Butanol Yield Research Articles

Effect of Endogenous and Exogenous Butyric Acid on Butanol Production From CO by Enriched Clostridia.

Butanol is a potential renewable fuel. To increase the selectivity for butanol during CO fermentation, exogenous acetic acid and ethanol, exogenous butyric acid or endogenous butyric acid from glucose fermentation have been investigated using CO as reducing power, with a highly enriched Clostridium sludge. Addition of 3.2 g/L exogenous butyric acid led to the highest 1.9 g/L butanol concentration with a conversion efficiency of 67%. With exogenous acetate and ethanol supply, the butanol concentration reached 1.6 g/L at the end of the incubation. However, the presence of acetic acid and ethanol favoured butanol production to 2.6 g/L from exogenous butyric acid by the enriched sludge. Finally, exogenous 14 g/L butyric acid yielded the highest butanol production of 3.4 g/L, which was also among the highest butanol concentration from CO/syngas fermentation reported so far. CO addition triggered butanol production from endogenous butyric acid (produced from glucose, Glucose + N2) with as high as 58.6% conversion efficiency and 62.1% butanol yield. However, no efficient butanol production was found from glucose and CO co-fermentation (Glucose + CO), although a similar amount of endogenous butyric acid was produced compared to Glucose + N2. The Clostridium genus occupied a relative abundance as high as 82% from the initial inoculum, while the Clostridia and Bacilli classes were both enriched and dominated in Glucose + N2 and Glucose + CO incubations. This study shows that the supply of butyric acid is a possible strategy for enhancing butanol production by CO fed anaerobic sludge, either via exogenous butyric acid, or via endogenous production by sugar fermentation.

Enhanced butanol production in Clostridium acetobutylicum by manipulating metabolic pathway genes

Biobutanol is naturally produced by solventogenic clostridia during acetone-butanol-ethanol (ABE) fermentation, which is hampered by low butanol titer, yield and productivity. Here, Clostridium acetobutylicum ATCC 55025 and its robust mutant HKKO with histidine kinase genes cac3319/cac0323 knockout were metabolically engineered to eliminate acetone formation and improve butanol production. By knocking out acetoacetate decarboxylase gene adc in ATCC 55025 and HKKO, the acetone titer decreased to 0.1 g/L and the butanol/ABE ratio increased by 39.0 % and 40.9 %, respectively. However, the butanol production was limited to ∼11 g/L. To improve butanol production, an alternative strategy was used by overexpressing acetaldehyde/alcohol dehydrogenase gene adhE2 in HKKO, and the mutant produced butanol of 19.7 g/L than that of 18.2 g/L from HKKO with high butanol productivity of 0.41 g/L/h and yield of 0.26 g/g. This study provides novel insights into the roles of adc and adhE2 on butanol biosynthesis and generates a hyper-butanol-producing strain.

Reduction wastewater discharge in second-generation acetone-butanol-ethanol (ABE) fermentation process by adsorptive removal of organic acids toward the broth recycling system

The treatment of high chemical oxygen demand (COD) organic wastewater is one of the major technical bottlenecks for the industrialization of second-generation biobutanol production. An effective strategy to decrease the wastewater discharge in biobutanol fermentation process is broth recycling (BR). Nevertheless, the conventional BR process suffered from server organic-acid by-products inhibition, causing low butanol yield. In this study, to solve the barrier, active carbon (AC) adsorption system was hybrid with the acetone-butanol-ethanol (ABE) fermentation and ex-situ product separation (ESPR) units. After fermentation and pervaporation, organic acids that remaining in the ABE-separated broth were partially adsorbed onto AC so that improve the continuity and efficiency of the BR process. The physicochemical characterizations of different types of commercial ACs and the organic acids adsorption efficiency was evaluated by analyzing the static and dynamic adsorptions kinetics. When adopting the DD-type AC (code C299105), the better adsorbent for organic acids removal, 87.50% of fermentative wastewater can be saved whilst the ABE concentrations were also kept constantly at 15.22 ± 0.45 g L−1 in the following cycles of fermentation, and the ABE productivity was almost no changed (0.21 ± 0.01 g L−1 h−1) after 3 cycles of operation. Therefore, the AC assisted ESPR process is an attractive method for ABE production with low wastewater discharge.

High butanol/acetone ratio featured ABE production using mixture of glucose and waste Pichia pastoris medium-based butyrate fermentation supernatant.

In this study, butanol (ABE) fermentations were implemented in a 7 L anaerobic fermentor, by directly using the mixture of glucose solution with the corn/waste Pichia pastoris medium-based butyrate fermentation supernatants (BFS II) as the co-substrate, followed by consecutively feeding of the BFS and concentrated glucose solution. When compared with the major index of ABE fermentation using 150g/L corn-based medium, butanol concentration could be maintained at high level of 12.7-12.8g/L, butanol/acetone (B/A) largely increased from ~ 2.0 to 4.4-5.0, butanol yield on total carbon sources increased from 0.32-0.34 to 0.39-0.41 (mol base) with a higher butyrate/glucose consumption ratio of 37%-53%. Efficient utilization of butyrate, SO42-, amino acids, oligosaccharides, etc. in BFS II and the intracellular NADH contributed to the ABE fermentation performance improvement. The proposed strategy could be considered as the second utilization of waste Pichia pastoris, which could save raw materials/operating costs, fully use the oligosaccharides/SO42- in BFS II to relieve the working loads in downstream waste water treatment process, and increase fermentation products diversity/flexibility to deal with the varied marketing prices and requirements.

Optimal Conditions for Butanol Production from Ethanol over MgAlO Catalyst Derived from Mg-Al Layer Double Hydroxides.

The MgAlO catalyst was obtained from thermal decomposition of the MgAl-LDH catalyst having Mg/Al molar ratio of 5. The catalytic Guerbet reaction of ethanol was investigated to determine the effect of WHSV and nitrogen flow rate on butanol production and product distribution. It was performed in a fixed-bed microreactor under continuous flow of vaporized ethanol mixed with N2. The MgAlO catalyst had high total basic sites and high total acid sites that were crucial for ethanol Guerbet reaction. The MgAlO catalyst showed the highest butanol selectivity at 300℃ under WHSV = 3.10 h-1 and nitrogen flow rate = 3,600 mL/h, and the highest butanol yield at 400℃ under WHSV = 3.10 h-1 and nitrogen flow rate = 900 mL/h. It can be summarized that in order to enhance the butanol yield, the low WHSV is preferred to increase the contact time of ethanol and catalyst under moderate temperature.

Optimization of a cathodic electro-fermentation process for enhancing co-production of butanol and hydrogen via acetone-butanol-ethanol fermentation of Clostridium beijerinckii

To develop an effective acetone-butanol-ethanol (ABE) fermentation process for the enhanced co-production of butanol and H2, a series of neutral red mediated cathodic electro-fermentation (CEF) processes of Clostridium beijerinckii NCIMB 8052 were conducted with/without exogenous butyric acid as the co-substrate of glucose. The cathode potential, NR dosage, glucose concentration and exogenous butyric acid concentration were optimized progressively. The results showed that the most favorable cathode potential, NR dosage, glucose concentration and exogenous butyric acid was -0.60 V (vs Ag/AgCl), 0.50 mmol/L, 30 g/L and 3.00 g/L, respectively. With the optimal condition, the metabolism in the CEF process of C. beijerinckii NCIMB 8052 was well manipulated, resulting the excellent production of butanol and H2 as high as 10.85 ± 0.45 g/L and 4.33 ± 0.40 L/L-medium, respectively. The excellent butanol yield of 0.40 ± 0.00 g/g-glucose (0.60 ± 0.01 mol-C/mol-C) and H2 yield of 161.69 ± 19.28 mL/g-glucose resulted an energy recovery (ER) and energy recovery efficiency (ERE) as high as 446.90 ± 21.39 kJ/L-medium and 95.85 ± 1.62%, respectively. The optimized CEF process demonstrates an effective approach to manipulating the microbial metabolism of Clostridia for the enhanced co-production of butanol and H2.

Renewable Butanol Production via Catalytic Routes.

Fluctuating crude oil price and global environmental problems such as global warming and climate change lead to growing demand for the production of renewable chemicals as petrochemical substitutes. Butanol is a nonpolar alcohol that is used in a large variety of consumer products and as an important industrial intermediate. Thus, the production of butanol from renewable resources (e.g., biomass and organic waste) has gained a great deal of attention from researchers. Although typical renewable butanol is produced via a fermentative route (i.e., acetone-butanol-ethanol (ABE) fermentation of biomass-derived sugars), the fermentative butanol production has disadvantages such as a low yield of butanol and the formation of byproducts, such as acetone and ethanol. To avoid the drawbacks, the production of renewable butanol via non-fermentative catalytic routes has been recently proposed. This review is aimed at providing an overview on three different emerging and promising catalytic routes from biomass/organic waste-derived chemicals to butanol. The first route involves the conversion of ethanol into butanol over metal and oxide catalysts. Volatile fatty acid can be a raw chemical for the production of butanol using porous materials and metal catalysts. In addition, biomass-derived syngas can be transformed to butanol on non-noble metal catalysts promoted by alkali metals. The prospect of catalytic renewable butanol production is also discussed.

Combining acetone-butanol-ethanol production and methyl orange decolorization in wastewater by fermentation with solid food waste as substrate

Simultaneously producing acetone-butanol-ethanol (ABE) and decolorizing Methyl orange (MO) by fermentation with solid food waste (SFW) was studied. During the batch fermentation with MO concentration from 100 mgL−1 to 1000 mgL−1, carbon source utilization efficiency was above 93% with yields of acetone, butanol and ethanol being 0.13 gg-1, 0.23 gg-1 and 0.04 gg-1, respectively. During the repeated fermentation with the bagasse as the cell immobilization material, carbon source utilization efficiencies were all above 90% in the first 5 cycles fermentation. The maximum butanol and ABE concentration were 12.8 gL-1 and 22.1 gL-1, respectively. The solvent yield of acetone, butanol and ethanol were around 0.13 gg-1, 0.25 gg-1 and 0.06 gg-1, respectively. The MO decolorization had no negative effect on the ABE production and the decolorization efficiencies were all above 99% during batch fermentation or repeated fermentation with electrons transferred from intracellular to extracellular by mediator with the help of azo reductase to decolorize MO.

Safety and Control Analysis of Hybrid Liquid-liquid Extraction and Divided Wall Column for Biobutanol Purification

Butanol is getting attention among the possible biofuels as an alternative to bioethanol. Compared to bioethanol, biobutanol shows less miscibility with water, less flammability and corrosivity, and it has the advantage of being able to directly replace gasoline in the car engines without any modifications. However, the production cost from the Acetone-Butanol-Ethanol (ABE) fermentation process remains high, mainly due to the low yield of butanol. The conventional recovery of butanol by distillation is a process of high energy consumption that has greatly restricted its industrial production. In this work different hybrid configurations, based on liquid-liquid extraction and dividing wall columns were examined. Sustainability was analysed based on green metrics, inherent security, and control properties through open and closed-loop policy using PI controllers. The results indicate that as long as the process is highly intensified, the sustainability and the inherent safety are improved but not necessarily the control properties. The best alternative was selected as a compromise between contrasting objective functions.

Butanol production from Saccharina japonica hydrolysate by engineered Clostridium tyrobutyricum: The effects of pretreatment method and heat shock protein overexpression

Macroalgal biomass is currently considered as a potential candidate for biofuel production. In this study, the effects of pretreatment method and heat shock protein overexpression were investigated for efficient butanol production from Saccharina japonica using engineered Clostridium tyrobutyricum. First, various pretreatment methods including acid hydrolysis, acid hydrolysis and enzymatic saccharification, and ultrasonic-assisted acid hydrolysis were employed to obtain the fermentable sugars, and the resulted hydrolysates were evaluated for butanol fermentation. The results showed that ultrasonic-assisted acid hydrolysate obtained the highest butanol yield (0.26 g/g) and productivity (0.19 g/L⋅h). Then, the effects of homologous or heterologous heat shock protein overexpression on butanol production and tolerance were examined. Among all the engineered strains, Ct-pMA12G exhibited improved butanol tolerance and enhanced butanol production (12.15 g/L butanol with a yield of 0.34 g/g and productivity of 0.15 g/L⋅h) from 1.8-fold concentrated S. japonica hydrolysate, which was the highest level ever reported for macroalgal biomass.

Studies on Three 1‐ O ‐acylglycerol‐3‐P Acyl transferases in Clostridium beijerinckii

The biosynthesis of phospholipids in bacteria begins with the acylation of sn-glycerol-3-P. The second step is the acylation of the resulting 1-O-acyl-glycerol-3-P catalyzed by PlsC to form phosphatidic acid. Clostridium beijerinckii has two genes annotated as plsC and a related third gene, plsD, of unknown function. We have used CRISPR-Cas9 to delete each of these genes and examined the effects on phospholipid chain compositions. Deletion of Cbei_5028 (plsC) had no effect on the composition of the acyl or alk-1-enyl chains of the lipids. Deletion of Cbei_1741 (plsC) resulted in polar lipids with fewer unsaturated chains in diacyl lipids and plasmalogens. Deletion of Cbei_0579 (plsD) resulted in longer chains in all the lipids examined. It appears that Cbei_1741 (PlsC) is selective for unsaturated fatty acyl chains and that Cbei_0579 (PlsD) is a 1-O-acyl-sn-glycerol-3-P acyltransferase selective for shorter acyl chains. Strain characterization experiments demonstrated that ∆Cbei_1741 mutant had slightly increased tolerance to butanol compared to the wild type, possibly due to the more saturated chains in diacyl lipids and plasmalogens in the cell membrane. More dramatically, ∆Cbei_1741 mutant grew much faster and produced 40% more butanol (with more glucose consumed) than the wild type in a regular batch fermentation (at 35 oC). Comparatively, both ∆Cbei_5028 and ∆Cbei_0579 mutant strains produced slightly lower butanol than wild type under this condition. Interestingly, ∆Cbei_0579 produced 40% more butanol than the wild type when the fermentation was conducted at 20 oC. While at higher temperature (40 oC), the butanol yield in ∆Cbei_0579 reached 0.32 g/g (33% higher than in wild type) although the final butanol titer was actually slightly lower than in wild type.

Disruption of hydrogenase gene for enhancing butanol selectivity and production in Clostridium acetobutylicum

Butanol production by solventogenic clostridia has long been complicated with the formation of acetone as a main byproduct, which causes a low product yield. Here, although some recent attempts failed to engineer the hydrogenase gene hydA in wild-type Clostridium, we successfully disrupted hydA with a plasmid pSY6 in C. acetobutylicum ATCC 55025. The hydA disrupted strain was able to produce 18.3 % more butanol with by-product acetone decreased by 31.2 %, indicating the inactivation of hydrogenase regulated redox balance for selective inhibition of acetone production. Exogenous supplementation of methyl viologen altered the carbon flux and further reduced acetone formation, with the maximum butanol yield of 0.28 g/g and butanol ratio of 84.0 %, respectively. The similar improvements of butanol yield and ratio were observed with corn stover as carbon source, indicating hydA disrupted strain had the potential for biobutanol production using lignocellulosic biomass. This study provides valuable understanding for regulation of redox and solventogenesis by engineering hydrogenase in Clostridium species towards the advanced production of butanol.

A neutral red mediated electro-fermentation system of Clostridium beijerinckii for effective co-production of butanol and hydrogen

To enhance the co-production of butanol and hydrogen by the acetone-butanol-ethanol (ABE) fermentation of Clostridium beijerinckii, a novel cathodic electro-fermentation (CEF) system was constructed with neutral red (NR) as electron mediator. With the mediation of NR, production of butanol and hydrogen from glucose in the CEF system achieved 5.49 ± 0.28 g/L and 3.74 ± 0.16 L/L, 569.5% and 325.0% higher than that in the open circuit (OC) system, respectively. The butanol and hydrogen yield of 0.30 ± 0.02 g/g and 206.53 ± 8.20 mL/g was 172.7% and 71.4% higher than that in the OC system, respectively. The effective co-production of butanol and hydrogen in the NR-mediated CEF system was attributed to the cooperation of the introduced polarized electrode and the additional NR. With the control of the polarized electrode, a feasible ORP was available for the effective hydrogen production. And the additional NR had induced more carbon source and electrons to the synthesis of butanol.

Modelling and Simulation of Biobutanol Fermentation by Clostridium saccharoperbutylacetonicum N1-4

Biobutanol exhibits high octane number, non-hygroscopic, non-corrosive and can be mixed with gasoline without engine modification. Furthermore, the use of biobutanol as a fuel can reduce the negative impact of environmental due to the use of fossil-based fuel. Therefore, research on the development of biobutanol is still very much needed, including research on modelling and simulation of biobutanol production by fermentation. With modelling and simulation, we can predict various biobutanol production behaviour. This paper deals with the modelling and simulation of the production of biobutanol from glucose in batch bioreactor using Clostridium saccharoperbutylacetonicum N1-4. C. saccharoperbutylacetonicum N1-4 exhibits a good growth ability on simple or complex sugars, such as glucose, sucrose, or starch. This study concerned with the kinetics models of biobutanol production as developed by Shinto based on the C. acetobutylicum metabolic pathway which consists of the glycolysis, acidogenesis and solventogenesis pathways with several simplification of reactions. In this study, modelling was carried out by adapting the rate parameter value of k13 indicating the rate constant of cell death. The adapted models were applied to determine the effect of the initial concentration of glucose and organic acids on the production of butanol and compared to the experimental data from previous research. Model simulations were performed by using MATLAB® software. The butanol yield was affected by the initial glucose concentration. The highest butanol yield of 77% was obtained at a glucose concentration of 22 g/L. The simulation results exhibit that the biobutanol yield decreases when the glucose concentration is higher than 22 g/L. These results are consistent with previous experimental data. The simulation results also show the effect of the concentration of acetic acid in the fermentation medium and can be optimized. From the simulation results, the presence of 7.5 g/L acetic acid can increase the biobutanol production by 20%. However, previous experimental data showed that a concentration of 2.5 g/L acetic acid was preferred. The simulation results also show the effect of the addition of butyric acid and lactic acid on the production of biobutanol.

Well-to-wake analysis of switchgrass to jet fuel via a novel co-fermentation of sugars and CO2

Lignocellulosic biomass such as switchgrass can be converted to n-butanol using fermentation, which can be further processed into jet fuel. Traditional acetone-butanol-ethanol (ABE) fermentation only converts sugars derived from switchgrass to ABE. Novel co-fermentation processes convert sugars and gas (CO2/H2) produced during fermentation into butanol, thus increasing ABE yields by 15.5% compared to traditional ABE fermentation. Herein, the environmental impact of a Switchgrass to Jet Fuel (STJ) pathway was assessed using life cycle assessment (LCA) from well-to-wake. LCAs were performed for greenhouse gas (GHG) emissions from jet fuel production via co-fermentation of sugars and gas for ideal and practical cases of ABE fermentation and seven other jet fuel pathways. The ideal case assumes 100% sugar recovery and 95% ABE yield. The practical case assumes 90% sugar recovery and an 80% ABE yield. Results are presented based on 100-year global warming potential (GWP) per MJ of jet fuel. Co-products were allocated using various methods. The increase in butanol yield via the co-fermentation technology reduced GWP-100 for the STJ pathway by 6.5% compared to traditional ABE fermentation. Similarly, the STJ pathway for the practical case with co-fermentation had 14.2%, 47.5%, 73.8%, and 44.4% less GWP-100 compared to HRJ, Fischer-Tropsch jet fuel from switchgrass, Fischer-Tropsch jet fuel from coal, and conventional petroleum jet fuel. The results demonstrate that the STJ pathway via co-fermentation has the potential to increase product yield while reducing GHG emissions compared to other jet fuel production pathways.

Capability of Immobilized Clostridium beijerinckii TISTR 1461 on Lotus Stalk Pieces to Produce Butanol from Sugarcane Molasses

Immobilized Clostridium beijerinckii TISTR 1461 was used to enhance the butanol production efficiency from sugarcane molasses. Lotus stalk (LS) pieces were used as carriers for cell immobilization. Sugarcane molasses containing 50 g/L of sugar supplemented with 1 g/L of yeast extract was found to be an appropriate medium for bacterial cell immobilization on the LS pieces. Carrier size (4, 12 and 20 mm in length) and carrier loading (1:15, 1:30 and 1:45, w/v) were optimized for high levels of butanol production using response surface methodology (RSM). The batch fermentation was carried out under anaerobic conditions in 1 L screw-capped bottles at 37 °C and an agitation rate of 150 rpm. It was found that the optimum conditions for the butanol production were the carrier size of 4 mm and carrier loading of 1:31 (w/v). Under these conditions, the butanol concentration (PB) was 12.89 g/L, corresponding to the butanol productivity (QB) of 0.36 g/L∙h and butanol yield (YB/S) of 0.36 g/g. These values were higher than those using free cells (PB, 10.20 g/L, QB, 0.28 g/L∙h and YB/S, 0.32 g/g). In addition, it was found that a 24 h incubation time for cell immobilization was appropriate for the immobilization process, which was confirmed by the results of the scanning electron microscope (SEM) and atomic force microscopy (AFM) images and specific surface area measurement. When the fermentation using the immobilized cells was carried out in a stirred-tank reactor (STR), column reactor (CR) and CR coupled with STR, the results showed that all reactors could be used to produce butanol production from the immobilized cells on LS pieces. However, the PB using CR and CR coupled with STR were only 75% and 45% of those using the screw-capped bottle and STR.

Orange peel extract enhanced sugar recovery and butanol production from potato peel by Clostridium acetobutylicum

ABSTRACT The present study is focused on biobutanol production from potato peel waste in dilute orange peel extract by Clostridium acetobutylicum MTCC 11274 at three concentrations of 20, 40, and 60 g/L as sole nutrient sources. The biomass and solvent production from potato peel were enhanced in orange peel extract with the concentration and improved the sugar recovery. The effect on solvent production from 60 g/L potato peel in dilute orange peel extract was relatively higher than 20 and 40 g/L. The butanol yield was improved from 0.04 to 0.35 g/g from 60 g/L potato peel waste in orange peel extract. The butyric acid concentration was highly correlated with butanol production. The Clostridium acetobutylicum MTCC 11274 showed no inhibitory effect from D-limonene in orange peel extract. The potato peel waste hydrolyzate prepared in orange peel extract enhanced the growth of Clostridium acetobutylicum MTCC 11274 and fermentation of acetone, butanol, and ethanol.

Catalytic conversion of ethanol to butanol over magnesium oxide catalysts

Several types of magnesium oxide (MgO) has been studied as catalyst for catalytic conversion of ethanol into butanol. The reactions have been conducted at 350 °C in batch reactor. MgO catalysts have been characterized by XRD, CO2-TPD, and N2-physisorption. It was found that of the highest yield of butanol was obtained from MgO synthesized from precipitation method. The highest butanol yield was 2.4% at our experimental conditions. This was attributed to its surface area and basicity.

Advances in the synthesis of biobutanol by consolidated bioprocessing from lignocellulose

Consolidated bioprocessing (CBP) is a multi-step process in a bioreactor, which completes hydrolase production, enzymatic hydrolysis, and microbial fermentation. It is considered to be the most promising process for the production of second-generation biofuels because of its simple steps and low cost. Due to the complexity of lignocellulose degradation and the butanol synthesis pathway, few wild microorganisms can directly utilize lignocellulose to synthesize butanol. With the development of synthetic biology, single-bacterium directly synthesizes butanol using lignocellulose by introducing a butanol synthesis pathway in the cellulolytic Clostridium. However, there are still some problems such as heavy metabolic load of single bacterium and low butanol yield. Co-culture can relieve the metabolic burden of single bacterium through the division of labor in different strains and can further improve the efficiency of butanol synthesis. This review analyzes the recent research progress in the synthesis of biobutanol using lignocellulose by consolidated bioprocessing from both the single-bacterium strategy and co-culture strategy, to provide a reference for the research of butanol and other biofuels.

Enhanced coproduction and trade-off of the hydrogen and butanol in the coupled system of pervaporation and repeated-cycle fixed-bed fermentation

In the study, a coupled system of pervaporation and repeated-cycle fixed bed fermentation with the bagasse as the carrier material was developed. Three repeated cycles of fermentation lasting for 540 h was efficiently carried out to coproduce hydrogen and butanol with the strain Clostridium acetobutylicum. From 1st cycle to 3rd cycle, the average hydrogen productivity and butanol productivity were around 130 mLL−1 h−1 and 0.15 gL−1 h−1, respectively. Average hydrogen yield and butanol yield were around 0.17 Lg−1 and 0.19 gg−1. The hydrogen and butanol production per unit cell weight were 27 Lg−1 and 31 gg−1, which were more than two times of those in a conventional repeated-cycle fermentation. The trade-off and competition between the hydrogen and butanol production was studied and analyzed. Compared with the pervaporation-free fermentation, the hydrogen yield was decreased by 14 % and butanol yield was increased by 27 % in the coupled system. Instantaneous solvent removal from the broth by pervaporation changed probably the metabolic pathway to more butanol production over hydrogen. From 1st cycle to 3rd cycle, the respiration heat generated during each fermentation cycle could cover 45 %, 24 % and 37 % of the total energy requirement of the pervaporation achieving the best use of the energy.