Simultaneous Treatment of Long-chain Monounsaturated Fatty Acid and n-3 Polyunsaturated Fatty Acid Decreases Lipid and Cholesterol Levels in HepG2 Cell.

The n-3 type polyunsaturated fatty acids (n-3PUFAs), including eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), from fish oil exhibit health benefits such as triacylglycerol- and cholesterol-lowering effects. Some pelagic fishes contain long-chain monounsaturated fatty acids (LC-MUFAs) such as eicosenoic acid (C20:1), which exert health-promoting effects. However, no study has evaluated beneficial effects of n-3PUFA and LC-MUFA combination. Here, we investigated effects of simultaneous treatment with n-3PUFA (EPA and DHA) and LC-MUFA (cis-5-C20:1 and cis-7-C20:1) and found that n-3PUFA and LC-MUFA combination significantly decreased lipid accumulation and reduced total cholesterol in HepG2 cells. Cholesterol level was significantly lower in DHA + cis-7-C20:1 group than in DHA + EPA group. These results suggest the importance of LC-MUFA as a functional molecule in fish oil.

Separation of eicosapentaenoic acid and docosahexaenoic acid by three-zone simulated moving bed chromatography.

Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are essential fatty acids for human body, which are widely used in the field of healthy food and medicine. Meanwhile, there are some differences in their physiological functions, such as "scavenger for blood vessel" of EPA and "brain protector" of DHA. In order to make full use of EPA and DHA, it is necessary to prepare their high-purity component. In this paper, EPA and DHA were separated and purified by three-zone simulated moving bed (SMB) chromatography with C18 used as stationary phase and ethanol-water as mobile phase. For the single column experiment, a separation unit of SMB, the effects of the ratio of ethanol to water, pH value and temperature on the separation were investigated. The equilibrium dispersion (ED) model was used to obtain the adsorption parameters of EPA and DHA by inverse method and genetic algorithm, and the accuracy of the adsorption parameters was verified by fitting the overloaded elution curves under different conditions. Based on the acquired nonlinear adsorption isotherms the complete separation region was found according to triangle theory. The effects of sample concentration, flow ratios of adsorption zone and rectification zone, and column distribution mode of SMB on the separation were investigated. Under the optimized SMB conditions, the experimental result was that without regard to the other components, the chromatographic purity and recovery values of EPA and DHA exceeded 99% with the productivity of 4.15 g/L/h, and the solvent consumption of 1.11 L/g.

The formation, stability of DHA/EPA nanoemulsion prepared by emulsion phase inversion method and its application in apple juice.

This study prepared edible docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) nanoemulsion using EPI (emulsion phase inversion) method. The method for preparing DHA and EPA nanoemulsions is safe, convenient, low in energy consumption and can be used for food production. Factors affecting particle size and stability during preparation were investigated. Based on the optimal particle size combination, stability studies including particle size and residual rates of DHA and EPA at different temperature, pH and metal ions. The results showed that the nanoemulsion had good stability at low temperature storage, near neutral pH and in the absence of transition metal ions such as Fe3+, Cu2+, Al3+. The experiment initially studied the effect of nanoemulsion on apple juice beverage on the basic properties of juice itself. It was feasible in practical application of edible nanoemulsion.

Novel Vibrio spp. Strains Producing Omega-3 Fatty Acids Isolated from Coastal Seawater.

Omega-3 long-chain polyunsaturated fatty acids (LC-PUFAs), such as eicosapentaenoic acid (EPA) (20:5n-3) and docosahexaenoic acid (DHA) (22:6n-3), are considered essential for human health. Microorganisms are the primary producers of omega-3 fatty acids in marine ecosystems, representing a sustainable source of these lipids, as an alternative to the fish industry. Some marine bacteria can produce LC-PUFAs de novo via the Polyunsaturated Fatty Acid (Pfa) synthase/ Polyketide Synthase (PKS) pathway, which does not require desaturation and elongation of saturated fatty acids. Cultivation-independent surveys have revealed that the diversity of microorganisms harboring a molecular marker of the pfa gene cluster (i.e., pfaA-KS domain) is high and their potential distribution in marine systems is widespread, from surface seawater to sediments. However, the isolation of PUFA producers from marine waters has been typically restricted to deep or cold environments. Here, we report a phenotypic and genotypic screening for the identification of omega-3 fatty acid producers in free-living bacterial strains isolated from 5, 500, and 1000 m deep coastal seawater from the Bay of Biscay (Spain). We further measured EPA production in pelagic Vibrio sp. strains collected at the three different depths. Vibrio sp. EPA-producers and non-producers were simultaneously isolated from the same water samples and shared a high percentage of identity in their 16S rRNA genes, supporting the view that the pfa gene cluster can be horizontally transferred. Within a cluster of EPA-producers, we found intraspecific variation in the levels of EPA synthesis for isolates harboring different genetic variants of the pfaA-KS domain. The maximum production of EPA was found in a Vibrio sp. strain isolated from a 1000 m depth (average 4.29% ± 1.07 of total fatty acids at 10 °C, without any optimization of culturing conditions).

A biphasic system based on guanidinium ionic liquid: Preparative separation of eicosapentaenoic acid ethyl ester and docosahexaenoic acid ethyl ester by countercurrent chromatography.

Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are high nutritional components. Evidence for unique effects of them is increasing. Further understanding of their independent biological functions urgently needs more efficient separation techniques. Nowadays, most of the commercially available fish oil products are the mixture of eicosapentaenoic acid ethyl ester (EPAEE) and docosahexaenoic acid ethyl ester (DHAEE). It will be convenient to directly separate esterified EPA and DHA without saponification pretreatment. However, it is of great challenge to separate EPAEE and DHAEE because of their extremely fat-soluble nature and the equivalent chain length rule. In this research, the suitability of green guanidinium ionic liquid (IL) in countercurrent chromatography (CCC) solvent system for the separation of them was evaluated for the first time. Compared with imidazolium IL and phosphonium IL, guanidinium IL based non-aqueous biphasic system showed more outstanding separation performance. The separation mechanism was elucidated in depth through quantum mechanical calculations. It was found that guanidinium IL acted a crucial role in the CCC separation, which resulted in difference of partition behavior of EPAEE and DHAEE via different hydrogen-bonding affinity. EPAEE and DHAEE were successfully separated by solvent system (n-heptane/methanol/propylguanidinium chloride ([C3Gun]Cl, 1:1:5%, v/v/m)) with high purity (>95%) in one step, which was not achieved beforehand. Moreover, an easy recycling procedure of IL had also been devised, which significantly reduced waste generated. It opens up a new way for reasonable design water-free two-phase solvent system for efficient separation of very non-polar lipid compounds.

Preparation of eicosapentaenoic acid ethyl ester from fish oil ethyl esters by continuous batch chromatography.

Fish oils are rich in eicosapentaenoic acid, which has the wide-ranging biological activities. The rapid and efficient separation of eicosapentaenoic acid ethyl ester from fish oils ethyl ester is still regarded as a challenge. In this study, we described an effective and flexible chromatography for eicosapentaenoic acid ethyl ester preparation, named continuous batch chromatography, which combined the batch chromatography with the continuous chromatographic operation mode. After continuous batch chromatography experiment, the recovery of eicosapentaenoic acid ethyl ester was 82.01%, the average relative purity and the relative highest purity of eicosapentaenoic acid ethyl ester were 97.82 and 98.98%. The productivity of continuous batch chromatography was 5.48 times higher than that of batch chromatography, while the solvent consumption of eicosapentaenoic acid ethyl ester was 78% of the batch chromatography. This study provided a reference for the separation of the targeted chemical component from multi-component mixtures.

Selective Extraction of ω-3 Fatty Acids from Nannochloropsis sp. Using Supercritical CO2 Extraction.

In this article, microalgae Nannochloropsis sp. was used for fatty acid (FA) extraction, using a supercritical fluid-carbon dioxide (SF-CO2) extraction method. This study investigated the influence of different pre-treatment conditions by varying the grinding speed (200-600 rpm), pre-treatment time (2.5-10 min), and mixing ratio of diatomaceous earth (DE) and Nannochloropsis sp. biomass (0.5-2.0 DE/biomass) on FAs extraction. In addition, the effect of different operating conditions, such as pressure (100-550 bar), temperature (50-75 °C), and CO2 flow rate (7.24 and 14.48 g/min) on eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) recovery, was analyzed. Experimental data evidenced that, keeping constant the extraction conditions, the pre-treatment step enhanced the FAs extraction yield up to 3.4 fold, thereby the maximum extracted amount of FAs (61.19 mg/g) was attained with the pre-treatment with a ratio of DE/biomass of 1 at 600 rpm for 5 min. Moreover, by increasing both SF-CO2 pressure and temperature, the selectivity towards EPA was enhanced, while intermediate pressure and lower pressure promoted DHA recovery. The highest amount of extracted EPA, i.e., 5.69 mg/g, corresponding to 15.59%, was obtained at 75 °C and 550 bar with a CO2 flow rate of 14.48 g/min, while the maximum amount of extracted DHA, i.e., ~0.12 mg/g, equal to 79.63%, was registered at 50 °C and 400 bar with a CO2 flow rate of 14.48 g/min. Moreover, the increased CO2 flow rate from 7.24 to 14.48 g/min enhanced both EPA and DHA recovery.

Separation of high-purity eicosapentaenoic acid and docosahexaenoic acid from fish oil by pH-zone-refining countercurrent chromatography.

The evidence for unique effects of eicosapentaenoic acid and docosahexaenoic acid is growing. Further understanding and exploration of their independent effects in the nutraceutical and pharmaceutical industry is calling for the more efficient separation techniques to overcome the equivalent chain length rule of fatty acids. In this study, free eicosapentaenoic and docosahexaenoic acid were successfully separated by pH-zone-refining countercurrent chromatography for the first time. The different solvent systems and the influence of retainer and eluter concentration on the separation efficiency were investigated. A two-phase solvent system composed of n-heptane/methanol/water (100:55:45, v/v) was selected with 50 mM of trifluoroacetic acid as retainer in the organic phase and 40 mM of ammonium hydroxide as an eluter in the aqueous phase for the separation of 500 mg of free fatty acids from a refined fish oil sample. 79.6 mg of eicosapentaenoic acid and 328.3 mg docosahexaenoic acid were obtained with the purities of 95.5 and 96.9% respectively determined by gas chromatography with mass spectrometry after methyl esterification. The scale-up separation of 1 g of samples from both refined and crude fish oil after urea complexation were also achieved successfully with a markedly increased concentration 150 mM of retainer, producing satisfactory yields and purities of targets.

Eicosapentaenoic Acid Extraction from Nannochloropsis gaditana using Carbon Dioxide at Supercritical Conditions.

This research shows that carbon dioxide supercritical fluid (CO2-SF) is an emerging technology for the extraction of high interest compounds for applications in the manufacturing of pharmaceuticals, nutraceuticals, and cosmetics from microalgae. The purpose of this study is to recover fatty acids (FAs) and, more precisely, eicosapentaenoic acid (EPA) from Nannochloropsis gaditana biomass by CO2-SF extraction. In the paper, the effect of mechanical pre-treatment was evaluated with the aim of increasing FAs recovery. Extraction was performed at a pressure range of 250⁻550 bars and a CO2 flow rate of 7.24 and 14.48 g/min, while temperature was fixed at 50 or 65 °C. The effect of these parameters on the extraction yield was assessed at each extraction cycle, 20 min each, for a total extraction time of 100 min. Furthermore, the effect of biomass loading on EPA recovery was evaluated. The highest EPA extraction yield, i.e., 11.50 mg/g, corresponding to 27.4% EPA recovery, was obtained at 65 °C and 250 bars with a CO2 flow rate of 7.24 g/min and 1.0 g biomass loading. The increased CO2 flow rate from 7.24 to 14.48 g/min enhanced the cumulative EPA recovery at 250 bars. The purity of EPA could be improved by biomass loading of 2.01 g, even if recovery was reduced.

Screening of Suitable Ionic Liquids as Green Solvents for Extraction of Eicosapentaenoic Acid (EPA) from Microalgae Biomass Using COSMO-RS Model.

Omega-3 poly unsaturated fatty acids (PUFA) particularly eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA), have many health benefits including reducing the risk of cancer and cardiovascular disease. Recently, the use of ionic liquids (ILs) in lipid extraction from microalgae provides the potential to overcome common drawbacks, offers several other benefits. To date, very limited researches are available to focus on extracting microalgae lipid and PUFA in particular by using ILs. The objective of current work is to screen the potential ILs that can be applied in EPA extraction. In this study, fast ILs screening was performed with the help of a conductor like screening model for real solvents (COSMO-RS) and the ILs with higher capacity values for use in extraction of EPA were compared. According to the results, the highest capacity for EPA extraction among 352 screened cation/anion combinations belongs to [TMAm][SO4]. It is expected to achieve a higher yield of EPA once applying this combination as the solvent in the process of extraction. ILs with small anions were observed to have higher capacities, as well possessing higher charge density compared to larger ones, and therefore, they are more preferable for extraction purposes. Moreover, shorter alkyl chain cations are preferred when using imidazolium-based IL, which agrees with experimental data.

Production of Lipids and Proteome Variation in a Chilean Thraustochytrium striatum Strain Cultured under Different Growth Conditions.

Total lipids and docosahexaenoic acid (DHA) production by a Chilean isolated thraustochytrid were evaluated under different growth conditions in shake flasks. The analyzed strain was identified as Thraustochytrium striatum according to an 18S rRNA gene sequence analysis. The strain (T. striatum AL16) showed negligible growth in media prepared with artificial seawater at concentrations lower than 50% v/v and pH lower than 5. Maltose and starch were better carbon sources for growth than glucose. DHA content of the biomass grown with maltose (60 g L-1) was doubled by increasing the agitation rate from 150 to 250 rpm. The DHA (0.8-6%) and eicosapentaenoic acid (0.2-21%) content in the total lipids varied depending on culture conditions and culture age. Lipid and DHA concentration increased (up to 5 g L-1 and 66 mg L-1, respectively) by regularly feeding the culture with a concentrated starch solution. Carotenoid accumulation was detected in cells grown with maltose or starch. Contrasting conditions of starch and glucose cultures were selected for comparative proteomics. Total protein extracts were separated by two-dimensional gel electrophoresis; 25 spots were identified using ESI-MS/MS. A protein database (143,006 entries) for proteomic interrogation was generated using de novo assembling of Thraustochytrium sp. LLF1b - MMETSP0199_2 transcriptome; 18 proteins differentially expressed were identified. Three ATP synthases were differentially accumulated in cultures with glucose, whereas malate dehydrogenase was more abundant in cells cultured with starch.

[Separation of eicosapentaenoic acid ethyl ester and docosahexaenoic acid ethyl ester by simulated moving bed chromatography].

Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are important ω -3 polyunsaturated fatty acids. Their physiological effects on humans are not exactly the same; therefore, the production of products with high-purity EPA or DHA monomers is significant. In this work, EPA ethyl ester (EPA-EE) and DHA ethyl ester (DHA-EE) were first separated using HPLC with poly(styrene-co-divinylbenzene) (PS/DVB) as the stationary phase. The effects of the mobile phase, PS/DVB particle diameter, and column temperature were systematically evaluated. The results showed that methanol is a suitable mobile phase, having a resolution of 2.75. By comparing resolutions, a PS/DVB particle diameter of 10 µ m was chosen; however, when the pressure drop of PS/DVB is considered, PS/DVB with a particle diameter of 20 µ m is more favorable for large-scale preparations. A column temperature of 40℃ was found to be the most feasible for maintaining efficient separation. Second, eight semi-preparative columns (150 mm×10 mm) of PS/DVB polymer were prepared for the simulated moving bed (SMB) chromatography; the homogeneity of these columns was perfect, with a relative total column porosity error of less than 1%. Finally, an EPA-EE and DHA-EE mixture was separated using the SMB chromatography, and the contents of the extract and the raffinate were determined using GC-FID. The effects of the flow rate of Zone Ⅱ and Zone Ⅲ, the flow rate of the feed, and the feed concentration were investigated. Under optimal conditions, EPA-EE and DHA-EE with favorable purities of 91.6% and 93.6%, respectively, were achievable. The recovery of the EPA-EE was 97.0% and the recovery of the DHA-EE was 91.6%. The productivity and solvent requirements were 5.97 g/(L\5h) and 1.52 L/g, respectively. Therefore, SMB chromatography is an attractive technology for the production of high-value products.

Effect of spray-drying with organic solvents on the encapsulation, release and stability of fish oil.

Fish-oil (FO) was encapsulated with hydroxypropylcelullose (HPC) by conventional spray-drying with water (FO-water) and solvent spray-drying with ethanol (FO-EtOH), methanol (FO-MeOH) and acetone (FO-Acet) in order to study the effect of the solvent on the encapsulation efficiency (EE), microparticle properties and stability of FO during storage at 40 °C. Results showed that FO-Acet presented the highest EE of FO (92.0%), followed by FO-EtOH (80.4%), FO-MeOH (75.0%) and FO-water (71.1%). A decrease of the dielectric constant increased the EE of FO, promoting triglyceride-polymer interactions instead of oil-in-water emulsion retention. FO release profile in aqueous model was similar for all FO-microparticles, releasing only the surface FO, according to Higuchi model. Oxidative stability of FO significantly improved by spray-drying with MeOH, both in surface and encapsulated oil fractions. In conclusion, encapsulation of FO by solvent spray-drying can be proposed as an alternative technology for encapsulation of hydrophobic molecules.

Optimization of Subcritical Water Extraction (SWE) of Lipid and Eicosapentaenoic Acid (EPA) from Nannochloropsis gaditana.

Microalgae are a promising source of omega-3. The purpose of this study was to extract lipid with a relatively high content of eicosapentaenoic acid (EPA) from Nannochloropsis gaditana using subcritical water extraction (SWE). The effects of different temperatures (156.1-273.9°C), extraction times (6.6-23.4 minutes), and biomass loadings (33-117 g algae/L) on the extraction yield were studied. From the optimization study using central composite design (CCD), quadratic models generated for lipid yield and EPA composition were considered to be significant models (p < 0.05). The predictive equations were also formed for lipid yield and EPA composition. The predicted optimum lipid yield and EPA composition at 236.54°C, 13.95 minutes, and 60.50 g algae/L were 18.278 wt% of total biomass and 14.036 wt% of total fatty acid methyl ester (FAME), respectively.

Oil and eicosapentaenoic acid production by the diatom Phaeodactylum tricornutum cultivated outdoors in Green Wall Panel (GWP®) reactors.

Phaeodactylum tricornutum is a widely studied diatom and has been proposed as a source of oil and polyunsaturated fatty acids (PUFA), particularly eicosapentaenoic acid (EPA). Recent studies indicate that lipid accumulation occurs under nutritional stress. Aim of this research was to determine how changes in nitrogen availability affect productivity, oil yield, and fatty acid (FA) composition of P. tricornutum UTEX 640. After preliminary laboratory trials, outdoor experiments were carried out in 40-L GWP® reactors under different nitrogen regimes in batch. Nitrogen replete cultures achieved the highest productivity of biomass (about 18 g m-2 d-1 ) and EPA (about 0.35 g m-2 d-1 ), whereas nitrogen-starved cultures achieved the highest FA productivity (about 2.6 g m-2 d-1 ). The annual potential yield of P. tricornutum grown outdoors in GWP® reactors is 730 kg of EPA per hectare under nutrient-replete conditions and 5,800 kg of FA per hectare under nitrogen starvation. Biotechnol. Bioeng. 2017;114: 2204-2210. © 2017 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc.

Long chain fatty acids and related pro-inflammatory, specialized pro-resolving lipid mediators and their intermediates in preterm human milk during the first month of lactation.

This study aimed to measure longitudinal quantities of the long chain fatty acids, their biologically active terminal metabolites and related intermediates (also called oxylipins) in preterm human milk expressed during the first month of lactation. In a prospective cohort, breast milk was collected throughout the first month of lactation in 30 women who delivered preterm infants. Eighteen bioactive lipids and their intermediates were quantified via solid phase extraction and LC-MS/MS. Analysis by GC-FID quantified the fatty acid precursors. Arachidonic acid (ARA) and docosahexaenoic acid (DHA) milk concentrations significantly declined throughout the first month. Oxylipin concentrations did not change during lactation. Positive associations existed between ARA and thromboxane B2, eicosapentaenoic acid and 18-hydroxyeicosapentaenoic acid, and between DHA and PDX and 14- and 17-hydroxydocosahexaenoic acids. DHA concentrations were 1.5 times higher and 14-HDHA was 1.7 times higher in milk from women taking DHA supplements. This investigation showed conditionally essential fatty acids, ARA and DHA, decreased in preterm milk, suggesting a need to supplement their intake for the breast milk-fed preterm infant. Positive associations between parent fatty acids, bioactive lipids and intermediates, as well as sensitivity of milk to maternal fatty acid intake, support consideration of a comprehensive approach to providing fatty acids for preterm infants through both maternal and infant supplementation.

The Association of Fatty Acid Levels and Gleason Grade among Men Undergoing Radical Prostatectomy.

BACKGROUND: Epidemiological data suggest that omega-6 (ω-6) fatty acids (FAs) may be associated with cancer incidence and/or cancer mortality, whereas ω-3 FAs are potentially protective. We examined the association of the ratio of ω-6 to ω-3 FA (ω-6:ω-3) and individual FA components with pathological results among men with prostate cancer (PCa) undergoing radical prostatectomy. METHODS: Sixty-nine men were included in the study. Components of ω-6 (linoleic acid (LA), arachidonic acid (AA), and dihomo-γ-linolenic acid (DGLA)) and ω-3 (docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA)) were analyzed by liquid chromatography/mass selective detector separation. Logistic regression analysis was performed to determine association of FA with pathological high grade (Gleason ≥4+3) disease. RESULTS: The were 35 men with low grade disease (Gleason ≤3+4) and 34 men with high grade disease. Men with low grade disease were significantly younger (58y vs 61y, p = 0.012) and had lower D'Amico clinical classification (p = 0.001) compared to men with high grade disease. There was no significant association of ω-6:ω-3 with high grade disease (OR 0.93, p = 0.78), however overall ω-6, ω-3, and individual components of ω-6 and ω-3 FAs except EPA were significantly associated with high grade disease (ω-6: OR 3.37, 95% CI: 1.27,8.98; LA: OR 3.33, 95% CI:1.24,8.94; AA: OR 2.93, 95% CI:1.24,6.94; DGLA: OR 3.21, 95% CI:1.28,8.04; ω-3: OR 3.47, 95% CI:1.22,9.83; DHA: OR 3.13, 95% CI:1.26,7.74). ω-6 and ω-3 FA components were highly correlated (Spearman ρ = 0.77). CONCLUSION: Higher levels of individual components of ω-6 and ω-3FAs may be associated with higher-grade PCa. IMPACT: Studies into the causative factors/pathways regarding FAs and prostate carcinogenesis may prove a potential association with PCa aggressiveness.

Drosophila Fed ARA and EPA Yields Eicosanoids, 15S-Hydroxy-5Z,8Z, 11Z, 13E-Eicosatetraenoic Acid, and 15S-Hydroxy-5Z,8Z,11Z,13E,17Z-Eicosapentaenoic Acid.

Drosophila melanogaster has been a widely used as a model system for its powerful genetic tools. However, it remains to be illustrated if Drosophila can be used to examine the biochemical and physiological metabolism of eicosanoids. Thus, the analysis on the metabolism of C20 polyunsaturated fatty acids (PUFA) in Drosophila was implemented with high performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS). Fatty acid (FA) analysis of the whole body, head, and thorax-abdomen in Drosophila showed C20 PUFA could only be found in Drosophila fed diets supplemented with eicosapentaenoic acid (EPA) and arachidonic acid (ARA), but not in Drosophila fed base diets. The C20 PUFA were found in abundance in the head. Drosophila fed ARA- and EPA-supplemented diets yielded 15S-hydroxy-5Z,8Z,11Z,13E-eicosatetraenoic acid [15(S)-HETE] and 15S-hydroxy-5Z,8Z,11Z,13E,17Z-eicosapentaenoic acid [15(S)-HEPE], respectively, while other sampled eicosanoids could not be detected. Similar results were obtained by incubating fly tissue supplemented with ARA or EPA. Furthermore, a genome sequence scan indicated that no gene encoding the key enzymes synthesizing eicosanoids were found in Drosophila. These findings demonstrate that Drosophila may possess a special lipid metabolic system, which is different from mammals.

Heterotrophic Production of Omega-3 Long-Chain Polyunsaturated Fatty Acids by Trophically Converted Marine Diatom Phaeodactylum tricornutum.

We have created via metabolic engineering a heterotrophic strain of Phaeodactylum tricornutum that accumulates enhanced levels of the high value omega-3 long chain polyunsaturated fatty acid (LC-PUFAs) docosahexaenoic acid (DHA). This was achieved by generation of transgenic strains in which the Δ5-elongase from Ostreococcus tauri was co-expressed with a glucose transporter from the moss Physcomitrella patens. This double transformant has the capacity to grow in the dark in liquid medium supplemented with glucose and accumulate substantial levels of omega-3 LC-PUFAs. The effects of glucose concentrations on growth and LC-PUFA production of wild type and transformed strains cultivated in the light and dark were studied. The highest omega-3 LC-PUFAs accumulation was observed in cultures grown under mixotrophic conditions in the presence of 1% glucose (up to 32.2% of total fatty acids, TFA). Both DHA and EPA are detected at high levels in the neutral lipids of transgenic cells grown under phototrophic conditions, averaging 36.5% and 23.6% of TFA, respectively. This study demonstrates the potential for P. tricornutum to be developed as a viable commercial strain for both EPA and DHA production under mixo- and heterotrophic conditions.

CYTOTOXIC EFFECTS OF THE RED SEA SOFT CORAL SARCOPHYTON TROCHELIOPHORUM.

The present study describes the in vitro cytotoxic effects of soft coral (Sarcophyton tiocheliophorum). Soft corals of genus Sarcophyton were reported to contain compounds that are active against brine shrimp and promote paclitaxel cytotoxicity in the human colon cancer Caco-2 cell line. The n-hexane extract of the soft coral Sarcophyton tiocheliophorum induced significant dose-dependent toxicity (LC50 96.7 ppm) compared with ethyl acetate (LC50. 120 ppm). We reported the most active cytotoxic level to be correspondence to LC50 values of 20.2, 59.2 ppm and 18.9 and 26 ppm. Accordingly, bio-assay guided fractionation was conducted to identi- fy the bioactive compounds. Arachidonic acid, eicosapentaenoic acid and docosahexaenoic acid were characterized based on GC-MS analyses. Our results demonstrate the value of marine products as a natural source of medicinally interesting cytotoxic compounds.