Photoperoxidation in isolated chloroplasts. I. Kinetics and stoichiometry of fatty acid peroxidation.

Reactive oxygen species and antioxidant machinery in abiotic stress tolerance in crop plants

http://directory.umm.ac.id/Data%20Elmu/jurnal/P/PlantScience/PlantScience_Elsevier/Vol151.Issue1.2000/5250.pdf

Oxidative stress and some antioxidant systems in acid rain-treated bean plants Protective role of exogenous polyamines

The occurrence of malondialdehyde (MDA), a secondary end product of the oxidation of polyunsatu- rated fatty acids, is considered a useful index of general lipid peroxidation A common method for measuring MDA, referred to as the thiobarbituric acid-reactive- substances (TBARS) assay, is to react it with thiobar- bituric acid (TBA) and record the absorbance at 532 nm However, many plants contain interfering compounds that also absorb at 532 nm, leading to overestimation of MDA values Extracts of plant tissues including purple eggplant (Solanum melongena L) fruit, carrot (Daucus carota L) roots, and spinach (Spinacia oleracea L) leaves were assessed for the presence of MDA and other non-MDA compounds absorbing at 532 nm A method described herein corrects for these interferences by subtracting the absorbance at 532 nm of a solution containing plant extract incubated without TBA from an identical solution containing TBA The reliability and eAciency of this spectrophotometric method was assessed by altering the relative ratios of exogenous MDA additions and/or extracts of red cabbage (Brassica oleracea L) leaves containing inter- fering compounds and then measuring MDA recovery Reliability was also validated through high-performance liquid chromatography and high-performance liquid chromatography-mass spectrometry techniques Results indicated that over 90% of exogenously added MDA could be recovered through the improved protocol If there were no corrections for interfering compounds, MDA equivalents were overestimated by up to 965% Interfering compounds were not detected in vegetables such as lettuce (Lactuca sativa L) and spinach which had low or negligible concentrations of anthocyanidin derivatives Comparisons between the TBARS method presented here and two currently accepted protocols indicated that the new modified method exhibits greater accuracy for quantifying TBA-MDA levels in tissues containing anthocyanins and/or other interfering com- pounds This modified protocol represents a facile and rapid method for assessment of lipid peroxidation in virtually all plant species that contain interfering com- pounds

Improving the thiobarbituric acid-reactive-substances assay for estimating lipid peroxidation in plant tissues containing anthocyanin and other interfering compounds

Water deficits cause a reduction in the rate of photosynthesis. Exposure to mild water deficits, when relative water content (RWC) remains above 70%, primarily causes limitation to carbon dioxide uptake because of stomatal closure. With greater water deficits, direct inhibition of photosynthesis occurs. In both cases limitation of carbon dioxide fixation results in exposure of chloroplasts to excess excitation energy. Much of this can be dissipated by various photoprotective mechanisms. These include dissipation as heat via carotenoids, photorespiration, CAM idling and, in some species, leaf movements and other morphological features which minimize light absorption. The active oxygen species superoxide and singlet oxygen are produced in chloroplasts by photoreduction of Oxygen and energy transfer from triplet excited chlorophyll to oxygen, respectively. Hydrogen peroxide and hydroxyl radicals can form as a result of the reactions of superoxide. All these species are reactive and potentially damaging, causing lipid peroxidation and inactivation of enzymes. They are normally scavenged by a range of antioxidants and enzymes which are present in the chloroplast and other subcellular compartments. When carbon dioxide fixation is limited by water deficit, the rate of active oxygen formation increases in chloroplasts as excess excitation energy, not dissipated fay the photoprotective mechanisms, is used to form superoxide and singlet oxygen. However, photorespiratory hydrogen peroxide production in peroxisomes decreases. Increased superoxide can be detected by EPR (electron paramagnetic resonance) in chloroplasts from droughted plants. Stiperoxide formation leads to changes suggestive of oxidative damage including lipid peroxidation and a decrease in ascorbate. These changes are not, however, apparent until severe water deficits develop, and they could also be interpreted as secondary effects of water deficit-induced senescence or wounding. Non-lethal water deficits often result in increased activity of superoxide dismutase, glutathione reductase and monodehydroascorbate reductase. Increased capacity of these protective enzymes may be part of a general antioxidative response in plants involving regulation of protein synthesis or gene expression. Since the capacity of these enzymes is also increased by other treatments which cause oxidative damage, and which alter the balance between excitation energy input and carbon dioxide fixation such as low temperature and high irradiance, it is suggested that water deficit has the same effect. Light levels that are not normally excessive do become excessive and photoprotective/antioxidative systems are activated. Some of the photoprotective mechanisms themselves could result in active oxygen formation. Photoinhibitory damage also includes a component of oxidative damage. During normally-encountered degrees of water deficit the capacity of the antioxidant systems and their ability to respond to increased active oxygen generation may be sufficient to prevent overt expression of damage. Desiccation-tolerant tissues such as bryophytes, lichens, spores, seeds, some algae and a few vascular plant leaves can survive desiccation to below 30-40% RWC, A component of desiccation damage in seeds and bacteria is oxygen-dependent. Desiccation causes oxidation of glutathione, a major antioxidant, and appearance of a free radical signal detected by EPR in a number of tissues suggesting that oxidative damage has occurred. In photosynthetic cells damage may arise from photooxidation. Disruption of membrane-bound electron tranport systems in partially hydrated tissue could lead to reduction of oxygen to superoxide. Oxidation of lipids and sulphydryl groups may also occur in dry tissue. Tolerant cells recover upon rehydration and arc able to reduce their glutathione pool. Non-tolerant species go on to show further oxidative damage including lipid peroxidation. It is difficult to attribute this subsequent damage to the cause or effect of death. Embryos in seeds lose desiccation tolerance soon after imbibition. This is associated with membrane damage that has been attributed to superoxide-mediated deesterification of phospholipids and loss of lipophilic antioxidants. These effects are discussed in relation to other mechanisms involved in desiccation tolerance. Contents Summary 27 I. Introduction 28 II. Generation of active oxygen and defence mechanisms in plant cells 29 III. The effect of water deficit on photosynthesis 31 IV. Mechanisms for active oxygen generation during water deficit 36 V. Evidence for oxidative damage during water deficit 39 VI. Desiccation 47 VII. Conclusions 52 Acknowledgements 53 References 53.

Robert L. Heath

Papers

Photoperoxidation in isolated chloroplasts. I. Kinetics and stoichiometry of fatty acid peroxidation.

Photoperoxidation in isolated chloroplasts. II. Role of electron transfer.

Effect of light on lipid peroxidation in chloroplasts.

Steady-state fluorescence of spinach chloroplasts and electron flow