Decay Kinetics Of Radicals Research Articles

Electrocatalytic amplification of coreactant electrochemiluminescence using redox mediators

Electrochemiluminescence (ECL) has emerged as a sensitive analytical technique with a wide range of applications. While recent studies have begun to explore the role of redox mediators to facilitate ECL systems, this work extends these foundational insights by developing a comprehensive theoretical framework that supports, generalizes, and rationalizes mechanistic pathways borrowed from molecular electrocatalysis. This advancement is demonstrated through the electrocatalytic amplification of coreactant ECL within the [Ru(bpy)3]2+/TPrA system, utilizing a water-soluble redox-active Ir(III) complex as an electrocatalyst. Our investigation unveils previously uncharted mechanisms that enable the enhancement of Ru complex ECL at low electrode potentials, crucially without requiring the direct oxidation of [Ru(bpy)3]2+ or TPrA at the electrode, thus offering a deeper understanding of ECL activation through moelcular electrocatalysis. Through an integrated approach combining electrochemical analysis, spectroscopic investigation, and finite element modeling, we elucidate how the redox mediator critically modulates ECL efficiency by controlling the kinetics of radical species production and decay. The mediation by the Ir(III) complex not only governs the generation of TPrA radicals and excited [Ru(bpy)3]2+ states, thereby enhancing ECL intensity, but also the conditions under which ECL can be quenched. Further exploration of the mediator's redox characteristics provides predictive insights into ECL behavior, underscoring the mediator's redox potential as pivotal in determining ECL onset, peak potential, and intensity. This refined understanding paves the way for tailoring ECL systems for enhanced performance in analytical, imaging, and biomedical applications through the judicious selection of redox mediators.

ЭПР-исследование кинетики распада свободных радикалов облученной сахарозы

Electron paramagnetic resonance (EPR) spectrometry makes it possible to study the formation of free radicals during the transfer of energy to the object. The decay kinetics of free radicals is an integral part of the study since this phenomenon directly interferes with the goals of accurate identification of the irradiation fact and dosimetry using EPR spectrometry. In this work, we have studied the time dependence of the intensity and characteristics of the EPR signal of analytical sucrose irradiated with gamma rays at doses from 0.3 to 9 kGy. It has been shown that irradiated sucrose gives a stable EPR peak after 60 days. A change in the spectral characteristics of the signal was recorded in the first 23 hours after irradiation. The signal intensity tends to increase in the first 72 hours after irradiation for a sample with an irradiation dose of 300 Gy and 48 hours for 1000 Gy. The dependence of the signal intensity on the absorbed dose is linear in the range of studied doses. The g-factor value at the intersection point of the derivative contour with the zero line is 2.013. Based on the data obtained, it can be assumed that sucrose (sugar) is one of the best candidates among solid radiation-sensitive materials for identifying the fact of irradiation using EPR spectrometry

Recombination of Poly(Acrylic Acid) Radicals in Acidic Aqueous Solutions: A Pulse Radiolysis Study

Carbon-centered radicals have been randomly generated on the chains of poly(acrylic acid), PAA, the simplest synthetic anionic polyelectrolyte, by pulse-irradiating its dilute, oxygen-free aqueous solutions by 6 MeV electron beam. In some experiments, oligo(acrylic acid), OAA, and propionic acid, PA, were used as PAA models. Recombination kinetics of PAA radicals has been followed by fast spectrophotometry. A strong pH dependence of radical lifetime on pH, and thus on the linear charge density due to deprotonated carboxylate groups, has been confirmed, while a weaker amplitude of pH dependence was observed for OAA and PA. Decay kinetics of PAA radicals in the protonated state, at pH 2, have been studied in some detail. At moderate doses of ionizing radiation, resulting in a moderate average initial number of radicals per chain, ZR0, the decay can be satisfactorily described by a second-order kinetic model, but a somewhat better fit is obtained by using a dispersive kinetics approach. While for a constant polymer concentration the reciprocal half-lives are proportional to the initial radical concentrations, such a data series for different PAA concentrations do not overlap, indicating that the overall radical concentration is not the decisive factor controlling the kinetics. Arranging all data, in the form of second-order rate constants, as a function of the average initial number of radicals per chain allows one to obtain a common dependence. The latter seems to consist of two parts: a horizontal one at low ZR0 and another one of positive slope at higher ZR0. This is interpreted as two kinetic regimes where two distinct reactions dominate, intermolecular and intramolecular recombination, respectively. Comparison of the low ZR0 data with calculations based on the translational diffusion model indicate that the latter is not the rate-controlling process in intermolecular recombination of polymer radicals; segmental diffusion is the more likely candidate.

Numerical Simulation of the Kinetics of Radical Decay in Single-Pulse High-Energy Electron-Irradiated Polymer Aqueous Solutions.

A new method for the numerical simulation of the radiation chemistry of aqueous polymer solutions is introduced. The method makes use of a deterministic approach combining the conventional homogeneous radiation chemistry of water with the chemistry of polymer radicals and other macromolecular species. The method is applied on single-pulse irradiations of aqueous polymer solutions. The speciation of macromolecular species accounts for the variations in the number of alkyl radicals per chain, molecular weight, and number of internal loops (as a consequence of an intramolecular radical-radical combination). In the simulations, the initial polymer molecular weight, polymer concentration, and dose per pulse (function of pulse length and dose rate during the pulse) were systematically varied. In total, 54 different conditions were simulated. The results are well in line with the available experimental data for similar systems. At a low polymer concentration and a high dose per pulse, the kinetics of radical decay is quite complex for the competition between intra- and intermolecular radical-radical reactions, whereas at a low dose per pulse the kinetics is purely second-order. The simulations demonstrate the limitations of the polymer in scavenging all the radicals generated by water radiolysis when irradiated at a low polymer concentration and a high dose per pulse. They also show that the radical decay of lower-molecular-weight chains is faster and to a larger extent dominated by intermolecular radical-radical reactions, thus explaining the mechanism behind the experimentally observed narrowing of molecular weight distributions.

Cellular responses to radical propagation from ion-implanted plasma polymer surfaces

Biomolecule-functionalization, through the presentation of biological motifs that promote optimal cellular responses, has the capacity to improve the tissue integration of biomedical devices and hence patients’ quality of life. Radical-functionalized plasma polymer films (rPPFs) readily immobilize bioactive molecules on exposure to a biomolecule-containing aqueous solution without the need for chemical reagents. However, the potential for damage to cells and tissues due to the high local concentration of radicals in freshly deposited radical-functionalized plasma polymer films is of concern. In this study, we compared a fresh (4 h post-deposition) rPPF with one that had been aged for 11 days to explore the effect of the different radical fluxes on cellular responses. Primary osteoblasts and MG63 bone osteosarcoma cells were used to determine whether rPPFs at early aging times exhibited radical-induced cytotoxicity. The aging behavior of the rPPFs demonstrated a connection between the radical decay kinetics and the surface chemistry and wettability. Significant increases in cell attachment and spreading compared to bare Ti were observed for both cell lineages on the rPPF surfaces. The proliferation assays showed equivalent proliferation rates on both the fresh and aged surfaces, and no evidence of cytotoxicity was observed. Overall, we demonstrated that the high flux of radicals emerging to the surface has minimal influence on the biocompatibility of radical-functionalized plasma polymer films.

Formation and decay of radicals during Vacuum-UV irradiation of poly(dimethylsiloxane)

Formation of radicals in a poly(dimethylsiloxane) (PDMS) elastomer upon 172 nm vacuum-UV (VUV) irradiation was investigated by electron paramagnetic resonance spectroscopy (EPR). Kinetics of radical buildup and decay was investigated under varying conditions and found to depend strongly on the degree of pre-modification of the polymer. Furthermore, radicals were observed only when oxygen was absent during in situ VUV irradiation. The EPR spectrum is dominated by a broad signal, assignable to unspecified carbon-centered radicals. Stationary radical concentrations during VUV irradiation in nitrogen atmosphere can reach 0.2 mol/L in an about 260 nm thick surface layer, consistent with the remaining carbon concentration in the modified PDMS as measured by XPS. Gas diffusion is not limiting kinetics of the modification process. Our findings are consistent with a mechanism where oxygen molecules from the atmosphere diffuse in PDMS and quench radical sites in the polymer.

The Effect of Low Pressure Plasma Polymerization Modes on the Properties of the Deposited Plasma Polymers

This work characterizes the properties of deposited plasma polymer that has been generated using three different low-pressure, low-temperature plasma modes: magnetron-enhanced audio frequency (15 kHz), magnetron-enhanced radio frequency (13.56 MHz), and radio frequency (13.56 MHz). A correlation between the applied plasma modes, the concentration of generated radicals in the deposited nanofilms, and the radical decay kinetics was found and is discussed. In addition, the role of the plasma driving frequency with respect to the properties of the films, in particular their density and their barrier properties, has been elucidated.

EPR and kinetic studies of overall free radical decay in Gamma-irradiated UHMWPE used in joint arthroplasty

Medical-grade UHMWPE is widely used in an artificial joint and is commonly treated with γ-irradiation for sterilization and crosslinking. A vacuum annealing treatment is essential in eliminating the radicals generated by γ-irradiation and in maintaining the long-term stability of the material. It is important to better understand the kinetics of radical decay in an annealing process. In this study, the kinetics of overall radical decay in a γ-irradiated UHMWPE with doses of 25, 50, and 100 kGy at various temperatures was investigated. The overall free radical concentration was monitored by EPR technique and found to exhibit a fast-to-slow transition over time, which has been successfully simulated by a parallel 2nd-order reaction model proposed herein. The model includes two simultaneous but isolated reactions to describe the fast radical decay in amorphous and the slow radical decay in crystalline regions. Results showed that irradiation dose has limited effect on reaction rate constant and their activation energies in amorphous and crystalline regions are approximately 3–4 and 12–14 kcal/mol, respectively. The current parallel 2nd-order reaction model can be served as a useful tool in predicting the residue radical level of a vacuum annealing process.

Hydroxyl radical-induced crosslinking and radiation-initiated hydrogel formation in dilute aqueous solutions of carboxymethylcellulose

Ionizing radiation causes chain scission of polysaccharides in the absence of crosslinking agents. It has been demonstrated before that degradation of carboxyalkylated polysaccharides may be prevented, despite presence of strong electrostatic repulsing forces between chains, at very high polymer concentration in water (paste-like state) when physical proximity promotes recombination of radiation-generated polymer radicals. In such conditions, crosslinking dominates over chain scission and covalent, macroscopic gels can be formed. In an approach proposed in this work, neutralizing the charges on carboxymethylcellulose (CMC) by lowering the pH results in retracting the electrostatic repulsion between chain segments and thus allows for substantial reduction of polymer concentration required to achieve gelation due to domination of crosslinking reactions. Electron-beam irradiation of aqueous solutions of low pH containing 0.5–2% CMC results in hydrogel formation with 70% yield, while both concentration and dose determine their swelling properties. Time-resolved studies by laser flash photolysis clearly indicate strong pH influence on decay kinetics of CMC radicals.

Decay Kinetics of Benzophenone Triplets and Corresponding Free Radicals in Soft and Rigid Polymers Studied by Laser Flash Photolysis

The kinetics of transients formed under photoexcitation of benzophenone (B) dissolved in three different polymers was studied by ns laser flash photolysis. These polymers were the soft rubbers poly (ethylene-co-butylene) (EB), polystyrene block-poly(ethylene-ran-butylene)-block-polystyrene (SEBS) and hard polystyrene (PS). We monitored the decay kinetics of triplet state (3)B(*) and of ketyl radicals BH(●). We observed exponential decay of (3)B(*) and two-stage decay kinetics of BH(●) in EB. The first stage is a fast cage recombination of a radical pair (BH(●), radical of polymer R(●)). The second slow stage of BH(●) decay follows the second-order law with a relatively high rate constant, which corresponds to recombination of BH(●) in a homogeneous liquid with a viscosity of only ~0.1 P (about five times of 2-propanol viscosity). Application of a magnetic field (MF) of 0.2 T leads to deceleration of both stages of BH(●) decay in EB by approximately 20%. Decay kinetics of both transients were observed in SEBS. There was no MF effect on BH(●) decay in SEBS. We only observed (3)B(*) in PS. Decay kinetics of (3)B(*) in this case were described as polychromatic dispersive first-order kinetics. We discuss the effects of polymer structure on transient kinetics and the MF effect.

ANTIOXIDANT POTENTIAL IN LIPOPHILIC AND HYDROPHILIC EXTRACTS FROM MEDICINAL HERBS (<i>SALVIA OFFICINALIS</i> AND <i>ECHINACEA ANGUSTIFOLIA</i>). A COMPARISON BETWEEN ASSAYS BASED ON ELECTRON PARAMAGNETIC RESONANCE AND SPECTROPHOTOMETRY

Recently, detection of antioxidant activity of plant extracts has caught the attention of researchers. Determination of antioxidant activity is usually performed using stable free radicals. A number of assays is available for screening the antioxidant activity of food constituents, but few of these methods are applicable to lipophilic substances. EPR is the analytical technique that directly measures free radicals making possible the analysis of turbid or highly colored samples. The aim of this study was to determine how the detection technique, plant species and the different growing conditions influenced the antioxidative activity of aqueous and lipid extracts from leaves. Sage (Salvia officinalis L.) and coneflower (Echinacea angustifolia DC.) grown in hydroponics and in soil, were analyzed. Radical decay kinetics, due to plant extracts, were monitored by an EPR spectrometer, equipped with a data acquisition system and a software package designed for analysis and simulation of spectra. Parameters obtained by the fitting of the curves allowed the calculation of the antioxidant potential. The radicals Fremy’s salt (hydrophilic) and 1,1-diphenyl-2-picrylhydrazyl (DPPH, lipophilic) were employed. Comparisons with data obtained by a spectrophotometric detection with the radical cation ABTS+ [2,2’-azinobis(3-ethylbenzothiazoline-6-sulfonic acid)] were also made. Fremy’s salt and DPPH gave 1st and 2nd order kinetics, respectively. EPR can give more reliable measurements of the antioxidant potential than spectrophotometric assay. The reduced sensitivity of spectrophotometric detection was showed both for sage and coneflower extracts. Antioxidant potential of sage resulted extraordinary high compared to coneflower and a different ratio of aqueous to lipid antioxidant potential in the two species could be monitored. Antioxidant potentials were also higher for coneflower grown in hydroponics than in soil. In addition to the technique adopted, the detection of antioxidant potential was influenced by the growing conditions of plants and hydroponics showed to be able to increase antioxidant potential of both coneflower lipophilic and hydrophilic extracts. By registration of EPR kinetics we can get reliable measurements of the antioxidant potential of plant extracts, avoiding the artifacts due to the appearance of other radicals resulting from oxidative processes.

Laser Flash Photolysis of Benzophenone in Polymer Films

With a nanosecond laser we studied flash photolysis of benzophenone (BP) dissolved in four different polymer films. We measured kinetics of decay of a triplet state of benzophenone (3)BP as well as kinetics of decay of benzophenone ketyl free radicals BPH(•). Polymer matrices have plenty of reactive C-H bonds, and the hydrogen abstraction by (3)BP leads to a formation of geminate pair which either recombines into molecular products or dissociates. Decay kinetics of (3)BP is well described by dispersive kinetics and in particular by the kinetic law suggested in Albery, W. J.; et al. J. Am. Chem. Soc. 1985, 107, 1854. We observed a broader distribution of rate constants in hard films. It was observed that the decay kinetics of transients radicals in the "hard" polymers is quite satisfactory described by the same law for dispersive kinetics. Kinetics of radicals decay in "soft" polymers is satisfactorily described as a diffusion-enhanced reaction. Effect of a hardness of polymer matrix on the measured kinetic parameters is discussed.

Measurements of superoxide radical concentration and decay kinetics in the Gulf of Alaska

Numerous laboratory studies have shown that superoxide radical ( O 2 − ) can have a profound influence on the redox speciation of dissolved metals, especially copper, manganese, and iron. We made direct measurements of O 2 − concentrations and decay kinetics in seawater samples collected from the Gulf of Alaska, using the recently developed method of methyl Cypridina luciferin analog (MCLA) chemiluminescence. Concentrations ranged from <0.02 to ∼0.6 nmol L −1, with higher concentrations typically observed in the upper 1–10 m of the water column. Pseudo-first-order decay rate coefficients ranged 0.002–0.02 s −1, and confirm that even in the open ocean, uncatalyzed dismutation is not the major fate of O 2 − . Calculated O 2 − production rates varied from <1 to ∼20 nmol L −1 h −1 and appear to be primarily of biological origin. These results represent the first direct measurements of O 2 − concentrations in a non-tropical region, and are broadly similar to recently published findings for the eastern equatorial Pacific, suggesting that biological production of O 2 − may be a common feature of the ocean’s surface waters. Our study was also the first to examine the reproducibility of measurements of in situ O 2 − concentrations and decay coefficients. Subsurface samples collected using GO-FLO bottles exhibited a systematic increase in concentration and decay rate coefficient with time, a possible artifact that should be considered in future studies. In contrast, measurements conducted using “fish” samples under low or no light conditions were found to be reliably reproducible, and demonstrated substantial rates of biological O 2 − production.

ESR investigations on γ-ray irradiated 3-methyl nylon 3

Electron spin resonance spectroscopy investigations on γ irradiated 3-methyl nylon 3 (poly-3-methyl β-alanine) are reported. The resonance spectra (recorded after the irradiation in nitrogen atmosphere has been stopped) have been attributed to the parallel and perpendicular components of a triplet line assigned to the delocalization of the uncoupled electron over an effective nuclear spin 1. It was suggested that this effective spin arises from the fast tunneling/rotation of a proton between two positions. The resonance spectra have been simulated with accuracy by using a simplified spin Hamiltonian and assuming Lorentzian-like resonance line shapes and axial asymmetry of the resonance line due to the trapping of free radicals in randomly oriented crystallites. The time evolution of free radicals in nitrogen atmosphere at room temperature has been analyzed. The decay kinetics of stable free radicals in 3-methyl nylon 3 (under inert atmosphere) has been investigated. It was shown that the radiation-induced radicals in inert atmosphere decay through a unimolecular reaction.

Kinetics of bimolecular radical decay in different polymeric matrices

AbstractUsing electron spin resonance spectroscopy, we have studied the kinetics of the bimolecular decay of methacrylic macroradicals in networked and unnetworked polymeric matrices. We have determined that macroradical decay in the entire range of temperatures (20–70°C) is definitely ordered to a second‐order kinetic reaction equation. We have estimated the activation parameters of the process. Using Smolukhovsky's and Einstein's equations and the fractal dependence of the conformation radius of the macroradical on its polymerization degree, we have obtained an equation for the constant of bimolecular radical decay under diffusion conditions. This equation is in good agreement with all the experimental data and possesses a commonness permitting its use in the analysis of bimolecular chain termination in different reaction zones via polymerization processes until high conversions. © 2007 Wiley Periodicals, Inc. J Appl Polym Sci 2007

ESR and FTIR studies on electron beam‐irradiated low‐density polyethylene blends

AbstractLow‐density polyethylene (LDPE) composites modified with a resin based on ethylene/methacrylic acid copolymer (surlyn) and/or citric acid were electron beam‐irradiated and investigated by electron spin resonance (ESR) at room temperature. ESR studies were carried out directly after irradiation and after various time intervals up to 72 h postirradiation. The irradiated samples showed the ESR spectrum of seven lines that was assigned to the formation of allyl radical. The nature and yield of the allyl radical of the different LDPE samples were analyzed as a function of time after irradiation. Also, the radical concentration, decay, decay rate, and kinetics of radical decay were evaluated. Fourier transform infrared (FTIR) analysis at a series of different temperatures upon cooling from room temperature to −175°C and the reverse heating to +125°C was also carried out. The structural changes while cooling and heating of LDPE samples were investigated using FTIR spectrometry. The results showed that cooling of unirradiated LDPE samples to −175°C results in a decrease of the intensities of IR bands. However, heating the samples from −175°C up to +125°C led to a consequence increase in the intensities of the IR bands. © 2006 Wiley Periodicals, Inc. J Appl Polym Sci 103: 3461–3469, 2007

Enzymatic oxidation of dipyridamole in homogeneous and micellar solutions in the horseradish peroxidase–hydrogen peroxide system

Enzymatic oxidation of dipyridamole (DIP) by horseradish peroxidase–hydrogen peroxide system (HRP–H 2O 2) in aqueous and micellar solutions was carried out. The reaction was monitored by optical absorption and fluorescence techniques. In aqueous solution at pH 7.0 and pH 9.0, the disappearance of the characteristic bands of DIP centered at 400 nm and 280 nm was observed. A new strong band at 260 nm is observed for the oxidation product (s) with shoulders at 322 nm and 390 nm. A non-fluorescent product is formed upon oxidation. In cationic cethyl trimethyl-1-ammonium chloride (CTAC) and zwitterionic 3-( N-hexadecyl- N, N-dimethylammonium) propane sulfonate (HPS) micellar solutions the same results are observed: three, well-defined, isosbestic points in the optical spectra suggest the transformation between two species. In anionic micellar sodium dodecylsulfate solution (SDS), the appearance of a new band centered around 506 nm was observed, associated to a solution color change from the usual yellow to deep blue/violet, characteristic of a radical species associated to the one-electron oxidation of DIP to its cation radical ( DIP· +), observed previously in electrochemical oxidation. Experiments of radical decay kinetics monitoring the absorbance change at 506 nm were performed and analyzed in the frame of a kinetic model taking into account the species both in homogeneous and micellar media. The reaction medium is composed of bulk solution, SDS micelle/solution interface and enzyme catalytic site (s). The variation of DIP· + concentration was analyzed assuming: (1) synthesis of DIP· + by HRP through one-electron oxidation; (2) decomposition of DIP· + by further one-electron oxidation; (3) direct two-electron oxidation of DIP by HRP; (4) bimolecular DIP· + disproportionation. The main results of the analysis are as follows: (1) kinetic data can be divided in two phases, an HRP active phase and another phase which proceeds in the absence of enzyme activity due to consumption of all H 2O 2; (2) the reactions of DIP· + formation, DIP· + decomposition and DIP two-electron oxidation are HRP concentration dependent; (3) since DIP· + formation constant seems to be overestimated, it is proposed that two-electron oxidation is another source of DIP· +, through the comproportionation reaction. Evidences for this reaction were also observed previously in electrochemical experiments; and (4) the kinetic analysis provides evidences that the bimolecular reaction of DIP· + takes place mainly in the absence of active HRP and in this phase the combination of, at least, two second-order kinetic processes is needed to model the experimental data. Our data suggest that HRP oxidizes DIP in general by a two-electron process or that the cation radical is very unstable so that the one-electron process is only detected in the presence of anionic surfactant, which stabilizes significantly the DIP· + intermediate.

Kinetics of radical formation and decay in photooxidation of 4-halophenols sensitized by 4-carboxybenzophenone in aqueous solutions

Kinetics of formation and recombination of radicals formed by quenching of the triplet state of 4-carboxybenzophenone (CB) with para-substituted phenol derivatives RC6H4OH (R = OMe, H, Cl, Br, I) in aqueous solutions was studied by nanosecond laser photolysis. At pH ≥ 5.4, quenching proceeds with high rate constants ((1–3)⋅109 L mol−1 s−1) through electron transfer to form the radical anion CB⋅− and radical cation RC6H4OH⋅+. The latter is transformed into the phenoxyl radical within ≤10 ns. At pH ≤ 8, the CB⋅− radical anion is protonated in a phosphate buffer with the rate constant increasing from 4⋅106 to 15⋅106 s−1 with a decrease in the pH from 8 to 5.4. The yield of radicals decreases from 100 to 13% as the atomic weight of halogen in the RC6H4OH molecule increases due to an increase in the probability of recombination of the primary triplet radical pair in the solvent cage and partial intersystem crossing in an encounter complex (3CB, RC6H4OH). The effect of heavy atom is also observed in the kinetics of volume recombination of the radicals, the magnitude of effect corresponds to the acceleration of the primary recombination of the triplet radical pair.

The Three Nitric-oxide Synthases Differ in Their Kinetics of Tetrahydrobiopterin Radical Formation, Heme-Dioxy Reduction, and Arginine Hydroxylation

The nitric-oxide synthases (NOSs) make nitric oxide and citrulline from l-arginine. How the bound cofactor (6R)-tetrahydrobiopterin (H4B) participates in Arg hydroxylation is a topic of interest. We demonstrated previously that H4B radical formation in the inducible NOS oxygenase domain (iNOSoxy) is kinetically coupled to the disappearance of a heme-dioxy intermediate and to Arg hydroxylation. Here we report single turnover studies that determine and compare the kinetics of these transitions in Arg hydroxylation reactions catalyzed by the oxygenase domains of endothelial and neuronal NOSs (eNOSoxy and nNOSoxy). There was a buildup of a heme-dioxy intermediate in eNOSoxy and nNOSoxy followed by a monophasic transition to ferric enzyme during the reaction. The rate of heme-dioxy decay matched the rates of H4B radical formation and Arg hydroxylation in both enzymes. The rates of H4B radical formation differed such that nNOSoxy (18 s(-1)) > iNOSoxy (11 s(-1)) > eNOSoxy (6 s(-1)), whereas the lifetimes of the resulting H4B radical followed an opposite rank order. 5MeH4B supported a three-fold faster radical formation and greater radical stability relative to H4B in both eNOSoxy and nNOSoxy. Our results indicate the following: (i) the three NOSs share a common mechanism, whereby H4B transfers an electron to the heme-dioxy intermediate. This step enables Arg hydroxylation and is rate-limiting for all subsequent steps in the hydroxylation reaction. (ii) A direct correlation exists between pterin radical stability and the speed of its formation in the three NOSs. (iii) Uncoupled NO synthesis often seen for eNOS at low H4B concentrations may be caused by the slow formation and poor stability of its H4B radical.

Interaction of 1,2,3-benzodithiazolyls (Herz radicals) with dioxygen

The interaction of 4,6-di-tert-butyl-1,2,3-benzodithiazolyl (Herz radical) 1 with O2 in a hydrocarbon solution leads to substituted N,N’-disulfinyl-2,2’-diaminodiphenyl disulfide 2, the radical decay kinetics corresponds to a self-termination reaction with a rate constant, which is a linear function of the concentration of dissolved O2.