Bubble columns generally consist of cylindrically shaped transparent vessels that are aerated by a gas distributor feeding gas bubbles with limited diameter and thus high gas/liquid exchange area into the system. Aeration does not only supply cultures
with carbon dioxide but also induces liquid movement and dispersion and thus contributes to a more equalized distribution of dissolved gasses and also prevents cells from settling. The superficial gas velocity (typical values between 0.01 and

0. 05 m/s (e. g., [53, 9]) affects the radial dispersion coefficient and together with the diameter of gas bubbles determines the interfacial area for mass transfer. In an attempt to improve mass transfer and radial dispersion a system with two different gas distributors was tested. A first gas sparger provides bubbles with a smaller diam­eter mainly for carbon dioxide supply. The second gas distributor provides bubbles with a larger diameter that induce turbulences for improved radial mixing and increased radial dispersion coefficients. At the same time this system should dimin­ish wall adhesion [16].

With regard to the geometrical structure of a bubble column photobioreactor there are several degrees of freedom. The ratio of length to diameter varies significantly. With regard to scale-up the diameter is limited by the light path length. The length of the column, in contrast, is limited by mass transfer and energetic con­siderations because a high hydrostatic pressure requires high power input for the aeration system [40]. A scale-up approach, in this case numbering-up, was demon­strated by the placement of several columns in specific distances and taking into consideration that column reactors mutually shade each other depending on the angle of incident light [9, 53].

A second cylinder can be installed in the center of the column to form an annular reactor [9]. Therewith, the dark liquid volume that does not contribute to the overall productivity is reduced. Simultaneously, the thickness of the column is adjusted to the short light path length and the scalability of the diameter is less restricted. Moreover, if the material chosen for the inner cylinder is transparent, this surface can additionally contribute to illumination of cultures. If energy input is not decisive for the economy of a process, additional light sources could be installed to illumi­nate cultures even from the inside of the annular reactor. However, this approach should not be taken into account for algae cultivation for the energy market.

The airlift principle can be applied to column reactors to improve axial transport. In this case, two interconnected compartments are separated in longitudinal direc­tion. Aeration in just one compartment, the riser, induces and upward flow, while the liquid volume with a lower gas hold up flows down in the downcomer section [41].

Depending on the specific design of the reactor, illumination and dimension of riser and downcomer one should keep in mind that light/dark cycles can be induced. Especially unfavorable slow cycles should be avoided [30] .

This section will explore the process engineering design for an industrial scale microalgal production plant for biodiesel production. The design will focus on four key process steps: biomass cultivation, dewatering, extraction and biodiesel produc­tion. Various unit operations for each process step are designed to compare their appropriateness, and recommendations are made to clearly define the unit opera­tions that best optimise the process.

This design study uses a potassium hydroxide (KOH) catalyst and methanol to syn­thesise FAME based on Sakai et al.’s [29] biodiesel production model, which states that 100 parts of oil and 40 parts of methanol with a KOH catalyst will produce 92 parts of FAME and 21.5 parts of crude glycerol. The study makes use of partially recycled methanol feedstock whose composition is 24 parts of recycled methanol and 16 parts of fresh methanol. Figure 8 shows the transesterification model. Applying this model to the oil yields from the solvent extraction gives a daily biod­iesel production of ~7,158 kg/day and an annual production of ~2,360 tonnes. The yield of glycerol as a byproduct is ~1,673 kg/day.

1.6 Process Economics

Whilst the production of biodiesel from microalgae has been shown to be technically feasible, the viability of microalgal biodiesel as a practical alternative will be ulti­mately determined by its ability to become cost competitive with the current fuels.

After defining the system boundary, all the relevant environmental, economic, and social impacts associated with the microalgae biodiesel supply chain stages have to be identified. These can be the following:

• Energy consumption

• Net GHG emissions

• Freshwater consumption

• Wastewater treatment

• Nutrients consumption (e. g. nitrates, phosphates, carbon source)

• Chemicals for oil extraction (e. g. n-hexane) and biodiesel production (e. g. CH3OH, NaOH)

• Residual biomass management

• Land use

• Potential chemical risk

• Net cash flow generated

• Employment

Then, the relative significance or insignificance of these impacts are identified (for each supply chain stage) based on the authors current knowledge of the pro­cesses involved. For example, energy consumption is a significant impact in many supply chain stages, since it is needed for algae cultivation, harvesting, biomass processing, oil extraction, and biodiesel production, while land use is more significant for microalgae cultivation than in the remaining stages. Freshwater con­sumption is significant for the cultivation stage unless wastewater from another source, which can also be used as a source of nutrients, is used instead to cultivate microalgae.

After a careful analysis of the system under study, involving the identification of the impacts on the domains of sustainability that are significant for each supply chain stage, candidate indicators are selected for the sustainability analysis of microalgae biodiesel. The selected indicators have to fulfill the following conditions [10]:

• Form a coherent set of quantifiable variables consistent with the principles of sustainability

• Be representative of the physical system under study

• Be clear, simple, unambiguous, and not biased

• Be independent of each other and form a small set

• Be directly and easily calculated from system data

The dimensionality of metrics (3D, 2D or 1D) can then be determined [19, 20]. For example, “energy intensity” is a sustainability or 3D indicator, since it takes into account aspects of the three sustainability dimensions. The higher it is, the more negative the impact is for the environment, because of the waste generated in energy production. Yet, as it is positive for the economy, because it is essential for value creation and higher standards of living, it has both positive and negative societal impacts, as future generations will be deprived of currently used sources of energy because of their depletion but lower emissions of pollutants will result from the consumption of biodiesel. Similarly, “land use intensity” contributes to soil degra­dation and biodiversity loss with a negative impact in the environment. It is positive for the economy, because of the value creation from crops produced. Yet, it may have a positive or a negative societal impact depending on how it contributes respec­tively to employment or land competition with other crops, in particular food crops. On the other hand “Contribution to Global Warming” also called as carbon foot­print, can be seen as directly related to global warming and to the environmental effects of it. Also, it is expected that it will create an economic impact because of carbon trading or carbon taxes, if GHG control regulations are in place, this way being a 2D indicator.

After the candidate indicators have been selected, prioritization of the set of indi­cators follows, based on technical input and data availability. Only if the informa­tion exists it is possible to quantify the indicators and perform the sustainability analysis. The conclusions from the analysis will be more reliable if the data are of good quality.

The set of indicators that are of high priority for the sustainability evaluation of microalgae biodiesel are the following:

1. Life cycle energy efficiency (dimensionless), 3D

2. Fossil energy ratio (dimensionless), 1D

3. Land use intensity (m2/MJ biodiesel/year), 3D

4. Contribution to global warming (kg CO2-eq/MJ fuel), 2D

Table 1 synthesizes the sustainability indicators selected for evaluating the sus­tainability of microalgae systems for biodiesel production.

Similar metrics have been proposed by other authors in a biofuels or conven­tional fuels context. For example, Pradhan et al. [ 13] compared four biodiesel energy balances using two indicators: the net energy ratio and the renewability fac­tor. Zhou et al. [22] proposed four indicators for a sustainability assessment of con­ventional fuels during their life cycles. Kim et al. [7] considered the land use change and GHG emissions associated with the production of biofuels.

The purpose of this article is to describe how to perform a sustainability analysis of microalgae biodiesel system comprising the entire supply chain, and also pro­pose specific sustainability indicators. To the authors’ knowledge, no full scale com­mercial plant for microalgae biodiesel exists at present. So, in the absence of commercially relevant data, the reliability of the values of the metrics used at pres­ent would be somewhat limited.

Although some suggestions can be found in literature [1, 6, 8, 9, 16, 21], there are no complete LCA studies with reliable data on biodiesel produced from microal­gae. Also, the majority of the published studies analyzed hypothetical scenarios.

For example, Kadam [6] conducted an LCA to compare the environmental impli­cations of electricity production via coal firing vs. coal/algae co-firing, using 50% of the flue gas from a 50 MW power station as the carbon source to grow microalgae. Results of this study show that when recycling CO2 toward microalgae production it is possible to achieve an overall life cycle CO2 saving of 36.7%.

Table 1 Indicators for the sustainability evaluation of microalgae biodiesel Indicator Definition

Life cycle energy It is the ratio of the total energy produced (energy output) to the total

efficiency (LCEE) energy consumed (energy input)

(dimensionless)

Lifecycle energy efficiency (LCEE) = energy output/energy input

The energy content of by-products may be accounted for in the energy output if they are used for energy production in substitution of fossil fuel or electricity. LCEE also called net energy ratio (NER) measures the relative amount of energy that ends up in the final fuel products

Fossil energy ratio It is the ratio between the amount of energy that goes into the final fuel (FER) product (fuel energy output) and the amount of fossil energy input

(dimensionless) (non-renewable energy) required for the fuel production

Fossil energy ratio (FER) = Fossil energy output/Fossil energy input

FER is also called the renewability factor (RF) since it measures the degree to which a given fuel is or is not renewable. Larger the value of FER less fossil energy is used (assumed to be non-renewable) for the same energy output. A FER greater than one can be used to replace the energy used in producing it. Also, theoretically FER can be infinite if no fossil energy is needed for the fuel production meaning it is “completely” renewable It measures the area of land occupied per unit energy of product

(e. g. the land needed for the biodiesel feedstocks cultivation, which affects biodiversity and life support functions)

It measures the potential contribution of different GHG emissions (e. g. CO2, CH4, N2O) to global warming (or greenhouse effect), expressed as equivalent CO2 emission per unit energy of fuel product

Contributiontoglobalwarming GWP; x E. t,

і

where Ei is the mass of greenhouse gas i emitted to the air and GWP i is the Global Warming Potential of the substance i The total GHG emissions ( E ) through the fuel life cycle are calculated as [3]

■ e „ + e. + e + e + e — e — e„

where e are emissions from the extraction or cultivation of raw

ec

materials; el are annualized emissions from carbon stock changes caused by land use change; ep are emissions from processing; etd are emissions from transport and distribution; eu are emissions from fuel usage; eccs are emission savings from carbon capture and sequestra­tion; eccr are emission savings from carbon capture and replacement; and eee are emission savings from excess electricity from cogeneration

Lardon et al. [8] performed an LCA on the production of biodiesel from Chlorella vulgaris, showing that when this algae is grown in nitrogen-deprived conditions and the oil is extracted directly from the wet biomass without the need for drying, the biodiesel would have a GWP lower than fossil diesel but higher than biodiesel produced from rape seed oil or palm oil.

Lehr and Posten [9] estimated the energy needed for operating a photo-bioreactor compared to the possible chemical energy harvested. According to these authors the economical feasibility of biofuel production by algae cannot be obtained in the short term. The outstanding problems of cost and efficiency of microalgae cultivation for biodiesel need critical attention.

Rodolfi et al. [16] evaluated the biomass productivity, lipid content, and lipid productivity of 30 microalgal strains cultivated in 250 mL flasks. They suggested that in order for microalgae cultures to become an economic, renewable, and car­bon-neutral source of transportation fuel, biofuels production needs to be combined with that of production of higher value co-products.

Clarens et al. [ 1] compared from a life cycle perspective, conventional crops (rapeseed, switch grass, and corn) with microalgae cultivation for biofuels production, concluding that microalgae have higher environmental impacts than these conven­tional crops in terms of energy use, GHG emissions, and water consumption regard­less of cultivation location. These authors suggested that to reduce the impacts, flue gas could be used as a carbon source for producing algae near power plants and also waste­water treatment could be combined with algae cultivation as a source of nutrients.

Stephenson et al. [21] investigated the life cycle global warming potential (GWP) and the fossil energy requirements, for a hypothetical operation in which biodiesel is produced from the freshwater microalgae C. vulgaris. These authors concluded that for a more environmentally sustainable cultivation of these algae in open ponds instead if closed photo-bioreactors, it should be possible to achieve a lipids produc­tivity target of 40 tons/ha/year. This way the GWP of microalgae would be about 80% lower than fossil diesel on a net energy content basis.

The utilization of microalgae, at least from a land use intensity point of view should be a viable option for substituting current feedstocks, as the former has much higher productivity when compared with existing feedstocks. However, the state of development at present precludes a more extensive utilization of microalgae for biodiesel production which could have a real impact in the fossil fuel market.

We can summarize the reasons for the delay of a more widespread usage of microalgae as a feedstock for biofuel production. First, there are still some hurdles to cross concerning their cultivation at large scale. Although some strains are already identified as promising for biofuel production, only a few of them have been attempted at an industrial scale. Critical information about the conditions in which microalgae give the highest yield of lipids or other components of interest is still lacking, especially the nutrient mix and sunlight necessary for inducing it [ 11 ] . Second, there are scaling problems when going from laboratory and pilot scale units to fully commercial plants. When growing microalgae in large open ponds, one has to ensure that all microorganisms receive an adequate amount of energy and nutri­ents, a difficult task when the cell concentration is very high. Third, other challenges are posed by the growth cycle of the microalgae, which should be better understood in order to know when it is the best time for harvesting them, and that during the decline or death phase microalgae cultures are more susceptible to potential contaminations by other organisms that will compete for food and space.

Potential solutions for these problems include the selective growth of particu­larly resistant strains or even their genetic engineering in order to produce species better fitted for biofuels production. Fourth, the microalgae harvesting and process­ing steps, before the biodiesel production, are still under development. As described above, due to high water content of the algal biomass, some of the processes may require high quantities of energy, making the production of biodiesel from microal­gae an energy intensive process, thus increasing its environmental impact [11 ] .

Notwithstanding the possible difficulties, microalgae are seen as one of the most viable options in the medium to far future. Some of the reasons are directly related to their physiology. As simple organisms, they can grow very fast and produce lipids among other metabolites of interest, only requiring water with a given salinity and pH, sunlight, carbon dioxide and a source of nitrogen and other nutrients. This is clearly an advantage over agricultural feedstocks and even future cellulosic raw materials, which normally require pesticides, fertilizers, tillage, and other treat­ments for their production. Also, due to their diversity the probability of finding or engineering the most adequate microalgae strain is high and is easier to do than with the more complex plants.

2 Conclusions

In this work a set of sustainability indicators is proposed in order to assess the utilization of microalgae as a feedstock for biodiesel production from a supply chain point of view, the supply chain stages being cultivation of microalgae, harvesting, and further processing for biodiesel production. Microalgae can be a sustainable option as a feedstock for biofuels production, combining high productivity with high oil content. In particular, the land use intensity is clearly smaller when com­pared to other feedstocks, minimizing the questions directly linked with land use, and the loss of biodiversity. Also, microalgae have the potential to be used in the production of other chemicals of high added value or integrated in existing indus­trial processes for other beneficial purposes, such as carbon sequestration or waste­water treatment. However, more research is still needed to develop more economical industrial production systems and to fully explore the microalgae potentials.

3.1.1 Organic Solvent Extraction

Intracellular chlorophyll has traditionally been extracted from microalgal biomass using organic solvents [49]. The extraction process involves the organic solvent molecules penetrating through the cell membrane and dissolving the lipids as well as the lipoproteins of chloroplast membranes. Parameters which affect the yield of chlorophyll extraction by organic solvents include polarity of the organic solvents, storage conditions of the microalgal biomass, extraction duration, and number of extraction steps [44, 47]. Since chlorophyll is easily oxidized, extraction yield is also affected by the formation of degradation products. Chlorophyll starts degrading as soon as their molecules are exposed to excess light, atmospheric oxygen, high temperature, and acidic or basic pH condition [11 ] .

Acetone, methanol, and ethanol are three of the most common solvents used to recover chlorophyll from microalgae. Table 3 summarizes key findings from previ­ous studies on organic solvent extraction of microalgal chlorophyll. Jeffrey et al.

[24] , Simon and Helliwell [49], Sartory and Grobbelaar [44] found methanol and ethanol to be more efficient at extracting chlorophyll from microalgal biomass than acetone. Simon and Helliwell [49] conducted their sonication-assisted chlorophyll extractions in an ice bath and in the dark to prevent the formation of degradation products. They found that, with sonication, methanol recovered three times more chlorophyll than 90% acetone. Despite these findings, acetone remains the select primary solvent for chlorophyll extractions due to its known propensity to inhibit any chlorophyll degradation.

Macias-Sanchez et al. [28] recently used dimethyl formamide (DMF) to extract chlorophyll from microalgae and revealed the superiority of this solvent compared to the more traditional organic solvents, such as methanol, ethanol, and acetone. Extraction using DMF did not require prior cellular disruption as pigments were completely extracted after a few steps of soaking. Additionally, the chlorophyll remained stable for up to 20 days when stored in the dark at 5°C [32, 47]. The high toxicity associated with DMF, however, substantially decreased its appeal as an extraction solvent.

CO

^4

Storage of the dried microalgal biomass at low temperatures (-18 or -20°C) was found to assist cell disruption and to promote chlorophyll extraction. In a study by Schumann et al. [47], freezing the biomass in liquid nitrogen followed by lyophili — sation and then storage at -18°C was found to be the optimal storage procedure.

Sartory and Grobbelaar [44] found the efficiency of chlorophyll extraction from fresh water microalgae to be optimal when the extraction was carried out at the solvent’s boiling point. It was shown that boiling the biomass for 3-5 min in metha­nol or acetone prior to 24-h extraction led to the complete recovery of chlorophyll a without the formation of any degradation products.

Such findings are contradictory to the general assumption that chlorophyll degraded upon slight temperature elevation.

The amount of chlorophyll extracted from a particular microalgal species was found to be highly dependent on its growth stage. Microalgae extracted in the sta­tionary growth phase were shown to have significantly higher amount of chloro­phyll a compared to the same species in the logarithmic phase [47] .

Phenols are an important group of natural products with antioxidant and other bio­logical activities. These compounds play an important role in algal cell defense against abiotic and biotic stress. Several authors have recently published results regarding the total phenol content and antioxidant activity of algae [40] . Cinnamic acid esters (n-butyl 3,5-dimethoxy-4-hydroxycinnamate and isopropyl 3,5-dimethoxy-

4- hydroxycinnamate) and methyl 3,4,5-trihydroxybenzoate were studied using 1H and 13C NMR in brown algae Spatoglossum variabile [46]. Some of the first polyphenols found in algae (Fucus and Ascophyllum spp.) were phlorotannins. They are formed from the oligomeric structures of phloroglucinol (1,3,5-trihydroxyben — zene) [137]. Also, some flavanone glycosides have been found even in fresh water algae [86] .

The main bioactivity associated to phenolic compounds is antioxidant activity, which is also the main bioactivity of algal and microalgal phenolics [89]. Duan et al. [28] have demonstrated that antioxidant potency of crude extract from red algae (Polysiphoma urceoiata) correlated well with the total phenolic content. Strong cor­relation also existed between the polyphenol content and DPPH radical scavenging activity of a seaweed (H. fusiformis) extract [177], Using electron spin resonance spectrometry and comet assay, Heo et al. [51] found that phenolic content in sea­weeds could raise up to 1,352 mg/g on dry weight basis. The content and profile of phenolic substances in marine algae vary with the species. In marine brown algae, a group of polymers called phlorotannins comprises the major phenolic compounds [20], such as fucols, phlorethols, fucophlorethols, fuhalols, and halogenated and sulfited phlorotannins. Takamatsu et al. [186] showed that bromophenols isolated from several red marine algae exhibited antioxidant activities. These findings sug­gest that phlorotannins, the natural antioxidant compounds found in edible brown algae, can protect food products against oxidative degradation as well as prevent and/or treat free radical-related diseases [89] .

Some algal phenolic compounds have been associated with anti-inflammatory activity, such as rutin, hesperidin, morin, caffeic acid, catechol, catechin, and epi — gallocatechin gallate, whose have been identified in Porphyra genus. Kazlowska et al. [79] have studied recently the phenolic compounds in Porphyra dentata, they identified catechol, rutin, and hesperidin in crude extract using HPLC-DAD. They demonstrated that the crude extract and the phenolic compounds inhibited the production of nitric oxide in LPS-stimulated RAW 264.7 cells. Their results indicate that catechol and rutin, but not hesperidin, are primary bioactive phenolic compounds in the crude extract to suppress NO production in LPS-stimulated macrophages via NF-kB-dependent iNOS gene transcription. Data also explained the anti-inflammatory use and possible mechanism of P. dentata in iNOS — implicated diseases.

Digestibility, or the amount of VS reduced (converted to biogas) during AD, is one of the most important characteristics of the feedstock. The amount of algal VS reduced varies in the range from 20 to 60% for most macro- and microalgae (Tables 16-19). Consequently, the conventional ADP is not able to convert all algal organic matter to biogas and a large fraction of energy is lost as low-value residues. Pretreatment of algal biomass is one of the strategies used for conditioning and increasing algal digest­ibility, methane yield, and degradation rate. Possible goals of pretreatment include:

• Disruption of cell wall

• Size reduction and increase of specific surface area of particulate biomass

• Crystallinity reduction of fiber materials (e. g., cellulose)

• Solubilization of recalcitrant and poorly biodegradable materials (e. g., hemicel — lulose, lignin)

• Partial hydrolysis of cell polymers

• Deactivation of toxic materials

The important requirements for pretreatment methods are to preserve the total organic matter content and to prevent the formation of inhibitory materials. Little is known about providing efficient solutions for increasing algal digestibility. A variety of pretreatment methods have been tested on waste-activated sludge (WAS), live­stock manure, pulp and paper residues, and lignocellulosic biomass [21, 189-193]. Similar methods can be potentially applied for algal biomass conditioning. Methods applied for biomass pretreatment can be classified into the following groups:

• Mechanical—grinding, milling, homogenization, ultrasonic treatment, liquid shear [194-205]

• Thermal—drying, steam pretreatment, hydrothermolysis [195, 206-209]

• Chemical—acid or alkali hydrolysis, ozonation, hydrogen peroxide treatment [198,208,210-213]

• Biological—temperature-phased AD enzymatic treatment [195, 198, 214, 215]

• Electrical—electro-Fenton [216,217]

• Irradiation—gamma-ray, electron-beam, microwave [218-222]

• Combination—thermochemical, wet oxidation [208, 211, 223-226]

Removal of 95% biochemical oxygen demand (BOD), 85% COD, 90% ammonia, >65% total nitrogen, and >99% of the pathogenic indicator microorganisms from municipal wastewater can be achieved in algal ponds [469]. The effluent quality is highly variable: BOD = 10-25 mg/L and COD = 50-85 mg/L [470]. Oswald and col­leagues developed and tested the Advanced Integrated Wastewater Pond System (AIWPS) [471-475]. This system consists of methane production in an advanced facultative pond, algal high rate pond, algal settling pond, and maturation pond.

This system provides a secondary effluent adequate for agricultural irrigation and has 4-5 times lower electrical power consumption per water flow compared to con­ventional activated sludge and extended aeration systems [476]. A conventional wastewater treatment system requires approximately one kWhr of electricity for aeration for the removal of 1 kg of BOD. In contrast, photosynthetic BOD oxidation does not require aeration but produces algal biomass that can be converted to roughly one kWh of electricity through ADP [477] .

Algal Cultivation in Anaerobic Digester Effluent

Recent developments allow ADP to be applied for the treatment of a wide range of wastewaters with organic contamination. But ADP has several drawbacks particu­larly high organic matter and ammonium concentrations in the AD effluent. Moreover, AD has low efficiency of phosphorus removal. A similar waste liquid is generated during AD of algae. This nutrient rich anaerobic effluent can serve as fertilizer for intensive algal production.

Green microalgae (Chlorella and Scenedesmus) cultivated in diluted dairy waste anaerobic digester effluent are able to switch from phototrophic to heterotrophic or mixotrophic growth, utilize native substrates present in effluent, and increase the biomass and triglyceride production rate [478]. Ammonia removal efficiency from anaerobic treated dairy wastewater reached 96% with a mixed green algae culture [479] and 99% with A. platensis [480]. Aragon reported the removal of 85% BOD, 75% COD, 80% ammonia, and >97% detergents during treatment of the anaerobic effluent from an urban wastewater treatment plant by two local algae species Scenedesmus acutus and C. vulgaris [481].

A “closed” system of methane generation from light energy via algal production and anaerobic digestion was described by Golueke and Oswald [109]. The liquid phase from the digester was used as culture media for algal growth. The average methane yield was 0.44 L/gVS, the maximum energy conversion efficiency from light to biomass was 3%, and the energy conversion efficiency for the entire unit was 2%. Ras and colleagues repeated the same experiment with C. vulgaris as the solar light capturing organism [482]. The methane yield was 0.24 L/gVS at an HRT of 28 days and an OLR of 0.7 gVS/L-day.

Ryther compared the productivity of G. tikvahiae and Ulva sp. in media enriched by AD effluent with the productivity in a conventional mineral enrichment medium [483]. Ulva sp. had a similar methane yield in both media, but G. tikvahiae had 50-75% lower productivity in an AD effluent-enriched medium compared to a control.

Moridis and Reagan [131] showed that depressurization-induced dissociation appears to be the most promising gas production strategy in Class 2 deposits. They proposed new well configurations to maximize production and alleviate a persistent problem of substantial secondary hydrate (or ice) formation in a narrow zone (r < 10 m) around the well. Using the properties and conditions representative of the Tigershark formation and producing at an initial constant mass rate of QM = 19.2 kg/s (=10,000 BPD), Moridis and Reagan [131] showed (Fig. 13) that (a) QM cannot be

Fig. 11 Gas production from a class 1 hydrate deposit. Left: evolution of (a) the rate of CH4 release from hydrate dissociation, (b) the rate of CH4 production at the well, and (c) the corresponding rate replenishment ratio over the 30-year production period. Right: evolution of (a) the cumulative CH4 volume released from hydrate dissociation, (b) the produced CH4 volume at the well, and (c) the corresponding volume replenishment ratio over the 30-year production period [129]

maintained constant during the production period (but has to decline), (b) the gas production rate is highly variable, (c) it is encumbered by a long initial lead time during which little gas is produced, but (d) it can reach levels as high as QP=4.8 x 105 m3/day (=17 MMSCFD), with an average gas production Qav over the 5,660-day period of simulation is about 2.2 x 105 m3/day (=7.8 MMSCFD). This study showed very high recovery from hydrate deposits, although economic and geomechanical considerations may limit total recovery. Similar results were obtained from the study of an oceanic Class 2 deposit in the Ulleung Basin of the Korean East Sea [136] and (b) a permafrost-associated deposit in the North slope [133, 134], leading to the observation that QP on the order of several MMSCFD is attainable in Class 2 deposits despite significant differences in reservoir tempera­ture, HBS thickness, and salinity.

The use of horizontal wells can substantially improve gas production from such deposits and reduce the initial period of low QP [137]. Conversely, Moridis and Kowalsky [128] determined that QP was too low to justify considering such accumula­tions as viable targets in the presence of permeable boundaries and/or with a deep WZ.

Robert J. Conrado, Chad A. Haynes, Brenda E. Haendler, and Eric J. Toone

Abstract Biofuels are by now a well-established component of the liquid fuels market and will continue to grow in importance for both economic and environmen­tal reasons. To date, all commercial approaches to biofuels involve photosynthetic capture of solar radiation and conversion to reduced carbon; however, the low efficiency inherent to photosynthetic systems presents significant challenges to scaling. In 2009, the US Department of Energy (DOE) Advanced Research Projects Agency-Energy (ARPA-E) created the Electrofuels program to explore the potential of nonphotosynthetic autotrophic organisms for the conversion of durable forms of energy to energy-dense, infrastructure-compatible liquid fuels. The Electrofuels approach expands the boundaries of traditional biofuels and could offer dramati­cally higher conversion efficiencies while providing significant reductions in requirements for both arable land and water relative to photosynthetic approaches. The projects funded under the Electrofuels program tap the enormous and largely unexplored diversity of the natural world, and may offer routes to advanced biofuels that are significantly more efficient, scalable and feedstock-flexible than routes based on photosynthesis. Here, we describe the rationale for the creation of the Electrofuels program, and outline the challenges and opportunities afforded by chemolithoautotrophic approaches to liquid fuels.

R. J. Conrado • E. J. Toone (*)

US Department of Energy Advanced Research Projects Agency (ARPA-E), 1000 Independence Avenue, SW, Washington, DC 20585, USA e-mail: Eric. Toone@hq. doe. gov

C. A. Haynes • B. E. Haendler

Booz Allen Hamilton, 955 L’Enfant Plaza North, SW Suite 5300, Washington, DC 20024, USA

J. W. Lee (ed.), Advanced Biofuels and Bioproducts, DOI 10.1007/978-1-4614-3348-4_38, 1037

© Springer Science+Business Media New York 2013

1 Introduction

Virtually all transportation-over land, air and sea—utilizes the energy stored in carbon-carbon and carbon-hydrogen bonds to provide motive force. Our use of this stored pool of solar energy can only be described as rapacious, and although the magnitude of the remaining resource is difficult to estimate, it is finite and its extrac­tion will become increasingly complex. The USA currently consumes roughly 19 million barrels of oil per day, over 70% of which is used for transportation [58]. Nearly half the oil consumed in the USA is imported, accounting for a third of the Nation’s trade deficit. In recent years, the annual cost of imported oil has exceeded $300 billion, in current dollars the equivalent of funding the entire Apollo Program twice every year.[14] Even this staggering amount significantly underestimates the true cost of imported oil, since a significant fraction of both defense and nondefense Federal spending is devoted to ensuring a stable supply of imported oil. Alternative approaches to liquid fuels are both a national and global imperative.

The sustainable production of energy-dense, infrastructure compatible liquid fuels requires the conversion of a durable form of energy-most plausibly solar radiation, but also geothermal, nuclear, or other forms of renewable energy—into stored chemical energy. The biological production of carbon-based liquid fuels requires three distinct steps: the capture of energy and transduction of that energy to a usable form by an organism; reduction of inorganic carbon to a fungible metabolic intermediate, typically in an oxidation state at or below zero; and the formation of carbon-carbon bonds to provide a fuel with convenient physical properties. Although the reduction of inorganic carbon can be achieved chemically (i. e., abiotically), the high efficiency formation of carbon-carbon bonds remains a significant challenge for the field of chemistry and purely chemical approaches to liquid fuels are not currently economically feasible at scale.

Instead, the production of liquid fuels relies primarily on terrestrial photosynthe­sis, in which solar radiation is assimilated through Photosystems I and II, and the captured energy is used to reduce and fix carbon through the Calvin-Benson — Bassham (CBB) cycle. In the CBB cycle, inorganic carbon is converted to glyceral — dehyde-3-phosphate; although this intermediate is the source of reduced carbon for myriad products through both primary and secondary metabolism, it is primarily converted to glucose and polymerized to various structural and storage polymers. These photosynthetic products are converted to liquid fuels either fermentatively or thermally, producing a variety of fuel molecules.

The overall efficiency of this process—from solar photons to liquid fuel— depends on the nature of both the photosynthetic organism and the means of conversion, but is certainly less than 1%. While photosynthesis is operationally facile-plants and photosynthetic organisms are autonomous-the process competes with other forms of agriculture for resources, in particular land, fresh water and essential nutri­ents (NPK and trace metals). The scalable, sustainable production of liquid fuels would be greatly enhanced by the development of processes that do not share agri­cultural factors of production and that offer greater conversion efficiencies than those of photosynthesis.

In 2009, the US Department of Energy (DOE) Advanced Research Projects Agency-Energy (ARPA-E) created the Electrofuels program to explore nonphoto­synthetic autotrophic organisms for the conversion of renewable energy to energy dense liquid fuels for transportation [2] . The approach expands the boundaries of traditional biofuels and could achieve dramatically higher overall conversion efficiencies while providing massive reductions in both arable land and water usage. If a series of technical challenges can be overcome, an Electrofuels system could have a higher utilization of refinery capacity due to feedstock flexibility and be free from fluctuations in feedstock supply and cost inherent to photosynthetic biofuels approaches. To achieve this vision, the Electrofuels program leverages foundational work in microbiology, genomics, metabolic engineering, and synthetic biology to create microorganisms that assimilate energy, fix inorganic carbon, and produce fuel molecules without photosynthesis (Fig. 1).

In this article, we describe the rationale for the Electrofuels program and con­sider the challenges to the economic viability of the approach.

2 Background

The USA is the global leader in the production of biofuels, in 2010 producing over 13 billion gallons of ethanol, or 9 billion gasoline gallon equivalents (GGE), from corn grain [48] . Further expansion of corn grain ethanol production is now con­strained by diminishing Congressional support for government subsidies (in 2010 the Volumetric Ethanol Excise Tax credit cost taxpayers approximately $6B), grow­ing concerns about the environmental impacts of increased corn production, and perceived impacts of converting food to fuel resources in the face of global popula­tion growth and rising food prices.

In order to address these and other issues, the Departments of Energy, Agriculture and Defense have made significant investments in advanced biofuels—biofuels derived from sustainable, nonfood resources such as agricultural residues, residues from forestry operations, food processing by-products, and municipal solid waste. In the near future, dedicated energy crops that grow on marginal or non-food- production land, including perennial grasses (e. g., switchgrass), woody species (e. g., willow), and aquatic macroalgae (e. g., Saccharina), will add to the supply of biomass feedstocks for the production of fuels.

Despite the many benefits offered by dedicated energy crops, the efficiency of energy capture and transduction by plants is remarkably low, calculated as the ratio

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e-, H2

of incident solar radiation to stored energy in chemical bonds. Under optimized environmental conditions C4 plants can capture up to 6% of the available solar energy, while C3 plants convert a maximum of 4.5% [70]. Including in the calcula­tion the seasonal growth of plants, the diversion of plant matter for growth, and the conversion efficiency of fixed carbon to liquid fuels, overall annual photon-to-fuel efficiency stands at 0.18% for US corn ethanol (Supp Calc 1) and 0.20% for Brazil sugarcane ethanol (Supp Calc 2). As a result, land resources are vastly under — utilitized, even for perennial crops. Additionally, biofuels feedstocks and energy crops have significant competition in open markets, and suffer from price fluctuations that limit the viability of this approach.

Photosynthetic microorganisms represent an alternative to terrestrial plants for the production of biofuels, and several algal and cyanobacterial approaches to fuels have also been developed for use in closed systems [30, 31, 49]. Such microorgan­isms offer several advantages over terrestrial plants, including genetic tractability and the ability to secrete fuel products. Genetic tractability affords the opportunity to directly engineer pathways to produce fuel without the need for secondary pro­cessing, while the ability to secrete obviates the need for harvest or biomass manip­ulation. Still, diurnal and annual sunlight variation makes continuous algal production difficult to control, slows microbial growth, and reduces productivity, each of which results in an increase in the cost of capital and the final fuel product. Fresh water requirements are also potentially problematic, although these concerns can be ameliorated to some extent through the use of salt-tolerant species and aggressive water capture techniques. The deployment at scale of genetically modified microorganisms also raises issues of containment, both to prevent acci­dental release and adventitious infection by wild-type strains.

In its broadest conception, the term “biofuels” implies the action of living organ­isms in one or more of the steps required to reduce inorganic carbon to an energy — dense form: the capture of energy and transduction of that energy to a usable form; the reduction of carbon from the +4 oxidation state; and the elaboration of that reduced carbon into a final fuel molecule. The diversity of the microbial world— and especially of the deep oceans—is staggering: the ocean contains 300,000 times more bacteria than there are stars in the visible universe [59, 61]. Fewer than 1% of these microbes have been identified and fewer than 0.1% of marine microorganisms have been cultured [18, 29] . This astonishing store of diversity offers tremendous opportunities for many branches of science, including energy transduction and car­bon fi xation.

Photosynthesis is but one of the approaches to carbon fixation found in nature; myriad life forms exist in ecological niches where both reduced carbon and light are nonexistent. Such organisms assimilate energy from other energy rich (reduced) species, including H2, H2S, NH3 . and reduced metals ions. A group of so-called electrotrophs are capable of accepting reducing equivalents directly as electric current [39]. Many chemolithoautotrophs use carbon fixation cycles other than the CBB cycle, including the reductive acetyl-CoA, the reductive citric acid, and 3-hydroxypropionate-4-hydroxybutyrate cycles; some of these pathways offer significant advantages over the CBB cycle [5]. Such organisms might serve as factories for the high-efficiency production of liquid fuels from renewable forms of energy.

To address these fundamental questions the Electrofuels program seeks routes to biofuels that surpass the inherent limitations of photosynthesis. At the core of the program are chemolithoautotrophic microorganisms, organisms capable of fixing and reducing inorganic carbon but that derive energy from a variety of inor­ganic substrates. Such microorganisms might produce renewable biofuels from solar electricity, either directly or through the agency of a soluble mediator. This solar electricity could come from photovoltaic cells that even now capture >20% of the total solar spectrum [ 25] ; or the energy could come from other renewable sources (hydro, wind, wave, tidal), or non-fossil-based heat (geothermal, concentrating solar, nuclear). The approaches considered under the Electrofuels program tap the enormous and largely unexplored diversity of the microbial world, and may offer routes to advanced biofuels significantly more efficient, scalable and feedstock- flexible than routes based on photosynthesis.