Both lovastatin (a fungal product) and a tocotrienol rich fraction (TRF(25), a mixture of tocols isolated from stabilized and heated rice bran containing desmethyl [d-P(21)-T3] and didesmethyl [d-P(25)-T3] tocotrienols) are potent hypocholesterolemic agents, although they suppress cholesterol biosynthesis by different mechanisms. To determine additive and/or synergistic effects of both agents, chickens were fed diets supplemented with 50 ppm TRF(25) or d-P(25)-T3 in combination with 50 ppm lovastatin for 4 weeks. Combinations of d-P(25)-T3 with lovastatin were found most effective in reducing serum total cholesterol and low-density lipoprotein (LDL) cholesterol compared to the control diet or individual supplements. The mixture of TRF(25)+lovastatin inhibited the activity of beta-hydroxy-beta-methylglutaryl coenzymeA reductase (21%) compared to lovastatin alone, which did not change its activity. Cholesterol 7alpha-hydroxylase activity was increased by lovastatin (11%) and by lovastatin plus TRF(25) (19%). TRF(25)+lovastatin decreased levels of serum total cholesterol (22%), LDL cholesterol (42%), apolipoprotein B (13-38%), triglycerides (19%), thromboxane B(2) (34%) and platelet factor 4 (26%), although high-density lipoprotein (HDL) cholesterol, and apolipoprotein A1 levels were unaffected. The mixture of TRF(25)+lovastatin showed greater effects than did the individual treatments alone, reflecting possible additive pharmacological actions. The effects, however, of the d-P(25)-T3/lovastatin combination were no greater than that of d-P(25)-T3 alone, possibly indicating that d-P(25)-T3 produced a maximum cholesterol lowering effect at the concentration used.
Publications
Antioxidant activity of tocopherols, tocotrienols, and gamma-oryzanol components from rice bran against cholesterol oxidation accelerated by 2,2′-azobis(2-methylpropionamidine) dihydrochloride
Xu Z, Hua N, Godber JS.
J Agric Food Chem. 2001 Apr;49(4):2077-81.
The antioxidant activities of vitamin E (alpha-tocopherol, alpha-tocotrienol, gamma-tocopherol, and gamma-tocotrienol) and gamma-oryzanol components (cycloartenyl ferulate, 24-methylenecycloartanyl ferulate, and campesteryl ferulate) purified from rice bran were investigated in a cholesterol oxidation system accelerated by 2,2′-azobis(2-methylpropionamidine) dihydrochloride. All components exhibited significant antioxidant activity in the inhibition of cholesterol oxidation. The highest antioxidant activity was found for 24-methylenecycloartanyl ferulate, and all three gamma-oryzanol components had activities higher than that of any of the four vitamin E components. Because the quantity of gamma-oryzanol is up to 10 times higher than that of vitamin E in rice bran, gamma-oryzanol may be a more important antioxidant of rice bran in the reduction of cholesterol oxidation than vitamin E, which has been considered to be the major antioxidant in rice bran. The antioxidant function of these components against cholesterol oxidation may contribute to the potential hypocholesterolemic property of rice bran.
Novel tocotrienols of rice bran suppress cholesterogenesis in hereditary hypercholesterolemic swine
Qureshi AA, Peterson DM, Hasler-Rapacz JO, Rapacz J.
J Nutr. 2001 Feb;131(2):223-30.
A tocotrienol-rich fraction (TRF(25)) and novel tocotrienols (d-P(21)-T3 and d-P(25)-T3) of rice bran significantly lowered serum and low density lipoprotein cholesterol levels in chickens. The present study evaluated the effects of novel tocotrienols on lipid metabolism in swine expressing hereditary hypercholesterolemia. Fifteen 4-mo-old genetically hypercholesterolemic swine were divided into five groups (n = 3). Four groups were fed a corn-soybean control diet, supplemented with 50 microg of either TRF(25), gamma-tocotrienol, d-P(21)-T3 or d-P(25)-T3 per g for 6 wk. Group 5 was fed the control diet for 6 wk and served as a control. After 6 wk, serum total cholesterol was reduced 32-38%, low density lipoprotein cholesterol was reduced 35-43%, apolipoprotein B was reduced 20-28%, platelet factor 4 was reduced 12-24%, thromboxane B(2) was reduced 11-18%, glucose was reduced 22-25% (P<0.01), triglycerides were reduced 15-19% and glucagon was reduced 11-17% (P<0.05) in the treatment groups relative to the control. Insulin was 100% greater (P<0.01) in the treatment groups than in the control group. Preliminary data (n = 1) indicated that hepatic activity of the 3-hydroxy-3-methylglutaryl-coenzyme A reductase was lower in the treatment groups, and cholesterol 7alpha-hydroxylase activity was unaffected. Cholesterol and fatty acid levels in various tissues were lower in the treatment groups than in control. After being fed the tocotrienol-supplemented diets, two swine in each group were transferred to the control diet for 10 wk. The lower concentrations of serum lipids in these four treatment groups persisted for 10 wk. This persistent effect may have resulted from the high tocotrienol levels in blood of the treatment groups, suggesting that the conversion of tocotrienols to tocopherols may not be as rapid as was reported in chickens and humans.
Nonantioxidant functions of alpha-tocopherol in smooth muscle cells
Azzi A, Breyer I, Feher M, Ricciarelli R, Stocker A, Zimmer S, Zingg J.
J Nutr. 2001 Feb;131(2):378S-81S.
Most tocopherols and tocotrienols, with the exception of alpha-tocopherol, are not retained by humans. This suggests that alpha-tocopherol is recognized uniquely; therefore, it may exert an exclusive function. alpha-Tocopherol possesses distinct properties that are independent of its prooxidant, antioxidant or radical-scavenging ability. alpha-Tocopherol specifically inhibits protein kinase C, the growth of certain cells and the transcription of the CD36 and collagenase genes. Activation events have also been seen on the protein phosphatase 2A (PP(2)A) and on the expression of other genes (alpha-tropomyosin and connective tissue growth factor). Neither ss-tocopherol nor probucol possessed the same specialty functions as alpha-tocopherol. Recently, we isolated a new ubiquitous cytosolic alpha-tocopherol binding protein (TAP). Its motifs suggest that it is a member of the hydrophobic ligand-binding protein family (CRAL-TRIO). TAP may also be involved in the regulation of cellular alpha-tocopherol concentration and alpha-tocopherol-mediated signaling.
Molecular aspects of α-tocotrienol antioxidant action and cell signalling
Lester Packer,Stefan U. Weber and Gerald Rimbach
J Nutr. 2001 Feb;131(2):369S-73S.
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.
Studies of LDL oxidation following alpha-, gamma-, or delta-tocotrienyl acetate supplementation of hypercholesterolemic humans
O'Byrne D, Grundy S, Packer L, Devaraj S, Baldenius K, Hoppe PP, Kraemer K, Jialal I, Traber MG.
Free Radic Biol Med. 2000 Nov 1;29(9):834-45.
In vitro tocotrienols (T3s) have potent vitamin E antioxidant activity, but unlike tocopherols can inhibit cholesterol synthesis by suppressing 3-hydroxy-3-methyl-glutarylCoA (HMG-CoA) reductase. Because hypercholesterolemia is a major risk factor for coronary artery disease and oxidative modification of low-density lipoprotein (LDL) may be involved in atherogenesis, we investigated whether daily supplements of placebo, or alpha-, gamma-, or delta- (alpha-, gamma-, or delta-) tocotrienyl acetates would alter serum cholesterol or LDL oxidative resistance in hypercholesterolemics in a double-blind placebo controlled study. Subjects were randomly assigned to receive placebo (n = 13), alpha- (n = 13), gamma- (n = 12), or delta- (n = 13) tocotrienyl acetate supplements (250 mg/d). All subjects followed a low-fat diet for 4 weeks, then took supplements with dinner for the following 8 weeks while still continuing diet restrictions. Plasma alpha- and gamma-tocopherols were unchanged by supplementation. Plasma T3s were undetectable initially and always in the placebo group. Following supplementation in the respective groups plasma concentrations were: alpha-T3 0.98 +/- 0.80 micromol/l, gamma-T3 0.54 +/- 0.45 micromol/l, and delta-T3 0.09 +/- 0.07 micromol/l. Alpha-T3 increased in vitro LDL oxidative resistance (+22%, p <.001) and decreased its rate of oxidation (p <. 01). Neither serum or LDL cholesterol nor apolipoprotein B were significantly decreased by tocotrienyl acetate supplements. This study demonstrates that: (i) tocotrienyl acetate supplements are hydrolyzed, absorbed, and detectable in human plasma; (ii) tocotrienyl acetate supplements do not lower cholesterol in hypercholesterolemic subjects on low-fat diets; and (iii) alpha-T3 may be potent in decreasing LDL oxidizability.
Oxidative damage to mitochondria in normal and cancer tissues, and its modulation
Kamat JP, Devasagayam TP.
Toxicology. 2000 Nov 30;155(1-3):73-82.
Cellular damage induced by reactive oxygen species (ROS) in normal tissues has been implicated in the etiology of several human ailments. Among the subcellular organelles, damage to mitochondria is considered crucial and can lead to cytotoxicity and cell death. However, the same damage, if it is selectively induced in cancer tissues can lead to its cure. Hence analyzing the mechanisms of such damage and its modulation may result in better prevention or cure. Using mitochondria derived from rat brain/liver as well as sarcoma 180 ascites cells, we have examined the mechanisms of damage to lipid, as assessed by different products of lipid peroxidation and to proteins, as determined by loss of enzyme activity and protein oxidation. Mechanisms involved, in terms of scavenging of ROS have been determined using pulse radiolysis for hydroxyl radical and histidine destruction assay for singlet oxygen. Various ROS were generated using gamma-radiation, photosensitization etc. under different conditions. Some novel porphyrins, with potential uses in photodynamic therapy also were used as photosensitizers. Our study shows that ROS can induce significant oxidative damage in mitochondria from both normal and tumor tissues and this can be inhibited by natural antioxidants like tocotrienols, nicotinamide and caffeine. Damage, on the other hand, can be enhanced by deuteration of the buffer and oxygenation. Our results hence demonstrated that mitochondria were sensitive to damage by ROS and its modulation may have potential uses in prevention of the disease in normal tissues; if damage can be selectively induced in tumor, it can lead to its regression.
Tocotrienols inhibit growth of ZR-75-1 breast cancer cells
Nesaretnam K, Dorasamy S, Darbre PD.
nt J Food Sci Nutr. 2000;51 Suppl:S95-103.
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.
Antiproliferative and apoptotic effects of tocopherols and tocotrienols on preneoplastic and neoplastic mouse mammary epithelial cells
McIntyre BS, Briski KP, Gapor A, Sylvester PW.
Proc Soc Exp Biol Med. 2000 Sep;224(4):292-301.
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.
Isolation and identification of novel tocotrienols from rice bran with hypocholesterolemic, antioxidant, and antitumor properties
Qureshi AA, Mo H, Packer L, Peterson DM.
J Agric Food Chem. 2000 Aug;48(8):3130-40
Two novel tocotrienols were isolated from stabilized and heated rice bran, apart from the known alpha-, beta-, gamma-, and delta-tocopherols andtocotrienols. These new tocotrienols were separated by HPLC, using a normal phase silica column. Their structures were determined by ultraviolet, infrared, nuclear magnetic resonance, circular dichroism, and high-resolution mass spectroscopies and established as desmethyl tocotrienol [3, 4-dihydro-2-methyl-2-(4,8,12-trimethyltrideca-3′(E),7′(E), 11′-trienyl)-2H-1-benzopyran-6-ol] and didesmethy tocotrienol [3, 4-dihydro-2-(4,8,12-trimethyltrideca-3′(E),7′(E), 11′-trienyl)-2H-1-benzopyran-6-ol]. These tocotrienols significantly lowered serum total and LDL cholesterol levels and inhibited HMG-CoA reductase activity in chickens. They had much greater in vitro antioxidant activities and greater suppression of B16 melanoma cell proliferation than alpha-tocopherol and known tocotrienols. Results indicated that the number and position of methyl substituents in tocotrienols affect their hypocholesterolemic, antioxidant, and antitumor properties.