Total Acetone-butanol-ethanol Research Articles

Acetone-Butanol-Ethanol (ABE) fermentation with clostridial co-cultures for enhanced biobutanol production

This study has addressed the issue of enhanced biobutanol production through Consolidated Bioprocessing, with the use of clostridial co-cultures combining C. acetobutylicum (Cac) and C. pasteurianum (Cpa). To begin with, the total Acetone-Butanol-Ethanol (ABE) solvent production and butanol selectivity of individual clostridial cultures grown on different carbon sources present in lignocellulosic hydrolysates. Thereafter, statistical optimization of ABE fermentation with co-culture system was carried out using Box-Behnken design. The optimization variables were Cac to Cpa inoculum ratio (X1 in mL mL−1), sodium concentration in the medium (X2 g L−1), and xylose/glucose ratio (X3 g g−1 L L−1), whereas response variables were butanol titre (Y1 in g L−1), biomass growth (Y2 OD600), total ABE titre (Y3 in g L−1). Maximum values of the response variables with the corresponding set of optimization parameters were as follows: Y1, max = 12.1 ± 0.45 (for X1 = 0.30, X2 = 5.90, X3 = 0), Y2, max = 4.15 ± 0.03 (for X1 = 0.3, X2 = 2.42, X3 = 0.14), Y3, max = 23.1 ± 0.55 (for X1 = 0.46, X2 = 6.36, X3 = 0). The study demonstrated a substantial ≈ 22% and 61% increment in butanol titre in co-culture system compared to individual fermentations with Cac and Cpa grown on pure glucose, respectively.

Effects of Clostridium beijerinckii and Medium Modifications on Acetone-Butanol-Ethanol Production From Switchgrass.

The presence of lignocellulose-derived microbial inhibitory compounds (LDMICs) in lignocellulosic biomass (LB) hydrolysates is a barrier to efficient conversion of LB hydrolysates to fuels and chemicals by fermenting microorganisms. Results from this study provide convincing evidence regarding the effectiveness of metabolically engineered C. beijerinckii NCIMB 8052 for the fermentation of LB-derived hydrolysates to acetone–butanol–ethanol (ABE). The engineered microbial strain (C. beijerinckii_SDR) was produced by the integration of an additional copy of a short-chain dehydrogenase/reductase (SDR) gene (Cbei_3904) into the chromosome of C. beijerinckii NCIMB 8052 wildtype, where it is controlled by the constitutive thiolase promoter. The C. beijerinckii_SDR and C. beijerinckii NCIMB 8052 wildtype were used for comparative fermentation of non-detoxified and detoxified hydrothermolysis-pretreated switchgrass hydrolysates (SHs) with and without (NH4)2CO3 supplementation. In the absence of (NH4)2CO3, fermentation of non-detoxified SH with C. beijerinckii_SDR resulted in the production of 3.13- and 2.25-fold greater quantities of butanol (11.21 g/L) and total ABE (20.24 g/L), respectively, than the 3.58 g/L butanol and 8.98 g/L ABE produced by C. beijerinckii_wildtype. When the non-detoxified SH was supplemented with (NH4)2CO3, concentrations were similar for butanol (9.5 compared with 9.2 g/L) and ABE (14.2 compared with 13.5 g/L) produced by C. beijerinckii_SDR and C. beijerinckii_wildtype, respectively. Furthermore, when C. beijerinckii_SDR and C. beijerinckii_wildtype were cultured in detoxified SH medium, C. beijerinckii_SDR produced 1.11- and 1.18-fold greater quantities of butanol and ABE, respectively, than when there was culturing with C. beijerinckii_wildtype. When the combined results of the present study are considered, conclusions are that the microbial strain and medium modifications of the fermentation milieu resulted in greater production of fuels and chemicals from non-detoxified LB hydrolysates.

ZIF-L based mixed matrix membranes for acetone-butanol-ethanol (ABE) recovery from diluted aqueous solution

Enrichment of bio-based platform compound ABE (acetone-butanol-ethanol) from extremely diluted raw medium is pivotal for the full utilization of ABE and selective conversion of them into value-added chemicals. Vapor permeation process through mixed matrix membranes (MMMs) demonstrates an energy-efficient solution to this challenge. Here, MMMs, by doping two-dimensional ZIF-L nanosheets with porous skeletons into an oleophilic matrix (poly-R-siloxane), were produced to synchronously enrich ABE from the simulated fermentation broth (total ABE of 3.0 wt% or lower, and composition of A: B: E = 3: 6: 1). The good compatibility of ZIF-L and polymer matrix renders MMMs with significantly improved flux and separation factor of organics over water. Typically, MMMs incorporating 5.0 wt% of ZIF-L fillers contribute to an increase of 72.3% and 106% in liquid flux and butanol/water separation factor, respectively, when compared with the pure polymer membranes. Through ZIF-L based MMMs, the water content in the permeant is controlled below 30 wt%, which is a prerequisite for utilization of ABE in the real bio-refinery industry.

Biological production of hydrogen and acetone- butanol-ethanol from sugarcane bagasse and rice straw using co-culture of Enterobacter aerogenes and Clostridium acetobutylicum

Production of hydrogen and acetone-butanol-ethanol (ABE) from hydrolysates of sugarcane bagasse (SCB) and rice-straw (RS) was carried out via dark fermentation using co-culture of Enterobacter aerogenes and Clostridium acetobutylicum. The best improvement in the yield of hydrogen and total ABE production were observed for alkaline pretreated RS at 100 °C for 60 min obtained from the highest total reducing sugars concentration of 40.7 g L−1 after the enzymatic hydrolysis. The hydrolysates of alkaline pretreated samples at 100 °C were efficiently converted to H2 by co-culture of Enterobacter aerogenes and Clostridium acetobutylicum with the highest hydrogen yield of 136.3 and 149.9 mL H2 g−1-VS of SCB and RS, representing 3.4 and 1.5-fold enhancement in the hydrogen production yield over that obtained from the pure cultures of Enterobacter aerogenes and Clostridium acetobutylicum, respectively. Mass balance and energy generation analysis indicate that the highest specific ABE and total energy production yields per gram untreated feedstock reached 0.12 g ABE and 5.5 kJ for SCB and 0.08 g ABE and 6.6 kJ for RS.

Two-step fermentation of cooked rice with Aspergillus oryzae and Clostridium acetobutylicum YM1 for biobutanol production

In Asia, uneaten cooked rice is the highest portion amongst many forms of food wastes that are thrown away. In order to make use of the thrown-away rice and potentially use it for liquid fuels, steamed Japanese rice was evaluated on biobutanol production through a two-step fermentation by amylase-producing Aspergillus oryzae, and solvent-producing Clostridium acetobutylicum YM1. The effects of sterilization and providing anaerobic conditions on solvent production in acetone-butanol-ethanol (ABE) fermentation cannot be underestimated. Several conditions, including aerobic, anaerobic, sterile, and non-sterile were investigated concerning the solvent production capability of Clostridium acetobutylicum YM1. The maximum solvent production was 11.02 ± 0.22 g/l butanol and 18.03 ± 0.34 g/l total ABE from 75 g/l dried rice. The results confirmed that the solvent production performance of the YM1 strain was not affected by the sterilization conditions. In particular, 10.91 ± 0.16 g/l butanol and 16.68 ± 0.22 g/l ABE were produced under non-sterile and aerobic conditions, which can reduce industrial-scale production costs.

Fluorinated PDMS membrane with anti-biofouling property for in-situ biobutanol recovery from fermentation-pervaporation coupled process

Hydrophobic pervaporation membranes such as polydimethylsiloxane (PDMS) have shown great potential for biofuels recovery from fermentation process, which however face the challenge of biofouling issue. In this work, a new kind of anti-biofouling PDMS membrane was developed by creating an ultra-low energy surface via a facile crosslinking reaction between fluorosilane and PDMS. The chemical properties and wettability of the membrane surface were characterized by IR, XPS and contact angle measurements, in which the effect of chemical groups on the surface free energy was studied. The performance of PDMS membranes were evaluated in a typical acetone-butanol-ethanol (ABE) fermentation-pervaporation coupled process. The results demonstrated that the introduction of fluoroalkyl groups highly reduced the surface energy of PDMS membrane, thereby achieving excellent hydrophobicity and lipophobicity at the same time, and successfully alleviating microbial adhesion onto the membrane. As a result, the fluorinated PDMS membrane exhibited excellent anti-biofouling property, as well as much higher stabilized total flux (0.74 vs 0.36 kg/m2h) and ABE separation factor (21.8 vs 7.1) than the pristine PDMS membrane as coupling fed-batch fermentation for 140 h. In addition, a significant enhancement in ABE productivity (e.g., 51% higher than batch fermentation) was obtained in the fluorinated PDMS membrane coupled fed-batch fermentation process.

Biobutanol production from municipal solid waste: Technical and economic analysis

Novel processes for the production of acetone-butanol-ethanol (ABE) from municipal solid waste (MSW) were developed and simulated using Aspen Plus®. In scenario 1, a conventional distillation system was used, while a gas stripping system was coupled with a fermenter in scenario 2. In scenario 3, pervaporation (PV) and gas stripping systems right after the fermentation reactor were applied. Gas stripping increased the total ABE produced while the addition of the PV module decreased the number of distillation columns from 6 to 2 as well as created 6.4% increments in the amount of butanol in comparison with scenario 1. Economical evaluation resulted in having payout periods of 15.9, 4.4, and 2.9 years for scenarios 1 to 3, respectively. These results show that using MSW as an inexpensive sugar-rich feedstock together with gas stripping PV system is a promising solution to overcome the major obstacles in the way of the ABE production.

Isolation and characterization of Clostridia from the feces of wild rabbit and swine for hemicellulosic acetone-butanol-ethanol (ABE) production

Isolation and characterization of solventogenic bacteria from animal feces were carried out. Ten samples were collected continually for 5 d from the feces of wild rabbit and wild swine in Jawaharlal Nehru Zoological Park, Hyderabad, India. Ten acetone-positive strains were selected for evaluation of their phenotypic and physiological characteristics. Two potential solvent-producing cultures were selected for 16S rRNA gene analysis. The culture isolated from the wild rabbit feces exhibited 97.3% similarity with Clostridium beijerinckii 8052, and the culture isolated from the wild swine feces exhibited 93.8% similarity with Clostridium saccharoperbutylacetonicum NI-4(HMT). The isolated strains utilized a wide range of carbohydrate substrates including glucose, fructose, maltose, xylose, arabinose, and glycerol. The major fermentation products from glucose, xylose, and arabinose were acetone, butanol, and ethanol (ABE). The total ABE concentration produced by strain YVU1 was 13.1 g/L from arabinose, 16.3 g/L from xylose, and 20.6 g/L from glucose. Strain YVU2 produced a total ABE concentration of 16.5 g/L, 18.3 g/L, and 22.4 g/L from arabinose, xylose, and glucose, respectively.

Enhancement of acid re-assimilation and biosolvent production in Clostridium saccharoperbutylacetonicum through metabolic engineering for efficient biofuel production from lignocellulosic biomass

In the clostridial acetone-butanol-ethanol (ABE) fermentation, the intermediate acetate and butyrate are re-assimilated for solvent production. Here, key genes in ABE pathways in Clostridium saccharoperbutylacetonicum N1-4 were overexpressed to enhance acid re-assimilation and solvent production. With the overexpression of sol operon, acid re-assimilation was enhanced, and ABE production was increased by 20%, with ethanol production increased by six times but almost no increase in butanol production. To further drive carbon flux for C4 metabolites and ultimate butanol production, key genes including hbd, thl, crt and bcd in butanol production pathway were further overexpressed. Compared to the control, butanol, acetone and total ABE production in the new strain was increased by 8%, 18%, and 12.4%, respectively. Finally, simultaneous saccharification and fermentation was carried out using acetate-pretreated switchgrass. 15.4 g/L total ABE (with a yield of 0.31 g/g) was produced in both engineered strains, which was significantly higher than the control.

Enhanced butanol production using Clostridium beijerinckii SE-2 from the waste of corn processing

Acetone-butanol-ethanol (ABE) fermentation from food processing waste is one way to reduce cost. In this study, corncob hydrolysate and corn steep liquor (CSL), two waste materials from corn processing industries, were used as carbon and nitrogen sources to yield butanol by using Clostridium beijerinckii SE-2. Media compositions favoring butanol production were investigated using statistical experimental designs. Media components were first screened using a fractional factorial experimental design. CSL and CH3COONH4 were found to be the significant variables among six factors, including the contents of CSL, CH3COONH4, K2HPO4-KH2PO4, MnSO4 · H2O, MgSO4 · 7H2O and FeSO4 · 7H2O. The two factors were further optimized by the steepest assent and central composite rotatable design. The validated experiments showed that the total ABE in the system was 19.22 g L−1 and the concentration of butanol could reach 11.65 g L−1 with the optimized medium, which was 42% higher than the initial medium. Scale-up fermentation with optimized medium in the 100 L bioreactor resulted in 20.29 g L−1 of ABE and 11.92 g L−1 of butanol. Moreover, C. beijerinckii SE-2 can use both glucose and xylose from corncob hydrolysate. In conclusion, our study suggested that the waste of the corn processing industry can be used as sources to produce butanol by Clostridium, and statistical experimental designs are a useful approach for optimizing media compositions for butanol production.

Hybrid pervaporation and salting-out for effective acetone-butanol-ethanol separation from fermentation broth

In this study, hybrid pervaporation/salting-out process was coupled with fed-batch acetone-butanol-ethanol (ABE) fermentation for high-titer solvents production. At the first-stage separation, 18.8 g/L of ethanol, 125.9 g/L of acetone and 199.5 g/L of butanol were obtained in the permeate of pervaporation. Model solution with similar solvents content to the permeate condensate was used for the second-stage salting-out process based on K3PO4 and KH2PO4, respectively. The effect of salting-out factor on ABE concentrations and recoveries in the aqueous and organic phase was compared and optimized. Results indicated that K3PO4 showed better salting-out performance. At the salting-out factor of 1.0, 99.01% of ABE was recovered. Correspondingly, 486.74 g/kg of butanol (805.52 g/kg of total ABE) could be tested in the organic phase. This novel hybrid process was a promising method for biobutanol production and separation.

Phenolic compounds removal from sweet sorghum grain for efficient biobutanol production without nutrient supplementation

Industrial scale production of biobutanol has been hampered by substrate cost and availability. Sweet sorghum grain is an inexpensive substrate for acetone-butanol-ethanol (ABE) production by Clostridium acetobutylicum. Amylolytic activity of C. acetobutylicum eliminates the need for the hydrolysis of starchy grain prior to fermentation. However, untreated grain contains phenolic compounds, i.e. tannins, which exhibit inhibitory effects against amylolytic activity and ABE fermentation. Less than 3g/L ABE was obtained from untreated sweet sorghum grain at different substrate concentrations. Concentration of 0.2mM gallic acid equivalent (GAE) of sorghum tannins was detected as the critical concentration which inhibits severely ABE fermentation. Applying a multi-stage hot water treatment resulted in tannins removal and significant enhancement in total ABE production up to 18g/L. For efficient butanol production from 40, 60, and 80g/L sorghum grain, hot water treatment with two, five, and six stages were found to be essential for efficient butanol production, respectively. Moreover, the amylolytic activity of C. acetobutylicum was inhibited by sorghum grain tannins, more than twice as high as the effects on the ABE fermentation pathway. Furthermore, unlike most substrates, sweet sorghum grain could provide all nutrients required for ABE fermentation, eliminating the need for supplementing expensive additional nutrients.

Fed-batch fermentation with intermittent gas stripping using immobilized Clostridium acetobutylicum for biobutanol production from corn stover bagasse hydrolysate

Corn stover bagasse hydrolysate was used as the substrate for fed-batch acetone-butanol-ethanol (ABE) fermentation and in situ product removal by intermittent gas stripping. The mutant strain Clostridium acetobutylicum ABE-P 1201 was inoculated for ABE production. As results, a final cumulative butanol concentration of 18.6g/L (total ABE: 28.3g/L) was produced from 97.6g/L of total fermentable sugars (75.3g/L of glucose and 22.3g/L of xylose), with an ABE yield of 83.2g/kg of CSB. The gas stripping condensate obtained in the integrated process contained 77–136g/L of butanol and 115–220g/L of total ABE.

New Insight into Sugarcane Industry Waste Utilization (Press Mud) for Cleaner Biobutanol Production by Using C. acetobutylicum NRRL B-527.

In the present study, press mud, a sugar industry waste, was explored for biobutanol production to strengthen agricultural economy. The fermentative production of biobutanol was investigated via series of steps, viz. characterization, drying, acid hydrolysis, detoxification, and fermentation. Press mud contains an adequate amount of cellulose (22.3%) and hemicellulose (21.67%) on dry basis, and hence, it can be utilized for further acetone-butanol-ethanol (ABE) production. Drying experiments were conducted in the temperature range of 60-120°C to circumvent microbial spoilage and enhance storability of press mud. Furthermore, acidic pretreatment variables, viz. sulfuric acid concentration, solid to liquid ratio, and time, were optimized using response surface methodology. The corresponding values were found to be 1.5% (v/v), 1:5g/mL, and 15min, respectively. In addition, detoxification studies were also conducted using activated charcoal, which removed almost 93-97% phenolics and around 98% furans, which are toxic to microorganisms during fermentation. Finally, the batch fermentation of detoxified press mud slurry (the sample dried at 100°C and pretreated) using Clostridium acetobutylicum NRRL B-527 resulted in a higher butanol production of 4.43g/L with a total ABE of 6.69g/L.

High-efficient n-butanol production by co-culturing Clostridium acetobutylicum and Saccharomyces cerevisiae integrated with butyrate fermentative supernatant addition

Butanol is not only an important chemical intermediate and solvent in pharmaceutical and cosmetics industries, but also considered as an advanced biofuel. Although species of the natural host Clostridium have been engineered, butanol titers in the anaerobe seem to be limited by its intolerance to butanol less than 13g/L. Here we aimed to develop a technology for enhancing butanol production by a co-culture system with butyrate fermentative supernatant addition. First, when adding 4.0g/L butyrate into the acetone-butanol-ethanol (ABE) fermentation broth with single-shot at 24h, the "acid crash" phenomenon occurred and the ABE fermentation performance deteriorated. Subsequently, we found that adding certain amino acids could effectively enhance butyrate re-assimilation, butanol tolerance and titer (from 11.1 to 14.8g/L). Additionally, in order to decrease the raw material cost, butyrate fermentative supernatant produced by Clostridium tyrobutyricum was applied to butanol production in the Clostridium acetobutylicum/Saccharomyces cerevisiae co-culture system, instead of adding synthetic butyrate. Final butanol and total ABE concentrations reached higher levels of 16.3 and 24.8g/L with increments of 46.8 and 37.8%, respectively. These results show that the proposed fermentation strategy has great potential for efficiently butanol production with an economic approach.

Development of a High-Efficiency Transformation Method and Implementation of Rational Metabolic Engineering for the Industrial Butanol Hyperproducer Clostridium saccharoperbutylacetonicum Strain N1-4.

This paper presents the first steps toward advanced genetic engineering of the industrial butanol producer Clostridium saccharoperbutylacetonicum strain N1-4 (HMT). In addition to providing an efficient method for introducing foreign DNA into this species, we demonstrate successful rational engineering for increasing solvent production. Examples of future applications of this work include metabolic engineering for improving desirable industrial traits of this species and heterologous gene expression for expanding the end product profile to include high-value fuels and chemicals.

Sustainable biobutanol production using alkali-catalyzed organosolv pretreated cornstalks

Biobutanol is an important alternative fuel source, but current processing methods suffer from low yields. This study aimed to develop and optimize a pretreatment protocol for biobutanol production from cornstalks. Fractionation of cornstalks into carbohydrate (cellulose and hemicellulose) and lignin was performed by alkali-catalyzed organosolv pretreatment (ACOS). After optimization of the process parameters, more than 80% of the total lignin was removed, with minimal hemicellulose degradation, at 110°C, 4% (w/w dry cornstalk) NaOH, 90min reaction time, and 60% (v/v) ethanol. After enzymatic hydrolysis, the maximum recovery of total monosaccharide was 83.7% (85.0% cellulose, 82.0% hemicellulose). In acetone-butanol-ethanol (ABE) fermentation, a slightly higher total ABE concentration (12.8g/L vs. 11.9g/L) was produced from the enzymatic hydrolysate, compared with that from a glucose control. The physical structure and chemical properties of alkali-catalyzed organosolv lignin (ACOSL) showed higher phenolic group content and antioxidant capacity compared with alkali lignin. ACOS pretreatment is an economical method for the production of fermentable monosaccharide and high-value lignin, for use in biofuel production.

Butanol production by Clostridium pasteurianum NRRL B-598 in continuous culture compared to batch and fed-batch systems

Butanol production by acetone–butanol–ethanol (ABE) fermentation is usually associated with Clostridium acetobutylicum. In this report, ABE fermentation using the little known, oxygen tolerant solventogenic strain, Clostridium pasteurianum NRRL B-598, in packed-bed continuous cultures is described and compared with suspended cells in continuous, batch and fed-batch processes. While the highest butanol and total ABE concentrations (8.3 and 12.3g/L respectively), were obtained in fed-batch fermentation, the highest solvent yield and productivity (32% and 0.73g/L/h), were achieved in packed-bed continuous fermentation at a dilution rate of 0.121/h. In these processes, the strain showed exceptional stability, as demonstrated not only by low deviations in process parameters in batch and fed-batch experiments but also long-term stability (over 700h, or more than 35 bioreactor residence times) in continuous fermentations.

Isolation of a Clostridium acetobutylicum strain and characterization of its fermentation performance on agricultural wastes

A new solvent-producing Clostridium has been isolated from soil used in intensive rice cultivation. The 16S rRNA analysis of the isolate indicates that it is closely related to Clostridium acetobutylicum, with a sequence identity of 96%. The new isolate, named C. acetobutylicum YM1, produces biobutanol from multiple carbon sources, including glucose, fructose, xylose, arabinose, glycerol, lactose, cellobiose, mannitol, maltose, galactose, sucrose and mannose. This isolate can also utilize polysaccharides such as starch and carboxylmethyl cellulose (CMC) for the production of biobutanol. The ability of isolate YM1 to produce biobutanol from agro-industrial wastes was also evaluated for rice bran, de-oiled rice bran, palm oil mill effluent and palm kernel cake. The highest concentration of biobutanol (7.27 g/L) was obtained from the fermentation medium containing 2% (w/v) fructose, with a total acetone–butanol–ethanol (ABE) concentration of 10.23 g/L. The ability of isolate YM1 to produce biobutanol from various carbon sources and agro-wastes indicates the promise of the use of this isolate for the production of biobutanol, a renewable energy resource, from readily available renewable feedstocks.

Feasibility of acetone–butanol–ethanol (ABE) fermentation from Amorphophallus konjac waste by Clostridium acetobutylicum ATCC 824

Using Amorphophallus konjac waste as a source of fermentable carbohydrate, the feasibility of acetone–butanol–ethanol (ABE) fermentation by Clostridium acetobutylicum ATCC 824 was investigated in this study. Two distinct combination strategies for the enzymatic hydrolysis and fermentation, simultaneous saccharification and fermentation (SSF) and separate hydrolysis and fermentation (SHF), for konjac waste fermentation were compared. β-Mannanase was added to facilitate the hydrolysis of glucomannan and to increase the fermentability of the substrate. The ABE concentration (6.64g/L) from SSF was less than that from SHF; the result from the control experiment was 4.34g/L of ABE. SHF obtained a higher production of total ABE solvents (10.95g/L) with 7.10g/L of butanol at a productivity rate of 0.30g/L/h. The effect of the bioactive alkaloid abundant in the konjac waste on ABE fermentation was evaluated. The results indicated that the alkaloid affected the ABE fermentation results when using C. acetobutylicum, and the effective extraction of alkaloids can help to increase the butanol concentration and shorten the fermentation period. Therefore, the utilization of Amorphophallus konjac waste (a low-cost agricultural resource) in ABE fermentation provides an alternative to increase the economic viability of ABE production.