Vitamina E - Coletânea de 67 resumos

Fabio Gonçalves Gonzales

Mais 67 referências para aprofundar os estudos ...

1.J Gerontol A Biol Sci Med Sci 2000 Jun;55(6):B280-5

Effects of tocotrienols on life span and protein carbonylation in Caenorhabditis elegans.

Adachi H, Ishii N.

Life Science Research Center, Lion Corporation, Kanagawa, Japan. hadachi@lion.co.jp

To assess the efficiency of tocotrienols against oxidative damage, we have demonstrated in a

model-system nematode, Caenorhabditis elegans, that tocotrienol administration reduced the

accumulation of protein carbonyl (a good indicator of oxidative damage during aging) and

consequently extended the mean life span (LS), but not the maximum LS. Conversely, alpha-

tocopherol acetate did not affect these parameters. As a way to evaluate the protective ability of

tocotrienols against oxidative stress, the life spans of animals administrated tocotrienols before or

after exposure to ultraviolet B-induced oxidative stress were measured. Ultraviolet B irradiation

shortened the mean LS of animals, whereas preadministration of tocotrienols recovered the mean

LS to that of unirradiated animals. Interestingly, postadministration also extended the mean LS

more than that of unirradiated animals, and administration through the LS conferred greater

protection. Thus, the administration of tocotrienols to animals results in a reduction of oxidative

stress risks. These data indicated that tocotrienols merit further investigation as possible agents for

antiaging and oxidative stress prevention. In addition, they suggest that C. elegans will continue to

provide provocative clues into the mechanisms of aging.

2. N Engl J Med 1993 May 20;328(20):1450-6

Vitamin E consumption and the risk of coronary heart disease in men.

Rimm EB, Stampfer MJ, Ascherio A, Giovannucci E, Colditz GA, Willett WC.

Department of Epidemiology, Harvard School of Public Health, Boston, MA 02115.

BACKGROUND. The oxidative modification of low-density lipoproteins increases their

incorporation into the arterial intima, an essential step in atherogenesis. Although dietary

antioxidants, such as vitamin C, carotene, and vitamin E, have been hypothesized to prevent

coronary heart disease, prospective epidemiologic data are sparse. METHODS. In 1986, 39,910

U.S. male health professionals 40 to 75 years of age who were free of diagnosed coronary heart

disease, diabetes, and hypercholesterolemia completed detailed dietary questionnaires that assessed

their usual intake of vitamin C, carotene, and vitamin E in addition to other nutrients. During four

years of follow-up, we documented 667 cases of coronary disease. RESULTS. After controlling for

age and several coronary risk factors, we observed a lower risk of coronary disease among men

with higher intakes of vitamin E (P for trend = 0.003). For men consuming more than 60 IU per day

of vitamin E, the multivariate relative risk was 0.64 (95 percent confidence interval, 0.49 to 0.83) as

compared with those consuming less than 7.5 IU per day. As compared with men who did not take

vitamin E supplements, men who took at least 100 IU per day for at least two years had a

multivariate relative risk of coronary disease of 0.63 (95 percent confidence interval, 0.47 to 0.84).

Carotene intake was not associated with a lower risk of coronary disease among those who had

never smoked, but it was inversely associated with the risk among current smokers (relative risk,

0.30; 95 percent confidence interval, 0.11 to 0.82) and former smokers (relative risk, 0.60; 95

percent confidence interval, 0.38 to 0.94). In contrast, a high intake of vitamin C was not associated

with a lower risk of coronary disease. CONCLUSIONS. These data do not prove a causal relation,

but they provide evidence of an association between a high intake of vitamin E and a lower risk of

coronary heart disease in men. Public policy recommendations with regard to the use of vitamin E

supplements should await the results of additional studies.

3. N Engl J Med 1993 May 20;328(20):1444-9

Vitamin E consumption and the risk of coronary disease in women.

Stampfer MJ, Hennekens CH, Manson JE, Colditz GA, Rosner B, Willett WC.

Channing Laboratory, Boston, MA 02115.

BACKGROUND. Interest in the antioxidant vitamin E as a possible protective nutrient against

coronary disease has intensified with the recognition that oxidized low-density lipoprotein may be

involved in atherogenesis. METHODS. In 1980, 87,245 female nurses 34 to 59 years of age who

were free of diagnosed cardiovascular disease and cancer completed dietary questionnaires that

assessed their consumption of a wide range of nutrients, including vitamin E. During follow-up of

up to eight years (679,485 person-years) that was 97 percent complete, we documented 552 cases of

major coronary disease (437 nonfatal myocardial infarctions and 115 deaths due to coronary

disease). RESULTS. As compared with women in the lowest fifth of the cohort with respect to

vitamin E intake, those in the top fifth had a relative risk of major coronary disease of 0.66 (95

percent confidence interval, 0.50 to 0.87) after adjustment for age and smoking. Further adjustment

for a variety of other coronary risk factors and nutrients, including other antioxidants, had little

effect on the results. Most of the variability in intake and reduction in risk was attributable to

vitamin E consumed as supplements. Women who took vitamin E supplements for short periods

had little apparent benefit, but those who took them for more than two years had a relative risk of

major coronary disease of 0.59 (95 percent confidence interval, 0.38 to 0.91) after adjustment for

age, smoking status, risk factors for coronary disease, and use of other antioxidant nutrients

(including multi-vitamins). CONCLUSIONS. Although these prospective data do not prove a

cause-and-effect relation, they suggest that among middle-aged women the use of vitamin E

supplements is associated with a reduced risk of coronary heart disease. Randomized trials of

vitamin E in the primary and secondary prevention of coronary disease are being conducted; public

policy recommendations about the widespread use of vitamin E should await the results of these

trials.

4. Lancet 1996 Mar 23;347(9004):781-6

Randomised controlled trial of vitamin E in patients with coronary disease:

Cambridge Heart Antioxidant Study (CHAOS)

Stephens NG, Parsons A, Schofield PM, Kelly F, Cheeseman K, Mitchinson MJ.

Department of Medicine, Cambridge University.

BACKGROUND: Vitamin E (alpha-tocopherol) is thought to have a role in prevention of

atherosclerosis, through inhibition of oxidation of low-density lipoprotein. Some epidemiological

studies have shown an association between high dietary intake or high serum concentrations of

alpha-tocopherol and lower rates of ischaemic heart disease. We tested the hypothesis that treatment

with a high dose of alpha-tocopherol would reduce subsequent risk of myocardial infarction (MI)

and cardiovascular death in patients with established ischaemic heart disease. METHODS: In this

double-blind, placebo-controlled study with stratified randomisation, 2002 patients with

angiographically proven coronary atherosclerosis were enrolled and followed up for a median of

510 days (range 3-981). 1035 patients were assigned alpha-tocopherol (capsules containing 800 IU

daily for first 546 patients; 400 IU daily for remainder); 967 received identical placebo capsules.

The primary endpoints were a combination of cardiovascular death and non-fatal MI as well as non-

fatal MI alone. FINDINGS: Plasma alpha-tocopherol concentrations (measured in subsets of

patients) rose in the actively treated group (from baseline mean 34.2 micromol/L to 51.1

micromol/L with 400 IU daily and 64.5 micromol/L with 800 IU daily) but did not change in the

placebo group. Alpha-tocopherol treatment significantly reduced the risk of the primary trial

endpoint of cardiovascular death and non-fatal MI (41 vs 64 events; relative risk 0.53 [95% Cl 0.34-

0.83; p=0.005). The beneficial effects on this composite endpoint were due to a significant

reduction in the risk of non-fatal MI (14 vs 41; 0.23 [0.11-0.47]; p=0.005); however, there was a

non-significant excess of cardiovascular deaths in the alpha-tocopherol group (27 vs 23; 1.18 [0.62-

2.27]; p=0.61). All-cause mortality was 36 of 1035 alpha-tocopherol-treated patients and 27 of 967

placebo recipients. INTERPRETATION: We conclude that in patients with angiographically

proven symptomatic coronary atherosclerosis, alpha-tocopherol treatment substantially reduces the

rate of non-fatal MI, with beneficial effects apparent after 1 year of treatment. The effect of alpha-

tocopherol treatment on cardiovascular deaths requires further study.

5. JAMA 2001 Mar 7;285(9):1178-82

Effects of vitamin E on lipid peroxidation in healthy persons.

Meagher EA, Barry OP, Lawson JA, Rokach J, FitzGerald GA.

Center for Experimental Therapeutics, 811 Biomedical Research Bldg II/III,

University of Pennsylvania, 421 Curie Blvd, Philadelphia, PA 19104-6160, USA.

CONTEXT: Oxidative stress may play a role in the development or exacerbation of many common

diseases. However, results of prospective controlled trials of the effects of antioxidants such as

vitamin E are contradictory. OBJECTIVE: To assess the effects of supplemental vitamin E on lipid

peroxidation in vivo in healthy adults. DESIGN: Randomized, double-blind, placebo-controlled

trial conducted March 1999 to June 2000. SETTING: A general clinical research center in a tertiary

referral academic medical center. PARTICIPANTS: Thirty healthy men and women aged 18 to 60

years. INTERVENTIONS: Participants were randomly assigned to receive placebo or alpha-

tocopherol dosages of 200, 400, 800, 1200, or 2000 IU/d for 8 weeks (n = 5 in each group),

followed by an 8-week washout period. MAIN OUTCOME MEASURES: Three indices of lipid

peroxidation, urinary 4-hydroxynonenal (4-HNE) and 2 isoprostanes, iPF(2alpha)-III and

iPF(2alpha)-VI, measured by gas chromatography/mass spectrometry and compared among the 6

groups at baseline, 2, 4, 6, and 8 weeks, and 1, 3, and 8 weeks after discontinuation. RESULTS:

Circulating vitamin E levels increased in a dose-dependent manner during the study. No significant

effect of vitamin E on levels of urinary 4-HNE or either isoprostane was observed. Mean (SEM)

baseline vs week 8 levels of iPF(2alpha)-III were 154 (20.1) vs 168 (22.3) pg/mg of creatinine for

subjects taking placebo; 165 (19.6) vs 234 (30.1) pg/mg for those taking 200 IU/d of vitamin E; and

195 (26.7) vs 213 (40.6) pg/mg for subjects taking 2000 IU/d. Corresponding iPF(2alpha)-VI levels

were 1.43 (0.6) vs 1.62 (0.4) ng/mg of

creatinine for subjects taking placebo; 1.64 (0.3) vs 1.24 (0.8) ng/mg for those taking 200 IU/d of

vitamin E; and 1.83 (0.3) vs 1.94 (0.9) ng/mg for those taking 2000 IU/d. Baseline vs week 8 levels

of 4-HNE were 0.5 (0.04) vs 0.4 (0.05) ng/mg of creatinine for subjects taking placebo; 0.4 (0.06)

vs 0.5 (0.02) ng/mg with 200 IU/d of vitamin E; and 0.2 (0.02) vs 0.2 (0.1) ng/mg with 2000 IU/d.

CONCLUSIONS: Our results question the rationale for vitamin E supplementation in healthy

individuals. Specific quantitative indices of oxidative stress in vivo should be considered as entry

criteria and for dose selection in clinical trials of antioxidant drugs and vitamins in human disease.

6. JAMA 1995 Jun 21;273(23):1849-54

Serial coronary angiographic evidence that antioxidant vitamin intake reduces progression of

coronary artery atherosclerosis.

Hodis HN, Mack WJ, LaBree L, Cashin-Hemphill L, Sevanian A, Johnson R, Azen SP.

Atherosclerosis Research Unit, University of Southern California School of Medicine, Los Angeles

90033, USA.

OBJECTIVE--To explore the association of supplementary and dietary vitamin E and C intake with

the progression of coronary artery disease. DESIGN--A subgroup analysis of the on-trial

antioxidant vitamin intake database acquired in the Cholesterol Lowering Atherosclerosis Study, a

randomized, placebo-controlled, serial angiographic clinical trial evaluating the risk and benefit of

colestipol-niacin on coronary artery disease progression. SETTING--Community-and university-

based cardiac catheterization laboratories. SUBJECTS--A total of 156 men aged 40 to 59 years with

previous coronary artery bypass graft surgery. INTERVENTION--Supplementary and dietary

vitamin E and C intake (nonrandomized) in association with cholesterol-lowering diet and either

colestipol-niacin or placebo (randomized). OUTCOME--Change per subject in the percentage of

vessel diameter obstructed because of stenosis (%S) determined by quantitative coronary

angiography after 2 years of randomized therapy on all lesions, mild/moderate lesions (< 50%S),

and severe lesions (> or = 50%S). RESULTS--Overall, subjects with supplementary vitamin E

intake of 100 IU per day or greater demonstrated less coronary artery lesion progression than did

subjects with supplementary vitamin E intake less than 100 IU per day for all lesions (P = .04) and

for mild/moderate lesions (P = .01). Within the drug group, benefit of supplementary vitamin E

intake was found for all lesions (P = .02) and mild/moderate lesions

(P = .01). Within the placebo group, benefit of supplementary vitamin E intake was not found. No

benefit was found for use of supplementary vitamin C exclusively or in conjunction with

supplementary vitamin E, use of multivitamins, or increased dietary intake of vitamin E or vitamin

C. CONCLUSIONS--These results indicate an association between supplementary vitamin E intake

and angiographically demonstrated reduction in coronary artery lesion progression. Verification

from carefully designed, randomized, serial arterial imaging end point trials is needed.

7. Eur Heart J 2001 Jan;22(2):103-4

Clinical, public health, and research implications of the Heart Outcomes

Prevention Evaluation (HOPE) Study.

Yusuf S.

8. Am J Clin Nutr 1996 Mar;63(3):377-85

Inverse relation between the concentration of low-density-lipoprotein vitamin E and severity of

coronary artery disease.

Regnstrom J, Nilsson J, Moldeus P, Strom K, Bavenholm P, Tornvall P, Hamsten A.

Department of Medicine, the King Gustaf V Research Institute, Karolinska Hospital, Stockholm,

Sweden.

Oxidation of low-density lipoprotein (LDL) is believed to play an important role in atherogenesis,

and antioxidant vitamins are thought to protect against coronary artery disease (CAD). We

investigated whether the vitamin E concentrations in serum and LDL were associated with the

severity of CAD as assessed by a semiquantitative scoring system in which coronary angiograms

are analyzed for the number and size of distinct stenotic lesions (global stenosis score). The study

group consisted of 64 consecutive male survivors of myocardial infarction aged < 45 y. Lipid-

adjusted serum and LDL vitamin E concentrations were significantly lower in the patients than in

35 age-matched male control subjects, whereas the absolute serum and LDL vitamin E

concentrations did not differ significantly. No associations were found between the serum

concentration or lipid-adjusted serum values of vitamin E and the stenosis score. In contrast,

significant inverse correlation was found between the LDL vitamin E concentration, whether

adjusted to the lipid (r=-0.477,P<0.001) or protein (r=-0.375, P<0.01) content of LDL, and the

global coronary stenosis score. We conclude that a low LDL vitamin E concentration might play a

role in the development of stenoses in coronary arteries and may contribute to clinically manifest

CAD.

9. N Engl J Med 1996 May 2;334(18):1156-62

Dietary antioxidant vitamins and death from coronary heart disease in postmenopausal women.

Kushi LH, Folsom AR, Prineas RJ, Mink PJ, Wu Y, Bostick RM.

Division of Epidemiology, University of Minnesota School of Public Health,

Minneapolis 55454-1015, USA.

BACKGROUND: The role of dietary antioxidant vitamins in preventing coronary heart disease has

aroused considerable interest because of the knowledge that oxidative modification of low-density

lipoprotein may promote atherosclerosis. METHODS. We studied 34,486 postmenopausal women

with no cardiovascular disease who in early 1986 completed a questionnaire that assessed, among

other factors, their intake of vitamins A, E, and C from food sources and supplements. During

approximately seven years of follow-up (ending December 31, 1992), 242 of the women died of

coronary heart disease. RESULTS. In analyses adjusted for age and dietary energy intake, vitamin

E consumption appeared to be inversely associated with the risk of death from coronary heart

disease. This association was particularly striking in the subgroup of 21,809 women who did not

consume vitamin supplements (relative risks from lowest to highest quintile of vitamin E intake,

1.0, 0.68, 0.71, 0.42, and 0.42; P for trend 0.008). After adjustment for possible confounding

variables, this inverse association remained (relative risks from lowest to highest quintile, 1.0, 0.70,

0.76, 0.32, and 0.38; P for trend, 0.004). There was little evidence that the intake of vitamin E from

supplements was associated with a decreased risk of death from coronary heart disease, but the

effects of high-dose supplementation and the duration of supplement use could not be definitely

addressed. Intake of vitamins A and C did not appear to be associated with the risk of death form

coronary heart disease. CONCLUSIONS. These results suggest that in postmenopausal women the

intake of vitamin E from food is inversely associated with the risk of death from coronary heart

disease and that such women can lower their risk without using vitamin supplements. By contrast,

the intake of vitamins A and C was not associated with lower risks of dying from coronary disease.

10. Am J Epidemiol 1994 Jun 15;139(12):1180-9

Antioxidant vitamin intake and coronary mortality in a longitudinal population study.

Knekt P, Reunanen A, Jarvinen R, Seppanen R, Heliovaara M, Aromaa A.

Social Insurance Institution, Helsinki, Finland.

Oxidation of lipoproteins is hypothesized to promote atherosclerosis and, thus, a high intake of

antioxidant nutrients may protect against coronary heart disease. The relation between the intakes of

dietary carotene, vitamin C, and vitamin E and the subsequent coronary mortality was studied in a

cohort of 5,133 Finnish men and women aged 30-69 years and initially free from heart disease.

Food consumption was estimated by the dietary history method covering the total habitual diet

during the previous year. Altogether, 244 new fatal coronary heart disease cases occurred during a

mean follow-up of 14 years beginning in 1966-1972. An inverse association was observed between

dietary vitamin E intake and coronary mortality in both men and women with relative risks of 0.68

(p for trend = 0.01) and 0.35 (p for trend < 0.01), respectively, between the highest and lowest

tertiles of the intake. Similar associations were observed for the dietary intake of vitamin C and

carotenoids among women and for the intake of important food sources of these micronutrients, i.e.,

of vegetables and fruits, among both men and women. The associations were not attributable to

confounding by major nondietary risk factors of coronary heart disease, i.e., age, smoking, serum

cholesterol, hypertension, or relative weight. The results support the hypothesis that antioxidant

vitamins protect against coronary heart disease, but it cannot be excluded that foods rich in these

micronutrients also contain other constituents that provide the protection.

11. Sources And Consumption Of Antioxidants In The Diet

Bieri J G

J Am Oil Chem Soc 61 (12). 1984. 1917-1918. 1984

12. J Nutr 1985 Jun;115(6):807-13

Oral alpha-tocopherol supplements decrease plasma gamma-tocopherol levels in humans.

Handelman GJ, Machlin LJ, Fitch K, Weiter JJ, Dratz EA.

In a cross-sectional survey of 86 elderly persons, it was observed that subjects with elevated plasma

alpha-tocopherol levels had depressed plasma gamma-tocopherol. Tocopherols were measured by

both reverse-phase and normal-phase high performance liquid chromatography (HPLC). When

eight human volunteers (age range 30-60) were given 1200 IU of all-rac-alpha-tocopherol daily for

8 wk, plasma gamma-tocopherol and beta-tocopherol decreased in all subjects. After

supplementation, gamma-tocopherol values were typically 30-50% of initial values, and alpha-

tocopherol values were typically 200-400% of initial values. These results suggest that intestinal

uptake and/or plasma transport make more efficient use of alpha-tocopherol than of gamma- or

beta-tocopherol. Moreover, the results indicate that the ratio of gamma- to alpha-tocopherol in

plasma would be a more satisfactory index to measure compliance in trials involving

supplementation with alpha-tocopherol.

13. J Intern Med 1996 Feb;239(2):111-7

Gamma, but not alpha, tocopherol levels in serum are reduced in coronary heart disease patients.

Ohrvall M, Sundlof G, Vessby B.

Department of Geriatrics, University of Uppsala, Sweden.

OBJECTIVES. Low concentrations of alpha tocopherol are claimed to be associated with an

increased prevalence of coronary heart disease. This study was undertaken to see whether

measurements of serum tocopherol concentrations can contribute to discrimination between

subjects with and without coronary heart disease. SETTING. All patients had been referred to the

department of cardiology of the University Hospital in Uppsala, Sweden. SUBJECTS. Male

patients (n = 69) below 60 years of age with coronary heart disease (CHD) and healthy age-matched

reference subjects (n = 138) were compared. RESULTS. Lipid-corrected alpha tocopherol

concentrations did not differ significantly between the groups, but the CHD group had a lower

mean concentration of gamma tocopherol and a higher alpha/gamma ratio. In a stepwise logistic

regression analysis, the LDL/HDL ratio was the best independent discriminator between the groups,

followed by the proportion of palmitic acid in the cholesterol esters and the alpha/gamma

tocopherol ratio. CONCLUSIONS. The lower gamma tocopherol concentration and the high ratio

between alpha and gamma tocopherol in the CHD group indicate a difference in antioxidative status

between CHD patients and healthy subjects. The lipid-lowering treatment of these CHD patients is

far from optimal.

14. Proc Natl Acad Sci U S A 1997 Apr 1;94(7):3217-22

Gamma-tocopherol traps mutagenic electrophiles such as NO(X) and complements alpha-

tocopherol: physiological implications.

Christen S, Woodall AA, Shigenaga MK, Southwell-Keely PT, Duncan MW, Ames BN.

Division of Biochemistry and Molecular Biology, University of California, Berkeley 94720, USA.

Peroxynitrite, a powerful mutagenic oxidant and nitrating species, is formed by the near diffusion-

limited reaction of .NO and O2.- during activation of phagocytes. Chronic inflammation induced by

phagocytes is a major contributor to cancer and other degenerative diseases. We examined how

gamma-tocopherol (gammaT), the principal form of vitamin E in the United States diet, and alpha-

tocopherol (alphaT), the major form in supplements, protect against peroxynitrite-induced lipid

oxidation. Lipid hydroperoxide formation in liposomes (but not isolated low-density lipoprotein)

exposed to peroxynitrite or the .NO and O2.- generator SIN-1 (3-morpholinosydnonimine) was

inhibited more effectively by gammaT than alphaT. More importantly, nitration of gammaT at the

nucleophilic 5-position, which proceeded in both liposomes and human low density lipoprotein at

yields of approximately 50% and approximately 75%, respectively, was not affected by the

presence of alphaT. These results suggest that despite

alphaT's action as an antioxidant gammaT is required to effectively remove the peroxynitrite-

derived nitrating species. We postulate that gammaT acts in vivo as a trap for membrane-soluble

electrophilic nitrogen oxides and other electrophilic mutagens, forming stable carbon-centered

adducts through the nucleophilic 5-position, which is blocked in alphaT. Because large doses of

dietary alphaT displace gammaT in plasma and other tissues, the current wisdom of vitamin E

supplementation with primarily alphaT should be reconsidered.

15. Proc Natl Acad Sci U S A 1993 Mar 1;90(5):1771-5

Gamma-tocopherol detoxification of nitrogen dioxide: superiority to alpha-tocopherol.

Cooney RV, Franke AA, Harwood PJ, Hatch-Pigott V, Custer LJ, Mordan LJ.

Cancer Research Center of Hawaii, University of Hawaii, Honolulu 96813.

In the vitamin E group, alpha-tocopherol is generally considered to be the most potent antioxidant

with the highest vitamin bioactivity, yet gamma-tocopherol is produced in greater amounts by many

plants and is the principal tocopherol in the United States diet. This report describes a fundamental

difference in the chemical reactivities of alpha-tocopherol and gamma-tocopherol with nitrogen

dioxide (NO2), which leads to the formation of a nitrosating agent from alpha-tocopherol, but not

from gamma-tocopherol. Nitric oxide (NO) is a major product of the reaction of gamma-tocopherol

with NO2, while alpha-tocopherol reacts with NO2 to form an intermediate tocopheroxide

analogue. The biological significance of gamma-tocopherol is suggested by limited epidemiological

data as well as the observation that it is a more potent inhibitor than alpha-tocopherol of neoplastic

transformation during the postinitiation phase in

3-methylcholanthrene-treated C3H/10T1/2 murine fibroblasts. This latter property suggests the

superiority of gamma-tocopherol in a mammalian biological assay and a role for endogenous NO

production in promotion of neoplastic transformation.

16. J Gerontol A Biol Sci Med Sci 2000 Jun;55(6):B280-5

Effects of tocotrienols on life span and protein carbonylation in Caenorhabditis elegans.

Adachi H, Ishii N.

Life Science Research Center, Lion Corporation, Kanagawa, Japan.

hadachi@lion.co.jp

To assess the efficiency of tocotrienols against oxidative damage, we have demonstrated in a

model-system nematode, Caenorhabditis elegans, that tocotrienol administration reduced the

accumulation of protein carbonyl (a good indicator of oxidative damage during aging) and

consequently extended the mean life span (LS), but not the maximum LS. Conversely, alpha-

tocopherol acetate did not affect these parameters. As a way to evaluate the protective ability of

tocotrienols against oxidative stress, the life spans of animals administrated tocotrienols before or

after exposure to ultraviolet B-induced oxidative stress were measured. Ultraviolet B irradiation

shortened the mean LS of animals, whereas preadministration of tocotrienols recovered the mean

LS to that of unirradiated animals. Interestingly, postadministration also extended the mean LS

more than that of unirradiated animals, and administration through the LS conferred greater

protection. Thus, the administration of tocotrienols to animals results in a reduction of oxidative

stress risks. These data indicated that tocotrienols merit further investigation as possible agents for

antiaging and oxidative stress prevention. In addition, they suggest that C. elegans will continue to

provide provocative clues into the mechanisms of aging.

17. Lipids 1995 Dec;30(12):1179-83

Antioxidant effects of tocotrienols in patients with hyperlipidemia and carotid stenosis.

Tomeo AC, Geller M, Watkins TR, Gapor A, Bierenbaum ML.

Kenneth L. Jordan Research Group, Montclair, New Jersey 07042, USA.

Antioxidants may have a role in the prevention of atherosclerosis. In thepresent trial, we

investigated the antioxidant properties of Palm Vitee, a gamma-tocotrienol-, and alpha-tocopherol

enriched fraction of palm oil, in patients with carotid atherosclerosis. Serum lipids, fatty acid

peroxides, platelet aggregation and carotid artery stenosis were measured over an 18-month period

in fifty patients with cerebrovascular disease. Change in stenosis was measured with duplex

ultrasonography. Ultrasound scans were done at six months, twelve months, and yearly thereafter.

Bilateral duplex ultrasonography revealed apparent carotid atherosclerotic regression in seven and

progression in two of the 25 tocotrienol patients, while none of the control group exhibited

regression and ten of 25 showed progression (P < 0.002). Serum thiobarbituric acid reactive

substances, an ex vivo indicator of maximal platelet peroxidation, decreased in the treatment group

from 1.08 0.70 to 0.80 0.55 microM/L (P < 0.05) after 12 mon, and in the placebo group, they

increased nonsignificantly from 0.99 0.80 to 1.26 0.54 microM/L. Both tocotrienol and placebo

groups displayed significantly attenuated collagen-induced platelet aggregation responses (P < 0.05)

as compared with entry values. Serum total cholesterol, low-density lipoprotein cholesterol, and

triglyceride values remained unchanged in both groups, as did the plasma high density lipoprotein

cholesterol values. These findings suggest that antioxidants, such as tocotrienols, may influence the

course of carotid atherosclerosis.

18. J Biol Chem 1993 May 25;268(15):11230-8

Tocotrienols regulate cholesterol production in mammalian cells by post-transcriptional suppression

of 3-hydroxy-3-methylglutaryl-coenzyme A reductase.

Parker RA, Pearce BC, Clark RW, Gordon DA, Wright JJ.

Department of Metabolic Diseases, Bristol-Myers Squibb Pharmaceutical Research Institute,

Princeton, New Jersey 08543.

Tocotrienols are natural farnesylated analogues of tocopherols which decrease hepatic cholesterol

production and reduce plasma cholesterol levels in animals. For several cultured cell types,

incubation with gamma-tocotrienol inhibited the rate of [14C]acetate but not [3H] mevalonate

incorporation into cholesterol in a concentration- and time-dependent manner, with 50% inhibition

at approximately 2 microM and maximum approximately 80% inhibition observed within 6 h in

HepG2 cells. 3-Hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase total activity and

protein levels assayed by Western blot were reduced concomitantly with the decrease in cholesterol

synthesis. In HepG2 cells, gamma-tocotrienol suppressed reductase despite strong blockade by

inhibitors at several steps in the pathway, suggesting that isoprenoid flux is not required for the

regulatory effect. HMG-CoA reductase protein synthesis rate was moderately diminished (57% of

control), while the degradation rate was increased 2.4-fold versus control (t1/2 declined from 3.73

to 1.59 h) as judged by [35S] methionine pulse-chase/immunoprecipitation analysis of HepG2 cells

treated with 10 microM gamma-tocotrienol. Under these conditions, the decrease in reductase

protein levels greatly exceeded the minor decrease in mRNA (23 versus 76% of control,

respectively), and the low density lipoprotein receptor protein was augmented. In contrast, 25-

hydroxycholesterol strongly cosuppressed HMG-CoA reductase protein and mRNA levels and the

low density lipoprotein receptor protein. Thus, tocotrienols influence the mevalonate pathway in

mammalian cells by post-transcriptional suppression of HMG-CoA reductase, and appear to

specifically modulate the intracellular mechanism for controlled degradation of the reductase

protein, an activity that mirrors the actions of the putative non-sterol isoprenoid regulators derived

from mevalonate.

19. Am J Clin Nutr 1991 Apr;53(4 Suppl):1021S-1026S

Lowering of serum cholesterol in hypercholesterolemic humans by tocotrienols (palmvitee).

Qureshi AA, Qureshi N, Wright JJ, Shen Z, Kramer G, Gapor A, Chong YH, DeWitt G, Ong A,

Peterson DM, et al.

Advanced Medical Research, Madison, WI 53719.

A double-blind, crossover, 8-wk study was conducted to compare effects of the tocotrienol-enriched

fraction of palm oil (200 mg palmvitee capsules/day) with those of 300 mg corn oil/d on serum

lipids of hypercholesterolemic human subjects (serum cholesterol 6.21-8.02 mmol/L).

Concentrations of serum total cholesterol (-15%), LDL cholesterol (-8%), Apo B (-10%),

thromboxane (-25%), platelet factor 4 (-16%), and glucose (-12%) decreased significantly only in

the 15 subjects given palmvitee during the initial 4 wk. The crossover confirmed these actions of

palmvitee. There was a carry over effect of palmvitee. Serum cholesterol concentrations of seven

hypercholesterolemic subjects (greater than 7.84 mmol/L) decreased 31% during a 4-wk period in

which they were given 200 mg gamma-tocotrienol/d. This indicates that gamma-tocotrienol may be

the most potent cholesterol inhibitor in palmvitee capsules. The results of this pilot study are very

encouraging.

20. Novel tocotrienols of rice bran modulate cardiovascular disease risk parameters of

hypercholesterolomic humans

Qureshi A.A.; Bradlow B.A.; Salser W.A.; Brace L.D. Dr. A.A. Qureshi, Advance Medical

Research, 8251 Raymond Road, Madison, WI 53719 United States

Journal of Nutritional Biochemistry ( J. NUTR. BIOCHEM. ) ( United States ) 1997 , 8/5 (290-298)

Tocotrienols inhibit cholesterol synthesis by post-transcriptionally suppressing beta-hydroxy-beta-

methylglutaryl-coenzyme A reductase activity. A double blind, 12-week study was performed to

investigate the effect of a novel tocotrienol-rich fraction (TRFinf 2$D5: obtained by molecular

distillation from specially processed rice bran oil) on cardiovascular disease risk factors of

hypercholesterolomic human subjects (serum total cholesterol >5.69 mmol/L). After acclimation to

an alcohol-free regimen (baseline) participants were assigned to the National Cholesterol Education

Program (NCEP) Step-1 diet (saturated fat <19%, total fat <30% of total calories and cholesterol

<7.76 mmol/L). The participants were evaluated after 4 weeks of exposure to the NCEP Step-1 diet:

one group of 21 participants was continued on the NCEP Step-1 diet for 4 weeks receiving an

additional 1.2 gm corn oil (placebo group) and a second group of 20 received 200 mg TRFinf 2$D5

dissolved in 1.0 gm corn oil (TRFinf 2$D5 group). Serum total cholesterol and LDL-cholesterol

levels of all the participants, stable during the baseline phase of the study, decreased 5% and 8%,

respectively, during the 4-week NCEP Step-1 diet. Placebo continuing on the NCEP Step-1 diet for

an additional 4 weeks experienced additional but modest decreases in serum total cholesterol (2%)

and LDL-cholesterol (3%), yielding significant (P<0.05) decreases when compared with the

baseline values. These responses confirm the cholesterol- lowering action of a low fat, low

cholesterol diet. Participants receiving TRFinf 2$D5 had 12% and 16% reductions (P<0.05) in total

cholesterol and LDL- cholesterol levels during the 4-week experimental phase; during the two

phases (NCEP Step-1 diet plus treatment) the serum total cholesterol and LDL- cholesterol levels of

these of these participants were decreased (P<0.05) by 17% and 24%, respectively, TRFinf 2$D5-

mediated decreases in Apo B(a), platelet factor 4 and thromboxane Binf 2 (15%, 17%, 14%, and

31%, respectively) were significant (P<0.05). There was no change in the levels of HDL-cholesterol

and apolipoprotein A-I by this treatment. The treatment also resulted in remarkable increases in the

levels of LDL-bound antioxidants, especially tocotrienols, which have substantially greater

antioxidant activity than vitamin E.

21. N Engl J Med 1990 Nov 8;323(19):1289-98

Regression of coronary artery disease as a result of intensive lipid-lowering therapy in men with

high levels of apolipoprotein B.

Brown G, Albers JJ, Fisher LD, Schaefer SM, Lin JT, Kaplan C, Zhao XQ, Bisson BD, Fitzpatrick

VF, Dodge HT.

Department of Medicine, University of Washington School of Medicine, Seattle.

BACKGROUND AND METHODS. The effect of intensive lipid-lowering therapy on coronary

atherosclerosis among men at high risk for cardiovascular events was assessed by quantitative

arteriography. Of 146 men no more than 62 years of age who had apolipoprotein B levels greater

than or equal to 125 mg per deciliter, documented coronary artery disease, and a family history of

vascular disease, 120 completed the 2 1/2-year double-blind study, which included arteriography at

base line and after treatment. Patients were given dietary counseling and were randomly assigned to

one of three treatments: lovastatin (20 mg twice a day) and colestipol (10 g three times a day);

niacin (1 g four times a day) and colestipol (10 g three times a day); or conventional therapy with

placebo (or colestipol if the low-density lipoprotein [LDL] cholesterol level was elevated).

RESULTS. The levels of LDL and high-density lipoprotein (HDL) cholesterol changed only

slightly in the conventional-therapy group (mean changes, -7 and +5 percent, respectively), but

more substantially among patients treated with lovastatin and colestipol (-46 and +15 percent) or

niacin and colestipol (-32 and +43 percent). In the conventional-therapy group, 46 percent of the

patients had definite lesion progression (and no regression) in at least one of nine proximal coronary

segments; regression was the only change in 11 percent. By comparison, progression (as the only

change) was less frequent among patients who received lovastatin and colestipol (21 percent) and

those who received niacin and colestipol (25 percent), and regression was more frequent (lovastatin

and colestipol, 32 percent; niacin and colestipol, 39 percent; P less than 0.005). Multivariate

analysis indicated that a reduction in the level of apolipoprotein B (or LDL cholesterol) and in

systolic blood pressure, and an increase in HDL cholesterol correlated independently with

regression of coronary lesions. Clinical events (death, myocardial infarction, or revascularization

for worsening symptoms) occurred in 10 of 52 patients assigned to conventional therapy, as

compared with 3 of 46 assigned to receive lovastatin and colestipol and 2 of 48 assigned to receive

niacin and colestipol (relative risk of an event during intensive treatment, 0.27; 95 percent

confidence interval, 0.10 to 0.77). CONCLUSIONS. In men with coronary artery disease who were

at high risk for cardiovascular events, intensive lipid-lowering therapy reduced the frequency of

progression of coronary lesions, increased the frequency of regression, and reduced the incidence of

cardiovascular events.

22. N Engl J Med 1983 Aug 18;309(7):385-9

Apolipoprotein A-I as a marker of angiographically assessed coronary-artery disease.

Maciejko JJ, Holmes DR, Kottke BA, Zinsmeister AR, Dinh DM, Mao SJ.

This study was designed to determine whether the plasma level of apolipoprotein A-I is a better

discriminator of angiographically documented coronary-artery disease than the level of high-

density-lipoprotein (HDL) cholesterol in male subjects. The level of plasma apolipoprotein A-I in

83 patients with coronary-artery disease was 96.7 4.2 mg per deciliter (mean S.E.M.), which was

significantly lower (P less than 0.0001) than the level in 25 patients without coronary-artery disease

(146.9 2.1 mg per deciliter). The levels of HDL cholesterol were also lower (P less than 0.0001) in

patients with coronary-artery disease (31.9 1.5 mg per deciliter) than in those without it (45.9 2.3

mg per deciliter). A stepwise discriminant analysis, however, indicated the superiority of

apolipoprotein A-I over HDL cholesterol in detecting coronary-artery disease. Furthermore, a linear

discriminant analysis suggested that although HDL cholesterol by itself was a discriminator of

coronary-artery disease, it did not provide a substantial increase in discriminatory value over that

provided by apolipoprotein A-I; in contrast,

apolipoprotein A-I levels added discriminatory value to the information obtained by measuring

HDL cholesterol alone. We conclude that apolipoprotein A-I by itself is more useful than HDL

cholesterol for identifying patients with coronary-artery disease.

23. Br Heart J 1988 Nov;60(5):397-403

High prevalence of hypertriglyceridaemia and apolipoprotein abnormalities in coronary artery

disease.

Barbir M, Wile D, Trayner I, Aber VR, Thompson GR.

Department of Medicine, Royal Postgraduate Medical School, Hammersmith Hospital, London.

Serum lipids and apolipoproteins A-I and B were measured in 174 men aged less than 60 with

angiographically confirmed coronary artery disease and in 572 healthy control men. Two thirds of

the patients had raised age-corrected values of fasting serum cholesterol and/or triglyceride and/or a

low high density lipoprotein (HDL) cholesterol compared with the controls. Eighteen (30%) of the

61 normolipidaemic patients had a concentration of serum apolipoprotein A-I below the 5th

percentile of 233 controls. In normolipidaemic patients on beta blockers the relative prevalence of

serum low density lipoprotein

(LDL)-apolipoprotein B values above the 95th percentile of 339 controls was significantly

increased. Discriminant function analysis showed that a raised concentration of serum triglyceride

was the best discriminant between patients and controls, with raised LDL-apolipoprotein B and

reduced apolipoprotein A-I coming second only to triglyceride in analyses where each was

separately compared with all the lipid variables. These associations were highly significant and

were independent of other influences, including beta blockade. These findings re-emphasise the

importance of hypertriglyceridaemia as a risk factor and confirm that apolipoprotein abnormalities

occur frequently in coronary disease, even in normolipidaemic patients.

24. Proc Natl Acad Sci U S A 2000 Oct 10;97(21):11494-9

Gamma-tocopherol and its major metabolite, in contrast to alpha-tocopherol, inhibit

cyclooxygenase activity in macrophages and epithelial cells.

Jiang Q, Elson-Schwab I, Courtemanche C, Ames BN.

Division of Biochemistry and Molecular Biology, University of California,

Berkeley, CA 94720, USA.

Cyclooxygenase-2 (COX-2)-catalyzed synthesis of prostaglandin E(2) (PGE(2)) plays a key role in

inflammation and its associated diseases, such as cancer and vascular heart disease. Here we report

that gamma-tocopherol (gammaT) reduced PGE(2) synthesis in both lipopolysaccharide (LPS)-

stimulated RAW264.7 macrophages and IL-1beta-treated A549 human epithelial cells with an

apparent IC(50) of 7.5 and 4 microM, respectively. The major metabolite of dietary gammaT, 2,7,8-

trimethyl-2-(beta-carboxyethyl)-6-hydroxychroman (gamma-CEHC), also exhibited an inhibitory

effect, with an IC(50) of approximately 30 microM in these cells. In contrast, alpha-tocopherol at 50

microM slightly reduced (25%) PGE(2) formation in macrophages, but had no effect in epithelial

cells. The inhibitory effects of gammaT and gamma-CEHC stemmed from their inhibition of COX-

2 activity, rather than affecting protein expression or substrate availability, and appeared to be

independent of antioxidant activity. gamma-CEHC also inhibited PGE(2) synthesis when exposed

for 1 h to COX-2-preinduced cells followed by the addition of arachidonic acid (AA), whereas

under similar conditions, gammaT required an 8- to 24-h incubation period to cause the inhibition.

The inhibitory potency of gammaT and gamma-CEHC was diminished by an increase in AA

concentration, suggesting that they might compete with AA at the active site of COX-2. We also

observed a moderate reduction of nitrite accumulation and suppression of inducible nitric oxide

synthase expression by gammaT in lipopolysaccharide-treated macrophages. These findings

indicate that gammaT and its major metabolite possess anti-inflammatory activity and that gammaT

at physiological concentrations may be important in human disease prevention.

25. J Am Coll Cardiol 1999 Oct;34(4):1208-15

Differential effects of alpha- and gamma-tocopherol on low-density lipoprotein oxidation,

superoxide activity, platelet aggregation and arterial thrombogenesis.

Saldeen T, Li D, Mehta JL.

Department of Forensic Medicine, University of Uppsala, Sweden.

OBJECTIVES: This study was designed to examine the differential effects of alpha- and gamma-

tocopherol on parameters of oxidation-antioxidation and thrombogenesis. BACKGROUND:

Experimental studies have shown that antioxidants, such as vitamin E (alpha-tocopherol), improve

atherosclerotic plaque stability and vasomotor function, and decrease platelet aggregation and

tendency to thrombus formation. METHODS: Sprague Dawley rats were fed chow mixed with

alpha- or gamma-tocopherol (100 mg/kg/day) for 10 days. A filter soaked in 29% FeCl3 was

applied around the abdominal aorta to study the patterns of arterial thrombosis. The aortic blood

flow was observed and continuously recorded using an ultrasonic Doppler flow probe. ADP-

induced platelet aggregation, low-density lipoprotein oxidation induced by phorbol 12-myristate 13-

acetate (PMA)-stimulated leukocytes, superoxide anion generation and superoxide dismutase

(SOD) activity were also measured. RESULTS: Both alpha- and gamma-tocopherol decreased

platelet aggregation and delayed time to occlusive thrombus (all p < 0.05 vs. control). Both alpha-

and gamma-tocopherol decreased arterial superoxide anion generation, lipid peroxidation and LDL

oxidation (all p < 0.05 vs. control), and increased endogenous SOD activity (p < 0.05). The effects

of gamma-tocopherol were more potent than those of alpha-tocopherol (p < 0.05).

CONCLUSIONS: This study indicates that both alpha- and gamma-tocopherol decrease platelet

aggregation and delay intraarterial thrombus formation, perhaps by an increase in endogenous

antioxidant activity. Alpha-tocopherol is significantly more potent than alpha-tocopherol in these

effects.

26. J Nutr Biochem 2001 Jun;12(6):318-329

Synergistic effect of tocotrienol-rich fraction (TRF(25)) of rice bran and lovastatin on lipid

parameters in hypercholesterolemic humans.

Qureshi AA, Sami SA, Salser WA, Khan FA.

Advanced Medical Research, 8251 Raymond Road, 53719, Madison, WI, USA

Tocotrienols exert hypocholesterolemic action in humans and animals. Lovastatin is widely used for

that purpose. Both agents work by suppressing the activity of beta-hydroxy-beta methylglutaryl

coenzyme A reductase through different mechanisms, post-transcriptional vs competitive inhibition.

A human study with 28 hypercholesterolemic subjects was carried out in 5 phases of 35 days each,

to check the efficacy of tocotrienol-rich fraction (TRF(25)) of rice bran alone and in combination

with lovastatin. After placing subjects on the American Heart Association (AHA) Step-1 diet (phase

II), the subjects were divided into two groups, A and B. The AHA Step-1 diet was continued in

combination with other treatments during phases III to V. Group A subjects were given 10 mg

lovastatin, 10 mg lovastatin plus 50 mg TRF(25), 10 mg lovastatin plus 50 mg alpha-tocopherol per

day, in the third, fourth, and fifth phases, respectively. Group B subjects were treated exactly to the

same protocol except that in the third phase, they were given 50 mg TRF(25) instead of

lovastatin.The TRF(25) or lovastatin plus AHA Step-1 diet effectively lower serum total cholesterol

(14%, 13%) and LDL-cholesterol (18%, 15% P < 0.001), respectively, in hypercholesterolemic

subjects. The combination of TRF(25) and lovastatin plus AHA Step-1 diet significantly reduces of

these lipid parameters of 20% and 25% (P < 0.001) in these subjects. Substitution of TRF(25) with

alpha-tocopherol produces insignificant changes when given with lovastatin. Especially significant

is the increase in the HDL/LDL ratio to 46% in group (A) and 53% (P < 0.002) in group (B). These

results are consistent with the synergistic effect of these two agents. None of the subjects reported

any side-effects throughout the study of 25-weeks. In the present study, the increased effectiveness

of low doses of tocotrienols (TRF(25)) as hypocholesterolemic agents might be due to a minimum

conversion to alpha-tocopherol. The report also describes in vivo the conversion of gamma-[4-3H]-,

and [14C]-desmethyl (d-P(21)-T3) tocotrienols to alpha-tocopherol.

27. Lipids 1993 Dec;28(12):1113-8

gamma-Tocotrienol as a hypocholesterolemic and antioxidant agent in rats fed atherogenic diets.

Watkins T, Lenz P, Gapor A, Struck M, Tomeo A, Bierenbaum M.

Kenneth L. Jordan Heart Fund, Montclair, New Jersey 07042.

This study was designed to determine whether incorporation of gamma-tocotrienol or alpha-

tocopherol in an atherogenic diet would reduce the concentration of plasma cholesterol,

triglycerides and fatty acid peroxides, and attenuate platelet aggregability in rats. For six weeks,

male Wistar rats (n = 90) were fed AIN76A semisynthetic test diets containing cholesterol (2% by

weight), providing fat as partially hydrogenated soybean oil (20% by weight), menhaden oil (20%)

or corn oil (2%). Feeding the ration with menhaden oil resulted in the highest concentrations of

plasma cholesterol, low and very low density lipoprotein cholesterol, triglycerides, thiobarbituric

acid reactive substances and fatty acid hydroperoxides. Consumption of the ration containing

gamma-tocotrienol (50 mg/kg) and alpha-tocopherol (500 mg/kg) for six weeks led to decreased

plasma lipid concentrations. Plasma cholesterol, low and very low density lipoprotein cholesterol,

and triglycerides each decreased significantly (P < 0.001). Plasma thiobarbituric acid reactive

substances decreased significantly (P < 0.01), as did the fatty acid hydroperoxides (P < 0.05), when

the diet contained both chromanols. Supplementation with gamma-tocotrienol resulted in similar,

though quantitatively smaller, decrements in these plasma values. Plasma alpha-tocopherol

concentrations were lowest in rats fed menhaden oil without either chromanol. Though plasma

alpha-tocopherol did not rise with gamma-tocotrienol supplementation at 50 mg/kg, gamma-

tocotrienol at 100 mg/kg of ration spared plasma alpha-tocopherol, which rose from 0.60 0.2 to 1.34

0.4 mg/dL (P < 0.05). The highest concentration of alpha-tocopherol was measured in plasma of

animals fed a ration supplemented with alpha-tocopherol at 500 mg/kg.(ABSTRACT

TRUNCATED AT 250 WORDS)

28. J Nutr 2001 Jan;131(1):161S-163S

Vitamin E: mechanisms of action as tumor cell growth inhibitors.

Kline K, Yu W, Sanders BG.

Division of Nutrition and. School of Biological Sciences, The University of Texas at Austin,

Austin, TX 78712, USA. k.kline@mail.utexas.edu

http://www.nutrition.org/cgi/content/full/ 131/1/161S?view=full&pmid=11208955

29. J Nutr 1994 May;124(5):607-14

The chemoprevention of cancer by mevalonate-derived constituents of fruits and vegetables.

Elson CE, Yu SG.

Department of Nutritional Sciences, University of Wisconsin, Madison 53706-1571.

Anutritive isoprenoid constituents of fruits, vegetables, cereal grains and essential oils exhibit a

spectrum of anticarcinogenic activities. The induction of hepatic Phase II detoxifying activities by

dietary isoprenoids appears to underlie their blocking action. The second anticarcinogenic action of

the dietary isoprenoids, suppression of the growth of chemically initiated and transplanted tumors

is, we suggest, secondary to the inhibition of mevalonate pathway activities. Mevinolin, a

competitive inhibitor of 3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase activity,

depletes cells of the intermediate products of the pathway that are required for the posttranslational

modification of proteins, a process giving the proteins lipophilic anchors that bind to membranes.

As a consequence, nuclear lamins and ras oncoproteins remain in nascent states, and cells do not

proliferate. gamma-Tocotrienol, perillyl alcohol, geraniol and d-limonene suppress hepatic HMG-

CoA reductase activity, a rate-limiting step in cholesterol synthesis, and modestly lower serum-

cholesterol levels of animals. These isoprenoids also suppress tumor growth. The HMG-CoA

reductase of neoplastic tissues differs from that of sterologenic tissues in being markedly resistant

to sterol feedback inhibition. Our review suggests that the mevalonate pathway of tumor tissues is

uniquely sensitive to the inhibitory actions of the dietary isoprenoids.

30. J Nutr 1999 Apr;129(4):804-13

Apoptosis and cell-cycle arrest in human and murine tumor cells are initiated by isoprenoids.

Mo H, Elson CE.

Department of Nutritional Sciences, University of Wisconsin-Madison, Madison, WI 53706, USA.

Diverse classes of phytochemicals initiate biological responses that effectively lower cancer risk.

One class of phytochemicals, broadly defined as pure and mixed isoprenoids, encompasses an

estimated 22,000 individual components. A representative mixed isoprenoid, gamma-tocotrienol,

suppresses the growth of murine B16(F10) melanoma cells, and with greater potency, the growth of

human breast adenocarcinoma (MCF-7) and human leukemic (HL-60) cells. beta-Ionone, a pure

isoprenoid, suppresses the growth of B16 cells and with greater potency, the growth of MCF-7, HL-

60 and human colon adenocarcinoma (Caco-2) cells. Results obtained with diverse cell lines

differing in ras and p53 status showed that the isoprenoid-mediated suppression of growth is

independent of mutated ras and p53 functions. beta-Ionone suppressed the growth of human colon

fibroblasts (CCD-18Co) but only when present at three-fold the concentration required to suppress

the growth of Caco-2 cells. The isoprenoids initiated apoptosis and, concomitantly arrested cells in

the G1 phase of the cell cycle. Both suppress 3-hydroxy-3-methylglutaryl CoA reductase activity.

beta-Ionone and lovastatin interfered with the posttranslational processing of lamin B, an activity

essential to assembly of daughter nuclei. This interference, we postulate, renders neosynthesized

DNA available to the endonuclease activities leading to apoptotic cell death. Lovastatin-imposed

mevalonate starvation suppressed the glycosylation and translocation of growth factor receptors to

the cell surface. As a consequence, cells were arrested in the G1 phase of the cell cycle. This

rationale may apply to the isoprenoid-mediated G1-phase arrest of tumor cells. The additive and

potentially synergistic actions of these isoprenoids in the suppression of tumor cell proliferation and

initiation of apoptosis coupled with the mass action of the diverse isoprenoid constituents of plant

products may explain, in part, the impact of fruit, vegetable and grain consumption on cancer risk.

31. J Nutr 1997 May;127(5):668-74

Isoprenoids suppress the growth of murine B16 melanomas in vitro and in vivo.

He L, Mo H, Hadisusilo S, Qureshi AA, Elson CE.

Department of Nutritional Sciences, University of Wisconsin, Madison 53706, USA.

Sundry mevalonate-derived constituents (isoprenoids) of fruits, vegetables and cereal grains

suppress the growth of tumors. This study estimated the concentrations of structurally diverse

isoprenoids required to inhibit the increase in a population of murine B16(F10) melanoma cells

during a 48-h incubation by 50% (IC50 value). The IC50 values for d-limonene and perillyl alcohol,

the monoterpenes in Phase I trials, were 450 and 250 micromol/L, respectively; related cyclic

monoterpenes (perillaldehyde, carvacrol and thymol), an acyclic monoterpene (geraniol) and the

end ring analog of beta-carotene (beta-ionone) had IC50 values in the range of 120-150

micromol/L. The IC50 value estimated for farnesol, the side-chain analog of the tocotrienols

(50 micromol/L) fell midway between that of alpha-tocotrienol (110 micromol/L) and those

estimated for gamma- (20 micromol/L) and delta- (10 micromol/L) tocotrienol. A novel tocotrienol

lacking methyl groups on the tocol ring proved to be extremely potent (IC50, 0.9 micromol/L). In

the first of two diet studies, experimental diets were fed to weanling C57BL female mice for 10 d

prior to and 28 d following the implantation of the aggressively growing and highly metastatic

B16(F10) melanoma. The isomolar (116 micromol/kg diet) and the Vitamin E-equivalent (928

micromol/kg diet) substitution of d-gamma-tocotrienol for dl-alpha-tocopherol in the AIN-76A diet

produced 36 and 50% retardations, respectively, in tumor growth (P < 0.05). In the second study,

melanomas were established before mice were fed experimental diets formulated with 2 mmol/kg d-

gamma-tocotrienol, beta-ionone individually and in combination. Each treatment increased (P <

0.03) the duration of host survival. Our finding that the effects of individual isoprenoids were

additive suggests the possibility that one component of the anticarcinogenic action of plant-based

diets is the tumor growth-suppressive action of the diverse isoprenoid constituents of fruits,

vegetables and cereal grains.

32. Nutr Cancer 1992;18(1):1-29

Fruit, vegetables, and cancer prevention: a review of the epidemiological evidence.

Block G, Patterson B, Subar A.

Dept. of Social and Administrative Health Sciences, School of Public Health,

University of California, Berkeley 94720.

Approximately 200 studies that examined the relationship between fruit and vegetable intake and

cancers of the lung, colon, breast, cervix, esophagus, oral cavity, stomach, bladder, pancreas, and

ovary are reviewed. A statistically significant protective effect of fruit and vegetable consumption

was found in 128 of 156 dietary studies in which results were expressed in terms of relative risk.

For most cancer sites, persons with low fruit and vegetable intake (at least the lower one-fourth of

the population) experience about twice the risk of cancer compared with those with high intake,

even after control for potentially confounding factors. For lung cancer, significant protection was

found in 24 of 25 studies after control for smoking in most instances. Fruits, in particular, were

significantly protective in cancers of the esophagus, oral cavity, and larynx, for which 28 of 29

studies were significant. Strong evidence of a protective effect of fruit and vegetable consumption

was seen in cancers of the pancreas and stomach (26 of 30 studies), as well as in colorectal and

bladder cancers (23 of 38 studies). For cancers of the cervix, ovary, and endometrium, a significant

protective effect was shown in 11 of 13 studies, and for breast cancer a protective effect was found

to be strong and consistent in a meta analysis. It would appear that major public health benefits

could be achieved by substantially increasing consumption of these foods.

33. Proc Soc Exp Biol Med 1999 Sep;221(4):294-311

Isoprenoid-mediated inhibition of mevalonate synthesis: potential application to cancer.

Elson CE, Peffley DM, Hentosh P, Mo H.

Department of Nutritional Sciences, College of Agricultural and Life Sciences, University of

Wisconsin-Madison, Madison, Wisconsin 53706, USA. elson@nutrisci.wisc.edu

Pure and mixed isoprenoid end products of plant mevalonate metabolism trigger actions that

suppress 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase activity. These actions

modulate HMG CoA reductase mRNA translation and the proteolytic degradation of HMG CoA

reductase. Such post-transcriptional events, we propose, are activated directly by acyclic

isoprenoids and indirectly by cyclic isoprenoids. Isoprenoids, acting secondarily to the dominant

transcriptional effector of sterologenesis, modestly lower cholesterol levels, if and only if,

sterologenesis is not repressed by a saturating imput of dietary cholesterol. An anomaly associated

with tumor growth-a sterol feedback-resistant HMG CoA reductase activity-ensures a pool of

sterologenic pathway intermediates. Such intermediates provide lipophilic anchors essential for

membrane attachment and biological activity of growth hormone receptors, nuclear lamins A and B,

and oncogenic ras. Tumor HMG CoA reductase retains high sensitivity to the

isoprenoid-mediated secondary regulation. Repression of mevalonate synthesis by plant-derived

isoprenoids reduces ras and lamin B processing, arrests cells in G1, and initiates cellular apoptosis.

This unique tumor cell-specific sensitivity allows isoprenoids to be used for tumor therapy, an

application emulating that of the statins, but one free of adverse effects. When evaluated at levels

provided by a typical diet, isoprenoids individually have no impact on cholesterol synthesis and

tumor growth. Nonetheless, isoprenoid-mediated activities are additive, and, sometimes synergistic.

Therefore, the combined actions of the estimated 23,000 isoprenoid constituents of plant materials,

acting in concert with other chemopreventive phytochemicals, may explain the lowered cancer risk

associated with a diet rich in plant products. In contrast, that lowering of cancer risk does not

correspond to supplemental intake of other dietary factors associated with fruits, vegetables, and

cereal grains, namely fiber, beta-carotene, vitamin C, and vitamin E, and only weakly to

supplemental folate.

34. J Nutr 1995 Jun;125(6 Suppl):1666S-1672S

Suppression of mevalonate pathway activities by dietary isoprenoids: protective roles in cancer and

cardiovascular disease.

Elson CE.

Department of Nutritional Sciences, University of Wisconsin-Madison 53706, USA.

Diet-cancer and diet-cardiovascular disease interrelationships may be explained by the mevalonate-

suppressive action of isoprenoid end products of plant secondary metabolism. Assorted

monoterpenes, sesquiterpenes, carotenoids and tocotrienols posttranscriptionally down regulate 3-

hydroxy-3-methylglutaryl coenzyme A reductase activity, a key activity in the sterologenic

pathway. The modest decrease in cholesterol synthesis is associated with a concomitant lowering of

low-density lipoprotein cholesterol. The reductase activity in tumor tissues differs from that of liver

in being resistant to sterol feedback regulation. Tumor reductase activity retains sensitivity to

posttranscriptional regulation. As a consequence, the isoprenoid-mediated suppression of

mevalonate synthesis depletes tumor tissues of two intermediate products, farnesyl pyrophosphate

and geranylgeranyl pyrophosphate, which are incorporated

posttranslationally into growth control-associated proteins. At 10-fold higher concentrations,

monoterpenes inhibit the protein isoprenyl transferases that catalyze this incorporation. At levels of

intake likely provided by a diet based on Food Pyramid guidelines, assorted isoprenoids decrease

cardiovascular disease risk and suppress the growth of initiated cells. At pharmacological levels of

intake, isoprenoids block the initiation phase of chemical carcinogenesis. Isoprenoids targeted to the

inhibition of the isoprenylation of oncogenic forms of ras proteins may offer a novel approach to

chemotherapy. Adjunctive isoprenoids might decrease the level of competitive inhibitors of 3-

hydroxy-3-methylglutaryl coenzyme A reductase required to manage hypercholesterolemia.

35. Nutrition 1993 May-Jun;9(3):229-32

Long-term administration of tocotrienols and tumor-marker enzyme activities during

hepatocarcinogenesis in rats.

Rahmat A, Ngah WZ, Shamaan NA, Gapor A, Abdul Kadir K.

Department of Biochemistry, Faculty of Medicine, Universiti Kebangsaan Malaysia,

Jalan Raja Muda Abdul Aziz, Kuala Lumpur.

The effects of long-term administration of tocotrienol on hepatocarcinogenesis in rats induced by

diethylnitrosamine (DEN) and 2-acetylaminofluorene (AAF) were investigated by determining the

activities of gamma-glutamyl transpeptidase (GGT), alkaline phosphatase (ALP), glutathione S-

transferases (GSTs), and glutathione (GSH) levels in blood and liver. Twenty-eight male 7- to 8-

wk-old Rattus norwegicus rats, weighing 120-160 g, were used in this study. The rats were divided

into four treatment groups: a control group on a basal diet, a group fed a basal diet supplemented

with tocotrienol (30 mg/kg food), a group treated with DEN/AAF, and a group treated with

DEN/AAF and fed a diet supplemented with tocotrienol (30 mg/kg food). Blood was collected

monthly, and GGT, ALP, and GSH levels were determined. The rats were killed after 9 mo, and the

livers were examined morphologically. Grayish white nodules (2/liver) were found in all the

DEN/AAF-treated rats (n = 10), but only one of the rats treated with DEN/AAF and supplemented

with tocotrienol (n = 6) had liver nodules. A significant increase in the level of blood and liver

GSH, ALP, and GGT activities was observed in the DEN/AAF-treated rats. Liver GSTs were

similarly increased with DEN/AAF treatment. Tocotrienol supplementation attenuated the impact of

the carcinogens in the rats.

36. Comp Biochem Physiol C 1993 Sep;106(1):237-40

Glutathione S-transferase and gamma-glutamyl transpeptidase activities in cultured rat hepatocytes

treated with tocotrienol and tocopherol.

Ong FB, Wan Ngah WZ, Shamaan NA, Md Top AG, Marzuki A, Khalid AK.

Jabatan Biokimia, Fakulti Perubatan, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul

Aziz, Kuala, Lumpur.

The effect of tocotrienol and tocopherol on glutathione S-transferase (GST) and gamma-glutamyl

transpeptidase (GGT) activities in cultured rat hepatocytes were investigated. 2. Tocotrienol and

tocopherol significantly decreased GGT activities at 5 days in culture but tocotrienol also

significantly decreased GGT activities at 1-2 days. 3. Tocotrienol and tocopherol treatment

significantly decreased GST activities at 3 days compared to the control but tocotrienol also

decreased GST activities at 1-3 days. 4. Tocotrienol showed a more pronounced effect at a dosage

of greater than 50 microM tocotrienol at 1-3 days in culture compared to the control.

37. J Nutr 1997 May;127(5):668-74

Isoprenoids suppress the growth of murine B16 melanomas in vitro and in vivo.

He L, Mo H, Hadisusilo S, Qureshi AA, Elson CE.

Department of Nutritional Sciences, University of Wisconsin, Madison 53706, USA.

Sundry mevalonate-derived constituents (isoprenoids) of fruits, vegetables and cereal grains

suppress the growth of tumors. This study estimated the concentrations of structurally diverse

isoprenoids required to inhibit the increase in a population of murine B16(F10) melanoma cells

during a 48-h incubation by 50% (IC50 value). The IC50 values for d-limonene and perillyl

alcohol, the monoterpenes in Phase I trials, were 450 and 250 micromol/L, respectively; related

cyclic monoterpenes (perillaldehyde, carvacrol and thymol), an acyclic monoterpene (geraniol) and

the end ring analog of beta-carotene (beta-ionone) had IC50 values in the range of 120-150

micromol/L. The IC50 value estimated for farnesol, the side-chain analog of the tocotrienols

(50 micromol/L) fell midway between that of alpha-tocotrienol (110 micromol/L) and those

estimated for gamma- (20 micromol/L) and delta- (10 micromol/L) tocotrienol. A novel tocotrienol

lacking methyl groups on the tocol ring proved to be extremely potent (IC50, 0.9 micromol/L). In

the first of two diet studies, experimental diets were fed to weanling C57BL female mice for 10 d

prior to and 28 d following the implantation of the aggressively growing and highly metastatic

B16(F10) melanoma. The isomolar (116 micromol/kg diet) and the

Vitamin E-equivalent (928 micromol/kg diet) substitution of d-gamma-tocotrienol for dl-alpha-

tocopherol in the AIN-76A diet produced 36 and 50% retardations, respectively, in tumor growth (P

< 0.05). In the second study, melanomas were established before mice were fed experimental diets

formulated with 2 mmol/kg d-gamma-tocotrienol, beta-ionone individually and in combination.

Each treatment increased (P < 0.03) the duration of host survival. Our finding that the effects of

individual isoprenoids were additive suggests the possibility that one component of the

anticarcinogenic action of plant-based diets is the tumor growth-suppressive action of the diverse

isoprenoid constituents of fruits, vegetables and cereal grains.

38. Lipids 1995 Dec;30(12):1139-43

Effect of tocotrienols on the growth of a human breast cancer cell line in culture.

Nesaretnam K, Guthrie N, Chambers AF, Carroll KK.

Palm Oil Research Institute of Malaysia, Kuala Lumpur, Malaysia.

The tocotrienol-rich fraction (TRF) of palm oil consists of tocotrienols and some alpha-tocopherol

(alpha-T). Tocotrienols are a form of vitamin E having an unsaturated side-chain, rather than the

saturated side-chain of the more common tocopherols. Because palm oil has been shown not to

promote chemically-induced mammary carcinogenesis, we tested effects of TRF and alpha-T on the

proliferation, growth, and plating efficiency (PE) of the MDA-MB-435 estrogen-receptor-negative

human breast cancer cells. TRF inhibited the proliferation of these cells with a concentration

required to inhibit cell proliferation by 50% of 180 microgram/mL whereas alpha-T had no effect at

concentrations up to 1000 microgram/mL as measured by incorporation of [3H]thymidine. The

effects of TRF and alpha-T also were tested in longer-term growth experiments, using

concentrations of 180 and 500 microgram/mL. We found that TRF inhibited the growth of these

cells by 50%, whereas alpha-T did not. Their effect on the ability of these cells to form colonies

also was studied, and it was found that TRF inhibited PE, whereas alpha T had no effect. These

results suggest that the inhibition is due to the presence of tocotrienols in TRF rather than alpha T.

39. Lipids 1998 May;33(5):461-9

Tocotrienols inhibit the growth of human breast cancer cells irrespective of estrogen receptor status.

Nesaretnam K, Stephen R, Dils R, Darbre P.

Division of Cell and Molecular Biology, School of Animal and Microbial Sciences, The University

of Reading, Whiteknights, England. sarnesar@porim.gov.my

Potential antiproliferative effects of tocotrienols, the major vitamin E component in palm oil, were

investigated on the growth of both estrogen-responsive (ER+) MCF7 human breast cancer cells and

estrogen-unresponsive (ER-) MDA-MB-231 human breast cancer cells, and effects were compared

with those of alpha-tocopherol (alphaT). The tocotrienol-rich fraction (TRF) of palm oil inhibited

growth of MCF7 cells in both the presence and absence of estradiol with a nonlinear dose-response

but such that complete suppression of growth was achieved at 8 microg/mL. MDA-MB-231 cells

were also inhibited by TRF but with a linear dose-response such that 20 microg/mL TRF was

needed for complete growth suppression. Separation of the TRF into individual tocotrienols

revealed that all fractions could inhibit growth of both ER+ and

ER- cells and of ER+ cells in both the presence and absence of estradiol. However, the gamma- and

delta-fractions were the most inhibitory. Complete inhibition of MCF7 cell growth was achieved at

6 microg/mL of gamma-tocotrienol/delta-tocotrienol (gammaT3/deltaT3) in the absence of estradiol

and 10 microg/mL of deltaT3 in the presence of estradiol, whereas complete suppression of MDA-

MB-231 cell growth was not achieved even at concentrations of 10 microg/mL of deltaT3. By

contrast to these inhibitory effects of tocotrienols, alphaT had no inhibitory effect on MCF7 cell

growth in either the presence or the absence of estradiol, nor on MDA-MB-231 cell growth. These

results confirm studies using other sublines of human breast cancer cells and demonstrate that

tocotrienols can exert direct inhibitory effects on the growth of breast cancer cells. In searching for

the mechanism of inhibition, studies of the effects of TRF on estrogen-regulated pS2 gene

expression in MCF7 cells showed that tocotrienols do not act via an estrogen receptor-mediated

pathway and must therefore act differently from estrogen antagonists. Furthermore, tocotrienols did

not increase levels of growth-inhibitory insulin-like growth factor binding proteins (IGFBP) in

MCF7 cells, implying also a different mechanism from that proposed for retinoic acid inhibition of

estrogen-responsive breast cancer cell growth. Inhibition of the growth of breast cancer cells by

tocotrienols could have important clinical implications not only because tocotrienols are able to

inhibit the growth of both ER+ and ER-phenotypes but also because ER+ cells could be growth-

inhibited in the presence as well as in the absence of estradiol. Future clinical applications of TRF

could come from potential growth suppression of ER+ breast cancer cells otherwise resistant to

growth inhibition by antiestrogens and retinoic acid.

40. Int J Food Sci Nutr 2000;51 Suppl:S95-103

Tocotrienols inhibit growth of ZR-75-1 breast cancer cells.

Nesaretnam K, Dorasamy S, Darbre PD.

Palm Oil Research Institute of Malaysia, PO Box 10620, Kuala Lumpur 50720, Malaysia.

The vitamin E component of palm oil provides a rich source of tocotrienols which have been shown

previously to be growth inhibitory to two human breast cancer cell lines: responsive MCF7 cells

and unresponsive MDA-MB-231 cells. Data presented here shows that the tocotrienol-rich fraction

(TRF) of palm oil and individual fractions (alpha, gamma and delta) can also inhibit the growth of

another responsive human breast cancer cell line, ZR-75-1. At low concentrations in the absence of

oestrogen tocotrienols stimulated growth of the ZR-75-1 cells, but at higher concentrations in the

presence as well as in the absence of oestradiol, tocotrienols inhibited cell growth strongly. As for

MCF7 cells, alpha-tocopherol had no effect on growth of the ZR-75-1 cells in either the absence or

presence of oestradiol. In studying the effects of tocotrienols in combination with antioestrogens, it

was found that TRF could further inhibit growth of ZR-75-1 cells in the presence of tamoxifen (10(-

7) M and 10(-8) M). Individual tocotrienol fractions (alpha, gamma, delta) could inhibit growth of

ZR-75-1 cells in the presence of 10(-8) M oestradiol and 10(-8) M pure antioestrogen ICI 164,384.

The immature mouse uterine weight bioassay confirmed that TRF could not exert oestrogen

antagonist action in vivo. These results provide evidence of wider growth-inhibitory effects of

tocotrienols beyond MCF7 and MDA-MB-231 cells, and with an oestrogen-independent

mechanism of action, suggest a possible clinical advantage in combining administration of

tocotrienols with antioestrogen therapy.

41. J Nutr 1997 Mar;127(3):544S-548S

Inhibition of proliferation of estrogen receptor-negative MDA-MB-435 and -positive MCF-7

human breast cancer cells by palm oil tocotrienols and tamoxifen, alone and in combination.

Guthrie N, Gapor A, Chambers AF, Carroll KK.

Department of Biochemistry, The University of Western Ontario, London, Canada.

Tocotrienols are a form of vitamin E, having an unsaturated isoprenoid side-chain rather than the

saturated side-chain of tocopherols. The tocotrienol-rich fraction (TRF) from palm oil contains

alpha-tocopherol and a mixture of alpha-, gamma- and delta-tocotrienols. Earlier studies have

shown that tocotrienols display anticancer activity. We previously reported that TRF, alpha-,

gamma- and delta-tocotrienols inhibited proliferation of estrogen receptor-negative MDA-MB-435

human breast cancer cells with 50% inhibitory concentrations (IC50) of 180, 90, 30 and 90

microg/mL, respectively, whereas alpha-tocopherol had no effect at concentrations up to 500

microg/mL. Further experiments with estrogen receptor-positive MCF-7 cells showed that

tocotrienols also inhibited their proliferation, as measured by [3H] thymidine incorporation. The

IC50s for TRF, alpha-tocopherol, alpha-, gamma- and delta-tocotrienols were 4, 125, 6, 2 and 2

microg/mL, respectively. Tamoxifen, a widely used synthetic antiestrogen inhibits the growth of

MCF-7 cells with an IC50 of 0.04 microg/mL. We tested 1:1 combinations of TRF, alpha-

tocopherol and the individual tocotrienols with tamoxifen in both cell lines. In the MDA-MB-435

cells, all of the combinations were found to be synergistic. In the MCF-7 cells, only 1:1

combinations of gamma- or delta-tocotrienol with tamoxifen showed a synergistic inhibitory effect

on the proliferative rate and growth of the cells. The inhibition by tocotrienols was not overcome by

addition of excess estradiol to the medium. These results suggest that tocotrienols are effective

inhibitors of both estrogen receptor-negative and -positive cells and that combinations with

tamoxifen should be considered as a possible improvement in breast cancer therapy.

42. Nutr Cancer 1999;33(1):26-32

Induction of apoptosis in human breast cancer cells by tocopherols and tocotrienols.

Yu W, Simmons-Menchaca M, Gapor A, Sanders BG, Kline K.

Department of Zoology, University of Texas at Austin 78712, USA.

The apoptosis-inducing properties of RRR-alpha-, beta-, gamma-, and delta-tocopherols, alpha-,

gamma-, and delta-tocotrienols, RRR-alpha-tocopheryl acetate (vitamin E acetate), and RRR-alpha-

tocopheryl succinate (vitamin E succinate) were investigated in estrogen-responsive MCF7 and

estrogen-nonresponsive MDA-MB-435 human breast cancer cell lines in culture. Apoptosis was

characterized by two criteria: 1) morphology of 4,6-diamidino-2-phenylindole-stained cells and

oligonucleosomal DNA laddering. Vitamin E succinate, a known inducer of apoptosis in several

cell lines, including human breast cancer cells, served as a positive control. The estrogen-responsive

MCF7 cells were more susceptible than the estrogen-nonresponsive MDA-MB-435 cells, with

concentrations for half-maximal response for tocotrienols (alpha, gamma, and delta) and RRR-

delta-tocopherol of 14, 15, 7, and 97 micrograms/ml, respectively. The tocotrienols (alpha, gamma,

and delta) and RRR-delta-tocopherol induced MDA-MB-435 cells to undergo apoptosis, with

concentrations for half-maximal response of 176, 28, 13, and 145 micrograms/ml, respectively.

With the exception of RRR-delta-tocopherol, the tocopherols (alpha, beta, and gamma) and the

acetate derivative of RRR-alpha-tocopherol (RRR-alpha-tocopheryl acetate) were ineffective in

induction of apoptosis in both cell lines when tested within the range of their solubility, i.e., 10-200

micrograms/ml. In summary, these studies demonstrate that naturally occurring tocotrienols and

RRR-delta-tocopherol are effective apoptotic inducers for human breast cancer cells.

43. Proc Soc Exp Biol Med 2000 Sep;224(4):292-301

Antiproliferative and apoptotic effects of tocopherols and tocotrienols on preneoplastic and

neoplastic mouse mammary epithelial cells.

McIntyre BS, Briski KP, Gapor A, Sylvester PW.

College of Pharmacy, University of Louisiana at Monroe, Monroe, Louisiana 71209-0470, USA.

Studies were conducted to determine the comparative effects of tocopherols and tocotrienols on

preneoplastic (CL-S1), neoplastic (-SA), and highly malignant (+SA) mouse mammary epithelial

cell growth and viability in vitro. Over a 5-day culture period, treatment with 0-120 microM alpha-

and gamma-tocopherol had no effect on cell proliferation, whereas growth was inhibited 50%

(IC50) as compared with controls by treatment with the following: 13, 7, and 6 microM tocotrienol-

rich-fraction of palm oil (TRF); 55, 47, and 23 microM delta-tocopherol; 12, 7, and 5 microM

alpha-tocotrienol; 8, 5, and 4 microM gamma-tocotrienol; or 7, 4, and 3 microM delta-tocotrienol in

CL-S1, -SA and +SA cells, respectively. Acute 24-hr exposure to 0-250 microM alpha- or gamma-

tocopherol (CL-S1, -SA, and +SA) or 0-250 microM delta-tocopherol (CL-S1) had no effect on cell

viability, whereas cell viability was reduced 50% (LD50) as compared with controls by treatment

with 166 or 125 microM delta-tocopherol in -SA and +SA cells, respectively. Additional LD50

doses were determined as the following: 50, 43, and 38 microM TRF; 27, 28, and 23 microM alpha-

tocotrienol; 19, 17, and 14 microM gamma-tocotrienol; or 16, 15, or 12 microM delta-tocotrienol in

CL-S1, -SA, and +SA cells, respectively. Treatment-induced cell death resulted from activation of

apoptosis, as indicated by DNA fragmentation. Results also showed that CL-S1, -SA, and +SA cells

preferentially accumulate tocotrienols as compared with tocopherols, and this may partially explain

why tocotrienols display greater biopotency than tocopherols. These data also showed that highly

malignant +SA cells were the most sensitive, whereas the preneoplastic CL-S1 cells were the least

sensitive to the antiproliferative and apoptotic effects of tocotrienols, and suggest that tocotrienols

may have potential health benefits in preventing and/or reducing the risk of breast cancer in women.

44. Lipids 2000 Feb;35(2):171-80

Antiproliferative and apoptotic effects of tocopherols and tocotrienols on normal mouse mammary

epithelial cells.

McIntyre BS, Briski KP, Tirmenstein MA, Fariss MW, Gapor A, Sylvester PW.

Colleges of Pharmacy, University of Louisiana, Monroe 71209-0470, USA.

Studies were conducted to determine the comparative effects of tocopherols and tocotrienols on

normal mammary epithelial cell growth and viability. Cells isolated from midpregnant BALB/c

mice were grown within collagen gels and maintained on serum-free media. Treatment with 0-120

microM alpha- and gamma-tocopherol had no effect, whereas 12.5-100m microM tocotrienol-rich

fraction of palm oil (TRF), 100-120 microM delta-tocopherol, 50-60 microM alpha-tocotrienol, and

8-14 microM gamma- or delta-tocotrienol significantly inhibited cell growth in a dose-responsive

manner. In acute studies, 24-h exposure to 0-250 microM alpha-, gamma-, and delta-tocopherol had

no effect, whereas similar treatment with 100-250 microM TRF, 140-250 microM alpha-, 25-100

microM gamma- or delta-tocotrienol significantly reduced cell viability. Growth-inhibitory doses of

TRF, delta-tocopherol, and alpha-, gamma-, and delta-tocotrieol were shown to induce apoptosis in

these cells, as indicated by DNA fragmentation. Results also showed that mammary epithelial cells

more easily or preferentially took up tocotrienols as compared to tocopherols, suggesting that at

least part of the reason tocotrienols display greater biopotency than tocopherols is because of

greater cellular accumulation. In summary, these findings suggest that the highly biopotent gamma-

and delta-tocotrienol isoforms may play a physological role in modulating normal mammary gland

growth, function, and remodeling.

45. J Natl Cancer Inst 1995 May 17;87(10):746-50

Comparison of the effects of a pure steroidal antiestrogen with those of tamoxifen in a model of

human breast cancer.

Osborne CK, Coronado-Heinsohn EB, Hilsenbeck SG, McCue BL, Wakeling AE,

McClelland RA, Manning DL, Nicholson RI.

Department of Medicine, University of Texas Health Science Center at San Antonio 78284-7884,

USA.

BACKGROUND: Tamoxifen, a nonsteroidal estrogen antagonist, is the most prescribed drug for

the treatment of breast cancer. The use of tamoxifen is limited, however, by the development of

resistance to this compound in most patients. Although tamoxifen behaves primarily as an estrogen

antagonist, it has agonist (or growth-stimulatory) activity as well. ICI 182,780 is a 7 alpha-

alkylsulfinyl analogue of estradiol lacking agonist activity. The absence of agonist activity may

make this steroidal antiestrogen superior to tamoxifen in suppressing tumor cell growth. PURPOSE:

We compared the inhibitory effects of ICI 182,780, tamoxifen, and estrogen withdrawal on the

growth of established tumors and on tumorigenesis in a model system that uses estrogen-dependent,

human MCF-7 breast tumor cells growing in athymic nude mice. We also studied the hormonal

responsiveness of tumors that became resistant to the two estrogen antagonists and the effects of

these drugs on estrogen-regulated gene expression. METHODS: MCF-7 cells were injected

subcutaneously into the flanks of castrated, female nude mice. The effects of repeated doses of

tamoxifen and ICI 182,780 (500 micrograms and 5 mg, respectively) on the growth of established

tumors (8-10 mm in size) were determined after supplemental estrogen was removed. The effects of

antiestrogen treatments on the process of tumorigenesis, in the absence of estrogen

supplementation, were determined by initiating drug administration on the same day as tumor cell

inoculation. To evaluate the hormonal responsiveness of tumors resistant to tamoxifen and ICI

182,780, 1-mm3 segments of the tumors were transplanted onto the flanks of new recipient mice,

which were then treated with estrogen or the antiestrogens--alone or in combination. Tumor growth

was monitored by measuring tumor volumes twice a week. Expression of the estrogen-responsive

genes, pLIV1 and pS2, in the tumors of treated animals was analyzed using blots of total cellular

RNA and complementary DNA probes. RESULTS: Treatment with ICI 182,780 suppressed the

growth of established tumors twice as long as treatment with tamoxifen or estrogen withdrawal.

Tumorigenesis, in the absence of supplemental estrogen, was delayed to a greater extent in ICI

182,780-treated mice than in tamoxifen-treated mice. ICI 182,780 was found to be more effective

than tamoxifen in reducing the expression of estrogen-regulated genes. Most tumors eventually

became resistant to ICI 182,780 and grew independently of estrogen. CONCLUSIONS: ICI

182,780 is a more effective estrogen antagonist than tamoxifen in the MCF-7 tumor cell/nude

mouse model system.

46. Semin Urol Oncol 1999 May;17(2):85-90

Vitamin E, alpha- and gamma-tocopherol, and prostate cancer.

Moyad MA, Brumfield SK, Pienta KJ.

Section of Urology, University of Michigan, Ann Arbor 48109-0330, USA.

Vitamin E is one of the most researched compounds in medicine. Vitamin E is actually a general

name for potentially eight different compounds, so supplements can contain several forms and

vitamin E in the diet also differs from the form found over the counter. There has been a strong

interest in this supplement in the prostate cancer arena primarily because of a Finnish study that

demonstrated a lower morbidity and mortality from this disease in men taking 50 mg of synthetic

(alpha-tocopherol) vitamin E daily. In addition, observations from laboratory and clinical studies

dealing with heart disease have found that gamma-tocopherol may also play a significant role in

prevention; therefore, we decided to test the ability of this compound (versus synthetic vitamin E)

to control the growth of a human prostate cancer cell line. Gamma-tocopherol was found to be

superior to alpha-tocopherol in terms of cell inhibition in vitro. Both forms of vitamin E (and

others) should be thoroughly evaluated in the future to provide the most effective chemoprevention

information to the patient.

47. J Natl Cancer Inst 2000 Dec 20;92(24):2018-23

Association between alpha-tocopherol, gamma-tocopherol, selenium, and subsequent prostate

cancer.

Helzlsouer KJ, Huang HY, Alberg AJ, Hoffman S, Burke A, Norkus EP, Morris JS, Comstock GW.

Department of Epidemiology, The Johns Hopkins School of Hygiene and Public Health, Baltimore,

MD 21205, USA. Khelzlso@jhsph.edu

BACKGROUND: Selenium and alpha-tocopherol, the major form of vitamin E in supplements,

appear to have a protective effect against prostate cancer. However, little attention has been paid to

the possible role of gamma-tocopherol, a major component of vitamin E in the U.S. diet and the

second most common tocopherol in human serum. A nested case-control study was conducted to

examine the associations of alpha-tocopherol, gamma-tocopherol, and selenium with incident

prostate cancer. METHODS: In 1989, a total of 10,456 male residents of Washington County, MD,

donated blood for a specimen bank. A total of 117 of 145 men who developed prostate cancer and

233 matched control subjects had toenail and plasma samples available for assays of selenium,

alpha-tocopherol, and gamma-tocopherol. The association between the micronutrient concentrations

and the development of prostate cancer was assessed by conditional logistic regression analysis. All

statistical tests were two-sided. RESULTS: The risk of prostate cancer declined, but not linearly,

with increasing concentrations of alpha-tocopherol (odds ratio (highest versus lowest fifth) = 0.65;

95% confidence interval = 0.32--1.32; P(trend) =.28). For gamma-tocopherol, men in the highest

fifth of the distribution had a fivefold reduction in the risk of developing prostate cancer than men

in the lowest fifth (P:(trend) =.002). The association between selenium and prostate cancer risk was

in the protective direction with individuals in the top four fifths of the distribution having a reduced

risk of prostate cancer compared with individuals in the bottom fifth (P(trend) =.27). Statistically

significant protective associations for high levels of selenium and alpha-tocopherol were observed

only when gamma-tocopherol concentrations were high. CONCLUSIONS: The use of combined

alpha- and gamma- tocopherol supplements should be considered in upcoming prostate cancer

prevention trials, given the observed interaction between alpha-tocopherol, gamma-tocopherol, and

selenium.

48. Circulation 1989 Sep;80(3):719-23

Lipoproteins and the pathogenesis of atherosclerosis.

Steinberg D, et al.

NO ABSTRACT AVAILABLE

49. J Clin Invest 1989 Oct;84(4):1086-95

Evidence for the presence of oxidatively modified low density lipoprotein in atherosclerotic lesions

of rabbit and man.

Yla-Herttuala S, Palinski W, Rosenfeld ME, Parthasarathy S, Carew TE, Butler S, Witztum JL,

Steinberg D.

Department of Medicine, University of California, San Diego, La Jolla 92093-0613.

Three lines of evidence are presented that low density lipoproteins gently extracted from human and

rabbit atherosclerotic lesions (lesion LDL) greatly resembles LDL that has been oxidatively

modified in vitro. First, lesion LDL showed many of the physical and chemical properties of

oxidized LDL, properties that differ from those of plasma LDL: higher electrophoretic mobility, a

higher density, higher free cholesterol content, and a higher proportion of sphingomyelin and

lysophosphatidylcholine in the phospholipid fraction. A number of lower molecular weight

fragments of apo B were found in lesion LDL, similar to in vitro oxidized LDL. Second, both the

intact apo B and some of the apo B fragments of lesion LDL reacted in Western blots with antisera

that recognize malondialdehyde-conjugated lysine and 4-hydroxynonenal lysine adducts, both of

which are found in oxidized LDL; plasma LDL and LDL from normal human intima showed no

such reactivity. Third, lesion LDL shared biological properties with oxidized LDL: compared with

plasma LDL, lesion LDL produced much greater stimulation of cholesterol esterification and was

degraded more rapidly by macrophages. Degradation of radiolabeled lesion LDL was competitively

inhibited by unlabeled lesion LDL, by LDL oxidized with copper, by polyinosinic acid and by

malondialdehyde-LDL, but not by native LDL, indicating uptake by the scavenger receptor(s).

Finally, lesion LDL (but not normal intimal LDL or plasma LDL) was chemotactic for monocytes,

as is oxidized LDL. These studies provide strong evidence that atherosclerotic lesions, both in man

and in rabbit, contain oxidatively modified LDL.

50. Proc Natl Acad Sci U S A 1993 Mar 1;90(5):1771-5

Gamma-tocopherol detoxification of nitrogen dioxide: superiority to alpha-tocopherol.

Cooney RV, Franke AA, Harwood PJ, Hatch-Pigott V, Custer LJ, Mordan LJ.

Cancer Research Center of Hawaii, University of Hawaii, Honolulu 96813.

In the vitamin E group, alpha-tocopherol is generally considered to be the most potent antioxidant

with the highest vitamin bioactivity, yet gamma-tocopherol is produced in greater amounts by many

plants and is the principal tocopherol in the United States diet. This report describes a fundamental

difference in the chemical reactivities of alpha-tocopherol and gamma-tocopherol with nitrogen

dioxide (NO2), which leads to the formation of a nitrosating agent from alpha-tocopherol, but not

from gamma-tocopherol. Nitric oxide (NO) is a major product of the reaction of gamma-tocopherol

with NO2, while alpha-tocopherol reacts with NO2 to form an intermediate tocopheroxide

analogue. The biological significance of gamma-tocopherol is suggested by limited epidemiological

data as well as the observation that it is a more potent inhibitor than alpha-tocopherol of neoplastic

transformation during the postinitiation phase in

3-methylcholanthrene-treated C3H/10T1/2 murine fibroblasts. This latter property suggests the

superiority of gamma-tocopherol in a mammalian biological assay and a role for endogenous NO

production in promotion of neoplastic transformation.

51. Proc Natl Acad Sci U S A 1997 Apr 1;94(7):3217-22

gamma-tocopherol traps mutagenic electrophiles such as NO(X) and complements alpha-

tocopherol: physiological implications.

Christen S, Woodall AA, Shigenaga MK, Southwell-Keely PT, Duncan MW, Ames BN.

Division of Biochemistry and Molecular Biology, University of California, Berkeley 94720, USA.

Peroxynitrite, a powerful mutagenic oxidant and nitrating species, is formed by the near diffusion-

limited reaction of .NO and O2.- during activation of phagocytes. Chronic inflammation induced by

phagocytes is a major contributor to cancer and other degenerative diseases. We examined how

gamma-tocopherol (gammaT), the principal form of vitamin E in the United States diet, and alpha-

tocopherol (alphaT), the major form in supplements, protect against peroxynitrite-induced lipid

oxidation. Lipid hydroperoxide formation in liposomes (but not isolated low-density lipoprotein)

exposed to peroxynitrite or the .NO and O2.- generator SIN-1 (3-morpholinosydnonimine) was

inhibited more effectively by gammaT than alphaT. More importantly, nitration of gammaT at the

nucleophilic 5-position, which proceeded in both liposomes and human low density lipoprotein at

yields of approximately 50% and approximately 75%, respectively, was not affected by the

presence of alphaT. These results suggest that despite alphaT's action as an antioxidant gammaT is

required to effectively remove the peroxynitrite-derived nitrating species. We postulate that

gammaT acts in vivo as a trap for membrane-soluble electrophilic nitrogen oxides and other

electrophilic mutagens, forming stable carbon-centered adducts through the nucleophilic 5-position,

which is blocked in alphaT. Because large doses of dietary alphaT displace gammaT in plasma and

other tissues, the current wisdom of vitamin E supplementation with primarily alphaT should be

reconsidered.

52. J Nutr 2001 Feb;131(2):369S-73S

Molecular aspects of alpha-tocotrienol antioxidant action and cell signalling.

Packer L, Weber SU, Rimbach G.

Department of Molecular and Cellular Biology, University of California, Berkeley, CA 94720-

3200, USA. packer@socrates.berkeley.edu

Vitamin E, the most important lipid-soluble antioxidant, was discovered at the University of

California at Berkeley in 1922 in the laboratory of Herbert M. Evans (Science 1922, 55: 650). At

least eight vitamin E isoforms with biological activity have been isolated from plant sources. Since

its discovery, mainly antioxidant and recently also cell signaling aspects of tocopherols and

tocotrienols have been studied. Tocopherols and tocotrienols are part of an interlinking set of

antioxidant cycles, which has been termed the antioxidant network. Although the antioxidant

activity of tocotrienols is higher than that of tocopherols, tocotrienols have a lower bioavailability

after oral ingestion. Tocotrienols penetrate rapidly through skin and efficiently combat oxidative

stress induced by UV or ozone. Tocotrienols have beneficial effects in cardiovascular diseases both

by inhibiting LDL oxidation and by down-regulating 3-hydroxyl-3-methylglutaryl-coenzyme A

(HMG CoA) reductase, a key enzyme of the mevalonate pathway. Important novel antiproliferative

and neuroprotective effects of tocotrienols, which may be independent of their antioxidant activity,

have also been described.

53. Free Radic Biol Med 1991;10(5):263-75

Free radical recycling and intramembrane mobility in the antioxidant properties of alpha-tocopherol

and alpha-tocotrienol.

Serbinova E, Kagan V, Han D, Packer L.

Department of Molecular and Cell Biology, University of California, Berkeley 94720.

d-Alpha-tocopherol (2R,4'R,8'R-Alpha-tocopherol) and d-alpha-tocotrienol are two vitamin E

constituents having the same aromatic chromanol "head" but differing in their hydrocarbon "tail":

tocopherol with a saturated and toctrienol with an unsaturated isoprenoid chain. d-Alpha-tocopherol

has the highest vitamin E activity, while d-alpha-tocotrienol manifests only about 30% of this

activity. Since vitamin E is considered to be physiologically the most important lipid-soluble chain-

breaking antioxidant of membranes, we studied alpha-tocotrienol as compared to alpha-tocopherol

under conditions which are important for their antioxidant function. d-Alpha-tocotrienol possesses

40-60 times higher antioxidant activity against (Fe2+ + ascorbate)- and (Fe2+ + NADPH)-induced

lipid peroxidation in rat liver microsomal membranes and 6.5 times better protection of cytochrome

P-450 against oxidative damage than d-alpha-tocopherol. To clarify the mechanisms responsible for

the much higher antioxidant potency of d-alpha-tocotrienol compared to d-alpha-tocopherol, ESR

studies were performed of recycling efficiency of the chromanols from their chromanoxyl radicals.

1H-NMR measurements of lipid molecular mobility in liposomes containing chromanols, and

fluorescence measurements which reveal the uniformity of distribution (clusterizations) of

chromanols in the lipid bilayer.

From the results, we concluded that this higher antioxidant potency of d-alpha-tocotrienol is due to

the combined effects of three properties exhibited by d-alpha-tocotrienol as compared to d-alpha-

tocopherol: (i) its higher recycling efficiency from chromanoxyl radicals, (ii) its more uniform

distribution in membrane bilayer, and (iii) its stronger disordering of membrane lipids which makes

interaction of chromanols with lipid radicals more efficient. The data presented show that there is a

considerable discrepancy between the relative in vitro antioxidant activity of d-alpha-tocopherol

and d-alpha-tocotrienol with the conventional bioassays of their vitamin activity.

54. Palm oil vitamin E protects against ischemia/reperfusion injury in the isolated perfused

Langendorff heart

Serbinova E.; Khwaja S.; Catudioc J.; Ericson J.; Torres Z.; Gapor A.; Kagan V.; Packer L.

Malaysia Palm Oil Research Institute, Peti Surat 10620,50720 Kuala Lumpur Malaysia Nutrition

Research ( NUTR. RES. ) ( United States ) 1992 , 12/SUPPL. (S203-S215)

We studied the effect of palm oil vitamin E on Langendorff perfused rat hearts subjected to 40

minutes of global ischemia. Our results demonstrated that palm oil vitamin E was more efficient in

the protection of isolated Langendorff heart against ischemia/reperfusion injury than tocopherol as

measured by its mechanical recovery. Palm oil vitamin E completely suppressed LDH enzyme

leakage from ischemic hearts, prevented the decrease in ATP and creatine phosphate levels and

inhibited the formation of endogenous lipid peroxidation products. Our data indicate that a palm oil

vitamin E mixture containing both alpha-tocopherol and alpha-tocotrienol may be more efficient

than alpha-tocopherol alone in the protection of the heart against oxidative stress induced by

ischemia-reperfusion.

55. Neurosci Lett 1995 Aug 11;195(3):179-82

Tocotrienols from palm oil as potent inhibitors of lipid peroxidation and protein oxidation in rat

brain mitochondria.

Kamat JP, Devasagayam TP.

Radiation Biology and Biochemistry Division, Bhabha Atomic Research Centre,

Bombay, India.

The tocotrienol-rich-fraction (TRF) from palm oil, being tried as a more economical and efficient

substitute for alpha-tocopherol, significantly inhibited oxidative damage in vitro to both lipids and

proteins in rat brain mitochondria induced by ascorbate-Fe2+, the free radical initiator

azobis(2-amidopropane)dihydrochloride (AAPH) and photosensitisation. The observed inhibitory

effect was both time- and concentration-dependent. At a low concentration of 5 microM, TRF can

significantly inhibit oxidative damage to both lipids and proteins. The inhibitory effect of TRF

seems to be mainly due to gamma-tocotrienol and to a lesser extent alpha- and delta-tocotrienols.

TRF was significantly more effective than alpha-tocopherol. This fraction from palm oil can be

considered a natural antioxidant supplement capable of protecting the brain against oxidative

damage and thereby from the ensuing adverse alterations.

56. Mol Cell Biochem 1997 May;170(1-2):131-7

Tocotrienols from palm oil as effective inhibitors of protein oxidation and lipid peroxidation in rat

liver microsomes.

Kamat JP, Sarma HD, Devasagayam TP, Nesaretnam K, Basiron Y.

Radiation Biology and Biochemistry Division, Bhabha Atomic Research Centre, Bombay, India.

Tocotrienols from palm oil showed significant ability to inhibit oxidative damage induced by

ascorbate-Fe2+ and photosensitization, involving different mechanisms, in rat liver microsomes.

The tocotrienol-rich fraction from palm oil (TRF), being tried as a more economical and efficient

substitute for alpha-tocopherol, showed time- and concentration-dependent inhibition of protein

oxidation as well as lipid peroxidation. It was more effective against protein oxidation. The extent

of inhibition by TRF varied with different peroxidation products such as conjugated dienes, lipid

hydroperoxides and thiobarbituric acid reactive substances (TBARS). Among the constituents of

TRF, gamma-tocotrienol was the most effective followed by its alpha- and delta-isomers. In

general, at a low concentration of 5 microM, TRF was able to prevent oxidative damage to

significant extent (37% inhibition of protein oxidation and 27-30% of lipid peroxidation at 1 h of

incubation). The protective ability of TRF (30.1% at 5 microM with TBARS formation) was

significantly higher than that of the dominant form of vitamin E, alpha-tocopherol (16.5% under

same conditions). Hence our studies indicate that this fraction from palm oil can be considered as an

effective natural antioxidant supplement capable of protecting cellular membranes against oxidative

damage.

57. Proc Natl Acad Sci U S A 1993 Mar 1;90(5):1771-5

Gamma-tocopherol detoxification of nitrogen dioxide: superiority to alpha-tocopherol.

Cooney RV, Franke AA, Harwood PJ, Hatch-Pigott V, Custer LJ, Mordan LJ.

Cancer Research Center of Hawaii, University of Hawaii, Honolulu 96813.

In the vitamin E group, alpha-tocopherol is generally considered to be the most potent antioxidant

with the highest vitamin bioactivity, yet gamma-tocopherol is produced in greater amounts by many

plants and is the principal tocopherol in the United States diet. This report describes a fundamental

difference in the chemical reactivities of alpha-tocopherol and gamma-tocopherol with nitrogen

dioxide (NO2), which leads to the formation of a nitrosating agent from alpha-tocopherol, but not

from gamma-tocopherol. Nitric oxide (NO) is a major product of the reaction of gamma-tocopherol

with NO2, while alpha-tocopherol reacts with NO2 to form an intermediate tocopheroxide

analogue. The biological significance of gamma-tocopherol is suggested by limited epidemiological

data as well as the observation that it is a more potent inhibitor than alpha-tocopherol of neoplastic

transformation during the postinitiation phase in 3-methylcholanthrene-treated C3H/10T1/2 murine

fibroblasts. This latter property suggests the superiority of gamma-tocopherol in a mammalian

biological assay and a role for endogenous NO production in promotion of neoplastic

transformation.

58. J Nutr 1985 Jun;115(6):807-13

Oral alpha-tocopherol supplements decrease plasma gamma-tocopherol levels in humans.

Handelman GJ, Machlin LJ, Fitch K, Weiter JJ, Dratz EA.

In a cross-sectional survey of 86 elderly persons, it was observed that subjects with elevated plasma

alpha-tocopherol levels had depressed plasma gamma-tocopherol. Tocopherols were measured by

both reverse-phase and normal-phase high performance liquid chromatography (HPLC). When

eight human volunteers (age range 30-60) were given 1200 IU of all-rac-alpha-tocopherol daily for

8 wk, plasma gamma-tocopherol and beta-tocopherol decreased in all subjects. After

supplementation, gamma-tocopherol values were typically 30-50% of initial values, and alpha-

tocopherol values were typically 200-400% of initial values. These results suggest that intestinal

uptake and/or plasma transport make more efficient use of alpha-tocopherol than of gamma- or

beta-tocopherol. Moreover, the results indicate that the ratio of gamma- to alpha-tocopherol in

plasma would be a more satisfactory index to measure compliance in trials involving

supplementation with alpha-tocopherol.

59. Proc Natl Acad Sci U S A 1996 Jun 11;93(12):6002-7

A new endogenous natriuretic factor: LLU-alpha.

Wechter WJ, Kantoci D, Murray ED Jr, D'Amico DC, Jung ME, Wang WH.

Laboratory of Chemical Endocrinology, Loma Linda University, CA 92350, USA.

For over three decades, renal physiology has sought a putative natriuretic hormone (third factor)

that might control the body's pool of extracellular fluid, an important determinant in hypertension,

congestive heart failure, and cirrhosis. In our search for this hormone, we have isolated several pure

natriuretic factors from human uremic urine that would appear, alone or in combination, to mark a

cluster of phenomena previously presumed to be that of a single ?natriuretic hormone.? This paper

reports the purification, chemical structure, and total synthesis of the first of these compounds,

LLU-alpha, which proved to be 2,7,8-trimethyl-2-(beta-carboxyethyl)-6-hydroxychroman,

presumably a metabolite of gamma-tocopherol. Both natural LLU-alpha and synthetic material are

identical (except for optical activity) with respect to structure and biological activity. It appears that

the natriuretic activity of LLU-alpha is mediated by inhibition of the 70 pS K+ channel in the apical

membrane of the thick ascending limb of the kidney.

60. Biol Pharm Bull 2000 Nov;23(11):1395-7

Production of LLU-alpha following an oral administration of gamma-tocotrienol or gamma-

tocopherol to rats

Hattori A, Fukushima T, Yoshimura H, Abe K, Ima K.

Department of Bio-Analytical Chemisty, Graduate School Pharmacelutical Sciences, the University

of Tokyo, Japan.

An oral administration of gamma-tocotrienol (gamma-T3) or gamma-tocopherol (gamma-Toc) to

male rats caused an increase of the concentration of 2,7,8-trimethyl-2-(beta-carboxyethyl)-6-

hydroxy chroman (LLU-alpha, gamma-CEHC), a natriuretic compound, in plasma with a Tmax of

9 h. The configuration at C-2 of LLU-alpha produced from gamma-T3 or gamma-Toc was assigned

as S-form by an HPLC equipped with a chiral column. These data indicated that LLU-alpha was

produced not only from gamma-Toc but also gamma-T3, without racemization at C-2

in rats.

61. Anal Biochem 2000 Jun 1;281(2):209-15

Occurrence and determination of a natriuretic hormone, 2,7,8-trimethyl-2-(beta-carboxyethyl)-6-

hydroxy chroman, in rat plasma, urine, and bile.

Hattori A, Fukushima T, Imai K.

Department of Bio-Analytical Chemistry, Graduate School of Pharmaceutical Sciences, University

of Tokyo, Japan.

The occurrence of a new natriuretic hormone, 2,7,8-trimethyl-2-(beta-carboxyethyl)-6-hydroxy

chroman (LLU-alpha, gamma-CEHC) in rat plasma was demonstrated and its concentration was

determined using a coupled-column HPLC with a fluorometric derivatization with 4-N,N-

dimethylaminosulfonyl-7-piperazino-2,1,3-benzoxadiazol e (DBD-PZ) followed by O-acetylation.

The concentration of LLU-alpha was 328 113 nM in rat plasma (N = 5). LLU-alpha was found in

not only urine, but also bile, suggesting an enterohepatic circulation in body. We also assigned the

configuration at C-2 of LLU-alpha in these biological fluids as (S)-form by an HPLC with a chiral

column. The LLU-alpha concentration decreased significantly by fasting for 3 days (P < 0.01).

These results support the possibility that LLU-alpha is produced from gamma-tocopherol in diet via

oxidative metabolism without racemization.

62. Clin Exp Hypertens 1999 Nov;21(8):1297-313

Effect of gamma-tocotrienol on blood pressure, lipid peroxidation and total antioxidant status in

spontaneously hypertensive rats (SHR).

Newaz MA, Nawal NN.

Faculty of Medicine, International Islamic University Malaysia. Newaz@iiu.edu.my

The aim of this study was to determine the effects of gamma tocotrienol on lipid peroxidation and

total antioxidant status of spontaneously hypertensive rats (SHR), comparing them with normal

Wistar Kyoto (WKY) rats. SHR were divided into three groups and treated with different doses of

gamma tocotrienol (gamma1, 15 mg/kg diet; gamma2, 30 mg/kg diet and gamma3, 150 mg/kg

diet). Normal WKY and untreated SHR were used as normal (N) and hypertensive control (HC).

Blood pressure were recorded every fortnightly for three months. At the end of the trial, animals

were killed and measurement of plasma total antioxidant status, plasma superoxide dismutase

(SOD) activity and lipid peroxide levels in plasma and blood vessels were carried out following

well established methods. Study shows that lipid peroxides were significantly higher in

hypertensive plasma and blood vessels compared to that of normal rats (Plasma- N: 0.060.01, HC:

0.130.008; p<0.001, B1. Vessels - N: 0.470.17, HC: 0.960.37; p<0.001). SOD activity was

significantly lower in hypertensive than normal rats (N = 148.5829.56 U/ml, HC = 110.0814.36

U/ml; p = 0.014). After three months of antioxidant trial with gamma-tocotrienol, it was found that

all the treated groups have reduced plasma lipid peroxides concentration but was only significant

for group gamma1 (gamma1: 0.1090.026, HC: 0.1320.008; p = 0.034). On the other hand, lipid

peroxides in blood vessels reduced significantly in all treated groups (gamma1; p<0.05, gamma2;

p<0.001, gamma3; p<0.005). All the three treated groups showed improve total antioxidant status

(p<0.001) significantly. SOD activity also showed significant improvement in all groups (gamma1:

p<0.001, gamma2: p<0.05, gamma3: p<0.001). Correlation studies showed that, total antioxidant

status (TAS) and SOD were significantly negatively correlated with blood pressure in normal rats

(p = 0.007; p = 0.008) but not in SHR control. This correlation regained in all three groups SHR's

after treatment with tocotrienol. Lipid peroxides in blood vessel and plasma showed a positive

correlation with blood pressure in normal and SHR control. This correlation also remains in treated

groups significantly except that in gamma3 where positive correlation with plasma lipid peroxide

was not significant. In conclusion it was found that antioxidant supplement of gamma-tocotrienol

may prevent development of increased blood pressure, reduce lipid peroxides in plasma and blood

vessels and enhanced total antioxidant status including SOD activity.

63. Am J Clin Nutr 1988 Sep;48(3):612-9

Safety of oral intake of vitamin E.

Bendich A, Machlin LJ.

Clinical Nutrition, Hoffman-La Roche Inc, Nutley, NJ 07110.

A review of the literature concerning the safety of oral intake of vitamin E indicated that the

toxicity of vitamin E is low. Vitamin E supplementation has resulted in inconsistent effects in

serum lipid and lipoprotein levels. Animal studies showed that vitamin E is not mutagenic,

carcinogenic, or teratogenic. In human studies with double-blind protocols and in large population

studies, oral vitamin E supplementation resulted in few side effects even at doses as high as 3200

mg/d (3200 IU/d).

64. Am J Clin Nutr 1995 Dec;62(6 Suppl):1510S-1516S

Safety of antioxidant vitamins and beta-carotene.

Diplock AT.

Division of Biochemistry and Molecular Biology, United Medical School, Guy's Hospital,

University of London, United Kingdom.

Epidemiologic evidence links high antioxidant status with low risk of degenerative disease. Optimal

intakes of antioxidants may not be achievable by diet alone; supplements may be taken, particularly

in subgroups of the population at high risk. It is thus necessary to ensure that antioxidant

supplements are safe and free from side effects. The toxicity of vitamin E is low; no mutagenic,

teratogenic, or carcinogenic effects are known and in double-blind studies in which large amounts

of vitamin E were used in humans, no side effects occurred. High concentrations are contraindicated

in subjects with vitamin K-associated blood coagulation disorders, and the toxicity in normal

subjects ingesting large amounts of vitamin E over long periods requires additional investigation.

Toxicity of beta-carotene also is low. Evidence from human toxicity trials is not available but there

is much circumstantial evidence that 15-50 mg/d is without side effects except for

hypercarotenemia in some subjects at high intakes. The findings of more lung cancer in subjects

who smoked and who were given 20 mg beta-carotene/d than in those given a placebo could be

influenced by the cancer being well advanced before beta-carotene administration. Massive

anecdotal evidence exists that vitamin C (at > or = 1 g/d) is safe. Exhaustive literature searches have

failed to reveal a controlled study of vitamin C toxicity in human subjects. Anxiety exists about

oxalate stone formation, uricosuria, vitamin B-12 destruction, mutagenicity, and iron overload, but

the consensus is that adverse effects do not occur in healthy subjects ingesting large amounts of

vitamin C.

65. Arch Intern Med 1996 May 13;156(9):925-35

Safety of antioxidant vitamins.

Meyers DG, Maloley PA, Weeks D.

Department of Internal Medicine, Kansas University Medical Center, Kansas City, USA.

As a result of the many scientific and popular press reports of the benefits of antioxidant vitamins

(vitamin A, beta-carotene, vitamin E, and ascorbic acid), it is estimated that 40% of the US

population is consuming vitamin supplements. The efficacy of these supplements is not yet proved,

and some have questioned their safety. Approximately 10 to 15 cases of vitamin A toxic reactions

are reported per year in the United States, usually at doses greater than 100,000 IU/d. No adverse

effects have been reported for beta-carotene. The frequency of vitamin E toxic reactions is not well

delineated, but case reports are few at dosages less than 3200 mg/d. Ascorbic acid toxic reactions

are rare at dosages less than 4 g/d. Despite a lack of clinical trial data, it seems that antioxidant

vitamins are safe, although prudence might dictate their avoidance by women of childbearing

potential, persons with liver disease or renal dysfunction, and those taking certain medications or

undergoing specific laboratory tests.

66. Am J Clin Nutr 1981 Sep;34(9):1701-5

Effect of vitamin E on prothrombin levels in warfarin-induced vitamin K deficiency.

Corrigan JJ Jr, Ulfers LL.

Rats rendered lightly vitamin K deficient with warfarin (0.01 mg/100 g, IP) and given the

equivalent of 1000 units of vitamin E/kg IM for 7 days, showed a marked reduction in functional

factor II activity, but normal factor II levels using Echis venom on coagulation analysis. In 12

humans receiving warfarin, vitamin E was administered in doses of 100 or 400 units/day orally for

4 wk. The results in these patients showed no significant change in the prothrombin time, factor II

coagulant activity, or factor II antigen (by electroimmunoassay). However, by using a ratio of factor

II coagulant activity to immunoreactive protein, significant reduction was observed when compared

to pretreatment ratios. These data suggest that vitamin E acts at the step mediated by vitamin K and

not in the synthesis of the factor II precursor. Although the administration of high doses of vitamin

E in animals, and possibly humans, with vitamin K deficiency potentiates the vitamin K deficiency,

this effect is not clinically obvious with 400 IU/day or less.

67. Am J Clin Nutr 1997 Feb;65(2):496-502

Erythrocyte vitamin E and plasma ascorbate concentrations in relation to erythrocyte peroxidation

in smokers and nonsmokers: dose response to vitamin E supplementation.

Brown KM, Morrice PC, Duthie GG.

Rowett Research Institute, Bucksburn, Scotland, United Kingdom. kmb@rri.sari.ac.uk

Many human degenerative diseases involve free radical processes that nutritional antioxidants may

ameliorate or prevent, but the optimum intake of such nutrients has yet to be established.

Requirement will depend in part on the level of exposure to exogenous and endogenous reactive

oxygen species. Smokers incur a sustained degree of oxidant stress from both of these sources,

increasing their requirements for vitamins E and C. Male smokers (n = 50) from a Scottish

population with habitually low vitamin E and vitamin C intakes consistently had lower plasma

ascorbate concentrations (P < 0.02) and greater susceptibility to hydrogen peroxide-stimulated

erythrocyte peroxidation in vitro (P < 0.001) than did nonsmokers (n = 50) from the same

population. Erythrocyte vitamin E concentrations increased in a dose-dependent manner during 20

wk of supplementation with 70, 140, 560, and 1050 mg D-alpha-tocopherol/d. In smokers each dose

was associated with a significant decrease in susceptibility of erythrocytes to peroxidation (P <

0.001). However, red cells of nonsmokers receiving the 1050-mg supplement had an increased

susceptibility to peroxidation. Moreover, prolonged supplementation with D-alpha-tocopherol in

nonsmokers induced a decline in plasma ascorbate concentration (P < 0.02) in association with an

increasing erythrocyte vitamin E uptake (P < 0.001), and in nonsmokers receiving 1050 mg, the

susceptibility to peroxidation also increased (P < 0.001). Thus, vitamin E may have prooxidant

activity in nonsmokers at high and prolonged intake