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[Display omitted] • C. kluyveri bioaugmentation is proposed to enhance caproic acid fermentation. • Caproic acid concentration and selectivity reached 12.89 g/L and 55.08 %. • Microbial community structure was shifted by bioaugmentation. • Bacteria involved in degrading straw and producing caproic acid were enriched. Producing caproic acid via carboxylate platform is an environmentally-friendly approach for treating lignocellulosic agricultural waste. However, its implementation is still challenged by low product yields and selectivity. A microbiome named cellulolytic acid-producing microbiome (DCB), proficient in producing cellulolytic acid, was successfully acquired and shows promise for producing high-level caproic acid. In this study, a bioaugmentation method utilizing Clostridium kluyveri is proposed to enhance caproic acid yield of DCB using rice straw. With exogenous ethanol, bioaugmentation with Clostridium kluyveri significantly improved the caproic acid concentration and selectivity by 7 times and 4.5 times, achieving 12.9 g/L and 55.1 %, respectively. The addition of Clostridium kluyveri introduced reverse β-oxidation pathway, a more efficient caproic acid production pathway. Meanwhile, bioaugmentation enriched the bacteria proficient in degrading straw and producing short-chain fatty acids, providing more substrates for caproic acid production. This study provides potential bioaugmentation strategies for optimizing caproic acid yield from lignocellulosic biomass. [ABSTRACT FROM AUTHOR]

[Display omitted] • The phaP gene was first introduced for the purpose of enhancing hydrogen production. • Among engineering strain, the phaP strain exhibited the highest hydrogen production. • The introduction of phaP led to a decrease in the transcription of chaperone genes. • The introduction of phaP enhanced hydrogen production 2.6-fold compared to wild type. In this study, the goal was to enhance the tolerance of Clostridium acetobutylicum ATCC 824 to biomass-based inhibitory compounds for biohydrogen production and evaluate various known genes that enhance the production of biochemicals in various hosts. The introduction of phaP , the major polyhydroxyalkanoate granule-associated protein that has been reported as a chaperone-like protein resulted in increased tolerance to inhibitors and leads to higher levels of hydrogen production, cell growth, and glucose consumption in the presence of these inhibitors. It was observed that the introduction of phaP led to an increase in the transcription of the hydrogenase gene, whereas transcription of the chaperone functional genes decreased compared to the wild type. Finally, the introduction of phaP could significantly enhance biohydrogen production by 2.6-fold from lignocellulosic hydrolysates compared to that of wild type. These findings suggested that the introduction of phaP could enhance growth and biohydrogen production, even in non-polyhydroxyalkanoate-producing strains. [ABSTRACT FROM AUTHOR]

[Display omitted] • Spent Substrate of Pleurotus ostreatus can act as potential biomass for biobutanol production. • Clostridium acetobutylicum MTCC 11274 produced ABE concentration of 10.80 ± 0.02 g/L. • Hybrid pre-treatment showed maximum biobutanol yield of 0.38 g/g. • SEM, FT-IR, XRD data confirmed the breakdown of lignin and hemicellulose. Addressing global energy demand, researchers sought eco-friendly biobutanol production from lignocellulosic waste biomass. In the present research work, five different pre-treatment methods viz., Microwave, Ultrasound, Alkali, Acid, and Hybrid, were investigated to explore its biobutanol production potential by utilizing Pleurotus ostreatus spent as substrate. The compositional and physico-chemical changes of the pre-treated Spent Mushroom Substrate (SMS) were assessed using SEM, FTIR, and XRD. Hybrid pre-treatment (Microwave, Alkali, Ultrasound) showed higher delignification when compared to conventional pre-treatment method. Hybrid pre-treated SMS resulted in higher total reducing sugars (521.53 ± 1.84 mg/g) than indigenous SMS (267.89 ± 1.53 mg/g). Fermentation of hybrid pre-treated SMS with Clostridium acetobutylicum MTCC 11274 produced the highest biobutanol concentration (9.84 ± 0.03 g/L) and yielded 0.38 ± 0.02 g/g of biobutanol. This study revealed that hybrid pre-treatment could be a promising solution for enhanced biobutanol production using SMS biomass. [ABSTRACT FROM AUTHOR]

• C. kluyveri managed to grow in a medium with different levels of available CO 2. • 11.0 g/L C 6 was reached from acetate and ethanol in a bioreactor with CO 2 supply. • There is a direct correlation between CO 2 concentration and C. kluyveri 's growth. • CO 2 uptake by C. kluyveri at bioreactor level was demonstrated. Chain elongation technology allows medium-chain fatty acids (MCFAs) production as an alternative to fossil resources. Clostridium kluyveri generates n -caproate primarily from ethanol and acetate, presumably requiring CO 2 for growth. Here, the impact of CO 2 on C. kluyveri was explored. Bottle studies revealed the bacterium's adaptability to low CO 2 levels, even in conditions with minimal dissolved NaHCO 3 (0.0003 M) and unfavorable pH (below 6) under 1 bar CO 2. Bioreactor investigations demonstrated a direct correlation between CO 2 availability and bacterial growth. The highest n -caproate production (11.0 g/L) with 90.1 % selectivity was achieved in a bioreactor with continuous CO 2 supply at 3 mL/min. Additional bottle experiments pressurized with 1 bar CO 2 and varying ethanol:acetate ratios (1:1, 2:1, 4:1) also confirmed CO 2 consumption by C. kluyveri. However, increasing the ethanol:acetate ratio did not enhance n -caproate selectivity, likely due to overly acidic pH conditions. These findings provide insights into chain-elongators responses under diverse conditions. [ABSTRACT FROM AUTHOR]

[Display omitted] • Liquid fraction containing glycerol was used for recycled corn stover pretreatment. • Dry cell weight of P. pastoris reached ∼ 40 g/L with glycerol as sole carbon source. • The titer of human lysozyme reached 1.3 g/L by P. pastoris with methanol induction. • Mixing pretreated corn stover with cassava as co-substrate to produce biobutanol. • Final butanol titer of 13.9 g/L was produced from mixed substrates under SSF. Glycerol organosolv pretreatment (GOP) is considered an efficient method to deconstruct lignocellulose for producing fermentable sugars. Herein, the liquid fraction containing glycerol after GOP was utilized for recycled pretreatment of corn stover (CS) for four cycles. Enzymatic yield of glucose after recycled pretreatment was enhanced by 2.4–3.5 folds compared with untreated CS. Meanwhile, residual glycerol was used as carbon source for cultivation of Pichia pastoris to obtain high cell-density, and a final titer of 1.3 g/L human lysozyme was produced by P. pastoris under low temperature methanol induction strategy. Additionally, the pretreated CS was mixed with cassava as fermentable substrates for butanol production by wild-type Clostridium acetobutylicum ATCC 824. Final butanol production of 13.9 g/L was obtained from mixed substrates (25%:75% of CS/cassava) at 10% solids loading by simultaneous saccharification and fermentation. Overall, integration of residual glycerol utilization and butanol production by microbial fermentation provided an efficient strategy for biorefinery. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • Clostridium strains were assessed for butanol production from agave hydrolysates. • 6.04 g/L of butanol was produced using Clostridium acetobutylicum ATCC 824. • Clostridium acetobutylicum ATCC 824 showed higher tolerance to inhibitors. • Co-culture was carried out using Bacillus subtilis and Clostridium acetobutylicum. • 8.28 g/L of butanol was produced in co-culture from agave hydrolysates. Abstract In this work, three Clostridium strains were tested for butanol production from Agave lechuguilla hydrolysates to select one for co-culturing. The agave hydrolysates medium was supplemented with nutrients and reducing agents to promote anaerobiosis. Clostridium acetobutylicum ATCC 824 had the highest butanol production (6.04 g/L) and was selected for further analyses. In the co-culture process, Bacillus subtilis CDBB 555 was used to deplete oxygen and achieve anaerobic conditions required for butanol production. The co-culture was prepared with C. acetobutylicum and B. subtilis without anaerobic pretreatment. Butanol production in co-culture from agave hydrolysates was compared with experiments using synthetic medium with glucose and a pure culture of C. acetobutylicum. The maximum butanol concentration obtained was 8.28 g/L in the co-cultured hydrolysate medium. Results obtained in the present work demonstrated that agave hydrolysates have the potential for butanol production using a co-culture of B. subtilis and C. acetobutylicum without anaerobic pretreatment. [ABSTRACT FROM AUTHOR]

Highlights • Proline addition improved the tolerance of C. acetobutylicum to multiple inhibitors. • The proline biosynthesis was enhanced by overexpressing proA , proB and proC. • The ability of scavenging reactive oxygen species in 824(proABC) was improved. • Undetoxified lignocellulosic hydrolysates can be used for butanol production. Abstract When lignocellulosic biomass was used for acetone-butanol-ethanol (ABE) fermentation, several lignocellulose-derived inhibitors, which are toxic to Clostridium acetobutylicum , were generated during acid hydrolysis process and seriously hindered the industrialization of lignocellulosic butanol. In this study, an engineered strain 824(proABC) with significantly improved tolerance to multiple lignocellulose-derived inhibitors (formic acid and phenolic compounds) was constructed by strengthening the proline biosynthesis. The engineered strain exhibited more effective synthesis ability of proline and scavenging ability of reactive oxygen species (ROS). Consequently, the butanol produced by 824(proABC) was 1-, 2.4- or 3.4-fold higher than that of the wild type strain when using the undetoxified hydrolysate of soybean straw, rice straw or corn straw as the substrate, respectively. Therefore, enhancing the proline biosynthesis can be used as an effective strategy to improve the tolerance of C. acetobutylicum to multiple lignocellulose-derived inhibitors, and 824(proABC) has great potential to produce butanol from undetoxified lignocellulosic hydrolysates. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • An appreciable amount of butanol as an advanced biofuel was produced from OFMSW. • Ethanol organosolv pretreatment was promising for ABE yield improvement. • The presence of inhibitors in OFMSW significantly affected ABE production. • Tannins inhibit the amylolytic activity of Clostridia , affecting ABE production. Abstract Municipal solid waste (MSW) was used as a source for biobutanol production via acetone, butanol, and ethanol (ABE) fermentation. Organosolv pretreatment was used for simultaneous extraction of inhibitors, particularly tannins, and pretreatment of lignocellulosic fraction prior to hydrolysis. The hydrolysates of the pretreated MSW contained appreciable amounts of sugars and soluble starch together with a tolerable amount of inhibitors for Clostridium acetobutylicum. The hydrolysate obtained from MSW pretreated with 85% ethanol at 120 °C for 30 min fermented to the highest ABE concentration of 13.06 g/L with the yield of 0.33 g/g carbon source. Through this process, 102.4 mg butanol, 40.16 mg acetone, and 13.14 mg ethanol were produced from each g of organic fraction of MSW (OFMSW). The pretreatment at mild conditions with higher ethanol concentration accompanied with the lowest glucose yield (0.145 g/g) and the highest starch recovery resulted in the uppermost ABE yield of 0.16 g/g OFMSW. [ABSTRACT FROM AUTHOR]

In this study, a compound nitrogen source, integrating the advantages of ammonium acetate and soybean meal, was proposed to further improve acetone-butanol-ethanol fermentation. Unfortunately, this compound nitrogen source was found to effectively inhibit cellular performance, as the introduction of NH 4 + significantly decreased the yield of butanol and total solvents by 34.78% and 35.14%, to only 6.62 g/L and 10.76 g/L, respectively. Meanwhile, the regulatory mechanism was further elucidated at different levels. As a result, the NH 4 + could down-regulate the transcriptional levels of key genes involved in butanol synthesis, and the activity of acetoacetyl-CoA/acyl-CoA transferase, and then decrease the accumulation of key intermediates. Therefore, ammonium acetate has a dual function in ABE fermentation, as it effectively improves ABE fermentation when it is the sole nitrogen source but significantly decreases fermentation performance in the presence of soybean meal, broadening the understanding of nitrogen regulation mechanism of C. acetobutylicum . [ABSTRACT FROM AUTHOR]

[Display omitted] • Engineered C. thermocellum compatible with CBP for ester production from poplar. • Discovered minor thioesterase activities of esterases limiting ester production. • Improved ester production by manipulating carbon and electron metabolisms. • Severity of poplar CELF pretreatment was positively correlated with ester biosynthesis. Consolidated bioprocessing (CBP) of lignocellulosic biomass uses cellulolytic microorganisms to enable enzyme production, saccharification, and fermentation to produce biofuels, biochemicals, and biomaterials in a single step. However, understanding and redirecting metabolisms of these microorganisms compatible with CBP are limited. Here, a cellulolytic thermophile Clostridium thermocellum was engineered and demonstrated to be compatible with CBP integrated with a Co-solvent Enhanced Lignocellulosic Fractionation (CELF) pretreatment for conversion of hardwood poplar into short-chain esters with industrial use as solvents, flavors, fragrances, and biofuels. The recombinant C. thermocellum engineered with deletion of carbohydrate esterases and stable overexpression of alcohol acetyltransferases improved ester production without compromised deacetylation activities. These esterases were discovered to exhibit promiscuous thioesterase activities and their deletion enhanced ester production by rerouting the electron and carbon metabolism. Ester production was further improved up to 80-fold and ester composition could be modulated by deleting lactate biosynthesis and using poplar with different pretreatment severity. [ABSTRACT FROM AUTHOR]

Hybrid process that integrated fermentation, pervaporation and esterification was established aiming to improve the economic feasibility of the conventional acetone-butanol-ethanol (ABE) fermentation process. Candida sp 99–125 cells were used as full-cell catalyst. The feasibility of batch and fed-batch esterification using the ABE permeate of pervaporation (ranging from 286.9 g/L to 402.9 g/L) as substrate were compared. Valuable butyl oleate was produced along with ethyl oleate. For the batch esterification, due to severe inhibition of substrate to lipase, the yield of butyl oleate and ethyl oleate were only 24.9% and 3.3%, respectively. In contrast, 75% and 11.8% of butyl oleate and ethyl oleate were obtained, respectively, at the end of the fed-batch esterification. The novel integration process provides a promising strategy for in situ upgrading ABE products. [ABSTRACT FROM AUTHOR]

The techniques for enhancing butanol production in ABE fermentation by Clostridium acetobutylicum generally focus on adding electron carrier to strengthen NADH synthesis, repressing hydrogenase by aerating CO, supplementing butyrate, etc. However, those methods suffer from the problems of total solvent decrease, high purification cost, using expensive supplemental substances, etc. In this study, we added small amount of electron receptors (Na 2 SO 4 /CaSO 4 , 2 g/L) into ABE fermentation broth: to alter electron/proton distributions in the intracellular electron transport shuttle system, directing more electron/proton pairs into NADH synthesis route; to stimulate intracellular accumulation of those amino acids favorable for cells survival/butanol synthesis. In ABE fermentation in a 7 L fermentor, adding 2 g/L Na 2 SO 4 could raise butanol concentration to a higher level of 12.96 g/L, which was 34.8% higher than that of the control. Addition of tiny amount cheap electron receptor would provide a new way to enhance bio-butanol production. [ABSTRACT FROM AUTHOR]

Present investigation explores the use of pineapple peel, a food industry waste, for acetone-butanol-ethanol (ABE) production using Clostridium acetobutylicum B 527. Proximate analysis of pineapple peel shows that it contains 35% cellulose, 19% hemicellulose, and 16% lignin on dry basis. Drying experiments on pineapple peel waste were carried out in the temperature range of 60–120 °C and experimental drying data was modeled using moisture diffusion control model to study its effect on ABE production. The production of ABE was further accomplished via acid hydrolysis, detoxification, and fermentation process. Maximum total sugar release obtained by using acid hydrolysis was 97 g/L with 95–97% and 10–50% removal of phenolics and acetic acid, respectively during detoxification process. The maximum ABE titer obtained was 5.23 g/L with 55.6% substrate consumption when samples dried at 120 °C were used as a substrate (after detoxification). [ABSTRACT FROM AUTHOR]

[Display omitted] • Caproate production increased by 170.52% in bioaugmented electro-fermentation system. • Synergy of A. woodii and C. kluyveri was observed in electro-fermentation system. • Colonization of C. kluyveri on the biocathode was promoted by the electric field. • Electron transfer capacity of biocathode was promoted by the electric field. As a chain elongation (CE) model strain, Clostridium kluyveri has been used in the studies of bioaugmentation of caproate production. However, its application in the novel electro-fermentation CE system for bioaugmentation is still unclear. In this study, the CE performances, with or without bioaugmentation and in conventional or electro-fermentation systems were compared. And the mechanism of electrochemical-bioaugmentation by constructing a co-culture of Acetobacterium woodii and Clostridium kluyveri were further verified. Results demonstrated that the bioaugmentation treatments have better CE performance, especially in electro-fermentation system, with a highest caproate concentration of 4.68 g·L-1. Mechanism analysis revealed that C. kluyveri responded to the electric field and emerged synergy with the acetogens, which was proved by the increases of C. kluyveri colonization and the acetogens abundance in biofilm and supported by the co-culture experiment. This study provides a novel insight of microbial synergy mechanism of C. kluyveri during CE bioaugmentation in electro-fermentation system. [ABSTRACT FROM AUTHOR]

• High hexanol production from CO gas was achieved with Clostridium carboxidivorans. • Oleyl alcohol (OA) was selected as the solvent for extractive fermentation. • The highest hexanol titer (5.06 g/L) was achieved with repeated OA addition. • Hexanol production was further enhanced (8.45 g/L) with repeated ethanol addition. This study achieved high production of hexanol via gas fermentation using Clostridium carboxidivorans P7 by extracting hexanol from the fermentation broth. The hexanol extraction efficiency and inhibitory effects on C. carboxidivorans P7 of 2-butyl-1-octanol, hexyl hexanoate and oleyl alcohol were examined, and oleyl alcohol was selected as the extraction solvent. Oleyl alcohol was added at the beginning of fermentation and during fermentation or a small volume of oleyl alcohol was repeatedly added during fermentation. The addition of a small volume of oleyl alcohol during fermentation was the most effective for CO consumption and hexanol production (5.06 g/L), yielding the highest known hexanol titer through any type of fermentation including gas fermentation. Hexanol production was further enhanced to 8.45 g/L with the repeated addition of oleyl alcohol and ethanol during gas fermentation. The results of this study will enable sustainable and carbon–neutral hexanol production via gas fermentation. [ABSTRACT FROM AUTHOR]

To enhance the butanol productivity and reduce the material cost, acetone, butanol, and ethanol fermentation by Clostridium acetobutylicum SE25 was investigated using batch, repeated-batch and continuous cultures in a fibrous bed bioreactor, where cassava flour was used as the substrate. With periodical nutrient supplementation, stable butanol production was maintained for about 360 h in a 6-cycle repeated-batch fermentation with an average butanol productivity of 0.28 g/L/h and butanol yield of 0.32 g/g-starch. In addition, the highest butanol productivity of 0.63 g/L / h and butanol yield of 0.36 g/g-starch were achieved when the dilution rate were investigated in continuous production of acetone, butanol, and ethanol using a fibrous bed bioreactor, which were 231.6% and 28.6% higher than those of the free-cell fermentation. On the other hand, this study also successfully comfirmed that the biofilm can provide an effective protection for the microbial cells which are growing in stressful environment. [ABSTRACT FROM AUTHOR]

Dark fermentation systems often show low H 2 yields and unstable H 2 production, as the result of the variability of microbial dynamics and metabolic pathways. Recent batch investigations have demonstrated that an artificial consortium of two anaerobic bacteria, Clostridium acetobutylicum and Desulfovibrio vulgaris Hildenborough, may redirect metabolic fluxes and improve H 2 yields. This study aimed at evaluating the scale-up from batch to continuous H 2 production in an up-flow anaerobic packed-bed reactor (APBR) continuously fed with a glucose-medium. The effects of various parameters, including void hydraulic retention time (HRTv), pH, and alkalinity, on H 2 production performances and metabolic pathways were investigated. The results demonstrated that a stable H 2 production was reached after 3–4 days of operation. H 2 production rates increased significantly with decreasing HRTv from 4 to 2 h. Instead, H 2 yields remained almost stable despite the change in HRTv, indicating that the decrease in HRTv did not affect the global metabolism. [ABSTRACT FROM AUTHOR]

In this study, different pretreatment methods were evaluated for modified the corn stalk bagasse and further used the pretreated bagasse as immobilized carrier in acetone-butanol-ethanol fermentation process. Structural changes of the bagasses pretreated by different methods were analyzed by Fourier transform infrared, crystallinity index and scanning pictures by electron microscope. And the performances of batch fermentation using the corn stalk based carriers were evaluated. Results indicated that the highest ABE concentration of 23.86 g/L was achieved using NaOH pretreated carrier in batch fermentation. Immobilized fermentation-pervaporation integration process was further carried out. The integration process showed long-term stability with 225–394 g/L of ABE solvents on the permeate side of pervaporation membrane. This novel integration process was found to be an efficient method for biobutanol production. [ABSTRACT FROM AUTHOR]

The metabolism of butanol producing bacteria Clostridium acetobutylicum was studied in chemostat with glucose limited conditions, butanol stimulus, and as a reference cultivation. COnstraint-Based Reconstruction and Analysis (COBRA) was applied using additional constraints from 13 C Metabolic Flux Analysis ( 13 C-MFA) and experimental measurement results. A model consisting of 451 metabolites and 604 reactions was utilized in flux balance analysis (FBA). The stringency of the flux spaces considering different optimization objectives, i.e. growth rate maximization, ATP maintenance, and NADH/NADPH formation, for flux variance analysis (FVA) was studied in the different modelled conditions. Also a previously uncharacterized exopolysaccharide (EPS) produced by C. acetobutylicum was characterized on monosaccharide level. The major monosaccharide components of the EPS were 40 n-% rhamnose, 34 n-% glucose, 13 n-% mannose, 10 n-% galactose, and 2 n-% arabinose. The EPS was studied to have butanol adsorbing property, 70 (butanol) mg (EPS) g −1 at 37 °C. [ABSTRACT FROM AUTHOR]

In this study, metabolic target reactions for strain engineering were searched via intracellular coenzyme A (CoA) metabolite analysis. The metabolic reactions catalyzed by thiolase (AtoB) and aldehyde-alcohol dehydrogenase (AdhE1) were considered potential rate-limiting steps. In addition, CoA transferase (CtfAB) was highlighted as being important for the assimilation of organic acids, in order to achieve high butanol production. Based on this quantitative analysis, the BEKW_E1AB-atoB strain was constructed by overexpressing the thl ( atoB ), adhE1, and ctfAB genes in Clostridium acetobutylicum strain BEKW, which has the phosphotransacetylase ( pta ) and butyrate kinase ( buk ) genes knocked out. After 100 h of continuous fermentation coupled with adsorptive ex situ butanol recovery, the concentrations found after considering desorption, yield, and productivity for the BEKW_E1AB-atoB strain were 55.7 g/L, 0.38 g/g, and 2.64 g/L/h, respectively. The level of butanol production achieved (2.64 g/L/h) represents the highest reported value obtained after adsorptive, long-term fermentation. [ABSTRACT FROM AUTHOR]

[Display omitted] • Heat-treated sludge with Clostridium butyricum produced H 2 at 58.57 ± 5.03 L/L-d. • Sludge-originated H 2 -producers rather than C. butyricum dominated the grown culture. • C. butyricum prevented the growth of Streptococcus , Lactobacillus and Megasphaera. • Initial abundance of Clostridium over 71.3% is recommended for a continuous culture. This study aimed to mitigate the instability in the start-up and continuous performance of dark fermentative biohydrogen production using heat-treated sludge by the addition of an exogenous H 2 -producing strain. Continuous fermentation augmented with Clostridium butyricum showed the highest average biohydrogen production rate (HPR) as 50.35 ± 2.56 and 58.57 ± 5.03 L/L-d with H 2 -producing butyric and acetic acid pathways, whereas the fermenters without bioaugmentation showed the termination of biohydrogen production in 3 days of continuous operation with non H 2 -producing lactic acid pathway and H 2 -consuming propionic acid pathway. The bioaugmentation blocked the growth of the competitors for hexose such as Streptococcus , Lactobacillus and Megasphaera , and provided H 2 -producer dominated microbiome with not only Clostridium butyricum , but also Clostridium puniceum and Clostridium neuense originated from heat-treated sludge. Bioaugmentation of a H 2 -producing strain would be a reliable dissemination strategy for dark fermentative biohydrogen production by minimizing the influence of seed sludge population. [ABSTRACT FROM AUTHOR]

Butanol is an important industrial chemical and an attractive transportation fuel. However, the deficiency of reducing equivalents NAD(P)H in butanol fermentation results in a large quantity of oxidation products, which is a major problem limiting the atom economy and economic viability of bio-butanol processes. Here, we integrated the butanol fermentation process with a NADH-generating, acetoin biosynthesis process to improve the butanol production. By overexpressing the α-acetolactate decarboxylase gene alsD from Bacillus subtilis in Clostridium acetobutylicum , acetoin yield was significantly increased at the cost of acetone. After optimization of fermentation conditions, butanol (12.9 g/L), acetoin (6.5 g/L), and ethanol (1.9 g/L) were generated by the recombinant strain, with acetone no more than 1.8 g/L. Thus, both mass yield and product value were greatly improved. This study demonstrates that reducing power compensation is effective to improve the atom economy of butanol fermentation, and provides a novel approach to improve the economic viability of bio-butanol production. [ABSTRACT FROM AUTHOR]

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 60 h, 14.17 and 21.11 g L −1 of butanol and acetone–butanol–ethanol (ABE) were produced from 68.89 g L −1 of total sugars, respectively, yielding 0.22 and 0.33 g g −1 of sugars. The maximum yield of butanol and ABE reached 15.4 g and 22.9 g per 100 g raw SB, respectively. This established process may have potential application for butanol production from SB. [ABSTRACT FROM AUTHOR]

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 24 g/L of ABE solvents was obtained from 59.6 g/L of initial sugar after 80 h of fermentation with gas stripping. Then, long-term of fed-batch fermentation with continuous gas stripping was further performed. 112.9 g/L of butanol, 44.1 g/L of acetone, 9.5 g/L of ethanol (total 166.5 g/L of ABE) was produced in overall 312 h of fermentation. At the same time, concentrated ABE product was obtained in the condensate of gas stripping. [ABSTRACT FROM AUTHOR]

A prominent delay with 12 h was encountered in the phase shift from acidogenesis to solventogenesis in butanol production when the substrate—glucose was replaced by cassava flour. To solve this problem, different phase of pH regulation strategies were performed to shorten this delay time. With this effort, the phase shift occurred smoothly and the fermentation time was shortened. Under the optimal conditions, 16.24 g/L butanol and 72 h fermentation time were achieved, which were 25.3% higher and 14.3% shorter than those in the case of without pH regulation. Additionally, the effect of CaCO 3 on “acid crash” and butanol production was also investigated. It was found that organic acids reassimilation would be of benefit to enhance butanol production. These results indicated that the simple but effective approach for acceleration of phase shift is a promising technique for shortening the fermentation time and improvement of butanol production. [ABSTRACT FROM AUTHOR]

Paper mill sludge (PS), a solid waste from pulp and paper industry, was investigated as a feedstock for acetone–butanol–ethanol (ABE) production by simultaneous saccharification and fermentation (SSF). ABE fermentation of paper sludge by Clostridium acetobutylicum required partial removal of ash in PS to enhance its enzymatic digestibility. Enzymatic hydrolysis was found to be a rate-limiting step in the SSF. A total of 16.4–18.0 g/L of ABE solvents were produced in the SSF of de-ashed PS with solid loading of 6.3–7.4% and enzyme loading of 10–15 FPU/g-glucan, and the final solvent yield reached 0.27 g/g sugars. No pretreatment and pH control were needed in ABE fermentation of paper sludge, which makes it an attractive feedstock for butanol production. The results suggested utilization of paper sludge should not only consider the benefits of buffering effect of CaCO 3 in fermentation, but also take into account its inhibitory effect on enzymatic hydrolysis. [ABSTRACT FROM AUTHOR]

Acetone is the major by-product in ABE fermentations, most researches focused on increasing butanol/acetone ratio by decreasing acetone biosynthesis. However, economics of ABE fermentation industry strongly relies on evaluating acetone as a valuable platform chemical. Therefore, a novel ABE fermentation strategy focusing on bio-acetone production by co-culturing Clostridium acetobutylicum / Saccharomyces cerevisiae with exogenous acetate addition was proposed. Experimental and theoretical analysis revealed the strategy could, enhance C. acetobutylicum survival oriented amino acids assimilation in the cells; control NADH regeneration rate at moderately lower level to enhance acetone synthesis but without sacrificing butanol production; enhance the utilization ability of C. acetobutylicum on glucose and direct most of extra consumed glucose into acetone/butanol synthesis routes. By implementing the strategy using synthetic or acetate fermentative supernatant, acetone concentrations increased to 8.27–8.55 g/L from 5.86 g/L of the control, while butanol concentrations also elevated to the higher levels of 13.91–14.23 g/L from 11.63 g/L simultaneously. [ABSTRACT FROM AUTHOR]

This study aimed to quantitatively evaluate the inhibitor tolerance of butanol-producing bacterium Clostridium acetobutylicum . The inhibitory effect of the inhibitors generated by acid pretreatment of biomass feedstock on butanol fermentation decreased in the order of formic acid > oxalic acid > furfural > 5-HMF > Na 2 SO 4 . C. acetobutylicum has a small tolerance range for furfural (1.06–2.6 g/L) and 5-HMF (1.99–2.3 g/L). However, the inhibitory effect of Na 2 SO 4 appears to have a wide range, with a chronic toxicity for C. acetobutylicum . All the results could explain, in quantitative manner, the instability of butanol fermentation with C. acetobutylicum from various acid-pretreated feedstocks caused by the fermentation inhibitors. [ABSTRACT FROM AUTHOR]

Sodium sulfide (SS) was added to the non-detoxified wheat straw hydrolysate for ABE fermentation by Clostridium acetobutylicum CICC8012. Biochemical measurements demonstrated that supplementation with SS promoted earlier and enhanced conversion of acid to ABE and led to a 27.48% improvement in sugar consumption, a 20.48% improvement in the sugar-based ABE yield, a 47.63% improvement in the butanol titer, and a 53.50% improvement in the ABE concentration. The response of C. acetobutylicum CICC8012 at the mRNA level was examined by a transcriptional analysis performed with RNA sequencing. The expression of genes involved in the membrane transport of carbohydrates, glycolysis, and ABE formation increased following SS-supplemented fermentation, whereas the expression of genes encoding enzymes involved in acid formation decreased, which indicates that supplemental SS affected the central fermentative pathway, down-regulated the metabolic flux toward the acid formation branches, and up-regulated the metabolic flux toward the ABE formation branches. [ABSTRACT FROM AUTHOR]

The role of pH control on biohydrogen production by co-culture of dark-fermentative Clostridium acetobutylicum and photofermentative Rhodobacter sphaeroides was studied. Single stage dark fermentation, photofermentation and hybrid co-culture systems were studied at different values of controlled and uncontrolled pH. Increasing pH during dark fermentation resulted in lower hydrogen production rate (HPR) and longer lag time for both controlled and uncontrolled conditions. However, it only slightly affected cumulative H 2 volume. Results have shown that pH control at pH 7.5 increased photofermentative hydrogen production from 0.966 to 2.502 L H 2 /L medium when compared to uncontrolled process. Fixed pH value has proven to be an important control strategy also for the hybrid process and resulted in obtaining balanced co-culture of dark and photofermentative bacteria. Control of pH at 7.0 was found optimum for bacteria cooperation in the co-culture what resulted in obtaining 2.533 L H 2 /L medium and H 2 yield of 6.22 mol H 2 /mol glucose. [ABSTRACT FROM AUTHOR]

[Display omitted] • Magnetite supplementation increased hydrogen production by 25.6%. • Cosupplementation of lactate and magnetite increased hydrogen production by 58.9%. • Enhanced hydrogen production was caused by acceleration of lactate utilization. • Magnetite upregulates the expression of key genes for hydrogen production. • Magnetite promotes electron-bifurcation by activating the etfAB complex. A conductive metal compound can be used as a catalyst for enhancing hydrogen production by dark fermentation. This study aimed to identify mechanisms of enhanced hydrogen production by magnetite supplementation. Experiments were performed with lactate and/or magnetite supplementation to confirm that the lactate-utilizing pathway is the key cause of enhanced hydrogen production. Also, ribonucleic acid sample was collected for monitoring gene regulation under each condition. Hydrogen production was significantly enhanced by approximately 25.6% and 58.9%, respectively, via magnetite alone and with lactate. Moreover, the expression of genes involved in hydrogen production, including pyruvate ferredoxin oxidoreductase, hydrogenase, and ferredoxin, via magnetite alone and with lactate was upregulated by 0.26, 0.71, and 3.50 and 1.06, 2.14, and 1.94 times, respectively. [ABSTRACT FROM AUTHOR]

[Display omitted] • Effect of various inoculum pretreatment on microbial community dynamics evaluated. • H 2 yield was 70% higher for acid and alkali-treated inoculum than untreated. • The combined impact of heat and CO 2 enriched > 90% spore-forming fermentative bacteria. • Clostridial-type fermentation dominated mesophilic H 2 production from food waste. This study systematically evaluated and compared different inoculum pretreatment methods to quickly select dark fermentative bacteria from anaerobic sludge for the bioconversion of food waste. The hydrogen (H 2) production rate was found to be highest for 'heat + CO 2 ′ treated inoculum at 140.75 ± 2.61 mL/L/h compared to control experiments (60.27 ± 2.61 mL/L/h). At the same time, H 2 yield was found to be highest for alkali-treated inoculum at 157.25 ± 7.62 mL/g of volatile solids (VS) added compared to control experiments (91.61 ± 1.93 mL/g VS). Analysis of organic acids suggests a Clostridial-type fermentation with acetate (0.52 to 1.60 g/L) and butyrate (1.69 to 2.42 g/L) being the major by-products. The microbial data analysis showed that Firmicutes (63.64–90.39%), Bacteroidota (1.16–21.88%), and Proteobacteria (2.09–9.93%) were dominant at the phylum level, whereas genus-level classification showed Clostridium sensu stricto 1 (6.37–42.63%), Streptococcus (1.87–28.96%), Prevotella (0.57–16.59%), and Enterococcus (0.56–14.51%) dominated under different experimental conditions. [ABSTRACT FROM AUTHOR]

[Display omitted] • Xylose pathway genes in C. tyrobutyricum were identified and verified. • The potential mechanisms of CCR in C. tyrobutyricum were proposed and confirmed. • Double knockout of hprK and xylR eliminated CCR in C. tyrobutyricum. • Undetoxified lignocellulosic hydrolysates were used as the substrates. Clostridium tyrobutyricum , a gram-positive anaerobic bacterium, is recognized as the promising butyric acid producer. But, the existence of carbon catabolite repression (CCR) is the major drawback for C. tyrobutyricum to efficiently use the lignocellulosic biomass. In this study, the xylose pathway genes were first identified and verified. Then, the potential regulatory mechanisms of CCR in C. tyrobutyricum were proposed and the predicted engineering targets were experimental validated. Inactivation of hprK blocked the CcpA-mediated CCR and resulted in simultaneous conversion of glucose and xylose, although xylose consumption was severe lagging behind. Deletion of xylR further shortened the lag phase of xylose utilization. When hprK and xylR were inactivated together, the CCR in C. tyrobutyricum was completely eliminated. Consequently, ATCC 25755/Δ hprK Δ xylR showed significant increase in butyrate productivity (1.8 times faster than the control) and excellent butyric acid fermentation performance using both mixed sugars (11.0–11.9 g/L) and undetoxified lignocellulosic hydrolysates (12.4–13.4 g/L). [ABSTRACT FROM AUTHOR]

This work deals with the specific butanol production rate of Clostridium acetobutylicum using xylose – a relevant fraction of lignocellulosic feedstock for biofuel production – as carbon source. The tests were carried out in a CSTR equipped with a microfiltration unit. The dilution rate ( D ) ranged between 0.02 and 0.22 h −1 and the ratio R between the permeate stream rate and the stream fed to the reactor ranged between 14% and 88%. The biomass present in the broth was identified as a heterogeneous cell population consisting of: acidogenic cells, solventogenic cells and spores. The results were processed to assess the concentration of acidogenic cells, solventogenic cells and spores. The specific butanol production rate was also assessed. The max butanol productivity was 1.3 g L −1 h −1 at D = 0.17 h −1 and R = 30%. A comparison between the results reported in a previous work carried out with lactose was made. [ABSTRACT FROM AUTHOR]

This study conducted batch experiments to evaluate the potential of butanol production from microalgae biodiesel residues by Clostridium acetobutylicum . The results indicated that with 90 g/L of glucose as the sole substrate the highest butanol yield of 0.2 g/g-glucose was found, but the addition of butyrate significantly enhanced the butanol yield. The highest butanol yield of 0.4 g/g-glucose was found with 60 g/L of glucose and 18 g/L of butyrate. Using microalgae biodiesel residues as substrate, C. acetobutylicum produced 3.86 g/L of butanol and achieved butanol yield of 0.13 g/g-carbohydrate via ABE fermentation, but the results indicated that approximately one third of carbohydrate was not utilized by C. acetobutylicum . Biological butanol production from microalgae biodiesel residues can be possible, but further research on fermentation strategies are required to improve production yield. [ABSTRACT FROM AUTHOR]

This work reports the results of a series of tests on the specific butanol production rate by Clostridium acetobutylicum continuous cultures. The tests were carried out using lactose as carbon source to mimic cheese-whey. A continuous stirred tank reactor equipped with a microfiltration unit was used. The dilution rate ( D ) ranged between 0.02 and 0.15 h −1 and the ratio R of the permeate stream rate to the stream fed to the reactor ranged between 14% and 95%. For each set of D and R values, the continuous cultures were characterized in terms of concentration of cells, acids and solvents. Results were processed to assess the concentration of acidogenic cells, solventogenic cells, spores and the specific butanol production rate. The max butanol productivity was 0.5 g L −1 h −1 at D = 0.1 h −1 and R = 95%. The butanol productivity referred to solventogenic cells was expressed as a function of concentration of lactose, acids and butanol. [ABSTRACT FROM AUTHOR]

Acetone–butanol–ethanol (ABE) fermentation was studied using acid-hydrolyzed xylan recovered from hardwood Kraft black liquor by CO 2 acidification as the only carbon source. Detoxification of hydrolyzate using activated carbon was conducted to evaluate the impact of inhibitor removal and fermentation. Xylose hydrolysis yields as high as 18.4% were demonstrated at the highest severity hydrolysis condition. Detoxification using active carbon was effective for removal of both phenolics (76–81%) and HMF (38–52%). Batch fermentation of the hydrolyzate and semi-defined P2 media resulted in a total solvent yield of 0.12–0.13 g/g and 0.34 g/g, corresponding to a butanol concentration of 1.8–2.1 g/L and 7.3 g/L respectively. This work is the first study of a process for the production of a biologically-derived biofuel from hemicelluloses solubilized during Kraft pulping and demonstrates the feasibility of utilizing xylan recovered directly from industrial Kraft pulping liquors as a feedstock for biological production of biofuels such as butanol. [ABSTRACT FROM AUTHOR]

To obtain native strains resistant to butanol toxicity, a new isolating method and serial enrichment was used in this study. With this effort, mutant strain SE36 was obtained, which could withstand 35 g/L (compared to 20 g/L of the wild-type strain) butanol challenge. Based on 16s rDNA comparison, the mutant strain was identified as Clostridium acetobutylicum . Under the optimized condition, the phase shift was smoothly triggered and fermentation performances were consequently enhanced. The maximum total solvent and butanol concentration were 23.6% and 24.3%, respectively higher than that of the wild-type strain. Furthermore, the correlation between butanol produced and the butanol tolerance was investigated, suggesting that enhancing butanol tolerance could improve butanol production. These results indicate that the simple but effective isolation method and acclimatization process are a promising technique for isolation and improvement of butanol tolerance and production. [ABSTRACT FROM AUTHOR]

Production of acetone–butanol–ethanol (ABE) from cassava was investigated with a fermentation–pervaporation (PV) coupled process. ABE products were in situ removed from fermentation broth to alleviate the toxicity of solvent to the Clostridium acetobutylicum DP217. Compared to the batch fermentation without PV, glucose consumption rate and solvent productivity increased by 15% and 21%, respectively, in batch fermentation–PV coupled process, while in continuous fermentation–PV coupled process running for 304 h, the substrate consumption rate, solvent productivity and yield increased by 58%, 81% and 15%, reaching 2.02 g/L h, 0.76 g/L h and 0.38 g/g, respectively. Silicalite-1 filled polydimethylsiloxane (PDMS)/polyacrylonitrile (PAN) membrane modules ensured media recycle without significant fouling, steadily generating a highly concentrated ABE solution containing 201.8 g/L ABE with 122.4 g/L butanol. After phase separation, a final product containing 574.3 g/L ABE with 501.1 g/L butanol was obtained. Therefore, the fermentation–PV coupled process has the potential to decrease the cost in ABE production. [ABSTRACT FROM AUTHOR]

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 27.8 h (free-cell fermentation) to 3.3 h (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. [ABSTRACT FROM AUTHOR]

Highlights: [•] Gas stripping model of ABE from model solutions was characterized. [•] Integrated fermentation system with gas stripping for in situ butanol recovery. [•] Fed-batch fermentation with solvent removal at 70°C has been demonstrated. [•] Permeating–heating–gas stripping greatly enhances butanol productivity and yield. [ABSTRACT FROM AUTHOR]

Highlights: [•] The growth of Clostridium acetobutylicum on xylose was characterized. [•] Tests were carried out in a CSTR operated under controlled pH. [•] The growth kinetic was expressed by means of a multiple product inhibition. [•] Biomass and product yields were expressed as a function of specific growth rate. [Copyright &y& Elsevier]

Highlights: [•] Butanol production from sugarcane juice by C. acetobutylicum JB200 was studied. [•] Cells were immobilized in a fibrous bed bioreactor for ABE fermentation. [•] ABE production reached 28g/L, the highest ever achieved in fermentation with sucrose. [•] This hyper butanol-producing strain was stable for 16 consecutive repeated batches. [•] This process can be used to economically produce butanol in a sugarcane biorefinery. [ABSTRACT FROM AUTHOR]

Highlights: [•] Sweet sorghum cuticular waxes (SSCW) were critical inhibitors of ABE fermentation. [•] SSCW militate against both acidogenesis and solventogenesis phase of ABE fermentation. [•] Very long-chain fatty acids, fatty alcohols and aldehydes were the main composition of SSCW. [•] More severe inhibition was got with the chain length of fatty acid decreased. [ABSTRACT FROM AUTHOR]