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20/07/2012 Effects of cultivation conditions and media composition on cell growth and lipid productivity of indigenous microalga Chlorella vulgaris ESP-31
Yeh KL, Chang JS.
Publication The growth and lipid productivity of an isolated microalga Chlorella vulgaris ESP-31 were investigated under different media and cultivation conditions, including phototrophic growth (NaHCO(3) or CO(2), with light), heterotrophic growth (glucose, without light), photoheterotrophic growth (glucose, with light) and mixotrophic growth (glucose and CO(2), with light). C. vulgaris ESP-31 preferred to grow under phototrophic (CO(2)), photoheterotrophic and mixotrophic conditions on nitrogen-rich medium (i.e., Basal medium and Modified Bristol's medium), reaching a biomass concentration of 2-5 g/l. The growth on nitrogen-limiting MBL medium resulted in higher lipid accumulation (20-53%) but slower growth rate. Higher lipid content (40-53%) and higher lipid productivity (67-144 mg/l/d) were obtained under mixotrophic cultivation with all the culture media used. The fatty acid composition of the microalgal lipid comprises over 60-68% of saturated fatty acids (i.e., palmitic acid (C16:0), stearic acid (C18:0)) and monounsaturated acids (i.e., oleic acid (C18:1)). This lipid composition is suitable for biodiesel production.
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20/07/2012 Biofilm establishment and heavy metal removal capacity of an indigenous mining algal-microbial consortium in a photo-rotating biological contactor
Orandi S, Lewis DM, Moheimani NR.
Publication An indigenous mining algal-microbial consortium was immobilised within a laboratory-scale photo-rotating biological contactor (PRBC) that was used to investigate the potential for heavy metal removal from acid mine drainage (AMD). The microbial consortium, dominated by Ulothrix sp., was collected from the AMD at the Sar Cheshmeh copper mine in Iran. This paper discusses the parameters required to establish an algal-microbial biofilm used for heavy metal removal, including nutrient requirements and rotational speed. The PRBC was tested using synthesised AMD with the multi-ion and acidic composition of wastewater (containing 18 elements, and with a pH of 3.5 ± 0.5), from which the microbial consortium was collected. The biofilm was successfully developed on the PRBC's disc consortium over 60 days of batch-mode operation. The PRBC was then run continuously with a 24 h hydraulic residence time (HRT) over a ten-week period. Water analysis, performed on a weekly basis, demonstrated the ability of the algal-microbial biofilm to remove 20-50 % of the various metals in the order Cu > Ni > Mn > Zn > Sb > Se > Co > Al. These results clearly indicate the significant potential for indigenous AMD microorganisms to be exploited within a PRBC for AMD treatment.
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20/07/2012 Oil accumulation is controlled by carbon precursor supply for fatty acid synthesis in Chlamydomonas reinhardtii
Fan J, Yan C, Andre C, Shanklin J, Schwender J, Xu C.
Publication Microalgal oils have attracted much interest as potential feedstocks for renewable fuels, yet our understanding of the regulatory mechanisms controlling oil biosynthesis and storage in microalgae is rather limited. Using Chlamydomonas reinhardtii as a model system, we showed here that starch, rather than oil, is the dominant storage sink for reduced carbon under a wide variety of conditions. In short-term treatments, significant amounts of oil were found to be accumulated concomitantly with starch only under conditions of N starvation, as expected, or in cells cultured with high acetate in otherwise standard growth medium. Time-course analysis revealed that oil accumulation under N starvation lags behind that of starch and rapid oil synthesis occurs only when carbon supply exceeds the capacity of starch synthesis. In the starchless mutant BAFJ5, blocking starch synthesis results in significant increases in the extent and rate of oil accumulation. In the parental strain, but not the starchless mutant, oil accumulation under N starvation was strictly dependent on available external acetate supply and the amount of oil increased steadily as the acetate concentration increased to the levels several-fold higher than that of the standard growth medium. Additionally, oil accumulation under N starvation is saturated at low light intensities and appears to be largely independent of de novo protein synthesis. Collectively, our results suggest carbon availability is a key metabolic factor controlling oil biosynthesis between starch and oil in Chlamydomonas.
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20/07/2012 Performance of a sand filter in removal of micro-algae from seawater in aquaculture production systems
Sabiri NE, Castaing JB, Massé A, Jaouen P.
Publication In this study, a sand filter was used to remove micro-algae from seawater feeding aquaculture ponds. A lab-scale sand filter was used to filter 30,000 cells/mL of Heterocapsa triquetra suspension, a non-toxic micro-alga that has morphological and dimensional (15-20 microm) similarities with Alexandrium sp., one of the smallest toxic micro-algae in seawater. Removal efficiency and capture mechanisms for a fixed superficial velocity (3.5 m/h) were evaluated in relation to size distribution and mean diameter of the sand. Various sands (average diameter ranging between 200 microm and 600 microm) were characterized and used as porous media. The structural parameters of the fixed beds were evaluated for each medium using experimental measurements of pressure drop as a function of superficial velocity over a range of Reynolds numbers covering Darcy's regime and the inertial regime. For a filtration cycle of six hours, the best efficiency (E = 90%) was obtained with the following sand characteristics: sieved sand with a range of grain diameter of 100 and 300 microm and a mean grain diameter equal to 256 microm. Results obtained show the influence of the size distribution of sand on the quality of retention of the micro-algae studied.
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20/07/2012 Enhancement of neutral lipid productivity in the microalga Isochrysis affinis Galbana (T-Iso) by a mutation-selection procedure
Bougaran G, Rouxel C, Dubois N, Kaas R, Grouas S, Lukomska E, Le Coz JR, Cadoret JP.
Publication Microalgae offer a high potential for energetic lipid storage as well as high growth rates. They are therefore considered promising candidates for biofuel production, with the selection of high lipid-producing strains a major objective in projects on the development of this technology. We developed a mutation-selection method aimed at increasing microalgae neutral lipid productivity. A two step method, based on UVc irradiation followed by flow cytometry selection, was applied to a set of strains that had an initial high lipid content and improvement was assessed by means of Nile-red fluorescence measurements. The method was first tested on Isochrysis affinis galbana (T-Iso). Following a first round of mutation-selection, the total fatty acid content had not increased significantly, being $262\pm 21\,{\rm mgTFA}\,{\rm (gC)}^{- {\rm 1}} $ for the wild type (WT) and $269\pm 49\,{\rm mgTFA}\,{\rm (gC)}^{- {\rm 1}} $ for the selected population (S1M1). Conversely, fatty acid distribution among the lipid classes was affected by the process, resulting in a 20% increase for the fatty acids in the neutral lipids and a 40% decrease in the phospholipids. After a second mutation-selection step (S2M2), the total fatty acid content reached $409\pm 64\,{\rm mgTFA}\,{\rm (gC)}^{- {\rm 1}} $ with a fatty acid distribution similar to the S1M1 population. Growth rate remained unaffected by the process, resulting in a 80% increase for neutral lipid productivity. Biotechnol. Bioeng. © 2012 Wiley Periodicals, Inc.
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20/07/2012 Nannochloropsis production metrics in a scalable outdoor photobioreactor for commercial applications
Quinn JC, Yates T, Douglas N, Weyer K, Butler J, Bradley TH, Lammers PJ.
Publication Commercial production of renewable energy feedstocks from microalgae will require reliable and scalable growth systems. Two and one half years of biomass and lipid productivity data were obtained with an industrial-scale outdoor photobioreactor operated in Fort Collins, Colorado (USA). The annualized volumetric growth rates for Nannochloropsis oculata (CCMP 525) and Nannochloropsis salina (CCMP 1776) were 0.16gL(-1)d(-1) (peak=0.37gL(-1)d(-1)) and 0.15gL(-1)d(-1) (peak=0.37gL(-1)d(-1)) respectively. The collective average lipid production was 10.7m(3)ha(-1)yr(-1) with a peak value of 36.3m(3)ha(-1)yr(-1). Results from this study are unique based on publication of biomass and corresponding lipid content combined with demonstration of energy savings realized through analysis of gas delivery requirements, water recycling from successive harvests with no effect on productivity, and culture stability through serial batch lineage data and chemotaxonomic analysis of fatty acid contents.
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20/07/2012 Membrane-based energy efficient dewatering of microalgae in biofuels production and recovery of value added co-products
Bhave R, Kuritz T, Powell L, Adcock D.
Publication The objective of this paper is to describe the use of membranes for energy efficient biomass harvesting and dewatering. The dewatering of Nannochloropsis sp. was evaluated with polymeric hollow fiber and tubular inorganic membranes to demonstrate the capabilities of a membrane-based system to achieve microalgal biomass of >150 g/L (dry wt.) and ∼99% volume reduction through dewatering. The particle free filtrate containing the growth media is suitable for recycle and reuse. For cost-effective processing, hollow fiber membranes can be utilized to recover 90-95% media for recycle. Tubular membranes can provide additional media and water recovery to achieve target final concentrations. Based on the operating conditions used in this study and taking into scale-up considerations, an integrated hollow fiber-tubular membrane system can process microalgal biomass with at least 80% lower energy requirement compared to traditional processes. Backpulsing was found to be an effective flux maintenance strategy to minimize flux decline at high biomass concentration. An effective chemical cleaning protocol was developed for regeneration of fouled membranes.
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20/07/2012 Obtaining lutein-rich extract from microalgal biomass at preparative scale
Fernández-Sevilla JM, Fernández FG, Grima EM.
Elément Lutein extracts are in increasing demand due to their alleged role in the prevention of degenerative disorders such as age-related macular degeneration (AMD). Lutein extracts are currently obtained from plant sources, but microalgae have been demonstrated to be a competitive source likely to become an alternative. The extraction of lutein from microalgae posesses specific problems that arise from the different structure and composition of the source biomass. Here is presented a method for the recovery of lutein-rich carotenoid extracts from microalgal biomass in the kilogram scale.
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20/07/2012 Ketocarotenoid biosynthesis in transgenic microalgae expressing a foreign β-C-4-carotene oxygenase gene
Vila M, Galván A, Fernández E, León R.
Publication Ketocarotenoids are obtained by the action of the β-carotene ketolase, which catalyses the addition of a keto-group at the C4 position of carotenoids β-ion-rings. Most microalgae and higher plants do not posses the carotene ketolase activity and consequently do not synthesize ketocarotenoids, which are highly demanded as feed supplements and as nutraceutical for human nutrition. Here we propose the use of the unicellular microalga Chlamydomonas reinhardtii to express the Bkt (β-carotene ketolase) gene from Haematococcus pluvialis and synthesize a new ketocarotenoid that the vegetative cells of Chlamydomonas do not synthesize in the natural way. The methodology needed to successfully achieve metabolic engineering of ketocarotenoids synthesis in Chlamydomonas is described in this chapter, including the construction of a vector containing the Bkt gene, transformation of Chlamydomonas, selection of transformants, and carotenoids analysis.
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20/07/2012 Pathways of carotenoid biosynthesis in bacteria and microalgae
Paniagua-Michel J, Olmos-Soto J, Ruiz MA.
Publication The carotenoids, a subfamily of the isoprenoids, are among the most widespread, ancient, diverse, and rich class of all natural products and biomolecules. Microorganisms, as well as microalgae and bacteria synthesize isoprenoids from isopentenyl diphosphate (IPP) and its isomer dimethylallyl diphosphate (DMAPP). For long time, mevalonic acid was assumed to be the only natural precursor for IPP and DMAPP in the cytosolic acetate/mevalonate pathway for the biosynthesis of sterols, sesquiterpenes, triterpenoids, and carotenoids. At present, it is accepted that the relatively new route, the methylerythritol 4-phosphate (MEP), or 1-deoxy-D: -xylulose-5-phosphate (DOXP) is the main pathway for the biosynthesis of plastidic isoprenoids, such as carotenoids, phytol (a side chain of chlorophylls), plastoquinone-9, isoprene, mono-, and diterpenes. Cytosolic isoprenoids (sterols) biosynthesized by MEP have been reported in eubacteria and algae (Chlorella, Chlamydomonas, Scenedesmus, and Dunaliella). This review summarizes current knowledge of the biosynthetic pathways leading to the formation of different isoprenoids and carotenoids in bacteria and microalgae. Particular attention was given to the last early steps of the biosynthesis of the key C(5)-precursor and the final steps of the biosynthesis of carotenoids including selected examples in microalgae and bacteria as well as the recent advances in genomics and metabolic engineering.
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20/07/2012 Co-pyrolysis characteristics of microalgae Chlorella vulgaris and coal through TGA
Chen C, Ma X, He Y.
Publication To find out an alternative of coal saving, a kind of microalgae, Chlorella vulgaris (C. vulgaris) which is widespread in fresh water was introduced into coal pyrolysis process. In this work, the pyrolysis experiments of C. vulgaris and coal blend (CCB) were carried out by TGA, and those of C. vulgaris and coal were also taken respectively as control groups. It was found that: the TG and DTG profiles of CCB were similar to C. vulgaris, but different from coal under various blending ratios; DTG profiles of CCB were different at several heating rates; interaction was observed between the solid phases of CCB; kinetic triplets were determined by the Kissinger-Akahira-Sunose (KAS), Flynn-Wall-Ozawa (FWO), and master-plots method, respectively. The results provide a reference for further study on co-pyrolysis of microalgae and coal to a certain extent.
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20/07/2012 Production of biofuels from pretreated microalgae biomass by anaerobic fermentation with immobilized Clostridium acetobutylicum cells
Efremenko EN, Nikolskaya AB, Lyagin IV, Senko OV, Makhlis TA, Stepanov NA, Maslova OV, Mamedova F, Varfolomeev SD.
Publication The purpose of this work was to study the possible use of pretreated biomass of various microalgae and cyanobacteria as substrates for acetone-butanol-ethanol (ABE) fermentation by Clostridium acetobutylicum cells immobilized into poly(vinyl alcohol) cryogel. To this end, the biochemical composition of photosynthetic microorganisms cultivated under various conditions was studied. The most efficient technique for pretreating microalgal biomass for its subsequent conversion into biofuels appeared to be thermal decomposition at 108 °C. For the first time the maximum productivity of the ABE fermentation in terms of hydrogen (8.5 mmol/L medium/day) was obtained using pretreated biomass of Nannochloropsis sp. Maximum yields of butanol and ethanol were observed with Arthrospira platensis biomass used as the substrate. Immobilized Clostridium cells were demonstrated to be suitable for multiple reuses (for a minimum of five cycles) in ABE fermentation for producing biofuels from pretreated microalgal biomass.
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20/07/2012 Effects of hydraulic and solids retention times on productivity and settleability of microbial (microalgal-bacterial) biomass grown on primary treated wastewater as a biofuel feedstock
Valigore JM, Gostomski PA, Wareham DG, O'Sullivan AD.
Publication High biomass productivity and efficient harvesting are currently recognized challenges in microbial biofuel applications. To produce naturally settleable biomass, combined growth of native microalgae and bacteria was facilitated in laboratory sequencing batch reactors (SBRs) using primary treated wastewater from the Christchurch Wastewater Treatment Plant (CWTP) in New Zealand. SBRs were operated under a simulated, local, summer climate (i.e., 925 μmol/m(2)/s of photosynthetically active radiation for 14.7 h per day at 21 °C mean water temperature) using 1.4- to 8-day hydraulic retention times (HRTs) to optimize growth. Solids retention times (SRTs) were varied from 4 to 40 days by discharging different ratios of supernatant and completely mixed culture. Biomass productivity up to 31 g/m(2)/day of solids was obtained, and it generally increased as retention times decreased. Biomass settleability was typically 70-95%, and the microbes aggregated into compact flocs as cultures aged up to four months. Due to a low lipid content of 10.5%, anaerobic digestion appeared to be the most appropriate biofuel conversion process with potential to generate 19,200 m(3)/ha/yr of methane based on settleable mixture productivity.
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20/07/2012 Continuous microalgae cultivation in a photobioreactor.
Tang H, Chen M, Simon Ng KY, Salley SO.
Publication New biomass sources for alternative fuels has become a subject of increasing importance as the nation strives to resolve the economic and strategic impacts of limited fossil fuel resources on our national security, environment, and global climate. Algae are among the most promising non-food-crop-based biomass feedstocks. However, there are currently no commercially viable microalgae-based production systems for biofuel production that have been developed, as limitations include less-than optimal oil content, growth rates, and cultivation techniques. While batch studies are critical for determining basic growth phases and characteristics of the algal species, steady-state studies are necessary to better understand and measure the specific growth parameters. This study evaluated the effects of dilution rate on microalgal biomass productivity, lipid content, and fatty acid profile under steady-state conditions with continuous illumination and carbon dioxide supplemention for two types of algae. Continuous cultures were conducted for more that 3 months. Our results show that the productivity of Chlorella minutissima varied from 39 to 137 mg/L/day (dry mass) when the dilution rate varied from 0.08 to 0.64 day(-1) . The biomass productivity of C. minutissima reached a maximum value (137 mg/L/day) at a dilution rate of 0.33 day(-1) , while the productivity of Dunaliella tertiolecta varied from 46 to 91 mg/L/day at a dilution rate of 0.17 to 0.74 day(-1) . The biomass productivity of D. tertiolecta reached a maximum value of 91 mg/L/day at a dilution rate of 0.42 day(-1) . Moreover, the lipid content had no significant change with various dilution rates. Biotechnol. Bioeng. © 2012 Wiley Periodicals, Inc.
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20/07/2012 A novel method to harvest microalgae via co-culture of filamentous fungi to form cell pellets
Zhang J, Hu B.
Publication While current approaches have limitations for efficient and cost-effective microalgal biofuel production, new processes, which are financially economic, environmentally sustainable, and ecologically stable, are needed. Typically, microalgae cells are small and grow individually. Harvest of these cells is technically difficult and it contributes to 20-30% of the total cost of biomass production. A new process of pelletized cell cultivation is described in this study to co-culture a filamentous fungal species with microalgae so that microalgae cells can be co-pelletized into fungal pellets for easier harvest. This new process can be applied to microalgae cultures in both autotrophic and heterotrophic conditions to allow microalgae cells attach to each other. The cell pellets, due to their large size, can be harvested through sieve, much easier than individual cells. This method has the potential to significantly decrease the processing cost for generating microagal biofuel or other products.
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20/07/2012 Oral administration of an enzymatic protein hydrolysate from the green microalga Chlorella vulgaris enhances the nutritional recovery of malnourished mice
Morris HJ, Carrillo OV, Alonso ME, Bermúdez RC, Almarales Á, Llauradó G, Lebeque Y, Fontaine R.
Publication This study examined the effects of oral administration of an enzymatic protein hydrolysate from green microalga Chlorella vulgaris (Cv-PH) on the nutritional recovery of malnourished Balb/c mice after a 3-day fasting period. Mice were refed with commercial diet supplemented or not supplemented with Cv-PH (500 mg/kg) for 8 days. Regardless of the diet used during refeeding, animal body weights and serum protein concentrations did not differ between groups. Mice given Cv-PH had a significant increase in hemoglobin concentrations. Most serum amino acid levels were similar in the control and Cv-PH animals. Starved mice refed with Cv-PH showed normal liver functions, as judged by liver weight, protein concentration, and the enzymatic activities of cholinesterase and arginase. Cv-PH increased DNA, protein content, and gut-mucosal weight. In addition, brush-border oligosaccharidase activities were also higher in the Cv-PH group. These findings suggest that Chlorella protein hydrolysate can be used to develop specific formulations suitable for pharmacologic nutrition.
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20/07/2012 Early detection of protozoan grazers in algal biofuel cultures
Day JG, Thomas NJ, Achilles-Day UE, Leakey RJ.
Publication Future micro-algal biofuels will most likely be derived from open-pond production systems. These are by definition open to "invasion" by grazers, which could devastate micro-algal mass-cultures. There is an urgent requirement for methodologies capable of early detection and control of grazers in dense algal cultures. In this study a model system employing the marine alga Nannochloropsis oculata was challenged by grazers including ciliates, amoebae and a heterotrophic dinoflagellate. A FlowCAM flow-cytometer was used to detect all grazers investigated (size range <20->80 μm in length) in the presence of algae. Detection limits were <10 cells ml(-1) for both "large" and "small" model grazers, Euplotes vannus (80 × 45 μm) and an unidentified holotrichous ciliate (~18 × 8 μm) respectively. Furthermore, the system can distinguish the presence of ciliates in N. oculata cultures with biotechnologically relevant cell densities; i.e. >1.4 × 10(8) cells ml(-1) (>0.5 g l(-1) dry wt.).
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20/07/2012 Cyanobacterial biofuel production
Machado IM, Atsumi S.
Publication The development of new technologies for production of alternative fuel became necessary to circumvent finite petroleum resources, associate rising costs, and environmental concerns due to rising fossil fuel CO(2) emissions. Several alternatives have been proposed to develop a sustainable industrial society and reduce greenhouse emissions. The idea of biological conversion of CO(2) to fuel and chemicals is receiving increased attention. In particular, the direct conversion of CO(2) with solar energy to biofuel by photosynthetic microorganisms such as microalgae and cyanobacteria has several advantages compared to traditional biofuel production from plant biomass. Photosynthetic microorganisms have higher growth rates compared with plants, and the production systems can be based on non-arable land. The advancement of synthetic biology and genetic manipulation has permitted engineering of cyanobacteria to produce non-natural chemicals typically not produced by these organisms in nature. This review addresses recent publications that utilize different approaches involving engineering cyanobacteria for production of high value chemicals including biofuels.
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20/07/2012 Analysis and Design of Photobioreactors for Microalgae Production II: Experimental Validation of a Radiation Field Simulator Based on a Monte Carlo Algorithm
Heinrich JM, Niizawa I, Botta FA, Trombert AR, Irazoqui HA.
Publication In a previous study, we developed a methodology to assess the intrinsic optical properties governing the radiation field in algae suspensions. With these properties at our disposal, a Monte Carlo simulation program is developed and used in this study as a predictive autonomous program applied to the simulation of experiments that reproduce the common illumination conditions that are found in processes of large scale production of microalgae, especially when using open ponds such as raceway ponds. The simulation module is validated by comparing the results of experimental measurements made on artificially illuminated algal suspension with those predicted by the Monte Carlo program. This experiment deals with a situation that resembles that of an open pond or that of a raceway pond, except for the fact that for convenience, the experimental arrangement appears as if those reactors were turned upside down. It serves the purpose of assessing to what extent the scattering phenomena are important for the prediction of the spatial distribution of the radiant energy density. The simulation module developed can be applied to compute the local energy density inside photobioreactors with the goal to optimize its design and their operating conditions.
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20/07/2012 Validation of thermally assisted hydrolysis and methylation-gas chromatography using a vertical microfurnace pyrolyzer for the compositional analysis of Fatty Acid components in microalgae
Ishida Y, Honda T, Mabuchi S, Sueno O.
Publication Thermally assisted hydrolysis and methylation-gas chromatography (THM-GC) in the presence of trimethylsulfonium hydroxide, using a vertical microfurnace pyrolyzer, was validated for the compositional analysis of fatty acid components in microalgae. The chromatograms of a microalga, Pavlova lutheri , obtained under optimized THM conditions clearly showed a series of fatty acid methyl esters including thermally labile polyunsaturated fatty acid components. On the basis of these peak areas, their chemical compositions were rapidly determined without using any tedious sample pretreatment with a precision of <8% relative standard deviation. Moreover, the compositions thus obtained were in good agreement with those obtained by the conventional technique involving solvent extraction. Finally, the THM-GC technique was applied for the compositional analysis of fatty acid components in a newly found microalga, Coccomyxa gloeobotrydiformis . The obtained data showed a high abundance (24 mol %) of α-linolenic acid components, suggesting its potential usefulness as feed sources and/or functional foods
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