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Aquatic phototrophs: efficient alternatives to land-based crops for biofuels
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Aquatic phototrophs: efficient alternatives to land-based crops for biofuels G Charles Dismukes1, Damian Carrieri1, Nicholas Bennette1, Gennady M Ananyev1 and Matthew C Posewitz2 To mitigate some of the potentially deleterious environmental and agricultural consequences associated with current landbased-biofuel feedstocks, we propose the use of biofuels derived from aquatic microbial oxygenic photoautotrophs (AMOPs), more commonly known as cyanobacteria, algae, and diatoms. Herein we review their demonstrated productivity in mass culturing and aspects of their physiology that are particularly attractive for integration into renewable biofuel applications. Compared with terrestrial crops, AMOPs are inherently more efficient solar collectors, use less or no land, can be converted to liquid fuels using simpler technologies than cellulose, and offer secondary uses that fossil fuels do not provide. AMOPs pose a new set of technological challenges if they are to contribute as biofuel feedstocks. Address 1 Department of Chemistry and Princeton Environmental Institute, Princeton University, Princeton, NJ 08544, USA 2 Department of Chemistry and Geochemistry, Colorado School of Mines, Golden, CO 80401, USA Corresponding author: Dismukes, G Charles ([email protected])
land-based-biofuel feedstocks, it is possible to include biofuels derived from aquatic microbial oxygenic photoautotrophs (AMOPs), more commonly known as cyanobacteria, algae, and diatoms, into the bioenergy portfolio. The potential of AMOPs as high-yield sources for lipids (20–50% dry wt) and fermentable biomass (starch and glycogen, 20–50% dry wt) was documented in research conducted by the National Renewable Energy Laboratory (NREL) and its contractors within the Aquatic Species Program (NREL–ASP) during the 1980s–1990s [4]. Recent workshops including the NREL–AFOSR workshop on Algal Oil for Jet Fuel Production held in February 2008 concluded that ethanol cannot substitute for energydense diesel for aviation fuels, and the latter demand cannot be met solely by terrestrial crops. Multiple commercial ventures involving collaborations between large oil companies and research institutions have recently emerged to produce biodiesel from AMOPs, including Chevron and NREL, Shell Oil and researchers at the University of Hawaii and other institutions, and British Petroleum and Arizona State University.
Current Opinion in Biotechnology 2008, 19:235–240 This review comes from a themed issue on Energy Biotechnology Edited by Lee R. Lynd Available online 6th June 2008 0958-1669/$ – see front matter # 2008 Elsevier Ltd. All rights reserved. DOI 10.1016/j.copbio.2008.05.007
Introduction The scientific community has recently focused considerable attention on developing viable renewable biofuels as leading alternatives to fossil energy in order to address the issue of global warming. However, recent headlines have generally condemned biofuels (e.g. [1]) owing to their potential to drive up food prices and exacerbate CO2 release through the forced clearing of natural ecosystems, which are as effective or more efficient in capturing CO2 [2,3]. The topic is emotionally charged, with divisive positions held by laypeople and scientists alike, making policy choices controversial and their economic and environmental outcomes potentially devastating. In order to minimize the potentially deleterious environmental and agricultural consequences associated with current www.sciencedirect.com
Herein we summarize the demonstrated productivity of mass-cultured AMOPs and examine both their potential and their limitations for the production of biomass and biofuel precursors. We consider options for their largescale cultivation in marine or non-arable lands that would augment or even enable a transition away from conventional biomass grown on farmlands.
Current commercial and laboratory productivity of various AMOPs There is extensive worldwide experience in commercial scale growth of food-grade AMOPS [5,6]. This experience is limited to the 5 ha scale using open ponds. Growth data from NREL and other reliable sources are summarized in Table 1 and S1 listing the solar biomass productivity (dry metric tons/ha yr) for selected AMOPS grown year-round at solar fluxes common to the American southwest, in stirred open pond reactors at (suboptimal) ambient temperatures, and with or without CO2 supplementation. Compared with genetic hybrid strains of corn grain, the annual biomass yield from native strains of AMOPs is 5.4– 10 fold greater, while the comparable gain for switchgrass is 2.5 (fertilized arable land) to 10 fold (fertilized mixed prairie grasses, footnote a, Table 1). We have converted these biomass yields to raw energy content (GJ/ha yr; footnote f, Table 1). The range for AMOPs is 700–1550, Current Opinion in Biotechnology 2008, 19:235–240
236 Energy Biotechnology
Table 1 Biomass and energy productivities of land-based plants and mass-cultured aquatic microbial oxygenic phototrophs (algae and cyanobacteria) Outdoor, [1_TD$IF]solar demonstrated values (except in parentheses)
Corn [2_TD$IF]grain
Sugarcane
Switchgrass and mixed prairie grasses
Productivity (dry metric tons/ha [4_TD$IF] yr) Productivity raw energy (GJ/ha yr) e Components [8_TD$IF]Nonrecalcitrant [9_TD$IF]carbohydrates (%) Lipids (%) Protein (%) Water usage (L/dry kg) Water [1_TD$IF]usage per energy (L/MJ) e
7 [25] 120 [25] [25,29] 70 4.5–6 6–12 565 [25] 33
73–87 [25] 1230–1460 [25] [25] 30 13
3.6–15[5_TD$IF]a [26,27] 61–255 [27] [25,30] 4.5–11.5 1–1.6
2.7 [28] 73 [28] [31]
89–118 [33] 5–7
50 [34] 3
3390 [35] 200
Rape [3_TD$IF]seeds
42
Tetraselmis suecica
Arthrospira (Spirulina) species
38–69[6_TD$IF]b [4] 700–1550[7_TD$IF]f [4] (11)g–(47) h (23)[10_TD$IF]g–(15) h (68)g–(28) h 310–570 [4] 18–34
27c, 60–70 d 550, 1230–1435 [6,8,32] 15g–(50) h 5g–(13) h 72g–(27) h
a
Mixed prairie grass data reported here involve field burning of the annual crop rather than harvesting, thereby enhancing productivity by selffertilization at the cost of eliminating biomass utilization (zero yield). [12_TD$IF]b Lower number demonstrated full year, upper number demonstrated in summer months in New Mexico. Monoraphidium minutum (MONOR2) was also used for growth experiments. [13_TD$IF]c Food grade in Mexico [6] or grown on seawater and urea rather than standard media in Italy [36]. [14_TD$IF]d With heated ponds, control of photoinhibition, and proper harvest timings [37]. [15_TD$IF]e Assuming heat of combustion, theoretical maximum energy content. [16_TD$IF]f Assuming heat of combustion energy similar to A. maxima 2 kJ/g (A. maxima combustion energy = 20.5 kJ/g, our measurement, unpublished). [17_TD$IF]g For nutrient sufficient conditions. [18_TD$IF]h For nutrient deplete conditions (low [19_TD$IF]nitrogen or phosphorous; silicon can also be depleted in diatoms (not listed here). Under these conditions, suboptimal growth conditions (not reported in this table) are expected [4].
achieved because they are – for multiple reasons – inherently more efficient solar energy converters. Firstly, the intrinsic solar energy conversion efficiency (ECE)1 is greater for AMOPs at 3–9% (observed in the field) [12,13] versus a theoretical maximum of 2.4% for C3 crops (switchgrass) and 3.7% for C4 crops (maize) across a full growing season based on calculated solar radiation intercepted by the leaf canopy [14]. The disparity in favor of AMOPs is even larger with increasing solar intensity, where multiple photoprotection mechanisms intervene that differentially protect terrestrial plants by diverting absorbed light to heat. Water and thermal stresses play major roles in exacerbating photoinhibition in terrestrial plants but are much less important in AMOPs [15,16]. Secondly, AMOPs thrive across a greater range of light environments, with respect to mean photon flux, than do higher plants. Thirdly, AMOPs are full canopy absorbers, having superior light capture efficiencies when integrated over their growth cycle (1–3 doublings day 1) compared with slower growing, sparsely seeded plants that attain full canopy for only a fraction of the year. Depending on the cultivation strategy, strain selection and location, AMOPs can be used to harvest solar energy using a dedicated culturing footprint year-round, in contrast to the majority of current agricultural crops. Improvement in ECE is one area of research needed for all photosynthetic crops. Preliminary research in this regard suggests that ECE
which can be compared with conventional biofuels: 120 for corn grain and 61–255 for switchgrass or mixed prairie grasses. This 6–12 fold increased energy yield for AMOPs is one of several advantages they offer as sources of biomass and biofuel precursors relative to terrestrial plants. Examples: The hypercarbonate-requiring cyanobacterium Arthrospira (Spirulina) maxima thrives in volcanic soda lakes at pH 9.5–11 and up to 1.2 M sodium carbonate, conditions that prevent most competing microbes. It grows to high cell densities in open pond reactors with low microbial contamination [7,8]. Related strains are grown for the health– food industry, where productivity rates reach 27 dry metric tons/ha yr (annual in Mexico). Heated outdoor open ponds have demonstrated annual yields of 60–70 dry metric tons/ha yr (in Israel) (Table 1). A salt-tolerant green algal strain of the genus Tetraselmis originating from the Great Salt Lake thrives at 6% salt, and a marine strain of this genus has been cultivated in open ponds in New Mexico with productivity rates of 38 (annual) and 69 (summer) dry metric tons/ha yr (Table 1). This value increases to 146 if grown in shallow outdoor flume bioreactors in summer months in Hawaii [9]. Several reports have corroborated considerably higher yields in closed, thermostated photobioreactors with optimal light conditions. Several aspects of AMOP physiology are relevant for evaluating their possible integration into renewable biofuel applications [10,11]: 1. Superior solar energy yields. The 6–12 fold energy yield advantage of AMOPs versus terrestrial plants can be Current Opinion in Biotechnology 2008, 19:235–240
1
Measured as energy released by complete combustion of biomass divided by the solar energy absorbed. www.sciencedirect.com
Aquatic phototrophs Dismukes et al. 237
can be enhanced by reducing chlorophyll antenna size for more efficient light utilization [17,18]. 2. Lack of recalcitrant biopolymers. AMOPS are buoyant aquatic microcells that do not require structural biopolymers essential for higher plant growth in terrestrial environments. The resulting absence of cellulose, hemi-cellulose, and lignin eliminates the need for pretreatments to breakdown cellulytic products (hydrolysis by acid and enzymatic cleavage by cellulase) and lowers the reactor temperature of their subsequent fermentation. This simplification of process and improved conversion efficiency bypass the most costly and inefficient steps for conversion of cellulose to ethanol [19]. Lignin cannot now be readily transformed to transportation fuels. This advantage of simpler biomass composition needs to be weighed against the added cost for removal of excess water in cases where, for example, preprocessing for lipid extraction requires dry cell mass. 3. Metabolic and ecological diversity. AMOPs exhibit enormous ecological, genotypic, and metabolic diversities with over 4000 distinct species of cyanobacteria (prokaryotes) and a comparable number of unicellular algae (eukaryotes) classified thus far. This diversity allows selection of genera/species that are adapted for growth in locally available aquifers (marine, hypersaline, thermophilic and freshwater), or have morphological features that allow cost-effective harvesting (filamentous, buoyant, or aggregate), or possess anaerobic metabolisms that enable production of hydrogen, ethanol, and/or organic acids by autofermentation before final biomass conversion/utilization. Moreover, several AMOPs are well characterized and advanced genetic techniques exist for engineering of strains with already demonstrated progress toward optimizing bioenergy production and identifying promising targets by genomic techniques [20]. 4. Biosynthetic control of chemical composition by nutrient and environmental stresses. The ability of AMOPs to direct the majority of photosynthetic reductant into metabolic pathways that synthesize the most amenable bioenergy precursors starch, glycogen, and lipids rather than cellulose and lignin offers a potential processing simplification and gain in the overall energy yield. Most AMOPs are rich sources of proteins when grown under nutrient replete conditions (Table 1). Unlike higher plants, they can be easily induced to alter their composition using nutritional or environmental stresses. Using these techniques, protein content can be converted to energy storage compounds such as carbohydrates (starch, glycogen, polysaccharides) or higher energy lipids (fatty acids, monoacylglycerols, diacylglycerols and triacylglycerols) in a speciesdependent manner. Many microalgae have the ability to produce substantial amounts (e.g. 20–50% dry cell weight) of triacylglycerols (TAG) as a storage lipid under photo-oxidative stress or other adverse environwww.sciencedirect.com
mental conditions [10]. Lipids contain twice the energy stored per C atom than do carbohydrates, which translates directly into a twofold increase in fuel energy content. Depletion of nitrate (–N), silicate (–Si) or phosphate (–P) from the growth medium has been shown to produce twofold to threefold increase in the relative amounts of starch (glycogen in cyanobacteria) or lipids in many AMOPs [4]. This shift in composition enables a growth strategy for enriching cells in specific energy precursors before cell harvesting. By judicious selection of the initial nutrient concentrations for growth, batch culturing can take advantage of this transition from rapidly-growing, protein-rich young cells to mature cells that are richer in biofuel precursors. 5. Contamination. The NREL–ASP project found that contamination of non-native AMOPs by native AMOP strains is a serious problem facing large-scale cultivation in open reactors [4]. Their high nutritional value also makes them easy targets for predation. Suggested improvements require the selection of robust strains of AMOPs that thrive in high salt concentrations or high pH where microbial contamination is low. These conditions are unsuitable for many bacterial and fungal contaminants [4,5]. Common natural aquifers that harbor robust AMOP communities include the Great Salt Lake and volcanic soda lakes. One such example, A. maxima, possesses the highest solar energy conversion of any measured oxygenic phototroph (AMOPs and terrestrial plants) [21]. Much of this energy is not used for biomass accumulation, but rather for excess ATP production essential for maintaining osmotic balance at high ionic strength. Consequently, such environments represent a trade-off between productivity and species competition. 6. Water usage and harvesting. Table 1 summarizes the water usage per unit weight biomass and per energy unit produced after annual recycling for AMOP cultivation based upon commercial scale open pond reactors. The volume is comparable or lower than that required to grow maize at an equivalent weight of corn grain, eightfold less than rape cultivation on fertilized land, but 6–10 fold more than needed by unfertilized switchgrass on non-arable land. This highlights a main limitation of AMOPS in comparison with switchgrass. The natural water source for many AMOPs is seawater, which is plentiful and can be substituted by deep saline aquifers in the interior [22] or nutrient-laden agricultural wastewater [23], while terrestrial biofuel crops require freshwater. Mass cultivation of AMOPs is currently done at coastal locations in open ponds and interior locations in desert climate using nutrientsupplemented seawater where evaporative losses dominate. Scale-up is under consideration using offshore sites bounded by wind-farm pylons. AMOPs grown in hypersaline aquifers impose additional constraints on water. However, the additional high Current Opinion in Biotechnology 2008, 19:235–240
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Figure 1
Areas needed for cultivation of three biomass sources. Each box represents the area needed to produce a sufficient amount of biomass to convert to liquid fuel to displace all gasoline used in the USA (2006 figures) on an energy basis. Data taken from ref 24.
salt and bicarbonate concentrations needed for their culturing could be obtained from the concentrated brine discarded during desalination of seawater. Water desalination facilities are in high demand globally and projections forecast escalating expansion [22]. The synergism of these resources should be an important consideration for future co-development of water desalination and biofuels from AMOPs. Harvesting of mass-cultured AMOPs constitutes a major technical challenge for unicellular types that do not aggregate. These cell morphologies require some form of chemically induced coagulation or forced flotation using compressed air bubbles that are costly or have environmental trade-offs [23]. However, many strains of AMOPs are filamentous that enable separation by simple straining, or contain gas vacuoles to aid in buoyancy, which allows isolation by skimming. For this reason this class of AMOPs is often chosen for commercial farming. Genetic tools are limited for filamentous AMOPs and need to be developed to allow this class of easy-to-harvest cell types to be more fully exploited for bioenergy farming. 7. Valuable by-products. A significant fraction of the residual biomass following lipid and carbohydrate extraction is protein. This is expected to pass through largely unaltered by the mild conditions used for fermentation to ethanol and possibly those for lipid Current Opinion in Biotechnology 2008, 19:235–240
extraction for biodiesel conversion. Taking a holistic view of AMOP energy processing, these by-products can be directed toward secondary markets, such as use of residual protein as animal feed (and mineral ashes in the case of gasification).
Land versus aquatic biofuels Figure 1 compares the areas needed for three biomass sources. Data for corn grain and switchgrass/mixed prairie grasses [24] are compared with AMOPs. Each box represents the area needed to produce a sufficient amount of biomass to produce liquid fuel to displace all gasoline used in the USA (2006 figures) on an energy basis. For corn and switchgrass, carbohydrate/cellulose content would be fermented to ethanol. This estimate does not include the requisite energy input for growth or ethanol production (about 80% of the energy in corn ethanol, assuming an optimistic figure). The switchgrass area assumes yet unspecified optimal technology for both the production of cellulytic feedstock and its subsequent fermentation to ethanol. For AMOPs, either fermentation to ethanol or conversion to biodiesel is an option depending upon species. We show two boxes at 30 and 70% to illustrate the upper and lower ranges expected for the conversion efficiency and relative energy content of diesel versus ethanol. The overall solar energy conversion to biofuel works out to about 0.05% for www.sciencedirect.com
Aquatic phototrophs Dismukes et al. 239
solar to ethanol from corn grain and roughly 0.5% for switchgrass to ethanol [24]. Comparatively, this value is about 0.5–1% for AMOPs to ethanol or biodiesel (ignoring the fossil fuel input in all cases).
Conclusions To realize the potential of AMOPs, significant research efforts are underway to overcome several bottlenecks hindering their widespread use, which include (a) cell harvesting, (b) culturing strategies that maintain relative monocultures and promote high photosynthetic conversion efficiencies, (c) metabolic control (either physiologically or genetically) of the accumulated biopolymers, (d) access to suitable aquifers, and (e) advanced biorefining techniques to isolate biofuel precursors in a cost-effective manner. Although obstacles to deployment exist, AMOPs offer a wide range of alternatives for ‘designer’ biomass and biofuel precursors, and research into the use of AMOPs as viable bioenergy feedstocks is in its infancy relative to more thoroughly investigated terrestrial feedstocks. They are inherently more efficient solar collectors, use less or no land, can be converted to liquid fuels using simpler technologies than cellulose, and have secondary uses that fossil fuels do not provide. Current research efforts into the use of AMOPs as bioenergy feedstocks will continue to leverage several of the assets inherent in these organisms. The coming years will probably provide new insights into novel ways to use AMOPs in economically viable biofuel processes.
Acknowledgements The authors thank Mike Seibert for help with a figure, Bob Williams and Lee Lynd for stimulating discussions, and Patricia Brletic and Lindsay Leone for collaboration with combustion calorimetry experiments. GCD and MCP research is supported by an AFOSR-MURI grant.
Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at doi:10.1016/j.copbio.2008.05.007.
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Aquatic phototrophs: efficient alternatives to land-based crops for biofuels G Charles Dismukes1, Damian Carrieri1, Nicholas Bennette1, Gennady M Ananyev1 and Matthew C Posewitz2 To mitigate some of the potentially deleterious environmental and agricultural consequences associated with current landbased-biofuel feedstocks, we propose the use of biofuels derived from aquatic microbial oxygenic photoautotrophs (AMOPs), more commonly known as cyanobacteria, algae, and diatoms. Herein we review their demonstrated productivity in mass culturing and aspects of their physiology that are particularly attractive for integration into renewable biofuel applications. Compared with terrestrial crops, AMOPs are inherently more efficient solar collectors, use less or no land, can be converted to liquid fuels using simpler technologies than cellulose, and offer secondary uses that fossil fuels do not provide. AMOPs pose a new set of technological challenges if they are to contribute as biofuel feedstocks. Address 1 Department of Chemistry and Princeton Environmental Institute, Princeton University, Princeton, NJ 08544, USA 2 Department of Chemistry and Geochemistry, Colorado School of Mines, Golden, CO 80401, USA Corresponding author: Dismukes, G Charles ([email protected])
land-based-biofuel feedstocks, it is possible to include biofuels derived from aquatic microbial oxygenic photoautotrophs (AMOPs), more commonly known as cyanobacteria, algae, and diatoms, into the bioenergy portfolio. The potential of AMOPs as high-yield sources for lipids (20–50% dry wt) and fermentable biomass (starch and glycogen, 20–50% dry wt) was documented in research conducted by the National Renewable Energy Laboratory (NREL) and its contractors within the Aquatic Species Program (NREL–ASP) during the 1980s–1990s [4]. Recent workshops including the NREL–AFOSR workshop on Algal Oil for Jet Fuel Production held in February 2008 concluded that ethanol cannot substitute for energydense diesel for aviation fuels, and the latter demand cannot be met solely by terrestrial crops. Multiple commercial ventures involving collaborations between large oil companies and research institutions have recently emerged to produce biodiesel from AMOPs, including Chevron and NREL, Shell Oil and researchers at the University of Hawaii and other institutions, and British Petroleum and Arizona State University.
Current Opinion in Biotechnology 2008, 19:235–240 This review comes from a themed issue on Energy Biotechnology Edited by Lee R. Lynd Available online 6th June 2008 0958-1669/$ – see front matter # 2008 Elsevier Ltd. All rights reserved. DOI 10.1016/j.copbio.2008.05.007
Introduction The scientific community has recently focused considerable attention on developing viable renewable biofuels as leading alternatives to fossil energy in order to address the issue of global warming. However, recent headlines have generally condemned biofuels (e.g. [1]) owing to their potential to drive up food prices and exacerbate CO2 release through the forced clearing of natural ecosystems, which are as effective or more efficient in capturing CO2 [2,3]. The topic is emotionally charged, with divisive positions held by laypeople and scientists alike, making policy choices controversial and their economic and environmental outcomes potentially devastating. In order to minimize the potentially deleterious environmental and agricultural consequences associated with current www.sciencedirect.com
Herein we summarize the demonstrated productivity of mass-cultured AMOPs and examine both their potential and their limitations for the production of biomass and biofuel precursors. We consider options for their largescale cultivation in marine or non-arable lands that would augment or even enable a transition away from conventional biomass grown on farmlands.
Current commercial and laboratory productivity of various AMOPs There is extensive worldwide experience in commercial scale growth of food-grade AMOPS [5,6]. This experience is limited to the 5 ha scale using open ponds. Growth data from NREL and other reliable sources are summarized in Table 1 and S1 listing the solar biomass productivity (dry metric tons/ha yr) for selected AMOPS grown year-round at solar fluxes common to the American southwest, in stirred open pond reactors at (suboptimal) ambient temperatures, and with or without CO2 supplementation. Compared with genetic hybrid strains of corn grain, the annual biomass yield from native strains of AMOPs is 5.4– 10 fold greater, while the comparable gain for switchgrass is 2.5 (fertilized arable land) to 10 fold (fertilized mixed prairie grasses, footnote a, Table 1). We have converted these biomass yields to raw energy content (GJ/ha yr; footnote f, Table 1). The range for AMOPs is 700–1550, Current Opinion in Biotechnology 2008, 19:235–240
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Table 1 Biomass and energy productivities of land-based plants and mass-cultured aquatic microbial oxygenic phototrophs (algae and cyanobacteria) Outdoor, [1_TD$IF]solar demonstrated values (except in parentheses)
Corn [2_TD$IF]grain
Sugarcane
Switchgrass and mixed prairie grasses
Productivity (dry metric tons/ha [4_TD$IF] yr) Productivity raw energy (GJ/ha yr) e Components [8_TD$IF]Nonrecalcitrant [9_TD$IF]carbohydrates (%) Lipids (%) Protein (%) Water usage (L/dry kg) Water [1_TD$IF]usage per energy (L/MJ) e
7 [25] 120 [25] [25,29] 70 4.5–6 6–12 565 [25] 33
73–87 [25] 1230–1460 [25] [25] 30 13
3.6–15[5_TD$IF]a [26,27] 61–255 [27] [25,30] 4.5–11.5 1–1.6
2.7 [28] 73 [28] [31]
89–118 [33] 5–7
50 [34] 3
3390 [35] 200
Rape [3_TD$IF]seeds
42
Tetraselmis suecica
Arthrospira (Spirulina) species
38–69[6_TD$IF]b [4] 700–1550[7_TD$IF]f [4] (11)g–(47) h (23)[10_TD$IF]g–(15) h (68)g–(28) h 310–570 [4] 18–34
27c, 60–70 d 550, 1230–1435 [6,8,32] 15g–(50) h 5g–(13) h 72g–(27) h
a
Mixed prairie grass data reported here involve field burning of the annual crop rather than harvesting, thereby enhancing productivity by selffertilization at the cost of eliminating biomass utilization (zero yield). [12_TD$IF]b Lower number demonstrated full year, upper number demonstrated in summer months in New Mexico. Monoraphidium minutum (MONOR2) was also used for growth experiments. [13_TD$IF]c Food grade in Mexico [6] or grown on seawater and urea rather than standard media in Italy [36]. [14_TD$IF]d With heated ponds, control of photoinhibition, and proper harvest timings [37]. [15_TD$IF]e Assuming heat of combustion, theoretical maximum energy content. [16_TD$IF]f Assuming heat of combustion energy similar to A. maxima 2 kJ/g (A. maxima combustion energy = 20.5 kJ/g, our measurement, unpublished). [17_TD$IF]g For nutrient sufficient conditions. [18_TD$IF]h For nutrient deplete conditions (low [19_TD$IF]nitrogen or phosphorous; silicon can also be depleted in diatoms (not listed here). Under these conditions, suboptimal growth conditions (not reported in this table) are expected [4].
achieved because they are – for multiple reasons – inherently more efficient solar energy converters. Firstly, the intrinsic solar energy conversion efficiency (ECE)1 is greater for AMOPs at 3–9% (observed in the field) [12,13] versus a theoretical maximum of 2.4% for C3 crops (switchgrass) and 3.7% for C4 crops (maize) across a full growing season based on calculated solar radiation intercepted by the leaf canopy [14]. The disparity in favor of AMOPs is even larger with increasing solar intensity, where multiple photoprotection mechanisms intervene that differentially protect terrestrial plants by diverting absorbed light to heat. Water and thermal stresses play major roles in exacerbating photoinhibition in terrestrial plants but are much less important in AMOPs [15,16]. Secondly, AMOPs thrive across a greater range of light environments, with respect to mean photon flux, than do higher plants. Thirdly, AMOPs are full canopy absorbers, having superior light capture efficiencies when integrated over their growth cycle (1–3 doublings day 1) compared with slower growing, sparsely seeded plants that attain full canopy for only a fraction of the year. Depending on the cultivation strategy, strain selection and location, AMOPs can be used to harvest solar energy using a dedicated culturing footprint year-round, in contrast to the majority of current agricultural crops. Improvement in ECE is one area of research needed for all photosynthetic crops. Preliminary research in this regard suggests that ECE
which can be compared with conventional biofuels: 120 for corn grain and 61–255 for switchgrass or mixed prairie grasses. This 6–12 fold increased energy yield for AMOPs is one of several advantages they offer as sources of biomass and biofuel precursors relative to terrestrial plants. Examples: The hypercarbonate-requiring cyanobacterium Arthrospira (Spirulina) maxima thrives in volcanic soda lakes at pH 9.5–11 and up to 1.2 M sodium carbonate, conditions that prevent most competing microbes. It grows to high cell densities in open pond reactors with low microbial contamination [7,8]. Related strains are grown for the health– food industry, where productivity rates reach 27 dry metric tons/ha yr (annual in Mexico). Heated outdoor open ponds have demonstrated annual yields of 60–70 dry metric tons/ha yr (in Israel) (Table 1). A salt-tolerant green algal strain of the genus Tetraselmis originating from the Great Salt Lake thrives at 6% salt, and a marine strain of this genus has been cultivated in open ponds in New Mexico with productivity rates of 38 (annual) and 69 (summer) dry metric tons/ha yr (Table 1). This value increases to 146 if grown in shallow outdoor flume bioreactors in summer months in Hawaii [9]. Several reports have corroborated considerably higher yields in closed, thermostated photobioreactors with optimal light conditions. Several aspects of AMOP physiology are relevant for evaluating their possible integration into renewable biofuel applications [10,11]: 1. Superior solar energy yields. The 6–12 fold energy yield advantage of AMOPs versus terrestrial plants can be Current Opinion in Biotechnology 2008, 19:235–240
1
Measured as energy released by complete combustion of biomass divided by the solar energy absorbed. www.sciencedirect.com
Aquatic phototrophs Dismukes et al. 237
can be enhanced by reducing chlorophyll antenna size for more efficient light utilization [17,18]. 2. Lack of recalcitrant biopolymers. AMOPS are buoyant aquatic microcells that do not require structural biopolymers essential for higher plant growth in terrestrial environments. The resulting absence of cellulose, hemi-cellulose, and lignin eliminates the need for pretreatments to breakdown cellulytic products (hydrolysis by acid and enzymatic cleavage by cellulase) and lowers the reactor temperature of their subsequent fermentation. This simplification of process and improved conversion efficiency bypass the most costly and inefficient steps for conversion of cellulose to ethanol [19]. Lignin cannot now be readily transformed to transportation fuels. This advantage of simpler biomass composition needs to be weighed against the added cost for removal of excess water in cases where, for example, preprocessing for lipid extraction requires dry cell mass. 3. Metabolic and ecological diversity. AMOPs exhibit enormous ecological, genotypic, and metabolic diversities with over 4000 distinct species of cyanobacteria (prokaryotes) and a comparable number of unicellular algae (eukaryotes) classified thus far. This diversity allows selection of genera/species that are adapted for growth in locally available aquifers (marine, hypersaline, thermophilic and freshwater), or have morphological features that allow cost-effective harvesting (filamentous, buoyant, or aggregate), or possess anaerobic metabolisms that enable production of hydrogen, ethanol, and/or organic acids by autofermentation before final biomass conversion/utilization. Moreover, several AMOPs are well characterized and advanced genetic techniques exist for engineering of strains with already demonstrated progress toward optimizing bioenergy production and identifying promising targets by genomic techniques [20]. 4. Biosynthetic control of chemical composition by nutrient and environmental stresses. The ability of AMOPs to direct the majority of photosynthetic reductant into metabolic pathways that synthesize the most amenable bioenergy precursors starch, glycogen, and lipids rather than cellulose and lignin offers a potential processing simplification and gain in the overall energy yield. Most AMOPs are rich sources of proteins when grown under nutrient replete conditions (Table 1). Unlike higher plants, they can be easily induced to alter their composition using nutritional or environmental stresses. Using these techniques, protein content can be converted to energy storage compounds such as carbohydrates (starch, glycogen, polysaccharides) or higher energy lipids (fatty acids, monoacylglycerols, diacylglycerols and triacylglycerols) in a speciesdependent manner. Many microalgae have the ability to produce substantial amounts (e.g. 20–50% dry cell weight) of triacylglycerols (TAG) as a storage lipid under photo-oxidative stress or other adverse environwww.sciencedirect.com
mental conditions [10]. Lipids contain twice the energy stored per C atom than do carbohydrates, which translates directly into a twofold increase in fuel energy content. Depletion of nitrate (–N), silicate (–Si) or phosphate (–P) from the growth medium has been shown to produce twofold to threefold increase in the relative amounts of starch (glycogen in cyanobacteria) or lipids in many AMOPs [4]. This shift in composition enables a growth strategy for enriching cells in specific energy precursors before cell harvesting. By judicious selection of the initial nutrient concentrations for growth, batch culturing can take advantage of this transition from rapidly-growing, protein-rich young cells to mature cells that are richer in biofuel precursors. 5. Contamination. The NREL–ASP project found that contamination of non-native AMOPs by native AMOP strains is a serious problem facing large-scale cultivation in open reactors [4]. Their high nutritional value also makes them easy targets for predation. Suggested improvements require the selection of robust strains of AMOPs that thrive in high salt concentrations or high pH where microbial contamination is low. These conditions are unsuitable for many bacterial and fungal contaminants [4,5]. Common natural aquifers that harbor robust AMOP communities include the Great Salt Lake and volcanic soda lakes. One such example, A. maxima, possesses the highest solar energy conversion of any measured oxygenic phototroph (AMOPs and terrestrial plants) [21]. Much of this energy is not used for biomass accumulation, but rather for excess ATP production essential for maintaining osmotic balance at high ionic strength. Consequently, such environments represent a trade-off between productivity and species competition. 6. Water usage and harvesting. Table 1 summarizes the water usage per unit weight biomass and per energy unit produced after annual recycling for AMOP cultivation based upon commercial scale open pond reactors. The volume is comparable or lower than that required to grow maize at an equivalent weight of corn grain, eightfold less than rape cultivation on fertilized land, but 6–10 fold more than needed by unfertilized switchgrass on non-arable land. This highlights a main limitation of AMOPS in comparison with switchgrass. The natural water source for many AMOPs is seawater, which is plentiful and can be substituted by deep saline aquifers in the interior [22] or nutrient-laden agricultural wastewater [23], while terrestrial biofuel crops require freshwater. Mass cultivation of AMOPs is currently done at coastal locations in open ponds and interior locations in desert climate using nutrientsupplemented seawater where evaporative losses dominate. Scale-up is under consideration using offshore sites bounded by wind-farm pylons. AMOPs grown in hypersaline aquifers impose additional constraints on water. However, the additional high Current Opinion in Biotechnology 2008, 19:235–240
238 Energy Biotechnology
Figure 1
Areas needed for cultivation of three biomass sources. Each box represents the area needed to produce a sufficient amount of biomass to convert to liquid fuel to displace all gasoline used in the USA (2006 figures) on an energy basis. Data taken from ref 24.
salt and bicarbonate concentrations needed for their culturing could be obtained from the concentrated brine discarded during desalination of seawater. Water desalination facilities are in high demand globally and projections forecast escalating expansion [22]. The synergism of these resources should be an important consideration for future co-development of water desalination and biofuels from AMOPs. Harvesting of mass-cultured AMOPs constitutes a major technical challenge for unicellular types that do not aggregate. These cell morphologies require some form of chemically induced coagulation or forced flotation using compressed air bubbles that are costly or have environmental trade-offs [23]. However, many strains of AMOPs are filamentous that enable separation by simple straining, or contain gas vacuoles to aid in buoyancy, which allows isolation by skimming. For this reason this class of AMOPs is often chosen for commercial farming. Genetic tools are limited for filamentous AMOPs and need to be developed to allow this class of easy-to-harvest cell types to be more fully exploited for bioenergy farming. 7. Valuable by-products. A significant fraction of the residual biomass following lipid and carbohydrate extraction is protein. This is expected to pass through largely unaltered by the mild conditions used for fermentation to ethanol and possibly those for lipid Current Opinion in Biotechnology 2008, 19:235–240
extraction for biodiesel conversion. Taking a holistic view of AMOP energy processing, these by-products can be directed toward secondary markets, such as use of residual protein as animal feed (and mineral ashes in the case of gasification).
Land versus aquatic biofuels Figure 1 compares the areas needed for three biomass sources. Data for corn grain and switchgrass/mixed prairie grasses [24] are compared with AMOPs. Each box represents the area needed to produce a sufficient amount of biomass to produce liquid fuel to displace all gasoline used in the USA (2006 figures) on an energy basis. For corn and switchgrass, carbohydrate/cellulose content would be fermented to ethanol. This estimate does not include the requisite energy input for growth or ethanol production (about 80% of the energy in corn ethanol, assuming an optimistic figure). The switchgrass area assumes yet unspecified optimal technology for both the production of cellulytic feedstock and its subsequent fermentation to ethanol. For AMOPs, either fermentation to ethanol or conversion to biodiesel is an option depending upon species. We show two boxes at 30 and 70% to illustrate the upper and lower ranges expected for the conversion efficiency and relative energy content of diesel versus ethanol. The overall solar energy conversion to biofuel works out to about 0.05% for www.sciencedirect.com
Aquatic phototrophs Dismukes et al. 239
solar to ethanol from corn grain and roughly 0.5% for switchgrass to ethanol [24]. Comparatively, this value is about 0.5–1% for AMOPs to ethanol or biodiesel (ignoring the fossil fuel input in all cases).
Conclusions To realize the potential of AMOPs, significant research efforts are underway to overcome several bottlenecks hindering their widespread use, which include (a) cell harvesting, (b) culturing strategies that maintain relative monocultures and promote high photosynthetic conversion efficiencies, (c) metabolic control (either physiologically or genetically) of the accumulated biopolymers, (d) access to suitable aquifers, and (e) advanced biorefining techniques to isolate biofuel precursors in a cost-effective manner. Although obstacles to deployment exist, AMOPs offer a wide range of alternatives for ‘designer’ biomass and biofuel precursors, and research into the use of AMOPs as viable bioenergy feedstocks is in its infancy relative to more thoroughly investigated terrestrial feedstocks. They are inherently more efficient solar collectors, use less or no land, can be converted to liquid fuels using simpler technologies than cellulose, and have secondary uses that fossil fuels do not provide. Current research efforts into the use of AMOPs as bioenergy feedstocks will continue to leverage several of the assets inherent in these organisms. The coming years will probably provide new insights into novel ways to use AMOPs in economically viable biofuel processes.
Acknowledgements The authors thank Mike Seibert for help with a figure, Bob Williams and Lee Lynd for stimulating discussions, and Patricia Brletic and Lindsay Leone for collaboration with combustion calorimetry experiments. GCD and MCP research is supported by an AFOSR-MURI grant.
Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at doi:10.1016/j.copbio.2008.05.007.
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