Fatty acid composition of macrophage phospholipids in mice fed fish or borage oil

The polyunsaturated fatty acid (PUFA) composition of murine peritoneal macrophage phospholipids was dramatically altered in vivo following the four-wk feeding of specific dietary oils. Fish oil (containing 20∶5n–3 and 22∶6n−3) feeding significantly increased macrophage 20∶5n−3, 22∶5n−3, and 22∶6n−3 (P<0.05), while borage oil (containing 18∶2n−6 and 18∶3n−6) increased (P<0.05) the macrophage 20∶3n−6/20∶4n−6 ratio, relative to safflower oil (containing 18∶2n−6) and hydrogenated coconut oil (containing 12∶0)-fed animals. The macrophage phospholipid PUFA profiles were compared with those of the liver, lung and spleen. The significance of the PUFA alterations is discussed.     

Influence of genotype on diet-induced changes in membrane phosphatidylcholine and phosphatidylethanolamine composition of splenocytes,liver nuclear envelope and liver mitochondria

Inbred congenic mice of strains MRL/Mp-lpr/lpr (lpr/lpr) and MRL/Mp-+/+ (+/+) were fed nutritionally adequate semipurified diets containing 20% (w/w) fat and differing in linoleic acid content. Levels of linoleic acid (18∶2n−6) and arachidonic acid (20∶4n−6) in phospholipids of splenocytes, liver mitochondria and liver nuclear envelopes were determined. Membranes of lpr/lpr mice exhibited significantly lower levels of 18∶2n−6 and 20∶4n−6 in phospholipids compared with the +/+ strain. The high linoleic acid diet increased incorporation of 18∶2n−6 and 20∶4n−6 in most phospholipid fractions of these membranes. These observations indicate that genotype as well as dietary 18∶2n−6 content significantly influenced incorporation of 18∶2n−6 and 20∶4n−6 into membrane phospholipids. The results also suggest that membrane compositional abnormalities found in the lpr/lpr mice, which develop lymphoma and age faster than +/+ mice, are not restricted to the immune system but also extend to other organs. Differences observed in phospholipid fatty acid composition in splenocytes and liver subcellular membranes for mice fed diets differing in linoleic acid content suggest that the early expression of the lpr gene resulting in progression of autoimmunity may be delayed through dietary manipulation.     

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.     

Effect of tetracosahexaenoic acid on the content and release of histamine, and eicosanoid production in MC/9 mouse mast cell

6,9,12,15,18,21-Tetracosahexaenoic acid (24∶6n−3) was isolated from a brittle star, Ophiura sarsi Lütken, at>95% purity to evaluate its physiological functions. The effects of 24∶6n−3 on the production of leukotriene (LT)-related compounds such as LTB₄, LTC₄ and 5-hydroxyeicosatetraenoic acid, and the accumulation and release of histamine in an MC/9 mouse mast cell line were studied. We found that 24∶6n−3 could inhibit the antigen-stimulated production of LT-related compounds as well as other n−3 polyunsaturated fatty acids (PUFA) such as eicosapentaenoic acid (20∶5n−3) and docosahexaenoic acid (22∶6n−3), which are major n−3 PUFA in fish oils; 24∶6n−3 was also shown to reduce the histamine content in MC/9 cells at 25 μM (27% reduction from the control), and the effect was diminished with increase of the fatty acid concentration (up to 100 μM). These two n−3 PUFA, 20∶5n−3 and 22∶6n−3, also reduced the histamine content (16 and 20% reduction at 25 μM, respectively), whereas arachidonic acid (20∶4n−6) increased it (18% increase at 25 μM). Spontaneous- and antigen-induced release of histamine was not influenced with these PUFA (at 25 μM). Ionophore-stimulated release of histamine was suppressed by the PUFA (13,9,15, and 11% reduction with 20∶4n−6, 20∶5n−3, 22∶6n−3, and 24∶6n−3, respectively). The patterns of the effects of 24∶6n−3 on the synthesis of eicosanoids and histamine content were more similar to those of 22∶6n−3 than 20∶5n−3. From these results, 24∶6n−3 can be expected to have anti-inflammatory activity and antiallergic activities similar to those of 22∶6n−3.     

Effects of dietary n−3 and n−6 polyunsaturated fatty acids on macrophage phospholipid classes and subclasses

This study examined the effects of n−3 and n−6 polyunsaturated fatty acid alimentation on murine peritoneal macrophage phospholipids. Mice were fed complete diets supplemented with either corn oil predominantly containing 18∶2n−6, borage oil containing 18∶2n−6 and 18∶3n−6, fish/corn oil mixture containing 18∶2n−6, 20∶5n−3 and 22∶6n−3, or fish/borage oil mixture containing 18∶2n−6, 18∶3n−6, 20∶5n−3 and 22∶6n−3. After two weeks, the fatty acid levels of glycerophosphoserines (GPS), glycerophosphoinositols (GPI), sphingomyelin (SPH), and of the glycerophosphocholine (GPC) and glycerophosphoethanolamine (GPE) phospholipid subclasses were determined. We found that mouse peritoneal macrophage GPC contain primarily 1-0-alkyl-2-acyl (range for the dietary groups, 24.6–30.5 mol %) and 1,2-diacyl (63.2–67.2 mol %), and that GPE contains 1-O-alk-1-enyl-2-acyl (40.9–47.4 mol. %) and 1,2-diacyl (44.2–51.2 mol %) subclasses. In general, fish oil feeding increased macrophage 20∶5n−3, 22∶5n−3 and 22∶6n−3 levels while simultaneously reducing 20∶4n−6 in GPS, GPI, GPE and GPC subclasses except for 1-O-alk-1′-enyl-2-acyl GPC. Administration of 18∶3n−6 rich diets (borage and fish/borage mixture) resulted in the accumulation of 20∶3n−6 (2-carbon elongation product of 18∶3n−6) in most phospholipids. In general, the novel combination of dietary 18∶3n−6 and n−3 PUFA produced the highest 20∶3n−6/20∶4n−6 phospholipid fatty acid ratios. This study demonstrates that marked differences in the responses of macrophage phospholipid classes and subclasses exist following dietary manipulation. The reduction of 20∶4n−6, while simultaneously increasing 30∶3n−6 and n−3 PUFA levels, may be important in relation to the putative beneficial effects of 20∶3n−6 and fish oil on macrophage eicosanoid and platelet activating factor (PAF) biosynthesis.     

Dietary manipulation of macrophage phospholipid classes: Selective increase of dihomogammalinolenic acid

Because alterations in the dietary content of fatty acids are an important method for modulating macrophage eicosanoid production, we have quantitated the levels of n−6 and n−3 polyunsaturated fatty acids in peritoneal macrophage individual phospholipids from mice fed diets (3 wk) with either safflower oil (SAF), predominantly containing 18∶2n−6, borage (BOR) containing 18∶2n−6 and 18∶3n−6, fish (MFO) containing 20∶5n−3 and 22∶6n−3, and borage/fish mixture (MIX) containing 18∶2n−6, 18∶3n−6, 20∶5n−3 and 22∶6n−3. Dietary n−3 fattya cids were readily incorporated into macrophage phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS) and phosphatidylinositol (PI). The increase in n−3 fatty acid levels was accompanied by a decrease in the absolute levels of 18∶2n−6, 20∶4n−6 and 22∶4n−6 in PC, PE and PS. Interestingly, PI 20∶4n−6 levels were not significantly lowered (P>0.05) in MIX and MFO macrophages relative to SAF and BOR. These data demonstrate the unique ability of this phospholipid to selectively maintain its 20∶4n−6 levels. In BOR and MIX animals, 20∶3n−6 levels were significantly increased (P<0.05) in all phospholipids relative to SAF and MFO. The combination of borage and fish oils (MIX diet) produced the highest 20∶3n−6/20∶4n−6 ratio in all phospholipids. These data show that the macrophage eicosanoid precursor levels of 20∶3n−6, 20∶4n−6 and n−3 acids can be selectively manipulated through the use of specific dietary regimens. This is noteworthy because an increase in phospholipid levels of 20∶3n−6 and 20∶5n−3, while concomitantly reducing 20∶4n−6, may have therapeutic potential in treating inflammatory disorders.     

Effect of temperature on the incorporation into phospholipid classes and metabolismvia desaturation and elongation of n−3 and n−6 polyunsaturated fatty acids in fish cells in culture

The incorporation of 18∶2n−6, 18∶3n−3, 20∶4n−6 and 20∶5n−3 was greater at 10°C than at 22°C in Atlantic salmon (AS), rainbow trout (RTG-2) and turbot (TF) cells. However, there were generally no significant differences between the amount of incorporation of all four polyunsaturated fatty acids (PUFA) into total lipid within a cell type at either 22°C or 10°C. The distributions of the PUFA between individual phospholipid classes at 22°C was essentially the same in AS and TF cells—with the C₁₈ PUFA the order of incorporation in these cells was phosphatidylcholine (PC) > phosphatidylethanolamine (PE) > phosphatidic acid/cardiolipin (PA/CL); with 20∶4n−6 the order was PE and phosphatidylinositol (PI)>PC; with 20∶5n−3, PE>PC. In RTG-2 cells at 22°C the distributions of the C₁₈ PUFA were similar to the other cell lines, but with 20∶4n−6 the order was PC>PI>PE, and with 20∶5n−3 it was PC>PE. At 10°C the incorporation of C₁₈ PUFA into PC increased and into PE and PA/CL decreased, in general, in all cell lines. Incorporation of 20∶5n−3 into PC and PE was increased and decreased at 10°C, respectively, in AS and TF cells, whereas in RTG-2 cells the changes at 10°C were opposite i.e., increased in PE and decreased in PC. With 20∶4n−6, incorporation into PC at 10°C was increased in all cell lines with decreased incorporation into PI in AS and RTG-2 cells and into PE in AS and TF cells, whereas incorporation of 20∶4n−6 into PE increased in RTG-2 cells. The metabolismvia desaturation and elongation of the n−3 PUFA was greater than that of the equivalent n−6 PUFA in all cell lines, irrespective of temperature. There was less conversion of the C₁₈ PUFA at 10°C than at 22°C in RTG-2 and TF cells, but the conversion of 18∶3n−3 by AS cells was increased at 10°C. Temperature had no effect on the conversion of the C₂₀ PUFA.     

Lipid and FA composition of the pearl oyster <Emphasis Type="Italic">Pinctada fucata martensii</Emphasis>: Influence of season and maturation

The lipid and FA composition of the total lipids of the pearl oyster Pinctada fucata martensii, in different seasons and in different areas, were analyzed to clarify its lipid physiology and to estimate the possible influence of its prey phytoplankton. TAG and sterols were the major components in the neutral lipids in all conditions, whereas high levels of phospholipids (PE and PC) were found in the polar lipids. The major FA in the TAG in all samples were 14∶0, 16∶0, and 18∶0 as saturated FA (saturates); 16∶1n−7, 18∶1n−9, and 18∶1n−7 as monoenoic FA (monoenes); and 20∶4n−6 (arachidonic acid: AA), 20∶5n−3 (EPA), and 22∶6n−3 (DHA) as PUFA. The major components found in the polar lipids were 16∶0 and 18∶0 as saturates; 22∶2n−9, 15 and 22∶2n−7, 15 as non-methylene-interrupted dienes (NMID), and AA, 22∶3n−6, 9, 15, EPA, and DHA as PUFA. Although it is a marine animal, characteristically high levels of AA were found in both the TAG and phospholipids. This result suggests that lipids of P. fucata may be influenced by those of its phytoplanktonic prey. The increase in levels of NMID from TAG to PE with a decrease in those of monoenes suggests that the tissues of this species are able to biosynthesize only the less unsaturated PUFA, such as NMID. In particular, NMID derivatives are considered to be biosynthesized in the PE; thus, they might play a particular role in the membrane, because NMID were characteristically localized only in the PE.     

Gas chromatography-chemical ionization-mass spectrometric fatty acid analysis of a commercial supercritical carbon dioxide lipid extract from New Zealand green-lipped mussel (Perna canaliculus)

Supercritical fluid extracts of New Zealand green-lipped mussels (NZGLM) have been suggested to have therapeutic properties related to their oil components. The large number of minor FA in NZGLM extract was characterized by a GC-CIMS/MS method that excels at identification of double-bond positions in FAME. The extract contained five major lipid classes: sterol esters, TAG, FFA, sterols, and polar lipids. The total FA content of the lipid extract was 0.664 g/mL. Fifty-three unsaturated FA (UFA) were fully identified, of which 37 were PUFA, and a further 21 UFA were detected for which concentrations were too low for assignment of double-bond positions. There were 17 saturated FA, with 14∶0, 16∶0, and 18∶0 present in the greatest concentration. The 10 n−3 PUFA detected included 20∶5n−3 and 22∶6n−3, the two main n−3 FA; n−3 PUFA at low concentrations were 18∶3, 18∶4, 20∶3, 20∶4, 21∶5, 22∶5, 24∶6, and 28∶8. There were 43 UFA from the n−4, n−5, n−6, n−7, n−8, n−9, n−10, n−11 families, with 16∶2n−4, 16∶1n−5, 18∶1n−5, 18∶2n−6, 20∶4n−6, 16∶1n−7, 20∶1n−7, 16∶1n−9, 18∶1n−9, and 20∶1n−9 being the most abundant. In general, we estimated that FAME concentrations greater than 0.05% (w/w) were sufficient to assign double-bond positions. In total, 91 FA were detected in an extract of the NZGLM, whereas previous studies of fresh flesh from the NZGLM had reported identification of 42 FA. These data demonstrate a remarkable diversity of NZGLM FA.     

Lipid class and fatty acid composition of the boreal soft coral Gersemia rubiformis

Total lipid, phospholipid, and FA composition and distribution of FA between polar lipids (PL) and neutral lipids (NL) were investigated in the boreal soft coral Gersemia rubiformis from the Bering Sea. The total lipids were mostly hydrocarbons and waxes (33.7%) and PL (33.1%). The content of monoalkyldiacylglycerols (9.7%) exceeded the content of TAG (6.7%). PC and PE constituted 31.4% and 25.6% of total phospholipids, respectively. Principal FA were 16∶0, 16∶1n−7, 18∶0, 18∶1n−9, 18∶1n−7, 20∶1n−7, 20∶4n−6, 20∶4n−3, 20∶5n−3 22∶5n−3, 22∶6n−3, 24∶5n−6, and 24∶6n−3. Most n−6 PUFA (52% of total FA) were associated with the PL fraction; this was especially true for arachidonic and tetracosapentaenoic acids. The NL were enriched with mono-, di-, trienoic, and n−3 PUFA. The variation in EPA levels in both NL and PL suggests an origin of this acid from lipids of diatoms consumed by the corals.     

FA composition of plasma,red blood cell,and liver phospholipids of newborn piglets fed trans isomers of alpha-linolenic acid

The effects of dietary cis and trans α-linolenic acid (18∶3n−3) on the FA composition of plasma, red blood cell, and liver phospholipids were studied in newborn piglets. Animals were fed for 14 d with one of three diets: a control diet (group A) containing cis 18∶3n−3 at a level of 2.0% of total FA, a diet (group B) in which a part of the 18∶3n−3 acid was isomerized (1.3% of cis 18∶3n−3 and 0.7% of trans 18∶3n−3), or a diet (group C) with 2.0% cis 18∶3n−3 and 0.7% trans 18∶3n−3. Feeding animals with diets containing trans 18∶3n−3 resulted in the presence of trans isomers of 18∶3n−3, trans isomers of EPA, and trans isomers of DHA in phospholipids; however, the level of total trans n−3 PUFA in tissues was less than 0.3% of total tissue FA. In group B, the reduction of dietary amounts of cis 18∶3n−3 was associated with a decrease in individual and total cis n−3 PUFA. In contrast, in group C there was no decrease in tissue n−3 PUFA despite the increased dietary level of trans 18∶3n−3. These results suggest that the isomerization of a part of dietary n−3 PUFA, leading to the reduction of their levels in the diet, could induce a decrease in n−3 PUFA in phospholipids. The physiological effects of trans PUFA are not known and should be considered in future studies.     

Linoleic acid-induced fatty acid changes in platelet and aorta of the rat: Effect of age and cholesterol

The influence of age and cholesterol on polyunsaturated fatty acids (PUFa) levels was studied in young and old male Sprague-Dawley rats. Animals were fed a fat-free diet supplemented with 10% (by wt) safflower oil with or without 1% cholesterol for 8 wk. As a result of cholesterol feeding, proportions of linoleic acid (18∶2n−6) and dihomo-γ-linolenic acid (30∶3n−6) were increased and and that of arachidonic acid (20∶4n−6) was decreased in the liver and platelet phospholipids in 64-wk-old rats, suggesting inhibitory effects of cholesterol on 20∶4n−6 synthesis from 18∶2n−6. The prominent age-dependent effect on the levels of PUFA was a retention of C−22 n−3 PUFA, accompanied by decreased C−22 n−6 PUFA and increased 20∶3n−6 in the liver and platelet phospholipids. Ratio of 20∶3n−6/20∶4n−6 increased in 64-wk-old rats regardless of dietary cholesterol, suggesting depressed Δ5-desaturase with age. In aorta phospholipids, 20∶3n−6 content and 20∶3n−6/20∶4n−6 ratio increased with cholesterol supplementation, but not with age. These results suggest that changes of PUFA composition of platelet phospholipids with age are closely linked with changes in liver phospholipids. The 20∶4n−6 content in both platelet and aorta phospholipids is kept constant, despite other n−6 and n−3 PUFA being affected by age.     

Metabolism of n−3 and n−6 fatty acids in Atlantic salmon liver: Stimulation by essential fatty acid deficiency

Oxidation, esterification, desaturation, and elongation of [1-¹⁴C]18∶2n−6 and [1-¹⁴C]18∶3n−3 were studied using hepatocytes from Atlantic salmon (Salmo salar I.) maintained on diets deficient in n−3 and n−6 polyunsaturated fatty acids (PUFA) or supplemented with n−3 PUFA. For both dietary groups, radioactivity from 18∶3n−3 was incoporated into lipid fractions two to three times faster than from 18∶2n−6, and essential fatty acids (FFA) deficiency doubled the incorporation. Oxidation to CO₂ was very low and was independent of substrate or diet, whereas oxidation to acid-soluble products was stimulated by EFA deficiency. Products from 18∶2n−6 were mainly 18∶3n−6, 20∶3n−6, and 20∶4n−6, with minor amounts of 20∶2n−6 and 22∶5n−6. Products from 18∶3n−3 were mainly 18∶4n−3, 20∶5n−3, and 22∶6n−3, with small amounts of 20∶3n−3. The percentage of 22∶6n−3 in the polar lipid fraction of EFA-deficient hepatocytes was fourfold higher than in n−3 PUFA-supplemented cells. This correlated well with our other results obtained after abdominal injection of [1-¹⁴C]18∶3n−3 and [1-¹⁴C]18∶2n−6. In hepatocytes incubated with [4,5-³H]-22∶6n−3, 20∶5n−3 was the main product. This retrocon-version was increased by EFA deficiency, as was peroxisomal β-oxidation activity. This study shows that 18∶2n−6 and 18∶3n−3 can be elongated and desaturated in Atlantic salmon liver, and that this conversion and the activity of retroconversion of very long chain PUFA is markedly enhanced by FFA deficiency.     

Alteration in mouse splenic phospholipid fatty acid composition and lymphoid cell populations by dietary fat

The fatty acid composition of diacyl- and alkylacylglycerophosphocholine (PC), phosphatidylinositol (PI), phosphatidylserine (PS), alkenylacyl-glycerophosphoethanolamine (aPE), and diacyl- and alkylacyl-glycerophosphoethanolamine (dPE) was assessed in isolated splenocytes from C3H/Hen mice fed one of four purified isocaloric diets for six weeks. Diets contained 20% by weight of either a high-linoleate sunflower oil (Hi 18∶2), a high-oleate sunflower oil (Hi 18∶1), a mixture of 17% menhaden fish oil and 3% high-linoleate sunflower oil (Hi n−3), or a mixture of 17% coconut oil and 3% high-linoleate sunflower oil (Hi SFA). Spleen weight and immune cell yield were significantly higher (P<0.05) in mice fed the Hi 18∶1 or the Hi n−3 diets compared with those fed the Hi 18∶2 and Hi SFA diets. Distinctive patterns of fatty acids were observed for each phospholipid in response to dietary fatty acids. Dietary fat significantly affected (P<0.05) total polyunsaturated fatty acids (PUFA) in PC and dPE, total saturated fatty acids (SFA) in PC, total monounsaturated fatty acids (MUFA), and n−3 PUFA in all phospholipid classes examined. In mice fed the Hi n−3 diet, n−3 PUFA were significantly elevated, whereas n−6 PUFA decreased in all of the phospholipids. In these mice, eicosapentaenoic acid (EPA) was the predominant n−3 PUFA in PC and PI, whereas docosahexaenoic acid (DHA) was the major n−3 PUFA in aPE and PS. Interestingly, the ratios of n−3/n−6 PUFA in the phospholipids from these mice were 3.2, 2.4, 1.8, 0.8 and 0.8 for aPE, PS, dPE, PC and PI, respectively. These data suggest a preferential incorporation of n−3 PUFA into aPE, PS and dPE over PC and PI.     

Dietary fish oil dose-response effects on ileal phospholipid fatty acids and contractility

We have reported that dietary fish oil (FO) leads to the incorporation of long-chain n−3 PUFA into the gut tissue of small animal models, affecting contractility, particularly of rat ileum. This study examined the FO dose response for the incorporation of n−3 PUFA into ileal tissue and how this correlated with in vitro contractility. Groups of ten to twelve 13-wk-old Wistar-Kyoto rats were fed 0, 1, 2.5, and 5% FO-supplemented diets balanced with sunflower seed oil for 4 wk, after which ileal total phospholipid FA were determined and in vitro contractility assessed. For the total phospholipid fraction, increasing the dietary FO levels led to a significant increase first evident at 1% FO, with a stepwise, nonsaturating six-fold increase in n−3 PUFA as EPA (20∶5n−3), DPA (docosapentaenoic acid, 22∶5n−3), and DHA, but mainly as DHA (22∶6n−3), replacing the n−6 PUFA linoleic acid (18∶2n−6) and arachidonic acid (20∶4n−6) over the dosage range. There was no difference in KCl-induced depolarization-driven contractility. However, a significant increase in receptor-dependent maximal contractility occurred at 1% FO for carbachol and at 2.5% FO for prostaglandin E₂, with a concomitant increase in sensitivity for prostaglandin E₂ at 2.5 and 5% FO. These results demonstrate that significant increases in ileal membrane n−3 PUFA occurred at relatively low doses of dietary FO, with differential receptor-dependent increases in contractility observed for muscarinic and prostanoid agonists.     

The Effects of dietary n−3/n−6 ratio on brain development in the mouse: a dose response study with long-chain n−3 fatty acids

This study examines the effects of the ratio of n−3/n−6 fatty acids (FA) on brain development in mice when longchain n−3 FA are supplied in the diet. From conception until 12 days after birth, B6D2F₁ mice were fed liquid diets, each providing 10% of energy from olive oil, and a further 10% from different combinations of free FA concentrates derived from safflower oil (18∶2n−6), and fish oil (20∶5n−3 and 22∶6n−3). The range of dietary n−3/n−6 ratios was 0,025, 0.5, 1.0, 2.0, and 4.0, with an n−6 content of greater than 1.5% of energy in all diets, and similar levels of total polyunsaturated fatty acids (PUFA). In an additional group of ratio 0.5, 18∶2n−6 was partially replaced by its δ6 desaturation product, 18∶3n−6. Biochemical analyses were conducted on 12-day-old pup brains, as well as on samples of maternal milk. No obvious effects on overall pup growth and development were observed, apart from a smaller litter size at ratio 1. Co-variance analysis indicated that increasing the n−3/n−6 ratio was associated with slightly smaller brains, relative to body weight. We found that 18∶2n−6 and 20∶5n−3 were the predominant n−6 and n−3 FA in the milk; in the brain these were 20∶4n−6 and 22∶6n−3, respectively. Increasing dietary n−3/n−6 ratios generally resulted in an increase in n−3 FA, with a corresponding decrease in n−6 FA. The n−3/n−6 ratio of the milk lipids showed a strong linear relationship with the diet, but in the brain the rate of increase tended to decrease beyond 0.5 (phosphatidylcholine, PC) and 0.25 (phosphatidylethanolamine, PE), such that there was a significant quadratic contribution to the relationship. The partial replacement of dietary 18∶2n−6 with 18∶3n−6 raised levels of 20∶4n−6 in milk, brain PC, and brain PE. These results indicate that the n−3/n−6 ratio of the phospholipids in the developing mouse brain responds maximally to maternal dietary long-chain n−3/n−6 ratios of between 0.25 and 0.5.     

Changes in phosphatidylcholine molecular species in the shrimp <Emphasis Type="Italic">Macrobrachium borellii</Emphasis> in response to a water-soluble fraction of petroleum

The effect of the water-soluble fraction (WSF) of crude oil on lipid contents, lipid classes, FA, and PC molecular species was studied in high-phospholipid (hepatopancreas) and low-phospholipid (egg) tissues of a freshwater crustacean. After a 21-d exposure to a sublethal concentration of WSF, a significant decrease in shrimp total lipids was observed, although no alterations could be detected in the hepatopancreas or egg lipid contents. TAG/phospholipid ratios increased in the hepatopancreas and decreased in the eggs, suggesting alterations either in the mobilization of TAG to phospholipid pools or in the energy balance. The FA composition of phosphoglycerides in the hepatopancreas and eggs was dominated by PUFA, whereas the n−3/n−6 ratio was not affected by WSF exposure, although there was a significant increase in hepatopancreas 18∶1n−9. Analysis of the PC molecular species by HPLC-ELSD showed the presence of 15 species, with 16∶0/18∶1, 18∶1/18∶2, 16∶0/20∶5, and 16∶1/20∶5 being the major species in the hepatopancreas. The PC molecular species in the eggs showed a different pattern, dominated by 16∶0/18∶1 and 18∶1/18∶2. Of the PC molecular species, 10 contained 22∶6n−3, 20∶5n−3, and 20∶4n−6. Small amounts of di-PUFA species were also found. Exposure to WSF altered the PC molecular species in both tissues. The four major hepatopancreas molecular species and most of the ones containing PUFA decreased. This was compensated for by an increase in 16∶1/18∶1 (152%) and 18∶1/18∶1 (50%). The two major egg PC molecular species decreased, whereas the PUFA-containing ones increased. The contrasting responses of both tissues of WSF contamination suggests the presence of different homeostatic mechanisms.     

Hyperphagia modifies FA profiles of plasma phospholipids,plasma FFA,and adipose tissue TAG

Hyperphagia was achieved by continuous intracerebroventricular infusion of a melanocortin receptor antagonist (HS024; Neosystem, Strasbourg, France) in rats. The effects of hyperphagia on FA composition and concentration of plasma phospholipids (PL), plasma FFA, and adipose tissue TAG were studied in rats for 8 d [short-term hyperphagia (STH); n=8], or 28 d [longterm hyperphagia (LTH); n=9]. The control rats were treated with artificial cerebrospinal fluid for 8 d (n=8) or 28 d (n=10). The rats were fed the same regular diet. In STH rats the plasma PL and fasting plasma FFA contained higher concentrations of saturated FA (SFA) and monounsaturated FA (MUFA), and plasma FFA contained lower n−6 PUFA than in the control rats. In LTH rats the plasma PL contained higher concentrations of SFA, MUFA, and n−3 PUFA and higher proportions of 16∶1n−7 and 18∶1n−9 at the expense of 18∶2n−6 than in the control rats. In LTH rats the abundant dietary intake of 18∶2n−6 did not enrich 18∶2n−6 of the plasma PL or adipose tissue TAG. In LTH rats the fasting plasma FFA contained more than twofold higher concentrations of SFA and MUFA, and higher proportions of 16∶1n−7 and 18∶1n−9 at the expense of 18∶2n−6 than in the control rats. This animal obesity model shows that LTH affects the FA composition and concentration of plasma PL, plasma FFA, and adipose tissue TAG, a result consistent with changes associated with increased risk of various diseases in humans. These results also demonstrate that LTH alters the FA composition of plasma PL and adipose tissue TAG in a way that does not reflect the FA composition of dietary fat.     

Major clofibrate effects on liver and plasma lipids are independent of changes in polyunsaturated fatty acid composition induced by dietary fat

The effects of clofibrate on the content and composition of liver and plasma lipids were studied in mice fed for 4 wk on diets enriched in n−6 or n−3 polyunsaturated fatty acids (PUFA) from sunflower oil (SO) or fish oil (FO), respectively; both oils were fed at 9% of the diet (dry weight basis). Only FO was hypolipidemic. Both oil regimes led to slightly increased concentrations of phospholipids (PL) and triacylglycerols (TG) in liver as compared with a standard chow diet containing 2% fat. Clofibrate promoted hypolipidemia only in animals fed SO. Its main effect was to enlarge the liver, such growth increasing the amounts of major glycerophospholipids while depleting the TG. SO and FO consumption changed the proportion of n−6 or n−3 PUFA in liver and plasma lipids in opposite ways. After clofibrate action, the PUFA of liver PL were preserved better than in the absence of oil supplementation. However, most of the drug-induced changes (e.g., increased 18∶1n−9 and 20∶3n−6, decreased 22∶6/20∶5 ratios) occurred inrrespective of lipids being rich in n−6 or n−3 PUFA. The concentration of sphingomyelin (SM), a minor liver lipid that virtually lacks PUFA, increased with the dietary oils, decreased with clofibrate, and changed its fatty acid composition in both situations. Thus. oil-increased SM had more 22∶0 and 24∶0 than clofibrate-decreased SM, which was significantly richer in 22∶1 and 24∶1.     

Composition and seasonal variation of fatty acids of <Emphasis Type="Italic">Diplodus vulgaris</Emphasis> L. from the Adriatic Sea

Muscle tissue from the common two-banded sea bream Diplodus vulgaris L. originating from the Adriatic Sea, Croatia, was analyzed. The FA composition of neutral (TAG) and polar (PE, PC, PI/PS) lipid classes was determined, as well as the lipid and water contents during winter and summer periods. Both the total lipid and water contents were higher in the winter period. We identified 16 different FA. The major constituents of the total FA in both seasons were saturates: palmitic (16∶0) and stearic acids (18∶0); monoenes: oleic (18∶1n−9) and palmitoleic acids (16∶1n−7); and polyunsaturates: arachidonic acid (20∶4n−6), EPA (20∶5n−3), and DHA (22∶6n−3), but their amounts and ratios differed significantly between the two seasons and between lipid fractions. The FA composition showed a noticeable pattern of seasonality that reflected fluctuations mainly in TAG. The diminution of the monounsaturated FA content in the summer was clearly followed by an increase in PUFA content. Diplodus vulgaris is a good source of natural n−3 PUFA and would therefore be suitable for inclusion in highly unsaturated low-fat diets.