Butanol is an aliphatic saturated alcohol having the molecular formula of C4H9OH Butanol can be used as an intermediate in chemical synthesis and as a solvent for a wide variety of chemical and textile industry applications Moreover, butanol has been considered as a potential fuel or fuel additive Biological production of butanol (with acetone and ethanol) was one of the largest industrial fermentation processes early in the 20th century However, fermentative production of butanol had lost its competitiveness by 1960s due to increasing substrate costs and the advent of more efficient petrochemical processes Recently, increasing demand for the use of renewable resources as feedstock for the production of chemicals combined with advances in biotechnology through omics, systems biology, metabolic engineering and innovative process developments is generating a renewed interest in fermentative butanol production This article reviews biotechnological production of butanol by clostridia and some relevant fermentation and downstream processes The strategies for strain improvement by metabolic engineering and further requirements to make fermentative butanol production a successful industrial process are also discussed Biotechnol Bioeng 2008;101: 209-228 © 2008 Wiley Periodicals, Inc

https://onlinelibrary.wiley.com/doi/pdf/10.1002/bit.22003

Fermentative butanol production by clostridia

Butanol is a very competitive renewable biofuel for use in internal combustion engines given its many advantages. In this review, the properties of butanol are compared with the conventional gasoline, diesel fuel, and some widely used biofuels, i.e. methanol, ethanol, biodiesel. The comparison of fuel properties indicates that n-butanol has the potential to overcome the drawbacks brought by low-carbon alcohols or biodiesel. Then, the development of butanol production is reviewed and various methods for increasing fermentative butanol production are introduced in detailed, i.e. metabolic engineering of the Clostridia, advanced fermentation technique. The most costive part of the fermentation is the substrate, so methods involved in renewed substrates are also mentioned. Next, the applications of butanol as a biofuel are summarized from three aspects: (1) fundamental combustion experiments in some well-defined burning reactors; (2) a substitute for gasoline in spark ignition engine; (3) a substitute for diesel fuel in compression ignition engine. These studies demonstrate that butanol, as a potential second generation biofuel, is a better alternative for the gasoline or diesel fuel, from the viewpoints of combustion characteristics, engine performance, and exhaust emissions. However, butanol has not been intensively studied when compared to ethanol or biodiesel, for which considerable numbers of reports are available. Finally, some challenges and future research directions are outlined in the last section of this review.

Progress in the production and application of n-butanol as a biofuel

1-Butanol, an important chemical feedstock and advanced biofuel, is produced by Clostridium species. Various efforts have been made to transfer the clostridial 1-butanol pathway into other microorganisms. However, in contrast to similar compounds, only limited titers of 1-butanol were attained. In this work, we constructed a modified clostridial 1-butanol pathway in Escherichia coli to provide an irreversible reaction catalyzed by trans-enoyl-coenzyme A (CoA) reductase (Ter) and created NADH and acetyl-CoA driving forces to direct the flux. We achieved high-titer (30 g/liter) and high-yield (70 to 88% of the theoretical) production of 1-butanol anaerobically, comparable to or exceeding the levels demonstrated by native producers. Without the NADH and acetyl-CoA driving forces, the Ter reaction alone only achieved about 1/10 the level of production. The engineered host platform also enables the selection of essential enzymes with better catalytic efficiency or expression by anaerobic growth rescue. These results demonstrate the importance of driving forces in the efficient production of nonnative products.

https://www.phys.sinica.edu.tw/TIGP-NANO/Course/2013_Fall/classnote/Seminar_172%20Shen%20AEM%202011.pdf

Driving Forces Enable High-Titer Anaerobic 1-Butanol Synthesis in Escherichia coli

A comparative view of metabolite and substrate stress and tolerance in microbial bioprocessing: From biofuels and chemicals, to biocatalysis and bioremediation

Developments in biobutanol production: New insights

The present invention relates to a method of producing high levels of butanol using a fermentation process that employs a mutant strain of Clostridium beijerinckii.

A method of producing butanol using a mutant strain of clostridium beijerinckii

The genus Clostridium contains widely diverse species which have been shown to be associated with foodborne illness as well as solvent production (Blaschek, 1999). C. perfringens is representative of a clostridial species which is associated with food poisoning, while non-toxinogenic C. beijerinckii has important industrial and biotechnological potential because of its ability to produce acetone and butanol during fermentation of carbohydrates. The development of genetic systems for the clostridia was recently reviewed (Blaschek and White, 1995). Electroporation induced transformation has been shown to have considerable potential for introducing DNA into a wide variety of gram+ and gram− bacterial species (Dower et al., 1992) and electroporation protocols are available for previously non-transformable clostridial strains (Chen et al., 1996). However, the transformation efficiencies appear to vary greatly among different species and strains. It has been suggested that DNase and restriction enzyme activity are common barriers to successful transformation of the clostridia. The cell wall may also represent a barrier to DNA uptake in C. perfringens (Kim and Blaschek, 1989). As we learn more about mechanisms and factors involved in transforming various bacteria, protocols for overcoming these obstacles will likely emerge. Electroporation protocols have been developed for the clostridia which are based on using polyethylene glycol (PEG) in the shocking menstruum. During electroporation-induced transformation, PEG may limit cytoplasmic leakage and, via exclusion of the aqueous phase, help to push the plasmid DNA into the cell and consequently increase cell viability and transformation efficiency.

Clostridium in Biotechnology and Food Technology

Verfahren zur herstellung von butanol unter verwendung eines mutierten stammes von clostridium beijerinckii

Dramatically elevated levels of butanol and acetone resulted in higher butanol and total solvent yields for hyperamylolytic Clostridium beijerinckii BA101 relative to the NCIMB 8052 parent strain grown in semidefined P2 medium containing either 6% glucose or STAR-DRI 5 maltodextrin. C. beijerinckii BA101 consistently produced on the order of 19 g of butanol per liter in 20-liter batch fermentations. This represents a greater than 100% increase in butanol concentration by the BA101 strain compared to the parent NCIMB 8052 strain. The kinetics of butanol production over time also indicate a more rapid rate of butanol production by BA101 in semidefined P2 medium containing glucose or maltodextrin. The lower levels of butyric and acetic acids produced over the course of the fermentation carried out by BA101 are consistent with an enhanced capacity for uptake and recycling of these acids. C. beijerinckii BA101 appears to more completely utilize carbohydrate compared to the 8052 strain. Carbon balance following fermentation by C. beijerinckii 8052 and BA101 indicates that sufficient carbon is available for the twofold increase in butanol concentration observed during BA101 fermentations. C. beijerinckii BA101 also has superior solvent production capacity during continuous culture fermentation in P2 medium containing 6% glucose. Volumetric solvent yields of 0.78 and 1.74 g/liter/h for BA101 and 0.34 and 1.17 g/liter/h for NCIMB 8052 were obtained at dilution rates of 0.05 and 0.20 h(sup-1), respectively. No drift towards acid synthesis (strain degeneration) was observed for up to 200 h (d = 0.05 h(sup-1)) and 100 h (d = 0.20 h(sup-1)).

Enhanced Butanol Production by Clostridium beijerinckii BA101 Grown in Semidefined P2 Medium Containing 6 Percent Maltodextrin or Glucose.

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A method of producing butanol using a mutant strain of clostridium beijerinckii

Clostridium in Biotechnology and Food Technology

Verfahren zur herstellung von butanol unter verwendung eines mutierten stammes von clostridium beijerinckii

Enhanced Butanol Production by Clostridium beijerinckii BA101 Grown in Semidefined P2 Medium Containing 6 Percent Maltodextrin or Glucose.