Recommended dietary allowance for vitamin C in the United States is also applicable to a population of young Japanese women

The recommended dietary allowance (RDA) for ascorbic acid (AA) in Canada and the United States has been set for several years at 75 mg/day for women 19-30 years old. Recently this level was questioned, and an increase to 90 mg/day was suggested. For Japanese women in the same age group, we found that the RDA for AA is currently 100 mg/day. Our goal was to determine which RDA is sufficient for maintaining a serum concentration of AA in young Japanese women above the lower reference limit of 7.0 mg/L. We measured serum AA concentrations by an ascorbate oxidase method in 176 healthy Japanese women (19-26 years old). We also performed an ROC analysis to estimate the optimal cutoff value for oral dosage to distinguish individuals with hypovitaminosis-C (<7.0 mg/L) from those with a normal serum AA. We evaluated the Japanese RDA using the 75 or 90 mg/day U.S. RDA and the weight ratio between Japanese and U.S. women, and discovered that the RDA value ranged between 66 and 79 mg/day. From the ROC analysis, we found that the optimal daily dosage of AA is approximately 75 mg/day. This value gave the highest efficiency, sensitivity, negative predictive value, and positive likelihood ratio, and the lowest negative likelihood ratio. Therefore, an RDA of 100 mg/day may be unnecessarily high for young Japanese women.     

Alteration in the glutathione,glutathione peroxidase,superoxide dismutase and lipid peroxidation by ascorbic acid in the skin of mice exposed to fractionated gamma radiation

BACKGROUND: In spite of the immense therapeutic gains produced by the fractionated irradiation (IR) regimen, radiation burden on the skin increases significantly. Protection of skin might enable use of higher radiation doses for better therapeutic gains. Ascorbic acid (AA), an essential ingredient of the human diet, is known to be a free radical scavenger and radioprotective agent. This study was undertaken to evaluate the effect of ascorbic acid on the radiation-induced changes in the status of glutathione (GSH), glutathione peroxidase (GPx), superoxide dismutase (SOD) and lipid peroxidation (LPx) in the skin of mice exposed to 10, 16 and 20 Gy of fractionated gamma radiation. METHODS: One group of the animals was administered daily with double distilled water (DDW), while the other group received 250 mg/kg b. wt. of ascorbic acid once daily, consecutively for 5, 8 or 10 days, before hemibody (below rib cage) exposure to 2 Gy/day of gamma-rays. Skin biopsies from both the groups were collected for the biochemical estimations. RESULTS: The irradiation of animals resulted in a dose-dependent decline in the activities of superoxide dismutase, glutathione peroxidase and glutathione contents. Ascorbic acid pretreatment resulted in a significant increase in the activities of both the enzymes and glutathione in the irradiated mouse skin. Normal concentrations of glutathione could not be restored even by day 6 post-irradiation. Conversely, lipid peroxidation increased in a dose-dependent manner in both the groups reaching a peak concentration by 3 h post-irradiation, while the ascorbic acid pretreatment inhibited the radiation-induced increase in lipid peroxidation. CONCLUSIONS: The ascorbic acid treatment arrested the decline in the activities of superoxide dismutase and glutathione peroxidase, glutathione contents and inhibited the radiation-induced lipid peroxidation in the skin of mice exposed to different doses of fractionated gamma radiation.     

Mitochondrial redox studies of oxidative stress in kidneys from diabetic mice

Chronic hyperglycemia during diabetes leads to increased production of reactive oxygen species (ROS) and increased oxidative stress (OS). Here we investigated whether changes in the metabolic state can be used as a marker of OS progression in kidneys. We examined redox states of kidneys from diabetic mice, Akita(/+) and Akita(/+);TSP1(-/-) mice (Akita mice lacking thrombospondin-1, TSP1) with increasing duration of diabetes. OS as measured by mitochondrial redox ratio (NADH/FAD) was detectable shortly after the onset of diabetes and further increased with the duration of diabetes. Thus, cryo fluorescence redox imaging was used as a quantitative marker of OS progression in kidneys from diabetic mice and demonstrated that alterations in the oxidative state of kidneys occur during the early stages of diabetes.     

S-adenosylmethionine prevents hepatic tocopherol depletion in carbon tetrachloride-injured rats

In various experimental models, S-adenosylmethionine (SAMe) has been shown to reduce liver injury by preventing depletion of glutathione, one of the antioxidant systems that plays a critical role in defence against oxidative stress. On the other hand, alpha-tocopherol may be decreased in liver diseases, and treatment with this vitamin reduces liver injury in CCl(4)-treated rats. Since there is a close relationship among the different antioxidant systems (mainly glutathione, alpha-tocopherol and ascorbic acid), we have assessed whether, as well as restoring hepatic glutathione content, SAMe has any effect on liver alpha-tocopherol and ascorbic acid levels in CCl(4)-injured rats. Four groups of seven male Wistar rats treated for 9 weeks were studied: rats induced to cirrhosis with CCl(4), rats induced to cirrhosis plus SAMe administration (10 mg x kg(-1) x day(-1)) and their respective controls. Liver samples were obtained for measuring levels of glutathione, alpha-tocopherol, ascorbic acid and thiobarbituric acid-reactive substances (TBARS), and hydroxyproline concentration as an index of collagen content. The hydroxyproline content was higher in CCl(4)-injured rats than in the control group (4.4+/-1.8 and 1.1+/-0.3 micromol/g respectively; P<0.05). In CCl(4)-injured rats, SAMe administration decreased collagen content (2.7+/-1.0 microl/g; P<0.05) and TBARS, and corrected glutathione depletion. alpha-Tocopherol was significantly lower in CCl(4)-injured rats than in controls (17.3+/-4.9 and 23.0+/-4.0 micromol/g respectively; P<0.05). By contrast, alpha-tocopherol levels were similar (23.8+/-5.1 micromol/g) in CCl(4)-injured rats receiving SAMe and in controls. In CCl(4)-injured rats, liver ascorbic acid was decreased in comparison with controls (4.9+/-1.8 and 8.2+/-1.0 micromol/g respectively; P<0.05), levels which were not replenished by SAMe (4.6+/-0.4 micromol/g). In conclusion, SAMe not only decreases fibrosis and protects against hepatic glutathione depletion, but has a further antioxidant effect of preventing alpha-tocopherol depletion in CCl(4)-injured rats.     

N-acetylcysteine improves the hemodynamics and oxidative stress in hypoxic newborn pigs reoxygenated with 100% oxygen

Neonatal asphyxia may lead to cardiac and renal complications perhaps mediated by oxygen free radicals. Using a model of neonatal hypoxia-reoxygenation, we tested the hypothesis that N-acetylcysteine (NAC) would improve cardiac function and renal blood flow. Eighteen piglets (aged 1-4 days old, weighing 1.4-2.2 kg) were anesthetized and acutely instrumented for continuous monitoring of pulmonary and renal artery flow (cardiac index [CI] and renal artery flow index [RAFI], respectively) and mean blood pressure. Alveolar hypoxia was induced for 2 h, followed by resuscitation with 100% oxygen for 1 h and 21% oxygen for 3 h. Animals were randomized to sham-operated, hypoxic control, and NAC treatment (i.v. bolus of 150 mg/kg given at 10 min of reoxygenation followed by 100 mg/kg per h infusion) groups. Myocardial and renal tissue glutathione content and lipid hydroperoxide levels were assayed, and histology was examined. After 2 h of hypoxia, all animals were acidotic (pH 6.96 +/- 0.04) and in cardiogenic shock with depressed renal blood flow. Upon reoxygenation, CI and RAFI increased but gradually deteriorated later. The NAC treatment prevented the decreased CI, stroke volume, mean blood pressure, systemic oxygen delivery, RAFI, and renal oxygen delivery at 2 to 4 h of reoxygenation observed in hypoxic controls (versus shams, all P < 0.05). The myocardial and renal tissue glutathione content was significantly higher in the NAC treatment group (versus controls). The CI and RAFI at 4 h of reoxygenation correlated with the tissue glutathione redox ratio (r = 0.5 and 0.6, respectively, P < 0.05). There were no significant differences in heart rate, pulmonary artery pressure, systemic oxygen uptake, and tissue lipid hydroperoxide levels between groups. No histologic injury was found in the heart or kidney. In this porcine model of neonatal hypoxia and 100% reoxygenation, NAC improved cardiac function and renal perfusion, with improved tissue glutathione content.     

Selenium diet-supplementation improves cocaine-induced myocardial oxidative stress and prevents cardiac dysfunction in rats

Chronic cocaine abuse causes cardiac dysfunction and induces oxidative stress. The goal of this study was to evaluate whether an enhanced antioxidant pool, induced by the administration of selenium, may prevent the myocardial dysfunction induced by cocaine. Cocaine was administered for 7 days (15 mg/kg/day, i.p.) to rats pretreated for 4 weeks with selenium (1.16 mg/L/day, p.o.). Cardiac function was evaluated by cardiac index and left ventricular (LV) fractional shortening (FS) measured by echocardiography. The redox ratio and enzymatic activities of glutathione peroxidase (GPX) and superoxide dismutase (SOD) were measured in the LV myocardium. Cocaine administration induced a cardiac dysfunction, as evidenced by a decrease in cardiac index and LV FS as well as by an increase in LV diameters. Moreover, antioxidant markers and redox ratio were altered in rats after cocaine exposure. Selenite supplementation induced a significant limitation of cardiac index and FS alterations observed after cocaine administration. This improvement in cardiac function was associated with a redox ratio recovery while SOD and GPX activities remained unchanged. Thus, selenite reversed both the oxidative stress and the contractile dysfunction induced by cocaine administration. These results suggest a major role of oxidative stress in the cocaine-induced cardiotoxicity.     

Plasma cysteine and glutathione are independent markers of postmethionine load endothelial dysfunction

OBJECTIVES: Oxidative stress caused by acute hyperhomocysteinemia impairs endothelial function in human arteries. We sought to identify markers of endothelial dysfunction during methionine-induced hyperhomocysteinemia. DESIGN AND METHODS: 35 subjects underwent flow-mediated dilation (FMD) of the brachial artery by high-resolution ultrasonography and fasting blood samples before and 3 h postmethionine load (PML). Clinical, conventional biochemical, and redox status (plasma total and reduced homocysteine, glutathione, cysteine, cysteinylglycine, ascorbic acid, alpha-tocopherol, free malondialdehyde, blood glutathione) data were sequentially entered into an univariate and multivariate stepwise linear regression analysis to evaluate their relation with the dependent variable FMD. RESULTS: Median [interquartile range] FMD decreased from 4.1% [2.8-6.3] to 3.2% [0.7-4.3] PML (P=0.02). At the multivariate analysis PML total cysteine (beta=-0.008, P=0.002) and glutathione (beta=0.21, P=0.005) were the only independent variables associated with FMD after methionine, adjusted for baseline FMD. CONCLUSIONS: Elevated plasma total cysteine and decreased plasma total glutathione levels were associated with abnormal FMD PML. Cysteine and glutathione are stronger markers of endothelial dysfunction than clinical and all other biochemical variables explored.     

Xanthine oxidase activity and blood glutathione redox ratio in infants and children with septic shock syndrome

OBJECTIVES: The possible role of xanthine oxidase (XO) activation in the signal transduction process during the septic shock syndrome was examined. The XO activity index after caffeine intake was assessed simultaneously with the blood glutathione redox ratio, a known parameter of oxidative stress. DESIGN AND SETTING: An investigational clinical study in a nine-bed pediatric intensive care unit. PATIENTS: Critically ill infants and children (n = 34) with systemic inflammatory response syndrome following infection, trauma or major surgery. Biochemical investigations (n = 54) were performed at various stages of the shock syndrome, characterized by pediatric risk of mortality and organ dysmetabolic scores. Controls consisted of 30 healthy children. MEASUREMENTS AND RESULTS: The in vivo XO activity index was measured as the urinary ratio of two metabolites of caffeine: 1-methyluric acid and 1-methylxanthine. The blood concentrations of oxidized (GSSG) and reduced glutathione (GSH) were determined. The XO activity index and redox ratio GSSG/GSH were highly increased in patients in shock dominated by the clinical symptoms of a proinflammatory response. A significantly lower XO activity index was found with an increased GSSG/ GSH in patients whose stage of shock was characteristic of an excessive anti-inflammatory response. The XO activity index and GSSG/ GSH were correlated closely with each other (r = 0.624, n = 54; p < 0.001), and were also related to the daily severity scores. CONCLUSION: Potent and simultaneous activation of the two redox systems strongly indicates a definite role of free radicals from XO in the overspill of the acute proinflammatory reaction of the shock syndrome, followed by a significant downregulation.     

L-2-Oxothiazolidine-4-carboxylic acid reverses endothelial dysfunction in patients with coronary artery disease.

The effective action of endothelium-derived nitric oxide (EDNO) is impaired in patients with atherosclerosis. This impairment has been attributed in part to increased vascular oxidative stress. EDNO action is improved by administration of ascorbic acid, a water-soluble antioxidant. Ascorbic acid is a potent free-radical scavenger in plasma, and also regulates intracellular redox state in part by sparing cellular glutathione. We specifically investigated the role of intracellular redox state in EDNO action by examining the effect of L-2-oxo-4-thiazolidine carboxylate (OTC) on EDNO-dependent, flow-mediated dilation in a randomized double-blind placebo-controlled study of patients with angiographically proven coronary artery disease. OTC augments intracellular glutathione by providing substrate cysteine for glutathione synthesis. Brachial artery flow-mediated dilation was examined with high-resolution ultrasound before and after oral administration of 4.5 g of OTC or placebo in 48 subjects with angiographically documented coronary artery disease. Placebo treatment produced no change in flow-mediated dilation (7.0+/-3.9% vs. 7.2+/-3.7%), whereas OTC treatment was associated with a significant improvement in flow-mediated dilation (6.6+/-4.4% vs. 11.0+/-6.3%; P = 0.005). OTC had no effect on arterial dilation to nitroglycerin, systemic blood pressure, heart rate, or reactive hyperemia. These data suggest that augmenting cellular glutathione levels improves EDNO action in human atherosclerosis. Cellular redox state may be an important regulator of EDNO action, and is a potential target for therapy in patients with coronary artery disease.     

Thiol and redox reactive agents exert different effects on glutathione metabolism in HeLa cell cultures.

Glutathione protects cells against oxidative damage, free radical damage and other types of toxicity. The aim of the present study was to investigate the impact on glutathione metabolism exerted by different thiol or redox reactive agents. Intracellular concentrations of glutathione in HeLa cell cultures were lowered after addition of agents mainly exerting oxidative stress (homocysteine and hydrogen peroxide), whereas thiol reactive oxidative agents (mercury ions, copper ions and hydroquinone) in concentrations not affecting cell growth seemed to stimulate the production of glutathione. Possibly, the thiol reactive agents decrease the concentration of glutathione, thereby stimulating further synthesis of glutathione, since glutathione synthesis is subject to feedback regulation by glutathione on gamma-glutamylcysteine synthase. Redox changes after addition of thiol and redox reactive agents were also investigated. Copper ions lowered the concentrations of reduced forms of all extracellular thiols and of intracellular reduced cysteine in HeLa cell cultures. The addition of mercury ions, hydroquinone, homocysteine or hydrogen peroxide did not change the proportions between reduced and total thiol concentrations. After addition of buthionine sulfoxime, the total concentrations of intra- and extracellular glutathione were markedly decreased and the ratio between reduced and total glutathione concentrations was lowered. However, both cysteine and homocysteine exhibited normal ratios between the concentrations of reduced and total thiols in the presence of buthionine sulfoxime. This finding could be due to other cellular antioxidants, such as thioredoxin, ascorbic acid or tocopherols, maintaining redox status of these thiols.     

Additional evidence of autoxidation as a possible mechanism of neutrophil locomotory dysfunction in blunt trauma

Previous studies in victims of blunt injury suggest that the observed neutrophil (PMN) locomotory dysfunction is, in part, due to autoxidation. To further clarify the occurrence and significance of autoxidation, we studied changes in levels of glutathione in PMN and of ascorbic acid and alpha-tocopherol in serum and blood cells of postsurgical and blunt trauma patients. Levels of total, reduced, and oxidized glutathione in PMN from trauma patients were similar to normal controls. Serum and cellular ascorbic acid and alpha-tocopherol levels dropped significantly after injury and remained below normal control levels during the 7 to 8-day study period. Low serum alpha-tocopherol was partially explainable on the basis of changes in serum lipids. When serum samples of trauma patients were thawed unprotected without pyrogallol, there was significant loss of recoverable alpha-tocopherol, whereas no significant losses occurred with unprotected thawed normal sera. Less total reducing capacity was observed in PMN of trauma patients compared with normal controls. These findings indicate that synthesis and regeneration capacity of glutathione are intact but that the levels of the consumable antioxidants, ascorbic acid, and alpha-tocopherol are compromised after injury. These results add further support to the hypothesis that autoxidation occurs in trauma.     

Vitamin C aspirin interactions in laboratory animals.

Ascorbic acid concentration has been determined in samples of plasma, leucocytes, urine, faeces and adrenal glands of guinea-pigs after administration of (i) 10, 25 or 100 mg ascorbic acid, (ii) 10 mg ascorbic acid plus 10, 25 or 50 mg aspirin and (iii) 25 mg aspirin and 25, 50 or 100 mg ascorbic acid. When the dose of aspirin was 25 mg or more, the transport of ascorbic acid into leucocytes was inhibited, the plasma concentration of vitamin C was elevated significantly and the excretion of ascorbic acid in the urine was increased in direct proportion to the aspirin dose. Ascorbic acid concentration in the adrenal glands was not significantly elevated after 3 h. When a constant dose of 25 mg of aspirin was given along with increasing doses of ascorbic acid both plasma and leucocyte ascorbic acid levels were elevated but not significantly after 2, 3 and 24 h. Urine ascorbic acid levels did not show any changes with the same doses.     

The use of water-soluble vitamins in viral hepatitis A]

To correct disbolism of vitamins C, B3, B6 and B12 due to viral hepatitis A (VHA), the patients received daily doses of the vitamins: 300 and 400 mg of ascorbic acid, 300 and 600 mg of calcium pantothenate or 90 and 180 mg of pantetin per os. 100 and 200 micrograms of cyanocobalamin alternating with 50 and 100 mg of pyridoxin was given intramuscularly every other day. The course lasted 3-4 weeks. The vitamin treatment promoted a positive trend in clinico-biochemical parameters, normalization of immunocompetent cell and serum immunoglobulins levels, of phagocytic reaction of peripheral blood neutrophils.     

Orally administered octacosanol improves some features of high fructose-induced metabolic syndrome in rats

We examined whether orally administered octacosanol, a long-chain aliphatic saturated alcohol, improves the features of high fructose-induced metabolic syndrome in rats. Five-week-old rats were fed a high fructose diet containing 60% fructose for 3 weeks. Then, the high fructose fed rats received a daily single oral administration of octacosanol (10 or 100 mg/kg body weight) with high fructose feeding for one week. Three- or four-week high fructose feeding increased insulin resistance, serum insulin, triglyceride, total cholesterol, free fatty acids, uric acid, and lipid peroxide concentrations, and hepatic triglyceride and cholesterol contents significantly and decreased serum high-density lipoprotein cholesterol and adiponectin concentrations significantly but did not affect blood pressure and hepatic lipid peroxide and reduced glutathione contents. Four-week high fructose feeding decreased hepatic ascorbic acid content significantly. Oral administration of octacosanol (10 or 50 mg/kg body weight) to high fructose-fed rats for the last 1-week fructose diet feeding attenuated these changes except serum insulin level and insulin resistance significantly and increased hepatic reduced glutathione content significantly. The higher dose of Oct decreased hepatic lipid peroxide content significantly. These results indicate that orally administered octacosanol improves dyslipidemia, hyperuricemia, hypoadiponectinemia, and oxidative stress associated with the features of high fructose-induced metabolic syndrome rats.     

Supplemental oxygen does not modulate responses to acetylcholine or ascorbic acid in the forearm of patients with congestive heart failure

Despite providing symptomatic relief in patients with congestive heart failure (CHF), supplemental oxygen (O(2)) has been demonstrated to increase total peripheral resistance. The present study investigated the possibility that O(2) inhalation reduces nitric oxide (NO) bioavailability, using endothelium-dependent (acetylcholine) and -independent (phentolamine) vasodilators, and the antioxidant ascorbic acid. Ten patients (nine male and one female) with primary left ventricular failure participated in the study. Forearm venous occlusion plethysmography was used to study blood flow responses to acetylcholine and the alpha-adrenergic antagonist phentolamine during inhalation of either room air or 100% O(2), with and without the simultaneous infusion of ascorbic acid. Neither O(2) inhalation (3.9+/-0.4 compared with 3.8+/-0.3 ml.min(-1).100 ml(-1)) nor ascorbic acid infusion (5.2+/-0.4 compared with 5.5+/-0.4 ml.min(-1).100 ml(-1)) affected resting forearm blood flow. The percentage increase from basal blood flow after acetylcholine infusion was not altered by either O(2) inhalation or ascorbic acid infusion (room air, 140+/-55%; O(2), 118+/-46%; ascorbic acid, 147+/-39%; ascorbic acid+O(2), 109+/-31%). O(2) inhalation did, however, reduce the dilation induced by phentolamine (room air, 131+/-24%; O(2), 80+/-14%; P<0.05). These data indicate that oxygen inhalation does not increase forearm vascular resistance. Secondly, preservation of reactivity to acetylcholine during O(2) inhalation suggests that degradation of NO by O(2)-derived free radicals is not enhanced. Attenuation of phentolamine-induced vasodilation during O(2) inhalation, however, implies increased adrenergic activity, which may possibly exacerbate the detrimental effects of elevated sympathetic activity in CHF.     

(E)‐2‐benzylidene‐4‐phenyl‐1,3‐diselenole has antioxidant and hepatoprotective properties against oxidative damage induced by 2‐nitropropane in rats

The in vitro effect of (E)‐2‐benzylidene‐4‐phenyl‐1,3‐diselenole (BPD) was evaluated through iron/EDTA‐induced thiobarbituric acid reactive species (TBARS) and reactive species (RS) determinations as well as of the scavenging 2,2′‐diphenyl‐1‐picrylhydrazyl (DPPH) radical quantification. BPD at the concentrations of 10 and 50 μΜ decreased RS and TBARS levels, respectively. The antioxidant activity was not related to the scavenging DPPH radical mechanism. A second objective of this study was to investigate the hepatoprotective action of BPD, administered by oral route, against oxidative damage induced by 2‐nitropropane (2‐NP) (100 mg/kg of body weight) in liver of rats. At the dose of 50 mg/kg, BPD protected against the increase in aspartate aminotransferase (AST), alanine aminotransferase (ALT), and lactate dehydrogenase (LDH) activities induced by 2‐NP. BPD (10 and 50 mg/kg) protected against the increase in TBARS levels and alkaline phosphatase (ALP) activity. Sections of liver from 2‐NP‐exposed rats presented intense infiltration of inflammatory cells and loss of cellular architecture. BPD (10 and 50 mg/kg) attenuated 2‐NP‐induced hepatic histological alterations. The inhibition of δ‐aminolevulinic dehydratase (δ‐ALA‐D), glutathione peroxidase (GPx), glutathione reductase (GR), and glutathione S‐transferase (GST) activities and the decreased GSH levels caused by 2‐NP were protected by BPD (50 mg/kg). Catalase activity and ascorbic acid levels were not altered by 2‐NP. These results demonstrated the antioxidant and hepatoprotective effects of BPD in liver of rats.     

Ionized calcium during plateletpheresis

The concentration of ionized calcium (Ca++) during platelepheresis was monitored when donors received 461 ± 95 (mean ± S.D.) ml of anticoagulant acid-citrate-dextrose (ACD), N.I.H. formula A. Most donors experienced mild subjective symptoms (perioral tingling) during reinfusion of autologous blood. The concentration of serum Ca++ before the procedure was 4.19 ± 0.203 (mean ± S.D.) mg/dl and it decreased to 3.27 ± 0.391 (mean ± S.D.) mg/dl after the procedure. Two of the 79 donors experienced more severe symptoms (nausea, lightheadedness) while ionized calcium was lower than 3 mg/dl. Mild hypocalcemic effects could be demonstrated by electrocardiography in most donors. When donors received 414 ± 28 (mean ± S.D.) ml of ACD, N.I.H. formula B, only some of them experienced perioral tingling during reinfusion of the autologous blood. The concentration of Ca++ before the procedure was 4.18 ± 0.14 (mean ± S.D.) mg/dl and it decreased to 3.66 ± 0.14 (mean ± S.D.) mg/dl after the procedure. These data indicated that ACD N.I.H. formula B promotes greater donor comfort and safety than does ACD, formula A. The data showed that hemodilution occurred immediately following the withdrawal of one unit of blood, prior to infusion of any intravenous fluids. The analysis of the data revealed also that both magnesium and calcium entered the intravascular compartment during plateletpheresis. Although the origin of the calcium ion entering the plasma could not be established, it appeared that at least part was derived from calcium-citrate complexes.     

Autonomic reflexes and vascular reactivity in experimental scurvy in man

Ascorbic acid is a required cofactor in the conversion of dopamine to norepinephrine in vitro, and the deficiency of this vitamin in guinea pigs is associated with degeneration of autonomic ganglion cells and with cardiac supersensitivity to norepinephrine. Because of these findings, we tested the hypothesis that ascorbic acid deficiency in man alters autonomic cardiovascular reflexes and vasomotor responses to adrenergic stimuli. We studied five normal volunteers who had been deprived of ascorbic acid for a period of 3 months; they had developed symptoms and signs of scurvy and their plasma levels of ascorbic acid averaged 0.178 ±SE 0.07 mg/100 ml. We repeated the studies after giving the subjects vitamin C for a period of 4 months; they had become asymptomatic and their plasma ascorbic acid had increased to an average of 1.68 ±0.151 mg/100 ml.     

Hemoglobin Function of Blood Stored at 4 C in ACD and CPD with Adenine and Inosine

Normal hemoglobin function depends on adequate erythrocyte levels of 2,3‐diphosphoglycerate (2,3‐DPG), a compound that is poorly maintained during blood bank storage in acid‐citrate‐dextrose (ACD). Since 2,3‐DPG is better maintained at the higher pH afforded by citrate‐phosphate‐dextrose (CPD), hemoglobin function was compared during storage in CPD and ACD. Further, hemoglobin function was studied in CPD blood containing adenine and inosine, compounds that provide metabolic energy and thus prolong the shelf‐life of blood, because they also effect the levels of 2,3‐DPG during storage. Hemoglobin function, expressed as the P₅₀ (the P₀₂ at 50 per cent oxygenation, an inverse but direct measure of oxygen affinity) is considerably better maintained during storage in CPD than in ACD. The hemoglobin function or P₅₀ of blood stored in CPD‐adenine is not maintained as well as blood stored in CPD without adenine, but the oxyhemoglobin dissociation curves show only a small difference when compared to the difference between ACD and CPD. Blood stored in CPD‐adenine with inosine, present initially or added at day 25, allows higher P₅₀ values late in storage, thus providing better hemoglobin function for more of the storage period.     

The effect of folic acid,vitamin B6 and vitamin B12 on the homocysteine levels in rabbits fed by methionine-enriched diets

Atherosclerosis is an important cause of cardiovascular morbidity and mortality in recent years. Hyperhomocysteinemia is recognized as an independent risk factor for premature atherosclerosis and venous thrombosis. It is suggested that administration of folic acid, vitamin B6 and vitamin B12 may decrease homocysteine levels. In our study, we induced hyperhomocysteinemia in rabbits by giving methionine and studied the effects of folic acid, vitamin B6 and vitamin B12 on homocysteine levels. A total of 40 (20 female, 20 male New Zealand rabbits) were divided into four groups, each consisting of 10 rabbits. Methionine (100 mg/kg/day), methionine (100 mg/kg/day) plus vitamin B6 (30 mg/kg/day), methionine (100 mg/kg/day) plus vitamin B12 (80 mg/kg/day) and methionine (100 mg/kg/day) plus folic acid (20 mg/kg/day) were given to the first, second, third and forth groups respectively. These rabbits were followed up for two months. We studied homocysteine levels on the 0, 20th, 40th and 60th days in all groups. In rabbits we induced hyperhomocysteinemia by giving methionine for 2 months. The decreases of homocysteine levels in the forth group were significant with respect to the second and third groups. Folic acid supplementation clearly resulted in a reduction of plasma homocysteine levels, whereas vitamin B12 was little effective and vitamin B6 failed to show an effect. We conclude that even folic acid treatment alone may be sufficient for decreasing negative effects of homocysteine.