Dietary fish oil dose- and time-response effects on cardiac phospholipid fatty acid composition

Fish consumption is associated with reduced cardiovascular mortality, and elevated myocardial long-chain n−3 polyunsaturated FA (PUFA) content is implicated in this cardioprotection. This study examined the dose and time responses for incorporation of n−3 PUFA into cellular membranes in rats fed fish oil (FO)-containing diets. For the time course study, rats were fed a 10% FO diet for periods ranging from 0 to 42 d, after which myocardial and erythrocyte membrane fatty acid composition was determined. For the dose response study, rats (n=3) were fed 0, 1.25, 2.5, 5, or 10% FO for 4 wk, with myocardial, erythrocyte, and skeletal muscle membrane FA determined. Myocardial DHA (22∶6n−3) levels doubled in 2 d, stabilizing at levels ≈200% higher than control after 28 d feeding with 10% FO. By comparison, DHA levels doubled after 4 wk of 1.25% FO feeding. In myocardium and skeletal muscle, EPA (20∶5n−3) levels remained low, but in erythrocytes EPA levels reached 50% of DHA levels. The n−3 PUFA were incorporated at the expense of n−6 PUFA in myocardium and skeletal muscle, whereas erythrocytes maintained arachidonic acid levels, and total n−3 PUFA incorporation was lower. This study shows that low doses of FO produce marked changes in myocardial DHA levels; maximal incorporation takes up to 28 d to occur; and while erythrocytes are a good indicator of tissue n−3 incorporation in stable diets, they vary greatly in their time course and pattern of incorporation.     

Effect of struture and form on the ability of plant sterols to inhibit cholesterol absorption in hamsters

We investigated the effect of three types of plant sterols (4-desmethylsterols, 4,4′-dimethylsterols, and pentacyclic triterpene alcohols) in three forms (free, esterified with FA, or with phenolic acids) on cholesterol absorption. Plant sterol fractions derived from soybean (99% 4-desmethylsterols), rice bran (70% 4,4′-dimethylsterols), or shea nut (89% pentacyclic triterpene alcohols) were fed to male hamsters (n=20/group) as free sterols or esterified with FA or phenolic acids (cinnamic or ferulic). Cholesterol absorption was measured after 5–8.5 (mean, 7) wk by a dual-isotope technique. Soybean sterol intake significantly reduced cholesterol absorption efficiency (23%) and plasma total cholesterol (11%). Rice bran sterols tended to lower cholesterol absorption efficiency by 7% and plasma total cholesterol by 5%, whereas shea nut sterols had no effect. In hamsters, dietary 4-desmethylsterols were more effective than 4,4′-dimethylsterols in lowering cholesterol absorption and levels of cholesterol in blood. Pentacyclic triterpene alcohols had no effect on the absorption of cholesterol or on its level in blood. Esterification with FA did not impair the ability of 4-desmethylsterols and 4,4′-dimethylsterols to inhibit cholesterol absorption, whereas esterification with phenolic acids reduced this ability. This study supports the use of 4-desmethylsterols, esterified with FA to increase solubility, as the most effective cholesterol-lowering plant sterols in the diet.     

Supplementation and delivery of n−3 fatty acids through spray-dried milk reduce serum and liver lipids in rats

Indian diets comprising staples such as cereals, millets, and pulses provide 4.8 energy % from linoleic acid (18∶2n−6) but fail to deliver adequate amounts of n−3 FA. Consumption of long-chain n−3 PUFA such as EPA (20∶5n−3) and DHA (22∶6n−3) is restricted to those who consume fish. The majority of the Indian population, however, are vegetarians needing additional dietary sources of n−3 PUFA. The present work was designed to use n−3 FA-enriched spray-dired milk powder to provide n−3 FA. Whole milk was supplemented with linseed oil to provide α-linolenic acid (LNA, 18∶3n−3), with fish oil to provide EPA and DHA, or with groundnut oil (GNO), which is devoid of n−3 PUFA, and then spray-dired. Male Wistar rats were fed the spray-dired milk formulations for 60 d. The rats given formulations containing n−3 FA showed significant increases (P<0.001) in the levels of LNA or EPA/DHA in the serum and in tissue as compared with those fed the GNO control formulation. Rats fed formulations containing n−3 FA had 30–35% lower levels of serum total cholesterol and 25–30% lower levels of serum TAG than control animals. Total cholesterol and TAG in the livers of rats fed the formulations containing n−3 FA were lower by 18–30% and 11–18%, respectively, compared with control animals. This study showed that spray-dried milk formulations supplemented with n−3 FA are an effective means of improving dietary n−3 FA intake, which may decrease the risk factors associated with cardiovascular disease.     

Dietary α-linolenic acid lowers postprandial lipid levels with increase of eicosapentaenoic and docosahexaenoic acid contents in rat hepatic membrane

This study was designed to examine the effects of dietary n−3 and n−6 polyunsaturated fatty acids (PUFA) on postprandial lipid levels and fatty acid composition of hepatic membranes. Male Sprague-Dawley rats were trained for a 3−h feeding protocol and fed one of five semipurified diets: one fat-free diet or one of four diets supplemented with 10% (by weight) each of corn oil, beef tallow, perilla oil, and fish oil. Two separate experiments were performed, 4-wk long-term and 4-d short-term feeding models, to compare the effects of feeding periods. Postprandial plasma lipid was affected by dietary fats. Triacylglycerol (TG) and total cholesterol levels were decreased in rats fed perilla oil and fish oil diets compared with corn oil and beef tallow diets. Hepatic TG and total cholesterol levels were also reduced by fish oil and perilla oil diets. Fatty acid composition of hepatic microsomal fraction reflected dietary fatty acids and their metabolic conversion. The major fatty acids of rats fed the beef tallow diet were palmitic, stearic, and oleic. Similarly, linoleic acid (LA) and arachidonic acid in the corn oil group, α-linolenic acid (ALA) and eicosapentaenoic acid (EPA) in the perilla oil group, and palmitic acid and docosahexaenoic acid (DHA) in the fish oil group were detected in high proportions. Both long- and short-term feeding experiments showed similar results. In addition, microsomal DHA content was negatively correlated with plasma lipid levels. Hepatic lipid levels were also negatively correlated with EPA and DHA contents. These results suggest that n−3 ALA has more of a hypolipidemic effect than n−6 LA and that the hypolipidemic effect of n−3 PUFA may be partly related to the increase of EPA and DHA in hepatic membrane.     

Effects of a water-soluble phytostanol ester on plasma cholesterol levels and red blood cell fragility in hamsters

The aim of this study was to assess the efficacy of a novel water-soluble phytostanol anolog, disodium ascorbyl phytostanyl phosphate (DAPP), on plasma lipid levels and red blood cell fragility in hamsters fed atherogenic diets. For 4 wk, 50 male Golden Syrian hamsters were fed a semipurified diet without added cholesterol (noncholesterol, group 1), or a semipurified diet with 0.25% cholesterol (cholesterol-control, group 2). Groups 3–5 were fed the cholesterol-control diet with an addition of 1% phytostanols (diet 3), 0.71% DAPP (DAPP 0.7%, diet 4), or 1.43% DAPP (DAPP 1.4%, diet 5). Diets 4 and 5 provided 0.5 and 1% phytostanols, respectively. Supplementation of 0.71 and 1.43% DAPP decreased plasma total cholesterol concentrations by 34 (P<0.001) and 46% (P<0.001), respectively, in comparison with the cholesterol-control group, whereas free stanols reduced (P=0.007) plasma cholesterol concentrations by 14%. Similarly, non-HDL-cholesterol concentrations were reduced by 39 (P<0.001) and 54% (P<0.001) in hamsters supplemented with DAPP 0.7% and DAPP 1.4%, respectively, relative to the cholesterol-control group. The hypocholesterolemic effect of DAPP 1.4% was threefold stronger than that of free stanols. In hamsters supplemented with DAPP 1.4%, plasma TG concentrations were 45% lower (P=0.018) than in cholesterol-control-fed hamsters, whereas no such beneficial effect was observed in the free stanol group. Erythrocyte fragility was unaffected by DAPP or free phytostanols. Results of the current study demonstrate that DAPP lowers cholesterol more efficiently than free stanols, without an adverse effect on erythrocyte fragility in hamsters.     

Dietary menhaden, seal, and corn oils differentially affect lipid and Ex vivo eicosanoid and thiobarbituric acid-reactive substances generation in the guinea pig

This investigation was carried out to characterize the effects of specific dietary marine oils on tissue and plasma fatty acids and their capacity to generate metabolites (prostanoids, lipid peroxides). Young male guinea pigs were fed nonpurified diet (NP), or NP supplemented (10%, w/w) with menhaden fish oil (MO), harp seal oil (SLO), or corn oil (CO, control diet) for 23 to 28 d. Only the plasma showed significant n−3 polyunsaturated fatty acid (PUFA)-induced reductions in triacylglycerol (TAG) or total cholesterol concentration. Proportions of total n−3 PUFA in organs and plasma were elevated significantly in both MO and SLO dietary groups (relative to CO), and in all TAG fractions levels were significantly higher in MO-than SLO-fed animals. The two marine oil groups differed in their patterns of incorporation of eicosapentaenoic acid (EPA). In guinea pigs fed MO, the highest levels of EPA were in the plasma TAG, whereas in SLO-fed animals, maximal incorporation of EPA was in the heart polar lipids (PL). In both marine oil groups, the greatest increases in both docosahexaenoic acid (22∶6n−3, DHA) and docosapentaenoic acid (22∶5n−3, DPA) relative to the CO group, were in plasma TAG, although the highest proportions of DHA and DPA were in liver PL and heart TAG, respectively. In comparing the MO and SLO groups, the greatest difference in levels of DHA was in heart TAG (MO>SLO, P<0.005), and in levels of DPA was in heart PL (SLO>MO, P<0.0001). The only significant reduction in proportions of the major n−6 PUFA, arachidonic acid (AA), was in the heart PL of the SLO group (SLO>MO=CO, P<0.005). Marine oil feeding altered ex vivo generation of several prostanoid metabolites of AA, significantly decreasing thromboxane A₂ synthesis in homogenates of hearts and livers of guinea pigs fed MO and SLO, respectively (P<0.04 for both, relative to CO). Lipid peroxides were elevated to similar levels in MO- and SLO-fed animals in plasma, liver, and adipose tissue, but not in heart preparations. This study has shown that guinea pigs respond to dietary marine oils with increased organ and plasma n−3 PUFA, and changes in potential synthesis of metabolites. They also appear to respond to n−3 PUFA-enriched diets in a manner that is different from that of rats.     

Specific modifications of phosphatidylinositol and nonesterified fatty acid fractions in cultured porcine cardiomyocytes supplemented with n-3 polyunsaturated fatty acids

Mechanisms for the antiarrhythmic effect of n−3 polyunsaturated fatty acids (PUFA) are currently being investigated using isolated cardiac myocytes. It is still not known whether the incorporation of n−3 PUFA into membrane phospholipids is a prerequisite for its protective action or if n−3 PUFA exert antiarrhythmic effects in their nonesterified form as demonstrated by recent studies. Adult porcine cardiomyocytes were grown in media supplemented with arachidonic acid (AA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA). After 24 h, analysis of total lipids showed that the myocytes were enriched with the respective fatty acids compared to control cells. Large proportions of all three fatty acids supplemented (69% AA, 72% DHA, and 66% EPA) remained unesterified. Fatty acid analysis of total phospholipids (PL) revealed that the incorporation of EPA and DHA, though small, was significantly different (P<0.05) from that of the control cells. The PL fraction was further separated into phosphatidylinositol (Pl), phosphatidylethanolamine, phosphatidylcholine, and phosphatidylserine to study the pattern of incorporation of the fatty acids in these fractions. It became apparent that EPA and DHA were selectively incorporated into the Pl fraction. This study demonstrates that in adult porcine cardiomyocytes, the n−3 PUFA supplementation selectively modulates two important lipid fractions, nonesterified fatty acid and Pl, which were implicated in the mechanisms of prevention of cardiac arrhythmias.     

Incorporation of n-3 fatty acids into plasma lipid fractions, and erythrocyte membranes and platelets during dietary supplementation with fish, fish oil, and docosahexaenoic acid-rich oil among healthy young men

The effects of n-3 fatty acid supplementation in the form of fresh fish, fish oil, and docosahexaenoic acid (DHA) oil on the fatty acid composition of plasma lipid fractions, and platelets and erythrocyte membranes of young healthy male students were examined. Altogether 59 subjects (aged 19–32 yr, body mass index 16.8–31.3 kg/m²) were randomized into the following diet groups: (i) control group; (ii) fish diet group eating fish meals five times per week [0.38±0.04 g eicosapentaenoic acid (EPA) and 0.67±0.09 g DHA per day]; (iii) DHA oil group taking algae-derived DHA oil capsules (1.68 g/d DHA oil group taking algae-derived DHA oil capsules (1.68 g/d DHA in triglyceride form); and (iv) fish oil group (1.33 g EPA and 0.95 g DHA/d as free fatty acids) for 14 wk. The fatty acid composition of plasma lipids, platelets, and erythrocyte membranes was analyzed by gas chromatography. The subjects kept 4-d food records four times during the study to estimate the intake of nutrients. In the fish diet, in DHA oil, and in fish oil groups, the amounts of n-3 fatty acids increased and those of n-6 fatty acids decreased significantly in plasma lipid fractions and in platelets and erythrocyte membranes. A positive relationship was shown between the total n-3 polyunsaturated fatty acids (PUFA) and EPA and DHA intake and the increase in total n-3 PUFA and EPA and DHA in all lipid fractions analyzed. DHA was preferentially incorporated into phospholipid (PL) and triglyceride (TG) and there was very little uptake in cholesterol ester (CE), while EPA was preferentially incorporated into PL and CE. The proportion of EPA in plasma lipids and platelets and erythrocyte membranes increased also by DHA supplementation, and the proportion of linoleic acid increased in platelets and erythrocyte membranes in the DHA oil group as well. These results suggest retroconversion of DHA to EPA and that DHA also interferes with linoleic acid metabolism.     

Accretion of Dietary Docosahexaenoic Acid in Mouse Tissues Did Not Differ between Its Purified Phospholipid and Triacylglycerol Forms

Recent studies suggest that dietary krill oil leads to higher omega-3 polyunsaturated fatty acids (n-3 PUFA) tissue accretion compared to fish oil because the former is rich in n-3 PUFA esterified as phospholipids (PL), while n-3 PUFA in fish oil are primarily esterified as triacylglycerols (TAG). Tissue accretion of the same dietary concentrations of PL- and TAG-docosahexaenoic acid (22:6n-3) (DHA) has not been compared and was the focus of this study. Mice (n = 12/group) were fed either a control diet or one of six DHA (1%, 2%, or 4%) as PL-DHA or TAG-DHA diets for 4 weeks. Compared with the control, DHA concentration in liver, adipose tissue (AT), heart, and eye, but not brain, were significantly higher in mice consuming either PL- or TAG-DHA, but there was no difference in DHA concentration in all tissues between the PL- or TAG-DHA forms. Consumption of PL- and TAG-DHA at all concentrations significantly elevated eicosapentaenoic acid (20:5n-3) (EPA) in all tissues when compared with the control group, while docoshexapentaenoic acid (22:5n-6) (DPA) was significantly higher in all tissues except for the eye and heart. Both DHA forms lowered total omega-6 polyunsaturated fatty acids (n-6 PUFA) in all tissues and total monounsaturated fatty acids (MUFA) in the liver and AT; total saturated fatty acid (SFA) were lowered in the liver but elevated in the AT. An increase in the DHA dose, independent of DHA forms, significantly lowered n-6 PUFA and significantly elevated n-3 PUFA concentration in all tissues. Our results do not support the claim that the PL form of n-3 PUFA leads to higher n-3 PUFA tissue accretion than their TAG form.     

Lipase-catalyzed incorporation of n−3 PUFA into palm oil

Two immobilized lipases, IM60 from Rhizomucor miehei and QLM from Alcaligenes sp., were used as biocatalysts for the modification of the FA composition of palm oil by incorporating n−3 PUFA. Acidolysis and interesterification reactions were conducted with hexane as organic solvent, and the products were analyzed by using GLC. After a 24-h incubation in hexane, there was an average incorporation of 20.8% EPA and 15.6% DHA into palm oil, respectively, while the percentages of palmitic and oleic acids in palm oil decreased by 28.8 and 11.8%, respectively. Higher EPA and DHA incorporation was obtained when EPAX (fish oil concentrate high in n−3 PUFA) was used in the ethyl ester form (interesterification reaction) than in the free acid form (acidolysis) in the presence of Lipozyme (IM60 lipase. Lipase QLM was found to discriminate against EPA, and it showed slightly better catalytic activity for DHA in the free acid form than in the ethyl ester form. Generally, as the mole ratio of the acyl donor to TAG increased, the percentage incorporation of EPA and DHA increased; however, reactions catalyzed by Lipozyme IM60 did not show increases in the incorporation beyond a TAG/EPAX mole ratio of 3. When limitations due to mass transfer were not a factor, an increase in the reactant amount also gave an increase in the percentage incorporation of the n−3 PUFA. Palm oil containing EPA and DHA was successfully produced and may be beneficial in certain food and nutritional applications.     

Separation of eicosapentaenoic acid and docosahexaenoic acid in fish oil by kinetic resolution using lipase

The objective of this study was to investigate the use of lipases as catalysts for separating eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in fish oil by kinetic resolution. Transesterification of various fish oil triglycerides with a stoichiometric amount of ethanol by immobilized Rhizomucor miehei lipase under anhydrous solvent-free conditions resulted in a good separation. When free fatty acids from the various fish oils were directly esterified with ethanol under similar conditions, greatly improved results were obtained. By this modification, complications related to regioselectivity of the lipase and nonhomogeneous distribution of EPA and DHA into the various positions of the triglycerides were avoided. As an example, when tuna oil comprising 6% EPA and 23% DHA was transesterified with ethanol, 65% conversion into ethyl esters was obtained after 24 h. The residual glyceride mixture contained 49% DHA and 6% EPA (8:1), with 90% DHA recovery into the glyceride mixture and 60% EPA recovery into the ethyl ester product. When the corresponding tuna oil free fatty acids were directly esterified with ethanol, 68% conversion was obtained after only 8h. The residual free fatty acids comprised 74% DHA and only 3% EPA (25:1). The recovery of both DHA into the residual free fatty acid fraction and EPA into the ethyl ester product remained very high, 83 and 87%, respectively.     

The content and composition of sterols and sterol esters in low erucic acid rapeseed (Brassica napus)

The low temperature crystallization technique for the enrichment of “minor” components, such as sterols and sterol esters, from vegetable oils was applied to low erucic acid rapeseed oils. The recovery of free sterols and sterol esters was estimated by use of¹⁴C-cholesterol and¹⁴C-cholesterol oleate. 80% of the free sterols and 45% of the sterol esters were recovered in the liquid fraction, while in two studies total recoveries were 95% and 99%, respectively. This technique showed some selectivity toward the sterol bound fatty acids when compared to direct preparative thin layer chromatography (TLC) of the crude oil. Gas liquid chromatography (GLC) analysis of the free and esterified sterols as TMS-derivatives showed very little selectivity in the enrichment procedure. The fatty acid patterns of the sterol esters demonstrated, however, a preference in the liquid fraction for those sterol esters which have a high linoleic and linolenic acid content. The content of free sterols was 0.3–0.4% and that of sterol esters 0.7–1.2% of the rapeseed oils in both winter and summer types of low erucic acid rapeseed (Brassica napus) when the lipid classes were isolated by direct preparative TLC of the oils. The free sterols in the seven cultivars or breeding lines analyzed were composed of 44–55% sitosterol, 27–36% campesterol, 17–21% brassicasterol, and a trace of cholesterol. The esterified sterols were 47–57% sitosterol, 36–44% campesterol, 6–9% brassicasterol, and traces of cholesterol and Δ5-avenasterol. The fatty acid patterns of these esters were characterized by ca. 30% oleic acid and ca. 50% linoleic acid, whereas these acids constitute 60% and 20%, respectively, of the total fatty acids in the oil. Little or no variation in sterol and sterol ester patterns with locality within Sweden was observed for the one cultivar of summer rapeseed investigated by the low temperature crystallization technique.     

The content and composition of sterols and sterol esters in sunflower and poppy seed oils

The composition and proportion of free sterols and sterol esters in crude sunflower and poppy seed oils were determined, using preparative thin layer chromatography followed by gas chromatography with cholesterol as an internal standard. Free sterols and sterol esters were also isolated in a liquid fraction obtained by low temperature crystallization (−80 C) of the oils and enriched with minor lipid classes. This enrichment procedure provided a liquid fraction suitable for studies of minor components in the oils. However, selectivity towards sterol esters was observed since sterols esterified to very long chain fatty acids (C₂₀–C₂₄) were preferentially retained in the precipitate. The proportions of free and esterified sterols were found to be 0.34 and 0.28%, respectively, in the sunflower oil, whereas the corresponding figures for poppy seed oil were 0.33% and 0.05%. Sunflower oil was characterized by a relatively high percentage of Δ7-sterols, preferentially obtained in the esterified fraction, and by very long chain saturated fatty acids of sterol esters. The sterols in poppy seed oil were composed almost entirely of campesterol, stigmasterol, sitosterol and Δ5-avenasterol, although their percentage distributions were remarkably different in the free and esterified fraction.     

Dietary manipulation of long-chain polyunsaturated fatty acids in the retina and brain of guinea pigs

High levels of n−6 docosapentaenoic acid (22∶5n−6) have been reported in the retina of guinea pigs fed commercially-prepared grain-based rations (commercial diet). In rats and monkeys, high levels of 22∶5n−6 are an indicator of n−3 polyunsaturated fatty acid (PUFA) deficiency. We have examined the fatty acid composition of the retina and brain in guinea pigs fed a commercial diet or one of three semi-purified diets containing three different levels of n−3 PUFA. The diets comprised a diet deficient in n−3 PUFA (semi-purified diet containing safflower oil), two diets containing α-linolenic acid (standard commercial laboratory diet and semi-purified diet containing canola oil), and a diet containing α-linolenic acid, eicosapentaenoic acid, and docosahexaenoic acid (DHA) (semi-purified diet containing canola oil, safflower oil, and fish oil). Two groups of guinea pigs were given the diets from day 1 to 4 wk or day 1 to 8 wk, when they were sacrificed and the retinal tissues were extracted and analyzed for PUFA content by gas-liquid chromatography. Fatty acid analyses of the retinal phospholipids of the four-week-old animals revealed that the group fed DHA (from the fish oil) had the highest level of DHA (32%), compared with values of 19 and 13% for the groups fed canola oil diet and commercial diet, respectively, and 2% for the group fed the diet deficient in n−3 PUFA. The levels of 22∶5n−6 in the retinal lipids were inversely related to the DHA values, being 0.6, 6.6, 11.4, and 20.6 for the fish oil, canola oil, commercial diet, and safflower oil diet groups, respectively. The long-chain PUFA profiles in the brain phospholipids of the four-week-old group were similar to those from the retina. The retinal PUFA values for the eight-week-old animals were similar to the four-week-old group. The safflower oil diet induced a greater deficit of DHA in retinal lipids than has been reported in rats and monkeys fed similar diets. The guinea pigs fed the commercial diet had retinal and brain PUFA patterns similar to that produced by n−3 PUFA-deficient diets in rats and monkeys. Guinea pigs fed the canola oil diet had significantly greater retinal DHA levels than those fed the commercial diet, but lower than those fed fish oil. The data suggest that the guinea pig has a reduced capacity for DHA synthesis from α-linolenic acid as compared with other mammals. Supplementation of guinea pig diets with fish oil produced high retinal and brain DHA levels and prevented the accumulation of 22∶5n−6.     

Different kinetic in incorporation and depletion of n−3 fatty acids in erythrocytes and leukocytes of mice

n−3 PUFA are well known for their anti-inflammatory effects. However, there has been only limited study on the kinetics of incorporation and depletion of n−3 PUFA in immune cells. In the present study we investigated the incorporation and depletion of n−3 PUFA in erythrocytes and leukocytes in mice during a 6-wk feeding period. Over the first 3-wk period (the incorporation period) the mice were fed a special diet with a high n−3/n−6 PUFA ratio. In the following 3-wk period (the depletion period) the mice were fed a standard chow diet. A linear incease of the concentration of EPA and DHA in erythrocyte membranes was observed during the incorporation period, whereas a stagnation was observed after the second week for leukocytes. The level of EPA did not fall to the background level after the depletion period, and the level of DHA was kept almost constant during the depletion period in the erythrocyte membranes. In leukocytes the concentration of both EPA and DHA decreased during the depletion period, but did not reach the background level after the 3-wk depletion. In conclusion, the kinetics of EPA and DHA in the different cells are different. The rate of incorporation is faster than that of depletion for n−3 PUFA. More n−3 PUFA can be incorporated into leukocytes in comparison with erythrocytes. The ratio of n−3/n−6 PUFA is more important than the amount of n−3 FA in changing the FA compositions of membrane lipids.     

Dietary intake of fish vs. formulations leads to higher plasma concentrations of n−3 fatty acids

The n−3 fatty acids from fish appear to be more efficacious, in terms of cardioprotection, than equivalent amounts provided as capsules. Volunteers were given, for 6 wk, either 100 g/d of salmon, providing 383 mg of EPA and 544 mg of DHA, esterified in glycerol lipids, or 1 or 3 capsules of fish oil/d, providing 150 mg of EPA and 106 mg of DHA or 450 mg of EPA and 318 mg of DHA, as ethyl esters. Further, we reevaluated data from a previous study carried out with the same design, i.e., with 3 and 6 capsules/d of fish oil, providing 1290 and 2580 mg/d EPA and 960 and 1920 mg/d DHA. Marked increments in plasma EPA and DHA concentrations (μg/mg total lipid) and percentages of total fatty acids were recorded at the end of treatment with either n−3 capsules or salmon. Net increments of EPA and DHA in plasma lipids were linearly and significantly correlated with the dose after capsule administration. Further, increments in plasma EPA and DHA concentration after salmon intake were significantly higher than after administration of capsules. The same increments would be obtained with at least two- and ninefold higher doses of EPA and DHA, respectively, if administered with capsules rather than salmon. We provide experimental evidence that n−3 fatty acids from fish are more effectively incorporated into plasma lipids than when administered as capsules and that increments in plasma concentrations of EPA and DHA given as capsules are linearly correlated with their intakes.     

Analysis of EPA and DHA and distinction between fish oils and concentrates

Fish oil is the major dietary source of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). Adequate dietary intake of these n‐3 PUFA is beneficial to reduce the risks of cardiovascular mortality, prostate cancer, and neurological disorders in children and adults. There is a surge in demand for fish oil in the functional food market. Microencapsulation of fish oil is the trend to improve its stability and sensory quality. The EPA and DHA content of the fish oil products may vary markedly from the label due to their susceptibility to oxidation. Quick and reliable methods other than the AOAC BF₃ method have been exploited to quantify EPA and DHA in the encapsulated fish oil and the microencapsulated powdered products such as infant formula. This article describes a method to differentiate the ethyl‐ester form from the TAG form of EPA and DHA in encapsulated fish oil which may be a mixture of natural triacylglycerol enriched with ethyl esterified EPA and DHA. A method is recommended due to the difference in apparent potency of these two esterified forms, which may be a concern in infant formula and elsewhere.     

Fractionation of urea-pretreated squid visceral oil ethyl esters

Ethyl esters of squid (Illex argentinus) visceral oil contained 11.8% eicosapentaenoic acid (EPA) and 14.9% docosahexaenoic acid (DHA). The esters were treated with urea to increase the contents of EPA and DHA. The non-urea complexing ethyl esters of squid visceral oil contained 28.2% EPA and 35.6% DHA. This mixture was fractionated by molecular distillation to further increase the EPA or DHA content. The fraction collected in the 110°C distillate had an EPA content of 39.0% with 0.26 g/100 g of cholesterol, while the 130°C distillate contained 65.6% DHA and 0.42 g/100 g of cholesterol. Ethyl esters prepared from visceral oil of squid Ommastrephes bartrami had 4.5% EPA and 12.7% DHA. After urea pretreatment, the EPA and DHA contents were raised to 10.1 and 30.0%, respectively. When this mixture was further fractionated by molecular distillation, 16.9% EPA with 0.35 g/100 g cholesterol was found in the 110°C distillate and 52.6% DHA with 0.70 g/100 g cholesterol was found in the 130°C distillate. Cholesterol in the squid visceral oil ethyl esters was concentrated in the final residue of molecular distillation when the polyunsaturated ethyl esters were enriched by the urea complexation method prior to molecular distillation. For example, the cholesterol content in the ethyl esters from O. bartrami squid visceral oil was 2.28 g/100 g originally. It was enriched to 64.15 g/100 g in the final residue from the molecular distillation.     

Replacement of dietary fish oil with increasing levels of linseed oil: Modification of flesh fatty acid compositions in Atlantic salmon (<Emphasis Type="Italic">Salmo salar</Emphasis>) 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 (r ²=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.     

Lipase selectivity toward fatty acids commonly found in fish oil

The main objective of this study was to compare the fatty acid selectivity of numerous commercially available lipases toward the most ubiquitous fatty acids present in fish oils in form of their corresponding ethyl esters. Special interest was taken in their ability to separate the n‐3 long‐chain polyunsaturated fatty acids (PUFA), mainly eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), from the more saturated fatty acids as well as exploiting the putative discrimination between these highly valuable n‐3 PUFA. Hydrolysis of sardine oil ethyl esters in a Tris buffer solution by 12 microbial lipases is described. The results reveal that all of the lipases strongly discriminate against the n‐3 PUFA and prefer the more saturated fatty acids as substrates. Most of the lipases discriminate between EPA and DHA in favor of EPA, however, 2 bacterial lipases from Pseudomonas were observed to prefer DHA to EPA. Digestive lipolytic enzymes isolated from salmon and rainbow trout intestines displayed reversed fatty acid selectivity when their fish oil triacylglycerol hydrolysis was studied. Thus, the n‐3 PUFA including EPA and DHA were observed to be hydrolyzed at a considerably higher rate than the more saturated fatty acids.