Stages Of Peroxidation Research Articles

Cemtirestat dimerization in liposomes and erythrocytes exposed to peroxyl radicals was reverted by thiol-disulfide exchange with GSH

In the model system of DOPC (1,2-dioleoyl-sn-glycero-3-phosphocholine) liposomes exposed to peroxyl radicals generated by the azoinitiator AAPH, cemtirestat (CMTI-SH) inhibited lipid peroxidation more efficiently than the natural antioxidant glutathione. In the concentrations 100 to 500 µM, both CMTI-SH and GSH induced distinct lag phases in the initial stages of lipid peroxidation yet GSH produced consistently shorter induction periods (about twice) than equimolar CMTI-SH. Moreover, concentration dependence of lipid peroxidation inhibition measured at the 80th minute, revealed about three times higher IC50 value for GSH compared to CMTI-SH. When the incubations prolonged till 180 min no further absorbance changes at 270 and 302 nm, respectively, occurred. After addition of the reducing agent tris(2-carboxyethyl)phosphine, the absorbance peak at 270 nm shifted back to 302 nm. These findings pointed to the presence of reducible CMTI-SH disulfide whose definite structure was confirmed by proving identity of TLC retention and spectral data with those of the synthesized CMTI disulfide. When CMTI-SH and GSH were present simultaneously in the liposomal incubations, the mixing effect on the induction period was synergistic rather than additive. This was explained by ability of GSH to reduce CMTI disulfide which was proved in separate experiments with an authentic CMTI disulfide prepared synthetically. This finding was also demonstrated by experiment with CMTI-disulfide to protect the erythrocytes against oxidative damage induced by peroxyl radicals. To conclude, CMTI-SH scavenges reactive oxygen species yielding CMTI disulfide while GSH maintains CMTI-SH in the reduced state. This finding was also demonstrated by experiment with CMTI-disulfide to protect the erythrocytes against oxidative damage induced by peroxyl radicals. CMTI-SH would thus represent the first line of the cellular defense against peroxyl radical mediated oxidative stress.

Unearthing of the Antidiabetic Potential of Aqueous Extract of Solanum betaceum Cav. Leaves.

Solanum betaceum Cav., commonly known as tamarillo or Brazilian tomato, belongs to the Solanaceae family. Its fruit is used in traditional medicine and food crops due to its health benefits. Despite the numerous studies involving the fruit, there is no scientific knowledge about the tamarillo tree leaves. In this work, the phenolic profile of aqueous extract obtained from S. betaceum leaves was unveiled for the first time. Five hydroxycinnamic phenolic acids were identified and quantified, including 3-O-caffeoylquinic acid, 4-O-caffeoylquinic acid, chlorogenic acid, caffeic acid and rosmarinic acid. While the extract displayed no effect on α-amylase, the extract inhibited the activity of α-glucosidase (IC50 = 1617 mg/mL), and it was particularly effective for human aldose reductase (IC50 = 0.236 mg/mL): a key enzyme in glucose metabolism. Moreover, the extract exhibited interesting antioxidant properties, such as a potent capacity to intercept the in vitro-generated reactive species O2•- (IC50 = 0.119 mg/mL) and •NO (IC50 = 0.299 mg/mL), as well as to inhibit the first stages of lipid peroxidation (IC50 = 0.080 mg/mL). This study highlights the biological potential of S. betaceum leaves. The scarcity of research on this natural resource underscores the need for additional studies in order to fully explore its antidiabetic properties and to promote the value of a species currently at risk of extinction.

Lentinula edodes, a Novel Source of Polysaccharides with Antioxidant Power.

The fruiting bodies of edible mushrooms represent an important source of biologically active polysaccharides. In this study, Lentinula edodes crude polysaccharides (LECP) were extracted in hot water, and their antioxidant and antiradical activities were investigated. The antioxidant activity of LECP was investigated against reactive species such as 1,1’-diphenyl-2-picrylhydrazyl, 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid, hydroxyl and superoxide anion radicals, reducing power with EC50 values of 0.51, 0.52, 2.19, 3.59 and 1.73 mg/mL, respectively. Likewise, LECP inhibited the lipid peroxidation induced in methyl linoleate through the formation of conjugated diene hydroperoxide and malondialdehyde. The main sugar composition of LECP includes mannose, galactose, glucose, fucose and glucuronic acid. Characterization by Fourier transform infrared spectroscopy and nuclear magnetic resonance determined that LECP was made up of α and β glycosidic bonds with a backbone of α-D-Glc, →6)-β-D-Glcp-(1→, →6)-α-D-Galp-(1→ and β-D-Manp-(1→ residues. The results showed that LECP can scavenge all reactive species tested in a concentration-dependent manner and with a protective effect in the initial and final stages of lipid peroxidation. The natural antioxidant activity of the LECP that was investigated strengthens the high medicinal and nutritional value of this mushroom.

Influence of oxidative, carbonyl, and nitrosative stresses on the course of chronic kidney disease (analytical review)

Oxidative stress is a phenomenon caused by an imbalance of peroxide homeostasis. The concept of oxidative stress was introduced in 1985 and has a clear connection with redox chemistry. The article provides a detailed description of molecular redox switches that control the response to oxidative stress. Four levels of protection of the cell by enzymatic antioxidants from the action of reactive oxygen species are described. It has been shown that antioxidant enzymes play a major role in antioxidant protection rather than low molecular weight antioxidant compounds. Four stages of lipid peroxidation are considered. Oxidative stress plays an important role in the pathogenesis of chronic kidney disease. Uraemia increases oxidative stress. Mitochondrial dysfunction is the most important cause of oxidative stress in acute and chronic kidney disease. The effect of molecular stress on the kidneys and the course of chronic kidney disease is described. Some information is presented in the form of tables for the readers’ convenience.

Deletion polymorphism of GSTT1 gene and lipid peroxidation hyperactivation in menopausal insomnia

A comparative analysis of the parameters of lipid peroxidation - antioxidant defense system in menopausal women with- and without insomnia, depending on the GSTT1 polymorphism. The study included 181 menopausal Caucasian women (ethnic group - Russians), who according to the results of the clinical-anamnestic examinations were divided into insomnia group (n=120, age 53.68±4.61 years, body mass index 26.47±3.73 kg/m2) and control group (n=61, age 52.64±4.91 years, body mass index 27.69±3.11 kg/m2). The insomnia group was formed according to the clinical guidelines for the diagnosis and treatment of chronic insomnia in adults, and scores of the Insomnia Severity Index. The study of the GSTT1 gene polymorphism was carried out using polymerase chain reaction. The lipid peroxidation - antioxidant system parameters were determined by spectrofluorophotometric methods. In the insomnia group, carriers of the normal GSTT1 genotype had higher diene conjugates with lower glutathione peroxidase activity as compared to controls. When comparing groups of women with the deletion in GSTT1, an increase in the content of substrates and lipid peroxidation products was observed at all stages of lipid peroxidation, oxidized glutathione, glutathione S-transferase activity with a decrease in the content of reduced glutathione and retinol in the insomnia group as compared to controls. In the insomnia group, carriers of the deletion genotype had a higher content of ketodienes/conjugated trienes and active products of thiobarbituric acid, a lower content of retinol and α-tocopherol as compared to the patients with normal GSTT1 genotype. Conclusions. Insomnia in the menopausal women is accompanied with the intensification of lipid peroxidation only in the carriers of the GSTT1 deletion genotype.

ESTIMATION OF LIPID PEROXIDATION IN TISSUES OF FISH FROM LAKES OF THE KENTI RIVER SYSTEM, REPUBLIC OF KARELIA

Lipid peroxidation in tissues of pike Esox lucius Linnaeus, 1758 and roach Rutilus ruti‑ lus (Linnaeus, 1758) from Lake Kostomukshskoe (storing tailings from the Kostomuksha ore mining and processing enterprise) was found to be more intensive than in fish living downstream the Kenti River. The content of lipid peroxidation products in the liver of fish from Lake Koivas was significantly lower than in fish living higher up the Kenti River. This may be due to significant differences in the concentration of ions in these water bodies. A high level of primary lipid peroxidation products – diene conjugates and diene ketones, was detected in the liver of roach and pike from the tailing pond, indicating the activation of the initial stages of peroxidation in fish tissues in response to a high level of minerals from the tailings pond. Among all the studied fish tissues the highest level of lipid peroxidation products was found in gills. This can be explained by the higher degree of oxygenation in this organ and, as a consequence, enhanced oxidation of membrane lipids in gills. The high mineralization of the Kostomuksha mining plant’s tailing dump creates an unusual, hypertonic habitat for freshwater fish. This affects, above all, the functioning of their gills, which regulate the body’s water-salt balance. Fine suspended solids in Lake Kostomukshskoe reduce the oxygenation of gills in roach and pike, thus affecting the rate of peroxidation processes. The tissue with the lowest content of lipid peroxidation products was muscles. The activity of peroxidation processes in muscles showed no significant variation among fish from different waterbodies. Fish from Lake Koivas (the farthest away from the industrial source) demonstrated the lowest degree of tissue peroxidation. The activity of peroxidation processes declines in the following sequence: gills-liver-muscles. The most pronounced variation in the content of lipid peroxidation products was found in the liver, presumably due to the high metabolic and detoxification activity of this organ. Based on the analysis of the studied parameters in the two fish species, pike is more resistant to industrial impact, most likely due to the characteristics of its ecology

Parameters of Oxidative Stress in Reproductive and Postmenopausal Mexican Women.

In the reproductive phase, women experience cyclic changes in the ovaries and uterus, and hormones regulate these changes. Menopause is the permanent loss of menstruation after 12 months of amenorrhea. Menopause is also linked to a decrease in estrogen production, causing an imbalance in oxidative stress. We aimed to compare the three stages of lipid peroxidation, protein oxidative damage, and total antioxidant capacity (TAC) between reproductive-aged women (RAW) and postmenopausal women (PMW) in Mexico. We carried out a cross-sectional study with 84 women from Mexico City, including 40 RAW and 44 PMW. To determine the oxidative stress of the participants, several markers of lipid damage were measured: dienes conjugates (DC), lipohydroperoxides (LHP), and malondialdehyde (MDA); exposure to protein carbonyl is indicative of oxidative modified proteins, and TAC is indicative of the antioxidant defense system. Biomarkers of oxidative stress were significantly lower in RAW vs. PMW. DC were 1.31 ± 0.65 vs. 1.7 ± 0.51 pmol DC/mg dry weight (p = 0.0032); LHP were 4.95 ± 2.20 vs. 11.30 ± 4.24 pmol LHP/mg dry weight (p < 0.0001); malondialdehyde was 20.37 ± 8.20 vs. 26.10 ± 8.71 pmol MDA/mg dry weight (p = 0.0030); exposure of protein carbonyl was 3954 ± 884 vs. 4552 ± 1445 pmol PC/mg protein (p = 0.042); and TAC was 7244 ± 1512 vs. 8099 ± 1931 pmol Trolox equivalent/mg protein (p = 0.027). PMW display significantly higher oxidative stress markers compared to RAW; likewise, PMW show a higher TAC.

Про-антиокcидантна рівновага в печінці та м’язах cтерляді за дії штучного гіпобіозу й анестезії

&lt;p&gt;The pro-antioxidant balance in the liver and muscles of sterlet under the influence of artificial carbonic dioxyde hibernation and anaesthesia have been studied. In experiments were used two-years old sterlet weighing 170–210 g which were grown under conditions of a closed water supply. Four groups of five copies of each fish were formed: Control group I (fish remained intact); Group II (the clove oil was used for fish anaesthesia, which was added to the water); Group III (fish were hibernated by carbon dioxide); Group IV - control of hibernation (after complete recovery from carbon dioxide hibernation, these groups of fish were returned to the pool of incubation shop for her follow–up). The research of intensity of the formation products of peroxide oxidation of lipids (POL) in the body of sterlet in the experimental conditions were conducted by determining the content of thiobarbituric acid active products (TBA–active products) in the muscles and liver that are generated at the final stages of lipid peroxidation. The content of malonic dialdehyde (MDA), one of the intermediate products of lipid peroxidation of membrane was determined photometrically by the concentration of colored complex formed by its reaction in the acidic environment of the two molecules of thiobarbituric acid (TBA). Catalase activity was determined by the ability of hydrogen peroxide to form the stable colored complex with Molybdenum salts. The results support the adaptive character in fluctuations in the antioxidant protection system by the actions of carbon dioxide hibernation and anaesthesia on the sterlet body. This also contributes to usage of these conditions in fish transportation at long distances without stress factors.&lt;/p&gt;

Concentration of hepatic vitamins A and E in rats exposed to chlorpyrifos and/or enrofloxacin

The aim of the study was to determine the level of antioxidant vitamins A and E in the liver of rats exposed to chlorpyrifos and/or enrofloxacin. Chlorpyrifos (Group I) was administered at a dose of 0.04 LD50 (6 mg/kg b.w.) for 28 days, and enrofloxacin (Group II) at a dose of 5 mg/kg b.w. for 5 consecutive days. The animals of group III were given both of the mentioned above compounds at the same manner as groups I and II, but enrofloxacin was applied to rats for the last 5 days of chlorpyrifos exposure (i.e. on day 24, 25, 26, 27 and 28). Chlorpyrifos and enrofloxacin were administered to rats intragastrically via a gastric tube. The quantitative determination of vitamins was made by the HPLC method. The results of this study indicated a reduction in the hepatic concentrations of vitamins A and E, compared to the control, which sustained for the entire period of the experiment. The four-week administration of chlorpyrifos to rats resulted in a significant decrease of vitamins in the initial period of the experiment, i.e. up to 24 hours after exposure. For vitamin A the maximum drop was observed after 24 hours (19.24%) and for vitamin E after 6 hours (23.19%). Enrofloxacin caused a slight (3-9%) reduction in the level of the analysed vitamins. In the chlorpyrifos-enrofloxacin co-exposure group reduced vitamins A and E levels were also noted, but changes in this group were less pronounced in comparison to the animals intoxicated with chlorpyrifos only. The decrease in the antioxidant vitamin levels, particularly noticeable in the chlorpyrifos- and the chlorpyrifos combined with enrofloxacin-treated groups, may result not only from the increase in the concentration of free radicals, but also from the intensification of the secondary stages of lipid peroxidation.

Interaction of wine anthocyanin derivatives with lipid bilayer membranes

This work deals with the capacity of various anthocyanin derivatives to insert lipid bilayer membrane. Malvidin-3-O-glucoside was studied in its various charge forms (flavylium cation, neutral and anionic quinonoid bases) as well as its deglycosylated, hydrated and conjugated derivatives. Based on molecular dynamics (MD) and COSMOmic simulations, membrane partitioning and crossing were evaluated. The free MD simulations provided molecular description of all intermolecular interactions driving penetration and orientation of these polyphenols in a model of DOPC lipid bilayer. Most of the derivatives are theoretically predicted to insert rather deep in the membrane i.e., embedded in between lipid chains, therefore being prone to scavenge both the initiation and propagation stages of lipid peroxidation. Here we also stress again the importance of the method used to evaluate atomic charge distribution to allow a correct description of membrane penetration.

Effect of frozen storage on fatty acid composition and changes in lipid content ofScomberomorus commersoniandCarcharhinus dussumieri

Department of Food Technology, College ofAgriculture, Tarbiat Modares University, Tehran, IranSummaryInvestigated were the changes in fatty acid composition,oxidation and enzymatic deterioration of lipids in frozen()30 C) fish fillets from the Persian Gulf. The narrow barredSpanish mackerel (Scomberomorus commersoni) and whitecheek shark (Carcharhinus dussumieri) were tested with storagetimes of 0, 1, 2, 3, 4, 5 and 6 months at )18 C. Statisticalresults showed that the major fatty acids among the saturatedand monounsaturated fatty acids of each fish species werepalmitic (C16:0) and oleic (C18:1n-9) acids, respectively. Bothlinoleic acid (C18:2n-6) and arachidonic acid (AA) (C20:4n-6)were predominant in total n-6 polyunsaturated fatty acids inboth mackerel and shark. The EPA (eicosapentaenoic acid;C20:5 n-3) and DHA (docosahexaenoic acid; C22:6 n-3) acidswere the major fatty acids among total n-3 acids in both fishes.During frozen storage, the PUFA (40.1 and 23.94%), n-3 (48and 42.83%), x3 ⁄ x6 (41.36 and 50%), PUFA ⁄ SFA (56 and42.23%) and EPA + DHA ⁄ C16 (55.55 and 46.66%) con-tents decreased in S. commersoni and C. dussumieri, respec-tively. Also peroxide, thiobarbituric acid (TBA) and free fattyacid (FFA) values significantly increased (P < 0.01) with thetime of storage.IntroductionMarine lipids have a high content of polyunsaturated fattyacids (PUFAs), particularly eicosapentaenoic (EPA; 20:5x-3)and docosahexaenoic acids (DHA; 22:6x-3) (Pazos et al.,2005; Bayir et al., 2006). There is strong evidence thatconsumption of fish is favourable to human health (Bayiret al., 2006). It is generally recognized that PUFA compositionmay vary among fish species.Degradation of PUFA by auto-oxidation during storageand the processing of fish oils and fatty fish easily leads toformation of volatiles associated with rancidity (Pazos et al.,2005). For this reason, freezing and frozen storage have largelybeen employed to retain fish sensory and nutritional properties(Lugasi et al., 2007). Although frozen storage of fish caninhibit microbial spoilage, the muscle proteins undergo anumber of changes (causing insolubility and formation ofaggregates) which modify their structural and functionalproperties. However, it is widely acknowledged that lipidoxidation may also be involved (Badii and Howell, 2002).Moreover, fish undergo quality changes during chilling andfrozen storage. The high degree of unsaturated lipids makesfish tissues highly susceptible to peroxidation and rapiddeterioration. Oxidative changes are mainly related to tasteand texture of the fish. In later stages of lipid peroxidation,changes in colour and nutritional value are observed (Dragoevet al., 1998).Our study was aimed at determining the fatty acid (FAs)composition and oxidative changes in fish fillets (narrow-barred Spanish mackerel and white cheek shark being the mostimportant fishes in the Persian Gulf: Fishery Statistics, 2005)caused by lipid peroxidation during frozen storage as animportant factor in quality control of frozen fishery products.Materials and methods

Structural and Functional Characteristics of Erythrocyte Membranes and Their Correction with Perftoran

We studied the effect of single intravenous injection of perftoran on the intensity of the initial stages of lipid peroxidation and structural and functional characteristics of erythrocyte membranes (cell model). Perftoran slightly activated the initial stages of lipid peroxidation and optimized structural and functional characteristics of erythrocyte membranes.

Molecular action of vitamin E in lipoprotein oxidation:: Implications for atherosclerosis

The oxidation theory of atherosclerosis proposes that the oxidative modification of low-density lipoproteins (LDL) plays a central role in the disease. Although a direct causative role of LDL oxidation for atherogenesis has not been established, oxidized lipoproteins are detected in atherosclerotic lesions, and in vitro oxidized LDL exhibits putative pro-atherogenic activities. α-Tocopherol (α-TOH; vitamin E), the major lipid-soluble antioxidant present in lipoproteins, is thought to be antiatherogenic. However, results of vitamin E interventions on atherosclerosis in experimental animals and cardiovascular disease in humans have been inconclusive. Also, recent mechanistic studies demonstrate that the role of α-TOH during the early stages of lipoprotein lipid peroxidation is complex and that the vitamin does not act as a chain-breaking antioxidant. In the absence of co-antioxidants, compounds capable of reducing the α-TOH radical and exporting the radical from the lipoprotein particle, α-TOH exhibits anti- or pro-oxidant activity for lipoprotein lipids depending on the degree of radical flux and reactivity of the oxidant. The model of tocopherol-mediated peroxidation (TMP) explains the complex molecular action of α-TOH during lipoprotein lipid peroxidation and antioxidation. This article outlines the salient features of TMP, comments on whether TMP is relevant for in vivo lipoprotein lipid oxidation, and discusses how co-antioxidants may be required to attenuate lipoprotein lipid oxidation in vivo and perhaps atherosclerosis.

Validation of the peroxidative indicators, cis-parinaric acid and parinaroyl-phospholipids, in a model system and cultured cardiac myocytes

cis-Parinaric acid is increasingly being used in eukaryotic cells as a very sensitive marker for the initial stages of lipid peroxidation. Despite the increased application of this probe, no extensive validation, especially in cellular systems, has been performed. cis-Parinaric acid can either be inserted freely into biomembranes or incorporated (bio)synthetically into lipids (parinaroyl-lipid). Therefore, a direct comparison was made between the peroxidative behaviour of the two parinaroyl probes and the endogenous polyunsaturated fatty acids arachidonic and linoleic acid, in both an artificial lipidic system and in cultured neonatal rat heart cells. Three different radical generating systems were used, i.e., hydrogen peroxide, cumene hydroperoxide and the thermo-labile 2,2′-azobis(2-amidinopropane hydrochloride) (AAPH). The data demonstrate that the peroxidation rate of cis-parinaric acid is higher than that of the parinaroyl, arachidonoyl and linoleoyl lipids. The latter three displayed comparable peroxidation rates, showing that the peroxidative decay of parinaroyl-lipid is a good marker for the degradation of endogenous polyunsaturated fatty acids. Experimental results using the freely inserted cis-parinaric acid could potentially lead to an overestimation of the inflicted damage and should be interpreted with care. In addition, a comparison was made with the measurement of conjugated dienes and malon dialdehyde as thiobarbituric acid reactive substances. The results demonstrate that measurement of conjugated dienes and malon dialdehyde only provide information on peroxidative processes in vitro, but are not suitable for in-depth studies in cultured cells. In contrast, the use of the parinaroyl probes is a suitable, straightforward, sensitive and reproducible method for detecting the initial stages of lipid peroxidation in living cells.

Site-specific effect of radical scavengers on the resistance of low density lipoprotein to copper-mediated oxidative stress: influence of α-tocopherol and temperature

The radical scavenging capacity of active nitroxide spin label radicals located at different depths in the surface monolayer of native and α-tocopherol enriched low density lipoprotein (LDL) has been evaluated at early stages of copper-mediated lipid peroxidation. Spin labels induced a concentration-dependent prolongation in lag time and a pronounced decrease in the initial rate of conjugated diene (CD) formation. These effects strongly argue for a protective, antioxidative action of spin labels, which in turn become destroyed with the extent of oxidation by radical recombination reactions. The results revealed that the decrease in spectral intensity proceeds at a higher rate for nitroxide radicals located in a more hydrophobic environment. The loss in spin label activity is accompanied by simultaneous α-tocopherol consumption and progresses rather independently of initial α-tocopherol content. The data provided no evidence that spin labels either save α-tocopherol or compete with it for radicals. The authors, therefore, deduce that due to enhanced accessibility and mobility, spin labels located in the interior of LDL eliminate lipid-derived radicals, which otherwise would promote lipid peroxidation. Lowering of temperature clearly below the core-lipid phase transition temperature of LDL exerts a significant effect on the kinetics of copper-induced LDL oxidation, whereas the characteristics of the radical scavenging mechanisms of the spin label molecules located in the surrounding phospholipid monolayer are conserved. Taken together, the susceptibility of LDL to primary oxidative stress conditions was efficiently retarded by small amounts of radical scavengers. This effect was more pronounced for nitroxide radicals embedded deeper in the phospholipid monolayer and was rather independent of α-tocopherol enrichment.

The Effect of Indolinic and Quinolinic Nitroxide Radicals on Trout Erythrocytes Exposed to Oxidative Stress

The purpose of this study was to evaluate the ability of indolinic and quinolinic nitroxide radicals to protect trout (Salmo irideus) erythrocytes against oxidative stress. By using laurdan as a fluorescence probe, it was observed that the nitroxides inhibited the shift towards a gel phase of liposomes prepared with phospholipids extracted from trout erythrocyte membranes prior to the hemolytic event. In addition, the presence of 100 μM nitroxides in these liposomes protected the latter against lipid peroxidation determined by monitoring conjugated diene formation. However, the short chain analogue of the indolinic nitroxide and the quinolinic nitroxide had a negative effect on trout hemolysis, contrary to what has already been observed in previous studies on human RBCs (red blood cells). The half-time (t1/2) of the hemolytic process was 174 ± 4.02 min for the former and 184 ± 4.30 min for the latter compared to the control, 283 ± 5.05 min. Furthermore, the nitroxides remarkably increased the autoxidation rate of both trout and human hemoglobin to met-Hb. Even though protection at the membrane level is conferred by the nitroxides during the early stages of lipid peroxidation, their antioxidative ability might be overwhelmed at a later stage by other mechanisms such as the increased autoxidation of hemoglobin in the presence of the nitroxides, thus giving a possible explanation for the early induction of hemolysis induced by the nitroxides. The superoxide scavenging ability of all the nitroxides used was also evaluated through chemiluminescence.

A rapid and simple screening test for potential inhibitors of tocopherol-mediated peroxidation of LDL lipids

We report a rapid and convenient method for screening potential inhibitors of the initiation of low density lipoprotein (LDL) lipid peroxidation. The method uses positively and negatively charged micelles of either cetyltrimethyl ammonium chloride or sodium dodecyl sulfate with added alpha-tocopherol. It is based on the capacity of an antioxidant to attenuate alpha-tocopheroxyl radicals, generated by irradiation of the alpha-tocopherol-containing micelles with UV light, and measured directly by electron spin resonance spectroscopy. To establish the reliability of the method, we compared the alpha-to-copheroxyl radical attenuating ability (TRAA) of 53 natural and synthetic potential antioxidants with their respective ability to inhibit the early stages of LDL lipid peroxidation initiated by a low flux of water-soluble peroxyl radicals. The relationship between the measured TRAA and corresponding LDL antioxidation activity was highly significant (P < 0.00005, Rank test). Thus, the potency of a co-antioxidant for LDLs alpha-tocopherol could be predicted with > 98% probability by the TRAA test alone. The results suggest that this relatively simple method represents an effective and simple screening test that can be used for large numbers of potential inhibitors of the early stages of LDL lipid oxidation.

An experimental model for the evaluation of lipid peroxidation in lens membranes.

Lipid peroxidation has been associated with a number of specific manifestations related both to lens aging and cataract development. The assessment of the effect of various naturally occurring prooxidants as well as the development of antioxidant strategies has often been limited by the lack of appropriate and simple experimental models. In this study we discuss the adaptation of a method based on the incorporation of a fluorescent probe (parinaric acid) into biological membranes to monitor early stages of lipid peroxidation. After establishing the appropriate conditions, the method can be successfully applied to study peroxidation in bovine lens membranes, allowing for the evaluation of the effect of several free radical generating systems, including the following metal-dependent initiators: ascorbate/ iron, hydrogen peroxide/copper and cumene hydroperoxide/ copper. The inhibitory effect of the chelating agent diethylene-triaminepenta-acetic acid and the competitive hydroxyl radical scavenger sorbitol, was consistently observed on parinaric acid degradation, on hydroxyl radical yield and on the amount of thiobarbituric acid reactive material produced. It could be shown that oxidative degradation of the probe gives direct information on lens membrane susceptibility to a specific peroxidation system. Parinaric acid can therefore be used as an efficient oxidation probe to evaluate oxidative damage inflicted to lens membranes by different systems, allowing also the evaluation of the antioxidant effect of various drugs including those with potential anticataractogenic effect.

Electrophoretic evidence for the impairment of complexes of the respiratory chain during iron/ascorbate induced peroxidation in isolated rat liver mitochondria

The impairment of the complexes of the respiratory chain was studied in isolated rat liver mitochondria under the conditions of an iron/ascorbate-mediated oxidative stress. Using blue native electrophoresis technique the NADH-ubiquinone oxidoreductase, ubiquinol-cytochrome- c oxidoreductase, cytochrome oxidase and ATP-synthetase were separated from mitochondrial samples at different stages of peroxidation and quantified by densitometry. In the second dimension the protein complexes were separated into their individual subunits by Tricine/SDS-electrophoresis. In relation to the time course of lipid peroxidation protein losses were moderate in the exponential phase and enhanced towards plateau phase of TBARS formation, when the intensity of staining for the native complexes became reduced by 84%, 69%, 63% and 24% for complexes I, III, V and IV, respectively, and a high molecular aggregation band as a putative marker of oxidative stress was formed. The decline of overall staining by 23%, a decrease in trichloroacetic acid precipitable protein and the formation of acid soluble primary amines suggest the occurrence of fragmentation or degradation processes. Apparently, the impairment of the respiratory chain complexes during peroxidation was not reflected in altered electrophoretic mobilities or specific losses of protein subunits of these innermitochondrial membrane components.

Cyclic voltammetry of lipophilic compounds in oil: Direct determination of lipid peroxide with a carbon past electrode

AbstractIn preliminary experiments, we demonstrated that a cyclic voltammogram of a lipophilic compound can be conveniently obtained with the past electrode that was made from graphite powder and an oil in which the compound is dissolved. This finding was applied to the direct determination of lipid peroxides. Oxygenation of lipids at 120°C was followed by the cathodic current created by the reduction of the peroxides. The validity of the method was investigated by comparison with conventional methods (peroxide value, thiobarbituric acid, and weight gain). The amount of peroxides in an oil and its cathodic current show a linear relation in the initial stages of peroxidation (ca. 12 h). This electrochemical method was applied successfully to the evaluation of the antioxidant activity of butylated hydroxytoluene in linseed oil.