Influence of sources of dietary vitamin E on the maternal transfer of alpha-tocopherol to fetal and neonatal guinea pigs as determined by a stable isotopic technique.

The accepted biological potencies of vitamin E (United States Phamacopeia, 1985) for 1 mg all-rac-alpha-tocopheryl acetate (synthetic form) is 1.00 IU and that of 1 mg (RRR)-alpha-tocopheryl acetate (natural form) is 1.36 IU. In the present study, a stable isotopic (2H) technique was employed to evaluate the bioavailability of natural v. synthetic forms of vitamin E and to determine whether the potency of the forms is the stated relationship of 1.36:1.00 (RRR)-alpha-tocopheryl acetate:all-rac-alpha-tocopheryl acetate. Sixty female in-bred guinea pigs received either 40 or 80 mg vitamin E/kg diet with equal levels of (RRR)-alpha-tocopheryl acetate and all-rac-alpha-tocopheryl acetate throughout gestation and lactation. At late-term pregnancy (day 50 or 60) and during early lactation, dams and their corresponding fetuses or neonates were killed and various tissues collected for subsequent alpha-tocopherol analysis. Vitamin E analysis of fetal and neonatal tissues indicated a substantial transfer of 2H-labelled alpha-tocopherol across the placenta and through the mammary gland. Total alpha-tocopherol concentrations were significantly influenced by tissue type and dose level, but not by stage of gestation or lactation. The relative bioavailability (d3:d6) across fetal and neonatal tissues was on average 1.81:1.00, with a range from 1.62:1.00 to 2.01:1.00. Maternal tissues had a mean ratio of 1.77:1.00. A higher relative bioavailability (P

Effect of chronic ethanol dosing on hepatic triglyceride and phospholipid profile and fatty acids in the guinea pig.

An alcohol feeding study was conducted with guinea pigs to evaluate the influence of alcohol upon hepatic triglyceride and total phospholipid profile as well as phospholipid fatty acids. Twenty-seven guinea pigs were randomly assigned into four groups consisting of a control and alcohol-treated group and each group carried over a 105- or 135-day period . Alcohol was administered via the drinking water starting with a 2.5% solution (v/v) and gradually increased to 12.5% (v/v) over a 30-day period and thereafter maintained continuously for either 75 or 105 days, respectively. Control guinea pigs received glucose via the drinking water to match isocalorically the alcohol given to the test animals. At the end of the 105- and 135-day periods, animals were sacrificed and livers collected. Hepatic triglycerides were significantly elevated by alcohol dosing, whereas total phospholipid fraction remained essentially unaltered. No significant time effect was observed on hepatic triglyceride and phospholipid profiles. In ethanol-fed guinea pigs, significant increases in percentages of 18:1 n-9 and 18:2 n-6 and decreases in 16:0, 20:3 n-6 and 20:4 n-6 were observed in hepatic total phospholipid fatty acid profile compared to controls. In addition, other polyenoic acids including 22:4 n-6, 22:5 n-6, 22:5 n-3, and 22:6 n-3 were found to be highly significantly depressed in alcohol-treated animals in comparison to the controls. This study provides important baseline lipid data on guinea pig responses to ethanol and provides a starting point for the use of the guinea pig as an experimental model.

Gender and exercise influence on tissue antioxidant vitamin status in rats.

Although gender differences in antioxidant status based largely on differing estrogen levels have been postulated, it is not known if other gender based differences in tissue antioxidants exist. This experiment examined whether gender based differences in tissue vitamin C and vitamin E concentration exist, and investigated the possibility of gender based differences in indices of tissue oxidative stress following an acute exercise bout. It was determined that female rats had significantly higher levels of vitamin E in liver and heart tissues than males and that males had significantly more vitamin C in the plantaris muscle than females. However, female rats also had less liver glutathione than males. Acute exercise resulted in significant and equal tissue oxidative stress in both genders as indicated by tissue glutathione status. With some exceptions, tissue vitamin C and vitamin E concentrations were generally unaffected by acute exercise in either gender. Hence, while some gender differences in tissue antioxidant status in rats are evident, these differences do not affect tissue indices of oxidative stress following acute exercise.

Estrogen administration, postexercise tissue oxidative stress and vitamin C status in male rats.

Estrogen can putatively act as an antioxidant and protect tissues from exercise-induced oxidative stress. To test the in vivo efficacy of estrogen, the effects of 2 weeks of daily estrogen (40 microg x kg(-1) body weight beta-estradiol 3-benzoate) injection on indices of immediate postexercise oxidative stress and antioxidant status were determined in adult male rats, with and without 8 weeks of prior dietary vitamin E deprivation. The treadmill running protocol (60 min at 21 m x min(-1), 12% grade) induced significant oxidative stress as indicated by muscle glutathione status. Estrogen administration had little effect on postexercise tissue glutathione status, superoxide dismutase and glutathione peroxidase activity, and vitamin E levels. Estrogen administration induced significant reductions in muscle, liver, and heart vitamin C concentrations following exercise, as well as in unexercised male rats. Tissue vitamin C loss was not directly mediated through liver glycogen or glutathione status. Thus, estrogen administration generally did not appear to influence postexercise tissue indices of oxidative stress or antioxidant status and may have contributed to a decline in overall antioxidant protection by inducing losses in tissue vitamin C content.

Vitamin E levels in gastric mucosa and indomethacin-induced gastric lesions in rats.

The role of vitamin E as a gastroprotective agent from indomethacin-induced damage was investigated in male rats with deficient or supplemented levels of vitamin E in their blood and tissues. Ten weeks of feeding a vitamin E deficient diet produced very low levels of the vitamin in all the tissues studied (below 1.2 micrograms/g tissue). Ten weeks of feeding a diet high in vitamin E content produced high levels of vitamin E in fundus (26 micrograms/g), antrum (18 micrograms/g), duodenum (21 micrograms/g), liver (41 micrograms/g) and plasma (11 micrograms/ml). Oral administration of indomethacin (15 mg/kg) to rats deficient or supplemented with vitamin E resulted in similar degrees of injuries to their gastric mucosa. These data together with previous results indicates that if vitamin E functions as an antioxidant in vivo, lipid peroxidation is not the main cause of gastric damage due to indomethacin.

Repletion and depletion of vitamin E in gastroduodenal mucosa.

Male rats were used to assess the effect of fasting and depletion and repletion rates on the levels of vitamin E in the liver and in mucosa from the fundus, antrum, and duodenum. Fasting for 24 h did not produce a significant decrease in vitamin E content in gastroduodenal mucosa. Feeding rats a diet low in vitamin E content for 10 weeks produced very low levels of the vitamin in all the tissues studied (below 2.4 micrograms/g tissue). Feeding rats a diet high in vitamin E content for 10 weeks produced high levels of vitamin E in the liver (64.5 micrograms/g) and in fundic (47.5), antral (36.9), and duodenal (24.5) mucosa. Calculated rates of depletion (microgram/g tissue/day) were very low in fundic and antral mucosa (0.2-0.7); duodenal mucosa (4.2) and liver (12.2) were the highest. Rates of repletion (microgram/g tissue/day) were lower in the antrum (1.2) and fundus (2.6); duodenal (4.2) and liver (7.6) were the highest. The findings of this study indicate that fasting or short-term dietary manipulation did not substantially change the vitamin E content of the gastroduodenal mucosa.

Tissue discrimination between dietary RRR-alpha- and all-rac-alpha-tocopherols in rats.

The discrimination between the natural form of vitamin E, RRR-alpha-tocopherol, and the synthetic form all-rac-alpha-tocopherol was measured in two experiments for plasma and tissues of rats. In Experiment A, the animals were fed RRR-alpha- or all-rac-alpha-tocopheryl acetate at two different levels, 0.035 and 0.200 g/kg diet, for 2.5 mo. In Experiment B, rats were first depleted of vitamin E for 7 wk then fed a diet containing 0.035 g/kg diet of RRR-alpha- or all-rac-alpha-tocopheryl acetate. In the latter experiment, rats were killed after 3, 7, 17, 31 or 45 d to study the time course of the discrimination. At the end of both experiments, levels of alpha-tocopherol were measured in plasma and tissues. The results showed a preferential uptake of the natural RRR-alpha-tocopherol by the majority of the tissues. The preference is small and in some tissues is not immediately manifested. The greatest discrimination in the tissues examined was found in red blood cells; the levels of alpha-tocopherol accumulated with the RRR-alpha-tocopherol diet were four to six times that accumulated with all-rac-alpha-tocopherol diet. For all other tissues, this ratio was between 1.0 and 1.5. These results indicate that tissues have a mechanism for alpha-tocopherol uptake, but this mechanism is not entirely specific for RRR-alpha-tocopherol.

Malonaldehyde determination in tissues and biological fluids by ion-pairing high-performance liquid chromatography.

A method for the analysis of malonaldehyde by ion pairing high-performance liquid chromatography is described. The method is direct; no thiobarbiturate chromogen formation is required, and sample preparation is simple. After deproteinization with 50% ethanol and removal of particulate by centrifugation samples were passed through a small silica amino column to remove contaminants. Diluted samples (20 microL) were injected onto an octadecylsilane column (25 cm x 4.6 mm ID, 5 micron) which is eluted with 30 mM sodium phosphate buffer, pH 6.5 containing 30% ethanol and 1 mM tetradecyltrimethylammonium bromide. Detection was accomplished by monitoring absorbance at 267 nm. The lower limit for reliable quantification was 5 pmol per injection. The method has been successfully applied to the quantification of malonaldehyde present in plasma, urine and tissues of rats kept under different dietary conditions as well as after in vivo treatment with CCl4 and iron-dextran. The method was also applied to the quantification of malonaldehyde during liver microsomal lipid peroxidation and was compared to the thiobarbituric acid test.

Vitamin C and vitamin E status in the spontaneously diabetic BB rat before the onset of diabetes.

Ascorbic acid (AA), dehydroascorbic acid (DHAA), and vitamin E were measured in tissues and plasma of 30 control and 30 spontaneously diabetic BioBreeding rats (BBdp) during development and before the onset of diabetes. At weaning, rats were fed an AIN-76 semisynthetic diet for 30, 64, or 113 days, after which plasma and tissues from 10 rats of each group were collected and analysed for AA, DHAA, and vitamin E. AA and DHAA levels were significantly increased in plasma and spleen of the diabetes-prone rats compared with those of the control group at 30 and 64 days, but the difference disappeared by 113 days. No differences were observed in liver, adrenals, thymus, and pancreas at any of the time periods. However, lower levels of vitamin E were observed in adrenal gland, thymus, and pancreas of the diabetes-prone rats. It is concluded that BBdp rats have an altered metabolism of AA, DHAA, and vitamin E, before the onset of diabetes. These changes could be due to genetic and physiological factors operating during development of this rat strain.

Kinetics of tissue RRR-alpha-tocopherol depletion and repletion. Effect of cold exposure.

Vitamin E was estimated in plasma and tissues of rats kept for three months on a low vitamin E diet or a high vitamin E diet. Some of the animals from each group were switched to the opposite diet, and the kinetics of uptake and depletion of vitamin E were followed 3, 8, and 15 days after the diet change. Some rats were also submitted to cold exposure (6 degrees C) for three days. During repletion plasma, red blood cells, liver, spleen, and adrenal gland were the only tissues that responded rapidly to the diet change; after three days, their vitamin E levels corresponded to that of the new diet. Heart, brain, lung, muscle, and thymus were slow in reacting to diet change. Fifteen days after the change in diet, white adipose tissue did not respond. The rate of repletion for all tissues was more rapid than the rate of depletion, but liver was the only tissue that after three days had vitamin E levels corresponding to the low-vitamin diet. Cold exposure for three days did not produce any significant change in the vitamin E content of any tissue, indicating that despite high oxygen consumption by the animal, vitamin E was not consumed or mobilized.

Ascorbic and dehydroascorbic acids status in rats fed diets varying in vitamin E levels.

Vitamin E, ascorbic and dehydroascorbic acids were determined in plasma and selected tissues of rats fed for 2-3 months different diets varying in vitamin E content. The diets contained as low as 5 IU for group LE, a normal amount of 50 IU for group NE and as high as 250 IU of vitamin E for group HE. Small increases in total vitamin C were observed only in liver, kidney, spleen and plasma with increased dietary levels of vitamin E, however, this was not followed by a substantial increment in the ascorbate/dehydroascorbate ratio. These differences were only observed between diets LE and HE; there was no correlation between vitamin E and vitamin C levels in the tissue. These data suggest that the interactions that readily take place in vitro between these two vitamins do not occur in vivo, probably due to the complexity of natural membranes.

Mechanisms of absorption, transport and tissue uptake of RRR-alpha-tocopherol and d-gamma-tocopherol in the white rat.

The metabolism of alpha- and gamma-tocopherol was studied in three groups of rats that were fed a modified AIN-76 diet containing normal (NE, 0.2 g alpha-tocopherol/kg), high (HE, 1.0 g alpha-tocopherol/kg) or low (LE, less than 0.02 alpha-tocopherol/kg) vitamin E for 3 mo. After 1, 2 and 3 d of an oral dose of 20 mg of alpha-tocopherol, gamma-tocopherol or both, the levels of the two vitamers were measured in plasma and tissues and in some cases in isolated microsomal and mitochondrial fractions from liver. Twenty-four hours after an oral dose of 20 mg gamma-tocopherol the levels of alpha-tocopherol in plasma and tissues remained constant and higher levels of gamma-tocopherol were found in tissues in which low alpha-tocopherol levels could be found such as in the LE group. In spite of this, it was enabled to remain there, after 2 and 3 d gamma-tocopherol had decreased levels in all tissues. When given in combination with alpha-tocopherol, the levels of gamma-tocopherol were lower than when gamma-tocopherol was given alone. Microsomes and mitochondria from livers of LE group bound five and nine times more alpha-tocopherol than gamma-tocopherol in rats dosed with equal amount of alpha- or gamma-tocopherol, respectively. These data indicate that the mechanisms that regulate the metabolism of vitamin E are highly specific for alpha-tocopherol. Moreover, the relative amount of alpha-tocopherol determined the levels of gamma-tocopherol in tissues. However, the retention of gamma-tocopherol in tissues did not depend on the presence of alpha-tocopherol.(ABSTRACT TRUNCATED AT 250 WORDS)

A highly sensitive high-performance liquid chromatography method for the estimation of ascorbic and dehydroascorbic acid in tissues, biological fluids, and foods.

A highly sensitive procedure for determining ascorbic acid (AA) and dehydroascorbic acid (DHAA) by high-performance liquid chromatography with electrochemical detection in biological fluids, tissues, and foods is described. AA is separated in a C18 reverse-phase column after extraction from the sample with metaphosphoric acid. An aliquot of 20 microliter of diluted extract is injected into the column for the estimation of AA. DHAA is indirectly estimated by converting it to AA after reduction with DL-homocysteine at pH 7.0-7.2 for 30 min at 25 degrees C. After dilution, a 20-microliter aliquot is injected into the column to obtain total vitamin C (AA + DHAA). The concentration of DHAA is calculated by subtraction. AA can be reproducibly quantified at concentrations as low as 50 pg/20 microliter of sample extract. The method described here used a specially designed mobile phase, gave greater stability and a noiseless baseline, and increased substantially the sensitivity and precision. The procedure is rapid, analysis being completed within 10 min after sample preparation, and has been successfully applied to biological fluids, tissues, and foods.

Effect of dietary vitamin E on the vitamin E status in the BB rat during development and after the onset of diabetes.

Weanling diabetes-prone BB rats were fed AIN-76 diets containing high (HE, 1 g/kg diet), basal (NE, 0.2 g/kg) or low (LE, trace) vitamin E and were killed at 21, 42 or 60 days of age. Plasma and tissues (adrenals, pancreas, spleen, thymus, liver, brown and white adipose tissue, muscle and testes) were analysed for vitamin E. Vitamin E levels reflected the level in the diet and no diabetic animals were detected at these times. In a second experiment, a total of 90 diabetes-prone BB rats were kept on diets LE and HE for 6 months or until they became diabetic. 11/45 on LE and 5/45 on HE became diabetic. Again, plasma and tissue levels of vitamin E reflected the levels in the diet with the exception of the thymus of diabetic rats fed the high vitamin E diet. Thymus vitamin E levels (microgram/g tissue) were 1.8 and 1.2 in LE-fed diabetics and asymptomatic rats, respectively; and 22.7 and 49.5 in HE-fed diabetics and asymptomatic rats, respectively. The last 2 values were significantly different (p less than 0.005). There were no other differences in plasma or tissue levels of vitamin E in these groups of animals. These findings suggest that high dietary vitamin E may decrease the incidence of diabetes in animals which are able to accumulate sufficient amounts of the vitamin in the thymus. Since the thymus plays a key role in the maturation of T cell populations, which appear to be altered in this disease, it seems possible that the protective effect may be exerted at this level.

Alpha- and gamma tocopherol concentrations in human serum.

Vitamin E (alpha- and gamma-tocopherol) and total lipid concentrations were determined in the serum of 451 healthy human male and female subjects ranging in age from 19 to 70 years. Mean serum alpha-tocopherol concentration (microgram/ml) was 8.82 +/- 2.84 (SD) and gamma-tocopherol concentration was 1.35 +/- 0.70. The concentration of alpha-tocopherol increased significantly with age, as well as with total lipid; gamma-tocopherol concentration remained unchanged with age. The data also indicated that subjects with higher serum alpha-tocopherol had lower gamma-tocopherol levels and vice versa for subjects with higher gamma levels. A close interrelationship and competition appeared to exist between these two vitamers.

Modification of sympathetic nervous system activity in rat tissues by dietary vitamin E.

Noradrenaline content and turnover rate were measured in sympathetically innervated tissues of adult male rats that were fed diets varying in alpha-tocopherol for 10 to 11 weeks. At the end of the feeding period, increasing amounts of vitamin E increased the levels of noradrenaline in liver and white adipose tissue but no differences were observed in heart and interscapular brown adipose tissue. By contrast, increased amounts of dietary vitamin E decreased both the rate constant for the decline of specific activity of noradrenaline with time and the noradrenaline turnover rate in heart and brown adipose tissue. No significant differences were observed in these parameters in liver and white adipose tissue. These results indicate that vitamin E in the diet may affect activity of the sympathetic nervous system.

Increased plasma and tissue levels of vitamin E in the spontaneously diabetic BB rat.

Increased plasma and tissue levels of vitamin E were found in spontaneously diabetic BB rats (D) as well as asymptomatic/diabetes-prone BB rats (AD) in comparison to levels in non-diabetic control rats (ND). Treatment of D rats with insulin for 30 days returned plasma and tissue values of vitamin E to control levels. The changes reported here could not be explained solely on the basis of variations in total lipid content of plasma. These data suggest the metabolism of vitamin E is altered in asymptomatic and spontaneously diabetic BB rats and this alteration returns to control values following insulin treatment. Furthermore, it might be speculated that these data indicate a relationship between vitamin E and insulin.

Distribution of alpha-tocopherol in human plasma lipoproteins.

Lipoproteins were removed from human plasma by ultracentrifugation at a density of 1.225. Three classes of lipoproteins were then separated by 4% Agarose-column chromatography: very low density lipoproteins (VLDL), low-density lipoproteins (LDL), and high-density lipoproteins (HDL). A sensitive high performance liquid chromatography method with a fluorescence detector was used to estimate alpha-tocopherol in plasma and in column eluates. Total plasma tocopherol was not significantly different in males (n = 6) and females (n = 6) and almost all of the vitamin was recovered in the isolated lipoproteins. Although LDL and HDL were the main carriers of alpha-tocopherol in both males and females, more tocopherol was found in LDL than in HDL in males but the opposite was true in females. The distribution of alpha-tocopherol in males was: VLDL, 8%; LDL, 59%; and HDL, 33% whereas that in females was VLDL, 2%; LDL, 42%; HDL, 56%. The distribution of protein in lipoprotein from males was: VLDL, 4%, LDL, 37%; and HDL, 59% and in females: VLDL, 2%; LDL, 25%; and HDL, 73%. The alpha-tocopherol concentration (expressed as microgram alpha-tocopherol/mg protein) in lipoproteins differed little between the sexes. The values in males were: VLDL, 7.0; LDL, 4.3; and HDL, 1.5, in females: VLDL, 3.9; LDL, 4.7; and HDL, 2.1. The data suggest that the different distribution of alpha-tocopherol in plasma lipoproteins in males and females is due to the different levels of proteins in those lipoprotein fractions. Overall, tocopherol and protein levels were highly correlated in HDL, a lower correlation was found in LDL.

Ascorbic acid in brown adipose tissue: effect of cold acclimation and high intake of the vitamin.

The ascorbic acid content of brown adipose tissue increased 62% when rats underwent cold acclimation; however, the increase was 126% when rats living in the cold were fed a basal diet supplemented with 2% ascorbic acid. These findings suggest a role for ascorbic acid in this tissue during nonshivering thermogenesis.