Kinetics of lipid peroxidation in compartmentalized systems initiated by a water-soluble free radical source

Kinetic rate laws arising from theoretical expectations for the oxidation of lipids initiated by water-soluble free radicals in compartmentalized systems under different experimental conditions are deduced. In particular, the predictions for the kinetic reaction orders in: (a) intra-particle oxidizable compound concentration (at fixed number of particles and particle size), alpha; (b) number of particles or analytical lipid concentration (at fixed intra-particle concentration and particle size), beta and (c) initiator, gamma, are obtained. The reaction orders beta and gamma are determined by the fraction of initiator derived radicals captured by the particles (f) and the mean number of chain carrying radicals per particle () when the system reaches the steady state condition. Predicted orders in initiator range from 0 ( = 0.5) to 0.5 (f-->1; > > 1), while the order in number of particles ranges between 0.5 (f-->1; > > 1) and 1. These predictions are tested by measuring the kinetic law for the oxidation of SUV's egg yolk phosphatidylcholine vesicles initiated by the thermal decomposition of ABAP. The results indicate that, under the conditions employed, beta = 0.68 +/- 0.05 and gamma = 0.46 +/- 0.04. These values are close to those expected for a system in which > > 1 and the efficiency of capture is relatively high. This last condition is confirmed by estimating the efficiency of capture from a comparison of induction times elicited by similar concentrations of Trolox and alpha-tocopherol.     

The role of iron in ferritin- and haemosiderin-mediated lipid peroxidation in liposomes.

Ferritin and haemosiderin were shown, by the measurement of malondialdehyde production and loss of polyunsaturated fatty acids, to stimulate lipid peroxidation in liposomes. At pH 7.4 ascorbate was additionally required to achieve peroxidation; however, peroxidation occurred at pH 4.5 in the presence of iron-proteins alone. The damage was completely inhibited by the incorporation of chain-breaking antioxidants (alpha-tocopherol and butylated hydroxytoluene) into the liposomes. Metal chelators (desferrioxamine and EDTA) also completely inhibited lipid peroxidation. These and further results indicate that, at pH 4.5, even in the absence of a reducing agent, iron is released from haemosiderin and can mediate oxidative damage to a lipid membrane.     

Administration of glutathione and lipid peroxidation induced during fasting

It is well known that lipid peroxidation may be initiated or exaggerated by conditions leading to hepatic GSH depletion or altered GSH/GSSG ratio. In our study we evaluated the effects of GSH administration on hepatic, bile and plasma GSH, GSSG and MDA in rats depleted of the tripeptide by a prolonged. fasting. An exteriorized biliary-duodenal fistula was established and GSH or saline solution was administered i.p. for a period of 6h. Rats treated with GSH exhibited an increased GSH and decreased GSSG biliary excretion. Whereas in control rats an opposite pattern was observed, namely enhanced GSSG and decreased GSH biliary excretion. While hepatic GSH and GSSG concentrations were comparable in the two groups, a significant increase in liver and plasma MDA production was found in controls compared to GSH treated rats. Our data suggest a protective role of GSH against the production of lipoperoxidation as evidenced by the decrease of hepatic, biliary and plasma MDA levels and by a decreased percentage of biliary GSSG. In addition, the significant increase of biliary GSH excretion, observed in rats treated with GSH compared to controls, may be due to an increased supply of the tripeptide which is known to be preferentially excreted into bile in the reduced form.     

Aging and lung antioxidant enzymes, glutathione, and lipid peroxidation in the rat.

The five major antioxidants enzymes, cytochrome oxidase (COX), GSH, and GSSG, and endogenous and in vitro stimulated lipid peroxidation (TBA-RS) were assayed in the lung of old (28 months) and young (9 months) adult rats due to the almost total absence of data of this kind in this tissue, which is normally exposed to relatively high pO2 throughout life. Catalase, selenium (Se)-dependent GSH peroxidase (GPx), GSH reductase, GSH, GSSG, GSSG/GSH, and in vivo and in vitro TBA-RS showed similar values in old and young animals. The decrease observed for non Se-dependent GPx disappeared when the values were expressed in relation to COX activity. Only superoxide dismutase showed a clear decrease when referred both to protein and COX activity. These results suggest that lung aging is not accelerated in old age due to a decrease in the antioxidant capacity of the tissue. Nevertheless, they are compatible with a continuous damage of the lung tissue by free radicals throughout the life span.     

Fe and Fe initiated peroxidation of sonicated and non-sonicated liposomes made of retinal lipids in different aqueous media

Retina is highly susceptible to oxidative damage due to its high content of polyunsaturated fatty acids (PUFAs), mainly docosahexaenoic acid (22:6 n3). Lipid peroxidation process is thought to be involved in many physiological and pathological events. Many model membranes can be used to learn more about issues that cannot be studied in biological membranes. Sonicated liposomes (SL) and non-sonicated liposomes (NSL) prepared with lipids isolated from bovine retina and characterized by dynamic light-scattering, were submitted to lipid peroxidation, under air atmosphere at 22 °C, with Fe²⁺ or Fe³⁺ as initiator, in different aqueous media. Conjugated dienes and trienes, determined by absorption at 234 and 270 nm respectively, and thiobarbituric acid-reactive substances were measured as a function of time. Peroxidation of SL or NSL initiated with 25 μM FeSO₄ in 20 mM Tris-HCl pH 7.4 resulted in an increase in TBARS production after a lag phase of 60 min. Incubation of both types of liposomes in water resulted in shortening of the lag phase at 30 min. When lipid peroxidation was performed in 0.15 M NaCl, lag phase completely disappeared. On the other hand, FeCl₃ (25 μM) induced a limited production of TBARS only just after 30 min of incubation. When Fe²⁺- or Fe³⁺-lipid peroxidation of both types of liposomes was carried out in water or 0.15 M NaCl, formation of conjugated dienes and conjugated trienes were higher than in reactions carried out in 20 mM Tris-HCl pH 7.4.Our results established that both liposome types were susceptible to Fe²⁺- and Fe³⁺-initiated lipid peroxidation. However, Fe²⁺ showed a clearly enhanced effect on peroxidation rate and steady state concentration of oxidation products.We verified that peroxidation of liposomes made of retinal lipids is affected not only by type of initiator but also by aqueous media. This model constitutes a useful system to study formation of lipid peroxidation intermediaries and products in an aqueous environment.     

The role of selenium in the regulation of lipid peroxidation in biological membranes and glutathione peroxidase activity

The antioxidative effect of selenium cannot be exclusively due to the functioning of the selenium-dependent glutathione peroxidase mechanism of utilization of various hydroperoxides. This hypothesis is based on the following experimental evidence. Selenium ions are readily incorporated into animal organs and tissues immediately after injection (2 hours) as well as into cell organelles and cytosol where they inhibit lipid peroxidation. The activity of glutathione peroxidase (EC 1.1.1.19) in rat liver and guinea pig cytosol is thereby unchanged but increases drastically after 12 hours reaching a maximum an the 3rd-4th day. The effectiveness of lipid peroxidation inhibition does not increase under these conditions. Although the glutathione peroxidase activity is absent in the nuclei and microsomes, exogenous selenium inhibits lipid peroxidation in these organelles. The activity of the rat liver cytosolic enzyme markedly exceeds that of its guinea pig counterpart. However, lipid peroxidation in guinea pig liver occurs less intensively than that in rat liver cytosol.     

Perhydroxyl radical (HOO.) initiated lipid peroxidation. The role of fatty acid hydroperoxides

It is demonstrated that the perhydroxyl radical (HOO., the conjugate acid of superoxide (O2-], initiates fatty acid peroxidation (a model for biological lipid peroxidation) by two parallel pathways: fatty acid hydroperoxide (LOOH)-independent and LOOH-dependent. Previous workers (Gebicki, J. M., and Bielski, B. H. J. (1981) J. Am. Chem. Soc. 103, 7020-7025) demonstrated that HOO., generated by pulse radiolysis, initiates peroxidation in ethanol/water fatty acid dispersions by abstraction of the bis-allylic hydrogen atom from a polyunsaturated fatty acid. Addition of O2 to the fatty acid radicals forms peroxyl radicals (LOO.s), the chain-propagating species of lipid peroxidation. In this work it is demonstrated that HOO., generated either chemically (KO2) or enzymatically (xanthine oxidase), is a good initiator of fatty acid peroxidation in linoleic acid ethanol/water dispersions; O2- serves only as the source of HOO., and HOO. initiation can be observed at physiologically relevant pH values. In contrast to the previous results, the initiating effectiveness of HOO. is related directly to the initial concentrations of LOOHs in the lipids to be peroxidized. This defines a LOOH-dependent mechanism for fatty acid peroxidation initiation by HOO., which parallels the previously established LOOH-independent pathway. Since the LOOH-dependent pathway is much more facile than the LOOH-independent pathway, LOOH is the kinetically preferred site of HOO. attack in these systems. Experiments comparing HOO./LOOH-dependent fatty acid peroxidation with transition metal- and peroxyl radical-initiated peroxidation rule out the participation of the latter two species as initiators, which defines the HOO./LOOH initiation system as mechanistically unique. LOOH product studies are consistent with either a direct or indirect hydrogen atom transfer between LOOH and HOO. to yield LOO.s, which propagate peroxidation. The LOOH-dependent pathway of HOO.-initiated fatty acid peroxidation may be relevant to mechanisms of lipid peroxidation initiation in vivo.     

The effects of ochratoxin A on lipid peroxidation and antioxidant enzymes: a protective role of melatonin

Various studies indicate that the mycotoxin ochratoxin A (OTA) is a carcinogenic, genotoxic, teratogenic, immunotoxic, and nephrotoxic agent. In the present study, the activities of some enzymes in the serum and liver of rats with ochratoxicosis and the effects of melatonin on these enzymes were investigated. Rats were divided into three equal groups, each consisting of eight rats; control, OTA (289 microg/kg per day) and OTA + melatonin groups for this study. In the OTA treated group, the level of lipid peroxidation (LPO) and the activities of glutathione peroxidase (GSH-Px) were increased in the liver and serum in comparison with the control group. The activities of catalase (CAT) and superoxide dismutase (SOD) were significantly changed in the serum when compared with controls. Our results showed structural tissue damage in the liver in OTA-treated rats. Melatonin decreased the OTA-induced damage to support the antioxidant defense system and/or with free radical scavenger action.     

The response of rat liver lipid peroxidation, antioxidant enzyme activities and glutathione concentration to the thyroid hormone.

1. In liver microsomes from hyperthyroid rats NADPH-dependent lipid peroxidation induces a hydroperoxide formation 56% higher than that in euthyroid ones. 2. The addition of 5 microM Fe2+ (or Fe3+) strongly decreases the hydroperoxide level in favour of that of TBA-reactive substances. Higher iron concentrations (30 microM) have no significant effect. 3. In hepatocytes from hyperthyroid rats CCl4-induced lipid peroxidation produces an amount of TBA-reactive substances four times higher than that in those from euthyroid rats. 4. In the liver of hyperthyroid rats a GSH concentration decrease (by about 35%) is found while the opposite occurs in the blood of the same animals where GSH increases 2.5 times. 5. It is shown that in the liver of hyperthyroid rats, besides higher lipid peroxidation, a more active defense mechanism is operating since both glutathione peroxidase and glutathione reductase specific activities are higher than in euthyroid rats.     

Protection of the monoamine oxidase in brain synaptosomes by fat- and water-soluble antioxidants during lipid peroxidation

The protective effects of alpha-tocopherol, carnosine and their mixtures on monoamine oxidase activity, accumulation of lipid peroxidation products, lipid fatty acid composition, hydrophobicity and microviscosity of synaptic membranes during lipid peroxidation were studied. It was shown that the protective efficiency is more higher when the mixture of water and liposoluble antioxidants was used.     

Effect of lysophosphatidylcholine on the radiation-induced lipid peroxidation in liposomes

The effect of lysophosphatidylcholine (LPC) on the lipid peroxidation process (LPO) in liposomes of rat liver phosphatidylcholine initiated by irradiation of 137Cs source was studied. The formation of diene conjugates (DC) is shown to increase dramatically on incorporation of more than 10% LPC into liposomes. The dependence of DC proportion on the irradiation dose is practically linear in the range of 0 to 5 kGy. The DC concentration in the liposomes without LPC grows at least to dose of 3.3 kGy and is unchanged on further irradiation. The malonic dialdehide accumulation follows the similar dependence. The LPC effect is neutralized by the incorporation of cholesterol into liposomes. The product of free radical fragmentation of LPC, palmitoxyacetone, practically has no influence on the DC concentration. The reasons of LPC effect on the irradiation initiated LPO in liposomes is discussed.     

Effect of splenosid on lipid peroxidation process and glutathione antioxidant system in rats exposed to fractionated radiation

The dynamics of lipid peroxidation, antioxidant glutathione system condition in blood and viscerals (brain, heart, liver, spleen) of rats which were fractionally irradiated (10 fractions) in the total dose 1.0 Gy and oxidative homeostasis increase of primary and secondary lipid peroxidation products and glutathione system disturbances were established in the irradiated rats. The administration of splenosid diminished the disturbances of oxidative homeostasis but does not completely normalize the latter. The administration of splenosid during the irradiation course and after its finishing is more effective than only during the irradiation course.     

Alterations in lipid peroxidation and antioxidant status in pregnancy with preeclampsia

Soy consumption is associated with a lower risk of atherosclerotic disease in the oriental population. Genistein is a soy isoflavone bearing estrogenic properties. Previous experiments in our laboratory demonstrated the potentiation of endothelium-independent relaxation of coronary artery by both estrogen and genistein. The potentiating effects of both estrogen and genistein were mediated through the cAMP-signaling pathway. We hypothesize that genistein could enhance protein kinase A (PKA) activity in porcine coronary artery smooth muscle, thereby offering a mechanism for the potentiation of vascular relaxation by genistein. In our study, a high concentration of genistein (10^−4.5 M) significantly increased PKA activity in porcine coronary artery rings. While genistein at 10^−5.5 M and forskolin at 10⁻⁷ M had no effect on PKA activity, the combination of the two compounds at the prescribed concentrations caused a significant increase in PKA activity. The increase in PKA activity by genistein was abolished by SQ 22536 (adenylate cyclase blocker), but not by NF 449 (Gs protein blocker) or ICI 182780 (estrogen receptor antagonist). Our results suggest that the action of genistein is mediated via adenylate cyclase, but does not appear to involve Gs protein or ICI 182780-sensitive estrogen receptor.     

Protective effect of glutathione and selenium against alloxan induced lipid peroxidation and loss of antioxidant enzymes in erythrocytes

Alloxan is a diabetogenic drug and is known to induce diabetes through generation of free radicals. The toxic oxygen species can be detoxified by antioxidant enzyme system and thus reduce the deleterious effect of lipid peroxidation. Erythrocytes exposed to alloxan induced lipid peroxidationin vivo as well asin vitro. Although alloxan treatment produced a deleterious effect on antioxidant enzymes, pretreatment with glutathione and selenium led to a recovery of the activities of superoxide dismutase and glutathione peroxidase. However, catalase activity increased on alloxan treatment. Alloxan reduced blood glucose level significantly within 60 min but thereafter a slow and steady rise was observed.     

The role of lipid peroxidation in liver damage

The consequences of the peroxidative breakdown of membrane lipids have been considered in relation to both the subcellular and tissue aspects of liver injury. Mitochondrial functions can be impaired by lipid peroxidation probably through the oxidation of pyridine nucleotides and the consequent alteration in the uptake of calcium. Several enzymatic functions of the endoplasmic reticulum are also affected as a consequence of peroxidative events and among these are the activities of glucose 6-phosphatase, cytochrome P-450 and the calcium sequestration capacity. Moreover, a release of hydrolytic enzymes from lysosomes and a decrease in the fluidity of plasma membranes can contribute to the liver damage consequent to the stimulation of lipid peroxidation. Extensive studies carried out in vivo and integrated with the use of isolated hepatocytes have shown that lipid peroxidation impairs lipoprotein secretion mainly at the level of the dismission from the Golgi apparatus, rather than during their assembly. However, such an alteration appears to give a late and not essential contribution to the fat accumulation. A more critical role is played by peroxidative reactions in the pathogenesis of acute liver necrosis induced by several pro-oxidant compounds as indicated by the protective effects against hepatocyte damage exerted by antioxidants. In addition, even in the cases where lipid peroxidation has been shown not to be essential in causing cell death there is evidence that it can still act synergistically with other damaging mechanisms in the amplification of liver injury.     

The effect of zinc on iron-induced lipid peroxidation in different lipid systems including liposomes and micelles

The effect of zinc on FeSO4/ascorbic acid-induced lipid peroxidation was measured by the thiobarbituric acid assay in various lipid systems including small unilamellar liposomes prepared from egg phosphatidylcholine (EPC), ionic micelles prepared from arachidonic acid (C20:4), non-ionic monocomponent micelles prepared from EPC-derived, methylated fatty acids, and an eicosatetrene emulsion. With the exception of C20:4 micelles, zinc inhibited lipid peroxidation in each of the above systems in a similar dose-related fashion, with 0.5 mM zinc having maximal effect. Gas-chromatographic fatty acid analysis too indicated a protective effect of zinc against FeCl3-induced lipid peroxidation in soybean PC vesicles, which do not contain C20:4 moieties. These findings, in particular the inhibition of lipid peroxidation in eicosatetrene emulsion, suggest that the presence of uncharged polar head groups, or packing of lipid molecules into ordered self-assemblages (membranes and micelles) have no critical influence on the antioxidant effect of zinc. The results with Fe2+ are compatible with the concept that zinc interferes with the formation of Fe2+-oxygen-enoic complexes. This mechanism, however, cannot account for the inhibition by zinc of the Fe#+-induced lipid peroxidation, suggesting the involvement of other types of zinc effects in these systems.     

Plasma lipid peroxidation and antioxidant levels in patients with rheumatoid arthritis

In recent years, a great number of studies have investigated the possible role of reactive oxygen species in the aetiology and pathogenesis of rheumatoid arthritis (RA). The aim of this study was to investigate plasma concentrations of vitamin E, beta-carotene, activities of glutathione peroxidase (GSH-Px) and catalase, levels of lipid peroxidation (MDA) and reduced glutathione (GSH) in 36 patients with rheumatoid arthritis (RA) and 22 healthy age-matched controls. The plasma activity of GSH-Px and catalase (p < 0.001), levels of GSH (p < 0.01), concentration of beta-carotene (p < 0.05) and vitamin E (p < 0.001), haemoglobin and hematocrit (p < 0.05) were significantly lower in patients with RA than in controls. The MDA levels (p < 0.01), C reactive protein, rheumatoid factor, anti-streptolysin-o values (p < 0.001), platelet count (p < 0.05) and erythrocyte sedimentation rate (p < 0.001) were higher in the patient group than in the control group. These results provide some evidence for a potential role of increased lipid peroxidation and decreased enzymic and non-enzymic antioxidants in RA by its inflammatory character. These results suggested that oxidant stress plays a very important role in the pathogenesis of RA.     

The role of iron in the initiation of lipid peroxidation

Iron is required for the initiation of lipid peroxidation. Evidence is presented that lipid peroxidation requires both Fe3+ and Fe2+, perhaps with oxygen to form a Fe3+-dioxygen-Fe2+ complex. Other mechanisms of initiation, mostly involving the iron-catalyzed formation of hydroxyl radical, are described and discussed from both theoretical and experimental view points.     

The role of lipid peroxidation in the mechanism of stress

A concept on the mechanism of stress response is substantiated proceeding from the data available in literature and obtained from the author's research made on the radiation stress model. The conception envisages that products of lipid peroxidation (LPO) appear as primary (under direct effect of a stress factor on tissues) and secondary (as a consequence of high- and long-term catecholamine++) mediators. Mobilization of stress-realizing systems in that process is regarded as an adequate response of the auto-oxidative++ system to the primary activation of LPO. Transformation of catecholamines into the factor of LPO stimulation (secondary) is a result of an increase in the relative role of the quinoid way to transform catecholamines in the case of their high concentration. Radical intermediates of the quinoid metabolism appear as LPO initiators. An important pathogenetic role of LPO activation in the stress mechanism substantiates expedience to use antioxidants as agents for prophylaxis and early treatment of stress-factor injuries.