Exploiting the potential of gas fermentation

G Model

ARTICLE IN PRESS

INDCRO-9248; No. of Pages 10

Industrial Crops and Products xxx (2016) xxx–xxx

Contents lists available at ScienceDirect

Industrial Crops and Products journal homepage: www.elsevier.com/locate/indcrop

Exploiting the potential of gas fermentation Stephanie Redl a,1 , Martijn Diender b,1 , Torbjørn Ølshøj Jensen a , Diana Z. Sousa b , Alex Toftgaard Nielsen a,∗ a b

Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Hørsholm, Denmark Laboratory of Microbiology, Wageningen University, Wageningen, The Netherlands

a r t i c l e

i n f o

Article history: Received 15 July 2016 Received in revised form 20 October 2016 Accepted 8 November 2016 Available online xxx Keywords: Syngas fermentation Biomass gasification Co-cultures Mixotrophy Acetogens Thermophiles

a b s t r a c t The use of gas fermentation for production of chemicals and fuels with lower environmental impact is a technology that is gaining increasing attention. Over 38 Gt of CO2 is annually being emitted from industrial processes, thereby contributing significantly to the concentration of greenhouse gases in the atmosphere. Together with the gasification of biomass and different waste streams, these gases have the potential for being utilized for production of chemicals through fermentation processes. Acetogens are among the most studied organisms capable of utilizing waste gases. Although engineering of heterologous production of higher value compounds has been successful for a number of acetogens, the processes are challenging due to the redox balance and the lack of efficient engineering tools. In this review, we address the availability of different gaseous feedstock and gasification processes, and we focus on the advantages of alternative fermentation scenarios, including thermophilic production strains, multi-stage fermentations, mixed cultures, as well as mixotrophy. Such processes have the potential to significantly broaden the product portfolio, increase the product concentrations and yields, while enabling the exploitation of alternative and mixed feedstocks. The reviewed processes also have the potential to address challenges associated with product inhibition and may contribute to reducing the costs of downstream processing. Given the widespread availability of gases, such processes will likely significantly impact the transition towards a more sustainable society. © 2016 Elsevier B.V. All rights reserved.

1. Introduction There is an increasing demand for processes that reduce carbonemissions and ensure carbon neutral and sustainable production of energy and commodities for the steadily growing population (Pachauri et al., 2014). Previous advances in the production of first generation biofuels have raised the feed vs. fuel debate. A promising technology that has gained increasing attention within recent years is gas fermentation, a process in which microorganisms anaerobically convert a gaseous substrate into biofuels and biochemicals. Several microorganisms have the ability to utilize CO2 and CO as

Abbreviations: 3HP, 3-hydroxypropionic acid; 4HB, 4-hydroxybutyrate; ATP, adenosine triphosphate; BDO, butanediol; GHG, greenhouse gas; MEK, methyl ethyl ketone; MSW, municipal solid waste; NADH/NAD+, nicotinamide adenine dinucleotide; NADPH/NADP+, nicotinamide adenine dinucleotide phosphate; NETL, National Energy Technology Laboratory; PFOR, pyruvate:ferredoxin oxidoreductase; PHA, polyhydroxyalkanoates; VFA, volatile fatty acids; VSS, volatile suspended solids; WLP, Wood-Ljungdahl pathway. ∗ Corresponding author. E-mail address: [email protected] (A.T. Nielsen). 1 These authors contributed equally.

energy and carbon source. Acetogenic bacteria are the most studied and have the greatest industrial potential, making use of the Wood-Ljungdahl pathway (WLP) to convert CO2 (and CO). Compared to the Calvin-Benson-Bassham cycle, which is used by plants, algae, cyanobacteria, purple bacteria, and some proteobacteria, the WLP is a highly energy efficient CO2 fixation pathway (Hawkins et al., 2013). Microorganisms employing the WLP are therefore relevant as biotechnological platforms for the production of biofuels and biochemicals from one-carbon compounds, and potentially decrease our dependence on fossil resources. Metabolic traits of gas-fermenting bacteria have been reviewed recently (Daniell et al., 2016; Dürre and Eikmanns, 2015; Latif et al., 2014). The scope of this article is to assess the potential of gas fermentation and ways to exploit this potential. We would like to draw attention to alternative production scenarios, including multi-stage processes, co-cultures, mixotrophy, and thermophilic production strains. To date, pure cultures of mesophilic strains are deployed, with a focus mainly on ethanol, and on 2,3-BDO production. The aforementioned alternative production scenarios on the contrary would broaden the spectrum of products that can be produced from CO and CO2 . Additionally, it would be possible to explore combina-

http://dx.doi.org/10.1016/j.indcrop.2016.11.015 0926-6690/© 2016 Elsevier B.V. All rights reserved.

Please cite this article in press as: Redl, S., et al., Exploiting the potential of gas fermentation. Ind. Crops Prod. (2016), http://dx.doi.org/10.1016/j.indcrop.2016.11.015

G Model INDCRO-9248; No. of Pages 10

ARTICLE IN PRESS S. Redl et al. / Industrial Crops and Products xxx (2016) xxx–xxx

2

tions of different industrial feedstock streams (gas and sugar) and to take advantage of variable process conditions. 2. Feedstock availability The present review focuses on the conversion of CO and CO2 as carbon source by gas fermenting microorganisms. Hereby, CO serves as carbon source and electron donor. When CO2 serves as sole carbon source, an additional electron donor is required. Below, we describe some of the sources of CO, CO2 , and electron donors, as well as their industrial availability. 2.1. Off-gases from industry, heat and energy generation CO- and CO2 -rich waste gases are an attractive substrate for gas fermentation. Many industrial processes produce large amounts of carbon-rich gases that are often left unused, thereby contributing to elevated concentrations of CO2 and CO in the atmosphere. In 2011, about 23% of the total CO2 emissions were derived from industrial processes (van der Hoeven, 2013), the second largest sector contributing to CO2 emissions, after electricity- and heatgeneration installations. More than 40% of the CO2 emissions in 2011 were derived from generation of electricity and heat (van der Hoeven, 2013), which worldwide relies heavily upon coal combustion (Gutmann, 2014). Overall, the anthropogenic CO2 emissions account to 38 Gt/year (Edenhofer et al., 2014). Industrial processes produce CO2 emissions through chemical reactions that do not involve combustion, of which the following three sub-sectors are the main-contributors: iron and steel (27%), non-metallic minerals (27%), and chemicals and petrochemicals (16%) (International Energy Agency, 2007). For example, 60% of the CO2 emissions from cement production come from inevitable chemical reactions in the process (Cement Sustainability Initiative, 2014). Those emissions cannot be prevented by heat and energy generation with renewables, thus alternative strategies for reducing GHG emission are required. According to the world steel association, 1.7 × 109 t of crude steel were produced worldwide in 2014 (World Steel Association, 2015a) and 1.9 tons CO2 are emitted per ton crude steel produced (World Steel Association, 2015b), which accounts to an annual CO2 emission of 3.2 × 109 t. In conclusion, large amounts of carbon rich off-gases are available and their conversion into chemicals and fuels has the potential to significantly decrease GHG emissions (Handler et al., 2015; Ou et al., 2013). Off-gases from electricity and heat generation, as well as industrial waste gases are supposedly a substrate that comes free of charge. Currently, gas fermentation processes closest to commercialization are based on industrial waste gases (LanzaTech, 2016). However, not all waste gases are equally suitable for microbial gas fermentation, since there are demands with regards to the continuity of the gas stream, the carbon content, as well as the purity of the gas. The CO and CO2 content of the off-gas is dependent on the sector, but is also heavily dependent on process parameters and can therefore vary between production sites. For example, the off-gas from power plants contains only 3–4% (gas-fired) to 13–14% (coalfired) CO2 , but process improvements such as chemical looping combustion and oxyfuel-technology (O2 -fired instead of air-fired) can increase the power-efficiency and the CO2 -content of the offgas (International Energy Agency, 2014). The off-gases of other industrial processes contain high percentage of CO2 , for example the production of ethylene oxide emitting nearly 100% CO2 (International Energy Agency, 2014). As it is the case for electricity and heat generation, the off-gas composition can be greatly influenced by the process parameters: for example, the CO2 content of

cement kiln waste gas increases from <50% CO2 (air-fired) to up to 100% when being O2 -fired (International Energy Agency, 2014). 2.2. Gasification of low-value carbonaceous materials to syngas The availability of substrate for gas fermentation is broadened immensely when considering the amount of feedstock that can be converted into carbon- and energy-rich gas streams via gasification. Gasification is defined as the “thermo-chemical conversion of carbonaceous feedstock to gaseous products through a partial oxidation process at elevated temperatures” (Mohammadi et al., 2011). Besides fossil fuels, there is a broad range of more sustainable options: lignocellulosic energy crops such as willow, switchgrass, etc. can serve as feedstock for syngas production. The use of lignocellulosic energy crops has the advantage that their prices are “more stable as they only participate in the energy market” (Daniell et al., 2016). Also algae is an abundant biomass that is suitable as feedstock for gasification (Azadi et al., 2015). Another option is lignocellulosic biomass waste, which is not suitable for food production or starch- and sugar-based production of chemicals and biofuels. The impact of this option is even more significant when taking into account that the lignin-fraction is not utilized in sugar-based production (both 1st and 2nd generation production). In wheat straw the lignin content is around 20% (Sheldon, 2014), and can be as high as 44.5% in some woody biomass (Vassilev et al., 2012). This kind of lignin-rich feedstock, suitable for gasification, accumulates as agricultural residues or as forestry by-products. Crop production (for food, feed, or production of 1st generation biofuels), generates large amounts of residue, with a residue/crop ratio of 1:1.4 for conventional crops (Kim and Dale, 2004). Another interesting source for lignocellulosic biofuels include residues from 2nd generation biofuels production. In the process of ethanol production from corn stover, for example, 5.9times (by mass) more lignocellulosic residues are generated than ethanol (McAloon et al., 2000). Municipal solid waste (MSW) or industrial waste could also serve as feedstock for gasification. A total of 251 × 106 t of MSW was generated in the US in 2012 after annual increases during the last decades (U. S. Environmental Protection Agency, 2014). Especially those parts of the complex waste that are of organic origin possess great potential as starting material for gasification. Although their carbon content is high, with 13% by mass (Staley and Barlaz, 2009), they are currently left unrecycled. In particular, the biodegradation of the aforementioned waste fraction has been identified as one of the main challenges for direct exploitation (Drzyzga et al., 2015). Additionally, sludge from waste water treatment could act as potential feedstock for gasification, but is of lesser relevance, since its carbon-content is relatively low (Drzyzga et al., 2015). The feedstock requirements for gasification technologies are generally considered flexible (Daniell et al., 2016). However, there are certain requirements, for example with regards to the moisture content of the feedstock (Piccolo and Bezzo, 2009). Additionally, the technology of large-scale gasifiers restricts the feed rate to around 2000 t per day (Griffin and Schultz, 2012). There are different gasification technologies available, and fluidized bed gasification is most suitable for large scale gas production, when taking throughput, costs, complexity, and efficiency into account (Alauddin et al., 2010; Mohammadi et al., 2011). The efficiency of conventional biomass gasification, when comparing the lower heating value of the produced syngas with that of the gasification feedstock, is around 85% for biomass and coal (Ptasinski, 2008). Currently, advances in biomass gasification technologies are made (Heidenreich and Foscolo, 2015), thus more efficient gasification technologies are likely to emerge. The syngas composition depends on the feedstock (TiquiaArashiro, 2014), but can be greatly influenced by the gasification

Please cite this article in press as: Redl, S., et al., Exploiting the potential of gas fermentation. Ind. Crops Prod. (2016), http://dx.doi.org/10.1016/j.indcrop.2016.11.015

G Model

ARTICLE IN PRESS

INDCRO-9248; No. of Pages 10

S. Redl et al. / Industrial Crops and Products xxx (2016) xxx–xxx

3

Table 1 Theoretical yields for the production of some compounds of interest from CO, CO2 /H2 , and CO/H2 respectively by C. ljungdahlii. CO

acetate acetone ethanol i-propanol butanol

CO2 /H2

CO/H2

mol/mol CO

g/g CO

mol ATP/mol producta

mol/mol CO2

g/g CO2

mol ATP/mol producta

mol/mol CO

g/g CO

mol ATP/mol producta

0.250 0.125 0.167 0.111 0.083

0.527 0.259 0.274 0.238 0.221

1.13 1.25 1.13 1.75 2.75

0.500 0.333 0.500 0.333 0.250

0.671 0.440 0.523 0.455 0.421

0.125 −0.750 −0.375 0.500 −0.250

0.5 0.25 0.5 0.25 0.25

1.054 0.518 0.822 0.536 0.662

0.625 0.250 0.125 0.500 0.750

a The estimated energy yield is displayed taking into account the minimal energy yield of the C. ljungdahlii metabolism (Diender et al., 2015; Schuchmann and Müller, 2014).

technology and gasification parameters used (Drzyzga et al., 2015; McKendry, 2002). Syngas typically contains 40–50% N2 , 15–20% H2 , 10–15% CO, 10–15% CO2 and 3–5% CH4 (McKendry, 2002). The syngas composition subsequently can be shifted towards a certain composition by gas cleaning and reforming, such as (reverse) water-gas-shift or gas drying. In mid-2016, 95 active gasification plants are operating worldwide (23 in the USA and 72 in the rest of the world) according to the NETL gasification plant database of the U.S. Department of Energy. However, only 16 plants utilize waste or biomass to date, the rest are still petro-fueled. The produced syngas is not yet utilized in gas fermentation processes, but is mainly channeled to the chemical industry for production of fuels, fertilizer, etc. (National Energy Technology Laboratory, 2016). The technology for biomass gasification for syngas production exists, but economic incentives, as well as the necessary infrastructure are required (Roddy, 2013). Until now, only few economic analyses have been published in the scientific literature with respect to fermentation of biomassderived syngas (Choi et al., 2010; Piccolo and Bezzo, 2009; Spath and Dayton, 2003).

2.3. CO and H2 as alternative carbon and electron sources in microbial processes Beyond the utilization of off-gases from industry or syngas generated by gasification of certain feedstock, there are alternative ways to supply the microorganisms with carbon and electrons. Due to its high energy content, CO is in general preferred by gas fermenting microorganisms over CO2 and H2 . Besides reverse water-gas shift reaction, some other methods make it possible to generate CO from CO2 . For example, light-driven processes can be used to perform splitting of CO2 into CO (Service, 2009; Woolerton et al., 2010). In microbial fermentation of C5 and C6 sugars, the substrate is reduced to CO2 . Therefore microbial processes such as production of 1st and 2nd generation biofuels provide an additional source of CO2 . The fermentation off-gases contain nearly 100% CO2 (International Energy Agency, 2014). For fermentation of C5/C6 sugars, the theoretical mass yield of the conversion is 0.51 g ethanol and 0.49 g CO2 per g sugar. As mentioned before, fixation of CO2 by microorganisms requires an additional electron source. The concept of photosynthesis, where electrons are made available by induction with light, can also be exploited for gas fermentation as cyanobacteria are naturally fixing CO2 through photosynthesis. Recently, Sakimoto et al. described a method for artificially photosensitizing the acetogen Moorella thermoacetica. In this hybrid approach, the cells were loaded with cadmium sulfide nanoparticles, which enabled acetate production via photosynthesis (Sakimoto et al., 2016). Some microorganisms can also directly accept electrons from an electrode for the reduction of fixed CO2 , in a process called electrosynthesis. The process is described to be far more efficient than

photosynthesis (Lovley and Nevin, 2013). However, electrosynthesis technology is still awaiting large-scale demonstration. Hydrogen can serve as an electron donor to enable fixation of CO2 for many microorganisms. In addition to the natural hydrogen content of syngas, there are several processes that can lead to the production of H2 . For instance, H2 can be produced by hydrogenogenic microorganisms (Das and Veziroˇglu, 2001). Another option is production of H2 by electrolysis of water, and technologies of several companies have reached commercial levels (Bertuccioli et al., 2014). Already about 4% of the global hydrogen production is derived from water electrolysis (Gandía et al., 2013). The required electricity could be generated by renewables, especially wind power and photovoltaics, to allow energy storage in form of hydrogen during times of excess energy production. In conclusion, there is a substantial amount of gaseous substrates, electron sources, and feedstock for gasification available. Making use of the available resources holds a promise for chemical and fuel production with reduced carbon emission and full exploitation of available resources such as biomass and renewable energy. However, the potential of gas fermentation and its role in the transition towards a bio-based economy can only be exploited if the processes are also economically attractive. Hence, the gaseous substrates have to be converted by the microbial production organisms to a product of sufficiently high value.

3. Status of current gas fermentation processes Bacteria fermenting CO2 - and CO-rich gases have a range of natural products which are mainly of interest as biocommodities or biofuels, and some are already on their way to being commercialized. Acetate is the typical product of acetogenic bacteria and advances have been made to improve the production of natural acetate producers by genetic modification (Straub et al., 2014). Butyrate and 2,3-BDO are other natural products of gas fermenting bacteria that are of interest as biocommodity (Bengelsdorf et al., 2013; Cho et al., 2014; Köpke et al., 2011). However, syngas fermentation processes closer to commercialization generally aim at the production of biofuels. Multiple acetogenic bacteria can naturally produce ethanol (Abubackar et al., 2011) or butanol (Dürre, 2016). Some other natural products of potential industrial interest have been described, such as 2-oxobutyrate (Daniell et al., 2016), hexanol (Daniell et al., 2016), polymers and polymer building blocks (Drzyzga et al., 2015), PHA (Brown, 2007), 3HP/4HB (Hawkins et al., 2013), 5-aminovulinic acid, and vitamin B12 (Wiegel, 1994). Table 1 shows the theoretical production yields for a range of relevant compounds, and significant production of certain biochemicals have already been achieved by gas fermentation as summarized in Table 2. Generally, the natural products that can be produced with higher yields have relatively low commercial value. This, combined with relatively high costs of fermentation and gasification reactors, makes such processes less attractive when compared to conventional production based on fossil resources or on 1st generation

Please cite this article in press as: Redl, S., et al., Exploiting the potential of gas fermentation. Ind. Crops Prod. (2016), http://dx.doi.org/10.1016/j.indcrop.2016.11.015

G Model

ARTICLE IN PRESS

INDCRO-9248; No. of Pages 10

S. Redl et al. / Industrial Crops and Products xxx (2016) xxx–xxx

4

Table 2 Examples of volumetric productivity, titer, and yield of some important gas fermentation target products. Volumetric productivity

Titer

Yield

References

Comments

479 mmol/l/day 7.92 mmol/l/day

833 mM 10 mM

n.a. n.a.

(Straub et al., 2014) (Kim et al., 2016)

11.92 mmol/l/day

97.7 mM

n.a.

(Xu et al., 2015)

A. woodii (ME), controlled batch Multistage process of carboxydotrophic, hydrogenogenic culture converting CO into H2 /CO2 , off-gas in second reactor converted mainly to acetate by the acetogen T. kivui Acetogenic enrichment culture grown on H2 /CO2

193 mmol/l/day

450 mM

0.14 mol/mol CO

(Richter et al., 2013)

0.25 mmol/l/day

5.5 mM

n.a.

(Xu et al., 2015)

n.a. 3.5 mmol/l/day

25.6 mM 6 mM

(Köpke and Liew, 2012) (Diender et al., 2016)

2.7 mmol/l/daya

8 mM

n.a. 0.074 mol/mol CO 0.0080 mol/mol CO

2 mmol/l/day

4 mM

2.1 mmol/l/daya

5 mM

Acetone

10.9 mmol/l/day

52 mM

n.a.

(Hoffmeister et al., 2016)

A.woodii (GM)

Butyrate

227 mmol/l/day

220 mM

n.a.

(Vasudevan et al., 2014)

8.5 mmol/l/day

11 mM

(Diender et al., 2016)

6.3 mmol/l/daya

6 mM

0.136 mol/mol CO 0.0135 mol/mol CO

Chain-elongating open mixed culture fed with effluent of reactor syngas fermenting C. ljungdahlii ERI-2 Co-culture of C. kluyveri and C. autoethanogenum C. carboxydivorans P7 at 25 ◦ C

14.7 mmol/l/day

8.6 mM

n.a.

(Vasudevan et al., 2014)

2.5 mmol/l/day

6 mM

(Diender et al., 2016)

4.6 mmol/l/daya

6 mM

0.074 mol/mol CO 0.0073 mol/mol CO

Acetate

Ethanol

Butanol

Hexanol

Caproate

a

0.05 mol/mol CO 0.0038 mol/mol CO

(Ramió-Pujol et al., 2015) (Diender et al., 2016) (Ramió-Pujol et al., 2015)

(Ramió-Pujol et al., 2015)

(Ramió-Pujol et al., 2015)

C. ljungdahlii ERI-2 Yield calculated from 28% of introduced CO present in carbon of ethanol Acetogen enrichment culture grown on H2 /CO2 C. autoethanogenum (GM) Co-culture of C. kluyveri and C. autoethanogenum C. carboxydivorans P7 at 25 ◦ C Co-culture of C. kluyveri and C. autoethanogenum C. carboxydivorans P7 at 25 ◦ C

Chain-elongating open mixed culture fed with effluent from reactor with syngas fermenting C. ljungdahlii ERI-2 Co-culture of C. kluyveri and C. autoethanogenum C. carboxydivorans P7 at 25 ◦ C

Converted from mmol/g protein/h to volumetric production rates using protein concentrations and maximal production rates reported.

bio-production (Piccolo and Bezzo, 2009; Wei et al., 2009). Therefore, attempts of producing higher value compounds from CO2 and CO gases with genetically engineered strains are being pursued. Two main approaches can be taken: engineering of an autotrophic strain to produce a heterologous compound, or engineering of a heterotrophic producer to utilize gaseous substrates. Initial success to the latter approach, conferring autotrophy on heterotrophic host organisms, has been achieved within the recent years. Engineering of CO2 fixation in a heterotroph was reported for the first time when the 3HP/4HB pathway was engineered into a hyperthermophilic host, Pyrococcus furiosus, for the production of n-butanol from H2 /CO2 (Keller et al., 2013). Efforts have also been made to engineer the WLP for CO into E. coli (Burk et al., 2009), however, only low rates have been reported (Daniell et al., 2016). Antonovsky et al. (2016) recently published a hybrid approach of rational metabolic engineering and laboratory evolution. The evolved E. coli strain employs a fully functional Calvin-Benson-Bassham cycle to fix CO2 , while energy and reduction equivalents are generated by another supplied compound. The developed strain has been the first report of a fully functional carbon fixation cycle in a heterologous host (Antonovsky et al., 2016). More research is focused on the engineering of gas-fermenting microorganisms. Successful heterologous production in Clostridium autoethanogenum, C. ljungdahlii, and Acetobacterium woodii has

been reported for acetone (Becker et al., 2009; Hoffmeister et al., 2016), n-Butanol (Dürre, 2016; Köpke et al., 2010), butyrate (Ueki et al., 2014), isoprene (Beck et al., 2013; Chen et al., 2013), isopropanol (Köpke et al., 2012), and MEK (Mueller et al., 2013). The recently published review by Liew et al. gives an excellent overview of the advances in genetic engineering of syngas-fermenting bacteria (Liew et al., 2016). Cueto-Rojas et al. gives an overview of the potential products which can be formed from gaseous (and nongaseous) substrates when taking the Gibbs free energy content per electron into account (Cueto-Rojas et al., 2015). 4. Alternative gas fermentation scenarios Progress is being made in the field of genetic modification of gas fermenting microorganisms. However, pathways (natural or heterologous) have to make it possible to harvest the energy that can be generated by product formation. The metabolism of syngas-fermenting bacteria operates at a thermodynamic limit (Schuchmann and Müller, 2014) and the ATP yield for a given product is dependent on the production strain (Bengelsdorf et al., 2013; Bertsch and Müller, 2015). The expression of heterologous pathways therefore often disturbs the energy balance of the cell. To fully exploit the potential of gas fermentation, alternative approaches such as multi-stage fermentation, mixed fermentation,

Please cite this article in press as: Redl, S., et al., Exploiting the potential of gas fermentation. Ind. Crops Prod. (2016), http://dx.doi.org/10.1016/j.indcrop.2016.11.015

G Model INDCRO-9248; No. of Pages 10

ARTICLE IN PRESS S. Redl et al. / Industrial Crops and Products xxx (2016) xxx–xxx

co-cultivation, mixotrophy, and thermophilic production strains therefore need to be considered. 4.1. Multi-stage fermentation As the formation of products from gas fermentation is influenced by environmental factors and interesting products are mainly produced under conditions non-optimal for growth, high concentrations of desired products can often not be obtained in a single processing step. Therefore, applying multiple processing stages could broaden the portfolio of products obtained from syngas fermentation together with higher end-product concentrations. One example is the combination of fermentation of easily accessible C5- and C6-compounds of the lignocellulosic biomass with syngas fermentation. The residual lignin-rich biomass, not converted via sugar fermentation, can be gasified, making previously inaccessible carbon available for syngas fermenters. Additionally, as mentioned above, CO2 off-gas from the sugar fermentation might be introduced to the gas fermentation, reducing waste emission and increasing the overall yield of the process. This process can be seen as an upgraded version of the sugar platform accessing the full range of carbon present in the initial material. Syngas fermentation by acetogens generally results in a mixture of both alcohols and acids under optimal growth conditions (Bertsch and Müller, 2015). Lowering pH and decreasing the concentration of yeast extract has shown to move the spectrum towards alcohol production (Abubackar et al., 2012). In addition, many microorganisms cannot tolerate the presence of CO at high concentrations, requiring partly removal of the toxic gas in order to operate efficiently. To produce interesting amounts of one type of product, and to do this efficiently, the fermentation process can be performed in multiple stages. In a study by Kim et al. (2016), acetate was produced as an end-product in a multistage fermentation process using steel mill off-gas as feedstock. In the first part of the process the hydrogenogenic Thermococcus onnurineus converted a large part of the CO fraction in the gas to H2 and CO2 . The resulting H2 /CO/CO2 mixture was subsequently fed into a reactor containing the homoacetogenic Thermoanaerobacter kivui, allowing for generation of acetate as the main end-product at a maximal volumetric production rate of 0.33 mmol/l/h. Absence of the initial biological gas-treatment step caused T. kivui not to grow in the mentioned study due to CO inhibition (Kim et al., 2016). Recently, T. kivui was shown to be capable of adapting to growth on CO up to 100% headspace composition, but this took several transfers with increasing CO pressures (Weghoff and Müller, 2016). A multistage approach can also be used for the production of ethanol, generating acetate in the first step, followed by solventogenesis in the second system (Grethlein and Jain, 1992). Richter et al. (2013) designed a two stage system for producing ethanol from syngas using C. ljungdahlii. The first reactor was employed using acetogenic conditions, producing acetate as the main endproduct. The broth was subsequently fed to a second fermenter, which employed a lower pH causing acetate to be converted to ethanol. An ethanol production rate of 0.37 g/l/h was achieved with this system with maximal concentrations up to 450 mM (Richter et al., 2013). Carbon and hydrogen recovery in the ethanol endproduct were 28% and 74%, respectively. Performance of different clostridial strains were tested in this system showing C. ljungdahlii PETC as the most optimal producer in this case (Martin et al., 2016). In addition to production of relatively simple products such as acetate and ethanol, longer chain VFAs could be produced via a similar multistage system. These VFAs are produced via the process of chain elongation, in which shorter chain VFAs are converted to longer chain VFAs via reverse ␤-oxidation (Angenent et al., 2016). Strains performing this metabolism often grow optimally around

5

pH 7 and lowering the pH significantly decreases their efficiency (Ganigué et al., 2016). Vasudevan and co-workers have shown that effluent from a syngas reactor, containing acetate and ethanol, could be used to feed a reactor performing chain elongation, indirectly obtaining elongated acids from syngas (Vasudevan et al., 2014). A pure culture of C. ljungdahlii strain ERI-2 was grown in the first stage in order to ferment the syngas, while a mixed culture obtained from a chain-elongating reactor was used in the second part of the system. The separation of these two processes allows both systems to function at optimal conditions, obtaining production rates for butyrate and caproate of 20 and 1.7 g/l/day, respectively. For the production of longer chain lipids (C16-C18) from syngas, Hu and coworkers constructed a two-stage system (Hu et al., 2016). This was done by growing M. thermoacetica in the first stage on syngas, mainly producing acetate as the final product. The outgoing broth was fed into an aerobic reactor containing a genetically engineered Yarrowia lipolytica, a widely used oleaginous yeast. The system accumulated up to 18 g/l C16-C18 lipids at a rate of 0.19 g/l/h. This system nicely shows how carbon from syngas can be fixated into more complex products by using a multi-stage system. In theory, such a two-stage system would allow for production of other complex products as the second reactor can contain different genetically altered strains. Not only biological processes can be coupled for production of bio-based chemicals. The integration of bio-based processes with chemical processes enables the production of chemicals rarely obtained from biological systems. A collaboration of LanzaTech with Invista and SK Innovation proposes production of 1,3-butadiene via a two-step process involving a biological and chemical step (Green Chemicals Blog, 2014; Invista, 2015). Butadiene precursors, 2,3-BDO and 1,3-BDO are generated biologically via syngas fermentation. Subsequently these products can be thermocatalytically dehydrated to form butadiene. Additionally, there are attempts to make production of butadiene a one-step process, involving an engineered biological catalyst. Both the one- and twostep process are still in the development stage, but these initiatives show the possibilities for combining different processes in order to expand the scope of syngas derived products. 4.2. Mixed cultures and defined co-cultures The utilization of open or defined mixed cultures has some advantages over pure culture approaches (Drzyzga et al., 2015): (i) microbial interactions can result in the production of different final products, (ii) some strains may detoxify the environment from a toxic compound or reduce the concentration of a certain product allowing for a more efficient conversion of the gas or increased product yield, (iii) cultures are more robust and resilient to changes in environmental conditions, (iv) less prone to contamination by foreign bacteria or collapse due to phage infection. Trials for syngas conversion by open mixed cultures resulted mainly in the production of methane, hydrogen, or acetate. In general, methanogens are poor CO utilizers (Daniels et al., 1977; Diender et al., 2015), and grow slowly on the substrate. In mixed cultures it was observed that methane could be produced indirectly from CO. Under thermophilic conditions, CO was first converted to hydrogen, subsequently acting as substrate for methanogenesis. Under mesophilic conditions it was observed that CO was mainly converted to acetate, which subsequently acted as substrate for methanogenesis. (Sipma et al., 2003, 2004). Growing methanogens in a mixed culture containing carboxydotrophs, such as acetogens and hydrogenogens, does not only enable production of intermediate substrates, but also for the removal of CO as a toxic compound. Open mixed cultures not adapted to growth on CO were shown to be able to convert syngas to biogas (Guiot

Please cite this article in press as: Redl, S., et al., Exploiting the potential of gas fermentation. Ind. Crops Prod. (2016), http://dx.doi.org/10.1016/j.indcrop.2016.11.015

G Model INDCRO-9248; No. of Pages 10 6

ARTICLE IN PRESS S. Redl et al. / Industrial Crops and Products xxx (2016) xxx–xxx

et al., 2011). After enrichment and adaptation of anaerobic sludge, a CO conversion efficiency of 75% was obtained. The overall system was mass transfer-limited, but a specific activity of 20 mmol CH4 /gVSS/day is expected to be achieved under thermophilic conditions. Complementation of traditional anaerobic digestion with gasification of the non-degraded fraction and subsequent fermentation of syngas could result in a significant increase in biogas yield, which for municipal waste can be up to six times higher than in traditional anaerobic digestion (Guiot et al., 2011). However, the additional energy investment for the gasification may reduce the energy efficiency of the overall process. In the work of Alves et al. (2013), a thermophilic enrichment produced mainly H2 and subsequently acetate. Predominant microorganisms in this culture included Desulfotomaculum, Thermincola, and Thermoanaerobacter (Alves et al., 2013). Another study on CO conversion to H2 by mixed cultures was performed by Liu et al. (2016). Here methanogens were inhibited with chloroform in order to obtain production of mainly hydrogen (Liu et al., 2016). CO conversion to hydrogen was 91% in a continuous reactor operated with gas recycling, with an average production rate of 6.1 mmol/gVSS/day. Acetate production from syngas by mixed cultures has also been pursued, obtaining concentrations of up to 23 g/l at a production rate of 0.72 g/l/day, and a maximal yield of 0.663 gacetate /gCO (Nam et al., 2016). Also mixtures of acetate and ethanol could be obtained by mixed cultures (Xu et al., 2015); however, their final concentrations being lower compared to other approaches. In both studies, pH was found to be of major influence on the production capacity of the mixed cultures, being optimal at pH 7. Recently, Ganigué et al. (2016) studied formation of higher alcohols by open mixed cultures fed with syngas. Chain elongated products were generated in these mixed cultures at higher pH while higher alcohols were formed at lower pH, showing that pH is an essential parameter for production of acids or alcohols in a mixed culture (Ganigué et al., 2016). Higher alcohols were also produced via mixed cultures by feeding syngas and longer chain VFAs (Liu et al., 2014). Carboxydotrophic bacteria, such as C. ljungdahlii, are able to convert carboxylic acids into their corresponding alcohols, using syngas as electron donor. Propionic acid, n-butyric acid, isobutyric acid, n-valeric acid, and ncaproic acid are converted into n-propanol, n-butanol, isobutanol, n-pentanol, and n-hexanol (Perez et al., 2013). Mixed cultures are more robust than pure cultures, but their complexity and potential competition within the system often results in a poor understanding of these systems and in a low yield of desired products. An alternative is to use defined synthetic cocultures, which are less complex while still having the benefits from microbial interactions. Defined co-cultures are currently largely applied in food industrial fermentation processes, such as production of dairy products and alcoholic beverages (Bader et al., 2010). In the case of syngas fermentation, Diender et al. (2016) described a co-culture of C. autoethanogenum and C. kluyveri, capable of producing butyrate, caproate, and their respective alcohols from CO as substrate. Butyrate, caproate, butanol, and hexanol were produced at rates of 8.5, 2.5, 3.5, and 2.0 mmol/l/day, respectively. Other configurations of defined co-cultures and bio-augmentation of open-culture systems might stimulate production of different biochemicals from syngas. Defined co-cultures have advantages over open cultures, which have to deal with competition for substrate and potential removal of interesting products. Such competition was observed in a sulfate reducing system where methanogens, sulfate reducers, and acetogens competed for utilization of H2 , resulting in lower efficiency of the overall system (Sipma et al., 2006). Next to applications in food industry, patents have been filed on the use of co-cultures in applications used for butanol production from syngas (Datta and Reeves, 2013) and production of biofuels using photosynthetic co-cultures (Contag, 2015). Table 2

gives an overview of achieved key production data of some relevant compounds by gas fermentation processes. The use of mixed and co-cultures shows that microbial interactions allow for the production of different end-products. Due to competition for substrate and complexity of mixed communities, co-cultures are in theory more efficient and easier to control. However, it remains to be assessed whether co-cultures are robust enough to maintain over long periods in continuous systems. An alternative use for co-cultures would be to act as bio-augmentation in open non-sterile systems. However, further study is required to assess feasibility of such an approach. 4.3. Mixotrophy Some organisms are able to use two or more carbon/electron sources simultaneously, as observed in some phototrophs using both CO2 and organic substrates at the same time for growth. During mixotrophic growth with sugars, electrons, and CO2 released from the fermentation can be fixed via the WLP. Fast et al. (2015) described an anaerobic, non-photosynthetic, mixotrophic fermentation in which 2 CO2 and 8 electrons from glycolysis are used in the WLP (pathway coupling), generating 3 acetyl-CoA instead of the 2 acetyl-CoA normally generated. This causes the fermentation to have a net CO2 exhaust of zero. Despite that the pathway from glycolysis to acetyl-CoA yields 8 electrons (the amount required in the WLP), an additional electron donor can be required as the glycolysis mainly yields low redox potential electrons (mainly in the form of NADH, −340 mV). Only when a pyruvate ferredoxin oxidoreductase (PFOR), generating reduced ferredoxin, is present, all 8 electrons can be stoichiometrically used in the WLP (Table 3). In other cases the more reduced electrons have to be derived from H2 (−414 mV) via bifurcation processes or directly from CO (−520 mV) in order to complete the WLP (Table 3). Mixotrophic growth with CO as additional substrate could also result in higher yields of more reduced products. Therefore, depending on the degree of reduction of the final end-product, the respective electron donor is either generated or has to be supplied (Fast et al., 2015). One of the challenges in mixotrophic cultivation is to prevent catabolite repression, causing one of the substrates not to be used in the presence of the other (Görke and Stülke, 2008). For the use of sugars and H2 /CO2 or CO, both the WLP and the glycolysis need to be active to have the desired effect. Toxic properties of CO might stimulate mixotrophy as its oxidation by the microorganisms can prevent inhibition, and might be necessary for preserving other catabolic routes active. An indirect application of the concept of mixotrophy is the co-utilization of lower value products from syngas-fermenting bacteria with sugars. Chotani et al. (2012) described for example a method to enhance isoprene production in E. coli by co-utilization of acetate and glucose (Chotani et al., 2012). 4.4. Overview of products from alternative gas fermentation scenarios Fig. 1 gives an overview of the product range that can be achieved with multistage fermentation, mixed cultures, and defined co-cultures in comparison to the products of pure cultures of natural and engineered hosts. Considering the approaches involving non-engineered syngas fermenters, multistage fermentation is currently the most suitable option for generating a relatively broad scope of products from gas waste streams. Especially options combining the conversion of syngas by native organisms with the generation of products by genetically engineered strains using the products of the syngas fermenters are interesting. These systems do however lack microbial interaction, which may be necessary for the production of different products like higher alcohols. An avenue to explore is the utilization of mixed and co-cultures for

Please cite this article in press as: Redl, S., et al., Exploiting the potential of gas fermentation. Ind. Crops Prod. (2016), http://dx.doi.org/10.1016/j.indcrop.2016.11.015

G Model

ARTICLE IN PRESS

INDCRO-9248; No. of Pages 10

S. Redl et al. / Industrial Crops and Products xxx (2016) xxx–xxx

7

Table 3 Stoichiometric conversion of different combinations of electron donors is displayed for the formation of acetyl-CoA or acetate. Pathway

WLP

Glycolysisd Glycolysis + WLP (in presence of PFOR) Glycolysis + WLPe (H2 ) Glycolysis + WLPe (CO)

Electron donor

4 CO 4 H2 2 H2 + 2 COb C6 sugar C6 sugar C6 sugar + 28 H2 C6 sugar + 4 CO

CO2 generated

2 −2 0 2 0 −14 2

Acetyl-CoA formation

Acetate formation

Number of molecules

ATP yielda

Number of molecules

ATP yielda

1 1 1 2 3 10 4

0.125 −0.875c −0.75c 2 1.125 −6 0.25

1 1 1 2 3 10 4

1.125 0.125c 0.625c 4 4.125 4 4.25

a For the WLP, energy yields can vary strongly per organism, depending on the type of enzymes present and final end-product. Here the estimated energy yield is displayed taking into account the minimal energy yield of the C. ljungdahlii metabolism (Diender et al., 2015; Schuchmann and Müller, 2014). b Data is displayed under the assumption that CO and H2 are co-utilized. c A negative ATP yield is obtained for the WLP up to formation of acetyl-CoA from H2 /CO2 or a syngas mixture rich in hydrogen, this yield becomes positive when acetate or acetate derived products (e.g. ethanol) are formed. d This reaction yields 8 electrons that need to be re-oxidized via other pathways (not included). e It is assumed that the reaction from pyruvate to acetyl-CoA is not catalysed by a PFOR enzyme and that the electrons derived from glycolysis and acetyl-CoA formation are all in the form of NADH and are reinvested in the WLP.

Fig. 1. Product portfolio of native and engineered acetogens utilizing CO, CO2 and H2 (grey arrows). Multi-stage fermentation (orange arrows) and mixed co-cultures (green arrows) broaden the range of potential products. TAGs: triacylglycerides. MEK: methyl ethyl ketone.

syngas conversion. An added value of mixed and co-cultures is the potential to utilize non-CO tolerant microbes, like methanogens, to grow on syngas by-products. Finally, mixotrophy is interesting for increasing the yield of already existing platforms or decreasing CO2 exhaust from these systems. However, suitable strains need to be selected or engineered to have multiple pathways active at the same time.

4.5. Thermophilic gas fermenting microbes Thermophilic production strains are known to have certain advantages over mesophiles and have attracted significant attention also for gas fermentation. The risk of contamination is reduced, since most contaminants are mesophiles (Bosma et al., 2013), and the oxygen solubility (although also the CO and H2 solubility) is reduced at elevated temperatures (Tiquia-Arashiro, 2014). Furthermore, the use of a thermophilic production host offers the opportunity of low cost downstream processing via condensation and distillation, since the product enters the gas phase and can be condensed from the gas stream leaving the reactor. Since the product does not accumulate in the fermentation broth, it solves problems with product toxicity and inhibition of production pathways.

Several anaerobic, carboxydotrophic thermophiles have been characterized to date (Henstra et al., 2007; Tiquia-Arashiro, 2014). The product spectrum of those thermophilic carboxydotrophs comprises acetate, formate, H2 S, H2 , and CH4 (Henstra et al., 2007; Tiquia-Arashiro, 2014). The first thermophilic acetogen to naturally produce ethanol was the strain Moorella sp. HUC22-1, published in 2004 (Sakai et al., 2005, 2004). Efforts are being made to develop the genetic toolbox of acetogenic thermophiles in order to broaden the range of products. Although production of industrially relevant heterologous products have not been reported so far, most progress is made with M. thermoacetica (Iwasaki et al., 2013; Kita et al., 2013a, 2013b). Recently, we analyzed a hypothetical process in which acetone is produced in a bubble column fermentation at 60 ◦ C from corn stover-derived syngas with a M. thermoacetica strain with regard to thermodynamic and economic aspects (manuscript in preparation). We found that the process has the potential to be economically interesting, if the high costs for biomass gasification are reduced. Fig. 2 shows compounds with relatively low boiling point, which may be interesting product candidates for thermophilic production strains. In M. thermoacetica, the generation of 1 acetyl-CoA requires 0.5 ATP (Eq. (1)); net ATP is only generated in the subsequent step of acetate formation. Acetyl-CoA formation from CO leads to a net

Please cite this article in press as: Redl, S., et al., Exploiting the potential of gas fermentation. Ind. Crops Prod. (2016), http://dx.doi.org/10.1016/j.indcrop.2016.11.015

G Model INDCRO-9248; No. of Pages 10

ARTICLE IN PRESS S. Redl et al. / Industrial Crops and Products xxx (2016) xxx–xxx

8

Fig. 2. Compounds with relatively low boiling point are interesting products for gas fermentation processes using thermophiles. The temperature range and optimum of the thermophilic acetogen M. thermoacetica (Drake and Daniel, 2004) is highlighted in orange.

Fig. 3. The ATP yield and the mol of CO2 consumed/produced based on the energy generation mechanism in M. thermoacetica proposed by Schuchmann and Müller, 2014. MEK: methyl ethyl ketone.

ATP balance of +0.5 ATP (Eq. (2)).

5. Conclusion

Acetyl-CoA from H2 /CO2 : 4 H2 + 2CO2 − > 1acetyl-CoA + 2 H2 O − 0.5ATP

(1)

Acetyl-CoA from CO : 4CO + 2 H2 O − > 1acetyl-CoA + 0.5ATP + 2CO2

(2)

For the production of 1 mol acetone, 2 acetyl-CoA are required. Fig. 3 shows the ATP yield per 1 mol of product. The ATP yield calculations are based on the energy conservation mechanism proposed by Schuchmann and Müller, 2014. For the production of ethanol, 1 acetyl-CoA is reduced to acetaldehyde and further to ethanol, with NADH as electron donor in each step. The 2 NAD+ can generate an additional 0.5 ATP. Acetone can be further reduced with NADH to isopropanol, which generates an additional 0.25 ATP. The production of isoprene via the mevalonate pathway requires 3 acetyl-CoA, 2 NADH, and 3 ATP. The energy requirement for isoprene production is relatively high, and 4 ATP are required per mol product when H2 /CO2 serves as substrate. For the production of methyl ethyl ketone (MEK), 2 acetyl-CoA are converted via pyruvate into acetolactate, which is decarboxylated into acetoin. Acetoin is further reduced with NADH into (R,S)-2,3-BDO, which undergoes conversion to MEK (Mueller et al., 2013). In comparison, the production of the native product of M. thermoacetica yields 1.5 ATP for the growth on CO, and 0.5 ATP for the growth on H2 /CO2 . Assuming that 0.5 ATP is the lower limit to maintain and gain the biomass, only the production of the more reduced compounds acetone, ethanol, isopropanol, and MEK from CO generates enough energy. For the other production scenarios, by-product formation would be necessary.

Vast amounts of industrial waste gases are currently emitted to the atmosphere. Together with gasification of other waste streams and underutilized fractions of biomass, these gases have the potential to serve as substrates for gas fermentation processes for the production of biochemicals. Such processes are attracting increasing attention since they may help to significantly curb the emission of greenhouse gases, while at the same time having the potential of decreasing the production cost of certain chemicals. A significant amount of research is currently focused on developing tools for metabolic engineering of novel acetogenic production organisms, and novel products are constantly being pursued. Heterologous production of higher value compounds has been successful for some strains, but the reactions are often challenging due to the redox balance, and mainly ethanol and BDO production is reaching commercial levels. Currently, there are only few examples of biomass-based gasification plants operating at a commercial scale, and none of these are reported to utilize syngas for gas fermentation processes. To investigate the economic aspect of these processes, it is therefore important to also conduct techno-economic studies. Alternative fermentation processes have the potential to address many of the challenges associated with production of biochemicals through gas fermentation. In this review we have focused on the recent development of processes utilizing for example multi-stage fermentation, mixed communities, defined co-cultures, mixotrophic processes, as well as thermophilic production organisms. Such production scenarios have the potential for significantly broadening the product portfolio, increasing production titers and yields, exploiting different and also mixed feedstock streams, removing inhibitors and decreasing the cost of downstream processing. The future will undoubtedly show further development of industrial processes from this rapidly emerging field of research.

Please cite this article in press as: Redl, S., et al., Exploiting the potential of gas fermentation. Ind. Crops Prod. (2016), http://dx.doi.org/10.1016/j.indcrop.2016.11.015

G Model INDCRO-9248; No. of Pages 10

ARTICLE IN PRESS S. Redl et al. / Industrial Crops and Products xxx (2016) xxx–xxx

Acknowledgements This work was supported by The Novo Nordisk Foundation and a Ph.D. grant from the People Programme (Marie Curie Actions) of the European Union Seventh Framework Programme FP7People-2012-ITN, under grant agreement No. 317058, “BACTORY”. Research of M. Diender and D.Z. Sousa is supported by a ERC grant (project 323009) of the European Union Seventh Framework Programme FP7 and a Gravitation grant (project 024.002.002) of the Netherlands Ministry of Education, Culture and Science and the Netherlands Science Foundation(NWO).

References Abubackar, H.N., Veiga, M.C., Kennes, C., 2011. Biological conversion of carbon monoxide: rich syngas or waste gases to bioethanol. Biofuels Bioprod. Biorefin. 5 (1), 93–114. Abubackar, H.N., Veiga, M.C., Kennes, C., 2012. Biological conversion of carbon monoxide to ethanol: effect of pH, gas pressure, reducing agent and yeast extract. Bioresour. Technol. 114, 518–522. Alauddin, Z.A.B.Z., Lahijani, P., Mohammadi, M., Mohamed, A.R., 2010. Gasification of lignocellulosic biomass in fluidized beds for renewable energy development: a review. Renew. Sustain. Energy Rev. 14 (9), 2852–2862. Alves, J.I., van Gelder, A.H., Alves, M.M., Sousa, D.Z., Plugge, C.M., 2013. Moorella stamsii sp. nov., a new anaerobic thermophilic hydrogenogenic carboxydotroph isolated from digester sludge. Int. J. Syst. Evol. Microbiol. 63 (11), 4072–4076. Angenent, L.T., Richter, H., Buckel, W., Spirito, C.M., Steinbusch, K.J.J., Plugge, C.M., Strik, D.P., Im Grootscholten, T., Buisman, C.J.N., Hamelers, H.V.M., 2016. Chain elongation with reactor microbiomes: open-culture biotechnology to produce biochemicals. Environ. Sci. Technol. 50 (6), 2796–2810. Antonovsky, N., Gleizer, S., Noor, E., Zohar, Y., Herz, E., Barenholz, U., Zelcbuch, L., Amram, S., Wides, A., Tepper, N., 2016. Sugar synthesis from CO2 in Escherichia coli. Cell 166 (1), 115–125. Azadi, P., Brownbridge, G., Mosbach, S., Inderwildi, O., Kraft, M., 2015. Simulation and life cycle assessment of algae gasification process in dual fluidized bed gasifiers. Green Chem. 17 (3), 1793–1801. ´ M.K., Bajpai, R., Stahl, U., 2010. Relevance of Bader, J., Mast-Gerlach, E., Popovic, microbial coculture fermentations in biotechnology. J. Appl. Microbiol. 109 (2), 371–387. Beck, Z.Q., Cervin, M.A., Chotani, G.K., Diner, B.A., Fan, J., Peres, C.M., Sanford, K.J., Scotcher, M.C., Wells, D.H., Whited, G.M., 2013. Recombinant anaerobic acetogenic bacteria for production of isoprene and/or industrial bio-products using synthesis gas. US20140234926 A1. Becker, U., Grund, G., Orschel, M., Doderer, K., Löhden, G., Brand, G., Dürre, P., Lederle, S., Bahl, H.J., Fischer, R.J., 2009. Zellen und Verfahren zur Herstellung von Aceton 10. EP2421960 A1. Bengelsdorf, F.R., Straub, M., Dürre, P., 2013. Bacterial synthesis gas (syngas) fermentation. Environ. Technol. 34 (13–14), 1639–1651. Bertsch, J., Müller, V., 2015. Bioenergetic constraints for conversion of syngas to biofuels in acetogenic bacteria. Biotechnol. Biofuels 8 (1), 1. Bertuccioli, L., Chan, A., Hart, D., Lehner, F., Madden, B., Standen, E., 2014. Development of Water Electrolysis in the European Union: Report for Fuel Cells and Hydrogen Joint Undertaking. http://www.fch.europa.eu/sites/ default/files/study%20electrolyser 0-Logos 0.pdf. Bosma, F., van der Oost, E., de Vos, John M., Willem van Kranenburg, R., 2013. Sustainable production of bio-based chemicals by extremophiles. Curr. Biotechnol. 2 (4), 360–379. Brown, R.C., 2007. Hybrid thermochemical/biological processing. Appl. Biochem. Biotechnol. 137 (1–12), 947–956. Schilling, Christophe, H., Burgard, A., Trawick, John, D., 2009. Methods and organisms for utilizing synthesis gas or other gaseous carbon sources and methanol. WO/2009/094485. Cement Sustainability Initiative, 2014. Cement Sector View on Industrial Innovation (accessed June 2016) https://www.iea.org/media/workshops/2014/ industryreviewworkshopoct/2 Session1 A CSICement 231014.pdf. Chen, W.Y., Liew, F., Koepke, M., 2013. Recombinant microorganisms and uses therefor. US 20130323820 A1. Cho, C., Jang, Y., Moon, H.G., Lee, J., Lee, S.Y., 2014. Metabolic engineering of clostridia for the production of chemicals. Biofuels Bioprod. Biorefin. 9 (2), 211–225. Choi, D., Chipman, D.C., Bents, S.C., Brown, R.C., 2010. A techno-economic analysis of polyhydroxyalkanoate and hydrogen production from syngas fermentation of gasified biomass. Appl. Biochem. Biotechnol. 160 (4), 1032–1046. Chotani, G.K., Nielsen, A.T., Vaviline, D.V., 2012. Methods for increasing microbial production of isoprene, isoprenoids, and isoprenoid precursor molecules using glucose and acetate co-metabolism. WO2013052914 A3. Contag, P.R., 2015. Organism co-culture in the production of biofuels. US8986962 B2. Cueto-Rojas, H.F., van Maris, A.J., Wahl, S.A., Heijnen, J.J., 2015. Thermodynamics-based design of microbial cell factories for anaerobic product formation. Trends Biotechnol. 33 (9), 534–546.

9

Dürre, P., Eikmanns, B.J., 2015. C1-carbon sources for chemical and fuel production by microbial gas fermentation. Curr. Opin. Biotechnol. 35, 63–72. Dürre, P., 2016. Butanol formation from gaseous substrates. FEMS Microbiol. Lett. 363 (6), 1–7. Daniell, J., Nagaraju, S., Burton, F., Köpke, M., Simpson, S.D., 2016. Low-carbon fuel and chemical production by anaerobic gas fermentation. Adv. Biochem. Eng. Biotechnol., 1–29. Daniels, L., Fuchs, G., Thauer, R.K., Zeikus, J.G., 1977. Carbon monoxide oxidation by methanogenic bacteria. J. Bacteriol. 132 (1), 118–126. Das, D., Veziroˇglu, T.N., 2001. Hydrogen production by biological processes: a survey of literature. Int. J. Hydrogen Energy 26 (1), 13–28. Datta, R., Reeves, A., 2013. Syntrophic co-culture of anaerobic microorganism for production of n-butanol from syngas. US20140206066 A1. Diender, M., Stams, A.J.M., Sousa, D.Z., 2015. Pathways and bioenergetics of anaerobic carbon monoxide fermentation. Front. Microbiol. 6, 1725. Diender, M., Stams, A.J.M., Sousa, D.Z., 2016. Production of medium-chain fatty acids and higher alcohols by a synthetic co-culture grown on carbon monoxide or syngas. Biotechnol. Biofuels 9 (1), 1. Drake, H.L., Daniel, S.L., 2004. Physiology of the thermophilic acetogen Moorella thermoacetica. Res. Microbiol. 155 (10), 869–883. Drzyzga, O., Revelles, O., Durante-Rodríguez, G., Díaz, E., García, J.L., Prieto, A., 2015. New challenges for syngas fermentation: towards production of biopolymers. J. Chem. Technol. Biotechnol. 90 (10), 1735–1751. Edenhofer, O., Pichs-Madruga, R., Sokona, Y., Farahani, E., Kadner, S., Seyboth, K., Adler, A., Baum, I., Brunner, S., Eickemeier, P., 2014. Climate change 2014: Mitigation of climate change. Working group III contribution to the fifth assessment report of the Intergovernmental Panel on Climate Change. UK and New York. U. S. Environmental Protection Agency, 2014. Municipal Solid Waste Generation, Recycling, and Disposal in the United States: Facts and Figures for 2012. https:// www.epa.gov/sites/production/files/2015-09/documents/2012 msw fs.pdf. Fast, A.G., Schmidt, E.D., Jones, S.W., Tracy, B.P., 2015. Acetogenic mixotrophy: novel options for yield improvement in biofuels and biochemicals production. Curr. Opin. Biotechnol. 33, 60–72. Görke, B., Stülke, J., 2008. Carbon catabolite repression in bacteria: many ways to make the most out of nutrients. Nat. Rev. Microbiol. 6 (8), 613–624. Gandía, L.M., Arzamendi, G., Diéguez, P.M., 2013. Renewable hydrogen energy: an overview. Renew. Hydrogen Technol., 1–17. ˜ Ganigué, R., Sánchez-Paredes, P., Baneras, L., Colprim, J., 2016. Low fermentation pH is a trigger to alcohol production, but a killer to chain elongation. Front. Microbiol. 7 (702), 1–11. Green Chemicals Blog, 2014. INVISTA and LanzaTech Expand Collaboration (accessed June 2016) http://greenchemicalsblog.com/2014/06/05/invista-andlanzatech-expand-collaboration/. Grethlein, A.J., Jain, M.K., 1992. Bioprocessing of coal-derived synthesis gases by anaerobic bacteria. Trends Biotechnol. 10, 418–423. Griffin, D.W., Schultz, M.A., 2012. Fuel and chemical products from biomass syngas: a comparison of gas fermentation to thermochemical conversion routes. Environ. Prog. Sustain. Energy 31 (2), 219–224. Guiot, S.R., Cimpoia, R., Carayon, G., 2011. Potential of wastewater-treating anaerobic granules for biomethanation of synthesis gas. Environ. Sci. Technol. 45 (5), 2006–2012. Gutmann, K., 2014. Europe’s Dirty 30: How the EU’s Coal-fired Power Plants Are Undermining Its Climate Efforts. CAN Europe, WWF European Policy Office, HEAL, the EEB and Climate. https://europeanclimate.org/wp-content/uploads/ 2014/07/Dirty-30-report-finale2.pdf. Handler, R.M., Shonnard, D.R., Griffing, E.M., Lai, A., Palou-Rivera, I., 2015. Life Cycle Assessments of ethanol production via gas fermentation: anticipated greenhouse gas emissions for cellulosic and waste gas feedstocks. Ind. Eng. Chem. Res. 55 (12), 3253–3261. Hawkins, A.S., McTernan, P.M., Lian, H., Kelly, R.M., Adams, M.W.W., 2013. Biological conversion of carbon dioxide and hydrogen into liquid fuels and industrial chemicals. Curr. Opin. Biotechnol. 24 (3), 376–384. Heidenreich, S., Foscolo, P.U., 2015. New concepts in biomass gasification. Prog. Energy Combust. Sci. 46, 72–95. Henstra, A.M., Sipma, J., Rinzema, A., Stams, A.J.M., 2007. Microbiology of synthesis gas fermentation for biofuel production. Curr. Opin. Biotechnol. 18 (3), 200–206. Hoffmeister, S., Gerdom, M., Bengelsdorf, F.R., Linder, S., Flüchter, S., Öztürk, H., Blümke, W., May, A., Fischer, R.-J., Bahl, H., 2016. Acetone production with metabolically engineered strains of Acetobacterium woodii. Metab. Eng. 36, 37–47. Hu, P., Chakraborty, S., Kumar, A., Woolston, B., Liu, H., Emerson, D., Stephanopoulos, G., 2016. Integrated bioprocess for conversion of gaseous substrates to liquids. P. Natl. Acad. Sci. U. S. A. 113 (14), 3773–3778. International Energy Agency, 2007. Tracking Industrial Energy Efficiency and CO2 Emissions, https://www.iea.org/publications/freepublications/publication/ tracking emissions.pdf. International Energy Agency, 2014. IEA Energy Training Week 2014: Carbon Capture and Storage. https://www.iea.org/media/training/presentations/ etw2014/course4and5/Day 2 Session 2 CCS.pdf. Invista, 2015. INVISTA and LanzaTech Make Breakthrough for Bio-derived Butadiene Production (accessed June 2016) http://www.invista.com/en/news/ pr-invista-and-lanzatech-make-breakthrough-for-bio-derived-butadieneproduction.html.

Please cite this article in press as: Redl, S., et al., Exploiting the potential of gas fermentation. Ind. Crops Prod. (2016), http://dx.doi.org/10.1016/j.indcrop.2016.11.015

G Model INDCRO-9248; No. of Pages 10 10

ARTICLE IN PRESS S. Redl et al. / Industrial Crops and Products xxx (2016) xxx–xxx

Iwasaki, Y., Kita, A., Sakai, S., Takaoka, K., Yano, S., Tajima, T., Kato, J., Nishio, N., Murakami, K., Nakashimada, Y., 2013. Engineering of a functional thermostable kanamycin resistance marker for use in Moorella thermoacetica ATCC 39073. FEMS Microbiol. Lett., 8–12. Köpke, M., Liew, F., 2012. Production of butanol from carbon monoxide by a recombinant microorganism. Patent WO2012/053905. Köpke, M., Held, C., Hujer, S., Liesegang, H., Wiezer, A., Wollherr, A., Ehrenreich, A., Liebl, W., Gottschalk, G., Dürre, P., 2010. Clostridium ljungdahlii represents a microbial production platform based on syngas. P. Natl. Acad. Sci. U. S. A. 107 (29), 13087–13092. Köpke, M., Mihalcea, C., Bromley, J.C., Simpson, S.D., 2011. Fermentative production of ethanol from carbon monoxide. Curr. Opin. Biotechnol. 22 (3), 320–325. Köpke, M., Simpson, S., Liew, F., Chen, W., 2012. Fermentation process for producing isopropanol using a recombinant microorganism. US20120252083 A1. Keller, M.W., Schut, G.J., Lipscomb, G.L., Menon, A.L., Iwuchukwu, I.J., Leuko, T.T., Thorgersen, M.P., Nixon, W.J., Hawkins, A.S., Kelly, R.M., 2013. Exploiting microbial hyperthermophilicity to produce an industrial chemical, using hydrogen and carbon dioxide. P. Natl. Acad. Sci. U. S. A. 110 (15), 5840–5845. Kim, S., Dale, B.E., 2004. Global potential bioethanol production from wasted crops and crop residues. Biomass Bioenergy 26 (4), 361–375. Kim, T.W., Bae, S.S., Lee, J.W., Lee, S.-M., Lee, J.-H., Lee, H.S., Kang, S.G., 2016. A biological process effective for the conversion of CO-containing industrial waste gas to acetate. Bioresour. Technol. 211, 792–796. Kita, A., Iwasaki, Y., Sakai, S., Okuto, S., Takaoka, K., Suzuki, T., Yano, S., Sawayama, S., Tajima, T., Kato, J., 2013a. Development of genetic transformation and heterologous expression system in carboxydotrophic thermophilic acetogen Moorella thermoacetica. J. Biosci. Bioeng. 115 (4), 347–352. Kita, A., Iwasaki, Y., Yano, S., Nakashimada, Y., Hoshino, T., Murakami, K., 2013b. Isolation of thermophilic acetogens and transformation of them with the pyrF and kanr genes. Biosci. Biotechnol. Biochem. LanzaTech, 2016. ArcelorMittal, LanzaTech and Primetals Technologies Announce Partnership to Construct Breakthrough D 87 m Biofuel Production Facility. http://www.lanzatech.com/arcelormittal-lanzatech-primetals-technologiesannounce-partnership-construct-breakthrough-e87m-biofuel-productionfacility/. Accessed June 2016. Latif, H., Zeidan, A.A., Nielsen, A.T., Zengler, K., 2014. Trash to treasure: production of biofuels and commodity chemicals via syngas fermenting microorganisms. Curr. Opin. Biotechnol. 27, 79–87. Liew, F., Martin, M.E., Tappel, R.C., Heijstra, B.D., Mihalcea, C., Köpke, M., 2016. Gas fermentation—a flexible platform for commercial scale production of low-carbon-fuels and chemicals from waste and renewable feedstocks. Front. Microbiol. 7, 694. Liu, K., Atiyeh, H.K., Stevenson, B.S., Tanner, R.S., Wilkins, M.R., Huhnke, R.L., 2014. Mixed culture syngas fermentation and conversion of carboxylic acids into alcohols. Bioresour. Technol. 152, 337–346. Liu, Y., Wan, J., Han, S., Zhang, S., Luo, G., 2016. Selective conversion of carbon monoxide to hydrogen by anaerobic mixed culture. Bioresour. Technol. 202, 1–7. Lovley, D.R., Nevin, K.P., 2013. Electrobiocommodities: powering microbial production of fuels and commodity chemicals from carbon dioxide with electricity. Curr. Opin. Biotechnol. 24 (3), 385–390. Martin, M.E., Richter, H., Saha, S., Angenent, L.T., 2016. Traits of selected Clostridium strains for syngas fermentation to ethanol. Biotechnol. Bioeng. 113 (3), 531–539. McAloon, A., Taylor, F., Yee, W., Ibsen, K., Wooley, R., 2000. Determining the cost of producing ethanol from corn starch and lignocellulosic feedstocks. National Renewable Energy Laboratory Report. McKendry, P., 2002. Energy production from biomass (part 3): gasification technologies. Bioresour. Technol. 83 (1), 55–63. Mohammadi, M., Najafpour, G.D., Younesi, H., Lahijani, P., Uzir, M.H., Mohamed, A.R., 2011. Bioconversion of synthesis gas to second generation biofuels: a review. Renew. Sustain. Energy Rev. 15 (9), 4255–4273. Mueller, A.P., Köpke, M., Nagaraju, S., 2013. Recombinant microorganisms and uses therefor. US 2013/0330809 A1. Nam, C.W., Jung, K.A., Park, J.M., 2016. Biological carbon monoxide conversion to acetate production by mixed culture. Bioresour. Technol. 211, 478–485. National Energy Technology Laboratory, 2016. Gasification Plant Databases (accessed June 2016) http://www.netl.doe.gov/research/coal/energy-systems/ gasification/gasification-plant-databases. Ou, X., Zhang, X., Zhang, Q., Zhang, X., 2013. Life-cycle analysis of energy use and greenhouse gas emissions of gas-to-liquid fuel pathway from steel mill off-gas in China by the LanzaTech process. Front Energy 7 (3), 263–270. Pachauri, R.K., Allen, M.R., Barros, V.R., Broome, J., Cramer, W., Christ, R., Church, J.A., Clarke, L., Dahe, Q., Dasgupta, P., 2014. Climate Change 2014: Synthesis Report. Contribution of Working Groups I, II and III to the Fifth Assessment Report of the Intergovernmental Panel on Climate Change. IPCC. Perez, J.M., Richter, H., Loftus, S.E., Angenent, L.T., 2013. Biocatalytic reduction of short-chain carboxylic acids into their corresponding alcohols with syngas fermentation. Biotechnol. Bioeng. 110 (4), 1066–1077.

Piccolo, C., Bezzo, F., 2009. A techno-economic comparison between two technologies for bioethanol production from lignocellulose. Biomass Bioenergy 33 (3), 478–491. Ptasinski, K.J., 2008. Thermodynamic efficiency of biomass gasification and biofuels conversion. Biofuels Bioprod. Biorefin. 2 (3), 239–253. ˜ Ramió-Pujol, S., Ganigué, R., Baneras, L., Colprim, J., 2015. Incubation at 25C prevents acid crash and enhances alcohol production in Clostridium carboxidivorans P7. Bioresour. Technol. 192, 296–303. Richter, H., Martin, M.E., Angenent, L.T., 2013. A two-stage continuous fermentation system for conversion of syngas into ethanol. Energies 6 (8), 3987–4000. Roddy, D.J., 2013. A syngas network for reducing industrial carbon footprint and energy use. Appl. Therm. Eng. 53 (2), 299–304. Sakai, S., Nakashimada, Y., Yoshimoto, H., Watanabe, S., Okada, H., Nishio, N., 2004. Ethanol production from H2 and CO2 by a newly isolated thermophilic bacterium, Moorella sp. HUC22-1. Biotechnol. Lett. 26 (20), 1607–1612. Sakai, S., Nakashimada, Y., Inokuma, K., Kita, M., Okada, H., Nishio, N., 2005. Acetate and ethanol production from H2 and CO2 by Moorella sp. using a repeated batch culture. J. Biosci. Bioeng. 99 (3), 252–258. Sakimoto, K.K., Wong, A.B., Yang, P., 2016. Self-photosensitization of nonphotosynthetic bacteria for solar-to-chemical production. Science 351 (6268), 74–77. Schuchmann, K., Müller, V., 2014. Autotrophy at the thermodynamic limit of life: a model for energy conservation in acetogenic bacteria. Nat. Rev. Microbiol., 809–821. Service, R.F., 2009. Solar fuels. Sunlight in your tank. Science (New York, NY) 326 (5959), 1472. Sheldon, R.A., 2014. Green and sustainable manufacture of chemicals from biomass: state of the art. Green Chem. 16 (3), 950–963. Sipma, J., Lens, P.N.L., Stams, A.J.M., Lettinga, G., 2003. Carbon monoxide conversion by anaerobic bioreactor sludges. FEMS Microbiol. Ecol. 44 (2), 271–277. Sipma, J., Meulepas, R.J., Parshina, S.N., Stams, A.J., Lettinga, G., Lens, P.N., 2004. Effect of carbon monoxide, hydrogen and sulfate on thermophilic (55 ◦ C) hydrogenogenic carbon monoxide conversion in two anaerobic bioreactor sludges. Appl. Microbiol. Biotechnol. 64 (3), 421–428. Sipma, J., Lettinga, G., Stams, A.J.M., Lens, P.N.L., 2006. Hydrogenogenic CO conversion in a moderately thermophilic (55 ◦ C) sulfate-fed gas lift reactor: competition for CO-Derived H2 . Biotechnol. Progr. 22 (5), 1327–1334. Spath, P.L., Dayton, D.C., 2003. Preliminary screening-technical and economic assessment of synthesis gas to fuels and chemicals with emphasis on the potential for biomass-derived syngas: (NREL/TP-510-34929). Staley, B.F., Barlaz, M.A., 2009. Composition of municipal solid waste in the United States and implications for carbon sequestration and methane yield. J. Environ. Eng. 135 (10), 901–909. Straub, M., Demler, M., Weuster-Botz, D., Dürre, P., 2014. Selective enhancement of autotrophic acetate production with genetically modified Acetobacterium woodii. J. Biotechnol. 178, 67–72. Tiquia-Arashiro, S.M., 2014. Thermophilic Carboxydotrophs and their Applications in Biotechnology. Springer, New York City, NY, pp. 3–978 (doi 10). Ueki, T., Nevin, K.P., Woodard, T.L., Lovley, D.R., 2014. Converting carbon dioxide to butyrate with an engineered strain of Clostridium ljungdahlii. mBio 5 (5), e01636–14. Vassilev, S.V., Baxter, D., Andersen, L.K., Vassileva, C.G., Morgan, T.J., 2012. An overview of the organic and inorganic phase composition of biomass. Fuel 94, 1–33. Vasudevan, D., Richter, H., Angenent, L.T., 2014. Upgrading dilute ethanol from syngas fermentation to n-caproate with reactor microbiomes. Bioresour. Technol. 151, 378–382. Weghoff, M.C., Müller, V., 2016. CO metabolism in the thermophilic acetogen Thermoanaerobacter kivui. Appl. Environ. Microbiol. 82 (8), 2312–2319. Wei, L., Pordesimo, L.O., Igathinathane, C., Batchelor, W.D., 2009. Process engineering evaluation of ethanol production from wood through bioprocessing and chemical catalysis. Biomass Bioenergy 33 (2), 255–266. Wiegel, J., 1994. Acetate and the potential of homoacetogenic bacteria for industrial alications. In: Acetogenesis. Springer, pp. 484–504. Woolerton, T.W., Sheard, S., Reisner, E., Pierce, E., Ragsdale, S.W., Armstrong, F.A., 2010. Efficient and clean photoreduction of CO2 to CO by enzyme-modified TiO2 nanoparticles using visible light. J. Am. Chem. Soc. 132 (7), 2132–2133. World Steel Association, 2015a. Steel Statistical Yearbook 2015. https://www. worldsteel.org/dms/internetDocumentList/bookshop/2015/Steel-StatisticalYearbook-2015/document/Steel%20Statistical%20Yearbook%202015.pdf. World Steel Association, 2015b. Sustainability Indicators (accessed June 2016) https://www.worldsteel.org/statistics/Sustainability-indicators.html. Xu, S., Fu, B., Zhang, L., Liu, H., 2015. Bioconversion of H2 /CO2 by acetogen enriched cultures for acetate and ethanol production: the impact of pH. World J. Microbiol. Biotechnol. 31 (6), 941–950. van der Hoeven, M., 2013. CO2 Emissions From Fuel Combustion: IEA Statistics. https://www.iea.org/publications/freepublications/publication/ CO2EmissionsFromFuelCombustionHighlights2015.pdf.

Please cite this article in press as: Redl, S., et al., Exploiting the potential of gas fermentation. Ind. Crops Prod. (2016), http://dx.doi.org/10.1016/j.indcrop.2016.11.015