-deepwell plate system was used to explore the impact of media and feed components on main quality attributes of a monoclonal antibody. Six CHO-S derived clonal cell lines expressing the same monoclonal antibody were tested in two different cell culture media with six components added at three different levels. The resulting 384 culture conditions including controls were simultaneously tested in fed-batch conditions, and process performance such as viable cell density, viability, and product titer were monitored. At the end of the culture, supernatants from each condition were purified and the product was analyzed for N-glycan profiles, charge variant distribution, aggregates, and low molecular weight forms. The screening described here provided highly valuable insights into the factors and combination of factors that can be used to modulate the quality attributes of a molecule. The approach also revealed specific intrinsic differences of the selected clonal cell lines - some cell lines were very responsive in terms of changes in performance or quality attributes, whereas others were less affected by the factors tested in this study. Moreover, it indicated to what extent the attributes can be impacted within the selected experimental design space. The outcome correlated well with confirmations performed in larger cell culture volumes such as small-scale bioreactors. Being fast and resource effective, this integrated high-throughput approach can provide information which is particularly useful during early stage cell culture development. © 2014 American Institute of Chemical Engineers.

  13. Optimal control for nonlinear dynamical system of microbial fed-batch culture (United States)

    Liu, Chongyang


    In fed-batch culture of glycerol bio-dissimilation to 1, 3-propanediol (1, 3-PD), the aim of adding glycerol is to obtain as much 1, 3-PD as possible. So a proper feeding rate is required during the process. Taking the concentration of 1, 3-PD at the terminal time as the performance index and the feeding rate of glycerol as the control function, we propose an optimal control model subject to a nonlinear dynamical system and constraints of continuous state and non-stationary control. A computational approach is constructed to seek the solution of the above model in two aspects. On the one hand we transcribe the optimal control model into an unconstrained one based on the penalty functions and an extension of the state space; on the other hand, by approximating the control function with simple functions, we transform the unconstrained optimal control problem into a sequence of nonlinear programming problems, which can be solved using gradient-based optimization techniques. The convergence analysis of this approximation is also investigated. Numerical results show that, by employing the optimal control policy, the concentration of 1, 3-PD at the terminal time can be increased considerably.

  14. Identification of genes whose expressions are enhanced or reduced in baker's yeast during fed-batch culture process using molasses medium by DNA microarray analysis. (United States)

    Shima, Jun; Kuwazaki, Seigo; Tanaka, Fumiko; Watanabe, Hajime; Yamamoto, Hideki; Nakajima, Ryoichi; Tokashiki, Tadaaki; Tamura, Hiromi


    Genes whose expression levels are enhanced or reduced during the cultivation process that uses cane molasses in baker's yeast production were identified in this study. The results showed that baker's yeast grown in molasses medium had higher fermentation ability and stress tolerance compared with baker's yeast grown in synthetic medium. Molasses apparently provided not only sugar as a carbon source but also provided functional components that enhanced or reduced expression of genes involved in fermentation ability and stress tolerance. To identify the genes whose expression is enhanced or reduced during cultivation in molasses medium, DNA microarray analysis was then used to compare the gene expression profile of cells grown in molasses with that of cells grown in synthetic medium. To simulate the commercial baker's yeast production process, cells were cultivated using a fed-batch culture system. In molasses medium, genes involved in the synthesis or uptake of vitamins (e.g., biotin, pyridoxine and thiamine) showed enhanced expression, suggesting that vitamin concentrations in molasses medium were lower than those in synthetic medium. Genes involved in formate dehydrogenase and maltose assimilation showed enhanced expression in molasses medium. In contrast, genes involved in iron utilization (e.g., siderophore, iron transporter and ferroxidase) showed enhanced expression in synthetic medium, suggesting that iron starvation occurred. The genes involved in the metabolism of amino acids also showed enhanced expression in synthetic medium. This identification of genes provides information that will help improve the baker's yeast production process.

  15. Fed-batch methanol feeding strategy for recombinant protein production by Pichia pastoris in the presence of co-substrate sorbitol. (United States)

    Celik, Eda; Calik, Pinar; Oliver, Stephen G


    Batch-wise sorbitol addition as a co-substrate at the induction phase of methanol fed-batch fermentation by Pichia pastoris (Mut(+)) was proposed as a beneficial recombinant protein production strategy and the metabolic responses to methanol feeding rate in the presence of sorbitol was systematically investigated. Adding sorbitol batch-wise to the medium provided the following advantages over growth on methanol alone: (a) eliminating the long lag-phase for the cells and reaching 'high cell density production' at t = 24 h of the process (C(X) = 70 g CDW/l); (b) achieving 1.8-fold higher recombinant human erythropoietin (rHuEPO) (at t = 18 h); (c) reducing specific protease production 1.2-fold; (d) eliminating the lactic acid build-up period; (e) lowering the oxygen uptake rate two-fold; and (f) obtaining 1.4-fold higher overall yield coefficients. The maximum specific alcohol oxidase activity was not affected in the presence of sorbitol, and it was observed that sorbitol and methanol were utilized simultaneously. Thus, in the presence of sorbitol, 130 mg/l rHuEPO was produced at t = 24 h, compared to 80 mg/l rHuEPO (t = 24 h) on methanol alone. This work demonstrates not only the ease and efficiency of incorporating sorbitol to fermentations by Mut(+) strains of P. pastoris for the production of any bio-product, but also provides new insights into the metabolism of the methylotrophic yeast P. pastoris.

  16. Conversion of a CHO cell culture process from perfusion to fed-batch technology without altering product quality. (United States)

    Meuwly, F; Weber, U; Ziegler, T; Gervais, A; Mastrangeli, R; Crisci, C; Rossi, M; Bernard, A; von Stockar, U; Kadouri, A


    During the development of a new drug product, it is a common strategy to develop a first-generation process with the aim to rapidly produce material for pre-clinical and early stage clinical trials. At a later stage of the development, a second-generation process is then introduced with the aim to supply late-stage clinical trials as well as market needs. This work was aimed at comparing the performance of two different CHO cell culture processes (perfusion and fed-batch) used for the production of a therapeutically active recombinant glycoprotein at industrial pilot-scale. The first-generation process was based on the Fibra-Cel packed-bed perfusion technology. It appeared during the development of the candidate drug that high therapeutic doses were required (>100mg per dose), and that future market demand would exceed 100 kg per year. This exceeded by far the production capacity of the first-generation process, and triggered a change of technology from a packed-bed perfusion process with limited scale-up capabilities to a fed-batch process with scale-up potential to typical bioreactor sizes of 15m(3) or more. The productivity per bioreactor unit volume (in product m(-3)year(-1)) of the fed-batch process was about 70% of the level reached with the first-generation perfusion process. However, since the packed-bed perfusion system was limited in scale (0.6m(3) maximum) compared to the volumes reached in suspension cultures (15m(3)), the fed-batch was selected as second-generation process. In fact, the overall process performance (in product year(-1)) was about 18-fold higher for the fed-batch compared to the perfusion mode. Data from perfusion and fed-batch harvests samples indicated that comparable product quality (relative abundance of monomers dimers and aggregates; N-glycan sialylation level; isoforms distribution) was obtained in both processes. To further confirm this observation, purification to homogeneity of the harvest material from both processes, followed


    Directory of Open Access Journals (Sweden)

    A.S. Silva


    Full Text Available Cephalosporin C, a b -lactam antibiotic, is the starting molecule for industrial production of semi-synthetic cephalosporins. The bioprocess for its production is carried out in batch stirred and aerated tank reactors utilizing strains of the filamentous fungus Cephalosporium acremonium. In this work a comparison was made between the processes of production of cephalosporin C in a conventional batch bioreactor, with synthetic medium containing glucose and sucrose, and in a fed-batch reactor at several flowrates of supplementary medium containing invert sucrose. In general, the fed-batch process was shown to be more efficient than the conventional batch one, and the process in which the lowest supplementation flowrate was used presented an antibiotic production significantly higher than those obtained under the other conditions.

  18. Production of savinase and population viability of Bacillus clausii during high-cell-density fed-batch cultivations

    DEFF Research Database (Denmark)

    Christiansen, Torben; Michaelsen, S.; Wumpelmann, M.


    feed profiles applied and, in addition, there was a time-dependent decrease in specific productivity. The specific glucose uptake rate increased with time for constant specific growth rate indicating that the maintenance requirements increased with time, possibly due to a decreasing K+ concentration......The growth and product formation of a Savinase-producing Bacillus clausii were investigated in high-cell-density fed-batch cultivations with both linear and exponential feed profiles. The highest specific productivity of Savinase was observed shortly after the end of the initial batch phase for all....... The physiological state of the cells was monitored during the cultivations using a flow cytometry assay based on the permeability of the cell membrane to propidium iodide. In the latter parts of the fed-batch cultures with a linear feed profile, a large portion of the cell population was found to have a permeable...

  19. Amino acid and glucose metabolism in fed-batch CHO cell culture affects antibody production and glycosylation

    DEFF Research Database (Denmark)

    Fan, Yuzhou; Jimenez Del Val, Ioscani; Müller, Christian


    optimization, especially media optimization. Gaining knowledge on their interrelations could provide insight for obtaining higher immunoglobulin G (IgG) titer and better controlling glycosylationrelated product quality. In this work, different fed-batch processes with two chemically defined proprietary media...... and glutamine concentrations and uptake rates were positively correlated with intracellular UDP-Gal availability. All these findings are important for optimization of fed-batch culture for improving IgG production and directing glycosylation quality....... energy and recombinant product, respectively. Accumulation of by-products such as NH4+ and lactate as a consequence of unbalanced nutrient supply to cell activities inhibits cell growth. The levels of Leu and Arg in the culture, which relate to cell growth and IgG productivity, need to be well controlled...

  20. Enhanced expression of the recombinant lethal factor of Bacillus anthracis by Fed-Batch culture. (United States)

    Gupta, P; Sahai, V; Bhatnagar, R


    High cell density cultivation has been one of the most effective ways to increase cell as well as the product yields. The structural gene for the 90-kDa lethal factor (LF) isolated from Bacillus anthracis was expressed as fusion protein with 6x histidine residues under the transcriptional regulation of the T5 promoter in Escherichia coli. Various strategies were tried to scale up the expression of the recombinant lethal factor by bioprocess optimization using fed batch culture technique in a 14 litre fermentor. The media, a defined mixture of salts, trace elements, vitamins, etc. along with a specified carbon source was used for the growth. The pH of the media was maintained at 6.8 while the temperature was changed from 37 to 28 degrees C during the cultivation. During the growth and induction phases, the DO was maintained above 20% by automatic control of agitation. The specific growth rate was controlled by utilizing an exponential feeding profile determined from mass balance equations. As a result of control of specific growth rate at two different levels, there was about twenty five fold increase in biomass compared to the biomass in the shake flask. E. coli cells yielded a soluble cytosolic protein with an apparent molecular mass of 90 kDa. The protein was purified to homogeneity using metal chelate affinity chromatography, followed by anion exchange on FPLC using Mono-Q column. In solution, trypsin cleaved protective antigen bound to native and recombinant LF with comparable affinity. The recombinant LF resembled the LF purified from B. anthracis in the macrophage lysis assay, using a murine macrophage cell line J774A.1 sensitive to anthrax toxin. It was possible to achieve a yield of 50 mg of the purified protein from 1 litre culture broth. Copyright 2001 Academic Press.

  1. Removal of polychlorinated dioxins by semi-aerobic fed-batch composting with biostimulation of "Dehalococcoides". (United States)

    Narihiro, Takashi; Kaiya, Shinichi; Futamata, Hiroyuki; Hiraishi, Akira


    A semi-aerobic, mesophilic, fed-batch composting (FBC) reactor loaded with household garbage was used to remove polychlorinated dibenzo-p-dioxins/dibenzofurans (PCDD/Fs). The reactor was packed with woodchips as the solid matrix and PCDD/F-contaminated soil or flyash and then operated at a waste-loading rate of 0.5 kg (wet wt) day(-1). All congeners of PCDD/Fs (initial concentration, 200-830 pmol g(-1) [dry wt]) were totally reduced during the over period of operation, with a half reduction time of 4 months. Direct cell counting and respiratory quinone profiling showed that the reactors at the fully acclimated stage harbored a high population density of bacteria (10(11) g(-1) [dry wt]) with members of the Actinobacteria predominating. Real-time quantitative PCR showed that the population of "Dehalococcoides" and its phylogenetic relatives of Chloroflexi as the possible dechlorinators varied between at the order of 10(7) to 10(8) g(-1) (dry wt). A "Dehalococcoides"-containing dechlorinating culture from the soil-treating reactor was successfully enriched with a model PCDD/F compound, fthalide. 16S rRNA gene-targeted PCR-denaturated gradient gel electrophoresis and clone library analyses showed that this culture comprised at least three major phylogenetic groups of bacteria, Acidaminobacter, "Dehalococcoides," and Rhizobium. These results suggest that the semi-aerobic FBC process is applicable for the bioremediation of PCDD/Fs and possibly other haloorganic compounds with the biostimulation of "Dehalococcoides" and its relatives as the potent dechlorinators.

  2. Development of a two-stage feeding strategy based on the kind and level of feeding nutrients for improving fed-batch production of L-threonine by Escherichia coli. (United States)

    Liu, Shuwen; Liang, Yong; Liu, Qian; Tao, Tongtong; Lai, Shujuan; Chen, Ning; Wen, Tingyi


    Fed-batch fermentation is the predominant method for industrial production of amino acids. In this study, we comprehensively investigated the effects of four kinds of feeding nutrients and developed an accurate optimization strategy for fed-batch production of L-threonine. The production of L-threonine was severely inhibited when cell growth ceased in the bath culture. Similarly, L-threonine production was also associated with cell growth in the carbon-, phosphate-, and sulfate-limited fed-batch cultures, but the accumulation of L-threonine was markedly increased because of the extended production time in the growth stage. Interestingly, auxotrophic amino acid (L-isoleucine)-limited feeding promoted L-threonine production over the non-growth phase. Metabolite analysis indicates that substantial production of acetate and glutamate and the resulting accumulation of ammonium may lead to the inhibition of L-threonine production. During the growth phase, the levels of L-isoleucine were accurately optimized by balancing cell growth and production with Pontryagin's maximum principle, basing on the relationship between the specific growth rate μ and specific production rate ρ. Furthermore, the depletion of L-isoleucine and phosphate at the end of the growth phase favored the synthesis of L-threonine in the subsequent non-growth phase. Combining the two-stage feeding profiles, the final L-threonine concentration and conversion rate were increased by 5.9- and 2.1-fold, respectively, compared to batch processes without feeding control. The identification of efficient feeding nutrient and the development of accurate feeding strategies provide potential guidelines for microbial production of amino acids.

  3. Development of Fed-Batch Cultivation Strategy for Efficient Oxytetracycline Production by Streptomyces rimosus at Semi-Industrial Scale

    Directory of Open Access Journals (Sweden)

    Elsayed Ahmed Elsayed


    Full Text Available ABSTRACTOxytetracycline (OTC production byStreptomyces rimosus was studied in batch and fed-batch cultures in shake flask and bioreactor levels using semi-defined medium. First, the effect of glucose concentration on OTC production and growth kinetics was studied intensively. The optimal glucose concentration in the medium was 15 g/L. Higher glucose concentrations supported higher biomass production by less volumetric and specific antibiotic production. Based on these data, cultivations were carried out at semi-industrial scale 15 L bioreactor in batch culture. At bioreactor level, cell growth and OTC production were higher compared to the shake flask culture by about 18 and 38%, respectively. During the bioreactor cultivation, glucose was totally consumed after only 48 h. Thus, the fed-batch experiment was designed for mono-glucose feeding and complete medium feeding to increase the OTC production by overcoming carbon limitations. The results showed that the fed-batch culture using constant glucose feeding strategy with rate of 0.33 g/L/h produced 1072 mg/L. On the other hand, feeding with complete medium resulted in 45% higher biomass but less OTC production by about 26% compared to mono-glucose fed culture. A further improvement in this process was achieved in by keeping the dissolved oxygen (DO value at 60% saturation by cascading the glucose feeding pump with the DO controller. The later feeding strategy resulted in higher antibiotic production, reaching 1414 mg/L after 108 h.

  4. Fed-batch culture of Escherichia coli for L-valine production based on in silico flux response analysis. (United States)

    Park, Jin Hwan; Kim, Tae Yong; Lee, Kwang Ho; Lee, Sang Yup


    We have previously reported the development of a 100% genetically defined engineered Escherichia coli strain capable of producing L-valine from glucose with a high yield of 0.38 g L-valine per gram glucose (0.58 mol L-valine per mol glucose) by batch culture. Here we report a systems biological strategy of employing flux response analysis in bioprocess development using L-valine production by fed-batch culture as an example. Through the systems-level analysis, the source of ATP was found to be important for efficient L-valine production. There existed a trade-off between L-valine production and biomass formation, which was optimized for the most efficient L-valine production. Furthermore, acetic acid feeding strategy was optimized based on flux response analysis. The final fed-batch cultivation strategy allowed production of 32.3 g/L L-valine, the highest concentration reported for E. coli. This approach of employing systems-level analysis of metabolic fluxes in developing fed-batch cultivation strategy would also be applicable in developing strategies for the efficient production of other bioproducts.

  5. Mixotrophic growth of Phaeodactylum tricornutum on fructose and glycerol in fed-batch and semi-continuous modes. (United States)

    Cerón-García, M C; Fernández-Sevilla, J M; Sánchez-Mirón, A; García-Camacho, F; Contreras-Gómez, A; Molina-Grima, E


    Mixotrophic cultures of Phaeodactylum tricornutum were carried out in bubble columns using fructose and glycerol in indoor fed-batch and semi-continuous modes. In the fed-batch cultures, different nutrient-addition strategies, combined with stepwise increments in the light intensity, were assayed. It was found that glycerol promoted significantly higher biomass productivity than fructose. A glycerol-induced photoinhibition that arrested the growth of P. tricornutun was also observed. As this was considered a limitation as regards transferring the fed-batch mode to outdoor conditions, this information was used to culture P. tricornutum in semi-continuous mode. Similar glycerol-induced photoinhibition was not observed in these cultures, even at highest dilution rates. Although the highest biomass (1.5 g L(-1) d(-1)) and EPA (40 mg L(-1) d(-1)) productivities found in the semi-continuous cultures were lower than those obtained photoautotrophically in outdoor photobioreactors, the findings showed that semi-continuous mode was an excellent candidate for transferring mixotrophic culture to an outdoor setting.

  6. The use of date waste for lactic acid production by a fed-batch culture using Lactobacillus casei subsp. rhamnosus

    Directory of Open Access Journals (Sweden)

    Aicha Nancib


    Full Text Available The production of lactic acid from date juice by Lactobacillus caseisubsp. rhamnosus in batch and fed-batch cultures has been investigated. The fed-batch culture system gave better results for lactic acid production and volumetric productivity. The aim of this work is to determine the effects of the feeding rate and the concentration of the feeding medium containing date juice glucose on the cell growth, the consumption of glucose and the lactic acid production by Lactobacillus casei subsp. rhamnosus in fed-batch cultures. For this study, two concentrations of the feeding medium (62 and 100 g/L of date juice glucose were tested at different feeding rates (18, 22, 33, 75 and 150 mL/h. The highest volumetric productivity (1.3 g/L.h and lactic acid yield (1.7 g/g were obtained at a feeding rate of 33 mL/h and a date juice glucose concentration of 62 g/L in the feeding medium. As a result, most of the date juice glucose was completely utilised (residual glucose 1 g/L, and a maximum lactic acid production level (89.2 g/L was obtained.

  7. The use of date waste for lactic acid production by a fed-batch culture using Lactobacillus casei subsp. rhamnosus. (United States)

    Nancib, Aicha; Nancib, Nabil; Boubendir, Abdelhafid; Boudrant, Joseph


    The production of lactic acid from date juice by Lactobacillus caseisubsp. rhamnosus in batch and fed-batch cultures has been investigated. The fed-batch culture system gave better results for lactic acid production and volumetric productivity. The aim of this work is to determine the effects of the feeding rate and the concentration of the feeding medium containing date juice glucose on the cell growth, the consumption of glucose and the lactic acid production by Lactobacillus casei subsp. rhamnosus in fed-batch cultures. For this study, two concentrations of the feeding medium (62 and 100 g/L of date juice glucose) were tested at different feeding rates (18, 22, 33, 75 and 150 mL/h). The highest volumetric productivity (1.3 g/L.h) and lactic acid yield (1.7 g/g) were obtained at a feeding rate of 33 mL/h and a date juice glucose concentration of 62 g/L in the feeding medium. As a result, most of the date juice glucose was completely utilised (residual glucose 1 g/L), and a maximum lactic acid production level (89.2 g/L) was obtained.

  8. A study on clavulanic acid production by Streptomyces clavuligerus in batch, fed-batch and continuous processes

    Directory of Open Access Journals (Sweden)

    A. B. Neto


    Full Text Available Clavulanic acid (CA is a potent inhibitor of beta-lactamases, enzymes that are responsible for the hydrolysis of beta-lactam antibiotics. It is a secondary metabolite produced by the filamentous aerobic bacterium Streptomyces clavuligerus in submerged cultivations. In the present work clavulanic acid production in batch, fed-batch and continuous bioreactors was studied with the objective of increasing productivity. The operating conditions: temperature, aeration and agitation, were the same in all cases, 28º C, 0.5 vvm and 800 rpm, respectively. The CA concentration obtained in the fed-batch culture, 404 mg L-1, was ca twice the value obtained in the batch culture, 194 mg L-1, while 293 mg L-1 was obtained in the continuous culture. The highest productivity was obtained in the continuous cultivation, 10.6 mg L-1 h-1, as compared with 8.8 mg L-1 h-1 in the fed-batch process and 3.5 mg L-1 h-1 in the batch process, suggesting that continuous culture of Streptomyces clavuligerus is a promising strategy for clavulanic acid production.

  9. Heterotrophic high cell-density fed-batch cultures of the phycocyanin-producing red alga Galdieria sulphuraria. (United States)

    Schmidt, Rikke Ankerstjerne; Wiebe, Marilyn G; Eriksen, Niels Thomas


    Growth and phycocyanin production in batch and fed-batch cultures of the microalga Galdieria sulphuraria 074G, which was grown heterotrophically in darkness on glucose, fructose, sucrose, and sugar beet molasses, was investigated. In batch cultures, specific growth rates and yields of biomass dry weight on the pure sugars were 1.08-1.15 day-1 and 0.48-0.50 g g-1, respectively. They were slightly higher when molasses was the carbon source. Cellular phycocyanin contents during the exponential growth phase were 3-4 mg g-1 in dry weight. G. sulphuraria was able to tolerate concentrations of glucose and fructose of up to 166 g L-1 (0.9 M) and an ammonium sulfate concentration of 22 g L-1 (0.17 M) without negative effects on the specific growth rate. When the total concentration of dissolved substances in the growth medium exceeded 1-2 M, growth was completely inhibited. In carbon-limited fed-batch cultures, biomass dry weight concentrations of 80-120 g L-1 were obtained while phycocyanin accumulated to concentrations between 250 and 400 mg L-1. These results demonstrate that G. sulphuraria is well suited for growth in heterotrophic cultures at very high cell densities, and that such cultures produce significant amounts of phycocyanin. Furthermore, the productivity of phycocyanin in the heterotrophic fed-batch cultures of G. sulphuraria was higher than is attained in outdoor cultures of Spirulina platensis, where phycocyanin is presently obtained.

  10. Model-based scale-up methodology for aerobic fed-batch bioprocesses: application to polyhydroxybutyrate (PHB) production. (United States)

    Monsalve-Bravo, Gloria Milena; Garelli, Fabricio; Mozumder, Md Salatul Islam; Alvarez, Hernan; De Battista, Hernan


    This work presents a general model-based methodology to scale-up fed-batch bioprocesses. The idea behind this approach is to establish a dynamics hierarchy, based on a model of the process, that allows the designer to determine the proper scale factors as well as at which point of the fed-batch the process should be scaled up. Here, concepts and tools of linear control theory, such as the singular value decomposition of the Hankel matrix, are exploited in the context of process design. The proposed scale-up methodology is first described in a bioprocesses general framework highlighting its main features, key variables and parameters. Then, it is applied to a polyhydroxybutyrate (PHB) fed-batch bioreactor and compared with three empirical criteria, that are traditionally employed to determine the scale factors of these processes, showing the usefulness and distinctive features of this proposal. Moreover, this methodology provides theoretical support to a frequently used empirical rule: scale-up aerobic bioreactors at constant volumetric oxygen transfer coefficient. Finally, similar process dynamic behavior and PHB production set at the laboratory scale are predicted at the new operating scale, while it is also determined that is rarely possible to reproduce similar dynamic behavior of the bioreactor using empirical scale-up criteria.

  11. Enhanced yield of medium-chain-length polyhydroxyalkanoates from nonanoic acid by co-feeding glucose in carbon-limited, fed-batch culture. (United States)

    Sun, Zhiyong; Ramsay, Juliana; Guay, Martin; Ramsay, Bruce


    Medium-chain-length polyhydroxyalkanoates (MCL-PHAs) were produced in carbon-limited, single-stage, fed-batch fermentations of Pseudomonas putida KT2440 by co-feeding nonanoic acid (NA) and glucose (G) to enhance the yield of PHA from NA. An exponential (mu=0.25 h(-1)) followed by a linear feeding strategy at a NA:G ratio of 1:1 (w/w) achieved 71 g l(-1) biomass containing 56% PHA. Although the same overall PHA productivity (1.44 g l(-1) h(-1)) was obtained when NA alone was fed at the same specific growth rate, the overall yield of PHA from NA increased by 25% (0.66 g PHA g NA(-1) versus 0.53 g g(-1)) with glucose co-feeding. Further increasing glucose in the feed (NA:G=1:1.5) resulted in a slightly higher yield (0.69 g PHA g NA(-1)) but lower PHA content (48%) and productivity (1.16 g l(-1) h(-1)). There was very little change in the PHA composition.

  12. Batch and fed-batch bioreactor studies for the enhanced production of glutaminase-free L-asparaginase from Pectobacterium carotovorum MTCC 1428. (United States)

    Kumar, Sanjay; Prabhu, Ashish A; Dasu, V Venkata; Pakshirajan, Kannan


    The effect of dissolved oxygen (DO) level and pH (controlled/uncontrolled) was first studied to enhance the production of novel glutaminase-free L-asparaginase by Pectobacterium carotovorum MTCC 1428 in a batch bioreactor. The optimum level of DO was found to be 20%. The production of L-asparaginase was found to be maximum when pH of the medium was maintained at 8.5 after 12 h of fermentation. Under these conditions, P. carotovorum produced 17.97 U/mL of L-asparaginase corresponding to the productivity of 1497.50 U/L/h. The production of L-asparaginase was studied in fed-batch bioreactor by feeding L-asparagine (essential substrate for production) and/or glucose (carbon source for growth) at the end of the reaction period of 12 h. The initial medium containing both L-asparagine and glucose in the batch mode and L-asparagine in the feeding stream was found to be the best combination for enhanced production of glutaminase-free L-asparaginase. Under this condition, the L-asparaginase production was increased to 38.8 U/mL, which corresponded to a productivity of 1615.8 U/L/h. The production and productivity were increased by 115.8% and 7.9%, respectively, both of which are higher than those obtained in the batch bioreactor experiments.

  13. In silico optimization and low structured kinetic model of poly[(R)-3-hydroxybutyrate] synthesis by Cupriavidus necator DSM 545 by fed-batch cultivation on glycerol. (United States)

    Spoljarić, Ivna Vrana; Lopar, Markan; Koller, Martin; Muhr, Alexander; Salerno, Anna; Reiterer, Angelika; Horvat, Predrag


    Glycerol was utilized by Cupriavidus necator DSM 545 for production of poly-3-hydroxybutyrate (PHB) in fed-batch fermentation. Maximal specific growth rates (0.12 and 0.3h(-1)) and maximal specific non-growth PHB production rate (0.16 g g(-1)h(-1)) were determined from two experiments (inocula from exponential and stationary phase). Saturation constants for nitrogen (0.107 and 0.016 g L(-1)), glycerol (0.05 g L(-1)), non-growth related PHB synthesis (0.011 g L(-1)) and nitrogen/PHB related inhibition constant (0.405 g L(-1)), were estimated. Five relations for specific growth rate were tested using mathematical models. In silico performed optimization procedures (varied glycerol/nitrogen ratio and feeding) has resulted in a PHB content of 70.9%, shorter cultivation time (23 h) and better PHB yield (0.347 g g(-1)). Initial concentration of biomass 16.8 g L(-1) and glycerol concentration in broth between 3 and 5 g L(-1) were decisive factors for increasing of productivity. Copyright © 2013 Elsevier B.V. All rights reserved.

  14. Physiological changes of Candida tropicalis population degrading phenol in fed batch reactor

    Directory of Open Access Journals (Sweden)

    Eliska Komarkova


    Full Text Available Candida tropicalis can use phenol as the sole carbon and energy source. Experiments regarding phenol degradations from the water phase were carried out. The fermentor was operated as a fed-batch system with oxistat control. Under conditions of nutrient limitation and an excess of oxygen the respiration activity of cells was suppressed and some color metabolites (black-brown started to be formed. An accumulation of these products inhibited the cell growth under aerobic conditions. Another impact was a decrease of the phenol hydroxylase activity as the key enzyme of the phenol degradation pathway at the end of the cell respiration activity. This decrease is linked with the above mentioned product inhibition. The cell death studied by fluorescent probe proceeded very slowly after the loss of the respiration activity. The starvation stress induced an increase of the endogenous respiration rate at the expense of phenol oxidation.Candida tropicalis pode utilizar fenol como única fonte de carbono e de energia. O fermentador foi operado em um sistema ''batelada-alimentada'' e controle oxidativo. Em condições limitantes de nutrientes e excesso de oxigênio a atividade respiratória das células foi suprimida e o calor do metabolismo pode ser formado. Uma acumulação desses produtos inibiu o crescimento das células em condições aeróbicas. Outro impacto foi um decréscimo da atividade fenol hidroxilase como enzima chave da degradação do fenol no final da atividade respirométrica. Essa redução está relacionada com os fatos acima mencionados. A morte da célula estudada por sonda de fluorescência ocorreu lentamente após a perda da atividade respiratória. O ''stress'' celular induziu um aumento na taxa de respiração endógena devido à oxidação fenólica.

  15. Production of ethyl alcohol by a repeated batch fermentation method. Kurikaeshi kaibun hakkoho ni yoru arukoru no seizoho

    Energy Technology Data Exchange (ETDEWEB)

    Saiki, T. (Tokyo (Japan)); Takagi, Y. (Chiba (Japan)); Shiba, M. (Kagoshima (Japan))


    This invention relates to a production method of ethyl alcohol from sugar and starch raw materials by a repeated batch fermentation method and aims to provide a method to produce ethyl alcohol efficiently by accurate repetition. Conventional batch fermentation methods require the experiences and skill to confirm an end point of the fermentation and have a problem in a control of repeated batch fermentation. This invention uses a gas flow meter of a velocity head measurement type and judges an end point of fermentation at a time when the amount of the exhaust gas flow from the fermentation is 0.35 to 0.1 m[sup 3] per hour and per 1 kl fermentation liquid or below and the next batch fermentation is started. By this invention, ethyl alcohol is produced efficiently from the raw materials with less damage of the yeast and efficient repeated batch fermentation is realized. 6 figs., 1 tab.

  16. Effect of Fed-Batch vs. Continuous Mode of Operation on Microbial Fuel Cell Performance Treating Biorefinery Wastewater

    Energy Technology Data Exchange (ETDEWEB)

    Pannell, Tyler C.; Goud, R. Kannaiah; Schell, Daniel J.; Borole, Abhijeet P.


    Bioelectrochemical systems have been shown to treat low-value biorefinery streams while recovering energy, however, low current densities and anode conversion efficiencies (ACE) limit their application. A bioanode was developed via enrichment of electroactive biofilm under fed-batch and continuous feeding conditions using corn stover-derived waste stream. The continuously-fed MFC exhibited a current density of 5.8 +/- 0.06 A/m2 and an ACE of 39% +/- 4. The fed-batch MFC achieved a similar current density and an ACE of 19.2%, however, its performance dropped after 36 days of operation to 1.1 A/m2 and 0.5%, respectively. In comparison, the ACE of the continuously-fed MFC remained stable achieving an ACE of 30% +/- 3 after 48 days of operation. An MFC treating a biorefinery stream post fuel separation achieved a current density of 10.7 +/- 0.1 A/m2 and an ACE of 57% +/- 9 at an organic loading of 12.5 g COD/L-day. Characterization of the microbial communities indicate higher abundance of Firmicutes and Proteobacteria and lower abundance of Bacteriodetes and a higher level of Geobacter spp. (1.4% vs. 0.2%) in continuously-fed MFC vs. fed-batch MFC. The results demonstrate that limiting substrate to the equivalent maximum current that the anode can generate, maintains MFC performance over a long term for high strength wastewaters, such as those generated in the biorefinery.

  17. Versatile modeling and optimization of fed batch processes for the production of secreted heterologous proteins with Pichia pastoris

    Directory of Open Access Journals (Sweden)

    Gasser Brigitte


    Full Text Available Abstract Background Secretion of heterologous proteins depends both on biomass concentration and on the specific product secretion rate, which in turn is not constant at varying specific growth rates. As fed batch processes usually do not maintain a steady state throughout the feed phase, it is not trivial to model and optimize such a process by mathematical means. Results We have developed a model for product accumulation in fed batch based on iterative calculation in Microsoft Excel spreadsheets, and used the Solver software to optimize the time course of the media feed in order to maximize the volumetric productivity. The optimum feed phase consisted of an exponential feed at maximum specific growth rate, followed by a phase with linearly increasing feed rate and consequently steadily decreasing specific growth rate. The latter phase could be modeled also by exact mathematical treatment by the calculus of variations, yielding the explicit shape of the growth function, however, with certain indeterminate parameters. To evaluate the latter, one needs a numerical optimum search algorithm. The explicit shape of the growth function provides additional evidence that the Excel model results in correct data. Experimental evaluation in two independent fed batch cultures resulted in a good correlation to the optimized model data, and a 2.2 fold improvement of the volumetric productivity. Conclusion The advantages of the procedure we describe here are the ease of use and the flexibility, applying software familiar to every scientist and engineer, and rapid calculation which makes predictions extremely easy, so that many options can be tested in silico quickly. Additional options like further biological and technological constraints or different functions for specific productivity and biomass yield can easily be integrated.

  18. Lipid production in batch and fed-batch cultures of Rhodosporidium toruloides from 5 and 6 carbon carbohydrates

    Directory of Open Access Journals (Sweden)

    Wiebe Marilyn G


    Full Text Available Abstract Background Microbial lipids are a potential source of bio- or renewable diesel and the red yeast Rhodosporidium toruloides is interesting not only because it can accumulate over 50% of its dry biomass as lipid, but also because it utilises both five and six carbon carbohydrates, which are present in plant biomass hydrolysates. Methods R. toruloides was grown in batch and fed-batch cultures in 0.5 L bioreactors at pH 4 in chemically defined, nitrogen restricted (C/N 40 to 100 media containing glucose, xylose, arabinose, or all three carbohydrates as carbon source. Lipid was extracted from the biomass using chloroform-methanol, measured gravimetrically and analysed by GC. Results Lipid production was most efficient with glucose (up to 25 g lipid L−1, 48 to 75% lipid in the biomass, at up to 0.21 g lipid L−1 h−1 as the sole carbon source, but high lipid concentrations were also produced from xylose (36 to 45% lipid in biomass. Lipid production was low (15–19% lipid in biomass with arabinose as sole carbon source and was lower than expected (30% lipid in biomass when glucose, xylose and arabinose were provided simultaneously. The presence of arabinose and/or xylose in the medium increased the proportion of palmitic and linoleic acid and reduced the proportion of oleic acid in the fatty acids, compared to glucose-grown cells. High cell densities were obtained in both batch (37 g L−1, with 49% lipid in the biomass and fed-batch (35 to 47 g L−1, with 50 to 75% lipid in the biomass cultures. The highest proportion of lipid in the biomass was observed in cultures given nitrogen during the batch phase but none with the feed. However, carbohydrate consumption was incomplete when the feed did not contain nitrogen and the highest total lipid and best substrate consumption were observed in cultures which received a constant low nitrogen supply. Conclusions Lipid production in R. toruloides was lower from arabinose and mixed

  19. Hydrogen production from formic acid in pH-stat fed-batch operation for direct supply to fuel cell. (United States)

    Shin, Jong-Hwan; Yoon, Jong Hyun; Lee, Seung Hoon; Park, Tai Hyun


    Enterobacter asburiae SNU-1 harvested after cultivation was used as a whole cell biocatalyst, for the production of hydrogen. Formic acid was efficiently converted to hydrogen using the harvested cells with an initial hydrogen production rate and total hydrogen production of 491 ml/l/h and 6668 ml/l, respectively, when 1 g/l of whole cell enzyme was used. Moreover, new pH-stat fed-batch operation was conducted, and total hydrogen production was 1.4 times higher than that of batch operation. For practical application, bio-hydrogen produced from formic acid using harvested cells was directly applied to PEMFC for power generation.

  20. Thermodynamics of metabolic pathways for penicillin production: Analysis of thermodynamic feasibility and free energy changes during fed-batch cultivation

    DEFF Research Database (Denmark)

    Pissarra, P.D.; Nielsen, Jens Bredal


    ) is an intermediate. It is found that the L-lysine pathway in P. chrysogenum is thermodynamically feasible and that the calculated standard Gibbs free energy values of the two enzymes controlling the pathway flux indicate that they operate far from equilibrium. It is therefore proposed that the regulation of alpha......-aminoadipate reductase by lysine is important to maintain a high concentration of alpha-aminoadipate in order to direct the carbon flux to penicillin production. Secondly the changes in Gibbs free energy in the penicillin biosynthetic pathway during fed-batch cultivation were studied. The analysis showed that all...

  1. Repeated batch fermentation from raw starch using a maltose transporter and amylase expressing diploid yeast strain. (United States)

    Yamakawa, Syun-ichi; Yamada, Ryosuke; Tanaka, Tsutomu; Ogino, Chiaki; Kondo, Akihiko


    We successfully demonstrated batch ethanol fermentation repeated ten times from raw starch with high ethanol productivity. We constructed a yeast diploid strain coexpressing the maltose transporter AGT1, alpha-amylase, and glucoamylase. The introduction of AGT1 allows maltose and maltotriose fermentation as well as the improvement of amylase activities. We also found that alpha-amylase activity during fermentation was retained by the addition of 10 mM calcium ion and that the highest alpha-amylase activity was 9.26 U/ml during repeated fermentation. The highest ethanol productivity was 2.22 g/l/h at the fourth batch, and after ten cycles, ethanol productivity of more than 1.43 g/l/h was retained, as was alpha-amylase activity at 6.43 U/ml.

  2. Electrochemical study of multi-electrode microbial fuel cells under fed-batch and continuous flow conditions

    KAUST Repository

    Ren, Lijiao


    Power production of four hydraulically connected microbial fuel cells (MFCs) was compared with the reactors operated using individual electrical circuits (individual), and when four anodes were wired together and connected to four cathodes all wired together (combined), in fed-batch or continuous flow conditions. Power production under these different conditions could not be made based on a single resistance, but instead required polarization tests to assess individual performance relative to the combined MFCs. Based on the power curves, power produced by the combined MFCs (2.12 ± 0.03 mW, 200 ω) was the same as the summed power (2.13 mW, 50 ω) produced by the four individual reactors in fed-batch mode. With continuous flow through the four MFCs, the maximum power (0.59 ± 0.01 mW) produced by the combined MFCs was slightly lower than the summed maximum power of the four individual reactors (0.68 ± 0.02 mW). There was a small parasitic current flow from adjacent anodes and cathodes, but overall performance was relatively unaffected. These findings demonstrate that optimal power production by reactors hydraulically and electrically connected can be predicted from performance by individual reactors. © 2013 Elsevier B.V. All rights reserved.

  3. Kinetic and stoichiometric characterization of organoautotrophic growth of Ralstonia eutropha on formic acid in fed-batch and continuous cultures. (United States)

    Grunwald, Stephan; Mottet, Alexis; Grousseau, Estelle; Plassmeier, Jens K; Popović, Milan K; Uribelarrea, Jean-Louis; Gorret, Nathalie; Guillouet, Stéphane E; Sinskey, Anthony


    Formic acid, acting as both carbon and energy source, is a safe alternative to a carbon dioxide, hydrogen and dioxygen mix for studying the conversion of carbon through the Calvin-Benson-Bassham (CBB) cycle into value-added chemical compounds by non-photosynthetic microorganisms. In this work, organoautotrophic growth of Ralstonia eutropha on formic acid was studied using an approach combining stoichiometric modeling and controlled cultures in bioreactors. A strain deleted of its polyhydroxyalkanoate production pathway was used in order to carry out a physiological characterization. The maximal growth yield was determined at 0.16 Cmole Cmole(-1) in a formate-limited continuous culture. The measured yield corresponded to 76% to 85% of the theoretical yield (later confirmed in pH-controlled fed-batch cultures). The stoichiometric study highlighted the imbalance between carbon and energy provided by formic acid and explained the low growth yields measured. Fed-batch cultures were also used to determine the maximum specific growth rate (μmax  = 0.18 h(-1) ) and to study the impact of increasing formic acid concentrations on growth yields. High formic acid sensitivity was found in R eutropha since a linear decrease in the biomass yield with increasing residual formic acid concentrations was observed between 0 and 1.5 g l(-1) .

  4. Screening and assessment of performance and molecule quality attributes of industrial cell lines across different fed-batch systems. (United States)

    Rouiller, Yolande; Bielser, Jean-Marc; Brühlmann, David; Jordan, Martin; Broly, Hervé; Stettler, Matthieu


    The major challenge in the selection process of recombinant cell lines for the production of biologics is the choice, early in development, of a clonal cell line presenting a high productivity and optimal cell growth. Most importantly, the selected candidate needs to generate a product quality profile which is adequate with respect to safety and efficacy and which is preserved across cell culture scales. We developed a high-throughput screening and selection strategy of recombinant cell lines, based on their productivity in shaking 96-deepwell plates operated in fed-batch mode, which enables the identification of cell lines maintaining their high productivity at larger scales. Twelve recombinant cell lines expressing the same antibody with different productivities were selected out of 470 clonal cell lines in 96-deepwell plate fed-batch culture. They were tested under the same conditions in 50 mL vented shake tubes, microscale and lab-scale bioreactors in order to confirm the maintenance of their performance at larger scales. The use of a feeding protocol and culture conditions which are essentially the same across the different scales was essential to maintain productivity and product quality profiles across scales. Compared to currently used approaches, this strategy has the advantage of speeding up the selection process and increases the number of screened clones for getting high-producing recombinant cell lines at manufacturing scale with the desired performance and quality.

  5. Fed-batch strategy for enhancing cell growth and C-phycocyanin production of Arthrospira (Spirulina) platensis under phototrophic cultivation. (United States)

    Xie, Youping; Jin, Yiwen; Zeng, Xianhai; Chen, Jianfeng; Lu, Yinghua; Jing, Keju


    The C-phycocyanin generated in blue-green algae Arthrospira platensis is gaining commercial interest due to its nutrition and healthcare value. In this study, the light intensity and initial biomass concentration were manipulated to improve cell growth and C-phycocyanin production of A.platensis in batch cultivation. The results show that low light intensity and high initial biomass concentration led to increased C-phycocyanin accumulation. The best C-phycocyanin productivity occurred when light intensity and initial biomass concentration were 300μmol/m(2)/s and 0.24g/L, respectively. The fed-batch cultivation proved to be an effective strategy to further enhance C-phycocyanin production of A.platensis. The results indicate that C-phycocyanin accumulation not only requires nitrogen-sufficient condition, but also needs other nutrients. The highest C-phycocyanin content (16.1%), production (1034mg/L) and productivity (94.8mg/L/d) were obtained when using fed-batch strategy with 5mM medium feeding. Copyright © 2014 Elsevier Ltd. All rights reserved.

  6. Fed-Batch Enzymatic Saccharification of High Solids Pretreated Lignocellulose for Obtaining High Titers and High Yields of Glucose. (United States)

    Jung, Young Hoon; Park, Hyun Min; Kim, Dong Hyun; Yang, Jungwoo; Kim, Kyoung Heon


    To reduce the distillation costs of cellulosic ethanol, it is necessary to produce high sugar titers in the enzymatic saccharification step. To obtain high sugar titers, high biomass loadings of lignocellulose are necessary. In this study, to overcome the low saccharification yields and the low operability of high biomass loadings, a fed-batch saccharification process was developed using an enzyme reactor that was designed and built in-house. After optimizing the cellulase and biomass feeding profiles and the agitation speed, 132.6 g/L glucose and 76.0% theoretical maximum glucose were obtained from the 60 h saccharification of maleic acid-pretreated rice straw at a 30% (w/v) solids loading with 15 filter paper units (FPU) of Cellic CTec2/g glucan. This study demonstrated that through the proper optimization of fed-batch saccharification, both high sugar titers and high saccharification yields are possible, even with using the high solids loading (i.e., ≥30%) with the moderate enzyme loading (i.e., high solids saccharification process in cellulosic fuel and chemical production.

  7. Influence of feeding conditions on clavulanic acid production in fed-batch cultivation with medium containing glycerol. (United States)

    Teodoro, Juliana C; Baptista-Neto, Alvaro; Cruz-Hernández, Isara L; Hokka, Carlos O; Badino, Alberto C


    First, the effect of different levels of nitrogen source on clavulanic acid (CA) production was evaluated in batch cultivations utilizing complex culture medium containing glycerol and three different levels of soy protein isolate (SPI). Cellular growth, evaluated in terms of the rheological parameter K, was highest with a SPI concentration of 30 g.L(-1) (4.42 g.L(-1) N total). However, the highest production of CA (380 mg.L(-1)) was obtained when an intermediate concentration of 20 g.L(-1) of SPI (2.95 g.L(-1) total N) was used. To address this, the influences of volumetric flow rate (F) and glycerol concentration in the complex feed medium (Cs(F)) in fed-batch cultivations were investigated. The best experimental condition for CA production was F=0.01 L.h(-1) and Cs(F)=120 g.L(-1), and under these conditions maximum CA production was practically twice that obtained in the batch cultivation. A single empirical equation was proposed to relate maximum CA production with F and Cs(F) in fed-batch experiments.

  8. Fed-batch production of hydrophobin RodB from Aspergillus fumigatus in host Pichia pastoris

    DEFF Research Database (Denmark)

    Pedersen, Mona Højgaard; Borodina, Irina; Frisvad, Jens Christian

    . The emulsifying property of rRodB was investigated using olive oil stained with Sudan black suspended in tris-buffer. The stability of oil micelles were studied by light microscopy. Results: Protein bands of expected size were detected by SDS-PAGE and western blotting in both the fermentation broth and excess...

  9. 玉米秸秆分批补料获得高还原糖浓度酶解液的条件优化%Optimization of corn stover hydrolysis by fed-batch process

    Institute of Scientific and Technical Information of China (English)

    宋安东; 任天宝; 张玲玲; 王风芹; 谢慧


    High-concentration sugars production from stover is an important perspective technology for the cellulosic ethanol industrialization.Fed-batch process is an effective way to achieve this goal in the fermentation industry.In this study, based on fed-batch process, high-concentration sugars were produced from pretreated corn stover by enzymatic hydrolysis.After being pretreated by the dilute sulphuric acid, the impacts of the ratio of solid raw material to liquid culture, the content of supplementary materials and the refilling time on the saccharification rate were investigated.Results showed that the initial ratio of solid raw material to liquid culture was 20% (W/V) and the initial concentrations of enzymes for xylanase, cellulose and pectinase were 220 U,6 FPU, and 50 U per gram of substrates, respectively.After 24 hours and 48 hours, 8% pretreated corn stovers were added respectively together with the additions of xylanase (20 U) and cellulose (2 FPU) per gram of substrates.After 72 hours, the final concentration of reducing sugar was increased to 138.5 g/L from 48.5 g/L of the non fed-batch process.The rate of enzyme hydrolysis of the raw material was 62.5% of the thoretieal value in the fed-batch process.This study demonstrated that the fed-batch process could significantly improve the concentration of reducing sugar.%木质纤维素高浓度还原糖水解液的获得是纤维乙醇产业化发展的方向.在发酵工业领域,分批补料法是实现这一目标的重要研究途径.本研究采用分批补料法对获得高浓度玉米秸秆酶解还原糖的条件进行了优化.以稀硫酸预处理的玉米秸秆为原料,考察了液固比、补加量与补加时间对分批补料糖化的影响.结果表明,秸秆高浓度酶解液条件的初始物料为20%(重量/体积),木聚糖酶220 U/g(底物),纤维素酶6 FPU/g(底物),果胶酶50 U/g(底物),在24 h、48 h后分批补加8%预处理后的物料,同时添加与

  10. Biosorption of copper(II) ions onto powdered waste sludge in a completely mixed fed-batch reactor: estimation of design parameters. (United States)

    Pamukoglu, Yunus; Kargi, Fikret


    Biosorption of Cu(II) ions onto pre-treated powdered waste sludge (PWS) was investigated using a fed-batch operated completely mixed reactor. Fed-batch adsorption experiments were performed by varying the feed flow rate ( 0.075-0.325 l h(-1)), feed copper (II) ion concentrations (50-300 mg l(-1)) and the amount of adsorbent (1-6 g PWS) using fed-batch operation. Breakthrough curves describing the variations of effluent copper ion concentrations with time were determined for different operating conditions. Percent copper ion removals from the aqueous phase decreased, but the biosorbed (solid phase) copper ion concentrations increased with increasing the feed flow rate and Cu(II) concentration. A modified Bohart-Adams equation was used to determine the biosorption capacity of PWS and the rate constant for Cu(II) ion biosorption. Adsorption rate constant in fed-batch operation was an order of magnitude larger than those obtained in adsorption columns because of elimination of mass transfer limitations encountered in the column operations while the biosorption capacity of PWS was comparable with powdered activated (PAC) in column operations. Therefore, a completely mixed reactor operated in fed-batch mode was proven to be more advantageous as compared to adsorption columns due to better contact between the phases yielding faster adsorption rates.

  11. Transcriptional response of P. pastoris in fed-batch cultivations to Rhizopus oryzae lipase production reveals UPR induction

    Directory of Open Access Journals (Sweden)

    Valero Francisco


    Full Text Available Abstract Background The analysis of transcriptional levels of the genes involved in protein synthesis and secretion is a key factor to understand the host organism's responses to recombinant protein production, as well as their interaction with the cultivation conditions. Novel techniques such as the sandwich hybridization allow monitoring quantitatively the dynamic changes of specific RNAs. In this study, the transcriptional levels of some genes related to the unfolded protein response (UPR and central metabolism of Pichia pastoris were analysed during batch and fed-batch cultivations using an X-33-derived strain expressing a Rhizopus oryzae lipase under control of the formaldehyde dehydrogenase promoter (FLD1, namely the alcohol oxidase gene AOX1, the formaldehyde dehydrogenase FLD1, the protein disulfide isomerase PDI, the KAR2 gene coding for the BiP chaperone, the 26S rRNA and the R. oryzae lipase gene ROL. Results The transcriptional levels of the selected set of genes were first analysed in P. pastoris cells growing in shake flask cultures containing different carbon and nitrogen sources combinations, glycerol + ammonium, methanol + methylamine and sorbitol + methylamine. The transcriptional levels of the AOX1 and FLD1 genes were coherent with the known regulatory mechanism of C1 substrates in P. pastoris, whereas ROL induction lead to the up-regulation of KAR2 and PDI transcriptional levels, thus suggesting that ROL overexpression triggers the UPR. This was further confirmed in fed-batch cultivations performed at different growth rates. Transcriptional levels of the analysed set of genes were generally higher at higher growth rates. Nevertheless, when ROL was overexpressed in a strain having the UPR constitutively activated, significantly lower relative induction levels of these marker genes were detected. Conclusion The bead-based sandwich hybridization assay has shown its potential as a reliable instrument for quantification of

  12. Multi-objective optimization of glycopeptide antibiotic production in batch and fed batch processes

    DEFF Research Database (Denmark)

    Maiti, Soumen K.; Eliasson Lantz, Anna; Bhushan, Mani


    as pareto optimal solutions. These solutions gives flexibility in evaluating the trade-offs and selecting the most suitable operating policy. Here, ε-constraint approach was used to generate the pareto solutions for two objectives: product concentration and product per unit cost of media, for batch and fed......Fermentation optimization involves potentially conflicting multiple objectives such as product concentration and production media cost. Simultaneous optimization of these objectives would result in a multiobjective optimization problem, which is characterized by a set of multiple solutions, knows...

  13. Enhanced poly(γ-glutamic acid) fermentation by Bacillus subtilis NX-2 immobilized in an aerobic plant fibrous-bed bioreactor. (United States)

    Xu, Zongqi; Feng, Xiaohai; Zhang, Dan; Tang, Bao; Lei, Peng; Liang, Jinfeng; Xu, Hong


    To enhance poly(γ-glutamic acid) (PGA) production, a novel aerobic plant fibrous-bed bioreactor (APFB) was constructed for immobilized fermentation. Based on the analysis of the kinetics of immobilized-cell fermentation using the APFB and conventional free-cell fermentation, immobilized-cell fermentation exhibited more efficient PGA production. Furthermore, repeated fed-batch cultures for PGA production were conducted to evaluate the stability of the APFB system. Average final PGA concentration and productivity of 71.21±0.83g/L and 1.246±0.008g/L/h were respectively achieved by cells immobilized in bagasse during APFB, which was reused eight times over a period of 457±18h. Analysis of the membrane phospholipids and the key enzyme activities indicated that APFB-adapted cells had better productivity than original cells. Thus, this study demonstrated the significant potential of the APFB culture system in future industrial applications.

  14. Glycoprofiling effects of media additives on IgG produced by CHO cells in fed-batch bioreactors

    DEFF Research Database (Denmark)

    Kildegaard, Helene Faustrup; Fan, Yuzhou; Wagtberg Sen, Jette


    . In this study, the effect on IgG N-glycosylation from feeding CHO cells with eight glycosylation precursors during cultivation was investigated. The study was conducted in fed-batch mode in bioreactors with biological replicates to obtain highly controlled and comparable conditions. We assessed charge......Therapeutic monoclonal antibodies (mAbs) are mainly produced by heterogonous expression in Chinese hamster ovary (CHO) cells. The glycosylation profile of the mAbs has major impact on the efficacy and safety of the drug and is therefore an important parameter to control during production...... heterogeneity and glycosylation patterns of IgG. None of the eight feed additives caused statistically significant changes to cell growth or IgG productivity, compared to controls. However, the addition of 20 mM galactose did result in a reproducible increase of galactosylated IgG from 14% to 25%. On the other...

  15. Modeling of growth and sporulation of Bacillus thuringiensis in an intermittent fed batch culture with total cell retention. (United States)

    Atehortúa, Paula; Alvarez, Hernán; Orduz, Sergio


    An extended dynamical model for growth and sporulation of Bacillus thuringiensis subsp. kurstaki in an intermittent fed-batch culture with total cell retention is proposed. This model differs from reported models, by including dynamics for natural death of cells and substrate consumption for cell maintenance. The proposed model uses sigmoid functions to describe these kinetic parameters. Equations for time evolution of substrate, vegetative, sporulated and total cell concentration were taken from previous works. Model parameters were determined from batch experimental data obtained in pilot plant. Parameter identification was developed in two stages: (1) coarse identification using a multivariable optimization with constraints algorithm, (2) fine identification by heuristic fit of model parameters looking for a minimal model error. The proposed model estimates adequate time evolution of the process variables with a mean error of 2.6% on substrate concentration and 6.7% on biomass concentration.

  16. Fingerprint detection and process prediction by multivariate analysis of fed-batch monoclonal antibody cell culture data. (United States)

    Sokolov, Michael; Soos, Miroslav; Neunstoecklin, Benjamin; Morbidelli, Massimo; Butté, Alessandro; Leardi, Riccardo; Solacroup, Thomas; Stettler, Matthieu; Broly, Hervé


    This work presents a sequential data analysis path, which was successfully applied to identify important patterns (fingerprints) in mammalian cell culture process data regarding process variables, time evolution and process response. The data set incorporates 116 fed-batch cultivation experiments for the production of a Fc-Fusion protein. Having precharacterized the evolutions of the investigated variables and manipulated parameters with univariate analysis, principal component analysis (PCA) and partial least squares regression (PLSR) are used for further investigation. The first major objective is to capture and understand the interaction structure and dynamic behavior of the process variables and the titer (process response) using different models. The second major objective is to evaluate those models regarding their capability to characterize and predict the titer production. Moreover, the effects of data unfolding, imputation of missing data, phase separation, and variable transformation on the performance of the models are evaluated.

  17. Expression and purification of recombinant human granulocyte colony-stimulating factor in fed-batch culture of Escherichia coli. (United States)

    Kim, Chang-Kyu; Choi, Jun-Ha; Lee, Seung-Bae; Lee, Sang-Mahn; Oh, Jae-Wook


    Granulocyte colony-stimulating factor (G-CSF) is a cytokine that has multiple roles in hematopoietic cells such as the regulation of proliferation and differentiation. Here, we describe fed-batch culture, refolding, and purification of rhG-CSF. The suitability of urea or sarcosine for solubilizing inclusion bodies (IBs) was tested. It was observed that urea is more efficient for solubilizing and refolding IBs than sarcosine is. The purity of rhG-CSF and the removal percentage of the rhG-CSF isoforms during purification were increased by pH 5.5 precipitation. The purity and the yield of purified rhG-CSF were 99% and 0.5 g of protein per liter culture broth, respectively. Our protocols of recombinant protein purification using ion exchange chromatography and semipreparative high performance liquid chromatography of pH-precipitated refolded solution may be informative to the industrial scale production of biopharmaceuticals.

  18. The use of dissolved oxygen-controlled, fed-batch aerobic cultivation for recombinant protein subunit vaccine manufacturing. (United States)

    Farrell, Patrick; Sun, Jacob; Champagne, Paul-Philippe; Lau, Heron; Gao, Meg; Sun, Hong; Zeiser, Arno; D'Amore, Tony


    A simple "off-the-shelf" fed-batch approach to aerobic bacterial cultivation for recombinant protein subunit vaccine manufacturing is presented. In this approach, changes in the dissolved oxygen levels are used to adjust the nutrient feed rate (DO-stat), so that the desired dissolved oxygen level is maintained throughout cultivation. This enables high Escherichia coli cell densities and recombinant protein titers. When coupled to a kLa-matched scale-down model, process performance is shown to be consistent at the 2L, 20L, and 200L scales for two recombinant E. coli strains expressing different protein subunit vaccine candidates. Additionally, by mining historical DO-stat nutrient feeding data, a method to transition from DO-stat to a pre-determined feeding profile suitable for larger manufacturing scales without using feedback control is demonstrated at the 2L, 20L, and 200L scales.

  19. Polyhydroxybutyrate production by direct use of waste activated sludge in phosphorus-limited fed-batch culture. (United States)

    Cavaillé, Laëtitia; Grousseau, Estelle; Pocquet, Mathieu; Lepeuple, Anne-Sophie; Uribelarrea, Jean-Louis; Hernandez-Raquet, Guillermina; Paul, Etienne


    Polyhydroxybutyrate (PHB) production directly by waste activated sludge (WAS) was investigated in aerobic fed-batch conditions using acetic acid as substrate. PHB production was induced by phosphorus limitation. WAS of different origin were tested with various degrees of phosphorus limitation and PHB contents of up to 70% (gCOD PHB/gCOD particulate) were obtained. This strategy showed the importance of maintaining cell growth for PHB production in order to increase PHB concentration and that the degree of phosphorus limitation has a direct impact on the quantity of PHB produced. Pyrosequencing of 16S rRNA transcripts showed changes in the active bacteria of the WAS microbial community as well as the acclimation of populations depending on sludge origin. The monitoring of the process appeared as the key factor for optimal PHB production by WAS. Different strategies are discussed and compared in terms of carbon yield and PHB content with the feast and famine selection process.

  20. Fed-batch cultivation of Arthrospira and Chlorella in ammonia-rich wastewater: Optimization of nutrient removal and biomass production. (United States)

    Markou, Giorgos


    In the present work the cyanobacterium Arthrospira platensis and the microalga Chlorella vulgaris were fed-batch cultivated in ammonia-rich wastewater derived from the anaerobic digestion of poultry litter. Aim of the study was to maximize the biomass production along with the nutrient removal aiming to wastewater treatment. Ammonia and phosphorus removals were very high (>95%) for all cultures investigated. Both microorganisms were able to remove volatile fatty acids to an extent of >90%, indicating that they were capable of mixotrophic growth. Chemical oxygen demand and proteins were also removed in various degrees. In contrast, in all cultures carbohydrate concentration was increased. The biochemical composition of the microorganisms varied greatly and was influenced by the indicate that the nutrient availability. A. platensis accumulated carbohydrates (≈ 40%), while C. vulgaris accumulated lipids (≈ 50%), rendering them interesting for biofuel production.

  1. Optimal fed batch experiment design for estimation of monod kinetics of Azospirillum brasilense: from theory to practice. (United States)

    Cappuyns, Astrid M; Bernaerts, Kristel; Smets, Ilse Y; Ona, Ositadinma; Prinsen, Els; Vanderleyden, Jos; Van Impe, Jan F


    In this paper the problem of reliable and accurate parameter estimation for unstructured models is considered. It is illustrated how a theoretically optimal design can be successfully translated into a practically feasible, robust, and informative experiment. The well-known parameter estimation problem of Monod kinetic parameters is used as a vehicle to illustrate our approach. As known for a long time, noisy batch measurements do not allow for unique and accurate estimation of the kinetic parameters of the Monod model. Techniques of optimal experiment design are, therefore, exploited to design informative experiments and to improve the parameter estimation accuracy. During the design process, practical feasibility has to be kept in mind. The designed experiments are easy to implement in practice and do not require additional monitoring equipment. Both design and experimental validation of informative fed batch experiments are illustrated with a case study, namely, the growth of the nitrogen-fixing bacteria Azospirillum brasilense.

  2. Optimization of the Production of Polygalacturonase from Aspergillus kawachii Cloned in Saccharomyces cerevisiae in Batch and Fed-Batch Cultures

    Directory of Open Access Journals (Sweden)

    Diego Jorge Baruque


    Full Text Available Polygalacturonases (PG; EC catalyze the hydrolysis of pectin and/or pectic acid and are useful for industrial applications such as juice clarification and pectin extraction. Growth and heterologous expression of recombinant Saccharomyces cerevisiae which expresses an acidic PG from Aspergillus kawachii has been studied in batch and fed-batch cultures. Kinetics and stoichiometric parameters of the recombinant yeast were determined in batch cultures in a synthetic medium. In these cultures, the total biomass concentration, protein concentration, and enzyme activity achieved were 2.2 g/L, 10 mg/L, and 3 U/mL, respectively, to give a productivity of 0.06 U/(mL·h. In fed-batch cultures, various strategies for galactose feeding were used: (i after a glucose growth phase, the addition of a single pulse of galactose which gave a productivity of 0.19 U/(mL·h; (ii after a glucose growth phase, a double pulse of galactose at the same final concentration was added, resulting in a productivity of 0.21 U/(mL·h; (iii a simultaneous feeding of glucose and galactose, yielding a productivity of 1.32 U/(mL·h. Based on these results, the simultaneous feeding of glucose and galactose was by far the most suitable strategy for the production of this enzyme. Moreover, some biochemical characteristics of the recombinant enzyme such as a molecular mass of ~60 kDa, an isoelectric point of 3.7 and its ability to hydrolyze polygalacturonic acid at pH=2.5 were determined.

  3. Differential Expression Of Small Rnas Under Chemical Stress And Fed-batch Fermentation In Escherichia Coli

    DEFF Research Database (Denmark)

    Rau, Martin Holm; Bojanovic, Klara; Nielsen, Alex Toftgaard;


    Introduction: Bacterial small RNAs (sRNAs) are often expressed in response to changing environmental conditions and function to modulate gene expression. Although chemical stress is routinely encountered in microbial processing applications, the cellular response and the involvement of sRNAs in t...

  4. The morphology of Ganoderma lucidum mycelium in a repeated-batch fermentation for exopolysaccharide production

    Directory of Open Access Journals (Sweden)

    Wan Abd Al Qadr Imad Wan-Mohtar


    Full Text Available The morphology of Ganoderma lucidum BCCM 31549 mycelium in a repeated-batch fermentation (RBF was studied for exopolysaccharide (EPS production. RBF was optimised for time to replace and volume to replace. G. lucidum mycelium showed the ability to self-immobilise and exhibited high stability for repeated use in RBF with engulfed pellets. Furthermore, the ovoid and starburst-like pellet morphology was disposed to EPS production in the shake flask and bioreactor, respectively. Seven RBF could be carried out in 500 mL flasks, and five repeated batches were performed in a 2 L bioreactor. Under RBF conditions, autolysis of pellet core in the shake flask and shaving off of the outer hairy region in the bioreactor were observed at the later stages of RBF (R4 for the shake flask and R6 for the bioreactor. The proposed strategy showed that the morphology of G. lucidum mycelium can withstand extended fermentation cycles.

  5. Repeated fermentation from raw starch using Saccharomyces cerevisiae displaying both glucoamylase and α-amylase. (United States)

    Yamakawa, Syun-ichi; Yamada, Ryosuke; Tanaka, Tsutomu; Ogino, Chiaki; Kondo, Akihiko


    A diploid yeast strain displaying both α-amylase and glucoamylase was developed for repeated fermentation from raw starch. First, the construct of α-amylase was optimized for cell surface display, as there have been no reports of α-amylase-displaying yeast. The modified yeast displaying both glucoamylase and α-amylase produced 46.5 g/l of ethanol from 200 g/l of raw corn starch after 120 h of fermentation, and this was 1.5-fold higher when compared to native α-amylase-displaying yeast. Using the glucoamylase and modified α-amylase co-displaying diploid strain, we repeated fermentation from 100g/l of raw starch for 23 cycles without the loss of α-amylase or glucoamylase activity. The average ethanol productivity and yield during repeated fermentation were 1.61 g/l/h and 76.6% of the theoretical yield, respectively. This novel yeast may be useful for reducing the cost of bio-ethanol production and may be suitable for industrial-scale bio-ethanol production.

  6. Dynamic lipidomic insights into the adaptive responses of Saccharomyces cerevisiae to the repeated vacuum fermentation. (United States)

    Zhou, Xiao; Zhou, Jian; Tian, Hongchi; Yuan, Yingjin


    Vacuum fermentation is utilized in a wide range of life science industries and biomedical R&D. Little is known, however, on the effects of the vacuum on the yeast, and in particular, on the yeast lipidome that plays a central role in maintaining cell membrane and other vital (yeast) cell functions. The present study evaluated the adaptive responses of Saccharomyces cerevisiae to repeated vacuum fermentation by lipidomic analysis. We employed gas chromatography coupled to time-of-flight mass spectrometry (GC-TOF-MS) and liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI/MS(n)) to quantify a total of 13 intermediate sterols and 139 phospholipid species of yeast cells. Principal components analysis found that the PI (phosphatidylinositol) 26:0, PI 28:0, PE (phosphatidylethanolamine) 32:1, and PE 34:1 were potential biomarkers to distinguish the vacuum fermentation process. Quantitative analysis showed that vacuum fermentation increased the synthesis of PI and the PC (phosphatidylcholine) species with short saturated acyl chains. The synthesis of PC via CDP-choline and turnover of PC were enhanced, instead of formation via methylation of PE. Additionally, increased PI at the expense of PE and PG (phosphatidylglycerol) was associated with enhancement of ethanol productivity. Vacuum fermentation caused eburicol accumulation, suggesting that vacuum can activate the branch of the ergosterol biosynthesis pathway. Eburicol decrease and PI increase contributed to recovery of cellular activities with oxygenating treatment. Ethanol productivity was increased by sixfold in vacuum-treated cells. These observations may allow the development of future mechanistic approaches to optimization of yeast fermentation under vacuum for bioindustry and life science applications. In particular, our findings on changes in lipid molecular species and the ergosterol biosynthesis pathway elucidate the defense responses of yeast cell membranes during the repeated

  7. Biorefinery development through utilization of biodiesel industry by-products as sole fermentation feedstock for 1,3-propanediol production. (United States)

    Chatzifragkou, Afroditi; Papanikolaou, Seraphim; Kopsahelis, Nikolaos; Kachrimanidou, Vasiliki; Dorado, Maria Pilar; Koutinas, Apostolis A


    Rapeseed meal (RSM) hydrolysate was evaluated as substitute for commercial nutrient supplements in 1,3-propanediol (PDO) fermentation using the strain Clostridium butyricum VPI 1718. RSM was enzymatically converted into a generic fermentation feedstock, enriched in amino acids, peptides and various micro-nutrients, using crude enzyme consortia produced via solid state fermentation by a fungal strain of Aspergillus oryzae. Initial free amino nitrogen concentration influenced PDO production in batch cultures. RSM hydrolysates were compared with commercial nutrient supplements regarding PDO production in fed-batch cultures carried out in a bench-scale bioreactor. The utilization of RSM hydrolysates in repeated batch cultivation resulted in a PDO concentration of 65.5 g/L with an overall productivity of 1.15 g/L/h that was almost 2 times higher than the productivity achieved when yeast extract was used as nutrient supplement.

  8. Expression of recombinant Pseudomonas stutzeri di-heme cytochrome c(4) by high-cell-density fed-batch cultivation of Pseudomonas putida

    DEFF Research Database (Denmark)

    Thuesen, Marianne Hallberg; Nørgaard, Allan; Hansen, Anne Merete


    The gene of the di-heme protein cytochrome c(4) from Pseudomonas stutzeri was expressed in Pseudomonas putida. High-yield expression of the protein was achieved by high-cell-density fed-batch cultivation using an exponential glucose feeding strategy. The recombinant cytochrome c(4) protein...

  9. A Single Dynamic Metabolic Model Can Describe mAb Producing CHO Cell Batch and Fed-Batch Cultures on Different Culture Media. (United States)

    Robitaille, Julien; Chen, Jingkui; Jolicoeur, Mario


    CHO cell culture high productivity relies on optimized culture medium management under fed-batch or perfused chemostat strategies enabling high cell densities. In this work, a dynamic metabolic model for CHO cells was further developed, calibrated and challenged using datasets obtained under four different culture conditions, including two batch and two fed-batch cultures comparing two different culture media. The recombinant CHO-DXB11 cell line producing the EG2-hFc monoclonal antibody was studied. Quantification of extracellular substrates and metabolites concentration, viable cell density, monoclonal antibody concentration and intracellular concentration of metabolite intermediates of glycolysis, pentose-phosphate and TCA cycle, as well as of energetic nucleotides, were obtained for model calibration. Results suggest that a single model structure with a single set of kinetic parameter values is efficient at simulating viable cell behavior in all cases under study, estimating the time course of measured and non-measured intracellular and extracellular metabolites. Model simulations also allowed performing dynamic metabolic flux analysis, showing that the culture media and the fed-batch strategies tested had little impact on flux distribution. This work thus paves the way to an in silico platform allowing to assess the performance of different culture media and fed-batch strategies.

  10. miRNA profiling of high, low and non-producing CHO cells during biphasic fed-batch cultivation reveals process relevant targets for host cell engineering. (United States)

    Stiefel, Fabian; Fischer, Simon; Sczyrba, Alexander; Otte, Kerstin; Hesse, Friedemann


    Fed-batch cultivation of recombinant Chinese hamster ovary (CHO) cell lines is one of the most widely used production modes for commercial manufacturing of recombinant protein therapeutics. Furthermore, fed-batch cultivations are often conducted as biphasic processes where the culture temperature is decreased to maximize volumetric product yields. However, it remains to be elucidated which intracellular regulatory elements actually control the observed pro-productive phenotypes. Recently, several studies have revealed microRNAs (miRNAs) to be important molecular switches of cell phenotypes. In this study, we analyzed miRNA profiles of two different recombinant CHO cell lines (high and low producer), and compared them to a non-producing CHO DG44 host cell line during fed-batch cultivation at 37°C versus a temperature shift to 30°C. Taking advantage of next-generation sequencing combined with cluster, correlation and differential expression analyses, we could identify 89 different miRNAs, which were differentially expressed in the different cell lines and cultivation phases. Functional validation experiments using 19 validated target miRNAs confirmed that these miRNAs indeed induced changes in process relevant phenotypes. Furthermore, computational miRNA target prediction combined with functional clustering identified putative target genes and cellular pathways, which might be regulated by these miRNAs. This study systematically identified novel target miRNAs during different phases and conditions of a biphasic fed-batch production process and functionally evaluated their potential for host cell engineering.

  11. Citric acid production from hydrolysate of pretreated straw cellulose by Yarrowia lipolytica SWJ-1b using batch and fed-batch cultivation. (United States)

    Liu, Xiaoyan; Lv, Jinshun; Zhang, Tong; Deng, Yuanfang


    In this study, crude cellulase produced by Trichoderma reesei Rut-30 was used to hydrolyze pretreated straw. After the compositions of the hydrolysate of pretreated straw were optimized, the study showed that natural components of pretreated straw without addition of any other components such as (NH4)2SO4, KH2PO4, or Mg(2+) were suitable for citric acid production by Yarrowia lipolytica SWJ-1b, and the optimal ventilatory capacity was 10.0 L/min/L medium. Batch and fed-batch production of citric acid from the hydrolysate of pretreated straw by Yarrowia lipolytica SWJ-1b has been investigated. In the batch cultivation, 25.4 g/L and 26.7 g/L citric acid were yields from glucose and hydrolysate of straw cellulose, respectively, while the cultivation time was 120 hr. In the three-cycle fed-batch cultivation, citric acid (CA) production was increased to 42.4 g/L and the cultivation time was extended to 240 hr. However, iso-citric acid (ICA) yield in fed-batch cultivation (4.0 g/L) was similar to that during the batch cultivation (3.9 g/L), and only 1.6 g/L of reducing sugar was left in the medium at the end of fed-batch cultivation, suggesting that most of the added carbon was used in the cultivation.

  12. Metabolic Control in Mammalian Fed-Batch Cell Cultures for Reduced Lactic Acid Accumulation and Improved Process Robustness

    Directory of Open Access Journals (Sweden)

    Viktor Konakovsky


    Full Text Available Biomass and cell-specific metabolic rates usually change dynamically over time, making the “feed according to need” strategy difficult to realize in a commercial fed-batch process. We here demonstrate a novel feeding strategy which is designed to hold a particular metabolic state in a fed-batch process by adaptive feeding in real time. The feed rate is calculated with a transferable biomass model based on capacitance, which changes the nutrient flow stoichiometrically in real time. A limited glucose environment was used to confine the cell in a particular metabolic state. In order to cope with uncertainty, two strategies were tested to change the adaptive feed rate and prevent starvation while in limitation: (i inline pH and online glucose concentration measurement or (ii inline pH alone, which was shown to be sufficient for the problem statement. In this contribution, we achieved metabolic control within a defined target range. The direct benefit was two-fold: the lactic acid profile was improved and pH could be kept stable. Multivariate Data Analysis (MVDA has shown that pH influenced lactic acid production or consumption in historical data sets. We demonstrate that a low pH (around 6.8 is not required for our strategy, as glucose availability is already limiting the flux. On the contrary, we boosted glycolytic flux in glucose limitation by setting the pH to 7.4. This new approach led to a yield of lactic acid/glucose (Y L/G around zero for the whole process time and high titers in our labs. We hypothesize that a higher carbon flux, resulting from a higher pH, may lead to more cells which produce more product. The relevance of this work aims at feeding mammalian cell cultures safely in limitation with a desired metabolic flux range. This resulted in extremely stable, low glucose levels, very robust pH profiles without acid/base interventions and a metabolic state in which lactic acid was consumed instead of being produced from day 1. With

  13. Improved docosahexaenoic acid production in Aurantiochytrium by glucose limited pH-auxostat fed-batch cultivation. (United States)

    Janthanomsuk, Panyawut; Verduyn, Cornelis; Chauvatcharin, Somchai


    Fed-batch, pH auxostat cultivation of the docosahexaenoic acid (DHA)-producing microorganism Aurantiochytrium B072 was performed to obtain high cell density and record high productivity of both total fatty acid (TFA) and DHA. Using glucose feeding by carbon excess (C-excess) and by C-limitation at various feeding rates (70%, 50% or 20% of C-excess), high biomass density was obtained and DHA/TFA content (w/w) was improved from 30% to 37% with a 50% glucose feed rate when compared with C-excess. To understand the biochemistry behind these improvements, lipogenic enzyme assays and in silico metabolic flux calculations were used and revealed that enzyme activity and C-fluxes to TFA were reduced with C-limited feeding but that the carbon flux to the polyketide synthase pathway increased relative to the fatty acid synthase pathway. As a result, a new strategy to improve the DHA to TFA content while maintaining relatively high DHA productivity is proposed. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. A new approach to ammonium sulphate feeding for fed-batch Arthrospira (Spirulina) platensis cultivation in tubular photobioreactor. (United States)

    Ferreira, Lívia S; Rodrigues, Mayla S; Converti, Attilio; Sato, Sunao; Carvalho, João Carlos M


    Arthrospira platensis was cultivated in tubular photobioreactor using different photosynthetic photon flux densities (PPFD) and protocols of (NH₄)₂SO₄ fed-batch supply. Results were evaluated by variance analysis selecting maximum cell concentration (X(m)), cell productivity (P(x)), nitrogen-to-cell conversion factor (Y(X/N)) and biomass, protein and lipid contents as responses. At PPFD of 120 and 240 μmol-photons/m² s, a parabolic profile of (NH₄)₂SO₄ addition aiming at producing biomass with 7% nitrogen content ensured X(m) values (14.1 and 12.2 g/L, respectively) comparable to those obtained with NaNO₃. At PPFD of 240 μmol-photons/m² s, P(x) (1.69 g/Ld) was 36% higher, although the photosynthetic efficiency (3.0%) was less than one-half that at PPFD of 120 μmol-photons/m² s. Biomass was shown to be constituted by about 35% proteins and 10% lipids, without any dependence on PPFD or kind of nitrogen source. These results highlight the possible use of (NH₄)₂SO₄ as alternative, cheap nitrogen source for A. platensis cultivation in tubular photobioreactors.

  15. Fed-batch cultivation of Arthrospira (Spirulina) platensis: potassium nitrate and ammonium chloride as simultaneous nitrogen sources. (United States)

    Rodrigues, M S; Ferreira, L S; Converti, A; Sato, S; Carvalho, J C M


    Arthrospiraplatensis was cultivated in minitanks at 13 klux, using a mixture of KNO(3) and NH(4)Cl as nitrogen source. Fed-batch daily supply of NH(4)Cl at exponentially-increasing feeding rate allowed preventing ammonia toxicity and nitrogen deficiency, providing high maximum cell concentration (X(m)) and high-quality biomass (21.85 mg chlorophyll g cells(-1); 20.5% lipids; 49.8% proteins). A central composite design combined to response surface methodology was utilized to determine the relationships between responses (X(m), cell productivity and nitrogen-to-cell conversion factor) and independent variables (KNO(3) and NH(4)Cl concentrations). Under optimum conditions (15.5mM KNO(3); 14.1mM NH(4)Cl), X(m) was 4327 mg L(-1), a value almost coincident with that obtained with only 25.4mM KNO(3), but more than twice that obtained with 21.5mM NH(4)Cl. A 30%-reduction of culture medium cost can be estimated when compared to KNO(3)-batch runs, thus behaving as a cheap alternative for the commercial production of this cyanobacterium.

  16. A process for energy-efficient high-solids fed-batch enzymatic liquefaction of cellulosic biomass. (United States)

    Cardona, M J; Tozzi, E J; Karuna, N; Jeoh, T; Powell, R L; McCarthy, M J


    The enzymatic hydrolysis of cellulosic biomass is a key step in the biochemical production of fuels and chemicals. Economically feasible large-scale implementation of the process requires operation at high solids loadings, i.e., biomass concentrations >15% (w/w). At increasing solids loadings, however, biomass forms a high viscosity slurry that becomes increasingly challenging to mix and severely mass transfer limited, which limits further addition of solids. To overcome these limitations, we developed a fed-batch process controlled by the yield stress and its changes during liquefaction of the reaction mixture. The process control relies on an in-line, non-invasive magnetic resonance imaging (MRI) rheometer to monitor real-time evolution of yield stress during liquefaction. Additionally, we demonstrate that timing of enzyme addition relative to biomass addition influences process efficiency, and the upper limit of solids loading is ultimately limited by end-product inhibition as soluble glucose and cellobiose accumulate in the liquid phase. Copyright © 2015 Elsevier Ltd. All rights reserved.

  17. Acrylamide synthesis using agar entrapped cells of Rhodococcus rhodochrous PA-34 in a partitioned fed batch reactor. (United States)

    Raj, Jog; Sharma, Nitya Nand; Prasad, Shreenath; Bhalla, Tek Chand


    The nitrile hydratase (Nhase) induced cells of Rhodococcus rhodochrous PA-34 catalyzed the conversion of acrylonitrile to acrylamide. The cells of R. rhodochrous PA-34 immobilized in 2% (w/v) agar (1.76 mg dcw/ml agar matrix) exhibited maximum Nhase activity (8.25 U/mg dcw) for conversion of acrylonitrile to acrylamide at 10 degrees C in the reaction mixture containing 0.1 M potassium phosphate buffer (pH 7.5), 8% (w/v) acrylonitrile and immobilized cells equivalent to 1.12 mg dcw (dry cell weight) per ml. In a partitioned fed batch reaction at 10 degrees C, using 1.12 g dcw immobilized cells in a final volume of 1 l, a total of 372 g of acrylonitrile was completely hydrated to acrylamide (498 g) in 24 h. From the above reaction mixture 87% acrylamide (432 g) was recovered through crystallization at 4 degrees C. By recycling the immobilized biocatalyst (six times), a total of 2,115 g acrylamide was produced.

  18. Fed-batch anaerobic valorization of slaughterhouse by-products with mesophilic microbial consortia without methane production. (United States)

    Pessiot, J; Nouaille, R; Jobard, M; Singhania, R R; Bournilhas, A; Christophe, G; Fontanille, P; Peyret, P; Fonty, G; Larroche, C


    This work aimed at setting up a fully instrumented, laboratory-scale bioreactor enabling anaerobic valorization of solid substrates through hydrogen and/or volatile fatty acid (VFA) production using mixed microbial populations (consortia). The substrate used was made of meat-based wastes, especially from slaughterhouses, which are becoming available in large amounts as a consequence of the growing constraints for waste disposal from meat industry. A reconstituted microbial mesophilic consortium without Archaebacteria (methanogens), named PBr, was cultivated in a 5-L anaerobic bioreactor on slaughterhouse wastes. The experiments were carried out with sequential fed-batch operations, including liquid medium removal from the bioreactor and addition of fresh substrate. VFAs and nitrogen were the main metabolites observed, while hydrogen accumulation was very low and no methane production was evidenced. After 1,300 h of culture, yields obtained for VFAs reached 0.38 g/g dry matter. Strain composition of the microbial consortium was also characterized using molecular tools (temporal temperature gradient gel electrophoresis and gene sequencing).

  19. Glutathione accumulation in ethanol-stat fed-batch culture of Saccharomyces cerevisiae with a switch to cysteine feeding. (United States)

    Nisamedtinov, Ildar; Kevvai, Kaspar; Orumets, Kerti; Rautio, Jari J; Paalme, Toomas


    Shot-wise supplementation of cysteine to a yeast culture is a common means of promoting glutathione (GSH) production. In the present work, we study the accumulation kinetics of cysteine, gamma-glutamylcysteine, and GSH and the expression of genes involved in GSH and sulfur metabolism in ethanol-stat fed-batch cultures as a result of switching to a medium enriched with cysteine and glycine. Supplementation in this fashion resulted in a rapid but short-term increase in the rate of GSH synthesis, while the expression of GSH1 decreased. Expression of GSH1 and GSH synthesis rate were observed to revert close to the base level after a few hours. These results indicate that, under such conditions, the control of GSH synthesis at higher concentrations occurred at the enzymatic, rather than the transcriptional level. The incorporation of cysteine into GSH was limited to approximately 40% of the theoretical yield, due to its requirement as a source of sulfur for protein synthesis under conditions whereby the sulfate assimilation pathway is down-regulated. This was supported by the expression profiles of genes involved in cysteine and homocysteine interconversion.

  20. Transformation of ferulic acid to vanillin using a fed-batch solid-liquid two-phase partitioning bioreactor. (United States)

    Ma, Xiao-kui; Daugulis, Andrew J


    Amycolatopsis sp. ATCC 39116 (formerly Streptomyces setonii) has shown promising results in converting ferulic acid (trans-4-hydroxy-3-methoxycinnamic acid; substrate), which can be derived from natural plant wastes, to vanillin (4-hydroxy-3-methoxybenzaldehyde). After exploring the influence of adding vanillin at different times during the growth cycle on cell growth and transformation performance of this strain and demonstrating the inhibitory effect of vanillin, a solid-liquid two-phase partitioning bioreactor (TPPB) system was used as an in situ product removal technique to enhance transformation productivity by this strain. The thermoplastic polymer Hytrel(®) G4078W was found to have superior partitioning capacity for vanillin with a partition coefficient of 12 and a low affinity for the substrate. A 3-L working volume solid-liquid fed-batch TPPB mode, using 300 g Hytrel G4078W as the sequestering phase, produced a final vanillin concentration of 19.5 g/L. The overall productivity of this reactor system was 450 mg/L. h, among the highest reported in literature. Vanillin was easily and quantitatively recovered from the polymers mostly by single stage extraction into methanol or other organic solvents used in food industry, simultaneously regenerating polymer beads for reuse. A polymer-liquid two phase bioreactor was again confirmed to easily outperform single phase systems that feature inhibitory or easily further degraded substrates/products. This enhancement strategy might reasonably be expected in the production of other flavor and fragrance compounds obtained by biotransformations.

  1. Combined data preprocessing and multivariate statistical analysis characterizes fed-batch culture of mouse hybridoma cells for rational medium design. (United States)

    Selvarasu, Suresh; Kim, Do Yun; Karimi, Iftekhar A; Lee, Dong-Yup


    We present an integrated framework for characterizing fed-batch cultures of mouse hybridoma cells producing monoclonal antibody (mAb). This framework systematically combines data preprocessing, elemental balancing and statistical analysis technique. Initially, specific rates of cell growth, glucose/amino acid consumptions and mAb/metabolite productions were calculated via curve fitting using logistic equations, with subsequent elemental balancing of the preprocessed data indicating the presence of experimental measurement errors. Multivariate statistical analysis was then employed to understand physiological characteristics of the cellular system. The results from principal component analysis (PCA) revealed three major clusters of amino acids with similar trends in their consumption profiles: (i) arginine, threonine and serine, (ii) glycine, tyrosine, phenylalanine, methionine, histidine and asparagine, and (iii) lysine, valine and isoleucine. Further analysis using partial least square (PLS) regression identified key amino acids which were positively or negatively correlated with the cell growth, mAb production and the generation of lactate and ammonia. Based on these results, the optimal concentrations of key amino acids in the feed medium can be inferred, potentially leading to an increase in cell viability and productivity, as well as a decrease in toxic waste production. The study demonstrated how the current methodological framework using multivariate statistical analysis techniques can serve as a potential tool for deriving rational medium design strategies. Copyright © 2010 Elsevier B.V. All rights reserved.

  2. Modelling Fungal Fermentations for Enzyme Production

    DEFF Research Database (Denmark)

    Albæk, Mads Orla; Gernaey, Krist; Hansen, Morten S.

    We have developed a process model of fungal fed-batch fermentations for enzyme production. In these processes, oxygen transfer rate is limiting and controls the substrate feeding rate. The model has been shown to describe cultivations of both Aspergillus oryzae and Trichoderma reesei strains in 550...


    Directory of Open Access Journals (Sweden)

    A. C. L. Horta


    Full Text Available Abstract The performance of an in-situ capacitance sensor for on-line monitoring of biomass concentration was evaluated for some of the most important microorganisms in the biotechnology industry: Escherichia coli, Saccharomyces cerevisiae, Pichia pastoris and Bacillus megaterium. A total of 33 batch and fed-batch cultures were carried out in a bench-scale bioreactor and biomass formation trends were followed by dielectric measurements during the growth phase as well as the induction phase, for 5 recombinant E. coli strains. Permittivity measurements and viable cellular concentrations presented a linear correlation for all the studied conditions. In addition, the permittivity signal was further used for inference of the cellular growth rate. The estimated specific growth rates mirrored the main trends of the metabolic states of the different cells and they can be further used for setting-up control strategies in fed-batch cultures.

  4. A Substrate Fed-Batch Biphasic Catalysis Process for the Production of Natural Crosslinking Agent Genipin with Fusarium solani ACCC 36223. (United States)

    Zhu, Yuyao; Zhao, Botao; Huang, Xiaode; Chen, Bin; Qian, Hua


    The natural crosslinking agent genipin has been applied widely in biomedicines and foods nowadays. Because of the special hemiacetal ring structure in its molecule, it can only be prepared by hydrolysis of geniposide according to biocatalysis. In this research, strategies including aqueous-organic biphasic catalysis and substrate fed-batch mode were adopted to improve the biocatalysis process of genipin. A 10 L ethyl acetate-aqueous biphasic system with geniposide fed-batch led to a satisfying genipin yield. With Fusarium solani ACCC 36223, 15.7 g/l genipin in the ethyl acetate phase was obtained, corresponding to space-time yields of 0.654 g l(-1) h(-1).

  5. Biotransformation of sweet lime pulp waste into high-quality nanocellulose with an excellent productivity using Komagataeibacter europaeus SGP37 under static intermittent fed-batch cultivation. (United States)

    Dubey, Swati; Singh, Jyoti; Singh, R P


    Herein, sweet lime pulp waste (SLPW) was utilized as a low- or no-cost feedstock for the production of bacterial nanocellulose (BNC) alone and in amalgamation with other nutritional supplements by the isolate K. europaeus SGP37 under static batch and static intermittent fed-batch cultivation. The highest yield (26.2±1.50gL(-1)) was obtained in the hot water extract of SLPW supplemented with the components of HS medium, which got further boosted to 38±0.85gL(-1) as the cultivation strategy was shifted from static batch to static intermittent fed-batch. BNC obtained from various SLPW medium was similar or even superior to that obtained with standard HS medium in terms of its physicochemical properties. The production yields of BNC thus obtained are significantly higher and fit well in terms of industrial scale production. Copyright © 2017 Elsevier Ltd. All rights reserved.


    Directory of Open Access Journals (Sweden)

    Mahmoud M. Berekaa


    Full Text Available Bacillus megaterium SW1-2 showed enhanced growth and polyhydroxybutyrate (PHB production during cultivation on date palm syrup (DEPS or sugar cane molasses. FT-IR and NMR spectroscopic analyses of the polymer accumulated during growth on DEPS revealed specific absorption peaks characteristic for PHB. 1.65 g/L of PHB (56.9% CDW was produced during growth on medium supplemented with 2 g/L of DEPS. Approximately, 36.1% CDW of PHB were recorded during growth on sugar cane molasses. Six runs of different fed-batch cultivation strategies were tested, the optimal run showed approximately 6.87-fold increase. Modified E2 medium was prefered recording 10.11 and 11.34 g/L of total PHB produced for runs 1 and 2, at the end of 96 h incubation period, respectively. Decrease in PHB was recorded during growth on complex medium (run 3 and run 4. In another independent optimization strategy, ten variables were concurrently examined for their significance on PHB production by Plackett-Burman statistical design for the first time. Among variables, DEPS-II and inoculum concentration followed by KH2PO4 and (NH42SO4 were found to be the most significant variables encourage PHB production. Indeed, DEPS-II or Fresh syrup is more significant than commercial syrup DEPS-I (p-value= 0.05. RPM, incubation period have highly negative effect on PHB production. Role of ago-industrial wastes, especially DEPS, in enhancement of PHB production was closely discussed.

  7. Real-time monitoring and control of the specific growth rate in yeast fed-batch cultures based on process analytical technology tools such as biocalorimetry or spectroscopy


    Schuler, Moira Monika


    Key features of bioprocesses, such as product quantity and quality, but also cell physiology can be related to the growth characteristics of the organism under study. The specific growth rate, a key variable, cannot be measured directly, but might be estimated and inferred from other measurable variables such as biomass, substrate or product concentrations. The present thesis reviews techniques for real-time estimation and control of the specific growth rate in microbial fed-batch cultures by...

  8. A multi-pronged investigation into the effect of glucose starvation and culture duration on fed-batch CHO cell culture

    DEFF Research Database (Denmark)

    Fan, Yuzhou; Jimenez Del Val, Ioscani; Müller, Christian;


    In this study, omics-based analysis tools were used to explore the effect of glucose starvation and culture duration on monoclonal antibody (mAb) production in fed-batch CHO cell culture to gain better insight into how these parameters can be controlled to ensure optimal mAb productivity and qual......In this study, omics-based analysis tools were used to explore the effect of glucose starvation and culture duration on monoclonal antibody (mAb) production in fed-batch CHO cell culture to gain better insight into how these parameters can be controlled to ensure optimal mAb productivity...... and quality. Titer and N-glycosylation of mAbs, as well as proteomic signature and metabolic status of the production cells in the culture were assessed. We found that the impact of glucose starvation on the titer and N-glycosylation of mAbs was dependent on the degree of starvation during early stationary...... phase of the fed-batch culture. Higher degree of glucose starvation reduced intracellular concentrations of UDP-GlcNAc and UDP-GalNAc, but increased the levels of UDP-Glc and UDP-Gal. Increased GlcNAc and Gal occupancy correlated well with increased degree of glucose starvation, which can be attributed...

  9. Change in turnover capacity of crude recombinant dye-decolorizing peroxidase (rDyP) in batch and fed-batch decolorization of Remazol Brilliant Blue R. (United States)

    Shakeri, M; Shoda, M


    Decolorization of the representative anthraquinone dye, Remazol Brilliant Blue R (RBBR) was assessed to determine the practical potential of crude recombinant dye-decolorizing peroxidase generated by Aspergillus oryzae (rDyP) in term of turnover capacity of rDyP. The turnover capacity, defined as the milligram of RBBR decolorized per unit of rDyP inactivated over the catalytic life time of rDyP, was quantified under condition by H(2)O(2) -mediated rDyP inactivation. In batch culture, equimolar batch addition of H(2)O(2) and RBBR yielded complete decolorization of RBBR by rDyP, with a turnover capacity of 4.75. In stepwise fed-batch addition of H(2)O(2), the turnover capacity increased to 5.76, and by increasing dye concentration, it reached 14.3. When H(2)O(2) was added in continuous fed-batch to minimize rDyP inactivation and 1.6 mM dye was added in stepwise fed-batch mode, the turnover capacity increased to 20.4. At this turnover capacity, 1 l of crude rDyP solution containing 5,000 U could decolorize up to 102 g RBBR in 650 min.

  10. Aroma improvement by repeated freeze-thaw treatment during Tuber melanosporum fermentation (United States)

    Xiao, Deng-Rong; Liu, Rui-Sang; He, Long; Li, Hong-Mei; Tang, Ya-Ling; Liang, Xin-Hua; Chen, Tao; Tang, Ya-Jie


    The aroma attributes of sulfurous, mushroom and earthy are the most important characteristics of the aroma of Tuber melanosporum. However, these three aroma attributes are absent in the T. melanosporum fermentation system. To improve the quality of the aroma, repeated freeze-thaw treatment (RFTT) was adopted to affect the interplay of volatile organic compounds (VOCs). Using RFTT, not only was the score on the hedonic scale of the aroma increased from the “liked slightly” to the “liked moderately” grade, but the aroma attributes of sulfurous, mushroom and earthy could also be smelled in the T. melanosporum fermentation system for the first time. A total of 29 VOCs were identified, and 9 compounds were identified as the key discriminative volatiles affected by RFTT. Amino acid analysis revealed that methionine, valine, serine, phenylalanine, isoleucine and threonine were the key substrates associated with the biosynthesis of the 9 key discriminative VOCs. This study noted that amino acid metabolism played an important role in the regulation of the aroma of the T. melanosporum fermentation system. PMID:26607288

  11. Aroma improvement by repeated freeze-thaw treatment during Tuber melanosporum fermentation. (United States)

    Xiao, Deng-Rong; Liu, Rui-Sang; He, Long; Li, Hong-Mei; Tang, Ya-Ling; Liang, Xin-Hua; Chen, Tao; Tang, Ya-Jie


    The aroma attributes of sulfurous, mushroom and earthy are the most important characteristics of the aroma of Tuber melanosporum. However, these three aroma attributes are absent in the T. melanosporum fermentation system. To improve the quality of the aroma, repeated freeze-thaw treatment (RFTT) was adopted to affect the interplay of volatile organic compounds (VOCs). Using RFTT, not only was the score on the hedonic scale of the aroma increased from the "liked slightly" to the "liked moderately" grade, but the aroma attributes of sulfurous, mushroom and earthy could also be smelled in the T. melanosporum fermentation system for the first time. A total of 29 VOCs were identified, and 9 compounds were identified as the key discriminative volatiles affected by RFTT. Amino acid analysis revealed that methionine, valine, serine, phenylalanine, isoleucine and threonine were the key substrates associated with the biosynthesis of the 9 key discriminative VOCs. This study noted that amino acid metabolism played an important role in the regulation of the aroma of the T. melanosporum fermentation system.

  12. Comparison of Different Strategies for Selection/Adaptation of Mixed Microbial Cultures Able to Ferment Crude Glycerol Derived from Second-Generation Biodiesel

    Directory of Open Access Journals (Sweden)

    C. Varrone


    Full Text Available Objective of this study was the selection and adaptation of mixed microbial cultures (MMCs, able to ferment crude glycerol generated from animal fat-based biodiesel and produce building-blocks and green chemicals. Various adaptation strategies have been investigated for the enrichment of suitable and stable MMC, trying to overcome inhibition problems and enhance substrate degradation efficiency, as well as generation of soluble fermentation products. Repeated transfers in small batches and fed-batch conditions have been applied, comparing the use of different inoculum, growth media, and Kinetic Control. The adaptation of activated sludge inoculum was performed successfully and continued unhindered for several months. The best results showed a substrate degradation efficiency of almost 100% (about 10 g/L glycerol in 21 h and different dominant metabolic products were obtained, depending on the selection strategy (mainly 1,3-propanediol, ethanol, or butyrate. On the other hand, anaerobic sludge exhibited inactivation after a few transfers. To circumvent this problem, fed-batch mode was used as an alternative adaptation strategy, which led to effective substrate degradation and high 1,3-propanediol and butyrate production. Changes in microbial composition were monitored by means of Next Generation Sequencing, revealing a dominance of glycerol consuming species, such as Clostridium, Klebsiella, and Escherichia.

  13. Comparison of Different Strategies for Selection/Adaptation of Mixed Microbial Cultures Able to Ferment Crude Glycerol Derived from Second-Generation Biodiesel (United States)

    Varrone, C.; Heggeset, T. M. B.; Le, S. B.; Haugen, T.; Markussen, S.; Skiadas, I. V.; Gavala, H. N.


    Objective of this study was the selection and adaptation of mixed microbial cultures (MMCs), able to ferment crude glycerol generated from animal fat-based biodiesel and produce building-blocks and green chemicals. Various adaptation strategies have been investigated for the enrichment of suitable and stable MMC, trying to overcome inhibition problems and enhance substrate degradation efficiency, as well as generation of soluble fermentation products. Repeated transfers in small batches and fed-batch conditions have been applied, comparing the use of different inoculum, growth media, and Kinetic Control. The adaptation of activated sludge inoculum was performed successfully and continued unhindered for several months. The best results showed a substrate degradation efficiency of almost 100% (about 10 g/L glycerol in 21 h) and different dominant metabolic products were obtained, depending on the selection strategy (mainly 1,3-propanediol, ethanol, or butyrate). On the other hand, anaerobic sludge exhibited inactivation after a few transfers. To circumvent this problem, fed-batch mode was used as an alternative adaptation strategy, which led to effective substrate degradation and high 1,3-propanediol and butyrate production. Changes in microbial composition were monitored by means of Next Generation Sequencing, revealing a dominance of glycerol consuming species, such as Clostridium, Klebsiella, and Escherichia. PMID:26509171

  14. Comparison of Different Strategies for Selection/Adaptation of Mixed Microbial Cultures Able to Ferment Crude Glycerol Derived from Second-Generation Biodiesel

    DEFF Research Database (Denmark)

    Varrone, Cristiano; Heggeset, T. M. B.; Le, S. B.


    Objective of this study was the selection and adaptation of mixed microbial cultures (MMCs), able to ferment crude glycerol generated from animal fat-based biodiesel and produce building-blocks and green chemicals. Various adaptation strategies have been investigated for the enrichment of suitable...... and stable MMC, trying to overcome inhibition problems and enhance substrate degradation efficiency, as well as generation of soluble fermentation products. Repeated transfers in small batches and fed-batch conditions have been applied, comparing the use of different inoculum, growth media, and Kinetic...... Control. The adaptation of activated sludge inoculum was performed successfully and continued unhindered for several months. The best results showed a substrate degradation efficiency of almost 100% (about 10 g/L glycerol in 21 h) and different dominant metabolic products were obtained, depending...

  15. Application of Multivariate Analysis Tools to Industrial Scale Fermentation Data

    DEFF Research Database (Denmark)

    Mears, Lisa; Nørregård, Rasmus; Stocks, Stuart M.;

    . 2014). This may be the case for fed-batch fermentation processes, where mechanistic modelling is challenging due to non-linear dynamics, and non-steady state operation. There is also a lack of sensors for key parameters which are considered to define the quality of the batch, such as product...

  16. Improvement of l-lactic acid productivity from sweet sorghum juice by repeated batch fermentation coupled with membrane separation. (United States)

    Wang, Yong; Meng, Hongyu; Cai, Di; Wang, Bin; Qin, Peiyong; Wang, Zheng; Tan, Tianwei


    In order to efficiently produce l-lactic acid from non-food feedstocks, sweet sorghum juice (SSJ), which is rich of fermentable sugars, was directly used for l-lactic acid fermentation by Lactobacillus rhamnosus LA-04-1. A membrane integrated repeated batch fermentation (MIRB) was developed for productivity improvement. High-cell-density fermentation was achieved with a final cell density (OD620) of 42.3, and the CCR effect was overcomed. When SSJ (6.77gL(-1) glucose, 4.51gL(-1) fructose and 50.46gL(-1) sucrose) was used as carbon source in MIRB process, l-lactic acid productivity was increased significantly from 1.45gL(-1)h(-1) (batch 1) to 17.55gL(-1)h(-1) (batch 6). This process introduces an effective way to produce l-lactic acid from SSJ.

  17. Mainstream partial nitritation and anammox in a 200,000 m3/day activated sludge process in Singapore: scale-down by using laboratory fed-batch reactor. (United States)

    Yeshi, Cao; Hong, Kwok Bee; van Loosdrecht, Mark C M; Daigger, Glen T; Yi, Png Hui; Wah, Yuen Long; Chye, Chua Seng; Ghani, Yahya Abd


    A laboratory fed-batch reactor has been used to study under controlled conditions the performance of partial nitritation/anammox for the 200,000 m(3)/day step-feed activated sludge process at the Changi Water Reclamation Plant, Singapore. The similarity of the concentrations of NH(4), NO(2), NO(3), PO(4), suspended chemical oxygen demand (sCOD), pH, and alkalinity (ALK) between the on-site process and laboratory reactor illustrates that the laboratory fed-batch reactor can be used to simulate the site performance. The performance of the reactor fed by primary effluent illustrated the existence of anammox and heterotrophic denitrification and apparent excessive biological phosphorus removal as observed from the site. The performance of the reactor fed by final effluent proved the presence of anammox process on site. Both the laboratory reactor and on-site process showed that higher influent 5-day biochemical oxygen demand/total nitrogen (BOD(5)/TN) (COD/TN) ratio increases the nitrogen removal efficiency of the process.

  18. Improvement of cloned [alpha]-amylase gene expression in fed-batch culture of recombinant Saccharomyces cerevisiae by regulating both glucose and ethanol concentrations using a fuzzy controller

    Energy Technology Data Exchange (ETDEWEB)

    Shiba, Sumihisa; Nishida, Yoshio; Park, Y.S.; Iijima, Shinji; Kobayashi, Takeshi (Nagoya Univ. (Japan). Dept. of Biotechnology)


    The effect of ethanol concentration on cloned gene expression in recombinant Saccharomyces cerevisiae strain 20B-12 containing one of two plasmids, pNA3 and pNA7, was investigated in batch cultures. Plasmids pNA3 and pNA7 contain the [alpha]-amylase gene under the control of the SUC2 or PGK promoter, respectively. When the ethanol concentration was controlled at 2 to 5 g/L, the gene expressions were two times higher than those at 20 g/L ethanol. To increase the gene expression by maintaining both the ethanol and glucose concentrations at low levels, a fuzzy controller was developed. The concentrations of glucose and ethanol were controlled simultaneously at 0.15 and 2 g/L, respectively, in the production phase using the fuzzy controller in fed-batch culture. The synthesis of [alpha]-amylase was induced by the low glucose concentration and maintained at a high level of activity by regulating the ethanol concentration at 2 g/L. The secretory [alpha]-amylase activities of cells harboring plasmids pNA3 and pNA7 in fed-batch culture were 175 and 392 U/mL, and their maximal specific activities 7.7 and 12.4 U/mg dry cells, respectively. These values are two to three times higher in activity and three to four times higher in specific activity than those obtained when glucose only was controlled.

  19. Fed-batch bioreactor process scale-up from 3-L to 2,500-L scale for monoclonal antibody production from cell culture. (United States)

    Yang, Jeng-Dar; Lu, Canghai; Stasny, Brad; Henley, Joseph; Guinto, Woodrow; Gonzalez, Carlos; Gleason, Joseph; Fung, Monica; Collopy, Brett; Benjamino, Michael; Gangi, Jennifer; Hanson, Melissa; Ille, Elisabeth


    This case study focuses on the scale-up of a Sp2/0 mouse myeloma cell line based fed-batch bioreactor process, from the initial 3-L bench scale to the 2,500-L scale. A stepwise scale-up strategy that involved several intermediate steps in increasing the bioreactor volume was adopted to minimize the risks associated with scale-up processes. Careful selection of several available mixing models from literature, and appropriately applying the calculated results to our settings, resulted in successful scale-up of agitation speed for the large bioreactors. Consideration was also given to scale-up of the nutrient feeding, inoculation, and the set-points of operational parameters such as temperature, pH, dissolved oxygen, dissolved carbon dioxide, and aeration in an integrated manner. It has been demonstrated through the qualitative and the quantitative side-by-side comparison of bioreactor performance as well as through a panel of biochemical characterization tests that the comparability of the process and the product was well controlled and maintained during the process scale-up. The 2,500-L process is currently in use for the routine clinical production of Epratuzumab in support of two global Phase III clinical trials in patients with lupus. Today, the 2,500 L, fed-batch production process for Epratuzumab has met all scheduled batch releases, and the quality of the antibody is consistent and reproducible, meeting all specifications, thus confirming the robustness of the process.

  20. A multi-pronged investigation into the effect of glucose starvation and culture duration on fed-batch CHO cell culture

    DEFF Research Database (Denmark)

    Fan, Yuzhou; Jimenez Del Val, Ioscani; Müller, Christian;


    In this study, omics-based analysis tools were used to explore the effect of glucose starvation and culture duration on monoclonal antibody (mAb) production in fed-batch CHO cell culture to gain better insight into how these parameters can be controlled to ensure optimal mAb productivity...... and quality. Titer and N-glycosylation of mAbs, as well as proteomic signature and metabolic status of the production cells in the culture were assessed. We found that the impact of glucose starvation on the titer and N-glycosylation of mAbs was dependent on the degree of starvation during early stationary...... phase of the fed-batch culture. Higher degree of glucose starvation reduced intracellular concentrations of UDP-GlcNAc and UDP-GalNAc, but increased the levels of UDP-Glc and UDP-Gal. Increased GlcNAc and Gal occupancy correlated well with increased degree of glucose starvation, which can be attributed...

  1. The fed-batch principle for the molecular biology lab: controlled nutrient diets in ready-made media improve production of recombinant proteins in Escherichia coli. (United States)

    Krause, Mirja; Neubauer, Antje; Neubauer, Peter


    While the nutrient limited fed-batch technology is the standard of the cultivation of microorganisms and production of heterologous proteins in industry, despite its advantages in view of metabolic control and high cell density growth, shaken batch cultures are still the standard for protein production and expression screening in molecular biology and biochemistry laboratories. This is due to the difficulty and expenses to apply a controlled continuous glucose feed to shaken cultures. New ready-made growth media, e.g. by biocatalytic release of glucose from a polymer, offer a simple solution for the application of the fed-batch principle in shaken plate and flask cultures. Their wider use has shown that the controlled diet not only provides a solution to obtain significantly higher cell yields, but also in many cases folding of the target protein is improved by the applied lower growth rates; i.e. final volumetric yields for the active protein can be a multiple of what is obtained in complex medium cultures. The combination of the conventional optimization approaches with new and easy applicable growth systems has revolutionized recombinant protein production in Escherichia coli in view of product yield, culture robustness as well as significantly increased cell densities. This technical development establishes the basis for successful miniaturization and parallelization which is now an important tool for synthetic biology and protein engineering approaches. This review provides an overview of the recent developments, results and applications of advanced growth systems which use a controlled glucose release as substrate supply.

  2. Enhancement of canthaxanthin production from Dietzia natronolimnaea HS-1 in a fed-batch process using trace elements and statistical methods

    Directory of Open Access Journals (Sweden)

    M. R. Nasri Nasrabadi


    Full Text Available Under fed-batch process conditions, the statistical analysis of trace elements was performed by application of Plackett-Burman design (for screening tests and response surface methodology (for predicting the optimal points to achieve the highest level of canthaxanthin production from Dietzia natronolimnaea HS-1. Plackett-Burman design was conducted on eleven trace elements (i. e., aluminum, boron, cobalt, copper, iron, magnesium, manganese, molybdenum, selenium, vanadium and zinc to select out elements that significantly enhance the canthaxanthin production of D. natronolimnaea HS-1. Plackett-Burman design revealed that Fe3+, Cu2+ and Zn2+ ions had the highest effect on canthaxanthin production of D. natronolimnaea HS-1 (P<0.05. These three elements were used for further optimization. By means of response surface methodology for the fed-batch process, the optimum conditions to achieve the highest level of canthaxanthin (8923±18 µg/L were determined as follow: Fe3+ 30 ppm, Cu2+ 28.75 ppm and Zn2+ 27 ppm.

  3. Butyric acid fermentation in a fibrous bed bioreactor with immobilized Clostridium tyrobutyricum from cane molasses. (United States)

    Jiang, Ling; Wang, Jufang; Liang, Shizhong; Wang, Xiaoning; Cen, Peilin; Xu, Zhinan


    Butyrate fermentation by immobilized Clostridium tyrobutyricum was successfully carried out in a fibrous bed bioreactor using cane molasses. Batch fermentations were conducted to investigate the influence of pH on the metabolism of the strain, and the results showed that the fermentation gave a highest butyrate production of 26.2 g l(-1) with yield of 0.47 g g(-1) and reactor productivity up to 4.13 g l(-1)h(-1) at pH 6.0. When repeated-batch fermentation was carried out, long-term operation with high butyrate yield, volumetric productivity was achieved. Several cane molasses pretreatment techniques were investigated, and it was found that sulfuric acid treatment gave better results regarding butyrate concentration (34.6+/-0.8 g l(-1)), yield (0.58+/-0.01 g g(-1)), and sugar utilization (90.8+/-0.9%). Also, fed-batch fermentation from cane molasses pretreated with sulfuric acid was performed to further increase the concentration of butyrate up to 55.2 g l(-1).

  4. Overcome of Carbon Catabolite Repression of Bioinsecticides Production by Sporeless Bacillus thuringiensis through Adequate Fermentation Technology. (United States)

    Ben Khedher, Saoussen; Jaoua, Samir; Zouari, Nabil


    The overcoming of catabolite repression, in bioinsecticides production by sporeless Bacillus thuringiensis strain S22 was investigated into fully controlled 3 L fermenter, using glucose based medium. When applying adequate oxygen profile throughout the fermentation period (75% oxygen saturation), it was possible to partially overcome the catabolite repression, normally occurring at high initial glucose concentrations (30 and 40 g/L glucose). Moreover, toxin production yield by sporeless strain S22 was markedly improved by the adoption of the fed-batch intermittent cultures technology. With 22.5 g/L glucose used into culture medium, toxin production was improved by about 36% when applying fed-batch culture compared to one batch. Consequently, the proposed fed-batch strategy was efficient for the overcome of the carbon catabolite repression. So, it was possible to overproduce insecticidal crystal proteins into highly concentrated medium.

  5. Overcome of Carbon Catabolite Repression of Bioinsecticides Production by Sporeless Bacillus thuringiensis through Adequate Fermentation Technology

    Directory of Open Access Journals (Sweden)

    Saoussen Ben Khedher


    Full Text Available The overcoming of catabolite repression, in bioinsecticides production by sporeless Bacillus thuringiensis strain S22 was investigated into fully controlled 3 L fermenter, using glucose based medium. When applying adequate oxygen profile throughout the fermentation period (75% oxygen saturation, it was possible to partially overcome the catabolite repression, normally occurring at high initial glucose concentrations (30 and 40 g/L glucose. Moreover, toxin production yield by sporeless strain S22 was markedly improved by the adoption of the fed-batch intermittent cultures technology. With 22.5 g/L glucose used into culture medium, toxin production was improved by about 36% when applying fed-batch culture compared to one batch. Consequently, the proposed fed-batch strategy was efficient for the overcome of the carbon catabolite repression. So, it was possible to overproduce insecticidal crystal proteins into highly concentrated medium.

  6. Efficient production of bioactive metabolites from Antrodia camphorata ATCC 200183 by asexual reproduction-based repeated batch fermentation. (United States)

    Li, Hua-Xiang; Lu, Zhen-Ming; Geng, Yan; Gong, Jin-Song; Zhang, Xiao-Juan; Shi, Jin-Song; Xu, Zheng-Hong; Ma, Yan-He


    Large-scale submerged fermentation (SmF) of Antrodia camphorata (A. camphorata) usually encounters challenges including tedious preparation of mycelial inoculum, long fermentation period (10-14 d), and poor repeatability. Here we developed an asexual reproduction-based repeated batch fermentation (RBF) process for bioactive metabolites production by A. camphorata ATCC 200183. Compared with traditional batch fermentation, production time was shortened to 58 d from 80 d (overall time for eight cycles) using the RBF process established in this study, and accordingly, the productivities of bioactive metabolites (including antrodins) were improved by 40-60%. Kinetic parameters (α is 2.1-18.7 times as β) indicated that the cell growth was the major contribution for bioactive metabolites production. The RBF shows excellent batch-repeatability (Pearson correlation coefficient of 0.998±0.001), together with advantages of energy-efficient, low cost, and labor-saving, RBF process can be implemented to SmF by other filamentous fungi.

  7. Optimization of simultaneous saccharification and fermentation conditions with amphipathic lignin derivatives for concentrated bioethanol production. (United States)

    Cheng, Ningning; Koda, Keiichi; Tamai, Yutaka; Yamamoto, Yoko; Takasuka, Taichi E; Uraki, Yasumitsu


    Amphipathic lignin derivatives (A-LDs) prepared from the black liquor of soda pulping of Japanese cedar are strong accelerators for bioethanol production under a fed-batch simultaneous enzymatic saccharification and fermentation (SSF) process. To improve the bioethanol production concentration, conditions such as reaction temperature, stirring program, and A-LDs loadings were optimized in both small scale and large scale fed-batch SSF. The fed-batch SSF in the presence of 3.0g/L A-LDs at 38°C gave the maximum ethanol production and a high enzyme recovery rate. Furthermore, a jar-fermenter equipped with a powerful mechanical stirrer was designed for 1.5L-scale fed-batch SSF to achieve rigorous mixing during high substrate loading. Finally, the 1.5L fed-batch SSF with a substrate loading of 30% (w/v) produced a high ethanol concentration of 87.9g/L in the presence of A-LDs under optimized conditions.

  8. Polymalic acid fermentation by Aureobasidium pullulans for malic acid production from soybean hull and soy molasses: Fermentation kinetics and economic analysis. (United States)

    Cheng, Chi; Zhou, Yipin; Lin, Meng; Wei, Peilian; Yang, Shang-Tian


    Polymalic acid (PMA) production by Aureobasidium pullulans ZX-10 from soybean hull hydrolysate supplemented with corn steep liquor (CSL) gave a malic acid yield of ∼0.4g/g at a productivity of ∼0.5g/L·h. ZX-10 can also ferment soy molasses, converting all carbohydrates including the raffinose family oligosaccharides to PMA, giving a high titer (71.9g/L) and yield (0.69g/g) at a productivity of 0.29g/L·h in fed-batch fermentation under nitrogen limitation. A higher productivity of 0.64g/L·h was obtained in repeated batch fermentation with cell recycle and CSL supplementation. Cost analysis for a 5000 MT plant shows that malic acid can be produced at $1.10/kg from soy molasses, $1.37/kg from corn, and $1.74/kg from soybean hull. At the market price of $1.75/kg, malic acid production from soy molasses via PMA fermentation offers an economically competitive process for industrial production of bio-based malic acid.

  9. Characteristics of human cell line, F2N78, for the production of recombinant antibody in fed-batch and perfusion cultures. (United States)

    Seo, Joon Serk; Min, Byung Sub; Kwon, Young-Bum; Lee, Soo-Young; Cho, Jong-Moon; Park, Keun-Hee; Yang, Yae Ji; Maeng, Ki Eun; Chang, Shin-Jae; Kim, Dong-Il


    A human hybrid cell line, F2N78, was developed by somatic fusion of HEK293 and Namalwa cells for the production recombinant biopharmaceutical proteins. In this study, we performed perfusion culture to verify its potential in culture process used for human cell expression platform. Cell viability could be maintained over 90% and high viable cell density was obtained at higher than 1.0 × 10(7) cells/mL by bleeding process in perfusion culture. The cells were adapted well in both culture modes, but there were apparent differences in protein quality. Compared to fed-batch culture, degalactosylated forms such as G0F and G0 as well as Man5 showed no significant increases in perfusion culture. In terms of charge variants, acidic peaks increased, whereas main peaks constantly decreased according to the length of culture period in both methods.

  10. Using a medium of free amino acids to produce penicillin g acylase in fed-batch cultivations of Bacillus megaterium ATCC 14945

    Directory of Open Access Journals (Sweden)

    R. G. Silva


    Full Text Available The production of penicillin G acylase (PGA, an important industrial enzyme from a wild strain of Bacillus megaterium using a pool of free amino acids as substrate was studied in a bench-scale bioreactor. Experiments carried out in shakers showed that the substitution of casein for free amino acids in the presence of cheese whey was the culture medium that provided the highest productivity. Several cultivations were carried out in a bioreactor operated in either batch or fed-batch mode. Batch runs showed that enzyme production is associated with microorganism growth. The following set of amino acids was preferentially consumed: Ala, Arg, Asp, Gly, Lys, Ser, Thr and Trp. On the other hand, the rates of consumption of His, Ile, Leu, Met, Phe, Pro, Tyr and Val were lower.

  11. Scale effect of anaerobic digestion tests in fed-batch and semi-continuous mode for the technical and economic feasibility of a full scale digester. (United States)

    Ruffino, Barbara; Fiore, Silvia; Roati, Chiara; Campo, Giuseppe; Novarino, Daniel; Zanetti, Mariachiara


    Methane production capacity in mesophilic conditions of waste from two food industry plants was assessed in a semi-pilot (6L, fed-batch) and pilot (300 L, semi-continuous) scale. This was carried out in order to evaluate the convenience of producing heat and electricity in a full scale anaerobic digester. The pilot test was performed in order to obtain more reliable results for the design of the digester. Methane yield, returned from the pilot scale test, was approximately 80% of that from the smaller scale test. This outcome was in line with those from other studies performed in different scales and modes and indicates the success of the pilot scale test. The net electricity produced from the digester accounted for 30-50% of the food industry plants' consumption. The available thermal energy could cover from 10% to 100% of the plant requirements, depending on the energy demand of the processes performed.

  12. Microencapsulation of Baker’s Yeast in Gellan Gum Beads Used in Repeated Cycles of Glucose Fermentation

    Directory of Open Access Journals (Sweden)

    Camelia Elena Iurciuc (Tincu


    Full Text Available The purpose of this work is to prepare ionically cross-linked (with CaCl2 gellan particles with immobilized yeast cells for their use in repeated fermentation cycles of glucose. The study investigates the influence of ionic cross-linker concentration on the stability and physical properties of the particles obtained before extrusion and during time in the coagulation bath (the cross-linker solution with different CaCl2 concentrations. It was found that by increasing the amount of the cross-linker the degree of cross-linking in the spherical gellan matrix increases, having a direct influence on the particle morphology and swelling degree in water. These characteristics were found to be very important for diffusion of substrate, that is, the glucose, into the yeast immobilized cells and for the biocatalytic activity of the yeast immobilized cells in gellan particles. These results highlight the potential of these bioreactors to be used in repeated fermentation cycles (minimum 10 without reducing their biocatalytic activity and maintaining their productivity at similar parameters to those obtained in the free yeast fermentation. Encapsulation of Saccharomyces cerevisiae into the gellan gum beads plays a role in the effective application of immobilized yeast for the fermentation process.


    Directory of Open Access Journals (Sweden)

    C. Cruz-Martínez


    Full Text Available AbstractNH4NO3 simultaneously provides a readily assimilable nitrogen source (ammonia and a reserve of nitrogen (nitrate, allowing for an increase in Arthrospira platensis biomass production while reducing the cost of the cultivation medium. In this study, a 22plus star central composite experimental design combined with response surface methodology was employed to analyze the influence of light intensity (I and the total amount of added NH4NO3 (Mt on a bench-scale tubular photobioreactor for fed-batch cultures. The maximum cell concentration (Xm, cell productivity (PX and biomass yield on nitrogen (YX/N were evaluated, as were the protein and lipid contents. Under optimized conditions (I = 148 μmol·photons·m-2·s-1 and Mt = 9.7 mM NH4NO3, Xm = 4710 ±34.4 mg·L-1, PX = 478.9 ±3.8 mg·L-1·d-1 and YX/N = 15.87 ±0.13 mg·mg-1 were obtained. The best conditions for protein content in the biomass (63.2% were not the same as those that maximized cell growth (I = 180 μmol·photons·m-2·s-1 and Mt = 22.5 mM NH4NO3. Based on these results, it is possible to conclude that ammonium nitrate is an interesting alternate nitrogen source for the cultivation of A. platensisin a fed-batch process and could be used for other photosynthetic microorganisms.

  14. A robust feeding strategy to maintain set-point glucose in mammalian fed-batch cultures when input parameters have a large error. (United States)

    Konakovsky, Viktor; Clemens, Christoph; Müller, Markus Michael; Bechmann, Jan; Herwig, Christoph


    Industrial CHO cell cultures run under fed-batch conditions are required to be controlled in particular ranges of glucose, while glucose is constantly consumed and must be replenished by a feed. The most appropriate feeding rate is ideally stoichiometric and adaptive in nature to balance the dynamically changing rate of glucose consumption. However, high errors in biomass and glucose estimation as well as limited knowledge of the true metabolic state challenge the control strategy. In this contribution, we take these errors into account and simulate the output with uncertainty trajectories in silico in order to control glucose concentration. Other than many control strategies, which require parameter estimation, our assumptions are founded on two pillars: (i) first principles and (ii) prior knowledge about the variability of fed-batch CHO cell culture. The algorithm was exposed to an in-silico Design of Experiments (DoE), in which variations of parameters were changed simultaneously, such as clone-specific behavior, precision of equipment and desired control range used. The results demonstrate that our method achieved the target of holding the glucose concentration within an acceptable range. A robust and sufficient level of control could be demonstrated even with high errors for biomass or metabolic state estimation. In a time where blockbuster drugs are queuing up for time slots of their production, this transferable control strategy that is independent of tedious establishment runs may be a decisive advantage for rapid implementation during technology transfer and scale up and decrease in campaign change over time. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:317-336, 2017. © 2017 American Institute of Chemical Engineers.

  15. On-line multi-analyzer monitoring of biomass, glucose and acetate for growth rate control of a Vibrio cholerae fed-batch cultivation. (United States)

    Navrátil, Marián; Norberg, Anna; Lembrén, Lars; Mandenius, Carl-Fredrik


    In situ near-infrared (NIR) spectroscopy and in-line electronic nose (EN) mapping were used to monitor and control a cholera-toxin producing Vibrio cholerae fed-batch cultivation carried out with a laboratory method as well as with a production method. Prediction models for biomass, glucose and acetate using NIR spectroscopy were developed based on spectral identification and partial-least squares (PLS) regression resulting in high correlation to reference data (standard errors of prediction for biomass, glucose and acetate were 0.20 gl(-1), 0.26 gl(-1) and 0.28 gl(-1)). A compensation algorithm for aerated bioreactor disturbances was integrated in the model computation, which in particular improved the prediction by the biomass model. First, the NIR data were applied together with EN in-line data selected by principal component analysis (PCA) for generating a trajectory representation of the fed-batch cultivation. A correlation between the culture progression and EN signals was demonstrated, which proved to be beneficial in monitoring the culture quality. It was shown that a deviation from a normal cultivation behavior could easily be recognized and that the trajectory was able to alarm a bacterial contamination. Second, the NIR data indicated the potential of predicting the concentration of formed cholera toxin with a model prediction error of 0.020 gl(-1). Third, the on-line biomass prediction based on the NIR model was used to control the overflow metabolism acetate formation of the V. cholerae culture. The controller compared actual specific growth rate as estimated from the prediction with the critical acetate formation growth rate, and from that difference adjusted the glucose feed rate.

  16. A multi-pronged investigation into the effect of glucose starvation and culture duration on fed-batch CHO cell culture. (United States)

    Fan, Yuzhou; Jimenez Del Val, Ioscani; Müller, Christian; Lund, Anne Mathilde; Sen, Jette Wagtberg; Rasmussen, Søren Kofoed; Kontoravdi, Cleo; Baycin-Hizal, Deniz; Betenbaugh, Michael J; Weilguny, Dietmar; Andersen, Mikael Rørdam


    In this study, omics-based analysis tools were used to explore the effect of glucose starvation and culture duration on monoclonal antibody (mAb) production in fed-batch CHO cell culture to gain better insight into how these parameters can be controlled to ensure optimal mAb productivity and quality. Titer and N-glycosylation of mAbs, as well as proteomic signature and metabolic status of the production cells in the culture were assessed. We found that the impact of glucose starvation on the titer and N-glycosylation of mAbs was dependent on the degree of starvation during early stationary phase of the fed-batch culture. Higher degree of glucose starvation reduced intracellular concentrations of UDP-GlcNAc and UDP-GalNAc, but increased the levels of UDP-Glc and UDP-Gal. Increased GlcNAc and Gal occupancy correlated well with increased degree of glucose starvation, which can be attributed to the interplay between the dilution effect associated with change in specific productivity of mAbs and the changed nucleotide sugar metabolism. Herein, we also show and discuss that increased cell culture duration negatively affect the maturation of glycans. In addition, comparative proteomics analysis of cells was conducted to observe differences in protein abundance between early growth and early stationary phases. Generally higher expression of proteins involved in regulating cellular metabolism, extracellular matrix, apoptosis, protein secretion and glycosylation was found in early stationary phase. These analyses offered a systematic view of the intrinsic properties of these cells and allowed us to explore the root causes correlating culture duration with variations in the productivity and glycosylation quality of monoclonal antibodies produced with CHO cells.

  17. Changes of the microbial population structure in an overloaded fed-batch biogas reactor digesting maize silage. (United States)

    Kampmann, Kristina; Ratering, Stefan; Geißler-Plaum, Rita; Schmidt, Michael; Zerr, Walter; Schnell, Sylvia


    Two parallel, stable operating biogas reactors were fed with increasing amounts of maize silage to monitor microbial community changes caused by overloading. Changes of microorganisms diversity revealed by SSCP (single strand conformation polymorphism) indicating an acidification before and during the pH-value decrease. The earliest indicator was the appearance of a Methanosarcina thermophila-related species. Diversity of dominant fermenting bacteria within Bacteroidetes, Firmicutes and other Bacteria decreased upon overloading. Some species became dominant directly before and during acidification and thus could be suitable as possible indicator organisms for detection of futurity acidification. Those bacteria were related to Prolixibacter bellariivorans and Streptococcus infantarius subsp. infantarius. An early detection of community shifts will allow better feeding management for optimal biogas production.

  18. Novel strategies for control of fermentation processes

    DEFF Research Database (Denmark)

    Mears, Lisa; Stocks, Stuart; Sin, Gürkan

    Bioprocesses are inherently sensitive to fluctuations in processing conditions and must be tightly regulated to maintain cellular productivity. Industrial fermentations are often difficult to replicate across production sites or between facilities as the small operating differences in the equipment...... of a fermentation. Industrial fermentation processes are typically operated in fed batch mode, which also poses specific challenges for process monitoring and control. This is due to many reasons including non-linear behaviour, and a relatively poor understanding of the system dynamics. It is therefore challenging...

  19. Development of a lipase fermentation process that uses a recombinant Pseudomonas alcaligenes strain

    NARCIS (Netherlands)

    Gerritse, G; Hommes, R.W J; Quax, Wim


    Pseudomonas alcaligenes M-l secretes an alkaline lipase, which has excellent characteristics for the removal of fatty stains under modern washing conditions. A fed-batch fermentation process based on the secretion of the alkaline lipase from P. alcaligenes was developed. Due to the inability of P. a

  20. Mode of operation in fermentation of dilute acid hydrolyzates

    Energy Technology Data Exchange (ETDEWEB)

    Liden, Gunnar [Chalmers Univ. of Technology, Goeteborg (Sweden). Dept. of Chemical Reaction Engineering; Gustafsson, Lena [Goeteborg Univ. (Sweden). Dept. of General and Marine Microbiology


    This report describes work done to find process solution for fermentation of dilute acid hydrolyzates. The work was carried out in the period July 1997 - December 1998. Financial support was initially given by the Swedish National Board of Technical Development, but as of July 1998 support was instead given by the Swedish National Energy Agency. The main findings and achievements of the project are summarized below: 1. It has been found possible to ferment strongly inhibiting hydrolyzates from both spruce and birch using fed-batch technique, i.e. by slowly feeding hydrolyzate to the fermentor. 2. A very promising on-line control algorithm for use in fed-batch fermentation, even with varying hydrolyzate composition, has been developed and tested for a range of different substrates. Previously unreported metabolites, of possible importance in the acute inhibition of glycolysis, have been identified in studies of kinetics of conversion of the inhibitors furfural and HMF in model synthetic media.

  1. Process Control for Production of Human-like Collagen in Fed-batch Culture of Escherichia coli BL 21

    Institute of Scientific and Technical Information of China (English)

    骆艳娥; 范代娣; 马晓轩; 王德伟; 米钰; 花秀夫; 李稳宏


    Recombinant E. coli BL 21 was cultivated in high cell density to produce human-like collagen. The effects of the feeding of nitrogen source, controlled by an auto on/off-feeding mode with two different cycles of 0.5 min and 4 rain intervals, oxygen-enrichment methods and inducement strength on the cell yield and human-like collagen production were investigated. The studies showed that nitrogen source feeding in fast cycle could result in higher human-like collagen production than that in slow cycle; and the feedback regulation of glucose, increase of the pressure of fermentation bioreactor, and supply of oxygen-enriched air could all increase cell yield and human-like collagen production. The effects of inducement strength on protein expression were found important. When OD600 reached 90—100, the cultivation temperature was increased to 42℃ to begin induction for 2—3 h, and then shifted to 39℃ for 5—6 h induction, the cell density and human-like collagen production could reach 96 g·L-1 [DCW (dry cell mass)] and 19.8% (g·g-1 DCW) respectively.

  2. A simple Pichia pastoris fermentation and downstream processing strategy for making recombinant pandemic Swine Origin Influenza a virus Hemagglutinin protein. (United States)

    Athmaram, T N; Singh, Anil Kumar; Saraswat, Shweta; Srivastava, Saurabh; Misra, Princi; Kameswara Rao, M; Gopalan, N; Rao, P V L


    The present Influenza vaccine manufacturing process has posed a clear impediment to initiation of rapid mass vaccination against spreading pandemic influenza. New vaccine strategies are therefore needed that can accelerate the vaccine production. Pichia offers several advantages for rapid and economical bulk production of recombinant proteins and, hence, can be attractive alternative for producing an effective influenza HA based subunit vaccine. The recombinant Pichia harboring the transgene was subjected to fed-batch fermentation at 10 L scale. A simple fermentation and downstream processing strategy is developed for high-yield secretory expression of the recombinant Hemagglutinin protein of pandemic Swine Origin Influenza A virus using Pichia pastoris via fed-batch fermentation. Expression and purification were optimized and the expressed recombinant Hemagglutinin protein was verified by sodium dodecyl sulfate polyacrylamide gel electrophoresis, Western blot and MALDI-TOF analysis. In this paper, we describe a fed-batch fermentation protocol for the secreted production of Swine Influenza A Hemagglutinin protein in the P. pastoris GS115 strain. We have shown that there is a clear relationship between product yield and specific growth rate. The fed-batch fermentation and downstream processing methods optimized in the present study have immense practical application for high-level production of the recombinant H1N1 HA protein in a cost effective way using P. pastoris.

  3. Long-term production of bioethanol in repeated-batch fermentation of microalgal biomass using immobilized Saccharomyces cerevisiae. (United States)

    El-Dalatony, Marwa M; Kurade, Mayur B; Abou-Shanab, Reda A I; Kim, Hoo; Salama, El-Sayed; Jeon, Byong-Hun


    Separate hydrolysis fermentation (SHF) and simultaneous saccharification fermentation (SSF) processes were studied for bioethanol production from microalgal biomass. SSF was selected as an efficient process to enhance the bioethanol yield through repeated-batches using immobilized yeast cells. Combined sonication and enzymatic hydrolysis of Chlamydomonas mexicana generated 10.5 and 8.48g/L of ethanol in SSF and SHF, respectively. Yeast utilized maximum portion of total reducing sugar (TRS) reaching a consumption efficiency of 91-98%. A bioethanol yield of 0.5g/g (88.2% of theoretical yield) and volumetric productivity of 0.22g/L/h was obtained after 48h of SSF. Immobilized yeast cells enabled repetitive production of ethanol for 7 cycles displaying a fermentation efficiency up to 79% for five consecutive cycles. The maximum ethanol production was 9.7g/L in 2nd-4th cycles. A total energy recovery of 85.81% was achieved from microalgal biomass in the form of bioethanol. Repeated-batch SSF demonstrated the possibility of cost-effective bioethanol production.

  4. A novel fed-batch based cultivation method provides high cell-density and improves yield of soluble recombinant proteins in shaken cultures

    Directory of Open Access Journals (Sweden)

    Glumoff Tuomo


    Full Text Available Abstract Background Cultivations for recombinant protein production in shake flasks should provide high cell densities, high protein productivity per cell and good protein quality. The methods described in laboratory handbooks often fail to reach these goals due to oxygen depletion, lack of pH control and the necessity to use low induction cell densities. In this article we describe the impact of a novel enzymatically controlled fed-batch cultivation technology on recombinant protein production in Escherichia coli in simple shaken cultures. Results The enzymatic glucose release system together with a well-balanced combination of mineral salts and complex medium additives provided high cell densities, high protein yields and a considerably improved proportion of soluble proteins in harvested cells. The cultivation method consists of three steps: 1 controlled growth by glucose-limited fed-batch to OD600 ~10, 2 addition of growth boosters together with an inducer providing efficient protein synthesis within a 3 to 6 hours period, and 3 a slow growth period (16 to 21 hours during which the recombinant protein is slowly synthesized and folded. Cell densities corresponding to 10 to 15 g l-1 cell dry weight could be achieved with the developed technique. In comparison to standard cultures in LB, Terrific Broth and mineral salt medium, we typically achieved over 10-fold higher volumetric yields of soluble recombinant proteins. Conclusions We have demonstrated that by applying the novel EnBase® Flo cultivation system in shaken cultures high cell densities can be obtained without impairing the productivity per cell. Especially the yield of soluble (correctly folded proteins was significantly improved in comparison to commonly used LB, Terrific Broth or mineral salt media. This improvement is thought to result from a well controlled physiological state during the whole process. The higher volumetric yields enable the use of lower culture volumes and can

  5. Codon optimization of xylA gene for recombinant glucose isomerase production in Pichia pastoris and fed-batch feeding strategies to fine-tune bioreactor performance. (United States)

    Ata, Özge; Boy, Erdem; Güneş, Hande; Çalık, Pınar


    The objectives of this work are the optimization of the codons of xylA gene from Thermus thermophilus to enhance the production of recombinant glucose isomerase (rGI) in P. pastoris and to investigate the effects of feeding strategies on rGI production. Codons of xylA gene from T. thermophilus were optimized, ca. 30 % of the codons were replaced with those with higher frequencies according to the codon usage bias of P. pastoris, codon optimization resulted in a 2.4-fold higher rGI activity. To fine-tune bioreactor performance, fed-batch bioreactor feeding strategies were designed as continuous exponential methanol feeding with pre-calculated feeding rate based on the pre-determined specific growth rate, and fed-batch methanol-stat feeding. Six feeding strategies were designed, as follows: (S1) continuous exponential methanol- and pulse- sorbitol feeding; (S2) continuous exponential methanol- and peptone- feeding; (S3) continuous exponential methanol- and pulse- mannitol feeding; (S4) continuous exponential methanol- and peptone- feeding and pulse-mannitol feeding; (S5) methanol-stat feeding by keeping methanol concentration at 5 g L(-1); and, (S6) methanol-stat feeding by keeping methanol concentration at 5 g L(-1) and pulse-mannitol feeding. The highest cell and rGI activity was attained as 117 g L(-1) at t = 66 h and 32530 U L(-1) at t = 53 h, in strategy-S5. The use of the co-substrate mannitol does not increase the rGI activity in methanol-stat feeding, where 4.1-fold lower rGI activity was obtained in strategy-S6. The overall cell yield on total substrate was determined at t = 53 h as 0.21 g g(-1) in S5 strategy.

  6. Novel micro-bioreactor high throughput technology for cell culture process development: Reproducibility and scalability assessment of fed-batch CHO cultures. (United States)

    Amanullah, Ashraf; Otero, Jose Manuel; Mikola, Mark; Hsu, Amy; Zhang, Jinyou; Aunins, John; Schreyer, H Brett; Hope, James A; Russo, A Peter


    With increasing timeline pressures to get therapeutic and vaccine candidates into the clinic, resource intensive approaches such as the use of shake flasks and bench-top bioreactors may limit the design space for experimentation to yield highly productive processes. The need to conduct large numbers of experiments has resulted in the use of miniaturized high-throughput (HT) technology for process development. One such high-throughput system is the SimCell platform, a robotically driven, cell culture bioreactor system developed by BioProcessors Corp. This study describes the use of the SimCell micro-bioreactor technology for fed-batch cultivation of a GS-CHO transfectant expressing a model IgG4 monoclonal antibody. Cultivations were conducted in gas-permeable chambers based on a micro-fluidic design, with six micro-bioreactors (MBs) per micro-bioreactor array (MBA). Online, non-invasive measurement of total cell density, pH and dissolved oxygen (DO) was performed. One hundred fourteen parallel MBs (19 MBAs) were employed to examine process reproducibility and scalability at shake flask, 3- and 100-L bioreactor scales. The results of the study demonstrate that the SimCell platform operated under fed-batch conditions could support viable cell concentrations up to least 12 x 10(6) cells/mL. In addition, both intra-MB (MB to MB) as well as intra-MBA (MBA to MBA) culture performance was found to be highly reproducible. The intra-MB and -MBA variability was calculated for each measurement as the coefficient of variation defined as CV (%) = (standard deviation/mean) x 100. The % CV values for most intra-MB and intra-MBA measurements were generally under 10% and the intra-MBA values were slightly lower than those for intra-MB. Cell growth, process parameters, metabolic and protein titer profiles were also compared to those from shake flask, bench-top, and pilot scale bioreactor cultivations and found to be within +/-20% of the historical averages.

  7. Characterization of the AlkS/P-alkB-expression system as an efficient tool for the production of recombinant proteins in Escherichia coli fed-batch fermentations

    NARCIS (Netherlands)

    Makart, Stefan; Heinemann, Matthias; Panke, Sven


    The availability of suitable, well-characterized, and robust expression systems remains an essential requirement for successful metabolic engineering and recombinant protein production. We investigated the suitability of the Pseudomonas putida GPo1-derived AlkS/P-alkB expression system in strictly a

  8. Enhanced growth and recombinant protein production of Escherichia coli by a perfluorinated oxygen carrier in miniaturized fed-batch cultures

    Directory of Open Access Journals (Sweden)

    Neubauer Peter


    Full Text Available Abstract Background Liquid perfluorochemicals (PFCs are interesting oxygen carriers in medicine and biotechnology with a high solubility for oxygen. They have been repeatedly used for improving oxygen transfer into prokaryotic and eukaryotic cell cultures, however their application is still limited. Here we show the great benefit of air/oxygen saturated perfluorodecalin (PFD for high cell density cultivation of Escherichia coli in microwell plates and their positive effect on the soluble production of a correctly folded heterologously expressed alcohol dehydrogenase. Results In EnBase® cultivations the best effect was seen with PFD saturated with oxygen enriched air (appr. 10 μM oxygen per ml when PFD was added at the time of induction. In contrast the effect of PFD was negligible when it was added already at the time of inoculation. Optimisation of addition time and content of loaded oxygen into the PFD resulted in an increased the cell density by 40% compared to control cultures, and correspondingly also the product yield increased, demonstrated at the example of a recombinant alcohol dehydrogenase. Conclusions PFCs are a valuable additive in miniaturized cell culture formats. For production of recombinant proteins in low cell density shaken cultures the addition of oxygen-enriched PFD makes the process more robust, i.e. a high product yield is not any more limited to a very narrow cell density window during which the induction has to be done. The positive effect of PFD was even more obvious when it was added during high cell density cultures. The effect of the PFD phase depends on the amount of oxygen which is loaded into the PFD and which thus is a matter of optimisation.

  9. Quantitative modeling of viable cell density, cell size, intracellular conductivity, and membrane capacitance in batch and fed-batch CHO processes using dielectric spectroscopy. (United States)

    Opel, Cary F; Li, Jincai; Amanullah, Ashraf


    Dielectric spectroscopy was used to analyze typical batch and fed-batch CHO cell culture processes. Three methods of analysis (linear modeling, Cole-Cole modeling, and partial least squares regression), were used to correlate the spectroscopic data with routine biomass measurements [viable packed cell volume, viable cell concentration (VCC), cell size, and oxygen uptake rate (OUR)]. All three models predicted offline biomass measurements accurately during the growth phase of the cultures. However, during the stationary and decline phases of the cultures, the models decreased in accuracy to varying degrees. Offline cell radius measurements were unsuccessfully used to correct for the deviations from the linear model, indicating that physiological changes affecting permittivity were occurring. The beta-dispersion was analyzed using the Cole-Cole distribution parameters Deltaepsilon (magnitude of the permittivity drop), f(c) (critical frequency), and alpha (Cole-Cole parameter). Furthermore, the dielectric parameters static internal conductivity (sigma(i)) and membrane capacitance per area (C(m)) were calculated for the cultures. Finally, the relationship between permittivity, OUR, and VCC was examined, demonstrating how the definition of viability is critical when analyzing biomass online. The results indicate that the common assumptions of constant size and dielectric properties used in dielectric analysis are not always valid during later phases of cell culture processes. The findings also demonstrate that dielectric spectroscopy, while not a substitute for VCC, is a complementary measurement of viable biomass, providing useful auxiliary information about the physiological state of a culture.

  10. Effect of postinduction nutrient feed composition and use of lactose as inducer during production of thermostable xylanase in Escherichia coli glucose-limited fed-batch cultivations. (United States)

    Ramchuran, Santosh O; Holst, Olle; Karlsson, Eva Nordberg


    Escherichia coli is a microorganism routinely used in the production of heterologous proteins. The overexpression of a xylanase (Xyn 10 A Delta NC), which originated from the thermophile Rhodothermus marinus cloned under the control of the strong T7/lac promoter in a defined medium (mAT) using a substrate-limited feed strategy, was however shown to impose a significant metabolic burden on host cells. This resulted in a decreased cell growth rate and ultimately also a decreased target protein production. The investigation hence centers on the effect of some selected nutrient feed additives (amino acid [Cys] or TCA-intermediates [citrate, succinate, malate]) used to relieve the metabolic burden imposed during the feeding and postinduction phases of these glucose-limited fed-batch cultivations. The use of either succinic acid or malic acid as feed-additives resulted in an increase in production of approximately 40% of the heterologous thermostable xylanase. Furthermore, use of lactose as an alternative inducer of the T7/lac promoter was also proven to be a suitable strategy that significantly prolonged the heterologous protein production phase as compared with induction using isopropyl beta-D-thiogalactopyranoside (IPTG).

  11. Efficient production of a thermophilic 2-deoxyribose-5-phosphate aldolase in glucose-limited fed-batch cultivations of Escherichia coli by continuous lactose induction strategy. (United States)

    Pei, Xiao-Lin; Wang, Qiu-Yan; Li, Cheng-Lu; Qiu, Xiao-Feng; Xie, Kai-Lin; Huang, Li-Feng; Wang, An-Ming; Zeng, Zhao-Wu; Xie, Tian


    The production of a thermophilic 2-deoxyribose-5-phosphate aldolases (DERA) in Escherichia coli BL21 under continuous lactose induction strategy was investigated. The process was combined with the exponential feeding method, controlling the feeding rate to maintain the specific growth rate at 0.15 h(-1). The results indicate that the lactose concentration in the feed medium affected directly the expression of the target protein. The use of 50 g/L in the feed medium resulted in the biomass concentration of 39.3 g DCW/L, and an expression level of above 30%, and the maximum final DERA concentration of 16,200 U/L. Furthermore, the acetate concentration remained at a low level in the fed-batch phase, less than 0.5 g/L. In conclusion, combining glucose feeding with lactose induction is a more powerful way to achieve high cell density cultures and to efficiently produce the thermophilic DERA. The results also indicate the potential industrial utility in the scale production of other recombinant proteins.

  12. Production of oleic acid ethyl ester catalyzed by crude rice bran (Oryza sativa lipase in a modified fed-batch system: problem and its solution

    Directory of Open Access Journals (Sweden)

    Indro Prastowo


    Full Text Available A fed-batch system was modified for the enzymatic production of Oleic Acid Ethyl Ester (OAEE using rice bran (Oryza sativa lipase by retaining the substrate molar ratio (ethanol/oleic acid at 2.05: 1 during the reaction. It resulted in an increase in the ester conversion up to 76.8% in the first 6 h of the reaction, and then followed by a decrease from 76.8% to 22.9% in 6 h later. Meanwhile, the production of water in the reaction system also showed a similar trend to the trend of ester production. The water was hypothesized to lead lipase to reverse the reaction which resulted in a decrease in both (water and esters in the last 6 h of the reaction. In order to overcome the problem, zeolite powders (25 and 50 mg/ml were added into the reaction system at 5 h of the reaction. As the result, final ester conversions increased drastically up to 90 - 95.7% (1.17 – 1.24 times. The addition also proved a hypothesis that the water was involved in reducing the ester conversion in the last 6 h of the reaction. Thus, the combination was effective to produce the high final ester conversion.

  13. Influence of ammonium sulphate feeding time on fed-batch Arthrospira (Spirulina) platensis cultivation and biomass composition with and without pH control. (United States)

    Rodrigues, Mayla Santos; Ferreira, Lívia Seno; Converti, Attilio; Sato, Sunao; de Carvalho, João Carlos Monteiro


    Previous work demonstrated that a mixture of NH(4)Cl and KNO(3) as nitrogen source was beneficial to fed-batch Arthrospira (Spirulina) platensis cultivation, in terms of either lower costs or higher cell concentration. On the basis of those results, this study focused on the use of a cheaper nitrogen source mixture, namely (NH(4))(2)SO(4) plus NaNO(3), varying the ammonium feeding time (T=7-15 days), either controlling the pH by CO(2) addition or not. A. platensis was cultivated in mini-tanks at 30°C, 156 μmol photons m(-2) s(-1), and starting cell concentration of 400 mg L(-1), on a modified Schlösser medium. T=13 days under pH control were selected as optimum conditions, ensuring the best results in terms of biomass production (maximum cell concentration of 2911 mg L(-1), cell productivity of 179 mg L(-1)d(-1) and specific growth rate of 0.77 d(-1)) and satisfactory protein and lipid contents (around 30% each).

  14. Optimization of the heterologous production of a Rhizopus oryzae lipase in Pichia pastoris system using mixed substrates on controlled fed-batch bioprocess. (United States)

    Arnau, Carolina; Ramon, Ramon; Casas, Carles; Valero, Francisco


    In this work a systematic study of the influence of methanol set-point and sorbitol feeding rate in fed-batch operation with a Pichia pastoris Mut(s) strain producing Rhizopus oryzae lipase is presented. Different experiments were made at a constant methanol set-point of 0.5, 2 and 4gl(-1), controlled by a predictive algorithm at two different sorbitol feeding rates to assure a constant specific growth rate of 0.01 and 0.02h(-1), by means of a pre-programmed exponential feeding rate strategy. Lipolytic activity, yields, productivity and specific productivity, but also specific growth, consumption and production rates were analyzed showing that the best values were reached when the methanol set-point was 2gl(-1) with a low influence of the constant specific growth rate tested. The sorbitol addition as a co-substrate during the induction phase avoids the severe decrease of the specific production rate obtained when methanol was used as a sole carbon source and it permits to achieve higher ROL production.

  15. Phototrophic cultivation of a thermo-tolerant Desmodesmus sp. for lutein production: effects of nitrate concentration, light intensity and fed-batch operation. (United States)

    Xie, Youping; Ho, Shih-Hsin; Chen, Ching-Nen Nathan; Chen, Chun-Yen; Ng, I-Son; Jing, Ke-Ju; Chang, Jo-Shu; Lu, Yinghua


    Four indigenous thermo-tolerant Desmodesmus sp. strains were examined for their ability to produce lutein. Among them, Desmodesmus sp. F51 was the best strain for this purpose. The medium composition, nitrate concentration and light intensity were manipulated to improve the phototrophic growth and lutein production of Desmodesmus sp. F51. It was found that a nitrogen-sufficient condition was required for lutein accumulation, while a high light intensity enhanced cell growth but caused a decrease in the lutein content. The best cell growth and lutein production occurred when the light intensity and initial nitrate concentration were 600 μmol/m(2)/s and 8.8 mM, respectively. The fed-batch cultivation strategy was shown to further improve lutein production. The highest lutein productivity (3.56±0.10 mg/L/d) and content (5.05±0.20 mg/g) were obtained when pulse-feeding of 2.2 mM nitrate was employed. This study demonstrated the potential of using Desmodesmus sp. F51 as a lutein producer in practical applications.

  16. Comparative study of four fed-batch propagation systems of beer yeast Estudio comparativo de cuatro sistemas de propagación de levadura cervecera por lote alimentado

    Directory of Open Access Journals (Sweden)

    Caicedo L.


    Full Text Available Beer yeast was propagated using batch culture, and a mathematical model was fitted to the resulting data. Intermittent, continuousintermittent, and high-density fed-batch techniques were used. The highest cell yield was found using the high density technique. Simulation also unveiled an effect of the feeding technique on cellular growth rate and yield. The high density technique increased the stoichiometric factor Yx/s.Se realizó la propagación por lotes de levadura cervecera y con base en estos resultados se ajustó un modelo matemático. Se llevaron a cabo cuatro fermentaciones con diferentes técnicas de alimentación y se confrontaron los resultados mediante simulación. Las técnicas estudiadas fueron alimentación puntual, alimentación continua-puntual y de alta densidad. La mayor concentración celular y factor de propagación se presentó con el cultivo de alta densidad. La simulación demostró que la alimentación continua o puntual afecta el comportamiento celular, ya sea sobre la velocidad específica y/o el factor estequiométrico Yx/s . Se encontró que la técnica de alta densidad aumenta el factor estequiométrico Yx/s.

  17. Combined effect of the methanol utilization (Mut) phenotype and gene dosage on recombinant protein production in Pichia pastoris fed-batch cultures. (United States)

    Cos, Oriol; Serrano, Alicia; Montesinos, José Luis; Ferrer, Pau; Cregg, James M; Valero, Francisco


    An important number of heterologous proteins have been produced in the methylotrophic yeast Pichia pastoris using the alcohol oxidase promoter. Two factors that drastically influence protein production and cultivation process development in this system are gene dosage and methanol assimilation capacity of the host strain (Mut phenotype). Using a battery of four strains which secrete a Rhizopus oryzae lipase (ROL), the combined effects of gene dosage and Mut phenotype on recombinant protein production in Pichia pastoris was studied in fed-batch cultures. Regarding the effect of phenotype, the specific productivity and the Y(P/X) were 1.29- and 2.34-fold higher for Mut(s)ROL single copy strain than for Mut+ROL single copy strain. On the contrary, the productivity of Mut+ROL single copy strain was 1.34-fold higher than Mut(s)ROL single copy strain. An increase in ROL gene dosage seems to negatively affect cell's performance in bioreactor cultures, particularly in Mut(s) strains. Overall, the Mut(s) strain may be still advantageous to use because it allows for easier process control strategies.

  18. Dynamics of hydrogen-producing bacteria in a repeated batch fermentation process using lake sediment as inoculum. (United States)

    Romano, Stefano; Paganin, Patrizia; Varrone, Cristiano; Tabacchioni, Silvia; Chiarini, Luigi


    In this study, we evaluated the effectiveness of lake sediment as inoculum for hydrogen production through dark fermentation in a repeated batch process. In addition, we investigated the effect of heat treatment, applied to enrich hydrogen-producing bacteria, on the bacterial composition and metabolism. Denaturing gradient gel electrophoresis and molecular cloning, both performed using the 16S rDNA gene as target gene, were used to monitor the structure of the bacterial community. Hydrogen production and bacterial metabolism were analysed via gas chromatography and high-performance liquid chromatography. Both treated and non-treated inocula were able to produce high amounts of hydrogen. However, statistical analysis showed a clear difference in their bacterial composition and metabolism. The heat treatment favoured the growth of different Clostridia sp., in particular of Clostridium bifermentans, allowing the production of a constant amount of hydrogen over prolonged time. These cultures showed both butyrate and ethanol fermentation types. Absence of heat treatment allowed species belonging to the genera Bacillus, Sporolactobacillus and Massilia to outgrow Clostridia sp. with a reduction in hydrogen production and a significant metabolic change. Our data indicate that lake sediment harbours bacteria that can efficiently produce hydrogen over prolonged fermentation time. Moreover, we could show that the heat treatment stabilizes the bacterial community composition and the hydrogen production.

  19. Novel approach of high cell density recombinant bioprocess development: Optimisation and scale-up from microlitre to pilot scales while maintaining the fed-batch cultivation mode of E. coli cultures

    Directory of Open Access Journals (Sweden)

    Rimšeliene Renata


    Full Text Available Abstract Background Bioprocess development of recombinant proteins is time consuming and laborious as many factors influence the accumulation of the product in the soluble and active form. Currently, in most cases the developmental line is characterised by a screening stage which is performed under batch conditions followed by the development of the fed-batch process. Performing the screening already under fed-batch conditions would limit the amount of work and guarantee that the selected favoured conditions also work in the production scale. Results Here, for the first time, high throughput multifactorial screening of a cloning library is combined with the fed-batch technique in 96-well plates, and a strategy is directly derived for scaling to bioreactor scale. At the example of a difficult to express protein, an RNase inhibitor, it is demonstrated that screening of various vector constructs and growth conditions can be performed in a coherent line by (i applying a vector library with promoters and ribosome binding sites of different strength and various fusion partners together with (ii an early stage use of the fed-batch technology. It is shown that the EnBase® technology provides an easy solution for controlled cultivation conditions in the microwell scale. Additionally the high cell densities obtained provide material for various analyses from the small culture volumes. Crucial factors for a high yield of the target protein in the actual case were (i the fusion partner, (ii the use of of a mineral salt medium together with the fed-batch technique, and (iii the preinduction growth rate. Finally, it is shown that the favorable conditions selected in the microwell plate and shake flask scales also work in the bioreactor. Conclusions Cultivation media and culture conditions have a major impact on the success of a screening procedure. Therefore the application of controlled cultivation conditions is pivotal. The consequent use of fed-batch

  20. Software sensor for primary metabolite production case of alcoholic fermentation

    Energy Technology Data Exchange (ETDEWEB)

    Roux, G.; Dahhou, B.; Queinnec, I. [Centre National de la Recherche Scientifique (CNRS), 31 - Toulouse (France)]|[Institut National des Sciences Appliquees (INSA), 31 - Toulouse (France); Goma, G. [Institut National des Sciences Appliquees (INSA), 31 - Toulouse (France)


    This paper investigate the application of an observer for state and parameter estimation to batch, continuous and fed batch fermentations for alcohol production taken as model for a primary metabolite production. This observer is provided to palliate the lack of suitable sensors for on-line biomass and ethanol concentrations measurements and to estimate the time varying specific growth rate. Estimates are obtained from an interlaced structure filter based on a `modified extended Kalman filter` by using on-line measurements of carbon dioxide outflow rate and substrate concentration. The filter algorithm was tested during batch, continuous and fed batch fermentation processes. The filter behaviour observed in the experiments gives good results with an agreement theory/practice. (authors) 18 refs.

  1. The study of glycerol-based fermentation and broth downstream by nanofiltration

    Directory of Open Access Journals (Sweden)

    Gryta Marek


    Full Text Available In this work, the glycerol fermentation was carried out using Citrobacter freundii bacteria. The influence of glycerol and metabolites concentrations, and the pH changes on the efficiency of 1,3-propanediol production, during batch and fed-batch processes, was presented. The nanofiltration was used for the separation of obtained post-fermentation solutions. The resulted 1,3-PD solutions were significantly desalted, which may facilitate further downstream processes during 1,3-PD production.

  2. Performance of the auxotrophic Saccharomyces cerevisiae BY4741 as host for the production of IL-1β in aerated fed-batch reactor: role of ACA supplementation, strain viability, and maintenance energy

    Directory of Open Access Journals (Sweden)

    Zueco Jesus


    Full Text Available Abstract Background Saccharomyces cerevisiae BY4741 is an auxotrophic commonly used strain. In this work it has been used as host for the expression and secretion of human interleukin-1β (IL1β, using the cell wall protein Pir4 as fusion partner. To achieve high cell density and, consequently, high product yield, BY4741 [PIR4-IL1β] was cultured in an aerated fed-batch reactor, using a defined mineral medium supplemented with casamino acids as ACA (auxotrophy-complementing amino acid source. Also the S. cerevisiae mutant BY4741 Δyca1 [PIR4-IL1β], carrying the deletion of the YCA1 gene coding for a caspase-like protein involved in the apoptotic response, was cultured in aerated fed-batch reactor and compared to the parental strain, to test the effect of this mutation on strain robustness. Viability of the producer strains was examined during the runs and a mathematical model, which took into consideration the viable biomass present in the reactor and the glucose consumption for both growth and maintenance, was developed to describe and explain the time-course evolution of the process for both, the BY4741 parental and the BY4741 Δyca1 mutant strain. Results Our results show that the concentrations of ACA in the feeding solution, corresponding to those routinely used in the literature, are limiting for the growth of S. cerevisiae BY4741 [PIR4-IL1β] in fed-batch reactor. Even in the presence of a proper ACA supplementation, S. cerevisiae BY4741 [PIR4-IL1β] did not achieve a high cell density. The Δyca1 deletion did not have a beneficial effect on the overall performance of the strain, but it had a clear effect on its viability, which was not impaired during fed-batch operations, as shown by the kd value (0.0045 h-1, negligible if compared to that of the parental strain (0.028 h-1. However, independently of their robustness, both the parental and the Δyca1 mutant ceased to grow early during fed-batch runs, both strains using most of the

  3. Application of simple fed-batch technique to high-level secretory production of insulin precursor using Pichia pastoris with subsequent purification and conversion to human insulin

    Directory of Open Access Journals (Sweden)

    Chugh Dipti


    Full Text Available Abstract Background The prevalence of diabetes is predicted to rise significantly in the coming decades. A recent analysis projects that by the year 2030 there will be ~366 million diabetics around the world, leading to an increased demand for inexpensive insulin to make this life-saving drug also affordable for resource poor countries. Results A synthetic insulin precursor (IP-encoding gene, codon-optimized for expression in P. pastoris, was cloned in frame with the Saccharomyces cerevisiae α-factor secretory signal and integrated into the genome of P. pastoris strain X-33. The strain was grown to high-cell density in a batch procedure using a defined medium with low salt and high glycerol concentrations. Following batch growth, production of IP was carried out at methanol concentrations of 2 g L-1, which were kept constant throughout the remaining production phase. This robust feeding strategy led to the secretion of ~3 gram IP per liter of culture broth (corresponding to almost 4 gram IP per liter of cell-free culture supernatant. Using immobilized metal ion affinity chromatography (IMAC as a novel approach for IP purification, 95% of the secreted product was recovered with a purity of 96% from the clarified culture supernatant. Finally, the purified IP was trypsin digested, transpeptidated, deprotected and further purified leading to ~1.5 g of 99% pure recombinant human insulin per liter of culture broth. Conclusions A simple two-phase cultivation process composed of a glycerol batch and a constant methanol fed-batch phase recently developed for the intracellular production of the Hepatitis B surface antigen was adapted to secretory IP production. Compared to the highest previously reported value, this approach resulted in an ~2 fold enhancement of IP production using Pichia based expression systems, thus significantly increasing the efficiency of insulin manufacture.

  4. Enhancing toxic protein expression in Escherichia coli fed-batch culture using kinetic parameters: Human granulocyte-macrophage colony-stimulating factor as a model system. (United States)

    Khasa, Yogender Pal; Khushoo, Amardeep; Mukherjee, Krishna Jyoti


    The kinetics of recombinant human granulocyte-macrophage colony-stimulating factor (hGM-CSF) expression was studied under the strong T7 promoter in continuous culture of Escherichia coli using complex medium to design an optimum feeding strategy for high cell density cultivation. Continuous culture studies were done at different dilution rates and the growth and product formation profiles were monitored post-induction. Recombinant protein expression was in the form of inclusion bodies with a maximum specific product formation rate (q(p)) of 63.5 mg g(-1) DCW h(-1) at a dilution rate (D) of 0.3 h(-1). The maximum volumetric product concentration achieved at this dilution rate was 474 mg l(-1), which translated a ~1.4 and ~1.75 folds increase than the values obtained at dilution rates of 0.2 h(-1) and 0.4 h(-1) respectively. The specific product yield (Y(P/x)) peaked at 138 mg g(-1) DCW, demonstrating a ~1.6 folds increase in the values obtained at other dilution rates. A drop in q(p) was observed within 5-6 h of induction at all the dilution rates, possibly due to protein toxicity and metabolic stress associated with protein expression. The data from the continuous culture studies allowed us to design an optimal feeding strategy and induction time in fed-batch cultures which resulted in a maximum product concentration of 3.95 g l(-1) with a specific hGM-CSF yield (Y(P/x)) of 107 mg g(-1) DCW.

  5. Influence of pH, temperature, and urea molar flowrate on Arthrospira platensis fed-batch cultivation: a kinetic and thermodynamic approach. (United States)

    Sánchez-Luna, Luis Dante; Bezerra, Raquel Pedrosa; Matsudo, Marcelo Chuei; Sato, Sunao; Converti, Attilio; de Carvalho, João Carlos Monteiro


    Arthrospira platensis was cultivated photoautotrophically at 6.0 klux light intensity in 5.0-L open tanks, using a mineral medium containing urea as nitrogen source. Fed-batch experiments were performed at constant flowrate. A central composite factorial design combined to response surface methodology (RSM) was utilized to determine the relationship between the selected response variables (cell concentration after 10 days, X(m), cell productivity, P(X), and nitrogen-to-cell conversion factor, Y(X/N)) and codified values of the independent variables (pH, temperature, T, and urea flowrate, K). By applying the quadratic regression analysis, the equations describing the behaviors of these responses as simultaneous functions of the selected independent variables were determined, and the conditions for X(m) and P(X) optimization were estimated (pH 9.5, T = 29 degrees C, and K = 0.551 mM/day). The experimental data obtained under these conditions (X(m) = 749 mg/L; P(X) = 69.9 mg/ were very close to the estimated ones (X(m) = 721 mg/L; P(X) = 67.1 mg/ Additional cultivations were carried out under the above best conditions of pH control and urea flowrate at variable temperature. Consistently with the results of RSM, the best growth temperature was 29 degrees C. The maximum specific growth rates at different temperatures were used to estimate the thermodynamic parameters of growth (DeltaH* = 59.3 kJ/mol; DeltaS* = -0.147 kJ/mol.K; DeltaG* = 103 kJ/mol) and its thermal inactivation (DeltaH(D) (o) = 72.0 kJ/mol; DeltaS(D) (o) = 0.144 kJ/mol.K; DeltaG(D) (o) = 29.1 kJ/mol).

  6. Life-cycle and cost of goods assessment of fed-batch and perfusion-based manufacturing processes for mAbs. (United States)

    Bunnak, Phumthep; Allmendinger, Richard; Ramasamy, Sri V; Lettieri, Paola; Titchener-Hooker, Nigel J


    Life-cycle assessment (LCA) is an environmental assessment tool that quantifies the environmental impact associated with a product or a process (e.g., water consumption, energy requirements, and solid waste generation). While LCA is a standard approach in many commercial industries, its application has not been exploited widely in the bioprocessing sector. To contribute toward the design of more cost-efficient, robust and environmentally-friendly manufacturing process for monoclonal antibodies (mAbs), a framework consisting of an LCA and economic analysis combined with a sensitivity analysis of manufacturing process parameters and a production scale-up study is presented. The efficiency of the framework is demonstrated using a comparative study of the two most commonly used upstream configurations for mAb manufacture, namely fed-batch (FB) and perfusion-based processes. Results obtained by the framework are presented using a range of visualization tools, and indicate that a standard perfusion process (with a pooling duration of 4 days) has similar cost of goods than a FB process but a larger environmental footprint because it consumed 35% more water, demanded 17% more energy, and emitted 17% more CO2 than the FB process. Water consumption was the most important impact category, especially when scaling-up the processes, as energy was required to produce process water and water-for-injection, while CO2 was emitted from energy generation. The sensitivity analysis revealed that the perfusion process can be made more environmentally-friendly than the FB process if the pooling duration is extended to 8 days. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1324-1335, 2016. © 2016 The Authors Biotechnology Progress published by Wiley Periodicals, Inc. on behalf of American Institute of Chemical Engineers.


    Directory of Open Access Journals (Sweden)

    Folly R.


    Full Text Available A fed-batch alcohol fermentation on a pilot plant scale with a digital supervisory control system was evaluated as an experimental application case study of an adaptive controller. The verification of intrinsically dynamic variations in the characteristics of the fermentation, observed in previous work, showed the necessity of an adaptive control strategy for controller parameter tuning in order to adjust the changes in the specific rates of consumption, growth and product formation during the process. Satisfactory experimental results were obtained for set-point variations and sugar feed concentration load changes in the manipulated inlet flow to the fermenter

  8. Cultivo mixotrófico da microalga Spirulina platensis em batelada alimentada Mixotrophic growth of Spirulina platensis in fed-batch mode

    Directory of Open Access Journals (Sweden)

    Adriana Muliterno


    Full Text Available A Spirulina platensis tem sido estudada devido a seu alto valor protéico, digestibilidade e por apresentar quantidades significativas de ácidos graxos poliinsaturados, vitaminas, fenólicos e ficocianina, podendo ser utilizada na alimentação humana. A utilização de nutrientes de baixo custo é um fator importante na produção da cianobactéria por possibilitar a redução de custos de processo. Objetivou-se com este trabalho estudar o cultivo mixotrófico da S. platensis por meio da adição de uma fonte orgânica de carbono (glicose em modo bateladaalimentada. Foi utilizado um Planejamento Fatorial Completo 2³ para o cultivo e as variáveis de estudo foram a concentração de glicose (0,5 gL-1 e 1,0 gL-1, a diluição do meio Zarrouk (50% e 75% e a iluminância (1800 lux e 3000 lux. A concentração celular máxima obtida foi de 5,38 gL-1 com uma velocidade específica máxima de crescimento de 0,0063 h-1, nas condições de 0,5 gL-1 de glicose, diluição do meio de 75% e iluminância de 3000 lux.The cyanobacterium Spirulina platensis has been studied due to its high content (~65% of highly digestible protein as well as significant amounts of polyunsaturated fatty acids, phenolics, vitamins, minerals and phycocyanin which could be useful in the human nutrition. The use of nutrients of low costs in the cyanobacterium growth could reduce the costs of production. We studied the fed-batch mixotrophic growth of the S. platensis in Zarrouk's medium with glucose (0.5 gL-1 and 1.0 gL-1 as carbon source and also investigated the effects of dilution (50% and 75%, with water and illumination (1,800 lux and 3,000 lux using a 2³ factorial design. The maximum celular concentration of 5.38 gL-1 and maximum specific growth rate of 0.0063 h-1 were obtained with a glucose concentration of 0.5 gL-1, 50% dilution and 1800 lux of illuminance.

  9. Determining the impacts of fermentative bacteria on wollastonite dissolution kinetics. (United States)

    Salek, S S; Kleerebezem, R; Jonkers, H M; Voncken, J H L; van Loosdrecht, M C M


    Silicate minerals can be a source of calcium and alkalinity, enabling CO2 sequestration in the form of carbonates. For this to occur, the mineral needs to be first dissolved in an acidifying process such as the biological process of anaerobic fermentation. In the present study, the main factors which govern the dissolution process of an alkaline silicate mineral (wollastonite, CaSiO3) in an anaerobic fermentation process were determined. Wollastonite dissolution kinetics was measured in a series of chemical batch experiments in order to be able to estimate the required amount of alkaline silicate that can neutralize the acidifying fermentation process. An anaerobic fermentation of glucose with wollastonite as the neutralizing agent was consequently performed in a fed-batch reactor. Results of this experiment were compared with an abiotic (control) fed-batch reactor in which the fermentation products (i.e. organic acids and alcohols) were externally supplied to the system at comparable rates and proportions, in order to provide chemical conditions similar to those during the biotic (fermentation) experiment. This procedure enabled us to determine whether dissolution of wollastonite was solely enhanced by production of organic acids or whether there were other impacts that fermentative bacteria could have on the mineral dissolution rate. The established pH profiles, which were the direct indicator of the dissolution rate, were comparable in both experiments suggesting that the mineral dissolution rate was mostly influenced by the quantity of the organic acids produced.

  10. Production of poly(malic acid) from sugarcane juice in fermentation by Aureobasidium pullulans: Kinetics and process economics. (United States)

    Wei, Peilian; Cheng, Chi; Lin, Meng; Zhou, Yipin; Yang, Shang-Tian


    Poly(β-l-malic acid) (PMA) is a biodegradable polymer with many potential biomedical applications. PMA can be readily hydrolyzed to malic acid (MA), which is widely used as an acidulant in foods and pharmaceuticals. PMA production from sucrose and sugarcane juice by Aureobasidium pullulans ZX-10 was studied in shake-flasks and bioreactors, confirming that sugarcane juice can be used as an economical substrate without any pretreatment or nutrients supplementation. A high PMA titer of 116.3g/L and yield of 0.41g/g were achieved in fed-batch fermentation. A high productivity of 0.66g/L·h was achieved in repeated-batch fermentation with cell recycle. These results compared favorably with those obtained from glucose and other biomass feedstocks. A process economic analysis showed that PMA could be produced from sugarcane juice at a cost of $1.33/kg, offering a cost-competitive bio-based PMA for industrial applications. Copyright © 2016 Elsevier Ltd. All rights reserved.

  11. L-Lactic acid production by combined utilization of agricultural bioresources as renewable and economical substrates through batch and repeated-batch fermentation of Enterococcus faecalis RKY1. (United States)

    Reddy, Lebaka Veeranjaneya; Kim, Young-Min; Yun, Jong-Sun; Ryu, Hwa-Won; Wee, Young-Jung


    Enterococcus faecalis RKY1 was used to produce l-lactic acid from hydrol, soybean curd residues (SCR), and malt. Hydrol was efficiently metabolized to l-lactic acid with optical purity of >97.5%, though hydrol contained mixed sugars such as glucose, maltose, maltotriose, and maltodextrin. Combined utilization of hydrol, SCR, and malt was enough to sustain lactic acid fermentation by E. faecalis RKY1. In order to reduce the amount of nitrogen sources and product inhibition, cell-recycle repeated-batch fermentation was employed, where a high cell mass (26.3g/L) was obtained. Lactic acid productivity was improved by removal of lactic acid from fermentation broth by membrane filtration and by linearly increased cell density. When the total of 10 repeated-batch fermentations were carried out using 100g/L hydrol, 150g/L SCR hydrolyzate, and 20g/L malt hydrolyzate as the main nutrients, lactic acid productivity was increased significantly from 3.20g/L/h to 6.37g/L/h.

  12. L-lactic acid production by Lactobacillus casei fermentation with corn steep liquor-supplemented acid-hydrolysate of soybean meal. (United States)

    Li, Zheng; Ding, Shaofeng; Li, Zhaopeng; Tan, Tianwei


    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.

  13. Aeration-Controlled Formation of Acid in Heterolactic Fermentations

    DEFF Research Database (Denmark)

    Adler-Nissen, Jens


    Controlled aeration of Leuconostoc mesenteroides was studied as a possible mechanism for control of the formation of acetic acid, a metabolite of major influence on the taste of lactic fermented foods. Fermentations were carried out in small scale in a medium in which growth was limited...... by the buffer capacity only. Ethanol and acetic acid formed during the fermentation were analyzed by rapid head space gas chromatography, and the ratio of the molar concentrations of these two volatiles quantitatively predicted the balance between the formation of acetic acid and lactic acid. The oxygen...... fermentation processes should be analyzed as fed-batch fermentations with oxygen as the limiting substrate. Addition of fructose in limited amounts leads to the formation of one half mole of acetic acid for each mole fructose, thus offering an alternative mechanism for controlling acetic acid formation....

  14. Enhancement of fermentative hydrogen production in an extreme-thermophilic (70°C) mixed-culture environment by repeated batch cultivation. (United States)

    Lu, Wenjing; Fan, Gaoyuan; Zhao, Chenxi; Wang, Hongtao; Chi, Zifang


    Repeated batch cultivation was applied to enrich hydrogen fermentative microflora under extreme-thermophilic (70°C) environment. Initial inoculums received from a hydrogen producing reactor fed with organic fraction of household solid wastes. In total seven transfers was conducted and maximum hydrogen yield reached 296 ml H(2)/g (2.38 mol/mol) glucose and 252 ml H(2)/g (2.03 mol/mol) for 1 and 2 g/l glucose medium, respectively. It was found that hydrogen production was firstly decreased and got increased gradually from third generation. Acetate was found to be the main metabolic by-product in all batch cultivation. Furthermore, the diversity of bacterial community got decreased after repeated batch cultivation. It was proved that repeated batch cultivation was a good method to enhance the hydrogen production by enriching the mixed cultures of dominant species.

  15. Ethanol fermentation integrated with PDMS composite membrane: An effective process. (United States)

    Fu, Chaohui; Cai, Di; Hu, Song; Miao, Qi; Wang, Yong; Qin, Peiyong; Wang, Zheng; Tan, Tianwei


    The polydimethylsiloxane (PDMS) membrane, prepared in water phase, was investigated in separation ethanol from model ethanol/water mixture and fermentation-pervaporation integrated process. Results showed that the PDMS membrane could effectively separate ethanol from model solution. When integrated with batch ethanol fermentation, the ethanol productivity was enhanced compared with conventional process. Fed-batch and continuous ethanol fermentation with pervaporation were also performed and studied. 396.2-663.7g/m(2)h and 332.4-548.1g/m(2)h of total flux with separation factor of 8.6-11.7 and 8-11.6, were generated in the fed-batch and continuous fermentation with pervaporation scenario, respectively. At the same time, high titre ethanol production of ∼417.2g/L and ∼446.3g/L were also achieved on the permeate side of membrane in the two scenarios, respectively. The integrated process was environmental friendly and energy saving, and has a promising perspective in long-terms operation.

  16. Learning control of fermentation process with an improved DHP algorithm☆

    Institute of Scientific and Technical Information of China (English)

    Dazi Li; Ningjia Meng; Tianheng Song


    Control of the fed-batch ethanol fermentation processes to produce maximum product ethanol is one of the key issues in the bioreactor system. However, ethanol fermentation processes exhibit complex behavior and nonlinear dynamics with respect to the cel mass, substrate, feed-rate, etc. An improved dual heuristic program-ming algorithm based on the least squares temporal difference with gradient correction (LSTDC) algorithm (LSTDC-DHP) is proposed to solve the learning control problem of a fed-batch ethanol fermentation process. As a new algorithm of adaptive critic designs, LSTDC-DHP is used to realize online learning control of chemical dynamical plants, where LSTDC is commonly employed to approximate the value functions. Application of the LSTDC-DHP algorithm to ethanol fermentation process can realize efficient online learning control in continuous spaces. Simulation results demonstrate the effectiveness of LSTDC-DHP, and show that LSTDC-DHP can obtain the near-optimal feed rate trajectory faster than other-based algorithms.

  17. Comparative study on processes of simultaneous saccharification and fermentation with high solid concentration for cellulosic ethanol production%高底物浓度纤维乙醇同步糖化发酵工艺的比较

    Institute of Scientific and Technical Information of China (English)

    常春; 王铎; 王林风; 马晓建


    The effects of various simultaneous saccharification fermentation (SSF) technologies on ethanol yield from cellulose were investigated. Using steam-exploded corn stalks as raw materials, five SSF technologies, including traditional SSF, SSF combined with preliminary enzymatic hydrolysis, fed-batch SSF coupled with preliminary enzymatic hydrolysis, SSF united with preliminary enzymatic hydrolysis coupling with vacuum separation and fed-batch SSF associated with preliminary enzymatic hydrolysis coupling with vacuum separation, were used for decomposition of the stalks. By comparing the fermentation process with higher solid concentration including 15% (mass) and 30% (mass), it was found that the fed-batch SSF combined with preliminary enzymatic hydrolysis is an effective way for raising substrate concentration. However, higher substrate concentration may lead to decrease of final ethanol yield. Moreover, SSF coupled with vacuum separation can weaken the product inhibition, and increase the ethanol yield. The ethanol productivity by the fed-batch SSF associated with preliminary enzymatic hydrolysis coupling with vacuum separation is 0. 40 g · L-1 · h-1, the highest value obtained, indicating that this technology can be a potential new technology.

  18. Commercialization of a novel fermentation concept. (United States)

    Mazumdar-Shaw, Kiran; Suryanarayan, Shrikumar


    Fermentation is the core of biotechnology where current methodologies span across technologies based on the use of either solid or liquid substrates. Traditionally, solid substrate fermentation technologies have been the widely practiced in the Far East to manufacture fermented foods such as soya sauce, sake etc. The Western World briefly used solid substrate fermentation for the manufacture of antibiotics and enzymes but rapidly replaced this technology with submerged fermentation which proved to be a superior technology in terms of automation, containment and large volume fermentation. Biocon India developed its enzyme technology based on solid substrate fermentation as a low-cost, low-energy option for the production of specialty enzymes. However, the limitations of applying solid substrate fermentation to more sophisticated biotechnology products as well as large volume fermentations were recognized by Biocon India as early as 1990 and the company embarked on a 8 year research and development program to develop a novel bioreactor capable of conducting solid substrate fermentation with comparable levels of automation and containment as those practiced by submerged fermentation. In addition, the novel technology enabled fed-batch fermentation, in situ extraction and other enabling features that will be discussed in this article. The novel bioreactor was christened the "PlaFractor" (pronounced play-fractor). The next level of research on this novel technology is now focused on addressing large volume fermentation. This article traces the evolution of Biocon India's original solid substrate fermentation to the PlaFractor technology and provides details of the scale-up and commercialization processes that were involved therein. What is also apparent in the article is Biocon India's commercially focused research programs and the perceived need to be globally competitive through low costs of innovation that address, at all times, processes and technologies that

  19. Optimal control of switched systems arising in fermentation processes

    CERN Document Server

    Liu, Chongyang


    The book presents, in a systematic manner, the optimal controls under different mathematical models in fermentation processes. Variant mathematical models – i.e., those for multistage systems; switched autonomous systems; time-dependent and state-dependent switched systems; multistage time-delay systems and switched time-delay systems – for fed-batch fermentation processes are proposed and the theories and algorithms of their optimal control problems are studied and discussed. By putting forward novel methods and innovative tools, the book provides a state-of-the-art and comprehensive systematic treatment of optimal control problems arising in fermentation processes. It not only develops nonlinear dynamical system, optimal control theory and optimization algorithms, but can also help to increase productivity and provide valuable reference material on commercial fermentation processes.

  20. Solid phase fermentation of Helianthus tuberosus for ethanol

    Energy Technology Data Exchange (ETDEWEB)

    Baerwald, G.; Hamad, S.H.


    The direct fermentation of pure inulin and hammer mill crushed Helianthus tuberosus tubers (topinambur, Jerusalem artichoke) was studied using two heat-tolerant yeasts, namely Kluyveromyces marxianus and Candida kefyr. A Saccharomyces cerevisiae was included in the study so as to compare the yields of these two yeasts with that of a commercial distiller's yeast. The inulin fermentation was carried out in an 18-L bioreactor using the fed-batch and the batch-fermentation methods. The final ethanol concentration was 6.1% (L/L) which represents 82% of the theoretical yield. Commercial scale experiments with hammer mill crushed tubers gave yields lower than those found in the laboratory: 69% of the theoretical yield for direct fermentation without enzyme addition, and about 91% when cellolytic enzymes were added.


    Directory of Open Access Journals (Sweden)

    Abdullah Mochamad Busairi


    Full Text Available Pineapple juice waste contains valuable components, which are mainly sucrose, glucose, and fructose. Recently, lactic acid has been considered to be an important raw material for the production of biodegradable lactide polymer. The fermentation experiments were carried out in a 3 litres fermentor (Biostat B Model under anaerobic condition with stirring speed of 50 rpm, temperature at 40oC, and pH of 6.00. Effect of feed concentration on lactic acid production, bacterial growth, substrate utilisation and productivity was studied. The results obtained from fed- batch culture fermentation showed that the maximum lactic acid productivity was 0.44 g/L.h for feed concentration of 90 g/L at 48 hours. Whereas the lactic acid productivity obtained from fed-batch culture was twice and half fold higher than that of batch culture productivity.  Buangan jus nanas mengandung komponen yang berharga terutama sukrosa, glukosa, dan fruktosa. Asam laktat adalah bahan baku yang terbaru dan penting untuk dibuat sebagai polimer laktat yang dapat terdegradasi oleh lingkungan. Percobaan dilakukan pada fermentor 3 liter (Model Biostat B di bawah kondisi anaerob dengan kecepatan pengadukan 50 rpm, temperatur 40oC, dan pH 6,00. Pengaruh konsentrasi umpan terhadap produksi asam laktat, pertumbuhan mikroba, pengggunaan substrat dan produktivitas telah dipelajari. Hasil yang didapatkan pada fermentasi dengan menggunakan sistem fed-batch menunjukkan bahwa produktivitas asam laktat maksimum adalah 0.44 g/L,jam dengan konsentrasi umpan, 90 g/L pada waktu 48 jam. Bahkan produktivitas asam laktat yang didapat pada kultur fed-batch lebih tinggi 2,5 kali dari pada proses menggunakan sistem batch

  2. Improving lactic acid productivity from wheat straw hydrolysates by membrane integrated repeated batch fermentation under non-sterilized conditions

    DEFF Research Database (Denmark)

    Zhang, Yuming; Chen, Xiangrong; Qi, Benkun


    Bacillus coagulans IPE22 was used to produce lactic acid (LA) from mixed sugar and wheat straw hydrolysates, respectively. All fermentations were conducted under non-sterilized conditions and sodium hydroxide was used as neutralizing agent to avoid the production of insoluble CaSO4. In order to e...

  3. 分批补料及缺氮培养对小球藻油脂产量的影响%Effects of Fed-batch and Nitrogen-deficient Culture on Lipid Yield of Chlorella sp.

    Institute of Scientific and Technical Information of China (English)

    葛珍珍; 王杰; 余晓斌


    目的 为了实现小球藻的高密度及高产油培养。方法 通过分析分批培养过程中藻细胞的生长曲线,葡萄糖消耗曲线,pH及溶氧变化曲线,以小球藻进行分批补料,待藻细胞达到一定的高密度后再进行缺氮培养以富集细胞内的油脂。结果 经过4次分批补料,小球藻的生物量达到了65.25g/L,然后进行缺氮培养12h,然后进行缺氮培养12h,小球藻的油脂含量由42.75%提高到63.82%,油脂含量达43.47g/L.结论 合理的分批补料明显地提高了小球藻的生物量。缺氮培养进一步提高了小球藻的油脂含量。%[Objective] This study was to realize high-density culture of Chlorella sp. as well as the culture with high lipid yield. [Method] Through analyzing the growth curve of Chlorella sp. cells, dextrose consumption curve, change curves of pH and dissolved oxygen, a fed-batch culture was conducted, followed by a nitrogen-deficient culture aiming at accumulating the lipids in Chlorella sp. cells when a high density of Chlorella sp. cells was obtained. [Result] After four batches of feeding were pro- vided, the biomass of Chlorella sp. reached up to 65.25 g/L, and the lipid content increased from 42.75% to 63.82% in Chlorella sp. cells, with the yield of 43.37 g/L in the following 12 hours of nitrogen-deficient culture. [Conclusion] Reasonable fed- batch can significantly improve the biomass of Chlorella sp., and the nitrogen-defi- cient culture further raises the lipid yield of Chlorella sp.

  4. 重组大肠杆菌BL21(pUC19 Hyp)产羟脯氨酸的补料分批培养%Production of hydroxyproline by fed-batch culture of novel recombinant Escherichia coli BL21(pUC19-Hyp)

    Institute of Scientific and Technical Information of China (English)

    袁春伟; 何艳春; 张胜利; 张震宇


    利用自主构建的组成型重组大肠杆菌BL21( pUC19 Hyp)为出发菌株,运用间歇流加、指数流加和恒速流加3种流加C源的方式进行补料分批培养。结果表明:在装液量为4 L的7 L发酵罐中,以0�30 g/min恒速流加为最优,在发酵44 h时,羟脯氨酸的质量浓度达到最高,为42�50 g/L,脯氨酸转化率为81%,此时细胞干质量为21�33 g/L,残糖质量浓度为0�17 g/L。 L 羟脯氨酸含量与摇瓶发酵时的1�39 g/L相比,提高了大约30倍,比日本株式会社的发酵产量提高了1�50 g/L,发酵过程中糖酸转化率约为4�0∶1。发酵液中的氨基酸分析结果表明,除脯氨酸、羟脯氨酸外的其他氨基酸质量浓度均低于0�1 g/L,发酵液中主要氨基酸为脯氨酸和羟脯氨酸。%The constitutive recombinant Escherichia coli, constructed in our lab, was used as the object of study. Three different ways of carbon source flow: intermittent flow addition, index flow addition and constant speed flow, were used in the process of supplementary fed-batch cultivation in the fermentor. The results showed that the optimal method was constant speed flow with 0�30 g/min. After 44 hours of fermentation, the concentration of L-hydroxyproline was the highest of 42�50 g/L;Conversion of proline was 81%; residual sugar concentration was 0�17 g/L. Compared with shaking flask fermentation with concentration of 1�39 g/L, L-hydroxyproline content increased about 30 times and at the same time. In the process of fermentation, the ratio of glucose consumption and product of L-hydroxyproline was about 4�0∶1. The analysis of amino acids in the fermentation liquid showed proline and L-hydroxyproline were the main amino acids.

  5. Scale-up of rifamycin B fermentation with Amycolatoposis mediterranei

    Institute of Scientific and Technical Information of China (English)

    金志华; 林建平; 岑沛霖


    Study of the effect of dissolved oxygen and shear stress on rifamycin B fermentation with A. mediterranei XC 9-25 showed that rifamycin B fermentation with Amycolatoposis mediterranei XC 9-25 needs high dissolved oxygen and is not very sensitive to shearing stress. The scale-up ofrifamycin B fermentation withA, mediterranei XC 9-25 from a shaking flask to a 15 L fermentor was realized by controlling the dissolved oxygen to above 25% of saturation in the fermentation process, and the potency of rifamycin B fermentation in the 15 L fermentor reached 10 g/L after 6-day batch fermentation. By continuously feeding glucose and ammonia in the fermentation process, the potency of rifamycin B fermentaion in the 15 L fermentor reached 18.67 g/L, which was 86.65% higher than that of batch fermentation. Based on the scale-up principle of constantly aerated agitation power per unit volume, the scale-up of rifamycin B fed-batch fermentation with continuous feed from a 15 L fermentor to a 7 m3 fermentor and further to a 60 m3 fermentor was realized successfully. The potency of rifamycin B fermentation in the 7 m3 fermentor and in the 60 m3 fermentor reached 17.25 g/L and 19.11 g/L, respectively.

  6. Scale-up of rifamycin B fermentation with Amycolatoposis mediterranei

    Institute of Scientific and Technical Information of China (English)

    金志华; 林建平; 岑沛霖


    Study of the effect of dissolved oxygen and shear stress on rifamycin B fermentation with A. mediterranei XC 9-25 showed that rifamycin B fermentation with Amycolatoposis mediterranei XC 9-25 needs high dissolved oxygen and is not very sensitive to shearing stress. The scale-up of rifamycin B fermentation withA. mediterranei XC 9-25 from a shaking flask to a 15 L fermentor was realized by controlling the dissolved oxygen to above 25% of saturation in the fermentation process, and the potency of rifamycin B fermentation in the 15 L fermentor reached 10 g/L after 6-day batch fermentation.By continuously feeding glucose and ammonia in the fermentation process, the potency of rifamycin B fermentaion in the 15 L fermentor reached 18.67 g/L, which was 86.65% higher than that of batch fermentation. Based on the scale-up principle of constantly aerated agitation power per unit volume, the scale-up ofrifamycin B fed-batch fermentation with continuous feed from a 15 L fermentor to a 7 m3 fermentor and further to a 60 m3 fermentor was realized successfully. The potency of rifamycin B fermentation in the 7 m3 fermentor and in the 60 m3 fermentor reached 17.25 g/L and 19.11 g/L, respectively.

  7. 微量营养素对玉米酵母流加培养的影响%Effect ofmicronutrients on fed-batch culture of yeast on maize medium

    Institute of Scientific and Technical Information of China (English)

    李竹生; 张新伟; 宋娜; 牛芳方


    The effects of different amounts of micronutrients on the feeding rate of sugar solution, dissolved oxygen, yeast cell concentration and yeast cell growth during the fed-batch culture of yeast on maize medium were studied. It was showed that the optimal amount of the micronutrients were as follows: CaCl2 28.8mg/L, FeSO4 14.4mg/L, thiamine hydrochloride 4.8mg/L, biotin 0.12mg/L, D-calcium pantothenate 6.4mg/L and myoinositol 120mg/L. The yield of yeast had increase by 80.9% with the addition of micronutrients.%文中研究了微量营养素的不同添加量对玉米酵母流加培养过程中糖液流加速率、溶氧量、酵母细胞浓度、酵母细胞生长率的变化,得出微量营养素的添加量为CaCl2 28.8mg/L,FeSO4 14.4mg/L,盐酸硫氨4.Smg/L,生物素0.12mg/L,D-泛酸钙6.4mg/L,肌醇120mg/L,玉米酵母细胞的产率提高了80.9%.

  8. Adaptive control of feed load changes in alcohol fermentation

    Energy Technology Data Exchange (ETDEWEB)

    Folly, R.; Berlim, R.; Salgado, A.; Franca, R.; Valdman, B. [Universidade Federal, Rio de Janeiro, RJ (Brazil). Escola de Quimica


    A fed-batch alcohol fermentation on a pilot plant scale with a digital supervisory control was evaluated as an experimental application case study of an adaptive controller. The verification of intrinsically dynamic variations in the characteristics of the fermentation, observed in previous work, showed the necessity of an adaptive control strategy for controller parameter tuning in order to adjust the changes in the specific rates of consumption, growth and product formation during the process. Satisfactory experimental results were obtained for set-point variations and sugar feed concentration load changes in the manipulated inlet flow to the fermenter. (author) 5 refs., 10 figs., 2 tabs.; e-mail: Valdman at H2O.EQ.UFRJ.BR

  9. Efficient calcium lactate production by fermentation coupled with crystallization-based in situ product removal. (United States)

    Xu, Ke; Xu, Ping


    Lactic acid is a platform chemical with various industrial applications, and its derivative, calcium lactate, is an important food additive. Fermentation coupled with in situ product removal (ISPR) can provide more outputs with high productivity. The method used in this study was based on calcium lactate crystallization. Three cycles of crystallization were performed during the fermentation course using a Bacillus coagulans strain H-1. As compared to fed-batch fermentation, this method showed 1.7 times higher average productivity considering seed culture, with 74.4% more L-lactic acid produced in the fermentation with ISPR. Thus, fermentation coupled with crystallization-based ISPR may be a biotechnological alternative that provides an efficient system for production of calcium lactate or lactic acid.

  10. Glucose and xylose co-fermentation of pretreated wheat straw using mutants of S. cerevisiae TMB3400. (United States)

    Erdei, Borbála; Frankó, Balázs; Galbe, Mats; Zacchi, Guido


    Wheat straw was pretreated and fermented to ethanol. Two strains, which had been mutated from the genetically modified Saccharomyces cerevisiae TMB3400, KE6-12 and KE6-13i, have been used in this study and the results of performance were compared to that of the original strain. The glucose and xylose co-fermentation ability was investigated in batch fermentation of steam-pretreated wheat straw (SPWS) liquid (undiluted, and diluted 1.5 and 2 times). Both strains showed improved xylose uptake in diluted SPWS liquid, and increased ethanol yields compared with the original TMB3400 strain, although xylitol formation also increased slightly. In undiluted SPWS liquid, however, only KE6-13i performed better than the original strain regarding xylose utilization. Fed-batch fermentation of 1.5 and 2 times diluted liquid was performed by adding the glucose-rich hydrolysates from enzymatic hydrolysis of the solid fraction of SPWS at a constant feed rate after 5 h of fermentation, when the glucose had been depleted. The modified strains showed improved xylose conversion; however, the ethanol yield was not significantly improved due to increased glycerol production. Fed-batch fermentation resulted in faster xylose utilization than in the batch cases. Copyright © 2012 Elsevier B.V. All rights reserved.

  11. Acetone-butanol-ethanol from sweet sorghum juice by an immobilized fermentation-gas stripping integration process. (United States)

    Cai, Di; Wang, Yong; Chen, Changjing; Qin, Peiyong; Miao, Qi; Zhang, Changwei; Li, Ping; Tan, Tianwei


    In this study, sweet sorghum juice (SSJ) was used as the substrate in a simplified ABE fermentation-gas stripping integration process without nutrients supplementation. The sweet sorghum bagasse (SSB) after squeezing the fermentable juice was used as the immobilized carrier. The results indicated that the productivity of ABE fermentation process was improved by gas stripping integration. A total 24g/L of ABE solvents was obtained from 59.6g/L of initial sugar after 80h of fermentation with gas stripping. Then, long-term of fed-batch fermentation with continuous gas stripping was further performed. 112.9g/L of butanol, 44.1g/L of acetone, 9.5g/L of ethanol (total 166.5g/L of ABE) was produced in overall 312h of fermentation. At the same time, concentrated ABE product was obtained in the condensate of gas stripping.


    Institute of Scientific and Technical Information of China (English)


    In batch cultures of Candida krusei, the effect of corn steep liquor (CSL) concentration was investigated. The result showed that glycerol yield was greatly affected by CSL concentration. Higher glycerol yield was attained when the initial CSL concentration in the medium was 9 g.L-1. The fermentation kinetics were studied and the parameters for cell growth, maintenance, and glycerol production were obtained. Material balance revealed by product formation during the growth phase and suggested a fed-batch culture strategy.

  13. Production of Welan Gum by Alcaligenes sp.NX-3 with Fed-batch Fermentation%Alcaligenes sp.NX-3产威兰胶的补料分批发酵工艺研究

    Institute of Scientific and Technical Information of China (English)

    李会; 李莎; 冯小海; 汪芙蓉; 徐虹


    在7.5 L发酵罐上考察了Alcaligenes sp.NX-3产威兰胶的发酵工艺.选用葡萄糖为碳源,通过分析比较不同初糖浓度下的细胞比生长速率和产物比合成速率,进一步研究了不同补料方式对产胶的影响.结果表明,采用分批补糖发酵工艺,威兰胶产量较分批发酵提高了13.6%,而且有效地缩短了发酵周期.在50 L发酵罐上进行补料分批发酵放大实验,威兰胶产量高达27.0 g/L,葡萄糖转化率由初始的44%提高到54%.

  14. Study on Monascus pigment production by fed-batch fermentation of glucose solution with immobilized Monascus purpureus%固定化红曲葡萄糖母液流加发酵红曲色素的研究

    Institute of Scientific and Technical Information of China (English)

    王克明; 何潇湘


    对固定化红曲(Monascus purpureus),在生物反应器中流加葡萄糖母液发酵生产红曲色素进行了研究,建立了简单的数学模型控制流加.结果表明:当总葡萄糖母液浓度为150g/L时,以90g/L初始葡萄糖母液开始发酵,当流加因子K=0.0013时,变速流加发酵组的色素浓度比非流加发酵组的色价提高32%.

  15. Stable acetate production in extreme-thermophilic (70°C) mixed culture fermentation by selective enrichment of hydrogenotrophic methanogens (United States)

    Zhang, Fang; Zhang, Yan; Ding, Jing; Dai, Kun; van Loosdrecht, Mark C. M.; Zeng, Raymond J.


    The control of metabolite production is difficult in mixed culture fermentation. This is particularly related to hydrogen inhibition. In this work, hydrogenotrophic methanogens were selectively enriched to reduce the hydrogen partial pressure and to realize efficient acetate production in extreme-thermophilic (70°C) mixed culture fermentation. The continuous stirred tank reactor (CSTR) was stable operated during 100 days, in which acetate accounted for more than 90% of metabolites in liquid solutions. The yields of acetate, methane and biomass in CSTR were 1.5 +/- 0.06, 1.0 +/- 0.13 and 0.4 +/- 0.05 mol/mol glucose, respectively, close to the theoretical expected values. The CSTR effluent was stable and no further conversion occurred when incubated for 14 days in a batch reactor. In fed-batch experiments, acetate could be produced up to 34.4 g/L, significantly higher than observed in common hydrogen producing fermentations. Acetate also accounted for more than 90% of soluble products formed in these fed-batch fermentations. The microbial community analysis revealed hydrogenotrophic methanogens (mainly Methanothermobacter thermautotrophicus and Methanobacterium thermoaggregans) as 98% of Archaea, confirming that high temperature will select hydrogenotrophic methanogens over aceticlastic methanogens effectively. This work demonstrated a potential application to effectively produce acetate as a value chemical and methane as an energy gas together via mixed culture fermentation.

  16. Kinetics of volatile metabolites during alcoholic fermentation of cane molasses by Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Cachot, T.; Mueller, M.; Pons, M.N. (Centre National de la Recherche Scientifique, 54 - Nancy (France). Lab. des Sciences du Genie Chimique)


    The kinetics of ethanol, acetaldehyde, ethyl acetate and fusel alcohols during alcoholic fermentations on cane molasses by Saccharomyces cerevisiae have been obtained via an in-situ gas membrane sensor connected to a gas chromatograph. Various operation parameters have been investigated such as inoculum rate, molasses concentration, operation mode (batch, fed-batch). The modification of fusel alcohols kinetics in response to addition of amino acids has been studied as well as the assimilation of two intermediary aldehydes (isovaleraldehyde and isobutyraldehyde) in the fusel alcohol synthesis pathway. (orig.).

  17. Improvement of productivity in acetic acid fermentation with Clostridium thermoaceticum

    Energy Technology Data Exchange (ETDEWEB)

    Shah, M.M.; Cheryan, M. [Univ. of Illinois, Urbana, IL (United States)


    Production of acetic acid by a mutant strain of Clostridium thermoaceticum was compared in three types of membrane cell-recycle bioreactors. A modified fed-batch bioreactor (where the product is partially removed at the end of fermentation, but the cells are retained), and a two-stage CSTR (with product being removed continuously and the cells being recycled from the second to the first stage) resulted in better performance than a one-stage CSTR or batch fermenter. The difference in performance was greater at higher acetate concentration. With 45 g/L of glucose in the feed, productivity was 0.75-1.12 g/L-h and acetic acid concentrations were 34-38 g/L. This is more than double the batch system. The nutrient supply rate also appeared to have a strong influence on productivity of the microorganism.

  18. Repeated-batch fermentation of lignocellulosic hydrolysate to ethanol using a hybrid Saccharomyces cerevisiae strain metabolically engineered for tolerance to acetic and formic acids. (United States)

    Sanda, Tomoya; Hasunuma, Tomohisa; Matsuda, Fumio; Kondo, Akihiko


    A major challenge associated with the fermentation of lignocellulose-derived hydrolysates is improved ethanol production in the presence of fermentation inhibitors, such as acetic and formic acids. Enhancement of transaldolase (TAL) and formate dehydrogenase (FDH) activities through metabolic engineering successfully conferred resistance to weak acids in a recombinant xylose-fermenting Saccharomyces cerevisiae strain. Moreover, hybridization of the metabolically engineered yeast strain improved ethanol production from xylose in the presence of both 30 mM acetate and 20mM formate. Batch fermentation of lignocellulosic hydrolysate containing a mixture of glucose, fructose and xylose as carbon sources, as well as the fermentation inhibitors, acetate and formate, was performed for five cycles without any loss of fermentation capacity. Long-term stability of ethanol production in the fermentation phase was not only attributed to the coexpression of TAL and FDH genes, but also the hybridization of haploid strains.

  19. Wastewater recycling technology for fermentation in polyunsaturated fatty acid production. (United States)

    Song, Xiaojin; Ma, Zengxin; Tan, Yanzhen; Zhang, Huidan; Cui, Qiu


    To reduce fermentation-associated wastewater discharge and the cost of wastewater treatment, which further reduces the total cost of DHA and ARA production, this study first analyzed the composition of wastewater from Aurantiochytrium (DHA) and Mortierella alpina (ARA) fermentation, after which wastewater recycling technology for these fermentation processes was developed. No negative effects of DHA and ARA production were observed when the two fermentation wastewater methods were cross-recycled. DHA and ARA yields were significantly inhibited when the wastewater from the fermentation process was directly reused. In 5-L fed-batch fermentation experiments, using this cross-recycle technology, the DHA and ARA yields were 30.4 and 5.13gL(-1), respectively, with no significant changes (P>0.05) compared to the control group, and the water consumption was reduced by half compared to the traditional process. Therefore, this technology has great potential in industrial fermentation for polyunsaturated fatty acid production. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Optimization of solid substrate fermentation of wheat straw. (United States)

    Abdullah, A L; Tengerdy, R P; Murphy, V G


    Optimal conditions for solid substrate fermentation of wheat straw with Chaetomium cellulolyticum in laboratory-scale stationary layer fermenters were developed. The best pretreatment for wheat straw was ammonia freeze explosion, followed by steam treatment, alkali treatment, and simple autoclaving. The optimal fermentation conditions were 80% (w/w) moisture content; incubation temperature of 37 degrees C; 2% (w/w) unwashed mycelial inoculum; aeration at 0.12 L/h/g; substrate thickness of 1 to 2 cm; and duration of three days. Technical parameters for this optimized fermentation were: degree of substance utilization, 27.2%; protein yield/substrate, 0.09 g; biomass yield/bioconverted substrate, 0.40 g; degree of bioconversion of total available sugars in the substrate, 60.5%; specific efficiency of bioconversion, 70.8%; and overall efficiency of biomass production from substrate, 42.7%. Mixed culturing of Candida utilis further increased biomass production by 20%. The best mode of fermentation was a semicontinuous fed-batch fermentation where one-half of the fermented material was removed at three-day intervals and replaced by fresh substrate. In this mode, protein production was 20% higher than in batch mode, protein productivity was maintained over 12 days, and sporulation was prevented.

  1. Whey fermentation by anaerobiospirillum succiniciproducens for production of a succinate-based animal feed additive (United States)

    Samuelov; Datta; Jain; Zeikus


    Anaerobic fermentation processes for the production of a succinate-rich animal feed supplement from raw whey were investigated with batch, continuous, and variable-volume fed-batch cultures with Anaerobiospirillum succiniciproducens. The highest succinate yield, 90%, was obtained in a variable-volume fed-batch process in comparison to 80% yield in a batch cultivation mode. In continuous culture, succinate productivity was 3 g/liter/h, and the yield was 60%. Under conditions of excess CO2, more than 90% of the whey-lactose was consumed, with an end product ratio of 4 succinate to 1 acetate. Under conditions of limited CO2, lactose was only partially consumed and lactate was the major end product, with lower levels of ethanol, succinate, and acetate. When the succinic acid in this fermentation product was added to rumen fluid, it was completely consumed by a mixed rumen population and was 90% decarboxylated to propionate on a molar basis. The whey fermentation product formed under excess CO2, which contained mainly organic acids and cells, could potentially be used as an animal feed supplement.

  2. Bioprocess Control in Microscale: Scalable Fermentations in Disposable and User-Friendly Microfluidic Systems

    Directory of Open Access Journals (Sweden)

    Müller Carsten


    Full Text Available Abstract Background The efficiency of biotechnological production processes depends on selecting the best performing microbial strain and the optimal cultivation conditions. Thus, many experiments have to be conducted, which conflicts with the demand to speed up drug development processes. Consequently, there is a great need for high-throughput devices that allow rapid and reliable bioprocess development. This need is addressed, for example, by the fiber-optic online-monitoring system BioLector which utilizes the wells of shaken microtiter plates (MTPs as small-scale fermenters. To further improve the application of MTPs as microbioreactors, in this paper, the BioLector technology is combined with microfluidic bioprocess control in MTPs. To realize a user-friendly system for routine laboratory work, disposable microfluidic MTPs are utilized which are actuated by a user-friendly pneumatic hardware. Results This novel microfermentation system was tested in pH-controlled batch as well as in fed-batch fermentations of Escherichia coli. The pH-value in the culture broth could be kept in a narrow dead band of 0.03 around the pH-setpoint, by pneumatically dosing ammonia solution and phosphoric acid to each culture well. Furthermore, fed-batch cultivations with linear and exponential feeding of 500 g/L glucose solution were conducted. Finally, the scale-up potential of the microscale fermentations was evaluated by comparing the obtained results to that of fully controlled fermentations in a 2 L laboratory-scale fermenter (working volume of 1 L. The scale-up was realized by keeping the volumetric mass transfer coefficient kLa constant at a value of 460 1/h. The same growth behavior of the E. coli cultures could be observed on both scales. Conclusion In microfluidic MTPs, pH-controlled batch as well as fed-batch fermentations were successfully performed. The liquid dosing as well as the biomass growth kinetics of the process-controlled fermentations

  3. Ethanol from Whey: Continuous Fermentation with a Catabolite Repression-Resistant Saccharomyces cerevisiae Mutant. (United States)

    Terrell, S L; Bernard, A; Bailey, R B


    An alternative method for the conversion of cheese whey lactose into ethanol has been demonstrated. With the help of continuous-culture technology, a catabolite repression-resistant mutant of Saccharomyces cerevisiae completely fermented equimolar mixtures of glucose and galactose into ethanol. The first step in this process was a computer-controlled fed-batch operation based on the carbon dioxide evolution rate of the culture. In the absence of inhibitory ethanol concentrations, this step allowed us to obtain high biomass concentrations before continuous fermentation. The continuous anaerobic process successfully incorporated a cell-recycle system to optimize the fermentor productivity. Under conditions permitting a low residual sugar concentration (fermentation of highly concentrated feed solutions (20%) was also demonstrated, but only with greatly diminished fermentor productivity (5.5 g liter h).

  4. Kinetics of High Cell Density Fed-batch Culture of Recombinant Escherichia coli Producing Human-like Collagen%重组大肠杆菌分批-补料高密度发酵生产类人胶原蛋白的动力学

    Institute of Scientific and Technical Information of China (English)

    花秀夫; 范代娣; 骆艳娥; 张兮; 施惠娟; 米钰; 马晓轩; 尚龙安; 赵桂仿


    The kinetics of batch and fed-batch cultures of recombinant Escherichia coli producing human-like collagen was investigated. In the batch culture, a kinetic model of a simple growth-association system was concluded without consideration of cell endogeneous metabolism. The cell lag time, the maximum specific growth rate and growth rates were set at (0.15, 0.2, 0.25h-1) by the method of pseudo-exponential feeding, and the expressions for the specific rate of substrate consumption, the growth kinetics and the product formation kinetics of each phase spectively. The model predictions are in good agreement with the experimental data.

  5. Fermentation and purification strategies for the production of betulinic acid and its lupane-type precursors in Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Czarnotta, Eik; Dianat, Mariam; Korf, Marcel


    from the bark of plane tree or birch. Here, we reengineered the reported betulinic acid pathway into S. cerevisiae and used this novel strain to develop efficient fermentation and product purification methods. Fed-batch cultivations with ethanol excess, using either an ethanol-pulse feed or controlling....../L and total triterpenoid concentrations of 854 mg/L, the highest concentrations reported so far. Purification of lupane-type triterpenoids with high selectivity and yield was achieved by solid-liquid extraction without prior cell disruption using polar aprotic solvents such as acetone or ethyl acetate...

  6. Bioethanol production from fermentable sugar juice. (United States)

    Zabed, Hossain; Faruq, Golam; Sahu, Jaya Narayan; Azirun, Mohd Sofian; Hashim, Rosli; Boyce, Amru Nasrulhaq


    Bioethanol production from renewable sources to be used in transportation is now an increasing demand worldwide due to continuous depletion of fossil fuels, economic and political crises, and growing concern on environmental safety. Mainly, three types of raw materials, that is, sugar juice, starchy crops, and lignocellulosic materials, are being used for this purpose. This paper will investigate ethanol production from free sugar containing juices obtained from some energy crops such as sugarcane, sugar beet, and sweet sorghum that are the most attractive choice because of their cost-effectiveness and feasibility to use. Three types of fermentation process (batch, fed-batch, and continuous) are employed in ethanol production from these sugar juices. The most common microorganism used in fermentation from its history is the yeast, especially, Saccharomyces cerevisiae, though the bacterial species Zymomonas mobilis is also potentially used nowadays for this purpose. A number of factors related to the fermentation greatly influences the process and their optimization is the key point for efficient ethanol production from these feedstocks.

  7. Bioethanol Production from Fermentable Sugar Juice (United States)

    Zabed, Hossain; Faruq, Golam; Sahu, Jaya Narayan; Azirun, Mohd Sofian; Hashim, Rosli; Nasrulhaq Boyce, Amru


    Bioethanol production from renewable sources to be used in transportation is now an increasing demand worldwide due to continuous depletion of fossil fuels, economic and political crises, and growing concern on environmental safety. Mainly, three types of raw materials, that is, sugar juice, starchy crops, and lignocellulosic materials, are being used for this purpose. This paper will investigate ethanol production from free sugar containing juices obtained from some energy crops such as sugarcane, sugar beet, and sweet sorghum that are the most attractive choice because of their cost-effectiveness and feasibility to use. Three types of fermentation process (batch, fed-batch, and continuous) are employed in ethanol production from these sugar juices. The most common microorganism used in fermentation from its history is the yeast, especially, Saccharomyces cerevisiae, though the bacterial species Zymomonas mobilis is also potentially used nowadays for this purpose. A number of factors related to the fermentation greatly influences the process and their optimization is the key point for efficient ethanol production from these feedstocks. PMID:24715820

  8. High cell density propionic acid fermentation with an acid tolerant strain of Propionibacterium acidipropionici. (United States)

    Wang, Zhongqiang; Jin, Ying; Yang, Shang-Tian


    Propionic acid is an important chemical with wide applications and its production via fermentation is of great interest. However, economic production of bio-based propionic acid requires high product titer, yield, and productivity in the fermentation. A highly efficient and stable high cell density (HCD) fermentation process with cell recycle by centrifugation was developed for propionic acid production from glucose using an acid-tolerant strain of Propionibacterium acidipropionici, which had a higher specific growth rate, productivity, and acid tolerance compared to the wild type ATCC 4875. The sequential batch HCD fermentation at pH 6.5 produced propionic acid at a high titer of ∼40 g/L and productivity of 2.98 g/L h, with a yield of ∼0.44 g/g. The product yield increased to 0.53-0.62 g/g at a lower pH of 5.0-5.5, which, however, decreased the productivity to 1.28 g/L h. A higher final propionic acid titer of >55 g/L with a productivity of 2.23 g/L h was obtained in fed-batch HCD fermentation at pH 6.5. A 3-stage simulated fed-batch process in serum bottles produced 49.2 g/L propionic acid with a yield of 0.53 g/g and productivity of 0.66 g/L h. These productivities, yields and propionic acid titers were among the highest ever obtained in free-cell propionic acid fermentation.

  9. Bio-Hydrogen Potential Of Easily Biodegradable Substrate Through Dark Fermentation

    Energy Technology Data Exchange (ETDEWEB)

    Baris Calli; Wesley Boenne; Karolien Vanbroekhoven [Flemish Institute for Technological Research (VITO), Boeretang 200, 2400 Mol, (Belgium)


    For hydrogen production through dark fermentation of glucose, a continuously stirred 1-liter bioreactor was inoculated with yard waste compost and operated at 55 C with fed-batch runs. The reducing pH was regulated automatically by using pH transmitter and kept constant at 5.4. In this way, no methane was generated in any of the fed-batch runs and H{sub 2} yield in the range of 0.25 to 1.75 mol H{sub 2}/mol glucose consumed was obtained by inhibiting methanogenic H{sub 2} consumption. Astonishingly, the highest H{sub 2} yield was achieved with fresh inoculum which was neither heat treated nor acclimated. However, yield was not steady and decreased due to shift in metabolic pathway from acido-genesis to ethanol fermentation subsequent to high H{sub 2} partial pressure. Effluent ethanol concentrations above 400 mg/l after high H{sub 2} yielding runs were indication of this metabolic shift. (authors)

  10. Comparison of different estimation techniques for biomass concentration in large scale yeast fermentation. (United States)

    Hocalar, A; Türker, M; Karakuzu, C; Yüzgeç, U


    In this study, previously developed five different state estimation methods are examined and compared for estimation of biomass concentrations at a production scale fed-batch bioprocess. These methods are i. estimation based on kinetic model of overflow metabolism; ii. estimation based on metabolic black-box model; iii. estimation based on observer; iv. estimation based on artificial neural network; v. estimation based on differential evaluation. Biomass concentrations are estimated from available measurements and compared with experimental data obtained from large scale fermentations. The advantages and disadvantages of the presented techniques are discussed with regard to accuracy, reproducibility, number of primary measurements required and adaptation to different working conditions. Among the various techniques, the metabolic black-box method seems to have advantages although the number of measurements required is more than that for the other methods. However, the required extra measurements are based on commonly employed instruments in an industrial environment. This method is used for developing a model based control of fed-batch yeast fermentations. Copyright © 2010 ISA. Published by Elsevier Ltd. All rights reserved.

  11. Simultaneous saccharification and fermentation of lignocellulosic residues pretreated with phosphoric acid-acetone for bioethanol production. (United States)

    Li, Hui; Kim, Nag-Jong; Jiang, Min; Kang, Jong Won; Chang, Ho Nam


    Bermudagrass, reed and rapeseed were pretreated with phosphoric acid-acetone and used for ethanol production by means of simultaneous saccharification and fermentation (SSF) with a batch and fed-batch mode. When the batch SSF experiments were conducted in a 3% low effective cellulose, about 16 g/L of ethanol were obtained after 96 h of fermentation. When batch SSF experiments were conducted with a higher cellulose content (10% effective cellulose for reed and bermudagrass and 5% for rapeseed), higher ethanol concentrations and yields (of more than 93%) were obtained. The fed-batch SSF strategy was adopted to increase the ethanol concentration further. When a higher water-insoluble solid (up to 36%) was applied, the ethanol concentration reached 56 g/L of an inhibitory concentration of the yeast strain used in this study at 38 degrees C. The results show that the pretreated materials can be used as good feedstocks for bioethanol production, and that the phosphoric acid-acetone pretreatment can effectively yield a higher ethanol concentration.

  12. Expression and purification of ELP-intein-tagged target proteins in high cell density E. coli fermentation

    Directory of Open Access Journals (Sweden)

    Wood David W


    Full Text Available Abstract Background Elastin-like polypeptides (ELPs are useful tools that can be used to non-chromatographically purify proteins. When paired with self-cleaving inteins, they can be used as economical self-cleaving purification tags. However, ELPs and ELP-tagged target proteins have been traditionally expressed using highly enriched media in shake flask cultures, which are generally not amenable to scale-up. Results In this work, we describe the high cell-density expression of self-cleaving ELP-tagged targets in a supplemented minimal medium at a 2.5 liter fermentation scale, with increased yields and purity compared to traditional shake flask cultures. This demonstration of ELP expression in supplemented minimal media is juxtaposed to previous expression of ELP tags in extract-based rich media. We also describe several sets of fed-batch conditions and their impact on ELP expression and growth medium cost. Conclusions By using fed batch E. coli fermentation at high cell density, ELP-intein-tagged proteins can be expressed and purified at high yield with low cost. Further, the impact of media components and fermentation design can significantly impact the overall process cost, particularly at large scale. This work thus demonstrates an important advances in the scale up of self-cleaving ELP tag-mediated processes.

  13. A comprehensive and quantitative review of dark fermentative biohydrogen production

    Directory of Open Access Journals (Sweden)

    Rittmann Simon


    Full Text Available Abstract Biohydrogen production (BHP can be achieved by direct or indirect biophotolysis, photo-fermentation and dark fermentation, whereof only the latter does not require the input of light energy. Our motivation to compile this review was to quantify and comprehensively report strains and process performance of dark fermentative BHP. This review summarizes the work done on pure and defined co-culture dark fermentative BHP since the year 1901. Qualitative growth characteristics and quantitative normalized results of H2 production for more than 2000 conditions are presented in a normalized and therefore comparable format to the scientific community. Statistically based evidence shows that thermophilic strains comprise high substrate conversion efficiency, but mesophilic strains achieve high volumetric productivity. Moreover, microbes of Thermoanaerobacterales (Family III have to be preferred when aiming to achieve high substrate conversion efficiency in comparison to the families Clostridiaceae and Enterobacteriaceae. The limited number of results available on dark fermentative BHP from fed-batch cultivations indicates the yet underestimated potential of this bioprocessing application. A Design of Experiments strategy should be preferred for efficient bioprocess development and optimization of BHP aiming at improving medium, cultivation conditions and revealing inhibitory effects. This will enable comparing and optimizing strains and processes independent of initial conditions and scale.

  14. Saccharomyces pastorianus as cell factory to improve production of fructose 1,6-diphosphate using novel fermentation strategies

    Directory of Open Access Journals (Sweden)

    Chiara Schiraldi


    Full Text Available Enzymatic phosphorylation of glucose with inorganic phosphate, mediated by permeabilized yeast cells, is one of the methods commonly used to manufacture fructose 1,6-diphosphate, a compound of pharmaceutical interest. This process requires high concentrations of yeast active biomass, that is the catalyst of bioconversion of glucose and inorganic phosphate into fructose 1,6-diphosphate. In this study we firstly describe the high cell density production of a brewer's Saccharomyces strain (Saccharomyces pastorianus DSM 6581, focusing on the optimization of medium composition and exploiting fed-batch strategies and novel technologies based on membrane bioreactors. In fed-batch fermentation an appropriate exponential feed profile was set up to maintain the glucose concentration in the bioreactor below 0.9 g·L-1, thus yielding reproducibly 58 g dry weight biomass per liter in 80 h fermentation, improving eight-fold batch processes output. In addition a higher final biomass density was reached when implementing a microfiltration strategy (70 g dry weight biomass, that led to a productivity of 2.1 gcdw·L-1·h-1, 2.4-fold the fed-batch one. Successively, this biomass was opportunely permeabilized and proved capable of catalyzing the bioconversion of glucose into fructose 1,6-diphosphate. Acting on critical parameters of the bioconversion (substrates molar ratio, catalyst concentration and permeabilization agent, fructose 1,6-diphosphate was produced, after 3 h of process, at 56.3 ± 1 g·L-1 with a yield of 80% of the theoretical value.

  15. Assessment of morphological changes of Clostridium acetobutylicum by flow cytometry during acetone/butanol/ethanol extractive fermentation. (United States)

    González-Peñas, Helena; Lu-Chau, Thelmo Alejandro; Moreira, Maria Teresa; Lema, Juan Manuel


    Acetone/butanol/ethanol (ABE) fermentation by Clostridium acetobutylicum was investigated in extractive fed-batch experiments. In conventional fermentations, metabolic activity ceases when a critical threshold products concentration is reached (~21.6 g solvents l(-1)). Solvents production was increased up to 36.6 and 37.2 g l(-1), respectively, using 2-butyl-1-octanol (aqueous to organic ratio: 1:0.25 v/v) and pomace olive oil (1:1 v/v) as extraction solvents. The morphological changes of different cell types were monitored and quantified using flow cytometry. Butanol production in extractive fermentations with pomace olive oil was achieved mainly by vegetative cells, whereas the percentage of sporulating cells was lower than 10%.

  16. Application of bipolar electrodialysis to E.coli fermentation for simultaneous acetate removal and pH control

    DEFF Research Database (Denmark)

    Wong, M.; Woodley, John; Lye, G.J.


    The application of bipolar electrodialysis (BPED) for the simultaneous removal of inhibitory acetate and pH control during E. coli fermentation was investigated. A two cell pair electrodialysis module, consisting of cation exchange, anion exchange and bipolar membranes with working area of 100 cm2...... each, was integrated with a standard 7 l stirred tank bioreactor. Results showed that BPED was beneficial in terms of in situ removal of inhibitory acetate and a reduction in the amount NH4OH used for pH control. In batch and fed-batch BPED fermentations, base additions were decreased by up to 50......% in both cases compared to electrodialysis (ED) fermentations with pH controlled at 6.7 ± 0.1. Consequently, the final biomass (34.2 g DCW l−1) and recombinant protein (5.5 g l−1) concentrations obtained were increased by up to 37 and 20%, respectively....

  17. Development and Appl ication of Continuous Packing-tower Surface Fermentation Device%填料塔式连续表面发酵装置的研制及应用

    Institute of Scientific and Technical Information of China (English)

    傅亮; 陈宇哲; 彭英


    The research developed a kind of continuous packing-tower surface fermentation device,in which the RF4 Gluconacetobacter xylinus strain fermented the broth with ethanol into acetic acid.A comparison of total acidity and ADH enzyme activity of fermentation broth in continuous packing-tower sur-face fermentation device and static flask by single-batch fermentation and fed-batch fermentation is made.The results show that the ADH enzyme activity of fermentation broth reaches to 4.13 × 10-3 U/g and the total acidity peaks to 7.07 g/dL on the 13th day by fed-batch fermentation in contin-uous packing-tower surface fermentation device,whose total acidity increased by 1 3% and the fermen-tation time shortened by 1 5% compared with static fed-batch fermentation.The total acidity increased by 7 1% compared with static single-batch fermentation.%研制了一种填料塔式连续发酵装置,以木醋杆菌(Gluconacetobacter xylinus )RF4为菌种,用含乙醇培养液进行醋酸发酵。比较了该填料表面发酵装置与静置三角瓶各自进行单批和分批发酵的产酸和乙醇脱氢酶的活力。结果表明:在连续填料发酵装置中进行分批补料发酵,发酵液中的乙醇脱氢酶活力为4.13×10-3 U/g,总酸度在第13天达到峰值7.07 g/dL。比静置分批补料发酵酸度提高13%,时间缩短15%,比静置单批发酵酸度提高71%。

  18. Fermentative hydrogen gas production using biosolids pellets as the inoculum source. (United States)

    Kalogo, Youssouf; Bagley, David M


    Biosolids pellets produced from anaerobically digested municipal wastewater sludge by drying to greater than 90% total solids at 110-115 degrees C for at least 75 min, were tested for their suitability as an inoculum source for fermentative hydrogen production. The hydrogen recoveries (mg gaseous H(2) produced as COD/mg added substrate COD) for glucose-fed batch systems were equal, 20.2-21.5%, between biosolids pellets and boiled anaerobic digester sludge as inoculum sources. Hydrogen recoveries from primary sludge were 2.4% and 3.5% using biosolids pellets and boiled sludge, respectively, and only 0.2% and 0.8% for municipal wastewater. Biosolids pellets should be a practical inoculum source for fermentative hydrogen reactors, although the effectiveness will depend on the wastewater treated.

  19. Heterologous fermentation of a diterpene from Alternaria brassisicola (United States)

    Arens, Julia; Bergs, Dominik; Mewes, Mirja; Merz, Juliane; Schembecker, Gerhard; Schulz, Frank


    A variety of different applications render terpenes and terpenoids attractive research targets. A promising but so far insufficiently explored family of terpenoids are the fusicoccanes that comprise a characteristic 5-8-5 fused tricyclic ring system. Besides herbicidal effects, these compounds also show apoptotic and anti-tumour effects on mammalian cells. The access to fusicoccanes from natural sources is scarce. Recently, we introduced a metabolically engineered Saccharomyces cerevisiae strain to enable the heterologous fermentation of the shared fusicoccane–diterpenoid precursor, fusicocca-2,10(14)-diene. Here, we show experiments towards the identification of bottlenecks in this process. The suppression of biosynthetic by-products via medium optimisation was found to be an important aspect. In addition, the fermentation process seems to be improved under oxygen limitation conditions. Under fed-batch conditions, the fermentation yield was reproducibly increased to approximately 20 mg/L. Furthermore, the impact of the properties of the terpene synthase on the fermentation yield is discussed, and the preliminary studies on the engineering of this key enzyme are presented. PMID:25379342

  20. Three-stage fermentation and kinetic modeling of bioflocculant by Corynebacterium glutamicum

    Institute of Scientific and Technical Information of China (English)

    Liang Shen; Zhongtao An; Qingbiao Li; Chuanyi Yao; Yajuan Peng; Yuanpeng Wang; Ruihua Lai; Xu Deng; Ning He


    Fermentation of bioflocculant with Corynebacterium glutamicum was studied by way of kinetic modeling. Lorentzian modified Logistic model, time-corrected Luedeking–Piret and Luedeking–Piret type models were pro-posed and applied to describe the cell growth, bioflocculant synthesis and consumption of substrates, with the correlation of initial biomass concentration and initial glucose concentration, respectively. The results showed that these models could well characterize the batch culture process of C. glutamicum at various initial glucose con-centrations from 10.0 to 17.5 g·L−1. The initial biomass concentration could shorten the lag time of cel growth, while the maximum biomass concentration was achieved only at the optimal initial glucose concentration of 16.22 g·L−1. A novel three-stage fed-batch strategy for bioflocculant production was developed based on the model prediction, in which the lag phase, quick biomass growth and bioflocculant production stages were sequentially proceeded with the adjustment of glucose concentration and dissolved oxygen. Biomass of 2.23 g·L−1 was obtained and bioflocculant concentration was enhanced to 176.32 mg·L−1, 18.62% and 403.63%higher than those in the batch process, respectively, indicating an efficient fed-batch culture strategy for bioflocculant production.

  1. Stable high-titer n-butanol production from sucrose and sugarcane juice by Clostridium acetobutylicum JB200 in repeated batch fermentations. (United States)

    Jiang, Wenyan; Zhao, Jingbo; Wang, Zhongqiang; Yang, Shang-Tian


    The production of n-butanol, a widely used industrial chemical and promising transportation fuel, from abundant, low-cost substrates, such as sugarcane juice, in acetone-butanol-ethanol (ABE) fermentation was studied with Clostridium acetobutylicum JB200, a mutant with high butanol tolerance and capable of producing high-titer (>20 g/L) n-butanol from glucose. Although JB200 is a favorable host for industrial bio-butanol production, its fermentation performance with sucrose and sugarcane juice as substrates has not been well studied. In this study, the long-term n-butanol production from sucrose by JB200 was evaluated with cells immobilized in a fibrous-bed bioreactor (FBB), showing stable performance with high titer (16-20 g/L), yield (∼ 0.21 g/g sucrose) and productivity (∼ 0.32 g/Lh) for 16 consecutive batches over 800 h. Sugarcane thick juice as low-cost substrate was then tested in 3 consecutive batches, which gave similar n-butanol production, demonstrating that JB200 is a robust and promising strain for industrial ABE fermentation. Copyright © 2014 Elsevier Ltd. All rights reserved.

  2. Fermentation strategies for 1,3-propanediol production from glycerol using a genetically engineered Klebsiella pneumoniae strain to eliminate by-product formation. (United States)

    Oh, Baek-Rock; Seo, Jeong-Woo; Heo, Sun-Yeon; Hong, Won-Kyung; Luo, Lian Hua; Son, Jun Ho; Park, Don-Hee; Kim, Chul-Ho


    We generated a genetically engineered Klebsiella pneumoniae strain (AK-VOT) to eliminate by-product formation during production of 1,3-propanediol (1,3-PD) from glycerol. In the present study, the glycerol-metabolizing properties of the recombinant strain were examined during fermentation in a 5 L bioreactor. As expected, by-product formation was completely absent (except for acetate) when the AK-VOT strain fermented glycerol. However, 1,3-PD productivity was severely reduced owing to a delay in cell growth attributable to a low rate of glycerol consumption. This problem was solved by establishing a two-stage process separating cell growth from 1,3-PD production. In addition, nutrient co-supplementation, especially with starch, significantly increased 1,3-PD production from glycerol during fed-batch fermentation by AK-VOT in the absence of by-product formation.

  3. Open fermentative production of L-lactic acid by Bacillus sp. strain NL01 using lignocellulosic hydrolyzates as low-cost raw material. (United States)

    Ouyang, Jia; Ma, Rui; Zheng, Zhaojuan; Cai, Cong; Zhang, Min; Jiang, Ting


    Highly efficient L-lactate production by a thermophilic strain Bacillus sp. NL01 was demonstrated in this study. Lignocellulosic hydrolyzates containing a high content of glucose, which was prepared from corn stover, was used as substrate for L-lactic acid production. The fermentation was carried out under open condition without sterilization and used NaOH as alkaline neutralizing reagent. In batch fermentation, 56.37 g l(-1) L-lactic acid was obtained from lignocellulosic hydrolyzates which contained the solid residues produced in enzymatic saccharification. In fed-batch fermentation, 75.03 g l(-1) L-lactic acid was obtained from lignocellulosic hydrolyzates supernatant. The yield was 74.5% and the average productivity was 1.04 g l(-1) h(-1). Crown Copyright © 2012. Published by Elsevier Ltd. All rights reserved.

  4. The preparation and ethanol fermentation of high-concentration sugars from steam-explosion corn stover. (United States)

    Xie, Hui; Wang, Fengqin; Yin, Shuangyao; Ren, Tianbao; Song, Andong


    In the field of biofuel ethanol, high-concentration- reducing sugars made from cellulosic materials lay the foundation for high-concentration ethanol fermentation. In this study, corn stover was pre-treated in a process combining chemical methods and steam explosion; the cellulosic hydrolyzed sugars obtained by fed-batch saccharification were then used as the carbon source for high-concentration ethanol fermentation. Saccharomyces cerevisiae 1308, Angel yeast, and Issatchenkia orientalis were shake-cultured with Pachysolen tannophilus P-01 for fermentation. Results implied that the ethanol yields from the three types of mixed strains were 4.85 g/100 mL, 4.57 g/100 mL, and 5.02 g/100 mL (separately) at yield rates of 91.6, 89.3, and 92.2%, respectively. Therefore, it was inferred that shock-fermentation using mixed strains achieved a higher ethanol yield at a greater rate in a shorter fermentation period. This study provided a theoretical basis and technical guidance for the fermentation of industrial high-concentrated cellulosic ethanol.

  5. Combined process for ethanol fermentation at high-solids loading and biogas digestion from unwashed steam-exploded corn stover. (United States)

    Wang, Zhen; Lv, Zhe; Du, Jiliang; Mo, Chunling; Yang, Xiushan; Tian, Shen


    A combined process was designed for the co-production of ethanol and methane from unwashed steam-exploded corn stover. A terminal ethanol titer of 69.8 g/kg mass weight (72.5%) was achieved when the fed-batch mode was performed at a final solids loading of 35.5% (w/w) dry matter (DM) content. The whole stillage from high-solids ethanol fermentation was directly transferred in a 3-L anaerobic digester. During 52-day single-stage digester operation, the methane productivity was 320 mL CH₄/g volatile solids (VS) with a maximum VS reduction efficiency of 55.3%. The calculated overall product yield was 197 g ethanol + 96 g methane/kg corn stover. This indicated that the combined process was able to improve overall content utilization and extract a greater yield of lignocellulosic biomass compared to ethanol fermentation alone.

  6. Study and management of atmospheric exhaust gas in acetic acid fermentation developing a new process for alcohol vinegar production; Etude et maitrise des rejets atmospheriques en vinaigrerie, developpement d'un procede nouveau de conduite de la fermentation acetique

    Energy Technology Data Exchange (ETDEWEB)

    Pochat Bohatier, C.


    The aim of the study was to examine emissions of volatile organic compounds (VOCs) during acetic (acid) fermentation. The first part of the study presents the methodology developed to reproduce production cycles for spirit vinegar and to analyse gas effluents. The second part describes the origin and quantification of the emissions (ethanol, acetic acid, acetaldehyde and ethyl acetate). The acetic acid is produced by bacterial metabolism while the ethyl acetate is a result of the chemical reaction of esterification. By modelling the emissions during batch processing we were able to identify the various parameters involved when VOCs are carried along by the fermentation gases. The quantities of ethyl acetate depend on the length of time the diluted alcohol is stored, and on its composition. By using a fed-batch method with a continuous supply of ethanol we could reduce alcohol emissions. The third part of the study develops the kinetics studies carried out to adapt the fed-batch process to acetic acid fermentation. The influence of ethanol, either in terms of deficiency or inhibition, is minimized between 8 and 16 g.1-1. A study of the growth rate of bacteria in relation to the amount of acetic acid showed that the latter was highly inhibitive. There is a critical concentration of acetic acid at which the growth of bacteria stops, and the death rate of the culture increases rapidly. The latter depends on the composition of the culture's medium; the corresponding pH of the concentration is between 2.25 and 2.28. By limiting the formation of ethyl acetate in the diluted alcohol and by controlling the concentration of ethanol at 16 g.l -1 per fermentation, the VOC emissions are reduced by 30% and the yield increases as a result. (author)

  7. The impact of pH inhomogeneities on CHO cell physiology and fed-batch process performance - two-compartment scale-down modelling and intracellular pH excursion. (United States)

    Brunner, Matthias; Braun, Philipp; Doppler, Philipp; Posch, Christoph; Behrens, Dirk; Herwig, Christoph; Fricke, Jens


    Due to high mixing times and base addition from top of the vessel, pH inhomogeneities are most likely to occur during large-scale mammalian processes. The goal of this study was to set-up a scale-down model of a 10-12 m(3) stirred tank bioreactor and to investigate the effect of pH perturbations on CHO cell physiology and process performance. Short-term changes in extracellular pH are hypothesized to affect intracellular pH and thus cell physiology. Therefore, batch fermentations, including pH shifts to 9.0 and 7.8, in regular one-compartment systems are conducted. The short-term adaption of the cells intracellular pH are showed an immediate increase due to elevated extracellular pH. With this basis of fundamental knowledge, a two-compartment system is established which is capable of simulating defined pH inhomogeneities. In contrast to state-of-the-art literature, the scale-down model is included parameters (e.g. volume of the inhomogeneous zone) as they might occur during large-scale processes. pH inhomogeneity studies in the two-compartment system are performed with simulation of temporary pH zones of pH 9.0. The specific growth rate especially during the exponential growth phase is strongly affected resulting in a decreased maximum viable cell density and final product titer. The gathered results indicate that even short-term exposure of cells to elevated pH values during large-scale processes can affect cell physiology and overall process performance. In particular, it could be shown for the first time that pH perturbations, which might occur during the early process phase, have to be considered in scale-down models of mammalian processes.

  8. The enhancement of butanol production by in situ butanol removal using biodiesel extraction in the fermentation of ABE (acetone-butanol-ethanol). (United States)

    Yen, Hong-Wei; Wang, Yi-Cheng


    High butanol accumulation is due to feedback inhibition which leads to the low butanol productivity observed in acetone-butanol-ethanol (ABE) fermentation. The aim of this study is to use biodiesel as an extractant for the in situ removal of butanol from the broth. The results indicate that adding biodiesel as an extractant at the beginning of fermentation significantly enhances butanol production. No significant toxicity of biodiesel on the growth of Clostridium acetobutylicum is observed. In the fed-batch operation with glucose feeding, the maximum total butanol obtained is 31.44 g/L, as compared to the control batch (without the addition of biodiesel) at 9.85 g/L. Moreover, the productivity obtained is 0.295 g/L h in the fed-batch, which is higher than that of 0.185 g/L h for the control batch. The in situ butanol removal by the addition of biodiesel has great potential for commercial ABE production. Copyright © 2012 Elsevier Ltd. All rights reserved.

  9. Formulation of fermentation media from flour-rich waste streams for microbial lipid production by Lipomyces starkeyi. (United States)

    Tsakona, Sofia; Kopsahelis, Nikolaos; Chatzifragkou, Afroditi; Papanikolaou, Seraphim; Kookos, Ioannis K; Koutinas, Apostolis A


    Flour-rich waste (FRW) and by-product streams generated by bakery, confectionery and wheat milling plants could be employed as the sole raw materials for generic fermentation media production, suitable for microbial oil synthesis. Wheat milling by-products were used in solid state fermentations (SSF) of Aspergillus awamori for the production of crude enzymes, mainly glucoamylase and protease. Enzyme-rich SSF solids were subsequently employed for hydrolysis of FRW streams into nutrient-rich fermentation media. Batch hydrolytic experiments using FRW concentrations up to 205 g/L resulted in higher than 90% (w/w) starch to glucose conversion yields and 40% (w/w) total Kjeldahl nitrogen to free amino nitrogen conversion yields. Starch to glucose conversion yields of 98.2, 86.1 and 73.4% (w/w) were achieved when initial FRW concentrations of 235, 300 and 350 g/L were employed in fed-batch hydrolytic experiments, respectively. Crude hydrolysates were used as fermentation media in shake flask cultures with the oleaginous yeast Lipomyces starkeyi DSM 70296 reaching a total dry weight of 30.5 g/L with a microbial oil content of 40.4% (w/w), higher than that achieved in synthetic media. Fed-batch bioreactor cultures led to a total dry weight of 109.8 g/L with a microbial oil content of 57.8% (w/w) and productivity of 0.4 g/L/h. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. Production of butanol from bagasse and molasses by mixed fermentation%甘蔗渣和糖蜜混合发酵制备燃料丁醇

    Institute of Scientific and Technical Information of China (English)

    杜腾飞; 郭亭; 汤艳; 贺爱永; 姜岷


    以抗逆突变株Clostridium beijerinckii IB4为研究对象,葡萄糖为C源,对其进行补料分批发酵过程的优化,同时将该优化工艺应用于甘蔗渣和糖蜜混合发酵制备燃料丁醇.结果表明:在5L发酵罐中,先加入作为还原糖的甘蔗渣酸解糖液10 g/L,16 h后补加甘蔗糖蜜30 g/L,于35 ℃、100 r/min发酵50 h,丁醇和总溶剂产量分别达到11.1和15.3 g/L,丁醇比例高达72.5%.%A tolerance mutant strain Clostridium beijerinckii IB4 was selected. Fed-batch fermentations were carried out in P2 fermentation medium with glucose, and the optimal fed-batch fermentation strategy was applied to butanol mixed fermentation from bagasse and molasses. Clostridium beijerinckii IB4 was cultured in a 5 L fermentor at 35 ℃ , 100 r/min for 50 h. The initial 10 g/L reducing sugar of acid hy-drolysate of bagasse was added, and 30 g/L of molasses was fed into fermentation medium after 16 h. Butanol and total solvents concentrations accumulated in the broth were 11.1 and 15. 3 g/L, respectively. Butanol proportion was up to 72. 5%.

  11. Metabolic approaches for the optimisation of recombinant fermentation processes. (United States)

    Cserjan-Puschmann, M; Kramer, W; Duerrschmid, E; Striedner, G; Bayer, K


    The aim of this work was the establishment of a novel method to determine the metabolic load on host-cell metabolism resulting from recombinant protein production in Escherichia coli. This tool can be used to develop strategies to optimise recombinant fermentation processes through adjustment of recombinant-protein expression to the biosynthetic capacity of the host-cell. The signal molecule of the stringent-response network, guanosine tetraphosphate (ppGpp), and its precursor nucleotides were selected for the estimation of the metabolic load relating to recombinant-protein production. An improved analytical method for the quantification of nucleotides by ion-pair, high-performance liquid chromatography was established. The host-cell response upon overexpression of recombinant protein in fed-batch fermentations was investigated using the production of human superoxide dismutase (rhSOD) as a model system. E. coli strains with different recombinant systems (the T7 and pKK promoter system) exerting different loads on host-cell metabolism were analysed with regard to intracellular nucleotide concentration, rate of product formation and plasmid copy number.

  12. Immobilized ethanol fermentation coupled to pervaporation with silicalite-1/polydimethylsiloxane/polyvinylidene fluoride composite membrane. (United States)

    Cai, Di; Hu, Song; Chen, Changjing; Wang, Yong; Zhang, Changwei; Miao, Qi; Qin, Peiyong; Tan, Tianwei


    A novel silicalite-1/polydimethylsiloxane/polyvinylidene fluoride hybrid membrane was used in ethanol fermentation-pervaporation integration process. The sweet sorghum bagasse was used as the immobilized carrier. Compared with the conventional suspend cells system, the immobilized fermentation system could provide higher ethanol productivity when coupled with pervaporation. In the long-term of operations, the ethanol productivity, separation factor, total flux and permeate ethanol concentration in the fed-batch fermentation-pervaporation integration scenario were 1.6g/Lh, 8.2-9.9, 319-416g/m(2)h and 426.9-597.2g/L, respectively. Correspondingly, 1.6g/Lh, 7.8-9.8, 227.8-395g/m(2)h and 410.9-608.1g/L were achieved in the continuous fermentation-pervaporation integration scenario, respectively. The results indicated that the integration process could greatly improve the ethanol production and separation performances.

  13. Evaluation of recent advances in butanol fermentation, upstream, and downstream processing

    Energy Technology Data Exchange (ETDEWEB)

    Qureshi, N.; Blaschek, H.P. [Biotechnology and Bioengineering Group, Univ. of Illinois, IL (United States)


    Four different processes for butanol production from corn, namely, batch fermentation and distillative recovery (BFDR), batch fermentation and pervaporative recovery (BFPR), fed-batch fermentation and pervaporative recovery (FBFPR), and immobilized cell continuous fermentation and pervaporative recovery (ICCFPR) were evaluated. Pervaporative recovery significantly reduces the cost of butanol production. Depending upon the byproduct credit, which is approximately 3.7 times that of the amount of butanol produced, BFDR, BFPR, FBFPR, and ICCFPR result in a butanol price of $0.55, $0.14-0.39, $0.12-0.37, and $0.11-0.36 x kg{sup -1}, respectively. The price of butanol was recently reported at $1.21 x kg{sup -1} by Chemical Marketing Reporter. It should be noted that all three components (acetone, butanol, and ethanol: ABE) diffuse through the pervaporation membrane. Further separation and purification of the solvents would require distillation, which has been considered in this exercise. This article also details the impact of byproduct credit, rate of return, and tax on butanol price. (orig.)

  14. 2,3-Butanediol recovery from fermentation broth by alcohol precipitation and vacuum distillation. (United States)

    Jeon, Sangjun; Kim, Duk-Ki; Song, Hyohak; Lee, Hee Jong; Park, Sunghoon; Seung, Doyoung; Chang, Yong Keun


    This study presents a new and effective downstream process to recover 2,3-butanediol (2,3-BD) from fermentation broth which is produced by a recombinant Klebsiella pneumoniae strain. The ldhA-deficient K. pneumoniae strain yielded about 90 g/L of 2,3-BD, along with a number of by-products, such as organic acids and alcohols, in a 65 h fed-batch fermentation. The pH-adjusted cell-free fermentation broth was firstly concentrated until 2,3-BD reached around 500 g/L by vacuum evaporation at 50°C and 50 mbar vacuum pressure. The concentrated solution was further treated using light alcohols, including methanol, ethanol, and isopropanol, for the precipitation of organic acids and inorganic salts. Isopropanol showed the highest removal efficiency, in which 92.5% and 99.8% of organic acids and inorganic salts were precipitated, respectively. At a final step, a vacuum distillation process enabled the recovery of 76.2% of the treated 2,3-BD, with 96.1% purity, indicating that fermentatively produced 2,3-BD is effectively recovered by a simple alcohol precipitation and vacuum distillation. Copyright © 2013 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  15. Prevention of GABA reduction during dough fermentation using a baker's yeast dal81 mutant. (United States)

    Ando, Akira; Nakamura, Toshihide


    γ-Aminobutyric acid (GABA) is consumed by yeasts during fermentation. To prevent GABA reduction in bread dough, a baker's yeast mutant AY77 deficient in GABA assimilation was characterized and utilized for wheat dough fermentation. An amber mutation in the DAL81 gene, which codes for a positive regulator of multiple nitrogen degradation pathways, was found in the AY77 strain. The qPCR analyses of genes involved in nitrogen utilization showed that transcriptional levels of the UGA1 and DUR3 genes encoding GABA transaminase and urea transporter, respectively, are severely decreased in the AY77 cells. The AY77 strain cultivated by fed-batch culture using cane molasses exhibited inferior gas production during dough fermentation compared to that of wild-type strain AY13. However, when fed with molasses containing 0.5% ammonium sulfate, the mutant strain exhibited gas production comparable to that of the AY13 strain. In contrast to the AY13 strain, which completely consumed GABA in dough within 5 h, the AY77 strain consumed no GABA under either culture condition. Dough fermentation with the dal81 mutant strain should be useful for suppression of GABA reduction in breads.

  16. Recovery of purified lactonic sophorolipids by spontaneous crystallization during the fermentation of sugarcane molasses with Candida albicans O-13-1. (United States)

    Yang, Xue; Zhu, Lingqing; Xue, Chaoyou; Chen, Yu; Qu, Liang; Lu, Wenyu


    Numerous studies have focused on how to obtain high yield of sophorolipids using low-cost materials as substrates, and there has been various work on the experimental methods for purifying lactonic sophorolipids. These studies have not yet obtained satisfied results in combining a low-cost fermentation process and the purification of lactonic sophorolipids. This study establishes a fed-batch fermentation process of purifying sophorolipids from Candida albicans O-13-1 using low-cost sugarcane molasses as the substrate. In the optimized conditions of this research, using sugarcane molasses as a substrate and product synthesis based on the temperature stage-controlled fermentation, our result indicates that sophorolipids production could reach 108.7 g/L. More importantly, lactonic sophorolipids can crystallize and precipitate during our established fermentation process. The structures and content of sophorolipids separated from the fermentation broth and sophorolipids crystallized in the fermentation broth were analyzed by a scanning electron microscope (SEM) and liquid chromatography-mass spectrometry (LC-MS). The fermentation process produced 90.5 g/L crystallized lactonic sophorolipids with 90.51% purity. This is an energy-saving and low-cost method to obtain such pure lactonic sophorolipids. Copyright © 2012 Elsevier Inc. All rights reserved.

  17. Optimization of construct design and fermentation strategy for the production of bioactive ATF-SAP, a saporin based anti-tumoral uPAR-targeted chimera. (United States)

    Errico Provenzano, Alfredo; Posteri, Riccardo; Giansanti, Francesco; Angelucci, Francesco; Flavell, Sopsamorn U; Flavell, David J; Fabbrini, Maria Serena; Porro, Danilo; Ippoliti, Rodolfo; Ceriotti, Aldo; Branduardi, Paola; Vago, Riccardo


    The big challenge in any anti-tumor therapeutic approach is represented by the development of drugs selectively acting on the target with limited side effects, that exploit the unique characteristics of malignant cells. The urokinase (urokinase-type plasminogen activator, uPA) and its receptor uPAR have been identified as preferential target candidates since they play a key role in the evolution of neoplasms and are associated with neoplasm aggressiveness and poor clinical outcome in several different tumor types. To selectively target uPAR over-expressing cancer cells, we prepared a set of chimeric proteins (ATF-SAP) formed by the human amino terminal fragments (ATF) of uPA and the plant ribosome inactivating protein saporin (SAP). Codon-usage optimization was used to increase the expression levels of the chimera in the methylotrophic yeast Pichia pastoris. We then moved the bioprocess to bioreactors and demonstrated that the fed-batch production of the recombinant protein can be successfully achieved, obtaining homogeneous discrete batches of the desired constructs. We also determined the cytotoxic activity of the obtained batch of ATF-SAP which was specifically cytotoxic for U937 leukemia cells, while another construct containing a catalytically inactive mutant form of SAP showed no activity. Our results demonstrate that the uPAR-targeted, saporin-based recombinant fusion ATF-SAP can be produced in a fed-batch fermentation with full retention of the molecules selective cytotoxicity and hence therapeutic potential.

  18. Evaluation of asymmetric polydimethylsiloxane-polyvinylidene fluoride composite membrane and incorporated with acetone-butanol-ethanol fermentation for butanol recovery. (United States)

    Xue, Chuang; Du, Guang-Qing; Chen, Li-Jie; Ren, Jian-Gang; Bai, Feng-Wu


    The polydimethylsiloxane-polyvinylidene fluoride (PDMS-PVDF) composite membrane was studied for its pervaporation performance to removal of butanol from butanol/ABE solution, fermentation broth as well as incorporated with acetone-butanol-ethanol (ABE) fermentation. The total flux and butanol titer in permeate through the PDMS-PVDF membrane were up to 769.6 g/m(2)h and 323.5 g/L at 80 °C, respectively. The butanol flux and total flux increased with increasing the feed temperature as well as the feed butanol titer. The butanol separation factor and butanol titer in permeate decreased slightly in the presence of acetone and ethanol in the feed due to their preferential dissolution and competitive permeation through the membrane. In fed-batch fermentation incorporated with pervaporation, butanol titer and flux in permeate maintained at a steady level with the range of 139.9-154.0 g/L and 13.3-16.3 g/m(2)h, respectively, which was attributed to the stable butanol titer in fermentation broth as well as the excellent hydrophobic nature of the PDMS-PVDF matrix. Therefore, the PDMS-PVDF composite membrane had a great potential in the in situ product recovery with ABE fermentation, enabling the economic production of biobutanol. Copyright © 2014 Elsevier B.V. All rights reserved.

  19. Bioethanol production from the dry powder of Jerusalem artichoke tubers by recombinant Saccharomyces cerevisiae in simultaneous saccharification and fermentation. (United States)

    Wang, Yi-Zhou; Zou, Shan-Mei; He, Mei-Lin; Wang, Chang-Hai


    It has been found that recombinant Saccharomyces cerevisiae 6525 can produce high concentration of ethanol in one-step fermentation from the extract of Jerusalem artichoke tubers or inulin. However, the utilization rate of raw materials was low and the fermentation process was costly and complicated. Therefore, in this study, after the optimum processing conditions for ethanol production in fed-batch fermentation were determined in flask, the recombinant S. cerevisiae 6525 was first used to produce ethanol from the dry powder of Jerusalem artichoke tubers in 5-L agitating fermentor. After 72 h of fermentation, around 84.3 g/L ethanol was produced in the fermentation liquids, and the conversion efficiency of inulin-type sugars to ethanol was 0.453, or 88.6 % of the theoretical value of 0.511. This study showed high feasibility of bioethanol industrial production from the Jerusalem artichoke tubers and provided a basis for it in the future.

  20. High temperature simultaneous saccharification and fermentation of starch from inedible wild cassava (Manihot glaziovii) to bioethanol using Caloramator boliviensis. (United States)

    Moshi, Anselm P; Hosea, Ken M M; Elisante, Emrode; Mamo, G; Mattiasson, Bo


    The thermoanaerobe, Caloramator boliviensis was used to ferment starch hydrolysate from inedible wild cassava to ethanol at 60°C. A raw starch degrading α-amylase was used to hydrolyse the cassava starch. During fermentation, the organism released CO2 and H2 gases, and Gas Endeavour System was successfully used for monitoring and recording formation of these gaseous products. The bioethanol produced in stoichiometric amounts to CO2 was registered online in Gas Endeavour software and correlated strongly (R(2)=0.99) with values measured by HPLC. The organism was sensitive to cyanide that exists in cassava flour. However, after acclimatisation, it was able to grow and ferment cassava starch hydrolysate containing up to 0.2ppm cyanide. The reactor hydrogen partial pressure had influence on the bioethanol production. In fed-batch fermentation by maintaining the hydrogen partial pressure around 590Pa, the organism was able to ferment up to 76g/L glucose and produced 33g/L ethanol.

  1. Online determination of viable biomass up to very high cell densities in Arxula adeninivorans fermentations using an impedance signal. (United States)

    Knabben, Ingo; Regestein, Lars; Grumbach, Carsten; Steinbusch, Sven; Kunze, Gotthard; Büchs, Jochen


    Up to now biomass has been measured online by impedance analysis only at low cell densities in yeast fermentations. As industrial fermentation processes focus, for example, on producing high target concentrations of biocatalysts or pharmaceutical proteins, it is important to investigate cell growth under high cell-density conditions. Therefore, for the first time, biomass has been measured online using impedance analysis in a 50L high-pressure stirred tank reactor. As model organism the yeast Arxula adeninivorans was cultivated in two chemically defined mineral media at a constant growth rate in fed-batch mode. To ensure aerobic culture conditions over the entire fermentation time, the fermentations were conducted at an elevated headspace overpressure of up to 9.5bar. The highest oxygen transfer rate value of 0.56molL(-1)h(-1) ever reported for yeast fermentations was measured in these investigations. Unlike previous findings, in this study a linear correlation was found between capacitance and biomass up to concentrations of 174gL(-1) dry cell weight.

  2. Probing control of fed-batch cultivations: analysis and tuning

    DEFF Research Database (Denmark)

    Åkesson, Mats Fredrik; Hagander, P.; Axelsson, J.P.


    Production of various proteins can today be made using genetically modified Escherichia coli bacteria. In cultivations of E. coli it is important to avoid accumulation of the by- product acetate. Formation of acetate occurs when the specific glucose uptake exceeds a critical value and can...... be avoided by a proper feeding strategy. A difficulty is that the critical glucose uptake often is poorly known and even time varying. We here analyze an approach for control of glucose feeding that enables feeding at the critical glucose uptake without prior information. The key idea is to superimpose...... a probing signal to the feed rate in order to obtain information used to determine if the feed rate should be increased or decreased. The main contribution of this paper is to derive guidelines for tuning of the probing controller. A sufficient condition for stability is presented. By introducing...

  3. Fed-Batch Feeding Strategies for Enzymatic Biodiesel Production

    DEFF Research Database (Denmark)

    Price, Jason Anthony; Nordblad, Mathias; Woodley, John


    of the differences in the interfacial and bulk concentrations of the enzyme. The model is then used to evaluate various feeding strategies to improve the enzymatic biodiesel production. The feeding strategies investigated, gave insight into how the methanol should be fed to potentially mitigate enzyme deactivation...... while improving the biodiesel yield. The best experimental results gave a yield of 703 .76 g FAME L-1 and a reactor productivity of 28.12 g FAME L-1 h-1. In comparison, to reach the same yield, the optimised two step feeding strategy took 6.25 hours less, which equates to an increase the reactor...

  4. A Comparative Study of Temperature Optimal Control in a Solid State Fermentation Process for Edible Mushroom Growing

    Directory of Open Access Journals (Sweden)

    K. J. Gurubel


    Full Text Available In this paper, optimal control strategies for temperature trajectory determination in order to maximize thermophilic bacteria in a fed-batch solid-state fermentation reactor are proposed. This process is modeled by nonlinear differential equations, which has been previously validated experimentally with scale reactor temperature profiles. The dynamic input aeration rate of the reactor is determined to increase microorganisms growth of a selective substrate for edible mushroom cultivation. In industrial practice, the process is comprised of three thermal stages with constant input air flow and three types of microorganisms in a 150-hour lapse. Scytalidium thermophilum and actinobacteria are desired in order to obtain a final biomass composition with acceptable microorganisms concentration. The Steepest Descent gradient algorithm in continuous time and the Gradient Projection algorithm in discrete-time are used for the process optimal control. A comparison of simulation results in the presence of disturbances is presented, where the resulting temperature trajectories exhibit similar tendencies as industrial data.

  5. Breeding of Coenzyme Q10 Produced Strain by Low-Energy Ion Implantation and Optimization of Coenzyme Q10 Fermentation

    Institute of Scientific and Technical Information of China (English)

    XU Dejun; ZHENG Zhiming; WANG Peng; WANG Li; YUAN Hang; YU Zengliang


    In order to increase the production efficiency of coenzyme Q10, the original strain Agrobacterium tumefaciens ATCC 4452 was mutated by means of Nitrogen ions implantation. A mutant strain, ATX 12, with high contents of coenzyme Q10 was selected. Subsequently, the conditions such as carbohydrate concentration, nitrogen source concentration, inoculum's size, seed age, aeration and temperature which might affect the production of CoQ10 were investigated in detail. Under optimal conditions, the maximum concentration of the intracellular CoQ10 reached 200.3 mg/L after 80 h fed-batch fermentation, about 245% increasing in CoQ10 production after ion implantation, compared to the original strain.

  6. [Drug development from natural fermentation products: establishing a manufacturing process which maximizes the potential of microorganisms]. (United States)

    Nagao, Koji; Ueda, Satoshi; Kanda, Munekazu; Oohata, Nobutaka; Yamashita, Michio; Hino, Motohiro


    Natural fermentation products have long been studied as attractive targets for drug discovery due to their amazing diverse, complex chemical structures and biological activities. As such, a number of revolutionary drugs developed from natural fermentation products have contributed to global human health. To commercialize a drug derived from natural fermentation products, an effective chemical entity must be identified and thoroughly researched, and an effective manufacturing process to prepare a commercial supply must be developed. To construct such a manufacturing process for tacrolimus and micafungin, the following studies were conducted: first, we focused on controlling the production of the tacrolimus-related compound FR900525, a fermentation by-product of tacrolimus which was critical for quality assurance of the drug substance. FR900525 production was reduced by using a mutant strain which produced more pipecolic acid, the biosynthesis material of tacrolimus, than the original strain. Then, to optimize the fermentation process of FR901379, an intermediate of micafungin, a fed-batch culture was adopted to increase FR901379 productivity. Additionally, FULLZONE(TM) impeller was installed into the scaled-up fermenter, reducing the agitation-induced damage to the mycelium. As a result, the mycelial form changed from filamentous to pellet-shaped, and the air uptake rate during fermentation was drastically improved. Finally, we conducted screening for FR901379 acylase-producing microorganisms, as FR901379 acylase is necessary to manufacture micafungin. We were able to easily discover FR901379 acylase-producing microorganisms in soil samples using our novel, convenient screening method, which involves comparing the difference in antibiotic activity between FR901379 and its deacylated product.

  7. Acetone-butanol-ethanol (ABE) fermentation using Clostridium acetobutylicum XY16 and in situ recovery by PDMS/ceramic composite membrane. (United States)

    Wu, Hao; Chen, Xiao-Peng; Liu, Gong-Ping; Jiang, Min; Guo, Ting; Jin, Wan-Qin; Wei, Ping; Zhu, Da-Wei


    PDMS/ceramic composite membrane was directly integrated with acetone-butanol-ethanol (ABE) fermentation using Clostridium acetobutylicum XY16 at 37 °C and in situ removing ABE from fermentation broth. The membrane was integrated with batch fermentation, and approximately 46 % solvent was extracted. The solvent in permeates was 118 g/L, and solvent productivity was 0.303 g/(L/h), which was approximately 33 % higher compared with the batch fermentation without in situ recovery. The fed-batch fermentation with in situ recovery by pervaporation continued for more than 200 h, 61 % solvent was extracted, and the solvent in penetration was 96.2 g/L. The total flux ranged from 0.338 to 0.847 kg/(m(2)/h) and the separation factor of butanol ranged from 5.1 to 27.1 in this process. The membrane was fouled by the active fermentation broth, nevertheless the separation performances were partially recovered by offline membrane cleaning, and the solvent productivity was increased to 0.252 g/(L/h), which was 19 % higher compared with that in situ recovery process without membrane cleaning.

  8. Assessment of the manufacturability of Escherichia coli high cell density fermentations. (United States)

    Perez-Pardo, M A; Ali, S; Balasundaram, B; Mannall, G J; Baganz, F; Bracewell, D G


    The physical and biological conditions of the host cell obtained at the end of fermentation influences subsequent downstream processing unit operations. The ability to monitor these characteristics is central to the improvement of biopharmaceutical manufacture. In this study, we have used a combination of techniques such as adaptive focus acoustics (AFA) and ultra scale-down (USD) centrifugation that utilize milliliter quantities of sample to obtain an insight into the interaction between cells from the upstream process and initial downstream unit operations. This is achieved primarily through an assessment of cell strength and its impact on large-scale disc stack centrifugation performance, measuring critical attributes such as viscosity and particle size distribution. An Escherichia coli fed-batch fermentation expressing antibody fragments in the periplasm was used as a model system representative of current manufacturing challenges. The weakening of cell strength during cultivation time, detected through increased micronization and viscosity, resulted in a 2.6-fold increase in product release rates from the cell (as measured by AFA) and approximately fourfold decrease in clarification performance (as measured by USD centrifugation). The information obtained allows for informed harvest point decisions accounting for both product leakages during fermentation and potential losses during primary recovery. The clarification performance results were verified at pilot scale. The use of these technologies forms a route to the process understanding needed to tailor the host cell and upstream process to the product and downstream process, critical to the implementation of quality-by-design principles.

  9. Reduction of foaming and enhancement of ascomycin production in rational Streptomyces hygroscopicus fermentation

    Institute of Scientific and Technical Information of China (English)

    Xing Xin; Haishan Qi; Jianping Wen; Xiaoqiang Jia; Yunlin Chen


    Foaming reduces the working volume and limits the biosynthesis of macrolide immunosuppressant ascomycin (FK520) in the batch fermentation process of Streptomyces hygroscopicus FS-35 in a 7.5 L bioreactor. To find the relation between FK520 production and foaming, effects of 10 fermentation parameters including organic acids and membrane permeability were investigated. The results suggest that acetate accumulation caused by short period oxygen deficiency and fast consumption of glucose is the reason for increased foaming and declined FK520 production. Therefore, a fed-batch fermentation strategy was developed to reduce the accumulation of ac-etate. After optimization, the maximum acetate concentration dropped from 320 mg·L−1 to 157 mg·L−1, de-creased by 50.8%, and the maximum foam height reduced from 5.32 cm to 3.74 cm, decreased by 29.7%, while the maximum FK520 production increased from 375 mg·L−1 to 421 mg·L−1, improved by 12%.

  10. Fermentation, fractionation and purification of streptokinase by chemical reduction method

    Directory of Open Access Journals (Sweden)

    M Niakan


    Full Text Available Background and Objectives: Streptokinase is used clinically as an intravenous thrombolytic agent for the treatment of acute myocardial infarction and is commonly prepared from cultures of Streptococcus equisimilis strain H46A. The objective of the present study was the production of streptokinase from strain H46A and purification by chemical reduction method."nMaterials and Methods: The rate of streptokinase production evaluated under the effect of changes on some fermentation factors. Moreover, due to the specific structure of streptokinase, a chemical reduction method employed for the purification of streptokinase from the fermentation broth. The H46A strain of group C streptococcus, was grown in a fermentor. The proper pH adjusted with NaOH under glucose feeding in an optimum temperature. The supernatant of the fermentation product was sterilized by filtration and concentrated by ultrafiltration. The pH of the concentrate was adjusted, cooled, and precipitated by methanol. Protein solution was reduced with dithiothreitol (DTT. Impurities settled down by aldrithiol-2 and the biological activity of supernatant containing streptokinase was determined."nResults: In the fed -batch culture, the rate of streptokinase production increased over two times as compared with the batch culture and the impurities were effectively separated from streptokinase by reduction method."nConclusion: Improvements in SK production are due to a decrease in lag phase period and increase in the growth rate of logarithmic phase. The methods of purification often result in unacceptable losses of streptokinase, but the chemical reduction method give high yield of streptokinase and is easy to perform it.

  11. Analysis of L-glutamic acid fermentation by using a dynamic metabolic simulation model of Escherichia coli. (United States)

    Nishio, Yousuke; Ogishima, Soichi; Ichikawa, Masao; Yamada, Yohei; Usuda, Yoshihiro; Masuda, Tadashi; Tanaka, Hiroshi


    Understanding the process of amino acid fermentation as a comprehensive system is a challenging task. Previously, we developed a literature-based dynamic simulation model, which included transcriptional regulation, transcription, translation, and enzymatic reactions related to glycolysis, the pentose phosphate pathway, the tricarboxylic acid (TCA) cycle, and the anaplerotic pathway of Escherichia coli. During simulation, cell growth was defined such as to reproduce the experimental cell growth profile of fed-batch cultivation in jar fermenters. However, to confirm the biological appropriateness of our model, sensitivity analysis and experimental validation were required. We constructed an L-glutamic acid fermentation simulation model by removing sucAB, a gene encoding α-ketoglutarate dehydrogenase. We then performed systematic sensitivity analysis for L-glutamic acid production; the results of this process corresponded with previous experimental data regarding L-glutamic acid fermentation. Furthermore, it allowed us to predicted the possibility that accumulation of 3-phosphoglycerate in the cell would regulate the carbon flux into the TCA cycle and lead to an increase in the yield of L-glutamic acid via fermentation. We validated this hypothesis through a fermentation experiment involving a model L-glutamic acid-production strain, E. coli MG1655 ΔsucA in which the phosphoglycerate kinase gene had been amplified to cause accumulation of 3-phosphoglycerate. The observed increase in L-glutamic acid production verified the biologically meaningful predictive power of our dynamic metabolic simulation model. In this study, dynamic simulation using a literature-based model was shown to be useful for elucidating the precise mechanisms involved in fermentation processes inside the cell. Further exhaustive sensitivity analysis will facilitate identification of novel factors involved in the metabolic regulation of amino acid fermentation.

  12. Multiple Objective Optimization and Optimal Control of Fermentation Processes

    Directory of Open Access Journals (Sweden)

    Mitko Petrov


    Full Text Available A multiple objective optimization is applied for finding an optimum policy of fed-batch processes of whey fermentation and L-lysine production. The multiple objective optimization problems are transformed to a standard problem of optimization with single objective function by a general utility function with weight coefficients for each single utility coefficient criteria. A combined algorithm is applied when solving the maximizing decision problem. The algorithm includes a method for random search of finding an initial point and a method based on the fuzzy sets theory, combined in order to find the best solution of the optimization problem. The application of the combined algorithm eliminates the main disadvantage of the used fuzzy optimization method, namely it decreases the number of discrete values of the control variables. Thus, the algorithm allows problems with larger scale to be solved. After this multiple optimization, the useful product quality rises and the residual substrate concentration at the end of the process decreases. In this way, the process productivity is increased.

  13. Optimization of fermentation medium for nisin production from ...

    African Journals Online (AJOL)


    production from Lactococcus lactis subsp. lactis using ... the optimal medium obtained by ANN-GA was located at the verge of the test region, a further Box- ...... Lactococcus lactis in fed-batch culture. Appl. Microbiol. Biot. 68: 322-. 326.

  14. 补料分批培养生产2-酮基-D-葡萄糖酸的研究%Studies on the Fermentative Production of 2-Keto-D-gluconic Acid by Pseudomonas fluorescens in Fed-batch Culture

    Institute of Scientific and Technical Information of China (English)

    孙文敬; 赵峰梅; 杨庆文; 郭金权; 秦丽; 刘敬泽


    研究了补糖对荧光假单胞菌(Pseudomonas fluorescens)AR4生产2-酮基-D-葡萄糖酸的影响,并在50kL发酵罐上进行了补料分批培养工业应用性试验.研究结果表明,采用补料分批培养方法,能有效提高发酵液中的产物浓度;开始补糖时发酵液中的残糖浓度以3%~6%为宜;补料分批培养方法及AR4菌株可用于2-酮基-D-葡萄糖酸的工业化规模生产中.

  15. 法夫酵母响应PH-stat的底物流加促进虾青素合成的研究%High cell density fermentation of Xanthophyllomyces dendrorhous and its overproducing of astaxanthin by means of ph-stat with glucose and ammonia fed-batch

    Institute of Scientific and Technical Information of China (English)

    梁新乐; 岑沛霖; 励建荣; 张虹; 张大中; 俞黎南


    本文主要研究了采用响应pH-stat的流加培养模式,进行Xanthophyllomyces dendrorhous高细胞密度培养及虾青素合成.在该补料间歇培养模式中,发酵过程pH的恒定控制采用2.78mol/L葡萄糖及3mol/L氨水来控制,同时起到流加葡萄糖的作用.试验结果表明,虾青素积累量于78h可达4.05mg/L,菌体干重在第84h获最大值14.62g/L.与不流加葡萄糖、氨水的间歇培养相比,虾青素含量和菌体干重分别增加2.89、5.22倍.

  16. Production of 1,3-propanediol by Klebsiella pneumoniae using raw ...

    African Journals Online (AJOL)



    Jan 16, 2012 ... In addition, the fed-batch culture using raw glycerol as the ... renewable carbon source for low-cost 1,3-PD production. ... medium. Fed-batch fermentation experiment ... Glucose was determined with immobilized glucose.

  17. Fermentation conditions that affect clavulanic acid production in Streptomyces clavuligerus: a systematic review

    Directory of Open Access Journals (Sweden)

    Hooi-Leng eSer


    Full Text Available The β-lactamase inhibitor, clavulanic acid is frequently used in combination with β-lactam antibiotics to treat a wide spectrum of infectious diseases. Clavulanic acid prevents drug resistance by pathogens against these β-lactam antibiotics by preventing the degradation of the β-lactam ring, thus ensuring eradication of these harmful microorganisms from the host. This systematic review provides an overview on the fermentation conditions that affect the production of clavulanic acid in the firstly described producer, Streptomyces clavuligerus. A thorough search was conducted using predefined terms in several electronic databases (PubMed, Medline, ScienceDirect, EBSCO, from database inception to June 30th 2015. Studies must involve wild-type Streptomyces clavuligerus, and full texts needed to be available. A total of 29 eligible articles were identified. Based on the literature, several factors were identified that could affect the production of clavulanic acid in S. clavuligerus. The addition of glycerol or other vegetable oils (e.g. olive oil, corn oil could potentially affect clavulanic acid production. Furthermore, some amino acids such as arginine and ornithine, could serve as potential precursors to increase clavulanic acid yield. The comparison of different fermentation systems revealed that fed-batch fermentation yields higher amounts of clavulanic acid as compared to batch fermentation, probably due to the maintenance of substrates and constant monitoring of certain entities (such as pH, oxygen availability, etc.. Overall, these findings provide vital knowledge and insight that could assist media optimization and fermentation design for clavulanic acid production in S. clavuligerus.

  18. Fermentation Conditions that Affect Clavulanic Acid Production in Streptomyces clavuligerus: A Systematic Review. (United States)

    Ser, Hooi-Leng; Law, Jodi Woan-Fei; Chaiyakunapruk, Nathorn; Jacob, Sabrina Anne; Palanisamy, Uma Devi; Chan, Kok-Gan; Goh, Bey-Hing; Lee, Learn-Han


    The β-lactamase inhibitor, clavulanic acid is frequently used in combination with β-lactam antibiotics to treat a wide spectrum of infectious diseases. Clavulanic acid prevents drug resistance by pathogens against these β-lactam antibiotics by preventing the degradation of the β-lactam ring, thus ensuring eradication of these harmful microorganisms from the host. This systematic review provides an overview on the fermentation conditions that affect the production of clavulanic acid in the firstly described producer, Streptomyces clavuligerus. A thorough search was conducted using predefined terms in several electronic databases (PubMed, Medline, ScienceDirect, EBSCO), from database inception to June 30th 2015. Studies must involve wild-type Streptomyces clavuligerus, and full texts needed to be available. A total of 29 eligible articles were identified. Based on the literature, several factors were identified that could affect the production of clavulanic acid in S. clavuligerus. The addition of glycerol or other vegetable oils (e.g., olive oil, corn oil) could potentially affect clavulanic acid production. Furthermore, some amino acids such as arginine and ornithine, could serve as potential precursors to increase clavulanic acid yield. The comparison of different fermentation systems revealed that fed-batch fermentation yields higher amounts of clavulanic acid as compared to batch fermentation, probably due to the maintenance of substrates and constant monitoring of certain entities (such as pH, oxygen availability, etc.). Overall, these findings provide vital knowledge and insight that could assist media optimization and fermentation design for clavulanic acid production in S. clavuligerus.

  19. Chlorine dioxide against bacteria and yeasts from the alcoholic fermentation (United States)

    Meneghin, Silvana Perissatto; Reis, Fabricia Cristina; de Almeida, Paulo Garcia; Ceccato-Antonini, Sandra Regina


    The ethanol production in Brazil is carried out by fed-batch or continuous process with cell recycle, in such way that bacterial contaminants are also recycled and may be troublesome due to the substrate competition. Addition of sulphuric acid when inoculum cells are washed can control the bacterial growth or alternatively biocides are used. This work aimed to verify the effect of chlorine dioxide, a well-known biocide for bacterial decontamination of water and equipments, against contaminant bacteria (Bacillus subtilis, Lactobacillus plantarum, Lactobacillus fermentum and Leuconostoc mesenteroides) from alcoholic fermentation, through the method of minimum inhibitory concentration (MIC), as well as its effect on the industrial yeast inoculum. Lower MIC was found for B. subtilis (10 ppm) and Leuconostoc mesenteroides (50 ppm) than for Lactobacillus fermentum (75 ppm) and Lactobacillus plantarum (125 ppm). Additionally, these concentrations of chlorine dioxide had similar effects on bacteria as 3 ppm of Kamoran® (recommended dosage for fermentation tanks), exception for B. subtilis, which could not be controlled at this Kamoran® dosage. The growth of industrial yeasts was affected when the concentration of chlorine dioxide was higher than 50 ppm, but the effect was slightly dependent on the type of yeast strain. Smooth yeast colonies (dispersed cells) seemed to be more sensitive than wrinkled yeast colonies (clustered cells/pseudohyphal growth), both isolated from an alcohol-producing unit during the 2006/2007 sugar cane harvest. The main advantage in the usage of chlorine dioxide that it can replace antibiotics, avoiding the selection of resistant populations of microorganisms. PMID:24031227

  20. Enhancement of Surfactin yield by improving the medium composition and fermentation process. (United States)

    Willenbacher, Judit; Yeremchuk, Wladimir; Mohr, Teresa; Syldatk, Christoph; Hausmann, Rudolf


    Surfactin is one of the most promising biosurfactants due to its extraordinary surface activity. Commonly, the well-established Cooper medium, a glucose-based mineral salt medium, is utilized for the microbial production of Surfactin. The current study investigated the enhancement of Surfactin yields by analyzing the effects of different glucose concentrations, next to the introduction of an alternative chelating agent and nitrogen source. The utilization of 8 g/L glucose, 0.008 mM Na3citrate and 50 mM (NH4)2SO4 increased Surfactin yields from 0.7 to 1.1 g/L during shake flask experiments applying Bacillus subtilis DSM10(T). Consequentially conducted shake flask experiments, employing five other Surfactin producer strains during cultivation in the former and enhanced version of the Cooper medium, suggest a general enhancement of Surfactin yields during application of the enhanced version of the Cooper medium. The enhancement of the medium composition is therefore most likely independent from the employed producer strain. The following utilization of the enhanced medium composition during fed-batch fermentation with integrated foam fractionation yielded 30 % more Surfactin in comparison to batch fermentations with integrated foam fractionation employing the former version of the Cooper medium.

  1. Online prediction for contamination of chlortetracycline fermentation based on Dezert-Smarandache theory

    Institute of Scientific and Technical Information of China (English)

    Jianwen Yang; Xiangguang Chen; Huaiping Jin


    Fermentative production of chlortetracycline is a complex fed-batch bioprocess. It generally takes over 90 h for cultivation and is often contaminated by undesired microorganisms. Once the fermentation system is contami-nated to certain extent, the product quality and yield wil be seriously affected, leading to a substantial economic loss. Using information fusion based on the Dezer–Smarandache theory, self-recursive wavelet neural network and unscented kalman filter, a novel method for online prediction of contamination is developed. Al state vari-ables of culture process involving easy-to-measure and difficult-to-measure variables commonly obtained with soft-sensors present their contamination symptoms. By extracting and fusing latent information from the chang-ing trend of each variable, integral and accurate prediction results for contamination can be achieved. This makes preventive and corrective measures be taken promptly. The field experimental results show that the method can be used to detect the contamination in time, reducing production loss and enhancing economic efficiency.

  2. Hydrophobic microspheres for in situ removal of 2-phenylethanol from yeast fermentation. (United States)

    Achmon, Yigal; Goldshtein, Jenny; Margel, Shlomo; Fishman, Ayelet


    The commercial production of the fragrance compound 2-phenylethanol (2-PE) from phenylalanine by yeast is limited by the accumulation of the toxic product, and therefore, in situ product removal techniques are required. We describe the use of hydrophobic polymethylmethacrylate (PMMA) microspheres of narrow size distribution of 1.53 ± 0.10 µm diameter for continuous removal of 2-PE from the fermentation medium by a mechanism of swelling. In shake flask experiments with conditions simulating 2-PE stress, a 10-fold increase in productivity was measured for systems containing >10% (w/v) microspheres. A 1 L fed-batch fermentation with 8% (w/v) of PMMA microspheres resulted in a total 2-PE concentration of 7.05 g/L, from which 5.40 g/L was incorporated inside the resin, implying 76% encapsulation. This ratio of 0.07 g/g of product per resin is among the highest reported to date. Scanning electron microscopy revealed a concomitant increase in sphere diameter confirming that swelling occurred.

  3. Membrane-mediated extractive fermentation for lactic acid production from cellulosic biomass

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Rongfu; Lee, Y.Y. [Auburn Univ., AL (United States)


    Lactic acid production from cellulosic biomass by cellulose and Lactobacillus delbrueckii was studied in a fermenter-extractor employing a microporous hollow fiber membrane (NIHF). This bioreactor system was operated under a fed-batch mode with continuous removal of lactic acid by an in situ extraction. A tertiary amine (Alamine 336) was used as an extractant for lactic acid. The extraction capacity of Alamine 336 is greatly enhanced by addition of alcohol. Long-chain alcohols serve well for this purpose since they are less toxic to micro-organism. Addition of kerosene, a diluent, was necessary to reduce the solvent viscosity. A solvent mixture of 20% Alamine 336,40% oleyl alcohol, and 40% kerosene was found to be most effective in the extraction of lactic acid. Progressive change of pH from an initial value of 5.0 down to 4.3 has significantly improved the overall performance of the simultaneous saccharification and extractive fermentation over that of constant pH operation. The change of pH was applied to promote cell growth in the early phase, and extraction in the latter phase. 20 refs., 10 figs., 1 tab.

  4. Cost-effective simultaneous saccharification and fermentation of l-lactic acid from bagasse sulfite pulp by Bacillus coagulans CC17. (United States)

    Zhou, Jie; Ouyang, Jia; Xu, Qianqian; Zheng, Zhaojuan


    The main barriers to cost-effective lactic acid production from lignocellulose are the high cost of enzymes and the ineffective utilization of the xylose within the hydrolysate. In the present study, the thermophilic Bacillus coagulans strain CC17 was used for the simultaneous saccharification and fermentation (SSF) of bagasse sulfite pulp (BSP) to produce l-lactic acid. Unexpectedly, SSF by CC17 required approximately 33.33% less fungal cellulase than did separate hydrolysis and fermentation (SHF). More interestingly, CC17 can co-ferment cellobiose and xylose without any exogenous β-glucosidase in SSF. Moreover, adding xylanase could increase the concentration of lactic acid produced via SSF. Up to 110g/L of l-lactic acid was obtained using fed-batch SSF, resulting in a lactic acid yield of 0.72g/g cellulose. These results suggest that SSF using CC17 has a remarkable advantage over SHF and that a potentially low-cost and highly-efficient fermentation process can be established using this protocol.

  5. Water-insoluble material from apple pomace makes changes in intracellular NAD⁺/NADH ratio and pyrophosphate content and stimulates fermentative production of hydrogen. (United States)

    Sato, Osamu; Suzuki, Yuma; Sato, Yuki; Sasaki, Shinsuke; Sonoki, Tomonori


    Apple pomace is one of the major agricultural residues in Aomori prefecture, Japan, and it would be useful to develop effective applications for it. As apple pomace contains easily fermentable sugars such as glucose, fructose and sucrose, it can be used as a feedstock for the fermentation of fuels and chemicals. We previously isolated a new hydrogen-producing bacterium, Clostridium beijerinckii HU-1, which could produce H2 at a production rate of 14.5 mmol of H2/L/h in a fed-batch culture at 37 °C, pH 6.0. In this work we found that the HU-1 strain produces H2 at an approximately 20% greater rate when the fermentation medium contains the water-insoluble material from apple pomace. The water-insoluble material from apple pomace caused a metabolic shift that stimulated H2 production. HU-1 showed a decrease of lactate production, which consumes NADH, accompanied by an increase of the intracellular pyrophosphate content, which is an inhibitor of lactate dehydrogenase. The intracellular NAD(+)/NADH ratios of HU-1 during H2 fermentation were maintained in a more reductive state than those observed without the addition of the water insoluble material. To correct the abnormal intracellular redox balance, caused by the repression of lactate production, H2 production with NADH oxidation must be stimulated.

  6. Comprehensive assessment of the L-lysine production process from fermentation of sugarcane molasses. (United States)

    Anaya-Reza, Omar; Lopez-Arenas, Teresa


    L-Lysine is an essential amino acid that can be produced by chemical processes from fossil raw materials, as well as by microbial fermentation, the latter being a more efficient and environmentally friendly procedure. In this work, the production process of L-lysine-HCl is studied using a systematic approach based on modeling and simulation, which supports decision making in the early stage of process design. The study considers two analysis stages: first, the dynamic analysis of the fermentation reactor, where the conversion of sugars from sugarcane molasses to L-lysine with a strain of Corynebacterium glutamicum is carried out. In this stage, the operation mode (either batch or fed batch) and operating conditions of the fermentation reactor are defined to reach the maximum technical criteria. Afterwards, the second analysis stage relates to the industrial production process of L-lysine-HCl, where the fermentation reactor, upstream processing, and downstream processing are included. In this stage, the influence of key parameters on the overall process performance is scrutinized through the evaluation of several technical, economic, and environmental criteria, to determine a profitable and sustainable design of the L-lysine production process. The main results show how the operating conditions, process design, and selection of evaluation criteria can influence in the conceptual design. The best plant design shows maximum product yield (0.31 g L-lysine/g glucose) and productivity (1.99 g/L/h), achieving 26.5% return on investment (ROI) with a payback period (PBP) of 3.8 years, decreasing water and energy consumption, and with a low potential environmental impact (PEI) index.

  7. Improving the production yield and productivity of 1,3-dihydroxyacetone from glycerol fermentation using Gluconobacter oxydans NL71 in a compressed oxygen supply-sealed and stirred tank reactor (COS-SSTR). (United States)

    Zhou, Xin; Zhou, Xuelian; Xu, Yong; Yu, Shiyuan


    In this study, a compressed oxygen gas supply was connected to a sealed aerated stirred tank reactor (COS-SSTR) bio-system, leading to a high-oxygen pressure bioreactor used to improve the bio-transformative performance in the production of 1,3-dihydroxyacetone (DHA) from glycerol using Gluconobacter oxydans NL71. A concentration of 301.2 ± 8.2 g L(-1) DHA was obtained from glycerol after 32 h of fed-batch fermentation in the COS-SSTR system. The volumetric productivity for this process was 9.41 ± 0.23 g L(-1) h(-1), which is presently the highest obtained level of glycerol bioconversion into DHA. These results show that the application of this bioreactor would enable microbial production of DHA from glycerol at the industrial scale.

  8. Mono-fermentation of glycerine - Fermentation of a substrate in a dominant amount. Final report; Monovergaerung von Glycerin - Vergaerung von einem Substrat in dominierender Menge. Schlussbericht

    Energy Technology Data Exchange (ETDEWEB)

    Erb, D.; Bueeler, E.; Spicher, M.


    The present study investigated the feasibility of a mono fermentation of the glycerine-fraction from biodiesel production. Part of the experiments took place in a single-stage, continuous system with 700 l usable volume. The maximum yield of biogas of the glycerine-fraction is 1100 l/l of glycerine, or 870 l/kg of glycerine, in continuous operation. The average methane content is 70 %. The adaptation rate of the biomass at the substrate of glycerine-fraction is high. Two or three days after starting the feed 100 % degradation rates will be achieved. The single-stage, continuous fermentation of the glycerine-fraction at 40 {sup o}C runs only stable at very low organic loading rate (0.65 kg oDM/(d m{sup 3})) and is therefore not currently economical. At higher organic loading rates (1.5 to 3.0 kg oDM/(d m{sup 3})) the adapted biomass collapsed after about 20 days due to massive instability of the process. A two-stage system with separate hydrolysis stage could probably allow a stable fermentation as search for literature has shown. Fed-batch experiments in the laboratory of the University of Waedenswil, Switzerland (ZHAW) demonstrated that the glycerine-fraction from biodiesel production is slightly better degradable than pure glycerine. The process dysfunctions arise because of the inhibition of intermediates resulting from the degradation of glycerine. At higher concentrations of 1.2-propanediol and 2.3-butanediol the degradation was incomplete. Further inhibitors can not be excluded. The failed stability of the process is not due to the lack of main nutrients or trace elements. (author)

  9. Growth kinetics and physiological behavior of co-cultures of Saccharomyces cerevisiae and Kluyveromyces lactis, fermenting carob sugars extracted with whey. (United States)

    Rodrigues, B; Lima-Costa, M E; Constantino, A; Raposo, S; Felizardo, C; Gonçalves, D; Fernandes, T; Dionísio, L; Peinado, J M


    Alcoholic fermentation of carob waste sugars (sucrose, glucose and fructose) extracted with cheese whey, by co-cultures of Saccharomyces cerevisiae and Kluyveromyces lactis has been analyzed. Growth and fermentation of S. cerevisiae in the carob-whey medium showed an inhibition of about 30% in comparison with water-extracted carob. The inhibition of K. lactis on carob-whey was greater (70%) when compared with the whey medium alone, due to osmolarity problems. Oxygen availability was a very important factor for K. lactis, influencing its fermentation performance. When K. lactis was grown alone on carob-whey medium, lactose was always consumed first, and glucose and fructose were consumed afterwards, only at high aeration conditions. In co-culture with S. cerevisiae, K. lactis was completely inhibited and, at low aeration, died after 3 days; at high aeration this culture could survive but growth and lactose fermentation were only recovered after S. cerevisiae became stationary. To overcome the osmolarity and K. lactis' oxygen problems, the medium had to be diluted and a sequential fermentative process was designed in a STR-3l reactor. K. lactis was inoculated first and, with low aeration (0.13vvm), consumed all the lactose in 48h. Then S. cerevisiae was inoculated, consuming the total of the carob sugars, and producing ethanol in a fed-batch regime. The established co-culture with K. lactis increased S. cerevisiae ethanol tolerance. This fermentation process produced ethanol with good efficiency (80g/l final concentration and a conversion factor of 0.4g ethanol/g sugar), eliminating all the sugars of the mixed waste. These efficient fermentative results pointed to a new joint treatment of agro-industrial wastes which may be implemented successfully, with economic and environmental sustainability for a bioethanol industrial proposal.

  10. Fermentation Industry. (United States)

    Grady, C. P. L., Jr.; Grady, J. K.


    Presents a literature review of wastes from the fermentation industry, covering publications of 1976-77. This review focuses on: (1) alcoholic beverage production; (2) pharmaceuticals and biochemicals production; and (3) biomass production. A list of 62 references is also presented. (HM)

  11. Fermentation Industry. (United States)

    Grady, C. P. L., Jr.; Grady, J. K.


    Presents a literature review of wastes from the fermentation industry, covering publications of 1976-77. This review focuses on: (1) alcoholic beverage production; (2) pharmaceuticals and biochemicals production; and (3) biomass production. A list of 62 references is also presented. (HM)

  12. Food Fermentation

    NARCIS (Netherlands)

    Nout, M.J.R.; Vos, de W.M.; Zwietering, M.H.


    The focus of this book is on state of the art technologies and scientific developments in academia and industry that contribute to the characterization and specification of fermentation starter microorganisms, to the present-day experimental approaches in product and process development and control,

  13. Efficient production of ethanol from waste paper and the biochemical methane potential of stillage eluted from ethanol fermentation. (United States)

    Nishimura, Hiroto; Tan, Li; Sun, Zhao-Yong; Tang, Yue-Qin; Kida, Kenji; Morimura, Shigeru


    Waste paper can serve as a feedstock for ethanol production due to being rich in cellulose and not requiring energy-intensive thermophysical pretreatment. In this study, an efficient process was developed to convert waste paper to ethanol. To accelerate enzymatic saccharification, pH of waste paper slurry was adjusted to 4.5-5.0 with H2SO4. Presaccharification and simultaneous saccharification and fermentation (PSSF) with enzyme loading of 40 FPU/g waste paper achieved an ethanol yield of 91.8% and productivity of 0.53g/(Lh) with an ethanol concentration of 32g/L. Fed-batch PSSF was used to decrease enzyme loading to 13 FPU/g waste paper by feeding two separate batches of waste paper slurry. Feeding with 20% w/w waste paper slurry increased ethanol concentration to 41.8g/L while ethanol yield decreased to 83.8%. To improve the ethanol yield, presaccharification was done prior to feeding and resulted in a higher ethanol concentration of 45.3g/L, a yield of 90.8%, and productivity of 0.54g/(Lh). Ethanol fermentation recovered 33.2% of the energy in waste paper as ethanol. The biochemical methane potential of the stillage eluted from ethanol fermentation was 270.5mL/g VTS and 73.0% of the energy in the stillage was recovered as methane. Integrating ethanol fermentation with methane fermentation, recovered a total of 80.4% of the energy in waste paper as ethanol and methane.

  14. 恒速补料与pH胁迫相结合提高透明质酸发酵产量的研究%Constant Speed Feeding and pH Stress Combination to Improve the Yield of Hyaluronic Acid Fermentation

    Institute of Scientific and Technical Information of China (English)

    邱平; 柯常毅; 邓益全


    通过对兽疫链球菌恒速补料培养与间歇性pH胁迫相组合提高透明质酸发酵产量的研究,发现通过恒速补料培养模式与间歇性的pH胁迫相结合的方式,能使菌体获得较高的菌体比生长速率与碳源利用率,同时转变细胞产能途径、提高细胞产能效率,有效提高HA 的发酵产量。该研究最终得到HA 的发酵产量为6.724g/L,比单一恒速补料培养产量提高了56.4%;比单一间歇性pH胁迫培养产量提高了25.8%,比对照提高了68.1%。%The Streptococcus zooepidemicus constant fed-batch culture and intermittent pH stress combination of hyaluronic acid fermentation yield, through constant batch culture mode and intermittent pH stress combination, can make the cell to obtain a higher specific cell growth rate and carbon source utilization rate, at the same time change the cell capacity approach, improve cell production efficiency, improve the yield of fermentation HA. The study finally obtained the fermentation yield of HA was 6.724g/L, than a single constant fed-batch production increased by 56.4%; pH stress than single intermittent training of production increased by 25.8%,increased by 68.1% than that of contrast.

  15. Optimization of Fermentation Conditions and Feed Batch Modes of Bt Using Brewery Waste Medium%利用啤酒废弃物培养Bt的条件与补料优化

    Institute of Scientific and Technical Information of China (English)



    Bt BRC-WLY| isolated from wastewater-digestive plants was cultivated in brewery-waste-medium fay submerged fermentation with the single factor study to optimize the fermentation conditions in the experiment and the affection of fermentation of Bt by different feeding modes and feeding composition of material, pretreatmented brewery yeast fluid (yeast fluid), KH2PO4 and feeding time on fermentation were also explored . The results showed that the optimal conditions comprised initial pH at 7.5 -8.0, 80 tnL of liquid volume in the 500 mL flask, fermentation temperature at 30 °C and 5% of inoculation volume. The optimal fed batch mode was fedding 10% (V/V) of yeast fluid after cultured for 8 hr, which resulted in spores yield, crystal weightiness, production intensity and yield of units sugar increased 8, 1.78, 0.98 and 3.07 times than those of fermentation without fed batched.%以污水源分离菌株Bt BRC-WLY1为供试菌,以啤酒废弃物为培养基,单因素实验优化发酵条件,并探讨补麦芽糖、预处理后的新鲜啤酒酵母液(下称酵母液)和KH2PO4的不同补料成分和补料时间对发酵水平的影响.结果显示,优化的最佳发酵条件是初始pH7.5~8.0、装液量80 mL、发酵温度30℃、接种量5%.最佳补料方式为发酵8h,加入10%的酵母液,与未补料对比,芽胞数、晶体干重、生产强度和单位糖产量分别提高了8、1.78、0.98、3.07倍.

  16. Enhancement of n-butanol production by in situ butanol removal using permeating-heating-gas stripping in acetone-butanol-ethanol fermentation. (United States)

    Chen, Yong; Ren, Hengfei; Liu, Dong; Zhao, Ting; Shi, Xinchi; Cheng, Hao; Zhao, Nan; Li, Zhenjian; Li, Bingbing; Niu, Huanqing; Zhuang, Wei; Xie, Jingjing; Chen, Xiaochun; Wu, Jinglan; Ying, Hanjie


    Butanol recovery from acetone-butanol-ethanol (ABE) fed-batch fermentation using permeating-heating-gas was determined in this study. Fermentation was performed with Clostridium acetobutylicum B3 in a fibrous bed bioreactor and permeating-heating-gas stripping was used to eliminate substrate and product inhibition, which normally restrict ABE production and sugar utilization to below 20 g/L and 60 g/L, respectively. In batch fermentation (without permeating-heating-gas stripping), C. acetobutylicum B3 utilized 60 g/L glucose and produced 19.9 g/L ABE and 12 g/L butanol, while in the integrated process 290 g/L glucose was utilized and 106.27 g/L ABE and 66.09 g/L butanol were produced. The intermittent gas stripping process generated a highly concentrated condensate containing approximately 15% (w/v) butanol, 4% (w/v) acetone, a small amount of ethanol (butanol solution [∼ 70% (w/v)] after phase separation. Butanol removal by permeating-heating-gas stripping has potential for commercial ABE production. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

  17. In situ butanol recovery from Clostridium acetobutylicum fermentations by expanded bed adsorption. (United States)

    Wiehn, Michael; Staggs, Kyle; Wang, Yuchen; Nielsen, David R


    Although butanol is a promising biofuel, its fermentative production suffers from inhibition caused by end product toxicity. The in situ removal of butanol from cultures via expanded bed adsorption offers an effective strategy for mitigating the effects of product toxicity while eliminating the need to clarify cultures via microfiltration. The hydrophobic polymer resin Dowex Optipore L-493 was found to be both an effective butanol adsorbent and suitable for use in expanded bed adsorption. Recirculation rates through the adsorption column were strongly correlated with and ultimately controlled rates of butanol uptake from the media which, reaching as high as 41.1 g/L h, easily exceed those of its production in a typical fermentation. Vacuum application with vapor collection was found to be an effective means of adsorbent regeneration, with an average of 81% butanol recovery possible, with butanol concentrations in the cold trap reaching as high as 85.8 g/L. Integration of expanded bed adsorption with a fed-batch Clostridium acetobutylicum ATCC 824 fermentation and its continuous operation for 38.5 h enabled the net production (i.e., in solution and adsorbed) of butanol and total solvent products at up to 27.2 and 40.7 g/L of culture, respectively, representing 2.2- and 2.3-fold improvements over conventional batch culture. While adsorbent biofouling was found to be minimal, further investigation of biofouling in longer-term studies will provide useful and further insight regarding the robustness of the process strategy. © 2013 American Institute of Chemical Engineers.

  18. Deployment Repeatability (United States)


    controlled to great precision, but in a Cubesat , there may be no attitude determination at all. Such a Cubesat might treat sun angle and tumbling rates as...could be sensitive to small differences in motor controller timing. In these cases, the analyst might choose to model the entire deployment path, with...knowledge of the material damage model or motor controller timing precision. On the other hand, if many repeated and environmentally representative

  19. A Multiple-objective Optimization of Whey Fermentation in Stirred Tank Bioreactors

    Directory of Open Access Journals (Sweden)

    Mitko Petrov


    Full Text Available A multiple-objective optimization is applied to find an optimal policy of a fed-batch fermentation process for lactose oxidation from a natural substratum of the strain Kluyveromyces marxianus var. lactis MC5. The optimal policy is consisted of feed flow rate, agitation speed, and gas flow rate. The multiple-objective problem includes: the total price of the biomass production, the second objective functions are the separation cost in downstream processing and the third objective function corresponds to the oxygen mass-transfer in the bioreactor. The multiple-objective optimization are transforming to standard problem for optimization with single-objective function. Local criteria are defined utility function with different weight for single-type vector task. A fuzzy sets method is applied to be solved the maximizing decision problem. A simple combined algorithm guideline to find a satisfactory solution to the general multiple-objective optimization problem. The obtained optimal control results have shown an increase of the process productiveness and a decrease of the residual substrate concentration.

  20. Purification of chondroitin precursor from Escherichia coli K4 fermentation broth using membrane processing. (United States)

    Schiraldi, Chiara; Carcarino, Immacolata Loredana; Alfano, Alberto; Restaino, Odile Francesca; Panariello, Andrea; De Rosa, Mario


    Recently the possibility of producing the capsular polysaccharide K4, a fructosylated chondroitin, in fed-batch experiments was assessed. In the present study, a novel downstream process to obtain chondroitin from Escherichia coli K4 fermentation broth was developed. The process is simple, scalable and economical. In particular, downstream procedures were optimized with a particular aim of purifying a product suitable for further chemical modifications, in an attempt to develop a biotechnological platform for chondroitin sulfate production. During process development, membrane devices (ultrafiltration/diafiltration) were exploited, selecting the right cassette cut-offs for different phases of purification. The operational conditions (cross-flow rate and transmembrane pressure) used for the process were determined on an ÄKTA cross-flow instrument (GE Healthcare, USA), a lab-scale automatic tangential flow filtration system. In addition, parameters such as selectivity and throughput were calculated based on the analytical quantification of K4 and defructosylated K4, as well as the major contaminants. The complete downstream procedure yielded about 75% chondroitin with a purity higher than 90%.

  1. Fermentation strategy for second generation ethanol production from sugarcane bagasse hydrolyzate by Spathaspora passalidarum and Scheffersomyces stipitis. (United States)

    Nakanishi, Simone C; Soares, Lauren B; Biazi, Luiz Eduardo; Nascimento, Viviane M; Costa, Aline C; Rocha, George Jackson M; Ienczak, Jaciane L


    Alcoholic fermentation of released sugars in pretreatment and enzymatic hydrolysis of biomass is a central feature for second generation ethanol (E2G) production. Saccharomyces cerevisiae used industrially in the production of first generation ethanol (E1G) convert sucrose, fructose, and glucose into ethanol. However, these yeasts have no ability to ferment pentose (xylose). Therefore, the present work has focused on E2G production by Scheffersomyces stipitis and Spathaspora passalidarum. The fermentation strategy with high pitch, cell recycle, fed-batch mode, and temperature decrease for each batch were performed in a hydrolyzate obtained from a pretreatment at 130°C with NaOH solution (1.5% w/v) added with 0.15% (w/w) of anthraquinone (AQ) and followed by enzymatic hydrolysis. The process strategy has increased volumetric productivity from 0.35 to 0.38 g · L(-1)  · h(-1) (first to third batch) for S. stipitis and from 0.38 to 0.81 g · L(-1)  · h(-1) for S. passalidarum (first to fourth batch). Mass balance for the process proposed in this work showed the production of 177.33 kg ethanol/ton of sugar cane bagasse for S. passalidarum compared to 124.13 kg ethanol/ton of sugar cane bagasse for S. stipitis fermentation. The strategy proposed in this work can be considered as a promising strategy in the production of second generation ethanol. Biotechnol. Bioeng. 2017;114: 2211-2221. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  2. An overview on fermentation, downstream processing and properties of microbial alkaline proteases. (United States)

    Gupta, R; Beg, Q K; Khan, S; Chauhan, B


    Microbial alkaline proteases dominate the worldwide enzyme market, accounting for a two-thirds share of the detergent industry. Although protease production is an inherent property of all organisms, only those microbes that produce a substantial amount of extracellular protease have been exploited commercially. Of these, strains of Bacillus sp. dominate the industrial sector. To develop an efficient enzyme-based process for the industry, prior knowledge of various fermentation parameters, purification strategies and properties of the biocatalyst is of utmost importance. Besides these, the method of measurement of proteolytic potential, the selection of the substrate and the assay protocol depends upon the ultimate industrial application. A large array of assay protocols are available in the literature; however, with the predominance of molecular approaches for the generation of better biocatalysts, the search for newer substrates and assay protocols that can be conducted at micro/nano-scale are becoming important. Fermentation of proteases is regulated by varying the C/N ratio and can be scaled-up using fed-batch, continuous or chemostat approaches by prolonging the stationary phase of the culture. The conventional purification strategy employed, involving e.g., concentration, chromatographic steps, or aqueous two-phase systems, depends on the properties of the protease in question. Alkaline proteases useful for detergent applications are mostly active in the pH range 8-12 and at temperatures between 50 and 70 degrees C, with a few exceptions of extreme pH optima up to pH 13 and activity at temperatures up to 80-90 degrees C. Alkaline proteases mostly have their isoelectric points near to their pH optimum in the range of 8-11. Several industrially important proteases have been subjected to crystallization to extensively study their molecular homology and three-dimensional structures.

  3. Evaluation of by-products from the biodiesel industry as fermentation feedstock for poly(3-hydroxybutyrate-co-3-hydroxyvalerate) production by Cupriavidus necator. (United States)

    García, I L; López, J A; Dorado, M P; Kopsahelis, N; Alexandri, M; Papanikolaou, S; Villar, M A; Koutinas, A A


    Utilization of by-products from oilseed-based biodiesel production (crude glycerol, rapeseed meal hydrolysates) for microbial polyhydroxyalkanoate (PHA) production could lead to the replacement of expensive carbon sources, nutrient supplements and precursors for co-polymer production. Batch fermentations in shake flasks with varying amounts of free amino nitrogen led to the production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (P(3HB-co-3HV)) with a 2.8-8% 3HV content. Fed-batch fermentations in shake flasks led to the production of 10.9g/L P(3HB-co-3HV) and a 55.6% P(3HB-co-3HV) content. NaCl concentrations between 2 and 6g/L gradually became inhibitory to bacterial growth and PHA formation, whereas in the case of K(2)SO(4), the inhibitory effect was observed only at concentrations higher than 20g/L. Differential scanning calorimetry (DSC), thermogravimetric analysis (TGA) and nuclear magnetic resonance ((13)C NMR) demonstrated that the incorporation of 3HV into the obtained P(3HB-co-3HV) lowered glass transition temperature, crystallinity and melting point as compared to polyhydroxybutyrate. Integrating PHA production in existing oilseed-based biodiesel plants could enhance the viability and sustainability of this first generation biorefinery. Copyright © 2012 Elsevier Ltd. All rights reserved.

  4. Metabolic process engineering of Clostridium tyrobutyricum Δack-adhE2 for enhanced n-butanol production from glucose: effects of methyl viologen on NADH availability, flux distribution, and fermentation kinetics. (United States)

    Du, Yinming; Jiang, Wenyan; Yu, Mingrui; Tang, I-Ching; Yang, Shang-Tian


    Butanol biosynthesis through aldehyde/alcohol dehydrogenase (adhE2) is usually limited by NADH availability, resulting in low butanol titer, yield, and productivity. To alleviate this limitation and improve n-butanol production by Clostridium tyrobutyricum Δack-adhE2 overexpressing adhE2, the NADH availability was increased by using methyl viologen (MV) as an artificial electron carrier to divert electrons from ferredoxin normally used for H2 production. In the batch fermentation with the addition of 500 μM MV, H2 , acetate, and butyrate production was reduced by more than 80-90%, while butanol production increased more than 40% to 14.5 g/L. Metabolic flux analysis revealed that butanol production increased in the fermentation with MV because of increased NADH availability as a result of reduced H2 production. Furthermore, continuous butanol production of ∼55 g/L with a high yield of ∼0.33 g/g glucose and extremely low ethanol, acetate, and butyrate production was obtained in fed-batch fermentation with gas stripping for in situ butanol recovery. This study demonstrated a stable and reliable process for high-yield and high-titer n-butanol production by metabolically engineered C. tyrobutyricum by applying MV as an electron carrier to increase butanol biosynthesis.

  5. 玉米螟高毒力生防菌株筛选与发酵工艺研究%Screening of Bacillus thuringiensis with High Virulence to Pyrausta nubilalis and Its Fermentation Processing

    Institute of Scientific and Technical Information of China (English)

    周荣华; 廖先清; 刘芳; 张志刚; 饶犇


    以玉米螟(Pyrausta nubilalis)为对象筛选出一株高毒力生防菌株苏云金芽孢杆菌(Bacillus thuringiensis)NBIC380。通过培养基优化,获得适宜的培养基配方D。补料发酵研究的结果表明,固定氮源浓度为80 g/L,发酵前期补料效果明显,补料速率为根据发酵不同阶段进行调整,可以取得更好的发酵结果,有利于提高产量,降低生产成本。%A biocontrol strain Bacillus thuringiensis NBIC380 with high virulence to Pyrausta nubilalis was obtained. Accord-ing to this strain, appropriate media formulations D was obtained by medium optimization. Fed-batch fermentation experiments showed fermentation pre-feeding effect was obvious by using 80 g/L nitrogen concentration. Feeding rate was adjusted accord-ing to the different stages of fermentation. This strategy could improve production and reduce production costs.

  6. Process Control for Production of Human-like Collagen in Fed-batch Culture of Escherichia coli BL 21%重组大肠杆菌高密度发酵生产类人胶原蛋白的过程控制研究

    Institute of Scientific and Technical Information of China (English)

    骆艳娥; 范代娣; 马晓轩; 王德伟; 米钰; 花秀夫; 李稳宏


    Recombinant E. Coli BL 21 was cultivated in high cell density to produce human-like collagen. The effects of the feeding of nitrogen source, controlled by an auto on/off-feeding mode with two different cycles of 0.5 min and 4 min intervals, oxygen-enrichment methods and inducement strength on the cell yield and human-like collagen production were investigated. The studies showed that nitrogen source feeding in fast cycle could result in higher human-like collagen production than that in slow cycle; and the feedback regulation of glucose, increase of the pressure of fermentation bioreactor, and supply of oxygen-enriched air could all increase cell yield and human-like collagen production. The effects of inducement strength on protein expression were found important. When OD600reached 90-100, the cultivation temperature was increased to 42℃ to begin induction for 2-3 h, and then shifted to 39C for 5-6 h induction, the cell density and human-like collagen production could reach 96 g·L- 1 [DCW (dry cell mass)] and 19.8% (g·g-1 DCW) respectively.

  7. Isolation of a novel high erythritol-producing Pseudozyma tsukubaensis and scale-up of erythritol fermentation to industrial level. (United States)

    Jeya, Marimuthu; Lee, Kyoung-Mi; Tiwari, Manish Kumar; Kim, Jung-Soo; Gunasekaran, Paramasamy; Kim, Sang-Yong; Kim, In-Won; Lee, Jung-Kul


    This study isolated a novel erythritol-producing yeast strain, which is capable of growth at high osmolarity. Characteristics of the strain include asexual reproduction by multilateral budding, absence of extracellular starch-like compounds, and a negative Diazonium blue B color reaction. Phylogenetic analysis based on the 26S rDNA sequence and physiological analysis indicated that the strain belongs to the species Pseudozyma tsukubaensis and has been named P. tsukubaensis KN75. When P. tsukubaensis KN75 was cultured aerobically in a fed-batch culture with glucose as a carbon source, it produced 245 g/L of erythritol, corresponding to 2.86 g/L/h productivity and 61% yield, the highest erythritol yield ever reported by an erythritol-producing microorganism. Erythritol production was scaled up from a laboratory scale (7 L fermenter) to pilot (300 L) and plant (50,000 L) scales using the dissolved oxygen as a scale-up parameter. Erythritol production at the pilot and plant scales was similar to that at the laboratory scale, indicating that the production of erythritol by P. tsukubaensis KN75 holds commercial potential.

  8. Paper sludge (PS) to bioethanol: Evaluation of virgin and recycle mill sludge for low enzyme, high-solids fermentation. (United States)

    Boshoff, Sonja; Gottumukkala, Lalitha Devi; van Rensburg, Eugéne; Görgens, Johann


    Paper sludge (PS) from the paper and pulp industry consists primarily of cellulose and ash and has significant potential for ethanol production. Thirty-seven PS samples from 11 South African paper and pulp mills exhibited large variation in chemical composition and resulting ethanol production. Simultaneous saccharification and fermentation (SSF) of PS in fed-batch culture was investigated at high solid loadings and low enzyme dosages. Water holding capacity and viscosity of the PS influenced ethanol production at elevated solid loadings of PS. High viscosity of PS from virgin pulp mills restricted the solid loading to 18% (w/w) at an enzyme dosage of 20 FPU/gram dry PS (gdPS), whereas an optimal solid loading of 27% (w/w) was achieved with corrugated recycle mill PS at 11 FPU/gdPS. Ethanol concentration and yield of virgin pulp and corrugated recycle PS were 34.2g/L at 66.9% and 45.5 g/L at 78.2%, respectively.

  9. Non-sterilized fermentative production of polymer-grade L-lactic acid by a newly isolated thermophilic strain Bacillus sp. 2-6.

    Directory of Open Access Journals (Sweden)

    Jiayang Qin

    Full Text Available BACKGROUND: The demand for lactic acid has been increasing considerably because of its use as a monomer for the synthesis of polylactic acid (PLA, which is a promising and environment-friendly alternative to plastics derived from petrochemicals. Optically pure L-lactic acid is essential for polymerization of PLA. The high fermentation cost of L-lactic acid is another limitation for PLA polymers to compete with conventional plastics. METHODOLOGY/PRINCIPAL FINDINGS: A Bacillus sp. strain 2-6 for production of L-lactic acid was isolated at 55 degrees C from soil samples. Its thermophilic characteristic made it a good lactic acid producer because optically pure L-lactic acid could be produced by this strain under open condition without sterilization. In 5-liter batch fermentation of Bacillus sp. 2-6, 118.0 g/liter of L-lactic acid with an optical purity of 99.4% was obtained from 121.3 g/liter of glucose. The yield was 97.3% and the average productivity was 4.37 g/liter/h. The maximum L-lactic acid concentration of 182.0 g/liter was obtained from 30-liter fed-batch fermentation with an average productivity of 3.03 g/liter/h and product optical purity of 99.4%. CONCLUSIONS/SIGNIFICANCE: With the newly isolated Bacillus sp. strain 2-6, high concentration of optically pure L-lactic acid could be produced efficiently in open fermentation without sterilization, which would lead to a new cost-effective method for polymer-grade L-lactic acid production from renewable resources.

  10. Probiotic fermented dairy products

    Directory of Open Access Journals (Sweden)

    Adnan Tamime


    Full Text Available Fermented dairy products are the most popular vehicle used in theindustry for the implantation of the probiotic microflora in humans. Therefore this paper provides an overview of new knowledge on probiotic fermented dairy products. It involves historical developments, commercial probiotic microorganisms and products, and their therapeutic properties, possibilities of quality improvement of different types of newly developed fermented dairy products together with fermented goat’s milk products.

  11. Fermented milk for hypertension

    DEFF Research Database (Denmark)

    Usinger, Lotte; Reimer, Christina; Ibsen, Hans


    Fermented milk has been suggested to have a blood pressure lowering effect through increased content of proteins and peptides produced during the bacterial fermentation. Hypertension is one of the major risk factors for cardiovascular disease world wide and new blood pressure reducing lifestyle...... interventions, such as fermented milk, would be of great importance....

  12. 表达重组咖啡豆α-半乳糖苷酶的酵母工程菌的高密度发酵%High Cell-Density Fermentation of Pichia pastoris Producing Recombinant Coffee Bean α-Galactosidase

    Institute of Scientific and Technical Information of China (English)

    高红伟; 贾延军; 鲍国强; 季守平; 檀英霞; 李素波; 章扬培; 宫锋


    用5 L发酵罐优化了重组咖啡豆α-半乳糖苷酶酵母工程菌pPIC9K-Gal/GS115(本室构建)的高密度发酵工艺.通过对发酵条件的优化,包括甘油补充量及补充时机、甲醇诱导量及诱导时机、溶氧控制、诱导时间等,重组咖啡豆α-半乳糖苷酶在毕赤酵母中得到了高效表达.利用所确定的最适条件进行发酵,菌体密度最终达到368 g/L以上,每批发酵液离心后可获得3.5 L的发酵上清,上清中的蛋白含量达到3 g/L以上,目的蛋白占上清总蛋白的50%以上,含量约为1.5 g/L,上清中α-半乳糖苷酶的活性维持在80 U/ml左右.确立工艺后又进行了3次发酵试验,证明了工艺的可行性和稳定性.为重组咖啡豆α-半乳糖苷酶在B→O血型改造和酶解大豆低聚糖方面的应用奠定了基础.%Based on the previous study of construction of Pichia pastoris engineering Strain ( pPIC9K-Gal/GSl15 ) expressing recombinant ( -galactosidase, this report described the optimal fermentation conditions of Pichia pastoris in a 5-liter-working-volume fer-mentor. 3 experiments were performed with various fermentation parameters, including the volume of inoculum seed, the volume and time of glycerol fed-batch,the volume and time of methanol fed-batch, dissolved oxygen, pH, agitation, temperature of glycerol batch phase and methanol fed-batch phase. For each experiment,0.4 liter pPIC9K-Gal/GS115 cells were inoculated into 4.5 liter basal salts medium. Then 3.5 liters of supernatant were harvested at the end of the fermentation. The results showed that the protein concentration in fermentation supernatant was 3 -4. 5 mg/ml,the activity of α-galactosidase was 80 U/ml,and the enzyme activity ratio was 24 - 30 U/ mg. Based on the above experiments, 3 fermentations were carried out and confirmed the feasibility and stability of the technology. Therefore, this fermentation research laid a foundation for recombinant α-galactosidase purification to obtain

  13. Optimization of Fermentation Conditions for the Production of Bacteriocin Fermentate (United States)


    FERMENTATION CONDITIONS FOR THE PRODUCTION OF BACTERIOCIN “ FERMENTATE ” by Anthony Sikes Wayne Muller and Claire Lee March 2015...From - To) October 2010 – November 2013 4. TITLE AND SUBTITLE OPTIMIZATION OF FERMENTATION CONDITIONS FOR THE PRODUCTION OF BACTERIOCIN “ FERMENTATE ...nisin and pediocin. Whey + yeast extract was the best performing whey fermentation media. The nisin producer strain Lactococcus. lactis ssp. lactis was

  14. Enhanced ethanol fermentation by engineered Saccharomyces cerevisiae strains with high spermidine contents. (United States)

    Kim, Sun-Ki; Jo, Jung-Hyun; Jin, Yong-Su; Seo, Jin-Ho


    Construction of robust and efficient yeast strains is a prerequisite for commercializing a biofuel production process. We have demonstrated that high intracellular spermidine (SPD) contents in Saccharomyces cerevisiae can lead to improved tolerance against various fermentation inhibitors, including furan derivatives and acetic acid. In this study, we examined the potential applicability of the S. cerevisiae strains with high SPD contents under two cases of ethanol fermentation: glucose fermentation in repeated-batch fermentations and xylose fermentation in the presence of fermentation inhibitors. During the sixteen times of repeated-batch fermentations using glucose as a sole carbon source, the S. cerevisiae strains with high SPD contents maintained higher cell viability and ethanol productivities than a control strain with lower SPD contents. Specifically, at the sixteenth fermentation, the ethanol productivity of a S. cerevisiae strain with twofold higher SPD content was 31% higher than that of the control strain. When the SPD content was elevated in an engineered S. cerevisiae capable of fermenting xylose, the resulting S. cerevisiae strain exhibited much 40-50% higher ethanol productivities than the control strain during the fermentations of synthetic hydrolysate containing high concentrations of fermentation inhibitors. These results suggest that the strain engineering strategy to increase SPD content is broadly applicable for engineering yeast strains for robust and efficient production of ethanol.

  15. Coproduction of acetic acid and electricity by application of microbial fuel cell technology to vinegar fermentation. (United States)

    Tanino, Takanori; Nara, Youhei; Tsujiguchi, Takuya; Ohshima, Takayuki


    The coproduction of a useful material and electricity via a novel application of microbial fuel cell (MFC) technology to oxidative fermentation was investigated. We focused on vinegar production, i.e., acetic acid fermentation, as an initial and model useful material that can be produced by oxidative fermentation in combination with MFC technology. The coproduction of acetic acid and electricity by applying MFC technology was successfully demonstrated by the simultaneous progress of acetic acid fermentation and electricity generation through a series of repeated batch fermentations. Although the production rate of acetic acid was very small, it increased with the number of repeated batch fermentations that were conducted. We obtained nearly identical (73.1%) or larger (89.9%) acetic acid yields than that typically achieved by aerated fermentation (75.8%). The open-cycle voltages measured before and after fermentation increased with the total fermentation time and reached a maximum value of 0.521 V prior to the third batch fermentation. The maximum current and power densities measured in this study (19.1 μA/cm² and 2.47 μW/cm², respectively) were obtained after the second batch fermentation.

  16. Optimization of Fermentation Conditions for Nisin Production by Lactococcus lactis N302%Nisin生产菌株Lactococcus lactis N302的发酵优化

    Institute of Scientific and Technical Information of China (English)

    李瑞青; 轩辕铮铮; 姜德洲; 苏俊杰; 徐海津; 张秀明; 乔明强


    对一株Nisin生产菌株Lactococcus lactis N302现有培养基进行了氮源替代,并采用Plackett-Burman(PB)法和中心复合设计(Central Composite Design)对影响其发酵生产Nisin的6个培养条件进行筛选优化.PB实验表明,蔗糖、初始pH值和酵母粉是影响Nisin效价的三个关键因素.对三因素进行中心复合设计,经响应面法优化分析(RSM)确定了L.Lactis N302发酵生产Nisin的最优条件为:蔗糖13.7g.L-1,初始pH值7.74,酵母粉25.7g.L-1,大豆蛋白胨10.0g.L-1,K2HPO410.0g.L-1,接种量3%.优化后Nisin效价较优化前提高了7.2%.小试(10 L)研究表明,分批发酵18h、补碱分批发酵16h菌株L.lactis N302单位Nisin效价最高,分别为4 597.03 IU.mL-1和8 773.34 IU.mL-1.%Nisin is a bacteriocin widely used in food industry as an effective food preservative. High nisin production was aimed by optimizing the fermentation conditions of Lactococcus lactis N302. First, soybean peptone was used the main nitrogen source of the culture medium instead of peptone. Then, the Plackett-Burman design (PB) and the path of steepest ascent method were applied to investigate the main factors that affect the yield of nisin, and to find the optimum region of the response. The results indicated that sucrose, initial pH value and yeast extract were the significant factors for nisin production. Central composite experimental design and response surface methodology (RSM) were further adopted to derive a statistical model for optimizing the fermentation conditions. The optimum fermentation conditions were found to be sucrose 13. 7 g · L-1, initial pH value 7. 74, yeast extract 25. 7 g · L-1, soybean peptone 10 g · L-1, K2HPO410 g · L-1, inoculum size 3%. The nisin yield increased by 7. 2% compared to the no-optimized conditions. Finally, 10 liter batch and pH fed-batch fermentation with the optimized conditions were carried out. The maximum nisin yield was achieved at 18 h for batch fermentation and 16 h for fed-batch

  17. Fermentative production of L-pipecolic acid from glucose and alternative carbon sources. (United States)

    Pérez-García, Fernando; Max Risse, Joe; Friehs, Karl; Wendisch, Volker F


    Corynebacterium glutamicum is used for the million-ton scale production of amino acids and has recently been engineered for production of the cyclic non-proteinogenic amino acid L-pipecolic acid (L-PA). In this synthetic pathway L-lysine was converted to L-PA by oxidative deamination, dehydration and reduction by L-lysine 6-dehydrogenase (deaminating) from Silicibacter pomeroyi and pyrroline 5-carboxylate reductase from C. glutamicum. However, production of L-PA occurred as by-product of L-lysine production only. Here, the author show that abolishing L-lysine export by the respective gene deletion resulted in production of L-PA as major product without concomitant lysine production while the specific growth rate was reduced due to accumulation of high intracellular lysine concentrations. Increasing expression of the genes encoding L-lysine 6-dehydrogenase and pyrroline 5-carboxylate reductase in C. glutamicum strain PIPE4 increased the L-PA titer to 3.9 g L(-1) , and allowed faster growth and, thus, a higher volumetric productivity of 0.08 ± 0.00 g L(-1) h(-1) respectively. Secondly, expression of heterologous genes for utilization of glycerol, xylose, glucosamine, and starch in strain PIPE4 enabled L-PA production from these alternative carbon sources. Third, in a glucose/sucrose-based fed-batch fermentation with C. glutamicum PIPE4 L-PA was produced to a titer of 14.4 g L(-1) with a volumetric productivity of 0.21 g L(-1) h(-1) and an overall yield of 0.20 g g(-1) . Copyright © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Pathway construction and metabolic engineering for fermentative production of ectoine in Escherichia coli. (United States)

    Ning, Yike; Wu, Xuejiao; Zhang, Chenglin; Xu, Qingyang; Chen, Ning; Xie, Xixian


    Ectoine is a protective agent and stabilizer whose synthesis pathway exclusively exists in select moderate halophiles. A novel established process called "bacterial milking" efficiently synthesized ectoine in moderate halophiles, however, this method places high demands on equipment and is cost prohibitive. In this study, we constructed an ectoine producing strain by introducing the ectoine synthesis pathway into Escherichia coli and improved its production capacity. Firstly, the ectABC gene cluster from Halomonas elongata was introduced into E. coli W3110 and the resultant strain synthesized 4.9g/L ectoine without high osmolarity. Subsequently, thrA encoding the bifunctional aspartokinase/homoserine dehydrogenase was deleted to weaken the competitive l-threonine branch, resulting in an increase of ectoine titer by 109%. Furthermore, a feedback resistant lysC from Corynebacterium glutamicum encoding the aspartate kinase was introduced to complement the enzymatic activity deficiency caused by thrA deletion and a 9% increase of ectoine titer was obtained. Finally, the promoter of ppc that encodes phosphoenolpyruvate carboxylase was replaced by a trc promoter, and iclR, a glyoxylate shunt transcriptional repressor gene, was deleted. The oxaloacetate pool, was thus reinforced and ectoine titer increased by 21%. The final engineered strain ECT05 (pTrcECT, pSTVLysC-CG) produced 25.1g/L ectoine by fed-batch fermentation in low salt concentration with glucose as a carbon source. The specific ectoine production and productivity was 0.8g/g DCW and 0.84gL(-)(1)h(-)(1) respectively. The overall ectoine yield was 0.11g/g of glucose.

  19. Repeat-until-success quantum repeaters (United States)

    Bruschi, David Edward; Barlow, Thomas M.; Razavi, Mohsen; Beige, Almut


    We propose a repeat-until-success protocol to improve the performance of probabilistic quantum repeaters. Conventionally, these rely on passive static linear-optics elements and photodetectors to perform Bell-state measurements (BSMs) with a maximum success rate of 50%. This is a strong impediment for entanglement swapping between distant quantum memories. Every time a BSM fails, entanglement needs to be redistributed between the corresponding memories in the repeater link. The key ingredients of our scheme are repeatable BSMs. Under ideal conditions, these turn probabilistic quantum repeaters into deterministic ones. Under realistic conditions, our protocol too might fail. However, using additional threshold detectors now allows us to improve the entanglement generation rate by almost orders of magnitude, at a nominal distance of 1000 km, compared to schemes that rely on conventional BSMs. This improvement is sufficient to make the performance of our scheme comparable to the expected performance of some deterministic quantum repeaters.

  20. 黄浆水液态深层发酵制醋工艺的研究%Vinegar production from yellow slurry water by submerged fermentation

    Institute of Scientific and Technical Information of China (English)

    张伟; 洪厚胜; 张庆文; 段许佳


    以白酒生产中间产物黄浆水为原料,结合液态深层发酵酿醋工艺,成功开发出风味独特的食醋.试验使用15L自吸式发酵罐补料分批培养,实验结果表明:在分割量35%时,周期24h以内,酸度高达7.6g/100mL,醋酸产率90%以上;成品醋不挥发酸比例25%.处理后各项指标均达到优级老陈醋标准,实现了黄浆水的综合利用,具有良好的实用价值和现实意义.%Using yellow slurry water (a by-product of Chinese liquor) as material, the vinegar with unique flavor was developed successfully by liquid submerged fermentation. In this study, the vinegar was fermented in a 15L self-aspirated acetator by semi-continuous fed-batch fermentation. The results showed that when the withdrawal rate was 35% and the withdrawal period was lower than 24h, the acidity reached 7.6g/100ml, the vinegar yield was over 90%, and the non-volatile acid content of finished vinegar was 25%. All indexes were up to the standards of top-grade aged vinegar after treatment. The utilization of yellow slurry water was realized, which was practical and meaningful.

  1. Cell-free supernatants obtained from fermentation of cheese whey hydrolyzates and phenylpyruvic acid by Lactobacillus plantarum as a source of antimicrobial compounds, bacteriocins, and natural aromas. (United States)

    Rodríguez-Pazo, Noelia; Vázquez-Araújo, Laura; Pérez-Rodríguez, Noelia; Cortés-Diéguez, Sandra; Domínguez, José Manuel


    Cheese whey hydrolyzates supplemented with phenylpyruvic acid (PPA) and commercial nutrients can be efficiently metabolized by Lactobacillus plantarum CECT-221 to biosynthesize some compounds with attractive applications in the food market. The main metabolites of cell-free extracts were antimicrobial compounds such as phenyllactic acid (PLA) and lactic acid (LA). The production of PLA by L. plantarum CECT-221 was evaluated in the Man-Rogosa-Sharpe broth supplemented with two biosynthetic precursors: phenylalanine or PPA. Using 30.5 mM PPA, the microorganism increased sevenfold the concentration of PLA producing 16.4 mM PLA in 46 h. A concentration of 40 mM PPA was a threshold to avoid substrate inhibition. The biosynthesis of whey hydrolyzates as a carbon source was enhanced by fed-batch fermentation of PPA; the average productivity of PLA increased up to 45.4 ± 3.02 mM after 120 h with a product yield of 0.244 mM mM(-1); meanwhile, LA reached 26.1 ± 1.3 g L(-1) with a product yield of 0.72 g g(-1). Cell-free fed-batch extracts charged in wells showed bacteriocin activity with halos of 7.49 ± 1.44 mm in plates inoculated with Carnobacterium piscicola and antimicrobial activity against Staphylococcus aureus (11.54 ± 1.14 mm), Pseudomonas aeruginosa (10.17 ± 2.46 mm), Listeria monocytogenes (7.75 ± 1.31 mm), and Salmonella enterica (3.60 ± 1.52 mm). Additionally, the analysis of the volatile composition of the headspace of this cell-free extract revealed that L. plantarum is a potential producer for natural aromas, such as acetophenone, with high price in the market. This is the first report of PLA production from cheese whey and PPA. The extracts showed bacteriocin activity and potential to be applied as an antimicrobial in the elaboration of safer foods.

  2. Food Technologies: Fermentation

    NARCIS (Netherlands)

    Nout, M.J.R.


    Fermentation refers to the use of microorganisms to achieve desirable food properties in the fermented food or beverage. Although the word ‘fermentation’ indicates ‘anaerobic metabolism,’ it is also used in a broader sense to indicate all anaerobic and aerobic microbiological and biochemical

  3. Asian fungal fermented food

    NARCIS (Netherlands)

    Nout, M.J.R.; Aidoo, K.E.


    In Asian countries, there is a long history of fermentation of foods and beverages. Diverse micro-organisms, including bacteria, yeasts and moulds, are used as starters, and a wide range of ingredients can be made into fermented foods. The main raw materials include cereals, leguminous seeds,

  4. Asian fungal fermented food

    NARCIS (Netherlands)

    Nout, M.J.R.; Aidoo, K.E.


    In Asian countries, there is a long history of fermentation of foods and beverages. Diverse micro-organisms, including bacteria, yeasts and moulds, are used as starters, and a wide range of ingredients can be made into fermented foods. The main raw materials include cereals, leguminous seeds, vegeta

  5. Enzymes in Fermented Fish. (United States)

    Giyatmi; Irianto, H E

    Fermented fish products are very popular particularly in Southeast Asian countries. These products have unique characteristics, especially in terms of aroma, flavor, and texture developing during fermentation process. Proteolytic enzymes have a main role in hydrolyzing protein into simpler compounds. Fermentation process of fish relies both on naturally occurring enzymes (in the muscle or the intestinal tract) as well as bacteria. Fermented fish products processed using the whole fish show a different characteristic compared to those prepared from headed and gutted fish. Endogenous enzymes like trypsin, chymotrypsin, elastase, and aminopeptidase are the most involved in the fermentation process. Muscle tissue enzymes like cathepsins, peptidases, transaminases, amidases, amino acid decarboxylases, glutamic dehydrogenases, and related enzymes may also play a role in fish fermentation. Due to the decreased bacterial number during fermentation, contribution of microbial enzymes to proteolysis may be expected prior to salting of fish. Commercial enzymes are supplemented during processing for specific purposes, such as quality improvement and process acceleration. In the case of fish sauce, efforts to accelerate fermentation process and to improve product quality have been studied by addition of enzymes such as papain, bromelain, trypsin, pepsin, and chymotrypsin. © 2017 Elsevier Inc. All rights reserved.

  6. Asian fungal fermented food

    NARCIS (Netherlands)

    Nout, M.J.R.; Aidoo, K.E.


    In Asian countries, there is a long history of fermentation of foods and beverages. Diverse micro-organisms, including bacteria, yeasts and moulds, are used as starters, and a wide range of ingredients can be made into fermented foods. The main raw materials include cereals, leguminous seeds, vegeta

  7. Food Technologies: Fermentation

    NARCIS (Netherlands)

    Nout, M.J.R.


    Fermentation refers to the use of microorganisms to achieve desirable food properties in the fermented food or beverage. Although the word ‘fermentation’ indicates ‘anaerobic metabolism,’ it is also used in a broader sense to indicate all anaerobic and aerobic microbiological and biochemical modific

  8. Pure Culture Fermentation of Brined Cucumbers. (United States)

    Etchells, J L; Costilow, R N; Anderson, T E; Bell, T A


    , pure culture fermentor which was suitable for gamma radiation, resistant to salt and acid, and which permitted repeated aseptic sampling of the fermenting brine, is illustrated and the specifications are given.

  9. Fermented milk for hypertension. (United States)

    Usinger, Lotte; Reimer, Christina; Ibsen, Hans


    Fermented milk has been suggested to have a blood pressure lowering effect through increased content of proteins and peptides produced during the bacterial fermentation. Hypertension is one of the major risk factors for cardiovascular disease world wide and new blood pressure reducing lifestyle interventions, such as fermented milk, would be of great importance. To investigate whether fermented milk or similar products produced by lactobacilli fermentation of milk proteins has any blood pressure lowering effect in humans when compared to no treatment or placebo. The Cochrane Central Register of Controlled Trials (CENTRAL), English language databases, including MEDLINE (1966-2011), EMBASE (1974-2011), Cochrane Complementary Medicine Trials Register, Allied and Complementary Medicine (AMED) (1985-2011), Food science and technology abstracts (1969-2011). Randomised controlled trials; cross over and parallel studies evaluating the effect on blood pressure of fermented milk in humans with an intervention period of 4 weeks or longer. Data was extracted individually by two authors, afterwards agreement had to be obtained before imputation in the review. A modest overall effect of fermented milk on SBP was found (MD -2.45; 95% CI -4.30 to -0.60), no effect was evident on DBP (MD -0.67; 95% CI -1.48, 0.14). The review does not support an effect of fermented milk on blood pressure. Despite the positive effect on SBP the authors conclude, for several reasons, that fermented milk has no effect on blood pressure. The effect found was very modest and only on SBP, the included studies were very heterogeneous and several with weak methodology. Finally, sensitivity and subgroup analyses could not reproduce the antihypertensive effect. The results do not give notion to the use of fermented milk as treatment for hypertension or as a lifestyle intervention for pre-hypertension nor would it influence population blood pressure.

  10. Fed-batch cultivation of the docosahexaenoic-acid-producing marine alga Crypthecodinium cohnii on ethanol

    NARCIS (Netherlands)

    Swaaf, de M.E.; Pronk, J.T.; Sijtsma, L.


    The heterotrophic marine microalga Crypthecodinium cohnii produces docosahexaenoic acid (DHA), a polyunsaturated fatty acid with food and pharmaceutical applications. So far, DHA production has been studied with glucose and acetic acid as carbon sources. This study investigates the potential of etha

  11. Calorimetric control of the specific growth rate during fed-batch cultures of Saccharomyces cerevisiae. (United States)

    Biener, Richard; Steinkämper, Anne; Horn, Thomas


    The specific growth rate of a Saccharomyces cerevisiae strain with glucose as limiting C-source was estimated from the measured heat flow produced by the cells. For the cultivation a standard 30 l laboratory bioreactor was used, which was extended in such a way that heat balancing is possible. The feed rate was adjusted by a feedforward/feedback controller such that the specific growth rate was kept on the desired set-point value. On the basis of experimental investigations it was demonstrated that the specific growth rate can be controlled at a given set point value below the critical value to prevent the production of growth-inhibitory ethanol due to the Crabtree effect. With this control strategy high biomass concentrations of more than 110 g l(-1) can be obtained. Copyright © 2012 Elsevier B.V. All rights reserved.

  12. An evolutionary strategy for fed-batch bioreactor optimization : concepts and performance

    NARCIS (Netherlands)

    Roubos, J.A.; Straten, van G.; Boxtel, van A.J.B.


    An evolutionary program, based on a real-code genetic algorithm (GA), is applied to calculate optimal control policies for bioreactors. The GA is used as a nonlinear optimizer in combination with simulation software and constraint handling procedures. A new class of GA-operators is introduced to obt

  13. Modeling and optimization of hairy root growth in fed-batch process. (United States)

    Mairet, Francis; Villon, Pierre; Boitel-Conti, Michèle; Shakourzadeh, Khalil


    This article proposes a feeding strategy based on a kinetic model to enhance hairy roots growth. A new approach for modeling hairy root growth is used, considering that there is no nutrient limitation thanks to an appropriate feeding, and the intracellular pools are supposed to be always saturated. Thus, the model describes the specific growth rate from extracellular concentration of the major nutrients and nutrient uptakes depend on biomass growth. An optimized feeding strategy was determined thanks to the model to maintain the major nutrient levels at their optimum assuming optimal initial concentrations. The optimal feed rate is computed in open loop using kinetic model prediction or in closed loop using conductivity measurements to estimate biomass growth. Datura innoxia was chosen as the model culture system. Shake flask cultures were used to calibrate the model. Finally, cultures in bioreactor were performed to validate the model and the control laws. Copyright 2010 American Institute of Chemical Engineers

  14. Analysis and control of proteolysis of a fusion protein in Pichia pastoris fed-batch processes. (United States)

    Jahic, Mehmedalija; Gustavsson, Malin; Jansen, Ann-Katrin; Martinelle, Mats; Enfors, Sven-Olof


    A fusion protein composed of a cellulose-binding module (CBM) from Neocallimastix patriciarum cellulase 6A and lipase B from Candida antarctica (CALB), was produced by Pichia pastoris Mut(+) in high-cell density bioreactor cultures. The production was induced by switching from growth on glycerol to growth on methanol. The lipase activity in the culture supernatant increased at an almost constant rate up to a value corresponding to 1.3 g x l(-1) of CBM-CALB. However, only about 40% of the product was of full-length according to Western blot analysis. This loss was due to a cleavage of the protein in the linker between the CBM and the CALB moieties. The cleavage was catalyzed by serine proteases in the culture supernatant. The CALB-moiety was subjected to further slow degradation by cell-associated proteolysis. Different strategies were used to reduce the proteolysis. Previous efforts to shorten the linker region resulted in a stable protein but with ten times reduced product concentration in bioreactor cultures (Gustavsson et al. 2001, Protein Eng. 14, 711-715). Addition of rich medium for protease substrate competition had no effect on the proteolysis of CBM-CALB. The kinetics for the proteolytic reactions, with and without presence of cells were shown to be influenced by pH. The fastest reaction, cleavage in the linker, was substantially reduced at pH values below 5.0. Decreasing the pH from 5.0 to 4.0 in bioreactor cultures resulted in an increase of the fraction of full-length product from 40 to 90%. Further improvement was achieved by decreasing the temperature from 30 to 22 degrees C during the methanol feed phase. By combining the optimal pH and the low temperature almost all product (1.5 g x l(-1)) was obtained as full-length protein with a considerably higher purity in the culture supernatant compared with the original cultivation.

  15. Cloning, multicopy expression and fed-batch production of Rhodotorula araucariae epoxide hydrolase in yarrowia lipolytica

    CSIR Research Space (South Africa)

    Ramduth, D


    Full Text Available cloned and functionally expressed in Y. lipolytica, under the control of a growth inducible hp4d promoter. The transformation experiments yielded only two positive multicopy transformants, which were assessed in flask cultures. The selected transformant...

  16. Degradation of chlorophenol mixtures in a fed-batch system by two ...

    African Journals Online (AJOL)


    Dec 16, 2010 ... sources is an approach that may result in significant cost reduc- tions in the ... tions to improve the industrial application of CP degradation (Mun et al., 2008) ... water is fed to the biological treatment unit either intermittently or ... et al., 2008) and several wastewaters contain PCP, present in mixtures together ...

  17. A framework for the systematic design of fed-batch strategies in mammalian cell culture. (United States)

    Kyriakopoulos, Sarantos; Kontoravdi, Cleo


    A methodology to calculate the required amount of amino acids (a.a.) and glucose in feeds for animal cell culture from monitoring their levels in batch experiments is presented herein. Experiments with the designed feeds on an antibody-producing Chinese hamster ovary cell line resulted in a 3-fold increase in titer compared to batch culture. Adding 40% more nutrients to the same feed further increases the yield to 3.5 higher than in batch culture. Our results show that above a certain threshold there is no linear correlation between nutrient addition and the integral of viable cell concentration. In addition, although high ammonia levels hinder cell growth, they do not appear to affect specific antibody productivity, while we hypothesize that high extracellular lactate concentration is the cause for the metabolic shift towards lactate consumption for the cell line used. Overall, the performance of the designed feeds is comparable to that of a commercial feed that was tested in parallel. Expanding this approach to more nutrients, as well as changing the ratio of certain amino acids as informed by flux balance analysis, could achieve even higher yields. © 2014 Wiley Periodicals, Inc.

  18. Sulfide oxidation at halo-alkaline conditions in a fed-batch bioreactor

    NARCIS (Netherlands)

    Bosch, van den P.L.F.; Beusekom, van O.C.; Buisman, C.J.N.; Janssen, A.J.H.


    A biotechnological process is described to remove hydrogen sulfide (H2S) from high-pressure natural gas and sour gases produced in the petrochemical industry. The process operates at halo-alkaline conditions and combines an aerobic sulfide-oxidizing reactor with an anaerobic sulfate (SO) and

  19. Sulfide oxidation at halo-alkaline conditions in a fed-batch bioreactor. (United States)

    van den Bosch, Pim L F; van Beusekom, Otto C; Buisman, Cees J N; Janssen, Albert J H


    A biotechnological process is described to remove hydrogen sulfide (H(2)S) from high-pressure natural gas and sour gases produced in the petrochemical industry. The process operates at halo-alkaline conditions and combines an aerobic sulfide-oxidizing reactor with an anaerobic sulfate (SO(4) (2-)) and thiosulfate (S(2)O(3) (2-)) reducing reactor. The feasibility of biological H(2)S oxidation at pH around 10 and total sodium concentration of 2 mol L(-1) was studied in gas-lift bioreactors, using halo-alkaliphilic sulfur-oxidizing bacteria (HA-SOB). Reactor operation at different oxygen to sulfide (O(2):H(2)S) supply ratios resulted in a stable low redox potential that was directly related with the polysulfide (S(x) (2-)) and total sulfide concentration in the bioreactor. Selectivity for SO(4) (2-) formation decreased with increasing S(x) (2-) and total sulfide concentrations. At total sulfide concentrations above 0.25 mmol L(-1), selectivity for SO(4) (2-) formation approached zero and the end products of H(2)S oxidation were elemental sulfur (S(0)) and S(2)O(3) (2-). Maximum selectivity for S(0) formation (83.3+/-0.7%) during stable reactor operation was obtained at a molar O(2):H(2)S supply ratio of 0.65. Under these conditions, intermediary S(x) (2-) plays a major role in the process. Instead of dissolved sulfide (HS(-)), S(x) (2-) seemed to be the most important electron donor for HA-SOB under S(0) producing conditions. In addition, abiotic oxidation of S(x) (2-) was the main cause of undesirable formation of S(2)O(3) (2-). The observed biomass growth yield under SO(4) (2-) producing conditions was 0.86 g N mol(-1) H(2)S. When selectivity for SO(4) (2-) formation was below 5%, almost no biomass growth was observed. (c) 2007 Wiley Periodicals, Inc.

  20. Enzyme feeding strategies for better fed-batch enzymatic hydrolysis of empty fruit bunch. (United States)

    Sugiharto, Yohanes Eko Chandra; Harimawan, Ardiyan; Kresnowati, Made Tri Ari Penia; Purwadi, Ronny; Mariyana, Rina; Andry; Fitriana, Hana Nur; Hosen, Hauna Fathmadinda


    Lignin inhibitory becomes a major obstacle for enzymatic hydrolysis of empty fruit bunch conducted in high solid loading. Since current technology required high enzyme loading, surfactant application could not effectively used since it is only efficient in low enzyme loading. In addition, it will increase final operation cost. Hence, another method namely "proportional enzyme feeding" was investigated in this paper. In this method, enzyme was added to reactor proportionally to substrate addition, different from conventional method ("whole enzyme feeding") where whole enzyme was added prior to hydrolysis process started. Proportional enzyme feeding could increase enzymatic digestibility and glucose concentration up to 26% and 12% respectively, compared to whole enzyme feeding for hydrolysis duration more than 40h. If enzymatic hydrolysis was run less than 40h (25% solid loading), whole enzyme feeding is preferable.