Succinate-ubiquinone reductase linked recycling of alpha-tocopherol in reconstituted systems and mitochondria: requirement for reduced ubiquinone.

Studies have demonstrated that accumulation of mitochondrial tocopheroxyl radical, the primary oxidation product of alpha-tocopherol, accompanies rapid consumption of tocopherol. Enzyme-linked electron flow lowers both the steady-state concentration of the radical and the consumption of tocopherol. Reduction of tocopheroxyl radical by a mitochondrial electron carrier(s) seems a likely mechanism of tocopherol recycling. Succinate-ubiquinone reductase (complex II) was incorporated into liposomes in the presence of tocopherol and ubiquinone-10. After inducing formation of tocopheroxyl radical, it was possible to show that reduced ubiquinone prevents radical accumulation and tocopherol consumption. There was no evidence of direct reduction of tocopheroxyl radical by succinate-reduced complex II. These reactions were also measured using ubiquinone-1 and alpha-C-6-chromanol (2,5,7,8-tetramethyl-2-(4'-methylpentyl)-6-chromanol) which are less hydrophobic analogues of ubiquinone-10 and alpha-tocopherol. Mitochondrial membranes were made deficient in ubiquinone but sufficient in alpha-tocopherol and were reconstituted with added quinone. With these membranes it was shown that mitochondrial enzyme-linked reduction of ubiquinone protects alpha-tocopherol from consumption, and there is a requirement for ubiquinone. This complements the observations made in liposomes and we propose that reduced mitochondrial ubiquinones have a role in alpha-tocopherol protection, presumably through efficient reduction of the tocopheroxyl radical.     

The role of ubiquinone in the respiratory chain of Acetobacter xylinum

1. Whole cells of Acetobacter xylinum were found to contain a quinone of the ubiquinone (coenzyme Q) group. The quinone was isolated from the cells and crystallized. It was identified by its physical, chemical and spectroscopic properties as a ubiquinone with 10 isoprene units (ubiquinone-10). No naphthaquinone was detected in the cells. 2. Cell-free extracts prepared by means of a French pressure cell were separated into three fractions by differential centrifugation. The ubiquinone was located predominantly in the particulate fraction sedimenting at 33000g, which also contained most of the NADH oxidase and malate oxidase activities. The concentration of ubiquinone-10 in extracts was similar to that of the flavoproteins and about three times the concentration of the individual cytochromes. 3. Aerobic incubations of crude extracts with either NADH or malate resulted in reduction of the endogenous ubiquinone-10 to steady-state concentrations of 55 and 40% of the total quinone respectively. In the presence of cyanide more than 95% of the endogenous ubiquinone-10 was reduced by either NADH or malate. 4. The initial rate of reduction of endogenous ubiquinone-10 by malate and the rate of ubiquinol oxidation, in A. xylinum extracts, were found to be compatible with the overall rate of malate oxidation with oxygen. 5. The effects of various respiratory inhibitors on the oxidation-reduction reactions of the endogenous quinone indicate that its position on the respiratory chain is between the malate flavoprotein dehydrogenase and the cytochrome chain.     

Dynamics of antioxidant action of ubiquinol: a reappraisal.

The dynamics of action of ubiquinol as an antioxidant against lipid peroxidation was reinvestigated and compared with that of alpha-tocopherol. It was found that ubiquinol was 2.5 and 1.9 times more reactive than alpha-tocopherol toward phenoxyl and peroxyl radicals, respectively, at 25 degrees C in ethanol and that it was capable of donating two hydrogen atoms toward oxygen radicals but that the apparent stoichiometric number decreased in the inhibition of lipid peroxidation, to even smaller than 1, due to its autoxidation. The autoxidation of ubiquinol proceeded even in the micelles and liposomal membranes in aqueous dispersions as well as in organic homogeneous solution. The apparent antioxidant activity of ubiquinol was smaller than that of alpha-tocopherol against lipid peroxidation in organic solution as judged from either rate of oxidation or duration of inhibition period. They exerted similar antioxidant potency against lipid peroxidation in the membranes and micelles in aqueous dispersions. The combination of ubiquinol and alpha-tocopherol was suggested to be effective.     

Phenotypes of fission yeast defective in ubiquinone production due to disruption of the gene for p-hydroxybenzoate polyprenyl diphosphate transferase

Ubiquinone is an essential component of the electron transfer system in both prokaryotes and eukaryotes and is synthesized from chorismate and polyprenyl diphosphate by eight steps. p-Hydroxybenzoate (PHB) polyprenyl diphosphate transferase catalyzes the condensation of PHB and polyprenyl diphosphate in ubiquinone biosynthesis. We isolated the gene (designated ppt1) encoding PHB polyprenyl diphosphate transferase from Schizosaccharomyces pombe and constructed a strain with a disrupted ppt1 gene. This strain could not grow on minimal medium supplemented with glucose. Expression of COQ2 from Saccharomyces cerevisiae in the defective S. pombe strain restored growth and enabled the cells to produce ubiquinone-10, indicating that COQ2 and ppt1 are functional homologs. The ppt1-deficient strain required supplementation with antioxidants, such as cysteine, glutathione, and alpha-tocopherol, to grow on minimal medium. This suggests that ubiquinone can act as an antioxidant, a premise supported by our observation that the ppt1-deficient strain is sensitive to H(2)O(2) and Cu(2+). Interestingly, we also found that the ppt1-deficient strain produced a significant amount of H(2)S, which suggests that oxidation of sulfide by ubiquinone may be an important pathway for sulfur metabolism in S. pombe. Ppt1-green fluorescent protein fusion proteins localized to the mitochondria, indicating that ubiquinone biosynthesis occurs in the mitochondria in S. pombe. Thus, analysis of the phenotypes of S. pombe strains deficient in ubiquinone production clearly demonstrates that ubiquinone has multiple functions in the cell apart from being an integral component of the electron transfer system.     

Effect of oxygen free radicals on ubiquinone in aqueous solution and phospholipid vesicles

The purpose of this study was to evaluate the direct effect of oxygen free radicals produced by ultrasonic irradiation on ubiquinone and to compare the efficiency with which the antioxidant can compete with these radicals when it is both in aqueous solution and within the lipid bilayer. The main product obtained after insonation of aqueous solutions of ubiquinone-0 was ubiquinol, moreover some degradation occurred. The direct electron donor responsible for most of the ubiquinol generated by ultrasonic irradiation appeared to be superoxide radical. Addition reactions of hydroxyl radicals with aromatic ring structure led probably to degradation products of ubiquinone, which were not identified. Experiments were also performed to evaluate the efficiency with which ubiquinone-3 could react with oxygen radicals when it was within the lipid bilayer. The effect of presence or absence of a net surface charge was studied selecting a suitable bilayer including dimyristylphosphatidic acid or stearylamine in uncharged dimyristylphosphatidylcholine vesicles. In these systems hydroxyl radicals did not represent a potential danger for the antioxidant, the reaction between superoxide and ubiquinone-3 instead was significant only in positively charged membranes and gave rise to ubiquinol. It is suggested that ubiquinone acts as an antioxidant by stopping the propagation reaction.     

Pleiotropic phenotypes of fission yeast defective in ubiquinone-10 production. A study from the abc1Sp (coq8Sp) mutant

We previously constructed two Schizosaccahromyces pombe ubiquinone-10 (or Coenzyme Q10) less mutants, which are either defective for decaprenyl diphosphate synthase or p-hydroxybenzoate polyprenyl diphosphate transferase. To further confirm the roles of ubiquinone in S. pombe, we examined the phenotype of the abc1Sp (coq8Sp) mutant, which is highly speculated to be defective in ubiquinone biosynthesis. We show here that the abc1Sp defective strain did not produce UQ-10 and could not grow on minimal medium. The abc1Sp-deficient strain required supplementation with antioxidants such as cysteine or glutathione to grow on minimal medium. In support of the antioxidant function of ubiquinone, the abc1Sp-deficient strain is sensitive to H2O2 and Cu2+. In addition, expression of the stress inducible ctt1 gene was much induced in the ubiquinone less mutant than wild type. Interestingly, we also found that the abc1-deficient strain as well as other ubiquinone less mutants produced a significant amount of H2S, which suggests that oxidation of sulfide by ubiquinone may be an important pathway for sulfur metabolism in S. pombe. Thus, analysis of the phenotypes of S. pombe ubiquinone less mutants clearly demonstrate that ubiquinone has multiple functions in the cell apart from being an integral component of the electron transfer system.     

Identification of ubiquinone-50 as the major methylated nonpolar lipid in human monocytes. Regulation of its biosynthesis via methionine-dependent pathways and relationship to superoxide production

Human blood monocytes incorporated the methyl group from methionine into their neutral lipids. The major methylated product was identified as ubiquinone-50 in monocytes, lymphocytes, and a variety of human tumor cell lines by several analytical procedures including TLC or high performance liquid chromatography and as ubiquinone-45 in a mouse tumor cell line. Up to three methyl groups were shown to be derived from methionine by mass spectrometry. The rate of synthesis of ubiquinone-50 by monocytes as assessed by measuring labeled methyl group incorporation was shown to be linear over a 3-h period. Degradation of ubiquinone proceeded slowly; 80% of the labeled compound persisted after 18 h. The dependence of ubiquinone-50 synthesis upon methionine concentration was established in monocytes, with an estimated apparent Km for methionine of about 20 microM. The tumor promoter, tetradecanoate phorbol acetate, a potent stimulator of superoxide anion (O2-) production in phagocytic cells, inhibited ubiquinone-50 synthesis at nanomolar concentrations in monocytes, but not in lymphocytes, under conditions where oxidation of methionine takes place. Degradation of the labeled ubiquinone was unaffected. Formylmethionylleucyl-phenylalanine, a chemoattractant peptide which stimulates O2- production in phagocytic cells, also inhibited ubiquinone-50 synthesis. The degree of inhibition by either stimulus was increased when the methionine concentration in the medium was low. These findings demonstrate that in human monocytes ubiquinone-50 biosynthesis is regulable and that methionine concentration modulates both its rate of synthesis and the inhibitory effects of two stimuli of O2- production.     

Purification of the 45 kDa, membrane bound NADH dehydrogenase of Escherichia coli (NDH-2) and analysis of its interaction with ubiquinone analogues

The NADH:ubiquinone reductase (NDH-2) of Escherichia coli was expressed as a His-tagged protein, extracted from the membrane fraction using detergent and purified by chromatography. The His-tagged NDH-2 was highly active and catalyzed NADH oxidation by ubiquinone-1 at rates over two orders of magnitude higher than previously reported. The purified, His-tagged NDH-2, like native NDH-2, did not oxidize deamino-NADH. Steady-state kinetics were used to analyze the enzyme's activity in the presence of different electron acceptors. High V(max) and low K(m) values were only found for hydrophobic ubiquinone analogues, particularly ubiquinone-2. These findings strongly support the notion that NDH-2 is a membrane bound enzyme, despite the absence of predicted transmembrane segments in its primary structure. The latter observation is in agreement with possible evolutionary relation between NDH-2 and water-soluble enzymes such as dihydrolipoamide dehydrogenase. There is currently no clear indication of how NDH-2 binds to biological membranes.     

Nature, intracellular distribution and formation of terpenoid quinones in Euglena gracilis

1. Light-grown cells of Euglena gracilis strain Z, var. bacillaris and 1224/5g contain phylloquinone, plastoquinone, alpha-tocopherol, alpha-tocopherolquinone and ubiquinone-9 (i.e. ubiquinone with 9 isoprene units/mol.). 2. The concentration (per g. dry wt.) of plastoquinone (and chlorophyll) in light-grown cells of strain Z was governed by the composition of the culture medium and age of the cells. Highest yields of plastoquinone were obtained under autotrophic conditions, the concentration reaching a maximum after 6-8 days' growth. The concentrations were less in heterotrophic media. The concentration of ubiquinone was relatively unaffected by the age of the cells or composition of the medium. 3. In light-grown cells of strain Z plastoquinone, alpha-tocopherolquinone and alpha-tocopherol were mainly localized in the chloroplast; ubiquinone was found to be in the mitochondria. 4. Etiolated (dark-grown) cells of strain Z contained no phylloquinone, plastoquinone or alpha-tocopherolquinone; alpha-tocopherol was present in lower concentrations compared with light-grown cells; ubiquinone concentrations were similar to those for light-grown cells. The presence of alpha-tocopherol in etiolated cells suggested that this chromanol was not entirely confined to the chloroplast. 5. On illumination of etiolated cells of strain Z the chloroplastidic components plastoquinone, alpha-tocopherolquinone and alpha-tocopherol were synthesized in step with chloroplast formation. Ubiquinone concentrations, as expected, were unaffected. 6. [2-(14)C]Mevalonic acid, the specific distal terpenoid precursor, was not incorporated into any of the terpenoid components examined. This was attributed to the impermeability of the cell wall to this compound, rather than to a novel pathway of terpenoid biosynthesis.     

The mode of action of lipid-soluble antioxidants in biological membranes: relationship between the effects of ubiquinol and vitamin E as inhibitors of lipid peroxidation in submitochondrial particles.

The effects of ubiquinol and vitamin E on ascorbate- and ADP-Fe3+-induced lipid peroxidation were investigated by measuring oxygen consumption and malondialdehyde formation in beef heart submitochondrial particles. In the native particles, lipid peroxidation showed an initial lag phase, which was prolonged by increasing concentrations of ascorbate. Lipid peroxidation in these particles was almost completely inhibited by conditions leading to a reduction of endogenous ubiquinone, such as the addition of succinate or NADH in the presence of antimycin. Lyophilization of the particles followed by three or four consecutive extractions with pentane resulted in a complete removal of vitamin E and a virtually complete removal of ubiquinone, as revealed by reversed-phase high pressure liquid chromatography. In these particles, lipid peroxidation showed no significant lag phase and was not inhibited by either increasing concentrations of ascorbate or conditions leading to ubiquinone reduction. Treatment of the particles with a pentane solution of vitamin E (alpha-tocopherol) restored the lag phase and its prolongation by increasing ascorbate concentrations. Treatment of the extracted particles with pentane containing ubiquinone-10 resulted in a restoration of the inhibition of lipid peroxidation by succinate or NADH in the presence of antimycin, but not the initial lag phase or its prolongation by increasing concentrations of ascorbate. Malonate and rotenone, which prevent the reduction of ubiquinone by succinate and NADH, respectively, abolished, as expected, the inhibition of the initiation of lipid peroxidation in both native and ubiquinone-10-supplemented particles. Reincorporation of both vitamin E and ubiquinone-10 restored both effects.(ABSTRACT TRUNCATED AT 250 WORDS)     

Antioxidant Activity of Ubiquinone-3 in Human Low Density Lipoprotein

The ability of ubiquinone-3, a short chain ubiquinone homologue, to prevent Cu²⁺ induced oxidation of human low density lipoprotein was investigated. The results are as follows: in the presence of ubiquinone-3 the extent of peroxidation, as determined by the formation of thiobarbituric acid reactive substances, was only one third of that found in its absence; the quinone can also prevent the fragmentation of apolipo-protein B-100 and the increase of the net negative surface charge of the particle.     

Action of quinolinic and indolinonic aminoxyls as radical-scavenging antioxidants.

The kinetic studies on the actions of quinolinic and indolinonic aminoxyls in the oxidation of lipid peroxidation induced by free radicals were carried out to evaluate their antioxidant activity. These aminoxyls showed a similar reactivity toward peroxyl radical with alpha-tocopherol. The antioxidant efficacies of aminoxyls against oxidation of methyl linoleate in homogeneous solution were smaller than that of alpha-tocopherol. Hydroxylamine, a reduced form of aminoxyl, possessed a comparative antioxidant efficacy with alpha-tocopherol and was capable of suppressing the consumption of alpha-tocopherol. Aminoxyls showed more potent antioxidant activity than alpha-tocopherol against the oxidation of methyl linoleate micelles induced by peroxyl radical or by a combination of copper ion and hydrogen peroxide. These results suggest that quinolinic and indolinonic aminoxyls may act as potent antioxidants against lipid peroxidation, especially in the presence of a good reductant which reduces aminoxyl radicals to hydroxylamines.     

Identification of regulatory sites in the biosynthesis of ubiquinone in the perfused rat heart

The biosynthesis of ubiquinone was studied in an isolated perfused beating heart preparation from adult male rats to determine rate-limiting steps in the biosynthetic pathway. The isolated heart could incorporate p-hydroxy[U-14C]benzoate into ubiquinones (ubiquinone-9 and -10) and two other lipids which were identified as 3-nonaprenyl 4-hydroxybenzoate and 3-decaprenyl 4-hydroxybenzoate. No other lipids could be detected. Addition of unlabeled mevalonolactone to the perfusate stimulated the rate of incorporation of p-hydroxy[U-14C]benzoate into 3-nonaprenyl 4-hydroxybenzoate and 3-decaprenyl 4-hydroxybenzoate. The level of radioactivity in these intermediates was much greater than that in ubiquinone-9 and -10. These results show that in the intact heart there is a large excess capacity to form postmevalonate isoprenoid precursors of ubiquinone and suggest a possible regulatory step at the premevalonate level. Moreover, the accumulation of prenylated derivatives of 4-hydroxybenzoic acid indicates further rate limitation at one or more of the subsequent steps in conversion of these intermediates to ubiquinone.     

Antioxidant activities of probucol against lipid peroxidations.

The antioxidant activities of probucol were measured in the oxidations of methyl linoleate in homogeneous solution and soybean phosphatidylcholine liposomal membranes and also of low-density lipoproteins. When an excess amount of probucol was reacted with galvinoxyl, the EPR spectrum of galvinoxyl disappeared and a new triplet EPR signal was found: g = 2.0058 and aH(2H) = 0.14 mT. The identical EPR spectrum was observed when probucol was reacted with tert-butoxyl radical generated from di-tert-butylperoxy oxalate. This EPR signal disappeared rapidly when reacted with either alpha-tocopherol or 6-O-palmitoyl-ascorbic acid. Probucol suppressed the free-radical-mediated oxidations of methyl linoleate in hexane and in acetonitrile, in a dose-dependent manner. Its antioxidant activity was 17.5-fold less than that of alpha-tocopherol in hexane. Probucol incorporated into soybean phosphatidylcholine liposomes suppressed its oxidation. The antioxidant activity of probucol was less than that of alpha-tocopherol, but the difference between the two antioxidant activities was smaller in the membranes than in homogeneous solution. Probucol also suppressed the oxidation of low-density lipoprotein. Interestingly, probucol suppressed the oxidation of LDL as efficiently as alpha-tocopherol, implying that physical factors as well as chemical reactivity are important in determining the overall activity of antioxidant in low-density lipoprotein.     

Glucagon treatment of rats activates the respiratory chain of liver mitochondria at more than one site

The rate of reduction of ferricyanide in the presence and absence of antimycin and ubiquinone-1 was measured using liver mitochondria from control and glucagon treated rats. Glucagon treatment was shown to increase electron flow from both NADH and succinate to ubiquinone, and from ubiquinone to cytochrome c. 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) was shown to inhibit the oxidation of glutamate + malate to a much greater extent than that of succinate or duroquinol. Spectral and kinetic studies confirmed that electron flow between NADH and ubiquinone was the primary site of action but that the interaction of the ubiquinone pool with complex 3 was also affected. The effects of various respiratory chain inhibitors on the rate of uncoupled oxidation of succinate and glutamate + malate by control and glucagon treated mitochondria were studied. The stimulation of respiration seen in the mitochondria from glucagon treated rats was maintained or increased as respiration was progressively inhibited with DCMU, 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone (DBMIB), 2-heptyl-4-hydroxyquinoline-n-oxide (HQNO) and colletotrichin, but greatly reduced when inhibition was produced with malonate or antimycin. These data were also shown to support the conclusion that glucagon treatment may cause some stimulation of electron flow through NADH dehydrogenase, succinate dehydrogenase and through the bc1 complex, probably at the point of interaction of the complexes with the ubiquinone pool. The effects of glucagon treatment on duroquinol oxidation and the inhibitor titrations could not be mimicked by increasing the matrix volume, nor totally reversed by aging of mitochondria. These are both processes that have been suggested as the means by which glucagon exerts its effects on the respiratory chain (Armston, A.E., Halestrap, A.P. and Scott, R.D., 1982, Biochim. Biophys. Acta 681, 429-439). It is concluded that an additional mechanism for regulating electron flow must exist and a change in lipid peroxidation of the inner mitochondrial membrane is suggested.     

Inhibition by capsaicin of NADH-quinone oxidoreductases is correlated with the presence of energy-coupling site 1 in various organisms

The NADH-ubiquinone reductase activity of the respiratory chains of several organisms was inhibited by capsaicin and dihydrocapsaicin, which are the pungent principles of red pepper. This inhibition was correlated with the presence of an energy transducing site in this segment of the respiratory chain. Where the NADH-quinone oxidoreductase segment involved an energy coupling site (e.g., in Paracoccus denitrificans, Escherichia coli, and Thermus thermophilus HB-8 membranes and bovine heart mitochondria), capsaicin acted as an inhibitor of ubiquinone reduction by NADH. In contrast, where this energy coupling site was absent (e.g., in Saccharomyces cerevisiae mitochondria and Bacillus subtilis membranes), there was no inhibition of NADH-ubiquinone reductase activity by capsaicin. The capsaicin inhibition of Paracoccus membranes was reversed by washing the membranes with medium containing bovine serum albumin. In the E. coli and Paracoccus membranes and bovine submitochondrial particles, capsaicin acted as a noncompetitive inhibitor for ubiquinone-1 at lower concentrations of ubiquinone-1 (less than 20 microM) and as a competitive inhibitor at higher concentrations of ubiquinone-1 (greater than 50 microM). In addition, the concentrations of capsaicin required for 50% inhibition of NADH oxidase activity of bovine submitochondrial particles were increased when ubiquinone-10 was added to the particles. The mechanism by which capsaicin inhibits the energy-transducing NADH-quinone oxidoreductase is discussed.     

The biomolecule ubiquinone exerts a variety of biological functions

The chemistry of ubiquinone allows reversible addition of single electrons and protons. This unique property is used in nature for aerobic energy gain, for unilateral proton accumulation, for the generation of reactive oxygen species involved in physiological signaling and a variety of pathophysiological events. Since several years ubiquinone is also considered to play a major role in the control of lipid peroxidation, since this lipophilic biomolecule was recognized to recycle alpha-tocopherol radicals back to the chain-breaking form, vitamin E. Ubiquinone is therefore a biomolecule which has increasingly focused the interest of many research groups due to its alternative pro- and antioxidant activity. We have intensively investigated the role of ubiquinone as prooxidant in mitochondria and will present experimental evidences on conditions required for this function, we will also show that lysosomal ubiquinone has a double function as proton translocator and radical source under certain metabolic conditions. Furthermore, we have addressed the antioxidant role of ubiquinone and found that the efficiency of this activity is widely dependent on the type of biomembrane where ubiquinone exerts its chain-breaking activity.     

Recycling and antioxidant activity of tocopherol homologs of differing hydrocarbon chain lengths in liver microsomes

Tocopherols (vitamin E) function as inhibitors of lipid peroxidation in biomembranes by donating a hydrogen atom to the chain propagating lipid radicals, thus giving rise to chromanoxyl radicals of the antioxidant. We have shown that alpha-tocopherol homologs differing in the lengths of their hydrocarbon side chains (alpha-Cn) manifest strikingly different antioxidant potencies in membranes. The antioxidant activity of tocopherol homologs during (Fe2+ + ascorbate)- or (Fe2+ + NADPH)-induced lipid peroxidation in rat liver microsomes increased in the order alpha-tocopherol (alpha-C16) less than alpha-C11 less than alpha-C6 less than alpha-C1. Chromanoxyl radicals generated from alpha-tocopherol and its more polar homologs by an enzymatic oxidation system (lipoxygenase + linolenic acid) can be recycled in rat liver microsomes by NAD-PH-dependent electron transport or by ascorbate. The efficiency of recycling increased in the same order: alpha-tocopherol (alpha-C16) less than alpha-C11 less than alpha-C6 less than alpha-C1. Thus the high efficiency of regeneration of short-chain homologs of vitamin E may account for their high antioxidant potency.     

Kinetics of the lipoperoxyl radical-scavenging activity of indicaxanthin in solution and unilamellar liposomes

The reaction of the phytochemical indicaxanthin with lipoperoxyl radicals generated in methyl linoleate methanol solution by 2,2'-azobis(2,4-dimethylvaleronitrile), and in aqueous soybean phosphatidylcholine unilamellar liposomes by 2,2'-azobis(2-amidinopropane)hydrochloride, was studied. The molecule acts as a chain-terminating lipoperoxyl radical scavenger in solution, with a calculated inhibition constant of 3.63 x 10(5) M(-1) s(-1), and a stoichiometric factor approaching 2. Indicaxanthin incorporated in liposomes prevented lipid oxidation, inducing clear-cut lag periods and decrease of the propagation rate. Both effects were concentration-dependent, but not linearly related to the phytochemical concentration. The consumption of indicaxanthin during liposome oxidation was remarkably delayed, the lower the concentration the longer the time-interval during which it remained in its native state. Indicaxanthin and alpha-tocopherol, simultaneously incorporated in liposomes, exhibited cooperative antioxidant effects and reciprocal protective interactions. The extent of synergism decreased at the increase of the ratio (indicaxanthin)/(alpha-tocopherol). A potential antioxidant mechanism of indicaxanthin is discussed in the context of the chemistry of the molecule, and of the possible reactivity of a short-lived intermediate.     

Coenzyme Q10 enrichment decreases oxidative DNA damage in human lymphocytes

Ubiquinol-10, the reduced form of coenzyme Q10, is a powerful antioxidant in plasma and lipoproteins. It has been suggested that endogenous ubiquinol-10 also exerts a protective role even towards DNA oxidation mediated by lipid peroxidation. Even though the antioxidant activity of coenzyme Q10 is mainly ascribed to ubiquinol-10, a role for ubiquinone-10 (the oxidized form), has been suggested not only if appropriate reducing systems are present. To investigate whether the concentration of ubiquinol-10 or ubiquinone-10 affects the extent of DNA damage induced by H2O2, we supplemented in vitro human lymphocytes with both forms of coenzyme Q10 and evaluated the DNA strand breaks by Comet assay. The exposure of lymphocytes to 100 microM H2O2 resulted in rapid decrease of cellular ubiquinol-10 content both in ubiquinol-10-enriched and in control cells, whereas alpha-tocopherol and beta-carotene concentration were unchanged. After 30 min from H2O2 exposure, the amount of DNA strand breaks was lower and cells' viability was significantly higher in ubiquinol-10-enriched cells compared with control cells. A similar trend was observed in ubiquinone-10-enriched lymphocytes when compared with control cells. Our experiments suggest that coenzyme Q10 in vitro supplementation enhances DNA resistance towards H2O2-induced oxidation, but it doesn't inhibit directly DNA strand break formation.