ascorbic-acid and sodium-nitrate

X-ray-based techniques are a powerful tool in structural biology but the radiation-induced chemistry that results can be detrimental and may mask an accurate structural understanding. In the crystallographic case, cryocooling has been employed as a successful mitigation strategy but also has its limitations including the trapping of non-biological structural states. Crystallographic and solution studies performed at physiological temperatures can reveal otherwise hidden but relevant conformations, but are limited by their increased susceptibility to radiation damage. In this case, chemical additives that scavenge the species generated by radiation can mitigate damage but are not always successful and the mechanisms are often unclear. Using a protein designed to undergo a large-scale structural change from breakage of a disulfide bond, radiation damage can be monitored with small-angle X-ray scattering. Using this, we have quantitatively evaluated how three scavengers commonly used in crystallographic experiments - sodium nitrate, cysteine, and ascorbic acid - perform in solution at 10°C. Sodium nitrate was the most effective scavenger and completely inhibited fragmentation of the disulfide bond at a lower concentration (500 µM) compared with cysteine (∼5 mM) while ascorbic acid performed best at 5 mM but could only reduce fragmentation by ∼75% after a total accumulated dose of 792 Gy. The relative effectiveness of each scavenger matches their reported affinities for solvated electrons. Saturating concentrations of each scavenger shifted fragmentation from first order to a zeroth-order process, perhaps indicating the direct contribution of photoabsorption. The SAXS-based method can detect damage at X-ray doses far lower than those accessible crystallographically, thereby providing a detailed picture of scavenger processes. The solution results are also in close agreement with what is known about scavenger performance and mechanism in a crystallographic setting and suggest that a link can be made between the damage phenomenon in the two scenarios. Therefore, our engineered approach might provide a platform for more systematic and comprehensive screening of radioprotectants that can directly inform mitigation strategies for both solution and crystallographic experiments, while also clarifying fundamental radiation damage mechanisms.

Topics: Ascorbic Acid; Crystallography, X-Ray; Cysteine; Disulfides; Free Radical Scavengers; Molecular Structure; Nitrates; Scattering, Small Angle; Solutions; Temperature

Forty-two New Zealand White male rabbits were housed individually in wire cages and randomly distributed among six experimental groups of seven rabbits each, during 16 to 61 weeks of age. There were three main nitrate groups: 0 (tap water), 350 and 700 ppm. Within the 700 ppm of nitrate, there were four subgroups, in which one group was used as control group and the other three groups were supplemented with either 200 ppm of ascorbic acid (vitamin (Vit) C), 200 ppm of Vit E with 0.2 ppm of selenium (Se) and 1000 ppm of probiotic. The nitrate was supplemented as a sodium nitrate. The aim is to test the ability of Vit C and Vit E, Se and probiotic on the deleterious effects (blood and seminal plasma biochemical constituents, semen quality and productive performance) of nitrate in drinking water. Rabbits given nitrate at 700 ppm had significantly lower plasma globulin, red blood cells (RBCs), hemoglobin (Hgb), packed cell volume % (PCV%) and total antioxidant capacity (TAC) than those given the other concentrations of nitrate. Vit C, Vit E with Se and probiotic resulted in significantly (P < 0.05) greater Hgb, RBCs, PCV% and TAC than those of bucks given water supplemented with only 700 ppm nitrate, but the aspartate aminotransferase and alanine aminotransferase concentrations in seminal plasma were lower. Testosterone in the blood plasma and the seminal plasma was significantly (P < 0.05) lower in rabbits given 700 ppm nitrate than in those given other concentrations of nitrate. Vit C, Vit E with Se and the probiotic significantly increased testosterone, fertility, number of offspring and total offspring weight of rabbits sired by bucks supplemented with 700 ppm of nitrate.

Topics: Animals; Antioxidants; Ascorbic Acid; Dietary Supplements; Dose-Response Relationship, Drug; Drinking Water; Hematologic Tests; Inactivation, Metabolic; Male; Nitrates; Probiotics; Rabbits; Reproduction; Selenium; Semen; Semen Analysis; Sperm Count; Sperm Motility; Testis; Vitamin E

A new strategy was explored for the visual determination of Cu(2+) using copper-catalysed in situ formation of Ag nanoparticles. In this method, only common reagents were used and the pre-synthesis and modification of nanoparticles are avoided. Ag(+) can form a milk-white suspension (AgBr) with Br(-) in an aqueous solution composed of AgNO(3), cetyltrimethylammonium bromide, ascorbic acid, bovine serum albumin, and NaNO(3). The reaction will be stopped by addition of Cu(2+), accompanied by a colour change from milk-white to orange or brilliant yellow. Cu(+) (the reduction product of Cu(2+)) consumes the dissolved O(2) and prevents the O(2) from oxidizing the newly reduced Ag atoms (by ascorbic acid) back to Ag(+), facilitating the further aggregation of Ag atoms to become Ag nanoparticles. The visible colour change was shown to be specific towards Cu(2+) over most other metal ions. The limit of detection was 0.75 μM Cu(2+) by the naked eye and 0.25 μM by spectrometer. Quantitation of Cu(2+) was achieved in a linear range from 0.25 to 2.0 μM. This method was validated by measuring real water and serum samples, giving results agreeing well with the data reported and measured by inductively coupled plasma mass spectrometry. The recovery was 95.6-106% for untreated tap water and 96.0-100% for resin-pre-treated water and serum samples.

Topics: Animals; Ascorbic Acid; Catalysis; Cattle; Cetrimonium; Cetrimonium Compounds; Copper; Ions; Mass Spectrometry; Metal Nanoparticles; Nitrates; Serum Albumin, Bovine; Silver; Temperature; Water

The likely protective effects of nitric oxide (NO) against ammonium toxicity were investigated in the submerged macrophyte Hydrilla verticillata. The plants were subjected to ammonium stress (3mM ammonium chloride) in the presence of sodium nitroprusside (SNP, 10 μM), an NO donor. Treatment with SNP significantly increased the NO content and partially reversed the ammonium-induced negative effects, including membrane damage and the decrease in levels of chlorophyll, malondialdehyde, glutathione and ascorbic acid. Further, SNP application increased the catalytic activities of ascorbate peroxidase, superoxide dismutase, guaiacol peroxidase, catalase and glutathione S-transferase, but decreased that of NADH-oxidase. Histochemical staining showed that SNP application caused a significant decrease in the levels of superoxides and hydrogen peroxide. In contrast, application of other breakdown products of SNP (10 μM sodium ferrocyanide, 10 μM sodium nitrite and 10 μM sodium nitrate) failed to show any protective effect. The results suggest that the increased intracellular NO, resulting from SNP application, improved the antioxidant capacity of H. verticillata plants in coping with ammonium-induced oxidative stress.

Topics: Antioxidants; Ascorbic Acid; Chlorophyll; Ferricyanides; Glutathione Transferase; Hydrocharitaceae; Hydrogen Peroxide; Malondialdehyde; Multienzyme Complexes; NADH, NADPH Oxidoreductases; Nitrates; Nitric Oxide; Nitroprusside; Oxidative Stress; Quaternary Ammonium Compounds; Sodium Nitrite; Superoxides

Electrochemically modified screen-printed carbon electrode (SPCE) has been prepared by electrodepositing nickel hexacyanoferrate(III) (NiHCF) onto the electrode surface using cyclic voltammetry (CV). The performance of NiHCF-SPCE sensor was characterized and optimized by controlling several operational parameters. The NiHCF film has been proven to remain stable after CV scanning from 0 to +1.0 V vs. Ag/AgCl in the pH range of 3 to 10 and is re-useable. The most favourable supporting electrolyte solution exhibiting the optimum electroanalytical performance of the NiHCF-SPCE sensor was found to be 0.2 mol/L sodium nitrate. The electrochemical response toward ascorbic acid (AA) and H2O2 in 0.2 mol/L sodium nitrate solution was studied by using CV and the results showed that both analytes were electrocatalytically oxidized at approximately +0.4 V, while H2O2 also revealed a reduction signal at -0.8 V vs. Ag/AgCl. The NiHCF-SPCE sensor exhibited highly linear response for AA and H2O2 in the examined concentration range from 5.0x10-5 to 1.5x10-3 mol/L and from 2.0x10-5 to 1.0x10-3 mol/L (at +0.4 V), with the correlation coefficients of 0.999 and 0.998, respectively. The reproducibility of the NiHCF-SPCE sensor was followed for the determination of AA by using four individual electrodes, and the relative standard deviation of CV peak currents varied between 0.9 % and 2.2 %. The proposed NiHCF-SPCE has been shown to be a very attractive electrochemical sensor for AA and H2O2, also in a view of inexpensive mass production of disposable single-use sensors. The NiHCF-SPCE sensor was tested by measuring AA in multivitamin tablets, with recoveries obtained between 94.4 % and 108.2 % (n=5).

Topics: Ascorbic Acid; Biosensing Techniques; Carbon; Dietary Supplements; Electrochemistry; Electrodes; Ferrocyanides; Humans; Hydrogen Peroxide; Hydrogen-Ion Concentration; Nickel; Nitrates

A 22-year-old woman with an initial diagnosis of 'ruptured ectopic pregnancy' and 'hemorrhagic shock' was sent to the operation room for surgical treatment. The mucocutaneous color was deeply cyanosed and the pulse oximeter oxygen saturation (SpO2) was only 86% after tracheal intubation (100% O2). 'Chocolate-brown' blood was observed and methemoglobinemia was considered. Then the arterial blood gas (ABG) sample was obtained, an intravenous infusion of methylene blue and vitamin C followed. The patient recovered quickly, and later two other patients with similar symptoms were treated in the same way. The success was due to a correct diagnosis accompanied with prompt treatment and quick recognition of the etiology.

Topics: Adult; Antidotes; Antioxidants; Ascorbic Acid; Blood Gas Analysis; Diagnosis, Differential; Female; Food Preservatives; Humans; Meat; Methemoglobinemia; Methylene Blue; Nitrates; Oxygen; Pregnancy; Pregnancy, Ectopic; Rupture; Shock, Hemorrhagic; Sodium Nitrite

By using the ESR spin trapping technique with the N-methyl-D-glucamine dithiocarbamate (MGD)2-Fe(II) complex, the generation of nitric oxide (NO), a gaseous free radical, was observed in NO spin trapping solution bubbled with the filtered main-stream of cigarette smoke. The ESR signal with a three-line spectrum characteristic of an NO radical, which was not observed immediately after bubbling of smoke, started rapidly increasing with time up to around 25 min after the last addition of ferrous ions Fe(II), and then slowly approached a peak value dependent on the burned cigarette mass and on the smoking speed. The production of NO was, however, much affected by air oxidation and enhanced by the addition of ascorbic acid. A certain concentration of sodium nitrite (NaNO2) solution, in which nitrite NO2- is assumed as the main origin of the NO, mimicked closely the time course of NO generation resulting from the smoke of one cigarette. The cigarette smoke that was passed through alkaline pyrogallol solution as a deoxidizer; however, it exhibited an unchanged intensity of NO signal throughout the measurement. These results strongly suggest that NO would be gradually reproduced from NO2- in the reductive aqueous solution containing excess Fe(II) through NO2, which is initially formed and is concomitantly oxidized from NO in cigarette smoke.

Topics: Antioxidants; Ascorbic Acid; Hydrogen Peroxide; Nicotiana; Nitrates; Nitric Oxide; Oxidation-Reduction; S-Nitroso-N-Acetylpenicillamine; Smoke; Solutions

In the present study, we investigated the cellular components that are involved in the release of nitric oxide (NO) from S-nitrosothiols and whether these components also modulate the activity of the nitrergic neurotransmitter in the rat gastric fundus. Electrical stimulation of nitrergic nerves induced frequency-dependent transient relaxations which were mimicked by exogenous NO. The S-nitrosothiols S-nitrosocysteine, S-nitrosoglutathione and S-nitroso-N-acetylpenicillamine induced concentration-dependent relaxations which were generally more sustained as compared to those to nitrergic nerve stimulation or NO. The relaxations to nitrergic nerve stimulation and those to NO were significantly enhanced by the copper(I) chelator neocuproine but not affected by the copper(II) chelator cuprizone. The relaxations to the S-nitrosothiols were significantly inhibited by neocuproine but not by cuprizone. The antioxidant ascorbate did not affect the tension of the muscle strip. However, in the presence of an S-nitrosothiol, ascorbate induced an immediate, sharp and transient relaxation that was significantly inhibited by a low concentration of neocuproine but not by cuprizone. Ascorbate did not induce a relaxation during short-train or prolonged nerve stimulation of the muscle strip. These results suggest that ascorbate interacts with copper to modulate the biological activity of S-nitrosothiols but not that of the nitrergic neurotransmitter. The differential effect of neocuproine indicates that S-nitrosothiols do not mediate the nitrergic neurotransmission of the rat gastric fundus. As neocuproine is to date the only compound that exerts an opposite effect on the biological activity of the nitrergic neurotransmitter and on that of S-nitrosothiols, it may be useful to elucidate the nature of the nitrergic neurotransmitter in the peripheral nervous system.

Topics: Adenosine Triphosphate; Animals; Ascorbic Acid; Chelating Agents; Cuprizone; Electric Stimulation; Gastric Fundus; Isometric Contraction; Male; Monoamine Oxidase Inhibitors; Muscle, Smooth; Neurotransmitter Agents; Nitrates; Nitric Oxide; Phenanthrolines; Rats; Rats, Wistar; Sulfhydryl Compounds

Guinea-pig tracheal strips were used to investigate whether activation of guanylate cyclase in the trachea can reduce the contractile responses of the smooth muscle. Guanylate cyclase was activated by glyceryl trinitrate and a combination of sodium nitrite and ascorbic acid. These activators inhibited tracheal smooth muscle contractions produced by acetylcholine, histamine and electrical field stimulation. However, in the presence of methylene blue, a guanylate cyclase inhibitor, tracheal smooth muscle contractions were not inhibited by the activators. But, in the presence of propranolol, which blocked inhibition mediated by beta-adrenoceptor, both glyceryl trinitrate and the sodium nitrite/ascorbic acid combination were still capable of inhibiting tracheal smooth muscle contractions. Additionally, methylene blue inhibited tracheal smooth muscle relaxation that was electrically induced. These results suggest that the inhibitory action mediated by activated guanylate cyclase may be a mechanism for regulating tracheal smooth muscle contractile responses.

Topics: Acetylcholine; Animals; Ascorbic Acid; Carcinogens; Electric Stimulation; Guanylate Cyclase; Guinea Pigs; Histamine; Male; Muscle Contraction; Muscle, Smooth; Nitrates; Nitroglycerin; Trachea; Vasodilator Agents

The endogenous formation of N-nitrosoproline (NPRO) and N-nitrosothioproline (NTPRO, N-nitrosothiazolidine-4-carboxylic acid) was studied by monitoring their excretion in the urine of guinea pigs given oral doses of 10 mg proline or thioproline after supplementation with 34 mg (0.4 mmol) sodium nitrate. In order to estimate the conversion of nitrate to nitrite, the animals were also supplemented with 3.5 mg (0.05 mmol) sodium nitrite instead of sodium nitrate. In animals fed commercial diets, the excretion of NPRO and NTPRO under supplementation with sodium nitrate was 2.0 micrograms and 28.7 micrograms/animal/day, respectively, whereas the excretion under supplementation with sodium nitrite was 0.7 micrograms and 13.3 micrograms/animal/day, respectively. The higher excretion of NTPRO than NPRO in each case shows that thioproline is more effective for nitrite trapping than proline. The animals supplemented with nitrate excreted more than twice the amounts of NPRO or NTPRO than those supplemented with nitrite. It is assumed, therefore, that more than 0.1 mmol nitrate is reduced to nitrite and takes part in the endogenous nitrosation of the guinea pig. When various concentrations of L-ascorbic acid (AsA), known to inhibit the formation of N-nitroso compounds, were also administered orally to animals immediately after supplementation with sodium nitrate, the NPRO excretion decreased with increasing AsA concentration. These data indicate that the guinea pig, which is unable to synthesize AsA as well as humans, may be an appropriate animal model for evaluation of the endogenous nitrosation ability of humans ingesting nitrate.

Topics: Animals; Ascorbic Acid; Ascorbic Acid Deficiency; Gastric Acidity Determination; Guinea Pigs; Hydrogen-Ion Concentration; Male; Nitrates; Nitrosamines; Nitrosation; Nitroso Compounds; Proline; Thiazoles; Thiazolidines

A relationship between ascorbic acid intake and N-nitrosoproline (NPRO) excretion in humans on a controlled diet was established. Seven healthy males were placed on a low nitrate, low ascorbic acid diet for 12 consecutive days. On days 3-12, a 5.24 mmol oral dose of sodium nitrate was administered in mid-afternoon, at least 2 h after the subject's last meal. On days 4-12, a 4.35 mmol oral dose of L-proline was administered 30 min after the nitrate dose. Ascorbic acid was given in amounts which increased daily from day 5 to day 10 (0.01-5.68 mmol; 1.76-1000 mg) with the proline. Total 24 h urines were assayed for nitrate, NPRO and total ascorbic acid. Nitrate balance was monitored using [15N]nitrate. Average endogenous nitrate synthesis was 1.28 +/- 0.43 mmol/day/person. NPRO excretion was reduced by 6 nmol/day when 0.05 mmol of ascorbic acid was administered. However, as much as 5.68 mmol ascorbic acid did not return NPRO excretion to levels observed before the nitrate and proline were administered. More than 10 times the ascorbic acid required to completely inhibit NPRO formation in vitro did not return NPRO excretion to baseline levels. These data indicate that endogenous nitrosation may be more facile than predicted by the in vitro chemistry.

Topics: Adult; Ascorbic Acid; Diet; Humans; Hydrogen-Ion Concentration; Kinetics; Male; Nitrates; Nitrosamines; Proline