Reactive Radical Intermediates Research Articles

Guest editors’ tribute to Fritz Pragst on the occasion of his 70th birthday

This issue is dedicated to Fritz Pragst on the occasion of his 70th birthday on September 9th, 2011. Fritz Pragst is an outstanding scientist who has succeeded to make two scientific carriers in rather different fields of chemistry, first in organic electrochemistry and then in forensic analysis. From 1961 to 1966, Pragst studied chemistry at Humboldt University, Berlin. He finished with a diploma work on the electrochemical reduction of porphyrines and porphyrine metal complexes. His Ph.D. thesis, defended in 1969, concerned the electrochemical oxidation of diazo compounds. In 1976, he has defended his doctor of science thesis which was entitled “Correlations between structure, electrochemical properties and electrochemical luminescence of organic compounds”. The electrochemically generated luminescence (ECL) is the topic to which he has substantially contributed to a number of original papers published in the 1970s and 1980s and for which he is best known in the community of electrochemists. In the last two decades, several reviews as well as monographs dealing with ECL were published; however, two important facets of ECL deserve a much higher attention than had been given so far. To both these facets, Fritz Pragst has considerably contributed: First, it is the use of electron transfer reactions (indirect oxidation or reduction) and/or energy transfer reactions (both in homogeneous phase) to generate selectively and exclusively triplet states as ECL intermediates. The interpretation of emission can be used not only for correction or confirmation of the theoretical energetic considerations but also for estimation of the triplet energies of the studied compounds in an alternative way and, based on known energetic criteria, for the determination of—otherwise unavailable—redox potentials of short-living radical species. Second, it is the mechanistic significance of ECL studies, based on the ECL principle, that the luminescence is a consequence of a strongly exoenergetic electron transfer reaction fulfilling fundamental energetic criteria; the observed emission serves also as an unambiguous proof of formation and existence of highly reactive radical intermediates. They often represent the missing link in investigated reaction mechanisms. This type of ECL is generated at one single potential at one single electrode (originally called “direct-current”) utilizing the follow-up splitting of the primary radical ion. It is a very sensitive alternative method for the interception of even highly unstable intermediates and for the proof of the twoelectron ECE (electrochemical–chemical–electrochemical step) reaction mechanism in electrochemistry. Today, this type of ECL is called ECL with coreactants. J. Ludvik (*) Adademy of Science of the Czech Rep., J. Heyrovský Institute of Physical Chemistry, Dolejskova 3, 182 23 Prague 8, Czech Republic e-mail: jiri.ludvik@jh-inst.cas.cz

Synthesis of Allylic Hydroperoxides and EPR Spin-Trapping Studies on the Formation of Radicals in Iron Systems as Potential Initiators of the Sensitizing Pathway

Many terpenes used as fragrance compounds autoxidize when exposed to air, forming allylic hydroperoxides that have the potential to be skin contact allergens. To trigger the immunotoxicity process that characterizes contact allergy, these hydroperoxides are supposed to bind covalently to proteins in the skin via radical pathways. We investigated the formation of reactive radical intermediates from 7-hydroperoxy-3,7-dimethylocta-1,5-dien-3-ol and 2-hydroperoxylimonene, responsible for the sensitizing potential acquired by autoxidized linalool and limonene. Both compounds were synthesized through new short and reproducible synthetic pathways. The hydroperoxide decomposition catalyzed by Fe(II)/Fe(III) redox systems, playing a key role in degradating peroxides in vivo, was examined by spin-trapping-EPR spectroscopy. Alkoxyl and carbon-centered free radicals derived from the hydroperoxides were successfully trapped by the spin-trap 5,5-dimethyl-1-pyrroline N-oxide, whereas peroxyl radicals were characterized by spin-trapping studies with 5-diethoxyphosphoryl-5-methyl-1-pyrroline N-oxide. Using liquid chromatography combined with mass spectrometry, we demonstrated the formation of adducts, via radical mechanisms induced by Fe(II)/Fe(III), between the hydroperoxides and N-acetylhistidine methyl ester, a model amino acid that is prone to radical reactions. Free radicals derived from these hydroperoxides can thus induce amino acid chemical modifications via radical mechanisms. The study of these mechanisms will help to understand the sensitizing potential of hydroperoxides.

Laccase: Microbial Sources, Production, Purification, and Potential Biotechnological Applications

Laccase belongs to the blue multicopper oxidases and participates in cross-linking of monomers, degradation of polymers, and ring cleavage of aromatic compounds. It is widely distributed in higher plants and fungi. It is present in Ascomycetes, Deuteromycetes and Basidiomycetes and abundant in lignin-degrading white-rot fungi. It is also used in the synthesis of organic substance, where typical substrates are amines and phenols, the reaction products are dimers and oligomers derived from the coupling of reactive radical intermediates. In the recent years, these enzymes have gained application in the field of textile, pulp and paper, and food industry. Recently, it is also used in the design of biosensors, biofuel cells, as a medical diagnostics tool and bioremediation agent to clean up herbicides, pesticides and certain explosives in soil. Laccases have received attention of researchers in the last few decades due to their ability to oxidize both phenolic and nonphenolic lignin-related compounds as well as highly recalcitrant environmental pollutants. It has been identified as the principal enzyme associated with cuticular hardening in insects. Two main forms have been found: laccase-1 and laccase-2. This paper reviews the occurrence, mode of action, general properties, production, applications, and immobilization of laccases within different industrial fields.

Selective photocatalytic aerobic bromination with hydrogen bromide via an electron-transfer state of 9-mesityl-10-methylacridinium ion

Photocatalytic bromination of aromatic hydrocarbons by molecular oxygen with hydrogen bromide occurs efficiently to produce monobrominated products selectively using 9-mesityl-10-methylacridinium ion (Acr+–Mes) as a photocatalyst under visible light irradiation. Both the product yield and selectivity for the bromination of 1,3,5-trimethoxybenzene were 100% with a quantum yield of 4.8%. The photocatalytic turnover number is 900 based on the initial concentration of Acr+–Mes. The reactive radical intermediates involved in the photocatalytic cycle have been successfully detected by laser flash photolysis measurements. The photocatalytic bromination is initiated by photoinduced electron transfer from the mesitylene moiety to the singlet excited state of acridinium ion, which results in formation of the electron-transfer state of Acr+–Mes (Acr˙–Mes˙+), followed by electron transfer from aromatic hydrocarbons to the mesitylene radical cation moiety and electron transfer from the acridinyl radical moiety to O2. The resulting radical cations of aromatic hydrocarbons react with Br− to produce the corresponding monobrominated products selectively.

Radical Chemistry in the Thermal Decomposition of Anisole and Deuterated Anisoles: An Investigation of Aromatic Growth

The pyrolyses of anisole (C(6)H(5)OCH(3)), d(3)-anisole (C(6)H(5)OCD(3)), and d(8)-anisole (C(6)D(5)OCD(3)) have been studied using a hyperthermal tubular reactor and photoionization reflectron time-of-flight mass spectrometer. Gas exiting the reactor is subject to an immediate supersonic expansion after a residence time of approximately 65 mus. This allows the detection of highly reactive radical intermediates. Our results confirm that the first steps in the thermal decomposition of anisole are the loss of a methyl group to form phenoxy radical, followed by ejection of a CO to form cyclopentadienyl radical (c-C(5)H(5)); C(6)H(5)OCH(3) --> C(6)H(5)O + CH(3); C(6)H(5)O --> c-C(5)H(5) + CO. At high temperatures (T(wall) = 1200 degrees C - 1300 degrees C) the c-C(5)H(5) decomposes to propargyl radical (CH(2)CCH) and acetylene; c-C(5)H(5) --> CH(2)CCH + C(2)H(2). The formation of benzene and naphthalene is demonstrated with 1 + 1 resonance-enhanced multiphoton ionization. Propargyl radical recombination is a significant benzene formation channel. However, we show the majority of benzene is formed by a ring expansion reaction of methylcyclopentadiene (C(5)H(5)CH(3)) resulting from methyl radical addition to cyclopentadienyl radical; CH(3) + c-C(5)H(5) --> C(5)H(5)CH(3) --> C(6)H(6) + 2H. The naphthalene is generated from cyclopentadienyl radical recombination; 2c-C(5)H(5) --> C(5)H(5)-C(5)H(5) --> C(10)H(8) + 2H. The respective intermediate amu 79 and 129 species associated with these reactions are detected, confirming the stepwise nature of the decompositions. These reactions are verified by pyrolysis studies of cyclopentadiene (C(5)H(6)) and C(5)H(5)CH(3) obtained from rapid thermal dissociation of the respective dimer compounds, as well as pyrolysis studies of propargyl bromide (BrCH(2)CCH).

Carbon nanotubes degraded by neutrophil myeloperoxidase induce less pulmonary inflammation

We have shown previously that single-walled carbon nanotubes can be catalytically biodegraded over several weeks by the plant-derived enzyme, horseradish peroxidase. However, whether peroxidase intermediates generated inside human cells or biofluids are involved in the biodegradation of carbon nanotubes has not been explored. Here, we show that hypochlorite and reactive radical intermediates of the human neutrophil enzyme myeloperoxidase catalyse the biodegradation of single-walled carbon nanotubes in vitro, in neutrophils and to a lesser degree in macrophages. Molecular modelling suggests that interactions of basic amino acids of the enzyme with the carboxyls on the carbon nanotubes position the nanotubes near the catalytic site. Importantly, the biodegraded nanotubes do not generate an inflammatory response when aspirated into the lungs of mice. Our findings suggest that the extent to which carbon nanotubes are biodegraded may be a major determinant of the scale and severity of the associated inflammatory responses in exposed individuals.

Adenosyl radical: reagent and catalyst in enzyme reactions.

Adenosine is undoubtedly an ancient biological molecule that is a component of many enzyme cofactors: ATP, FADH, NAD(P)H, and coenzyme A, to name but a few, and, of course, of RNA. Here we present an overview of the role of adenosine in its most reactive form: as an organic radical formed either by homolytic cleavage of adenosylcobalamin (coenzyme B(12), AdoCbl) or by single-electron reduction of S-adenosylmethionine (AdoMet) complexed to an iron-sulfur cluster. Although many of the enzymes we discuss are newly discovered, adenosine's role as a radical cofactor most likely arose very early in evolution, before the advent of photosynthesis and the production of molecular oxygen, which rapidly inactivates many radical enzymes. AdoCbl-dependent enzymes appear to be confined to a rather narrow repertoire of rearrangement reactions involving 1,2-hydrogen atom migrations; nevertheless, mechanistic insights gained from studying these enzymes have proved extremely valuable in understanding how enzymes generate and control highly reactive free radical intermediates. In contrast, there has been a recent explosion in the number of radical-AdoMet enzymes discovered that catalyze a remarkably wide range of chemically challenging reactions; here there is much still to learn about their mechanisms. Although all the radical-AdoMet enzymes so far characterized come from anaerobically growing microbes and are very oxygen sensitive, there is tantalizing evidence that some of these enzymes might be active in aerobic organisms including humans.

ChemInform Abstract: The Chemistry of Reactive Radical Intermediates in Combustion and the Atmosphere

AbstractChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

Decolorization of C.I. Reactive Black 5 in UV/TiO 2, UV/oxidant and UV/TiO 2/oxidant systems: A comparative study

This study evaluated the effectiveness of decolorizing C.I. Reactive Black 5 (RB5) in UV/TiO 2, UV/oxidant and UV/TiO 2/oxidant systems in the presence of oxidants H 2O 2, Na 2S 2O 8, NaBrO 3, and NaIO 4. The effects of pH, oxidant dosage and ethanol addition on decolorization were determined. Decolorization rate constants ( k) were of pseudo-first-order in all systems. In UV/TiO 2 and UV/oxidant systems, the k values followed the order pH 4 > pH 7 > pH 10. At pH 7 with 1 mM of oxidant added, the k values followed the order UV/NaIO 4 (0.377 min −1) > UV/Na 2S 2O 8 (0.156 min −1) > UV/H 2O 2 (0.105 min −1) > UV/NaBrO 3 (0.101 min −1) > UV (0.002 min −1). In UV/oxidant systems, decolorization effectiveness increased with increasing Na 2S 2O 8 and NaBrO 3 doses but reached optima with NaIO 4 and H 2O 2 at 1 and 6 mM, respectively. Addition of oxidants to UV/TiO 2 resulted in quenching of conduction band electrons and generation of reactive radical intermediates, enhancing decolorization. In UV/TiO 2/oxidant systems, decolorization rate constants varied with oxidant following the order NaIO 4 (0.558 min −1) > Na 2S 2O 8 (0.044 min −1) > NaBrO 3 (0.036 min −1) > H 2O 2 (0.035 min −1) > absence of oxidant (0.033 min −1). In UV/oxidant systems, addition of ethanol inhibited decolorization, which indicated the involvement of hydroxyl radicals in the decolorization pathway with UV/H 2O 2 and UV/Na 2S 2O 8 systems, as well as probable involvement of other radicals with UV/NaBrO 3 and UV/NaIO 4 systems. Direct oxidation by photogenerated holes was likely significant in UV/TiO 2 and UV/TiO 2/oxidant systems.

Tin-free radical cyclization reactions initiated by visible light photoredox catalysis

Herein, we report an advancement in the application of visible light photoredox catalysts in a classic free radical mediated reaction, cyclization onto unactivated pi-systems. The reactive radical intermediate is generated by the single electron reduction of an activated C-Br bond by an electron-rich redox catalyst afforded by a visible light induced catalytic cycle.

Kinetics of oxidation of bilirubin and its protein complex by hydrogen peroxide in aqueous solutions

A comparative study of oxidation reactions of bilirubin and its complex with albumin was carried out in aqueous solutions under the action of hydrogen peroxide and molecular oxygen at different pH values. Free radical oxidation of the pigment in both free and bound forms at pH 7.4 was shown not to lead to the formation of biliverdin, but to be associated with the decomposition of the tetrapyrrole chromophore into monopyrrolic products. The effective and true rate constants of the reactions under study were determined. It was assumed that one possible mechanism of the oxidation reaction is associated with the interaction of peroxyl radicals and protons of the NH groups of bilirubin molecules at the limiting stage with the formation of a highly reactive radical intermediate. The binding of bilirubin with albumin was found to result in a considerable reduction in the rate of the oxidation reaction associated with the kinetic manifestation of the protein protection effect. It was found that the autoxidation of bilirubin by molecular oxygen with the formation of biliverdin at the intermediate stage can be observed with an increase in the pH of solutions.

Fate of monoterpenes in near-critical water and supercritical alcohols assisted by microwave irradiation

The rearrangement of alpha- and beta-pinene was studied under microwave irradiation in near-critical water and supercritical lower aliphatic alcohols, with the aim of identifying the pathway of alpha- and beta-pinene isomerization. Generally, two pathways occur, pyrolysis on the one and acidolysis on the other hand, whereby acidolysis is predominant in the case of near-critical water and the second pathway is favored for experiments employing supercritical alcohols. The different behavior of these two structurally related solvents is attributed to the increased availability of protons if water is heated to 270 degrees C and 80 bar, thus enhancing the autoprotolysis of water. The application of alcohols instead furnished rearrangement products clearly attributed to thermal pyrolysis route by the formation of radical reaction intermediates.

Electron Transfer Photoredox Catalysis: Intramolecular Radical Addition to Indoles and Pyrroles

The utilization of the photoredox catalyst, tris(2,2'-bipyridyl)ruthenium dichloride, and a household light bulb to effect radical cyclizations onto indoles and pyrroles at room temperature is reported. A reactive free radical intermediate is generated via the reduction of an activated C-Br bond by the single electron reductant, Ru(I), generated in a visible light induced photocatalytic cycle. This system represents an expansion of the application of photoredox catalysis in conventional free radical processes.

Specific local induction of DNA strand breaks by infrared multi-photon absorption

Highly confined DNA damage by femtosecond laser irradiation currently arises as a powerful tool to understand DNA repair in live cells as a function of space and time. However, the specificity with respect to damage type is limited. Here, we present an irradiation procedure based on a widely tunable Er/Yb : fiber femtosecond laser source that favors the formation of DNA strand breaks over that of UV photoproducts by more than one order of magnitude. We explain this selectivity with the different power dependence of the reactions generating strand breaks, mainly involving reactive radical intermediates, and the direct photochemical process leading to UV-photoproducts. Thus, localized multi-photon excitation with a wavelength longer than 1 µm allows for the selective production of DNA strand breaks at sub-micrometer spatial resolution in the absence of photosensitizers.

Photolysis of Dibenzyl Ketones Sorbed on MFI Zeolites in the Presence of Spectator Molecules: Cage Effects, Kinetics, and External Surface Sites Characterization

Over the past two decades, the photolytic reactions of dibenzyl ketones sorbed on zeolites have been investigated. The reported results are consistent with a supramolecular model that takes into account the physical and chemical nature of the structure of the zeolites and their effect on the reactive radical intermediates produced by photolysis of adsorbed molecules. The model incorporates various phenomena such as surface coverage, external and internal sorption, surface diffusion, radical sieving, and the resulting product distributions. This account reports direct evidence for the validation of the model through FT-IR spectroscopy and through a new method for "titrating" the binding sites via EPR spectroscopy. It is shown that it is possible to adjust and modulate the photolytic product distribution by varying the parameters of the system. The effects of co-adsorbed spectator molecules with different polarities, namely water, pyridine, and benzene, on the photolysis of o-methyldibenzyl ketone and dibenzyl ketone sorbed on MFI zeolites is examined. This study provides insights into a displacement mechanism caused by spectator molecules and further demonstrates how the product distribution of photolysis of sorbed ketones can be controlled. The kinetics of persistent radicals formed by photolysis of ketones sorbed on zeolites is directly monitored over time by EPR, providing a measure of the lifetime of these reactive organic intermediates. Finally, measurement of Langmuir isotherms was employed to provide classical evidence for the model.

Determination of reactive oxygen species generated in laccase catalyzed oxidation of wood fibers from Chinese fir ( Cunninghamia lanceolata) by electron spin resonance spectrometry

The aim of the present study was to determine whether the radical reaction intermediates – reactive oxygen species (ROS) were formed during the laccase-catalyzed oxidation of wood fibers from Chinese fir ( Cunninghamia lanceolata) and to quantify tentatively its production with electron spin resonance (ESR) spectrometry. To investigate the activation pathways triggered by laccase, ESR spin-trapping techniques using N- tert-butyl-α-phenylnitrone (PBN) as spin trap followed by ethyl acetate extraction were employed to identify and quantify the free radical intermediates. ROS such as the superoxide and hydroxyl radical was detected and quantified in the laccase catalyzed oxidation of wood fibers, suggesting that ROS is the main free radical intermediates for laccase reaction. Based on the findings of the presence of ROS and previous literature on the free radical reaction of laccase oxidation of wood fibers, a possible reaction mechanism involving ROS-mediated attack on the domains of lignin which is not directly accessible for the enzyme and solubilized low-molecular mass lignins which function as reactive compounds like adhesives and may cling back to the fiber surface, could accordingly describe laccase-catalyzed oxidation of Chinese fir wood fibers.

Degradation of methylene blue in aqueous dispersion of hollow titania photocatalyst: Study of reaction enhancement by various electron scavengers

Submicrometer-sized titania hollow spheres have been synthesized by employing sulfonated polystyrene latex particles (averaged diameter: 200 nm) as a template in conjunction with the sol–gel method. Utilization of 0.20 mol/L titanium tetrabutoxide and 25 g/L latex led to the formation of anatase particles having shell thickness of about 15 nm and void diameter of about 150 nm. Photocatalytic activity of the titania hollow spheres was examined by focusing on its enhancement by electron scavengers in the photocatalytic decomposition of methylene blue (MB). The electron scavengers employed were inorganic oxidants such as ClO − 3, BrO − 3, IO − 4, H 2O 2, and S 2O 2− 8. Differences in electronegativity, atomic radius of the halogens, and the number of highly reactive radical and nonradical intermediates were proven to be important criteria for an electron scavenger to yield high efficiency in the MB photodecomposition. Based on the results, the effect of the oxidants used were found to be in the order of S 2 O 8 2 − > IO 4 − > BrO 3 − > H 2 O 2 > ClO 3 − .

One-electron oxidation of catecholamines generates free radicals with an in vitro toxicity correlating with their lifetime

One-electron oxidation of dopamine by ferricyanide generates a highly reactive free radical intermediate that inactivates the V-type H +-ATPase proton pump in catecholamine storage vesicles, i.e., the driving force in both the vesicular uptake and the storage of catecholamines, in a cell-free in vitro model system at pH 7.0. Electron paramagnetic resonance spectroscopy revealed that a radical with g = 2.0045, formed by this oxidation, was relatively long-lived ( t 1/2 obs = 79 s at pH 6.5 and 25°C). Experimental evidence is presented that the observed radical most likely represents dopamine semiquinone free radical, although an o-quinone free radical cannot be ruled out. Oxidation of noradrenaline and adrenaline by ferricyanide generated similar isotropic radicals, but of shorter half-lives (i.e., 43 and 5.3 s, respectively), and the efficacy of inactivation of the H +-ATPase correlated with the half-life of the respective catecholamine free radical (i.e., dopamine > noradrenaline ≫ adrenaline). Thus, the generation of relatively long-lived semiquinone free radicals, although at low concentrations, in dopaminergic and noradrenergic neurons may represent a common mechanism of cytotoxicity linked to neurodegeneration of the respective neurons related to Parkinson disease.

Assembly and Protection of the Radical Enzyme, Methylmalonyl-CoA Mutase, by Its Chaperone

MeaB is a recently described P-loop GTPase that plays an auxiliary role in the reaction catalyzed by the radical B12 enzyme, methylmalonyl-CoA mutase. Defects in the human homologue of MeaB result in methylmalonic aciduria, but the role of this protein in coenzyme B12 assimilation and/or utilization is not known. Methylmalonyl-CoA mutase catalyzes the isomerization of methylmalonyl-CoA to succinyl-CoA that uses reactive radical intermediates that are susceptible to oxidative inactivation. In this study, we have examined the influence of MeaB on the kinetics of the reaction catalyzed by methylmalonyl-CoA mutase and on the thermodynamics of cofactor binding. MeaB alone has a modest effect on the affinity of the mutase for the 5'-deoxyadenosylcobalamin (AdoCbl) cofactor, increasing it 2-fold from 404 +/- 71 to 210 +/- 22 nM. However, in the presence of GDP, the affinity for the cofactor decreases 5-fold to 1.89 +/- 0.33 microM, while in the presence of guanosine 5'(beta-gamma imino)triphosphate, a nonhydrolyzable analogue of GTP, the binding of AdoCbl to the mutase is not detected. Protection against oxidative inactivation of the mutase by MeaB is dependent upon the presence of nucleotides with the MeaB/GDP and MeaB/GTP complexes decelerating the rate of formation of oxidized cofactor by 3- and 15-fold, respectively. This study suggests that MeaB functions in the GTP-dependent assembly of holomethylmalonyl-CoA mutase and subsequent protection of radical intermediates during catalysis.

Peroxynitrite-induced oxidation and nitration products of guanine and 8-oxoguanine: Structures and mechanisms of product formation

Peroxynitrite induces DNA base damage predominantly at guanine (G) and 8-oxoguanine (8-oxoG) nucleobases via oxidation reactions. Nitration products are also observed, consistent with the generation of radical intermediates that can recombine with the (.)NO(2) formed during peroxynitrite degradation. The neutral G radical, G(.), reacts with (.)NO(2) to yield 8-nitroguanine (8-nitroG) and 5-nitro-4-guanidinohydantoin (NI), while for 8-oxoG we have proposed a reactive guanidinylidene radical intermediate. The products generated during peroxynitrite-mediated 8-oxoG oxidation depend on oxidant flux, with dehydroguanidinohydantoin (DGh), 2,4,6-trioxo-[1,3,5]triazinane-1-carboxamidine (CAC) and NO(2)-DGh predominating at high fluxes and spiroiminodihydantoin (Sp), guanidinohydantoin (Gh) and 4-hydroxy-2,5-dioxo-imidazolidine-4-carboxylic acid (HICA) predominating at low fluxes. Both product sets are observed at intermediate fluxes. It is therefore important in model systems to ensure that the relative concentrations are well controlled to minimize competing reactions that may not be relevant in vivo. Increasingly sophisticated systems for modeling peroxynitrite production in vivo are being developed and these should help with predicting the products most likely to be formed in vivo. Together with the emerging information on the genotoxic and mutational characteristics of the individual oxidation products, it may be found that the extent of tissue damage, mutational spectra and, hence, cancer risk may change as a function of peroxynitrite fluxes as different product combinations predominate.