L.A. McEvoy

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.

Optimising lipid nutrition in first-feeding flatfish larvae

Although global production of flatfish has increased in recent years, both in terms of numbers of fish and diversification into new species, problems still remain with low survival rates and difficulties with metamorphosis. This short review highlights some advances made in optimising lipid nutrition in an attempt to overcome some of these problems. Copepod nauplii are the best live prey for first-feeding flatfish larvae. Rotifers can provide a useful method of essential fatty acid delivery but Artemia are poorer in this regard, especially at first feeding. Copepods are nutritionally beneficial due to their naturally high levels of the essential highly unsaturated fatty acids (HUFA), 20:5n-3 (eicosapentaenoic acid; EPA) and 22:6n-3 (docosahexaenoic acid; DHA), which are predominantly in the form of phospholipids. Rotifers can be enriched with fish oil emulsions to provide compositions similar to copepods, while enriched Artemia are difficult to enrich with high levels of DHA and the HUFA tend to be located in triglycerides rather than phospholipids. There is considerable evidence that the superior efficacy of copepods and rotifers is largely due to the digestibility and availability of HUFA supplied as pre-formed phospholipids. In addition to the essentiality of EPA and DHA, the requirement for 20:4n-6 (arachidonic acid; ARA) should also be considered. Improvements in dorsal pigmentation in turbot and halibut can be achieved by providing ratios of DHA/EPA of >2:1 but, perhaps more importantly, an EPA/ARA ratio of >5:1. This suggests that eicosanoids are involved in the control of pigmentation and this is further supported by the use of enrichments containing 18:3n-6, the elongation product of which (20:3n-6), is a potent inhibitor of ARA-derived eicosanoid formation, as is EPA.

Growth, survival, lipid composition and pigmentation of turbot (Scophthalmus maximus) larvae fed live-prey enriched in Arachidonic and Eicosapentaenoic acids

Abstract Turbot larvae were fed live-prey enriched with different levels of arachidonic (ARA) and eicosapentaenoic (EPA) acids to study the effects of these fatty acids on body composition and pigmentation success. Significantly reduced pigmentation was obtained in those fish fed medium and high ARA diets for 43 days. Growth and survival were the same for all groups. The incorporation of ARA and EPA in fish eyes, brains, livers and carcasses reflected the percentage of these fatty acids in the diets. ARA accumulation was similar in all tissues, but brain accumulated EPA was less efficient than the other tissues examined. A highly significant, negative correlation was found between the %ARA in turbot juvenile brain total lipids and pigmentation success. A weaker, positive correlation was found between brain EPA and pigmentation. Increasing dietary ARA affected the fatty acid composition of turbot brain phosphoglycerides more than increasing dietary EPA, especially in phosphatidylinositol (PI) and phosphatidylethanolamine (PE). A negative relationship was found between percentage normal pigmentation and ARA levels in brain phosphatidylcholine (PC), PE and phosphatidylserine (PS). Elevated levels of ARA in PI also resulted in malpigmented juveniles, but EPA:ARA ratios ≥1 in PI were associated with normal pigmentation. We conclude that, given a sufficiency of dietary docosahexaenoic acid (DHA), the optimum dietary level of EPA is not a function of DHA, but of dietary ARA.

Lipid and fatty acid composition of normal and malpigmented Atlantic halibut (Hippoglossus hippoglossus) fed enriched Artemia: a comparison with fry fed wild copepods

Atlantic halibut larvae originating from a single spawn of one female and one male were fed either Super Selco™ enriched, enrichment-grade Artemia nauplii or extensively grown zooplankton. The feeding experiment was conducted in 1.5-m3 outdoor tanks to evaluate if diet affected pigmentation. Pigmentation rates were found to be higher in the zooplankton-fed fish: 99.2% compared to 66.4% in the Artemia-fed halibut. Lipid and fatty acid analyses were performed on five malpigmented Artemia-fed fry, five normally pigmented Artemia-fed fry and 10 copepod-fed fry after 43 days feeding. Lipid class analysis was carried out on the eyes and carcasses of the halibut fry by HPTLC and densitometry. Fatty acid analyses of the same tissues were performed by gas liquid chromatography. Copepod-fed fry showed significantly higher levels of docosahexaenoic acid (22:6n−3; DHA) than their Artemia-fed counterparts (54.9% and 15.8% DHA in eye phosphatidylethanolamine (PE), respectively). Ratios of DHA: eicosapentaenoic acid (20:5n−3; EPA) were also higher in the copepod-fed fish: 11.1, 4.1 and 2.3 in, respectively, PE, phosphatidylcholine (PC) and triglyceride (TAG) in the eye lipid fractions of copepod-fed fry compared to 0.7, 0.3 and 0.1 in the same lipid of eyes of Artemia-fed fry. Within the Artemia-fed population, DHA and EPA levels were slightly, but significantly, raised in the eye PC fractions of normally pigmented fish compared to malpigmented ones. However, there was no significant difference in DHA:EPA ratios between normal and malpigmented fish fed Artemia. EPA: arachidonic acid (ARA) ratios were higher in zooplankton-fed fish but there was no significant difference in these ratios between normal and malpigmented Artemia-fed halibut.

Lipid conversions during enrichment of Artemia

Abstract Artemia nauplii were enriched for 24 h with radiolabelled fatty acid ethyl esters and then starved for a subsequent period of 24 h. Analyses of the distribution of radioactivity in lipids from samples taken at the end of the enrichment period and after the subsequent starvation showed that the ethyl esters were readily converted into other lipid classes, mainly triacylglycerols, during assimilation by the nauplii. The proportions of radioactivity recovered in free fatty acids and phospholipids increased during the starvation period indicating the mobilisation of fatty acids from triacylglycerols for use in catabolism and in the formation of biomembrane lipids. The distribution pattern of radioactivity from [U– 14 C ]22:6n−3 in the fatty acids of the nauplii demonstrates that Artemia are capable of retroconverting 22:6n−3 to 20:5n−3.

Two novel Anemia enrichment diets containing polar lipid

Abstract Two novel Anemia enrichment diets were tested against control diets of Super Selco (SS) (Artemia Systems, INVE, Ghent) and bakers yeast. The first experimental diet consisted of tuna orbital oil emulsified with 12% herring roe polar lipid (P-T). The second test diet comprised liposomes incorporating the phosphatidylcholine (PC) fraction of herring roe. Enrichment efficiency was measured in terms of lipid class and fatty acid composition of enriched nauplii, with particular attention being paid to PC content, docosahexaenoic acid (DHA) levels and DHA:eicosapentaenoic acid (EPA) ratios. The P-T emulsion yielded Artemia nauplii with 14% DHA (% total fatty acids) and a DHA:EPA ratio of 1.8, significantly higher than in nauplii fed the other three diets. Polar lipid levels in both P-T and SS enriched nauplii were similar (32.4 mg g−1 and 34.7 mg g−1 dry body weight (DBW) of nauplii, respectively). Lower %DHA and DHA:EPA ratios were obtained in liposome-fed nauplii (2% and 0.3, respectively) However, they yielded significantly higher levels of naupliar polar lipid (40.1 mg g−1 DBW) than those fed the two oil emulsion treatments. The significance of these findings is discussed.

Autoxidation of oil emulsions during the Artemia enrichment process

Abstract Autoxidation in three Artemia enrichment emulsions (cod liver oil, tuna orbital oil, and Super Selco (Artemia Systems N.V., Belgium)) were monitored over a 23-h period. Percentage concentrations of docosahexaenoic acid (DHA, 22:6n − 3), eicosapentaenoic acid (EPA, 20:5n-3), total polyun-saturated, monounsaturated and saturated fatty acids, as well as DHA:EPA ratios were measured in (a) the enriched Artemia nauplii (b) control enrichment media from which Artemia were absent and (c) enrichment media in which Artemia were present. Concentrations of thiobarbituric acid reactive substances (TBARS) were also assayed in the three emulsions. Polyunsaturated fatty acid levels in all the control oil emulsions remained steady over the 23-h enrichment period. However, there were significant relative decreases in DHA, EPA and total polyunsaturated fatty acids in all enrichment media containing Artemia , with a corresponding significant increase in total saturated and monounsaturated fatty acids in the final stages of the enrichment period. Furthermore, percentage and absolute concentrations of DHA and the DHA:EPA ratios in the enriched Artemia decreased significantly in the 23-h sample from the tuna orbital oil and Super Selco emulsions compared to their respective 18-h and 19-h samples. The significance of these results in view of the common aquacultural practice of employing Artemia enrichment periods of 24 h is discussed.

Effects of diet on fatty acid composition of body zones in larvae of the sea bass Dicentrarchus labrax: a chemometric study

Larvae of the sea bass Dicentrachus labrax were fed four Artemia sp. diets for 28 d. Three were nauplii enriched with emulsions of polyunsaturated fatty acids, and the fourth nauplii enriched with bakers yeast. At the end of the experimental period, the fatty acids of the bodies, heads and eyes of the larvae were analysed. A multivariate statistical method (discriminant analysis, DA) applied to the data revealed anatomical as well as dietary fatty acid pattern-discrimination. We propose here the use of discriminant analysis as a pattern-recognition method that will help to integrate the fatty acid information obtained in nutritional studies.