Effect of dietary n−6 and n−3 polyunsaturated fatty acids on peroxidizability of lipoproteins in steers

The susceptibility of major plasma lipoproteins to lipoperoxidation was studied in relation to the FA composition of their neutral and polar lipids in steers given PUFA-rich diets. Two trials used, respectively, 18 (“sunflower” experiment, S) or 24 (“linseed” experiment, L) crossbred Salers x Charolais steers. Each involved three dietary treatments over a 70-d period: a control diet (CS or CL diets) consisting of hay and concentrate, or the same diet supplemented with oilseeds (4% diet dry matter) fed either as seeds (SS or LS diets) or continuously infused into the duodenum (ISO or ILO diets). Compared with control diets, ISO and ILO treatments tended to decrease the resistance time of LDL and HDL classes to peroxidation, mainly owing to the enrichment of their polar and neutral lipids with PUFA. With diets SS and LS, sensitivity of major lipoprotein classes (LDL, light and heavy HDL) was not affected because ruminal hydrogenation of dietary PUFA decreased their incorporation into lipoparticles. ISO and ILO treatments induced a more important production of conjugated dienes and hydroperoxides generated by peroxidation in the three lipoprotein classes due to the higher amounts of PUFA esterified in lipids of the core and the hydrophilic envelope of particles. The production of malondialdehyde (MDA) increased in steers fed linseed supplements, indicating that MDA production did not occur with linoleic acid provided by sunflower oil supplements. Thus, plasma peroxidation of PUFA generates toxic products in steers fed diets supplemented with PUFA and can be deleterious for the health of the animal during long-term treatment.     

A function for α-tocopherol: Stabilization of the microsomal membrane from radical attack during TPNH-dependent oxidations

Events accompanying electron transport in the membrane fraction of liver and other tissues have led us to propose a specific function for α-tocopherol based on a sequence of biochemical changes we observed to occur in these membranes and on pertinent information from other laboratories. The activity of a membrane-bound enzyme system (TPNH oxidase) which involves transport of electrons from substrate to oxygen, has been shown to promote simultaneous formation of peroxide functions on the β position polyunsaturated fatty acids (PUFA) of phospholipids in the membrane. The phospholipid peroxides then undergo a chain cleavage reaction producing phospholipids containing a variety of carbonyl moieties in the β position. The process results in marked alteration of the membrane structure. During the overall reaction α-tocopherol present in the membrane is converted to a compound more polar than tocopheryl quinone and the conversion is dependent on the same enzymic factors promoting the phospholipid alterations. The membrane alteration process is enhanced in microsomes from animals fed diets containing relatively high levels of PUFA or diets low in α-tocopherol, and is diminished by low levels of dietary PUFA or relatively high levels of α-tocopherol. The experimental data indicate that enzymic electron transport associated with TPNH oxidation by the microsomal membrane involves free radical functions. The latter apparently can promote extensive peroxidative alterations of phospholipids that result in structural changes in the membrane unless adequate α-tocopherol is present in this organelle. This system appears to be part of the microsomal drug metabolizing system.     

Suppression of growth in a leukemic T cell line by n−3 and n−6 polyunsaturated fatty acids

Proliferation in a lekemic T cell line (Jurkat) was suppressed in a dose dependent manner by n−6 and n−3 polyunsaturated fatty acids (PUFA) added to the culture medium. At high concentrations, PUFA have a cytotoxic effect on Jurkat cells. The inhibitory effect of the PUFA was not due to production of prostaglandins, and lipid peroxidation was only partly responsible. In addition to production of peroxides and aldehydes, lipid peroxidation also reduced the plasmalogen levels in these cells. The antioxidant α-tocopherol blocked lipid peroxidation and restored the plasmalogen levels to normal. α-Tocopherol did not totally restore cell proliferation although the MDA-like products in these cultures (supplemented with PUFA) were reduced to control level. Cultures supplemented with n−6 PUFA seemed to respond better to α-tocopherol than n−3 PUFA. This suggests that n−6 PUFA may exert their growth inhibitory effect predominantly via lipid peroxidation while different mechanisms might be operating for the n−3 PUFA.     

Dietary antioxidants in young swine

Young swine obtained by hysterectomy were fed purified diets low in vitamin E and supplemented with d-α-tocopheryl acetate and ethoxyquin (Santoquin^R). It was demonstrated that with very low levels of polyunsaturated fatty acids (PUFA) in the diet, both tocopherol and Santoquin protected the tissues of the pig from increased thiobarbituric acid (TBA) values and from increased hemolysis usually associated with low vitamin E status. When the dietary PUFA were increased to levels over 5%, the supplements of tocopherol and Santoquin protected against increased TBA values of tissue homogenates, but not against increased hemolysis of erythrocytes, even when blood serum showed substantial amounts of tocopherol. Some of the interrelationships of dietary PUFA and α-tocopherol were demonstrated. It was shown that for each 1% of peroxidized corn oil added to the diet above 4%, roughly 100 mg of d-α-tocopheryl acetate was necessary to protect the pigs from erythrocyte hemolysis. The failure to reach a “zero” TBA value in vitamin E-deficient swine tissue homogenates substantiated the theory ofin vivo lipid autoxidation, and the increased TBA values of incubated tissue homogenates demonstratedin vitro lipid autoxidation in tissues not protected by a biological antioxidant. Presented at the AOCS meeting in Toronto, Canada, 1962. Supported by grants from the Monsanto Chemical Co., St. Louis, Mo., and The Hormel Foundation.     

Plasma and lipoprotein lipid peroxidation in humans on sunflower and rapessed oil diets

The effects of natural mixed diets on lipid peroxidation were investigated in humans. In the first study, 59 subjects were fed a rapeseed oil-based diet rich in monounsaturated fatty acids (MUFA) and a sunflower oil-based diet rich in polyunsaturated fatty acids (PUFA) in a cross-over manner for three and a half weeks. The lipid peroxidation products in plasma were determined by measuring conjugated dienes and malondialdehyde (MDA). In a second study, plasma thiobarbituric acid reactive substances (TBARS), lipid hydroperoxides, and the susceptibility of very low density lipoprotein + low-density lipoprotein (LDL) toin vitro oxidation were measured from subjects fed similar MUFA and PUFA diets for six week diets. No significant differences in plasma MDA or conjugated diene concentrations were found after the rapeseed oil diet or the sunflower oil diet in Study 1. In the second study, a small but significant decrease (P<0.05) in both lipid hydroperoxides and TBARS was observed in the LDL fraction after the sunflower oil diet. Thein vitro oxidation gave opposite results, showing increased oxidation after the sunflower oil diet. Despite a high intake of α-tocopherol during the oil peroids, no increase in plasma α-tocopherol was noticed in either study. The results suggest that moderate changes in the fatty acid composition in the Western-type diet may be adequate to affect lipoprotein susceptibility to oxidationin vitro, but there is considerable disparity with some indices ofin vivo lipid peroxidation.     

An enzymatic protective mechanism against lipid peroxidation damage to lungs of ozone-exposed rats

The effects of whole animal exposure to ozone and of dietary α-tocopherol on the occurrence in rat lung of lipid peroxidation and alteration of the activity of enzymes important in detoxification of lipid peroxides were studied. Exposure to 0.7 and 0.8 ppm ozone continuously for 5 and 7 days, respectively, significantly elevated the concentration of TBA reactants, primarily malonaldehyde, produced by lipid peroxidation, as well as the activities of glutathione (GSH) peroxidase, GSH reductase and glucose-6-phosphate (G-6-P) dehydrogenase. As a logarithmic function of dietary α-tocopherol (0, 10.5, 45, 150 and 1500 mg/kg), the increase in formation of malonaldehyde and the increase in activities of GSH peroxidase and G-6-P dehydrogenase were partially inhibited. The activity of GSH reductase was not affected by dietary α-tocopherol. The concentration of malonaldehyde and the activity of GSH peroxidase in lung were linearly correlated (p<0.001). This study confirmed the occurrence of lipid peroxidation in the lung during ozone exposure and revealed an enzymatic mechanism against damage. An apparent compensation mechanism is that with increased lipid peroxides there is increased activity of GSH peroxidase, which in turn increases lipid peroxide catabolism. The increased activities of GSH reductase and G-6-P dehydrogenase also function in the protective chain by providing increased levels of GSH and NADPH, respectively. Postdoctoral fellow of the American Society for Clinical Nutrition sponsored by the National Vitamin Foundation.     

Effect of antioxidants on lipid peroxidation in iron-loaded rats

Indirect evidence has suggested that lipid peroxidation is associated with iron overload in vivo. As a measure of lipid peroxidation, pentane expired in the breath of rats loaded with an accumulated dose of either 100 mg or 186–200 mg of iron injected intraperitoneally as iron dextran was measured over a 7 to 8 week period, and the effect on pentane production of feeding antioxidant-supplemented diets was determined. By the seventh week of feeding the diets, rats fed 0.3% L-ascorbic acid produced 17% less (P=0.03) pentane than did rats fed the basal antioxidant-deficient diet, whereas rats fed 0.004% dl-α-tocopherol acetate produced 92% less (P<0.001). After being fed the basal diet for 7 weeks, iron-loaded rats produced 76±9 pmol pentane/100 g body wt/min. When synthetic antioxidants were added to the diet at a concentration of 0.25%, the order of effectiveness in decreasing pentane production after 1 week was: N,N′-diphenyl-p-phenylenediamine > ethoxyquin > butylated hydroxyanisole > butylated hydroxytoluene > propyl gallate ∼ no antioxidant. After removal of either ethoxyquin or N,N′-diphenyl-p-phenylenediamine from the diets for 1 week, pentane production increased to a high level. The total amount of lipid soluble fluorophores in individual spleens of rats fed N,N′-diphenyl-p-phenylenediamine, ethoxyquin, dl-α-tocopherol acetate, ascorbic acid and no antioxidant were correlated significantly with the corresponding total integrated amount of pentane produced by the individual rats over the 7 to 8 week period. This study has provided some of the most direct evidence to date that lipid peroxidation is associated with iron overload in vivo.     

Dietary comparison of conjugated linolenic acid (9 cis, 11 trans, 13 trans) and α-tocopherol effects on blood lipids and lipid peroxidation in alloxan-induced diabetes mellitus in rats

The present study investigated the dietary effect of conjugated linolenic acid (CLnA) on lipid profiles and lipid peroxidations in alloxan-induced diabetes mellitus in rats. Diabetic rats were fed with 20% sunflower oil (diabetic control), sunflower oil supplemented with 0.5% CLnA, sunflower oil supplemented with 0.15% α-tocopherol, and sunflower oil containing 0.25% CLnA+0.15% α-tocopherol. The results demonstrated that 0.5% CLnA, 0.15% α-tocopherol, and 0.25% CLnA+0.15% α-tocopherol each on supplementation significantly lowered total cholesterol and non-HDL-cholesterol in comparison with the diabetic control group. The TAG level was significantly lowered in both the 0.15% α-tocopherol and 0.25% CLnA+0.15% α-tocopherol groups. LDL lipid peroxidation and erythrocyte membrane lipid peroxidation were reduced significantly in each of the experimental groups vs. the control group. The CLnA+α-tocopherol diet induced a greater reduction in membrane lipid and liver lipid peroxidation than the α-tocopherol diet alone. In conclusion, dietary CLnA exerts antioxidant activity as evidenced by reduced lipid peroxidation in chemically induced diabetes mellitus.     

The relationship between fatty acid peroxidation and α-tocopherol consumption in isolated normal and transformed hepatocytes

The response of normal and transformed rat hepatocytes to oxidative stress was investigated. Isolated normal rat hepatocytes and differentiated hepatoma cells (the Fao cell line was derived from the Reuber H 35 rat hepatoma) in suspension were incubated with the ADP/Fe³⁺ chelate for 30 min at 37°C. Membrane lipid oxidation was assessed by measuring (i) free malondialdehyde (MDA) production by a high-performance liquid chromatography (HPLC) procedure, (ii) membrane fatty acid disappearance as judged by capillary gas chromatography, and (iii) α-tocopherol oxidation as determined by HPLC and electrochemical detection. The addition of iron led to increased MDA production in normal as well as in transformed cells, and to simultaneous consumption of polyunsaturated fatty acids (PUFA) and α-tocopherol. In addition, in Fao cells more α-tocopherol was consumed during lipid peroxidation while less PUFA was oxidized. Lipid peroxidation was lower in tumoral hepatocytes than in normal cells. This could be due to a difference in membrane lipid composition because of a lower PUFA content and a higher α-tocopherol level in Fao cells. During oxidation, Fao cells produced 1.5 to 2 times less MDA than normal cells, while in the tumoral cells the amount of oxidized PUFA having 3 or more double bonds was 7 to 8 times lower. Therefore, measuring MDA alone as an index of lipid peroxidation did not allow for proper comparison of the membrane lipid oxidizability of transformed cellsvs. the membrane lipid oxidizability of normal cells.     

Dietary linoleic acid and the fatty acid profiles in rats fed partially hydrogenated marine oils

The influence of the linoleic acid levels of diets containing partially hydrogenated marine, oils (HMO) rich in isomeric 16∶1, 18∶1, 20∶1 and 22∶1 fatty acids on the fatty acid profiles of lipids from rat liver, heart and adipose tissue was examined. Five groups of rats were fed diets containing 20 wt% fat−16% HMO+4% vegetable oils. In these diets, the linoleic acid contents varied between 1.9% and 14.5% of the dietary fatty acids, whereas the contents oftrans fatty acids were 33% in all groups. A sixth group was fed a partially hydrogenated soybean oil (HSOY) diet containing 8% linoleic acid plus 32%trans fatty acids, mainly 18∶1, and a seventh group, 20% palm oil (PALM), with 10% linoleic acid and notrans fatty acids. As the level of linoleic acid in the HMO diets increased from 1.9% to 8.2%, the contents of (n−6) polyunsaturated fatty acids (PUFA) in the phospholipids increased correspondingly. At this dietary level of linoleic acid, a plateau in (n−6) PUFA was reached that was not affected by further increase in dietary 18∶2(n−6) up to 14.5%. Compared with the HSOY- or PALM-fed rats, the plateau value of 20∶4(n−6) were considerably lower and the contents of 18∶2(n−6) higher in liver phosphatidylcholines (PC) and heart PC. Heart phosphatidylethanolamines (PE) on the contrary, had elevated contents of 20∶4(n−6), but decreased 22∶5(n−6) compared with the PALM group. All groups fed HMO had similar contents oftrans fatty acids, mainly 16∶1 and 18∶1, in their phospholipids, irrespective of the dietary 18∶2 levels, and these contents were lower than in the HSOY group. High levels of linoleic acid consistently found in triglycerides of liver, heart and adipose tissue of rats fed HMO indicated that feeding HMO resulted in a reduction of the conversion of linoleic acid into long chain PUFA that could not be overcome by increasing the dietary level of linoleic acid.     

Gender and dietary fat affect α-tocopherol status in F344/N rats

For four weeks, groups of eight male and eight female F344/N rats were fed diets containing 15.5, 20, 30 or 40% of energy (en%) as fat. The fat was composed of corn oil and beef tallow with 9 en% from linoleate in all diets. Females had greater mean hepatic α-tocopherol levels, whereas males had greater plasma α-tocopherol and cholesterol concentrations. In males, the plasma ratio of α-tocopherol/cholesterol was significantly greater than in females (P<0.05). Plasma α-tocopherol increased with increasing en% fat (r=0.51,P<0.001) in both sexes, but dietary fat did not alter hepatic α-tocopherol levels. These results suggest that plasma α-tocopherol may serve as a biomarker of total dietary fat intake and that in F344/N rats gender differences affect α-tocopherol and cholesterol status.     

The effect of dietary vitamin E supply and a moderately oxidized oil on activities of hepatic lipogenic enzymes in rats

Diets rich in polyunsaturated fatty acids (PUFA) are well known to suppress hepatic lipogenic enzymes compared to fat-free diets or diets rich in saturated fatty acids. However, the mechanism underlying suppression of lipogenic enzymes is not quite clear. The present study was undertaken to investigate whether lipid peroxidation products are involved in suppression of lipogenic enzymes. Therefore, an experiment with growing male rats assigned to six groups over a period of 40 d was carried out. Rats received semisynthetic diets containing 9.5% coconut oil and 0.5% fresh soybean oil (coconut oil diet, peroxide value 5.1 meq O₂/kg oil), 10% fresh soybean oil (fresh soybean oil diet, peroxide value 0.5 meq O₂/kg oil), or 10% thermally treated soybean oil (oxidized soybean oil diet, peroxide value 74 meq O₂/kg oil). To modify the antioxidant state of the rats, we varied the vitamin E supply (11 and 511 mg α-tocopherol equivalents per kg of diet) according to a bi-factorial design. Food intake and body weight gain were not influenced by dietary fat and vitamin E supply. Activities of hepatic lipogenic enzymes were markedly influenced by the dietary fat. Feeding either fresh or oxidized soybean oil diets markedly reduced activities of fatty acid synthase, (FAS), acetyl CoA-carboxylase, (AcCX), glucose-6-phosphate dehydrogenase, (G6PDH), 6-phosphogluconate dehydrogenase, and ATP citrate lyase (ACL) relative to feeding the coconut oil diet. Moreover, feeding oxidized soybean oil slightly, but significantly, lowered activities of FAS, AcCX, and ACL compared to feeding fresh soybean oil. Activities of hepatic lipogenic enzymes were reflected by concentrations of triglycerides in liver and plasma. Rats fed the coconut oil diet had markedly higher triglyceride concentrations in liver and plasma than rats consuming fresh or oxidized soybean oil diets, and rats fed oxidized soybean oil had lower concentrations than rats fed fresh soybean oil. The vitamin E supply of the rats markedly influenced concentrations of thiobarbituric acid-reactive substances in liver, but it did not influence activities of hepatic lipogenic enzymes. Because the vitamin E supply had no effect, and ingestion of an oxidized oil had only a minor effect, on activities of hepatic lipogenic enzymes, it is strongly suggested that neither exogenous nor endogenous lipid peroxidation products play a significant role in the suppression of hepatic lipogenic enzymes by diets rich in PUFA. Therefore, we assumed that dietary PUFA themselves are involved in regulatio of hepatic lipogenic enzymes. Nevertheless, the study shows that ingestion of oxidized oils, regardless of the vitamin E supply, also affects hepatic lipogenesis, and hence influences triglyceride levels in liver and plasma.     

Effect of dietary carnosine on plasma and tissue antioxidant concentrations and on lipid oxidation in rat skeletal muscle

The effect of dietary carnosine supplementation on plasma and tissue carnosine and α-tocopherol concentrations and on the formation of thiobarbituric acid reactive substances (TBARS) in rat skeletal muscle homo-genates was evaluated. Plasma, heart, liver and hind leg muscle was obtained from rats fed basal semipurified diets or basal diets containing carnosine (0.0875%), α-tocopheryl acetate (50 ppm), or carnosine (0.0875%) plusα-tocopheryl acetate (50 ppm). Dietary carnosine supplementation did not increase carnosine concentrations in heart, liver and skeletal muscle. Dietary supplementation with both carnosine and α-tocopherol increased carnosine concentrations in liver 1.56-, 1.51- and 1.51-fold as compared with diets lacking carnosine, α-tocopherol or both carnosine and α-tocopherol, respectively. Dietary supplementation with both carnosine and α-tocopherol also increased α-tocopherol concentrations in heart and liver 1.38-fold and 1.68-fold, respectively, as compared to supplementation with α-tocopherol alone. Dietary supplementation with carnosine, α-tocopherol or both car-nosine and α-tocopherol was effective in decreasing the formation of TBARS in rat skeletal muscle homogenate, with dietary α-tocopherol and α-tocopherol plus carnosine being more effective than dietary carnosine alone. The data suggest that dietary supplementation with carnosine and α-tocopherol modulates some tissue carnosine and α-tocopherol concentrations and the formation of TBARS in rat skeletal muscle homogenates.     

Lipid composition and peroxide levels of mucosal cells in the rat large intestine in relation to dietary fat

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.     

Positive association between plasma antioxidant capacity and n−3 PUFA in red blood cells from women

PUFA are susceptible to oxidation. However, the chain-reaction of lipid peroxidation can be interrupted by antioxidants. Whether an increased concentration of PUFA in the body leads to decreased antioxidant capacity and/or increased consumption of antioxidants is not known. To elucidate the relationship between plasma total antioxidant capacity (TAC), the concentration of antioxidant vitamins, and the proportion of PUFA in red blood cells (RBC), plasma TAC was measured by a Trolox equivalent antioxidant capacity assay in blood samples from 99 Icelandic women. Concentrations of tocopherols and carotenoids in the plasma were determined by HPLC, and the FA composition of RBC total lipids was analyzed by GC. Plasma TAC and the plasma concentration of α-tocopherol correlated positively with the proportion of total n−3 PUFA, 20∶5n−3, and 22∶6n−3 in RBC, whereas the plasma lycopene concentration correlated negatively with the proportion of total n−3 PUFA and 20∶5n−3. On the other hand, plasma TAC correlated negatively with the proportion of n−6 PUFA in RBC. Plasma TAC also correlated positively with the plasma concentration of α-tocopherol, alcohol consumption, and age. Both the plasma concentration of α-tocopherol and age correlated positively with the proportion of n−3 PUFA in RBC; however, n−3 PUFA contributed independently to the correlation with plasma TAC. Because the proportion of n−3 PUFA in RBC reflects the consumption of n−3 PUFA, these results suggest that dietary n−3 PUFA do not have adverse effects on plasma TAC or the plasma concentration of most antioxidant vitamins.     

Ratios of polyunsaturated fatty acids to α-tocopherol in tissues of rats fed corn or soybean oils

This study was part of a larger experiment designed to assess the vitamin E adequacy of corn and soybean oils in relation to their polyunsaturated fatty acids (PUFA). Young male rats were fed a semipurified diet containing 20% corn or soybean oil and adequate selenium. After 8 and 12 weeks, animals were sacrificed, and 7 tissues analyzed for α- and γ-tocopherols and for fatty acids. Calculations were made of the molar ratios of total polyunsaturated fatty acids/α-tocopherol, and also of all polyunsaturated fatty acids, except linoleate, designated polyunsaturated fatty acids>18∶2, to α-tocopherol. It is proposed that the latter ratio may have more significance, physiologically, than when linoleic acid also is considered. Tissues from rats fed corn oil had slightly more favorable (lower) ratios than did tissues from rats fed soybean oil. In both groups, the molar polyunsaturated fatty acids>18∶2/α-tocopherol ratio was lowest for heart and lung, intermediate for muscle and testis, and highest for liver, kidney, and adipose tissue. Since both corn and soybean oils provide adequate vitamin E as determined by several biochemical and physiological parameters, adequate molar ratios of polyunsaturated fatty acids>18∶2/α-tocopherol were: lung, 400; heart and leg muscles, 700; testis, 1100; liver and kidney, 1500–2000; and adipose tissue, 2000.     

Rat vitamin E status and heart lipid peroxidation: Effect of dietary α-Linolenic acid and marine n−3 fatty acids

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.     

Effect of γ-tocopherol on formation of nonvolatile lipid degradation products during frying of potato chips in triolein

Formation of undesirable odors and flavors during food processing operations is an important problem for the food industry. To determine the effect of γ-tocopherol on these negative attributes of fried food, we fried potato chips in triolein with 0, 100, or 400 ppm γ-tocopherol. Triolein extracted from potato chips was sampled for residual γ-tocopherol and nonvolatile degradation products after the chips were aged. RP-HPLC coupled to atmospheric pressure chemical ionization MS and size-exclusion chromatography was used to analyze, samples for degradation products in the triolein absorbed in potato chips as well as the fryer, triolein. MS results showed that γ-tocopherol reduced the production of nonvolatile degradation products in the triolein absorbed by the potato chips and in the triolein in the fryer. Fryer oil samples and extracted potato chip oils with 400 ppm γ-tocopherol had a significantly lower production of degradation compounds than did samples with 100 ppm γ-tocopherol. Both fryer oils and potato chips containing 100 ppm γ-tocopherol had significantly fewer nonvolatile degradation products than did the samples without γ-tocopherol. These nonvolatile compounds are known precursors of negative odors and flavor compounds produced during the frying and aging of foods.     

Cholesterol oxidation in meat from chickens fed α-tocopherol-and β-carotene-supplemented diets with different unsaturation grades

The production of B-ring and side-chain oxysterols was evaluated in meat from chickens fed diets differing by the kind of oil or fat added. The effect of supplementary levels of natural antioxidants, as α-tocopherol and β-carotene, on the meat cholesterol oxidative stability was also studied. Lard, sunflower and olive oil were used as dietary fat. Raw and cooked meats were analyzed for oxysterols, and cholesterol was also quantified. Oxysterol analyses were carried out by combining the use of solid-phase extraction, thin-layer chromatography, capillary gas chromatography, and capillary gas chromatography-mass spectrometry. Oxysterols were detected within the 0.1–0.5 μg/g range in raw meat. Cooking increased the oxysterol content of the meat, and levels as high as 5 μg/g muscle tissue were observed. B-Ring oxysterols were mainly produced: the α-and the β-epoxycholesterols, the 7α-and 7β-hydroxycholesterols, and the 7-ketocholesterol. The results showed that the meat from the chickens fed the olive oil-based diet containing α-tocopherol at 200 mg/kg of diet presented the best cholesterol oxidative stability. A positive effect could not be found for dietary β-carotene administered at levels of 15 and 50 mg/kg of diet. Furthermore, a significant decrease in the tissue cholesterol content was observed with the olive and the sunflower oil-based diets.     

Reduced glutathione effects on α-tocopherol concentration of rat liver microsomes undergoing NADPH-dependent lipid peroxidation

Factors involved in reduced glutathione (GSH) and vitamin E-mediated inhibition of NADPH-dependent rat liver microsomal lipid peroxidation were examined. Lipid peroxidation was monitored over a time-course of 180 min by thiobarbituric acid reactive product formation. The addition of 5 mM GSH to the reaction system containing microsomes from rats fed a diet supplemented with 150 IU/kg of α-tocopherol acetate for eight weeks produced a lag in peroxidation of >30 min. This effect was not observed for microsomes prepared from rats fed a diet deficient in vitamin E. Indeed, a prooxidant effect of 5 mM GSH was observed in assays containing microsomes from rats fed a diet deficient in vitamin E. The inhibition by GSH of lipid peroxidation in microsomes prepared from livers of vitamin E supplemented rats was not restricted by its availability, for it was found that approximately 92% of the GSH remained in the reduced form after 60 min. Additional experiments revealed that the α-tocopherol content of peroxidizing microsomes decreased rapidly in the absence of GSH. The addition of 5 mM GSH to the assay system markedly depressed the loss of microsomal α-tocopherol. The results ofin vivo labeling of liver microsomes with [¹⁴C] α-tocopherol demonstrated that i) GSH addition to thein vitro peroxidizing medium reduced the disappearance of α-tocopherol, and ii) a compound that interfered with the determination of α-tocopherol was separated by HPLC and was not an oxidation product of α-tocopherol. A portion of the microsomal¹⁴C-labeled α-tocopherol was converted to an unidentified product with HPLC retention characteristics that was similar, but not identical, to α-tocopherol quinone.