In this study the effects of vitamin E deficiency and supplementation on bone calcification were determined using 4-month-old female Sprague-Dawley rats. The rats weighed between 180 and 200 g. The study was divided in three parts. In experiment I the rats were given normal rat chow (RC, control group), a vitamin E deficient (VED) diet or a 50% vitamin E deficient (50%VED) diet. In experiment 2 the rats were given VED supplemented with 30 mg/kg palm vitamin E (PVE30), 60 mg/kg palm vitamin E (PVE60) or 30 mg/kg pure alpha-tocopherol (ATF). In experiment 3 the rats were fed RC and given the same supplements as in experiment 2. The treatment lasted 8 months. Vitamin E derived from palm oil contained a mixture of ATF andtocotrienols. Rats on the VED and 50%VED diets had lower bone calcium content in the left femur compared to the RC group (91.6 +/- 13.3 mg and 118.3 +/- 26.0 mg cf 165.7 +/- 15.2 mg; P < 0.05) and L5 vertebra (28.3 +/- 4.0 mg and 39.5 +/- 6.2 mg compared with 51.4 +/- 5.8 mg; P < 0.05). Supplementing the VED group with PVE60 improved bone calcification in the left femur (133.6 +/- 5.0 mg compared with 91.6 +/- 13.3 mg; P < 0.05) and L5 vertebra (41.3 +/- 3.3 mg compared with 28.3 +/- 4.0 mg; P < 0.05) while supplementation with PVE30 improved bone calcium content in the L5 vertebra (35.6 +/- 3.1 mg compared with 28.3 +/- 4.0 mg; P < 0.05). However, supplementation with ATF did not change the lumbar and femoral bone calcium content compared to the VED group. Supplementing the RC group with PVE30, PVE60 or ATF did not cause any significant changes in bone calcium content. In conclusion, vitamin E deficiency impaired bone calcification. Supplementation with the higher dose of palm vitamin E improved bone calcium content, but supplementation with pure ATF alone did not. This effect may be attributed to the tocotrienol content of palm vitamin E. Therefore, tocotrienols play an important role in bone calcification.

Cardiovascular disease, in particular coronary artery disease (CAD), remains the most important cause of morbidity and mortality in developed countries and, in the near future, more so in the developing world. Atherosclerotic plaque formation is the underlying basis for CAD. Growth of the plaque leads to coronary stenosis, causing a progressive decrease in blood flow that results in angina pectoris. Acute myocardial infarction and unstable angina were recently recognised as related to plaque rupture, not progressive coronary stenosis. Acute thrombus formation causes an abrupt coronary occlusion. The characteristics of the fibrin cap, contents of the plaque, rheological factors and active inflammation within the plaque contribute to plaque rupture. Oxidative processes are important in plaque formation. Oxidized low density lipoproteins (LDL) but not unoxidized LDL is engulfed by resident intimal macrophages, transforming them into foam cells which develop into fatty streaks, the precursors of the atherosclerotic plaque. Inflammation is important both in plaque formation and rupture. Animal studies have shown that antioxidants reduce plaque formation and lead to plaque stabilisation. In humans, high intakes of antioxidants are associated with lower incidence of CAD, despite high serum cholesterol levels. This observation suggests a role for inflammation in CAD and that reducing inflammation using antioxidants may ameliorate these processes. Men and women with high intakes of vitamin E were found to have less CAD. Vitamin E supplementation was associated with a significant reduction in myocardial infarction and cardiovascular events in the incidence of recurrent myocardial infarction. In the hierarchy of evidence in evidence-based medicine, data from large placebo-controlled clinical trials is considered necessary. Results from various mega-trials have not shown benefits (nor adverse effects) conferred by vitamin E supplementation, suggesting that vitamin E has no role in the treatment of CAD. These results do not seem to confirm, at the clinical level, the effect of antioxidants against active inflammation during plaque rupture. However, a closer examination of these studies showed a number of limitations, rendering them inconclusive in addressing the role of vitamin E in CAD prevention and treatment. Further studies that specifically address the issue of vitamin E in the pathogenesis of atherosclerosis and in the treatment of CAD need be performed. These studies should use the more potent antioxidant property of alpha-tocotrienol vitamin E.

Vitamin E, the most important lipid-soluble antioxidant, was discovered at the University of California at Berkeley in 1922. Since its discovery, studies of the constituent tocopherols and tocotrienols have focused mainly on their antioxidant properties. In 1991 Angelo Azzi’s group (Boscoboinik et al. 1991a,b) first described non-antioxidant cell signalling functions for alpha-tocopherol, demonstrating that vitamin E regulates protein kinase C activity in smooth muscle cells. At the transcriptional level, alpha-tocopherol modulates the expression of the hepatic alpha-tocopherol transfer protein, as well as the expression of liver collagen alphal gene, collagenase gene and alpha-tropomyosin gene. Recently, a tocopherol-dependent transcription factor (tocopherol-associated protein) has been discovered. In cultured cells it has been demonstrated that vitamin E inhibits inflammation, cell adhesion, platelet aggregation and smooth muscle cell proliferation. Recent advances in molecular biology and genomic techniques have led to the discovery of novel vitamin E-sensitive genes and signal transduction pathways.

Vitamin C, tocopherols, and tocotrienols in berries of wild and cultivated sea buckthorn (Hippophaë rhamnoides L.) of different origins and harvesting dates were determined with HPLC. Wild berries of subsp. sinensis, native to China, contained 5-10 times more vitamin C in the juice fraction than the berries of subsp. rhamnoides from Europe and subsp. mongolica from Russia (4-13 vs 0.02-2 g/L juice). Genetic background and berry-harvesting date were two primary factors determining the vitamin C content in the berries. Crossing different subspecies influenced the vitamin C content to some extent. For bushes cultivated in southwest Finland, the best berry-harvesting date for high vitamin C content was the end of August. The seeds of subsp. sinensis contained less tocopherols and tocotrienols (average 130 mg/kg) compared with seeds of subsp. rhamnoides (average 290 mg/kg) and mongolica (average 250 mg/kg). The fruit flesh of sinensis berries had contents of tocopherols and tocotrienols 2-3 times higher than those found in the other two subspecies (120 mg/kg vs 40 mg/kg in rhamnoides and 50 mg/kg in mongolica). The fresh whole berries of subsp. sinensis were clearly the best source of total tocopherols and tocotrienols. The total content of tocopherols and tocotrienols in the soft parts of the berries reached the maximum level around early- to mid-September, whereas the content in seeds continued to increase until the end of November. The excellent combination of the highest content of vitamin C and tocopherols and tocotrienols makes the berries of subsp. sinensis an optimal raw material for nutritional investigation as a candidate for functional foods with special antioxidative properties.

We have previously shown that alpha-tocotrienol (alpha-T3), a vitamin E analogue and HMG CoA reductase (HMGR) inhibitor, markedly inhibited monocyte-endothelial cell adhesion, a process that was reversed with the addition of mevalonate intermediates involved in protein prenylation. Since delta-T3 and gamma-T3 possess greater HMGR inhibition than alpha-T3, we postulated that these analogues might have a greater effect on protein prenylation, and thus on monocyte adhesion and endothelial adhesion molecule expression in comparison to alpha-T3. Hence, we pursued to investigate the effect of various analogues of tocotrienol (alpha, gamma, delta) on monocytic cell adhesion and expression of adhesion molecules using a human umbilical vein endothelial cell-line, EA.hy926, as the model system. Relative to alpha-T3, delta-T3 displayed a more profound inhibitory effect on monocytic cell adherence using a 15 micromol/L concentration within 24 h (delta: 42 +/- 5%; alpha: 26 +/- 8% vs. control). This inhibitory action was reversed by co-incubation with farnesol and geranylgeraniol, suggesting a role for prenylated proteins in the regulation of monocyte adhesion. To further evaluate the effect of tocotrienols on the vascular endothelium, we measured the surface expression of adhesion molecules. Compared to alpha-T3, delta-T3 markedly inhibited the expression of VCAM-1 (delta: 57 +/- 6%; alpha: 37 +/- 10% vs. control) and E-selection (delta: 36 +/- 3%; alpha: 18 +/- 6% vs. control) in TNF-alpha activated endothelial cells. The above result suggests that delta-T3 is a potent and effective agent for the reduction of cellular adhesion molecule expression and monocytic cell adherence.