Academic literature on the topic 'Omega 3 (n-3) long chain (LC) polyunsaturated fatty acids (PUFA)'

Aims: Adequate intake of long-chain (LC) omega-3 polyunsaturated fatty acids ( n-3 PUFAs) is considered important for cardiovascular health. However, the effects of LC n-3 PUFAs on the risk of heart failure (HF) remain unclear. This systematic review and meta-analysis aimed to determine the role of LC n-3 PUFAs in the incidence of HF. Materials and Methods: Electronic databases were searched for studies up to 31 July 2021. Studies were included for the meta-analysis if they reported the adjusted associations between different dietary intakes or circulating concentrations of LC n-3 PUFAs and the risk of HF. A random-effect model was used to calculate the pooled estimated hazard ratios (HRs) and 95% confidence intervals (CIs) for higher LC n-3 PUFA concentrations. Results: Thirteen studies were included in the meta-analysis. Eight studies comprising 316,698 individuals (11,244 incident HF cases), with a median follow-up of 10.7 years, showed that a higher dietary intake of LC n-3 PUFAs was associated with a lower risk of HF (highest versus lowest quintile: HR = 0.84, 95% CI = 0.75–0.94). Six studies, comprising 17,163 participants (2520 HF cases) with a median follow-up of 9.7 years, showed that higher circulating LC n-3 PUFA concentrations were associated with a lower risk of HF (highest versus lowest quintile: HR = 0.59, 95% CI = 0.39–0.91). Higher circulating docosahexaenoic acid concentrations were associated with a decreased risk of HF (top versus bottom quintile: HR = 0.44, 95% CI = 0.26–0.77). The associations between eicosapentaenoic acid (HR = 0.58, 95% CI = 0.26–1.25), docosahexaenoic acid (HR = 0.66, 95% CI = 0.24–1.82), and the risk of HF were not significant. Conclusion: High LC n-3 PUFA concentrations measured by dietary intake or circulating biomarkers are associated with a lower risk of developing HF.

The health benefits of diets containing rich sources of long-chain omega-3 polyunsaturated fatty acids (n-3 LC-PUFA) are well documented and include reductions in the risk of several diseases typical of Western societies. The dietary intake of n-3 LC-PUFA has also been linked to fertility, and there is abundant evidence that a range of ejaculate traits linked to fertility in humans, livestock and other animals depend on an adequate intake of n-3 LC-PUFA from dietary sources. However, relatively few studies have explored how n-3 LC-PUFA influence reproductive fitness, particularly in the context of sexual selection. Here, we show that experimental reduction in the level of n-3 LC-PUFA in the diet of guppies ( Poecilia reticulata ) depresses a male's share of paternity when sperm compete for fertilization, confirming that the currently observed trend for reduced n-3 LC-PUFA in western diets has important implications for individual reproductive fitness.

The health benefits of a diet rich in omega-3 long chain polyunsaturated fatty acids (n-3 LC-PUFA) no longer need to be proven. However, while health authorities attempt to increase the consumption of the n-3 LC-PUFAs eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), data from the latest intake surveys demonstrate that EPA and DHA consumption is still too low. A push towards greater sustainability, and a rise in vegetarianism are pushing manufacturers to move from traditional fish oils towards alternative sources. Microalgae oils provide a source of n-3 LC-PUFA with a lower environmental impact and are produced using processes that limit damage to the oils. This review aims to report on oleaginous microalgae strains available for n-3 LC-PUFA production, the processes used for their growth and the extraction and refining processes for their oils. It also addresses the challenges inherent in these products and their fabrication, and some of the novel characteristics of microalgal oils, including their very high n-3 LC-PUFA content and the chemical structure of their triglycerides, that lead to exciting opportunities in their use as functional food ingredients.

Omega-3 long-chain polyunsaturated fatty acids (LC-PUFA) play a central role in neuronal growth and in the development of the human brain, and a deficiency of these substances has been reported in children with attention deficit hyperactive disorder (ADHD). In this regard, supplementation with omega-3 polyunsaturated fatty acids is used as adjuvant therapy in ADHD. Seafood, particularly fish, and some types of nuts are the main dietary sources of such fatty acids in the Spanish diet. In order to assess the effect of the intake of common foods containing high amounts of omega-3 polyunsaturated fatty acids, a food frequency questionnaire was administered to parents of children with ADHD (N = 48) and to parents of normally developing children (control group) (N = 87), and the intake of dietary omega-3 LC-PUFA, such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), was estimated. Children with ADHD consumed fatty fish, lean fish, mollusks, crustaceans, and chicken eggs significantly less often (p < 0.05) than children in the control group. The estimated daily omega-3 LC-PUFA intake (EPA + DHA) was significantly below that recommended by the public health agencies in both groups, and was significantly lower in children with ADHD (p < 0.05, Cohen’s d = 0.45) compared to normally developing children. Dietary intervention to increase the consumption of fish and seafood is strongly advised and it is especially warranted in children with ADHD, since it could contribute to improve the symptoms of ADHD.

Omega-3 long chain polyunsaturated fatty acids (ω3 LC-PUFAs) such as eicosapentaenoic acid (EPA; 20:5ω3) and docosahexaenoic acid (DHA; 22:6ω3) are important fatty acids for human health. These ω3 LC-PUFAs are produced from their ω3 precursors by a set of desaturases and elongases involved in the biosynthesis pathway and are also converted from ω6 LC-PUFA by omega-3 desaturases (ω3Ds). Here, we have investigated eight ω3-desaturases obtained from a cyanobacterium, plants, fungi and a lower animal species for their activities and compared their specificities for various C18, C20 and C22 ω6 PUFA substrates by transiently expressing them in Nicotiana benthamiana leaves. Our results showed hitherto unreported activity of many of the ω3Ds on ω6 LC-PUFA substrates leading to their conversion to ω3 LC-PUFAs. This discovery could be important in the engineering of EPA and DHA in heterologous hosts.

AbstractConsiderable evidence exists for marked beneficial effects of omega-3 long-chain polyunsaturated fatty acids (LC-PUFA) during pregnancy. The omega-3 LC-PUFA docosahexaenoic acid (DHA) is incorporated in large amounts in fetal brain and other tissues during the second half of pregnancy, and several studies have provided evidence for a link between early DHA status of the mother and visual and cognitive development of her child after birth. Moreover, the supplementation of omega-3 LC-PUFA during pregnancy increases slightly infant size at birth, and significantly reduces early preterm birth before 34 weeks of gestation by 31%. In our studies using stable isotope methodology in vivo, we demonstrated active and preferential materno-fetal transfer of DHA across the human placenta and found the expression of human placental fatty acid binding and transport proteins. From the correlation of DHA values with placental fatty acid transport protein 4 (FATP 4), we conclude that this protein is of key importance in mediating DHA transport across the human placenta. Given the great importance of placental DHA transport for infant outcome, further studies are needed to fully appreciate the effects and optimal strategies of omega-3 fatty acid interventions in pregnancy, dose response relationships, and the potential differences between subgroups of subjects such as women with gestational diabetes or other gestational pathology. Such studies should contribute to optimize substrate intake during pregnancy and lactation that may improve pregnancy outcome as well as fetal growth and development.

The aim of this research was to evaluate the nutritional enhancement of omega-3 long-chain polyunsaturated fatty acid (n-3 LC-PUFA) composition of edible lamb Longissimus thoracis et lumborum muscle, heart, kidney, and liver in response to dietary supplementation of lot-fed lambs with or without omega-3 oil fortified pellets. The hypothesis tested was that fortifying feedlot pellets with omega-3 oil will enhance the human health beneficial n-3 LC-PUFA composition of edible lamb muscle tissue and organs. Seventy-five Tattykeel Australian White lambs exclusive to the MARGRA brand, with an average body weight of 30 kg at six months of age, were randomly assigned to the following three dietary treatments of 25 lambs each, and lot-fed as a cohort for 47 days in a completely randomized experimental design: (1) Control grain pellets without oil plus hay; (2) Omega-3 oil fortified grain pellets plus hay; and (3) Commercial whole grain pellets plus hay. All lambs had ad libitum access to the basal hay diet and water. Post-slaughter fatty acid composition of the Longissimus thoracis et lumborum muscle, liver, kidney, and heart were determined using thee gas chromatography–mass spectrophotometry technique. Results indicated significant variations (p < 0.05) in fatty acid profiles between tissues and organs. Omega-3 oil fortified pellets significantly (p < 0.05) increased ≥C20 n-3 LC-PUFA (C20:5n-3 eicosapentaenoate, EPA + C22:5n3 docosapentaenoate, DPA + C22:6n3 docosahexanoate DHA); C18:3n-3 alpha-linolenate, ALA; C18:2 conjugated linoleic acid, CLA; total monounsaturated fatty acids, MUFA; polyunsaturated fatty acids, PUFA contents; and reduced the ratio of omega-6 to omega-3 fatty acids in all lamb organs and tissues without impacting shelf-life. The findings demonstrate that the inclusion of omega-3 oil in feedlot diets of lambs enhances the human health beneficial omega-3 long-chain polyunsaturated fatty acid profiles of edible muscle tissue and organs without compromising meat quality.

Abstract Context The effects of dietary intake of different fatty acids and pharmacological use of fatty acids, specifically long-chain n-3 polyunsaturated fatty acids (LC n-3 PUFAs), on cardiovascular health and atherosclerotic cardiovascular disease (ASCVD) prevention have been examined in a large number of observational studies and clinical trials. This review summarizes recent data and discusses potential mechanisms. Evidence acquisition The review is based on the authors’ knowledge of the field supplemented by a PubMed search using the terms seafood, fish oil, saturated fatty acids, omega-3 fatty acids, eicosapentaenoic acid, docosahexaenoic acid, polyunsaturated fatty acids, monounsaturated fatty acids, and ASCVD. Evidence synthesis We mainly discuss the recent clinical trials that examine the effects of different types of dietary fatty acids and pharmacological use of n-3 PUFA products on ASCVD prevention and the potential mechanisms. Conclusions While replacement of dietary saturated fat with unsaturated fat, polyunsaturated fat in particular, or intake of LC n-3 PUFA–rich seafood has generally shown benefit for ASCVD prevention and is recommended for cardiovascular benefits, data on effects of n-3 PUFA products on ASCVD health are inconsistent. However, recent clinical trials support benefits of prescription EPA in ASCVD prevention. n-3 PUFAs may contribute to ASCVD prevention through multiple mechanisms, including lowering plasma triglyceride levels, anti-inflammatory effects, antithrombotic effects, and effects on endothelial function.

Omega-3 polyunsaturated fatty acids (n-3 PUFA) are termed essential fatty acids because they cannot be synthesized de novo by humans due to the lack of delta-12 and delta-15 desaturase enzymes and must therefore be acquired from the diet. n-3 PUFA include α-linolenic acid (ALA, 18:3n-3), eicosapentaenoic (EPA, 20:5n-3), docosahexaenoic (DHA, 22:6n-3), and the less recognized docosapentaenoic acid (DPA, 22:5n-3). The three long-chain (≥C20) n-3 PUFA (n-3 LC-PUFA), EPA, DHA, and DPA play an important role in human health by reducing the risk of chronic diseases. Up to the present time, seafood, and in particular, fish oil-derived products, have been the richest sources of n-3 LC-PUFA. The human diet generally contains insufficient amounts of these essential FA due largely to the low consumption of seafood. This issue provides opportunities to enrich the content of n-3 PUFA in other common food groups. Milk and milk products have traditionally been a major component of human diets, but are also among some of the poorest sources of n-3 PUFA. Consideration of the high consumption of milk and its processed products worldwide and the human health benefits has led to a large number of studies targeting the enhancement of n-3 PUFA content in dairy products. The main objective of this review was to evaluate the major strategies that have been employed to enhance n-3 PUFA content in dairy products and to unravel potential knowledge gaps for further research on this topic. Nutritional manipulation to date has been the main approach for altering milk fatty acids (FA) in ruminants. However, the main challenge is ruminal biohydrogenation in which dietary PUFA are hydrogenated into monounsaturated FA and/or ultimately, saturated FA, due to rumen microbial activities. The inclusion of oil seed and vegetable oil in dairy animal diets significantly elevates ALA content, while the addition of rumen-protected marine-derived supplements is the most effective way to increase the concentration of EPA, DHA, and DPA in dairy products. In our view, the mechanisms of n-3 LC-PUFA biosynthesis pathway from ALA and the biohydrogenation of individual n-3 LC-PUFA in ruminants need to be better elucidated. Identified knowledge gaps regarding the activities of candidate genes regulating the concentrations of n-3 PUFA and the responses of ruminants to specific lipid supplementation regimes are also critical to a greater understanding of nutrition-genetics interactions driving lipid metabolism.

Marine sources of long chain omega-3 polyunsaturated fatty acids (LC n-3 PUFA) are in high demand for use in health supplements. Mass cultivated marine microalgae is a promising and sustainable source of LC n-3 PUFA, which relieves pressure on natural fish stocks. The lipid class profile from cultivated photosynthetic algae differ from the marine organisms currently used for the production of LC n-3 PUFA. The objective of this study was to compare in vitro intestinal digestion of oil extracted from the cold-adapted marine diatom Porosira glacialis with commercially available LC n-3 PUFA supplements; cod liver oil, krill oil, ethyl ester concentrate, and oil from the copepod Calanus finmarchicus (Calanus® oil). The changes in the free fatty acids and neutral and polar lipids during the enzymatic hydrolysis were characterized by liquid and gas chromatography. In Calanus® oil and the Ethyl ester concentrate, the free fatty acids increased very little (4.0 and 4.6%, respectively) during digestion. In comparison, free fatty acids in Krill oil and P. glacialis oil increased by 14.7 and 17.0%, respectively. Cod liver oil had the highest increase (28.2%) in free fatty acids during the digestion. Monounsaturated and saturated fatty acids were more easily released than polyunsaturated fatty acids in all five oils.

Tilapia are freshwater fish that have become important in aquaculture and as a stable global source of seafood due to their ability to thrive in different environments. However, tilapia are sometimes considered nutritionally undesirable due to their high n-6 to n-3 fatty acid ratios. A market study was conducted first to determine fatty acid compositions in tilapia fillets in different US markets. Then a research was conducted to enhance nutritional value of tilapia by improving the n-3 and oleic acid contents in fish fillets without compromising fish growth or feed conversion ratios. Feeds were formulated with combinations of high and low n-6, n-3, and oleic acid levels using soybean oil, fish oil, algae oil, and high-oleic sunflower oil. Then 12 diets, including a commercial diet, were assigned to 24 tanks, each with 25 tilapia per tank. A Recirculating Aquaculture System (RAS) was used to grow the fish for 8 weeks. Fatty acid compositions of tilapia fillets were determined and samples were vacuum packed and stored at -10oC and -20oC to test oxidative degradation and fatty acid compositional changes. The market survey data showed that there were significant differences in fatty acid composition, lipid content, and n-6:n-3 fatty acid ratios depending on the country of origin. Samples from USA had ideal n-6:n-3 ratios (1.3 ±0.85) while samples from Southeast Asia had higher n-6:n-3 fatty acids ratio (6.6 ±0.54). Algae oil incorporation significantly increased DHA level while fish oil incorporation significantly increased both EPA and DPA. High-oleic sunflower oil based diets improved oleic acid levels and reduced linoleic acid compared to the soybean oil based diets. Sensory evaluation indicated that lipid source did not significantly impact preference or overall fillet quality, including texture. Interestingly, a survey showed people were interested in value-added tilapia, and would pay up to 30% more for nutritionally enhanced fish compared to the $5.00/lb fresh fillet price currently available in supermarkets. There was no observable oxidation during long term frozen storage. The oxidation study proved that value-addition would not be compromised during the long term storage conditions, even under temperature abuse. It is possible to improve tilapia nutritional quality through diet to provide consumers with value-added products that maintain quality during frozen storage.
Ph. D.

Les acides gras polyinsaturés (AGPIs) à longue chaine oméga-3, essentiels à la santé et au développement chez l'homme, sont les précurseurs de médiateurs lipidiques jouant des rôles importants au maintien de l'homéostasie tissulaire. Ces métabolites oxygénés, et collectivement appelés oxylipines, sont impliqués dans la régulation de nombreux processus physiopathologiques tels que l'inflammation ou le cancer. De nouvelles sources alternatives et durables d'AGPIs n-3 étant requises pour répondre à la demande mondiale croissante, les microalgues, producteurs primaires d'AGPIs n-3, représentent une source naturelle d'intérêt pour la production de ces macromolécules à haute valeur ajoutées. Dans ce contexte, le présent travail visait à évaluer le potentiel de la microalgue pico-eucaryote Ostreococcus comme source alternative d'AGPIs n-3 et de leurs oxylipines dérivées. Cette étude a démontré que les microalgues du genre Ostreococcus contiennent de forts taux d'AGPIs, les oméga-3 étant majoritaires. En particulier, les cellules d'Ostreococcus ont montré de forts taux de DHA qui restaient relativement stables au cours de la croissance des cultures ainsi qu’avec des conditions de température, intensités lumineuses et salinité très variables. La biomasse d'Ostreococcus a montré un ensemble d'oxylipines dérivées d’AGPIs n-3 et n-6. Deux mono-hydroxy acides dérivés du DHA, les 17-HDoHE et 14-HDoHE, se sont avérés majoritaires dans les cellules d’Ostreococcus, et ce, indépendamment de la souche et des conditions de culture. En outre, des approches de génie génétique ont été réalisées avec succès pour augmenter les concentrations cellulaires d’oxylipines
Omega-3 long-chain polyunsaturated fatty acids (LC-PUFAs) that are essential to human health and development are precursors of lipid mediators that play important roles for tissue homeostasis. These metabolites derived from lipid oxidation processes and collectively named oxylipins, are involved in the regulation of various physiopathological processes including inflammation and cancer. As the global consumer needs for n-3 LC-PUFAs is increasing the fishes market will likely not be sufficient and new alternative sources of n-3 LC-PUFAs are needed. Microalgae are an interesting natural source as primary producers of n-3 LC-PUFAs and therefore, a possible source of these high-values added macromolecules. In this context, the present work aimed to evaluate the potential of the green picoeukaryote Ostreococcus as a source of n-3 PUFAs and derived oxylipins. This study clearly revealed microalgae of the genus Ostreococcus contain high levels of PUFAs, the omega-3 being predominant over the omega-6. Particularly, Ostreococcus cells showed high docosahexaenoic acid (DHA, C22:6 n-3) levels that remained fairly stable throughout the growth cycle and under various temperature, light intensity and salinity stress conditions. The biomass of Ostreococcus showed an array of oxylipins derived from PUFAs from the n-3 and n-6 series. In particular, two monohydroxy acids derived from DHA, 17-HDoHE and 14-HDoHE, were found to be predominant in Ostreococcus cells regardless the strain or the culture conditions tested. Furthermore, genetic engineering approach was successfully used to increase oxylipins content

La résistance des cellules tumorales à la chimiothérapie constitue une cause majeure d’échec des traitements anticancéreux. Des études précliniques montrent que les acides gras polyinsaturés oméga-3 à longues chaînes (AGPIn-3LC), apportés par l’alimentation, améliorent l’efficacité des chimiothérapies sans majorer les effets secondaires. Cette thèse a eu pour but d’identifier les mécanismes moléculaires impliqués dans l’augmentation de la sensibilité des cellules tumorales mammaires au docétaxel. Nous avons montré que le docétaxel induit un mécanisme de résistance via l’activation des voies de signalisation PKC/ERK et Akt impliquées dans la prolifération et la survie cellulaires. La modification de l’environnement lipidique membranaire par la supplémentation en AGPIn-3LC inhibe ces voies de signalisation et augmente l’efficacité du docétaxel dans des lignées tumorales mammaires et dans un modèle préclinique de tumeurs mammaires autochtones chez le rongeur. De plus, dans ce modèle in vivo, nous avons identifié une autre cible moléculaire régulée par les AGPIn-3LC : l’épiréguline, membre de la famille EGF. Les AGPIn-3LC bloquent l’induction de l’épiréguline par le VEGF dans les cellules endothéliales et induisent un remodelage de la vascularisation tumorale. Outre un effet direct des AGPIn-3LC sur les cellules tumorales, les AGPIn- 3LC agissent sur le microenvironnement tumoral. Ces travaux de thèse apportent des arguments supplémentaires pour l’utilisation des AGPIn-3LC comme molécules adjuvantes pour lutter contre la résistance des tumeurs mammaires aux agents anticancéreux
Chemotherapy-resistant tumor cells are a major cause of cancer treatment failure. Preclinical studies show that polyunsaturated omega-3 long chain fatty acids (AGPIn-3LC), provided by food, improve the efficacy of chemotherapy without increasing side effects. AGPIn-3LCs are incorporated in cancer and stromal cells. This thesis aimed to identify molecular mechanisms involved in the increased sensitivity of mammary tumor cells to docetaxel. We have shown that docetaxel induces a resistance mechanism via activation of PKC/ERK and Akt pathways involved in cell proliferation and survival. Modification of the membrane lipid environment by AGPIn-3LCs supplementation inhibits these signaling pathways and increases the efficacy of docetaxel in mammary tumor cell lines and in a preclinical rodent model of native mammary tumors. Moreover, in this mammary tumor model we have found another molecular target regulated by AGPIn-3LCs: epiregulin, a member of the EGF family. AGPIn-3LCs inhibit epiregulin-VEGF induced in endothelial cells and induce a remodeling of tumor vasculature. Furthermore, AGPIn-3LCs act on the tumor microenvironment directly. This thesis work provides additional arguments for the use of AGPIn-3LCs as adjuvant molecules to reduce the resistance of breast tumors to anticancer agents

Yellowtail Kingfish (Seriola lalandi) (YTK) are carnivorous marine finfishes that are commercially farmed in Australia. YTK present the greatest opportunity for expansion of the Australian aquaculture industry, but improved diet formulations and feed conversion ratios are essential for production gains and economic upscaling. Lipids constitute a major cost component of aquafeeds but lipid composition has not been optimised for YTK. The purpose of this research was to increase understanding of how YTK utilise dietary lipids, and to improve feed conversion efficiency and product quality for human consumers. Fish oil (FO) as a dietary lipid source is central to this research as YTK require dietary omega 3 (n-3) long chain polyunsaturated fatty acids (LC PUFA) from FO for healthy development and growth, but FO is limited and less economically sustainable than other types of oil/lipid. The first study presented in this thesis sought to benchmark the fatty acid composition of wild YTK compared to aquacultured YTK. Tissue total lipid content was on average 4-times higher in aquacultured than wild YTK, with significantly higher concentrations of total saturated, omega 9, omega 7 and omega 6 fatty acids in tissues, but n-3 LC PUFA concentrations were not significantly different in the white muscle of wild and aquacultured YTK. The second and third studies were carried out with YTK grown in tanks using aquafeeds with varying lipid formulations. Generally, the fatty acid composition of aquacultured fish is reflective of the composition of aquafeeds, however this was not always the case for YTK in the following experiments. The key findings were that YTK have the capacity to spare in full Docosahaexonoic Acid (DHA) in white muscle at the expense of oleic acid (18:1n-9) when dietary levels of n-3 LC PUFA were <1.6 g 100 g-1 feed and that the digestibility of saturated fatty acids decreased with increasing chain length. Both of these findings could be used to manipulate dietary formulations and improve utilisation of n-3 LC PUFA. The fourth study investigated the potential for finishing diets to be utilised to modify the tissue fatty acid composition of YTK prior to harvest. Results showed significant changes in white muscle n-3 LC PUFA over 33 days at warm water temperatures, however further research was recommended to optimise the duration of finishing periods under a range of conditions. It was also recommended that the n-3 LC PUFA content of YTK feeds be closely monitored with strict lower limits set to ensure optimal product quality. The fifth and final study validated a method for the quantification of bioactive free fatty acid and oxylipin levels in YTK blood plasma. The approach was then used to determine the effects of dietary levels of n-3 LC PUFA on plasma free fatty acids and oxylipin bioactives. This method provides a new tool for aquaculture nutritionists to assess the impact of changes to YTK aquafeed formulations. In summary, this thesis has provided insight into the factors that affect fatty acid utilisation in YTK which have the potential to positively influence future aquafeed formulations, while also providing new methods to investigate lipid metabolism in the future.
Thesis (Ph.D.) -- University of Adelaide, School of Agriculture, Food and Wine, 2019

Circulating lipids play an important role in human physiology and pathophysiology. Lipids, as the major components in various cellular membranes, are involved in homeostatic regulation, particularly in relation to immune function and inflammatory mechanisms. With the growing global prevalence of lifestyle-related diseases, including obesity, diabetes, cardiovascular disorders and cancers, dietary lipids have received a great attention. Long-chain omega-3 polyunsaturated fatty acids (LC n-3 PUFA) have been associated with a broad range of health benefits. The three main LC n-3 PUFA are eicosapentaenoic acid (EPA, 20:5n-3), docosahexaenoic acid (DHA, 22:6n-3) and docosapentaenoic acids (DPA, 22:5n-3). Fish oil and krill oil are currently the most available sources of EPA and DHA as over-the-counter supplements, although other marine sources such as algae oil are also rich in EPA and DHA. Krill oil, derived from Antarctic krill (Euphausia Superba), is rich in EPA and DHA found in phospholipids (predominantly phosphatidylcholine) rather than triacylglycerol (TAG), in which EPA and DHA in fish oil are found. Krill oil also contains astaxanthin, a carotenoid contributing to its red colour which may also have beneficial health effects (Barros et al. 2014a, Pashkow et al. 2008). Despite a number of studies examining the effects of krill oil compared with fish oil on the incorporation of LC n-3 PUFA into different tissues, the outcomes have been conflicting, which might be associated with the different study designs using different chemical forms of fish oil and/or different doses of LC n-3 PUFA, and focusing at different target tissues. The research presented in this thesis consists of nine chapters covering a literature review (Chapter two) and two intervention studies in humans (Chapters four, five, six and seven) which have examined the effect of krill oil compared with fish oil on the incorporation of LC n-3 PUFA into plasma lipid fractions. There were a postprandial and a longer-term (30 days) intervention studies, and both clinical studies were randomised crossover designs involved healthy women (n = 10 and n = 11, respectively). All participants were instructed to maintain the habitual dietary intake and habitual physical activity throughout the interventions. The aim of the postprandial study was to compare the incorporation of LC n-3 PUFA into the plasma and circulating lipids in plasma and chylomicron fractions from five capsules (1 g each) of krill oil compared with five capsules (1g each) of fish oil and 5 g of the olive oil (control) over a 5-hour postprandial period. The second study aimed to investigate the longer-term effect of krill oil supplementation (containing 1,269 mg/d of LC n-3 PUFA including EPA, DHA and DPA) on the plasma LC n-3 PUFA, plasma circulating TAG and inflammatory biomarkers compared with fish oil supplementation (containing the closest possible match to these fatty acids from the capsules, 1,441 mg/d) over a 30-day intervention period. In both studies, lipidomics, was applied to identify the differences in plasma lipid molecular responses between krill oil and fish oil supplementation. Using this technique, a number of plasma lipid classes, and lipid molecular species containing EPA and DHA were identified and quantified. In the 5-hour postprandial study (Chapters four and five), there were no significant differences in the levels of TAG or cholesterol in plasma or chylomicron between the three study oil interventions, although the expected increases in chylomicron TAG were observed in all groups. In comparison to the olive oil, both krill oil (containing 907 mg of LC n-3 PUFA) and fish oil (containing 1,441 mg of LC n-3 PUFA) supplementation significantly increased the level of plasma EPA, which plateaued after three hours; there were no significant differences in the plasma EPA levels between krill oil and fish oil supplementation groups. There were no significant changes in either DHA or DPA between the three groups. Krill oil, with a lower dose of EPA in this study, showed a similar incorporation outcome of EPA into plasma lipids as fish oil. Given that there were 31% less EPA from krill oil, these results indicate a differential extent of incorporation of EPA between krill oil and fish oil, suggesting that EPA from krill oil may be more efficiently incorporated into the plasma than fish oil. The advanced technique for lipidomics was performed by high-performance liquid chromatography-mass spectrometer analysis (HPLC MS/MS), which was able to identify and quantify changes in various lipid molecular species containing LC n-3 PUFA in both the postprandial and the longer-term studies. Therefore, the HPLC MS/MS facilitated a comparison between differences in the individual lipid molecular species between krill oil and fish oil supplementation. A more sensitive setting of HPLC MS/MS was applied to the postprandial data than the longer-term data, based on the settings applied by the research laboratory at Baker Heart and Diabetes Institute where these analyses were conducted. In Chapter five, the postprandial plasma lipidomic changes are reported at hours zero (baseline), 3 and 5. A total of 29 lipid classes (≥ 500 pmol/mL) (for example: TAG, diacylglycerol (DAG), phosphatidylcholine (PC), cholesterol esther (CE)) were identified; six of these including O-linked phosphatidylethanolamine classes had significantly greater the incremental area under the curve from baseline (net iAUC 0-5 h) after krill oil supplementation compared with fish oil supplementation. Over the postprandial period, 56 EPA-containing and 76 DHA-containing molecular species (for example 16:0-20:5-PC, 16:0-18:1-20:5-TAG, 16:0-22:6-PC, 16:0-18:1-22:6-TAG) were significantly increased after both krill oil and fish oil supplementation. There were 33 phospholipid molecular species containing EPA, and 16 of these molecular species, including six ether-phospholipid molecular species had significantly greater increased net iAUC 0-5 h after krill oil than fish oil supplementation. In contrast, for TAG and DAG molecular species containing EPA, seven out of a total of 21 showed significantly increased net iAUC 0-5 h for fish oil compared with krill oil. Put simply, the EPA from krill oil was associated with increases in phospholipid EPA-molecular species, while the EPA from fish oil was associated with increased TAG and DAG EPA-molecular species. There were 49 phospholipid molecular species containing DHA, and 11 of these including six ether-phospholipid molecular species, had significantly greater increased net iAUC 0-5 h after krill oil supplementatin than fish oil supplementation. In a total of 61 AA-containing molecular species (for example 16:0-20:4-PC, 16:1-20:4-DAG) identified, there were 51 phospholipid molecular species containing AA, and seven of these including six ether-phospholipid molecular species, had significantly greater increased net iAUC 0-5 h after krill oil supplementation than fish oil. A novel finding from this postprandial study was that there was a consistent trend that ether-phospholipid classes (O-linked (containing an alkyl bond) or P-linked (containing an alkenyl bond) phosphatidylcholine and phosphatidylethanolamine) were significantly increased after krill oil supplementation, but decreased after fish oil supplementation. Consistently, it was found that EPA- and DHA-containing ether-phosphatidylethanolamines were significantly increased after the krill oil supplementation, but decreased after the fish oil supplementation. While the significance of this finding is not clear, it is worth noting that plasma levels of O- and P-linked phosphatidylethanolamine have been reported to be decreased in a number of disease states including Alzheimer’s disease. Little is known about the origin of these ether-phospholipids in plasma, but the fact that krill oil increased their post-prandial levels and fish oil decreased them is a clear differentiation between these two omega-3 oils. In the longer-term study (Chapters six and seven), EPA, DHA and DPA were significantly increased after both krill oil and fish oil supplementation over the 30-day period (p < 0.001). The main response to the 30-day krill oil supplementation was that the increase of plasma EPA level was significantly greater in the net iAUC 0-30 d than that of fish oil supplementation (p < 0.05). Both krill oil and fish oil significantly reduced plasma TAG over the intervention period (p < 0.05 and p < 0.01, respectively), but no significant differences were observed between the two groups. Over the 30-day intervention period, some plasma pro-inflammatory cytokines including IL-1β, IL-10, IL-4 and IL-5 (p ≤ 0.05) were significantly reduced after krill oil supplementation, while no such changes were found after fish oil supplementation. In Chapter seven, the long-term lipidomic changes (≥ 500 pmol/mL), at days zero (baseline), 15 and 30, are reported. Twenty three EPA-containing and 46 DHA-containing molecular species were significantly increased after both krill oil and fish oil supplementation over the 30-day supplementation period. Among EPA-, DHA-, and DPA-containing molecular species, there were 20 cases of net iAUC 0-30 d significant differences between the two supplementation. Fourteen of these molecular species in phospholipid species, including 12 ether-phospholipid species, had significantly greater increased net iAUC 0-30 d after krill oil than fish oil (p ≤ 0.05) supplementation. Consistently, it was found that EPA- and DHA-containing ether-phospholipid species, including six ether-phosphatidylethanolamines, were significantly increased after the krill oil supplementation, and decreased after the fish oil supplementation. The changes in the ether-phospholipids in the long-term trial were consistent with the changes described in the postprandial trial (Chapter five). These results support strongly the differentiation between krill oil and fish oil although there are still many unanswered questions flowing from this novel finding. What is known about plasmalogens is that they play a role in anti-inflammatory response, which might be linked to the significant decrease in pro-inflammatory cytokines observed in the present study. Overall, both postprandial and longer-term studies demonstrated that EPA from krill oil is efficiently incorporated into plasma, has a similar effect on the plasma TAG-lowering and a greater efficacy on the plasma inflammatory biomarkers when compared with fish oil. No previous studies have investigated plasma lipidomic responses to krill oil and fish oil supplementation in humans. There were significant increases in molecular species containing EPA and DHA following supplementation with krill oil and fish oil over both the postprandial and the longer-term periods. The plasma lipidomic changes of net iAUC over both intervention periods were significantly different between krill oil and fish oil supplementation, particularly for phospholipids (krill oil resulted in a greater increase than fish oil) and TAG (fish oil resulted in a greater increase than krill oil, as described in Chapter five). A novel aspect identified in this study was that krill oil increased ether-phospholipids, particularly ether-linked phosphatidylethanolamine, whereas fish oil decreased ether-phospholipids. The biological relevance of this novel lipidomic finding has yet to be fully explored.

The potential for supplemented omega-3 polyunsaturated fatty acids (PUFA) to increase sow reproductive performance when supplied from isocaloric diets containing low levels (3 – 6 g/kg of diet) of fish oil as a partial replacement for tallow was investigated. In the first experiment, there was an increase of 1 piglet live born (P < 0.05) to sows at the subsequent parity fed a supplemented diet before farrowing and during lactation. In contrast, litter size was unaffected when gilts were fed a supplemented diet with fish oil during puberty and early pregnancy. Furthermore in gilts, increasing the duration or level of supplementation did not improve litter size or embryo survival, possibly due to their inherently high level of fertility (82% embryo survival). In subsequent experiments, the effect of omega-3 supplementation on reproduction was evaluated in older parity sows known to have an inherently lower level of fertility compared with gilts. In parity 4 – 7 sows fed a supplemented diet prefarrowing and during lactation continuing to mating, embryo survival at 23 d was increased (Omega-3 70% vs 61% in Controls; P =0.054), without affecting ovulation rate. Subsequent experiments examined the response when supplemented diets were fed either during lactation continuing to mating; or after mating and during early gestation; or across both periods from lactation through to early gestation. Litter size born was maximised in the subsequent parity in sows fed fish oil diets from lactation to early gestation for 28 d, with the response being greatest in higher parity sows (+0.7 live born; and +0.9 total born, P < 0.05). In the following experiment this increases was associated with a 19% increase in embryo survival with omega-3 supplementation (P = 0.061). There was no effect on live weight or backfat during lactation; litter weight gain; piglet wean weight; and sow intake when gilts or sows were fed supplemented diets. The increase in embryo survival and litter size consistently observed in the sow studies was associated with increases in the omega- 3 PUFA eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) and were independent of energy intake or energy metabolism. Partially replacing tallow (wt/wt) with 3 – 6 kg levels of fish oil did not change plasma levels of the essential omega-6 fatty acids, linoleic acid (LIN) and arachidonic acid (ARA). Using in-vitro cultures of granulosa cells it was demonstrated that progesterone production is increased with prostaglandin E3 and there was evidence for PGE₃ to enhance the steroidogenic response to PGE₂. It is proposed that specific long-chain omega-3 fatty acids increases embryo survival in older sows due to improved oocyte quality and/or embryo development, possibly through synergistic activities of PGE₂ and PGE₃ on progesterone levels in the local ovarian- uterine circulation. Supplementation of diets with EPA and DHA from fish oil offers pig producers a nutritional approach to improve sow litter size in older parities thereby increasing longevity and lifetime performance.
Thesis (Ph.D.) -- University of Adelaide, School of Paediatrics and Reproductive Health, 2012

Omega-3 (n-3) long chain polyunsaturated fatty acids (LCPUFA), particularly eicosapentaenoic acid (EPA, 20:5n-3) and docosahexaenoic acid (DHA, 22:6n-3), are important for normal health as well as growth and brain development in humans. These fatty acids can be consumed in the diet directly, or synthesised from short chain PUFA consumed in the diet. Fish, particularly in species with a high fat content like salmon, are a major source of these beneficial fatty acids in the human diet. Fish production from aquaculture continues to expand due to a growing human population and demand for fish. Currently there is a reliance on fish oil and fish meal derived exclusively from wild fish as the primary lipid and protein source in fish feeds. Depleted wild fish stocks have made this source of n-3 LCPUFA unsustainable and alternative sources of n-3 LCPUFA are required to fill the void. Most animal species can convert the plant derived 18 carbon (C18) n-6 linoleic acid (LA, 18:2 n-6) and n-3 α-linolenic acid (ALA, 18:3 n-3) to 20 and 22 carbon (C20-C22) LCPUFA by using a series of enzymes to extend and alter the saturation level. There are two types of enzymes responsible for desaturating and elongating fatty acids are desaturases and elongases. The genes associated with these processes appear to be regulated by the extremes of dietary PUFA intake but the extent is currently unclear. This thesis is aimed to examine the effect of dietary PUFA on tissue n-3 LCPUFA levels in animals (rat and fish) after the consumption of diets with increasing levels of ALA, and to investigate whether the expression of desaturases and elongases is involved in the regulation of lipid metabolism and therefore LCPUFA biosynthesis. Furthermore, this thesis also investigated the potential enzyme functions of barramundi Δ6 desaturase and elongase using a yeast heterologous system. Experiments showed that while high ALA diets consistently produced higher levels of n-3 LCPUFA in rat tissues than low ALA diets, mRNA abundance of the Δ6 desaturase (FADS2) and elongase 2 (ELOVL2) genes were increased only in animals fed the low PUFA reference diet compared to those fed diets with adequate to high PUFA levels. There was no correlation between the gene expression of desaturases, elongases or transcription factors and the levels of EPA, docosapentaenoic acid (DPA, 22:5 n-3) or DHA in rat blood, liver and other tissues as a result of feeding increasing levels of ALA. In barramundi however, while vegetable oils induced significant increases in mRNA abundance of FADS2 and ELOVL genes compared with those fed the fish oil-based commercial diet, the tissue EPA, DPA and DHA levels were not increased. It is therefore hypothesised that the enzyme activity of barramundi Δ6 desaturase was low and therefore limited the effectiveness of the enzymes in the LCPUFA pathway to produce EPA and DHA. Furthermore, a large amount of variation between individual fish in DHA levels among those fed the vegetable oil-based diets was found, and this may provide a possibility for a future breeding program of barramundi for better DHA production. Barramundi FADS2 and ELOVL genes were also cloned into yeast cells and performed functional expression of the two enzymes. Results revealed that the barramundi Δ6 desaturase also showed Δ8 desaturase activity and the elongase showed a broad range of fatty acid specificity with the greatest activity with EPA. In addition, a significant amount of the desaturation and elongation fatty acid products could be detected in the culture medium at various time points after the addition of fatty acid substrates, and that it was important to take the levels of fatty acids in the medium into account when it came to calculating enzyme activity.
Thesis (Ph.D.) -- University of Adelaide, School of Agriculture, Food and Wine, 2011

Research Doctorate - Doctor of Philosophy (PhD)
The consumption of foods rich in saturated fats has been associated with elevated blood lipid levels and consequently with risk for numerous chronic diseases, such as coronary heart disease. However, understanding the effects of replacing saturated fat in the diet is complex, and the health effects of reducing saturated fat consumption clearly depend on what substitutions are made. Furthermore, studies using animal models have demonstrated that dietary saturated fats raise triglyceride levels only when the diet is deficient in omega-3 polyunsaturated fatty acids (n-3PUFA). The n-3PUFA are known for their potential to help in managing hyperlipidaemia for the prevention of coronary heart disease, as well as for their anti-inflammatory, anti-arrhythmic and anti-aggregatory potential. In addition, research in human and animal models has shown competition for important enzymes in the metabolism of omega-6 polyunsaturated fatty acids (n-6PUFA) and n-3PUFA, with high consumption of n-6PUFA leading to an increase in their metabolism at the expense of n-3PUFA. This leads to an increased production of n-6PUFA derived eicosanoids, which are pro-inflammatory and pro-aggregatory, in contrast with those derived from n-3PUFA, which are less inflammatory and aggregatory. Therefore, we hypothesised that consumption of saturated fats, would not adversely influence coronary heart disease risk factors (blood lipid levels, lipoprotein profiles, platelet aggregation and inflammation) when the diet was balanced with an adequate intake of n-3PUFA. Moreover, we hypothesized that the health benefits obtained with the consumption of n-3PUFA would be maximised by including foods rich in saturated fat and reducing the consumption of vegetable oils (rich in n-6PUFA) in the diet. Our first aim, addressed in chapter 3, was to establish the basis for our hypothesis, analysing the literature on saturated fatty acids to identify the contradictions in the literature to date and to highlight the gaps in knowledge gained from previous interventional and epidemiological studies. We have observed that although many studies have associated saturated fatty acids with hyperlipidaemia and cardiovascular disease risk factors, there is still much contradiction on the subject with not all studies finding the same association. The key studies relating saturated fat consumption and heart health made no mention about the presence or absence of n-3PUFA as a possible confounding factor. This may have been due to the lack of knowledge about the existence of n-3PUFA or an inability to determine n-3PUFA concentration in most of the early studies. Therefore the missing link in the research on cardiovascular disease risk and dietary fats could be an ignorance of the interactions between different dietary fats and the effect of this interaction. In Chapters 4a and 4b we aimed to determine if LCn-3PUFA and the other dietary fats interact during digestion, absorption, re-esterification into triglycerides and assembly into chylomicrons to modulate circulating lipid levels postprandially. In a randomised cross-over design, we investigated the effect of feeding meals rich in either saturated fatty acids or n-6PUFA in conjunction with LCn-3PUFA on plasma lipid (triglycerides and total, low density lipoprotein and high density lipoprotein cholesterol) and fatty acid levels. The postprandial lipemic response and fatty acid kinetics were similar after the consumption of both meals and suggest that the competition between n-3 and n-6PUFA may be a longer term phenomenon, not just a postprandial effect. The aim of Chapter 5 was then to determine if there were interactions between LCn-3PUFA and other dietary fats in the longer term (6 weeks). Therefore, in a randomized parallel design intervention we investigated the longer-term effects of LCn-3PUFA supplementation in subjects consuming diets enriched in either saturated fatty acids or n-6PUFA, on blood lipid profiles and on the incorporation of fatty acids into plasma and erythrocyte lipids. Long chain omega-3 polyunsaturated fatty acids were incorporated to a greater extent into the plasma and erythrocyte lipids of subjects consuming the saturated fat rich diet compared to the n-6PUFA rich diet, although total and low density lipoprotein (LDL) cholesterol were also increased. Plasma samples of the subjects who completed the intervention in chapter 5 were then further analysed in chapter 6 for lipoprotein profiles, with the aim of determining if the increase in plasma cholesterol levels was due to changes in the lipoprotein particle concentration or size. The increase in LDL cholesterol was due to an increase in the less atherogenic, large, buoyant LDL particles rather than the small, dense LDL particles. In chapter 7, the aim was to determine if pre-supplementation rather than co-supplementation with LCn-3PUFA would improve the effect of the major dietary fat groups on plasma lipids and lipoprotein profiles. Therefore, in a randomized parallel design clinical intervention, we examined the effect of increasing the omega-3 index of subjects before randomizing them to a diet rich in either saturated fatty acids or n-6PUFA. The diet rich in saturated fatty acids increased, while the diet rich in n-6PUFA decreased, total and LDL cholesterol, independently of LCn-3PUFA supplementation. However, the saturated fatty acid rich diet caused a further increase in plasma and erythrocyte LCn-3PUFA compared to the n-6PUFA rich diet. In conclusion, the results presented in this thesis demonstrate that the background dietary fat is a determinant of the degree of incorporation of LCn-3PUFA into plasma and tissue lipids. The consumption of a saturated fat rich diet did indeed cause an increase in plasma cholesterol levels. However, the rise in circulating cholesterol levels following saturated fat consumption is accompanied by an increase in the less atherogenic LDL particle size, when the LCn-3PUFA status is adequate, which is likely to reduce the detrimental effects. In addition, there was a concurrent increase in incorporation of LCn-3PUFA into plasma and erythrocytes, which may have benefits, independent of cholesterol or blood lipids. Hence, this thesis paves the way for further research to examine the impact of increased plasma and tissue LCn-3PUFA levels as a result of saturated fat consumption with adequate LCn-3PUFA intakes, on cardiovascular health risk indicators, such as inflammation, hypertension, platelet aggregation and endothelial function.

Microalgae are considered a promising alternative source of omega-3 long chain-polyunsaturated fatty acids (ω-3 LC-PUFAs) since they are the primary producers of eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids in the marine environment. Extraction methods commonly used for the isolation of these bioactives are based on conventional techniques, which imply the use of high volumes of organic solvents and high energy requirements, producing health and environmental problems. In this sense, greener alternatives need to be developed to meet the global consumer demand for natural ingredients and eco-friendly products. In this work, the use of ultrasonic-assisted enzymatic extraction (UAEE) technology in combination with environmentally friendly solvents was evaluated as a green strategy to efficiently extract the omega-3 lipids from the microalga Nannochloropsis gaditana. The microalgal biomass was pretreated with different commercial enzymes (Viscozyme® L, Celluclast® 1.5 L, and Saczyme® Yield) and the results were compared with the traditional Folch method (2:1 chloroform/methanol). A promising extraction approach was developed using Saczyme® Yield and ethanol as solvent, achieving a lipid yield of 25.7% ± 0.5, comparable to the results obtained with the traditional method (27.3% ± 0.7) (p > 0.05). Similar omega-3 content was found by GC-MS analysis for both lipid extracts (30.2% ± 2.9 and 29.2% ± 1.0 for the green and the traditional method, respectively), showing that the environmentally friendly approaches did not negatively affect the fatty acid profile. Additionally, the bioactivity of the produced lipids was investigated by a spectrum of in vitro cell-based assays measuring potential endpoints of interest like cytotoxicity, antioxidant and anti-inflammatory activities. In conclusion, this work provides relevant results for new eco-friendly extraction approaches to produce functional omega-3 lipids with potential applications in the food industry, avoiding the use of toxic solvents and reducing the environmental impact.

Very-long-chain fatty acids (VLC-FA) have a chain length of ≥24 carbon atoms. They are generally not provided through dietary sources but generated endogenously, involving chain elongation of LC-FA by ELOVL4. There is emerging substantial evidence suggesting they play important roles in tissues where they are naturally found, including retina, skin, testis, and brain. Mutations in ELOVL4 have been associated with several tissue-specific conditions, suggesting these FA may be involved in the disease pathology. A lack of availability of these fatty acids for dietary interventions has however, until recently, made them difficult to investigate. After identifying VLC-FA in fish oil and developing a method for concentrating n-3 VLC-PUFAs in kg scale, our research team have conducted feeding trials to determine if they are taken up directly from diet through supplementation, and their effect on development and maturation of skin tissue. Salmon fed different dietary levels of the concentrate were analysed for tissue fatty acid composition by GC and histology by H&E and Von Kossa staining. After establishing a clear tissue-specific uptake, we conducted in-vitro trials where we observed promising effects by incubating skin cells from human and Atlantic salmon with n-3 VLC-PUFA concentrate in scratch assay and cell migration trials. The in-vitro results show improved cell migration, which is in line with our in-vivo findings and demonstrates a promising effect on skin tissue development, maturation, and skin cell migration. Here we will present our data and discuss the relevance of this in skin biology. As VLC-FA potentially play a critical role in skin barrier function and skin biology, understanding these FAs may lead to improvements in treatment of dermatological diseases and conditions.

The essential dietary fatty acid α-linolenic acid (ALA) can be converted into anti-inflammatory 18 carbon oxylipins or into longer chain n€‘3 polyunsaturated fatty acids (PUFAs) such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). The partitioning of ALA between these alternative metabolic fates is not understood. To address this, peripheral blood CD3+ T cells from healthy volunteers (18-30 years; n=10) were cultured for 48h, with or without concanavalin A (10µg/ml) in 10% (v/v) pooled donor plasma with low ALA (20 µM) or high ALA (40 µM) concentrations (1:10 [13C]€‘labelled/unlabelled). [13C]ALA metabolites were detected either by GC-isotope ratio mass spectrometry for intracellular PUFA or by LC-MS/MS for oxylipins in cell culture supernatant. The ratio of the labelled metabolites hydroxyoctatrienoic acid ([13C]HOTrE) and dihydroxyoctadecaenoic acid ([13C]DiHODE) to [13C]ALA were 1.8±0.2 / 7.2±1.1 and 0.9±0.2 / 4.3±0.6 for low / high ALA, respectively, compared to the eicosatrienoic acid ([13C]20:3n€‘3) to [13C]ALA ratio of 0.002±0.0001 / 0.02±0.003 in stimulated T cells. Results from unstimulated cells were similar. Furthermore, oxylipins from all PUFA precursors were analysed in the culture supernatant of the T cells. The ratio of oxylipin concentrations in high compared to low ALA cultures was 1.4±0.1 for EPA-derived dihydroxyeicosatetraenoic acid (DiHETE), 5.6±0.9 for DHA-derived dihydroxydocosapentaenoic acid (DiHDPE), 7.9±2.7 for resolvin RvE1 and 2.0±0.3 for resolvin RvD1. The total oxylipin profile was not altered significantly by mitogen stimulation. These findings show that ALA is used primarily by T cells for constitutive production of anti-inflammatory lipid mediators rather than synthesis of longer chain PUFA. Further, ALA addition changes the secreted oxylipins towards a less-inflammatory profile. This has implications for understanding the effects of dietary PUFA on immune function.

Microalgae is currently receiving strong consideration as a potential biofuel feedstock to help meet the advanced biofuels mandate of the 2007 Energy Independence and Security Act because of its theoretically high yield (gallons/acre/year) in comparison to current terrestrial feedstocks. Microalgae lipids can be converted into a variety of biofuels including fatty acid methyl esters (e.g. biodiesel), renewable diesel, renewable gasoline or synthetic paraffinic aviation kerosene. For algal methyl ester biodiesel, fuel properties will be directly related to the fatty acid composition of the lipids produced by the given microalgae strain. Several microalgae species under consideration for wide scale cultivation, such as Nannochloropsis, produce lipids with fatty acid compositions containing substantially higher quantities of long chain-polyunsaturated fatty acids (LC-PUFA) in comparison to terrestrial feedstocks. It is expected that increased levels of LC-PUFA will be problematic in terms of meeting all of the current ASTM specifications for biodiesel. For example, it is well known that oxidative stability decreases with increasing levels of LC-PUFA. However, these same LC-PUFA fatty acids, such as eicosapentaenoic acid (EPA: C20:5) and docosahexaenoic acid (DHA: C22:6) are known to have high nutritional value thereby making separation of these compounds economically attractive. Given the uncertainty in the future value of these LC-PUFA compounds and the economic viability of the separation process, the goal of this study was to examine the oxidative stability of algal methyl esters with varying levels of EPA and DHA. Tests were conducted using a Metrohm 743 Rancimat with automatic induction period determination following ASTM D6751 and EN 14214 standards, which call for induction periods of at least 3 hours and 6 hours, respectively. Tests were conducted at a temperature of 110°C and airflow of 10 L/h with model algal methyl ester compounds synthesized from various sources to match the fatty acid compositions of several algae strains subjected to varying removal amounts of roughly 0 to 100 percent LC-PUFA. In addition, tests were also conducted with real algal methyl esters produced from multiple sources. The bis-allylic position equivalent (BAPE) was calculated for each fuel sample to quantify the level of unsaturation. The induction period was then plotted as a function of BAPE, which showed that the oxidative stability varied exponentially with the amount of LC-PUFA. The results suggest that removal of 45 to 65 percent of the LC-PUFA from Nannochloropsis-based algal methyl esters would be sufficient for meeting existing ASTM specifications for oxidative stability.

Photoautotrophic microalgae are a novel source of biomass rich in lipids containing nutritionally interesting n-3 long-chain polyunsaturated fatty acids (n-3 LC-PUFA). However, microalgae incorporated into foods as whole biomass may lead to limited n-3 LC-PUFA uptake by the human body. Incorporation of microalgal oil instead of whole biomass is an interesting alternative. The extraction efficiency of these lipids can be enhanced by adding a cell disruption step during lipid extraction. Ultrasound assisted extraction (UAE) to enhance lipid recovery from microalgae has been covered in recent literature. UAE is used to speed up lipid extraction for analytical purposes or for industrially applicable lipid extraction of microalgae. In addition, the ultrasonication and lipid extraction by organic solvents of microalgal biomass has been performed in different set-ups: simultaneous ultrasonication and lipid extraction or ultrasonication and subsequent lipid extraction, ultrasonication of wet biomass or dry biomass. Often, no reference method is included to which the lipid extraction efficiency using UAE is compared and the total lipid content of the biomass is not always specified making it impossible to calculate the extraction efficiency. Therefore, the effectiveness of this cell disruption technique for industrial microalgae processing has not consistently been proven in literature. This study investigates the effect of UAE of Nannochloropsis sp. biomass on the lipid extraction efficiency and the lipid quality, expressed as free fatty acid content and peroxide value. The effect of UAE on wet and dry biomass in the absence or presence of organic solvents is compared. The effect of the solvent system, ultrasonication power and ultrasonication time is studied.The results show that the lipid extraction efficiency increases after UAE. The presence of some solvent systems leads to higher lipid extraction efficiency when UAE is performed on dry biomass compared to wet biomass while for other solvent systems, this is reversed.

In industrial fish filleting, around 30–70 % of the total weight of the fish end up as side streams (often called by- or co-products), such as the head, backbone, caudal fin, skin, and intestines. Currently, these fractions are dedicated to low value uses as fodder meals or mink feed, even if they contain significant amounts of protein, long chain (LC) n-3 polyunsaturated fatty acids (PUFA), and other nutrients such as vitamins and minerals. However, most fish processors mix their side streams, not least when it comes to small pelagic species like herring. This practice limits use of the side streams for food production since the raw material gets very complex, and since blood, enzymes and lipids from e.g., the viscera and head parts easily contaminate the cleaner parts like the backbones and tails, accelerating e.g., their oxidative or enzymatic degradation. In the present study, lipid oxidation in ice-stored sorted and minced herring fractions (head, backbone, viscera+belly flap, tail, fillet) from spring and fall, and its association with endogenous pro-oxidants, antioxidants and lipid substrates were investigated. Peroxide value (PV) and thiobarbituric acid reactive substances (TBARS) had increased significantly in all fractions after 1 day, but for both seasons, the most rapid PV and TBARS development occurred in head, which also had highest hemoglobin (Hb) levels and lipoxygenases (LOX) activity. Viscera+belly flap was overall the most stable part, and also had the highest -tocopherol content. Pearson correlation analyses across all five fractions confirmed a significant impact of Hb, LOX and -tocopherol on the lipid oxidation susceptibility, while content of total iron, copper, lipids or PUFA provided no significant correlation. Overall, the study showed which pro-oxidants that should be inhibited or removed to succeed with value adding of herring filleting side streams along with the fillet itself.

Various unicellular algae produce omega-3 (w3) very-long-chain polyunsaturated fatty acids (VLC-PUFA), which are rarely found in higher plants. In this research and other studies from our laboratories, it has been demonstrated that the marine unicellular alga Nannochloropsis (Eustigmatophyceae) can be used as a reliable and high quality source for the w3 VLC-PUFA eicosapentaenoic acid (EPA). This alga is widely used in mariculture systems as the primary component of the artificial food chain in fish larvae production, mainly due to its high EPA content. Furthermore, w3 fatty acids are essential for humans as dietary supplements and may have therapeutic benefits. The goal of this research proposal was to understand the physiological and biochemical mechanisms which regulate the synthesis and accumulation of glycerolipids enriched with w3 VLC-PUFA in Nannochloropsis. The results of our studies demonstrate various aspects of lipid synthesis and its regulation in the alga: 1. Variations in lipid class composition imposed by various environmental conditions were determined with special emphasis on the relative abundance of the molecular species of triacylglycerol (TAG) and monogalactosyl diacylglycerol (MGDG). 2. The relationships between the cellular content of major glycerolipids (TAG and MGDG) and the enzymes involved in their synthesis were studied. The results suggested the importance of UDP-galactose diacylglycerol galactosyl (UDGT) in regulation of the cellular level of MGDG. In a current effort we have purified UDGT several hundredfold from Nannochloropsis. It is our aim to purify this enzyme to near homogeneity and to produce antibodies against this enzyme in order to provide the tools for elucidation of the biochemical mechanisms that regulate this enzyme and carbon allocation into galactolipids. 3. Our in vitro and in vivo labeling studies indicated the possibility that phosphatidylcholine (PC) and phosphatidylethanolamine (PE) are associated with desaturation of the structural lipids, whereas shorter chain saturated fatty acids are more likely to be incorporated into TAG. 4. Isolation of several putative mutants of Nannochloropsis which appear to have different lipid and fatty acid compositions than the wild type; a mutant of a special importance that is devoid of EPA was fully characterized. In addition, we could demonstrate the feasibility of Nannochloropsis biomass production for aquaculture and human health: 1) We demonstrated in semi-industrial scale the feasibility of mass production of Nannochloropsis biomass in collaboration with the algae plant NBT in Eilat; 2) Nutritional studies verified the importance algal w3 fatty acids for the development of rats and demonstrated that Nannochloropsis biomass fed to pregnant and lactating rats can benefit their offspring.