bluuebunny/crossref_metadata_2025 huggingface dataset:
Rat vitamin E status and heart lipid peroxidation: Effect of dietary α‐Linolenic acid and marine n−3 fatty acids
Authors: Javouhey‐Donzel Anne , Guenot Lucien , Maupoil Véronique , Rochette Luc , Rocquelin Gérard
Published: November 2006

Three groups of sixteen male rats each were fed semipurified diets containing 15% by weight of lipid for a period of 4 wk. The diets contained the same amount of polyunsaturated fatty acids (PUFA) (20% of total fatty acids) and saturated fatty acids (19% of total fatty acids). Dietary PUFA were represented exclusively by linoleic acid (18∶2 diet), or 10% linoleic acid and 10% linolenic acid (18∶3 diet), or 10% linoleic acid and 10% long‐chain n−3 fatty acids (LCn−3 diet). The overall amount of vitamin E was similar in the three diets,i.e, 140, 133 and 129 mg/kg diet, respectively. Following appropriate extraction, tocopherol levels in heart, liver, brain, adipose tissue (AT) and plasma were measured by high‐performance liquid chromatography. The level of vitamin E in the heart decreased with n−3 PUFA diets, most markedly with LCn−3 PUFA. Liver and AT vitamin E contents also decreased with n−3 PUFA diets when expressed as μg/mg total lipids and μg/mg phospholipids, respectively. Total plasma vitamin E was lower in rats fed the LCn−3 diet, but there was no significant difference when expressed as μg/mg total lipids. Brain vitamin E was not affected by the various diets.In vitro cardiac lipid peroxidation was quantified by the thiobarbituric acid reactive substances (TBARS) test. Heart homogenates were incubated at 37°C for 15 and 30 min in both the absence (uninduced) or presence (induced) of a free radical generating system (1 mM xanthine, 0.1 IU per mL xanthine oxidase, 0.2 mM/0.4 mM Fe/ethylenediaminetetraacetic acid). TBARS release was time‐independent but significantly higher when LCn−3 fatty acids were fed to rats in either the uninduced or induced system. The study demonstrated that n−3 PUFA diets can influence vitamin E status of rats even in short‐term experiments and can change the susceptibility of the heart toin vitro lipid peroxidation.
A multivariate optimization of triacylglycerol analysis by high‐performance liquid chromatography
Authors: Bergqvist Magnus H. J. , Kaufmann Peter
Published: November 2006

In the present study we have used statistical experimental design and multivariate optimization to formally optimize a reversed‐phase high‐performance liquid chromatography method for the analysis of triacylglycerol molecular species of natural oils. The optimal conditions found were, on an octadecylsilan‐column, from acetonitrile/isooctane (90∶10, vol/vol) to acetonitrile/ethanol/isooctane (40∶35∶25, by vol), at a column temperature of 50°C and a flowrate of 1.5mL/min using a negative exponential gradient profile. Several examples of separations of natural seed and animal oils,i.e., soybean oil, rapeseed oil, palm oil, linseed oil, tallow and fish oil, are given. A version of the equivalent carbon number concept, utilizing the Snyder polarity index, was used to identity the molecular species.
Regio‐ and stereochemical analysis of trihydroxyoctadecenoic acids derived from linoleic acid 9‐ and 13‐hydroperoxides
Authors: Hamberg Mats
Published: November 2006

The methyl esters of 9S,10S,13R‐trihydroxy‐11E‐octadecenoic acid, 9S,10R,13S‐trihydroxy‐11E‐octadecenoic acid, and 9S,10R,13R‐trihydroxy‐11E‐octadecenoic acid were prepared from 9S‐hydroperoxy‐10E,12Z‐octadecadienoic acidvia the epoxy alcohols methyl 10R,11R‐epoxy‐9S‐hydroxy‐12Z‐octadecenoate and methyl 10S,11S‐epoxy‐9S‐hydroxy‐12Z‐octadecenoate. The trihydroxyesters, as well as four stereoisomeric methyl 9,12,13‐trihydroxy‐10E‐octadecenoates earlier prepared [Hamberg, M.,Chem. Phys. Lipids 43, 55–67 (1987)], were characterized by thin‐layer chromatography, gas‐liquid chromatography, mass spectrometry, and by chemical degradation. Availability of these chemically defined trihydroxyoctadecenoates made it possible to design a method for regio‐ and stereochemical analysis of 9,10,13‐ and 9,12,13‐trihydroxyoctadecenoic acids obtained from various sources. Application of the method revealed that the mixture of 9,10,13‐ and 9,12,13‐trihydroxyoctadecenoic acids formed during autoxidation of linoleic acid in aqueous medium contained comparable amounts of the sixteen possible regio‐ and stereoisomers. Furthermore, hydrolysis of the allylic epoxy alcohol, methyl 9S,10R‐epoxy‐13S‐hydroxy‐11E‐octadecenoate, yielded a major trihydroxyoctadecenoate,i.e., methyl 9S,10S,13S‐trihydroxyl‐11E‐octadecenoate, together with smaller amounts of methyl 9S,10R,13S‐trihydroxy‐11E‐octadecenoate, methyl 9S,12R,13S‐trihydroxy‐10E‐octadecenoate, and methyl 9S,12S,13S‐trihydroxy‐10E‐octadecenoate.
Comparative specificity of plasma lecithin: Cholesterol acyltransferase from ten animal species
Authors: Grove Douglas , Pownall Henry J.
Published: November 2006

The molecular specificities of plasma lecithin:cholesterol acyltransferase (LCAT) from ten animal species have been compared. Using a reassembled high density lipoprotein containing a mixture of phosphatidylcholines, the relative rates of liberation of different species of cholesteryl ester were measured. All but two species of LCAT clustered according to one of three patterns of substrate specificity. The LCAT from six species, including human, did not transfer highly polyunsaturated fatty acyl chains. In addition, human LCAT transesterified saturated fatty acyl chains more effectively than unsaturated fatty acyl chains. We conclude that the structures of the active sites of the enzymes differ, and that this may be related to size constraints that prevent efficient binding of large bulky phosphatidylcholines.
Activation and solubilization by triton X‐100 of membrane‐bound phospholipase D of rat brain
Authors: Kanoh Hiroyuki , Kanaho Yasunori , Nozawa Yoshinori
Published: November 2006

In the present study, we examined the ability of detergents to stimulate and solubilize phospholipase D (PLD) of a particulate fraction of rat brain. PLD activity was assayed by measuring the [3H]choline produced from the exogenous substrate dipalmitoyl phosphatidyl[3H]choline (dipalmitoyl [3H]PC). In the absence of detergents, PLD activity was not detectable. Of the detergents examined, Triton X‐100 was found to markedly enhance PLD activity, whereas other detergents including sodium doexycholate, sodium cholate, CHAPS and Lubrol‐PX caused only a small, if any increase in activity. Enhancement by Triton X‐100 was maximal at 0.1–0.2% (w/v) and decreased at higher concentrations. The optimal pH was 7.1–7.3. Both Ca2+ and Mg2+ inhibited enzyme activity stimulated by Triton X‐100 in a concentration‐dependent manner. Triton X‐100 effectively solubilized PLD from the particulate fraction of rat brain; more than 70% of the activity of the particulate fraction was extracted by 0.5–1.0% (w/v) Triton X‐100. Furthermore, when the PLD activities in brains of three different species (rat, rabbit and bovine) were measured under optimal conditions, the activities were found to differ greatly. PLD activity was highest in rabbit brain, followed by rat and bovine brains; the activity in bovine brain was extremely low compared to the activities in rat and rabbit brains. We conclude that Triton X‐100 is potentially useful for the purification of PLD and that rabbit and rat brains are the preferred sources.
Lipid composition and peroxide levels of mucosal cells in the rat large intestine in relation to dietary fat
Authors: Turini M. E. , Thomson A. B. R. , Clandinin M. T.
Published: November 2006

To examine whether dietary fat alters membrane lipid composition and peroxidation of polyunsaturated fatty acids in “non‐proliferative” and “proliferative” cells in the large intestine, Sprague‐Dawley rats were fed diets providing a polyunsaturated‐to‐saturated fatty acid ratio of 1.2 or 0.3 at a high or low level of fat intake for a 25‐day period. Cell populations were isolated and the effect of dietary fat on membrane polyunsaturated fatty acid content and peroxide levels was determined. Neither fat level nor fatty acid composition of diet influenced total cholesterol, total phospholipids, and percentage of phospholipid classes in membrane phospholipids. Feeding the high fat and/or high polyunsaturated‐to‐saturated fatty acid ratio diet increased polyunsaturated fatty acid content of mucosal cell phospholipids. Increase in polyunsaturated fatty acid content was paralleled by a decrease in the monounsaturated fatty acid content of mucosal cell phospholipids. Membrane content of total saturated fatty acids was not significantly affected by diet. Variation in phospholipid fatty acid composition between “non‐proliferative” and ”proliferative” cells was observed. Lipid peroxide levels in mucosal cell lipid fractions were altered by dietary fat treatment. Animals fed high fat diets, compared to groups fed low fat diets, exhibited higher membrane peroxide levels when results are expressed as nmol/mg protein. Higher peroxide levels were observed in mucosal cells for rats fed high polyunsaturated‐to‐saturated fatty acid ratio diets when results were expressed per nmol of phospholipid. It is concluded that changes in fat level and fatty acid composition of the diet alters the mucosal cell membrane lipid composition in the rat large intestine and influences susceptibility of mucosal cell lipid to peroxidation. Further research is required to delineate which dietary factors—fat level, polyunsaturated‐to‐saturated fatty acid ratio, or both—have a primary influence on the degree of lipid peroxidation.
Effect of dietary fat on diabetes‐induced changes in liver microsomal fatty acid composition and glucose‐6‐phosphatase activity in rats
Authors: Venkatraman J. T. , Pehowich D. , Singh B. , Rajotte R. V. , Thomson A. B. R. , Clandinin M. T.
Published: November 2006

Experimental diabetes may manifest itself in a defect in liver microsomal fatty acid desaturation and increased activity of glucose‐6‐phosphatase (G‐6‐Pase). The present study was designed to determine whether these changes could be normalized by a change in the dietary fat consumed. Control and streptozotocin‐induced diabetic rats were fed nutritionally adequate diets which varied in fatty acid composition. Fatty acid analysis of liver microsomal phospholipids revealed that non‐diabetic control animals fed saturated fat (beef tallow) or a diet high in ω3 fatty acids (fish oil) exhibited a significantly higher level of 18∶2ω6 and a lower level of 20∶4ω6 in the phosphatidylcholine and phosphatidylethanolamine fractions compared with diabetic animals. Control and diabetic animals fed the high linoleic acid diet had similar levels of 18∶2ω6 in the microsomal phosphatidylcholine and phosphatidylserine fractions. Microsomal G‐6‐Pase activity was higher in diabetic than in control animals. Activity of G‐6‐Pase was lower in microsomes of control animals fed the soybean oil or the fish oil diet, but was not significantly reduced in diabetic animals fed high polyunsaturated fats. Blood glucose levels were similar in control groups fed the different diets, but the plasma hemoglobin A1c level was lower in diabetic animals fed the soybean oil diet. Cholesterol and triglyceride levels were lower in diabetic animals fed the fish oil‐based diet. The results suggest that dietary fat manipulation has the potential to change at least some of the abnormalities in the microsomal membrane in experimental diabetes.
The effect of dietary linoleic acid on the fatty acid composition of individual phospholipid and lipoxygenase products from gills and leucocytes of Atlantic salmon (Salmo salar)
Authors: Gordon Bell J. , Raynard Robert S. , Sargent John R.
Published: November 2006

Diets containing either fish oil or sunflower oil, both of which supplied the minimum required level of n−3 fatty acids, were given to Atlantic salmon (Salmo salar) postsmolts for a period of 16 weeks. In fish fed sunflower oil, the phospholipids of gills showed increased 18∶2n−6 (2–13‐fold), 20∶2n−6 (4.5–12‐fold) and 20∶−6 (2–8‐fold). In addition, phosphatidylethanolamine had increased 20∶4n−6 (1.5‐fold). Changes of a similar magnitude were observed in the phospholipids of blood leucocytes except that, in addition, 20∶4n−6 was elevated in phosphatidylserine (1.7‐fold) and phosphatidylinositol (1.4‐fold). Both tissues showed a general decrease in phospholipid 20∶5n−3 (up to 3‐fold), which caused an increase in 20∶4n−6/20∶5n−3 ratio (1.3–6‐fold). The elongation and desaturation products of 20∶4n−6, 22∶4n−6 and 22∶5n−6 were not increased as a result of feeding sunflower oil. When isolated gill cells were stimulated with the calcium ionophore A23187, 12‐hydroxy‐8,10,14,17‐eicosapentaenoic acid (12‐HEPE) was the major lipoxygenase product from salmon given fish oil. 12‐HEPE was significantly reduced in salmon given sunflower oil. When stimulated with A23187, the lipoxygenase products derived from whole blood of fish given sunflower oil showed decreased levels of leukotriene B5, 12‐HEPE and 12‐hydroxy‐5,8,10,14‐eicosatetraenoic acid.
Effect of chronic glucagon administration on lipoprotein composition in normally fed, fasted and cholesterol‐fed rats
Authors: Guettet Catherine , Rostaqui Najmuddin , Mathe Denis , Lecuyer Bernard , Navarro Nicole , Jacotot Bernard
Published: November 2006

Male adult Wistar rats received daily (at 9 a.m. and 5 p.m.) 10 μg of zinc‐protamine glucagon by subcutaneous injection for 8 days. Plasma cholesterol levels were decreased by 36% in fed rats, 33% in cholesterol‐fed rats and by 55% in fasted rats. Lipoproteins were separated into 22 fractions by ultracentrifugation using a density gradient. Glucagon administration decreased the cholesterol content in all lipoproteins except low density lipoprotein (LDL1) (1.006–1.040) and very low density lipoprotein (VLDL) from cholesterol‐fed rats. The main decrease (−57 to −81%) was observed in 1.050–1.100 g/mL lipoproteins (LDL2 and HDL2), which contained a large amount of apo E, while HDL3 cholesterol was not affected. Triacylglycerol levels were decreased only in chylomicrons and VLDL (−70%) of fed and cholesterol‐fed rats, while plasma and lipoprotein triacylglycerol levels were not changed in fasted rats treated with glucagon. In normally fed rats glucagon administration increased by 42% the fractional catabolic rate of [125I]HDL2 while the absolute catabolic rate appeared to be unchanged. Glucagon seems to be a potent hypolipidemic agent affecting mainly the apo E‐rich lipoproteins. Its chronic administration limits lipoprotein accumulation which occurs upon cholesterol feeding.
The effect of dietary zinc deficiency on the lipid composition of the rat erythrocyte membrane
Authors: Driscoll Eric R. , Bettger William J.
Published: November 2006

The effect of dietary zinc deficiency in the rat on the lipid composition of the erythrocyte membrane was determined. Weanling male Wistar rats were fed an egg whitebased diet containing <1.0 mg Zn/kg dietad libitum. Control rats were either pair‐fed orad libitum‐fed the basal diet suppelemented with 100 mg Zn/kg diet. A zinc refed group was fed the −Zn diet until day 18 and then pairfed the +Zn diet until day 21. The voluntary feed restriction associated with dietary zinc deficiency resulted in erythrocyte membranes that had depressed phospholipid/protein and elevated cholesterol/phospholipid ratios. Similarly, all feed restricted groups had elevated 22‐carbon n−3 polyunsaturated fatty acid (PUFA) and depressed 22‐carbon n−6 PUFA concentrations in alkenylacyl and diacyl glycerophosphoethanolamine, phosphatidylserine and phosphatidylcholine; they also had depressed 24∶2n−6 levels in sphingomyelin. The relative concentrations of phospholipids in the membrane was similar between −Zn and +Zn (ad libitum) groups; however, the −Zn group had significantly less phosphatidylserine relative to +Zn (pair‐fed) controls.
Effect of dietary α‐linolenate/linoleate balance on endotoxin‐induced hepatitis in mice
Authors: Watanabe Shiro , Okuyama Harumi
Published: November 2006

Mice were fed diets with three different ratios of α‐linolenate (18∶−3) to linoleate (18∶2n−6), and the severity of hepatitis during endotoxic shock was compared. Dietary enrichment with α‐linolenate increased the severity of hepatitis and the mortality induced by lipopolysaccharide (LPS) in combination with D‐galactosamine (GalN). Differences in the dietary α‐linolenate/linoleate balance were mainly reflected in the levels of arachidonate and eicosapenatenoate in liver phospholipids. Pretreatement of mice with indomethacin was found to also enhance the severity of GalN/LPS‐hepatitis. This indicated that cyclooxygenase products of arachidonate may suppress the development of GalN/LPS‐hepatitis. The enhancement by high α‐linolenate diets was not observed when a lethal dose of LPS in the absence of GalN was given. Our results indicate that there are pathophysiological conditions of endotoxin‐induced hepatitis under which cyclooxygenase products of arachidonate play protective roles.
Headspace gas chromatography of volatile lipid peroxidation products from human red blood cell membranes
Authors: Frankel E. N. , Tappel A. L.
Published: November 2006

An improved headspace capillary gas chromatographic (GC) method was developed to measure the oxidative susceptibility of human red blood cell (RBC) membranes. This method analyzed volatile peroxidation products of both n−6 (hexanal and pentane) and n−3 (propanal) polyunsaturated fatty acids. Oxidative susceptibility tests were standardized by incubating in a sealed 10‐mL headspace bottle 0.25 or 1 mL of human RBC membrane in 40 mM phosphate buffer for 1 hr at 37°C with a mixture of Fe++, ascorbic acid and H2O2. Sodium dodecyl sulfate increased significantly the amount of hexanal measured by headspace GC. By this standard headspace method, in one series of red blood cell membranes (RBCM) samples a four‐fold variation in oxidative susceptibility was observed in RBCM from blood freshly drawn from six healthy subjects. In another series of RBCM samples a sixteen‐fold variation in oxidative susceptibility was noted in frozen RBCM from blood freshly drawn from five healthy subjects. Correlation between hexanal formation and polyunsaturated fatty acids (PUFA) depletion provided good evidence that under these standard conditions hexanal is exclusively derived from the oxidation of arachidonic acid. Hydroperoxides of arachidonic acid are more readily formed and decomposed than those of linoleic acid in the presence of Fe++, ascorbic acid and H2O2 to produce hexanal as the main product that can be readily analyzed by headspace GC. This method may provide a useful tool to study susceptibility toward lipid peroxidative damage in human RBC membranes.
Transformation of fatty acid hydroperoxides by alkali and characterization of products
Authors: Gardner Harold W. , Simpson Thomas D. , Hamberg Mats
Published: November 2006

It has previously been determined that (13S,9Z,11E)‐13‐hydroperoxy‐9,11‐octadecadienoic acid was mainly converted into (13S,9Z,11E)‐13‐hydroxy‐9,11‐octadecadienoic acid by 5 N KHO with preservation of the stereochemistry of the reactant [Simpson, T.D., and Gardner, H.W. (1993)Lipids 28, 325–330]. In addition, about 20–25% of the reactant was converted into several unknown by‐products. In the present work it was confirmed that the stereochemistry was conserved during the hydroperoxy‐diene to hydroxydiene transformation, but also, novel by‐products were identified. It was found that after only 40 min reaction (9Z)‐13‐oxo‐trans‐11,12‐epoxy‐9‐octadecenoic acid accumulated to as much as 7% of the total. Later, (9Z)‐13‐oxo‐trans‐11,12‐epoxy‐9‐octadecenoic acid began to disappear, and several other compounds continued to increase in yield. Two of these compounds, 2‐butyl‐3,5‐tetradecadienedioic acid and 2‐butyl‐4‐hydroxy‐5‐tetradecenedioic acid, were shown to originate from (9Z)‐13‐oxo‐trans‐11,12‐epoxy‐9‐octadecenoic acid, and they accumulated up to 2–3% each after 4 to 6 h. Some other lesser products included 11‐hydroxy‐9,12‐heptadecadienoic acid, 3‐hydroxy‐4‐tridecenedioic acid, 13‐oxo‐9,11‐octadecadienoic acid and 12,13‐epoxy‐11‐hydroxy‐9‐octadecenoic acid. Except for the latter two, most or all of the compounds could have originated from Favorskii rearrangement of the early product, (9Z)‐13‐oxo‐trans‐11,12‐epoxy‐9‐octadecenoic acid, through a cyclopropanone intermediate.
Dynamics of iron‐ascorbate‐induced lipid peroxidation in charged and uncharged phospholipid vesicles
Authors: Fukuzawa Kenji , Seko Takayuki , Minami Kazushi , Terao Junji
Published: November 2006

Peroxidation of egg yolk phosphatidylcholine (egg PC) liposomes was induced by addition of ascorbic acid (AsA) and Fe(II) in the presence of a trace of autoxidized egg PC (PC−OOH), but not in the absence of PC−OOH. PC−OOH was degraded upon addition of AsA and Fe(II) but not of either one alone. The results suggest that PC−OOH is necessary to initiate lipid peroxidation by AsA/Fe(II). AsA oxidation in the bulk water phase was also associated with an increase in lipid peroxidation by AsA/Fe(II) in the presence of PC−OOH, but not in the absence of PC−OOH. Furthermore, the spin probe 12‐NS [12‐(N‐oxyl‐4,4′‐dimethyloxazolidin‐2‐yl)stearic acid], which labels the hydrophobic region of dimyristoyl phosphatidylcholine (DMPC) liposomal membranes, was degraded upon addition of AsA and Fe(II) in the presence of PC−OOH, but not in the absence of PC−OOH. These results indicate that the “induction message” that is associated with decreases of PC−OOH and AsA in the initiation step of lipid peroxidation must be transferred from the membrane surface to the inner hydrophobic membrane region. AsA in the bulk phase was oxidized faster and more extensively upon its addition together with Fe(II) to egg PC liposomes than to DMPC liposomes, though the initial content of PC−OOH in the former was 5–10 times lower than in the latter. This suggests that, in egg PC liposomes, the OOH‐groups of new PC−OOH generated in the inner membrane regions must become accessible from the surface, enabling reaction with AsA/Fe(II) which in turn would result in an extensive decrease in AsA. By contrast, in DMPC liposomes, that do not generate PC−OOH, AsA is only oxidized slightly in connection with the degradation of the PC−OOH initially present. The effect of surface charges on the membrane surface was also studied to obtain further information on the initiation step of lipid peroxidation. The rate of lipid peroxidation by AsA/Fe(II) or Fe(III) decreased in the order, egg PC liposomes ≫negatively charged egg PC liposomes containing dicetylphosphate>positively charge egg PC liposomes containing stearylamine. The rate of associated AsA oxidation was in the order, egg PC liposomes≫egg PC/stearylamine liposomes>egg PC/dicetylphosphate liposomes. However, in DMPC liposomes that do not generate PC−OOH, the rates of AsA oxidation associated with the reductive cleavage of PC−OOH by AsA/Fe(II) and coupled with the reduction of Fe(III) to Fe(II) were in the order, DMPC liposomes =DMPC/stearylamine liposomes≫DMPC/dicetylphosphate liposomes. These differences in the rates of lipid peroxidation, depending on differences in membrane charge, are discussed in relation to two properties of AsA: (i) its antioxidant property through trapping of lipid radicals and (ii) its prooxidant properties (a) by being an effective iron chelator thus altering the reactivity of iron with oxygen and peroxides and (b) by being an iron reductant and providing a source of Fe(II).
Role of lipid structure in the activation of phospholipase A2 by peroxidized phospholipids
Authors: McLean Larry R. , Hagaman Karen A. , Davidson W. Sean
Published: November 2006

The time course of hydrolysis of a mixed phospholipid substrate containing bovine liver 1,2‐diacyl‐sn‐glycero‐3‐phosphocholine (PC) and 1,2‐diacyl‐sn‐glycero‐3‐phosphoethanolamine (PE) catalyzed byCrotalus adamanteus phospholipase A2 was measured before and after peroxidation of the lipid substrate. The rate of hydrolysis was increased after peroxidation by an iron/adenosine diphosphate (ADP) system; the presence of iron/ADP in the assay had a minimal inhibitory effect. The rate of lipid hydrolysis was also increased after the substrate was peroxidized by heat and O2. Similarly, peroxidation increased the rate of hydrolysis of soy PC liposomes that did not contain PE. In order to minimize interfacial factors that may result in an increase in rate, the lipids were solubilized in Triton X‐100. In mixtures of Triton with soy PC in the absence of PE, peroxidation dramatically increased the rate of lipid hydrolysis. In addition, the rate of hydrolysis of the unoxidizable lipid 1‐palmitoyl‐2‐[1‐14C]oleoyl PC incorporated into PC/PE liposomes was unaffected by peroxidation of the host lipid. These data are consistent with the notions that the increase in rate of hydrolysis of peroxidized PC substrates catalyzed by phospholipase A2 is due largely to a preference for peroxidized phospholipid molecules as substrates and that peroxidation of host lipid does not significantly increase the rate of hydrolysis of nonoxidized lipids.
Effect of age and α‐linolenic acid deficiency on Δ6 desaturase activity and liver lipids in rats
Authors: Dinh T. K. L. , Bourre J. M. , Durand G.
Published: November 2006

The combined effects of age and of diet deficient in n−3 fatty acids on Δ6 desaturation of linoleic acid and on lipid fatty acid composition were studied in the liver of the rat at 2, 6, 12, 18 and 24 mon of age. The profiles of Δ6 desaturase activity and fatty acid composition were studied in the deficient rats refed, at these different ages either with 18∶3n−3 (mixture of peanut and rapeseed oils) or with 20∶5n−3+22∶6n−3 (fish oil) diets for 2, 4, 8 or 12 wk. Results showed that the liver Δ6 desaturation activity in the control rats remained high at 2 and 6 mon, decreased by 30% from 6 to 12 mon, and then remained stable from 12 to 24 mon. In the deficient rats, this activity remained high during the entire period studied. Thus, the profile of liver Δ6 desaturase activity after puberty was not related to age only; it also depended on the polyunsaturated fatty acid (PUFA) n−6 and n−3 balance in the diet. In the controls, in parallel with the Δ6 desaturase activity, PUFA metabolism could be divided into three periods: a “young” period, and “old age” period, separated by a period of transition between 6 and 12 mon. Recovery from PUFA n−3 deficiency occurred at all ages but in a different manner depending on whether the rats were “young” or “old”. Recovery was faster if long‐chain n−3 PUFA rather than α‐linolenic acid were supplied in the diet.
Differential utilization of eicosapentaenoic acid and docosahexaenoic acid in human plasma
Authors: Hodge Jason , Sanders Kerrie , Sinclair Andrew J.
Published: November 2006

It has recently been shown that the ω3 fatty acid status in humans can be predicted by the concentration of eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids in plasma phospholipids [Bjerve, K.S., Brubakk, A.M., Fougner, K.J., Johnsen, H., Midjthell, K., and Vik, T. (1993)Am. J. Clin. Nutr., in press]. In countries with low intake of ω3 fatty acids, the level of EPA in plasma phospholipids is often only about one‐fifth the concentration of DHA. The purpose of this study was to investigate whether this difference in the concentration of these two fatty acids was due to a selective loss of EPA relative to DHA or to a lower dietary intake of EPA. Seven female volunteers ingested four grams of MaxEPA daily for 2 wk and in the following 4 wk they ate a diet almost completely devoid of the long‐chain ω3 fatty acids. The concentrations of the ω3 fatty acids in the plasma cholesteryl esters, triglycerides and phospholipids and the high density lipoprotein phospholipids were examined at weekly intervals throughout the study. There was a more rapid rise in the concentration of EPA than in DHA levels in the supplementation period in all lipid fractions, but there was a disproportionate rise in DHA relative to EPA in the plasma lipids compared with the ratio in the supplement. In the depletion phase there was a rapid disappearance of EPA from all fractions, such that pre‐trial levels were reached by one week post‐supplementation. The disappearance of DHA was slower, particularly for the plasma phospholipids: at 4 wk post‐supplementation, the DHA concentration in this fraction was still 40% above the pre‐trial value. It is suggested that the low plasma EPA values relative to DHA are the result of increased β‐oxidation of EPA and/or low dietary intake, rather than a rapid conversion of EPA to DHA. One practical result of this experiment is that, compared with DHA, the maintenance of increased EPA levels in plasma (and therefore tissues) would require constant inputs of EPA due to its more rapid loss from the plasma.
Dietary α‐linolenic acid alters tissue fatty acid composition, but not blood lipids, lipoproteins or coagulation status in humans
Authors: Kelley Darshan S. , Nelson Gary J. , Love James E. , Branch Leslie B. , Taylor Peter C. , Schmidt Perla C. , Mackey Bruce E. , Iacono James M.
Published: November 2006

We examined the effect of dietary α‐linolenic acid (ALA) on the indices of lipid and coagulation status and on the fatty acid composition of serum and peripheral blood mononuclear cell (PBMNC) lipids in ten healthy men (age 21–37 yr) who consumed all their meals at the Western Human Nutrition Research Center for 126 d. There was a stabilization period of 14 d at the start when all 10 subjects consumed the basal diet (BD) containing 23.4 energy percent (en%) fat and two intervention periods of 56 d each. During the first intervention period, 5 subjects consumed the BD containing 23.4 en% fat, and 5 subjects consumed a diet providing 6.3% calories from α‐linolenic acid [flaxseed oil (FSO) diet containing 28.8 en% fat]. Diets were crossed over between the two groups during the second intervention period. Feeding the FSO diet did not nignificantly alter serum triglycerides, cholesterol, highdensity lipoproteins, low‐density lipoproteins, apoprotein A‐I and apoprotein B when compared to the corresponding values in the subjects fed the BD, nor was there any effect of the FSO diet on the bleeding time, prothrombin time and partial prothrombin time for these subjects. Feeding the ALA‐containing diet did cause a significant increase in ALA concentration in serum (P<0.001) and PBMNC lipids (P<0.05). It also caused a significant increase (P<0.05) in the eicosapentaenoic and docosapentaenoic acid contents of PBMNC lipids, and a decrease (P<0.01) in linoleic and eicosatrienoic acid contents of serum lipids. Thus, dietary ALA, fed for 56 d at 6.3% of calories, had no effect on plasma triglyceride or very low density lipoprotein levels or the common risk factors associated with atherosclerosis, although these parameters have been reported by others to be influenced by fatty acids, such as palmitic or linoleic acids, in the diet. Dietary ALA did significantly alter the fatty acid composition of plasma and PBMNC.
Cholesterol synthesis and esterification in isolated enterocytes: Regulation by cholesterol and cholestyramine feeding
Authors: Iglesias José , Gonzalez‐Pacanowska Dolores , Marco Carmen , Garcia‐Peregrin Eduardo
Published: November 2006

The purpose of the present study was to investigate the physiological control of the main regulatory enzymes of cholesterol metabolism in isolated enterocytes obtained from chick duodenum, jejunum and ileum. Cholesterol feeding resulted in an inhibition of 3‐hydroxy‐3‐methylglutaryl‐CoA reductase and mevalonate 5‐pyrophosphate decarboxylase, while cholestyramine feeding increased reductase activity in all the regions studied and decarboxylase activity only in duodenum. Cholesterol feeding markedly increased acyl‐CoA:cholesterol acyltransferase, but the effects of cholestyramine were less clear. The effects on transferase activity cannot be due to differences in the availability of acyl‐CoA as exogenous substrate as no significant differences were found in acyl‐CoA hydrolase activity after any of the dietary treatments. The effects of cholesterol feeding were related to changes in the cholesterol content of epithelial cells, whereas in the case of cholestyramine this relationship was less apparent.
2‐alkenyl‐4,4‐dimethyloxazolines as derivatives for the structural elucidation of iosmeric unsaturated fatty acids
Authors: Luthria Devanand L. , Sprecher Howard
Published: November 2006

Several types of unsaturated fatty acid methyl esters were converted into 4,4‐dimethyloxazoline (DMOX) derivatives and analyzed by mass spectrometry to further evaluate the feasibility of using this derivative for locating the positions of double bonds in isomeric fatty acids. Five isomeric 20‐carbon tetraenoic acids were analyzed in which the fourcis double bonds were systematically moved from the 4,7,10,13‐ to the 8,11,14,17‐positions. It was possible to locate the positions of all four double bonds in the 7,10,13,16‐ and 8,11,14,17‐isomers by appropriate ions differing by 12 atomic mass units. In a similar way the three terminal double bonds in the 4,7,10,13‐, 5,8,11,14‐ and 6,9,12,15‐isomers could be assigned. Odd‐numbered ions atm/z 139, 153 and 167 which are accompanied by an even mass ion at 138, 152 and 166, respectively, are diagnostic for DMOX derivatives of acids with their first double bond, respectively, at positions 4, 5 and 6. It was thus possible to assign the location of all four double bonds in these three isomers. A comparison of the spectra of the DMOX derivatives of 17,17,18,18‐d4vs. 9,10,12,13‐d4 linoleic acid suggests that double bonds preferentially migrate toward the polar end of the molecule prior to fragmentation. The merit of using DMOX derivatives to locate double‐bond positions in mono‐ and dicarboxylic acids, produced during β‐oxidation of polyunsaturated fatty acids, was evaluated. The spectra of 3‐cis‐ and 4‐cis‐decenoic acids differ as do the spectra of 8‐carbon dicarboxylic acids with their double bonds at positions 3 and 4.
Time response of cholesterol synthesis inhibition by compactin‐related compunds.In vitro quantitation of the “Escape Phenomenon”
Authors: Sviridov Dmitri D. , Endo Akira , Pavlov Michael Y. , Repin Vadim S.
Published: November 2006

The time course of the inhibition of cholesterol synthesis by low and high doses of mevinolin and monacolin X were studied in normal human skin fibroblasts, fibroblasts without low density lipoprotein receptor and HepG2 hepatome cells. Low doses of the inhibitors (0.2 ng/mL) caused a sharp decrease in the rate of cholesterol synthesis during the firt 2–3 h, which gradually increased to about 40% during the next 6 h. Further incubation led to a decrease or stabilization of the cholesterol synthesis rate. High doses of the drugs (100 mg/mL) strongly inhibited cholesterol synthesis during the first 2–3 h, followed by a moderate increase during the next 20 h. No drug or tissue selectivity was observed.
Oxidative modification of triglyceride‐rich lipoproteins in hypertriglyceridemic rats following magnesium deficiency
Authors: Gueux Elyett , Cubizolles Christine , Bussière Laurence , Mazur Andrzej , Rayssiguier Yves
Published: November 2006

Hypertriglyceridemia observed in magnesium (Mg)‐deficient rats was associated with a significant increase in the very low‐density lipoprotein (VLDL) plus low‐density lipoprotein (LDL) fractions. The results fromin vitro copper‐induced lipid peroxidation, expressed in terms of conjugated dienes and thiobarbituric acid reactive substances content, showed that VLDL+LDL particles from Mg‐deficient rats were more susceptible to oxidative damage than lipoproteins from control rats. These results suggest that the mechanism responsible for the atherogenicity and tissue damage characteristic of Mg deficiency may be mediated by an increased susceptibllity of triglyceride‐rich lipoproteins to peroxidation in hypertriglyceridemic animals.
Trp89 in the lid ofHumicola lanuginosa lipase is important for efficient hydrolysis of tributyrin
Authors: Holmquist Mats , Martinelle Mats , Clausen Ib Groth , Patkar Shamkant , Svendsen Allan , Hult Karl
Published: November 2006

To determine whether Trp89 located in the lid of the lipase (EC 3.1.1.3) fromHumicola lanuginosa is important for the catalytic property of the enzyme, site‐directed mutagenesis at Trp89 was carried out. The kinetic properties of wild type and mutated enzymes were studied with tributyrin as substrate. Lipase variants in which Trp89 was changed to Phe, Leu, Gly or Glu all showed less than 14% of the activity compared to that of the wild type lipase. The Trp89Glu mutant was the least active with only 1% of the activity seen with the wild type enzyme. All Trp mutants had the same binding affinity to the tributyrin substrate interface as did the wild type enzyme. Wild type lipase showed saturation kinetics against tributyrin when activities were measured with mixed emulsions containing different proportions of tributyrin and the nonionic alkyl polyoxyethylene ether surfactant, Triton DF‐16. Wild type enzyme showed a Vmax=6000±300 mmol·min−1·g−1 and an apparent Km=16±2% (vol/vol) for tributyrin in Triton DF‐16, while the mutants did not show saturation kinetics in an identical assay. The apparent Km for tributyrin in Triton DF‐16 was increased as the result of replacing Trp89 with other residues (Phe, Leu, Gly or Glu). The activities of all mutants were more sensitive to the presence of Triton DF‐16 in the tributyrin substrate than was wild type lipase. The activity of the Trp89Glu mutant was decreased to 50% in the presence of 2 vol% Triton DF‐16 compared to the activity seen with pure tributyrin as substrate. Wild type lipase and all mutants except Trp89Glu had the same affinity for the substrate interface formed by 15.6 vol% tributyrin in Triton DF‐16. The Trp89Glu mutant showed a lower affinity than all the other lipase variants for the interface of 15.6 vol% tributyrin in Triton DF‐16. The study showed that Trp89 located in the lid ofH. lanuginosa lipase is important for the efficient hydrolysis of tributyrin and that this residue plays a role in the catalytic steps after adsorption of the lipase to the substrate interface.
Central nervous system demyelinating diseases and increased release of cholesterol into the urinary system of rats
Authors: Nicholas Harold J. , Taylor John
Published: November 2006

The question of what happens to cholesterol in the adult central nervous system during its slow turnover has been addressed using rats with brain and spinal cord labeled with [4‐14C]cholesterol upon intracerebral injection of labeled cholesterol into rats at 10–12 days of age. At six months after injection,14C was found only in the brain and spinal cord and was slowly releasedvia the rat's urine. When labeled rats were given demyelinating agents (triethyl tin chloride, hexachlorophene, sodium cyanide) and when experimental allergic encephalomyelitis was induced, a measurable increase in urinary14C label above control levels was found. It was concluded that there is a direct relationship between the experimental demyelination induced and the increased release of cholesterol metabolites into urine. The study suggests that a clinical method could be developed to determine the rate of central nervous system demyelination by measuring the amount of urinary cholesterol metabolites.
The immunosuppressive substance 2‐chloro‐2‐deoxyadenosine modulates lipoprotein metabolism in a murine macrophage cell line (P388 cells)
Authors: Lechleitner Monika , Auer Bernhard , Zilian Udo , Hoppichler Fritz , Schirmer Michael , Föger Bernhard , Geisen Francoise , Patsch Josef R. , Konwalinka Günther
Published: November 2006

A recently developed immunosuppressive substance, 2‐chloro‐2‐deoxyadenosine (2‐CdA), was reported to inhibit monocyte functions at low concentration. Because macrophages play a key role in the formation of atherosclerotic plaques, it was of interest to study the effect of 2‐CdA on cellular lipid metabolism. For this purpose we have used a macrophage cell line (P388) to perform incubation studies in the presence of acetylated low density lipoprotein (Ac‐LDL) and 2‐CdA. The addition of 2‐CdA, in concentrations ranging from 5–20 nM, induced a dose‐dependent decrease in cellular cholesterol content and in the amount of extracellular [14C]oleic acid (OA) incorporated into the cholesteryl ester (CE) fraction. The effect was maximized at 20 nM 2‐CdA with an 86% reduction in cholesterol esterification compared to controls (P<0.008). To evaluate the mechanism of interaction of 2‐CdA with cellular lipid metabolism, deoxycytidine (dCyt) and 3‐methoxybenzamide (3‐MOB), substances known to antagonize the effect of 2‐CdA in different ways, were co‐administered with 2‐CdA. dCyt, a competitive inhibitor of dCyt kinase, which catalyzes phosphorylation to the active metabolite, antagonized the effects of 20 nM 2‐CdA, producing significantly greater incorporation of extracellular [14C]OA into the CE fraction than in the presence of 2‐CdA alone (P<0.0086). Co‐incubation with 2‐CdA and the poly‐ADP‐ribose synthetase inhibitor 3‐MOB, which is known to render cells resistant to 2‐CdA toxicity by preventing cellular nicotinamide adenine dinucleotide (NAD)‐ and adenosine triphosphase‐depletion, also reversed the effect of 2‐CdA on lipid accumulation. However, incubation of P388 cells with 20 nM 2‐CdA did not result in a decrease in cellular NAD content. As 20 nM 2‐CdA showed no effect on intracellular cholesterol synthesis based on measurement by 3‐hydroxy‐3‐methylglutaryl coenzyme A reductase activity, the decrease in cellular cholesterol content and in [14C]OA incorporation seems to be primarily due to an interference with Ac‐LDL metabolism.
Dietary polyunsaturated fat decreases interaction between low density lipoproteins and arterial proteoglycans
Authors: Manning James M. , Gebre Abraham K. , Edwards Iris J. , Wagner Williams D. , Rudel Lawrence L. , Parks John S.
Published: November 2006

Polyunsaturated dietary fat (n−3 and n−6) results in less atherosclerosis in monkeys compared to lard (Parks, J.S., Kaduck‐Sawyer, J., Bullock, B.C., and Rudel, L.L.,Arteriosclerosis 10, 1102–1112; Rudel, L.L., Parks, J.S., Johnson, F.L., and Babiak, J.,J. Lipid Res. 27, 465–474, 1986). We hypothesized that this was due, in part, to a decreased reactivity of low density lipoproteins (LDL) with arterial proteoglycans (PG). To test this hypothesis, cynomolgus monkeys were fed diets containing lard, safflower oil (n−6 polyunsaturated; Poly), menhanden fish oil (FO), or oleic acid‐rich safflower oil (oleinate; Mono) for 14 mon, and plasma LDL were isolated and characterized. Several properties of LDL thought to be important in the interaction of LDL with arterial PG were measured including LDL particle size, chemical composition, sialic acid content, density distribution, apolipoprotein E (apoE) content and cholesteryl ester transition temperature. Plasma LDL cholesterol concentrations (mg/dL) after 14 mon of diet consumption averaged (mean±SEM): FO (366±45), Lard (352±27), Poly (279±24), and Mono (230±43). The composition of LDL was similar among diet groups except that FO LDL were relatively depleted of cholesteryl ester and enriched in protein and were smaller in size. LDL sialic acid content was similar among diet groups (4.5–5.0 μg/mg LDL protein). The LDL apoE/B molar ratio, a measure of the apoE content per LDL particle averaged: Mono (3.0±1.0), Poly (2.0±0.1), Lard (1.8±0.5), and FO (1.0±0.2). The FO group had a lower proportion (13%) of the apoE enriched d=1.015–1.025 g/mL subfraction of LDL than did the other diet groups (31–45%). The transition temperature of the LDL cholesteryl esters was below body temperature for the FO and Poly groups (36°C) and above for the Lard and Mono groups (40–44°C). The percentage of LDL cholesterol that formed insoluble complexes with arterial chondroitin sulfate PG averaged: Mono (29±4%), Lard (18±3%), Poly (14±3%), and FO (7±2%). Among all diet groups, there was a significant positive correlation (r=0.54) between LDL‐PG complex formation and LDL apoE/B molar ratio. We conclude that dietary FO and Poly result in LDL that are less reactive with arterial PG compared to Lard or Mono fats. While FO appears to decrease PG binding by decreasing the apoE content and amount of the largest LDL subfraction, Poly fat appears to affect LDL‐PG interactions by other mechanisms. Decreased LDL‐PG interactions may lead to decreased atherosclerosis in animals fed polyunsaturated dietary fat.
Effect of dietary linseed oil on tumoricidal activity and eicosanoid production in murine macrophages
Authors: Hubbard Neil E. , Chapkin Robert S. , Erickson Kent L.
Published: November 2006

Diets that contain high levels of n−3 fatty acids from fish oil have been shown to significantly effect macrophage cytolytic capacity, tumor necrosis factor alpha production and eicosanoid production. The present study was undertaken to determine whether n−3 fatty acids from vegetable origin [linseed oil (LIN)] would have the same effects on murine macrophage tumoricidal capacity and eicosanoid production as would fish oil. Mice were fed for three weeks diets that contained 10% (wt/wt) of either LIN, which is high in linolenic acid (18∶3n−3), menhaden fish oil (MFO), which is high in eicosapentaenoic (20∶5n−3) and docosahexaenoic (22∶6n−3) acids, or safflower oil (SAF), which is high in linoleic acid (18∶2n−6).In vivo‐ orin vitro‐activated macrophages were assessed for select functions. As expected, macrophages from mice fed LIN and MFO produced significantly lower levels of both prostaglandins and leukotriene C4 when compared with macrophages from mice fed SAF. In addition, LIN and MFO macrophages were able to synthesize leuko‐triene C5, which could not be produced by macrophages from mice fed SAF. The effects of LIN, however, were not as pronounced as those of MFO. With respect to specific functions, macrophages from mice fed LIN did not have altered cytolytic capacity when compared with macrophages from mice fed SAF and activatedin vitro with either lipopolysaccharide (LPS) alone for 24 h or with LPS plus interferon gamma (IFNγ) for 5 h. Diet did not significantly alter tumoricidal capacity of macrophages activated completelyin vivo either. Specific binding of macrophages to tumor targets, nitric oxide production and the production of tumor necrosis factor alpha were found to be unaltered by LIN when compared with SAF. The results are not consistent with a general n−3 effect, as LIN could not effect the functions comparable to MFO. The results also suggest that a change in eicosanoid production may not be sufficient to modulate tumoricidal activity in macrophages.
Inhibition of cytolytic T lymphocyte activity by oxysterols
Authors: Küçük Ömer , Stoner‐Picking Jeanette , Yachnin Stanley , Gordon Leo I. , Williams R. Michael , Lis Leonard J. , Westerman Maxwell P.
Published: November 2006

The objective of this study was to investigate the effects of oxysterols (OS), namely 5α‐hydroxy‐6‐ketocholestanol, 6‐ketocholestanol and 25‐hydroxycholesterol, on specific cell‐mediated cytotoxicity by C57BL/6 spleen cells against P815‐X2 (a DBA/2 mastocytoma) target cells. Cytolytic T lymphocytes (CTL) were generated by intraperitoneally injecting C57BL/6 mice with P815‐X2 tumor cells 10 d prior to the cytotoxicity experiments. Preincubation of CTL with 10−5 M 5α‐hydroxy‐6‐ketocholestanol and 6‐ketocholestanol for 45 min in lipoprotein‐depleted medium resulted in an inhibition of cytolytic activity (73 and 43%, respectively) as measured by 4‐h51Cr release. At a concentration of 5×10−6 M, 5α‐hydroxy‐6‐ketocholestanol inhibited CTL activity by 65%, whereas 6‐ketocholestanol did not elicit any inhibition. By contrast, 25‐hydroxycholesterol did not inhibit CTL at either concentration, although it is known to be a potent inhibitor of 3‐hydroxy‐3‐methyl‐glutaryl‐coenzyme A reductase, the rate‐limiting enzyme in the cholesterol biosynthetic pathway. When CTL were preincubated with OS in lipoprotein‐replete medium, there was no inhibition of CTL activity at the respective concentrations. The results suggest that the inhibition of CTL activity upon short‐term incubation with OS is not due to the inhibition of cholesterol synthesis, but may be due to the insertion of OS into the plasma membrane to replace cholesterol and alteration of membrane physical properties.