Optimization of Direct Conversion of Wet Algae to Biodiesel under Super-critical Methanol Conditions

Because conventional processes of biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> production from microalgae are both energy and cost intensive, researchers are seeking alternative methods that would minimize these costs, enabling large-scale microalgae-based biodiesel production. The method studied by Patil et al. (2011) in this paper was the direct conversion of wet algae to biodiesel in supercritical methanol conditions (SCM). Traditional processes require the drying of wet algal biomass, the extraction of the oil with solvents, and the catalyzed conversion of the algal oil to biodiesel. SCM conditions allow for a single-step process that could circumvent these expensive steps, thus greatly lessening biodiesel production costs. The researchers sought in this study to characterize the optimal conditions under which biodiesel could be created in this manner, varying reaction time, temperature, and wet algae to methanol (wt./vol.) ratio. They used response surface methodology (RSM) to analyze the results and found that reactions run at a temperature of 255 °C for 25 minutes with a 1:9 wt./vol. ratio produced the best results, representing a potential economical and efficient method of biodiesel production. —Karen de Wolski
Patil P., Gude V., Mannarswamy A., Deng S., Cook P., Munson-McGee S., Rhodes I., Lammers P., Nirmalakhandan N<!–[if supportFields]>XE “nitrogen, N”<![endif]–><!–[if supportFields]><![endif]–><!–[if supportFields]> XE “nitrogen” <![endif]–><!–[if supportFields]><![endif]–>., 2011. Optimization of direct conversion of wet algae to biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> under supercritical methanol conditions. Bioresource Technology 102, 118–122.

Patil et al. (2011) set out to elucidate the optimal conditions for the one-step reaction of biodiesel<!–[if supportFields]>XE “biodiesel”<![endif]–><!–[if supportFields]><![endif]–> formation from wet algae with SCM conditions. This process avoids the drying, extraction, and catalyzed conversion processes which represent great costs in traditional biodiesel production. Under SCM conditions, water is used as a co-solvent that accelerates conversion of fats and oils to fatty acid methyl esters (FAMEs) and increases solubility and acidity. The process set forth in this study produces FAMEs from polar phospholipids, free fatty acids (FFAs), and triglycerides by reducing polarity of high energy algal molecules while increasing fluidity and volatility. This allows for a single-step process in which extraction and transesterification of wet algal biomass are carried out simultaneously, requiring modest temperatures and relatively low energy input. This process has been conducted successfully for vegetable oils at about half the cost of conventional transesterification methods, and the researchers in this study sought to both demonstrate that it could be carried out for algal oil and to elucidate optimal reaction conditions.
          Patil et al. first characterized the algal samples through various chemical analyses. Lipid extraction of the Nannochloropsis<!–[if supportFields]> XE “Nannochloropsis” <![endif]–><!–[if supportFields]><![endif]–> sp<!–[if supportFields]> XE “Nannochloropsis sp”<![endif]–><!–[if supportFields]><![endif]–>ecies resulted in triglyceride content at 37.72%, other non-polar hydrocarbons/isoprenoids at 8.72%, and polars, glycolipids, and phospholipids at 3.54%. The researchers also used thin layer chromatography, densitometry, and scanning electron microscopy to further characterize the algal sample. Additionally, an FTIR spectra showed this particular algal species to be highly aliphatic and to have hydroxyl, carboxyl, and carbonyl groups, all identified by specific absorption bands. The triglyceride biosynthetic pathway in microalgae is thought to consist of the formation of acetyl coenzyme A in the cytoplasm, the elongation and desaturation of fatty acid carbon chain, and the subsequent biosynthesis of triglycerides. Methanol’s intermolecular hydrogen bonding is significantly decreased in the supercritical state, reducing its polarity and dielectric constant, and allowing the alcohol to solvate non-polar triglycerides. This results in a single phase lipid/methanol mixture and produces FAMEs and diglycerides that can be transesterified into methyl ester and monoglyceride and eventually glycerol.
          The researchers identified the wet algae to methanol (wt./vol.) ratio, reaction temperature, and reaction time as being the most critical variables affecting product FAME content. They utilized RSM to analyze these variables, a statistical analysis involving a three factorial subset that allows for accurate approximation of true error and significance. Wet algae to methanol ratios between 1:4 and 1:12, reaction times between 10 and 30 minutes, and temperatures between 240 and 260 °C were used in a total of 28 experimental runs. Additionally, Patil et al. implemented a general second order linear model with a deconstructionist approach to facilitate parametric evaluation for the predicted response surface and a least square method to predict the values of the involved parameters.
          For the actual experiment, 4 g samples of wet algae paste were run through non-catalytic SCM in a micro-reactor under a matrix of the previously described reaction conditions at a constant pressure of 1200 psi. The organic contents containing the non-polar lipids were isolated and analyzed by gas chromatography-mass spectroscopy (GC-MS) with methyl heptadecanoate as an internal standard. The FAME content of the final product could then be calculated by comparing the integrals of the FAME peaks with integrals of the standard peak. A general linear model, least squares, and ANOVA<!–[if supportFields]> XE “ANOVA”<![endif]–><!–[if supportFields]><![endif]–> were conducted to analyze the effects of the varied reaction parameters.
          The regression analysis showed all three parameters to significantly influence FAME content, confirmed by both P-values and the correlation coefficient (R2=0.921). The researchers created graphs showing response contours of FAME yield against temperature and wet algae to methanol ratio at the three time intervals. The regression coefficients show that reaction time positively affects response up to 255 °C, while higher temperatures are not conducive to transesterification reactions, possibly due to decomposition<!–[if supportFields]> XE “decomposition” <![endif]–><!–[if supportFields]><![endif]–> of oil/lipids and alkyl esters. The ratio of wet algae to methanol had a positive effect on yield up to 1:9, but negatively influenced yield at higher ratios. This may be explained by the reversible reaction being shifted forward as a result of increased contact area between methanol and lipids. This parameter can also interact with reaction temperature to reduce FAME yield due to either FAME decomposition or the reduction of the critical temperature of the reactant/product. High reaction times allowed for completion of the transesterification reaction and thus higher FAME yields. This effect was especially notable at the wet algae to methanol ratio of 1:9 at 255 °C. The experimental analysis and RSM study showed maximum yields under the aforementioned conditions with a reaction time of 25 minutes.
          The researchers were also interested in the elemental composition of the algal samples. They therefore subjected raw and residual samples to scanning electron microscopy to produce the elemental spectra. These results showed that the algal cell wall structure was disturbed and fragmented under the SCM condition and that the algal biomass was thermally degraded due to high unsaturated fatty acid content.
          The algal biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> samples were analyzed by GC-MS to quantify the product. The FAME content was calculated by comparing the FAME peak integrals to the internal standard peak integrals. The algal biodiesel was found to have a large proportion of mono and poly unsaturated FAMEs. The ATR-FTIR spectra of the algal biodiesel were compared to the ATR-FTIR spectra of camelina biodiesel and petro-diesel. The main components of diesel are aliphatic hydrocarbons, which were observed by various peaks on all three spectra.
          The authors conclude that this single-step biodiesel<!–[if supportFields]>XE “biodiesel”<![endif]–><!–[if supportFields]><![endif]–> production process shows great promise in its shorter reaction time, simple product purification, and maximum FAME conversion. It requires lower energy input than conventional methods, and the process can be successfully optimized by RSM, representing the potential for efficient, relatively low-cost biodiesel production. 

Variables Affecting the In Situ Transes-terification of Microalgae Lipids

While microalgae show great potential as a future source of biomass for biofuel production, there still exist several obstacles for commercial production associated largely with the high costs of biomass production and fuel conversion routes. The majority of current research into fuel conversion involves the extraction of lipids from biomass and their subsequent conversion to fatty acid alkyl esters (FAAE) and glycerol. This research has traditionally used alkaline catalysts for the transesterification process, but these are limited because they result in partial saponification when used with oil reactants that have free fatty acid (FFA) content above 0.5% w/w. The use of hydrochloric and sulfuric acid as catalysts has been explored as a low-cost alternative that is not affected by FFA content. Ehimen et al. (2010) sought to determine how to maximize the reaction rate by varying conditions, including temperature, reacting alcohol volume, reaction time, and moisture content. They found that fatty acid methyl ester (FAME) conversions rates were positively correlated with volume and temperature, and that equilibrium FAME conversions approached asymptotic reaction time limits of 8 hours for almost all temperatures investigated. —Karen de Wolski
Ehimen E., Sun Z., Carrington C., 2010. Variables affecting the in situ transesterification of microalgae lipids. Fuel 89, 677–684.
Ehimen et al. (2010) researched how certain reaction variables affect biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> production from microalgae lipids in an acid-catalyzed in situ transesterification process. While most research into microalgae-based biodiesel production has focused on alkaline catalyzed transesterification, this type of catalyst is not viable for large-scale commercial biodiesel production because it results in saponification when used with oils that have an FFA content above 0.5% w/w. The use of inorganic acids as catalysts for this reaction has been explored, as biodiesel producing transesterification and esterifiction reactions can be catalyzed in this manner. Previous studies have shown that acidic catalysts result in higher fatty acid methyl esters (FAME) yields than alkaline catalysts, and these yields can be optimized under certain reaction conditions.
          In order to reduce biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> production costs, several alternatives to conventional transesterification reactions have been explored. This study focused on “in situ” transesterification, a process which reduces cost by eliminating the need for the solvent extraction step, as the biomass oil is converted directly to FAAE. This method could be particularly efficient for microalgae-based biodiesel production because the alcoholysis of the oil in the biomass directly increases biodiesel yields when derived from microalgae biomass oil. Because microalgae lipids tend to have a high FFA content, acid catalysts were used in this study, which sought to find optimal reaction conditions for microalgae-based in situ transesterification. The researchers varied reacting alcohol volume, temperature, reaction time, and process mixing, seeking how to create a reaction which would have the lowest process costs and the best biodiesel yield.
          Microalgae oil was extracted from dried Chlorella biomass via previously established methods. Several analyses were conducted on the oil sample, including specific gravity (SG), gas chromatography, and titration to determine acid value and FFA content. The researchers carried out the in situ transesterification process with a sulphuric acid catalyst at the different alcohol volumes in question. The SGs of the extracted FAME products were measured to determine the extent of conversion during reaction, as a decreasing SG signifies that the reaction has reached equilibrium conversion between microalgae lipids and methyl esters. The extent of conversion was further verified by a GC, and a calibration curve of the relationship between SG and FAME conversion was created.
          After setting up the in situ transesterification reaction, several conditions were varied to maximize output. Five different methanol volumes and four different temperatures were tried, and the FAME product yields and SGs were measured as described previously. Additionally, the researchers tested for the effect of reaction time at each temperature, running a total of eight different reaction times at each experimental temperature. The researchers were also interested in how moisture content and stirring affects the in situ transesterification process. They therefore studied the effects of moisture by air drying and oven drying Chlorella samples to nine different moisture contents and subjecting these samples to the process at a constant temperature and alcohol volume. They then subjected the in situ transesterification process to four stirring variations at a constant temperature and alcohol volume and analyzed the product yields as previously described.
          The Chlorella grown under the culture conditions of this study was found to have a total transesterifiable lipid fraction of 0.276 g oil/g biomass. The SG of the extracted oil was measured at 0.914 at 25 °C. Using the GC analysis of the extracted oil and the chemical equation of the reaction (MMoil=[3MMFA+ MMglycerol] –3MMwater), average molecular mass of the oil was calculated to be 880 g/mol. A calibration curve was obtained by correlating concentration of FAME species and SG of purified product. This curve covers a range of 0% conversion (pure extracted oil) to 92.22% conversion (lowest measured SG), and could therefore be used to predict corresponding percentage FAME conversions for the other reactions run in this study. A strong negative correlation between SG and percent FAME conversion was demonstrated by this curve. The acid value of the oil was found to be 10.21 mg KOH/g, and the FFA content was calculated at 5.11%. Because this is a relatively high FFA content, the use of an acidic catalyst for in situ transesterification reactions was justified for this microalgae oil.
          The results of the experiments varying temperature and alcohol volume show that
increasing both variables generally increases microalgae oil conversion to FAME. However, no significant trends were observed for temperature levels of 60 and 90 °C. For the experiments investigating the interaction of reaction time and temperature, asymptotic FAME conversion values were not reached in the 12 hour time boundary of this study at room temperature. Highest equilibrium conversion levels were reached in the shortest time (70% at 15 min, 90% at 1 hour) for the 90 °C reaction temperature. Similar asymptotic values at reaction times of 2 and 4 hours were found for the 60 and 90°C conditions, and the researchers state that 60 °C may therefore be ideal because process heating and pressure requirements could be inhibited at the higher temperature. The authors were interested in the effect of moisture content on FAME conversion because the drying process represents a significant cost in biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> production. However, the results indicate that drying cannot be avoided, as there was a strong negative correlation between moisture content and FAME conversion. The researchers also sought whether stirring could be avoided as a cost-cutting measure. While stirring the reaction for only one hour did produce high equilibrium conversion levels, the yield was still only 91.3% of a continuously stirred system. Intermittent one hour stirring produced results closer to the continuously stirred reaction, but nevertheless fell short, and the investigators therefore concluded that stirring (at least intermittently) is likely a necessary cost of biodiesel production for the in situ transesterification process.
          The authors conclude that increasing temperature, reaction time, and alcohol volume may favor biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> production for this particular process. However, energetic cost of the recovery of excess alcohol reactants and increases in FAME purification requirements could potentially limit any cost reductions achievable by these variables. Therefore, further optimization of this process, including studying filtration, evaporation, and extraction, should be studied to make in situ transesterification a viable biodiesel production method. 

Microalgae Cultivation in a Wastewater Dominated by Carpet Mill Effluents for Biofuel Applications

The current global target for biofuel feedstock crop production by 2030 would demand approximately 180 km3 of water, a demand that could be severely limiting given the overall worldwide depletion of freshwater sources. Because microalgae are a promising option for future biofuel production, finding ways to cultivate and harvest them with relatively little freshwater is both valuable and necessary. Chinnasamy et al. (2010) studied the feasibility of growing microalgae in wastewater consisting primarily of carpet mill effluents. This is especially appealing because it would not only greatly reduce the freshwater demand of microalgae cultivation, it would also serve to remove contaminants from the wastewater itself. The researchers found that a consortium of 15 native algal isolates could grow in treated wastewater with >96% nutrient removal, and 63.9% of their oil could be converted into biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–>. Chinnasamy et al. conclude that, while these results are promising, more research is needed to elucidate the mechanisms of anaerobic<!–[if supportFields]> XE “anaerobic” <![endif]–><!–[if supportFields]><![endif]–> digestion<!–[if supportFields]> XE “anaerobic digestion” <![endif]–><!–[if supportFields]><![endif]–> and thermochemical liquefaction in order for this process to be economically viable. —Karen de Wolski
Chinnasamy S., Bhatnagar A., Hunt R., Das KC., 2010. Microalgae cultivation in a wastewater dominated by carpet mill effluents for biofuel applications. Bioresource Technology 101, 3097–3105.

Chinnasamy et al. sought to determine the feasibility of microalgae cultivation in wastewater primarily consisting of carpet mill effluents. Annual worldwide domestic and industrial water consumption between 1987 and 2003 was estimated at 325 billion m3 and 665 billion m3 respectively. Approximately 247 million tons of algal biomass and 37 million tons of oil could be created if 50% of the wastewater from this consumed water was used for algae production. Due to the variation in wastewater composition, strains of microalgae able to grow in varying environments must be found if the technology is to advance. The wastewater from carpet mills is rich in phosphorous and nitrogen<!–[if supportFields]> XE “nitrogen” <![endif]–><!–[if supportFields]><![endif]–>, and the researchers therefore wanted to examine how microalgae could grow in the water and remove the contaminants. They additionally wanted to determine how consortium (multi-species) based technology functions for nutrient removal and biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> production.
          Wastewater consisting primarily of carpet mill effluent was collected from a utility company in Dalton, Georgia. In order to minimize temporal variation effects, wastewater was collected in large batches for all four seasons. Water was also collected from the treatment facility to enable the characterization of treated versus untreated wastewater. The algal taxa present and biovolume present in the samples were identified via standard protocol. The water samples were incubated in growth conditions to induce algal growth. The algae were then isolated by serial dilution and incubated on BG11 agar plates, from which individual colonies were selected and maintained. Thirteen microalgal strains and a consortium of wastewater isolates were identified and screened, and two freshwater and two marine forms and the consortium were selected for the timescale batch study.
                Several different experiments were carried out within the study. First, biomass production and nutrient removal of the consortium was examined by growing the consortium in flasks of filtered and sterilized wastewater as a nutrient medium under two different levels of CO2 (ambient and 6%) and temperature (15 and 25 °C) conditions. Because the researchers also wanted to know the potential of cultivating the consortium in open ponds, the consortium was also cultivated in treated wastewater in four raceway ponds. The algae were harvested, dried, and the lipids were extracted for biomass analysis. Biodiesel was produced from crude microalgae oil via acid transesterification and base transesterification. The biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> was then analyzed with gas chromatography. Biomass of the harvested cells was quantified through filtration, and lipid content was measured gravimetrically with an automated extraction system. Additionally, total nitrogen<!–[if supportFields]> XE “nitrogen” <![endif]–><!–[if supportFields]><![endif]–> and phosphorous were determined via a persulfate method.
          Because Chinnasamy et al. sought to examine temporal variation in nutrient concentration in wastewater, biochemical oxygen demand, chemical oxygen demand, total suspended solids, and several other parameters were measured in both treated and untreated water throughout the seasons. It was found that there are sufficient nutrients in both treated and untreated wastewater to support algae growth. About 27 species of green algae, 20 species of cyanobacteria<!–[if supportFields]> XE “cyanobacteria” <![endif]–><!–[if supportFields]><![endif]–>, and eight species of diatoms were found in the treated and untreated wastewater, with green algae and cyanobacteria dominating both water types in all seasons. The observed variations in responses of different species to different environmental conditions led the researchers to believe that parameters aside from nutrient availability are the most important in determining species composition.
          Several different species, including the consortium, were grown in treated and untreated wastewater and standard growth medium. Several strains showed significant growth in both wastewater types. Marine forms were able to grow in treated and untreated water without any supplements, indicating themselves as having possibly the highest potential for growth in wastewater for biofuel in the future. The species with the highest growth in the preliminary screening were subjected to a time-scale study in treated and untreated carpet industry wastewater. Overall, the best performer was the consortium grown in treated wastewater, with the potential to generate 4060 L of oil ha–1 year–1. The researchers also calculated that it could produce ample biofuel (3860 L of oil ha–1 year–1) when cultivated in untreated wastewater. For the experiment examining consortium growth in different CO2 and temperature conditions, it was found that microalgae cultivated in 6% CO2 at 25 °C had the highest biomass productivity. The consortium’s performance was enhanced in treated wastewater. After 72 hours of incubation, nitrate and phosphate were almost completely removed from the growth medium, indicating high nutrient removal abilities. Biomass of the consortium algae grown in the raceway ponds was quantified and analyzed. Interestingly, the consortium had a high protein content (54.6%) and low lipid and carbohydrate content. When algal lipid content is lower than 40%, energetic cost of harvest can outweigh energetic added value of lipid recovery. Therefore, direct energy recovery may be necessary in algae with low levels of lipids.
          To examine the viability of consortium algae-based biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> production, crude algal oil was extracted from the biomass, chemically analyzed, and converted into biodiesel. The crude algal oil had a free fatty acid content of about 50%, a trait not conducive to biodiesel conversion. However, the total acid esterification showed a product yield of about 70.9%, with losses mainly resulting from oil impurities. While the biodiesel produced had a higher linolenic acid content (27.9%) than is normally acceptable (12%), the researchers speculate that the quality of the fuel could be improved by deriving some biomass from other non-food feedstocks.
          The results of this study show that algal oil from mixed cultures of native algae is a feasible source of biostock for biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> production. It will be necessary to find economical methods of crude oil refinement to minimize product impurities, and this remains a large obstacle. Despite the low lipid content of the consortium in this study, biomass recovery via thermochemical liquefaction could enhance recovery rates and reduce energy expenditure for algae cultures with low lipid content. Therefore, thermochemcial liquefaction should be studied and developed to make wastewater-cultivated consortium microalgae a commercially feasible process of biodiesel production. 

Cost-effectiveness Analysis of Algae Energy Production in the EU

The transition towards renewable energy is being largely driven by the increase in global energy demand, challenges to energy security, and climate change. In the EU, fossil fuels constitute about 98% of energy consumption within the transportation sector. While biofuels are seen as a possible substitute for fossil fuels, having environmental, economic, and political benefits, first generation biofuels have come under fire<!–[if supportFields]> XE “fire” <![endif]–><!–[if supportFields]><![endif]–> because of their negative effects, including deforestation<!–[if supportFields]> XE “deforestation” <![endif]–><!–[if supportFields]><![endif]–>, water and soil degradation, and threats to food security. The European Environment Agency, doubting their ability to satisfy the EU’s energy needs, has proposed to suspend biofuels until a study has been conducted that weighs the costs and benefits of their production and utilization. These criticisms of first generation biofuels do not, however, apply to other potential biofuel alternatives and their external environmental benefits. In this study, Kovacevic and Wesseler (2010) completed a cost-effectiveness analysis of the possible utilization of algal biomass as a biofuel source. They took into account externalities, including emissions, impact on food prices, and pesticide<!–[if supportFields]> XE “pesticide” <![endif]–><!–[if supportFields]><![endif]–> use. They concluded that the development of biotechnology and the elevation of crude oil prices would lead to algal biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> outcompeting other fuels, but a substantial capital investment will be necessary to make this a reality.—Karen de Wolski
Kovacevic V., Wesseler J., 2010. Cost-effectiveness analysis of algae energy production in the EU. Energy Policy 38, 5749–5757.

Kovacevic and Wesseler sought to quantify both the direct financial and the external cost-effectiveness of producing algae as a biofuel source in the EU. While microalgae are seen as having great potential in this arena, one of the largest obstacles is capital investment required to develop the necessary biotechnologies.
          The two characteristics of microalgae most appealing for utilization as an energy source are their high biomass yield relative to other terrestrial plants, and their high utilization of CO. Microalgae’s efficient conversion of solar energy and utilization of water, carbon, and nutrients leads to a yield 7–31 times higher than palm oil, the best oil-yielding terrestrial feedstock. Recent interest in microalgae has led to an array of research, with studies indicating a vast spectrum of possible yields dependent on various factors and conditions. While biotechnology advancement is seen as a major pathway for improving yield, Kovacevic and Wesseler indicate that this development is not sufficient. Rather, systems must be optimized for all growing conditions if lipid yields are to be high enough on a large scale. Currently, the only successful mass culture productions of microalgae have been achieved with species tolerant to extreme conditions, a limitation that will need to be overcome if microalgae are to be a major fuel source.
          In this study, the researchers compared algal biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> with rapeseed biodiesel<!–[if supportFields]> XE “rapeseed biodiesel” <![endif]–><!–[if supportFields]><![endif]–> and fossil fuels in the EU-25 transportation sector. Annuities (expressed per GJ of fuel energy delivered to the gas station) were used to aggregate the private and external costs and benefits of each fuel option for the cost-effectiveness analysis, and represented average annual cost to society accounting for production costs, environmental benefits and costs, energy security, and food price impacts. The European Commission’s established biofuel target of 10% vehicle biofuel use by 2020 was used as the time frame and production scale for the comparison. This proposal projects 1.48 EJ of biofuel utilization by the target year. Because discount rates for biofuels vary between 3% and 8%, 5% was chosen as the social discount in this study, although 2% and 8% were also applied. Discount rate is important because high discount rates would probably favor fossil fuels over biofuels because biofuels require higher investment.
          The researchers calculated the private production costs of the various fuels in question by choosing certain growing conditions and production processes. They assume favorable conditions (high solar radiation, optimal climate, low altitude, proximal seawater source) as would be found in southwestern and eastern Spain and southeastern Italy. They define three cases: a base case, a low-yield case, and a high-yield case. Kovacevic and Wesseler calculated land requirement through previous microalgal studies, and accounted for the relatively low opportunity cost of land use for algal production. They assumed a basic production unit of 400 ha with paddle wheel mixing, pure CO2 mixing, and anaerobic<!–[if supportFields]> XE “anaerobic” <![endif]–><!–[if supportFields]><![endif]–> digestion<!–[if supportFields]> XE “anaerobic digestion” <![endif]–><!–[if supportFields]><![endif]–> for nutrient recycling, with carbon supplied from nearby coal<!–[if supportFields]> XE “coal” <![endif]–><!–[if supportFields]><![endif]–> plants. Water supply is assumed to be from seawater sources, with replenishment from freshwater. The cost of the pipe network for water transport and pumping is based on other known engineering projects. The transesterification process of biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> production is assumed to be equal to the rapeseed biodiesel<!–[if supportFields]> XE “rapeseed biodiesel” <![endif]–><!–[if supportFields]><![endif]–> process. The researchers also calculated the cost of fuel distribution, considering the differences in energy content between diesel and biodiesel.
          Rapeseed biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> production costs were generated based on current rapeseed growth and processing. Germany, France, Poland, and the UK are the largest EU rapeseed producers, constituting 72% of total EU rapeseed production. The projected 30% yield improvement by 2020 was accounted for, and maximum land utilization by rapeseed oil was calculated to be 29.1 Mha, as compared to 1.35 Mha for microalgae. Fossil fuel production costs were also calculated based on gas station prices and taxes. Three cases were estimated due to the volatility of crude oil prices, and elasticity between crude oil and fuels price were applied for diesel and gasoline<!–[if supportFields]> XE “gasoline” <![endif]–><!–[if supportFields]><![endif]–>.
          The researchers next calculated the social cost of fuel utilization by considering external costs, especially environmental effects. These included costs of CO2, methane<!–[if supportFields]> XE “methane (CH4)” <![endif]–><!–[if supportFields]><![endif]–>, nitrous oxide<!–[if supportFields]> XE “nitrous oxide (N2O)” <![endif]–><!–[if supportFields]><![endif]–>, volatile organic compounds, particulate matter, sulfur dioxide, and nitrogen<!–[if supportFields]> XE “nitrogen” <![endif]–><!–[if supportFields]><![endif]–> oxide emissions. In order to calculate greenhouse gas (GHG) emissions, three drivers were considered: land use change (only applicable for biofuels), fuel distribution and dispensing, and fuel combustion (only fossil fuels). The effect of land use change on GHG emissions accounted for changes in nitrogen emissions from fertilizers, changes in methane emissions from livestock, and cropland conversion. Conversion of grassland<!–[if supportFields]> XE “grassland” <![endif]–><!–[if supportFields]><![endif]–> for rapeseed cultivation increases GHG emissions because of fertilizer use, while conversion of cropland to algal ponds decreases GHG emissions. Additionally, energy input for processing algal and rapeseed biodiesel<!–[if supportFields]> XE “rapeseed biodiesel” <![endif]–><!–[if supportFields]><![endif]–> production were considered in the calculation. Distribution and dispensing costs were based on previous estimates, and applied for all fuel types. GHGs from combustion were calculated only for fossil fuels because they release carbon.
          Next, the impact on food prices of the different fuel types was considered. It was assumed that microalgae biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> would have no impact on food prices because of their small land requirement and ability to use marginal land. Rapeseed requires a significant amount of land, and it was estimated that it would necessitate reallocating 70%, 55%, and 54 % of total wheat, barley, and maize<!–[if supportFields]> XE “maize” <![endif]–><!–[if supportFields]><![endif]–> acreage respectively across Germany, France, Poland, and the UK. The researchers also account for the costs of fertilizer<!–[if supportFields]> XE “fertilizer” <![endif]–><!–[if supportFields]><![endif]–> and pesticide<!–[if supportFields]> XE “pesticide” <![endif]–><!–[if supportFields]><![endif]–> leaching, comparing cost of purchase to theoretical environmental cost. Because energy security has become a major issue surrounding oil dependency, security of supply for biofuels was derived from the EC Biofuels progress report (2007).
          The entire analysis is presented as a comparison of annuities. The social cost for the base case of algal biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> is 52.3 ϵ/GJ, with significant cost differences for the alternative cases. Rapeseed oil and fossil fuel had total social costs of 36.0 and 15.8 ϵ/GJ respectively. However, the private costs dominated the cost structure of algal biodiesel production, while the externalities constitute a larger percentage of total cost for rapeseed and fossil fuels. The social cost of fossil fuels increases significantly when crude oil prices rise ($100–$200 /barrel). The private costs of both biofuel types are largely due to biomass production costs, with carbon supply and water supply being the primary factors. Food prices impact and GHG emissions are the greatest external costs of rapeseed biodiesel<!–[if supportFields]> XE “rapeseed biodiesel” <![endif]–><!–[if supportFields]><![endif]–> and fossil fuels, while only algal biodiesel has external benefit in GHG emissions mitigation.
          Kovacevic and Wesseler conclude that fossil fuels have the lowest private utilization costs while algal biodiesel<!–[if supportFields]> XE “biodiesel” <![endif]–><!–[if supportFields]><![endif]–> has the highest cost. This makes sense given the current infrastructure supporting fossil fuel energy production. Rapeseed oil is also relatively supported within the current fuel system. However, there are scenarios in which algal biodiesel can outcompete rapeseed biodiesel<!–[if supportFields]> XE “rapeseed biodiesel” <![endif]–><!–[if supportFields]><![endif]–> and fossil fuels, even with high production costs. This change would come as a result of biotechnology development, increasing crude oil prices, and high carbon pricing. Algal biodiesel has significantly lower external costs than the other analyzed fuel types. It is therefore concluded that environmental costs can be game changers when examining social costs. If environmental impacts are highly prioritized and enable support for biotechnology development through policy change and investment, microalgae could become the lowest cost fuel to society. 

Life Cycle Energy and Greenhouse Gas Emissions for an Ethanol Production Process Based on Blue-Green Algae

Microalgae-based biofuel production has become a major focus of renewable energy development. Much of this research has centered on biodiesel created from harvested algae. In this paper, Luo et al. (2010) investigated the life cycle energy use and greenhouse emissions for a different method of microalgae-based biofuel production in which ethanol synthesized via intracellular photosynthetic pathways is collected directly from un-harvested cyanobacteria (blue-green algae). This method is advantageous in that it has lower energy costs and water usage than that of harvest-based microalgae biodiesel production. Luo et al. used already available information for this process to complete the necessary engineering calculations and determine life cycle impacts. They found that this extraction method has significantly lower (67–87% less) greenhouse gas (GHG) emissions than gasoline, and that the net life cycle energy consumption can be as low as 0.20 MJ/MJEtOH, indicating that this process has great promise as a biofuel production method that can meet regulatory requirements and decrease environmental impact.—Karen de Wolski
Luo D., Hu Z., Choi D., Thomas V., Realff M., Chance R., 2010. Life cycle energy and greenhouse gas emissions for an ethanol production process based on blue-green algae. Environmental Science Technology 44, 8670–8677.

Luo et al. completed a life cycle analysis of blue-green algae-based ethanol production by calculating projected energy usage and GHG emissions using approximate parameters. The ethanol-producing blue-green algae are genetically enhanced photoautotrophic cyanobacteria that are cultivated in CO2 and fertilizer supplemented seawater in closed photobioreactors (PBR). Because the microalgae require CO2, the PBRs could be located near a fossil-fuel power plant or industrial facility that outputs CO2. The calculations in this paper are done under this assumption. When mature, the algae produce ethanol that diffuses into the growth culture. Dilute ethanol-freshwater solution is extracted from the growth medium and purified to fuel grade (99.7% pure) through a series of separation processes. One of the primary determinants of energy usage for this process is initial concentration of ethanol, as higher starting concentrations require less energy for purification. The researchers used concentrations between 0.5% and 5% (weight percent) as parameters for their calculations because 0.5% is too dilute to be cost effective and 5% would likely allow for economical recovery. The life cycle analysis additionally accounts for production and disposal of the PBRs, mixing in the bioreactors, biomass disposal, fertilizer production and transportation, ethanol separation (vapor compression steam stripping, vapor compression distillation, and molecular sieve), ethanol transportation, and combustion.
          Because Luo et al. did their calculations based on a range of initial ethanol concentrations, they had to apply ethanol separation processes that would be efficient for the entire range. Standard column distillation would be applicable to the upper end of the range, but is inefficient below 5%. Therefore, they used vapor compression steam stripping (VCSS) as the first separation method, which would concentrate the ethanol between 5% and 30%. For the second separation step, they used vapor compression distillation (VCD), which would result in a 94% ethanol concentration. For the final step (99.7% purification), the researchers chose molecular sieve dehydration. They used an equation accounting for mass flow rate and heat of evaporation to calculate energy required for vaporization in a steam-stripping column. Assuming an 80% efficiency rate for heat exchange and a 1% initial concentration, they found that the net heat input would be 0.18 MJ/MJEtOH. The stripper column requires electrically-powered steam compression, and the researchers estimated the work required for this process through an equation accounting for the gas constant, temperature, pressure, and adiabatic coefficient. For the 1% concentration reference case and 38% efficient electricity production, this value was estimated to be 0.11MJ/MJEtOH. For the molecular sieves stage of purification, heat requirement was found to be between 1 and 2 MJ/kgEtOH. The purification process accounts for the highest amount of energy consumption of this entire ethanol production method.
          The energy usage of several other processes was additionally estimated. PBRs must be mixed to ensure uniformity of algae and nutrients, and the energy requirement was estimated to be about 0.056MJ/MJEtOH for a 1% initial ethanol concentration. Compressor-mediated oxygen removal was calculated to require 0.0001 MJ/MJEtOH. Water must be pumped into the system and sterilized, and 3 mol of water are needed per 1 mol of ethanol. The energy requirements of water pumping and sterilization were therefore estimated. Energy consumed during delivery of CO2 into the PBRs, fertilizer transportation and production, bioreactor production, and waste biomass disposal were additionally calculated.
          Luo et al. also performed a GHG emissions life cycle analysis of this ethanol production process. Cellulosic renewable fuels must have less than 40% GHG emissions than petroleum-derived fuels according to the U.S. Energy Independence and Security Act. However, the act allows for funding for those fuels that have less than 20% of the greenhouse emissions of petroleum-derived fuel, making this figure the goal for biofuel development. The researchers considered grid electricity and on-site combined heat and power (CHP) from gas turbines as two possible electricity sources. It was estimated that grid electricity would produce 700 g CO2e/kWh, which would mean 13.5 CO2e/MJEtOH for on-site electricity alone. If on-site heat process heat was provided via natural gas, it would mean an additional 11 g CO2e/MJEtOH. The researchers found that CHP could greatly reduce GHG emissions in comparison to grid electricity due to increased energy efficiency. Emissions could be further reduced significantly if solar power were to be used for process heat supply.
          The researchers conclude that the 20% goal can be reached through this process if initial ethanol concentrations are sufficiently high. For a grid electricity and natural gas powered system, this value would have to be 4–4.5%. For a natural gas CHP system, the value could be 1.0–1.2%. The figure decreases to 0.8% if solar heating is used at 80% heat exchange efficiency. This manner of ethanol production has many advantages, including the ability to locate facilities on nonagricultural land, and the relatively low energy, fertilizer, and water requirements in comparison to harvest-based microalgae biofuel production. If initial ethanol concentrations can be made reliably high, the life cycle net energy input and GHG emissions are low enough to meet new regulatory standards. 

Enhancement Effect of Ethyl-2-methyl Acetoacetate on Triacylglycerols Production by a Freshwater Microalga, Scenedesmus sp. LX1

Declining fossil fuel reserves have led to the necessity of developing alternative, sustainable fuel sources. One of the most promising possibilities is microalgae, as they are a renewable resource and have relatively high lipid productivity compared to conventional biofuel feedstock sources. A foreseeable obstacle to the advancement of this technology is the high cost of microalgae-based biodiesel production. Xin et al. (2010) sought to elucidate whether the antialgal allelochemical ethyl-2-methyl acetoacetate (EMA) could induce high lipid accumulation in microalgal cells, thus making the biodiesel production process more efficient and cost-effective. They conducted the experiment on a microalgae species known to have high lipid content, Scenedesmus sp. LX1, and measured the biomass and lipid productivity post-EMA exposure. They found that EMA could increase triacylglycerol (TAG)  lipid and TAG productivity content by 79% and 40% respectively in this species, indicating EMA as a possible method of reducing the costs of microalgae biodiesel production.—Karen de Wolski
Xin, L., Hong-ying H., Jia Y., Yin-hu W., 2010. Enhancement effect of ethyl-2-methyl acetoacetate on triacylglycerols production by a freshwater microalga, Scenedesmus sp. LX1. Bioresource Technology 101, 9819-9821.

Xin et al. sought to experimentally determine whether the addition of the antialgal allelochemical EMA could induce higher TAG content per lipid and TAG productivity in the microalgae species Scenesdesmus sp. LX1. EMA, a compound isolated from the reed Phragmites communis, has been found to inhibit the growth of Chlorella pyrenoidosa and Microcystis aeruginosa, and the researchers therefore believed that, as an environmental stressor, EMA may result in higher lipid content in microalgae. They chose Scenesdesmus sp. LX1 as the experimental subject because it had been previously isolated and is known to have high lipid content. The microalgae were grown in different concentrations of EMA culture and the TAG content per lipid and TAG productivity were measured and calculated.
          All microalgae cultures were grown in the same light intensity, light/dark periods, humidity, and temperature. The researchers added EMA to the growth medium in four different concentrations (0, 0.25, 0.5, 1.0, and 2.0 mg/L). Densities of the microalgae cultures were determined by measuring optical density (OD650) every 24 hours. These data were then extrapolated to a logistic model to find algal growth. Biomass and total lipid content were also measured. The experimenters then dissolved the dried lipids in isopropyl alcohol, and they estimated TAGs content by an enzymatic colorimetric method. Significant difference analyses were carried out by Independent-Samples t-tests.
          Xin et al. found that none of the experimental EMA concentrations had a significant effect on microalgae growth.  Through linear regression analysis, they also calculated that carrying capacity of culture was not significantly affected by different EMA concentrations. Lipid content per biomass and TAGs content per lipid were measured after 19 days of cultivation. Different EMA concentration did not induce significantly different biomasses (all about 30%). However, it was found that TAG accumulation was significantly higher (p<0.01) in microalgae grown in EMA concentrations of 0.5, 1.0, and 2.0 mg/L (34.8%, 78.3%, and 79.1% respectively). Additionally, TAG productivity was significantly greater in the microalgae samples grown in 1.0 and 2.0 mg/L EMA concentrations.
          These results are promising because they indicate that microalgae productivity can be enhanced by EMA without hindering growth rate. This could potentially greatly reduce costs of microalgae biodiesel production. The researchers conclude that, while the exact chemical mechanisms need to be further studied, this experiment partially elucidates a possible method of overcoming the cost-barrier of wide-scale microalgae cultivation. 

Life-Cycle Analysis on Biodiesel Production from Microalgae: Water Footprint and Nutrients Balance

The combination of both federal government-mandated and individual state renewable energy standards, and the increasing body of evidence that feedstock-based biofuels are unsustainable, have led to the critical evaluation of the feasibility of microalgae as an alternative biofuel source. While microalgae show great promise in their relatively low land requirements, high growth rate, and CO2 absorption abilities, there remain outstanding questions regarding water consumption rates, especially in comparison to current feedstocks. In their 2011 study, Yang et al. use known measurements of algal growth parameters, such as evaporation rate, growth rate, and nutrient usage, to quantify the water footprint and nutrient balance of Chlorella vulgaris, a species of microalgae.  They found that microalgae are competitive with traditional feedstocks in terms of total water footprint, and that freshwater and nutrient consumption could be significantly reduced by using seawater and waste water as the base water source. The researchers additionally analyzed spatial variation of microalgae growth in terms of solar radiation and temperature, and found that the water footprint would be lowest in the states of Florida, Hawaii, and Arizona.—Karen de Wolski
Yang J., Xu M., Hu Q., Sommerfeld M., Chen Y., 2011. Life-cycle analysis on biodiesel production from microalgae: water footprint and nutrients balance. Bioresource Technology 102, 159–165.

Yang et al. outline the feasibility of microalgae as a biofuel source in terms of water footprint and nutrient usage by drawing from previously gathered metrics to estimate the water and nutrient consumption of Chlorella vulgaris. The 2007 Energy Independence and Security Act  requires that renewable fuel production increase to 36 billion gallons per year by 2022. The first generation biofuels derived from corn and sugarcane (Brazil) are limited in that they require significant arable land, increase food prices internationally, and do not necessarily significantly reduce carbon emissions. Microalgae is promising as an alternative fuel source because of its high growth rate, smaller land usage (15–300 times more oil per land unit), high lipid content, and CO2 absorption abilities. However, microalgae are not yet grown on a mass scale, and several outstanding questions remain regarding the life-cycle impacts of large-scale cultivation and biodiesel production.  This study seeks to elucidate the water footprint of biodiesel production from microalgae through quantitative measurements and comparisons with current biodiesel production from feedstocks. The researchers calculate the differences between using seawater, wastewater, and freshwater as a culture base, and they additionally account for nutrient usage and the effects of solar radiation and temperature variation in microalgae cultivation.
Microalgae biodiesel production necessarily entails culture, harvest, drying, extraction, and esterification. Microalgae are initially grown in culture water (sea, waste, or fresh) in an open pond which must be constantly replenished with freshwater to accommodate evaporation and maintain salinity. When sufficiently grown, the microalgae are harvested, dried, and the lipids are extracted to be esterified for biodiesel. Culture water can be partially recycled directly back into a culture pond, and/or discharged into a wastewater treatment system.
Evaporation is the main source of water loss during the culture process, and the authors used lake evaporation rate to approximate open pond evaporation. Microalgal growth is affected by temperature and solar radiation, and data from the national solar radiation database were usd to make the appropriate calculations. Additionally, culture ponds need to be supplied with nutrients, and necessary nutrient concentrations were based on measurements from previous studies. Both harvesting and drying, the second and third steps of the process respectively, are quantified by solid content (ratio of microalgae to water) and recovery rate (ratio of harvested mass to mass after culture). These values were derived from several already established parameters. Because the extraction and esterification of microalgae is similar to that of soybeans, this study substituted water usage rates for biodiesel production from soybean oil (2–10 liters water used per liter of biodiesel produced) for the analogous water usage of biodiesel production from microalgae-derived oil.
The researchers found that, in the absence of recycling harvested water, the water footprint of microalgae biodiesel production is 3726 kg-water/kg-biodiesel. This value can be decreased to 591 kg-water/kg-biodiesel if all harvest water is recycled. The amount of harvest water recycled does not affect the water footprint of the other production processes. Additionally, they found that using seawater or wastewater can reduce the life-cycle freshwater usage by up to 90%. Harvest water recycling can also decrease nutrient (nitrogen, phosphorous, potassium, magnesium, and sulfur) usage by approximately 55%. The use of sea/wastewater for algal culture can additionally reduce nitrogen usage by 94% and abolish the necessity of adding potassium, magnesium and sulfur. A sensitivity analysis revealed evaporation rate, algal lipid content, and slurry content as the most sensitive parameters, while variations in these factors show growth rate to be the most sensitive factor.
When these results are compared to the water footprint of feedstock-based biofuel, microalgae are demonstrated to be extremely competitive. The authors discuss how differences in species could cause significant variation in these parameters. Increases in both lipid content and growth rate, the two most important parameters, result in the reduction of the water footprint. These two parameters are, however, usually negatively related to each other, and vary depending on species.  This study calculated the water footprint for eleven other species using the same methodology and found that the water footprint could be 1–6 times higher than that of C. vulgaris.
Geographic variations in solar radiation, temperature, and evaporation must be accounted for when estimating microalgae growth. While microalgae tend to prosper in high temperature/high solar radiation environments, both of these factors are proportionally related to evaporation rate, and therefore cause an increase in water footprint. Taking these factors into account, the researchers calculated that Florida, Hawaii, and Arizona would have the lowest water footprints for microalgae biodiesel production in the United States.
They conclude that, in terms of water footprint and nutrient usage, microalgae could be a feasible alternative to current feedstocks for biodiesel production. The advancement of technology, especially of photobioreactors for cultured growth, could enhance water conservation and increase cultivation efficiency. The use of sea/waste water for culture water would decrease water usage by 90% and greatly reduce the need for nutrient supplementation. Phosphate could prove to be a limiting factor, as it is not found in either of the aforementioned water types, and global phosphate sources are decreasing. However, experimentation with different microalgae species types and phosphate-rich water could overcome this barrier. While much remains to be done before microalgae can be used as a global renewable energy source, freshwater and nutrient usage does not appear to be a limiting obstacle.  

Microalgae for Biodiesel Production and Other Applications: a Review

As fossil fuel reserves decline and atmospheric CO2 concentration increases, the need to find renewable sustainable energy sources has become pressing. The current generation of biofuels, derived from food crops and oil seeds, has relatively low negative environmental effects from its consumption, but it is limited in its economic feasibility because it relies on drawing from food supplies. Microalgae is becoming increasingly viewed as an alternative source of biomass for biodiesel creation, as it has many biological and economic advantages. In their 2010 review article, Mata et al. outline the viability of microalgae as a biofuel source, discussing areas that need to be considered and further substantiated before microalgae can be used for wide scale fuel production. Additionally, they highlight other ways that microalgae may be used to ameliorate various climate and health problems, including flue gas COemission mitigation, wastewater purification, and medicinal applications. They conclude that, while having significant potential as an economically and environmentally feasible source of biodiesel, there remains a substantial amount of scientific research and technological advancement that must be completed before microalgae can be cultivated and processed on a large-scale.—Karen de Wolski
Mata T., Martins A., Caetano N. 2010. Microalgae for biodiesel production and other applications: a review. Renewable and Sustainable Energy Reviews 14, 217–232.

 Mata et al. emphasize the dire need to find renewable and sustainable energy alternatives, listing the negative effects of global warming and high atmospheric CO2 concentration and implicating the burning of fossil fuels as the main source of greenhouse gas (GHG) emissions. Biofuels are illustrated as a more environmentally friendly alternative, as they have lower combustion emissions per unit than diesel and gasoline. However, current biofuels are limited because they draw from foodstuffs, and they are therefore not only expensive but also lead to an increase in food prices while lowering food availability worldwide. Biodiesel, comprising 82% of total biofuels production and derived from vegetable oils and animal fats, cannot meet the current market demands, as both land availability and feed stocks are insufficient. In order to meet demand and ameliorate global warming, it is therefore necessary to utilize an alternative biomass source that is less costly to cultivate and has lower land requirements. Microalgae is currently under investigation as such a source, as it can provide feedstock for multiple biofuels, including biodiesel, ethanol, and methane, while having relatively low cultivation requirements. Mata et al. focus primarily on microalgae for biodiesel production, outlining what is already known about microalgae for this use, and what still needs to be elucidated in order to utilize it on a large-scale.
          It is estimated that 50,000 species of microalgae exist, and these prokaryotic and eukaryotic organisms are able to thrive in a variety of harsh conditions with high growth and reproduction rates. Extensive collections of algal species have been gathered at many research institutions, the largest being 4000 strains of 1000 species at the University of Coimbra. Microalgae are relatively easy to grow, requiring little attention, able to utilize non potable water, and needing 49–132 times less land than rapeseed or soy. They reproduce rapidly, completing a growth cycle every few days, and different species are adapted to live in different conditions, making them a versatile organism for cultivation. Microalgae as a source of biodiesel are additionally advantageous in that they can be simultaneously used to remove CO2 from industrial flue gases by bio-fixation, remove contaminants from wastewater, and provide other useful compounds for various industrial and pharmaceutical purposes. The large-scale production of biodiesel from microalgae could therefore serve multiple positive functions while meeting energy demands.
          While the large-scale culture of microalgae began in the 1960s in Japan, study of them as a renewable energy source has been limited by insufficient funding. An R&D program in the United States, carried out between 1978 and 1996, concluded that microalgae as a low-cost biodiesel source was technically feasible, but would require long term investigation and development to achieve. This project began to identify algal strains with particularly high lipid content for oil extraction and to screen for genetic variability between algal isolates, but the research was curtailed in 1995 by a funding cut. Microalgae research has been brought back to the forefront recently by high crude oil prices and the prevalent need to find less environmentally devastating sources of energy. Most contemporary studies focus on the genetic engineering of microalgae to optimize cultivation success and oil production, with an especial concentration on maximizing lipid content.
          Current microalgae to biodiesel production processes consist of cell growth in a production unit, cell separation from the growth media, and lipid extraction. From these lipids, biodiesels can be created by similar methods to those for other biofuel feedstocks. A key step in cultivation is site selection, which must take into account carbon and light availability, and the metabolic requirements of the algal species of interest. Some species have proven difficult to cultivate at high volume, and light and temperature are the most limiting factors for successful growth. Additionally, salinity, turbulence, and contaminants must be controlled as required by each individual species. The authors list a series of studies by different researchers investigating the effects of various growth conditions, including pH, and the concentrations of CO2,  iron, and nitrogen. It is emphasized that different species have relatively individualized requirements, and these must be understood if microalgae cultivation is to be maximized.
          Biomass recovery, or “harvesting,” can make up to 30% of biomass production cost, but no single method has proven to be optimal. The development of such a method does, however, remain an important area of interest. Harvesting can include sedimentation, centrifugation, filtration, flocculation, and flotation. The two main criteria for harvest method selection are identified as desired product quality and acceptable moisture level. Sedimentation can be used for lower quality products, while the more costly centrifugation produces higher value products. Processing is also an expensive production cost, and it consists of dehydration and cell disruption for metabolite release. Several extraction methods, including solvents, ultrasound, and microwave are currently used or under investigation. Biodiesels are produced from the lipids by a transesterifaction reaction, in which tricglycerides are converted to esters (biodiesel) and glycerol (by-product) through a multiple step chemical reaction. Current industrial transesterification processes are carried out in a stirred reactor in batch mode, although improvements have been proposed that would enable continuous mode production, allowing for decreased reaction time.
          Other considerations for microalgae cultivation include culture system type and operation mode. Open-culture systems, such as lakes, are easier and cheaper to build and operate. They are, however, more difficult to regulate and therefore could be limited in large-scale microalgae cultivation abilities. Closed-culture systems, or photo-bioreactors (PBR), while much more expensive, are more flexible, allowing for optimization of pH, temperature, evaporation levels, and CO2 loss. It has been shown that PBRs have higher volumetric productivity and cell concentrations than open-culture systems, but congruent areal productivity, and the competition between the two technologies is therefore not necessarily as important as the genetic engineering of microalgae to appropriately fit each system type. The second aforementioned cultivation consideration, operation mode, centers on batch versus continuous PBR operation. Continuous mode offer higher degree of control over conditions, and hence can produce more reliable and higher quality results. However, different bio-reaction types are better suited than others to this operation type, and this therefore needs to be understood when designing a cultivation process for any given microalgae species.
          The review ends with a brief discussion of other potential microalgae applications. Flue gases from power plants account for over 7% of world CO2 emissions, and microalgae could be used to diminish these emissions through natural bio-fixation processes. Microalgae could also be used for waste water treatment, as they require many common contaminants (nitrogen and phosphorous) as nutrient sources. Additionally, many species contain chemical compounds that can be used for both industrial and health applications. These include pigments, antioxidants, vitamins, and food additives. These compounds could be extracted from the microalgae processed for biodiesel production, and could therefore potentially offer multiple simultaneous benefits. Several nutritional supplements have also been found in microalgae, including sterols and carotenoids, which could prove highly beneficial for human health. There is also promising evidence to show that microalgae culture could be a significant source of food for aquatic animal rearing. Microalgae can be utilized to culture zooplankton, the food source of farmed crustaceans and finfish. Further understanding of the nutritional value of microalgae could hence be widely applied to aquaculture.

          The authors conclude that microalgae as a biodiesel source has great promise as a sustainable and environmentally friendly alternative to the current food crop and seed oil derived biodiesels. While much is already understood about the processes required for mass cultivation, much remains unknown. A significant amount of investment and research will therefore be necessary to develop this technology. Given the importance of reducing GHG emissions and supplementing fossil fuels, this capital investment into microalgae development could prove vital for human energy sustainability.