A re-look at the biochemical strategies to enhance butanol production

Butanol produced from renewable feedstock is defined as an emerging biofuel and biochemical. Research efforts made during the last three decades on biochemical production of butanol via conventional ABE (acetone-ethanol-butanol) fermentation has tried to bring biobutanol close to competition with petrobutanol. However, each new effort of development has been often countered by new challenges, confining biobutanol production mostly to the laboratory scale. This review provides a systematic, comparative analysis of different steps in biochemical production of butanol and identifies the counteractive aspects and challenges to overcome. A special emphasis is given on process inhibitors, applied detoxification techniques, chemical supplements and research & development in industry in order to enhance and update ABE fermentation and make it cost effective. Biobutanol future lies in utilization of inexpensive cellulose enriched lignocellulosic hydrolysates and hyper-butanol producing bacteria, combined with specific detoxification techniques and followed by efficient continuous fermentation technologies together with in situ product recovery.     

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.     

Optimization for simultaneous enhancement of biobutanol and biohydrogen production

Acetone-butanol-ethanol (ABE) fermentation guarantees a sustainable route for biohydrogen and biobutanol production. This research work is committed towards the enhancement of biohydrogen and biobutanol production by single and multi-parameter optimization for the improvement of substrate energy recovery using C. saccharoperbutylacetonicum. Single parameters optimization (SPO) manifested that headspace of 60% (v/v) and butyric acid supplementation of 9 g/L and temperatures of 30 °C and 37 °C were suitable for obtaining maximum biohydrogen and biobutanol production, respectively. The interaction between these parameters was further evaluated by implementing a 5-level 3-factor Central Composite Design (CCD). In the present study, a central composite design was employed to enhance the biohydrogen and biobutanol production. In addition, the experimental results were analyzed by response surface methodology (RSM) and artificial intelligence (AI) techniques such as artificial neural network (ANN). The prediction capability of RSM was further compared with ANN for predicting the optimum parameters that would lead to maximum biohydrogen and biobutanol production. ANN yielded higher values of biohydrogen and biobutanol. ANN was found to be superior as compared to RSM in terms of prediction accuracy for both biohydrogen and biobutanol because of its higher coefficient of determination (R²) and lower root mean square error (RMSE) value. Process temperature (32.65 °C), headspace (58.21% (v/v)) and butyric acid supplementation (9.16 g/L) led to maximum substrate energy recovery of 78% with biohydrogen and biobutanol production of 5.9 L/L and 16.75 g/L, respectively. Process parameter optimization led to a significant increase in substrate energy recovery from Biphasic fermentation.     

Feasibility of reed for biobutanol production hydrolyzed by crude cellulase

The aim of this study was to efficiently utilize reed for both cellulase and biobutanol production. The unprocessed cellulase blend produced under solid-state fermentation using reed as the substrate showed a similar reducing sugar yield using Whatman filter paper to the commercial enzyme blend (38.61%). Organosolv pretreatment method could efficiently reduce hemicellulose (29.3%–14.6%) and lignin (17.2%–14.1%) content and increase cellulose content (42.5%–62.3%) from reed. Enzymatic hydrolysis of organosolv-pretreated reed using the crude cellulase with enzyme loading of 25 FPU/g reed, 20% solid content at 50 °C and pH 5.5 resulted in a reed hydrolysate containing 40.01 g/L glucose and 3.55 g/L xylose after 72 h. Fermentation of the hydrolysate medium by Clostridium acetobutylicum produced 9.07 and 14.24 g/L of biobutanol and ABE with yield of 0.21 g/g and 0.33 g/g, respectively. This study proved that crude cellulase complex produced under solid state fermentation and organsolv pretreatment can efficiently provide reed hydrolysate that can be converted to biobutanol without any commercial cellulase usage.     

Recovery of butanol from fermentation broth by gas stripping

This is an overview of the butanol (usually called acetone butanol ethanol, ABE) fermentation in various types of reactor systems and recovery by gas stripping. Gas stripping is a simple technique which does not require expensive apparatus, does not harm the culture, does not remove nutrients and reaction intermediates and reduces butanol toxicity (inhibition). As a result of butanol removal by gas stripping, concentrated sugar solutions can be used to produce butanol/ABE. Compared to sugar utilization of 30 gl⁻¹ in a control batch reactor, sugar utilization of 199 gl⁻¹ has been reported with 69.7 gl⁻¹ solvent production. In fed-batch reactors concentrated sugar solutions (350 gl⁻¹) have been used. Additionally, the process of ABE production results in concentrated product streams containing 9.1–120 gl⁻¹ butanol/ABE. In the integrated ABE production and recovery systems, selectivities of 4–30.5 have been reported.     

Developments in biobutanol production: New insights

Biobutanol will become an attractive, economic and sustainable fuel as petroleum oil leads towards expensive fuel due to diminishing oil reserves and an increase of green house gases in the atmosphere. The major challenges in biobutanol production are low butanol titer, availability of compatible feedstocks, and product inhibition. These hurdles are being resolved using several genetic engineering techniques, metabolic engineering strategies, and promising integrated continuous fermentation processes with efficient product recovery techniques (like gas stripping). Adequate success in utilizing renewable and cost-effective cellulosic materials as feedstocks has opened up novel grounds for the advancement in economic biobutanol production. In this direction, Clostridium beijerinckii is being explored as promising strain to produce biobutanol from cellulosic materials. Moreover, high biobutanol titer is being focused through genetic modifications of Clostridia and non-Clostridia organisms (e.g., Escherichia coli, Saccharomyces cerevisiae, Pseudomonas putida, and Bacillus subtilis) in both aerobic and anaerobic fermentation. Further, application of various novel genetic tools and genome sequencing of hyper-butanol-producing Clostridial organism will enhance the scope of genetic engineering for biobutanol production. Therefore, consolidation of academic and industrial research towards economic synthesis of biobutanol illustrates the possibility of substantial breakthrough in future. In this review, we focus on (i) selection of suitable bacterial strain (ii) availability of cheaper biomass to produce butanol (iii) metabolic engineering strategies of various microorganisms (iv) attempts at process development and (v) biobutanol recovery techniques that provide future direction of economical biobutanol fermentation.     

Advances in biohydrogen production processes: An approach towards commercialization

Biological H₂ production has an edge over its chemical counterpart mainly because it is environmentally benign. Despite having simpler technology, higher evolution rate of H₂ and the wide spectrum of substrate utilization, the major deterrent of anaerobic dark fermentation process stems from its lower achievable yields. Theoretically, the maximum H₂ yield is 4 mol H₂/mol glucose when glucose is completely metabolized to acetate or acetone in the anaerobic process. But it is somewhat difficult to achieve the complete degradation of glucose to carbon dioxide and H₂ through anaerobic dark fermentation. Moreover, this yield appears too low to be economically viable as an alternative to the existing chemical or electrochemical processes of hydrogen generation. Intensive research studies have already been carried out on the advancement of these processes, such as the development of genetically modified microorganism, improvement of the reactor designs, use of different solid matrices for the immobilization of whole cells, development of two-stage processes, and higher H₂ production rates. Maximum H₂ yield is found to be 5.1 mol H₂/mol glucose. However, major bottlenecks for the commercialization of these processes are lower H₂ yield and rate of H₂ production. Competent microbial cultures are required to handle waste materials efficiently, which are usually complex in nature. This will serve dual purposes: clean energy generation and bioremediation. Scale-up studies on fermentative H₂ production processes have been done successfully. Pilot plant trials of the photo-fermentation processes require more attention. Use of cheaper raw materials and efficient biological H₂ production processes will surely make them more competitive with the conventional H₂ generation processes in near future.     

Sub-dominant bacteria as keystone species in microbial communities producing bio-hydrogen

Nowadays mixed cultures are considered as a serious alternative to pure cultures in biotechnological processes. Mixed cultures can use various organic substrates and be operated under unsterile and continuous conditions. Although most of studies dealing with fermentative hydrogen production have focused on dominant species, sub-dominant bacteria can also have a significant effect despite their low abundance. The determination of their exact ecological role is essential for better understanding microbial metabolic networks in mixed cultures.     

Biological hydrogen production; fundamentals and limiting processes

Biological hydrogen production has been known for over a century and research directed at applying this process to a practical means of hydrogen fuel production has been carried out for over a quarter century. The various approaches that have been proposed and investigated are reviewed and critical limiting factors identified. The low energy content of solar irradiation dictates that photosynthetic processes operate at high conversion efficiencies and places severe restrictions on photobioreactor economics. Conversion efficiencies for direct biophotolysis are below 1% and indirect biophotolysis remains to be demonstrated. Dark fermentation of biomass or wastes presents an alternative route to biological hydrogen production that has been little studied. In this case the critical factor is the amount of hydrogen that can be produced per mole of substrate. Known pathways and experimental evidence indicates that at most 2–3 mol of hydrogen can be obtained from substrates such as glucose. Process economics require that means be sought to increase these yields.     

Enhanced hydrogen production in a UASB reactor by retaining microbial consortium onto carbon nanotubes (CNTs)

Biohydrogen and subsequent biomethane generation from biomass is a promising strategy for renewable energy supply, because this combination can lead to higher energy recovery efficiency and faster fermentation than single methane fermentation. Microbial consortium control by retaining hydrogen-producers through the addition of microbial carriers is an alternative to constructing hydrogen-producing reactors. Here we report the use of carbon nanotubes (CNTs) as microbial carriers to enhance microbial retention and the production of biohydrogen. Laboratory-scale upflow anaerobic sludge blanket (UASB) reactors with CNTs at 100 mg/L achieved a maximal hydrogen production rate of 5.55 L/L/d and a maximal hydrogen yield of 2.45 mol/mol glucose. Compared to frequently used activated carbon (AC) particles, CNTs resulted in quicker startup and better performance of hydrogen fermentation in UASB reactors. Scanning electron microscopy (SEM) and pyrosequencing results revealed that the reactor with CNTs led to a high proportion of hydrogen-producing bacteria among the microbial consortium, which endowed the microbes with strong flocculation capacity and hydrogen productivity.     

Fermentative hydrogen production: Principles, progress, and prognosis

Dark fermentative hydrogen production is an attractive route to the renewable production of hydrogen for a number of reasons. At least in its initial employment, it would use readily available waste streams as substrate. The required reactors would probably be relatively simple in design and based on technology that is already well known and widely used. The metabolic pathways involved are well understood and are reviewed here. A large amount of research has focused on factors affecting hydrogen yields during fermentation of various pure and waste substrates by either defined bacterial cultures or natural microbial flora and some of the pertinent highlights are discussed. Finally, known fermentation pathways can deliver at most 4H₂/glucose, at best a 33% yield. Four different strategies to extract more hydrogen or energy have been proposed and are currently being investigated. The current progress in this direction is presented.     

Biohythane production from organic waste: Recent advancements,technical bottlenecks and prospects

The availability of fossil fuels is a major factor that determines the economy of a country. However, possible exhaustion of fossil fuel deposits as well as increased pollution, and other adverse effects on the environment has prompted us to search for alternative fuels. This resulted in the development of hythane, a blend of hydrogen with methane, at concentrations of 10%–30%. The breakdown of organic substrates using sequential dark fermentation (DF) and anaerobic digestion (AD) leads to biohythane production. The quality and quantity of biohythane can be improved by altering the following aspects: selection, development, and/or genetic engineering of suitable microbial consortium; the use of cheap, appropriate substrates; improved design of bioreactors; and the implementation of two-stage fermentation system. This review focusses on the mechanism of biohythane production and the different aspects involved in increasing both its production rate and quality. A comparative study has also been done to demonstrate the superiority of biohythane over other biofuels.     

Bio-butanol vs. bio-ethanol: A technical and economic assessment for corn and switchgrass fermented by yeast or Clostridium acetobutylicum

Fermentation-derived butanol is a possible alternative to ethanol as a fungible biomass-based liquid transportation fuel. We compare the fermentation-based production of n-butanol vs. ethanol from corn or switchgrass through the liquid fuel yield in terms of the lower heating value (LHV). Industrial scale data on fermentation to n-butanol (ABE fermentation) or ethanol (yeast) establishes a baseline at this time, and puts recent advances in fermentation to butanol in perspective. A dynamic simulation demonstrates the technical, economic and policy implications.The energy yield of n-butanol is about half that of ethanol from corn or switchgrass using current ABE technology. This is a serious disadvantage for n-butanol since feedstock costs are a significant portion of the fuel price. Low yield increases n-butanol's life-cycle greenhouse gas emission for the same amount of LHV compared to ethanol. A given fermenter volume can produce only about one quarter of the LHV as n-butanol per unit time compared to ethanol. This increases capital costs. The sometimes touted advantage of n-butanol being more compatible with existing pipelines is, according to our techno-economic simulations insufficient to alter the conclusion because of the capital costs to connect plants via pipeline.     

Energy balance of dark anaerobic fermentation as a tool for sustainability analysis

A process aimed at producing energy needs to produce more energy than the energy necessary to run the process itself in order to be energetically sustainable. In this paper, an energy balance of a batch anaerobic bioreactor has been defined and calculated, both for different operative conditions and for different reactor scales, in order to analyze the sustainability of hydrogen production through dark anaerobic fermentation. Energy production in the form of hydrogen and methane, energy to warm up the fermentation broth, energy loss during fermentation and energy for mixing and pumping have been considered in the energy balance. Experimental data and literature data for mesophilic microorganism consortia have been used to calculate the energy balance. The energy production of a mesophilic microorganism consortium in a batch reactor has been studied in the 16–50 °C temperature range. The hydrogen batch dark fermentation resulted to only have a positive net production of energy over a minimal reactor dimension in summer conditions with an energy recovery strategy. The best working temperature resulted to be 20 °C with 20% of available energy. Hydrogen batch dark fermentation may be coupled with other processes to obtain a positive net energy by recovering energy from the end products of hydrogen dark fermentation. As an example, methane fermentation has been considered to energetically valorize the end products of hydrogen fermentation. The combined process resulted in a positive net energy over the whole range of tested reactor dimension with 45–90% of available energy.     

Simultaneous enzymatic saccharification and ABE fermentation using pretreated oil palm empty fruit bunch as substrate to produce butanol and hydrogen as biofuel

Simultaneous saccharification and acetone–ethanol–butanol (ABE) fermentation was conducted in order to reduce the number of steps involved in the conversion of lignocellulosic biomass into butanol. Enzymatic saccharification of pretreated oil palm empty fruit bunch (OPEFB) by cellulase produced 31.58 g/l of fermentable sugar. This saccharification was conducted at conditions similar to the conditions required for ABE fermentation. The simultaneous process by Clostridium acetobutylicum ATCC 824 produced 4.45 g/l of ABE with butanol concentration of 2.75 g/l. The butanol yield of 0.11 g/g and ABE yield of 0.18 g/g were obtained from this simultaneous process as compared to the two-step process (0.10 g/g of butanol yield and 0.14 g/g of ABE yield). In addition, the simultaneous process also produced higher cumulative hydrogen (282.42 ml) than to the two-step process (222.02 ml) after 96 h of fermentation time. This study suggested that the simultaneous process has the potential to be implemented for the integrated production of butanol and hydrogen from lignocellulosic biomass.     

Critical assessment of anaerobic processes for continuous biohydrogen production from organic wastewater

Production of biohydrogen using dark fermentation has received much attention owing to the fact that hydrogen can be generated from renewable organics including waste materials. The key to successful application of anaerobic fermentation is to uncouple the liquid retention time and the biomass retention time in the reactor system. Various reactor designs based on biomass retention within the reactor system have been developed. This paper presents our research work on bioreactor designs and operation for biohydrogen production. Comparisons between immobilized-cell systems and suspended-cell systems based on biomass growth in the forms of granule, biofilm and flocs were made. Reactor configurations including column- and tank-based reactors were also assessed. Experimental results indicated that formation of granules or biofilms substantially enhanced biomass retention which was found to be proportional to the hydrogen production rate. Rapid hydrogen-producing culture growth and high organic loading rate might limit the application of biofilm biohydrogen production, since excessive growth of fermentative biomass would result in washout of support carrier. It follows that column-based granular sludge process is a preferred choice of process for continuous biohydrogen production from organic wastewater, indicating maximum hydrogen yield of 1.7 mol-H₂/mol-glucose and hydrogen production rate of 6.8 L-H₂/L-reactor h.     

Biotechnology in petroleum recovery: The microbial EOR

Biotechnology has played a significant role in enhancing crude oil recovery from the depleted oil reservoirs to solve stagnant petroleum production, after a three-stage recovery process employing mechanical, physical and chemical methods. Biotechnologically enhanced oil recovery processes, known as microbial enhanced oil recovery (MEOR), involve stimulating indigenous reservoir microbes or injecting specially selected consortia of natural bacteria into the reservoir to produce specific metabolic events that lead to improved oil recovery. This also involves flooding with oil recovery agents produced ex situ by industrial or pilot scale fermentation. This paper essentially reviews the operating mechanisms and the progress made in enhanced oil recovery through the use of microbes and their metabolic products. Improvement in oil recovery by injecting solvents and gases or by energizing the reservoir microflora to produce them in situ for carbonate rock dissolution and reservoir re-pressurization has been enunciated. The role of biosurfactants in oil mobilization through emulsification and that of biopolymers for selective plugging of oil-depleted zones and for biofilm formation have been delineated. The spoil sport played by sulfate-reducing bacteria (SRB) in MEOR has also been briefly reviewed. The importance of mathematical models used in predicting the applicability of an MEOR strategy and the microbial growth and transport has been qualitatively discussed. The results of some laboratory studies and worldwide field trials applying ex situ and in situ MEOR technologies were compiled and interpreted. However, the potential of the MEOR technologies has not been fully realized due to poor yield of the useful microbial metabolic products, growth inhibition by accumulated toxic metabolites and longer time of incubation. A complete evaluation and assessment of MEOR from an engineering standpoint based on economics, applicability and performance is required to further improve the process efficiency for writing more success stories. Thus, this review attempts to address almost all the issues concerning the MEOR, its past and recent trends and its future prospect and directions.     

A review on bioconversion processes for hydrogen production from agro-industrial residues

Hydrogen is a renewable gas, efficient to produce energy that makes it a suitable alternative and effective solution for a carbon-free environment. Unlike other fossil fuels, combustion of hydrogen does not produce toxic compounds, such as greenhouse gases, carbon monoxide, hydrocarbons, etc., resulting in less environmental pollution. Agro-industrial residues contain several lignocelluloses that favor the growth of microorganisms to produce valuable products such as hydrogen. Of the diverse techniques in hydrogen production, bioconversion proves to be an efficient method in permuting agro-industrial residues into hydrogen. This review provides detailed information on the bioconversion processes and factors involved in hydrogen production from agro-industrial residues including different fermentation processes such as dark fermentation and photo-fermentation, and fuel cell systems such as microbial electrolysis cell and microbial fuel cell. Different pretreatment techniques to enhance the availability of lignocellulose for hydrogen production have been elaborated in this review. Various factors including pH, temperature and nutrient composition of feed, affecting the production efficiency and purity of the products during fermentation have been discussed.     

Quest for sustainable bio‐production and recovery of butanol as a promising solution to fossil fuel

Biobutanol has conventionally been generated by fermentation of carbohydrates derived from biomass (starch or sugar‐based feedstock, such as corn) using Clostridia strains (mainly C. beijerinckii and C. acetobutylicum) under anaerobic conditions in batch mode. Under these premises, it has been tough for the acetone–butanol–ethanol fermentation to compete with petro‐butanol production from an energy efficiency and material consumption standpoint. Challenges for butanol production from biomass comprised high cost of feedstock, scarcity of hyper‐butanol producing bacteria and low butanol yield, volumetric productivity and titre, leading to high water usage and separation‐purification costs. This article is an up‐to‐date review on several under explored sections, such as optimization of fermenter feed, microbial culture responsible for solvent production (co‐culture techniques and electro‐biochemical process), latest recovery techniques and the studies integrating in situ continuous fermentation processes. Biobutanol refinery way forward should build upon the use of low‐cost lignocellulosic matter and zero cost organic wastes and by‐products from food, agriculture, forestry, fermentation and paper industries as feedstock; optimized fermentation of such diversified feed with appropriate hyper‐butanol producing strains in biofilm reactors and integration of fermentation step with hybrid high butanol‐selective recovery techniques. Copyright © 2015 John Wiley & Sons, Ltd.     

Production of hydrogen and methane by one and two stage fermentation of food waste

Anaerobic digestion is an attractive process for generation of hydrogen and methane, which involves complex microbial processes on decomposition of organic wastes and subsequent conversion of metabolic intermediates to hydrogen and methane. Comparative performance of a sequential hydrogen and methane fermentation in two stage process and methane fermentation in one stage process were tested in batch reactor at varying ratios of feedstock to microbial inoculum (F/M) under mesophilic incubation. F/M ratios influence biogas yield, production rate, and potential. The highest H₂ and CH₄ yields of 55 and 94 mL g⁻¹ VS were achieved at F/M of 7.5 in two stage process, while the highest CH₄ yield of 82 mL g⁻¹ VS in one stage process was observed at the same F/M. Acetic and butyric acids are the main volatile fatty acids (VFAs) produced in the hydrogen fermentation stage with the concentration range 10–25 mmol L⁻¹. Little concentrations of VFAs were accumulated in methane fermentation in both stage processes. Total energy recovery in two stage process is higher than that in one stage by 18%. This work demonstrated two stage fermentation achieved a better performance than one stage process.