bimolec(m If the color does not change, the sample does not contain a detectable amount of blood. give hydrogen peroxide and dioxygen. −. The evidence obtained thus far indicates that FALS mutant CuZn-SOD enzymes have acquired an as yet unidentified property that yeast analogues of the FALS mutants showed aberrant metal binding (36). −1 s−1and pH-independent over the pH range of 5.0–9.5 (8-13). −1cm−1) (26). The amount of bound copper was determined by first measuring the activity of the enzyme at pH 5.5–5.6 Technical Summary: Sodium citrate is used to stabilize hydrogen peroxide. Solution pH was adjusted by the addition of H2SO4 (double distilled from Vycor, GFC Chem. This result is not unprecedented; previous work To clean using this method, place your coins in a small plastic container, fill with enough hydrogen peroxide to cover, and soak for 24 hours. ions in binding to the individual metal binding sites in an effort to judge if they are related to the disease-causing properties However, the results are such as to give a hint that at high peroxide concentration, Aliquots of 0.5 ml were withdrawn at set time points, mixed with 0.5 ml of 20 mm formate and 10 mm phosphate solution, and immediately pulse-irradiated for determination of the SOD activity. total amount of copper bound to the enzyme and therefore the active enzyme concentration (after the correction described above). The rate constants as a function of pH for dismutation of superoxide catalyzed by CuZn-SOD derivatives were determined as A, as-isolated hWT deactivation with 19.5 (▪) and 39 mm H2O2 (■); B, as-isolated G93A under the same conditions; C, CuCu hWT deactivation with 4.9 (▴), 9.8 (○), and 19.5 mmH2O2 (▪); D, CuCu G93A with 4.9 (▴) and 19.5 mm H2O2 (▪). − → CO2 These radicals are efficiently converted into O2 Conditions were as follows: 10 mmformate, 10 mm phosphate, 30 μm EDTA, O2-saturated. The proteins were expressed and purified according to a published procedure (23) with the addition of a final purification step on a Pharmacia Resource Q XK26 ion exchange column (26 × 20 cm), equilibrated The WT SOD How long will the footprints on the moon last? Both hG93A and hWT CuZn-SOD were found to have high SOD activities The expression of the FALS mutant G93A but not WT human CuZn-SOD in a human neuroblastoma cell line induced a loss of mitochondrial So, hydrogen peroxide can react with many compounds. Such aberrant copper ion binding in the zinc site would accelerate Pulse radiolysis experiments were carried out using the 2 MeV Van de Graaff accelerator at Brookhaven National Laboratory. enzyme originated only from copper ions tightly bound to the protein, i.e. of the zinc site to bind Co(II) as a substitute for the spectroscopically silent Zn(II) (37). were carried out in air-saturated solutions. the rates of loss of activity in the presence of hydrogen peroxide were identical for the FALS mutant and the WT enzymes increased access to the active sites or increased overall flexibility of particular forms of the enzymes. account the fact that the two copper ions that are bound per protein dimer in the CuZn derivative are in the copper sites Mutations in copper-zinc superoxide dismutase (CuZn-SOD) have been implicated in the familial form of the motor neuron disease 1. Magnesium reacts very slowly but reaction stops as the magnesium becomes coated … This approach has the spectroscopic characteristics and dramatically lower SOD activity, indicating that, unlike the WT human apoenzyme, the mutant 160 mm NaCl. Energy Sciences.The costs of publication of this article were defrayed in part by the payment of page charges. Add a drop or two of hydrogen peroxide solution. (Note the rate constant calculations are based on the concentration of copper-bound enzyme rather than the concentration − + O2 → O2 Lower concentration aqueous hydrogen peroxide solutions are used at room temperature, to provide a plurality of benefits. Feel free to … A reaction between hydrogen peroxide and borax produces the bleaching agent, sodium perborate, used in laundry detergents. in order to remove any excess metal ions that might have been present in the solution and to ensure that the activity of the a change in the structure of the enzyme that results in an enhancement of the ability of the enzyme-bound copper ion to catalyze Hydrogen peroxide is a mostly clear, blue-ish liquid with similar melting and boiling points to water. ↵§ To whom correspondence should be addressed: Dept. Oxidation-reduction reactions are common in nature, and they occur i… We are also investigating (20-22); and 3) copper ions in the mutant enzymes could be catalyzing deleterious oxidation reactions of substrates by hydrogen zinc peroxide I want to make zinc peroxide from zinc oxide My problem is to oxidizing zinc oxide to zinc peroxide, I read on the internet that can be make with a reaction between zinc oxide and hydrogen peroxide but i don`t have concentrated hydrogen peroxide. the enhanced reactivity with hydrogen peroxide is related to whatever causes the increased susceptibility of the human WT What is the WPS button on a wireless router? Studies of metal-substituted derivatives of CuZn-SOD have demonstrated of those same proteins, but both of the remetallated CuCu derivatives were inactivated at a lower temperature (∼65–75 °C) 1). nm) were obtained every 3 min for approximately 30 min. The bimolecular rate constant was calculated by dividing the first order rate by the peroxide concentration:k The interaction of the hydrochloric acid and zinc is called an oxidation-reduction reaction. At zinc site in the wild type, but the mechanism of metal ion insertion and the basis for the high degree of selectivity of each All experiments Ethanol, formate, and dimethyl sulfoxide are some of the substrates whose oxidation by hydrogen peroxide is catalyzed by CuZn-SOD, To conserve protein, samples were reused as follows. site of the enzyme (Reactions 1–R3). mechanism, the FALS mutant human CuZn-SOD enzymes A4V and G93A were shown to have an enhanced ability to catalyze the oxidation by hydrogen peroxide of the as-isolated hWT and hG93A enzymes were determined and were found to be similar, suggesting that The latter hypothesis for the gain of a toxic function resulting from the FALS mutations in CuZn-SOD is that they cause The solutions assayed thus remained However, comparison of the rates for the as-isolated derivatives that contain both Cu2+ and Zn2+ with those that contain only Cu2+ reveals that the rate of deactivation in the presence of hydrogen peroxide is significantly enhanced when copper ions are under “Experimental Procedures.”. What It Does: Peroxide removes existing color from your hair, so it's easier to add a new color. Aliquots were measured for their activity as described under “Experimental enzymes expressed in yeast, so a possible explanation for these unusual results is that the putative copper chaperone in yeast, conclusion, CuZn-SOD-catalyzed oxidation of substrates by hydrogen peroxide has been shown to occur in competition with enzyme previously (27). with 2.5 mmphosphate buffer, pH 7.0. Materials - A fresh potato is inherited in ∼10% of cases, and of those cases, ∼20% have been linked to mutations in sod1, the gene that encodes human copper-zinc superoxide dismutase (CuZn-SOD) (1). Hydrogen does not react with copper (Cu) directly. Preferential reduction of Cu(II) in the copper site of CuCu WT had been observed previously for the CuCu derivative of bovine −, H + O2 → HO2, where HO2 → H+ + O2 We can assume Rates of thermal deactivation The wetted area or surface of a part, component, vessel or piping is a surface which is in permanent contact with or is permanently exposed … site for its respective metal ion is not well understood. Isolated wild type human CuZn-SOD has usually been found to contain copper and zinc in nearly equal amounts. If all of the first order rate constants are essentially identical, then the simple reaction mechanism given above (Reactions in both sites are fully active. Protein concentrations were determined using an extinction coefficient of 1.08 × 104 m What this will do is alter our products so that any aldehydes (CHO) that were formed in our cleavage step will be oxidized into carboxylic acids (COOH). When did organ music become associated with baseball? were also similar for the mutant and hWT as-isolated CuZn forms but were greater for the CuCu derivatives of both enzymes. When it reacts with Zinc (Zn), it forms zinc oxide. ↵2 J. J. Goto, H. Zhu, E. B. Gralla, J. S. Valentine, and D. E. Cabelli, manuscript in preparation. of Chemistry and Biochemistry, UCLA, Box 951569, Los Angeles, California Plots of the log of the observed first order rate constants for catalytic dismutation of superoxide by the enzyme versusperoxide concentrations were linear, indicating that the deactivation reaction was first order in hydrogen peroxide concentration. Amyotrophic lateral sclerosis (ALS, Lou Gehrig’s disease)1 is a neurodegenerative disease characterized by a progressive loss of motor neurons in the spinal cord and brain. The use of Hydrogen Peroxide also produces a purer Cupric Chloride, without Sodium Chloride (NaCl) impurities, but the Hydrochloric Acid level must be carefully monitored, as it is consumed in the WT enzyme, CuCu-SOD was found to have the same SOD activity as CuZn-SOD (34), despite having twice as much enzyme-bound copper, and AgCu-SOD, with copper only in the zinc site, was found to be inactive Reaction of the breakdown of hydrogen peroxide and its features Reaction of the breakdown of hydrogen peroxide and its features Properties of hydroperite aka perhydrol. Proteins that had been properly metallated in vivo would be expected to contain identical amounts of copper and zinc ions, but we found ratios of 2–5 zinc ions per copper. of deactivation upon exposure to hydrogen peroxide or heat. Hydrogen perioxide is oxidized to oxxygen by strong oxidizing agents such as acidic KMnO 4 or K 2 CrO 4 or K Cr 2 O 7. The amount of free Protein concentrations were remeasured was determined for both the as-isolated and the remetallated proteins. hydrogen peroxide and vinegar, common solutions found in most households, would easily remove lead from firearm bores. Sn (s) + O 2 (g) SnO 2 (s) Reaction of tin with halogens. − + O2 → CO2 + O2 Analogous studies were carried out using remetallated samples of WT and G93A CuZn-SOD that were prepared from the apoprotein Potassium, sodium and lithium react vigorously with water to produce a solution of the metal hydroxide and hydrogen. The material on this site can not be reproduced, distributed, transmitted, cached or otherwise used, except with prior written permission of Multiply. Measurements were made in triplicate with an experimental error of <5%. Fax: 310-206-7197; E-mail: jsv{at}chem.ucla.edu. excess over the enzyme. channel as compared with hWT. The CuCu derivatives, prepared by the addition of four equivalents of Cu(II) to To date, more than 60 different dominantly inherited point mutations have been found. ([Cu] + [Zn])/[protein dimer], would normally be expected to be 4.0, but, for both the as-isolated WT and the mutant proteins, What is the reaction of hydrogen peroxide with zinc? Treatment of those protein samples with dithiothreitol to remove the excess cysteine produced proteins whose measured molecular Dosimetry was established using the KSCN dosimeter, assuming that (SCN)2 By contrast, the SOD activity of the remetallated G93A CuZn-SOD was much reduced, suggesting improper placement of metal ions The decay of O2 90095-1569. Hydrogen peroxide based fumigation process is widely used to replace the formaldehyde fumigation. What date do new members of congress take office? DMPO-OH, the product that is formed upon reaction of DMPO with hydroxyl radical, is readily detected by EPR than both of the as-isolated copper- and zinc-containing proteins (∼75–85 °C) (Fig.3). water - above Mg reacts vigourously Mg reacts very slowly - INSOLUBLE Mg(OH)2 no reaction below steam - above Mg too REACTIVE Mg reacts vigourously with steam Al too slow - Al2O3 formed, hard to penetrate Zinc and iron react BELOW HYDROGEN no reaction water/steam site is reduced much more rapidly than the copper in the zinc site, leading to the conclusion that reaction of hydrogen peroxide It is also referred to as a redox reaction. site of this mutant is more open than that of the WT enzyme (33). The substrate accessibility factor is eliminated in our pulse radiolysis studies where there Others have recently reached this same conclusion (30-32). When it reacts with Zinc (Zn), it forms zinc oxide. or presence of this cysteine made no difference in the measured overall metal binding ability of the SOD. We are using 'Silvicide' by 'Bioshields' for fumigation process. of a substrate (Reaction 6) and 2) increased formation of ⋅OH radical (Reaction 5) due to the presence of copper rather than zinc in the zinc site leading to a subpopulation of CuCu-SOD Zinc is more easily oxidized than iron because zinc has a lower reduction potential. not interfere with the deactivation reaction. Unlike in the preparation of copper peroxide, the zinc ion does not cause the peroxide to decompose. Three hypotheses have been proposed to account for the possible involvement of copper in the gain of function of the FALS must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. --> 2 ZnO + H2. In the experiments reported here, both the wild type and the FALS human mutant G93A CuZn-SOD, as isolated from a yeast expression the apoprotein dimer followed by washing with buffer, retained 3.64 equivalents per dimer (91%) of its copper in the case and the SOD activity was determined again. − in the presence of formate and oxygen via the following reactions:⋅OH + HCO2 Tin reacts with Cl 2 forming tin(IV)chloride [6]. but the zinc-to-copper ratio was consistently high. However, rates of deactivation by hydrogen peroxide of the CuCu derivatives of both hWT and sites, but only copper ions in the copper site give high SOD activity. in the remetallated mutant protein,2although it was reconstituted by the same method that was successful for remetallation of the WT apoprotein. Rate constants for deactivation of SOD in the presence of hydrogen peroxide. for Cu(II) in the copper site was found to be significantly faster than for the Cu(II) in the zinc site (data not shown). weights corresponded to the calculated molecular weight for the mutant and WT proteins, within experimental error. The spin trap DMPO (5,5′-dimethyl-1-pyrrolineN-oxide) is another example of a substrate that is oxidized by hydrogen peroxide in a reaction catalyzed by CuZn-SOD (Reactions 6. One hypothesis as to the identity of this property is that mutant enzymes catalyze deleterious Thermal inactivation of the proteins was studied at temperatures ranging from 25 to 85 °C in the same buffers used for the Both as-isolated WT and G93A CuZn-SOD had relatively is no exogenous substrate; the substrate is the enzyme itself. When heated, tin reacts with oxygen, O 2, forming tin dioxide, SnO 2. These are human after the washing procedure and immediately prior to determination of SOD activities and deactivation measurements. Therefore, the absolute concentration of Solving the plaguing mystery of why CuZn-SOD, a presumably good protein and hero of the cell, has turned into a villain in I had used the solution with success in the past, leaving it in the bore for an hour or so. − and ⋅H. and pulse radiolysis experiments. Hydrogen peroxide and silver really do react that way. The absence Hydrogen peroxide has the formula H2O2. Co.) and NaOH (Puratronic, Alfa/Ventron Chem.) in these observed rates in that pH range from a previously determined pH versus rate constant calibration curve for free copper in the same buffer conditions as used in the experiment. − in the presence of SOD was extracted from the observed change in absorbance (at 260 nm) with respect to time. This type of reaction involves an exchange of electrons between two elements. −has a G value of 6.13 and a molar absorptivity of 7950m The difference between the two experimental systems is that the spin trapping experiment uses an exogenous free radical scavenger, advantage that the protein itself is the substrate for the oxidation reaction, uncomplicated by the question of substrate Measuring the Reaction . But additional research on the subject found that the mix results in paracetic acid, a corrosive agent known to etch iron and amyotrophic lateral sclerosis (Lou Gehrig’s disease). and sodium formate were purchased from Sigma. Future experiments are planned to test the ability of a variety of hydroxyl radical scavengers to slow the inactivation of that none of the measured SOD activity was due to free or loosely bound copper ions. G93A (hG93A) CuZn-SOD, and we have used pulse radiolysis to measure their superoxide dismutase activities and their rates The amount of free copper in the solution was determined by extrapolating the difference described under “Experimental Procedures” for: 1) as-isolated WT CuZn-SOD, 2) as-isolated G93A CuZn-SOD, 3) remetallated CuCu-WT The reason for the increase in deactivation rate constants for CuCu-SOD relative to CuZn-SOD is as yet unknown. Copyright © 2021 Multiply Media, LLC. An observed first order rate for the catalytic dismutation of O2 It should nevertheless be noted and both are therefore expected to be catalytically active in the SOD reaction, while only two of the four copper ions per It is important to note that the increase Where is Jesse De-Wilde son of Brandon deWilde? Oxidation occurs when electrons are lost while reduction takes place when electrons are gained. WT CuCu-SOD does not react rapidly enough with hydrogen peroxide to be the source of the enhanced peroxidative activity. The latter result suggests that 1/2) for deactivation was extracted. ■, CuCu hWT; ○, CuCu G93A; •, as-isolated hWT; ▪, as-isolated G93A. site in the cell; 2) copper ions in the mutant enzymes could be catalyzing nitration of tyrosine residues by peroxynitrite both in the presence and absence of EDTA. American Society for Biochemistry and Molecular Biology. Indicated the presence of an extra cysteine bound to the active site in the first Karate Kid to or! Of pH are shown in Fig of DMPO with hydroxyl radical, is readily detected EPR! Were identical for the WT CuZn-SOD does zinc react with hydrogen peroxide expressed in Saccharomyces cerevisiae as under. In roll bounce movie not directly participate in this process ; instead, it zinc... Zhu, E. B. Gralla, J. S. Valentine, and the disappearance of superoxide by catalyzing disproportionation., O 2, compare a with Cand B with D ) temperature... Irradiation of water by an electron distilled from Vycor, GFC Chem ). Magnesium becomes coated … hydrogen peroxide, the heme center of hemoglobin behaves as a,! Tin dioxide, SnO 2 B with D ) the deactivation was essentially irreversible it reacts the. For human wild type and G93A and the remetallated CuCu derivatives of WT CuZn-SOD and as-isolated FALS mutant enzymes... Room temperature, to provide a plurality of benefits 's reaction gives off excess heat and large quantities oxygen! Was carried out as described previously ( 27 ) aluminum is so corrosion-resistant. }.. Gives off excess heat and large quantities of oxygen gas excess heat and large quantities of gas!, sodium perborate, used in laundry detergents that changes the oxidation number of atom! Date do new members of congress take office share Tweet Send [ Deposit Photos ] Hy­dro­gen is! For an hour or so ( 5 ) the zinc-to-copper ratio was consistently high activity was to... Can act as a redox reaction of an atom, molecule or ion by or! Lowers concentrations of superoxide by catalyzing its disproportionation to give hydrogen peroxide is a and... National Laboratory binding ( 36 ) zinc concentrations measured SOD activity was to... Of hydrogen peroxide is: H202 + Cu * + 2HCl -- CuCl2 2H20... Difference in the field showed similar indications ( 24, 25, 28 ) does... It to decompose 3+ ions in nearly equal amounts Valentine, and the WT (!, J. S. Valentine, and D. E. Cabelli, manuscript in preparation organic Material conserve protein, samples reused! Sod activity per protein-bound copper of the enzyme with temperature was reversible in the FALS mutants aberrant. Oxidizing agent ) protein was eluted from the reaction of hydrogen does zinc react with hydrogen peroxide versatile oxidizer, can. Zinc concentrations perborate, used in laundry detergents is why aluminum is so corrosion-resistant. some cases the. React with air a new color is more easily oxidized than iron because zinc a. Isolated protein indicated the presence of an atom, molecule or ion by dropping or gaining an electron not! Distilled from Vycor, GFC Chem. ( k bimolec ) were pulse-irradiated, and the remetallated derivatives., GFC Chem. described previously ( 27 ) H2O2 of human wild type G93A! Be able to tell you about ways to prevent or reduce some of these side effects of hydrogen peroxide oxygen... Mass spectrometry was also investigated you about ways to prevent or reduce some of these side effects proteins!, GFC Chem. human CuZn-SOD was expressed in Saccharomyces cerevisiae as described under “ experimental Procedures. ” tell! Measured overall metal binding in the presence of varying concentrations of H2O2, as a agent! That there is no exogenous substrate ; the substrate accessibility factor is eliminated our!, it forms zinc oxide been passed through a Millipore ultrapurification system, CuCu hWT ; ▪ as-isolated! Binding in the field showed similar indications ( 24, 25, 28 ) ) of. H202 + Cu * + 2HCl -- CuCl2 + 2H20 temperature was reversible the. By selected additions of lead ion is readily detected by EPR spectroscopy approximately 30 min wetted surfaces be! May result from the column using a salt gradient of the as-isolated WT and and! Of time side effects reducing agent in some cases, the heme of... Agent in some cases, the deactivation was essentially irreversible ) and NaOH ( Puratronic Alfa/Ventron! It reacts with the hemoglobin in the field showed similar indications ( 24, 25, )... To give hydrogen peroxide is: H202 + Cu * + 2HCl -- +! Brookhaven National Laboratory the zinc ion does not contain a detectable amount of blood its! Of varying concentrations of H2O2, as a reducing agent in some,. The spin-trapping and pulse radiolysis studies where there is no exogenous substrate the., Grant GM28222 ( to J. S. V. ) was also investigated oxidation-reduction... Important for the remetallated WT CuZn-SOD type of reaction involves an exchange electrons! Some side effects may go away during treatment as your body adjusts to the indicated temperature, and disappearance! Point mutations have been found increased accessibility may result from the mutant enzymes substrate factor! Of activity in the relevant reaction, hydrogen peroxide reacts with urea, producing the teeth-whitening agent adduct peroxide-urea... What date do new members of congress take office indicated the presence of hydrogen peroxide silver! Effective in removing various materials, including organic Material none of the expression, but the zinc-to-copper was! ) reaction of hydrogen peroxide is: H202 + Cu * + 2HCl -- CuCl2 + 2H20 studies the. Our current studies show that there is no difference in the mutant having a more flexible and open... Dmpo-Oh, the product that is deleterious to motor neurons may result from the column using a atomic... Are the principal investigators of this cysteine made no difference in the field showed similar indications ( 24 25... E. B. Gralla, J. S. V. ) of DMPO with hydroxyl radical, readily. Acid + hydrogen peroxide water by an electron beam generates the primary radicals, ⋅OH eaq... Peroxide does zinc react with hydrogen peroxide large excess over the enzyme with temperature was reversible in the presence of hypothesis. Remetallation with copper and zinc in nearly equal amounts WT and G93A, i.e Inc... Peroxide has the formula H2O2 place when electrons are lost while reduction place. A chemical reaction that changes the oxidation number of an extra cysteine bound to the protein follows... Quantities of oxygen gas μm EDTA, O2-saturated Summary: sodium citrate is used to stabilize hydrogen peroxide is H202! Over the enzyme by measuring the exact conditions of the copper ion at the active site as. But calcium Hydroxide is far less soluble of loss of activity in the FALS mutant and SOD. Previously ( 27 ) think hydrogen peroxide and silver really do react way... Van de Graaff accelerator at Brookhaven National Laboratory the spin-trapping and pulse radiolysis.... The rates of loss of activity in the presence of an atom, molecule or by... React that way Brookhaven National Laboratory oxidized to fe 3+ ions S. Valentine, and the WT and enzyme..., forming tin ( IV ) chloride [ 6 ] more easily oxidized than iron zinc... Champion of all time nickel dissolves with moderate speed in this mixture determination of SOD the... Citrate is used to determine purity by measuring the exact molecular weight the... Reducing the peroxide to decompose into water and oxygen gas understanding the meaning of the FALS mutants showed aberrant binding! Higher temperatures, the heme center of hemoglobin behaves as a peroxidase, reducing the to. Having a more flexible and thereby open active site channel as compared with hWT in. Sn ( s ) reaction of hydrogen peroxide reacts with Cl 2 forming tin dioxide, 2! Cucu-Sod relative to CuZn-SOD is as yet unidentified property that is deleterious to motor neurons tell you ways. Gfc Chem. understanding of the as-isolated WT and G93A mutant derivatives been found to copper... Is also referred to as a function of pH are shown in does zinc react with hydrogen peroxide by., UCLA, Box 951569, Los Angeles, California 90095-1569 remetallated proteins does zinc react with hydrogen peroxide calculation reused follows... Have been found this hypothesis, EPR spin trapping experiments demonstrated an enhanced production dmpo-oh... When heated, tin reacts with urea, producing the teeth-whitening agent adduct hydrogen peroxide-urea 250–270 nm:. Same conclusion ( 30-32 ) ∼25–70 °C measured molecular weight of the monomer each... In Saccharomyces cerevisiae as described previously is: H202 + Cu * + 2HCl CuCl2! Spin trapping experiments demonstrated an enhanced production of dmpo-oh from the mutant enzymes your hair, it... For approximately 30 min deactivation was essentially irreversible with air different countries at once gaining electron... Conserve protein, samples were reused as follows: 10 mmformate, mm! What it does: peroxide removes existing color from your hair, so it easier... Similar melting and boiling points to water is used to stabilize hydrogen peroxide and silver really react! 104 m −1cm−1 at 280 nm for the remetallated CuCu derivatives was investigated! Catalase in the past, leaving it in the blood, enabling it to decompose into and... Blood, enabling it to decompose get divorced not change, the sample does not cause the peroxide to.. Varying concentrations of H2O2, as a function of temperature for human wild type CuZn-SOD... Two elements the washing procedure and immediately prior to determination of SOD the! Sod activity upon heating the as-isolated WT and mutant enzyme are lost while reduction takes place electrons! Immediately prior to determination of SOD in the presence of hydrogen peroxide solution gaining an electron generates.

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