Gamma-tocopherol attenuates moderate but not severe colitis and suppresses moderate colitis-promoted colon tumorigenesis in mice.

Inflammation can promote colon cancer. Mechanistic studies indicate that γ-tocopherol (γT), a major form of vitamin E in diets, has anti-inflammatory and anticancer properties. Here we investigated the effectiveness of γT and a mixture of tocopherols against colitis and colitis-promoted colon tumorigenesis in male BALB/c mice. γT or mixed tocopherols (at 0.1% diet) did not show any effect on colon tumorigenesis induced by azoxymethane (AOM, 10mg/kg) with three cycles of dextran sodium sulfate (DSS at 1.5-2.5%). γT failed to exhibit protection of severe colitis caused by three cycles of DSS at 2.5%. In contrast, when AOM-initiated carcinogenesis was promoted by relatively mild colitis induced by one-cycle DSS (1.5%), γT, but not mixed tocopherols, suppressed total multiplicity of macroscopic adenomas (P=0.06) and large adenomatous polyps (>2mm(2), P<0.05) by 60 and 85%, respectively. γT also significantly decreased tumor multiplicity (>2mm(2)) induced by AOM with two cycles of 1.5% DSS even when dietary supplementation was started after AOM injection. Consistently, γT but not mixed tocopherols attenuated DSS (1.5%)-induced colon inflammation and damage as well as formation of atypical glandular hyperplasia. Mice supplemented with tocopherols had high fecal excretion of 13'-carboxychromanol, a long-chain vitamin E metabolite shown to have potent anti-inflammatory activities. Our study demonstrates that γT is able to alleviate moderate but not severe colitis and its promoted tumorigenesis, and indicates that inflammation severity should be considered in evaluating anticancer effectiveness of chemoprevention agents.

Comparison of the intestinal absorption and bioavailability of γ-tocotrienol and α-tocopherol: in vitro, in situ and in vivo studies.

The aim of this work was to compare the intestinal absorption kinetics and the bioavailability of γ-tocotrienol (γ-T3) and α-tocopherol (α-Tph) administered separately as oil solutions to rats in vivo. Also, to explain the significant difference in the oral bioavailability of the compounds: (1) the release profiles using the dynamic in vitro lipolysis model, (2) the intestinal permeability and (3) carrier-mediated uptake by Niemann-Pick C1-like 1 (NPC1L1) transporter were examined. Absolute bioavailability studies were conducted after oral administration of γ-T3 or α-Tph prepared in corn oil to rats. In situ rat intestinal perfusion with ezetimibe (a NPC1L1 inhibitor) was performed to compare intestinal permeability. The in vitro interaction kinetics with NPC1L1 was examined in NPC1L1 transfected cells. While the in vitro release studies demonstrated a significantly higher release rate of γ-T3 in the aqueous phase, the oral bioavailability of α-Tph (36%) was significantly higher than γ-T3 (9%). Consequent in situ studies revealed significantly higher intestinal permeability for α-Tph compared with γ-T3 in rats. Moreover, the NPC1L1 kinetic studies demonstrated higher Vmax and Km values for α-Tph compared with γ-T3. Collectively, these results indicate that intestinal permeability is the main contributing factor for the higher bioavailability of α-Tph. Also, these results emphasize the potentially important role of intestinal permeability in the bioavailability of γ-T3, suggesting that enhancing its permeability would increase its oral bioavailability.

Gamma-tocotrienol and gamma-tocopherol are primarily metabolized to conjugated 2-(beta-carboxyethyl)-6-hydroxy-2,7,8-trimethylchroman and sulfated long-chain carboxychromanols in rats.

The metabolism of gamma-tocotrienol (gamma-TE) and gamma-tocopherol (gamma-T) was investigated in human A549 cells and in rats. Similar to gamma-T, A549 cells metabolized gamma-TE to sulfated 9'-, 11'-, and 13'-carboxychromanol and their unconjugated counterparts. After 72-h incubation with the cells, 90% of long-chain carboxychromanols in the culture media from gamma-TE, but <45% from gamma-T, were in the sulfated form. The formation of these metabolites was further investigated in rats gavaged by gamma-TE at 10 or 50 mg/kg, gamma-T at 10 mg/kg, or tocopherol-stripped corn oil in controls. Six hours after a single dosing, the supplemented rats had increased plasma concentrations of 13'-carboxychromanol and sulfated 9'-, 11'-, 13'-carboxychromanol, whereas none of these metabolites were detectable in the controls. Sulfated 11'-carboxychromanol was the most abundant long-chain metabolite in gamma-TE-supplemented rats. Sulfatase/glucuronidase hydrolysis revealed for the first time that >88% 2-(beta-carboxyethyl)-6-hydroxychroman (gamma-CEHC), the terminal beta-oxidation metabolite, was in the conjugated form in the plasma. In all groups, conjugated gamma-CEHC accounted for >75% of total metabolites, whereas free CEHC was a minor metabolite. At 10 mg/kg, the plasma concentrations of total metabolites from gamma-TE-supplemented rats were higher (P < 0.05) than those from gamma-T-fed rats. These results demonstrate that in rats, conjugation such as sulfation occurs parallel to beta-oxidation in the liver and is quantitatively important to vitamin E metabolism. Conjugated long-chain carboxychromanols may be novel excreted metabolites during supplementation. Our data also provide in vivo evidence that gamma-TE is more extensively metabolized than gamma-T.

Nanoemulsions of an anti-oxidant synergy formulation containing gamma tocopherol have enhanced bioavailability and anti-inflammatory properties.

The present study investigated whether MicroFluidizer Processor-based nanoemulsions of an antioxidant synergy formulation (ASF), containing delta, alpha and gamma tocopherol influenced inflammation and bioavailability in CD-1 mice. Croton oil was applied to all animals' right ear lobe to induce inflammation. Auricular thickness was measured after 2 and 6h after the various treatments. The animal plasma and ear lobes were collected and frozen for bioavailability and cytokine analyses. The ASF nanoemulsions of alpha, delta, or gamma tocopherol significantly reduced auricular thickness compared to control (57, -57, and -71%, respectively) and blank nanoemulsion (-50, -50, -67%, respectively). Relative to the suspensions of ASF, only the nanoemulsion of ASF containing gamma tocopherol significantly reduced auricular thickness (-60%), whereas the 40% reduction with nanoemulsions of delta tocopherol compared to suspension was not statistically significant. Auricular concentrations of cytokines TNF-alpha and IL-1 alpha were significantly reduced in mice treated only with ASF nanoemulsions of gamma tocopherol compared to control (-53, -46%, respectively) and blank nanoemulsion (-52, -46%, respectively). Auricular thickness was significantly associated with tissue TNF-alpha (r=0.539, p<0.001) and IL-1 alpha concentrations (r=0.404, p=0.01). Bioavailability for gamma and delta was dramatically enhanced (2.2- and 2.4-folds) with the nanoemulsion compared to suspensions. Only the plasma gamma tocopherol concentration was significantly associated with auricular thickness (r=-0.643, P=0.001). In conclusion, nanoemulsions of ASF containing gamma, alpha, and delta tocopherol, have enhanced anti-inflammatory properties and increased bioavailability, with gamma tocopherol, in particular compared to their suspensions.

A pilot study on the safety and efficacy of a novel antioxidant rich formulation in patients with cystic fibrosis.

BACKGROUND: Pancreatic insufficiency and a diminished bile acid pool cause malabsorption of important essential nutrients and other dietary components in cystic fibrosis (CF). Of particular significance is the malabsorption of fat-soluble antioxidants such as carotenoids, tocopherols and coenzyme Q(10) (CoQ(10)). Despite supplementation, CF patients are often deficient in these compounds, resulting in increased oxidative stress, which may contribute to adverse health effects. This pilot study was designed to evaluate the safety of a novel micellar formulation (CF-1) of fat-soluble nutrients and antioxidants and to determine its efficacy in improving plasma levels of these compounds and reducing inflammatory markers in induced sputum. METHODS: Ten CF subjects, ages 8 to 45 years old, were given orally 10 ml of the CF-1 formulation daily for 56 days after a 21-day washout period in which subjects stopped supplemental vitamin use except for a standard multivitamin. Plasma obtained at -3, 0 (baseline), 1, 2, 4, and 8 weeks was assayed for beta-carotene, gamma-tocopherol, retinol, and CoQ(10) as well as for safety parameters (comprehensive metabolic panel and complete blood count). In addition, pulmonary function was measured and induced sputum was assayed for markers of inflammation and quantitative bacterial counts both prior and during dosing. RESULTS: No serious adverse effects, laboratory abnormalities or elevated nutrient levels (above normal) were identified as related to CF-1. Supplementation with CF-1 significantly increased beta-carotene levels at all dosing time points when compared to screening and baseline. In addition, gamma-tocopherol and CoQ(10) significantly increased from baseline in all subjects. Induced sputum myeloperoxidase significantly decreased and there was a trend toward decreases in PMN elastase and total cell counts with CF-1. There was a significant inverse correlation between the antioxidant levels and induced sputum changes in IL-8 and total neutrophils. Lung function and sputum bacterial counts were unchanged. CONCLUSION: The novel CF-1 formulation safely and effectively increased plasma levels of important fat-soluble nutrients and antioxidants. In addition, improvements in antioxidant plasma levels were associated with reductions in airway inflammation in CF patients.

Bioavailability of a novel, water-soluble vitamin E formulation in malabsorbing patients.

In cystic fibrosis (CF), pancreatic insufficiency and a diminished bile acid pool cause malabsorption of important nutrients and dietary components leading to deficiency, poor nutritional status, and oxidative stress. Of particular significance is the malabsorption of fat-soluble nutrients and antioxidants, which are important for normal immune and neurologic function. Patients with CF often are deficient in these compounds despite supplementation with the current standard of care therapy. The objective was to compare the pharmacokinetic profile of this water-soluble vitamin E formulation (Aqua-E) with an oil-based softgel formulation in a malabsorbing patient population. Patients with CF who had documented malabsorption were recruited for participation in this pharmacokinetic study. Patients who met inclusion and exclusion criteria discontinued vitamin E supplementation, except for that in a multivitamin, for 7 to 21 days before the day of dosing. Patients were randomized to a single dose of 20 ml of Aqua-E or three oil-based softgels, which contained equivalent amounts of tocopherols. Blood was drawn from patients at time 0, 2, 4, 8, 24, 48, and 168 hr and analyzed for tocopherols. Eight patients were enrolled in the study and randomized to Aqua-E or softgels. The primary outcome, the absorption of gamma-tocopherol in Aqua-E (AUC=115 micro g/ml(*)hr), was significantly greater than that of oil-based softgels (AUC=25.3 micro g/ml(*)hr; P=0.013). Total-tocopherols (alpha+gamma+delta) in Aqua-E (AUC=294 micro g/ml(*)hr) showed a strong trend toward increased absorption compared with that of oil-based softgels (AUC=117 micro g/ml(*)hr; P=0.09). In conclusion, this novel, water-soluble formulation showed a marked and statistically significant increase in absorption of gamma-tocopherol in malabsorbing patients with CF compared with an oil-based formulation.

How an increased intake of alpha-tocopherol can suppress the bioavailability of gamma-tocopherol.

alpha-Tocopherol is the only form of vitamin E in vitamin supplements, whereas gamma-tocopherol is the predominant form of vitamin E in the US diet. gamma-Tocopherol has beneficial properties as an anti-inflammatory and possibly anti-atherogenic and anticancer agent. Excess a-tocopherol taken in supplements causes a reduction of gamma-tocopherol concentration in plasma. The biochemical mechanism of this effect, which is important to human nutrition, has recently been elucidated.

Vitamin E attenuates acute lung injury in sheep with burn and smoke inhalation injury.

INTRODUCTION: A decrease in alpha-tocopherol (vitamin E) plasma levels in burn patients is typically associated with increased mortality. We hypothesized that vitamin E supplementation (alpha-tocopherol) would attenuate acute lung injury induced by burn and smoke inhalation injury. MATERIALS AND METHODS: Under deep anesthesia, sheep (33 +/- 5 kg) were subjected to a flame burn (40% total body surface area, third degree) and inhalation injury (48 breaths of cotton smoke, < 40 degrees C). Half of the injured group received alpha-tocopherol (1000 IU vitamin E) orally, 24 h prior to injury. The sham group was neither injured nor given vitamin E. All three groups (n = 5 per group) were resuscitated with Ringer's lactate solution (4 ml/kg/%burn/24 h), and placed on a ventilator (PEEP = 5 cmH2O; tidal volume = 15 ml/kg) for 48 h. RESULTS: Plasma alpha-tocopherol per lipids doubled in the vitamin E treated sheep. Vitamin E treatment prior to injury largely prevented the increase in pulmonary permeability index and moderated the increase in lung lymph flow (52.6 +/- 6.2 ml/min, compared with 27.3 +/- 6.0 ml/min, respectively), increased the PaO2/FiO2 ratio, ameliorated both peak and pause airway pressure increases, and decreased plasma conjugated dienes and nitrotyrosine. CONCLUSIONS: Pretreatment with vitamin E ameliorated the acute lung injury caused by burn and smoke inhalation exposure.

Failure of vitamin E in clinical trials: is gamma-tocopherol the answer?

Oxidative stress and inflammation play a crucial role in atherosclerosis. However, prospective clinical trials of dietary antioxidants with anti-inflammatory properties, such as alpha-tocopherol (AT), have not yielded positive results. AT supplementation decreases gamma-tocopherol (GT) levels. GT is an antioxidant with potent anti-inflammatory activity, and plasma GT levels are inversely associated with cardiovascular diseases. Thus, studies using pure GT, alone or in conjunction with AT, will elucidate its utility in cardiovascular disease prevention.

gamma-Tocopherol biokinetics and transformation in humans.

BACKGROUND: The uptake and biotransformation of gamma-tocopherol (gamma-T) in humans is largely unknown. Using a stable isotope method we investigated these aspects of gamma-T biology in healthy volunteers and their response to gamma-T supplementation. METHODS: A single bolus of 100 mg of deuterium labeled gamma-T acetate (d(2)-gamma-TAC, 94% isotopic purity) was administered with a standard meal to 21 healthy subjects. Blood and urine (first morning void) were collected at baseline and a range of time points between 6 and 240 h post-supplemetation. The concentrations of d(2) and d(0)-gamma-T in plasma and its major metabolite 2,7,8-trimethyl-2-(b-carboxyethyl)-6-hydroxychroman (-gamma-CEHC) in plasma and urine were measured by GC-MS. In two subjects, the total urine volume was collected for 72 h post-supplementation. The effects of gamma-T supplementation on alpha-T concentrations in plasma and alpha-T and gamma-T metabolite formation were also assessed by HPLC or GC-MS analysis. RESULTS: At baseline, mean plasma alpha-T concentration was approximately 15 times higher than gamma-T (28.3 vs. 1.9 micromol/l). In contrast, plasma gamma-CEHC concentration (0.191 micromol/l) was 12 fold greater than alpha-CEHC (0.016 micromol/l) while in urine it was 3.5 fold lower (0.82 and 2.87 micromol, respectively) suggesting that the clearance of alpha-CEHC from plasma was more than 40 times that of gamma-CEHC. After d(2)-gamma-TAC administration, the d(2) forms of gamma-T and gamma-CEHC in plasma and urine increased, but with marked inter-individual variability, while the d(0) species were hardly affected. Mean total concentrations of gamma-T and gamma-CEHC in plasma and urine peaked, respectively, between 0-9, 6-12 and 9-24 h post-supplementation with increases over baseline levels of 6-14 fold. All these parameters returned to baseline by 72 h. Following challenge, the total urinary excretion of d(2)-gamma-T equivalents was approximately 7 mg. Baseline levels of gamma-T correlated positively with the post-supplementation rise of (d(0) + d(2)) - gamma - T and gamma-CEHC levels in plasma, but correlated negatively with urinary levels of (d(0) + d(2))-gamma-CEHC. Supplementation with 100 mg gamma-TAC had minimal influence on plasma concentrations of alpha-T and alpha-T-related metabolite formation and excretion. CONCLUSIONS: Ingestion of 100mg of gamma-TAC transiently increases plasma concentrations of gamma-T as it undergoes sustained catabolism to CEHC without markedly influencing the pre-existing plasma pool of gamma-T nor the concentration and metabolism of alpha-T. These pathways appear tightly regulated, most probably to keep high steady-state blood ratios alpha-T to gamma-T and gamma-CEHC to alpha-CEHC.