Binding of FAD to cytochrome b558 is facilitated during activation of the phagocyte NADPH oxidase, leading to superoxide production

The superoxide-producing phagocyte NADPH oxidase can be reconstituted in a cell-free system. The activity of NADPH oxidase is dependent on FAD, but the physiological status of FAD in the oxidase is not fully elucidated. To clarify the role of FAD in NADPH oxidase, FAD-free full-length recombinant p47(phox), p67(phox), p40(phox), and Rac were prepared, and the activity was reconstituted with these proteins and purified cytochrome b(558) (cyt b(558)) with different amounts of FAD. A remarkably high activity, over 100 micromol/s/micromol heme, was obtained in the oxidase with purified cyt b(558), ternary complex (p47-p67-p40(phox)), and Rac. From titration with FAD of the activity of NADPH oxidase reconstituted with purified FAD-devoid cyt b, the dissociation constant K(d) of FAD in cyt b(558) of reconstituted oxidase was estimated as nearly 1 nm. We also examined addition of FAD on the assembly process in reconstituted oxidase. The activity was remarkably enhanced when FAD was present during assembly process, and the efficacy of incorporating FAD into the vacant FAD site in purified cyt b(558) increased, compared when FAD was added after assembly processes. The absorption spectra of reconstituted oxidase under anaerobiosis showed that incorporation of FAD into cyt b(558) recovered electron flow from NADPH to heme. From both K(d) values of FAD and the amount of incorporated FAD in cyt b(558) of reconstituted oxidase, in combination with spectra, we propose the model in which the K(d) values of FAD in cyt b(558) is changeable after activation and FAD binding works as a switch to regulate electron transfer in NADPH oxidase.     

A rapid method to quantify pro-oxidant activity in cultures of wood-decaying white-rot fungi

A new, rapid method for evaluation of lipid peroxidation promoting (pro-oxidant) activity in cultures of wood-decaying fungi was developed. The method is based on measurement of the rate of oxygen consumption in the reaction of linoleic acid peroxidation initiated by fungal culture filtrates. The liquid cultures of the white-rot fungi Bjerkandera adusta and Phanerochaete chrysosporium grown on wheat straw-containing glucose-peptone-corn steep liquor medium possessed significant levels of the pro-oxidant activity. Other white-rot fungi producing manganese peroxidase (MnP) were also found to show the activity. MnP demonstrated a crucial role as the major pro-oxidant agent in the fungal cultures. The total pro-oxidant activity may be considered as net result of the peroxidation by MnP and the inhibition by antioxidant compounds present in the fungal culture fluids.     

Mechanism of the antioxidant to pro-oxidant switch in the behavior of dehydroascorbate during LDL oxidation by copper(II) ions

Oxidised low density lipoprotein (LDL) may be involved in the pathogenesis of atherosclerosis. We have therefore investigated the mechanisms underlying the antioxidant/pro-oxidant behavior of dehydroascorbate, the oxidation product of ascorbic acid, toward LDL incubated with Cu(2+) ions. By monitoring lipid peroxidation through the formation of conjugated dienes and lipid hydroperoxides, we show that the pro-oxidant activity of dehydroascorbate is critically dependent on the presence of lipid hydroperoxides, which accumulate during the early stages of oxidation. Using electron paramagnetic resonance spectroscopy, we show that dehydroascorbate amplifies the generation of alkoxyl radicals during the interaction of copper ions with the model alkyl hydroperoxide, tert-butylhydroperoxide. Under continuous-flow conditions, a prominent doublet signal was detected, which we attribute to both the erythroascorbate and ascorbate free radicals. On this basis, we propose that the pro-oxidant activity of dehydroascorbate toward LDL is due to its known spontaneous interconversion to erythroascorbate and ascorbate, which reduce Cu(2+) to Cu(+) and thereby promote the decomposition of lipid hydroperoxides. Various mechanisms, including copper chelation and Cu(+) oxidation, are suggested to underlie the antioxidant behavior of dehydroascorbate in LDL that is essentially free of lipid hydroperoxides.     

Priming of the neutrophil respiratory burst involves p38 mitogen-activated protein kinase-dependent exocytosis of flavocytochrome b558-containing granules

The respiratory burst of human neutrophils is primed by a number of pro-inflammatory stimuli, including tumor necrosis factor-alpha (TNFalpha) and lipopolysaccharide (LPS); however, the mechanism of priming remains unknown. LPS has been shown previously to increase membrane expression of flavocytochrome b(558), a component of the NADPH oxidase. This study shows that TNFalpha also increases membrane expression of flavocytochrome b(558). Mitogen-activated protein kinase (MAPK) modules have been implicated in the action of priming agents. Pharmacologic inhibitors of MAPKs, SB203580 and PD098059, revealed that priming of the respiratory burst and up-regulation of flavocytochrome b(558) are dependent on p38 MAPK but not on extracellular-signal regulated kinase (ERK). TNFalpha and LPS primed respiratory burst activity and increased membrane expression of CD35 and CD66b, specific markers of secretory vesicles and specific granules that contain flavocytochrome b(558), with similar time courses and concentration dependences. These processes also required p38 MAPK but were independent of ERK. TNFalpha failed to prime respiratory burst activity or to increase membrane CD35 expression in enucleated neutrophil cytoplasts. These data suggest that one mechanism by which TNFalpha and LPS prime neutrophil respiratory burst activity is by increasing membrane expression of flavocytochrome b(558) through exocytosis of intracellular granules in a process regulated by p38 MAPK.     

Inactivation of NADP+-dependent isocitrate dehydrogenase by lipid peroxidation products

Membrane lipid peroxidation processes yield products that may react with proteins to cause oxidative modification. Recently, we demonstrated that the control of cytosolic and mitochondrial redox balance and oxidative damage is one of the primary functions of NADP₊-dependent isocitrate dehydrogenase (ICDH) through to supply NADPH for antioxidant systems. When exposed to lipid peroxidation products, such as malondialdehyde (MDA), 4-hydroxynonenal (HNE) and lipid hydroperoxide, ICDH was susceptible to oxidative damage, which was indicated by the loss of activity and the formation of carbonyl groups. The structural alterations of modified enzymes were indicated by the change in thermal stability, intrinsic tryptophan fluorescence and binding of the hydrophobic probe 8-anilino 1-napthalene sulfonic acid. Upon exposure to 2,2'-azobis(2-amidinopropane) hydrochloride (AAPH), which induces lipid peroxidation in membrane, a significant decrease in both cytosolic and mitochondrial ICDH activities were observed in U937 cells. Using immunoprecipitation and immunoblotting, we were able to isolate and positively identify HNE adduct in mitochondrial ICDH from AAPH-treated U937 cells. The lipid peroxidation-mediated damage to ICDH may result in the perturbation of the cellular antioxidant defense mechanisms and subsequently lead to a pro-oxidant condition.     

Glutathione Cycle Impairment Mediates Aβ-induced Cell Toxicity

Membrane lipid peroxidation processes yield products that may react with proteins to cause oxidative modification. Recently, we demonstrated that the control of cytosolic and mitochondrial redox balance and oxidative damage is one of the primary functions of NADP⁺-dependent isocitrate dehydrogenase (ICDH) through to supply NADPH for antioxidant systems. When exposed to lipid peroxidation products, such as malondialdehyde (MDA), 4-hydroxynonenal (HNE) and lipid hydroperoxide, ICDH was susceptible to oxidative damage, which was indicated by the loss of activity and the formation of carbonyl groups. The structural alterations of modified enzymes were indicated by the change in thermal stability, intrinsic tryptophan fluorescence and binding of the hydrophobic probe 8-anilino 1-napthalene sulfonic acid. Upon exposure to 2,2′-azobis(2-amidinopropane) hydrochloride (AAPH), which induces lipid peroxidation in membrane, a significant decrease in both cytosolic and mitochondrial ICDH activities were observed in U937 cells. Using immunoprecipitation and immunoblotting, we were able to isolate and positively identify HNE adduct in mitochondrial ICDH from AAPH-treated U937 cells. The lipid peroxidation-mediated damage to ICDH may result in the perturbation of the cellular antioxidant defense mechanisms and subsequently lead to a pro-oxidant condition.     

Lipid peroxidation and 4-hydroxy-2-nonenal formation by copper ion bound to amyloid-beta peptide

The lipid peroxidation product 4-hydroxy-2-nonenal (HNE) is proposed to be a toxic factor in the pathogenesis of Alzheimer disease. The primary products of lipid peroxidation are phospholipid hydroperoxides, and degraded reactive aldehydes, such as HNE, are considered secondary peroxidation products. In this study, we investigated the role of amyloid-beta peptide (A beta) in the formation of phospholipid hydroperoxides and HNE by copper ion bound to A beta. The A beta1-42-Cu2+ (1:1 molar ratio) complex showed an activity to form phospholipid hydroperoxides from a phospholipid, 1-palmitoyl-2-linoleoyl phosphatidylcholine, through Cu2+ reduction in the presence of ascorbic acid. The phospholipid hydroperoxides were considered to be a racemic mixture of 9-hydroperoxide and 13-hydroperoxide of the linoleoyl residue. When Cu2+ was bound to 2 molar equivalents of A beta(1-42) (2 A beta1-42-Cu2+), lipid peroxidation was inhibited. HNE was generated from one of the phospholipid hydroperoxides, 1-palmitoyl-2-(13-hydroperoxy-cis-9, trans-11-octadecadienoyl) phosphatidylcholine (PLPC-OOH), by free Cu2+ in the presence of ascorbic acid through Cu2+ reduction and degradation of PLPC-OOH. HNE generation was markedly inhibited by equimolar concentrations of A beta(1-40) (92%) and A beta(1-42) (92%). However, A beta(1-42) binding 2 or 3 molar equivalents of Cu2+ (A beta1-42-2Cu2+, A beta1-42-3Cu2+) acted as a pro-oxidant to form HNE from PLPC-OOH. These findings suggest that, at moderate concentrations of copper, A beta acts primarily as an antioxidant to prevent Cu2+-catalyzed oxidation of biomolecules, but that, in the presence of excess copper, pro-oxidant complexes of A beta with Cu2+ are formed.     

Differing features of proteins in membranes may result in antioxidant or prooxidant action: opposite effects on lipid peroxidation of alcohol dehydrogenase and albumin in liposomal systems

SUMMARY

The influence of 3 thiol-containing compounds, bovine serum albumin (fatty acid free: BSA), glutathione (GSH) and yeast alcohol dehydrogenase (YADH) on lipid peroxidation in multilamellar liposomes, prepared from ox-brain phospholipid, was investigated. Thiol-compounds were added either before liposome formation, or after liposome formation; and their effects compared to a positive control. Bovine serum albumin (BSA), an acidic hydrophilic protein, displays a small, concentration dependent, antioxidant effect when added to preformed liposomes. A much larger antioxidant effect was observed when the BSA was entrapped inside the liposome, by adding BSA just prior to liposome preparation. In contrast, a Zn²⁺ containing redox enzyme, YADH, a basic hydrophobic membrane-associating protein, displays a large pro-oxidant effect at much lower concentrations especially when entrapped inside the liposome. This was observed also with GSH; but per mole of -SH, YADH was about 18 times as powerful a pro-oxidant perhaps because of structural changes to the membrane. Oxidized glutathione and N-acetylcysteine were also pro-oxidant (cysteine and cystine showed little effect). Formation of thiyl radicals may occur in the presence of iron ions with these pro-oxidant sulphur-containing compounds. Partial protection against lipid peroxidation was observed with EDTA, desferrioxamine and protoporphyrin (IX), potent iron-chelating agents.     

L-Homocysteine and L-homocystine stereospecifically induce endothelial nitric oxide synthase-dependent lipid peroxidation in endothelial cells

Atherothrombotic cardiovascular disease associated with hyperhomocysteinemia has been proposed to result, at least in part, from increased vascular oxidative stress. Here we characterize one mechanism by which homocyteine may induce a vascular cell type-specific oxidative stress. Our results show that L-homocysteine at micromolar levels stereospecifically increases lipid peroxidation in cultured endothelial cells, but not in vascular smooth muscle cells or when medium is incubated in the absence of cells. Consistent with these observations, homocysteine also increases the formation of intracellular reactive oxygen species. The pro-oxidant effect of homocysteine can be fully replicated by an equivalent concentration of homocystine (i.e., an oxidized form of homocysteine), but not with cysteine or glutathione. Homocyst(e)ine-dependent lipid peroxidation is independent of H(2)O(2) and alterations in glutathione peroxidase activity, but dependent on superoxide. Mechanistically, the pro-oxidant effect of homocysteine appears to involve endothelial nitric oxide synthase (eNOS), as it is blocked by the eNOS inhibitor L-N(G)-nitroarginine methyl ester. Thus, homocyst(e)ine actively promotes oxidative stress in endothelial cells via an eNOS-dependent mechanism.     

P67-phox-mediated NADPH oxidase assembly: imaging of cytochrome b558 liposomes by atomic force microscopy

NADPH oxidase activity depends on the assembly of the cytosolic activating factors, p67-phox, p47-phox, p40-phox, and Rac with cytochrome b(558). The transition from an inactive to an active oxidase complex induces the transfer of electrons from NADPH to oxygen through cytochrome b(558). The assembly of oxidase complex was studied in vitro after reconstitution in a heterologous cell-free assay by using true noncontact mode atomic force microscopy. Cytochrome b(558) was purified from neutrophils and Epstein-Barr virus-immortalized B lymphocytes and incorporated into liposomes. The effect of protein glycosylation on liposome size and oxidase activity was investigated. The liposomes containing the native hemoprotein purified from neutrophils had a diameter of 146 nm, whereas after deglycosylation, the diameter was reduced to 68 nm, although oxidase activity was similar in both cases. Native cytochrome b(558) was used after purification in reconstitution experiments to investigate the topography of NADPH oxidase once it was assembled. For the first time, atomic force microscopy illustrated conformational changes of cytochrome b(558) during the transition from the inactive to the active state of oxidase; height measurements allow the determination of a size of 4 nm for the assembled complex. In the processes that were studied, p67-phox displayed a critical function; it was shown to be involved in both assembly and activation of oxidase complex while p47-phox proceeded as a positive effector and increased the affinity of p67-phox with cytochrome b(558), and p40-phox stabilizes the resting state. The results suggest that although an oligomeric structure of oxidase machinery has not been demonstrated, allosteric regulation mechanisms may be proposed.     

Lipid peroxidation in mitochondrial inner membranes. I. An integrative kinetic model

An integrative mathematical model was developed to obtain an overall picture of lipid hydroperoxide metabolism in the mitochondrial inner membrane and surrounding matrix environment. The model explicitly considers an aqueous and a membrane phase, integrates a wide set of experimental data, and unsupported assumptions were minimized. The following biochemical processes were considered: the classic reactional scheme of lipid peroxidation; antioxidant and pro-oxidant effects of vitamin E; pro-oxidant effects of iron; action of phospholipase A₂, glutathione-dependent peroxidases, glutathione reductase and superoxide dismutase; production of superoxide radicals by the mitochondrial respiratory chain; oxidative damage to proteins and DNA. Steady-state fluxes and concentrations as well as half-lives and mean displacements for the main metabolites were calculated. A picture of lipid hydroperoxide physiological metabolism in mitochondria in vivo showing the main pathways is presented. The main results are:(a) perhydroxyl radical is the main initiation agent of lipid peroxidation (with a flux of 10⁻⁷Ms⁻¹); (b) vitamin E efficiently inhibits lipid peroxidation keeping the amplification (kinetic chain length) of lipid peroxidation at low values (10); (c) only a very minor fraction of lipid hydroperoxides escapes reduction via glutathione-dependent peroxidases; (d) oxidized glutathione is produced mainly from the reduction of hydrogen peroxide and not from the reduction of lipid hydroperoxides.     

Antioxidant and pro-oxidant effects of lycopene in comparison with beta-carotene on oxidant-induced damage in Hs68 cells

Lycopene has become a focal point in recent research following clinical trials that suggest that beta-carotene may promote lung cancer in smokers. Because lycopene only differs from beta-carotene in lacking the beta-ionone structure, and beta-carotene is known to have pro-oxidant activity in vitro, we sought to determine whether lycopene might also have pro-oxidant activity in vitro in comparison with beta-carotene. Human foreskin fibroblasts (Hs68 cells) were first enriched with 10 and 20 μM lycopene or beta-carotene for 1 hr followed by incubation with various oxidants. Lipid peroxidation was measured as thiobarbituric acid-reactive substances (TBARS) released into the medium and DNA damage was measured as formation of comet and 8-hydroxy-2'-deoxyguanosine. The results showed that lycopene at 20 μM significantly decreased levels of TBARS induced by ferric nitrilotriacetate (Fe/NTA) but enhanced levels of TBARS induced by a lipid-soluble radical generator (2,2'-azobis[2,4-dimethylvaleronitrile]; AMVN). Both the antioxidant and pro-oxidant effects of lycopene tended to be dose-dependent. beta-Carotene at 20 μM did not significantly decrease TBARS induced by Fe/NTA but significantly increased TBARS induced by AMVN. Lipid peroxidation induced by a water-soluble radical generator 2,2'-azobis(2-amidinopropane)dihydrochloride was not significantly affected by either lycopene or beta-carotene. Neither lycopene nor beta-carotene affected DNA damage or changes in cell morphology induced by any of the three oxidants tested. The present study in Hs68 cells demonstrates that lycopene can be either an antioxidant or a pro-oxidant depending on the oxidants used, and that lycopene and beta-carotene behave similarly under the in vitro oxidative conditions. Although it is unclear whether lycopene may have pro-oxidant activity in vivo, our results caution that it may be premature to undertake clinical trials with lycopene.     

Disorders of equilibrium between pro- and antioxidant metalloprotein action in blood of albino rats after gangliosympathectomy and its combination with acute acoustic stress]

Unilateral ganglionsympathetectomy-removal of right upper cervical sympathetic ganglia and its combination with acute acoustic stress results in intensification of process of free radical oxidation of lipids and balance disturbance between components of antioxidative and pro-oxidative system of blood. Changes of composition and activity Cu, Zn-superoxide dismutase, catalase activity and contents of catalase, ceruloplasmin, transferrin and metalloproteins pro-oxidative action such as (cytochromes b558(I), b558(II), b558(III), b558(IV)) as well as O2-producing lipoprotein suprol and cytochromes b5 are evidence of compensating-adoptive function disturbances with according changes in blood, relatively to adaptive-trophic function of sympathetic system, its regulative role, in system of tissue metabolic conversion, which is directed, in particular, on compensation of anti-radical protection of cells.     

Radiation protection by the ocimum flavonoids orientin and vicenin: mechanisms of action

In previous studies, flavonoids, orientin and vicenin, that were isolated from the leaf extract of Ocimum sanctum, were found to protect mice against radiation injury. Several flavonoids are known to be good antioxidants. Therefore, the effect of orientin and vicenin on radiation-induced lipid peroxidation in vivo and their antioxidant activity in vitro were studied. Adult mice were injected intraperitoneally with 50 microgram/kg of orientin or vicenin and exposed whole-body to 3 Gy of gamma radiation. Lipid peroxidation was measured in the liver 15 min to 8 h postirradiation. The antioxidant activity of orientin/vicenin (10-500 microM) was studied by measuring inhibition of hydroxyl radicals generated by the Fenton reaction (Fe(3+)-EDTA-ascorbic acid-H(2)O(2)) in vitro. The compounds were also tested for possible pro-oxidant and iron chelation activities at the above concentrations in the in vitro system. Orientin and vicenin provided almost equal protection against radiation-induced lipid peroxidation in mouse liver. Both compounds showed a significantly greater free radical-inhibiting activity in vitro than DMSO. Neither orientin nor vicenin showed any pro-oxidant activity at the concentrations tested. Both compounds inhibited free radical formation in the absence of EDTA. Free radical scavenging appears to be a likely mechanism of radiation protection by these flavonoids.     

The intensity of lipid peroxidation and enzymatic antioxidative activity in potato leaves under action of drought and polystimulin K

The influence long-term soil drought and potato plants treatment by synthetic analog of cytokinin--polystimulin K on intensity of lipid peroxidation processes and enzymatic antioxidative activity have been investigated. It has been found, that the drought induced the shift of prooxidative-antioxidative balance in respect of lipid peroxidation activation in the potato leaves. It was accompanied by the increase of the ethylene output, membrane permeability, as well as decrease of the lipids content and increase in the enzymatic antioxidative activity (catalase and peroxidase). It is shown, that the intensity of peroxidation processes was higher in budding phases, while enzymatic antioxidative activity was higher in flowering phases in potato plants. Plant exogenous treatment by polystimulin K induced both the decrease in peroxidate oxidation processes, stabilization of catalase and peroxidase activity, as well as the increase in potato resistance to drought.     

NADPH-dependent lipid peroxidation capacity in unfixed tissue sections: characterization of the pro-oxidizing conditions and optimization of the histochemical detection

Summary Factors which influence the iron-stimulated lipid peroxidation in rat liver have been studied by incubating unfixed cryostat sections with a pro-oxidant system and using an optimized histochemical detection method for lipid peroxidation products with 3-hydroxy-2-naphthoic acid hydrazide and Fast Blue B. We used a method that was slightly different from the one described previously. The final reaction product was exclusively localized in the cytoplasm of liver parenchymal cells with a homogeneous distribution within the liver lobule. The absorbance maximum, as measured cytophotometrically, was found to be 550 nm. Maximum lipid peroxidation was observed when the pro-oxidant system contained 0.2 mm NADPH, 1 mm ADP and 15 μm FeCl₂. Some reaction product was found when NADPH was omitted. Iron concentrations higher than 180 μm prevented the formation of lipid peroxidation products in certain areas of the sections, whereas ADP concentrations higher than 1 mm inhibited the reaction in the whole section. A pH dependency was also observed, with the highest lipid peroxidation at pH 7.2. Optimum lipid peroxidation was induced by incubating for 30 min at 37°C with the pro-oxidant system. A linear relationship was found between the thickness of the sections (up to 20 μm) and the amount of lipid peroxidation products. The addition of scavengers of O2- (superoxide dismutase), hydrogen peroxide (catalase) and OH · (mannitol) to the first step medium did not affect the amount of final reaction product. These findings appear to confirm the hypothesis proposed for events occurring in isolated microsomes, leading to the formation of hydroperoxides and ultimately lipid peroxidation-derived carbonyls. The present method is a useful tool for studying the capacity of lipid peroxidation in tissues under different (patho)physiological conditions.     

Copper supplementation in humans does not affect the susceptibility of low density lipoprotein to in vitro induced oxidation (FOODCUE project)

The oxidative modification of low-density lipoprotein cholesterol (LDL) has been implicated in the pathogenesis of atherosclerosis. Copper (Cu) is essential for antioxidant enzymes in vivo and animal studies show that Cu deficiency is accompanied by increased atherogenesis and LDL susceptibility to oxidation. Nevertheless, Cu has been proposed as a pro-oxidant in vivo and is routinely used to induce lipid peroxidation in vitro. Given the dual role of Cu as an in vivo antioxidant and an in vitro pro-oxidant, a multicenter European study (FOODCUE) was instigated to provide data on the biological effects of increased dietary Cu. Four centers, Northern Ireland (coordinator), England, Denmark, and France, using different experimental protocols, examined the effect of Cu supplementation (3 or 6 mg/d) on top of normal Cu dietary intakes or Cu-controlled diets (0.7/1.6/6.0 mg/d), on Cu-mediated and peroxynitrite-initiated LDL oxidation in apparently healthy volunteers. Each center coordinated its own supplementation regimen and all samples were subsequently transported to Northern Ireland where lipid peroxidation analysis was completed. The results from all centers showed that dietary Cu supplementation had no effect on Cu- or peroxynitrite-induced LDL susceptibility to oxidation. These data show that high intakes (up to 6 mg Cu) for extended periods do not promote LDL susceptibility to in vitro-induced oxidation.     

Dose-dependent modulation of systemic lipid peroxidation and activity of anti-oxidant enzymes by vitamin E in the rat

The objective of this work was to examine the time-dependent pro-oxidant versus antioxidant effect of various doses of vitamin E used commonly in experimental studies. Erythrocyte activity of superoxide dismutase (SOD), glutathione peroxidase (GPX), catalase (CAT) and plasma lipid peroxidation levels were investigated following biweekly intramuscular administration of 100, 300 and 600 mg/kg of vitamin E at a baseline time point, and additionally at 2, 4 and 6 weeks after initiating treatment. Vitamin E had an antioxidant effect when administered at low doses over short time periods, and increased the activity of antioxidant enzymes. At higher doses and over longer time periods, it increased the level of lipid peroxidation, and attenuated the activity of antioxidant enzymes. These results suggest that time-dependent variations in vitamin E effects should be considered in design and interpretation of experimental antioxidant studies, as well as during clinical trials.     

Antioxidative activity of natural products from plants

A variety of flavonoids, lignans, an alkaloid, a bisbenzyl, coumarins and terpenes isolated from Chinese herbs was tested for antioxidant activity as reflected in the ability to inhibit lipid peroxidation in rat brain and kidney homogenates and rat erythrocyte hemolysis. The pro-oxidant activities of the aforementioned compounds were assessed by their effects on bleomycin-induced DNA damage. The flavonoids baicalin and luteolin-7-glucuronide-6'-methyl ester, the lignan 4'-demethyldeoxypodophyllotoxin, the alkaloid tetrahydropalmatine, the bisbenzyl erianin and the coumarin xanthotoxol exhibited potent antioxidative activity in both lipid peroxidation and hemolysis assays. The flavonoid rutin and the terpene tanshinone I manifested potent antioxidative activity in the lipid peroxidation assay but no inhibitory activity in the hemolysis assay. The lignan deoxypodophyllotoxin, the flavonoid naringin and the coumarins columbianetin, bergapten and angelicin slightly inhibited lipid peroxidation in brain and kidney homogenates. It is worth stressing that the compounds with antioxidant effects in this assay, with the exception of tetrahydropalmatin and tanshinone I, have at least one free aromatic hydroxyl group in structure. Obviously, the aromatic hydroxyl group is very important for antioxidative effects of the compounds. None of the compounds tested exerted an obvious pro-oxidant effect.     

Effect of various thiamine supplies of the body on the enzyme activity in the metabolism of xenobiotics and lipid peroxidation in rat liver microsomes

The effect of different thiamine supply of the organism on the enzymic activity in metabolism of xenobiotics and the processes of the lipid peroxidation in the liver microsomes with the application of phenobarbital, an inductor of microsomes' enzymes are studied in experiments on Wistar albino male rats. It is established that deficit of vitamin B1 increases the activity in most of processes studied in microsomes and also the intensity of lipids' peroxidation. Phenobarbital enhances the activity of microsomal oxidation irrespectively of vitamin B1 supply, whereas peroxidation of lipids is activated by phenobarbital only in animals fed on physiological doses of vitamin B1. The N-demethylation rate of dimethylaniline in experiments in vitro is inhibited by high doses of thiamine (150 microM), its derivatives inhibited this process in low concentrations (15 microM) as well.