Ascorbyl Radical Research Articles

Structural, Antioxidant, and Protein/DNA-Binding Properties of Sulfate-Coordinated Ni(II) Complex with Pyridoxal-Semicarbazone (PLSC) Ligand

The pyridoxal-semicarbazone (PLSC) ligand and its transition metal complexes have shown significant biological activity. In this contribution, a novel nickel(II)-PLSC complex, [Ni(PLSC)(SO4)(H2O)2], was obtained, and its structure was determined by X-ray crystallographic analysis, FTIR, and UV-VIS spectroscopy. The sulfate ion is directly coordinated to the central metal ion. The intermolecular stabilization interactions were examined using Hirshfeld surface analysis. The crystal structure was optimized by a B3LYP functional using two pseudopotentials for nickel(II) (LanL2DZ and def2-TZVP) together with a 6-311++G(d,p) basis set for non-metallic atoms. The experimental and theoretical bond lengths and angles were compared, and the appropriate level of theory was determined. The stabilization interactions within the coordination sphere were investigated by the Quantum Theory of Atoms in Molecules (QTAIM). The antioxidant activity towards hydroxyl and ascorbyl radicals was measured by EPR spectroscopy. The interactions between Human Serum Albumin (HSA) and the complex were examined by spectrofluorimetric titration and a molecular docking study. The mechanism of binding to DNA was analyzed by complex fluorescence quenching, potassium iodide quenching, and ethidium bromide displacement studies in conjunction with molecular docking simulations.

Short communication: Ascorbyl/ascorbate ratio as a marker of oxidative stress in larvae (Apis mellifera) exposed to Paenibacillus larvae.

Paenibacillus larvae, the causal agent of American foulbrood disease (AFB), affects Apis mellifera larvae and can induce oxidative stress by overproduction of radical oxygen species (ROS). This study aimed to assess the oxidative stress levels in larvae exposed to three different strains of P. larvae through their diet by examining the ascorbyl radical (A) to ascorbate anion (AH¯) ratio. The results revealed that larvae inoculated with P. larvae exhibited a lower value of this index compared to uninoculated ones. Interestingly, the level of A remained constant, while the concentration of AH¯ increased. Said increase correlated with the virulence of the specific P. larvae strain used in the inoculation. These findings suggest a potential link between AH¯ molecules and a defense response in A. mellifera larvae against infection, consistent with their resistance to P. larvae (LD50).

Chemical Composition, Antioxidant Potential, and Nutritional Evaluation of Cultivated Sorghum Grains: A Combined Experimental, Theoretical, and Multivariate Analysis.

Sorghum grain (Sorghum bicolor L. Moench) is a gluten-free cereal with excellent nutritional value and is a good source of antioxidants, including polyphenols, as well as minerals with proven health benefits. Herein, the phenolic composition, elemental profile, and antioxidant activity of sixteen food-grade sorghum grains (S1-S16) grown under agroecological conditions in Serbia were determined. Nine phenolic compounds characteristic of sorghum grains, such as luteolinidin, 5-methoxyluteolinidin, luteolidin derivative, luteolidin glucoside, apigeninidin, 7-methoxyapigeninidin, apigeninidin glucoside, and cyanidin derivative, were quantified. The antioxidant potential of the analyzed sorghum grains was evaluated by UV/Vis (DPPH, ABTS, and FRAP) and Electron Paramagnetic Resonance spectroscopy (hydroxyl and ascorbyl radical scavenging assays). The content of macro- and microelements was determined by Inductively Coupled Plasma Optical Emission spectroscopy. Theoretical daily intakes of selected major and trace elements were assessed and compared with the Recommended Daily Allowance or Adequate Intake. Sample S8 had the highest amount of phenolic compounds, while S4, S6, and S8 exhibited the strongest antioxidative potential. The sorghum studied could completely satisfy the daily needs of macro- (K, Mg, and P) and microelements (Se, Zn, Fe). Pattern recognition techniques confirmed the discrimination of samples based on phenolic profile and elemental analysis and recognized the main markers responsible for differences between the investigated samples. The reaction between hydroxyl radicals and luteolinidin/apigeninidin was investigated by Density Functional Theory and thermodynamically preferred mechanism was determined.

Antioxidant and free radicals species in the aqueous humor of patients with age-related cataract

Introduction/Objective. Age-related cataract is a significant cause of visual impairment worldwide. Oxidative damage and the effects of free radical species are considered important in the etiopathogenesis of cataracts. The aim of this study was to evaluate the antioxidative capacity and oxidative stress in the aqueous humor (AH) according to age and cataracts maturity. Methods. Clinical and biochemical researches were carried out in 55 patients with age-related cataract. According to the cataract maturity, patients were classified into incipient (cortical ? group C, 18 patients; nuclear ? group N, 20 patients; mature ? group M, 17 patients). In order to evaluate the impact of age patients within each group were divided into Group I (65?69 years) and Group II (70 ? years). The antioxidant activity of AH was measured by the reduction power method and the activity of glutathione peroxidase (GPx) spectrophotometrically. Changes in the concentrations of hydroxyl and ascorbyl radicals were detected by electron spin resonance spectroscopy. Results. Both reduction power and GPx activity were significantly (p < 0.001) reduced in group N compared to group C and in group M compared to group N. Concentrations of hydroxyl (29.45 ? 1.01% in group C, 38.12 ? 1.29% in group N, and 74.14 ? 2.52% in group M) and ascorbyl radicals (26.12 ? 0.89% in group C, 41.15 ? 1.39% in group N, and 83.56 ? 2.84% in group M) increased significantly (p < 0.05) with progression of age-related cataract. Significant negative correlation (r = -0.759, p < 0.05) was determined between concentrations of HO? and content of GPx. Conclusion. Our research proved that the level of oxidative stress in the AH is significantly affected during aging and cataract progression, The obtained data support the hypothesis that during aging, depending on the maturity of the cataract, the antioxidant capacity in the AH decreases due to an increase in the concentration of reactive HO?.

Oxidative stress coping capacity (OSC) value: Development and validation of an in vitro measurement method for blood plasma using electron paramagnetic resonance spectroscopy (EPR) and vitamin C.

Oxidative stress as a driver of disease is reinforcing the trend towards supplementation with antioxidants. While antioxidants positively influence the redox status when applied at physiological doses, higher concentrations may have pro-oxidative effects. Precise assessment methods for testing the supply of antioxidants are lacking. Using in-situ-irradiation as stressor and electron paramagnetic resonance (EPR) spectroscopy as readout system for formed radicals, a stress response assessment method was developed, using protein solutions and plasma samples from transfusion medicine. The method was validated in a double-blind placebo-controlled in vivo cross-over pilot study in blood plasma samples of individuals before and after vitamin C supplementation. Reference measurements were performed for the exogenous antioxidants β-carotene and vitamin C, and glutathione as an endogenous representative. Malondialdehyde was studied for oxidative stress indication. Protein solutions without antioxidants showed a linear increase in radical concentration during irradiation. The in-vitro-addition of vitamin C or plasma samples from subjects displayed two slopes (m1, m2) for radical production, whereby m1 represented the amount of antioxidants and proteins, m2 only the protein content. These two slopes in combination with the intervening transition area (T) were used to calculate the oxidative stress coping capacity (OSC), which correlated positively with vitamin C concentration in blood plasma, while oxidative stress biomarkers showed only fluctuations within their reference ranges. Furthermore, a selective radical quenching mechanism for vitamin C was observed: the proportion of reactive oxygen species (ROS) in the plasma samples was degraded in dependence to the vitamin C concentration ingested. The proportion of lipid oxygen species (LOS) remained stable while the ascorbyl radical increased with higher vitamin C intake. OSC may represent a sensitive method to detect treatment effects on the redox status in vivo in future validation and treatment studies, and potentially in clinical routine.

Ascorbate Is a Primary Antioxidant in Mammals.

Ascorbate (vitamin C in primates) functions as a cofactor for a number of enzymatic reactions represented by prolyl hydroxylases and as an antioxidant due to its ability to donate electrons, which is mostly accomplished through non-enzymatic reaction in mammals. Ascorbate directly reacts with radical species and is converted to ascorbyl radical followed by dehydroascorbate. Ambiguities in physiological relevance of ascorbate observed during in vivo situations could be attributed in part to presence of other redox systems and the pro-oxidant properties of ascorbate. Most mammals are able to synthesize ascorbate from glucose, which is also considered to be an obstacle to verify its action. In addition to animals with natural deficiency in the ascorbate synthesis, such as guinea pigs and ODS rats, three strains of mice with genetic removal of the responsive genes (GULO, RGN, or AKR1A) for the ascorbate synthesis have been established and are being used to investigate the physiological roles of ascorbate. Studies using these mice, along with ascorbate transporter (SVCT)-deficient mice, largely support its ability in protection against oxidative insults. While combined actions of ascorbate in regulating epigenetics and antioxidation appear to effectively prevent cancer development, pharmacological doses of ascorbate and dehydroascorbate may exert tumoricidal activity through redox-dependent mechanisms.

Development of hemp seed oil nanoemulsions loaded with ascorbyl palmitate: Effect of operational parameters, emulsifiers, and wall materials

The perceived health properties of hemp seed oil, as one of the few plant-basedsources of omega-3 and omega-6 fatty acids with an ideal ratio of 1:3, suggest its incorporation in food-grade emulsions to improve its water solubility and oxidative stability. The current research's main aim was nanoemulsification of hemp seed oil using the oil-in-water emulsification method followed by ultrasonication. The entrapment efficiency of the nanoemulsions for antioxidant ascorbyl palmitate and its impact on oxidative stability of the oil was also evaluated. Gum arabic: maltodextrin in 75:25 ratio could result in nanoemulsion with entrapment efficiency of 97.10 % for ascorbyl palmitate and radical scavenging activity of oil-soluble bioactives of 92.13 %. Moreover, incorporation of ascorbyl palmitate could effectively retard the oxidation, specifically in nanoemulsions containing gum Arabic. The optimum formulation of nanoemulsion having an average droplet size of 293 nm can be applied as an ideal vegetarian source of omega-3 fatty acids.

OXIDATIVE BALANCE IN BRAIN AFTER CHRONIC EXPOSURE TO ARSENIC

The effect of chronic As exposure in brain, the specific oxidants production and oxidative stress still needs more attention. In the present study, male wistar rats were chronically exposed to As bythe drinking water during 60 days. The rats receiving a dose of 10 mg As/l showed a 9- and 18-fold increase in the content of As in blood and brain, respectively, as compared to the control group. Histopathological changes were detected in brain samples isolated from animals treated with Asas compared to control animals. The oxidation rate of 2ʹ,7ʹ-Dichlorofluorescein diacetate (DCFH-DA) showed no significant changes after the treatment. A significant increase (13%) in the steady state concentration of ascorbyl radical (A•) was determined in the brain isolated from animals exposed to 50 mg As/l. The lipid radical (LR•) generation rate and the content of malondialdehyde (MDA) were increased by 34% and 32%, respectively in brain isolated from rats receiving 50 mg As/l. The content of ascorbate (AH-) in brain was not affected by the exposure to As.However, the content of glutathione (GSH) and the lipophilic antioxidant α-tocopherol (α-T), were significantly decreased after As supplementation, as compared to control brains. The A•/AH- content ratio in rat brain showed no changes associated to the As supplementation. Nevertheless, a significant increase of 44% and 55% was determined in the LR•/α-T and MDA/α-T content ratios, respectively, suggesting that the oxidative imbalance in the lipophilic cellular medium could be the primarily effect of As associated to membrane damage.

Kinetics and mechanistic aspects of superoxide radical-mediated transformation of ascorbate

Ascorbate (Asc) has long been known to be an antioxidant against reactive oxygen species (ROS) for cytotoxicity and human diseases. However, its reported reactivity with superoxide radical (O2•−), an important ROS involved in aerobic metabolism system, exhibits great discrepancy; its mechanistic mode of actions on the molecular level is also largely debatable. Here, we investigated the reaction of Asc and O2•− by a combination of experimental and theoretical approaches. The rate constant measured by a competitive technique method was determined to be 3.3 × 104 M−1 s−1 under physiological condition. Mechanistically, our theoretical result indicated that O2•− abstracts a hydrogen atom on the enolic hydroxyl group forming ascorbyl radical (Asc•), which is unreactive with O2•− or Asc• itself. However, with different protonation state of Asc• (i.e., AscH•) in acidic environment, its reactivity was demonstrated to be significantly altered. AscH• can react with O2•−via hydrogen atom abstraction and single electron transfer and/or undergo self−disproportionation. These complementary results provided fundamental principles for the ecotoxicological function and environmental potentials of Asc against ROS.

Dose-dependent scavenging activity of the ultra-short-acting β1-blocker landiolol against specific free radicals.

Landiolol, a highly cardioselective ultra-short-acting β1-blocker, prevents perioperative atrial fibrillation associated with systemic inflammation and oxidative stress. We evaluated the direct scavenging activity of landiolol against multiple free radical species. Nine free radical species (hydroxyl, superoxide anion, ascorbyl, tert-butyl peroxyl, tert-butoxyl, singlet oxygen, 2,2-diphenyl-1-picrylhydrazyl, nitric oxide, and tyrosyl radicals) were directly quantified using an X-band ESR spectrometer with the spin-trapping method. IC50 and reaction rate constants were estimated from the dose-response curve for each free radical. Landiolol scavenged six of the free radical species examined: hydroxyl radical (IC50 = 0.76 mM, k landiolol = 1.4 × 10‍10 M‍-1 s‍-1, p<0.001), superoxide anion (58 mM, 2.1 M‍-1 s‍-1, p = 0.044), tert-butoxyl radical (4.3 mM, k landiolol/k CYPMPO = 0.77, p<0.001), ascorbyl free radical (0.31 mM, p<0.001), singlet oxygen (0.69 mM, k landiolol/k 4-OH ‍TEMP = 2.9, p<0.001), and nitric oxide (15 mM, 1.7 × 10 M‍-1 s‍-1, p<0.001). This study is the first to report that landiolol dose-dependently scavenges multiple free radical species with different reaction rate constants. These results indicate the potential clinical application of landiolol as an antioxidative and anti-inflammatory agent in addition to its present clinical use as an anti-arrhythmic agent.

Ascorbate-and iron-driven redox activity of Dp44mT and Emodin facilitates peroxidation of micelles and bicelles

BackgroundIron (Fe)-induced oxidative stress leads to reactive oxygen species that damage biomembranes, with this mechanism being involved in the activity of some anti-cancer chemotherapeutics. MethodsHerein, we compared the effect of the ligand, di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT), or the potential ligand, Emodin, on Fe-catalyzed lipid peroxidation in cell membrane models (micelles and bicelles). These studies were performed in the presence of hydrogen peroxide (H2O2) and the absence or presence of ascorbate. ResultsIn the absence of ascorbate, Fe(II)/Emodin mixtures incubated with H2O2 demonstrated slight pro-oxidant properties on micelles versus Fe(II) alone, while the Fe(III)-Dp44mT complex exhibited marked antioxidant properties. Examining more physiologically relevant phospholipid-containing bicelles, the Fe(II)- and Fe(III)-Dp44mT complexes demonstrated antioxidant activity without ascorbate. Upon adding ascorbate, there was a significant increase in the peroxidation of micelles and bicelles in the presence of unchelated Fe(II) and H2O2. The addition of ascorbate to Fe(III)-Dp44mT substantially increased the peroxidation of micelles and bicelles, with the Fe(III)-Dp44mT complex being reduced by ascorbate to the Fe(II) state, explaining the increased reactivity. Electron paramagnetic resonance spectroscopy demonstrated ascorbyl radical anion generation after mixing ascorbate and Emodin, with signal intensity being enhanced by H2O2. This finding suggested Emodin semiquinone radical formation that could play a role in its reactivity via ascorbate-driven redox cycling. Examining cultured melanoma cells in vitro, ascorbate at pharmacological levels enhanced the anti-proliferative activity of Dp44mT and Emodin. Conclusions and general significanceAscorbate-driven redox cycling of Dp44mT and Emodin promotes their anti-proliferative activity.

The Mimic Enzyme Properties of Au@PtNRs and the Detection for Ascorbic Acid Based on Their Catalytic Properties

Being superior to natural enzymes, nanoenzymes are drawing a great deal of attention in the field of biosensing. Herein, we developed an ultrasensitive, stable and selective colorimetric assay having dual functionalities of Au-tipped Pt nanorods (NRs). The optical and catalytic properties of Au-tipped Pt NRs were monitored using a spectrophotometer and the chromogenic substrate 3, 3′, 5, 5′-tetramethylbenzidine (TMB) in the presence of H2O2, respectively. We found that Au-tipped Pt NRs exhibited excellent peroxidase-like activity, which decomposed hydrogen peroxide (H2O2) into oxygen (O2). The produced O2 oxidized the chromogenic substrate into a blue color product. The oxidation rate of the chromogenic substrate could be monitored using a spectrophotometer at 652 nm. Notably, the peroxidase-like activity of Au-tipped Pt NRs decreased in the presence of ascorbic acid (AA). The produced O2 preferentially reacted with AA, generating ascorbyl radicals (AA·) instead of oxidizing TMB, and thereby decreased the oxidation rate of TMB. Based on this inhibitory property, a selective colorimetric assay was developed using Au-tipped Pt NRs for the detection of AA. This work offers a novel detection method for AA.

One electron oxidation of ascorbic acid facilitated by ionic liquid-doped poly (3, 4-ethylenedioxythiophene) as artificial enzyme

Abstract Ascorbic acid (AA) play vital roles in various physiological processes and its two-electron oxidation pathway is typically documented. In this study, we reported that ionic liquid (IL) 1-ethyl-3-methylimidazolium bis(trifluoromethyl sulfonyl) imide served as unique dopant to tune the surface electrical properties of poly (3, 4-ethylenedioxythiophene) (PEDOT), with a charge reversal from initially positively charged surface to partially negative after IL doping, as evidenced by cyclic voltammetry tests using redox probes (Ru(NH3)6Cl3 and K3Fe(CN)6). The IL-doped PEDOT were characterized via SEM, AFM and XPS techniques and functionalized as artificial enzyme to enable precisely one electron oxidation of ascorbic acid toward ascorbyl radical. For two other interferences, uric acid (UA) and dopamine (DA), IL-doped PEDOT displayed one-electron oxidation behavior as well. The fabricated IL-doped PEDOT artificial enzyme enabled the simultaneous electrochemical sensing of AA, DA and UA with wide linear ranges (50–1000 μM for AA, 2.5–1000 μM for DA and 4–350 μM for UA). The underlying catalytic mechanism was approached with molecular calculation, revealing that strong π-π and hydrogen bond interactions at the molecular level contributed to the enhanced catalytic effect. In general, this study innovatively reveals the inherent catalytic activity of IL-doped conductive polymer as artificial enzyme for sensing application.

AN EXPERIMENTAL AND THEORETICAL STUDY OF THE REACTIVITY OF SELECTED CATECHOLAMINES AND THEIR PRECURSORS TOWARDS ASCORBYL RADICAL

Ascorbyl radical is often used as a biomarker of oxidative stress in human organism. It is a product of the antioxidant activity of ascorbic acid and it is not expected to react further with biologically important molecules. For the first time, the reactivity of catecholamines and their precursors was investigated theoretically and experimentally towards ascorbyl radical and the main structural parameters governing activity were analyzed. It was shown that catechol moiety plays an important role, which classifies norepinephrine and 3,4-dihydroxyphenylacetic acid as the most reactive when compared to homovanillic acid, vanillylmandelic acid, and octopamine. DFT methods have been employed to determine the most probable mechanism of the reaction. Based on the change in reaction enthalpy it was concluded that Hydrogen Atom Transfer (HAT) is a preferred mechanism both in water and pentyl ethanoate. The stabilization interactions in ascorbic acid, its anion and radical are carefully analyzed in order to understand the preferability of the mentioned mechanism. The stability of the ascorbyl radical is explained in detail. The results prove that ascorbyl radical is not just a product of antioxidant activity, but also a potentially harmful species in an organism.

Capping 1,3-propanedithiol to boost the antibacterial activity of protein-templated copper nanoclusters

We have prepared copper nanoclusters (Cu NCs) in the presence of bovine serum albumin (BSA) and 1,3-propanedithiol (PDT). The PDT/BSA–Cu NCs possess great activities against different types of bacteria, including non-multidrug-resistant bacteria (Escherichia coli, Salmonella Enteritidis, Pseudomonas aeruginosa, and Staphylococcus aureus) and multidrug-resistant bacteria (methicillin-resistant S. aureus). Their minimal inhibitory concentration (MIC) values are at least 242-fold and 10-fold lower than that of the free PDT and BSA–Cu NCs, respectively. The PDT/BSA–Cu NCs are strongly bound to the bacterial membrane, in which they induce the generation of ascorbyl (Asc) and perhydroxyl (HOO) radicals that result in disruption of their membrane integrity. At a concentration of 100-fold higher than their MIC for Escherichia coli, the PDT/BSA–Cu NCs exhibit negligible cytotoxicity towards the tested mammalian cells and show insignificant hemolysis. We have further demonstrated that low-cost PDT/BSA–Cu NCs-coated carbon fiber fabrics (CFFs) are effective against antibacterial growth, showing their great potential for antifouling applications.

Liposomal integration method for assessing antioxidative activity of water insoluble compounds towards biologically relevant free radicals: example of avarol

The liposomal integration method, in conjunction with electron paramagnetic resonance (EPR) spectroscopy, has been presented for the investigation of antioxidant activity of selected water-insoluble compound towards biologically relevant free radicals. This method was applied to avarol, a sesquiterpenoid hydroquinone isolated from the marine sponge Dysidea avara. The antioxidant activity of water-insoluble avarol towards •OH, O2•− and NO• radicals was attained by its incorporation into the DPPC liposomes bilayer, and towards ascorbyl radicals in the organic solvent. Avarol’s activity towards •OH, O2•−, NO• and ascorbyl radicals was 86.2%, 50.9%, 23.6% and 61.8%, respectively, showing its significant radical scavenging potential.

Ascorbyl stearate stimulates cell death by oxidative stress-mediated apoptosis and autophagy in HeLa cervical cancer cell line in vitro.

In this study, Asc-s was evaluated for anti-cancer effect using cervical cancer cells (HeLa). Results determine that Asc-s treatment-induced dose-dependent inhibition of proliferation of HeLa cells and induced apoptosis. Flow-cytometry analysis shows Asc-s treatment-induced accumulation of cells at sub-G0/G1 stage of cell cycle and induced apoptosis as confirmed by DAPI, propodium iodide, and acridine staining in HeLa cells. Asc-s entered the cells and metabolized to ascorbate and stearate moieties, increased membrane permeability, and decreased membrane fluidity in HeLa cells. Asc-s treatment-induced dose-dependent increase in autophagy protein LC3-II, mRNA levels and decreased Nrf-2 levels in HeLa cells. It is hypothesized that both ascorbyl radical and stearoyl moieties of Asc-s induced cytotoxicity by generating reactive oxygen species (ROS) and modulating membrane fluidity/permeability leading to apoptosis/autophagy of HeLa cells. Thus, our findings demonstrate that Asc-s as anti-proliferative and apoptosis inducing compound in cervical cancer cells.

Ascorbyl and hydroxyl radical generation mediated by a copper complex adsorbed on gold.

This work presents the results obtained for a thioether derivative of bipyridine, (E,Z)-1-(4'-methyl-[2,2'-bipyridine]-4-yl)-N-(4(methylthio)phenyl)methanimine (4-mbpy-Bz-SMe), and its copper complex [CuII(4-mbpy-Bz-SMe)2]2+. Electronic spectra acquired at 183 K of the cuprous complex [CuI(4-mbpy-Bz-SMe)2]+ generated in situ indicated the formation of the peroxodicopper compound {[CuII(4-mbpy-Bz-SMe)2]2(μ-O22-)}2+. A gold electrode modified with [CuII(4-mbpy-Bz-SMe)2]2+ (Au/[Cu]) was fully characterized by SERS spectroscopy, electrochemistry and impedance spectroscopy thus showing adsorption occurs through the sulfur atom of the 4-mbpy-Bz-SMe moieties. DNA cleavage assays showed the copper complex, in solution and adsorbed on gold, degrades DNA if reducing conditions are maintained, i.e. ascorbic acid (H2AA) in solution or applied potentials more negative than 0.12 V vs. Ag/AgCl (CuI form). The electron paramagnetic resonance (EPR) spectra obtained for the electrolyzed solution (Eapl = -0.2 V, no H2O2) and for the solution containing [CuII(4-mbpy-Bz-SMe)2]2+ and H2O2 showed hydroxyl radical, HO˙, generation had occurred. The cyclic voltammograms obtained with H2AA in solution at Au/[CuII(4-mbpy-Bz-SMe)2]2+ as the working electrode showed a one-electron reaction leading to the ascorbyl radical (HA˙), which was detected by EPR. The current assigned to the electrode oxidation of HA˙ to AA decreased with the addition of catalase, a scavenger of H2O2, meaning peroxide is involved in the mechanism.

Gain of function conferred by selenocysteine: catalytic enhancement of one-electron transfer reactions by thioredoxin reductase.

Selenocysteine (Sec) is the 21st amino acid in the genetic code and it is present in a small number of proteins where it replaces the much more commonly used amino acid cysteine (Cys). It is both more complicated and bioenergetically costly to insert Sec into a protein in comparison to Cys, and this cost is most likely compensated by a gain of function to the enzyme/protein in which it is incorporated. Here we investigate one such gain of function, the enhancement of one-electron transfer catalysis. We compared the ability of Sec-containing mouse mitochondrial thioredoxin reductase (mTrxR2) to catalyze the reduction of bovine cytochrome c, ascorbyl radical, and dehydroascorbate in comparison to Cys-containing thioredoxin reductases from D. melanogaster (DmTrxR) and P. falciparum (PfTrxR). The Sec-containing mTrxR2 was able to reduce all three substrates, while the Cys-containing enzymes had little or no activity. In addition, we constructed Cys➔Sec mutants of DmTrxR and PfTrxR and found that this substitution resulted in a gain of function, as these mutant enzymes were now able to catalyze the reduction of these substrates. We also found that in the case of PfTrxR, reduction of cytochrome c was enhanced five-fold in a truncated PfTrxR in which the C-terminal redox center was removed. This shows that some of the ability of thioredoxin reductase to reduce this substrate comes from the flavin coenzyme. We also discuss a possible mechanism by which Sec-containing thioredoxin reductase reduces dehydroascorbate to ascorbate by two sequential, one-electron reductions, in part catalyzed by Sec.

Micellized Tris(bipyridine)ruthenium Catalysts Affording Preparative Amounts of Hydrated Electrons with a Green Light-Emitting Diode.

We have explored alkyl substitution of the ligands as a means to improve the performance of the title complexes in photoredox catalytic systems that produce synthetically useable amounts of hydrated electrons through photon pooling. Despite generating a super-reductant, these electron sources only consume the bioavailable ascorbate and are driven by a green light-emitting diode (LED). The substitutions influence the catalyst activity through the interplay of the quenching parameters, the recombination rate of the reduced catalyst OER and the ascorbyl radical across the micelle-water interface, and the quantum yield of OER photoionization. Laser flash photolysis yields comprehensive information on all these processes and allows quantitative predictions of the activity observed in LED kinetics, but the latter method provides the only access to the catalyst stability under illumination on the timescale of the syntheses. The homoleptic complex with dimethylbipyridine ligands emerges as the optimum that combines an activity twice as high with an undiminished stability in relation to the parent compound. With this complex, we have effected dehalogenations of alkyl and aryl chlorides and fluorides, hydrogenations of carbon-carbon double bonds, and self- as well as cross-coupling reactions. All the substrates employed are impervious to ordinary photoredox catalysts but present no problems to the hydrated electron as a super-reductant. A particularly attractive application is selective deuteration with high isotopic purity, which is achieved simply by using heavy water as the solvent.