Inhibition of key enzymes of carbohydrate metabolism in regenerating mouse liver by ascorbic acid.

The effect of ascorbic acid on the key enzymes of carbohydrate metabolism e.g. hexokinases, phosphofructokinase, pyruvate kinase, lactate dehydrogenase, glucose-6-phosphate dehydrogenase and malic enzyme was determined in regenerating mouse liver. All the enzymes showed a significant increase in the activity during regeneration. Ascorbic acid reduced the activities of the enzymes in regenerating liver. A decrease in liver weight in ascorbic acid treated animals may be correlated with its effect on these enzymes as glycolytic pathway is the main source of energy required by the dividing cells.     

Effect of the carcinogenic tyrosine metabolite p-hydroxyphenyllactic acid on the ascorbic acid concentration in the organs and blood of mice

The effect of cancerogenic tyrosine metabolite, p-hydroxyphenyllactic acid, on the concentration of ascorbic acid in the organs and blood of mice has been studied. p-Hydroxyphenyllactic acid was demonstrated to decrease considerably ascorbic acid concentration in the liver, adrenal glands and blood of mice. The above phenomenon and the previous data on tyrosine aminotransferase induction by p-hydroxyphenyllactic acid suggest the existence of two interdependent mechanisms of cancerogenic tyrosine metabolite (p-hydroxyphenyllactic acid) accumulation.     

Possible mechanisms responsible for the increased ascorbic acid content of Plasmodium vinckei-infected mouse erythrocytes

The possible mechanisms underlying the acquisition of an increased ascorbic acid content by mouse erythrocytes containing the malarial parasite Plasmodium vinckei were investigated. Ascorbic acid was taken up readily by parasitized red blood cells but not by controls, whilst its partly oxidized form, dehydroascorbic acid, entered both. The uptake of both ascorbic acid and dehydroascorbic acid into erythrocytes was increased as a result of malarial infection. Lysates prepared from parasitized red blood cells reduced exogenous dehydroascorbic acid to ascorbic acid at a higher rate than control red blood cell lysates; this difference was abolished following dialysis of the lysates, a process which removes endogenous reduced glutathione (GSH). The rates of chemical and enzymatic reduction of dehydroascorbic acid to ascorbic acid by GSH were of similar magnitude, thus calling into question the existence of a specific dehydroascorbate reductase in erythrocytes and parasites. These observations suggest that the increased uptake of dehydroascorbic acid into parasitized red blood cells may be a result of enhanced dehydroascorbate-reducing capacity, whilst the presence of the parasite induces a selective increase in the permeability of the erythrocyte plasma membrane to ascorbic acid. The endogenous ascorbic acid content of livers obtained from infected mice was 55% below the normal concentration and its relative rate of destruction during incubation in vitro was enhanced in comparison with that of control livers. Furthermore, the capacity of liver homogenates to synthesize ascorbic acid from glucuronic acid was greatly reduced in infected mice. Therefore it is unlikely that the increase in ascorbic acid content of parasitized red blood cells is a consequence of increased biosynthesis and release of ascorbic acid by the host liver. We have not been able to exclude the possibility that the malarial parasite itself may be capable of de novo synthesis of ascorbic acid.     

The effect of molybdenum on ascorbic acid metabolism in rats

1. The effect of dietary molybdenum on the growth rate and also on ascorbic acid metabolism in rats was studied. An excess of dietary molybdenum resulted in growth retardation and loss of weight. Tolerance to molybdenum was affected by the nature of the molybdenum salt administered. 2. Molybdenum ingestion altered certain aspects of ascorbic acid metabolism in rats. The conversion of d-glucuronolactone into l-ascorbic acid in vitro and the oxidative breakdown of l-ascorbic acid by liver enzymes decreased with high molybdenum intakes. The activity of liver uronolactonase was slightly inhibited. The activities of l-gulonate dehydrogenase and l-gulonate decarboxylase were not affected appreciably. 3. Molybdenum supplementation of the control diet resulted in an increase in ascorbic acid content of spleen and adrenal gland, and in a marked decrease in the urinary excretion of ascorbic acid and glucuronic acid. The implications of these findings are discussed.     

Therapeutic potentials of combined use of DMSA with calcium and ascorbic acid in the treatment of mild to moderately lead intoxicated mice

The aim of this study was to explore the therapeutic efficacies of combined use of meso-2,3-dimercaptosuccinic acid (DMSA) with calcium and ascorbic acid in the treatment of mild to moderately lead-intoxicated mice. Female albino mice were exposed to lead by drinking water contaminated with 0.1% (moderate lead exposure) or 0.05% (mild lead exposure) lead acetate. After the cessation of lead exposure, mice were supplemented by gavage with saline solution, 50 mg/kg body weight (b.w) DMSA, 100 mg/kg b.w DMSA, calcium and ascorbic acid, or 50 mg/kg b.w DMSA and calcium as well as ascorbic acid, respectively. Atomic absorption spectrophotometric method was used to analyze lead levels in blood, bone, liver, kidney and brain. Activities of blood δ-aminolevulinic acid dehydratase (ALAD) were determined by colorimetric method. DMSA supplemented alone could reduce lead levels in both soft tissues and bone and reverse lead-inhibited activities of blood ALAD in mild to moderately lead-intoxicated mice. On the other hand, combined use of DMSA with calcium and ascorbic acid achieved better therapeutic efficacies in mobilizing lead in blood, liver and kidney, and reversing lead-inhibited activities of blood ALAD in moderately lead intoxicated mice than DMSA supplemented alone. Moreover, the better therapeutic efficacies were also found in mildly lead intoxicated mice in mobilizing lead in blood and bone achieved by combined use of DMSA with calcium and ascorbic acid. Combined use of DMSA with calcium and ascorbic acid seems to be the better choice in the treatment of mild to moderate lead-intoxication.     

Effects of ascorbic acid on cadmium-induced oxidative stress and performance of broilers

The effects of cadmium on performance, antioxidant defense system, liver and kidney functions, and cadmium accumulation in selected tissues of broiler chickens were studied. Whether the possible adverse effects of cadmium would reverse with the antioxidant ascorbic acid was also investigated. Hence, 4 treatment groups (3 replicates of 10 chicks each) were designed in the study: control, ascorbic acid, cadmium, and cadmium plus ascorbic acid. Cadmium was given via the drinking water at a concentration of 25 mg/L for 6 wk. Ascorbic acid was added to the basal diet at 200 mg/kg either alone or with cadmium. Cadmium decreased the body weight (BW), body weight gain (BWG), and feed efficiency (FE) significantly at the end of the experiment, wheras its effect on feed consumption (FC) was not significant. Cadmium increased the plasma malondialdehyde (MDA) level as an indicator of lipid peroxidation and lowered the activity of blood superoxide dismutase (SOD). Liver function enzymes, aspartate amino transferase (AST), alanine aminotransferase (ALT), lactate dehydrogenase (LDH), and gamma glutamyl transferase (GGT) activities were not changed by cadmium. Cadmium ingestion did not alter serum creatinine levels. Although the serum cadmium level was not elevated, cadmium mainly accumulated in the kidneys, liver, pancreas, and muscle. Ascorbic acid supplementation resulted in a reduction of MDA level previously increased by cadmium and a restoration in SOD activity. However, ascorbic acid did not ameliorate the growth inhibitory effect of cadmium nor did it prevent accumulation of cadmium in analyzed tissues. These data indicate that oxidative stress, induced by cadmium, plays a role in decreasing the performance of broilers and that dietary supplementation by ascorbic acid might be useful in reversing the lipid peroxidation induced by cadmium and partly alleviating the adverse effect of cadmium on performance of broilers.     

Role of exogenous ascorbic acid in tissue status of ascorbic acid-2-sulphate in guinea pigs.

Guinea pigs were given ascorbic acid orally in two doses; a low and a high dose. The tissue levels of ascorbic acid-2-sulphate was estimated in these animals after 15 days of feeding and a subsequent deprivation period of 15 days. The specific activity of the enzymes ascorbic acid sulphotransferase and ascorbic acid-2-sulphate sulphohydrolase was studied. During higher ascorbic acid intake, the activity of ascorbic acid sulphotransferase was increased, whereas ascorbic acid-2-sulphate sulphohydrolase showed a decreased activity. But when ascorbic acid intake was lowered or ceased, the activity of the above enzymes showed a reverse pattern. Possible reasons for the lack of antiscorbutic activity of ascorbic acid-2-sulphate in guinea pigs is discussed.     

Effect of large doses of ascorbic acid on the hepatic and extra-hepatic drug-metabolizing enzymes in guinea pig

Water solubility and non-toxic properties of ascorbic acid are taken as criteria for beneficial effects of large doses of the vitamin. In the present study, male guinea pigs, dosed daily with 15, 30 or 50 mg/100g body weight for 10 weeks, demonstrated no differences in effect on liver and lung weights, body growth and microsomal protein contents of liver and lung when compared with controls. When guinea pigs were fed excessive ascorbic acid, there was a small non-significant increase (p less than 0.05) in hepatic and pulmonary cytochrome P-450, and significant increase (p less than 0.05) in hepatic cytochrome b5 which was accompanied with a significant increase in arylhydrocarbon hydroxylase activity in the two organs. Activity of NADPH-dependent cytochrome c-reductase was decreased in liver and remained unaffected in lung and colon. Drug detoxifying enzymes responded in different ways to increased intake of ascorbic acid. Activity of UDP-glucuronyltransferase remained unchanged on feeding excessive ascorbic acid, whereas glutathione S-transferase was decreased significantly in liver and was unaltered in lung and colon. Reduced glutathione was decreased only in the lung. The observed changes in drug activating and detoxifying enzymes appear to be important from drug pharmacokinetics and carcinogenesis point of view.     

The total ascorbic acid content and its variation with seasonal and nutritional factors in some common cirripedes

An investigation has been made of the ascorbic acid content, its seasonal changes, and the effect of starvation in several cirripedes. In Balanus balanoides (L.) and B. balanus (L.) there is an abrupt fall in ascorbic acid content which coincides with copulation. After fertilization values are low until the animals begin feeding again in March–April. Seasonal changes are not so clearly marked in Chthamalus spp.

60% of the ascorbic acid of Balanus balanoides is lost in the first month of starvation after which the rate of loss is reduced. In spite of different lengths of time for embryonic development 77% of the original ascorbic acid content of the eggs is lost in both B. balanoides and B. balanus.

Ascorbic acid has been demonstrated histologically in bodies of B. balanoides. Experiments with homogenates of isolated bodies, using additions of substrates and co-factors known to be required for in vitro synthesis, suggest very strongly that the tissues are incapable of synthesizing ascorbic acid unless at a very low level. Addition of the microsomal enzymes from a rabbit liver preparation to an homogenate with substrate and co-factors gave ascorbic acid synthesis. This suggests a “lesion” analogous to that found in other organisms known to be incapable of synthesizing ascorbic acid.     

Elevated oxidative stress and sensorimotor deficits but normal cognition in mice that cannot synthesize ascorbic acid

Oxidative stress is implicated in the cognitive deterioration associated with normal aging as well as neurodegenerative disorders such as Alzheimer's and Parkinson's diseases. We investigated the effect of ascorbic acid (vitamin C) on oxidative stress, cognition, and motor abilities in mice null for gulono-γ-lactone oxidase (Gulo). Gulo−/− mice are unable to synthesize ascorbic acid and depend on dietary ascorbic acid for survival. Gulo−/− mice were given supplements that provided them either with ascorbic acid levels equal to- or slightly higher than wild-type mice (Gulo-sufficient), or lower than physiological levels (Gulo-low) that were just enough to prevent scurvy. Ascorbic acid is a major anti-oxidant in mice and any reduction in ascorbic acid level is therefore likely to result in increased oxidative stress. Ascorbic acid levels in the brain and liver were higher in Gulo-sufficient mice than in Gulo-low mice. F₄-neuroprostanes were elevated in cortex and cerebellum in Gulo-low mice and in the cortex of Gulo-sufficient mice. All Gulo−/− mice were cognitively normal but had a strength and agility deficit that was worse in Gulo-low mice. This suggests that low levels of ascorbic acid and elevated oxidative stress as measured by F₄-neuroprostanes alone are insufficient to impair memory in the knockouts but may be responsible for the exacerbated motor deficits in Gulo-low mice, and ascorbic acid may have a vital role in maintaining motor abilities.     

Comparative tissue ascorbic acid studies in fishes

Comparative tissue ascorbic acid levels in four species of major carp viz., Labeo rohila, L. calbasu, Cirrhina tnrigala and Catla catla , were investigated. The ascorbic acid level was found to be the highest in the spleen in the four species studied (range 430–380 μg/g) followed by the anterior (adrenal) kidney, gonads, liver, renal kidney, brain and/or eye. Heart and blood had the lowest levels (range 26–18 μg/ml) amongst the tissues studied. Overall tissue ascorbic acid levels were the highest in L. rohita and the lowest in C. mrigala . Investigation on seasonal variations in blood and kidney ascorbic acid levels of Notopterus notopterus revealed peak levels in spring (February-April) and the lowest levels in the postspawning period (August-September).     

Protective effect of ascorbic acid on cadmium-induced hypertension and vascular dysfunction in mice

Cadmium (Cd) is one of the most important environmental pollutants that cause a number of adverse health effects in humans and animals. Recent studies have shown that Cd-induced oxidative damage within the vascular tissues results in vascular dysfunction. The current study was aimed to investigate whether ascorbic acid could protect against Cd-induced vascular dysfunction in mice. Male ICR mice were received CdCl₂ (100 mg/l) via drinking water for 8 weeks alone or received ascorbic acid supplementation at doses of 50 and 100 mg/kg/day for every other day. Results showed that Cd administration increased arterial blood pressure and blunted the vascular responses to vasoactive agents. These alterations were related to increased superoxide production in thoracic aorta, increased urinary nitrate/nitrite, increased plasma protein carbonyl, elevated malondialdehyde (MDA) concentrations in plasma and tissues, decreased blood glutathione (GSH), and increased Cd contents in blood and tissues. Ascorbic acid dose-dependently normalized the blood pressure, improved vascular reactivities to acetylcholine (ACh), phenylephrine (Phe) and sodium nitroprusside (SNP). These improvements were associated with significant suppression of oxidant formation, prevention of GSH depletion, and partial reduction of Cd contents in blood and tissues. The findings in this study provide the first evidence in pharmacological effects of ascorbic acid on alleviation of oxidative damage and improvement of vascular function in a mouse model of Cd-induced hypertension and vascular dysfunction. Moreover, our study suggests that dietary supplementation of ascorbic acid may provide beneficial effects by reversing the oxidative stress and vascular dysfunction in Cd-induced toxicity.     

A newly established strain of spontaneously hypertensive rat with a defect of ascorbic acid biosynthesis.

To investigate the effects of ascorbic acid deficiency on the pathogenesis of hypertension and/or its complications, we established a rat strain with both genetic hypertension and a defect of ascorbic acid biosynthesis. The od gene (L-gulono-gamma-lactone oxidase gene) of the ODS (Osteogenic Disorder Shionogi) rat, which is a rat mutant unable to synthesize ascorbic acid, was introduced into spontaneously hypertensive rats (SHR), and a novel congenic strain, SHR-od, was established. SHR-od showed scurvy when fed an ascorbic acid-free diet. Systolic blood pressure of male SHR-od began to increase at 9 weeks of age and reached 190-200 mmHg at 20 weeks of age. In 25-week-old SHR-od, ascorbic acid deficiency when fed an ascorbic acid-free diet for 6 weeks caused a remarkable reduction of blood pressure to lower than 110 mmHg. The wall to lumen ratio of the testicular artery in ascorbic acid-deficient SHR-od was lower than that of the control rats. When rats were fed a diet supplemented with ascorbic acid (300 mg/kg), ascorbic acid concentration in SHR-od was lower in the serum and liver than that in ODS rats. These results indicate that ascorbic acid could be closely related to the development of hypertension in SHR-od. We believe that SHR-od will be a useful model for experimental studies on hypertension and its complications, since all of them suffer from hypertension spontaneously and the level of ascorbic acid deficiency in these rats could be controlled at will both in concentration and duration.     

Ascorbic acid and aging in the rat. Uptake of ascorbic acid by teeth and concentration of various forms of ascorbic acid in different organs

1. The uptake of ascorbic acid in vitro by the teeth of rats showed a gradual decrease with age, indicating that the uptake may be related to collagen synthesis as in bone. 2. The concentration of total free ascorbic acid in various organs declined with age, but the rate of decline was different in different organs. In the spleen, however, it increased until maturity and then declined. 3. This decrease may be due to one or both of the following reasons: (a) the permeability of different tissues may decrease at different rates for ascorbic acid, or (b) the requirement for ascorbic acid may decrease at different rates. 4. The bound ascorbic acid declined with age in the skin, kidney, liver and brain after the age of 10-12 weeks, and in the spleen after the age of 26 weeks. 5. The concentration of dehydroascorbic acid and dioxogulonic acid declined with age in the skin.     

Scavenging of superoxide radical by ascorbic acid

Using acetaldehyde and xanthine oxidase as the source of suPeroxide radical, the second order rate constant for the reaction between ascorbic acid and superoxide radical was estimated to be 8.2 X 10⁷ M^-1 s^-1. In rats, the average tissue concentration of ascorbic acid was of the order of 10^-3 M and that of superoxide dismutase was of the order of 10^-6 M. So, taking together both the rate constants and the tissue concentrations, the efficacy of ascorbic acid for scavenging superoxide radical in animal tissues appears to be better than that of suPeroxide dismutase. The significance of ascorbic acid as a scavenger of superoxide radical has been discussed from the point of view of the evolution of ascorbic acid synthesizing capacity of terrestrial vertebrates.     

Pulmonary edema and ascorbic acid loss.

Loss of ascorbic acid from lung and pulmonary edema were produced in mice by intravenous injection of either adrenaline or noradrenaline (5 mumol/kg). While adrenalectomy performed before noradrenaline administration reduced the degree of pulmonary edema, a prior dose of hexamethonium accentuated this effect. Given alone, hexamethonium caused both loss of ascorbic acid and pulmonary edema. The results show that although endogenous catecholamines can potentiate the pulmonary edema produced by either adrenaline or noradrenaline, they play no specific role in the ascorbic acid loss. The evidence suggests that lung ascorbic acid levels are decreased following the development of pulmonary edema, irrespective of how it was caused.     

Detection of in vivo genotoxicity of endogenously formed N-nitroso compounds and suppression by ascorbic acid,teas and fruit juices

The genotoxicity of endogenously formed N-nitrosamines from secondary amines and sodium nitrite (NaNO(2)) was evaluated in multiple organs of mice, using comet assay. Groups of four male mice were orally given dimethylamine, proline, and morpholine simultaneously with NaNO(2). The stomach, colon, liver, kidney, urinary bladder, lung, brain, and bone marrow were sampled 3 and 24 h after these compounds had been ingested. Although secondary amines and the NaNO(2) tested did not yield DNA damage in any of the organs tested, DNA damage was observed mainly in the liver following simultaneous oral ingestion of these compounds. The administration within a 60 min interval also yielded hepatic DNA damage. It is considered that DNA damage induced in mouse organs with the coexistence of amines and nitrite in the acidic stomach is due to endogenously formed nitrosamines. Ascorbic acid reduced the liver DNA damage induced by morpholine and NaNO(2). Reductions in hepatic genotoxicity of endogenously formed N-nitrosomorpholine by tea polyphenols, such as catechins and theaflavins, and fresh apple, grape, and orange juices were more effective than was by ascorbic acid. In contrast with the antimutagenicity of ascorbic acid in the liver, ascorbic acid yielded stomach DNA damage in the presence of NaNO(2) (in the presence and absence of morpholine). Even if ascorbic acid acts as an antimutagen in the liver, nitric oxide (NO) formed from the reduction of NaNO(2) by ascorbic acid damaged stomach DNA.     

Effects of soyabean and ascorbic acid on experimental carcinogenesis

1. Ultrastructural changes in liver tissue of mice fed nitrosamine precursors, dibutylamine and nitrite, were observed. 2. The protective effect of soyabean in a diet containing nitrosamine precursors was demonstrated. 3. Liver tissue was examined to investigate the anticarcinogenicity of ascorbic acid. 4. The significance of soyabean and ascrobic acid in counteracting the potential hazards due to nitrosamine precursors is discussed.     

Cytochrome oxidase and ascorbic acid oxidase activities in cereal plants

Cytochrome oxidase and ascorbic acid oxidase activities were investigated in rye, wheat, barley and oat plants. The variations in the activity of both enzymes was followed in the course of the initial 28 days of growth, as well as at the phase of milk ripeness, namely in the cytoplasmic and mitochondrial cell fractions of roots, leaves and spikes. Both enzymes were active in all measurements. Cytochrome oxidase mostly exhibited a higher activity than ascorbio acid oxidase. The activity of the former enzyme was substantially higher in the mitochondrial fraction of leaves, roots and spikes of the four experimental plants in comparison with the cytoplasmic fraction. On the contrary, the ascorbic acid oxidase activity varied in both cell fractions according to the plant species, organ and growth phase. The variations in the activity of both enzymes exhibited on the whole a course similar to that of the respiration rate. During the first 14 to 21 days of growth the enzyme activities increased up to the maximum. This was thon followed at first by a rapid, later on by a slow decrease. The course of variations in the enzyme activities was, with certain exceptions, alike in all the four plant species investigated.     

Biosynthesis of ascorbic acid by extant actinopterygians

Polypterus senegalus , the longnose gar Lepisosteus osseus and the bowfin Amia calva had gulonolactone oxidase activity in the kidney and thus can synthesize ascorbic acid de novo . The enzyme activity was associated with the microsomal fraction. The common carp Cyprinus carpio and the goldfish Carassius auratus had no gulonolactone oxidase activity. Antibodies directed against white sturgeon gulonolactone oxidase showed cross-reactivity with lake sturgeon, bowfin and longnose gar kidney enzymes, but not with enzymes from Polypterus , sea lamprey, and tadpole kidney or pig liver. Given cross-reactivity, gulonolactone oxidase relatedness matched actinopterygian phylogeny, and suggested homology of the character throughout fishes. Modern teleosts may have lost the ability to synthesize ascorbic acid since the late Triassic as a result of a single reversal in the founding population. Wild bowfin and longnose gar exhibited high ascorbate concentrations in liver and spleen when compared with the teleosts rainbow trout Oncorhynchus mykiss and common carp fed vitamin C-supplemented diets.