Membrane Lipid Oxidation Research Articles

Solid-phase microextraction–gas chromatography–mass spectrometry method validation for the determination of endogenous substances: Urinary hexanal and heptanal as lung tumor biomarkers

Hexanal and heptanal are endogenous aldehydes coming from membrane lipid oxidation, found in lung cancer patients’ blood, and suggested as lung tumor biomarkers. Here the urinary matrix was investigated instead of blood and the difficulties related to the determination of endogenous substances in biological matrices were faced by developing an external calibration HS-SPME/GC/MS method. The methodology was validated according to international validation procedures and it was verified analyzing unknown biological samples from cancer patients and healthy subjects. Percentage accuracy and precision, ranging from −11.25 to 10.85% and from 0.45 to 4.46%, respectively, were obtained, together with limits of detection (LODs) and lower limits of quantification (LLOQs) of 0.11 and 0.23 pg μL −1 for hexanal and of 0.10 and 0.21 pg μL −1 for heptanal. Analytes percentage recoveries (66.3%, hexanal and 70.5%, heptanal) and stability were evaluated. No analytes degradation was found at room temperature, while the remarkable analytes loss found after 1 month storage suggests analyzing biological samples within a week from storage. Results coming from the analysis of unknown biological samples showed no evident differences of heptanal urinary excretion between lung cancer patients and healthy subjects (0.22–0.95 and 0.21–0.69 pg μL −1, respectively), while hexanal urinary concentrations in cancer patients (0.24–4.36 pg μL −1) were slightly higher than those found in control group ones (0.23–1.26 pg μL −1). The obtained results highly suggest to do further investigations in order to collect statistically significant biological data to discriminate between the pathological state of lung cancer patients and physiological conditions of healthy subjects, using the simple, rapid and cheap method here reported for the quantification of urinary aldehydes.

Perturbation of Rat Heart Plasma Membrane Fluidity Due to Metabolites of Permethrin Insecticide

Due to increased global use, acute and chronic exposures to pyrethroid insecticides in humans are of clinical concern. Pyrethroids have a primary mode of action that involves interference with the sodium and calcium channels in excitable cells, which may include cardiac myocytes. Here, we investigated the possible cardiac toxicity of permethrin metabolites (METP), 3-phenoxy-benzyl alcohol (3PBA), 3-phenoxy-benzaldehyde (3PBALD), and 3-phenoxybenzoic acid (3PBACID). Plasma membrane fluidity, polarity, lipid, and protein oxidation were studied in isolated rat heart cells. Laurdan was chosen as probe to detect the lateral mobility and polarity of its environment and thus water penetration into the hydrophobic part of the bilayer, while 1,6-diphenyl-1,3,5-hexatriene permits to measure changes in fluidity in the inner part of the bilayer. Results show that METP can change membrane fluidity at different depths of the bilayer according to their partition coefficient. Consequently, 3PBA, at all concentration used, decreases membrane fluidity and polarity in the hydrophilic-hydrophobic region of the bilayer, and similar effect was observed with 20μM 3PBALD or 10 or 20μM 3 PBACID. Membrane dynamics in the hydrophobic core resulted decreased by 3PBALD, while it was increased by 20μM 3PBACID. All METP increase protein and lipid oxidation, and the peroxidative lipid damage decreases with the type of METP produced during the transformation pathway from parent compound to 3PBACID. Consequently, 3PBA induced the highest lipid peroxidation, while 3PBACID was the stronger inducer of protein damage.

Instant coffee with high chlorogenic acid levels protects humans against oxidative damage of macromolecules

Coffee is among the most frequently consumed beverages. Its consumption is inversely associated to the incidence of diseases related to reactive oxygen species; the phenomenon may be due to its antioxidant properties. Our primary objective was to investigate the impact of consumption of a coffee containing high levels of chlorogenic acids on the oxidation of proteins, DNA and membrane lipids; additionally, other redox biomarkers were monitored in an intervention trial. The treatment group (n=36) consumed instant coffee co-extracted from green and roasted beans, whereas the control consumed water (800 mL/P/day, 5 days). A global statistical analysis of four main biomarkers selected as primary outcomes showed that the overall changes are significant. 8-Isoprostaglandin F2α in urine declined by 15.3%, 3-nitrotyrosine was decreased by 16.1%, DNA migration due to oxidized purines and pyrimidines was (not significantly) reduced in lymphocytes by 12.5 and 14.1%. Other markers such as the total antioxidant capacity were moderately increased; e.g. LDL and malondialdehyde were shifted towards a non-significant reduction. The oxidation of DNA, lipids and proteins associated with the incidence of various diseases and the protection against their oxidative damage may be indicative for beneficial health effects of coffee.

Adaptational changes in the lipids and fatty acid profile of the cell and thylakoid membrane of rice plants exposed to sunlight

Adaptational changes occurring in the lipids and fatty acids of the cell and the thylakoid membrane in response to high light treatment, was studied in 30days old rice (Oryza sativa L. cv. Jyothi) plants grown under low (150-200μmolm(-2)s(-1)) or moderate (600-800μmolm(-2)s(-1)) light conditions. Results were compared with rice plants grown in high (1200-2200μmolm(-2)s(-1)) light conditions. Exposure of rice plants and isolated chloroplast to high light, resulted in an increase in the amount of malonaldehyde, indicating oxidation of membrane lipids. Qualitative and quantitative changes in the phosphoglycolipids and quantitative changes in neutral lipids were observed in rice plants grown under the different growth conditions. A few of the phosphoglycolipids and neutral lipids were present exclusively in plants grown at low or moderate or high light, indicating requirement of different type of lipid composition of rice plants in response to their different growth irradiances. However, no significant quantitative changes were observed in the different saturated and unsaturated fatty acid groups of total lipids in low, moderate and high light grown rice plants, as a result of exposure to high light. No qualitative changes in the fatty acid composition due to difference in growth irradiance or high light treatment were seen. The changes observed in the phosphoglycolipids and neutral lipid composition of cell and thylakoid membrane of low, moderate and high light grown rice plants in response to high light, are probably the result of physiological changes in the rice plants, to sustain optimum structure and function of the cell and thylakoid membrane to maintain active physiological functions to endure high light conditions.

Oxidative alterations of the olfactory bulb and entorhinal cortex in early stages of Alzheimer's disease

derivative. Results: The present study indicates that oxidative stress resulting from the treatment of H2O2 can be inhibited in the presence of our oxazine derivative, in a dose-dependent manner. Based on our findings, it seems that in this cytoprotection, inhibition of NF-kB is the main factor that protects neurons against oxidative stress induced caspase-3 activity. In addition, upregulation of HO-1 and g-GCS through Nrf2 pathway may present another possible mechanism of action of this compound. It was also found that oxazine derivative interferes the oxidation of membrane lipids, one of the primary events in oxidative cellular damage. While treatment with H2O2 increased MDA level to about 170%, compared to control, pretreatment of PC12 cells with oxazine derivative decreased MDA content to about 102%, compared to control. On the other hand, the antioxidant enzymes those serve as a detoxifying system to prevent damage caused by ROS was greatly increased in the presence of oxazine derivative. Conclusions: In summary, our results clearly indicate that our oxazine derivative prevent the H2O2-induced cell death in differentiated PC12 cells, dose-dependently. It seems that in this cytoprotection, Nrf2 and NF-kB are the main factors which protect neurons against oxidative stressinduced apoptosis.

Comparison of glutathione peroxidase 1 (GPx1) and peroxiredoxin 6 (Prdx6) in protection against lung oxidative stress in a perfused lung model

Prdx6 plays an important role in lung anti‐oxidant defense, presumably due to its unique ability to reduce phospholipid hydroperoxides. Our previous studies with intact mice and isolated endothelial cells showed greater sensitivity to oxidant stress of Prdx6 null compared to wild type and GPx1 null. We now used isolated perfused mouse lungs to compare their response to 1 hr treatment with t‐butyl hydroperoxide (t‐BOOH, 5 mM) or paraquat (PQ, 1 mM) added to the perfusate. Lung injury as indicated by protein in the broncoalveolar lavage fluid (BALF), LDH in the lung perfusate, and protein carbonyls in the lung homogenate was increased significantly in Prdx6 null compared to GPx1 null lungs. Lipid peroxidation analyzed by TBARS and 8‐isoprostanes in the BALF and lung homogenate as well as by 235nm fluorescence (conjugated dienes), Fe2+‐xylenol orange, and DPPP assays was significantly greater in Prdx6 null compared to GPx1 null lungs. Analysis of expression levels of oxidant related genes using a PCR‐based Superarray (SA Biosciences) indicated no significant differences between the Prdx6 and the GPx1 null, except for the target genes. We conclude that Prdx6 null are significantly more sensitive to oxidant stress as compared to GPx1 null, suggesting that the ability of Prdx6 to scavenge phospholipid hydroperoxides and to repair membrane lipid oxidation can account for its major role in antioxidant defense.

Molecular Cloning and Expression of Sequence Variants of Manganese Superoxide Dismutase Genes from Wheat

Reactive oxygen species (ROS) are very harmful to living organisms due to the potential oxidation of membrane lipids, DNA, proteins, and carbohydrates. transformed E.coli strain QC 871, superoxide dismutase (SOD) double-mutant, with three sequence variant MnSOD1, MnSOD2, and MnSOD3 manganese superoxide dismutase (MnSOD) gene isolated from wheat. Although all QC 871 transformants grown at <TEX>$37^{\circ}C$</TEX> expressed mRNA of MnSOD variants, only MnSOD2 transformant had functional SOD activity. MnSOD3 expressed active protein when grown at <TEX>$22^{\circ}C$</TEX>, however, MnSOD1 did not express functional protein at any growing and induction conditions. The sequence comparison of the wheat MnSOD variants revealed that the only amino acid difference between the sequence MnSOD2 and sequences MnSOD1 and 3 is phenylalanine/serine at position 58 amino acid. We made MnSOD2S58F gene, which was made by altering the phenylalaine to serine at position 58 in MnSOD2. The expressed MnSOD2S58F protein had functional SOD activity, even at higher levels than the original MnSOD2 at all observed temperatures. These data suggest that amino acid variation can result in highly active forms of MnSOD and the MnSOD2S58F gene can be an ideal target used for transforming crops to increase tolerance to environmental stresses.

Acteoside protects endothelial cells against free radical-induced oxidative stress.

The protective effect of acteoside against membrane lipid oxidation and free radical-mediated impairment of endothelial function was investigated. Results showed that iron-mediated oxidative modification of the cell membrane in cultured bovine pulmonary endothelial cells (PAECs) was significantly attenuated by acteoside as measured by thiobarbituric acid-reactive substances (TBARS). Fenton's reagent (H2O2/Fe2+) was used to generate hydroxyl radicals (*OH) and induce oxidative stress. Acteoside not only effectively minimized the loss of cell viability induced by hydroxyl radicals in cultured endothelial cells but also countered the free radical-induced destruction of the endothelium-dependent relaxation to acetylcholine in rat aorta. Furthermore, acteoside showed a dose-dependent scavenging effect of 1,1-diphenyl-2-picryl-hydrazyl (DPPH) radicals and appeared to be the most efficient in comparison with the four reference compounds (alpha-tocopherol, vitamin C, probucol and resveratrol). These data suggested that acteoside protects the cell from oxidative stress and that scavenging of free radicals could be a key mechanism contributing to the cytoprotective effect of acteoside.

Protective effect of Amphiroa dilatata on ROS induced oxidative damage and MMP expressions in HT1080 cells

Antioxidative and matrix metalloproteinase (MMP) inhibitory effects of methanolic extract from A. dilatata were investigated in HT1080 cells. Treatment with A. dilatata extract suppressed the intensity of electron spin resonance signal on the generation of DPPH radicals in a cell-free system and intracellular reactive oxygen species (ROS) formation by 2′,7′-dichlorofluorescin diacetate (DCFH-DA). Radical simulated oxidation of membrane proteins and lipids were also inhibited by treatment with A. dilatata extract in a concentration-dependent manner. In addition, a protective effect on oxidative damage of purified genomic DNA was observed in the presence of A. dilatata extract. These results revealed that A. dilatata has an excellent scavenging ability on ROS-induced oxidative damage. Moreover, this extract reduced the expression levels of MMP-2 and -9 in gelatin zymography, and RT-PCR analysis without any cytotoxic influence. Therefore, A. dilatata has a promising potential as a valuable source to be used as ingredient in functional foods, related to oxidative damage.

Enhancing the natural defences and barrier protection of aquaculture species

The promotion of nutritional strategies that optimize natural defence mechanisms in fish is of critical importance in producing robust juveniles and adult fish. These animals are more capable of minimizing the impact of opportunistic pathogen attack, thus improving liveability and performance characteristics. The importance of the piscine gastrointestinal tract as a major endocrine and osmoregulatory organ is well reported as is its function as a defensive barrier to pathogen attack. Investigations using the inclusion of a specific structural form of mannan oligosaccharide have been shown to improve the performance parameters, immune status, and gut morphology and improve an important aspect of barrier protection through the enhancement of mucal production in a number of aquaculture species. The selenium status of an animal is pivotal in determining the success of the innate and adaptive immune response of the animal, and the use of an organic selenium source, in the form of a selenoyeast, has been shown to improve enzyme function and tissue uptake. The antioxidant role of many of the selenoptroteins and the role of selenium in the glutathione peroxidase enzyme pathways involved in the control of oxidative stress is critical if oxidative damage and cell membrane lipid peroxidation are to be prevented. The use of these compounds as feed additives has important implications for health management in commercial aquaculture facilities. Further research is needed to evaluate the benefits offered by a range of commercial products.

Application of inter simple sequence repeat (ISSR marker) to detect genotoxic effect of heavy metals on Eruca sativa (L.)

As an influence of the Mediterranean diet, Eruca sativa (rocket salad) is eaten all over the world in salads and soups. It belongs to plant order Capparales (glucosinolate-containing species) and it is from the family Brassicaceae. Predominantly, the leaves of this species is eaten raw or cooked, although flowers are also consumed. Assessment of environmental contamination on ecology (plant) at molecular and population levels is important in risk quantification and remediation study. Heavy metal toxicity in plants is to induce oxidative stress linked to oxidation of proteins and membrane lipids but also to alterations of DNA damage response. E. sativa has been investigated in our study which is of agronomical importance and widely used in European countries. We studied three heavy metals Zn, Pb and Cd which showed a dose-dependent effect on radicle and coleoptile lengths of E. sativa. The radicle length was more affected than the coleoptiles length under all concentration tested plant. The ranking of genotoxic potencies in all three heavy metals was in the descending order: Cd

Microcin J25 triggers cytochrome c release through irreversible damage of mitochondrial proteins and lipids

We previously showed that the antimicrobial peptide microcin J25 induced the over-production of reactive oxygen species with the concomitant release of cytochrome c from rat heart mitochondria via the opening of the mitochondrial permeability transition pore. Here, we were able to demonstrate that indeed, as a consequence of the oxidative burst, MccJ25 induces carbonylation of mitochondrial proteins, which may explain the irreversible inhibition of complex III and the partial inhibition of superoxide dismutase and catalase. Moreover, the peptide raised the levels of oxidized membrane lipids, which triggers the release of cytochrome c. From in silico analysis, we hypothesize that microcin would elicit these effects through interaction with heme c1 at mitochondrial complex III. On the other hand, under an excess of l-arginine, MccJ25 caused nitric oxide overproduction with no oxidative damage and a marked inhibition in oxygen consumption. Therefore, a beneficial anti-oxidative activity could be favored by the addition of l-arginine. Conversely, MccJ25 pro-oxidative–apoptotic effect can be unleashed in either an arginine-free medium or by suppressing the nitric oxide synthase activity.

Sargassum thunbergii extract inhibits MMP-2 and -9 expressions related with ROS scavenging in HT1080 cells

Antioxidative and matrix metalloproteinase (MMP) inhibitory effects of methanolic extract from Sargassum thunbergii were investigated in HT1080 cells. This extract suppressed the electron spin resonance (ESR) signal intensity on generation of 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals and intracellular reactive oxygen species (ROS) formation by 2′,7′-dichlorofluorescin diacetate (DCFH-DA) method. Also, treatment with this extract inhibited radical simulated oxidation of membrane lipids and proteins in a dose-dependent manner. These results revealed that S. thunbergii extract has excellent scavenging abilities in ROS-induced damage. Moreover, this extract led to the reduction of the expression levels of MMP-2 and -9 in gelatin zymography, reverse transcription polymerase chain reaction (RT-PCR) and Western bolt analysis. Systematic separation by diverse chromatographic methods led to the isolation of sargahydroquinoic acid and sargachromanols ( 1– 3). Polyunsaturated fatty acids were identified by gas chromatographic analysis. Therefore, S. thunbergii can be developed as therapeutic potential antioxidant and MMP inhibitor, closely related to ROS, which can be traceable to sargahydroquinoic acid, sargachromanols, and polyunsaturated fatty acids contained in S. thunbergii.

Two Disaccharides and Trimethylamine N-Oxide Affect Aβ Aggregation Differently, but All Attenuate Oligomer-Induced Membrane Permeability

Interaction between aggregates of amyloid beta protein (Abeta) and membranes has been hypothesized by many to be a key event in the mechanism of neurotoxicity associated with Alzheimer's disease (AD). Proposed membrane-related mechanisms of neurotoxicity include ion channel formation, membrane disruption, changes in membrane capacitance, and lipid membrane oxidation. Recently, osmolytes such as trehalose have been found to delay Abeta aggregation in vitro and reduce neurotoxicity. However, no direct measurements have separated the effects of osmolytes on Abeta aggregation versus membrane interactions. In this article, we tested the influence of trehalose, sucrose and trimethylamine-N-oxide (TMAO) on Abeta aggregation and fluorescent dye leakage induced by Abeta aggregates from liposomes. In the absence of lipid vesicles, trehalose and sucrose, but not TMAO, were found to delay Abeta aggregation. In contrast, all of the osmolytes significantly attenuated dye leakage. Dissolution of preformed Abeta aggregates was excluded as a possible mechanism of dye leakage attenuation by measurements of Congo red binding as well as hydrogen-deuterium exchange detected by mass spectrometry (HX-MS). However, the accelerated conversion of high order oligomers to fibril caused by vesicles did not take place if any of the three osmolytes presented. Instead, in the case of disaccharide, osmolytes were found to form adducts with Abeta, and change the dissociation dynamics of soluble oligomeric species. Both effects may have contributed to the observed osmolyte attenuation of dye leakage. These results suggest that disaccharides and TMAO may have very different effects on Abeta aggregation because of the different tendencies of the osmolytes to interact with the peptide backbone. However, the effects on Abeta membrane interaction may be due to much more general phenomena associated with osmolyte enhancement of Abeta oligomer stability and/or direct interaction of osmolyte with the membrane surface.

Paraoxonase genes and susceptibility to ALS

In this issue of Neurology ®, Wills et al.1 report that they found no association between paraoxonase genes and amyotrophic lateral sclerosis (ALS). Remarkably, 19 of the 23 authors of that article had previously published evidence that paraoxonase gene ( PON1 ) single nucleotide polymorphisms (SNPs) are associated with an increased risk of ALS. The story started in the 1991 Gulf War, when reports of a possible excess of ALS cases among deployed troops generated controversy.2–3 Although electrophysiologic studies found no increase of ALS or any neuromuscular disease in the Gulf War or other modern wars,4 the controversy persists.5–6 The paraoxonases are encoded by a three-gene cluster on chromosome 7 ( PON1 , PON2 , and PON3 ). These enzymes are involved in detoxifying products of oxidative stress, which is presumed to be important in the pathogenesis of ALS. Because they protect against the oxidation of neuronal membrane lipids, defects in these genes could be a mechanism by which genetic susceptibility to toxic environmental factors come about. Also, one ALS risk factor is exposure to organophosphate insecticides, chemicals that are metabolized by an oxidative process. In 2006,7 association studies suggested that PON genes are involved in ALS susceptibility but the same genes are also activated in stroke, Alzheimer disease, Parkinson disease, and other conditions. Wills et al. used data from six candidate gene studies; …

Hyperglycemia Promotes Membrane Cholesterol Crystalline Domain Formation Through Lipid Peroxidation: Inhibition with Atorvastatin Metabolite

Insulin resistance and poor glycemic control contribute to atherogenesis through the chemical and structural modification of cell membrane lipids. The direct contribution of glucose to these membrane alterations, however, is not well understood. In this study, small angle x-ray diffraction and spectrophotometry were used to examine the autoxidative effects of glucose on lipid oxidation and structural organization in model membranes comprised of dilinoleoylphosphatidylcholine (DLPC) and cholesterol. Membranes were prepared at cholesterol-to-phospholipid (C/P) mole ratios ranging from 0.2 to 0.8 in order to model physiologic and hyperlipidemic conditions. Changes in membrane lipid organization and unit cell periodicity (d-space) were correlated with lipid hydroperoxide (LOOH) concentration measured at 24 hr intervals. The effects of glucose on lipid peroxidation were more pronounced at elevated levels of membrane cholesterol, with LOOH levels 20% higher at 0.8 C/P than at 0.2 C/P. At 0.6 C/P, glucose treatment resulted in a concentration-dependent increase in LOOH formation as compared to control. These changes corresponded to a reduction in membrane bilayer width (51 A to 49 A) and the progressive formation of highly-ordered cholesterol crystalline domains (d-space value of 34 A). Treatment with atorvastatin hydroxy metabolite, a statin with scavenger antioxidant properties, inhibited the membrane-altering effects of hyperglycemia in a dose-dependent manner, even at elevated cholesterol levels. These data demonstrate that glucose directly stimulates lipid peroxidation and subsequent changes in membrane structure, including the formation of immiscible cholesterol crystalline domains. Insights from this study may also serve as a model for better understanding the membrane structural changes associated with diabetes and related complications.

Lipid and protein oxidation in hepatic homogenates and cell membranes exposed to bile acids

Cholestasis occurs in a variety of hepatic diseases and causes damage due to accumulation of bile acids in the liver. The aim was to investigate the effect of several bile acids, i.e. chenodeoxycholic, taurochenodeoxycholic, deoxycholic, taurodeoxycholic, ursodeoxycholic, lithocholic and taurolithocholic (TLC), in inducing oxidative damage. Hepatic tissue of male Sprague-Dawley rats was incubated with or without 1 mM of each bile acid, with or without 0.1 mM FeCl3 and 0.1 mM ascorbic acid for the purpose of generating free radicals. Several bile acids increased lipid and protein oxidation, with TLC being the most pro-oxidative (657% and 175% in homogenates and 350% and 311% in membranes, respectively). TLC also enhanced iron-induced oxidative stress to lipids (21% in homogenates and 29% in membranes) and to proteins (74% in membranes). This enhancement was dose- and time-dependent and was reduced by melatonin. These results suggest that bile acids differentially mediate hepatic oxidative stress and may be involved in the physiopathology of cholestasis.

Airborne Particulate Matter and Human Health: Toxicological Assessment and Importance of Size and Composition of Particles for Oxidative Damage and Carcinogenic Mechanisms

Air pollution has been considered a hazard to human health. In the past decades, many studies highlighted the role of ambient airborne particulate matter (PM) as an important environmental pollutant for many different cardiopulmonary diseases and lung cancer. Numerous epidemiological studies in the past 30 years found a strong exposure-response relationship between PM for short-term effects (premature mortality, hospital admissions) and long-term or cumulative health effects (morbidity, lung cancer, cardiovascular and cardiopulmonary diseases, etc). Current research on airborne particle-induced health effects investigates the critical characteristics of particulate matter that determine their biological effects. Several independent groups of investigators have shown that the size of the airborne particles and their surface area determine the potential to elicit inflammatory injury, oxidative damage, and other biological effects. These effects are stronger for fine and ultrafine particles because they can penetrate deeper into the airways of the respiratory tract and can reach the alveoli in which 50% are retained in the lung parenchyma. Composition of the PM varies greatly and depends on many factors. The major components of PM are transition metals, ions (sulfate, nitrate), organic compound, quinoid stable radicals of carbonaceous material, minerals, reactive gases, and materials of biologic origin. Results from toxicological research have shown that PM have several mechanisms of adverse cellular effects, such as cytotoxicity through oxidative stress mechanisms, oxygen-free radical-generating activity, DNA oxidative damage, mutagenicity, and stimulation of proinflammatory factors. In this review, the results of the most recent epidemiological and toxicological studies are summarized. In general, the evaluation of most of these studies shows that the smaller the size of PM the higher the toxicity through mechanisms of oxidative stress and inflammation. Some studies showed that the extractable organic compounds (a variety of chemicals with mutagenic and cytotoxic properties) contribute to various mechanisms of cytotoxicity; in addition, the water-soluble faction (mainly transition metals with redox potential) play an important role in the initiation of oxidative DNA damage and membrane lipid peroxidation. Associations between chemical compositions and particle toxicity tend to be stronger for the fine and ultrafine PM size fractions. Vehicular exhaust particles are found to be most responsible for small-sized airborne PM air pollution in urban areas. With these aspects in mind, future research should aim at establishing a cleared picture of the cytotoxic and carcinogenic mechanisms of PM in the lungs, as well as mechanisms of formation during internal engine combustion processes and other sources of airborne fine particles of air pollution.

Oxidative stress responses in bivalves ( Scrobicularia plana, Cerastoderma edule) from the Oued Souss estuary (Morocco)

The aim of this work was to study the responses of oxidative-stress biomarkers (catalase: CAT, thiobarbituric acid reactive substances: TBARS) in bivalves ( Scrobicularia plana and Cerastoderma edule) from the Oued Souss estuary (Morocco). This estuary, subjected to large amounts of sewage discharges and industrial effluents, do not receive since November 2002 any waste outlets due to their connection to a wastewater purification plant. High CAT and TBARS values were recorded in both species collected before the implantation of wastewater treatment. These results confirm that these bivalves were submitted to various contaminants generating oxidative stress and membrane lipid peroxidation. The diminution of these two biomarkers following the setting up of the purification plant indicates probably a reduction in contaminants in the Oued Souss estuary. We recommend the use of CAT and TBARS as oxidative-stress biomarkers in S. plana and C. edule.

Antioxidant effects of dihydroquercetin and rutin in peroxidase reactions catalyzed by cytochrome c

The combination of cytochrome c (Cyt c) and cardiolipin is known to catalyze free radical oxidation of membrane lipids, thereby triggering the cascade of reactions that lead to programmed cell death (apoptosis). It was interesting to estimate the possibility of controlling this reaction with the use of inhibitors of free radical reactions (antioxidants). The effects of bovine heart cardiolipin (BCL) and synthetic 1,1′,2,2′-tetraoleyl cardiolipin (TOCL) on the Cyt c peroxidase activity detected by luminol-enhanced chemiluminescence (CL) were studied. The CL amplitude substantially increased in the presence of TOCL and BCL in the ranges of the cardiolipin to Cyt c molar ratios from 0 to 30 and from 0 to 50, respectively, and insignificantly increased upon further increase in these ratios to 640: 1 and 320: 1. This dependence of the CL amplitude on the cardiolipin to protein ratio may have been accounted for by saturation of the protein surface with cardiolipin molecules reached at a certain molar ratio. The effects of the antioxidants rutin and dihydroquercetin (DHQ) on the peroxidase activity of Cyt c in combination with BCL was studied in the presence of the CL activator coumarine C-525 at a BCL to Cyt c molar ratio of 32: 1. In both cases, the peroxidase activity decreased as the antioxidant content of the system increased. A 50% inhibition of the reaction was observed at DHQ and rutin concentrations of 10 and 3 μM, respectively. It is supposed that the use of antioxidants will make it possible to control the formation of lipid radicals on membrane-bound Cyt c, which is the key stage of apoptosis.