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.     

Higher Increase in Plasma DHA in Females Compared to Males Following EPA Supplementation May Be Influenced by a Polymorphism in ELOVL2: An Exploratory Study

Young adult females have higher blood docosahexaenoic acid (DHA), 22:6n-3 levels than males, and this is believed to be due to higher DHA synthesis rates, although DHA may also accumulate due to a longer half-life or a combination of both. However, sex differences in blood fatty acid responses to eicosapentaenoic acid (EPA), 20:5n-3 or DHA supplementation have not been fully investigated. In this exploratory analysis, females and males (n = 14–15 per group) were supplemented with 3 g/day EPA, 3 g/day DHA, or olive oil control for 12 weeks. Plasma was analyzed for sex effects at baseline and changes following 12 weeks' supplementation for fatty acid levels and carbon-13 signature (δ¹³C). Following EPA supplementation, the increase in plasma DHA in females (+23.8 ± 11.8, nmol/mL ± SEM) was higher than males (−13.8 ± 9.2, p < 0.01). The increase in plasma δ¹³C-DHA of females (+2.79 ± 0.31, milliUrey (mUr ± SEM) compared with males (+1.88 ± 0.44) did not reach statistical significance (p = 0.10). The sex effect appears driven largely by increased plasma DHA in the AA genotype of females (+58.8 ± 11.5, nmol/mL ± SEM, n = 5) compared to GA + GG in females (+4.34 ± 13.5, n = 9) and AA in males (−29.1 ± 17.2, n = 6) for rs953413 in the ELOVL2 gene (p < 0.001). In conclusion, EPA supplementation increases plasma DHA levels in females compared to males, which may be dependent on the AA genotype for rs953413 in ELOVL2.     

EPA,but not DHA,decreases mean platelet volume in normal subjects

The first indication of platelet activation is an increase in mean platelet volume (MPV). n−3 FA are known to inhibit platelet function and to reduce the risk for coronary heart disease. The purpose of this study was to determine the effects of FPA and DHA on MPV. Healthy subjects received olive oil placebo for 4 wk and then were randomly assigned to receive 4g of ethyl esters of either safflower oil (n=11), EPA (n=10), or DHA (n=12) for 4 wk. At the end of placebo run-in and treatment periods, MPV (fL; mean±SEM) and platelet count (PLT-CT; 10³/μL blood) were measured in the basal state and after ex vivo stimulation with collagen (10 μg/mL), cold (4°C), and heat (37°C). Unlike DHA, EPA lowered MPV as compared with safflower oil (7.2±0.1 vs. 7.5±0.1 fL; P<0.05) and raised PLT-CT (211±18 vs. 192±18 10³/μL; P<0.05) in the fasting state. Collagen and cold significantly increased MPV whereas heat lowered MPV regardless of treatment. All stimuli decreased PLT-CT. EPA significantly increased platelet EPA (0.2±0.1 vs. 3.3±0.4%) and docosapentaenoic acid (DPA; 2.2±0.3 vs. 2.9±0.3%) concentrations, but not DHA. DHA treatment significantly increased DHA (1.4±0.2 vs. 4.1±0.5%) and DPA (2.0±0.4 vs. 3.0±0.4%) concentrations, but not EPA. In conclusion, EPA, but not DHA, reduces platelet activation, an early step in platelet aggregation.     

Purification of docosahexaenoic acid from tuna oil by a two-step enzymatic method: Hydrolysis and selective esterification

Purification of docosahexaenoic acid (DHA) was attempted by a two-step enzymatic method that consisted of hydrolysis of tuna oil and selective esterification of the resulting free fatty acids (FFA). When more than 60% of tuna oil was hydrolyzed with Pseudomonas sp. lipase (Lipase-AK), the DHA content in the FFA fraction coincided with its content in the original tuna oil. This lipase showed stronger activity on the DHA ester than on the eicosapentaenoic acid ester and was suitable for preparation of FFA rich in DHA. When a mixture of 2.5 g tuna oil, 2.5 g water, and 500 units (U) of Lipase-AK per 1 g of the reaction mixture was stirred at 40°C for 48 h, 83% of DHA in tuna oil was recovered in the FFA fraction at 79% hydrolysis. These fatty acids were named tuna-FFA-Ps. Selective esterification was then conducted at 30°C for 20 h by stirring a mixture of 4.0 g of tuna-FFA-Ps/lauryl alcohol (1:2, mol/mol), 1.0 g water, and 1,000 U of Rhizopus delemar lipase. As a result, the DHA content in the unesterified FFA fraction could be raised from 24 to 72 wt% in an 83% yield. To elevate the DHA content further, the FFA were extracted from the reaction mixture with n-hexane and esterified again under the same conditions. The DHA content was raised to 91 wt% in 88% yield by the repeated esterification. Because selective esterification of fatty acids with lauryl alcohol proceeded most efficiently in a mixture that contained 20% water, simultaneous reactions during the esterification were analyzed qualitatively. The fatty acid lauryl esters (L-FA) generated by the esterification were not hydrolyzed. In addition, L-FA were acidolyzed with linoleic acid, but not with DHA. These results suggest that lauryl DHA was generated only by esterification.     

n‐3 PUFA Induce Microvascular Protective Changes During Ischemia/Reperfusion

Ischemia/reperfusion (I/R) injury can occur in consequence of myocardial infarction, stroke and multiple organ failure, the most prevalent cause of death in critically ill patients. I/R injury encompass impairment of endothelial dependent relaxation, increase in macromolecular permeability and leukocyte‐endothelium interactions. Polyunsaturated fatty acids (n‐3 PUFA), such as eicosapentaenoic acid (EPA, 20:5n‐3) and docosahexaenoic acid (DHA, 22:6n‐3) found in fish oil have several anti‐inflammatory properties and their potential benefits against I/R injury were investigated using the hamster cheek pouch preparation before and after ischemia. Before the experiments, hamsters were treated orally with saline, olive oil, fish oil and triacylglycerol (TAG) and ethyl ester (EE) forms of EPA and DHA at different daily doses for 14 days. Fish oil restored the arteriolar diameter to pre ischemic values during reperfusion. At onset and during reperfusion, Fish oil and DHA TAG significantly reduced the number of rolling leukocytes compared to saline and olive oil treatments. Fish oil, EPA TAG and DHA TAG significantly prevented the rise on leukocyte adhesion compared to saline. Fish oil (44.83 ± 3.02 leaks/cm²), EPA TAG (31.67 ± 2.65 leaks/cm²), DHA TAG (41.14 ± 3.63 leaks/cm²), and EPA EE (30.63 ± 2.25 leaks/cm²), but not DHA EE (73.17 ± 2.82 leaks/cm²) prevented the increase in macromolecular permeability compared to saline and olive oil (134.80 ± 1.49 and 121.00 ± 4.93 leaks/cm², respectively). On the basis of our findings, we may conclude that consumption of n‐3 polyunsaturated fatty acids, especially in the triacylglycerol form, could be a promising therapy to prevent microvascular damage induced by ischemia/reperfusion and its consequent clinical sequelae.     

Docosahexaenoic Acid Supplementation Promotes Erythrocyte Antioxidant Defense and Reduces Protein Nitrosative Damage in Male Athletes

The aim of this study was to determine the influence of long‐term docosahexaenoic acid (DHA) dietary supplementation on the erythrocyte fatty acid profile and oxidative balance in soccer players after training and acute exercise. Fifteen volunteer male athletes (age 20.0 ± 0.5 years) were randomly assigned to a placebo group that consumed an almond‐based beverage (n = 6), or to an experimental group that consumed the same beverage enriched with DHA (n = 9) for 8 weeks. Blood samples were taken in resting conditions at the beginning and after 8 weeks of nutritional intervention and training in resting and in post‐exercise conditions. Oxidative damage markers (malonyldialdehyde, carbonyl and nitrotyrosine indexes) and the activity and protein level of antioxidant enzymes (catalase, superoxide dismutase, glutathione reductase and peroxidase) were assessed. The results showed that training increased antioxidant enzyme activities in erythrocytes. The experimental beverage increased DHA from 34.0 ± 3.6 to 43.0 ± 3.6 nmol/10⁹ erythrocytes. DHA supplementation increased the catalytic activity of superoxide dismutase from 1.48 ± 0.40 to 10.5 ± 0.35 pkat/10⁹ erythrocytes, and brought about a reduction in peroxidative damage induced by training or exercise. In conclusion, dietary supplementation with DHA changed the erythrocyte membrane composition, provided antioxidant defense and reduced protein peroxidative damage in the red blood cells of professional athletes after an 8‐week training season and acute exercise.     

Formation of genotoxic dicarbonyl compounds in dietary oils upon oxidation

Dietary oils—tuna, salmon, cod liver, soybean, olive, and corn oils—were treated with accelerated storage conditions (60°C for 3 and 7 d) and a cooking condition (200°C for 1 h). Genotoxic malonaldehyde (MA), glyoxal, and methylglyoxal formed in the oils were analyzed by GC. Salmon oil produced the greatest amount of MA (1070±77.0 ppm of oil) when it was heated at 60°C for 7 d. The highest formation of glyoxal was obtained from salmon oil heated at 60°C for 3 d. More glyoxal was found from salmon and cod liver oils when they were heated for 3 d (12.8±1.10 and 7.07±0.19 ppm, respectively) than for 7d (6.70±0.08 and 5.94±0.38 ppm, respectively), suggesting that glyoxal underwent secondary reactions during a prolonged time. The amount of methyglyoxal formed ranged from 2.03±0.13 (cod liver oil) to 2.89±0.11 ppm (tuna oil) in the fish oils heated at 60°C for 7 d. Among vegetable oils, only olive oil yielded methylglyoxal (0.61±0.03 ppm) under accelerated storage conditions. When oils were treated under cooking conditions, the aldehydes formed were comparable to those formed under accelerated storage conditions. Fish oils produced more MA, glyoxal, and methylglyoxal than did vegetable oils because the fish oils contained higher levels of long-chain PUFA, such as EPA and DHA, than did the vegetable oils. A statistically significant correlation (P<0.05) between the α-tocopherol content and the oxidation parameters was obtained from only MA and fish oils heated at 60°C for 3 d.     

Fish Oil Supplementation During Lactation: Effects on Cognition and Behavior at 7 Years of Age

Early accumulation of n-3 long-chain PUFA (LCPUFA) in the brain may contribute to differences in later cognitive abilities. In this study, our objective was to examine whether fish oil (FO) supplementation during lactation affects processing speed, working memory, inhibitory control, and socioemotional development at 7 years. Danish mothers (n = 122) were randomized to FO [1.5 g/d n-3 LCPUFA] or olive oil (OO) supplementation during the first 4 months of lactation. The trial also included a high-fish intake (HFI) reference group (n = 53). Ninety-eight children were followed-up with an assessment of processing speed, an age-appropriate Stroop task, and the Strength and Difficulties Questionnaire at 7 year. A group effect of the intervention (FO vs. OO) was found in prosocial behavior scores; this negative effect was carried by the boys. Exploratory analyses including all participants revealed the speed of processing scores were predicted by maternal n-3 LCPUFA intake during the intervention period (negative relation) and maternal education (positive relation). Stroop scores indicative of working memory and inhibitory control were predicted by infant erythrocyte DHA status at 4 months of age (negative relation). Early fish oil supplementation may have a negative effect on later cognitive abilities. Speed of processing and inhibitory control/working memory are differentially affected, with speed of processing showing effects of fish oil intake as a whole, whereas inhibitory control/working memory was related more specifically to DHA status.     

Lipid peroxidation during n−3 fatty acid and vitamin E supplementation in humans

The purpose of this study was to investigate in healthy humans the effect of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) intake, alone or in combination with dL-α-tocopherol acetate (vitamin E) supplements on lipid peroxidation. Eightly men were randomly assigned in a double-blind fashion to take daily for 6 wk either menhaden oil (6.26 g, n−3 fatty acids) or olive oil supplements with either vitamin E (900 IU) or its placebo. Antioxidant vitamins, phospholipid composition, malondialdehyde (MDA), and lipid peroxides were measured in the plasma at baseline and week 6. At the same time, breath alkane output was measured. Plasma α-tocopherol concentration increased in those receiving vitamin E (P<0.0001). In those supplemented with n−3 fatty acids, EPA and DHA increased in plasma phospholipids (P<0.0001) and plasma MDA and lipid peroxides increased (P<0.001 and P<0.05, respectively). Breath alkane output did not change significantly and vitamin E intake did not prevent the increase in lipid peroxidation during menhaden oil supplementation. The results demonstrate that supplementing the diet with n−3 fatty acids resulted in an increase in lipid peroxidation, as measured by plasma MDA release and lipid peroxide products, which was not suppressed by vitamin E supplementation.     

Concentration of docosahexaenoic acid in glyceride by hydrolysis of fish oil withcandida cylindracea lipase

In an attempt to concentrate the content of DHA (docosahexaenoic acid) in a glyceride mixture containing triglyceride, diglyceride and monoglyceride, fish oil was hydrolyzed with six kinds of microbial lipase. After the hydrolysis, free fatty acid was removed and fatty acid components of the glyceride mixtures were analyzed. When the hydrolysis withCandida cylindracea lipase was 70% complete, the DHA content in the glyceride mixture was three times more than that in the original fish oil. The EPA (eicosapentaenoic acid) content became almost 70% of the original fish oil. Hydrolysis with other lipases did not result in an increase in the DHA content in the glyceride mixtures. Hydrolysis of DHA-rich tuna oil (DHA content is about 25%) withCandida cylindracea lipase resulted in 53% DHA in the glyceride mixture. The EPA content, however, remained close to that of the original tuna oil. In this report, the acyl chain specificity of lipases is evaluated in terms of hydrolysis resistant value (HRV). HRV is the ratio between the DHA contents in the glyceride mixture of hydrolyzed oil and original oil. HRV clearly indicates differences in hydrolysis between DHA and other fatty acids (e.g., saturated and monoenoic acids).     

Combined Urea Complexation and Argentated Silica Gel Column Chromatography for Concentration and Separation of PUFAs from Tuna Oil: Based on Improved DPA Level

Eicosapentaenoic acid (EPA, 20:5n‐3), docosapentaenoic acid (DPA) isomers (22:5n‐6 and 22:5n‐3) and docosahexaenoic acid (DHA, 22:6n‐3) derived from tuna oil were concentrated by three stages of urea fractionation at various crystallization temperatures and different fatty acid/urea ratios. Thereafter, polyunsaturated fatty acids concentrate containing comparatively enriched DPA levels was purified by argentated silica gel column chromatography. A product containing 22.2 ± 0.6 % EPA, 4.6 ± 0.0 % DPAn‐6, 5.9 ± 0.1 % DPAn‐3 and 42.3 ± 1.2 % DHA was obtained at 1:1.6 fatty acid/urea ratio (w/w) by crystallization at ?8 °C for 16 h, ?20 °C for 8 h, and ?8 °C for 16 h. A DPA isomer concentrate containing 26.1 ± 0.5 % DPAn‐6 and 22.3 ± 0.4 % DPAn‐3 was achieved by argentated silica gel chromatography in the 6 % acetone/n‐hexane solvent fraction (v/v), and the recovery of both fatty acids was 66.1 ± 3.2 and 70.7 ± 2.2 %, respectively. Furthermore, 91.9 ± 2.5 % EPA and 99.5 ± 2.1 % DHA with recoveries of 47.8 ± 2.0 and 56.7 ± 3.3 %, respectively, were obtained in various fractions.     

Purification of docosahexaenoic acid by selective esterification of fatty acids from tuna oil with Rhizopus delemar lipase

To purify docosahexaenoic acid (DHA), we attempted the selective esterification of fatty acids originating from tuna oil with lipases. Tuna oil was hydrolyzed in NaOH-ethanol solution, and the resulting fatty acid mixture [DHA, 23.2%; named tuna-free fatty acid (FFA)] was used as a starting material. Rhizopus delemar which acted lightly on DHA, was a suitable catalyst for the selective esterification of tuna-FFA, and lauryl alcohol was the best substrate. The reaction proceeded most effectively when a mixture of 2.4 g lauryl alcohol/tuna-FFA (2:1, mol/mol), 0.6 g water, and 600 U Rhizopus lipase was incubated at 30°C for 20 h with stirring at 500 rpm. Under these conditions 72% of tuna-FFA was esterified, and 84% of DHA was recovered in the unesterified fatty acid fraction. The DHA content in the fatty acid fraction rose from 23 to 73% with this reaction. To further elevate the DHA content, the unesterified fatty acids were extracted, and then esterified again under the same conditions. By this repeated esterification, DHA was purified to 89% with a recovery of 71% of its initial content.     

Transforming growth factor beta in human milk does not change in response to modest intakes of docosahexaenoic acid

Long-chain polyunsaturated fatty acids have been associated with aspects of immune regulation including cytokine production. The purpose of this study was to investigate the effect of maternal dietary supplementation with tuna oil, rich in docosahexaenoic acid (DHA), on the concentration of transforming growth factor beta 1 (TGFβ1) and TGFβ2 in breast milk. In this randomized, dietary intervention trial, mothers of term infants consumed a daily supplement of 2000 mg oil containing either placebo (n=40), 300 mg DHA (n=40), or 600 mg DHA (n=40). The DHA increase in milk and plasma was proportional to dietary DHA. There was no relationship between milk DHA status and TGFβ1 and TGFβ2 levels.     

Influence of formulas with borage oil or borage oil plus fish oil on the arachidonic acid status in premature infants

Several studies have reported that feeding γ-linolenic acid (GLA) has resulted in no increase in arachidonic acid (AA) in newborns. This result was ascribed to the eicosapentaenoic acid (EPA)-rich fish oil used in these formulas. Docosahexaenoic acid (DHA) sources with only minor amounts of EPA are now available, thus the addition of GLA to infant formulas might be considered an alternative to AA supplementation. Sixty-six premature infants were randomized to feeding one of four formulas [ST: no GLA, no long-chain polyunsaturated fatty acids; BO: 0.6% GLA (borage oil); BO + FOLOW: 0.6% GLA, 0.3% DHA, 0.06% EPA; BO + FOHIGH: 0.6% GLA, 0.3% DHA, 0.2% EPA] or human milk (HM, nonrandomized) for 4 wk. Anthropometric measures and blood samples were obtained at study entry and after 14 and 28 d. There were no significant differences between groups in anthropometric measures, tocopherol, and retinol status at any of the studied time points. The AA content of plasma phospholipids was similar between groups at study start and decreased significantly until day 28 in all formulafed groups, but not in the breast-fed infants [ST: 6.6±0.2%, BO: 6.9±0.3%, BO + FOLOW: 6.9±0.4%, BO + FOHIGH: 6.7±0.2%, HM: 8.6±0.5%, where values are reported as mean ±standard error; all formulas significantly different (P≤0.05) from HM]. There was no significant influence of GLA or fish oil addition to the diet. GLA had only a very limited effect on AA status which was too small to obtain satisfactory concentrations (concentrations similar to breast-fed babies) under the circumstances tested. The effect of GLA on AA is independent of the EPA and DHA content in the diet within the dose ranges studied.     

Effect of dietary fish oil on blood levels of free fatty acids,ketone bodies and triacylglycerol in humans

Although the reduction of serum triacylglycerol concentrations by dietary fish oil is a well-known effect, the exact mechanism of this effect has not been previously studied in human subjects. Therefore, the aim of this study was (i) to examine the effect of short-term fish oil supplementation on blood concentrations of ketone bodies, free fatty acids and triacylglycerol in healthy humans and (ii) to verify whether the observed relationships between these variables would be consistent with reduced lipolysis and/or enhanced hepatic fatty acid oxidation after fish oil supplementation. Twenty subjects (21–23 years, normal liver function tests) were randomly divided into two groups to supplement their usual diet with either 30 g/d of fish oil (n=11) or olive oil (n=9). Venous blood samples were drawn after an overnight fast, before and after 1, 3 and 7 d of fish oil/olive oil supplementation. Blood concentrations of triacylglycerol and free fatty acids decreased consistently after fish oil supplementation; the reduction was already significant after one day of fish oil (P<0.001 for triacylglycerol andP=0.01 for free fatty acids). In contrast, neither of these blood values changed after olive oil supplementation (P>0.10). No significant changes in glucose, insulin or ketone body levels were observed in either group after supplementation. After fish oil, but not after olive oil supplementation, the ratio of blood ketone body levels to free fatty acid levels increased significantly (P<0.05). Furthermore, after fish oil supplementation only, free fatty acid levels were significantly correlated with levels of ketone bodies (day 7 of supplementation: r=0.90,P<0.001) and triacylglycerol (maximum value on day 3: r=0.77,P<0.01). These findings suggest that reduced lipolysis and increased hepatic β-oxidation/ketogenesis may contribute to reduced triacylglycerol levels after ω3 fatty acid supplementation in humans. Turnover studies are needed in order to further quantitate these processes.     

Eicosapentaenoic acid is primarily responsible for hypotriglyceridemic effect of fish oil in humans

The aim of this study was to determine whether eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA), or both, were responsible for the triglyceride (TG)-lowering effects of fish oil. EPA (91% pure) and DHA (83% pure), a fish oil concentrate (FOC; 41% EPA and 23% DHA) and an olive oil (OO) placebo (all ethyl esters) were tested. A total of 49 normolipidemic subjects participated. Each subject was given placebo for 2–3 wk and one of the n-3 supplements for 3 wk in randomized, blinded trials. The target n-3 fatty acid (FA) intake was 3 g/day in all studies. Blood samples were drawn twice at the end of each supplementation phase and analyzed for lipids, lipoproteins, and phospholipid FA composition. In all groups, the phospholipid FA composition changed to reflect the n-3 FA given. On DHA supplementation, EPA levels increased to a small but significant extent, suggesting that some retroconversion may have occurred. EPA supplementation did not raise DHA levels, however, FOC and EPA produced significant decreases in both TG and very low density lipoprotein (VLDL) cholesterol (C) levels (P<0.01) and increases in low density lipoprotein (LDL) cholesterol levels (P<0.05). DHA supplementation did not affect cholesterol, triglyceride, VLDL, LDL, or high density lipoprotein (HDL) levels, but it did cause a significant increase in the HDL₂/HDL₃ cholesterol ratio. We conclude that EPA appears to be primarily responsible for TG-lowering (and LDL-C raising) effects of fish oil.     

Dose-Dependent Effects of Docosahexaenoic Acid Supplementation on Blood Lipids in Statin-Treated Hyperlipidaemic Subjects

The objective of the study was to evaluate potential benefits of docosahexaenoic acid (DHA) rich fish oil supplementation as an adjunct to statin therapy for hyperlipidaemia. A total of 45 hyperlipidaemic patients on stable statin therapy with persistent elevation of plasma triglycerides (averaging 2.2 mmol/L) were randomised to take 4 g/day (n = 15) or 8 g/day (n = 15) of tuna oil or olive oil (placebo, n = 15) for 6 months. Plasma lipids, blood pressure and arterial compliance were assessed initially and after 3 and 6 months in 40 subjects who completed the trial. Plasma triglycerides were reduced 27% by 8 g/day DHA-rich fish oil (P < 0.05) but not by 4 g/day when compared with the placebo and this reduction was achieved by 3 months and was sustained at 6 months. Even though total cholesterol was already well controlled by the statin treatment (mean initial level 4.5 mmol/L), there was a further dose-dependent reduction with fish oil supplementation (r = −0.344, P < 0.05). The extent of total cholesterol reduction correlated (r = −0.44) with the initial total cholesterol levels (P < 0.005). In the subset with initial plasma cholesterol above 3.8 mmol/L, plasma very low density lipoprotein (VLDL), intermediate-density lipoprotein (IDL) and low-density lipoprotein (LDL) were isolated and assayed for cholesterol and apolipoprotein B (apoB) at the commencement of the trial and at 3 months of intervention. Fish oil tended to lower cholesterol and apoB in VLDL and raise both in LDL. There were no changes in IDL cholesterol, IDL apoB and high-density lipoprotein cholesterol. The results demonstrate that DHA-rich fish oil supplementation (2.16 g DHA/day) can improve plasma lipids in a dose-dependent manner in patients taking statins and these changes were achieved by 3 months. Fish oil in addition to statin therapy may be preferable to drug combinations for the treatment of combined hyperlipidaemia.     

A randomized controlled trial of the effect of fish oil supplementation in late pregnancy and early lactation on the n−3 fatty acid content in human breast milk

The aim of this research was to investigate the effect of fish oil supplementation, in the third trimester of pregnancy and early lactation period of healthy pregnant Danish women. Forty-four pregnant women were randomly allocated to fish oil supplementation (1.3 g EPA and 0.9 g DHA per day) from week 30 of gestation (FO-group) or to a control regimen (olive oil or no oil; controls). The FO-group was randomly subdivided into women stopping fish oil supplementation at delivery [FO(pregn)], and women continuing supplementation for an, additional 30 d [FO(pregn/lact)]. Thirty-six women agreed to collect milk samples at days 4, 16, and 30 postpartum. The FA composition of the milk samples was determined by GLC. At days 4, 16, and 30 in lactation, FO(pregn/lact) women (n=12) had, respectively 2.3 (P=0.001), 4.1 (P=0.001), and 3.3 (P=0.001) times higher mean contents of LCPUFA(n−3) in their breast milk compared with controls (n=13), and 1.7 (P=0.005), 2.8 (P=0.001), and 2.8 (P=0.001), times higher LCPUFA(n−3) contents, respectively, at these days compared with FO(pregn) women (n=11). The latter group did not differ significantly from controls with regard to LCPUFA(n−3) content in the breast milk. Similar results were obtained when analyzing separately for effects on the milk content of DHA. Dietary supplementation with 2.7 g LCPUFA(n−3) per day from week 30 of gestation and onward more than tripled the LCPUFA(n−3) content in early breast milk; supplementation limited to pregnancy only was much less effective.     

Purification of Free DHA by Selective Esterification of Fatty Acids from Tuna Oil Catalyzed by Rhizopus oryzae Lipase

Tuna fish oil contains 25–30 % docosahexaenoic acid (DHA) and is one of the richest sources of DHA. The present paper investigates the enrichment of DHA by selective esterification of fatty acids obtained from hydrolysis of tuna fish oil catalyzed by Rhizopus oryzae lipase (ROL). The fatty acid mixture obtained after hydrolysis of tuna fish oil, referred to as tuna-FFA contained 26 % DHA. For purification/concentration of DHA in free fatty acids, selective esterification of the fatty acid mixtures with butanol was carried out using ROL in a water-organic solvent system. The best reaction parameters found in this study were pH 7, temperature 35 °C, agitation speed 800 rpm and a fatty acid to solvent (iso-octane) ratio of 1:1.32 (w/v). Also, the effects of other parameters such as type of alcohol, type of enzyme, alcohol to fatty acid ratio, enzyme to fatty acid ratio were studied to determine the most suitable reaction conditions. Exactly 76.2 % of tuna-FFA was esterified in 24 h, under the most suitable reaction conditions and the DHA content in the fatty acid fraction rose from 26 to 86.9 % with 80 % recovery of DHA, after selective esterification. The DHA content of fatty acids in butyl esters was found to be 13.6 %.     

Docosahexaenoic acid supplementation in vegetarians effectively increases omega-3 index: A randomized trial

Low red blood cell (RBC) membrane content of FPA+DHA (hereafter called omega-3 index) has recently been described as an indicator for increased risk of death from coronary heart disease. The relationship between plasma and RBC FA, focusing on omega-3 index, and the response to DHA supplementation were investigated in a double-blind, randomized, placebo-controlled, intervention study. Healthy vegetarians (87 f, 17 m) consumed daily a microalgae oil from Ulkenia sp. (0.94 g DHA/d) or olive oil (placebo) for 8 wk. DHA supplementation significantly increased DHA in RBC total lipids (7.9 vs. 4.4 wt%), in RBC PE (12.1 vs. 6.5 wt%), in RBC PC (3.8 vs. 1.4 wt%), and in plasma phospholipids (PL) (7.4 vs. 2.8 wt%), whereas EPA levels rose to a much lesser extent. Microalgae oil supplementation increased the omega-3 index from 4.8 to 8.4 wt%. After intervention, 69% of DHA-supplemented subjects (but no subject of the placebo group) reached an omega-3 index above the desirable value of 8 wt%. Omega-3 index and EPA+DHA levels in RBC PE, RBC PC, and plasma PL were closely correlated (r always >0.9). We conclude that an 8-wk supplementation with 0.94 g DHA/d from microalgae oil achieves a beneficial omega-3 index of ≥8% in most subjects with low basal EPA+DHA status. RBC total FA analyses can be used instead of RBC lipid fraction analyses for assessing essential FA status, e.g., in clinical studies.