Time course of vitamin C distribution and absorption after oral administration in SMP30/GNL knockout mice

Objective

Because vitamin C (VC) has multiple metabolic and antioxidant functions, we investigated the movement of VC throughout the tissues of senescence marker protein-30 (SMP30)/gluconolactonase (GNL) knockout (KO) mice.

Methods

SMP30/GNL KO mice, which cannot synthesize VC in vivo, were divided into two groups: VC sufficient and VC deficient. Starting at 2 mo of age, both groups had free access to water containing 1.5 and 0.0375 g/L of VC for 1 mo.

Results

The average rate of VC retention in 20 tissues of VC-deficient SMP30/GNL KO mice was only 13.7% of that in VC-sufficient mice. Tissues that retained over 20% of VC were the cerebellum, white fat, testes, eyeballs, and pancreas, and those with less than 5% VC were the kidneys and heart. These results clearly indicate the different VC retention capacities among tissues. Next, we examined the time course of VC distribution and absorption in VC-deficient SMP30/GNL KO mice. After oral VC administration, VC content in the liver and kidney peaked at 3 h and then decreased. VC content in the lungs, adrenal glands, skin, white fat, and pancreas peaked at 6 h and in the cerebellum, cerebrum, skeletal muscles, eyeballs, thyroid gland, and testes at 12 h.

Conclusion

In this study, we found that exogenous VC administered orally in VC-deficient SMP30/GNL KO mice was distributed at distinctly different rates within individual tissues. The SMP30/GNL KO mice used in this study are a useful animal model that provides unique opportunities for investigating VC movement and metabolism in the entire body.     

Bone degeneration and its recovery in SMP30/GNL-knockout mice

Senescence marker protein-30 (SMP30) decreases androgen-independently with aging and is a lactone-hydrolyzing enzyme gluconolactonase (GNL) that is involved in vitamin C biosynthesis. In the present study, bone properties of SMP30/GNL knockout (KO) mice with deficiency in vitamin C synthesis were investigated to reveal the effects of SMP30/GNL and exogenous vitamin C supplementation on bone formation. Mineral content (BMC) and mineral density (BMD) of the mandible and femur of SMP30/GNL KO and wild-type mice at 2 and 3 months of age with or without vitamin C supplementation were measured by dual-energy X-ray absorptiometry. Body and bone weight of both age groups decreased and became significantly lower than those of wild-type mice. The bones of SMP30/GNL KO mice were rough and porous, with BMC and BMD significantly below wild-type. Oral supplementation with vitamin C eliminated differences in body weight, bone weight, BMC, and BMD between SMP30/GNL KO and wild-type mice at each age. These results indicate that bone degeneration in SMP30/GNL KO mice was caused by lack of vitamin C, and that this mouse strain is an appropriate model for bone metabolism in humans, which have no ability to synthesize vitamin C.     

Effect of ascorbic acid intake on tissue dehydroascorbic acid in mice

The effect of ascorbic acid intake on tissue levels of ascorbic acid, dehydroascorbic acid and the ratio of dehydroascorbic acid to ascorbic acid in mice was studied. In general, the trend of changes in tissue concentrations was: ascorbic acid > dehydroascorbic acid ratio of dehydroascorbic acid to ascorbic acid. Mice fed a diet with 1% ascorbic acid had significantly higher concentration of dehydroascorbic acid in the kidney, lung and spleen than did control mice fed an ascorbic acid-free diet. Mice fed a diet with 5% ascorbic acid had elevated levels of dehydroascorbic acid in the brain, kidney, liver, lung and spleen. The kidney and lung had the greatest increase in dehydroascorbic acid concentration, suggesting that these two organs may be important sites for catabolism and elimination of ascorbic acid. In comparison with the corresponding control values, the ratio of dehydroascorbic acid to ascorbic acid was higher in the lung, not different in the liver and spleen, and lower in the kidney of mice fed a diet with 1 or 5% added ascorbic acid. These ratios were higher in the brain of mice fed a diet with 5% added ascorbic acid than in mice fed the ascorbic-acid-free diet. No apparent physiological abnormality in these animals was observed. These effects were stereospecific. Exogenous erythorbic acid, D-isoascorbic acid, a stereoisomer of ascorbic acid, increased dehydroascorbic acid equivalents (the sum of dehydroascorbic and dehydroerythorbic acid) in the kidney, lung, and spleen but the ratios of dehydroascorbic acid plus dehydroerythorbic acid to ascorbic acid plus erythorbic acid were essentially unaffected. A large glucose intake (1 or 5% in the diet) did not have an effect on levels of tissue ascorbic acid or dehydroascorbic acid.     

Effect of dietary ascorbic acid intake on tissue vitamin C in mice

The effect of graded levels of dietary ascorbic acid on blood and tissue ascorbic acid levels in mice has been studied. Six levels of dietary ascorbic acid (0, 0.076, 0.5, 1, 5 and 8%) were used. Plasma ascorbic acid rose as dietary ascorbic acid intake increased from 1 to 8%. Mice fed a diet with 5 or 8% added ascorbic acid had significantly higher levels of ascorbic acid in the heart, kidney, lung, muscle and spleen than did control mice fed an ascorbic acid-free diet. Mice fed a diet with 1% added ascorbic acid had elevated ascorbic acid levels in the heart, kidney, lung and spleen. No significant change was observed in ascorbic acid level in the brain, adrenal gland or leukocytes in any of the experimental groups. Ascorbic acid level in the eyes was only slightly higher in mice fed a diet containing 8% added ascorbic acid than in control mice. The observation that the kidney had the greatest increase in ascorbic acid content suggests that the kidney may be a very important organ not only in elimination but also in catabolism of this vitamin. A diet containing 0.5 or 0.076% added ascorbic acid did not significantly increase ascorbic acid content in any of the organs studied. Mice fed a diet with 0.076% added ascorbic acid had slightly, but statistically significantly, lower levels of ascorbic acid in the liver, lung, muscle and spleen that control mice. Mice fed a diet with 0.5% added ascorbic acid had a lower ascorbic acid content in the liver and muscle than the controls.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of ascorbic acid on guinea pig adrenal adenylate cyclase activity and plasma cortisol

Intra-adrenal ascorbic acid (AsA) concentrations exert a braking or modulating effect upon steroid release. Because changes in steroidogenesis are mediated through alterations in adenylate cyclase activity (ACL), the effect of varied AsA plasma concentrations on guinea pig adrenal ACL activity and plasma cortisol was studied. Forty-two male guinea pigs were randomly allocated to the following seven groups: controls, scorbutic, and groups given 0.1, 5, 10, 20 or 100 mg ascorbic acid/100 g body weight, respectively. Scorbutic animals had very low levels of AsA in comparison to control animals. Plasma AsA levels increased as AsA dose increased. The levels of AsA in the group given 0.1 mg AsA were higher than in controls. Basal adenylate cyclase activity did not vary significantly among animal groups. In contrast, values for NaF-stimulated ACL activity showed a progressive decrease with increasing AsA doses. A highly significant correlation was found between decreasing ACL activity and increasing plasma AsA concentrations. On the other hand, NaF-responsive ACL activity was higher in scorbutic animals than in any other group. Higher mean cortisol values were found in the scorbutic group than in the controls, correlating with high levels of NAF-stimulated ACL activity. Higher mean cortisol values were also found in the group given 0.1 mg AsA although ACL activity in this group was not affected. This finding, coupled with reduced ACL activity in these groups, is consistent with the inhibitory effect of a megadose of AsA on production of cortisol from the adrenal. The above data may suggest that differing plasma concentrations of AsA regulate in vivo steroidogenesis by altering the activity of the membrane-bound enzyme adenylate cyclase.     

Effect of several xenobiotics on the activities of enzymes affecting ascorbic acid synthesis in rats

The dietary addition of several xenobiotics, such as PCB, DDT, aminopyrine, chloretone, BHT and BHA, caused significant increases in the ascorbic acid in urine and liver of rats. The administration of all types of xenobiotics used in the present experiments increased the activity of hepatic UDP-glucose dehydrogenase (1.3-2.8-fold), and the administration of PCB, DDT, BHT or BHA significantly increased the activity of hepatic UDP-glucuronyl transferase (2.2-13.1-fold). The activity of beta-glucuronidase was slightly increased with feeding of PCB, DDT, chloretone or aminopyrine. However, the activity of hepatic UDP-glucuronic acid pyrophosphatase, the conversion of D-glucuronic acid or D-glucuronolactone into L-ascorbic acid and the activity of hepatic L-gulonolactone oxidase did not increase with the administration of PCB or DDT. It is suggested that the increases in the activities of UDP-glucose dehydrogenase and UDP-glucuronyl transferase would have a major role in the stimulation of ascorbic acid synthesis in xenobiotic treated rats.     

Effect of ascorbic acid and vitamin E on biochemical changes associated with vitamin E deficiency in rats

Weanling male Sprague Dawley rats were fed a vitamin E and C-free basal diet with or without supplementation of 100 IU vitamin E per kg diet. After 20 weeks, the vitamin E-deficient rats were divided into four groups, six in each group, and received supplemental ascorbic acid and/or vitamin E by tube feeding daily for 7 days: Group I, 30 mg ascorbic acid/100 g body wt.; Group II, 0.03 mg RRR-alpha-tocopheryl acetate/100 g body wt.; Group III, 30 mg ascorbic acid and 0.03 mg RRR-alpha-tocopheryl acetate/100 g body wt.; and Group IV, placebo. The six control rats (Group V) received placebo. The rats were sacrificed, blood and liver samples were collected for biochemical determinations. Vitamin E deficiency significantly increased erythrocyte (RBC) spontaneous hemolysis, liver thiobarbituric acid (TBA) value, activities of glutamateoxaloacetate transaminase (GOT), pyruvate kinase (PK), and creatine phosphokinase (CPK) in plasma, and significantly lowered plasma vitamin E levels and glutathione peroxidase (GPX) activities. Tube-feeding ascorbic acid for 7 days produced partial reversal effect on liver TBA values, activities of plasma PK, GOT, CPK, and plasma vitamin E levels but not on RBC hemolysis and plasma GPX activity. Tube feeding both ascorbic acid and vitamin E showed similar partial reversal effect as feeding vitamin E alone on all the parameters stated above. The results suggest that ascorbic acid may spare the metabolism of vitamin E and partially reverse the changes in some of the biochemical parameters characteristic of vitamin E deficiency.     

二硫化碳染毒对ApoE基因敲除小鼠和C57BL/6J小鼠脂肪酸代谢的影响

目的 研究二硫化碳(CS2)染毒对ApoE基因敲除小鼠和C57BL/6J小鼠脂肪酸代谢的影响.方法 将24只雄性ApoE基因敲除小鼠随机分为CS2染毒正常饮食组、CS2未染毒正常饮食组、CS2染毒高脂饮食组、CS2未染毒高脂饮食组;24只C57BU6J雄性小鼠也按同样的方式分成4组;每组6只.将染毒组以浓度为1 g/m3的CS2进行静式吸入染毒,5 h/d,5 d/周,共2周.收集小鼠全血,采用酸催化甲酯化方法对脂肪酸进行衍生化,并用气质联用(GC-MS)方法比较染毒前后脂肪酸含量.结果 C57BL/6J小鼠染毒高脂饮食组花生酸含量明显低于C57BL/6J小鼠未染毒高脂饮食组,ApoE基因敲除小鼠染毒正常饮食组花生四烯酸含量明显低于ApoE基因敲除小鼠未染毒正常饮食组,ApoE基因敲除小鼠染毒高脂饮食组γ-亚麻酸含量明显高于ApoE基因敲除小鼠未染毒高脂饮食组,差异均有统计学意义(P＜0.05).结论 CS2染毒可以引起小鼠脂肪酸代谢紊乱,CS2可能对动脉粥样硬化等心血管疾病有影响.     

Effect of ascorbic acid on calcium elimination in humans

The long-term and short-term influence of large oral doses of ascorbic acid on the urinary excretion of calcium has been investigated. In the first experiment, daily doses of a total of 10 g of ascorbic acid were administered to healthy human subjects. Daily urinary samples of these subjects were collected before and during the treatment, and calcium contents of these samples were measured. Among the 22 subjects, 19 experienced no significant changes in urinary calcium levels during the ingestion of ascorbic acid, one subject experienced an increase, two had a decline. These changes in urinary calcium levels were relatively small and were within the changes from consuming normal diets. In the second experiment, urinary samples of 46 healthy subjects were collected during a period of 8 hours after the ingestion of 2 g of ascorbic acid (33 times the U.S. RDA). A significant increase in mean urinary calcium excretion from 48.2 +/- 25.1 mg to 58.3 +/- 28.0 mg in the 8-h time period was observed. Mean urinary volume and phosphorus were unchanged. Calcium levels of the initially low excretors were significantly elevated while the change in urinary calcium levels of the initially high excretors was not statistically significant following the administration of ascorbic acid. The results suggest that ascorbic acid has a short-term effect on the regulation of the absorption and metabolism of calcium in humans.     

Effect of prolonged marginal ascorbic acid deficiency on lenticular levels of antioxidants and lipid peroxide in guinea pigs

We examined the effect of prolonged marginal ascorbic acid deficiency of the levels of antioxidants and lipid peroxide in lenses of guinea pigs in order to clarify lenticular antioxidant status under ascorbic acid deficiency. Male guinea pigs aged 4 weeks were given a scorbutic diet (20 g/animal per day) with either marginally deficient ascorbic acid (0.5 mg/animal per day) or sufficient ascorbic acid (1 g/animal per day) in drinking water for 3 and 6 months. The deficient group showed no lens opacity during the administration period. The deficient group had 62.3 and 53.9% of lenticular ascorbic acid content in the sufficient group at 3 and 6 months of ascorbic acid deficiency, respectively. There were no differences in lenticular contents of reduced glutathione and thiobarbituric acid reactive substances, an index of lipid peroxidation, between both groups at 3 and 6 months of ascorbic acid deficiency, while the deficient group tended to have higher lenticular vitamin E content than the sufficient group. The deficient group had higher serum vitamin E concentration than the sufficient group at 3 and 6 months of ascorbic acid deficiency. These results indicate that lenticular antioxidant status is maintained well in guinea pigs with prolonged marginal ascorbic acid deficiency, which may result in no cataract formation.     

Effect of ascorbic acid on plasma alcohol clearance

The effects of short-term and long-term ascorbic acid supplements on plasma alcohol clearance were studied in 13 clinically healthy male subjects. Two dose levels of alcohol, 0.5 and 0.8 g/kg body weight, were used. Blood samples were taken at zero time, 0.5 hours, then hourly up to 6 hours after alcohol consumption for the measurement of plasma alcohol and ascorbic acid levels, red-cell reduced glutathione level, and plasma alanine aminotransferase activity. At both dosages of alcohol, short-term as well as long-term pretreatment with ascorbic acid significantly enhanced the clearance of plasma alcohol. Although long-term ascorbic acid pretreatment resulted in better alcohol clearance, no significant difference in alcohol clearance was found between short-term and long-term ascorbic acid pretreatment. The two dose levels of alcohol had no significant effect on the red-cell reduced glutathione concentration or plasma alanine aminotransferase activity.