Bioreductive Activation of Quinones: Redox Properties and Thiol Reactivity

Redox properties and thiol reactivity are central to the therapeutic and toxicological properties of qui-nones. The use of other physicochemical parameters to establish predictive relationships for redox properties of quinones is discussed. and attention drawn to situations where such relationships may be unreliable. The rates of reaction of semiquinone radicals with oxygen, including those of chemotherapeutic agents such as mitomycin and the anthracyclines. can be predicted with reasonable confidence from the redox properties. The reactions of quinones with thiols such as glutathione produces reduced quinones and radicals. but the reactions are complex and all the features are not well understood     

Reactivity of ubiquinones and ubiquinols with free radicals

The reactivity of quinones 1–4 and of the corresponding quinols 5–8 towards carbon- and oxygen-centred radicals were studied. All quinones bearing at least one nuclear position free, readily react with alkyl and phenyl radicals to afford the alkylated quinones 12–24; however, quinones 1 and 3 reacted with 2-cyano-2-propyl radical to yield products (the mono- and di-ethers 9–11) derived from the attack on the carbonylic oxygen. The reactions carried out on quinones with the benzoyloxy radical led to no reaction products and in the case of Q₁₀, the isoprenic chain also remained unchanged. Quinols 5–8 reacted only with oxygencentred radicals (benzoyloxy and 2-cyano-2-propylperoxy radicals) to give the corresponding quinones. The isoprenic chain of Q₁₀ did not undergo attack even with peroxy radicals. Carbon-centred radicals resulted unable to abstract hydrogen from the studied quinols.     

Free radical generation by redox cycling of estrogens

Natural and synthetic estrogens elicit normal hormonal responses in concentrations in a clearly defined yet low range. At elevated doses, metabolic reactions of the phenolic moiety, while harmless at low levels, may become the predominant biochemical activity and may exert deleterious effects. These metabolic pathways, such as i) oxidation of estrogens to catechol estrogens and further to their respective quinones, and ii) free radical generation by redox cycling between catechol estrogens or diethylstilbestrol and their quinones, are investigated for their influence in physiological or pathophysiological processes. In this review, the in vitro capacity of various enzymes to oxidize estrogen hydroquinones to quinones or to reduce corresponding quinones to hydroquinones is evaluated. The in vivo activities of enzymes supporting redox cycling of estrogens and free radical generation is correlated with induction of kidney tumors in Syrian hamsters. Concomitant changes in activities in quinone reductase and other detoxifying enzymes in kidneys of hamsters treated with estrogen support a role of free radicals in the induction of tumors by estrogen. Free radical damage to protein and possibly to DNA in kidneys of estrogen-treated hamsters may be used as markers of free radical action in vivo.     

Redox and addition chemistry of quinoid compounds and its biological implications

The overall biological activity of quinones is a function of the physico-chemical properties of these compounds, which manifest themselves in a critical bimolecular reaction with bioconstituents. Attempts have been made to characterize this bimolecular reaction as a function of the redox properties of quinones in relation to hydrophobic or hydrophilic environments. The inborn physico-chemical properties of quinones are discussed on the basis of their reduction potential and dissociation constants, as well as the effect of environmental factors on these properties. Emphasis is given on the effect of methyl-, methoxy-, hydroxy-, and glutathionyl substituents on the reduction potential of quinones and the subsequent electron transfer processes. The redox chemistry of quinoid compounds is surveyed in terms of a) reactions involving only electron transfer, as those accomplished during the enzymic reduction of quinones and the non-enzymic interaction with redox couples generating semiquinones, and b) nucleophilic addition reactions. The addition of nucleophiles, entailing either oxidation or reduction of the quinone, are exemplified in reactions with oxygen- or sulfur nucleophiles, respectively. The former yields quinone epoxides, whereas the latter yields thioether-hydroquinone adducts as primary molecular products. The subsequent chemistry of these products is examined in terms of enzymic reduction, autoxidation, cross-oxidation, disproportionation, and free radical interactions. The detailed chemical mechanisms by which quinoid compounds exert cytotoxic, mutagenic and carcinogenic effects are considered individually in relation to redox cycling, alterations of thiol balance and Ca++ homeostasis, and covalent binding.     

Radical-radical reactions of superoxide: a potential route to toxicity

Superoxide reacts with many radicals, such as phenoxyl radicals, at near diffusion-controlled rates. These reactions are usually considered to be repair processes and have received little biological attention. However, addition of superoxide to give hydroperoxides and secondary oxidation products can also occur. The relative contributions of addition and repair vary depending on the properties of the phenol. With tyrosine, addition to give tyrosine hydroperoxide predominates, but in peptides the efficiency of hydroperoxide formation depends on the proximity of free amine groups. Radicals from other phenolic compounds, such as alpha-tocopherol and serotonin, also undergo addition reactions with superoxide. Physiologically, these reactions are likely to be more significant than dimerization when both radicals are generated together. They warrant attention as potential contributors to superoxide toxicity.     

Spin Trapping of Inorganic Radicals

The use of nitrose compounds and nitrones as spin traps for the detection of short-lived inorganic radicals is discussed. To a certain degree nitrones and nitroso compounds are complementary. While nitroso compounds are superior with respect to spin trapping metal-centred radicals, nitrones form more persistent spin adducts with most small inorganic radicals.

Erroneous results may be obtained when hydrolysis and redox reactions involving the spin adducts are ignored. Spin trapping of pseudohalide radicals (·N_j· ·CN, ·SCN) are discussed in more detail.     

Spin Trapping of Inorganic Radicals

The use of nitrose compounds and nitrones as spin traps for the detection of short-lived inorganic radicals is discussed. To a certain degree nitrones and nitroso compounds are complementary. While nitroso compounds are superior with respect to spin trapping metal-centred radicals, nitrones form more persistent spin adducts with most small inorganic radicals.

Erroneous results may be obtained when hydrolysis and redox reactions involving the spin adducts are ignored. Spin trapping of pseudohalide radicals (·N_j· ·CN, ·SCN) are discussed in more detail.     

Apparent inhibition of photoredox reactions of magnesium octaethylporphyrin at the lipid bilayer-water interface by neutral quinones.

Neutral quinones rapidly equilibrate across the lipid bilayer, hereby rendering the photoeffects seen in pigmented bilayers sensitive to the redox properties at both interfaces. The lack of photoeffect by quinones themselves and their apparent quenching reactions with aqueous acceptors is thus explained. An aqueous donor is needed on one side to break the symmetry and to allow vectorial electron transfer to be recorded. It is concluded that the neutral quinone accumulates on the polar side of the interface with respect to the hydrophobic pigment. The system may allow the study of kinetics of proton transfer accompanying the redox reactions of the quinones.     

Role of quinone reductases in extracellular redox cycling in lichenized ascomycetes

Our previous work showed that many lichenized Ascomycetes can generate hydroxyl radicals using quinone-based extracellular redox cycling. During cycling, hydroquinones must be formed and subsequently regenerated from quinones using a quinone reductase (QR). However, we also showed that no simple correlation exists between QR activity and rates of hydroxyl radical formation. To further investigate the role of QR in hydroxyl radical formation, three model lichen species, Leptogium furfuraceum, Lasallia pustulata and Peltigera membranacea were selected for further investigation. All possessed QR activity and could metabolize quinones, and both Leptogium furfuraceum and Lasallia pustulata actively produced hydroxyl radicals. By contrast, P. membranacea produced almost no hydroxyl radicals, and although the lichen readily metabolized quinones, no hydroquinone production was detected. Peltigera had laccase (LAC) activity that was c. 50 times higher than in the other two species, suggesting that LAC rapidly oxidizes the hydroquinones, preventing radical formation deriving from auto-oxidation. It appears that in some lichens hydroxyl radical formation is blocked by the presence of high redox enzyme activity. QR from P. didactyla was studied further and found to display similar properties to the enzyme from free-living fungi, although it possessed an unusually high molecular mass (c. 62 kDa).     

Molecular mechanism of metal-independent decomposition of lipid hydroperoxide 13-HPODE by halogenated quinoid carcinogens

Halogenated quinones are a class of carcinogenic intermediates and newly identified chlorination disinfection by-products in drinking water. 13-Hydroperoxy-9,11-octadecadienoic acid (13-HPODE) is the most extensively studied endogenous lipid hydroperoxide. Although it is well known that the decomposition of 13-HPODE can be catalyzed by transition metal ions, it is not clear whether halogenated quinones could enhance its decomposition independent of metal ions and, if so, what the unique characteristics and similarities are. Here we show that 2,5-dichloro-1,4-benzoquinone (DCBQ) could markedly enhance the decomposition of 13-HPODE and formation of reactive lipid alkyl radicals such as pentyl and 7-carboxyheptyl radicals, and the genotoxic 4-hydroxy-2-nonenal (HNE), through the complementary application of ESR spin trapping, HPLC–MS, and GC–MS methods. Interestingly, two chloroquinone–lipid alkoxyl conjugates were also detected and identified from the reaction between DCBQ and 13-HPODE. Analogous results were observed with other halogenated quinones. This represents the first report that halogenated quinoid carcinogens can enhance the decomposition of the endogenous lipid hydroperoxide 13-HPODE and formation of reactive lipid alkyl radicals and genotoxic HNE via a novel metal-independent nucleophilic substitution coupled with homolytic decomposition mechanism, which may partly explain their potential genotoxicity and carcinogenicity.     

Thiyl radicals and induction of protein degradation

Thiyl radicals are important intermediates in the redox biology and chemistry of thiols. These radicals can react via hydrogen transfer with various C-H bonds in peptides and proteins, leading to the generation of carbon-centered radicals, and, potentially, to irreversible protein damage. This review summarizes quantitative information on reaction kinetics and product formation, and discusses the significance of these reactions for protein degradation induced by thiyl radical formation.     

Molecular mechanisms of quinone cytotoxicity

Quinones are probably found in all respiring animal and plant cells. They are widely used as anticancer, antibacterial or antimalarial drugs and as fungicides. Toxicity can arise as a result of their use as well as by the metabolism of other drugs and various environmental toxins or dietary constituents. In rapidly dividing cells such as tumor cells, cytotoxicity has been attributed to DNA modification. However the molecular basis for the initiation of quinone cytotoxicity in resting or non-dividing cells has been attributed to the alkylation of essential protein thiol or amine groups and/or the oxidation of essential protein thiols by activated oxygen species and/or GSSG. Oxidative stress arises when the quinone is reduced by reductases to a semiquinone radical which reduces oxygen to superoxide radicals and reforms the quinone. This futile redox cycling and oxygen activation forms cytotoxic levels of hydrogen peroxide and GSSG is retained by the cell and causes cytotoxic mixed protein disulfide formation. Most quinones form GSH conjugates which also undergo futile redox cycling and oxygen activation. Prior depletion of cell GSH markedly increases the cell's susceptibility to alkylating quinones but can protect the cell against certain redox cycling quinones. Cytotoxicity induced by hydroquinones in isolated hepatocytes can be attributed to quinones formed by autoxidation. The higher redox potential benzoquinones and naphthoquinones are the most cytotoxic presumably because of their higher electrophilicty and thiol reactivity and/or because the quinones or GSH conjugates are more readily reduced to semiquinones which activate oxygen.     

Hydroxyl radical generation by oligonucleotide derivatives of anthracycline antibiotic and synthetic quinone.

For the first time the covalent binding of anticancer anthracycline drugs and their potential synthetic analogs to oligonucleotides of different sequences is proposed for obtaining site-specific DNA scission in systems in vitro and in vivo. New compounds such as daunomycin (Dm) and synthetic naphthoquinone (NQ), covalently bound to the heptadeoxynucleotide of pCCAAACA (Dm-pN7) and decadeoxythymidilate (pT10p-NQ), have been obtained. These oligonucleotide derivatives can form specific complexes with complementary oligonucleotide sequences; these compounds and their complementary complexes can be reduced by purified NADPH-cytochrome P-450 reductase. Using the spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO), it has been shown that in aerobic conditions Dm-pN7 and pT10p-NQ are capable of generating OH radicals with and without complementary oligonucleotides. The chemical stability of the compounds in redox reactions has been studied. Oligonucleotide derivatives of natural and synthetic quinones capable of generating OH radicals seem to be a promising tool for site-specific scission of DNA in solution and in cells.     

Free radicals in physiology and pathology.

Free radicals are molecules with odd number of electrons and a high instability. Free radicals, which can occur in both organic (i.e., quinones) and inorganic molecules (i.e., O2-), are very reactive and their reactions are critical for the normal activity of a wide spectrum of biologic processes. They are also produced in the catalytic action of a variety of cellular enzymes and electron transport processes and are implicated in a number of physiologic and pathologic processes. Organisms can be exposed to free radicals in many ways other than through the processes of normal metabolism. Irradiation of organisms with electromagnetic radiation generates primary radicals (e-aq, OH., and H.), which can then undergo secondary reactions with dissolved O2 or with cellular solutes. In addition, a wide variety of environmental agents (drugs capable of redox cycling, and xenobiotics that can form free radical metabolites) including the aging process cause free radical damage to cells. This review deals with the reactions they can undergo and discusses the free radicals related to toxicology.     

Peroxidase catalyzed oxidation of naturally-occurring phenols and hardwood lignins

Horseradish peroxidase and hydrogen peroxide form phenoxy radicals from 4-substituted-2,6-dimethoxyphenols, milled wood lignin and alkali lignins. A number of factors governing this reaction are examined. Side chain cleavage to quinones is the principal disproportionation reaction of these radicals. Catalysis by UV light and inhibition by quinones is observed. Aerobic oxidation of phenols is catalyzed by small amounts of hydrogen peroxide. Lignin substrates are degraded by the same oxidation mechanism as are the simple phenolic substrates.     

The aprotic electrochemistry of quinones

Quinones play important roles in biological electron transfer reactions in almost all organisms, with specific roles in many physiological processes and chemotherapy. Quinones participate in two-electron, two-proton reactions in aqueous solution at equilibrium near neutral pH, but protons often lag behind the electron transfers. The relevant reactions in proteins are often sequential one electron redox processes without involving protons. Here we report the aprotic electrochemistry of the two half-couples, Q/Q^.- and Q^.?/Q⁼, of 11 parent quinones and 118 substituted 1,4-benzoquinones, 91 1,4-naphthoquinones, and 107 9,10-anthraquinones. The measured redox potentials are fit quite well with the Hammett para sigma (σ_para) parameter. Occasional exceptions can involve important groups, such as methoxy substituents in ubiquinone and hydroxy substituents in therapeutics. These can generally be explained by reasonable conjectures involving steric clashes and internal hydrogen bonds. We also provide data for 25 other quinones, 2 double quinones and 15 non-quinones, all measured under similar conditions.     

Effect Of Glutathione On The Redox Transitions Of Naphthohydroquinone Derivatives Formed During Dt-Diaphorase Catalysis

The oxidation of GSH coupled to the redox transitions of 1, Cnaphthoquinone derivatives during DT-diaphorase catalysis was examined. The quinones studied included 1,4-naphthoquinone and its dimethoxy-and hydroxy derivatives and were selected according to their different ability to undergo nucleophilic addition with GSH and the dual effect of superoxide dismutase on hydroquinone autoxidation

GSH was oxidized to GSSG during the redox transitions of the above quinones, regardless of their substitution pattern. This effect was accompanied by an increase of total O₂ consumption, indicating the ability of GSH to support quinone redox cycling. The values for the relationship [O₂]_consumed[GSSG]_formde were, with every quinone examined, above unity. thus pointing to the occurrence of autoxidation reactions other than those involved during GSSG formation

These results are discussed in terms of the functional group chemistry of the quinones and the ther-modynamic properties of the reactions involved in the reduction of the semi- to the hydro-quinone by GSH     

Reactions of glutathione and glutathione radicals with benzoquinones.

The reactions of glutathione (GSH) and glutathione radicals with a series of methyl-substituted 1,4-benzoquinones and 1,4-benzoquinone have been studied. It was found that by mixing excess benzoquinone with glutathione at pH above 6.5, the products formed were complex and unstable. All of the other experiments were carried out at pH 6.0, where the main product was stable for several hours. Stopped-flow analysis allowed the measurement of the rates of the rapid reactions between GSH and the quinones, and the products were monitored by High Performance Liquid Chromatography (HPLC). The rates of the reactions vary by five orders of magnitude and must be influenced by steric factors as well as changes in the redox states. It was observed that simple hydroquinones were not formed when the different benzoquinones were mixed with excess GSH and suggests that the initial reaction is addition/reduction rather than electron transfer. In the presence of excess quinone, the hydroquinone of the glutathione conjugate is oxidized back to its quinone. The rates of the reaction were measured. By using the technique of pulse radiolysis, it was possible to measure the reduction of the quinones by GSSG.- and the oxidation of hydroquinones by GS(.). It is proposed that the appearance of GSSG in reactions of quinones with glutathione could be due to oxidation of the hydroquinone by oxygen and the subsequent superoxide or H2O2 promoting the oxidation of GSH to GSSG.     

Free radical scavenging reactions and antioxidant activity of embelin: biochemical and pulse radiolytic studies

Embelin (from Embelia ribes) is a component of herbal drugs and possess wide range of medicinal properties. These properties may be, in part, due to scavenging of oxidizing free radicals. In this context, free radical scavenging reactions and antioxidant activity of embelin (2,5-dihydroxy-3-undecyl-1,4-benzoquinone) have been studied. It has been found to scavenge DPPH radical and inhibit hydroxyl radical induced deoxyribose degradation. It has been also found to inhibit lipid peroxidation and restore impaired Mn-superoxide dismutase in rat liver mitochondria. Further, kinetics and mechanism of the reactions of embelin with hydroxyl, one-electron oxidizing, organo-haloperoxyl and thiyl radicals have been studied using nanosecond pulse radiolysis technique. Its redox potential has been also evaluated with cyclic voltammetry. These studies suggest that embelin can act as a competitive antioxidant in physiological conditions.     

Cytochrome c reduction by semiquinone radicals can be indirectly inhibited by superoxide dismutase

The inhibition by superoxide dismutase of cytochrome c reduction by a range of semiquinone radicals has been studied. The semiquinones were produced from the parent quinones by reduction with xanthine and xanthine oxidase. Most of the quinones studied were favored over O₂ as the enzyme substrate, and in air as well as N₂, semiquinone radicals rather than superoxide were produced and they caused the cytochrome c reduction. With all but one of the quinones (benzoquinone), cytochrome c reduction in air was inhibited by superoxide dismutase, but the amount of enzyme required for inhibition was up to 100 times greater than that required to inhibit reduction by superoxide. It was highest for the quinones with the highest redox potential. These results demonstrate how superoxide dismutase can inhibit cytochrome c reduction by species other than superoxide. They can be explained by the dismutase displacing the equilibrium: semiquinone + O₂ ? quinone + O₂^? to the right, thereby allowing the forward reaction to out-compete other reactions of the semiquinone. The implication from these findings that superoxide dismutase-inhibitable reduction of cytochrome c may not be a specific test for superoxide production is discussed.