Metabolism and functions of lipids and fatty acids in teleost fish

Lipids and their constituent fatty acids are, along with proteins, the major organic constituents of fish, and they play major roles as sources of metabolic energy for growth including reproduction, and movement including migration. Furthermore, the fatty acids of fish lipids are rich in ω3 long chain, highly unsaturated fatty acids (n-3 HUFA) that have particularly important roles in animal nutrition, including fish and human nutrition, reflecting their roles in critical physiological processes. Indeed, fish are the most important food source of these vital nutrients for man Thus, the long standing interest in fish lipids stems from their abundance and their uniqueness. This review attempts to summarise our present state of knowledge of various aspects of the basic biochemistry, metabolism and functions of fatty acids, and the lipids they constitute part of, in fish, seeking where possible to relate that understanding as much to fish in their natural environment as to farmed fish. In doing so, it highlights the areas that require to be investigated in greater depth and also the increasing application of molecular technologies in fish lipid metabolism which will fascilitate further advances through molecular biological and genetic techniques including genomics and proteomics

Cytosolic cholesterol ester hydrolase in adrenal cortex

Cholesterol ester hydrolase (CEH) in adrenocortical cytosol was known to be phosphorylated and activated, in response to ACTH in a cAMPdependent protein kinase mediated process. The purification of CEH from bovine adrenocortical cytosol was attempted. The use of detergents to solubilise the enzyme from lipid-rich aggregates was investigated and sodium cholate was found to be effective. A purification procedure using cholate solubilised enzyme was developed. The detergent interfered in the operation of the ion-exchange columns, and pure enzyme was not obtained. Hydrophobic chromatography was studied but it was found to be unsuitable for the purification of CEH under the conditions investigated. An affinity chromatography technique was developed using columns of glass beads coated non-covalently with cholesterol oleate. Some positive results were obtained but the low capacity of the columns combined with the low concentration of the enzyme in the tissue cytosol prevented further study of the purified activity. A further purification procedure utilising a non-ionic detergent and gradient sievorptive chromatography resulted in a 150-fold purification of CEH from bovine adrenocortical cytosol with a recovery of about 25%. The specific activity of the partially purified enzyme (CEHQ2 preparation) was approximately 60 nmol oleic acid produced.min 3.mg protein Labelling studies using the CEHQ2 preparation, [γ-32P] ATP and [1,3-3H] DFP suggested that the enzyme activity was associated with a protein with Mr approximately 84 000. A phosphoprotein phosphatase with Mr 35 000 wTas purified from bovine adrenocortical cytosol to a state approaching homogeneity. The purified 32 phosphatase was active when measured towards P-phosphoprotexn and p-nitrophenyl phosphate. The role of this phosphoprotein phosphatase in the modulation of cytosolic CEH activity was investigated, but the enzyme did not deactivate partially-purified CEH from bovine adrenocortical cytosol. Cytosolic CEH in rat adrenal was found to exhibit a diurnal variation in activity. The enzyme activity was significantly higher during the dark phase. Serum corticosterone concentration reflected this variation in CEH activity. In vivo experiments including acute ACTH administration and dexamethasone suppression of pituitary ACTH secretion suggested that ACTH was involved in the production and maintenance of the diurnal rhythm of rat adrenal cytosolic CEH activity. The protein components of bovine adrenocortical and rat adrenal lipid droplets were investigated. The lipid droplets from both species contained a major protein subunit with Mr 40 000 and several other minor proteins. The protein profile was similar in both species. The Mr 40 000 apoprotein was able to be phosphorylated in rat lipid droplets but not in bovine lipid droplets. The delipidated apoprotein was phosphorylated in both cases. Bovine adrenocortical lipid droplets contained about 4% by weight protein, but CEH was found to be only a minor component of the protein fraction

Omega-3 long-chain polyunsaturated fatty acids and aquaculture in perspective

In the 40 years since the essentiality of polyunsaturated fatty acids (PUFA) in fish was first established by determining quantitative requirements for 18:3n−3 and 18:2n−6 in rainbow trout, essential fatty acid (EFA) research has gone through distinct phases. For 20 years the focus was primarily on determining qualitative and quantitative EFA requirements of fish species. Nutritional and biochemical studies showed major differences between fish species based on whether C18 PUFA or long-chain (LC)-PUFAwere required to satisfy requirements. In contrast, in the last 20 years, research emphasis shifted to determining “optimal” levels of EFA to support growth of fish fed diets with increased lipid content and where growth expectations were much higher. This required greater knowledge of the roles and functions of EFA inmetabolismand physiology, and howthese impacted on fish health and disease. Requirement studies were more focused on early life stages, in particular larval marine fish, defining not only levels, but also balances between different EFAs. Finally, a major driver in the last 10–15 years has been the unavoidable replacement of fish oil and fishmeal in feeds and the impacts that this can have on n−3 LC-PUFA contents of diets and farmed fish, and the human consumer. Thus, dietary n−3 infish feeds can be defined by three levels. Firstly, the minimumlevel required to satisfy EFA requirements and thus prevent nutritional pathologies. This level is relatively small and easy to supply even with today's current high demand for fish oil. The second level is that required to sustainmaximumgrowth and optimumhealth in fish being fed modern high-energy diets. The balance between different PUFA and LC-PUFA is important and defining them is more challenging, and so ideal levels and balances are still not well understood, particularly in relation to fish health. The third level is currently driving much research; how can we supply sufficient n−3 LC-PUFA tomaintain these nutrients in farmed fish at similar or higher levels than in wild fish? This level far exceeds the biological requirements of the fish itself and to satisfy it we require entirely new sources of n−3 LCPUFA. We cannot rely on the finite and limited marine resources that we can sustainably harvest or efficiently recycle. We need to produce n−3 LC-PUFA de novo and all possible options should be considered

An approach to study the nutritional requirements of the bluefin tuna (Thunnus thynnus thynnus L.)

The research required in relation to the nutrition of bluefin tuna (BFT) is great. In previous trials in the domestication of large scombrids and in current capture-fattening schemes, nutritional studies have been few. Therefore, virtually nothing is presently known about the nutritional requirements for these species. Nutritional factors are important in any domestication programme in a variety of areas. Correct nutrition is a vital factor in the production of a successful broodstock with high fecundity and fertility producing large numbers of high quality eggs. Reproductive control itself is affected by nutritional factors such as lipid/energy content which can influence sexual maturation in other fish species. Successful larval rearing of marime fish is highly dependent upon suitable diets, whether live prey species or artificial, and their precise composition particularly in relation to fatty acids is an area that still demands much research for all marine species. Clues to the nutritional requirements of any animal can be obtained by looking at the natural food for that animal, in this instance, the natural prey species of the BFT and also by determining the composition of wild caught animals, both prey and predator. The latter certainly being a source of information in terms of lipids and fatty acid requirements. The few data available from previous trials can also add to the overall view. In this section we will briefly review the literature on what is known about the nutrition of the large scombrids and also, importantly, place the problems of their feeding and nutrition in a global perspective

Tuna nutrition and feeds: current status and future perspectives

Aquaculture is providing an ever-increasing proportion of fish in the human food basket prompting a search for new species to expand the range available to consumers. Large tunids/scombrids have long-since been a very valuable resource providing not only high quality protein, but also a rich source of the highly beneficial omega-3 (or n-3) long-chain polyunsaturated fatty acids including eicosapentaenoic and, especially docosahexaenoic acids in the human diet. Consequently, there is considerable interest worldwide in developing the culture of large tunids, including Atlantic northern bluefin tuna (Thunnus thynnus), Pacific bluefin tuna (Thunnus orientalis), southern bluefin tuna (Thunnus maccoyii) and yellowfin tuna (Thunnus albacares). Nutrition is vital to this development, playing key roles in reproductive success, including the establishment of successful broodstock producing high quality eggs and larvae, and ultimately the cost-effective production of nutritious seafood. This review summarises the rather fragmentary data that compromise the current state-of-the-art in relation to tuna nutrition and the development of artificial, formulated feeds for these species. In highlighting the various considerable challenges that feed development will pose, we discuss the future perspectives for tuna culture in terms of both fish and human nutrition and welfare, against the background of diminishing global marine resources

Could an El Niño event put dietary supplies of n-3 long-chain polyunsaturated fatty acids (EPA and DHA) in jeopardy

The beneficial effects of n-3 long-chain polyunsaturated fatty acids, especially EPA and DHA, for human health are widely recognised. Intakes of these fatty acids in many parts of Europe fall far short of the recommended intake, especially in young people. The most robust evidence from human health studies demonstrating beneficial effects is that for cardiovascular disease. Other beneficial effects have been reported for inflammatory diseases such as rheumatoid arthritis, Crohn’s disease and ulcerative colitis, cognitive and visual development in pre-term infants, and on various psychological/behavioural and psychiatric disorders. For European consumers fish is the main source of EPA and ­­DHA, especially oily fish, and in particular farmed Atlantic salmon. With evidence of a strong El Niño type event occurring in the Pacific currently (2015/16), fish stock biomasses have drastically declined and permitted fishing heavily reduced. Subsequent fish oil shortages have resulted in increased substitution of fish oil with vegetable oil in feeds for farmed salmon. A significant fall in EPA and DHA in these fish and a reduction in intake by consumers are expected in the short term. Avoiding fish oil shortages causing reductions in EPA and DHA intake by consumers in the future may be possible through the production of DHA-rich algal biomass if the cost can be reduced. Genetically modified crops which might also be a source of these fatty acids are unlikely to be acceptable for salmon feeds in Europe in the near future

Essential fatty acid deficiency in freshwater fish: the effects of linoleic, alpha-linolenic, gamma-linolenic and stearidonic acids on the metabolism of [1-14C]18:3n-3 in a carp cell culture model

The desaturation of [1-14C]18:3n-3 to 20:5n-3 and 22:6n-3 is enhanced in an essential fatty acid deficient cell line (EPC-EFAD) in comparison with the parent cell line (EPC) from carp. In the present study, the effects of competing, unlabeled C18 polyunsaturated fatty acids (PUFA), linoleic (18:2n-6), α-linolenic (18:3n-3), γ-linolenic (18:3n-6) and stearidonic (18:4n-3) acids, on the metabolism of [1-14C]18:3n-3 were investigated in EPC-EFAD cells in comparison with EPC cells. The incorporation of [1-14C]18:3n-3 in both cell lines was significantly reduced by competing C18 PUFA, with the rank order being 18:4n-3 > 18:3n-3 = 18:2n-6 > 18:3n-6. In the absence of competing PUFA, radioactivity from [1-14C]18:3n-3 in EPC cells was predominantly recovered in phosphatidylethanolamine followed by phosphatidylcholine. This pattern was unaffected by competing n-6PUFA, but n-3PUFA reversed this pattern as did essential fatty acid deficiency in the presence of all competing PUFA. The altered lipid class distribution was most pronounced in cells supplementedwith 18:4n-3. Competing C18 PUFA significantly decreased the proportions of radioactivity recovered in 22:6n-3, pentaene and tetraene products, with the proportions of radioactivity recovered in 18:3n-3 and 20:3n-3 increased, in both cell lines. However, the inhibitory effect of competing C18 PUFA on the desaturation of [1-14C]18:3n-3 was significantly greater in EPC-EFAD cells. The magnitude of the inhibitory effects of C18 PUFA on [1-14C]18:3n-3 desaturation was dependent upon the specific fatty acid with the rank order being 18:4n-3 >18:3n-3 >18:2n-6, with 18:3n-6 having little inhibitory effect on the metabolism of [1-14C]18:3n-3 in EPC cells. The differential effects of the C18 PUFA on [1-14C]18:3n-3 metabolism were consistent with mass competition in combination with increased desaturation activity in EPC-EFAD cells and the known substrate fatty acid specificities of desaturase enzymes. However, the mechanism underpinning the greater efficacy with which the unlabeled C18 PUFA competed with [1-14C]18:3n-3 in the desaturation pathway in EPC-EFAD cells was unclear

Polyunsaturated fatty acid metabolism in a cell culture model of essential fatty acid deficiency in a freshwater fish, carp (Cyprinus carpio)

Proliferation of an essential fatty acid deficient cell line from carp (EPC-EFAD; epithelioma papillosum carp-essential fatty acid deficient) is stimulated by supplementing the cells with C20, but not C18 polyunsaturated fatty acids (PUFA). It is hypothesized that the differential ability of the PUFA to stimulate proliferation of the EPC-EFAD cells may be related to the extent of the cells’ ability to desaturate and elongate C18 PUFA. In the present study, the metabolism of 14C-labeled C18 and C20 PUFA was investigated in EPC-EFAD cells in comparison with normal EPC cells. The incorporation of all the PUFA was significantly greater in EPC-EFAD cells but the rank order, 20:5n-3 > 18:3n-3 = 18:2n-6 >20:4n-6 was the same in both cell lines. The proportion of radioactivity from all labeled PUFA recovered in phosphatidylethanolamine and total polar lipids was significantly lower in EPC-EFAD cells compared to EPC cells, whereas the proportion of radioactivity recovered in all the other phospholipid classes and total neutral lipid was greater in EPC-EFAD cells. Both cell lines desaturated[1-14C]18:3n-3 and [1-14C]20:5n-3 to a greater extent than the corresponding (n-6) substrates but the desaturation of all the 14C-labeled PUFA was significantly greater in EPC-EFAD cells compared to EPC cells. The results showed that, although essential fatty acid deficiency had several significant effects on PUFA metabolism in EPC cells, the fatty acid desaturation/elongation pathway was not impaired in EPC-EFAD cells and so they can desaturate 18:3n-3 to 20:5n-3 and 22:6n-3, and 18:2n-6 to 20:4n-6. However, 20:4n-3 and 20:3n-6, and not 20:4n-6 and 20:5n-3, were the predominant C20 PUFA produced by the elongation and desaturation of [1-14C]18:3n-3 and [1-14C]18:2n-6, respectively. Therefore, the previously reported inability of 18:3n-3 and 18:2n-6, compared to 20:5n-3 and 20:4n-6, to stimulate proliferation of the cells is apparently not due to a general deficiency in the fatty acid desaturation pathway in EPC-EFAD cells but may be related to potential differences in eicosanoid profiles in cells supplemented with C18 PUFA compared to C20 PUFA

Effects of dichlorvos and formalin on fatty acid metabolism of rainbow trout (Oncorhynchus mykiss) skin cells in primary culture

The effects of sub-lethal doses of dichlorvos and formalin, antimicrobial/parasitic agents used in aquaculture, on lipid composition and metabolism of rainbow trout skin cells in primary culture were investigated. [1-14C]Stearic (18:0), [1-14C]linoleic (18:2n-6) and [1-14C]linolenic (18:3n-3) acids were used as tracers to determine effects on fatty acid incorporation and metabolism. Formalin increased cell numbers and reduced the lipid content of the cells and the incorporation of radioactive fatty acids. The effects of dichlorvos were qualitatively similar but quantitatively less. Formalin induced relatively small but significant changes in lipid class composition including a decreased proportion of phosphatidycholine with increased proportions of phosphatidylethanolamine and phosphatidylserine. Dichlorvos had no significant effect on lipid class compositions. The trout primary skin cells expressed substantial Δ9, Δ6 and Δ5 fatty acyl desaturase activities. Although, as expected, the cells were more active towards [1-14C]18:3n-3, the cells were unusually active towards [1-14C]18:2n-6. Both dichlorvos and, especially, formalin appeared to significantly inhibit Δ9 and Δ6 desaturase. Changes in the distribution of radioactivity between individual phospholipid classes was also influenced by formalin and dichlorvos, and this may be related to changes in desaturase activity. This study has shown that topically active agents used in aquaculture, formalin and dichlorvos, had a range of effects on the rainbow trout skin cell cultures that may affect cell proliferation and lipid and fatty acid metabolism. Both agents significantly inhibited desaturation of fatty acids, particularly of 18:2n-6 to 20:4n-6 and, as 20:4n-6 is a major eicosanoid precursor in fish and considering the importance of eicosanoids in the biochemistry of skin ,it is suggested that these agents may have direct effects on fish skin that may have important consequences for fish health in general

Impact of sustainable feeds on omega-3 long-chain fatty acid levels in farmed Atlantic salmon, 2006–2015

As the global population and its demand for seafood increases more of our fish will come from aquaculture. Farmed Atlantic salmon are a global commodity and, as an oily fish, contain a rich source of the health promoting long-chain omega-3 fatty acids, eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids. Replacing the traditional finite marine ingredients, fishmeal and fish oil, in farmed salmon diets with sustainable alternatives of terrestrial origin, devoid of EPA and DHA, presents a significant challenge for the aquaculture industry. By comparing the fatty acid composition of over 3,000 Scottish Atlantic salmon farmed between 2006 and 2015, we find that terrestrial fatty acids have significantly increased alongside a decrease in EPA and DHA levels. Consequently, the nutritional value of the final product is compromised requiring double portion sizes, as compared to 2006, in order to satisfy recommended EPA + DHA intake levels endorsed by health advisory organisations. Nevertheless, farmed Scottish salmon still delivers more EPA + DHA than most other fish species and all terrestrial livestock. Our findings highlight the global shortfall of EPA and DHA and the implications this has for the human consumer and examines the potential of microalgae and genetically modified crops as future sources of these important fatty acids