The Na+K+ATPase activity in cultured human fibroblasts with an elevated phospholipid triene: tetraene ratio

Human skin fibroblasts were cultured at low density for 11 days in MCDB 110, 0.4% fetal bovine serum, a mitogen mixture, and were supplemented with 18∶2n−6 or 18∶1n−9 as a fatty acid-albumin complex. The cells cultured with the 18∶2n−6 supplement had a 20∶3n−9/20∶4n−6 ratio of 0.29±0.07; the 18∶1n−9 supplemented cells had a ratio of 1.51±0.27. There was less than 4% difference in total growth of the cell population under the two culture conditions. The cells supplemented with 18∶2n−9 had similar levels of protein/cell, K⁺/mg cell protein and functional Na⁺K⁺ATPase activity.     

Structures,Stabilities and Electronic Properties of Endo- and Exohedral Dodecahedral Silsesquioxane (T<Subscript>12</Subscript>-POSS) Nanosized Complexes with Atomic and Ionic Species

The structures of endohedral complexes of the polyhedral oligomeric silsesquioxane (POSS) cage molecule (HSiO_3/2)₁₂, with both D _2d and D _6h starting cage symmetries, containing the atomic or ionic species: Li⁰, Li⁺, Li⁻, Na⁰, Na⁺, Na⁻, K⁰, K⁺, K⁻, F⁻, Cl⁻, Br⁻, He, Ne, Ar were optimized by density functional theory using B3LYP and the 6-311G(d,p) and 6-311 ++G(2d,2p) basis sets. The exohedral Li⁺, Na⁺, K⁺, K⁻, F⁻, Cl⁻, Br⁻, He, Ne, Ar complexes, were also optimized. The properties of these complexes depend on the nature of the species encapsulated in, or bound to, the (HSiO_3/2)₁₂ cage. Noble gas (He, Ne and Ar) encapsulation in (HSiO_3/2)₁₂ has almost no effect on the cage geometry. Alkali metal cation encapsulation, in contrast, exhibits attractive interactions with cage oxygen atoms, leading to cage shrinkage. Halide ion encapsulation expands the cage. The endohedral X@(HSiO_3/2)₁₂ (X = Li⁺, Na⁺, K⁺, F⁻, Cl⁻, Br⁻, He and Ne) complexes form exothermically from the isolated species. The very low ionization potentials of endohedral Li⁰, Na⁰, K⁰ complexes suggest that they behave like “superalkalis”. Several endohedral complexes with small guests appear to be viable synthetic targets. The D _2d symmetry of the empty cage was the minimum energy structure in accord with experiment. An exohedral fluoride penetrates the D _6h cage to form the endohedral complex without a barrier.     

Dietary N−6 and N−3 fatty acids and salt-induced hypertension in the borderline hypertensive rat

This study examined the effects of salt-loading on blood pressure (BP) development in the borderline hypertensive rat (BHR) and its modification by dietary n−3 and n−6 fatty acids. In experiment 1, 4 groups (n=10/group) of male BHR receiving 1% NaCl as a replacement for tap water were placed on chow enriched with either olive oil (OL), sunflower oil (SUN), evening primrose oil (EPO) or fish oil for 6 weeks. BP, heart rate, body weight, water, Na⁺ and K⁺ intake and urinary output were measured weekly. SUN and fish oil reduced the pressor response to salt seen vs the OL group by 50%, and EPO abolished the pressor response, reducing BP below control levels. The BP response was unrelated to either food intake or water and electrolyte intake and excretion. In experiment 2, male BHR received water +/− 18∶3n−6 (0.04 mg/hr in OL via ip pump) or 1% NaCl +/− 18∶3n−6 (n=12/group) for 12 weeks, followed by 2 weeks recovery on tap water. Salt increased BP, and 18∶3n−6 decreased this response, but had no effect on animals receiving tap water. In experiment 3, effects of 3 doses of 18∶3n−6 (0.04, 0.08, and 0.12 mg/hr) on the pressor response to 1% NaCl were examined. All doses reduced the BP response to salt vs controls with no dose-response. These data suggest that the BHR is genetically salt-sensitive, and that dietary n−3 and n−6 fatty acids can attenuate the cardiovascular response to salt in this model.     

Pyloric ceca are significant sites of newly synthesized 22∶6n−3 in rainbow trout (<Emphasis Type="Italic">Oncorhynchus mykiss</Emphasis>)

In this pulse-chase study, rainbow trout fed a diet containing deuterated (D₅) (17,17,18,18,18)-18∶3n−3 ethyl ester accumulated D₅-22∶6n−3 in pyloric ceca to a greater extent than in liver 2 d post-dose. The ratio of newly synthesized D₅-22∶6n−3 in ceca to that in liver 2 d after feeding D₅-18∶3n−3 was 4.7±1.2 when expressed as per mg tissue and 5.2±2.4 when expressed as per mg protein. The amount of D₅-22∶6n−3 in ceca then declined whereas that in liver and blood increased, with the ratio of ceca to liver falling to 1.7 and 1.4, respectively, by day 5 and approaching unity by day 9. A crude cecal mucosa fraction contained 123±50 ng D₅-22∶6n−3/mg protein/mg D₅-18∶3n−3 eaten 2 d after feeding the tracer, compared with 35±21 ng D₅-22∶6n−3/mg protein/mg D₅-18∶3n−3 eaten in liver. Three days later the amount in cecal mucosa had fallen by one-third and that in liver had increased threefold. Most of the D₅-18∶3n−3 was catabolized very rapidly. The ratio of D₅-18∶3n−3 to 21∶4n−6 (a relatively inert FA marker) in the diet was 4.0, but this fell to 0.30 in ceca and ca. 0.8 in liver, blood, and whole carcass one day after feeding. These results indicate that ceca are active in the synthesis of 22∶6n−3 and the oxidation of 18∶3n−3.     

Fatty acid composition of the adipose tissue and yolk lipids of a bird with a marine-based diet, the emperor penguin (Aptenodytes forsteri)

The emperor penguin (Aptenodytes forsteri) is an Antarctic seabird feeding mainly on fish and therefore has a high dietary intake of n-3 polyunsaturated fatty acids. The yolk is accumulated in the developing oocyte while the females are fasting, and a large proportion of the fatty acid components of the yolk lipids are derived by mobilization from the female's adipose tissue. The fatty acid composition of the total lipid of the yolk was characterized by high levels of n-3 polyunsaturated fatty acids. However, it differed in several respects from that of the maternal adipose tissue. For example, the proportions of 14∶0, 16∶1n−7, 20∶1n−9, 22∶1n−9, 20∶5n−3, and 22∶6n−3 were significantly greater in adipose tissue than in yolk. Thus adipose tissue lipids contained 7.6±0.3% and 8.0±0.3% (wt% of total fatty acids; mean ±SE; n=5) of 20∶5n−3 and 22∶6n−3, respectively, whereas the yolk total lipid contained 1.6±0.1 and 5.5±0.3% of these respective fatty acids. The proportions of 16∶0, 18∶0, 18∶1n−9, 18∶2n−6, and 20∶4n−6 were significantly lower in the adipose tissue than in the yolk lipids. The proportions of triacylglycerol, phospholipid, free cholesterol, and cholesteryl ester in the yolk lipid were, respectively, 67.0±0.2, 25.4±0.3, 5.3±0.2, and 1.8±0.2% (wt% of total yolk lipid). The proportions of 20∶4n−6, 20∶5n−3, 22∶5n−3, and 22∶6n−3 were, respectively, 5.7±0.3, 2.8±0.2, 1.4±0.1, and 11.7±0.5% in phospholipid and 0.4±0.0, 1.2±0.1, 0.8±0.1 and 3.6±0.3% in triacylglycerol. About 95% of the total vitamin E in the yolks was in the form of α-tocopherol with γ-tocopherol forming the remainder. Two species of carotenoids, one identified as lutein, were present.     

Dietary fat effects on hepatic lipid peroxidation and enzymes of H2O2 metabolism and NADPH Generation

The purpose of this study was to determine the effects of dietary fat quantity and fatty acid composition on hepatic H₂O₂-metabolizing systems, activities of NADPH-generating enzymes and lipid peroxidation. Onemonth-old male C57BL/6J mice were fed one of six diets: (i) 5% fat, rich in 18∶2n−6 fatty acid (5% N−6); (ii) 20% fat, rich in 18∶3n−3 (N−3); (iii) 20% fat, rich in 18∶2n−6 (N−6); (iv) 20% fat, rich in 18∶1n−9 (N−9); (v) 20% fat, rich in saturated fatty acids (SAT); and (vi) 20% fat, deficient in essential fatty acids (EFAD); for 11 wk. Comparisons between animal groups receiving different fat quantities showed that activities of glucose-6-phosphate dehydrogenase (G6PDH, EC 1.1.1.49) and malic enzyme (ME, EC 1.1.1.40) and the levels of conjugated dienes were significantly lower in the N−6 than in 5% N−6 group. Conversely, activities of catalase (CAT, EC 1.11.1.6) and seleniumglutathione peroxidase (SeGSHPx, EC 1.11.1.9) were higher in the N−6 than in 5% N−6 group. Among the five dietary groups receiving 20% fat but differing in fatty acid composition, CAT activity was lower in the N−9 group, SeGSHPx activity was lower in the EFAD group, and glutathione reductase (GSSGR, EC 1.6.4.2) activity was higher in the N−6 than in the N−3, N−9, SAT and EFAD groups. The EFAD group had much higher levels of total lipids and conjugated dienes, as well as activities of NADPH-generating enzymes, including G6PDH, ME and isocitrate dehydrogenase (EC 1.1.1.42), than the other four high-fat groups. The hepatic levels of malondialdehyde were not different among the five groups fed 20% fat. In the EFAD group, higher hepatic lipid content can be attributed to higher activities of NADPH-generating enzymes, and the elevation of conjugated diene levels may be related to increased oxygenation of 20∶−6 (Mead acid)via the lipoxygenase/cyclooxygenase pathway. In short, both dietary fat quantity and fatty acid composition selectively affected hepatic H₂O₂-metabolizing systems, activities of NADPH-generating enzymes and lipid peroxidation status.     

Biosynthesis and tissue deposition of docosahexaenoic acid (22∶6n−3) in rainbow trout (Oncorhynchus mykiss)

Rainbow trout (Oncorhynchus mykiss) weighing ca. 5 g and previously acclimated for 8 wk on a diet comprising vegetable oil (11%), fish meal (5%), and casein (48%) as the major constituents were fed a pulse of diet containing deuterated (D₅) (17,17,18,18,18)-18∶3n−3 ethyl ester. The synthesis and tissue distribution of D₅-22∶6n−3 was determined 3,7,14, 24, and 35 d after the pulse. The whole-body accumulation of D₅-22∶6n−3 was linear over the first 7 d, corresponding to a rate of 0.54±0.12 μg D₅-22∶6n−3/g fish/mg D₅-18∶3n−3 eaten/d. Maximal accretion of D₅-22∶6n−3 was 4.3±1.2 μg/g fish/mg of D₅-18∶3n−3 eaten after 14 d. The amount of D₅-22∶6n−3 peaked in liver at day 7, in brain and eyes at day 24, and plateaued after day 14 in visceral and eye socket adipose tissue and in the whole fish. The majority of D₅-22∶6n−3 was found in the carcass (remaining tissues minus the above tissues analyzed separately) at all times. On a per milligram lipid basis, liver and eyes had the highest concentration of D₅-22∶6n−3. The experimental diet also contained 21∶4n−6 ethyl ester as a marker to estimate the amount of food eaten by individual fish. From such estimates it was calculated that the great majority of the D₅-tracer was catabolized, with the combined recovery of D₅-18∶3n−3 plus D₅-22∶6n−3 being 2.6%. The recovery of 21∶4n−6 was 57.6%. The concentration of 22∶6n−3 in the fish decreased during the 13-wk period, and the amount of 22∶6n−3 synthesized from 18∶3n−3 was only about 5% of that obtained directly from the fish meal in the diet.     

Linoleic acid uptake by isolated enterocytes: Influence of α-linolenic acid on absorption

In a previous study we showed that intestinal uptake of α-linolenic acid (18∶3n−3) was carrier-mediated and we suggested that a plasma membrane fatty acid protein was involved in the transport of long-chain fatty acids. To further test this hypothesis, the mechanism of linoleic acid (18∶2n−6) uptake by isolated intestinal cells was examined using a rapid filtration method and 20 mM sodium taurocholate as solubilizing agent. Under these experimental conditions transport of [1-¹⁴C]linoleic acid monomers in the concentration range of 2 to 2220 nM was saturable with a V_m of 5.1±0.6 nmol/mg protein/min and a K_m of 183±7 nM. Experiments carried out in the presence of metabolic inhibitors, such as 2,4-dinitrophenol and antimycin A, suggested that an active, carriermediated mechanism was involved in the intestinal uptake of this essential fatty acid. The addition of excess unlabeled linoleic acid to the incubation medium led to a 89% decrease in the uptake of [1-¹⁴C]linoleic acid, whiled-glucose did not compete for transport into the cell. Other long-chain polyunsaturated fatty acids added to the incubation mixture inhibited linoleic acid uptake by more than 80%. The presence of α-linolenic acid (18∶3n−3) in the incubation medium caused the competitive inhibition (K_i=353 nM) of linoleic acid uptake. The data are compatible with the hypothesis that intestinal uptake of both linoleic, and α-linolenic acid is mediated by a membrane carrier common to long-chain fatty acids.     

Influence of temperature and high dietary linoleic acid content on esterification,elongation, and desaturation of PUFA in atlantic salmon hepatocytes

The esterification, desaturation, and elongation of [1-¹⁴C]18∶3n−3, [1-¹⁴C]18∶2n−6, and [1-¹⁴C]20∶5n−3 at 5 and at 12°C were studied using cultivated hepatocytes from Atlantic salmon. The salmon were fed diets, in which 0, 50, or 100% of the supplementary fish oil had been replaced by soybean oil, for 950 day-degrees at 5 and 12°C. The endogenous percentage of 18∶2n−6 in hepatocyte lipids was 2% in cells from fish fed a diet with 100% of the supplemental lipid from fish oil, and it was slightly less than 25% in cells from fish fed the diet with 100% of the supplemental lipid from soybean oil. Furthermore, the percentages of 20∶3n−6 and 20∶4n−6 were significantly higher in hepatocytes from fish fed on soybean oil than they were in those of fish fed on fish oil. The percentages of 20∶5n−3 and 22∶6n−3, on the other hand, were lower. The endogenous levels of n−6 FA were not significantly correlated with the total amounts of radiolabeled FA esterified in hepatocyte lipids. The main radiolabeled products formed from 18∶2n−6 were 20∶2n−6 and 20∶3n−6. The level of the important eicosanoid precursor 20∶4n−6 was twice as high in hepatocyte phospholipids from fish fed the 100% soybean oil diet as it was in hepatocytes from fish fed the diet with 100% of supplemental lipid from fish oil. The main products formed from 18∶3n−3 were 20∶4n−3, 20∶5n−3, and 22∶6n−3. High levels of dietary 18∶2n−6 do allow, or even seem to increase, the production of 22∶6n−3 from 18∶3n−3 in hepatocytes. The main products formed from 20∶5n−3 were 22∶5n−3 and 22∶6n−3. The production of 22∶6n−3 from 20∶5n−3 was higher at 5°C than at 12°C. The percentage of 24∶5n−3 was higher at 5°C than it was at 12°C, as was the ratio of 24∶5 to 22∶5. These results suggest that the elongation rate of 22∶5n−3 to 24∶5n−3 is higher at the lower temperature.     

Molecular species of 1-O-alk-1′-enyl-2-acyl-, 1-O-alkyl-2-acyl- and 1,2-diacyl glycerophospholipids in Japanese oysterCrassostrea gigas (Thunberg)

Molecular species of 1-O-alk-1′-enyl-2-acyl-, 1-O-alkyl-2-acyl-, and 1,2-diacyl-sn-glycero-3-phosphoethanolamine (EPL) andsn-glycero-3-phosphocholine (CPL) of Japanese oysterCrassostrea gigas were analyzed by selectedion monitoring gas chromatography/mass spectrometry using electron impact ionization. The characteristic fragment ions, [RCH=CH+56]⁺ due to the alkenyl residue in thesn-1 position and [RCO+74]⁺ due to the acyl residue in thesn-2 position of alkenylacylglycerols, [R+130]⁺ due to the alkyl residue in thesn-1 position and [RCO+74]⁺ due to the acyl residue in thesn-2 position of alkylacylglycerols, [RCO+74]⁺ due to the acyl residues in thesn-1 and/orsn-2 positions of diacylglycerols, and [M−57]⁺ being indicative of the corresponding molecular weight, were used for structural assignments. For alkenylacyl EPL and CPL, 19 and 16 molecular species were determined, respectively. Two molecular species, 18∶0alkenyl-22∶6n−3 and 18∶0-alkenyl-22∶2-non-methylene interrupted diene (NMID), amounted to 53.2% and 47.9%, respectively. The alkylacyl EPL and CPL consisted of 16 and 20 molecular species, respectively, and the prominent components were 18∶0alkyl-22∶2NMID, 20∶1alkyl-20∶1n−11 (27.4%) and 20∶1alkyl-20∶2NMID (16.3%) in the former, and 16∶0alkyl-20∶5n−3 (23.0%) and 16∶0alkyl-22∶6n−3 (21.6%) in the latter. For the diacyl EPL and CPL, 14 and 51 molecular species were determined, respectively. The major molecular species were 18∶0–20∶5n−3 (37.4%), 16∶0–20∶5n−3 (14.2%) and 18∶1n−7–22∶2NMID (13.2%) in the former, and 16∶0–20∶5n−3 (33.4%) and 16∶0–22∶6n−3 (22.3%) in the latter. It was found that there were significant differences in the molecular species between the alkylacyl and diacyl EPL and the alkylacyl and diacyl CPL; the number of molecular species was larger in CPL than in EPL, while the number of total carbons and double bonds of the major molecular species were larger in the EPL than in the CPL. Alkenylacyl EPL were similar to alkenylacyl CPL in molecular species composition.     

Lithium-metal potential in Li<Superscript>+</Superscript> containing ionic liquids

Impedance spectroscopy studies of the interface between lithium and ionic liquid (IL) showed the formation of a film (solid electrolyte interface, SEI), protecting metal from its further dissolution. Consequently, the potential of metallic lithium immersed in an electrolyte containing Li⁺ cations may be described as a Li|SEI|Li⁺ system, rather than simply Li/Li⁺. The potential of lithium-metal in a series of ionic liquids (and in a number of molecular liquids) containing Li⁺ cation (0.1 M) was measured versus the Ag|(Ag⁺ 0.01 M, cryptand 222 0.1 M, in acetonitrile) reference. The lithium-metal potential (E(Li|SEI|Li⁺)) was ca. −2.633 ± 0.017 V in ILs based on the [N(CF₃SO₂)₂ ^– ] anion, while −2.848 ± 0.043 V in ILs containing [BF₄ ^– ] anion (the difference is ca. 200 mV). In the case of ILs based on the triflate anion ([CF₃SO₃ ^– ]), the cation of ionic liquid also influences the E(Li|SEI|Li⁺) value: it was ca. −1.987 ± 0.075 V for imidazolium based cations and much lower (−2.855 V) for the pyrrolidinium based cation. In ionic liquid based on the imidazolium cation and hexafluorophosphate anion ([PF₆ ^– ]), the Li/SEI/Li⁺ potential was −2.245 V. The Li|SEI|Li⁺ potential measured in cyclic carbonates was −2.780 ± 0.069 V while in dimethylsulfoxide showed the lowest value of ca. −3.285 V. The measured potentials were also expressed versus the formal potential of the ferrocene/ferrocinium redox couple, obtained from cyclic voltammetry.     

Effect of 18∶1n−9, 20∶5n−3, and 22∶6n−3 on lipid accumulation and secretion by atlantic salmon hepatocytes

We have studied the effects of dietary FA on the accumulation and secretion of [³H]glycerolipids by salmon hepatocytes in culture. Atlantic salmon were fed diets supplemented with either 100% soybean oil (SO) or 100% fish oil (FO), and grew from an initial weight of 113±5 g to a final weight of 338 ±19 g. Hepatocytes were isolated from both dietary groups and incubated with [³H]glycerol in an FA-free medium; a medium supplemented with 0.75 mM of one of three FA—18∶1n−9, 20∶5n−3, or 22∶6n−3—or a medium supplemented with 0.75 mM of the sulfur-substituted FA analog tetradecylthioacetic acid (TTA), which cannot undergo β-oxidation. Incubations were allowed to proceed for 1,2,6, or 24 h. The rate of the secretion of radioactive glycerolipids with no FA added was 36% lower from hepatocytes isolated from fish fed the FO diet than it was from hepatocytes isolated from fish fed the SO diet. Hepatocytes incubated with 18∶1n−9 secreted more [³H]TAG than when incubated with no FA, whereas hepatocytes incubated with 20∶5n−3 or TTA secreted less labeled TAG than when incubated with no FA. This observation was independent of the feeding group. Hepatocytes incubated with 22∶6n−3 secreted the highest amounts of total [³H]glycerolipids compared with the other treatments, owing to increased secretion of phospholipids and mono- and diacylglycerols (MDG). In contrast, the same amounts of [³H]TAG were secreted from these cells as from cells incubated in an FA-free medium. The lipid-lowering effect of FO is thus independent of 22∶6n−3, showing that 20∶5n−3 is the FA that is responsible for the lipid-lowering effect. The ratio of TAG to MDG in lipids secreted from hepatocytes to which 20∶5n−3 or TTA had been added was lower than that in lipids secreted from hepatocytes incubated with 18∶1n−9 or 22∶6n−3, suggesting that the last step in TAG synthesis was inhibited. Morphometric measurements revealed that hepatocytes incubated with 20∶5n−3 accumulated significantly more cellular lipid than cells treated with 18∶1n−9, 22∶6n−3, TTA, or no treatment. The area occupied by mitochondria was also greater in these cells. The present study shows that dietary FO reduces TAG secretion from salmon hepatocytes and that 20∶5n−3 mediates this effect.     

Chain elongation of polyunsaturated fatty acids by vascular endothelial cells: Studies with arachidonate analogues

This study has utilized radiolabeled analogues of arachidonic acid to study the substrate specificity of elongation of long-chain polyunsaturated fatty acids. Human umbilical vein endothelial cells were incubated for 2–72 hr in medium supplemented with 0.9–2.6 μM [¹⁴C]fatty acid, and cellular glycerolipids were analyzed by gas-liquid chromatography with radioactivity detection. Elongation of naturally occurring C₂₀ polyunsaturated fatty acids occurred with eicosapentaenoate (20∶5(n−3))>Mead acid (20∶3(n−9))>arachidonate (20∶4(n−6)). Chain length markedly influenced the extent of elongation of 5,8,11,14-tetraenoates (18∶4>19∶4>20∶4>21∶4); effects of initial double bond position were also observed (6,9,12,15–20∶4>4,7,10,13–20∶4. Neither 5,8,14- nor 5,11,14–20∶3 was elongated to the extent of 5,8,11–20∶3. Differences between polyunsaturated fatty acids were observed both in the initial rates and in the maximal percentages of elongation, suggesting that the content of cellular C₂₀ and C₂₂ fatty acids may represent a balance between chain elongation and retroconversion. Umbilical vein endothelial cells do not exhibit significant desaturation of either 22∶4(n−6) or 22∶5(n−3). By contrast, incubation with 5,8,11,14-[¹⁴C]18∶4(n−4) resulted in formation of both [¹⁴C]20∶5(n−4) and [¹⁴C]22∶5(n−4). The respective time courses for the appearances of [¹⁴C]22∶5(n−4) and [¹⁴C]20∶5(n−5) suggests Δ6 desaturation of [¹⁴C]22∶4(n−4) rather than Δ4 desaturation of [¹⁴C]20∶4(n−4).     

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.     

Influence of diet on conversion of14C1-linolenic acid to docosahexaenoic acid in the rat

¹⁴C₁-Linolenic acid was incorporated into lipids of hearts, livers, and carcasses of male rats. We studied the influence of diet composition on extent and distribution of radioactivity. A CHOW diet, a purified, essential fatty acid (EFA)-deficient diet, a purified control diet, and EFA-deficient diets with four fatty acid supplements were used. Supplements of 18∶2n−6, 20∶4n−6, 18∶3n−3, and 22∶6n−3 were given as single doses. Radioactivities in liver phosphatidyl ethanolamines (PE), phosphatidyl cholines, and neutral lipids were measured. The distribution of radioactivity among the fatty acids in liver phospholipids was determined. Rats on CHOW diet incorporated far less radioactivity than any other group into lipids of hearts and livers. Most of the activity in livers was recovered as 20∶5n−3 and 22∶6n−3 in all rats. In EFA-deficient rats, the radioactivity in 22∶6n−3 of liver PE was still increasing 36 hr after¹⁴C₁-linolenic acid had been administered. The n−6 supplements (18∶2n−6 and 20∶4n−6) seemed to reduce the conversion of 20∶4n−3 to 20∶5n−3 (desaturation), whereas the n−3 supplements (18∶3n−3 and 22∶6n−3) reduced the conversion of 20∶5n−3 to 22∶5n−3 (elongation). Formation of 22∶6n−3 may be controlled by 22∶6n−3 itself at the elongation of 20∶5n−3 to 22∶5n−3.     

Effects of dietary supplementation of rapeseed oil on metabolism of [1-<Superscript>14</Superscript>C]18∶1n−9, [1-<Superscript>14</Superscript>C]20∶3n−6, and [1-<Superscript>14</Superscript>C]20∶4n−3 in atlantic salmon heaptocytes

Atlantic salmon were fed fish meal-based diets supplemented with either 100% fish oil (FO) or 100% rapeseed oil (RO) from an initial weight of 85 g to a final average weight of 280 g. The effects of these diets on the capacity of Atlantic salmon hepatocytes to elogate, desaturate, and esterify [1-¹⁴C]18∶1n−9 and the immediate substrates for the Δ⁵ desaturase, [1-¹⁴C]20∶3 n−6 and [1-¹⁴C]20∶4n−3, were investigated. Radiolabeled 18∶1n−9 was mainly esterified into cellular TAG, whereas the more polyunsaturated FA, [1-¹⁴C]20∶3n−6 and [1-¹⁴C]20∶4n−3, were primarily esterified into cellular PL. More of the elongation product, [1-¹⁴C]20∶1n−9, was produced from 18∶1n−9 and more of the desaturation and elongation products, 22∶5n−6 and 22∶6n−3, were produced from [1-¹⁴C]20∶3n−6 and [1-¹⁴C]20∶4n−3, respectively, in RO hepatocytes than in FO hepatocytes. Further, we studied whether increased addition of [1-¹⁴C]18∶1n−9 to the hepatocyte culture media would affect the capacity of hepatocytes to oxidize 18∶1n−9 to acid-soluble products and CO₂. An increase in exogenous concentration of 18∶1n−9 from 7 to 100 μM resulted in a nearly twofold increase in the amount of 18∶1n−9 that was oxidized. The conversion of 20∶4n−3 and 20∶3n−6 to the longer-chain 22∶6n−3 and 22∶5n−6 was enhanced by RO feeding in Atlantic salmon hepatocytes. The increased capacity of RO hepatocytes to produce 22∶6n−3 was, however, not enought to achieve the levels found in FO hepatocytes. Our data further showed that there were no differences in the hepatocyte FA oxidation capacity and the lipid deposition of carcass and liver between the two groups.     

Effects of dietary n−3 polyunsaturated fatty acids on phospholipid composition and calcium transport in mouse cardiac sarcoplasmic reticulum

The effects of dietary n−3 and n−6 polyunsaturated fatty acids on the fatty acid composition of phospholipid, Ca⁺⁺· Mg⁺⁺ ATPase and Ca⁺⁺ transport activities of mouse sarcoplasmic reticulum were investigated. Mice were fed a 2 weight percent fat diet containing either 0.5 weight percent ethyl esters of 18∶3n−3, 20∶5n−3 or 22∶6n−3 as a source of n−3 polyusaturated fatty acid or 0.5 weight percent safflower oil as a cource of n−6 polyunsaturated fatty acid for 10 days. Olive oil (2 weight percent) was used as a control diet. Although feeding n−6 polyunsaturated fatty acid induced very little modifications of the phospholipid sarcoplasmic reticulum fatty acid composition, feeding n−3 polyunsaturated fatty acid altered it markedly. Inclusion of 18∶−3, 20∶5n−3 or 22∶6n−3 in the diet caused an accumulation of 22∶6n−3, which replaced 20∶4n−6 and 18∶2n−6 in phospholipid sarcoplasmic reticulum. The saturated fatty acids were significantly increased with a concurrent reduction of 18∶1n−9. These changes in the fatty acid composition resulted in a decrease in the values of the n−6/n−3 polyunsaturated fatty acid ratio and a decrease in the ratio of 20 carbon to 22 carbon fatty acids esterified in the phospholipid sarcoplasmic reticulum. This was associated with a decrease in Ca⁺⁺ uptake by n−3 polyunsaturated fatty acid enriched sarcoplasmic reticulum vesicles as compared with n−6 fatty acid and control diet sarcoplasmic reticulum vesicles. However, neither the affinity for Ca⁺⁺ nor the maximal velocity of ATP hydrolysis activity of Ca⁺⁺·MG⁺⁺ ATPase were altered by the different diets. The data suggest that the incorporation of 22∶6n−3 and/or the decrease of 20∶4n−6 plus 18∶2n−6 in the phospholipid sarcoplasmic reticulum may affect the membrane lipid bilayer structure and make it more permeable to Ca⁺⁺.     

Comparison of the effects of dietary alpha-linolenic,stearidonic, and eicosapentaenoic acids on production of inflammatory mediators in mice

The effects of dietary stearidonic acid (18∶4n−3) on inflammatory mediator release in whole blood and splenocytes was investigated in Balb/c mice, and the effects were compared with those of two other n−3 PUFA: α-linolenic acid (18∶3n−3) and EPA (20∶5n−3). TAG mixtures containing 10% of 18∶4n−3, 18∶3n−3, or 20∶5n−3 as the respective sole n−3 PUFA were enzymatically synthesized. Diets containing synthesized TAG mixtures were fed to Balb/c mice for 3 wk. The release of prostaglandin E₂ (PGE₂) and tumor necrosis factor (TNF) were measured in whole blood and splenocytes stimulated with lipopolysaccharide. In whole blood, the production of INF was suppressed by all dietary n−3 PUFA (18∶3n−3, 18∶4n−3, and 20∶5n−3) as compared with the control diet, which contained TAG prepared from safflower oil. PGE₂ production was not significantly changed. Differences among the n−3 PUFA (18∶3n−3), 18∶4n−3, and 20∶5n−3) were not observed. In splenocytes, PGE₂ production was suppressed by dietary n−3 PUFA, but TNF production was not. GC analysis of plasma and splenocyte FA profiles showed an increase in the levels of 20∶4n−3, 20∶5n−3, and 22∶6n−3 in mice fed the diet containing 18∶4n−3.     

CO<Subscript>2</Subscript> retention ability on alkali cation exchanged titanium silicate,ETS-10

ETS-10 was ion exchanged by various alkali cations (Li⁺, Na⁺, K⁺, Rb⁺ and Cs⁺) and the BET surface area and pore volume was exactly consistent with cationic size; that is, in the order of Li⁺ > Na⁺ > K⁺ > Rb⁺ > Cs⁺. It was observed that a single point adsorption capacity was inversely proportional to cationic size. The largest CO₂ capacity was observed for Li⁺-ETS-10 and it is attributed to greater cation–quadrupole interactions with CO₂ than larger cation. The results also suggests that as the CO₂ loading is increased, the accessibility of adsorbing CO₂ to framework basic O⁻ sites should have become difficult with the increase in cationic size due to the blocking effect by extra-framework CO₂-M⁺. The slight decrease in the slope of adsorption capacity with temperature, especially beyond 373 K for Li⁺-ETS-10 and K⁺-ETS-10 suggests that the adsorption of CO₂ on small alkali cation exchanged-ETS-10 at high temperature is somewhat associated with basic oxygen anion sites in framework due to the existence of large pore. The CO₂-TPD results show that the amount of desorbed CO₂ at higher temperature was proportionally increased due to the increased basicity of oxygen anions in framework. It also shows that the desorption temperature associated with alkali cations in extra-framework (corresponding to low temperature desorption peak) has been lowered with the increase in cationic size, indicating weak cation–quadrupole interactions with CO₂ for larger cations.     

The effect of dietary supplementation with eicosapentaenoic acid on the phospholipid and fatty acid composition of erythrocytes of marmoset

Adult male marmoset monkeys were fed eicosapentaenoic acid (20∶5n−3) as the ethyl ester in diets containing either 32% (reference diet, no added cholesterol) or 7% (atherogenic diet with 0.2% added cholesterol) linoleic acid (18∶2n−6) for 30 wk. No changes were seen in the level of phosphatidylcholine (PC) or phosphatidylethanolamine (PE) but minor changes were observed in both the sphingomyelin (SPM) and phosphatidylinositol plus phosphatidylserine (PI+PS) fractions of erythrocyte lipids. The extent of total n−3 fatty acid incorporation into membrane lipids was higher in atherogenic diets (polyunsaturated/monounsaturated/saturated (P/M/S) ratio 0.2∶0.6∶1.0) than reference diets (P/M/S ratio 1∶1∶1) and this was true for both PE (33.4±1.0%vs 24.3±1.1%) and PC (9.3±0.5%vs 4.9±0.3%). Although suitable controls for cholesterol effects were not included in the study, earlier results obtained with marmosets lead us to believe such effects were probably small. Regardless of basic diet (atherogenic, reference), 20∶5n−3 was preferentially incorporated into PE (10.8±0.2%, 6.0±0.02%) while smaller amounts were incorporated into PC (6.9±0.4%, 3.2±0.2%). The major n−3 polyunsaturated fatty acid found in PE in response to dietary 20∶5n−3 was the elongation metabolite 22∶5n−3 in both the atherogenic (17.7±0.7%) and reference (14.3±1.0%) dietary groups; 22∶6n−3 levels were less affected by diet (4.7±0.3% and 3.9±0.2%, respectively). The results can be interpreted to indicate an inverse relationship between the amount of dietary 18∶2n−6 and incorporation of 20∶5n−3 into erythrocyte membrane phospholipids regardless of whether the major dietary n−3 fatty acid was α-linolenate (18∶3n−3) or 20∶5n−3. This interpretation is supported by theoretical calculations.