Vitamins represent precursor molecules for the majority of cofactors essential for various enzyme activities. Is the expression of proteins that require a specific cofactor for their function correlated to its availability? Examples are presented and discussed of the involvement of vitamin-derived cofactors in the regulatory mechanisms controlling gene expression in bacteria and eukaryotes at the transcriptional and trans-lational level.

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Healthy individual were given 2g of vitamin C per day for 2 months. Whole blood iron, ascorbic acid, hemoglobin, and serum ceruloplasmin were determined at the beginning, and 1 or 2 months after the start of the experiment. The concentration of ascorbic acid was observed to increase significantly in the blood, while blood iron, hemoglo-bin, and serum ceruloplasmin levels significantly increased at the end of the 1st month, but decreased to control levels at the end of the 2nd month. Male albino guinea pigs were administered 8, 180, and 360mg of vitamin C per day for 2 months. Liver ferritin iron, liver copper, serum copper, and serum ceruloplasmin levels significantly decreased, but there was no significant change in hemosiderin iron while blood ascorbic acid signifi-cantly increased at the end of the 2 month period. There was no significant change in serum iron and hematocrit levels. These results suggest that vitamin C has an antagonistic effect on copper metabolism in guinea pigs but not in humans either on copper or iron metabolisms.

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Male guinea pigs were fed during 5 weeks with diets differing only in vitamin C content: low (33 mg/kg diet), medium (660mg/kg), and high (13, 200mg/kg). Heart vitamin C was strongly depend-ent on dietary vitamin C and heart vitamin E showed a trend to increase as a function of the vitamin C level in the diet. The low vitamin C diet decreased body weight gain, food intake, and heart malondialdehyde without changing lipid peroxidation, whereas the high vitamin C in-creased oxidized glutathione and glutathione peroxidase and decreased body growth. A tendency to show higher levels of all the first-line antioxidants reduced glutathione, uric acid, superoxide dismutase, cata-lase, and glutathione peroxidase at extreme (high or low) dietary levels of vitamin C was observed. The guinea pig heart showed capacity for enzymatic but not for non-enzymatic in vitro lipid peroxidation. It is concluded that dietary vitamin C supplementation is able to increase the global antioxidant capacity of the heart tissue.

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Long-term administration of a β-carotene preparation de-rived from Dunaliella bardawil, a β-carotene-rich algae, was studied in healthy young male volunteers. The daily administration of 60mg of the β-carotene preparation (30mg of all-trans β-carotene and 30mg of 9-cis β-carotene) was performed and β-carotene concentrations were deter-mined in the plasma, red blood cells (RBC), platelets (PLT), and mononuclear cells (MN). The all-trans β-carotene level increased four-, two-, and threefold the baseline in plasma, PLT, and MN, respectively. Basal levels of 9-cis β-carotene in plasma, PLT, and MN were low and found as one-tenth, one-fifth, and one-fifth of all-trans β-carotene, which increased three-, two-, and 1.5-fold the baseline, respectively. Plasma and RBC α-tocopherol levels were not changed by the intake of β-carotene. No side effects or toxicities were documented in any of the subjects during the administration period. In conclusion, the bioavailability of β-carotene derived from Dunaliella bardawil was preferential for all-trans β-carotene, although a small amount of the 9-cis form was detected in the plasma and blood cells.

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Mice fed a Spirulina platensis diet showed increased numbers of splenic antibody-producing cells in the primary immune response to sheep red blood cells (SRBC). However, immunoglobulin G (IgG)-antibody production in the secondary immune response was hardly affect-ed. The percentage of phagocytic cells in peritoneal macrophages from the mice fed S. platensis diet, as well as the proliferation of spleen cells by either concanavalin A (Con A) or phytohemagglutinin (PHA) was significantly increased. Addition of a hot-water extract of S. platensis (SHW) to an in vitro culture of spleen cells markedly increased prolifera-tion of these cells, whereas culture of thymus cells was scarcely affected. The Spirulina extract also significantly enhanced interleukin-1 (IL-1) production from peritoneal macrophages. Addition to the in vitro spleen cell culture of SHW as well as the supernatant of macrophages stimulated with SHW resulted in enhancement of antibody production, that is, an increase of the number of PFC. These results suggest that Spirulina enhances the immune response, particularly the primary response, by stimulating macrophage functions, phagocytosis, and IL-1 production.

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In our previous study, globin was found to be an effective dietary source for increasing bone mineral density (BMD) and mechani-cal strength. In this study, the bioavailability of the globin preparation was examined to clarify the mechanism of increase in bone density and strength. Six-week-old Sprague-Dawley female rats were ovariectomized and were fed on a low Ca diet for 30 days to produce the experimental osteoporotic rats. Thereafter they were divided into two groups. The BMD and the mechanical strength of bone of the rat group, whose diet was supplemented with globin, were significantly higher than those of the control group. The levels of the serum calcitonin and the bone-type alkaline phosphatase (Alp) activity in serum and bone were also higher, and the tartrate-resistant acid phosphatase (Tr-Acp) activity in serum and bone was lower in the globin group. Moreover, the bone morphogenetic protein activity in bone in the globin group was found to be greater. From these results, it is concluded that alimentary globin is effective for the acceleration of bone formation and the prevention of bone resorption.

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The aim of this study was to investigate whether there is variation in blood pressure when we reapportion the percentage of total daily salt intake consumed at each of three regular meals. The study was conducted on seven clinically healthy normotensive female subjects who, in Stages LH and DH, consumed two-thirds of the normal daily salt intake (12 g/day) at lunchtime or at dinnertime, respectively. The total daily amounts of nutrients and dietary salt were similar in Stage-R (regulated salt intake), Stage-LH (a high-salt intake at lunchtime), and Stage-DH (a high-salt intake at dinnertime). The blood pressure response to the variation of sodium content in the meals was examined by means of noninvasive automated blood pressure monitoring (ABPM-630; readings every 15 or 30 min over 48h) and chronobiologic analysis. A significant shift of blood pressure circadian rhythm was observed in Stage-LH. Additionally, the 24-h mean level of blood pressure significantly increased when the prevailing salt intake was at lunch, and significantly decreased when the prevailing intake was at dinner. These opposite responses corroborate the view that the blood pressure susceptibility of human beings to salt intake varies during the day, showing its maximal expression at midday. Such a time-dependent sensitivity may be exploited for better nutritional prevention and treatment of arterial hypertension by reappor-tioning the salt intake so that two-thirds is consumed at dinner.

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Antioxidant biofactor: AOB® is a unique processed grain food. It is a yellow-green powder. It contains the following extracts: germ extracts, soybean, rice bran, tear grass, sesame, wheat, citron, green tea, green leaf extract, and malted rice. These materials were slowly roasted under a powdered oure at less than 60°C and fermented with Aspergillus oryzae over 3 days to transform each ingredient into low molecular weight substances. These conditions were different by each material, environmental humidity and temperature. It probably contains a variety of substances having antioxidant activity including flavonoids, α-tocopherol, vitamin C, and tannins. We investigated its antioxidative properties using electron spin resonance (ESR) and autoxidation of rat brain homogenates. The superoxide, hydroxyl radical, and the stable free radical, diphenyl p-picrylhydrazyl (DPPH) radical scavenging activity of AOB® was investigated using ESR spectrometry. In an in vitro study, a suspension of AOB® was added directly to a superoxide generating system (hypoxanthine-xanthine oxidase; HX/XO) and investigated using 5, 5-dimethyl-l-pyrroline-N oxide (DMPO) as a spin trapping agent. At final concentrations of 0.01, 0.05, and 0.1 mg/ml, AOB® dose-dependent scav-enging activity was observed as 0.103, 0.619, and 1.369 U/ml, respectively. A concentration of 1.0mg/ml completely scavenged DMPO-OOH signals; 1.0 mg/ml of AOB® inhibited the DMPO-OH signal generated by Fenton's reaction, but its inhibitory effect was not competitive, and was inhibition of the Fenton's reaction. 1.0, 3.0, and 5.0 mg/ml of AOB® were significantly inhibited the DPPH radical. In an in vivo study, rats were fed AOB® orally at doses of 1 or 5g/day for 24 h or for 3 days and the superoxide scavenging activity was measured in plasma. With the admin-istration of 1g/day for 3 days, the superoxide scavenging activity was about 1.8 times that of the control group fed a basal diet; 1.5 times the control with 5 g/day for 1 day, and 2.6 times the control with 5 g/day for 3 days, all of which represented significant increases in superoxide scav-enging activity. AOB® strongly inhibited the autoxidation of rat brain homogenates in vitro in a dose-dependent manner. However, each ingre-dient before roast and fermentation little inhibited lipid peroxidation. Roasting and fermentation with A. oryzae way be important to transform each ingredient into low molecular weight substances. Therefore, it was suggested that AOB® possesses strong antioxidant and free radical scav-enging activities.

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We studied the coagulative and fibrinolytic activity in in-trinsic or extrinsic hyperlipidemia using 4-week-old male Wistar rats. Intrinsic hyperlipidemia was induced by a cholesterol-free high-fructose diet (HFD) and extrinsic hyperlipidemia, by a high-cholesterol diet (HCD) for 14 days. In intrinsic hyperlipidemic rats fed on the HFD, serum lipids were significantly increased as compared with the levels in control rats fed on a standard diet. An apparent increase in plasma fibrinogen level and coagulant factor XIII activity was also observed in HFD rats. In extrinsic hyperlipidemic rats fed on the HCD, significant increases in plasma fibrinogen level compared with that of control rats were found with the increases in serum lipids. Activities of antithrombin III and α₂-plasmin inhibitor in HFD-fed rats significantly increased compared with those of control and HFD rats. There was a significant positive correlation between plasma fibrinogen and serum total cholester-ol, free cholesterol, or phospholipid in diet-induced hyperlipidemia (p< 0.01). Because of the increase in coagulant XIII activity in HFD-fed rats and the increase in α₂-plasmin inhibitor activity in HCD-fed rats, both diet-induced hyperlipidemic rats were shown to have enhanced coagula-tive activity compared with the control rats. These results suggest that the HFD as well as the HCD causes a pre-hypercoagulative state due to the increase in plasma fibrinogen level and activities in other coagulative and fibrinolytic factors.

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An improved agar plate method of biotin bioassay using Lactobacillus plantarum ATCC 8014 and bromocresol purple was estab-lished to determine biotin levels in human serum and urine. Samples were treated with 4.5 N H₂SO₄ to liberate free biotin, autoclaved for 1 h and neutralized by 4.5N NaOH, then 10μl was added to wells in each plate. The biotin levels were measured in 190 serum and 59 urine samples, and the means were 2.7±0.53ng/ml and 12.4±5.56ng/mg of creatinine, re-spectively. The intra-assay coefficient varience (CV) were 3.2 (n=20) and 1.3% (n=23), respectively. The recovery of biotin added (10ng/ml) to serum was 110.7%, and to urine was 99.6%. These findings suggest that this assay is sufficiently accurate and reproducible for routine use in the clinical laboratory. The excretion of orally administered biotin was also demonstrated by the method.

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The undigested fraction of soybean protein (UDF) exerts a markedly greater hypocholesterolemic effect than soybean protein itself in rats. The present study was undertaken to confirm the effect in hamsters, a more appropriate animal model for human cholesterol metabolism. Hamsters were given diets containing UDF at a nitrogen level equivalent to the 20% casein diet. Dietary fats, at the 10% level, were perilla oil and safflower oil. There was apparently no increase in the serum and liver cholesterol levels in both groups of animals cholesterol-enriched diets that had been fed for 38 days. Fecal excretion of neutral and acidic steroid tended to be higher in the perilla oil group than in the safflower oil group. The perilla oil group significantly increased 20: 5n-3 in liver phosphatidyl-choline and phosphatidylethanolamine accompanying a decrease in 20:4n-6. Such changes were not so evident in liver phosphatidylinositol. The production of leukotriene B₄ and the concentration of prostaglandin E₂ in the spleen were higher in the safflower oil group than in the perilla oil group. Thus, the hypocholesterolemic effect of the undigested fraction of soybean protein was apparently reproduced even in hamsters. Dietary fat-induced changes in lipid parameters in hamsters resembled those observed in rats.

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Phosphatidylgenipin, synthesized via the transphospha-tidylation reaction of 1, 2-dipalmitoyl-3-sn-phosphatidylcholine to genipin by phospholipase D, was found to react with L-phenylalanine in chloro-form and gave a clear blue solution. This blue solution was also formed in following organic solvents: ethanol, ethyl acetate, diethyl ether, ben-zene, and hexane. However, genipin and L-phenylalanine did not give any colored product under the same conditions. The blue pigment resulted from phosphatidylgenipin and L-phenylalanine showed λ_max at 615nm in chloroform, and had a similar blue color to an aqueous solution of the natural blue pigment “gardenia blue.” This is an example for the preparation of a hydrophobic pigment from a phosphatidyl derivative of a water-soluble compound.

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