John R. Sargent

Recent developments in the essential fatty acid nutrition of fish

Abstract Because of competitive interactions in the metabolism of polyunsaturated fatty acids, tissue and bodily requirements for each of the three dietary essential fatty acids in marine fish, 22:6 n −3, 20:5 n −3 and 20:4 n −6, cannot be meaningfully considered in isolation. Rather, it is necessary to consider requirements in relative as well as absolute amounts, i.e., in terms of the ratio of 22:6 n −3:20:5 n −3:20:4 n −6. This is illustrated by recent research in our laboratories which has suggested that the optimal dietary ratio of 22:6 n −3:20:5 n −3 in sea bass larvae is circa 2:1 with the optimal dietary ratio of 20:5 n −3:20:4 n −6 being circa 1:1. The optimal dietary ratio of 22:6 n −3:20:5 n −3 in turbot and halibut larvae is similarly circa 2:1 but the optimal dietary ratio of 20:5 n −3:20:4 n −6 in these species is 10:1 or greater. In addition, studies with salmon parr point to dietary 18:3 n −3 and 18:2 n −6 being important in determining the optimal tissue ratio of 20:5 n −3:20:4 n −6 for successful parr–smolt transition. We deduce that differences in essential fatty acid requirements for different species of fish reflect different dietary and metabolic adaptations to different habitats and consider how such knowledge can be exploited to develop improved diets for fish, especially in their early stages of development.

Lipid nutrition of marine fish during early development: current status and future directions

Abstract Research on the dietary requirements of marine fish larvae has evolved from considerations of optimal dietary levels of n −3 HUFA to considerations of optimal dietary ratios of the two principal HUFAs, 22:6 n −3 and 20:5 n −3, and more recently to considerations of optimal dietary levels and ratios of all three dietary essential fatty acids, 22:6 n −3, 20:5 n −3 and 20:4 n −6. Our present understanding of the requirements and optimal dietary balance of 22:6 n −3, 20:5 n −3 and 20:4 n −6 is reviewed. Limitations of enriching live feed are considered, particularly from the point of view of achieving an optimal balance between levels of phospholipids and triacylglycerols in enriched live feeds that generate an optimal blend of essential fatty acids and energy-yielding fatty acids. It is concluded that the ideal marine fish larval diet is one containing circa 10% of the dry weight as n −3 HUFA-rich, marine phospholipids with less than 5% triacylglycerols, as exemplified by the lipid compositions of marine fish egg yolk, marine fish larvae themselves and their natural zooplankton prey. Such diets provide 22:6 n −3, 20:5 n −3 and 20:4 n −6 in the desired levels and ratios and simultaneously satisfy known requirements for phospholipids, inositol and choline. Approaches to developing marine fish larval diets more closely resembling this “gold standard” diet are considered.

Requirements, presentation and sources of polyunsaturated fatty acids in marine fish larval feeds

Abstract Current procedures for optimising the presentation of dietary polyunsaturated fatty acid (PUFA) to marine fish larvae are reviewed in relation to the advantages and disadvantages of using (a) single-cell eukaryotic organisms or, (b) purified oils, as primary sources of these essential nutrients in larval production systems. For the former option (a), phototrophic and heterotrophic organisms can both be used to advantage and current knowledge of the origins and functions of PUFA in marine organisms can help identify suitable organisms. For the latter option (b), control of PUFA peroxidation by various antioxidant systems is essential. Progress in this field requires definition of the optimal dietary ratio of 22:6( n −3): 20:5( n −3): 20:4( n −6); the significance of phospholipid vs. neutral lipid in larval diets, and larval requirements for antioxidant vitamins.

Arachidonic acid in aquaculture feeds: current status and future opportunities

The importance of arachidonic acid (20:4n6, ARA) in fish nutrition has tended to be overlooked in preference to eicosapentaenoic (20:5n3, EPA) and docosahexaenoic acids (22:6n3, DHA), probably due to the predominance of the latter two HUFA in fish tissues. However, despite the abundance of EPA and DHA in fish tissues, the importance of ARA as the primary eicosanoid precursor has been recognised for some time. Only very recently has the relative importance of ARA been given due acknowledgement and already the potential benefits to fish physiology and biochemistry of optimising the ARA nutrition of fish are becoming clear. In the coming decade, the role of dietary ARA should be the focus of considerably more research activity than has occurred to date. The results will undoubtedly emphasise why this ubiquitous HUFA cannot be underestimated in fish nutrition. D 2003 Elsevier Science B.V. All rights reserved.

Analyses of lipids and fatty acids in ripe roes of some Northwest European marine fish

Lipid class analyses and fatty acid analyses of neutral and polar lipids were carried out on ripe roes of herring, cod, haddock, whiting, saithe, sand eel and capelin. Total lipid was 10–26% of roe dry weight. The species with the highest total lipid, sand eel and capelin, also had the highest percentage of neutral lipid in total lipid, 77% and 49% respectively. In the other species, phospholipids accounted for 62–77% of roe total lipid. Both the neutral lipids, and especially the phospholipids, of all species were very unsaturated because of high concentrations of (n−3) polyunsaturated fatty acids (PUFA), frequently amounting to 50% of the total egg lipid. Phosphatidylcholine (PC) and phosphatidylethanolamine (PE) had similar fatty acid compositions in all species, with an average ratio (n−3)/(n−6) of ca. 20∶1. Phosphatidylinositol (PI) consistently had high concentrations of 18∶0 and 20∶4 (n−6) with an average ratio of (n−3)/(n−6) of 1.8∶1. Requirements for high levels of (n−3) PUFA in the embryonic and early larval development stages of marine fish are suggested as is a special role for the 20∶4(n−6) in PI.

A vertebrate fatty acid desaturase with Δ5 and Δ6 activities

Δ5 and Δ6 fatty acid desaturases are critical enzymes in the pathways for the biosynthesis of the polyunsaturated fatty acids arachidonic, eicosapentaenoic, and docosahexaenoic acids. They are encoded by distinct genes in mammals and Caenorhabditis elegans. This paper describes a cDNA isolated from zebrafish (Danio rerio) with high similarity to mammalian Δ6 desaturase genes. The 1,590-bp sequence specifies a protein that, in common with other fatty acid desaturases, contains an N-terminal cytochrome b5 domain and three histidine boxes, believed to be involved in catalysis. When the zebrafish cDNA was expressed in Saccharomyces cerevisiae it conferred the ability to convert linoleic acid (18:2n-6) and α-linolenic acid (18:3n-3) to their corresponding Δ6 desaturated products, 18:3n-6 and 18:4n-3. However, in addition it conferred on the yeast the ability to convert di-homo-γ-linoleic acid (20:3n-6) and eicosatetraenoic acid (20:4n-3) to arachidonic acid (20:4n-6) and eicosapentaenoic acid (20:5n-3), respectively, indicating that the zebrafish gene encodes an enzyme having both Δ5 and Δ6 desaturase activity. The zebrafish Δ5/Δ6 desaturase may represent a component of a prototypic vertebrate polyunsaturated fatty acids biosynthesis pathway.

Lipids and life strategy of Arctic Calanus

Abstract The three Arctic Calanus species, C. finmarchicus (Gunnerus, 1765), C. glacialis (Jaschov, 1955), and C. hyperboreus, are the most important herbivores in Arctic seas in terms of species biomass. They play a key role in the lipid-based energy flux in the Arctic, converting low-energy carbohydrates and proteins in ice algae and phytoplankton into high-energy wax esters. In this paper we review the over-wintering strategy, seasonal migration, stage development, life span, feeding strategy, body size, lipid biochemistry and the geographic distribution of the three dominant Calanus species in Arctic waters. We then relate these parameters to other biotic and abiotic factors, such as the timing of the Arctic phytoplankton and ice algae bloom, sea ice cover and climate variability. We also present new data on fatty acid and fatty alcohol content in the three Calanus species in addition to reviewing the available literature on these topics. These data are analysed for species homogeneity and geographic grouping. The dominance of diatom fatty acid trophic markers in the lipids of Calanus underpins the importance of diatoms as Arctic primary producers, even if dinoflagellates and Phaeocystis pouchetii can also be important food sources for the calanoid copepods. We conclude that the Arctic Calanus species are herbivores, engineered to feed on the Arctic bloom, and that the timing of the bloom is the most important factor in determining the life strategies of the individual species.

Rapeseed oil as an alternative to marine fish oil in diets of post-smolt Atlantic salmon (Salmo salar): changes in flesh fatty acid composition and effectiveness of subsequent fish oil “wash out”

Abstract Duplicate groups of Atlantic salmon post-smolts were fed five practical-type diets in which the added oil was 100% fish oil (FO)/0% rapeseed oil (0% RO), 90% FO/10% RO (10% RO), 75% FO/25% RO (25% RO), 50% FO/50% RO (50% RO) or 100% RO for 16 weeks. After sampling, the remaining fish were switched to a commercial grower diet containing FO as the only added lipid for a further 12 weeks. There were no effects of the inclusion of RO on growth or feed conversion. Fatty acid compositions of flesh total lipid showed significant increases in 18:1n-9, 18:2n-6 and 18:3n-3 with increasing inclusion of RO. The percentages of flesh 20:5n-3 and 22:6n-3 were significantly reduced in fish fed 10%, 25% and 50% RO, compared to fish fed 0% RO, and the percentages in fish fed 100% RO were significantly lower than all other treatments. Following transfer to the FO washout diet, percentages of flesh 20:5n-3 were not significantly different among treatments after 4 weeks while percentages of 22:6n-3 were restored after 12 weeks. However, even after 12 weeks, percentages of flesh 18:2n-6 were still significantly higher in fish previously fed 50% and 100% RO compared to the other treatments, although the final values were reduced by 48% and 65%, respectively, following the 12-week washout period. This study suggests that RO is a potential substitute for FO in Atlantic salmon culture, but that percentages of the n-3 highly unsaturated fatty acid (HUFA), 20:5n-3 and 22:6n-3, are significantly reduced by feeding RO above 50% of added oil. However, percentages of these two HUFA can be restored by feeding a diet containing FO for a period of 12 weeks.

Effects of purified diets containing different combinations of arachidonic and docosahexaenoic acid on survival, growth and fatty acid composition of juvenile turbot (Scophthalmus maximus)

The objective of this study was to determine the relative essential fatty acid (EFA) growth-promoting activities of pure arachidonic (AA, 20:4n−6) and docosahexaenoic (DHA, 22:6n−3) acids and various concentrations of these two acids in the diet of juvenile turbot (Scophthalmus maximus). Casein-based, semi-purified diets containing 15% fish oil or 14% hydrogenated coconut oil/oleic acid (1:1) supplemented with 1% 20:4n−6, 1% 22:6n−3 or 1% of various combinations of these two acids were fed to duplicate groups of 26 juvenile turbot for 11 weeks. In this trial, feeding the diet containing 20:4n−6 as the only highly unsaturated fatty acid (HUFA) resulted in higher growth and survival than any of the mixtures of the two fatty acids or 22:6n−3 alone. The diet containing 22:6n−3 as the sole HUFA resulted in the lowest growth and survival of all dietary treatments. The control diet with 15% fish oil resulted in a greater growth rate than any of the pure HUFA-supplemented diets. There was a significant effect of dietary lipid on the somatic index of the brain but not heart, kidney or liver. The percentage of lipid in the liver, but not of heart, brain, eyes, gills or kidney, was influenced by dietary lipid, with the highest percentage in fish supplemented with DHA alone. After 11 weeks, the 20:4n−6 and 22:6n−3 levels in whole-body total lipids were strongly influenced by the content of these fatty acids in the diets. The relative effect of dietary levels of these two fatty acids on their content in fish lipids varied considerably among the various organs and tissues of the fish that were analyzed. Brain and eye lipids were generally highest in 22:6n−3 while gill and kidney lipids were consistently higher in 20:4n−6 than the other organs analyzed. The effect of dietary 20:4n−6 on the content of that HUFA in organ lipid was greatest in gill and liver. The greatest impact of dietary 22:6n−3 level on content of that acid in organ lipid was seen in gill and kidney. There were also significant effects of dietary HUFA content on organ lipid levels of saturated, mono-unsaturated fatty acids and other members of the n−3 and n−6 PUFA, and HUFA series. The present study suggests that the EFA growth-promoting activity of arachidonic acid provides strong support for the contention that dietary 20:4n−6 is essential for juvenile turbot.

Replacement of dietary fish oil with increasing levels of linseed oil: Modification of flesh fatty acid compositions in Atlantic salmon (Salmo salar) using a fish oil finishing diet

Five groups of salmon, of initial mean weight 127±3 g, were fed increasing levels of dietary linseed oil (LO) in a regression design. The control diet contained capelin oil (FO) only, and the same oil was blended with LO to provide the experimental diets. After an initial period of 40 wk, all groups were switched to a finishing diet containing only FO for a further 24 wk. Growth and flesh lipid contents were not affected by dietary treatment. The FA compositions of flesh total lipids were linearly correlated with dietary FA compositions (r2=0.88–1.00, P<0.0001). LO included at 50% of added dietary lipids reduced flesh DHA and EPA (20∶5n−3) concentrations to 65 and 58%, respectively, of the concentrations in fish fed FO. Feeding 100% LO reduced flesh DHA and EPA concentrations to 38 and 30%, respectively, of the values in fish fed FO. Differences between diet and flesh FA concentrations showed that 16∶0, 18∶1n−9, and especially DHA were preferentially retained in flesh, whereas 18∶2n−6, 18∶3n−3, and 22∶1n−11 were selected against and presumably utilized for energy. In fish previously fed 50 and 100% LO, feeding a finishing diet containing FO for 16 wk restored flesh DHA and EPA concentrations, to ≈80% of the values in fish fed FO throughout. Flesh DHA and EPA concentrations in fish fed up to 50% LO were above recommended intake values for humans for these EFA. This study suggests that LO can be used as a substitute for FO in seawater salmon feeds and that any reductions in DHA and EPA can be largely overcome with a finishing diethigh in FO before harvest.