Urinary α-carboxyethyl hydroxychroman can be used as a predictor of α-tocopherol adequacy, as demonstrated in the Energetics Study.

BACKGROUND: Other than the in vitro erythrocyte hemolysis test, no valid biomarkers of vitamin E status currently exist. OBJECTIVE: We hypothesized that the urinary vitamin E metabolite α-carboxyethyl hydroxychroman (α-CEHC) could serve as a biomarker. DESIGN: The relations between urinary α-CEHC, plasma α-tocopherol, and vitamin E intakes were assessed by using a previously validated multipass, Web-based, 24-h self-administered dietary recall, and we concurrently collected plasma and 24-h urine samples from 233 participants of both sexes. RESULTS: Median vitamin E intakes were 9.7 mg α-tocopherol/d. Intakes were correlated with plasma α-tocopherol (R = 0.40, P < 0.001) and urinary α-CEHC (R = 0.42, P < 0.001); these correlations were essentially unchanged after multivariate adjustments. On the basis of multiple regression analysis, urinary α-CEHC excretion increased by ~0.086 µmol/g creatinine (95% CI: 0.047, 0.125) for every 1-mg (2.3-µmol) increase in dietary α-tocopherol. Urinary α-CEHC excretion remained at a plateau (median: 1.39 µmol/g creatinine) until dietary intakes of α-tocopherol exceeded 9 mg α-tocopherol/d. The inflection point at which vitamin E metabolism increased was estimated to be at an intake of 12.8 mg α-tocopherol/d. Daily excretion of >1.39 µmol α-CEHC/g creatinine is associated with a greater than adequate α-tocopherol status, as evidenced by increased vitamin E metabolism and excretion. CONCLUSION: Thus, urinary α-CEHC is a valid biomarker of α-tocopherol status that can be used to set a value for the Estimated Adequate Requirement of vitamin E.

Novel metabolites and roles for α-tocopherol in humans and mice discovered by mass spectrometry-based metabolomics.

BACKGROUND: Contradictory results from clinical trials that examined the role of vitamin E in chronic disease could be a consequence of interindividual variation, caused by factors such as xenobiotic use. Cometabolism of vitamin E with other pharmaceutical products could affect the bioavailability of the drug. Thus, it is necessary to understand fully the metabolic routes and biological endpoints of vitamin E. OBJECTIVE: The objective was to uncover novel metabolites and roles of vitamin E in humans and mouse models. DESIGN: Human volunteers (n = 10) were fed almonds for 7 d and then an α-tocopherol dietary supplement for 14 d. Urine and serum samples were collected before and after dosing. C57BL/6 mice (n = 10) were also fed α-tocopherol-deficient and -enriched diets for 14 d. Urine, serum, and feces were collected before and after dosing, and liver samples were collected after euthanization. Ultraperformance liquid chromatography electrospray ionization time-of-flight mass spectrometry and multivariate data analysis tools were used to analyze the samples. RESULTS: Three novel urinary metabolites of α-tocopherol were discovered in humans and mice: α-carboxyethylhydroxychroman (α-CEHC) glycine, α-CEHC glycine glucuronide, and α-CEHC taurine. Another urinary metabolite, α-CEHC glutamine, was discovered in mice after α-CEHC gavage. Increases in liver fatty acids and decreases in serum and liver cholesterol were observed in mice fed the α-tocopherol-enriched diet. CONCLUSION: Novel metabolites and metabolic pathways of vitamin E were identified by mass spectrometry-based metabolomics and will aid in understanding the disposition and roles of vitamin E in vivo.

Wheat germ supplementation of a low vitamin E diet in rats affords effective antioxidant protection in tissues.

BACKGROUND: Oxidative stress is implicated in the etiology of many diseases, but most of clinical trials failed to demonstrate beneficial effects of antioxidant supplementation. METHODS: In the present experiment, we assessed the mean-term effect of wheat germ supplementation, as a dietary source of vitamin E, on antioxidant protection in rat. RESULTS: Feeding rats a 20% wheat germ diet significantly increased plasma and liver vitamin E levels, compared to the low vitamin E basal diet. Concurrently, wheat germ diet consumption strongly decreased the susceptibility of heart and liver lipids to oxidation, as well as the plasma. Wheat germ feeding did not change triglycerides (TG) nor total cholesterol concentrations in plasma or liver, resulting in higher vitamin E/TG ratio compared to controls. Similar results were found with a diet in which wheat germ oil provided the same amount of vitamin E. CONCLUSIONS: Wheat germ appears thus very effective to improve antioxidant defense status, especially in tissues, irrespective of modifications of lipids status.

Urinary aldehydes as indicators of lipid peroxidation in vivo.

The excretion of malondialdehyde (MDA), lipophilic aldehydes and related carbonyl compounds in rat and human urine was investigated. MDA was found to be excreted mainly in the form of two adducts with lysine, indicating that its predominant reaction in vivo is with the lysine residues of proteins. Adducts with the phospholipid bases serine and ethanolamine and the nucleic acid bases guanine and deoxyguanosine also were found. Except for the adduct with deoxyguanosine (dG-MDA), the excretion of these compounds increased with peroxidative stress imposed in the form of vitamin E deficiency or the administration of iron or carbon tetrachloride. Marked differences in the concentration of dG-MDA in different tissues were correlated with their content of fatty acids having three or more double bonds, the putative source of MDA. Fourteen nonpolar and eleven polar lipophilic aldehydes and other carbonyl compounds were identified as their 2,4-diphenylhydrazine derivatives in rat urine. The excretion of five nonpolar and nine polar compounds was increased under conditions of peroxidative stress. The profile of lipophilic aldehydes obtained for human urine resembled that for rat urine. Except for a reported 4-hydroxynon-2-enal conjugate with mercapturic acid, the conjugated forms of the lipophilic aldehydes excreted in urine remain unidentified. Aldehyde excretion is influenced by numerous factors that affect the formation of lipid peroxides in vivo such as energy status, physical activity and environmental temperature, as well as by wide variations in the intake of peroxides in the diet. Consequently, urinalysis for aldehydic products of lipid peroxidation is an unreliable indicator of the general state of peroxidative stress in vivo.

Dietary oxidized frying oil enhances tissue alpha-tocopherol depletion and radioisotope tracer excretion in vitamin E-deficient rats.

Rats fed a diet containing 15% oxidized frying soybean oil (OFO) have been shown to have significantly lower tissue alpha-tocopherol (alpha-T) concentration than rats fed a 15% fresh soybean oil diet. To examine the turnover of alpha-tocopherol, a depletion-repletion experiment and a radioisotope tracer study were conducted. Two groups of male weanling Long-Evans rats were fed vitamin E-deficient diets containing either 15% OFO or 15% vitamin E-stripped fresh soybean oil (control). After 9 wk of depletion, rats fed the OFO diet had significantly higher plasma pyruvate kinase (PK) activity and lower concentrations of alpha-T in RBC, adrenal gland, heart, kidney, liver, spleen, testis and muscle compared with controls (P < 0.05), indicating that the vitamin E-deficient status was aggravated by feeding the OFO diet. After 12 wk, the depleted rats were intraperitoneally injected with a dose of all-rac-alpha-T (2.5 mg/rat, dissolved in Vitamin E-stripped corn oil) every other day. Three doses were administered to each rat during the 1-wk repletion period. Plasma PK activity decreased in both groups (P < 0.05) after repletion but that of the OFO rats was still significantly higher than that of the control group. The repleted OFO gorup also had significantly lower alpha-T concentration in adrenal gland, epididymal fat, liver and spleen than the repleted control group. Two rats from each group that had been vitamin E-depleted for 16 wk were injected intraperitoneally with a single dose of 5-methyl-14C-RRR-alpha-T (740 kBq/kg body weight). During the week after dosing, the radioactivity excreted in urine and feces of the OFO group was 1.3- and 1.7-fold, respectively, that of the control group. Tissue retention of radioactivity was also lower in the OFO rats than in the control rats. The results suggest that more of the alpha-T in the body was catabolized or turned over in rats fed the OFO-containing diet.

Increased formation and degradation of malondialdehyde-modified proteins under conditions of peroxidative stress.

The effect of increased in vivo lipid peroxidation on excretion of the main urinary metabolites of malondialdehyde (MDA) was investigated. peroxidative stress in the form of vitamin E deficiency or the administration of iron nitrilotriacetate or carbon tetrachloride was imposed on rats fed an MDA-free diet. Significant increases were observed in excretion of the lysine-MDA adduct epsilon-propenal lysine, its N-acetyl ester, and free MDA. Under the conditions imposed, the increments in excretion of the lysine adducts reflect increased peroxidative modification of tissue proteins in vivo. These adducts also were found to be the main forms of MDA excreted in human urine. Reacting 14C-bovine serum albumin (BSA) with MDA resulted in its accelerated proteolysis in vitro by soluble enzyme preparations derived from human erythrocytes and rat liver mitochondria. The increments observed were similar to those reported for the hydrolysis of BSA following its exposure to hydroxyl radicals. The results show that lipid peroxidation in vivo results in peroxidative damage to tissue proteins and indicate that such proteins are subject to an accelerated rate of proteolysis.

Identification of a deoxyguanosine-malondialdehyde adduct in rat and human urine.

In an ongoing study, rat and human urine have been examined for the presence of malondialdehyde (MDA) derivatives as indicators of the nature of lipid peroxidative damage caused by this compound in vivo. MDA in urine was found to be present mainly in the form of two lysine adducts, one acetylated and the other unacetylated, reflecting in vivo reactions with tissue proteins. Two minor metabolites were identified as adducts with the phospholipid bases serine and ethanolamine and a third one as an adduct with the nucleic acid base guanine. The identification of an MDA adduct with deoxyguanosine (dG-MDA) among the products of hydrolysis of rat liver DNA suggested the possible occurrence of this compound in urine. In the present study dG-MDA was identified in rat and in human urine, and a high-performance liquid chromatographic method utilizing fluorescence detection was developed for its estimation. The method is sensitive to 1 pmol of dG-MDA and requires a minimum of 1 mL of rat urine or 5 mL of human urine. Its rate of excretion by five-week-old rats (28.54 +/- 2.28 nmol/kg/24 h) (mean +/- SEM) was higher than that for nine-week-old rats (6.29 +/- 1.02) and much higher than that for adult humans (0.40 +/- 0.05). The results indicate that, as reported for 8-hydroxy-deoxyguanosine, dG-MDA excretion is related to metabolic rate. Excretion of dG-MDA by the rat, like the excretion of total MDA, declines during growth on a body weight basis at a rate similar to the decrease in resting energy metabolism.(ABSTRACT TRUNCATED AT 250 WORDS)

The effect of iron overload on urinary excretion of immunoreactive prostaglandin E2.

The effect of in vivo lipid peroxidation on the excretion of immunoreactive prostaglandin E2 (PGE2) in the urine of rats was studied. Weanling, male Sprague-Dawley rats were fed a vitamin E-deficient diet containing 10% tocopherol-stripped corn oil (CO) or 5% cod liver oil (CLO) with or without 40 mg dl-alpha-tocopheryl acetate/kg. To induce a high, sustained level of lipid peroxidation, some rats were injected intraperitoneally with 100 mg of iron as iron dextran after 10 days of feeding. Iron overload stimulated in vivo lipid peroxidation in rats, as measured by the increase in expired ethane and pentane. Dietary vitamin E reversed this effect. Rats fed the CLO diet excreted 9.5-fold more urinary thiobarbituric acid-reactive substances (TBARS) than did rats fed the CO diet. Iron overload increased the excretion of TBARS in the urine of rats fed the CO diet, but not in urine of rats fed the CLO diet. Dietary vitamin E decreased TBARS in the urine of rats fed either the CO or the CLO diet. Iron overload decreased by 40% the urinary excretion of PGE2 by rats fed the CO diet, and dietary vitamin E did not reverse this effect. Iron overload had no statistically significant effect on urinary excretion of PGE2 by rats fed the CLO diet. A high level of lipid peroxidation occurred in iron-treated rats, as evidenced by an increase in alkane production and in TBARS in urine in this study, and by an increase in alkane production by slices of kidney from iron-treated rats in a previous study [V. C. Gavino, C. J. Dillard, and A. L. Tappel (1984) Arch. Biochem. Biophys. 233, 741-747]. Since PGE2 excretion in urine was not correlated with these effects, lipid peroxidation appears not to be a major factor in renal PGE2 flux.

Urinary malondialdehyde as an indicator of lipid peroxidation in the diet and in the tissues.

Although malondialdehyde (MDA) is extensively metabolized to CO2, small amounts are nevertheless excreted in an acid-hydrolyzable form in rat urine. In this study, urinary MDA was evaluated as an indicator of lipid peroxidation in the diet and in the tissues. MDA was released from its bound form(s) in urine by acid treatment and determined as the TBA-MA derivative by HPLC. MDA excretion by the rat was found to be responsive to oral administration of the Na enol salt and to peroxidation of dietary lipids. Urinary MDA also increased in response to the increased lipid peroxidation in vivo produced by vitamin E deficiency and by administration of iron nitrilotriacetate. Chronic feeding of a diet containing cod liver oil led to increases in MDA excretion which were not completely eliminated by fasting or feeding a peroxide-free diet, indicating that there was increased lipid peroxidation in vivo. MDA excretion was not responsive to Se deficiency or CCl4 administration. DPPD, a biologically active antioxidant, but not BHA, a non-biologically active antioxidant, prevented the increase in MDA excretion in vitamin E deficient animals. The results indicate that MDA excretion can serve as an indicator of the extent of lipid peroxidation in the diet and, under conditions which preclude a dietary effect, as an index of lipid peroxidation in vivo.