Recent Advances in the Development of a Diagnostic Test for Irradiated Foodstuffs

Recent advances in the development and application of diagnostic tests for irradiated foodstuffs are reviewed. Exposure of water, the major chemical constituent of most foodstuffs to a source of ionising radiation initially generates the highly reactive radical species H', -OH and e^- (aq) which react very rapidly with a wide variety of biological molecules. The detection of foodstuffs subjected to irradiation processing requires the identification and/or quantification of ‘unnatural’ chemical species (i.e. those not usually formed by normal metabolic processes) produced by the attack of ·OH radical or e^-(aq) on suitable ‘target’ molecules. Modern methods for the analysis of a series of these ‘unnatural’ products arising from the interaction of radiolytically-generated ·OH radical or e^-(aq) with polyunsaturated fatty acids, DNA, aromatic compounds and other biologically important scavenger molecules are examined. It is concluded that the analytical test to be conducted is highly dependent on the nature of the foodstuff to be tested.     

A Water-Soluble Quaternary Ammonium Analog of α-Tocopherol,that Scavenges Lipoperoxyl,Superoxyl and Hydroxyl Radicals

The new water-soluble ammonium-analog of α-tocopherol (vitamin E) (compound1: 3, 4-dihydro-6-hydroxy-N, N, N-2, 5, 7, 8-heptamethyl-2H-1-benzopyran-2-ethanaminium 4-methylbenzenesulfonate) and its tertiary amine derivative (compound2: 3, 4-dihydro-2-(2-dimethylaminoethyl)-2, 5, 7, 8-tetramethyl-2H-1-benzopyran-6-ol hydrochloride) were investigated as scavengers of oxygen-derived free radicals. Compounds 1 and 2 were at least 40 times more potent inhibitors of Fe-driven heart microsomal lipid peroxidation than Trolox. While the α-tocopherol analogs had the same potency as scavengers of xanthine/xanthine oxidase-generated superoxyl radicals, the thiol compounds D, L-penicillamine and N-2-mercaptopropionyl glycine reacted at a much slower rate. The O-acetyl derivatives of compounds 1 and 2 were not scavengers of superoxyl radicals. Considerable differences between the α-tocopherol analogs were observed in their competition with 2-deoxyribose for hydroxyl radicals (OH.). Compound 2 was equipotent with Trolox and thiourea, whereas the reactivity of these substances was diminished by more than 30% as compared to compound 1 Although showing lower reactivity, the O-acetyl derivatives of compounds 1 and 2 were active nevertheless as OH-scavengers.

The previously reported high potency of compound 1 in reducing infarct size during myocardial ischemia/reperfusion appears to be due to its radical-scavenging properties, likely to be enhanced by its previously described cardioselectivity.     

Exercise training and oxidative stress in the elderly as measured by antipyrine hydroxylation products

Effects of 12 wk exercise training on oxidative stress were examined in elderly humans. We measured oxidative stress during a 45 min cycling test by using antipyrine hydroxylation products. Antipyrine breakdown is independent of blood flow to the liver, which is important during exercise. Furthermore, antipyrine reacts quickly with hydroxyl radicals to form para- and ortho-hydroxyantipyrine. Ortho-hydroxyantipyrine is not formed in man through the mono-oxygenase pathway of cytochrome P450. Twenty subjects (9 women; 60 ± 3 y) participated in the training program. Thirteen subjects (5 women; 64 ± 7 y) served as inactive controls. Subjects trained, twice a week for 1h, at a fitness center. After 12 wk, maximal oxygen uptake (p < .005) and workload capacity (p < .001) were only significantly elevated in the training group. After 12 wk, both groups observed no change in the ratios of antipyrine hydroxylates, para- and ortho-hydroxy-antipyrine, to native antipyrine. Furthermore, no differences were observed within or between groups in the exercise-induced increase in the plasma level of thiobarbituric acid reactive species. In conclusion, 12-wk training had no effect on exercise-induced oxidative stress in elderly humans as measured by free radical reaction products of antipyrine. Despite the fact that training in elderly humans improves functional capacity, it appears not to compromise antioxidant defense mechanisms.     

黄鳝清除氧自由基作用的研究

采用化学发光法建立四个活性氧体外模型分析黄鳝粘液、血液、粗多糖清除氧自由基和抑制脂质过氧化作用。结果表明:黄鳝粘液、血液和粗多糖具有清除超氧阴离子自由基(O2·)、羟自由基(·OH)、过氧化氢(H2O2)和抑制脂质过氧化(LPO)作用。清除O2·的IC50分别为5.10±2.68μg/mL、3.62±1.56μg/mL、7.19±1.19μg/mL;清除·OH的IC50分别为5.86±1.54μg/mL、3.36±1.36μg/mL、7.93±0.50μg/mL;清除H2O2的IC50分别为6.91±1.29μg/mL、5.92±0.39μg/mL、8.21±0.61μg/mL;抑制LPO的IC50分别为8.11±0.83μg/mL、6.90±0.51μg/mL、7.62±1.01μg/mL。提示黄鳝血液清除氧自由基作用最明显,粘液次之,最弱为粗多糖。     

Antioxidative property of T-0970, a new ureidophenol derivative

We investigated the antioxidative property of T-0970, a newly synthesized ureidophenol derivative. The inhibitory effect of T-0970 on spontaneous lipid peroxidation in rat brain was 10 times greater than those of well-known antioxidants such as butylhydroxytoluene (BHT), probucol and α-tocopherol. T-0970 also showed dose-dependent free radical scavenging activities in vitro for both superoxide anions and hydroxyl radicals. The radical-scavenging potencies of T-0970 were about 10–30 times stronger than those of BHT. We evaluated the in vivo antioxidative ability of T-0970 in the animal model of acute oxidative tissue injury in rats. Intraperitoneal injection of ferric nitrilotriacetate (Fe/NTA) caused an acute and remarkable increase in the level of thiobarbituric acid-reactive substances (TBARS) in both plasma and the liver, and also resulted in a considerable elevation of the plasma levels of GOT and GPT indicative of hepatic injury. Both oral and intravenous administration of T-0970 dose-dependently depressed these diagnostic parameters. These results indicate that T-0970 may have a therapeutic potential in various diseases associated with oxidative tissue injury.     

4-Mercaptoimidazoles derived from the naturally occurring antioxidant ovothiols 1. Antioxidant properties

4-Mercaptoimidazoles derived from the naturally occurring family of antioxidants, the ovothiols, were assayed for their antioxidant properties. These compounds are powerful HOCI scavengers, more potent than the aliphatic thiol N-acetylcysteine. They react slowly with hydrogen peroxide with second order rate constants of 0.13–0.89 M^-1 s^-1. Scavenging of hydroxyl radical occurs at a diffusion-controlled rate (k = 2.0–5.0 × 10¹⁰ M^-1 s^-1) for the most active compounds, which are also able to inhibit copper-induced LDL peroxidation. The combination of radical scavenging and copper chelating properties may explain the inhibitory effects on LDL peroxidation. Two molecules of mercaptoimidazole can chelate a copper ion and form a square planar complex detected by EPR. Compounds bearing an electron-withdrawing group on position 2 of the imidazole ring are the most potent antioxidant molecules in this series.     

罗汉果不同溶剂提取物抗氧化及清除活性氧自由基作用

以水、甲醇、乙醇和乙酸乙酯为溶剂,对罗汉果干果进行提取,分别采用磷钼酸铵体系、邻苯三酚自氧化体系、Fenton反应体系和卵黄脂质过氧化体系测定各种提取物的总抗氧化性能、超氧阴离子自由基和羟基自由基清除性能及其抗脂质过氧化作用。结果表明,四种溶剂提取物均具较强的抗氧化性和活性氧自由基清除性能,其能力的大小顺序为:乙酸乙酯提取物>水提物>甲醇提取物>乙醇提取物。     

DNA Polymerase Activity Associated with the Membrane Fraction of E. coli

Spheroplasts were disrupted with 0.2% Brij 58 and the separation of intact cells, spheroplasts, disrupted spheroplasts, fragmented membrane, and supernatant was performed on a linear 40~55% sucrose gradient. About half an amount of nucleic acid components was distributed in disrupted spheroplast fractions, while only a small amount of protein components was found in these fractions.

DNA polymerase in the fragmented membrane fraction incorporated ³H-TTP more rapidly than that in the supernatant fraction for the first 5 to 6 min, and then the incorporation rate decreased, while DNA polymerase in the supernatant fraction incorporated ³H-TTP linearly up to 20 min when native DNA was used as a primer. The former required native DNA as a primer and showed little activity towards denatured DNA, while the latter incorporated ³H-TTP at a similar rate to both the primer DNA’s.

DNA polymerase of the fragmented membrane fraction synthesized various sizes of DNA from short to a size of primer when native DNA was used as a primer, while when denatured DNA was used, products were only short. DNA polymerase of the supernatant fraction synthesized various sizes of DNA when both native and denatured DNA’s were used as primers.     

Spin Trapping of Free Radicals Formed During Peroxidation of Sarcolemmal Membranes (SLM)

The generation of free radicals in a superoxide (O₂-)driven Fe⁺³ catalysed reactions with isolated myocytic sarcolemma using electron spin resonance was investigated. Incubation of highly purified canine myocytic sarcolemma in the presence of the spin trap, 2-methyl-2-nitrosopropane (MNP). followed by the addition of dihydroxyfurmarate (DHF) and Fe⁺³-ADP resulted in the generation and detection of radical adducts of this spin trap. Spin trapping of the alkyl radicals with 2-methyl-2-nitrosopropane led to the identification of methyl radical adduct following exposure to DHF/Fe⁺³-ADP. With sarcolemma and the alkyl nitroso compound, the only radical product trapped was the methyl radical formed by β-scission of alkoxyl radical. The participation of hydroperoxide-derived radicals in this system verified that the decomposition of unsaturated hydroperoxy fatty acid does proceed via a free radical mechanism.     

渗透胁迫下稻苗中铁催化的膜脂过氧化作用

在－０．７ＭＰａ渗透胁迫下,水稻幼苗体内和Ｈ２Ｏ２大量产生,Ｆｅ２＋积累,膜脂过氧化作用加剧。水稻幼苗体内Ｆｅ２＋含量与膜脂过氧化产物ＭＤＡ含量呈极显著的正相关。外源Ｆｅ２＋、Ｆｅ３＋、Ｈ２Ｏ２、Ｆｅ２＋＋Ｈ２Ｏ２、ＤＤＴＣ均能刺激膜脂过氧化作用,而铁离子的螯合剂ＤＴＰＡ则有缓解作用。ＯＨ的清除剂苯甲酸钠和甘露醇能明显地抑制渗透胁迫下Ｆｅ２＋催化的膜脂过氧化作用。这都表明渗透胁迫下水稻幼苗体内铁诱导的膜脂过氧化作用主要是由于其催化Ｆｅｎｔｏｎ型Ｈａｂｅｒ－Ｗｅｉｓｓ反应形成ＯＨ所致。     

Role of hydroxyl radical in superoxide induced microsomal lipid peroxidation: Protective effect of anion channel blocker

Treatment of bovine pulmonary artery smooth muscle microsomes with the superoxide radical generating system hypoxanthine plus xanthine oxidase stimulated iron release, hydroxyl radical production and lipid peroxidation. Pretreatment of the microsomes with deferoxamine or dime thy lthiourea markedly inhibited lipid peroxidation, and prevented hydroxyl radical production without appreciably altering iron release. The superoxide radical generating system did not alter the ambient superoxide dismutase activity. However,addition of exogenous superoxide dismutase prevented superoxide radical induced iron release,hydroxyl radical production and lipid peroxidation. Simultaneous treatment of the microsomes with deferoxamine, dimethylthiourea or superoxide dismutase prevented hydroxyl radical production and liqid peroxidation. While deferoxamine or dimethylthiourea did not appreciably alter iron release, superoxide dismutase prevented iron release. However, addition of deferoxamine, dimethylthiourea or superoxide dismutase even 2 min after treatment did not significantly inhibit lipid peroxidation, hydroxyl radical production and iron release. Pretreatment of microsomes with the anion channel blocker 4,4’- dithiocyano 2,′- disulphonic acid stilbine did not cause any discernible change in chemiluminiscence induced by the superoxide radical generating system but markedly inhibited lipid peroxidation without appreciably altering iron release and hydroxial radical production.     

Characterization of dehydromonacolin-MV2 from Monascus purpureus mutant

AIM: Characterization of dehydromonacolin-MV2, a bioactive metabolite isolated from Monascus purpureus mutant (CFR 410-11). METHODS AND RESULTS: Chloroform extract of rice, fermented with a hyperpigment-producing mutant of M. purpureus (CFR 410-11) was found to contain metabolites that inhibited the growth of Bacillus, Pseudomonas and Streptococcus in agar gel diffusion assays. The extract inhibited lipid peroxidation and scavenged 2,2-diphenyl-1-pycrylhydrazyl and hydroxyl radicals. The active compound purified by silica gel column chromatography was characterized by NMR. The carbon, proton and 2D HSQCT assignments identified dehydromonacolin-MV2 as the bioactive metabolite. CONCLUSIONS: Dehydromonacolin-MV2 apparently originated in the mutant by hydroxylation and oxidation of monacolin-J, an intermediate of monacolin biosynthetic pathway. SIGNIFICANCE AND IMPACT OF THE STUDY: Identification of the production of dehydromonacolin-MV2 by M. purpureus mutant (CFR 410-11) is new to literature. Bioactive properties of the compound suggested its pharmaceutical applications.     

衰老叶片和叶绿体中H_2O_2的累积与膜脂过氧化的关系

在自然衰老和ABA处理的叶片和叶绿体中活性氧H_2O_2均比对照明显增高。外加H_2O_2刺激水稻叶绿体膜脂过氧化作用。叶绿体的丙二醛含量随H_2O_2浓度、光照时间、光照强度及叶绿体完整性而变化。AsA、GSH、SOD、甘露醇和过氧化氢酶对外源H_2O_2引起的膜脂过氧化有缓解作用,Fe~(2+)有刺激作用。而H_2O_2对叶绿体过氧化损伤主要是转化为OH之故。     

On the Characteristics of the Visible Chemiluminescence Following Free Radical Lipid Peroxidation

The characteristics of the visible luminescence that follows the lipid peroxidative process were investigated either in the autoxidation of rat brain homogenates or in the azo-bis-arnidinopropane initiated lipid peroxidation of erythrocyte plasma membranes and liver microsomes. In these systems the luminescence decay observed after total inhibition of the lipid peroxidation is not an iron-catalyzed process, and follows a complex kinetics comprising fast and slow components. The slow component of the decay lasts for several hours at 27°C and amounts to nearly half of the total intensity measured prior to the inhibition of the oxidative process by propyl gallate. The addition of thiols (diethyldithiocarbamate, penicillamine or dithiothreitol) to a lipid peroxidizing system inhibits the chain oxidation and catalyzes the dark decomposition of one (or several) of the luminescence precursors, following first order kinetics. The effect of temperature on the slow luminescence decay corresponds to an activation energy of 18.5kcal/mol.     

On the Characteristics of the Visible Chemiluminescence Following Free Radical Lipid Peroxidation

The characteristics of the visible luminescence that follows the lipid peroxidative process were investigated either in the autoxidation of rat brain homogenates or in the azo-bis-arnidinopropane initiated lipid peroxidation of erythrocyte plasma membranes and liver microsomes. In these systems the luminescence decay observed after total inhibition of the lipid peroxidation is not an iron-catalyzed process, and follows a complex kinetics comprising fast and slow components. The slow component of the decay lasts for several hours at 27°C and amounts to nearly half of the total intensity measured prior to the inhibition of the oxidative process by propyl gallate. The addition of thiols (diethyldithiocarbamate, penicillamine or dithiothreitol) to a lipid peroxidizing system inhibits the chain oxidation and catalyzes the dark decomposition of one (or several) of the luminescence precursors, following first order kinetics. The effect of temperature on the slow luminescence decay corresponds to an activation energy of 18.5kcal/mol.     

Antioxidant properties of a North American ginseng extract

A North American ginseng extract (NAGE) containing known principle ginsenosides for Panax quinquefolius was assayed for metal chelation, affinity to scavenge DPPH-stable free radical, and peroxyl (LOO·) and hydroxyl (·OH) free radicals for the purpose of characterizing mechanisms of antioxidant activity. Dissociation constants (Kd) for NAGE to bind transition metals were in the order of Fe²⁺ > Cu²⁺ > Fe³⁺ and corresponded to the affinity to inhibit metal induced lipid peroxidation. In a metal-free linoleic acid emulsion, NAGE exhibited a significant (p 0.05) concentration (0.01-10 mg/mL) dependent mitigation of lipid oxidation as assessed by the ammonium thiocyanate method. Similar results were obtained when NAGE was incubated in a methyl linoleate emulsion containing haemoglobin catalyst and assessed by an oxygen electrode. NAGE also showed strong DPPH radical scavenging activity up to a concentration of 1.6 mg/mL (r² = 0.996). Similar results were obtained for scavenging of both site-specific and non site-specific ·OH, using the deoxyribose assay method. Moreover, NAGE effectively inhibited the non site-specific DNA strand breakage caused by Fenton agents, and suppressed the Fenton induced oxidation of a 66 Kd soluble protein obtained from mouse brain over a concentration range of 2-40 mg/mL. These results indicate that NAGE exhibits effective antioxidant activity in both lipid and aqueous mediums by both chelation of metal ions and scavenging of free radicals.     

Biological activities of 7-dehydrocholesterol-derived oxysterols: implications for Smith-Lemli-Opitz syndrome

Smith-Lemli-Opitz syndrome (SLOS) is a metabolic and developmental disorder caused by mutations in the gene encoding the enzyme 7-dehydrocholesterol reductase (Dhcr7). This reductase catalyzes the last step in cholesterol biosynthesis, and levels of 7-dehydrocholesterol (7-DHC), the substrate for this enzyme, are elevated in SLOS patients as a result of this defect. Our group has previously shown that 7-DHC is extremely prone to free radical autoxidation, and we identified about a dozen different oxysterols formed from oxidation of 7-DHC. We report here that 7-DHC-derived oxysterols reduce cell viability in a dose- and time-dependent manner, some of the compounds showing activity at sub-micromolar concentrations. The reduction of cell survival is caused by a combination of reduced proliferation and induced differentiation of the Neuro2a cells. The complex 7-DHC oxysterol mixture added to control Neuro2a cells also triggers the gene expression changes that were previously identified in Dhcr7-deficient Neuro2a cells. Based on the identification of overlapping gene expression changes in Dhcr7-deficient and 7-DHC oxysterol-treated Neuro2a cells, we hypothesize that some of the pathophysiological findings in the mouse SLOS model and SLOS patients might be due to accumulated 7-DHC oxysterols.     

Improvement of the method to estimate the relative reaction rate constants of hydroxyl radical with polyphenols using ESR spin trap: X-ray irradiation of water with a flowing system

UV-photolysis of hydrogen peroxide is a useful technique to produce hydroxyl radical. However, it is not an appropriate method to estimate the reactivity of polyphenols with hydroxyl radicals because many of the polyphenol derivatives also absorb the UV-light to generate hydroxyl radicals. In this study, X-ray irradiation of water with a flowing system was applied to estimate the reactivity of hydroxyl radicals with polyphenols using electron spin resonance (ESR) spin trap. The obtained relative reaction rates reasonably agreed with previous data by pulse radiolysis. This method will be a useful technique to estimate the reactivity of antioxidants including polyphenols with hydroxyl radicals.     

Recent advances in studies of the mechanisms of microbial degradation of lignins

Major advances in our understanding of the biochemical and enzymological mechanisms of lignin biodegradation have been made in the past three years. Research has principally involved two ligninolytic microorganisms, the white rot fungus Phanerochaete chrysosporium and the actinomycete Streptomyces viridosporus. Research has been centred on attempts to identify the microbial catalysts that mediate lignin decay in these two microbes. Emphasis has been on studies concerned with isolating specific lignin catabolic enzymes and/or reduced forms of oxygen involved in attacking the lignin polymer. The possibility that lignin degradation might be non-enzymatic and mediated by extracellular reduced oxygen species such as hydrogen peroxide (H₂O₂), superoxide (O₂∪c-|_.), hydroxyl radical (·OH) or singlet oxygen (¹O₂) has been investigated with both microorganisms. Using methods which have not always been unequivocal, the question of involvement of reduced oxygen species in lignin degradation by P. chrysosporium has been examined exhaustively. Evidence for the involvement of H₂O₂ is conclusive. However, there is little evidence to support the involvement of other extracellular reduced oxygen species, including ·OH, directly in the process of lignin degradation. Scavenger studies have been inconclusive because of questions of their specificity. If activated oxygen species are involved, the activated oxygen is probably held within the active site of an enzyme molecule. With S. viridosporus, scavenger studies also strongly indicate that extracellular reduced oxygen species are not involved in lignin degradation since scavengers generally do not significantly affect the ligninolytic system. The involvement of specific enzymes in lignin degradation by both P. chrysosporium and S. viridosporus has now been confirmed. With P. chrysosporium, ligninolytic enzymes recently discovered include extracellular non-specific peroxidases and oxygenases. They show numerous activities including dehydrogenative, peroxidatic, oxygenative and C_α?C_β cleavages of lignin side chains. At least one P. chrysosporium enzyme, a unique H₂O₂-requiring oxygenase, has been purified to homogeneity. Evidence has been presented to show that S. viridosporus also produces a ligninolytic enzyme complex involved in demethylation of lignin's aromatic rings and in the oxidation of lignin side chains and cleavage of β-tether linkages within the polymer. The combined activites of these enzymes generate water-soluble polymeric modified lignin fragments, which are then slowly degraded further by S. viridosporus. The β-ether cleaving enzyme complex is probably membrane associated, but it is not extracellular. These first isolations of ligninolytic enzymes have changed the course of basic research on lignin biodegradation. New research priorities are already emerging and include enzyme purifications, kinetic studies, enzyme reaction mechanism studies and screenings for more enzymes. In addition, genetic studies are being carried out with both P. chrysosporium and S. viridosporus. Genetic manipulations include not only classical mutagenesis techniques, but also recombinant DNA techniques such as protoplast fusion. This latter technique has already been used to generate overproducers of the ligninolytic enzyme complex in S. viridosporus and it has been successfully used to recombine mutant strains of P. chrysosporium.     

Chilling-enhanced photooxidation: The production,action and study of reactive oxygen species produced during chilling in the light

Chilling-enhanced photooxidation is the light- and oxygen-dependent bleaching of photosynthetic pigments that occurs upon the exposure of chilling-sensitive plants to temperatures below approximately 10 °C. The oxidants responsible for the bleaching are the reactive oxygen species (ROS) singlet oxygen (¹O₂), superoxide anion radical (O ₂ ,hydrogen peroxide (H₂O₂), the hydroxyl radical (OH·), and the monodehydroascorbate radical (MDA) which are generated by a leakage of absorbed light energy from the photosynthetic electron transport chain. Cold temperatures slow the energy-consuming Calvin-Benson Cycle enzymes more than the energy-transducing light reactions, thus causing leakage of energy to oxygen. ROS and MDA are removed, in part, by the action of antioxidant enzymes of the Halliwell/Foyer/Asada Cycle. Chloroplasts also contain high levels of both lipid- and water-soluble antioxidants that act alone or in concert with the HFA Cycle enzymes to scavenge ROS. The ability of chilling-resistant plants to maintain active HFA Cycle enzymes and adequate levels of antioxidants in the cold and light contributes to their ability to resist chilling-enhanced photooxidation. The absence of this ability in chilling-sensitive species makes them susceptible to chilling-enhanced photooxidation. Chloroplasts may reduce the generation of ROS by dissipating the absorbed energy through a number of quenching mechanisms involving zeaxanthin formation, state changes and the increased usage of reducing equivalents by other anabolic pathways found in the stroma. During chilling in the light, ROS produced in chilling-sensitive plants lower the redox potential of the chloroplast stroma to such a degree that reductively-activated regulatory enzymes of the Calvin Cycle, sedohepulose 1,7 bisphosphatase (EC 3.1.3.37) and fructose 1,6 bisphosphatase (EC 3.1.3.11), are oxidatively inhibited. This inhibition is reversible in vitro with a DTT treatment indicating that the enzymes themselves are not permanently damaged. The inhibition of SBPase and FBPase may fully explain the inhibition in whole leaf gas exchange seen upon the rewarming of chilling-sensitive plants chilled in the light. Methods for the study of ROS in chilling-enhanced photooxidation and challenges for the future are discussed.Abbreviations ASP ascorbate-specific peroxidase - -TH reduced -tocopherol - DTT dithiothreitol - FBP fructose 1,6 bisphosphate - FBPase fructose 1,6 bisphosphatase (EC 3.1.3.11) - HFA Cycle the Halliwell/Foyer/Asada Cycle responsible for the enzymatic removal of ROS in the chloroplast stroma - MDA monodehydroascorbate radical - MDAR monodehydroascorbate reductase - ROS reactive oxygen species - SBP sedohepulose 1,7 bisphosphate - SBPase sedohepulose 1,7 bisphosphatase (EC 3.1.3.37) - SOD superoxide dismutase