Tocochromanols are potent lipid-soluble antioxidants and essential nutrients for human health. Genetic engineering techniques were used to develop soybeans with enhanced vitamin E levels, including tocotrienols, which are not found in soybean. The gene encoding rice homogentisate geranylgeranyl transferase (HGGT) was overexpressed in soybeans using seed-specific and constitutive promoters. The association between abundance of vitamin E isomers and antioxidant activity was investigated during seed germination. With the exception of β-tocotrienol, all vitamin E isomers were detected in germinating seeds expressing OsHGGT. The antioxidant properties of germinating seed extracts were determined using 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2′-azinobis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) radicals and lipid peroxidation (TBARS). Compared with intact wild-type seeds, transgenic seeds showed increases in radical scavenging of 5.4-17 and 23.2-35.3% in the DPPH and ABTS assays, respectively. Furthermore, the lipid peroxidation levels were 2.0-4.5-fold lower in germinating seeds from transgenic lines than in wild-type seeds. Therefore, it appears that the antioxidant potential of transgenic oil-producing plants such as soybean, sunflower, and corn may be enhanced by overexpressing OsHGGT during seed germination.

The biosynthesis of the tocotrienol and tocopherol forms of vitamin E is initiated by prenylation of homogentisate. Geranylgeranyl diphosphate (GGDP) is the prenyl donor for tocotrienol synthesis, whereas phytyl diphosphate (PDP) is the prenyl donor for tocopherol synthesis. We have previously shown that tocotrienol synthesis is initiated in monocot seeds by homogentisate geranylgeranyl transferase (HGGT). This enzyme is related to homogentisate phytyltransferase (HPT), which catalyzes the prenylation step in tocopherol synthesis. Here we show that monocot HGGT is localized in the plastid and expressed primarily in seed endosperm. Despite the close structural relationship of monocot HGGT and HPT, these enzymes were found to have distinct substrate specificities. Barley (Hordeum vulgare cv. Morex) HGGT expressed in insect cells was six times more active with GGDP than with PDP, whereas the Arabidopsis HPT was nine times more active with PDP than with GGDP. However, only small differences were detected in the apparent Km values of barley HGGT for GGDP and PDP. Consistent with its in vitro substrate properties, barley HGGT generated a mixture of tocotrienols and tocopherols when expressed in the vitamin E-null vte2-1 mutant lacking a functional HPT. Relative levels of tocotrienols and tocopherols produced in vte2-1 differed between organs and growth stages, reflective of the composition of plastidic pools of GGDP and PDP. In addition, HGGT was able to functionally substitute for HPT to rescue vte2-1-associated phenotypes, including reduced seed viability and increased fatty acid oxidation of seed lipids. Overall, we show that monocot HGGT is biochemically distinct from HPT, but can replace HPT in important vitamin E-related physiological processes.

BACKGROUND/OBJECTIVES: Vitamin E is an essential fat-soluble vitamin that has been shown to induce favorable effects on animal and human immune systems. The objective of this study was to assess the effects of tocotrienol-rich fraction (TRF) supplementation on immune response following tetanus toxoid (TT) vaccine challenge in healthy female volunteers.

SUBJECTS/METHODS: In this double-blinded, placebo-controlled clinical trial, participants were randomly assigned to receive either placebo (control group) or 400 mg of TRF (study group) supplementation daily. Over the 2-month period of the study, volunteers were asked to attend three clinical sessions (that is, on days 0, 28 and 56) and blood samples were obtained from the volunteers during the follow-up. On day 28, all volunteers were also vaccinated with the TT vaccine (20 Lf) intramuscularly.

RESULTS:The results from the clinical trial showed that TRF supplementation significantly increased the total vitamin E level in the plasma of the TRF-supplemented volunteers compared with the placebo group, indicating overall compliance. Volunteers supplemented with TRF showed a significantly (P < 0.05) enhanced production of interferon-γ and interleukin (IL)-4 by the mitogen or TT-stimulated leukocytes compared with the control group. Volunteers from the TRF group produced significantly (P < 0.05) lower amounts of IL-6 compared with the placebo group. Anti-TT IgG production was also significantly (P < 0.05) augmented in the TRF-supplemented group compared with the placebo group.

CONCLUSIONS: We conclude that TRF has immunostimulatory effects and potential clinical benefits to enhance immune response to vaccines.

AIMS: Vitamin E is an antioxidant that may protect bone against oxidative stress-induced osteoporosis. This in vitro study was conducted to determine the protective effects of a-tocopherol and γ-tocotrienol on osteoblasts, the bone forming cells, against oxidative stress.

MATERIALS AND METHODS: Toxicity tests were carried out on hydrogen peroxide (H(2)O(2)), a-tocopherol and γ-tocotrienol and their inhibitory concentration 50 (IC(50)) on osteoblasts were determined if any. Osteoblast cultures were then pretreated with different concentrations of a-tocopherol or γ-tocotrienol for 24 hours before incubated with the IC50 of H(2)O(2) for 2 hours. Cell viability was determined by using MTS assay to compare the protective effects of both vitamin E on osteoblast exposed to H(2)O(2).

RESULTS: The IC(50) after 2 hours and 24 hours incubation time for H(2)O(2) were 490 µM and 280 µM respectively. γ-Tocotrienol was found to be toxic to osteoblasts with the IC(50) of 290 µM after 24 hours incubation time while a-tocopherol was not toxic to osteoblasts at any doses. However, γ-tocotrienol was able to protect osteoblasts from H(2)O(2) toxicity at low concentration (1 µM), whereras a-tocopherol was not able to offer protection against H2O2 toxicity.

CONCLUSIONS: γ-tocotrienol was found to be toxic to osteoblasts at high concentrations but at much lower concentration, it has better antioxidant activity than a-tocopherol to protect osteoblasts from oxidative stress.