The production of hydroxyl radical from copper(I) complex systems of bleomycin and tallysomycin: comparison with copper(II) and iron(II) systems.

In contrast with BLM(or TALM)-Cu(II) complex system, Cu(I)-O₂ system of BLM(or TALM) as well as the corresponding Fe(II) system evidently produces reactive oxygen radicals as detected by ESR spin trapping. The sulfhydryl compound strongly prevented the generation of hydroxyl radical in BLM(or TALM)-Cu(I)-O₂ system. TALM forms metal complexes similar to BLM. The action mechanism of BLM and TALM has been proposed to be substantially same.     

The cationic plastoquinone radical of the chloroplast water splitting complex: Hyperfine splitting from a single methyl group determines the EPR spectral shape of Signal II

The proposal that EPR Signal II in spinach chloroplasts is due to a plastoquinone cation radical (O'Malley, P.J. and Babcock, G.T. (1983) Biophys. J. 41, 315a) has been investigated in further detail. The similarity in spectral shape between Signal II and the 2-methyl-5-isopropylhydroquinone cation radical is shown to arise from hyperfine coupling to one methyl group for both radicals. A well-resolved four line EPR spectrum of approximate relative intensity 1:3:3:1 for membrane orientation parallel and perpendicular to the applied magnetic field direction also indicates that the partially resolved structure of Signal II is due to hyperfine interaction with one methyl group, i.e., the 2-CH₃ group of the plastoquinone cation radical. The ENDOR band observed for this coupling is similar to that observed for methyl group bands of model quinone radicals. The principal hyperfine tensor values obtained for the methyl group interactions are A_⊥ = 27.2 MHz and $\text{A}{}_{\mathrm{}}\text{= 31.4}\text{MHz}$. The large isotropic coupling value (28.6 MHz) of the plastoquinone cation radical's 2-methyl group in vivo indicates that the antisymmetric orbital is the sole contributor to the spin-density distribution of Signal II. The orientation data also suggest that the plastoquinone cation radical is oriented such that the C-CH₃ bond direction, and hence the aromatic ring plane, lies perpendicular to the membrane plane.     

Identification of free radical species derived from caffeic acid and related polyphenols

Polyphenols are widely distributed in various fruits, vegetables and seasonings. It is well known that they have several physiological effects due to their antioxidative activities. Their activities depend on structural characteristics that favour the formation of their corresponding stable radicals. During the examination at which pH values, the polyphenol radicals are stabilized, we confirmed that polyphenol radicals were stabilized in NaHCO₃/Na₂CO₃ buffer (pH 10) rather than in physiological pH region. Then, we measured electron spin resonance (ESR) spectra at pH 10 to examine the characteristics of free radical species derived from caffeic acid (CA) with an unsaturated side chain, dihydrocaffeic acid (DCA) with a saturated side chain, chlorogenic acid (ChA) and rosmarinic acid (RA). In analyzing the radical structures, ESR simulation, determinations of macroscopic and microscopic acid dissociation constants and molecular orbital (MO) calculation were performed. In CA, the monophenolate forms were assumed to participate in the formation of free radical species, while in DCA, the diphenol form and the monophenolate forms were presumed to contribute to the formation of free radical species. On the basis of the results, we propose the possible structures of the free radical species formed from polyphenols under alkaline conditions.     

Crystal structures and magnetic properties of two complexes synthesized from manganese and halogenophenyl-substituted nitronyl nitroxide

One-dimensional complex (1), [Mn(hfac)₂(NITPhF)]₂ and one binuclear radical complex (2), [Mn(hfac)₂(NITPhBr)]_∞ have been synthesized. Here hfac stands for hexafluoroacetylacetonate, NITPhF for 2-(4′-fluorophenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide, and NITPhBr for 2-(4′-bromophenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide). All the two precursors were prepared and characterized by single-crystal X-ray diffraction analysis, IR, and magnetic analysis. In complex 1, the NITPhF radical acts as a bridge ligand linking two Mn(II) ions through the oxygen atom of the N-O group to form cyclic dimer. The dimers further connect two oxygen atoms of uncoordinated nitroxides of two adjacent radicals and yield one-dimensional chain sections. Instead in complex 2, the Mn(II) ions are bridged by the NITPhBr radicals through their N-O groups giving infinite one-dimensional chains. Magnetic susceptibility measurements indicate that both complex 1 and 2 behave ferrimagnetically. The Mn(II) ions interact antiferromagnetically with the direct bonding nitroxide group of the radicals although the structures of two complexes are different. The magnetic behaviors can be satisfactorily explained on the basis of the structural data.     

The enhancement of oxidative DNA damage by anti-diabetic metformin,buformin, and phenformin,via nitrogen-centered radicals

Metformin (N,N-dimethylbiguanide), buformin (1-butylbiguanide), and phenformin (1-phenethylbiguanide) are anti-diabetic biguanide drugs, expected to having anti-cancer effect. The mechanism of anti-cancer effect by these drugs is not completely understood. In this study, we demonstrated that these drugs dramatically enhanced oxidative DNA damage under oxidative condition. Metformin, buformin, and phenformin enhanced generation of 8-oxo-7,8-dihydro-2’-deoxyguanosine (8-oxodG) in isolated DNA reacted with hydrogen peroxide (H₂O₂) and Cu(II), although these drugs did not form 8-oxodG in the absence of H₂O₂ or Cu(II). An electron paramagnetic resonance (EPR) study, utilizing alpha-(4-pyridyl-1-oxide)-N-tert-butylnitrone and 3,3,5,5-tetramethyl-1-pyrroline-N-oxide as spin trapping agents, showed that nitrogen-centered radicals were generated from biguanides in the presence of Cu(II) and H₂O₂, and that these radicals were decreased by the addition of DNA. These results suggest that biguanides enhance Cu(II)/H₂O₂-mediated 8-oxodG generation via nitrogen-centered radical formation. The enhancing effect on oxidative DNA damage may play a role on anti-cancer activity.     

Enhancement of adriamycin toxicity by iron chelates is not a free radical mechanism

The possible involvement of metal ions and free radicals in the cytotoxic mechanism of Adriamycin (ADR) was investigated, using a model system ofEscherichia coli cells. It is shown thatE. coli mediated the production of free radicals under anaerobic (ADR-semiquinone) and aerobic (superoxide) conditions. ADR-induced loss of colony-forming ability was enhanced by the addition of iron (Fe) chelates. These observations suggested that a Fenton-type free radical mechanism was responsible for ADR toxicity. However, the mortality rate was essentially unchanged by the exclusion of oxygen. It was also unaffected by the addition of H₂O₂, catalase, or chelating agents. Cu(II), Zn(II) or Mg(II) had no effect on ADR toxicity. ADR and iron chelates did not induce measurable amounts of DNA strand-breaks. These observations suggest a mechanism of ADR-induced cell killing that is enhanced by Fe chelates, but does not directly involve oxygen-derived free radicals.     

Vibrational spectroscopy to study the properties of redox-active tyrosines in photosystem II and other proteins

Tyrosine radicals play catalytic roles in essential metalloenzymes. Their properties—midpoint potential, stability…—or environment varies considerably from one enzyme to the other. To understand the origin of these properties, the redox tyrosines are studied by a number of spectroscopic techniques, including Fourier transform infrared (FTIR) and resonance Raman (RR) spectroscopy. An increasing number of vibrational data are reported for the (modified-) redox active tyrosines in ribonucleotide reductases, photosystem II, heme catalase and peroxidases, galactose and glyoxal oxidases, and cytochrome oxidase. The spectral markers for the tyrosinyl radicals have been recorded on models of (substituted) phenoxyl radicals, free or coordinated to metals. We review these vibrational data and present the correlations existing between the vibrational modes of the radicals and their properties and interactions formed with their environment: we present that the ν_7a(C-O) mode of the radical, observed both by RR and FTIR spectroscopy at 1480-1515 cm⁻¹, is a sensitive marker of the hydrogen bonding status of (substituted)-phenoxyl and Tyr, while the ν_8a(C-C) mode may probe coordination of the Tyr to a metal. For photosystem II, the information obtained by light-induced FTIR difference spectroscopy for the two redox tyrosines Tyr_D and Tyr_Z and their hydrogen bonding partners is discussed in comparison with those obtained by other spectroscopic methods.     

Photochemical Properties of Camptothecin in the Presence of Copper(II) Ions: The Role of Radicals as Prospective Species in Photodynamic Therapy

Camptothecin (CPT) is an anticancer drug that inhibits topoisomerase I (Topo I) by forming a ternary DNA-CPT-Topo I complex. However, it has also been shown that UVA-irradiated CPT in the absence of Topo I produces significant DNA damage to cancer cells. In this work, we explored and identified free radicals generated in these processes. From the low-temperature EPR spectrum of Cu(II)-CPT complex, a proximity between Cu(II) ion and 20-hydroxy group of lactone E ring of CPT is proposed. Upon irradiation (λ = 365 nm) of the Cu(II)-CPT complex in de-oxygenated dimethylsulfoxide (DMSO), the EPR signal of Cu(II) measured in situ at room temperature shows formal first-order exponential decay with a formal half-life of 11 min. By the use of a specific Cu(I) chelating agent, neocuproine, it was shown that, during this process, Cu(II) is reduced to Cu(I). The loss in EPR signal intensity of the Cu(II)-CPT complex upon irradiation is accompanied by the appearance of a new EPR signal at g ≈ 2.0022. Application of the spin trap nitrosodurene (ND) revealed that the main radical product formed upon continuous irradiation of CPT in DMSO solutions is the hydroxyl radical (trapped in DMSO as the ^•CH₃ adduct) and superoxide radical. Application of 2,2,6,6-tetramethyl-4-piperidinol has revealed that irradiation of CPT in aerated DMSO solution also leads to formation of singlet oxygen (¹O₂). Our spectroscopic experiments indicate that CPT is a promising photosensitizer and that radicals and singlet oxygen generated upon illumination play a central role in DNA cleavage and in the induction of apoptosis in cancer cells.     

The kinetics of the reaction of nitrogen dioxide with iron(II)- and iron(III) cytochrome c

The reactions of NO₂ with both oxidized and reduced cytochrome c at pH 7.2 and 7.4, respectively, and with N-acetyltyrosine amide and N-acetyltryptophan amide at pH 7.3 were studied by pulse radiolysis at 23 °C. NO₂ oxidizes N-acetyltyrosine amide and N-acetyltryptophan amide with rate constants of (3.1±0.3)×10⁵ and (1.1±0.1)×10⁶ M⁻¹ s⁻¹, respectively. With iron(III)cytochrome c, the reaction involves only its amino acids, because no changes in the visible spectrum of cytochrome c are observed. The second-order rate constant is (5.8±0.7)×10⁶ M⁻¹ s⁻¹ at pH 7.2. NO₂ oxidizes iron(II)cytochrome c with a second-order rate constant of (6.6±0.5)×10⁷ M⁻¹ s⁻¹ at pH 7.4; formation of iron(III)cytochrome c is quantitative. Based on these rate constants, we propose that the reaction with iron(II)cytochrome c proceeds via a mechanism in which 90% of NO₂ oxidizes the iron center directly—most probably via reaction at the solvent-accessible heme edge—whereas 10% oxidizes the amino acid residues to the corresponding radicals, which, in turn, oxidize iron(II). Iron(II)cytochrome c is also oxidized by peroxynitrite in the presence of CO₂ to iron(III)cytochrome c, with a yield of ~60% relative to peroxynitrite. Our results indicate that, in vivo, NO₂ will attack preferentially the reduced form of cytochrome c; protein damage is expected to be marginal, the consequence of formation of amino acid radicals on iron(III)cytochrome c.     

Ascorbic acid induced degradation of beta-glucan: Hydroxyl radicals as intermediates studied by spin trapping and electron spin resonance spectroscopy

The formation of hydroxyl radicals in beta-glucan solutions treated with ascorbic acid and iron(II) was demonstrated by ESR spin trapping based methods. Two different spin traps were tested, namely DMPO which is commonly used to detect hydroxyl radicals, and POBN often used to detect carbon centered radicals. The experiments performed showed that the presence of iron(II) with DMPO led to low DMPO-OH adduct stability and further to DMPO dimerization. The level of hydroxyl radicals formed during the beta-glucan radical mediated degradation was evaluated using two ESR spin trapping methods based on the use POBN together with either 2% (v/v) EtOH or DMSO. The addition of ascorbic acid together with iron(II) in beta-glucan solution led to an immediate maximal production of hydroxyl radicals while the presence of ascorbic acid alone led to a progressive production of radical. Further hydroxyl radicals were found to be formed when iron(II) was added alone in beta-glucan solutions. The viscosity loss observed in the three last mentioned beta-glucan solutions were found to relate with the formation of hydroxyl radicals. These data confirm the involvement of hydroxyl radical in the beta-glucan degradation.     

Catecholamines Enhance Dihydrolipoamide Dehydrogenase Inactivation by the Copper Fenton System. Enzyme Protection by Copper Chelators

Catecholamines (CAs: epinephrine, norepinephrine, dopamine, L-DOPA, 6-hydroxydopamine) and o-diphenols (DOPAC and catechol) enhanced dihydrolipoamide dehydrogenase (LADH) inactivation by Cu(II) /H₂O₂ (Cu-Fenton system). The inhibition of LADH activity correlated with Cu(II), H₂O₂ and CA concentrations. Similar inhibitions were obtained wit! the assayed CAs and o-diphenols. CAs enhanced HO radical production by Cu(II) /H₂O₂, as demonstrated by benzoate hydroxylation and deoxyribose oxidation; LADH counteracted the pro-oxidant effect of CAs by scavenging hydroxyl radicals. Captopril, dihydrolipo amide, dihydrolipoic acid, DL-dithiothreitol, GSSG, try-panothione and histidine effectively preserved LADH from oxidative damage, whereas N-acetylcysteine, N-(2-mercaptopropionylglycine) and lipoamide were less effective protectors. Catalase (though neither bovine serum albumin nor superoxide dismutase) protected LADH against the Cu(II)/H₂O₂/CAs systems. Dena tured catalase protected less than the native enzyme, its action possibly depending on Cu-binding. LADH in creased and Captopril inhibited epinephrine oxidation by Cu(II)/H₂O₂ and Cu(II). The summarized evidence supports the following steps for LADH inactivation: (1) reduction of LADH linked-Cu(II) to Cu(I) by CAs; (2) production of HO^* from H₂O₂ by LADH-linked Cu(I) (Haber-Weiss reaction) and (3) oxidation of aminoacid residues at the: enzyme active site by site-specifically generated HO^* radicals. Hydrogen peroxide formation from CAs autoxidation may contribute to LADH inactivation.     

Pulse-radiolytic investigations of catechols and catecholamines II. Reactions of Tiron with oxygen radical species

Transient spectra and kinetic data of Tiron (1,2-dihydroxybenzene-3,5-disulphonic acid) are reported, obtained after pulse-radiolytic oxidation by hydroxyl radicals (°OH), superoxide anions (O₂^?) or a combination of both oxygen radicals. The rate constant with °OH radicals was determined at 1.0·10⁹ M^?1·s^?1. Contrary to a previous report (Greenstock, C.L. and Miller, R.W. (1975) Biochim. Biophys. Acta 396, 11–16), the rate constant with O₂^? of 1.0·10⁷ M^?1·s^?1 is lower by one order of magnitude; also the semiquinone absorbs at 300 nm rather than at 400 nm. The ratio of the rate constants with °OH and O₂^? of 100 again demonstrates that any oxidation reaction by the latter radical is unspecific due to the more efficient reaction of °OH radicals, leading to the same products with catechol compounds.     

Hydroxyl radical production from hydrogen peroxide and enzymatically generated paraquat radicals: Catalytic requirements and oxygen dependence

The ability of paraquat radicals (PQ^+.) generated by xanthine oxidase and glutathione reductase to give H₂O₂-dependent hydroxyl radical production was investigated. Under anaerobic conditions, paraquat radicals from each source caused chain oxidation of formate to CO₂, and oxidation of deoxyribose to thiobarbituric acid-reactive products that was inhibited by hydroxyl radical scavengers. This is in accordance with the following mechanism derived for radicals generated by γ-irradiation [H. C. Sutton and C. C. Winterbourn (1984) Arch. Biochem. Biophys.235, 106–115] PQ^+. + Fe³⁺ (chelate) → Fe²⁺ (chelate) + PQ⁺⁺ H₂O₂ + Fe²⁺ (chelate) → Fe³⁺ (chelate) + OH^? + OH.. Iron-(EDTA) and iron-(diethylenetriaminepentaacetic acid) (DTPA) were good catalysts of the reaction; iron complexed with desferrioxamine or transferrin was not. Extremely low concentrations of iron (0.03 μm) gave near-maximum yields of hydroxyl radicals. In the absence of added chelator, no formate oxidation occurred. Paraquat radicals generated from xanthine oxidase (but not by the other methods) caused H₂O₂-dependent deoxyribose oxidation. However, inhibition by scavengers was much less than expected for a reaction of hydroxyl radicals, and this deoxyribose oxidation with xanthine oxidase does not appear to be mediated by free hydroxyl radicals. With O₂ present, no hydroxyl radical production from H₂O₂ and paraquat radicals generated by radiation was detected. However, with paraquat radicals continuously generated by either enzyme, oxidation of both formate and deoxyribose was measured. Product yields decreased with increasing O₂ concentration and increased with increasing iron(DTPA). These results imply a major difference in reactivity between free and enzymatically generated paraquat radicals, and suggest that the latter could react as an enzyme-paraquat radical complex, for which the relative rate of reaction with Fe³⁺ (chelate) compared with O₂ is greater than is the case with free paraquat radicals.     

Oxygen-copper (II) interplay in the repair of semi-oxidized urate by quercetin bound to human serum albumin

The 1:1 complex of copper (II) and human serum albumin (HSA) slowly reacts with radiolytically generated ^•O₂^- radical-anion at a rate constant of 6.1×10₆ M_-1 s_-1. Absorbance and fluorescence spectroscopies demonstrate that addition of an equimolar portion of quercetin (QH₂) to the solution of the copper (II)-HSA complex induces a relocalization of the copper resulting in a ternary copper (II)-QH₂-HSA complex. This form of quercetin slowly oxidizes in air-saturated solutions. A 10-fold excess urate, a plasma antioxidant, cannot displace copper (II) bound to HSA. In N₂O-saturated solutions the ternary complex form of QH₂ can repair the urate radical with a rate constant of 2.7×10₆ M_-1 s_-1 by an electron transfer reaction similar to that observed in the absence of copper (II). In O₂-saturated solutions and in the absence of copper, HSA-bound QH₂ fails to repair the urate radical because of the fast competitive reaction of ^•O₂^- with urate radicals. However, addition of equimolar copper (II) restores the electron transfer from QH₂ to the urate radical. These contrasting results are tentatively explained either by an enhanced reactivity of copper (II) with ^•O₂^- in the ternary complex or by direct production of quercetin radicals via a copper-catalyzed reduction of the ^•O₂^- radicals by QH₂.     

Oxidation of uric acid in rheumatoid arthritis: is allantoin a marker of oxidative stress?

Free radicals are implicated in many diseases including atherosclerosis, cancer and also in rheumatoid arthritis. Reaction of uric acid with free radicals, such as hydroxyl radical and hypochlorous acid (HOCl) results in allantoin production. In this study, we measured the serum allantoin levels, oxidation products of uric acid, as a marker of free radical generation in rheumatoid arthritis. Fasting blood samples were obtained from 21 rheumatoid patients and 15 healthy controls. In this study, the serum allantoin and uric acid levels were measured by a gas chromatography-mass spectrometry method and the ratios were calculated. The mean allantoin and uric acid levels and ratios in the patient group were 22.1±11.3, 280.5±65.0 and 8.0±3.7 μM, while in the control group they were 13.6±6.3, 278.3±53.6 and 4.9±2.1 μM, respectively. The effects of gender, age, menopausal status, duration of disease and medications on serum allantoin and uric acid levels of the patient and control groups were studied. Our results suggest that uric acid acts as a free radical scavenger and thus is converted to allantoin. Increased allantoin levels suggest the possible involvement of free radicals in rheumatoid arthritis.     

Radical scavenging ability of some compounds isolated from Piper cubeba towards free radicals

The purpose of this study was to identify the antioxidant activity of 16 compounds isolated from Piper cubeba (CNCs) through the extent of their capacities to scavenge free radicals, hydroxyl radical (HO^?), superoxide anion radical () and 2,2‐diphenyl‐1‐picrylhydrazyl radical (DPPH^?), in different systems. Electron paramagnetic resonance (EPR) and 5,5‐dimethyl‐1‐pyrroline‐N‐oxide, DMPO, as the spin trap, and chemiluminescence techniques were applied. Using the Fenton‐like reaction [Fe(II) + H₂O₂], CNCs were found to inhibit DMPO? OH radical formation ranging from 5 to 57% at 1.25 mmol L^?1 concentration. The examined CNCs also showed a high DPPH antiradical activity (ranging from 15 to 99% at 5 mmol L^?1 concentration). Furthermore, the results indicated that seven of the 16 tested compounds may catalyse the conversion of superoxide radicals generated in the potassium superoxide/18‐crown‐6 ether system, thus showing superoxide dismutase‐like activity. The data obtained suggest that radical scavenging properties of CNCs might have potential application in many plant medicines. Copyright © 2010 John Wiley & Sons, Ltd.     

New Aspects in the Free-Radical Chemistry of Pyrimidine Nucleobases

The contribution will cover three aspects:

i) It has been known for some time that OH radicals and H atoms react with the pyrimidines by adding to the C(5)-C(6) double bond, but only the u.v.-spectra of the sum of these radicals have been reported so far. It will be shown how to arrive at the individual spectra of the C(5) and the C(6) adduct radicals.

ii) α-Hydroxyalkyl radicals are known to inactivate biologically active DNA. In contrast to the electrophilic radicals H and OH they are nucleophilic and the hydroxymethyl radicals add exclusively at the C(6) position of 1,3-dimethyluracil (k ～ 10⁴dm³ mol^?1 s^?1). In the corresponding thymine derivative this reaction also occurs, but one third of the hydroxymethyl radicals abstract an H-atom from the C(5)-methyl group thereby forming an allylic radical. In the course of these reactions pyrimidines with an exocyclic double bond are formed. These products react much more rapidly with hydroxymethyl radicals than the starting material leading to highly hydroxymethylated material at very low doses.

iii) The direct effect of ionizing radiation which would produce a pyrimidine base radical cation can be mimicked by reacting the pyrimidine with SO₄^?, a very good electron acceptor. In water, the radical cation of 1,3-dimethyluracil is rapidly (t_1/2 2μs) converted into the C(5) OH adduct radical. In the presence of peroxodisulphate a chain reaction sets in which leads to the cis-glycol.

The relevance of these findings to radiobiological aspects of nucleic acid research will be discussed.     

Effects of Free Radicals on the Fluidity of Myocardial Membranes

Free radicals, including superoxide anions (O₂^?_?), hydroxyl radical (HO'), and hypohalite radical (OCl'), as well as oxidants such as hydrogen peroxide (H₂O₂) and hypochlorous acid (HOCl), have been indicated in the pathogenesis of myocardial ischemic and reperfusion injury. In this report, we compared the integrity of the myocardial membrane when exposed to these free radicals/oxidants. Isolated rat heart membrane preparations were exposed to chemically generated free radicals with or without their respective scavengers. Membrane fluidity was monitored by fluorescence polarization using the diphenylhexatriene probe, as well as by electron spin resonance (ESR) spectroscopy using 2,2,6,6-tetramethyl piperidine-n-oxyl as the spin labeling agent. HO', H₂O₂, and OCl' + HOCl increased the fluorescence polarization (FP) and microvis-cosity significantly by 1.7-fold, 1.8-fold, and 1.7-fold, respectively, as compared to an only 1.2– fold increase in FP by O₂^?_? O₂^?_? did not alter the fatty acid profiles of the membrane phospholipids. However, HO' and H₂O₂ reduced the arachidonic acid contents in phosphatidylcholine (PC), phosphatidylethanolamine (PE), and phosphatidylinositol (PI). These radicals also stimulated the lipid peroxidation by several-fold, while that by O₂^?_? was only insignificant. These results suggest that HO' and H₂O₂ decreased the membrane fluidity and induced lipid peroxidation by releasing the arachidonic acid from PC, PE. and PI.     

Radical production by hydrogen peroxide/bicarbonate and copper uptake in mammalian cells: modulation by Cu(II) complexes

The presence of the bicarbonate/carbon dioxide pair is known to accelerate the transition metal ion-catalysed oxidation of various biotargets. It has been shown that stable Cu(II) complexes formed with imine ligands that allow redox cycling between Cu(I) and Cu(II) display diverse apoptotic effects on cell cultures. It is also reported that Cu(II)-tetraglycine can form a stable Cu(III) complex. In the present study, radical generation from H₂O₂ and H₂O₂/HCO₃⁻ in the presence of these two different classes of Cu(II) complexes was evaluated by monitoring the oxidation of dihydrorhodamine 123 and NADH and by the quantitative determination of thiobarbituric acid reactive substances (TBARs method). Cu(II)-imine complexes produced low levels of reactive species whereas Cu(II)-Gly-derived complexes, as well as the free Cu(II) ion, produced oxygen-derived radicals in significantly larger amounts. The effects of these two classes of complexes on mammalian tumour cell viability were equally distinct, in that Cu(II)-imine complexes caused apoptosis, entered in cell and remained almost unaffected in high levels whilst, at the same concentrations, Cu(II)-Gly peptide complexes and Cu(II) sulphate stimulated cell proliferation, with the cell managing copper efficiently. Taken together, these results highlight the different biological effects of Cu(II) complexes, some of which have been recently studied as anti-tumour drugs and radical system generators, and also update the effects of reactive oxygen species generation on cell cycle control.     

Enhancement of Lipid Peroxidation by Indole-3-Acetic Acid and Derivatives: Substituent Effects

The peroxidation of liposomes by a haem peroxidase and hydrogen peroxide in the presence of indole-3-acetic acid and derivatives was investigated. It was found that these compounds can accelerate the lipid peroxidation up to 65 fold and this is attributed to the formation of peroxyl radicals that may react with the lipids, possibly by hydrogen abstraction. The peroxyl radicals are formed by peroxidase-catalyzed oxidation of the enhancers to radical cations which undergo cleavage of the carbon-carbon bond on the side-chain to yield CO2 and carbon-centred radicals that rapidly add oxygen. In competition with decarboxylation, the radical cations deprotonate reversibly from the Nl position. Rates of decarboxylation,pK_a values and rate of reaction with the peroxidase compound I indicate consistent substituent effects which, however, can not be quantitatively related to the usual Hammett or Brown parameters. Assuming that the rate of decarboxylation of the radical cations taken is a measure of the electron density of the molecule (or radical), it is found that the efficiency of these compounds as enhancers of lipid peroxidation increases with increasing electron density, suggesting that, at least in the model system, the oxidation of the substrates is the limiting step in causing lipid peroxidation.