Biology REVIEW

Review (Narrative) Challenge and Potential of Biofuels from Algae

Michael E. Gordon, PhD; Andrew Thomas Cook, PhD

SUMMARY Algae biofuels could give a viable alternative to fossil fuels; however, this tech- nology has to overcome several of hurdles before it can contend in fuel market and be deployed. The main challenges faced are production management, har- vesting, coproduct development, fuel extraction, strain isolation, nutrient sourc- ing and utilization, refining and residual utilization. The solution to these hurdles is simple but needs time and effort. The affordable growth densities, high growth rates and high oil contents are reasons to invest notable capital to convert alga into biofuels. Furthermore, bioprospecting (in which natural diversi- ty is used to increase ), breeding and classical genetics (for trait iden- tification and improvement), bioengineering (in which synthetic biology is used to improve traits) and fuel are some of the techniques under research that can make algal biofuel competitive with fossil fuel. No technology is without its challenges; same is the case with this new technology, as we lean more to it we discover more challenges and obstacles. Even given these uncertainties, it is believed that fuel production from algae will be widely deployable and priced competitive within the next 7 to 10 years, however we still need expand our un- derstanding of this wonderful organism so we can expand our ability to engineer them for the specific task of developing a brand new energy industry.■

KEYWORDS Algae; Fuel; Biotransformation; Energy; Sustainable development

Sci Insigt. 2016; 2016:e00264. doi:10.15354/si.16.re207

Author Affiliations: Author affili- ations are listed at the end of this article. Correspondence to: Dr. Michael E. Gordon, PhD, Division of Biotechniques, BioFront Co., Liverpool St, Sydney NSW 2000, Australia Email: [email protected]

Copyright © 2016 Insights Publisher. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Gordon & Cook. Biofuels from Algae Review

lgae biofuels could give a viable alternative to fossil current technology is US$300–$2,600, compared with $40– fuels; however, this technology has to overcome $80 (2009) for fossil fuel (4), though all of the estimates for A several hurdles before it can contend in fuel market cost of a barrel of alga oil in specific regions reach as low and be deployed. These challenges are strain identification as $84 (5). The upper estimates are more common and al- and improvement, in terms of oil productivity and crop pro- most like our own estimates, and exclude algal oil from the tection, nutrient and resource allocation and use, and also present fuel market. The challenges and techniques to the production of co-products to enhance the productivity of tackle this economic discrepancy are mentioned in this arti- the whole system. Though there is abundant excitement cle. concerning the potential of algae biofuels, much work con- tinues to be needed in the field. This article discusses major challenges to economic algae biofuels at scale, and im- BENEFITS OF MICROALGAL BIOFUELS proves the main focus of the scientific community to deal with these challenges and move algae biofuels from prom- Microalgae are a diverse cluster of unicellular organisms ise to reality. that have the potential to supply a range of solutions for our liquid transportation fuel needs through variety of avenues.

Algae species grow in a wide selection of aquatic environ- IMPORTANCE AND CHALLENGES OF ALGAL ments, from fresh through saturated saline . Algae use BIOFUELS carbon dioxide, and are liable for over 40 percent of the worldwide carbon fixation, with the bulk of this productivi- The global economy needs fossil hydrocarbons to operate, ty coming from marine microalgae (6). Algae can reproduce from manufacturing plastics and fertilizers to providing the biomass very quickly, with some species doubling in as few energy required for transportation, lighting and heating. as six hours h, and a lot of them exhibiting 2 doublings per With our increasing population and growing economy, day (7). All algae have the capability to supply energy-rich there will be enhanced fuel use. As countries improve their oils, and variety of microalga species are found to naturally gross domestic product per capita, data suggest that their store high oil levels in their total dry biomass (8). For in- fuel use will increase, and competition for these restricted stance, some Botryococcus spp. are known to that have up resources will increase. Additionally, there comes increas- to half of their dry mass stored as long-chain hydrocarbons ing atmospheric carbon dioxide concentration, and therefore (9). With probably innumerable species, algal diversity the potential for vital greenhouse -mediated global cli- provides researchers several choices for distinguishing pro- mate change (1), that currently appears to have an effect on duction strains and conjointly provides sources for genetic all components of the planet. Finally, petroleum, which is data that may be used to improve these production strains. partly derived from ancient alga deposits, is a limited re- The micro algal species being investigated as potential bio- source that may eventually run out or become too costly to fuel producers originate from the groups whose ancestral recover (2). These factors are driving the exploration of re- relationships are considerably broader than most of land newable energy sources which will replace fossil fuels, and plants, providing a wealth of genetic diversity (10). The permit larger access to fuel resources for all nations, while groups most frequently thought of when discussing micro- greatly reducing carbon emissions into the atmosphere. A algae are prymnesiophytes, diatoms, green algae, golden variety of technologies are examined as renewable energy brown, eustigmatophytes and blue-green algae (11), and sources and, though no single strategy is probably going to members from all of these groups are examined as potential supply a complete answer, it looks possible that a mixture fuel production strains. However, it ought to be noted that of ways may be utilized that may considerably make us in- blue-green algae are not considered as algae but a category dependence on fossil fuels (3). The challenge that still re- of photosynthetic bacteria. mains is to develop renewable energy industries that oper- ate sustainably and might be price competitive with existing CHALLENGES FOR ALGAL BIOFUELS energy choices.

The affordable growth densities, high growth rates and high PRESENT ECONOMIC REALITY OF LIQUID oil contents are solid reasons to invest notable capital to convert alga into biofuels. However, for alga to mature as FUELS an economically viable platform to offset fossil oil and, consequently, mitigate carbon dioxide release, there are a There are significant challenges to creating biofuels capable few of hurdles to beat like where and how to grow these of competing with fossil fuel. Certainly, a premium worth is algae for maximized oil extraction and increased fuel pro- bonded for clean fuels (fuels that have a 50 percent lower cessing. The biofuels production chain tells that the main carbon dioxide cradle-to-grave footprint than petroleum); challenges faced are production management, harvesting, but, calculable prices of a barrel of algae-based fuel using coproduct development, fuel extraction, strain isolation,

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Gordon & Cook. Biofuels from Algae Review nutrient sourcing and utilization, refining and residual bio- by efficiencies in gas exchange and a demand for supple- mass utilization. mental cooling (17). Despite the advantage of minimizing contamination and raising productivity, it's still not clear Improving Oil Extraction whether or not PBRs can ever become cost competitive Oil extraction is another challenge that can most simply be with open systems. Despite the growth strategy used, addressed from the engineering aspect. There are 3 major substantial enhancements over current technologies for ex- methods for extracting oil from algae: supercritical carbon tracting oil from and harvesting algae ought to be created, dioxide fluid extraction, oil press/expeller and hexane ex- and coordinated efforts are required to couple improved traction (12). These technologies have all been used suc- production strains with engineering advancements. cessfully; however they are comparatively costly, either in terms of equipment required or energy needed to extract the oil. Luckily, all are amenable to engineering enhancements. NUTRIENT CHALLENGE Once extracted, because crude algal oil is chemically iden- tical to crude oil, the challenges related to alga oil conver- Algae need nutrients, light, water and a carbon supply, most sion to usable liquid fuels are same as those already frequently carbon dioxide, for economic growth. The nutri- managed by crude oil firms, though improved catalysts are ents necessary for growth of algae are phosphorous, nitro- needed to boost gasoline production from bio-oil (13). Ow- gen, iron and sulfur. The nutrient demand necessary for al- ing to these similarities, it appears reasonable to assume gal growth is neglected most of times, algae is pretty effi- that partnership between major oil firms and algal produc- cient at sequestering those nutrients when present in their ing firms are possible, since these firms have expertise in- atmosphere (18). Changes in nutrient load and algae growth creasing downstream processing efficiencies. has been studied extensively in terms of coastal regions and of , however not as heavily in terms of productivity in large-scale cultivation (19). If terrestrial ag- Water Use riculture is a model for a few of the challenges for algal cul- Water is doubtless a significant limiting factor in algal tivation, then providing ample nutrients for large-scale algal growth. Protoctist growth into non arable land requires wa- growth may be a vital challenge. Fertilizer or Micro- and ter; luckily, several of those places have considerable alka- macro-nutrient supplements, or account for vital costs with- line or underneath them, resulting in in the current terrestrial agriculture business (20), and bio- a supply of nonpotable water that's appropriate for growth fuels don't seem to be expected to be an exception. The uti- of the many algal species. Surprisingly, algae maturing in lization of fertilizers has been increasing globally. Sadly, open have water needs per unit area like that of cot- several fertilizer elements are generated from fossil fuels or ton or wheat, however lower than that of corn, to refill the mined and, as such, they're not renewable (21). Algae, just water lost in evaporation (for an outline of water needs of like plants, need sources of phosphorus, nitrogen and potas- terrestrial plants utilized in biofuel production see (14)). It's sium, these are the most important elements of agricultural imperative once considering broad preparation of alga, to fertilizers, and large-scale cultivation can impact these al- think about water use to avoid a future ‘water versus fuel’ ready restricted supplies. Additionally, optimum growth of discussion. Though substantial alkaline reserves are availa- the many algal species needs chelated iron and sulfur. ble, water remains an issue for algal fuel production and is needed to be thought about rigorously as the business ex- pands. CROP PROTECTION

MAKING ALGAL GROWTH AND HARVESTING Much like terrestrial monocultures, pests and pathogens MORE EFFICIENT invade multicellular algal monocultures, and thus, crop pro- tection could be a major challenge to algal pool sustainabil-

ity. Distinctive strains immune to pathogens, together with Improved engineering can create a major impact on alga several different methods, will need to be used. These biofuel production. These enhancements include economi- methods might include engineering specific pest resistance cal methods for nutrient circulation and exposure, and have into production species that have sturdy growth characteris- been reviewed before (15). In brief, there are important tics and important lipid composition. Alternative approach- challenges for engineers to either develop photo bioreactors es might include utilizing multiple species, which may be (PBRs) that are cheap enough for large-scale production, or adequate to the slow spread of specific pests and minimize for engineers and biologists to combine forces to develop crop loss in multicellular algal facilities. species that grow normally in affordable open systems (16).

PBRs have benefits over open systems as they can more efficiently maintain axenic cultures, and may maintain more Competition with Petroleum controlled growth environments, which can result in in- With current estimates of algal-based biofuels starting from crease in productivity; but contained systems are challenged $300–$2,600 USD per barrel acquired by current technolo-

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Gordon & Cook. Biofuels from Algae Review gy, technical hurdles ought to be overcome to reduce this production. This presents an implausible chance, however price. A number of these enhancements will come from im- also a major challenge. To maneuver a species into an ap- proving growth methods and engineering, as mentioned plied pipeline when the initial species identification is com- previously, however enhancements may also come from pleted, vital physiological, biochemical and genetic charac- optimizing the utilization of the whole algal organism. The terization should occur. This characterization includes es- price of a barrel of algal biofuel when production goes to tablishing best growth conditions (i.e nutrient levels, pH large scale is tough to extrapolate from the current small and temperature), growth characterization (i.e., production facilities, system enhancements will definitely growth rate and density of the final culture), and analysis of bring prices down. substance accumulation. In parallel, genomics will offer insight into metabolic pathways that are present in these Growth of Algal Strains species and supply a foundation for future metabolic engi- A key characteristic for deciding the viability of new identi- neering. The United States government-sponsored Aquatic fied algal species as a potential fuel yielding strains is their Species Program provided characterization of hundreds of growth. Growth rates and the growth densities should be species, from the Nineteen Seventies through to the mid- characterized in terms of real-world growth conditions in- 1990s (23). This work has been done in a number of labora- stead of laboratory conditions. These conditions are often tories round the world. However, there remains a section of simulated to some extent in the laboratory by utilization of biology that's mostly undiscovered. General descriptions of appropriate media, combining condition and light regimes; the few algae classes in which one or more species has been but, production estimates so far, that are based on laborato- characterised follow. ry conditions in confined photo bioreactors, will ought to be recalculated for industrially scalable systems before they Breeding and Classical Genetics: Trait Identification and can be thought of as significant source for biofuel produc- Improvement tion, whether they are open ponds or advanced photo biore- Artificial selection of desired traits in agricultural plants has actors. While helpful information can be gained from these most likely been occurring since the dawn of agriculture. kinds of investigations, for assessing true economic viabil- The ancient methods are identification of traits of interest ity, and for alga to be promoted forward from the bench to from the present diversity of the crop species, and selective the pond, all aspects of protoctist physiology should be as- combining these traits through interbreeding. Directed and sessed under real-world conditions, instead of the theoreti- productive breeding of some species of microalgae is feasi- cally results under perfect conditions. ble, however very little is known concerning the sexual Optimization of algal species growth in open ponds is lifecycle of the vast spread of species. This is partially due a key element of reaching economic viability, and remains a to the large diversity within the alga groups, from the dip- serious challenge for the industry. Identifying species that loid diatoms to the haploid chlorophyceae, specific process- grow well in these conditions should be the center of atten- es occurring in one species of algae don't seem to be appli- tion of analysis in all laboratories. Algae can grow in a very cable to other species. However, much more limiting is wide range of temperatures, with growth being restricted general lack of knowledge on the complete lifecycle of a primarily by nutrient accessibility and light. Light provides vast species of alga, principally due to their aqueous lifecy- the energy for carbon fixation, and is changed to energy cle and tiny size. There's potential to improve algae for the through chemical process, providing the building blocks for production biofuels by developing the tools for selective biofuel production. Algae growth rates are typically re- breeding and utilizing these to transfer traits that have an stricted by light penetration into the ponds from light ab- effect on biofuels, between closely related species, or to sorption by the water and self-shading, and these con- enhance specific strains of any one of algal species. These straints are major deciding factors of algal pond depth. The breeding methods have a benefit, in that polygenic traits are identification of production species that have evolved to often transferred between strains; but, nowadays mutagene- capture shorter wavelengths of light, which have larger wa- sis and molecular genetics are at the forefront of algal strain ter depth penetration, will enable e algal pond depth to be improvement. enhanced beyond the present 15–30 cm (22). This can de- crease the overall land needed for algal biofuel production. Bioengineering: Using Synthetic Biology to Improve

Traits POTENTIAL OF BIOFUELS FROM ALGAE Identification of a perfect, non-modified biofuel organism that fits into the current mechanism for harvesting, purifica-

tion and extraction, and is economically viable, is possible; Bioprospecting: Utilizing Natural Diversity to Increase but, a more likely situation is identification of a range of Productivity species in which each have one or some of these desirable Algae are some very diverse organisms that contain thou- traits. These traits, once engineered into a strain, are also sands of well-known species. The good diversity of algal adequate to lead to an economically viable production strain. species provides a large range of starting strains for fuel

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Additional to strain enhancements in fuel production, utiliz- provide biojet fuels or biogasoline (25). For maximizing ing genes known from other algal species might allow im- fuel production and minimizing prices, knowing the precise proved expression of heterologous proteins, which either lipid content in terms of carbon chain length is very im- have immense value as a coproduct, protein or enzymatical- portant, this can result in optimized chemical modification. ly manufacture a high-value coproduct. Both these ways are Lipid composition is often determined through lipid extrac- being investigated to enhance the economics of algae biofu- tion, fractionation and followed by mass spectrum analysis. els.

Improving Fuel Molecules CONCLUSIONS Algae are often harnessed to provide variety of molecules, which will be used for fuels. the present trend is to use the This article discusses methods to create algae-based fuels fatty acids that are naturally made in alga, since several prices competitive with fossil fuel. Bioprospecting is of im- species manufacture these lipids at a considerable propor- portance to spot algal species that have desired traits (e.g. tion of their total mass. Further discussion of lipid produc- growth densities, high lipid content, growth rates, and/or tion in algae can be reviewed elsewhere (24). In short, for the presence of valuable co-products), when growing on fatty acid production, acetyl CoA is changed to malonyl inexpensive media. Despite the potential of this strategy, CoA. Malonyl moiety from the malonyl CoA to an acyl the foremost possible situation is that bioprospecting won't carrier protein (ACP) is transferred by Fatty acid synthase determine species that are price competitive with fossil fuel, (FAS), generating malonyl-ACP. As a result, condensation and subsequent genetic engineering and breeding are going reactions catalyzed by FAS elongate the growing acyl chain, to be needed to bring these strains to economic viability. whereas the fatty acids are modified by elongases and de- The potential of engineering algal cells is starting to be real- saturase. Most alga species accumulate fatty acids with ized, from improvements in lipid biogenesis and rising crop ranging in length of 16–20 carbons, with no to as several as protection, to manufacturing valuable proteins and co- four of five sites of unsaturation. Though for various algal products. No technology is without its challenges; same is species fatty acid composition varies, once extracted, its the case with this new technology, as we lean more to it we chemical composition isn't thought to be totally different discover more challenges and obstacles. Even given these from the chemical composition of mixes presently found in uncertainties, it is believed that fuel production from algae sweet light-weight fossil oil. The enzymes concerned with will be widely deployable and priced competitive within the production of oil are also able to be manipulated; therefore, next 7–10 years, however we still need expand our under- increasing the production oil producing species and also the standing of this wonderful organism so we can expand our ensuing that the oil produced can be is used after being ability to engineer them for the specific task of developing a transesterified as diesel or can endure carbon cracking to brand new energy industry.■

Acquisition, analysis, or interpretation of data: ARTICLE INFORMATION Funding/Support: N/A. Both of authors. Author Affiliations: Division of Ocean Bio- Drafting of the manuscript: Cook. Role of the Funder/Sponsor: The funding Critical revision of the manuscript for im- techniques, BioFront Co., Liverpool St, Sydney supporters had no role in submitting and pub- portant intellectual content: Both of authors. lishing the work. NSW 2000, Australia (Gordon). Department of Statistical analysis: N/A. Environmental Biology, Macquarie University, Obtained funding: N/A. How to Cite This Paper: Gordon ME, Cook AT. Sydney NSW 2109, Australia (Cook). Administrative, technical, or material support: Challenge and potential of biofuels from Al- Gordon. gaet. 2016; 2016:e00264. Author Contributions: Dr. Gordon had full Study supervision: Gordon. Digital Object Identifier (DOI): access to all of the data in the study and takes http://dx.doi.org/10.15354/si.16.re207. responsibility for the integrity of the data and Conflict of Interest Disclosures: All authors the accuracy of the data analysis. declared no competing interests of this manu- Article Submission Information: Received, Study concept and design: Both of authors. script submitted for publication. July 07, 2016; Revised: August 01, 2016; Ac- cepted: August 20, 2016.

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