Continuous Production Of Butanol Research Articles

High-rate continuous n-butanol production by Clostridium acetobutylicum from glucose and butyric acid in a single-pass fibrous-bed bioreactor.

Biobutanol produced in acetone-butanol-ethanol (ABE) fermentation at batch mode cannot compete with chemically derived butanol because of the low reactor productivity. Continuous fermentation can dramatically enhance productivity and lower capital and operating costs, but are rarely used in industrial fermentation because of increased risks of culture degeneration, cell washout, and contamination. In this study, cells of the asporogenous Clostridium acetobutylicum ATCC55025 were immobilized in a single-passfibrous-bed bioreactor (FBB) for continuous production of butanol from glucose and butyrate at various dilution rates. Butyric acid in the feed medium helped maintaining cells in the solventogenic phase for stable continuous butanol production. At a dilution rate of 1.88 h-1 , butanol was produced at 9.55 g/L, with a yield of 0.24 g/g and productivity of 16.8 g/L/h, which was the highest productivity ever achieved for biobutanol fermentation and an 80-fold improvement over the conventional ABE fermentation. The extremely high productivity was attributed to the high density of viable cells (~100 g/L at >70% viability) immobilized in the fibrous matrix, which also enabled the cells to better tolerate butanol and butyric acid. The FBB was stable for continuous operation for an extended period of over 1 month.

Preparation of polymer of intrinsic microporosity composite membranes and their applications for butanol recovery

ABSTRACTWe fabricated novel composite membranes composed of a polymer of intrinsic microporosity (PIM‐1) and carbon black (CB) nanoparticles functionalized with the silane coupling agent aminopropyl triethoxysilane to recover butanol from aqueous solutions by pervaporation (PV). Scanning electron microscopy showed that the composite membranes were dense and defect free and had good adhesion with substrates. Compared with the those of pristine PIM‐1 membranes, the water contact angles of the composite membranes increased from around 86° to more than 90°; this confirmed the improvement of the hydrophobicity. The swelling degree of the 6 wt % CB‐filled PIM‐1 membranes dropped 23%; this indicated an increase in the swelling resistance. Furthermore, the PV results show experimentally that the incorporation of the functionalized CBs into the PIM‐1 matrix considerably improved both the permeability and selectivity to butanol. At a 4 wt % CB content, the optimum separation performance, with a separation factor of 19.7 and a permeation flux of 1116 g m−2 h−1, was achieved in an aqueous solution containing 5 wt % butanol at 30°C. It was noteworthy that the as‐fabricated membranes exhibited a good separation stability. This is a step forward in terms of continuous butanol production with hybrid membranes in fermentation processes. © 2018 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2019, 136, 46912.

Continuous butanol production by Clostridium acetobutylicum in a series of packed bed reactors

Continuous butanol production by Clostridium acetobutylicum in a series of packed bed reactors

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.

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.

Wood pulp as an immobilization matrix for the continuous production of isopropanol and butanol

The study was focused on developing a continuous method to produce an alcohol mixture suitable to be used as a gasoline supplement. The immobilized column reactor with wood pulp fibers was successfully used for the continuous production of butanol and isopropanol using Clostridium beijerinckii DSM 6423. A sugar mixture (glucose, mannose, galactose, arabinose and xylose) representing lignocellulose hydrolysate was used as a substrate for the production of solvents. The effect of dilution rate on solvent production was studied during continuous operation. The maximum total solvent concentration of 11.99g/l was obtained at a dilution rate of 0.16h(-1). The maximum solvent productivity (5.58g/lh) was obtained at a dilution rate of 1.5h(-1). The maximum solvent yield of 0.45g/g from sugar mixture was observed at 0.25h(-1). The system will be further used for the solvent production using wood hydrolysate as a substrate.

Engineering E. coli strain for conversion of short chain fatty acids to bioalcohols

BackgroundRecent progress in production of various biofuel precursors and molecules, such as fatty acids, alcohols and alka(e)nes, is a significant step forward for replacing the fossil fuels with renewable fuels. A two-step process, where fatty acids from sugars are produced in the first step and then converted to corresponding biofuel molecules in the second step, seems more viable and attractive at this stage. We have engineered an Escherichia coli strain to take care of the second step for converting short chain fatty acids into corresponding alcohols by using butyrate kinase (Buk), phosphotransbutyrylase (Ptb) and aldehyde/alcohol dehydrogenase (AdhE2) from Clostridium acetobutylicum.ResultsThe engineered E. coli was able to convert butyric acid and other short chain fatty acids of chain length C3 to C7 into corresponding alcohols and the efficiency of conversion varied with different E. coli strain type. Glycerol proved to be a better donor of ATP and electron as compared to glucose for converting butyric acid to butanol. The engineered E. coli was able to tolerate up to 100 mM butyric acid and produced butanol with the conversion rate close to 100% under anaerobic condition. Deletion of native genes, such as fumarate reductase (frdA) and alcohol dehydrogenase (adhE), responsible for side products succinate and ethanol, which act as electron sink and could compete with butyric acid uptake, did not improve the butanol production efficiency. Indigenous acyl-CoA synthetase (fadD) was found to play no role in the conversion of butyric acid to butanol. Engineered E. coli was cultivated in a bioreactor under controlled condition where 60 mM butanol was produced within 24 h of cultivation. A continuous bioreactor with the provision of cell recycling allowed the continuous production of butanol at the average productivity of 7.6 mmol/l/h until 240 h.ConclusionsE. coli engineered with the pathway from C. acetobutylicum could efficiently convert butyric acid to butanol. Other short chain fatty acids with the chain length of C3 to C7 were also converted to the corresponding alcohols. The ability of engineered strain to convert butyric acid to butanol continuously demonstrates commercial significance of the system.

Continuous butanol fermentation from xylose with high cell density by cell recycling system

A continuous butanol production system with high-density Clostridium saccharoperbutylacetonicum N1-4 generated by cell recycling was established to examine the characteristics of butanol fermentation from xylose. In continuous culture without cell recycling, cell washout was avoided by maintaining pH>5.6 at a dilution rate of 0.26h−1, indicating pH control was critical to this experiment. Subsequently, continuous culture with cell recycling increased cell concentration to 17.4gL−1, which increased butanol productivity to 1.20gL−1h−1 at a dilution rate of 0.26h−1 from 0.529gL−1h−1 without cell recycling. The effect of dilution rates on butanol production was also investigated in continuous culture with cell recycling. Maximum butanol productivity (3.32gL−1h−1) was observed at a dilution rate of 0.78h−1, approximately 6-fold higher than observed in continuous culture without cell recycling (0.529gL−1h−1).

Processes and Bioreactor Designs for Butanol Production from Lignocellulosic Biomass

Butanol is a chemical, solvent and a good liquid fuel, which can be produced from lignocellulosic biomass. This paper provides a review on the processes involved in butanol production from lignocellulosic biomass. Processes reviewed include: metabolic pathways of most commonly used microorganisms, lignocellulosic biomass as a potential feedstock for butanol production and bioreactor designs for batch and continuous butanol production. Butanol production from lignocellulosic biomass via different bioreactor designs is compared in terms of butanol concentration, butanol yield and butanol productivity. Factors affecting the fermentative butanol production are summarized and discussed: microbial inhibitors, acetate addition, butyrate addition, hydrogen production, culture stability, anaerobic conditions, agitation, pH, and temperature.

Continuous butanol production with reduced byproducts formation from glycerol by a hyper producing mutant of Clostridium pasteurianum

Butanol, a four-carbon primary alcohol (C(4)H(10)O), is an important industrial chemical and has a good potential to be used as a superior biofuel. Bio-based production of butanol from renewable feedstock is a promising and sustainable alternative to substitute petroleum-based fuels. Here, we report the development of a process for butanol production from glycerol, which is abundantly available as a byproduct of biodiesel production. First, a hyper butanol producing strain of Clostridium pasteurianum was isolated by chemical mutagenesis. The best mutant strain, C. pasteurianum MBEL_GLY2, was able to produce 10.8 g l(-1) butanol from 80 g l(-1) glycerol as compared to 7.6 g l(-1) butanol produced by the parent strain. Next, the process parameters were optimized to maximize butanol production from glycerol. Under the optimized batch condition, the butanol concentration, yield, and productivity of 17.8 g l(-1), 0.30 g g(-1), and 0.43 g l(-1) h(-1) could be achieved. Finally, continuous fermentation of C. pasteurianum MBEL_GLY2 with cell recycling was carried out using glycerol as a major carbon source at several different dilution rates. The continuous fermentation was run for 710 h without strain degeneration. The acetone-butanol-ethanol productivity and the butanol productivity of 8.3 and 7.8 g l(-1) h(-1), respectively, could be achieved at the dilution rate of 0.9 h(-1). This study reports continuous production of butanol with reduced byproducts formation from glycerol using C. pasteurianum, and thus could help design a bioprocess for the improved production of butanol.

Development of high-speed and highly efficient butanol production systems from butyric acid with high density of living cells of Clostridium saccharoperbutylacetonicum

Living cells are alive and have the butanol-producing ability but not much proliferation under nitrogen source-limited condition. We investigated various butanol production systems with high density of living cells of Clostridium saccharoperbutylacetonicum N1-4 supplemented with methyl viologen (MV) as an electron carrier and nutrient dosing for activity regeneration. In continuous butanol production with high density of living cells, butanol yield was drastically increased from 0.365 C-mol/C-mol with growing cells to 0.528 C-mol/C-mol at a dilution rate of 0.85 h⁻¹, being increased with the butanol to total solvent ratio. This yield was increased to 0.591 C-mol/C-mol by adding 0.01 mM MV. MV addition increased not only butanol yield but also butanol concentration and productivity as compared to those without MV addition. However, living cells lost their activity with incubation time, which lowered the operational stability of the system. Therefore, to maintain constant stability, activity regeneration was carried out with high density of living cells and MV. This system produced butanol at high concentration (9.40 g l⁻¹) and productivity (7.99 g l⁻¹ h⁻¹) for approximately 100 h with maintenance of considerably high yield of butanol (0.686 C-mol/C-mol). Thus, we established a high-speed and highly efficient butanol production system.

Continuous Butanol Production Using Suspended and Immobilized Clostridium beijerinckii NCIMB 8052 with Supplementary Butyrate

Butanol production by Clostridium beijerinckii NCIMB 8052 was investigated using both batch and continuous cultures containing suspended or immobilized cells. In the batch reactor, the initial addition of acetate and butyrate into the culture media was found not only to enhance solvent production but also to affect the ratio of acetone/butanol, which might result from the metabolic changes in solvent production. Furthermore, the addition of butyrate to the medium prevented strain degeneration during an extended subculturing, significantly induced butanol production (11.2 g/L butanol versus 0.45 g/L butanol with and without 36 mM butyrate, respectively), and shifted the acetone/butanol ratio to 1:3, which resulted in a higher yield (0.45 g of butanol/g of glucose) when compared to other studies. The beneficial effects of butyrate were also observed in continuous reactor tests containing suspended cells as the solvent production was maintained over more than 300 h of continuous operation. During a continuous butanol production with immobilized cells, using porous hydrophilic media and a dilution rate of 0.04 h−1, the overall butanol productivity and yield were 0.40 g L−1 h−1 and 0.44 g of butanol/g of glucose, respectively, which are approximately twice the values seen in a continuous reactor with suspended cells. Moreover, the butanol production was maintained over 150 days without apparent degeneration, even in the presence of high butanol concentrations (10−13 g/L). These results validate the effectiveness of producing butanol with an immobilized cell system supplemented with butyrate.

Continuous production of butanol by Clostridium acetobutylicum immobilized in a fibrous bed bioreactor.

We explored the influence of dilution rate and pH in continuous cultures of Clostridium acetobutylicum. A 200-mL fibrous bed bioreactor was used to produce high cell density and butyrate concentrations at pH 5.4 and 35degreesC. By feeding glucose and butyrate as a cosubstrate, the fermentation was maintained in the solventogenesis phase, and the optimal butanol productivity of 4.6 g/(L h) and a yield of 0.42 g/g were obtained at a dilution rate of 0.9 h-1 and pH 4.3. Compared to the conventional acetone-butanol-ethanol fermentation, the new fermentation process greatly improved butanol yield, making butanol production from corn an attractive alternative to ethanol fermentation.

Continuous production of butanol from starch-based packing peanuts.

Acetone, butanol, ethanol (ABE, or solvents) were produced from starch-based packing peanuts in batch and continuous reactors. In a batch reactor, 18.9 g/L of total ABE was produced from 80 g/L packing peanuts in 110 h of fermentation. The initial and final starch concentrations were 69.6 and 11.1 g/L, respectively. In this fermentation, ABE yield and productivity of 0.32 and 0.17 g/(L h) were obtained, respectively. Compared to the batch fermentation, continuous fermentation of 40 g/L of starchbased packing peanuts in P2 medium resulted in a maximum solvent production of 8.4 g/L at a dilution rate of 0.033 h-1. This resulted in a productivity of 0.27 g/(L h). However, the reactor was not stable and fermentation deteriorated with time. Continuous fermentation of 35 g/L of starch solution resulted in a similar performance. These studies were performed in a vertical column reactor using Clostridium beijerinckii BA101 and P2 medium. It is anticipated that prolonged exposure of culture to acrylamide, which is formed during boiling/autoclaving of starch, affects the fermentation negatively.

Bacterial conversion of molasses to acetone and butanol

In cooperation with the company Copersucar (Brazil), several variants of a fermentation system for the continuous production of butanol and acetone from high-test or invert molasses were developed. These fermentation systems involve a relatively economic batch fermentation requiring little investment, using a continuous culture as the inoculation culture, as well as a modern two-stage continuous culture with cell recycling. For example, 13.3 g·1−1 of solvent (acetone and butanol) are produced with a productivity of 3.3 g·1−1 ·h−1 by two-stage continuous molasses fermentation with cell recycling in the second stage. High-test molasses is converted completely into the products. Butanol and acetone production from molasses is economic in Brazil and the construction of a production plant is planned.

Continuous production of butanol from a glucose/xylose mixture with an immobilized cell system coupled to pervaporation.

Glucose and xylose can be converted continuously and simultaneously to butanol using immobilized cells. Xylose is only converted when glucose is completely consumed and the biocatalyst is not fully inhibited by butanol. With pervaporation for in-situ butanol recovery the complete conversion of a feed containing 60 kg/m3 of mixed sugars is achieved.

A simple model for the continuous production of butanol by immobilized clostridia: I: Clostridium beyerinckii on glucose

A simple model has been developed to describe the continuous production of butanol and isopropanol from glucose by Clostridium beyerinckii cells immobilized in calcium alginate beads. The model contains linear butanol inhibition kinetics and assumes the biomass to be homogeneously distributed throughout the alginate beads. For design purposes this model, containing five parameters, satisfactorily predicts the values of three output variables (the concentrations of glucose, butanol and isopropanol) as a function of three input variables (the glucose feed concentration, the dilution rate and the fraction of beads in the reactor). For process design a high bead fraction and well mixed conditions seem advantageous.

A simple model for the continuous production of butanol by immobilized Clostridia: II: Clostridium species DSM 2152 on glucose and whey permeate

The kinetic model developed for the continuous production of butanol by immobilized Clostridia (Chem. Eng. J., 32 (1986) B43) was tested for Clostridium species DSM 2152, immobilized in calcium alginate beads. The model described the butanol production rates adequately both on a model substrate, glucose, and on a technical substrate, whey permeate. For both substrates the r max values were found to be the same (about 55 kg substrate m −3 alginate h −1), but the maximal butanol concentration C B,max was much lower on whey permeate media than on glucose media (4.7 and 7.4 kg m −3). The operational stability of the immobilized Clostridium species was good; experiments lasted for 1000 – 1800 h without loss of activity and without disruption of the calcium alginate beads.

Continuous butanol production from whey permeate with immobilized Clostridium beyerinckii LMD 27.6

The aim of this study was to find the conditions necessary for the continuous butanol production from whey permeate with Clostridium beyerinckii LMD 27.6, immobilized in calcium alginate beads. The influence of three parameters on the butanol production was investigated: the fermentation temperature, the dilution rate (during start-up and at steady state) and the concentration of calcium ions in the fermentation broth. It was found that both a fermentation temperature of 30° C and a dilution rate of 0.1 h-1 or less during the start-up phase are required to achieve continuous butanol production from whey permeate. Butanol can be produced continuously from whey permeate in reactor productivities sixteen times higher than those found in batch cultures with free C. beyerinckii cells on whey media.

Continuous production of butanol with immobilized cells ofClostridium acetobutylicum

Spores ofClostridium acetobutylicum were immobilized in calcium alginate. An active gel preparation was obtained after outgrowth of the spores to vegetative cells within the gel matrix. A 100 mL column containing the immobilized cells was used for continuous production. At steady-state conditions the productivity of butanol was 67 g/L reactor volume/day.