Production of Jet Fuel Range Hydrocarbons as a Coproduct of Algal

Department of Marine Chemistry and Geochemistry, Woods Hole Oceanographic Institution, Woods Hole, Massachusetts 02543, United States. Energy Fuels , ...
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Production of Jet Fuel Range Hydrocarbons as a Coproduct of Algal Biodiesel by Butenolysis of Long-Chain Alkenones Gregory W. O’Neil,*,† Aaron R. Culler,† John R. Williams,† Noah P. Burlow,† Garrett J. Gilbert,† Catherine A. Carmichael,‡ Robert K. Nelson,‡ Robert F. Swarthout,‡ and Christopher M. Reddy‡ †

Department of Chemistry, Western Washington University, Bellingham, Washington 98225, United States Department of Marine Chemistry and Geochemistry, Woods Hole Oceanographic Institution, Woods Hole, Massachusetts 02543, United States

‡

S Supporting Information *

ABSTRACT: Long-chain (35−40 carbons) alkenones are a unique class of lipids biosynthesized by certain species of algae including the industrially grown marine microalgae Isochrysis. Their structures are characterized by a very long linear carbon-chain with trans double bonds and a methyl or ethyl ketone. A method is presented for the isolation of pure alkenones from Isochrysis biomass in parallel with biodiesel production. Yields for the isolated alkenones and biodiesel relative to the starting dry Isochrysis biomass were routinely 3.5 and 12% (w/w), respectively. Alkenones were then converted to smaller hydrocarbon fragments (jetfuel range) by cross-metathesis with 2-butene (butenolysis) using several commercial ruthenium-based metathesis initiators. Butenolysis with the second-generation Hoveyda-Grubbs catalyst occurred rapidly at 4 °C, yielding near quantitative conversion within 30 min to a mixture containing mostly 8-decen-2-one (C10), 2,9-undecadiene (C12), and 2-heptadecene (C17) as both cisand trans isomers based on analysis by comprehensive two-dimensional gas chromatography. temperatures, alkenones are more highly polyunsaturated,13,14 and the proportion of diunsaturated isomers of the C37 methyl alkenone (the so-called “unsaturation index”) has been widely adopted by geochemists as a proxy for past sea surface temperatures.15−20 Isochrysis is one of several species of haptophyte marine microalgae including the widely distributed coccolithophore Emiliania huxleyi and the closely related species Gephyrocapsa oceanica that biosynthesize alkenones and the related alkenoates and alkenes, known collectively as PULCA.21 Alkenones are thought to reside in cytoplasmic lipid bodies and can be more abundant than TAGs especially in the stationary growth phase.21,22 Under N or P limitation, up to 10−20% of cell C in the stationary phase is accumulated as alkenones.23,24 Evolutionarily, alkenones may have been favored over TAGs because their trans double bond geometry provides a more photostable form of energy storage.21 This unusual geometry has also been suggested to contribute to their limited degradation by grazers in surface waters of the ocean.21 Haptophytes had been included in several reviews25−28 and other reports29,30 related to biofuels, but until recently, alkenones were not discussed in any of these studies. In one study of 55 microalgal species for biodiesel production, Isochrysis galbana proved to be average in both biomass yield and lipid productivity.25 Average lipid content in 15 nutrientreplete and nitrogen-deficient cultures was 25 and 29% dry weight, respectively. We were attracted to haptophytes in part because Isochrysis is one of only a select number of algal species farmed industrially,

1. INTRODUCTION Replacing petroleum-derived liquid transportation fuels with alternatives from renewable sources continues to drive a large volume of ongoing research.1 This is in part a response to the United States Environmental Protection Agency Renewable Fuels Standard program that has committed the United States to replacing 36 billion gallons of its transportation fuel with fuel from renewable sources by 2022.2 Plant-based sugars and oils (i.e., carbohydrates and acylglycerols, respectively, plus their derivatives) have dominated the current industry.3 Primary examples include fermentation of sugars to produce ethanol that is often blended with gasoline and the conversion of oils into fatty acid methyl esters (FAMEs) as a drop-in replacement for petrodiesel.4,5 The demand and consumption of these fuels has increased dramatically in recent years, which is linked to rising oil prices, energy and national security concerns, along with other environmental considerations like climate change. Increased usage, however, can create certain food vs fuel controversies when the renewable feedstock is an edible crop.6 In response, various alternatives are being pursued including cellulosic plant biomass7 and algae.8 Nonetheless, the target for these approaches continues to be primarily sugars and oils. Although advances in conversion technologies can provide novel access to fuels from these substrates (e.g., “renewable diesel” by hydrotreatment of oils9), others have emphasized the importance of a diversified portfolio to meet the economic and ecological requirements of sustainable biofuels.10 Recently, we have focused on Isochrysis as they produce a unique and promising class of lipids known as long-chain alkenones.11,12 These compounds are unlike the cis-unsaturated methylene interrupted fatty acid components of triacylglycerols (TAGs) as they typically have two to four trans alkenes occurring at 7-carbon intervals (Figure 1). At colder growth © XXXX American Chemical Society

Received: November 21, 2014 Revised: January 8, 2015

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Figure 1. Structures of a common alkenone produced by Isochyrsis sp. (a) and common FAME methyl linoleate (b). Nomenclature for both compounds is number of carbons:number of double bonds; however, note that the configuration of double bonds is different.

Table 1. Comparison of Fatty Acid, Alkenone, and Botryococcene Structures for Consideration As a Biofuel Feedstock fatty acids algae source carbon number double bonds heteroatoms

alkenones

botryococcenes

various linear, 14−22

E. hux., G. oceanica, Isochrysis linear, 35−41

B. braunii branched, 30−34

0−6 methylene-interupted cis-disubstituted alkenes carboxylic acid

1−3 trans disubstituted separated by 5 methylenes methyl or ethyl ketone

6 mono-, di-, and trisubstituted separated by 2−3 carbons none

the first report with experimental data for an alkenone-toproduct process and one of only a few examples demonstrating combined coproduct and biodiesel production from an algal feedstock.39

harvested for purposes of mariculture31−33 and, therefore, representative of the scale necessary for biofuel production. In an effort to produce biodiesel (FAME) from Isochrysis, we recently described results from the acid-catalyzed esterification of the total hexanes extract (“hexane algal oil”).11 This material contained a significant amount of alkenones (14% w/w) and contamination by these high-melting (∼70 °C) components is detrimental to cold flow fuel properties. More recently, a method for producing an alkenone-free Isochrysis biodiesel was reported using a saponification/separation procedure.12 Although this sequence adds additional steps to the overall process, in addition to supplying a superior quality biodiesel, it also generates an alkenone-rich neutral lipid fraction as a potential secondary product stream. We argue that alkenones represent a potentially fruitful and as yet unexplored renewable carbon source with structures particularly well suited for a number of catalytic processes. Specifically, alkenones feature long olefinic carbon-chains, favorable carbon-to-hydrogen ratios, and few heteroatoms (i.e., no sulfur or nitrogen and C:O ∼ 37−40:1, ref Figure 1).34−36 Key differences relative to fatty acids include a much longer hydrocarbon backbone, more widely spaced trans double bonds, and a ketone functional group (Table 1). Alkenones are also fundamentally different than the terpenoid botryococcenes that have received significant attention as a potential algal biofuel source,37 despite the noted slow growth habit of B. braunii, which poses concerns about its suitability as a biofuel feedstock.38 PULCAs thus represent an unexplored source of renewable carbon biosynthesized by robust algal species that could provide access to a unique suite of products unobtainable from other oil feedstocks. Herein, we provide proof-of-concept for the use of olefin metathesis as an alkenone-conversion strategy to produce a primarily C10-, C12-, and C17-hydrocarbon mixture (kerosene/ jet fuel boiling range) as a coproduct with biodiesel produced from the marine microalgae Isochrysis. The butenolysis reaction employs commercial ruthenium-based metathesis-initiators, occurs very rapidly at low temperature, and cleanly delivers a predictable product mixture. Additionally, we used comprehensive two-dimensional gas chromatography (GC × GC) to analyze the complex mixture of products obtained by directly reacting the mixture of alkenones isolated from commercially obtained Isochrysis, highlighting the powerful resolving capabilities of this analytical technique. The results represent

2. EXPERIMENTAL SECTION 2.1. Microalgae. The marine microalgae Isochrysis sp. “T-iso” was purchased from Reed Mariculture (strain CCMP1324) (Santa Cruz, CA) who has grown this species for nearly 20 years as a primary feed in shellfish and shrimp hatcheries. The algae were grown in greenhouse ponds under natural sunlight in a modified F/2 media. Average water temperatures were 18 to 20 °C. Approximately 2 kg of wet biomass (20% biomass w/w) was freeze-dried yielding dry Isochrysis sp. biomass as a dark-brown solid. 2.2. Separation of Fatty Acids and Neutral Lipids from the Hexane Algal Oil. Dry Isochrysis biomass was extracted in 30 to 50 g batches with hexanes in a Soxhlet extraction apparatus. The Soxhlet was cycled for 24−48 h until the solvent was a faint yellow. The hexanes were removed with a rotary evaporator and the weight of the hexanes-extractable material (referred as “hexane algal oil”) was recorded (typically ∼10 g from a 50 g dry biomass extraction). Fatty acids and neutral lipids were then separated from the hexane algal oil using our previously reported method.12 Briefly, the algal oil was treated with KOH at 60 °C for 3 h. The resulting saponified acylglycerols were selectively partitioned into water while the neutral lipids extracted with hexanes. Reacidification of the aqueous phase with HCl and extraction with hexanes produced the free fatty acids (FFAs). The overall mass recovery for combined FFAs and neutral lipids from the algal oil is typically quantitative (40% neutral lipids + 60% FFAs). 2.3. Isolation and Purification of Alkenones from the Neutral Lipids. Neutral lipids (10 g) were dissolved in a minimal amount of dichloromethane and flushed through silica gel (230−400 mesh, 100 g) with pressure using dichloromethane (approximately 150 mL) as eluent. Solvent was then removed on a rotary evaporator and the resulting orange-colored solid was recrystallized in hexanes to give pure alkenones (typically 4 g) as a white solid. 2.4. Alkenone Butenolysis, General Procedure. 2-Butene (0.2 mL, 15 equiv) was condensed in a reaction flask at −78 °C under a nitrogen atmosphere. Alkenones (100 mg), methyl stearate (methyl octadecanoate) (56 mg), dichloromethane or toluene (1.0 mL), and catalyst (2 mol %, 2−3 mg) were then added and the resulting heterogeneous mixture was placed in a refrigerator (4 °C) or ice bath (0 °C) for the allotted time. Reactions conducted were quenched with ethyl vinyl ether (0.9 mL, 50 equiv) and stirred for 15 min before concentrating on a rotary evaporator and analyzing by 1H NMR and gas chromatography. B

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Energy & Fuels 2.5. Analysis by 1H Nuclear Magnetic Resonance (1H NMR) Spectroscopy. 1H NMR spectra of the purified alkenones and butenolysis reaction mixtures were obtained under ambient conditions using CDCl3 as solvent, which also served as internal reference (shift value of residual proton at 7.26 ppm). 2.6. Analysis by One-Dimensional Gas Chromatography with Flame Ionization Detection (GC−FID) and Gas Chromatography−Mass Spectrometry (GC−MS). The purified alkenones and butenolysis reactions were analyzed on a Hewlett-Packard 5890 Series II GC−FID. Samples (1 μL) were injected cool-on-column and separated on a 100% dimethyl polysiloxane capillary column (Restek Rtx-IMS, 30 m length, 0.25 mm I.D., 0.25 μm film thickness) with H2 as the carrier gas at a constant flow of 5 mL min−1. The GC oven was programmed from 70 °C (7 min hold) and ramped at 6 °C min−1 to 320 °C (15 min hold). Percent conversions for the butenolysis reactions were determined by comparison of integration ratios for combined alkenones (rt = 44−48 min) to methyl stearate (retention time = 27.5 min) relative to a starting alkenone/methyl stearate standard mixture. Select samples were also analyzed by GC−MS on an Agilent 6890 GC with a 5973 MSD. Splitless 1 μL sample injections, were separated on a DB-XLB capillary column (60 m × 0.25 mm × 0.25 μm film thickness) using helium as the carrier gas (10.5 psi constant pressure), and the following GC temperature program: 4 min at 40 °C and ramped to 320 at 5 °C/min (held 15 min). 2.7. Analysis by Comprehensive Two-Dimensional Gas Chromatography with Flame Ionization Detection (GC × GC−FID) and Time-of-Flight Mass Spectrometer (GC × GC− TOF). Select butenolysis reaction mixtures were analyzed by GC × GC−FID and GC × GC−TOF MS according to previous described methodologies40 (for details of the analysis methods see the Supporting Information).

Figure 2. Isochrysis extraction and fractionation scheme with yields given in parentheses for the different products obtained from each step.

Table 2. Results from Butenolysis Reactions of Alkenone Mixtures Isolated from Isochrysis

3. RESULTS 3.1. Isolation and Purification of Alkenones As a Byproduct of Biodiesel Production. Prior to attempting any reactions, we isolated and purified the alkenones from the neutral lipid fraction containing other compounds including pigments such as chlorophylls and carotenes.41 This was challenging due to the low solubility of alkenones in a variety of organic solvents (e.g., hexanes, diethyl ether, acetone, ethyl acetate). After some optimization, the dark-colored pigmentcontaining material could be removed by flushing the dissolved neutral lipids through silica using a minimal amount of dichloromethane (DCM) as eluent. Upon removal of the solvent, the resulting orange-colored solid was further purified by recrystallization with hexanes affording analytically pure alkenones as a white solid. This procedure generally resulted in 40% isolated yield (w/w) from the neutral lipids or 3.2% of the Isochrysis dry biomass (Figure 2), which is close to the total alkenone content of 5% that we determined previously.11 Analysis of the purified alkenones by gas chromatography and comparison to standards revealed the presence of C37:3 (42%), C37:2 (27%), C38:2 (23%), and C38:3 (5%) alkenones with the small remainder accounted by C39:3 (1%) and C39:2 (2%) (where C[number]:[number] refers to the number of carbon atoms:number of double bonds, see Figure 1). 3.2. Cross-Metathesis of Alkenones with 2-Butene. Results from the cross-metathesis reactions of isolated alkenones with 2-butene using Grubbs’ first- (Ru−I) and second-generation (Ru−II) catalysts42 and Ru-HG43 are summarized in Table 2.

entrya

catalyst

solvent, 2-butene

time

% conversionb

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17

Ru-HG Ru-HG Ru−II Ru−I Ru-HG Ru-HG Ru-HG Ru-HG Ru-HG Ru-HG Ru−I Ru−I Ru-HG Ru-HG Ru-HG Ru-HG Ru-HG

DCM, cis-butene DCM, trans-butene DCM, cis-butene DCM, cis-butene DCM, cis-butene DCM, cis-butene DCM, trans-butene DCM DCM DCM DCM DCM PhMe PhMe PhMe DCM/trans-butene DCM/trans-butene

18 h 18 h 18 h 18 h 3h 1h 1h 10 min 20 min 30 min 3h 6h 10 min 20 min 30 min 15 min 30 min

100 100 100 70.0 100 100 100 62.9 98.8 99.5 9.5 16.7 52.5 84.4 98.3 37.4 95.5

a

All reactions were performed by adding alkenones (100 mg) to condensed 2-butene (15 equiv) at −78 °C followed by solvent (1 mL) and catalyst. The flask was then sealed and placed in a refrigerator (4 °C, Entries 1−7) or ice bath (0 °C, Entries 8−17) for the indicated time before quenching with ethyl vinyl ether (50 equiv) and concentrating in vacuo. bSamples were completely dissolved in DCM before analysis by GC−FID. Percent conversions for the butenolysis reactions were determined by comparing the integration ratios for combined alkenones to methyl stearate (inert internal standard) relative to a starting alkenone/methyl stearate reference mixture. For those reactions reported as 100% conversion, no alkenone signal was detectable by GC−FID. (See Figure 3).

map”.44 Many others have stressed the importance of valueadded coproducts to improve the economic viability of algalbiofuel programs.45−47 In line with these recommendations and as part of an advanced chemical prospecting strategy, one focus in our group has been the development of methods for converting alkenones to higher-value chemicals and fuels. Olefin metathesis has long been embraced by the synthetic organic and polymer communities,48,49 often used to create

4. DISCUSSION Coproducts are cited as one of the key reasons for exploring algae as a source of biofuels in the United States Department of Energy (DOE) “National Algal Biofuels Technology RoadC

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Energy & Fuels Scheme 1. Cross-Metathesis and the Reverse Ethenolysis

capped alkene products that are less reactive toward selfmetathesis.59 Patel and co-workers reported the rapid and high-yielding cross-metathesis reaction of methyl oleate (methyl (9Z)octadecenoate) with 2-butene using the second-generation Hoveyda−Grubbs catalyst (Ru-HG) to produce methyl 9undecenoate and 2-undecene (Scheme 3).60 Applied to longchain alkenones, certain fundamental differences between the alkenones and FAMEs made success of this reaction uncertain (refer to Figure 1 and Table 1). First, again the alkenones contain trans alkenes as opposed to the more metathesisreactive cis-configured double bonds found in FAMEs. Second, alkenones have limited solubility in the organic solvents used to perform olefin metathesis, particularly at the cold temperatures required to condense 2-butene (trans-2-butene, bp = 0 °C). Therefore, it was unclear whether the alkenones would even dissolve and, if so, whether the catalyst would engage the alkenone trans double bonds at these low temperatures All butenolysis reactions were performed using an excess of 2-butene (15 equiv, calculated as 5 equiv per alkene for the most abundant (37:3) alkenone in the starting mixture) to drive the equilibrium toward products and 2 mol % (calculated as above) of the catalyst. After 18 h at 4 °C, the alkenones were consumed when using catalysts Ru−II and Ru-HG (Entries 1− 3, Table 2), whereas Ru−I gave only 70% conversion under these same conditions (Entry 4). Patel and co-workers reported very low conversions (95% conversion (Entry 17). DCM was chosen as a solvent for these reactions as it had demonstrated the greatest alkenone solubility,11 although its use is undesirable for any “green” process.63 We therefore examined the reaction in toluene (PhMe), a more tolerated solvent that showed some alkenone solubility and is often used in olefin metathesis reactions. Reactions performed in toluene gave lower F

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Figure 6. GC × GC−TOF chromatogram “plan view” of the alkenone butenolysis products mixture showing separation of compounds into different subclasses and identification of several trace unexpected products including hexadecane (C16), octadecene (C18), and 2,10-dodecadiene (m/z 166).

hydrocarbon analysis, petroleum research, and oil spill science64 as it has many advantages over one-dimensional GC: its higher chromatographic resolution increases the signal-to-noise ratio and compounds are separated based on two physical properties (e.g., vapor pressure and polarity depending on choice of column stationary phase), leading to a grouping of chemical classes in a GC × GC chromatogram. Coupling of GC × GC with a flame ionization detector (FID) allows for the quantification of numerous unidentified compounds because most hydrocarbons have similar response factors.65 Coupled to a time-of-flight mass spectrometer (TOFMS), the enhanced resolution and increased signal-to-noise afforded by GC × GC allows for more accurate spectral identification of many compounds.66,67 Specific alkenones in our samples were identified by their mass spectrum, comparison to published elution order on gas chromatographic columns,68 textbook descriptions of alkenones,68 and other more recent studies detailing alkenone structure analysis.69,70 Relative amounts of individual alkenones were determined by GC−FID and these values correlated well with those previously reported for the same Isochrysis strain used in our study (Table 3).71 On the basis of this alkenone profile, we can then predict the products from our butenolysis reaction. For instance, each of the 37 and 38 alkenones should produce 2-heptadecene (2) and two equivalents of 2,9undecadiene (3) (refer to Scheme 3). Butenolysis of the 37 and 39 alkenones would similarly give 3 along with 8-decen-2one (1). When the relative alkenone percentages is considered, this would then give the distribution outlined in Table 3 with 1, 2, and 3 accounting for 83% of the products. Figure 5 is a typical GC × GC−FID chromatogram of the butenolysis products obtained by reaction of our alkenone mixture with cis-2-butene using catalyst Ru-HG. For reaction times down to 30 min in DCM at 0 °C, the butenolysis was complete (refer to Entry 3, Table 2).72 Each of the expected major products 1, 2, and 3 can be clearly identified.

Additionally, for both 1 and 3, two peaks are clearly visible with integration ratios from the GC × GC−FID of 3.9:1 that we have assigned as the trans- and cis-isomers, respectively. This is based in part on the earlier work from Patel and co-workers who also reported a 4:1 trans:cis ratio for 2-undecene obtained by butenolysis of methyl oleate.60 Three peaks in our alkenone butenolysis product mixture were identified with m/z = 152 in a ratio of 17.9:7.5:1 that have been assigned to the three possible isomers for 2 (E,E-, E,Z- and Z,Z-). Additional signals include cis- and trans-9-undecen-3-one (m/z = 168) obtained from the 38:3 ethyl alkenone contained in our sample (refer to Figure 1) and catalyst-derived 1-isopropoxy-2-(propenyl)benzene (m/z = 176). Altogether the ratio of butenolysis products 1:2:3 by GC × GC−FID analysis was typically 1:2.0:2.5 respectively, which is slightly different than what was predicted in Table 3 (1:2.3:3.4). Closer inspection of the GC × GC−TOF chromatogram data revealed several unexpected products compared to those presented in Table 3 that might help explain this discrepancy (vide infra). As is typical of GC × GC data, certain regions of the chromatogram contain different subclasses of compounds. For instance, using drilling mud samples containing a series of linear alkenes (“n-alkenes”) as a standard reference,73 an “nalkene” region could be identified containing not only the expected C17 and C19, but also trace C16 and C18 olefins (Figure 6). For each, two peaks were observed with peak area ratios of approximately 1:4, consistent with our previous cis- and transisomer assignments.74 2-Octadecene could have arisen from our sample containing very small amounts of a 38-methyl or 39ethyl alkenone (for a list of alkenone structures see Supporting Information). By a similar argument, hexadecene formation could have been formed from a methyl C36 alkenone that we did not detect in our sample nor has a C36 alkenone been reported for Isochrysis elsewhere. Alternatively some double bond isomerization occurred during the course of the crossmetathesis which has been reported for metathesis reactions of G

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Energy & Fuels other aliphatic systems.75 It is interesting, however, that only C16−C19 alkenes were detected for our butenolysis conducted at both short (e.g., 30 min) and longer (18 h) reaction times rather than the larger range of alkenes that could be envisioned from an isomerization process. Another possibility is that Isochrysis biosynthesizes trace amounts of alkenones with differing double bond positions. This would also perhaps explain the peak with m/z = 166 in the diene region of the chromatogram that we have tentatively identified as 2,10dodecadiene. Efforts are ongoing to characterize completely and better understand the mixture of products generated in this and other related reactions as it relates to both product use and alkenone structure elucidation.

and drilling mud. This material is available free of charge via the Internet at http://pubs.acs.org/.

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Corresponding Author

*E-mail: [email protected]. Telephone: (360) 650-6283. Fax: (360) 650-2826. Address: Department of Chemistry, Western Washington University, 516 High Street, Bellingham, WA, 98225. Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS Financial support from the National Science Foundation (CHE-1151492) is gratefully acknowledged.

CONCLUSION Alkenones are a well-known and much studied class of lipids produced by several species of marine algae including Isochrysis that have largely gone unnoticed as a potential renewable chemical feedstock. They can be isolated as a coproduct with biodiesel and owing to their long olefinic hydrocarbon chains are presumably amenable to a number of catalytic conversion technologies. We have demonstrated that alkenones are reactive toward olefin metathesis and their cross-metathesis with 2butene produces a C10−C19 hydrocarbon mixture containing predominantly 2,9-undecadiene (43% predicted). The butenolysis reaction uses commercially available ruthenium-based metathesis initiators, occurs rapidly at low temperature, and delivers a predictable mixture of hydrocarbon fragments. It is known that alkenone unsaturation is sensitive to algae growing conditions.15,16 Other more sophisticated techniques like viral infection76 might also prove capable of controlling the alkenone unsaturation profile to ultimately fine-tune the products obtained by metathesis. There is no prior work on optimizing alkenone production from Isochrysis sp. However, as a coproduct of an industrial biofuel process, even at 5% (or 3.5% pure from our method) of the biomass, this would represent a significant amount of alkenones. For instance, in the U.S., consumption of petroleum-derived products is approximately 10 L per person per day (20 million bbl/day,77 319 million people,78 159 L/bbl). Replacing 10% of that quantity with an Isochrysis-derived biodiesel, thus, would require producing about 1 kg of algal biofuel/person/day. We typically obtain 12% (w/w) of a pure biodiesel (FAME) from dry Isochrysis biomass, meaning one would need 8.33 kg of Isochrysis/person/day. Using our total and purified alkenone yields, this would correspond to potentially 0.3−0.4 kg of alkenones/person/day, or 30−40% of the biodiesel production volume. Utilization of this material in some capacity (e.g., fuels or specialty chemicals), therefore, could significantly increase the overall value of the algal biomass. The current price of Isochrysis is approximately $400/kg, sold by a handful of vendors and controlled by the economics of its current market as shellfish feed.79 Using this value, the Isochrysis lipid-derived fuels we have described are far from cost-competitive (> $10,000/gal). Future work is aimed at assessing the commercial viability of these and other haptophyte-based biofuels.

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AUTHOR INFORMATION

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REFERENCES

(1) For a recent review see: Naika, S. N.; Goud, V. V.; Rout, P. K.; Dalai, A. K. Renew. Sust. Energy Rev. 2010, 14, 578−597. (2) “Renewable Fuels Standard”, United States Environmental Protection Agency. http://www.epa.gov/otaq/renewablefuels/ 420f07019.pdf (accessed November 17, 2014). (3) Liquid Transportation Fuels from Coal and Biomass: Technological Status, Costs, and Environmental Impacts (America’s Energy Future), America’s Energy Future Panel on Alternative Liquid Transportation Fuels; National Academy of Sciences, National Academy of Engineering, National Research Council, National Academies Press: Washington, DC, 2009. (4) Danner, H.; Braun, R. Chem. Soc. Rev. 1999, 28, 395−405. (5) The Biodiesel Handbook, 2nd ed.; Knothe, G., Van Gerpen, J., Eds.; AOCS Press: Urbana, IL, 2010. (6) Tenenbaum, D. J. Environ. Health Perspect. 2008, 116, A254− A257. (7) Brown, T. R.; Brown, R. C. Biofuels, Bioprod., Biorefin. 2013, 7, 235−245. (8) Knothe, G. Prog. Energy Combust. Sci. 2010, 36, 364−373. (9) Foley, P. M.; Beach, E. S.; Zimmerman, J. B. Green Chem. 2011, 13, 1399−1405. (10) Inderwildi, O. R.; King, D. A. Energy Environ. Sci. 2009, 2, 343− 346. (11) O’Neil, G. W.; Carmichael, C. A.; Goepfert, T. J.; Fulton, J. M.; Knothe, G.; Lau, C. P-L.; Lindell, S. R.; Mohammady, N. G-E.; Van Mooy, B. A. S.; Reddy, C. M. Energy Fuels 2012, 26, 2434−2441. (12) O’Neil, G. W.; Knothe, G.; Williams, J. R.; Burlow, N. P.; Culler, A. R.; Corliss, J. M.; Carmichael, C. A.; Reddy, C. M. Energy Fuels 2014, 28, 2677−2683. (13) Volkman, J. K.; Eglinton, G.; Corner, E. D. S. Phytochem. 1980, 19, 2619−2622. (14) Conte, M. H.; Thompson, A.; Lesley, D.; Harris, R. P. Geochim. Cosmochim. Acta 1998, 62, 51−68. (15) Brassell, S. C.; Eglinton, G.; Marlowe, I. T.; Pflaumann, U.; Sarnthein, M. Nature 1986, 320, 129−133. (16) Marlowe, I. T.; Brassell, S. C.; Eglinton, G.; Green, J. C. Org. Geochem. 1984, 6, 135−141. (17) Prahl, F. G.; Wakeham, S. G. Nature 1987, 330, 367−369. (18) Eglinton, G.; Bradshaw, S. A.; Rosell, A.; Sarnthein, M.; Pflaumann, U.; Tiedemann, R. Nature 1992, 356, 423−426. (19) Müller, P. J.; Kirst, G.; Ruhland, G.; von Storch, I.; Rosell-Melé. Geochim. Cosmochim. Acta 1998, 62, 1757−1772. (20) Volkman, J. K.; Barrerr, S. M.; Blackburn, S. I.; Sikes, E. L. Geochim. Cosmochim. Acta 1995, 59, 513−520. (21) Eltgroth, M. L.; Watwood, R. L.; Wolfe, G. V. J. Phycol. 2005, 41, 1000−1009. (22) E. Huxleyi has been shown to biosynthesize alkenones but no TAGs: Volkman, J. K.; Everitt, D. A.; Allen, D. I. J. Chromatogr. 1986, 356, 147−162.

ASSOCIATED CONTENT

* Supporting Information S

Alkenone structures, 1H NMR spectra of isolated alkenones and butenolysis product mixture, butenolysis GC−FID chromatograms, additional GC × GC−TOF data, and comparison of GC × GC “n-alkene” region for butenolysis H

DOI: 10.1021/ef502617z Energy Fuels XXXX, XXX, XXX−XXX

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DOI: 10.1021/ef502617z Energy Fuels XXXX, XXX, XXX−XXX