Typical composition of syngas and other potential gas streams from various sources
With diminishing global reserves of crude oil and increasing demand, especially from developing countries, the pressure on oil supply will grow. Although the 2007-2010 financial crisis brought down the price of crude oil (per barrel) from a record peak of US$145 in July 2008, factors such as recovering global economies and political instability in the Middle East have restored the price of crude oil to the US$100 mark. At current rate of consumption, the global reserves of petroleum are predicted to be exhausted within 50 years [1, 2]. This, coupled with the deleterious environmental impacts that result from accumulating atmospheric CO2 from the burning of fossil fuels, the development of affordable, and environmentally sustainable fuels is urgently required. Many countries have responded to this challenge by legislating mandates and introducing policies to stimulate research and development (R&D) and commercialization of technologies that allow the production of low cost, low fossil carbon emitting fuels. For instance, the European Union (EU) has mandated member countries to a target of deriving 10% of all transportation fuel from renewable sources by 2020 . Between 2005 and 2010, renewable energies such as solar, wind, and biofuels have been increasing at an average annual rate of 15-50% . Renewable energy accounted for an estimated 16% of global final energy consumption in 2009 .
Biofuels have been defined as solid (bio-char), liquid (bioethanol, biobutanol, and biodiesel) and gaseous (biogas, biosyngas, and biohydrogen) fuels that are mainly derived from biomass . Liquid biofuels provided a small but growing contribution towards worldwide fuel usage, accounting for 2.7% of global road transport fuels in 2010 . The world’s largest producer of biofuels is the United States (US), followed by Brazil and the EU . In 2009, US and Brazil accounted for approximately 85% of global bioethanol production while Europe generated about 85% of the world’s biodiesel . The global market for liquid biofuels (bioethanol and biodiesel) increased dramatically in recent years, reaching US$83 billion in 2011 and is projected to US$139 billion by 2021 .
The use and production of biofuels has a long history, starting with the inventors Nikolaus August Otto and Rudolph Diesel, who already envisioned the use of biofuels such as ethanol and natural oils when developing the first Otto cycle combustion and diesel engines . While fermentative production of ethanol has been used for thousands of years, mainly for brewing beer starting in Mesopotamia 5000 B.C., fermentative production of another potential biofuel butanol, has only been discovered over the last century, but had significant impact. During the World War 1, Chaim Weizmann successfully applied a process called ABE (acetone-butanol-ethanol) fermentation using
Traditionally sugar substrates derived from food crops such as sugar cane, corn (maize) and sugar beet have been the preferred feedstocks for the production of biofuels. However, world raw sugar prices have witnessed significant volatility over the last decade or so, ranging from US$216/ton in year 2000 to a 30 year high of US$795/ton in February 2011 due to global sugar deficits and crop shortfall . This has created uncertainty and raised sustainability issues about its use as a feedstock for large scale biofuel production. This review aims to shed light on the use of syngas and industrial waste gas as feedstocks, and the emerging field of gas fermentation to generate not only biofuels, but also other high-value added products. The advantages of gas fermentation over conventional sugar-based fermentation and thermochemical conversions, and their flexibility in utilizing a spectrum of feedstocks to generate syngas will be discussed. The biochemistry, genetic and energetic background of the microorganisms that perform this bioconversion process will be critically examined, together with recent advances in systems biology and synthetic biology that offer growing opportunities to improve biocatalysts in terms of both the potential products that can be produced and their process performance. The key processes such as gasification, bioreactor designs, media formulation, and product recovery will be analysed. Finally, the state of commercialization of gas fermentation will be highlighted and an outlook will be provided.
2. Advantages of gas fermentation
The production of first generation biofuels relies on food crops such as sugar beet, sugar cane, corn, wheat and cassava as substrates for bioethanol; and vegetable oils and animal fats for biodiesel. Although years of intense R&D have made methods of bioethanol production (typically using the yeast
These arguments have stimulated the search for so-called second generation biofuels, which utilize non-food lignocellulose biomass such as wood, dedicated energy crops, agricultural residues and municipal solid wastes as feedstocks. Biomass consists of cellulose, hemicellulose and lignin, and the latter of which is extremely resistant to degradation. One approach to unlocking the potential in this abundant feedstock is to separate the lignin from the carbohydrate fraction of the biomass via extensive pre-treatment of the lignocellulose involving, for example, steam-explosion and/or acid hydrolysis. These pre-treatments are designed to allow the carbohydrate portion of the biomass to be broken down into simple sugars, for example by enzymatic hydrolysis using exogenously added cellulases to release fermentable sugars . Such approaches have been found to be expensive and rate limiting [6, 12, 13]. Alternatively, processes using cellulolytic microorganisms (such as
Microorganisms such as acetogens, carboxytrophs and methanogens are able to utilize the CO2 + H2, and/or CO available in such syngas as their sole source of carbon and energy for growth as well as the production of biofuels and other valuable products. However, only acetogens are described to synthesize metabolic end products that have potentials as liquid transportation fuels. While biological processes are generally considered slower than chemical reactions, the use of these microbes to carry out syngas fermentation offers several key advantages over alternative thermo-chemical approaches such as the Fischer-Tropsch’ process (FTP). First, microbial processes operate at ambient temperatures and low pressures which offer significant energy and cost savings. Second, the ambient conditions and irreversible nature of biological reactions also avoid thermodynamic equilibrium relationships and allow near complete conversion efficiencies [14, 15]. Third, biological conversions are commonly more specific due to high enzymatic specificities, resulting in higher product yield with the formation of fewer by-products. Fourth, unlike traditional chemical catalysts which require a set feed gas composition to yield desired product ratios or suite, microbial processes have freedom to operate for the production ofthe same suite of products across a wider range of CO:H2 ratios in the feed gas . Fifth, biocatalysts exhibit a much higher tolerance to poisoning by tars, sulphur and chlorine than inorganic catalysts [6, 16]. However, some challenges have been identified for syngas fermentation to be commercialized, including gas mass transfer limitations, long retention times due to slow cell growth, and lower alcohol production rates and broth concentrations. Recent progress and development to remedy these issues will be highlighted in this review.
3. Feedstock and gasification
Due to the flexibility of the microbes to ferment syngas with diverse composition, virtually any carbonaceous materials can be used as feedstock for gasification. Non-food biomass that can be employed as feedstock for gasification includes agricultural wastes, dedicated energy crops, forest residues, and municipal organic wastes, or even glycerol and feathers [16-20]. Biomass is available on a renewable basis, either through natural processes or anthropogenic activities (e.g. organic wastes). It has been estimated that out of a global energy potential from modern biomass of 250 EJ per year in 2005, only 9 EJ (3.6%) was used for energy generation . The use of existing waste streams such as municipal organic waste also differentiate itself from other feedstocks such as dedicated energy crops because these wastes are available today at economically attractive prices, and they are often already aggregated and require less indirect land use. Alternatively, gasification of non-biomass sources such as coal, cokes, oil shale, tar sands, sewage sludge and heavy residues from oil refining, as well as reformed natural gas are commonly applied as feedstocks for the FTP and can also be used for syngas fermentation [15, 21]. Furthermore, some industries such as steel manufacturing, oil refining and chemical production generate large volume of CO and/or CO2 rich gas streams as wastes. Tapping into these sources using microbial fermentation process essentially convert existing toxic waste gas streams into valuable commodities such as biofuels. The overall process of gas fermentation is outlined in Figure 1.
Prior to gasification, biomass generally needs to go through a pre-treatment process encompassing drying, size reduction (e.g. chipping, grinding and chopping), pyrolysis, fractionation and leaching depending on the gasifier configuration [22, 23]. This upstream pre-treatment process can incur significant capital expense and add to the overall biomass feedstock cost, ranging from US$16-70 per dry ton . Gasification is a thermo-chemical process that converts carbonaceous materials to gaseous intermediates at elevated temperature (600-1000oC), in the presence of an oxidizing agent such as air, steam or oxygen [16, 22]. The resulting syngas contains mainly CO, CO2, H2 and N2, with varying amounts of CH4, water vapour and trace amount of impurities such as H2S, COS, NH3, HCl, HCN, NOx, phenol, light hydrocarbons and tar [17, 22, 24]. The composition and amount of impurities of syngas depends on the feedstock properties (e.g. moisture, dust and particle size), gasifier type and operational conditions (e.g. temperature, pressure, and oxidant) [17, 22]. Table 1 summarizes typical composition of syngas and other potential gas streams derived from various sources.
4. Fixed bed gasifier
Depending on the direction of the flows of carbonaceous fuel and oxidant (air or steam), fixed bed gasifier can be further categorized into updraft or downdraft reactor. In the updraft (counter-current) version of the fixed bed gasifier, biomass enters from the top while gasifying agent from the bottom. The biomass moves down the reactor through zones of drying (100oC), pyrolysis (300oC), gasification (900oC) and finally oxidation zone (1400oC) . Although this mode of gasifier is often associated with high tar content in the exit gas, recent advances in tar cracking demonstrated that very low tar level is achievable . The direct heat exchange of the oxidizing agent with the entering fuel feed results in low gas exit temperature and hence high thermal efficiency [18, 23]. The downdraft (co-current) gasifier has very similar design as the updraft reactor, except the carbonaceous fuel and oxidizing agent flow in the same direction. In comparison to the updraft gasifier, the downdraft reactor has lower tar content in the exit gas but exhibit lower thermal efficiency . Due to the size limitation in the constriction (where most of the gasification occurs) of the reactor, this mode of gasifier is considered unsuitable for large scale operation .
5. Fluidized bed reactor
In fluidized bed reactor, the carbonaceous fuel is mixed together with inert bed material (e.g. silica sand) by forcing fluidization medium (e.g. air and/or steam) through the reactor. The inert bed facilitates better heat exchange between the fuel materials, resulting in nearly isothermal operation conditions and high feedstock conversion efficiencies [18, 22]. The maximum operating temperature of the gasifier is typically around 800 - 900oC, which is limited by the melting point of the bed material . Furthermore, the geometry of the reactor and excellent mixing properties also means that fluidized bed reactors are suitable for up-scaling [18, 22]. Due to these properties, fluidized bed reactor is currently the most commonly used gasifier for biomass feedstock . However, this mode of gasifier is not suitable for feedstocks with high levels of ash and alkali metals because the melting of these components causes stickiness and formation of bigger lumps, which ultimately negatively affect the hydrodynamics of the reactor .
|Composition vol%, dry basis||Ref.|
|Coke oven gas||5.6||1.4||55.4||4.3||28.4||4.9|||
|Partial oxidation of heavy fuel oil||47||4.3||46||1.4||0.3||1|||
|Hardwood chips + 20 wt%liquid crude glycerol||19.73||11.67||19.38||NR*||3.82||NR*|||
|Steam reforming of natural gas||15.5||8.1||75.7||0.2||0.5||0|||
|Steam reforming of Naphtha||6.7||15.8||65.9||2.6||6.3||2.7|||
|Demolition wood + sewage sludge||10.53||15.02||8.02||60.46||3.19||2.78|||
|Kentucky bluegrass straw||12.9||17.4||2.6||64.2||2.1||0.8|||
6. Entrained flow reactor
Entrained flow reactor is the preferred route for large scale gasification of coal, petcoke and refinery residues because of high carbon conversion efficiencies and low tar production . This mode of gasifier does not require inert bed material but relies on feeding the feedstocks co-currently with oxidizing agent at high velocity to achieve a pneumatic transport regime . At operating temperature of 1200-1500oC, this method is able to convert tars and methane, resulting in better syngas quality . Importantly this technology requires the feedstocks to be pulverised into fine particles of ~50 µm before feeding, which is not a major issue for coal but very difficult and costly for biomass sources [18, 22].
7. Microbes and biochemistry of gas fermentation
Acetogens are defined as obligate anaerobes that utilize the reductive acetyl-CoA pathway for the reduction of CO2 to the acetyl moiety of acetyl-coenzyme A (CoA), for the conservation of energy, and for the assimilation of CO2 into cell carbon . In addition to the reductive acetyl-CoA pathway, four other biological pathways are known for complete autotrophic CO2 fixation: the Calvin cycle, the reductive tricarboxylic acid (TCA) cycle, the 3-hydroxypropionate/malyl-CoA cycle and the 3-hydroxypropionate/4-hydroxybutyrate cycle . Since the earlier atmosphere of earth was anoxic and the acetyl-CoA pathway is biochemically the simplest among the autotrophic pathways (the only linear pathway, whereas the other four pathways are cyclic), it has been postulated to be the first autotrophic process on earth [35, 36]. The reductive acetyl-CoA pathway is also known as the ‘Wood-Ljungdahl’ pathway, in recognition of the two pioneers, Lars G. Ljungdahl and Harland G. Wood, who elucidated the chemical and enzymology of the pathway using
|H2/CO2, CO||Acetate||30||6.8||Available||[41, 42]|
|H2/CO2, CO||Acetate, ethanol||37||8.0-8.5||[44, 45]|
|H2/CO2, CO||Acetate, ethanol, butyrate, butanol||37||6||[47-49]|
|H2/CO2, CO||Acetate||30||8.3||Under construction||[50-52]|
|H2/CO2, CO||Acetate, ethanol, 2,3-butanediol, lactate||37||5.8-6.0||[27, 53]|
|H2/CO2, CO||Acetate, ethanol, butyrate, butanol, lactate||38||6.2||Draft||[54, 55]|
|H2/CO2, CO||Acetate, ethanol, butyrate||25-30||5.8- 6.9||[55-57]|
|CO||Acetate, formate||37||NR||[50, 58, 59]|
|H2/CO2, CO||Acetate, ethanol, 2,3-butanediol, lactate||37||6||Available||[27, 62, 63]|
|H2/CO2, CO||Acetate, ethanol, 2,3-butanediol, lactate||37||6.3|||
|H2/CO2, CO||Acetate, ethanol, butyrate||37-40||5.4-7.5||[55, 56]|
|H2/CO2, CO||Acetate||38-39||7.0-7.2||Available||[41, 69]|
|H2/CO2, CO||Acetate, butyrate||36-38||7.3|||
The Acetyl-CoA pathway is essentially a terminal electron-accepting process that assimilates CO2 into biomass . It constitutes an
The reducing equivalents required for fixation of CO2 carbon into acetyl-CoA come from the oxidation of molecular hydrogen under chemolithoautotrophic growth, or NADH and reduced ferredoxin under heterotrophic growth . An extensive review by Calusinska
The direction of the hydrogenase reaction is directed by the redox potential of the components able to interact with the enzyme. Hydrogen evolution occurs when electron donor is available, whereas the presence of electron acceptor results in hydrogen oxidation . Hydrogenases can be classified into three phylogenetically distinct classes of metalloenzymes: [NiFe]-, [FeFe]-, and [Fe]-hydrogenases . In
Most acetogens are also able to utilize another gas carbon monoxide (CO). In contrast to CO2, CO can serve as both a source of carbon btut also as source of electrons such that hydrogen is not necessarily required. With a CO2/CO reduction potential of -524 to -558mV, CO is approximately 1000-fold more capable of generating extremely low potential electrons than NADH, capable of reducing cellular electron carriers such as ferredoxin and flavodoxin [38, 82]. The reducing equivalents generated from CO oxidation can be coupled to reduction of CO2 into acetate, butyrate and/or methane, evolution of molecular hydrogen from protons, reduction of nitrate/nitrite, reduction of sulfur species and reduction of aldehydes into alcohols [35, 83]. However, relatively few microorganisms are able to utilize CO as sole carbon and energy source, probably due to growth inhibition from sensitivity of their metalloproteins and hydrogenases towards CO [38, 83]. During exponential growth of
The operation of this water gas shift reaction is the biochemical basis for the tremendous flexibility that acetogens have in terms of input gas composition. Via this reaction these organisms can flexibly use CO or H2 as a source of electrons.Recently, some acetogens such as
Under chemolithoautotrophic conditions, acetogenesis must not only fix carbon but also conserve energy. Approximately 0.1 mol of ATP is required for generation of 1g of dry biomass in anaerobes . Acetyl-CoA is an energy rich molecule that through the combined actions of Pta (phosphotransacetylase) and Ack (acetate kinase), one ATP can be generated via substrate level phosphorylation (SLP). However, the activation of formate to 10-formyl-H4folate in the methyl-branch of Acetyl-CoA pathway consumes one ATP so no net gain in ATP is achieved via this mechanism [35, 75]. Furthermore, the reduction of CO2 to the carbonyl group also requires energy, estimated at one third of ATP equivalent . Recent advances indicated that other modes of energy conservation such as electron transport phosphorylation (ETP) or chemiosmotic processes that are coupled to the translocation of protons or sodium ions are implicated in acetogens. Acetogens such as
In an attempt to generate an autotrophic
8. Products of gas fermentation
Acetyl-CoA generated via the Wood-Ljungdahl pathway serves as key intermediate for synthesis of cell mass as well as products. All acetogens are described to produce acetate, in order to gain energy via SLP to compensate for the energy invested in activating formate in the
Due to historical roles in ABE fermentation, organisms like
Similar to sugar- and starch-utilizing ABE Clostridia, acetogens such as
For Clostridia such as acetogen
Under low extracellular pH of 4-4.5, the secreted undissociated acetic acid (p
The reassimilation of acetate and butyrate into the respective acyl-CoA and acetoacetate is catalyzed by acetoacetyl-CoA:acetate/butyrate CoA transferase (CtfA and CtfB) [110, 117, 118]. Acetoacetate is deconstructed by acetoacetate decarboxylase (Adc) into acetone and CO2. This enzyme is missing in acetogenic
The final step of solventogenesis utilizes Adh to reduce acetaldehyde and butyraldehyde into ethanol and butanol, respectively. For ethanol synthesis, transposon mutagenesis and enzymatic assay in
Pyruvate is a central molecule for anabolism and it is predominantly generated from glycolysis during heterotrophic growth. But under autotrophic growth, this four carbon molecule can be synthesized by PFOR and potentially also the pyruvate-formate lyase (PFL). Two variants of PFOR were reported in
9. Strain improvement and metabolic engineering
The genomes of several solventogenic Clostridia, including gas fermenting species, have been sequenced since 2001 [54, 62, 109, 119, 123, 130], and an array of transcriptomic [100, 116, 121, 131, 132], proteomic  and systems analysis [133, 134] are being made increasingly available. However, the generation of stable recombinant Clostridia has been severely hindered by the difficulties encountered introducing foreign DNA into cells and a lack of established genetic tools for this genera of bacteria. In comparison to starch-utilizing Clostridia, very little information is available for metabolic engineering of acetogens. Although this section describes recent advances in the development of genetic tools for mostly sugar-utilizing Clostridia, these techniques are highly relevant and applicable to the closely related acetogenic Clostridia for biofuels or chemical production via gas fermentation.
The ideal microbial catalyst for industrial scale gas fermentation might exhibit the following traits: high product yield and selectivity, low product inhibition, no strain degeneration, asporogenous, prolonged cell viability, strong aero-tolerance, high biomass density and efficient utilization of gas substrates. These can be achieved by directed evolution, random mutagenesis and/or targeted genetic engineering. Traditionally, chemical mutagenesis [135-137] and adaption strategies [138, 139] have been deployed to select for these traits. However, these strategies are limited and often come with the expense of unwanted events. First attempts of targeted genetic modification of Clostridia were made in the early 1990s by the laboratory of Prof. Terry Papoutsakis [140-142]. While these pioneering efforts relied on use of plasmids for (over)expression of genes in
Antisense RNA (asRNA) has been employed to down-regulate genes. Here, single stranded RNA binds to a complementary target mRNA and prevents translation by hindering ribosome-binding site interactions . For instance, this method has been used to knockdown
Several homologous recombination methods have been developed for integration or knock-out of genes in a range of sugar-utilizing Clostridia. In early stage, knockout mutants were almost exclusively generated from single crossover events that could revert back to wild-type [146-152], with stable double crossovers only observed in rare cases [153, 154]. For
ClosTron utilizes the specificity of mobile group II intron Ll.
The same laboratory recently also developed another method for integration of DNA into the genome. Termed Allele-Coupled Exchange (ACE), this approach does not employ a counter selective marker to select for the rare second recombination event. Rather, it utilizes the activation or inactivation of gene(s) that result in a selectable phenotype, and asymmetrical homology arms to direct the order of recombination events . Remarkably, the whole genome of phage lambda (48.5kb minus a 6kb region) was successfully inserted into the genome of
For reverse engineering, mainly transposon mutagenesis has been utilized. Earlier efforts of transposon mutagenesis were demonstrated in
While there is still a lack of some other essential metabolic engineering tools such as efficient inducible promoters, the array of available tools that enabled significant improvements to the ABE process and cellulolytic Clostridia fermentations as summarized in Table 3.
|Plasmid overexpression of butanol biosynthetic genes from ||Produced 2 mM butanol from syngas|||
|Plasmid overexpression of butanol biosynthetic genes from ||Produced 26 mM butanol using steel mill gas|||
|Plasmid expression of native ||Increased alcohol tolerance|||
|Plasmid overexpression of acetone operon from ||Produced up to 140 µM acetone using gas||[169, 170]|
|Inactivation of ||Produced same amount of butanol as control but relatively more ethanol, corresponding to a total alcohol tolerance of 21.2 g/l|||
|Inactivation of ||Produced 716 mM ethanol by diverting C4 products|||
|Inactivation of ||Reduction in acetate kinase activity by more than 97% resulted in 80% less acetate produced but similar final solvent amount|||
|Inactivation of ||Produced 216 mM butanol|||
|Inactivation of ||Showed 4 fold increase in ethanol yield (122 mM instead of 28 mM)|||
|Inactivation of||Generated 8.5 times higher ethanol yield (56.4 mM) than wild type (6.5 mM)|||
|Plasmid overexpression of a synthetic acetoneoperon (||Produced 85 mM isopropanol|||
|Genome insertion of ||Converted acetone into 28 mM isopropanol without affecting the yield of other fermentation products|||
|Plasmid overexpression of ||Produced 8.9 mM isobutanol by diverting 2-ketoacid intermediates|||
|Plasmid expression of native ||Produced 70 mg/l riboflavin and 190 mM butanol|||
|Plasmid overexpression of glutathione ||Improved aero- and solvent-tolerance|||
|Plasmid overexpression of chaperone ||Showed 85% decrease in butanol inhibition and 33% increase in solvent yield|||
|Plasmid expression of ||Increased incorporation of CO2 into extracellular products|||
|Knockdown hydrogenase ||Significantly reduced hydrogen uptake activity to 13% (relative to control strain)|||
In contrast, to date only a limited number of acetogenic Clostridia have been successfully modified. Pioneering work in this area has been undertaken in the laboratory of Prof. Peter Dürre.
Besides the classical Clostridial butanol pathway (which constitutes genes
10. Fermentation and product recovery
10.1. Bioreactor design
An optimum gas fermentation system requires efficient mass transfer of gaseous substrates to the culture medium (liquid phase) and microbial catalysts (solid phase). Gas-to-liquid mass transfer has been identified as the rate-limiting step and bottleneck for gas fermentation because of the low aqueous solubility of CO and H2, respectively at only 77% and 68% of that of oxygen (on molar basis) at 35oC . Hence, a bioreactor design that delivers sufficient gas-to-liquid mass transfer in an energy-efficient manner at commercial scale for gas fermentation represents a significant engineering challenge. A brief overview of reactor configurations reported in gas fermentation operations is given below.
10.2. Continuous Stirred Tank Reactor (CSTR)
In continuous stirred tank reactor (CSTR), gas substrates are continuously fed into the reactor and mechanically sheared by baffled impellers into smaller bubbles, which has greater interfacial surface area for mass transfer . In addition, finer bubbles have a slower rising velocity and a longer retention time in the aqueous medium, resulting in higher gas-to-liquid mass transfer . Fermentation reactions using
10.3. Bubble column reactor
In contrast to CSTR, gas mixing in bubble column reactor is achievable by gas sparging, without mechanical agitation. This reactor configuration has fewer moving parts, and consequently has a lower associated capital and operational costs while exhibiting good heat and mass transfer efficiencies, making it a good candidate for large scale gas fermentation . However, excessive level of gas inflow for enhanced mixing have been cited as an issue that leads to heterogeneous flow and back-mixing of the gas substrates [16, 17].
10.4. Immobilized cell column reactor
One of the key challenges of gas fermentation is cell density. Immobilization of microbes through crosslinking or adsorption to insoluble biosupport materials and the subsequent packing within the column offers a range of benefits . These include high cell densities, plug flow operation, high mass transfer rate via direct contact between microbe and gas, reduction of retention time, and operation without mechanical agitation [14, 16]. However, channelling issues may arise when the microbe overgrows and completely fill the interstitial space. Due to limitations in column dimensions and packing, this reactor configuration lacks flexibility to operate in various gas fermentation conditions [14, 16].
10.5. Trickle-bed reactor
Trickle-bed reactor is a gas- or liquid- continuous reactor consisting of packed bed, which liquid culture trickles down through packing media containing suspended or immobilized cells [16, 24, 187]. The gas substrate is delivered either co-currently or counter-currently to the liquid flow, and no mechanical agitation is required .In this reactor format, low gas and liquid flow rates are typically applied, generating relatively low pressure drops . Trickle-bed reactor was found to exhibit excellent gas conversion rates and higher productivities than CSTR and bubble column reactor .
11. Gas fermentation parameters
11.1. Gas composition
The gas composition and its impurities can have an impact on the productivity of the gas fermentation process. Greater molar ratio of H2:CO allows greater efficiency in the conversion of the carbon from CO into products such as ethanol, because reducing equivalents are generated from oxidation of H2 (rather than CO). However, CO is also a known inhibitor of hydrogenase which can affect utilization of H2 during fermentation. In
11.2. Substrate pressure
The partial pressure of syngas components have a major influence on microbial growth and product profiles because the enzymes involved are sensitive to substrate exposure . Due to the low solubility of CO and H2 in water, the growth of dense bacterial cell cultures can face mass transfer limitations, so increasing the partial pressure of gaseous substrates can help alleviate this problem. For instance, studies in which the CO partial pressure (PCO) increased from 0.35 to 2.0 atm showed that this resulted in a 440% increase in maximum cell density, a significant increase in ethanol productivity and a decrease in acetate production in
11.3. Medium formulation
Although acetogens are able of utilizing CO and CO2/H2 as carbon and energy source, other constituents such as vitamins, trace metal elements, minerals and reducing agents are also required for maintenance of high metabolic activity [16, 113]. Studies indicated that formation of ethanol in solventogenic Clostridiais non-growth associated and limitation of growth by reducing availability of carbon-, nitrogen- and phosphate- nutrients shift the balance from acidogenesis to solventogenesis [113, 200, 201]. Optimization of medium formulation for
A low redox potential is necessary for strict anaerobes to grow, hence reducing agents such as sodium thioglycolate, ascorbic acid, methyl viologen, benzyl viologen, titanium (III)–citrate, potassium ferricyanide, cysteine-HCl and sodium sulfide are commonly added to fermentation medium [14, 16, 204]. Furthermore, the addition of reducing agent directs the electron and carbon flow towards solventogenesis by enhancing the availability of reducing equivalents to form NADH for alcohol production [16, 205]. Excessive addition of reducing agents can cause slower microbial growth due to reduced ATP formation from acetogenesis so it is important to determine the optimum concentration of reducing agents [14, 16]. The sulfur containing gases (e.g. H2S) present in syngas are toxic to chemical catalysts but can be beneficial for microbial catalysts by reducing medium redox potential, stimulate redox sensitive enzymes such as CODH, and promote alcohol formation [206, 207].
11.4. Medium pH
Like other organisms, acetogens have a limited range of pH for optimal growth so the pH of the fermentation medium needs to be closely controlled. The extracellular pH directly influences the intracellular pH, membrane potential, proton motive force, and consequently substrate utilization and product profile [208, 209]. In most studies, lowering pH medium divert carbon and electron flow from cell and acid formation towards alcohol production [113, 209-211]. By applying this knowledge, Gaddy and Clausen performed a two-stage CSTR syngas fermentation systems using
The optimum temperature for mesophilic acetogens are between 30-40°C, while thermophilic acetogens grow best between 55 and 58°C. The fermentation temperature not only affects substrate utilization, growth rate and membrane lipid composition of the acetogens, but also gas substrate availability because gas solubility increases with decreasing temperature [24, 211]. “
12. Cell separation and product recovery
To retain high cell densities in reactor, microbes can be grown as biofilm attached to carrier material. Planktonic cells can be retained in the fermentation broth by installing solid/liquid separators such as membranous ultra-filtration units, spiral wound filtration systems, hollow fibres, cell-recycling membranes and centrifuges [214-216]. The concentrations of solvents from gas fermentation rarely exceed 6% [w/v] so a cost- and energy- efficient product recovery process is required. Furthermore, acetogens also exhibit low resistance towards solvents like ethanol [217, 218] and butanol [219, 220] so an
12.1. Liquid-liquid extraction
In liquid-liquid extraction, a water-insoluble organic extractant is mixed with the fermentation broth . Because solvents are more soluble in the organic phase than in the aqueous phase, they get selectively concentrated in the extractant.Although this technique does not remove water or nutrients from the fermentation broth, some gaseous substrates might be removed because CO and H2 have much higher solubility in organic solvents than water [222, 223]. Oleyl alcohol has been the extractant of choice due to its relatively non-toxicity .
Liquid-liquid extraction is associated with several problems including toxicity to the microbes, formation of emulsion, and the accumulation of microbes at the extractant and fermentation broth interphase . In an attempt to remediate these problems, perstraction was developed and this technique employs membrane to separate the extractant from the fermentation broth. This physical barrier prevent direct contact between the microbe and the toxicity of extractant, but it can also limit the rate of solvent extraction and is susceptible to fouling [219, 221]
In a product recovery technique termed pervaporation, a membrane that directly comes in contact with fermentation broth is used to selectively remove volatile compounds such as ethanol and butanol [219, 222]. The volatile compounds diffuse through the membrane as vapour and are then collected by condensation. To facilitate volatilization of permeates into vapour, a partial pressure difference across the membrane is usually maintained by applying a vacuum or inert gas (e.g. N2) across the permeate side of the membrane . Polydimethylsiloxane (PDMS) is the current material of choice for the membrane, but other materials such as poly(1-trimethylsilyl-1-propyne) (PTMSP), hydrophobic zeolite membranes, and composite membranes have also been investigated .
12.4. Gas stripping
Gas stripping is an attractive product recovery method for gas fermentation because the exit gas stream from the bioreactor can be used for
The growing commercial interests in using gas fermentation as a platform for biofuels production is evident in the recent spike in patent fillings within the field . A 2009 report compared mass and energy conversion efficiencies from a process engineering standpoint between enzymatic hydrolysis fermentation of lignocellulose, syngas fermentation and FTP . The authors concluded that while syngas fermentation offers a range of advantages such as low pretreatment requirement and low energy requirement for bioconversion, the technology is severely limited by low ethanol productivity . Another report documented the techno-economic analysis of gas fermentation and concluded that the selling price of ethanol using this technology would still be significantly higher than gasoline in 2009 . In contrast, Griffin and Schultz recently compared the production of ethanol from CO-rich gas using thermo-chemical route and biological gas fermentation route . The authors concluded that gas fermentation offers superior fuel yield per volume of biomass feed, carbon conversion to fuel, energy efficiency and lower carbon emissions relative to the thermo-chemical approach to bioethanol production.
Ethanol and butanol are the most attractive fuel products from current gas fermentation but other by-products such as 2,3-butanediol, acetic acid and butyric acid are also valuable commodities that have the potential to provide significant additional revenue streams, setting off costs for biofuel production. 2,3-butanediol is a high value commodity which can be used to synthesize chemical products such as 1,3-butanediane, methyl ethyl ketone, and gamma butyrolactone, with a combined potential market value of $43 billion . Acetic acid is an important precursor for synthesis of polymers while butyric acid can be used as a flavouring agent in the food industry [229, 230]. With the development of advanced genetic tools for expansion of product range, the industry might witness an increasing emphasis on the production of high-value commodities in addition to biofuels.
Several companies are actively engaged in the development of the gas fermentation technology and some are approaching commercialization. Bioengineering Resources Inc (BRI) founded by Prof. James Gaddy of University of Arkensas, Fayetteville, an early pioneer in the investigation of gas fermentation at scale, was the first company to explore the potential of gas fermentation for industrial bioethanol production. BRI was acquired by chemical company INEOS and rebranded as INEOS Bio (www.ineosbio.com). A pilot-scale facility in Arkansas has been operated since 2003 using several isolates of
Founded in 2006, Coskata Inc. (www.coskata.com) is a US-based company that has reported achieving ethanol yields of 100 gallons per dry ton of wood biomass in a semi-commercial facility in Pennsylvania . The company licensed several microbial strains from the University of Oklahoma , which has filed patents and journal publications for acetogens such as
LanzaTech is a NZ/US based company that has developed a gas fermentation technology to utilize industrial off-gases from steel making and other sources, as well as syngas produced from biomass as feedstocks. The company has reported the development of a proprietary Clostridial biocatalyst that is able to convert the CO-rich waste gas with minimal gas conditioning into bioethanol and the platform chemical 2,3-butanediol. The use of industrial off-gases as feedstock not only helps to reduce the carbon footprint of the steel-making operations but also allows the production of valuable commodities without the costs associated with feedstock gasification. The company has estimated that up to 30 billion gallon of bioethanol per year can be produced from the CO-rich off gases produced through steel manufacturers globally . Founded in 2005, LanzaTech has successfully demonstrated bioethanol production at a pilot plant at BlueScope Steel in Glenbrook, NZ, since 2008 and the company has recently started operating its 100,000 gallon bioethanol per year demonstration facility in Shanghai, China, using waste gas collected from an adjacent steel mill plant owned by its partner Baosteel Group [243, 244]. LanzaTech is planning to build a commercial facility with the capacity to produce 50 million gallon of bioethanol per annum in China by 2013 . The recent acquisition of a biorefinery facility developed by the US-based gasification technology company Range Fuels in Georgia, and a milestone signing of its first commercial customer, Concord Enviro Systems (India), highlighted LanzaTech’s intention to utilize MSW and lignocellulosic waste as feedstocks for biofuel and chemical production [243, 244].
One of the fundamental factors that govern the environmental and economical sustainability of biofuel production is feedstock. Through gasification, a spectrum of renewable non-food feedstock such as agricultural wastes, dedicated energy crops, forest residues, and MSW can be converted into syngas. This article presents a detailed examination of gas fermentation technology in capturing the carbon and energy from syngas and produce biofuels and chemicals. In comparison to indirect fermentation of lignocellulose via enzymatic hydrolysis, and thermo-chemical FTP, gas fermentation offers several advantages such as good product yield and selectivity, operation in ambient conditions, high tolerance to gas impurities, and elimination of expensive pre-treatment steps and costly enzymes. Furthermore, some industries such as steel mill, natural gas steam reforming, oil refining and chemical production generate large volumes of CO-rich off-gas. Gas fermentation can access these existing feedstocks and generate valuable products from these while reducing carbon emissions. Pivotal to gas fermentation is acetogens such as
The main challenges associated with commercialization of gas fermentation have been identified as gas-to-liquid mass transfer limitation, product yield, substrate utilization efficiency, low biomass density and product recovery. Further development of bioreactor is necessary to improve the availability of gas substrates and maintain high cell density for higher productivity. Improvement in integrated product recovery technology is also essential to lower the costs of product recovery and alleviate product inhibition. Gas fermentation appears to be mature enough for commercialization since several companies have already demonstrated their technologies at pilot scale and are moving towards commercialization in the near future.
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