L Arginine And Vitamin C

L Arginine And Vitamin C

• Research Article
• Open Access
• Published: 24 February 2012

Tetrahydrobiopterin, L-Arginine and Vitamin C Act Synergistically to Decrease Oxidative Stress, Increase Nitric Oxide and Improve Blood Flow after Induction of Hindlimb Ischemia in the Rat

Molecular Medicine volume 18,pages 676–684 (2012)Cite this article

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Abstract

Nitric oxide (NO) derived from endothelial nitric oxide synthase (eNOS) is a potent vasodilator and signaling molecule that plays an essential role in vascular remodeling of collateral arteries and perfusion recovery in response to hindlimb ischemia. In ischemic conditions, decreased NO bioavailability was observed because of increased oxidative stress, decreased L-arginine and tetrahydrobiopterin. This study tested the hypothesis that dietary cosupplementation with tetrahydrobiopterin (BH4), L-arginine, and vitamin C acts synergistically to decrease oxidative stress, increase nitric oxide and improve blood flow in response to acute hindlimb ischemia. Rats were fed normal chow, chow supplemented with BH4 or L-arginine (alone or in combination) or chow supplemented with BH4 + L-arginine + vitamin C for 1 wk before induction of unilateral hindlimb ischemia. Cosupplementation with BH4 + L-arginine resulted in greater eNOS expression, Ca²⁺-dependent NOS activity and NO concentration in gastrocnemius from the ischemic hindlimb, as well as greater recovery of foot perfusion and more collateral artery enlargement than did rats receiving either agent separately. The addition of vitamin C to the BH4 + L-arginine regimen did further increase these dependent variables, although only the increase in eNOS expression reached statistical significances. In addition, rats given all three supplements demonstrated significantly less Ca²⁺-independent activity, less nitrotyrosine accumulation, greater glutathione:glutathione disulfide (GSH:GSSG) ratio and less gastrocnemius muscle necrosis, on both macroscopic and microscopic levels. In conclusion, co-supplementation with BH4 + L-arginine + vitamin C significantly increased vascular perfusion after hindlimb ischemia by increasing eNOS activity and reducing oxidative stress and tissue necrosis. Oral cosupplementation of L-arginine, BH4 and vitamin C holds promise as a biological therapy to induce collateral artery enlargement.

Introduction

Endothelial dysfunction is a hallmark of peripheral artery disease (PAD) (1). Central to the development of endothelial dysfunction, regardless of its cause, is a reduction in the bioavailability of nitric oxide (NO) derived from endothelial nitric oxide synthase (eNOS). Three fundamental mechanisms can compromise NO bioavailability: loss of eNOS expression, loss of eNOS-derived NO production (that is, functional inactivation of eNOS) and inactivation of NO by a superoxide anion (O₂ ⁻) to form peroxynitrite (OONO⁻) (2,3). It is likely that all three mechanisms contribute to the endothelial dysfunction characteristic of PAD because increased oxidative stress is a common antecedent in the pathogenesis of this disease and can reduce NO bioavailability.

At least two characteristics of eNOS render it susceptible to oxidative stress. First, eNOS transcription, posttranslational modification and trafficking to the caveolae are attenuated by the accumulation of reactive oxygen species (ROS) within the endothelial cell (4). Second, the eNOS cofactor tetrahydrobiopterin (BH4) is highly susceptible to oxidation (5). BH4 maintains eNOS in its functional dimeric form; in the absence of BH4, eNOS becomes uncoupled so that the electron flux is diverted away from the L-arginine binding site and instead reduces molecular oxygen, generating O₂ ⁻ (6). This circumstance initiates a vicious cycle, wherein eNOS catalytic activity produces O₂ ⁻, not NO, worsening existent oxidative stress.

These molecular characteristics of eNOS predict that several therapeutic options might prove effective for the treatment of PAD, namely dietary supplementation with an antioxidant, or with the eNOS substrate L-arginine, or with the eNOS cofactor BH4. Vitamin C, or L-ascorbic acid, is a potent antioxidant and has been shown to preserve BH4 levels and enhance endothelial NO production in vitro (7). ONOO⁻ reacts with BH4 6–10 times faster when in the presence of ascorbate. The intermediate product of the reaction between ONOO⁻ and BH4 is the trihydrobiopterin radical (BH3*), which is also reduced back to BH4 by ascorbate. Thus, ascorbate does not protect BH4 from oxidation but rather recycles the BH3 radical back to BH4 (8). Vitamin C levels are low in PAD patients (9), and acute (10) or short-term (11) vitamin C supplementation reduces PAD symptoms; however, cross-sectional epidemiological surveys have failed to find a clear link between long-term vitamin C intake and PAD symptoms or disease progression (12,13). L-arginine supplementation showed exciting promise in short-term studies of PAD (14,15), but this effect was not observed in a subsequent long-term study by the same group (16). BH4 improves eNOS-dependent vasodilation in long-term smokers and patients with type 2 diabetes, conditions associated with increased oxidative stress (17,18). To our knowledge, BH4 has not been specifically evaluated as a therapeutic modality in PAD.

An important gap in our understanding of BH4, L-arginine and L-ascorbic acid in the prevention and treatment of PAD is the potential synergistic effect of combined therapy, and there is convincing evidence to suggest that such an approach would prove successful. For example, supplementation with L-arginine alone might prove deleterious in the face of endothelial oxidative stress inasmuch as the resultant increase in eNOS catalytic activity might generate O₂ ⁻, not NO, if BH4 levels were reduced by oxidation. Thus, cosupplementation of L-arginine with L-ascorbic acid and BH4 might enhance the therapeutic outcome by reducing oxidant stress and preserving eNOS in its functional dimeric form, respectively. This action would enhance eNOS-derived NO production and, by quenching existent O₂ ⁻, reduce NO inactivation by its reaction with O₂ ⁻.

In our previous work (19), we observed decreased eNOS expression, decreased bioavailable NO and increased oxidative stress in hindlimb ischemic rats, and oral supplementation of BH4 increased the beneficial effect of eNOS gene transfer. The goal of this study was to test the hypothesis that combined dietary supplementation with BH4, L-arginine and vitamin C will act synergistically to improve hindlimb perfusion recovery and preservation of tissue integrity in response to severe hindlimb ischemia. To this end, we generated severe hindlimb ischemia in the rat by means of femoral artery excision. Measured dependent variables included gastrocnemius muscle eNOS expression and activity, hindlimb laser Doppler perfusion and collateral artery enlargement, and gastrocnemius nitrotyrosine accumulation and tissue necrosis.

Materials and Methods

Materials

Dietary supplements included the following: BH4 (10 mg/kg/d; Schircks Laboratories, Jonas, Switzerland); L-arginine, provided as L-arginine α-ketoglutarate (hereafter L-arginine; 88.5 mg/kg/d; Body Tech, North Bergen, NJ, USA); and L-ascorbic acid (that is, vitamin C; 88.5 mg/kg/d, Sigma-Aldrich, St. Louis, MO, USA). The dose of tetrahydrobiopterin was selected on the basis of a published report of its use in rats (35). And the dose of L-arginine and vitamin C was on the basis of their clinical dose in patients.

Animals

All protocols were approved by the Institutional Animal Care and Use Committee at the University of California, San Francisco, and University of Massachusetts Medical School. Adult male Sprague-Dawley rats weighing 265–285 g (Charles River Laboratories, Wilmington, MA, USA) were maintained in a clean housing facility on a 12-h light/dark cycle.

Preparation

Severe ischemia was induced in the left hindlimb. The femoral artery was ligated between the inguinal ligament and popliteal fossa, and the ligated section and its branches were excised. This procedure was carried out under anesthesia with 2% isofluorane. The untreated right hindlimb served as an internal control for each rat. An additional group of sham-operated rats were also used for selected assays. These rats underwent isolation of the femoral artery in the left hindlimb under 2% isofluorane anesthesia, but the artery was left intact.

Study Design

All rats were fed standard chow in powder form and water ad libitum (Deans Feeds, Redwood City, CA, USA). Animals were randomly selected to receive normal chow (control), chow with a single added supplement (BH4 or L-arginine), chow supplemented with BH4 + L-arginine or chow supplemented with BH4 + L-arginine + L-ascorbic acid. Dietary supplementation was commenced 7 d before the induction of hindlimb ischemia and was continued until sacrifice of the animal. Chow was replaced every 2 d. The time of sacrifice varied with the measured endpoint under consideration, as described below.

Western Blotting

Rats were sacrificed 14 d after induction of ischemia for measurement of gastrocnemius muscle eNOS, phosphorylated eNOS (p-eNOS) and nitrotyrosine expression. The timing of death was selected on the basis of previous work that demonstrated maximal postischemic change in these variables at this time (19). Samples were homogenized in liquid nitrogen and transferred to NP-40 lysis buffer, comprised of 50 mmol/L HEPES (pH 7.5), 150 mmol/L NaCl, 10% glycerol, 1.5 mmol/L MgCl₂, 1 mmol/L ethylenediaminetetraacetic acid (EDTA), 100 mmol/L NaF, 1% NP 40, 1 mmol/L phenylmethylsulfonyl fluoride (PMSF) and 1 g/mL aprotinin. The lysates were centrifuged, the supernatant recovered and the protein concentration determined (Pierce Biotechnology, Rockford, IL, USA). A total of 100 µg protein per sample was separated on a 7.5% or 12% sodium dodecyl sulfate-polyacrylamide gel for determination of eNOS or nitrotyrosine, respectively, and then electroblotted on nitrocellulose membranes (Bio-Rad, Hercules, CA, USA). Membranes were incubated overnight at 4°C with mouse monoclonal anti-nitrotyrosine (Cayman Chemical, Ann Arbor, MI, USA; 1:1,000), mouse monoclonal anti-eNOS (BD Transduction Laboratories, San Diego, CA, USA; 1:1,000) or rabbit anti-p-eNOS (cell signaling) and then incubated for 2 h with horseradish peroxidase-conjugated anti-mouse or rabbit IgG antibody (Pierce Biotechnology; 1:5,000). Immunoreactive bands were visualized using the enhanced chemiluminescence system (Amersham, Arlington Heights, IL, USA). Band density was quantitated by standard densitometry, and the intensity of the band of interest was expressed as a function of the α-tubulin band.

eNOS Activity

Rats were sacrificed 14 d after induction of ischemia for determination of gastrocnemius muscle NOS activity. The timing of sacrifice was selected on the basis of previous work that demonstrated maximal postischemic change in this variable at this time (19). Muscles were homogenized in ice-cold buffer (250 mmol/L Tris-HCl, pH 7.4, 10 mmol/L EDTA, 10 mmol/L EGTA [ethylene glycol-bis|β-aminoethyl ether}-N, N, N′,N′-tetraacetic acid]) and centrifuged, and the protein in the supernatant was adjusted to 5 µg/mL. Samples were incubated in 10 mmol/L NADPH, 1 µCi/µL ¹⁴C-Arg, 6 mmol/L CaCl₂, 50 mmol/L Tris-HCl (pH 7.4), 6 µmol/L BH4, 2 µmol/L flavin adenine dinucleotide (FAD) and 2 µmol/L flavin mononucleotide (FMN) for 30 min at 37°C. The reaction was stopped with 400 µL of 50 mmol/L HEPES, pH 5.5, and 5mmol/L EDTA. Identical samples were prepared without CaCl₂, and all reactions were performed in duplicate. The radioactivity of the sample eluate was measured and expressed as carboxypeptidase M (CPM)/µg protein. The Ca²⁺-dependent NOS activity, which corresponds to the sum of the endothelial and neural NOS isoforms, was calculated by subtracting the NOS activity measured in the absence of CaCl₂ from the NOS activity measured in the presence of CaCl₂. The Ca²⁺-independent NOS activity corresponds to inflammatory isoform of NOS, or iNOS.

NOx (Nitrite + Nitrate) Assay

The left gastrocnemius muscles after ischemic d 14 were collected and homogenized in phosphate-buffered saline (PBS). After centrifugation, ultrafilter tissue homogenates were passed through a 10-kDa molecular weight cutoff filter. Filtrate was used for nitrite and nitrate concentration measurement according to the protocol of the Nitrate/Nitrite fluorometric assay kit (Cayman Chemical).

Assay of GSH:GSSG Ratio

The left gastrocnemius muscles were homogenized and deproteinated. And then the supernatant was used for the measurement of glutathione disulfide (GSSG) and glutathione (GSH) according to the Glutathione assay kit (Cayman Chemical).

Hindlimb Perfusion

Hindlimb blood flow was determined by means of laser Doppler imaging (Moor Instruments, Devon, UK). Flow was measured preoperatively, immediately after arterial excision, and then 3, 7, 14, 21, 28, 35 and 42 d after induction of ischemia. Scans were obtained during inhalation of 1% isofluorane while core body temperature was maintained between 36.8 and 37.2°C. Scans were repeated three times, and the average for each rat was determined. Data were expressed as the ratio of ischemic to non-ischemic hindlimb.

Angiograms

Angiograms were performed 42 d after induction of ischemia. Barium sulfate (2.5 mL; EZPaqe, Merry X-Ray, South San Francisco, CA, USA) was infused into the infrarenal aorta after ligation of the proximal aorta and inferior vena cava during inhalation of 2% isofluorane. A grid was superimposed over the film between the greater trochanter of the femur to the patella. The number of intersections between contrast-filled vessels and gridlines was determined independently by three blinded observers. The angioscore was calculated as the average ratio of intersections to the total number of gridlines. Within the experimental setting of this study, the angioscore is a marker of collateral artery enlargement; thus, as collateral arteries dilate and remodel in response to femoral artery excision, their diameters increase, enhancing their visibility on the X-ray film, thus increasing the angioscore (19).

Nitroblue Tetrazolium Staining to Detect Muscle Necrosis

The left gastrocnemius muscle was removed 7 d after induction of ischemia. The timing of sacrifice was selected on the basis of previous work that demonstrated maximal postischemic muscle necrosis at this time (19). The muscle was cut transversely into three 2-mm sections. Two sections were used for nitroblue tetrazolium (NBT) staining, while the third was frozen (−80°C) in optimal cutting temperature (OCT) embedding compound. Sections for NBT staining were incubated in PBS containing 0.033% NBT (Fisher Biotech, Austin, TX, USA) and 0.133% NADH (Roche Diagnostics, Indianapolis, IN, USA) at 21°C for 10 min. The samples were then fixed in 4% paraformaldehyde for 24 h. The areas of viable tissue, indicated by dark blue color, and nonviable tissue, indicated by white color, were measured by quantitative image analysis (ImagePro; Media Cybernetics, Bethesda, MD, USA). Data were expressed as the ratio of nonviable tissue to total tissue area. Measurements were made on the four exposed cut surfaces, and the average was taken and used as a single data point for each animal. The frozen section was used to prepare cryosections (10 µm) for hematoxylin and eosin (H&E) staining to evaluate histological integrity of the muscle, as well as to detect the presence of an inflammatory infiltrate.

Statistical Analysis

Analyses were carried out by means of analysis of variance (ANOVA). Post hoc Student-Newman-Keuls tests were carried out if the ANOVA F statistic was significant to determine sites of difference within the ANOVA format. Probability values less <0.05 were accepted as significant for all statistical calculations.

Results

Effects of Oral BH4, L-Arginine, and Vitamin C on Gastrocnemius eNOS and p-eNOS Expression

Dietary supplementation of BH4, L-arginine and vitamin C significantly affected eNOS and p-eNOS expression in the ischemic gastrocnemius muscle (Figure 1). Rats given single supplementation with BH4 or L-arginine demonstrated similar levels of eNOS expression, and these levels were significantly greater than that of rats fed normal chow. Rats given two (BH4 + L-arginine) or three (BH4 + L-arginine + vitamin C) supplements also had greater levels of eNOS expression than rats given single supplements. Hence, the combination of BH4 and L-arginine had an additive effect on eNOS expression, and the addition of vitamin C provided an additional beneficial effect. However, increased p-eNOS expression was only observed in BH4 + L-arginine- and BH4 + L-arginine + vitamin C-fed rats.

Figure 1

Effects of BH4, L-arginine (L-arg) and vitamin C (VitC) on eNOS and phosphorylated eNOS expression in the ischemic gastrocnemius muscle. Muscle was harvested 14 d after the induction of hindlimb ischemia, and supernatants from muscle homogenates were used in these assays. (A) Expression of eNOS was increased by all dietary additives, although the greatest increase was noted in rats that received BH4 + L-arginine + vitamin C. (B) Expression of p-eNOS was increased in rats fed BH4 + L-arginine or BH4 + L-arginine + vitamin C. Control rats were fed a standard diet, whereas the four treatment groups consumed diets supplemented with additives noted in the bar graph. Concentrations of these additives are noted in the text. Data are mean ± standard deviation (sd); n = 5–6. *p < 0.05 versus control; p < 0.05 versus BH4 or L-arginine groups; ‡p < 0.05 versus BH4 + L-arginine group.

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Effects of Oral BH4, L-Arginine, and Vitamin C on Gastrocnemius Ca²⁺-Dependent NOS Activity

Dietary supplements affected Ca²⁺-dependent NOS activity in the ischemic gastrocnemius muscle (Figure 2A). Rats given a single dietary supplement (BH4 or L-arginine) demonstrated Ca²⁺-dependent NOS activities that were similar to each other and greater than that of rats fed normal chow. Rats given two (BH4 + L-arginine) or three (BH4 + L-arginine + vitamin C) supplements displayed significantly greater Ca²⁺-dependent NOS activity than rats given a single dietary supplement. The combination of BH4 and L-arginine generated an additive effect. The addition of vitamin C further increased Ca²⁺-dependent NOS activity, although it did not reach statistical significance.

Figure 2

Effects of BH4, L-arginine (L-arg) and vitamin C (VitC) on NOS activity in the ischemic gastrocnemius muscle. Muscle was harvested 14 d after the induction of hindlimb ischemia, and supernatants from muscle homogenates were used in these assays. (A) Ca²⁺-dependent NOS activity was increased by all dietary additives, although the greatest increase was noted in mice receiving BH4 + L-arginine or BH4 + L-arginine + vitamin C. Data are mean ± sd; n = 5–6. *p < 0.05 versus control; p < 0.05 versus BH4 or L-arginine groups; ‡p < 0.05 versus BH4 + L-arginine group. (B) NOx (nitrite + nitrate) was increased in mice receiving BH4 + L-arginine or BH4 + L-arginine + vitamin C. Data are mean ± sd; n = 5. *p < 0.05 versus control; p < 0.05 versus BH4 or L-arginine groups. (C) Ca²⁺-independent NOS activity was less in mice receiving BH4 + L-arginine or BH4 + L-arginine + vitamin C than in control mice. Data are mean ± sd; n = 5–6. *p < 0.05 versus control. In all graphs, control rats were fed normal chow and underwent hindlimb ischemia, whereas the four treatment groups consumed diets supplemented with additives noted in the bar graph. Concentrations of these additives are noted in the text.

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Effects of Oral BH4, L-Arginine and Vitamin C on Gastrocnemius NOx Levels

The final products of NO in vivo are nitrite (NO₂ ⁻) and nitrate (NO₃ ⁻). NOx is the sum of NO₂ ⁻ and NO₃ ⁻ and it is the best index of total NO production. The NOx concentration in ischemic gastrocnemius was significantly increased from rats fed BH4 and L-arginine or BH4, L-arginine and vitamin C (Figure 2B). There were no changes in rats fed with either BH4 and L-arginine alone.

Effects of Oral BH4, L-Arginine and Vitamin C on Gastrocnemius Ca²⁺-Independent NOS Activity

Ca²⁺-independent NOS activity was greater in the ischemic gastrocnemius from rats fed normal chow than in the ischemic gastrocnemius of dietary fed rats (Figure 2C). Rats fed a single supplement (BH4 or L-arginine), or the combination of these two agents, demonstrated Ca²⁺-independent NOS activity statistically similar to the normal chow group (that is, there was no beneficial effect noted in these dietary intervention groups). However, rats provided with three dietary supplements (BH4 + L-arginine + vitamin C) demonstrated Ca²⁺-independent NOS activity levels in the ischemic gastrocnemius muscle that were lower than the other dietary intervention groups.

Effects of Oral BH4, L-Arginine and Vitamin C on Gastrocnemius Oxidative Stress

Dietary supplementation affected nitrotyrosine accumulation and the ratio of GSH versus GSSG in the ischemic gastrocnemius muscle (Figure 3). Rats given a single dietary supplement (BH4 or L-arginine), or the combination of these dietary agents, displayed similar nitrotyrosine levels. These levels were significantly less than the level noted in rats fed normal chow. Rats provided with all three dietary supplements (BH4 + L-arginine + vitamin C) exhibited a nitrotyrosine level significantly less than rats given a single supplement (BH4 or L-arginine) or the combination of these two agents. The ratio of reduced versus oxidized glutathione (GSH:GSSG) was also measured as another index of oxidative stress in the ischemic gastrocnemius muscle. GSH:GSSG ratio was increased in the ischemic gastrocnemius of rats fed the two dietary combination (BH4 + L-arginine) or the three dietary combination (BH4 + L-arginine + vitamin C). Addition of vitamin C had a beneficial effect in increasing GSH:GSSG ratio. Taken together, three dietary supplementation (BH4 + L-arginine + vitamin C) would be much more effective in decreasing ischemic muscles oxidative stress after induction of hindlimb ischemia in rats.

Figure 3

Effects of BH4, L-arginine (L-arg) and vitamin C (VitC) on oxidative stress in the ischemic gastrocnemius muscle. Muscle was harvested 14 d after the induction of hindlimb ischemia, and supernatants from muscle homogenates were used in these assays. (A) Nitrotyrosine expression was decreased by all dietary additives, although this decrease was greatest in rats receiving BH4 + L-arginine + vitamin C. (B) The GSH:GSSG ratio was increased in rats receiving BH4 + L-arginine; the addition of vitamin C to this regimen caused an additional increase in this ratio. Data are mean ± sd; n = 5–6. *p < 0.05 versus control, BH4 or L-arginine groups; †p < 0.05 versus BH4 + L-arginine groups.

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Effects of BH4, L-Arginine and Vitamin C on Hindlimb Perfusion

Dietary supplementation significantly increased the recovery of hindlimb perfusion after induction of severe ischemia, and this effect was regimen and time dependent (Figure 4A). Rats given a single supplement (BH4 or L-arginine) demonstrated similar degree of perfusion recovery in the foot, and this level was also similar to that noted in rats fed normal chow. Rats provided with two (BH4 + L-arginine) or three (BH4 + L-arginine + vitamin C) supplements showed significantly greater recovery of foot perfusion than rats fed normal chow or rats given a single dietary supplement. This difference was evident at the later phase of recovery, on d 21, 28 or 42 after induction of ischemia, which was also the time of maximal collateral artery wall remodeling (20). A similar pattern was noted for collateral artery angioscores determined on d 42 after induction of ischemia. Hence, the angioscore was significantly greater in rats given two (BH4 + L-arginine) or three (BH4 + L-arginine + vitamin C) supplements than in rats fed normal chow or in rats given a single supplement (Figures 4B, C).

Figure 4

Effects of BH4, L-arginine and vitamin C on perfusion recovery after induction of hindlimb ischemia. (A) Laser Doppler perfusion imaging (LDPI) data, expressed as the ratio of blood flow from the ischemic to nonischemic hindlimbs, was determined before, immediately after and then serially over the ensuing 6 wks. Group identity is shown in the color key. Data are mean ± sd; n = 6. *p < 0.05 for BH4 + L-arginine or BH4 + L-arginine + vitamin C groups versus all other groups. (B) The angioscore, determined 42 d after the induction of hindlimb ischemia and calculated as described in the text, was greater in rats receiving BH4 + L-arginine or BH4 + L-arginine + vitamin C than in all other groups. Data are mean ± sd; n = 6. *p < 0.05 for BH4 + L-arginine or BH4 + L-arginine + vitamin C groups versus all other groups. (C) Representative angiogram from each study group.

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Effects of Oral BH4, L-Arginine and Vitamin C on Gastrocnemius Muscle Necrosis

The extent of gastrocnemius necrosis was affected by the provision of dietary supplements. Rats given a single supplement (BH4 or L-arginine) or the combination of these agents manifest a similar degree of gastrocnemius muscle necrosis. Moreover, the extent of necrosis noted in these dietary intervention groups was similar to that noted in rats fed normal chow (that is, these dietary regimens did not improve postischemic muscle integrity). However, rats provided with all three dietary supplements (BH4 + L-arginine + vitamin C) had significantly less gastrocnemius necrosis than rats fed normal chow, rats provided with a single dietary supplement or rats given the combination of BH4 + L-arginine. This difference was evident on macroscopic and microscopic levels. The percentage of the cut surface of the ischemic gastrocnemius muscle that was necrotic, determined by NBT staining, was significantly less in the BH4 + L-arginine + vitamin C group than in all other groups (Figures 5A, B). Groups fed normal chow, or supplemented with BH4, L-arginine, or both agents, demonstrated similar histological evidence of severe necrosis: muscle nuclei were nearly absent, intramyofiber vacuolization was substantial and the distance between myofibers was large (Figure 5C). A pronounced inflammatory infiltrate was also present in these groups. In contrast, the BH4 + L-arginine + vitamin C group demonstrated good preservation of muscle histology and only a limited inflammatory cell infiltrate.

Figure 5

The effects of BH4, L-arginine (L-arg) and vitamin C (VitC) on necrosis in the ischemic gastrocnemius muscle. (A) Gastrocnemius muscle was removed 7 d after induction of hindlimb ischemia. Transverse sections of this muscle were stained with nitroblue tetrazolium to determine the ratio of necrotic versus viable surface area. This ratio was less in rats receiving BH4 + L-arginine + vitamin C than in all other groups. Data are mean ± sd; n = 6. *p < 0.05 for BH4 + L-arginine + vitamin C groups versus all other groups. (B) Representative images from each study group. (C) Histological sections of gastrocnemius muscle taken from the ischemic hindlimb 7 d after the induction of hindlimb ischemia were stained with H&E. Representative sections are shown. Note the loss of myofibers and the intense cellular infiltrate in the control, BH4 and L-arginine groups. Best preservation of muscle architecture was consistently observed in rats receiving BH4 + L-arginine + vitamin C.

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Discussion

The following study hypothesis was supported by our findings: dietary co-supplementation with tetrahydrobiopterin (BH4), L-arginine and vitamin C acts synergistically to decrease oxidative stress, increase nitric oxide and thereby improve perfusion and tissue recovery in response to acute hindlimb ischemia more than supplementation with a single supplement. Interestingly, two patterns of effect emerged. Cosupplementation with BH4 + L-arginine increased the dependent variables eNOS and p-eNOS expression, Ca²⁺-dependent NOS activity, foot perfusion and the collateral artery angioscore more than the addition of either component separately, whereas the addition of vitamin C provided a further beneficial effect on these variables, although only eNOS expression reached statistical significance. In addition, co-administration of all three dietary supplements had a significantly greater effect than BH4 or L-arginine, given individually or in combination, when the dependent variables of Ca²⁺-independent NOS activity, oxidative stress or muscle necrosis were measured.

eNOS and p-eNOS expression, Ca²⁺-dependent NOS activity, foot perfusion and the collateral artery angioscore are linked by established cause-and-effect relationships. eNOS-derived NO is a potent vasodilator (2); hence, the increased eNOS expression and activity present in the BH4 + L-arginine group should result in an NO-dependent increase in foot perfusion, and this expectation was realized by our findings. Moreover, eNOS-derived NO is a critical determinant in the response to hindlimb ischemia (21–23). This effect is direct, on the basis of the vasodilator effect of NO (24), but also indirect, insofar as eNOS-derived NO is critical to collateral artery remodeling. These effects include mobilization of endothelial progenitor cells from bone marrow and their subsequent homing to the ischemic hindlimb (25). Once there, endothelial progenitor cells participate in postischemic arteriogenesis, the process wherein existing collateral arteries undergo remodeling designed to restore vascular conductance (26). This process was evidenced by the increased angioscore, a marker of collateral artery enlargement, in rats provided with L-arginine + BH4 dietary supplements.

Vitamin C likely exerted its beneficial effects in this study through a variety of molecular mechanisms. In its capacity as an antioxidant, it enhances NO bioavailability by quenching O₂ ⁻, thus limiting the inactivation of NO that occurs when O₂ ⁻ and NO combine to produce OONO⁻ (3). Vitamin C also stabilizes existing BH4 (8) and increases endothelial BH4 synthesis (27), thus minimizing eNOS "uncoupling," which in turn lessens generation of O₂ ⁻ by eNOS and reduces vascular oxidative stress (7). However, BH4 is itself a potent antioxidant (7), and administration of exogenous BH4 has been established to increase endothelial BH4 levels (28). Moreover, L-arginine directly stimulates eNOS expression (29), enhances eNOS activity by a receptor-dependent G protein-linked process (30) and limits the inhibitory effect of asymmetric dimethylarginine on eNOS-derived NO production (31). We propose that under the experimental conditions imposed by hindlimb ischemia, addition of vitamin C to BH4 + L-arginine significantly decreased oxidative stress and accordingly decreased tissue necrosis. It may also restore BH4 or NO levels, since we observed an increase in eNOS activity and p-eNOS expression, although it did not reach statistical difference because of the number of samples analyzed, dosage of vitamin C or time of data collection. Cosupplementation with BH4 + L-arginine may have sufficiently restored endothelial BH4 levels and thereby intracellular redox balance. Hence, the addition of vitamin C did not further p-eNOS expression or activity, and hence NO bioavailability.

Ca²⁺-independent NOS activity and tissue nitrotyrosine accumulation were significantly lower in rats receiving all three dietary supplements than in rats receiving BH4 or L-arginine, or the combination of the two. When measured by methods used herein, Ca²⁺-independent NOS activity is an authentic reflection of iNOS activity, inasmuch as the assay was conducted in vitro, in the absence of shear stress, which can activate eNOS in the absence of Ca²⁺ via phosphorylation (32). The marked elevation of iNOS activity in rats fed normal chow indicates the presence of postischemia inflammation, which is also evidenced by the cellular inflammatory infiltrate in this group. Nitrotyrosine accumulation is indicative of OONO⁻-induced damage (33), and it is interesting that the group that exhibited the least amount of tissue damage (rats provided with all three supplements) also had the least nitrotyrosine accumulation. Moreover, rats in the triple therapy group demonstrated a virtual absence of nitrotyrosine. We interpret the present findings to indicate that muscle injury after ischemia is not entirely contingent upon loss of perfusion, inasmuch as cosupplementation with the antioxidant vitamin C clearly reduced tissue injury and eliminated nitrotyrosine accumulation, but did not affect perfusion or collateral artery enlargement. Instead, we speculate that vitamin C provided an antioxidant effect that limited tissue injury generated by inflammatory cells for which action depends in part on oxidant production (for example, neutrophils and macrophages). This effect could be direct, due to the antioxidant activity of vitamin C, or indirect, due to the beneficial effect of vitamin C on BH4 levels (8,27), insofar as BH4 also exhibits potent antioxidant activity (7).

Although it is well established that endothelial dysfunction related to vascular oxidative stress is a critical factor in PAD pathogenesis, dietary supplementation with L-arginine or antioxidants, such as vitamin C, have had equivocal effects on long-term outcome (12,13,16). Dietary supplementation with L-arginine alone has a beneficial effect when given acutely (that is, via intravenous infusion [14] or for short duration [2 months] [15]), and these clinical results are consistent with the positive effects observed in rats provided with dietary L-arginine. However, long-term administration of L-arginine (6 months) not only failed to demonstrate a beneficial effect, but resulted in a degree of eNOS-dependent vascular reactivity significantly less than that of the placebo group (16). The present findings demonstrated increased iNOS activity after induction of ischemia. If a similar circumstance is present in PAD, then the singular dietary supplementation with L-arginine, the substrate for all NOS isoforms, might serve to worsen vascular inflammation, a critical participant in the pathogenesis of PAD (1). Vitamin C reduces vascular inflammation (34), improves redox balance (11) and eNOS-dependent vascular reactivity (10), but these effects have only been evaluated on a short-term basis, whereas retrospective cross-sectional studies have failed to confirm that dietary supplementation with antioxidants improves PAD outcome (12,13). We interpret the present findings to indicate that provision of BH4 + L-arginine + vitamin C acting synergystically might prove a useful therapeutic alternative in PAD treatment. To this end, the use of sapropterin dihydrochloride, a synthetic form of (6R)-L-erythro-5,6,7,8-tetrahydrobiopterin recently approved for the treatment of phenylalanine hydroxylase deficiency (35), might provide a practical means for the provision of BH4.

In conclusion, cosupplementation with BH4 + L-arginine + vitamin C resulted in increased eNOS activity and NO concentration as well as greater foot blood flow recovery than rats receiving normal chow or either agent separately. The addition of vitamin C to the BH4 + L-arginine regimen further reduced oxidative stress and tissue injury in ischemic muscles (Figure 6). The clearly superior outcome of rats provided with BH4 + L-arginine + vitamin C warrants investigation of a cosupplementation strategy as a therapeutic alternative in PAD.

Figure 6

Schematic illustration of possible mechanism of three dietary combined regimen can increase NO bioavailability and decrease oxidative stress, accordingly increase blood flow recovery and reduce tissue necrosis. A⁻*, ascorbate; AH⁻, dehydroascorbate.

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Disclosure

The authors declare that they have no competing interests as defined by Molecular Medicine, or other interests that might be perceived to influence the results and discussion reported in this paper.

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Acknowledgments

This work was supported by National Institutes of Health, National Heart, Lung, and Blood Institute grant RO-1 HL-75353 (to LM Messina), as well as grants from the Pacific Vascular Research Foundation and the Wayne and Gladys Valley Foundation (to LM Messina).

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Affiliations

1. Department of Surgery, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, Massachusetts, 01655, USA

   Jinglian Yan, Guodong Tie & Louis M. Messina

Corresponding author

Correspondence to Louis M. Messina.

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Yan, J., Tie, G. & Messina, L.M. Tetrahydrobiopterin, L-Arginine and Vitamin C Act Synergistically to Decrease Oxidative Stress, Increase Nitric Oxide and Improve Blood Flow after Induction of Hindlimb Ischemia in the Rat. Mol Med 18, 676–684 (2012). https://doi.org/10.2119/molmed.2011.00103

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• Received: 22 March 2011

• Accepted: 21 February 2012

• Published: 24 February 2012

• Issue Date: April 2012

• DOI : https://doi.org/10.2119/molmed.2011.00103

Keywords

• Hindlimb Ischemia
• Decrease Oxidative Stress
• Increase Nitric Oxide
• Nitrotyrosine Accumulation
• eNOS Expression

L Arginine And Vitamin C

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Is There Vitamin C In Breast Milk

Is There Vitamin C In Breast Milk

A Guide to Vitamin C Serums

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For 70 years Vitamin C has been one of the biggest weapons in the skin care industry. It's used to make cleansers, moisturizers, lotions, masks, and serums. So what is this powerful vitamin? How can it benefit you? Why should you use serums that contain Vitamin C? We're here to answer all of those burning questions in this complete guide and reveal the many benefits Vitamin C serums offer for your skin.

What Are Vitamin C Serums?

There are many variations of Vitamin C, but the most popular is ascorbic acid, a common ingredient in skincare products. However, all the variations of Vitamin C have anti-inflammatory benefits.

Vitamin C Serums are products that contain a high level of Vitamin C. They're used to treat wrinkles, sagging skin, lighten dark or red spots, prevent breakouts and even your skin tone. Basically, they battle all of those annoying skin issues. There are loads of products out there that claim they'll save your skin, but some can cause damage. Vitamin C is one of the most revered ingredients and conclusive research has shown how effective it can be.

How Are Vitamin C Serums Made?

Well so far we've talked about how incredible Vitamin C is and it sounds like a dream come true doesn't it? It's not all good. In fact, the mighty vitamin is unstable when it's exposed to air and light. Other ingredients need to be used to stabilize it and allow it to deliver amazing results. The serums are combined with ferulic acid and Vitamin E. According to researchers the perfect mixture is 15% Vitamin C with 1% Vitamin E and 0.5% ferulic acid. This makes Vitamin C perform to the best of its abilities, without damaging your skin.

What Does Vitamin C Serum Do for Your Face?

Boosts Collagen Production: Collagen keeps your skin firm and prevents sagging. Environmental factors such as lifestyle choices and pollution can increase the elasticity of your skin, so it's important you try to increase your collagen production.

Hydrates Your Skin: Dry skin is a common issue, but Vitamin C can help to give your skin that much-needed moisture boost. Remember, it doesn't work immediately so you need to keep applying the serum to see results.

Brightens Your Complexion: Dark spots on your skin are caused by the overproduction of melanin. Vitamin C decreases the production and lightens the dark spots to even out your complexion.

Reduces Redness and Inflammation: Conditions such as Rosacea leave many people searching for a magic cure. Vitamin C helps facial redness and inflammation by reducing the appearance of broken capillaries.

Why Should You Be Using Vitamin C Serum?

Don't think Vitamin C serums are just beneficial for your face. They can also shield you from sun damage and reduce stretch marks.

Saves You From The Sun: Prolonged exposure to UV rays can cause long-term damage to your skin. Luckily, Vitamin C is a powerful antioxidant that reduces red sports and prevents sunburn from spreading.

Fades Acne Scars: Acne plagues most of us at some point and we look forward to being free. Most of us get stuck with some scars but Vitamin C helps to fade scars and even out any discoloration.

Reduces Stretch Marks: Yes, Vitamin C serum can even help prevent those unsightly stretch marks by tightening your skin.

Are you ready to change your skin? Add a Vitamin C serum to your beauty regimen and enjoy a glowing complexion.

Is There Vitamin C In Breast Milk

Source: https://www.bloglines.com/article/a-guide-to-vitamin-c-serums?utm_content=params%3Ao%3D740010%26ad%3DdirN%26qo%3DserpIndex

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How Many Vitamin C Pills Should I Take A Day

How Many Vitamin C Pills Should I Take A Day

This is a reader-friendly overview of Vitamin C. For more details, see our health professional fact sheet on Vitamin C.

For information on vitamin C and COVID-19, see Dietary Supplements in the Time of COVID-19.

What is vitamin C and what does it do?

Vitamin C, also known as ascorbic acid, is a water-soluble nutrient found in some foods. In the body, it acts as an antioxidant, helping to protect cells from the damage caused by free radicals. Free radicals are compounds formed when our bodies convert the food we eat into energy. People are also exposed to free radicals in the environment from cigarette smoke, air pollution, and ultraviolet light from the sun.

The body also needs vitamin C to make collagen, a protein required to help wounds heal. In addition, vitamin C improves the absorption of iron from plant-based foods and helps the immune system work properly to protect the body from disease.

How much vitamin C do I need?

The amount of vitamin C you need each day depends on your age. Average daily recommended amounts for different ages are listed below in milligrams (mg).

Life Stage	Recommended Amount
Birth to 6 months	40 mg
Infants 7–12 months	50 mg
Children 1–3 years	15 mg
Children 4–8 years	25 mg
Children 9–13 years	45 mg
Teens 14–18 years (boys)	75 mg
Teens 14–18 years (girls)	65 mg
Adults (men)	90 mg
Adults (women)	75 mg
Pregnant teens	80 mg
Pregnant women	85 mg
Breastfeeding teens	115 mg
Breastfeeding women	120 mg

If you smoke, add 35 mg to the above values to calculate your total daily recommended amount.

What foods provide vitamin C?

Fruits and vegetables are the best sources of vitamin C. You can get recommended amounts of vitamin C by eating a variety of foods including the following:

• Citrus fruits (such as oranges and grapefruit) and their juices, as well as red and green pepper and kiwifruit, which have a lot of vitamin C.
• Other fruits and vegetables—such as broccoli, strawberries, cantaloupe, baked potatoes, and tomatoes—which also have vitamin C.
• Some foods and beverages that are fortified with vitamin C. To find out if vitamin C has been added to a food product, check the product labels.

The vitamin C content of food may be reduced by prolonged storage and by cooking. Steaming or microwaving may lessen cooking losses. Fortunately, many of the best food sources of vitamin C, such as fruits and vegetables, are usually eaten raw.

What kinds of vitamin C dietary supplements are available?

Most multivitamins have vitamin C. Vitamin C is also available alone as a dietary supplement or in combination with other nutrients. The vitamin C in dietary supplements is usually in the form of ascorbic acid, but some supplements have other forms, such as sodium ascorbate, calcium ascorbate, other mineral ascorbates, and ascorbic acid with bioflavonoids. Research has not shown that any form of vitamin C is better than the other forms.

Am I getting enough vitamin C?

Most people in the United States get enough vitamin C from foods and beverages. However, certain groups of people are more likely than others to have trouble getting enough vitamin C:

• People who smoke and those who are exposed to secondhand smoke, in part because smoke increases the amount of vitamin C that the body needs to repair damage caused by free radicals. People who smoke need 35 mg more vitamin C per day than nonsmokers.
• Infants who are fed evaporated or boiled cow's milk, because cow's milk has very little vitamin C and heat can destroy vitamin C. Cow's milk is not recommended for infants under 1 year of age. Breast milk and infant formula have adequate amounts of vitamin C.
• People who eat a very limited variety of food.
• People with certain medical conditions such as severe malabsorption, some types of cancer, and kidney disease requiring hemodialysis.

What happens if I don't get enough vitamin C?

Vitamin C deficiency is rare in the United States and Canada. People who get little or no vitamin C (below about 10 mg per day) for many weeks can get scurvy. Scurvy causes fatigue, inflammation of the gums, small red or purple spots on the skin, joint pain, poor wound healing, and corkscrew hairs. Additional signs of scurvy include depression as well as swollen, bleeding gums and loosening or loss of teeth. People with scurvy can also develop anemia. Scurvy is fatal if it is not treated.

What are some effects of vitamin C on health?

Scientists are studying vitamin C to understand how it affects health. Here are several examples of what this research has shown.

Cancer prevention and treatment

People with high intakes of vitamin C from fruits and vegetables might have a lower risk of getting many types of cancer, such as lung, breast, and colon cancer. However, taking vitamin C supplements, with or without other antioxidants, doesn't seem to protect people from getting cancer.

It is not clear whether taking high doses of vitamin C is helpful as a treatment for cancer. Vitamin C's effects appear to depend on how it is administered to the patient. Oral doses of vitamin C can't raise blood levels of vitamin C nearly as high as intravenous doses given through injections. A few studies in animals and test tubes indicate that very high blood levels of vitamin C might shrink tumors. But more research is needed to determine whether high-dose intravenous vitamin C helps treat cancer in people.

Vitamin C dietary supplements and other antioxidants might interact with chemotherapy and radiation therapy for cancer. People being treated for cancer should talk with their oncologist before taking vitamin C or other antioxidant supplements, especially in high doses.

Cardiovascular disease

People who eat lots of fruits and vegetables seem to have a lower risk of cardiovascular disease. Researchers believe that the antioxidant content of these foods might be partly responsible for this association because oxidative damage is a major cause of cardiovascular disease. However, scientists aren't sure whether vitamin C itself, either from food or supplements, helps protect people from cardiovascular disease. It is also not clear whether vitamin C helps prevent cardiovascular disease from getting worse in people who already have it.

Age-related macular degeneration (AMD) and cataracts

AMD and cataracts are two of the leading causes of vision loss in older people. Researchers do not believe that vitamin C and other antioxidants affect the risk of getting AMD. However, research suggests that vitamin C combined with other nutrients might help slow AMD progression.

In a large study among older people with AMD who were at high risk of developing advanced AMD, those who took a daily dietary supplement with 500 mg vitamin C, 80 mg zinc, 400 IU vitamin E, 15 mg beta-carotene, and 2 mg copper for about 6 years had a lower chance of developing advanced AMD. They also had less vision loss than those who did not take the dietary supplement. People who have or are developing the disease might want to talk with their doctor about taking dietary supplements.

The relationship between vitamin C and cataract formation is unclear. Some studies show that people who get more vitamin C from foods have a lower risk of getting cataracts. But further research is needed to clarify this association and to determine whether vitamin C supplements affect the risk of getting cataracts.

The common cold

Although vitamin C has long been a popular remedy for the common cold, research shows that for most people, vitamin C supplements do not reduce the risk of getting the common cold. However, people who take vitamin C supplements regularly might have slightly shorter colds or somewhat milder symptoms when they do have a cold. Using vitamin C supplements after cold symptoms start does not appear to be helpful.

Can vitamin C be harmful?

Taking too much vitamin C can cause diarrhea, nausea, and stomach cramps. In people with a condition called hemochromatosis, which causes the body to store too much iron, high doses of vitamin C could worsen iron overload and damage body tissues.

The daily upper limits for vitamin C include intakes from all sources—food, beverages, and supplements—and are listed below:

Life Stage	Upper Limit
Birth to 12 months	Not established
Children 1–3 years	400 mg
Children 4–8 years	650 mg
Children 9–13 years	1,200 mg
Teens 14–18 years	1,800 mg
Adults	2,000 mg

Does vitamin C interact with medications or other dietary supplements?

Vitamin C dietary supplements can interact or interfere with medicines that you take. Here are several examples:

• Vitamin C dietary supplements might interact with cancer treatments, such as chemotherapy and radiation therapy. It is not clear whether vitamin C might have the unwanted effect of protecting tumor cells from cancer treatments or whether it might help protect normal tissues from getting damaged. If you are being treated for cancer, check with your healthcare provider before taking vitamin C or other antioxidant supplements, especially in high doses.
• In one study, vitamin C plus other antioxidants (such as vitamin E, selenium, and beta-carotene) reduced the heart-protective effects of two drugs taken in combination (a statin and niacin) to control blood-cholesterol levels. It is not known whether this interaction also occurs with other statins. Healthcare providers should monitor lipid levels in people taking both statins and antioxidant supplements.

Tell your doctor, pharmacist, and other healthcare providers about any dietary supplements and medicines you take. They can tell you if those dietary supplements might interact or interfere with your prescription or over-the-counter medicines or if the medicines might interfere with how your body absorbs, uses, or breaks down nutrients.

Vitamin C and healthful eating

People should get most of their nutrients from food and beverages, according to the federal government's Dietary Guidelines for Americans. Foods contain vitamins, minerals, dietary fiber and other components that benefit health. In some cases, fortified foods and dietary supplements are useful when it is not possible to meet needs for one or more nutrients (e.g., during specific life stages such as pregnancy). For more information about building a healthy dietary pattern, see the Dietary Guidelines for Americans and the U.S. Department of Agriculture's MyPlate.

Where can I find out more about vitamin C?

Disclaimer

This fact sheet by the Office of Dietary Supplements (ODS) provides information that should not take the place of medical advice. We encourage you to talk to your healthcare providers (doctor, registered dietitian, pharmacist, etc.) about your interest in, questions about, or use of dietary supplements and what may be best for your overall health. Any mention in this publication of a specific product or service, or recommendation from an organization or professional society, does not represent an endorsement by ODS of that product, service, or expert advice.

How Many Vitamin C Pills Should I Take A Day

Source: https://ods.od.nih.gov/factsheets/VitaminC-Consumer/

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Green Apple Vitamin C Content

Green Apple Vitamin C Content

Researchers in the United Kingdom say a study on eyesight shows diet and environmental factors are more important than genetics in lowering risk of cataracts.

Eating an apple a day may keep the doctor away, but eating oranges might do the same for cataracts.

In a study published today in the journal Ophthalmology, researchers in the United Kingdom said a higher dietary intake of vitamin C might significantly reduce the risk of developing cataracts.

The study, led by scientists at King's College London, is the first to conclude that dietary and environmental factors play a larger role than genetics in the development of cataracts.

"The findings of this study could have significant impact, particularly for the aging population globally by suggesting that simple dietary changes such as increased intake of fruits and vegetables as part of a healthier diet could help protect them from cataracts," Dr. Chris Hammond, professor of ophthalmology at King's College, consultant eye surgeon and lead author of the study, said in a statement.

Read More: What Is a Cataract? »

The researchers estimated genetic factors account for 35 percent of the difference in cataract progression. Environment and lifestyle account for 65 percent.

To study the impact diet has on cataracts, the researchers tracked the progression of the eye condition in 324 pairs of female twins from the United Kingdom.

The scientists examined digital images of the women's eye lenses when they were about 60 years old. They then studied the same type of images 10 years later.

They kept track of the women's intake of vitamins A, B, C, D, and E. They also tracked their intake of copper, manganese, and zinc using a food questionnaire.

The researchers said the women who ingested more vitamin C initially had a 20 percent reduced risk of developing cataracts. After 10 years, that risk had decreased by 33 percent.

The researchers noted that there was little risk reduction in the women who took vitamin supplements. Instead the preventative effects appeared to be obtained only by eating foods rich in vitamin C.

Dr. Ravi D. Goel, an ophthalmologist from New Jersey who is also a clinical instructor at Wills Eye Hospital in Pennsylvania, said the study provides helpful information for patients and doctors.

"These are novel findings for patients going forward," Goel, a spokesperson for the American Academy of Ophthalmology, told Healthline. "This is a helpful tool for patient education."

Read More: Americans Spend Billions on Vitamins and Supplements That Don't Work »

Cataracts occur when the lens of the eye becomes cloudy due to oxidation over a long period of time.

The researchers said the fluids that bathe the eye are rich in vitamin C, which helps stop the lens from oxidizing.

The dietary intake of vitamin C helps prevent cataracts by increasing the amount of this vitamin in the eye fluid.

The researchers added that smoking and diabetes also are risk factors for certain kinds of cataracts, so a balanced diet and healthy lifestyle are important.

"Healthy diets are always an advantage for patients," added Goel.

Goel also said vitamin C has already been shown to help slow the progression of age-related macular degeneration.

This latest information on cataracts adds to vitamin C's attributes. "It helps overall eye health," he said.

The researchers did note that their observational study has its limitations as it only involved women who were aged 60 years and older.

However, the researchers believe the information could also be relevant for male patients.

Cataracts are the leading cause of blindness in the world, affecting about 20 million people, according to statistics from the World Health Organization (WHO). Cataracts also affect 24 million Americans over the age of 40.

The condition can cause blurry vision, glare, poor night vision, and sensitivity to light.

Initially, better lighting and glasses may help ease some of the symptoms, but as cataracts progress surgery is sometimes needed.

Read More: Diabetes and Blurry Vision: What You Need to Know »

Green Apple Vitamin C Content

Source: https://www.healthline.com/health-news/vitamin-c-may-reduce-risk-of-cataracts

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Fermented Sauerkraut Vitamin C

Fermented Sauerkraut Vitamin C

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Fermented Sauerkraut Kept Raw

The Saving Essentials of Raw Fermented Sauerkraut

Ships carried precious cargo but scurvy spoiled the sailors. Death at sea was overwhelming. What could be the answer? Raw fermented sauerkraut with an abundance of vitamin C and digestive enzymes? Raw sauerkraut offers what for our health?

Sauerkraut (the German term for sour cabbage) is indeed considered to have been a real factor in overcoming the devastating and deathly disease of scurvy. What are the secrets of this super food? Does its vitality hold important nutrients for us today?

Vitamin C was deficient in the foods the sailors were eating. That was a major cause of the scurvy issue. Sauerkraut definitely made a deference. You see, fermenting cabbage escalates the vitamin C and antioxidant levels seriously. Studies show that there can be up to a 20 or more times increase of vitamin C resulting from this fermentation.

Common colds and flu prevention is often pursued by a better supply of vitamin C. Why choke down those extra boluses of supplements? Just eat real raw food full of it! Prepare your body for the challenges of the viruses long before they present themselves. Sauerkraut – savvy prevention against the common cold.

Fermented Sauerkraut Must Be Raw for Real Values

So is just eating sauerkraut the answer? Is all sauerkraut of equal value? You can pick it up off the canned goods isle in the supermarket, find it at the local orchard sealed in a glass jar, or pick up a pack out of the freezer at the local butcher shop. What's the best? This is a million dollar question when thinking of health benefits associated with sauerkraut.

First of all, remember that real uncooked kraut is a living food. If it is cooked, it has been heated and sterilized, thus killing the digestive enzymes and life of the kraut. It still will taste good but will lack the great stash of probiotics found in the raw, fermented uncooked form.

Another important fact to consider in your selection of sauerkraut is whether it contains vinegar. Vinegar-made kraut is not fermented and will not give you probiotics. But raw food is living food. Dine in the delight of the delicate vitality of small-farmed, raw, local, and lively!

Eat Raw Fermented Sauerkraut — Not Capsules of Probiotics

Raw, fermented kraut contains up to 100 times more probiotics than a probiotic supplement. But remember, you must be select unpasteurized, raw, uncooked, sauerkraut to get these good bugs for your body's health.

Research tells us that consuming it may prevent cancer. Furthermore, the probiotics in the sauerkraut attack cancer cells. A few spoonsful each day could certainly be worthy of consideration for a daily diet.

Enzyme Rich Foods Provide the Most

Raw sauerkraut is full of viable digestive enzymes. Enzyme rich foods help us digest our food and get the most vital nutrients from it. Digestive enzymes are valuable for our health in reducing inflammation and relieving pain. And so, eat enzyme rich foods; not dead pills.

Uncooked Sauerkraut for Better Vitamin B12 Assimilation

Vitamin B12 is so important for our nervous systems. But vitamin B12 assimilation into our bodies can be a problem. Low levels of this important vitamin to start with may be because we try to ingest dead pills instead of eating living food. We must have proper enzymes such as intrinsic factor for vitamin B12 assimilation into our bodies. And biotin is a B vitamin that helps with vitamin B12 assimilation and other B vitamins. Biotin is produced in the fermentation process; another reason to eat raw, fermented, uncooked kimchi.

This is also a good source of Vitamin K which is needful for quality bone health. Besides the vitamin K source, it is rich in calcium. Magnesium is also present and is quality for bone building.

A diet for weight loss should include some sauerkraut. When you need a snack, spoon out a bit of it. It can satisfy your hunger.

Raw Sauerkraut. Full of bioavailable nutrients from Raw Fermentation. Real food with digestive enzymes. Those are important facts to keep in mind when preparing food. Don't settle for the sterilized, lifeless forms of it. Vive in Raw Vitality!

Look for Swiss Villa Raw Sauerkraut

Swiss Villa wishes to thank you for considering goods offered via our food hub. We desire to assist small farms and start-up businesses to be sustainable by assisting them in marketing their high quality, nutrient rich foods. We understand uncooked kraut as we offer is one of the enzyme rich foods that can provide consumers with healthful eating. Thanks for your support.

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Fermented Sauerkraut Vitamin C

Source: https://www.swissvillallc.com/raw-sauerkraut/

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