Hydrogen peroxide pretreatment alters the activity of antioxidant enzymes and protects chloroplast ultrastructure in heat-stressed cucumber leaves

To elucidate the physiological mechanism of heat stress mitigated by exogenous hydrogen peroxide (H₂O₂) further, seedlings of Cucumis sativus cv Lvfeng no. 6 were subjected to two temperatures (42/38 and 25/18 °C) after pretreatment with 1.5 mM H₂O₂. We investigated whether exogenous H₂O₂ could protect chloroplast ultrastructure under heat stress (42/38 °C) and whether the protective effect was associated with the regulation of antioxidant enzymes. The heat condition disintegrated the membranes of 71.4% chloroplasts in the leaf cells and resulted in the elevated levels of malondialdehyde (MDA) and endogenous H₂O₂. When H₂O₂ pretreatment was combined with the heat stress, the abnormal chloroplasts occurred at a rate of 33.3%, and the contents of MDA and endogenous H₂O₂ were decreased. Heat stress and exogenous H₂O₂ both increased the activities of antioxidant enzymes such as glutathione peroxidase (GSH-Px, EC 1.11.1.9), dehydroascorbate reductase (DHAR, EC 1.8.5.1), monodehydroascorbate reducatase (MDHAR, EC 1.6.5.4), and glutathione reductase (GR, EC 1.6.4.2). The combined effect of heat stress and H₂O₂ pretreatment led to higher activity of antioxidant enzymes including superoxide dismutase (SOD, EC 1.15.1.1), ascorbate peroxidase (APX, EC 1.11.1.11), GSH-Px, DHAR, MDHAR and GR in comparison to the heat treatment alone. We propose that exogenous H₂O₂ increases antioxidant enzyme activities in cucumber leaves, decreases lipid peroxidation, and thus protects the ultrastructure of chloroplasts under heat stress.     

Stimulation of ROS-scavenging systems in squash (Cucurbita pepo L.) plants by compost supplementation under normal and low temperature conditions

The beneficial effect of compost, the final product of aerobic biodegradation of organic matter, on growth, lipid peroxidation [as malondialdehyde (MDA], hydrogen peroxide (H₂O₂) and superoxide anion (O₂^•−), activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX) and glutathione reductase (GR), as well as reduced ascorbate (ASC) and glutathione (GSH) and their oxidized forms was investigated in squash (Cucurbita pepo L. cv. Eskandarany) plants exposed to normal and low temperature (LT) conditions. LT stress of 8 °C significantly reduced the plant growth of untreated plants, but compost alleviated the adverse effect of stress and significantly increased the fresh and dry weights under normal and stress conditions. LT also induced accumulation of H₂O₂ and O₂^•− and resulted in increased lipid peroxidation, pointing out to cellular oxidative stress. Under compost application, such reactive oxygen species (ROS) and peroxidized lipids were markedly reduced, but SOD, CAT, APX and GR activities, key enzymes of ROS-scavenging systems, were significantly increased. Data also indicated that there were general reductions in total ascorbate and glutathione pool in LT control plants, but compost-treated ones considerably have maintained higher levels of such redox metabolites. Significantly higher ratios of ASC/DHA (dehydroascorbate) and GSH/GSSG (glutathione disulfide) were generally found in compost-treated plants than in untreated-ones. It is evident that compost induced enhancement of LT tolerance was related to up-regulation of enzymatic and non-enzymatic antioxidant systems. Such enhancement would eventually protect plant cells from LT-induced oxidative stress reactions via scavenging ROS.     

Induction of disease resistance and ROS metabolism in navel oranges by chitosan

The objective of this work was to evaluate how disease resistance and reactive oxygen species (ROS) metabolism in harvested navel oranges (Citrus sinensis L. Osbeck) may be affected by chitosan. Fresh navel oranges were treated with 2% chitosan or 0.5% glacial acetic acid (control) solution for 1 min, and some were inoculated with Penicillium italicum and Penicillium digitatum. Then, the fruit were stored at 20 °C and 85–95% RH. Treatment with 2% chitosan significantly reduced the disease incidence and the lesion diameter compared with control fruit. This treatment effectively enhanced the activities of peroxidase (POD) and superoxide dismutase (SOD), and levels of glutathione (GSH) and hydrogen peroxide (H₂O₂), inhibited the activities of catalase (CAT) and the decreases of ascorbate (AsA) content during navel orange fruits storage. Ascorbate peroxidase (APX) activity in the navel orange fruit was induced slightly by the chitosan treatment during 14–21 days storage. However, glutathione reductase (GR) activity in the fruit was not enhanced by the chitosan treatment. These results indicated that chitosan treatment could induce the navel orange fruit disease resistance by regulating the H₂O₂ levels, antioxidant enzyme and ascorbate–glutathione cycle.     

外源过氧化氢对干旱胁迫下温室黄瓜叶绿体超微结构和抗氧化酶的影响

 以温室黄瓜品种‘春光2号’为试材, 用100 g·L ^{- 1}聚乙二醇处理模拟干旱胁迫, 研究外源过氧化氢是否通过提高叶片抗氧化酶的活性来减轻干旱胁迫对叶绿体膜的伤害。结果表明: 干旱胁迫处理48 h后, 叶绿体肿胀变圆, 叶绿体膜和类囊体模糊不清。与干旱胁迫前没有经过预处理的幼苗相比, 经外源过氧化氢预处理后再干旱胁迫48 h的幼苗中大部分的叶绿体结构正常, 叶片的超氧化物歧化酶、过氧化氢酶、愈创木酚过氧化物酶、抗坏血酸过氧化物酶、单脱氢抗坏血酸还原酶、脱氢抗坏血酸还原酶和谷胱苷肽还原酶等抗氧化物酶的活性以及抗坏血酸和还原型谷胱苷肽的含量明显增高, 内源过氧化氢含量、超氧阴离子的产生速率和丙二醛的含量则显著降低。所以, 外源过氧化氢预处理能够提高抗氧化物的活性,降低活性氧的积累, 从而缓解干旱对黄瓜叶绿体膜的伤害。     

Effect of brassinosteroids on drought resistance and abscisic acid concentration in tomato under water stress

The effect of brassinosteroid (BR) on relative water content (RWC), stomatal conductance (g_s), net photosynthetic rate (PN), intercellular CO₂ concentration (Ci), lipid peroxidation level, activities of antioxidant enzymes and abscisic acid concentration (ABA) in tomato (Lycopersicon esculentum) seedlings under water stress was investigated. Two tomato genotypes, Mill. cv. Ailsa Craig (AC) and its ABA-deficient mutant notabilis (not), were used. Water stress was achieved by withholding water and both the AC and not plants were treated with 1 μM 24-epibrassinolide (EBR) or distilled water as a control. The RWC, g_s, Ci and PN were significantly decreased under water stress. However, EBR treatment significantly alleviated water stress and increased the RWC and PN. EBR application also markedly increased the activities of antioxidant enzymes (catalase, ascorbate peroxidase and superoxide dismutase) while it decreased g_s, Ci and the contents of H₂O₂ and malondialdehyde (MDA). Interestingly, ABA concentration in AC and not plants was markedly elevated after EBR treatment although the increasing rate and amplitude of ABA in not plants treated by EBR was significantly lower than those in AC plants. Our study suggested that amelioration of the drought stress of tomato seedlings may be caused by EBR-induced elevation of endogenous ABA concentration and/or the activities of antioxidant enzymes.     

Photosynthetic characteristics and protective mechanisms against photooxidation during high temperature stress in two citrus species

Exposure of Satsuma mandarin (Citrus unshiu Marc.) and Navel orange (Citrus sinensis Osbeck) plants to high temperature (38 °C) led to reductions of the net photosynthetic rate (Pn), the photorespiration rate (Pr), the quantum efficiency CO₂ assimilation (ΦCO₂${\Phi }_{\text{C}{\text{O}}_2}$), the maximal photochemical efficiency of PS2 (Fv/Fm), the photochemical quenching (qP) and the quantum efficiency of PS2 photochemistry (Φ_PS2), whereas the minimal fluorescence yield (Fo) and the non-photochemical quenching (qN) increased. Increase in the value of Pr/Pn and ΦPS₂/ΦCO₂${\Phi }_{\text{PS}2}/{\Phi }_{\text{C}{\text{O}}_2}$ was attributed to the greater decrease in Pn and ΦCO₂${\Phi }_{\text{C}{\text{O}}_2}$ than Pr and Φ_PS2. In addition, the superoxide radical (O₂⁻) production, the H₂O₂ concentration and the activities of antioxidant enzymes such as the superoxide dismutase (SOD, EC 1.15.1.1), the ascorbate peroxidase (APX, EC 1.11.1.11), the dehydroascorbate reductase (DHAR, EC 1.8.5.1) and the catalase (CAT, EC 1.11.1.6) were raised. On the other hand, the chlorophyll concentration in leaves decreased during high temperature stress. These results suggest that decline in Pn related to inactivation of PS2 reaction centers may be due to the enhanced number of active oxygen species in the citrus leaves. The water–water cycle may play a role in limiting the degree of photodamage caused by high temperature. Lower O₂⁻ production rate, the H₂O₂ concentration and the antioxidant enzymes activity were observed in high temperature tolerant species of citrus. The exogenous active oxygen scavenger ascorbic acid (Asc) enhanced the ability to clear the O₂⁻ in citrus plants, and quicken the recovery of photosynthetic apparatus.     

Changes in antioxidative system and membrane damage of lettuce in response to salinity and boron toxicity

Individual and combined effects of salinity and B toxicity on growth, the major antioxidant enzymes (superoxide dismutase, SOD; catalase, CAT; ascorbate peroxidase, APX) activities, ascorbic acid, proline, and H₂O₂ accumulation, and stomatal resistance (SR), malondialdehyde (MDA), membrane permeability (MP) and the concentrations of sodium (Na), chloride (Cl) and boron (B) of lettuce were investigated. Boron toxicity and salinity reduced growth of lettuce plants. Under B toxicity, B concentration of the plants was increased, but in the presence of NaCl, the concentration of B was significantly reduced. Sodium and Cl concentrations were increased in B + NaCl and NaCl treatments. Membrane damage was more pronounced in NaCl and B + NaCl treatments. Stomatal resistance of the plants was significantly increased by salinity treatments. The accumulation of proline and ascorbic acid was the highest in the B + NaCl treatment. In general, stress conditions significantly increased H₂O₂ and antioxidant enzyme (SOD, CAT and APX) activities. The present results indicate that stomatal closure is an important response of lettuce against NaCl and B + NaCl stress. Furthermore NaCl and B + NaCl toxicity-induced oxidative stress in lettuce resulting in lipid peroxidation and membrane damage. Increased antioxidant enzyme activities and also accumulation of ascorbic acid and proline are involved in order to overcome B- and NaCl-induced oxidative stress.     

Magnesium deficiency induced oxidative stress and antioxidant responses in mulberry plants

The aim of the study was to implicate induction of oxidative stress and antioxidative responses with the effects of Mg deficiency in mulberry plants. Mulberry (Morus alba L.) cv. Kanva-2 plants grown in hydroponics were subjected to deficiency of Mg. Mg-deficient plants developed visible symptoms—deep interveinal chlorotic mottling and necrosis in the older and middle leaves. The decreases in the dry matter yield of plants and concentrations of sugars and starch in the leaves of Mg-deficient plants are suggestive of decreased photosynthetic activity. Mg-deficiency decreased concentrations of photosynthetic pigments, and increased concentrations of H₂O₂ and ascorbate and activities of antioxidative enzymes—peroxidase (POD), ascorbate peroxidase (APX) and superoxide dismutase (SOD). The results suggest induction of oxidative stress by enhancing generation of ROS and inducing alterations in redox status, accompanied by activation of antioxidant machinery including induction of some new SOD isoforms in Mg-deficient mulberry plants. Despite significant increase in H₂O₂, lipid peroxidation was decreased in Mg-deficient plants.     

Ascorbate levels and the activity of key enzymes in ascorbate biosynthesis and recycling in the leaves of 22 Chinese persimmon cultivars

The objective of this study was to determine diversities of ascorbic acid (AsA) and activities of enzymes involved in AsA metabolism, and to determine their relationships to AsA content in the leaves of Chinese persimmon (Diospyros kaki) cultivars. Results showed that there were huge differences not only in the contents of AsA and glutathione (GSH) but also in the enzyme activities of l-galactono-1,4-lactone dehydrogenase (GLDH, EC 1.3.2.3), dehydroascorbate reductase (DHAR, EC 1.8.5.1), monodehydroascorbate reductase (MDHAR, EC 1.6.5.4), ascorbate peroxidase (APX, EC 1.11.1.11) and glutathione reductase (GR, EC 1.6.4.2) in the leaves of 22 Chinese persimmon cultivars. Control of AsA levels mainly depended on the capacity for biosynthesis and GLDH activity played a main role in determining AsA levels in the leaves of persimmon. Both MDHAR and DHAR, MDHAR in particular, also played an important role in maintaining AsA redox state by reducing oxidized AsA back to AsA. But contents of GSH and hydrogen peroide (H₂O₂) and activity of APX were not significantly correlated with AsA content in the leaves of persimmon.     

Effect of silicon on antioxidant and stomatal response of two grapevine (Vitis vinifera L.) rootstocks grown in boron toxic,saline and boron toxic-saline soil

We investigated individual and combined effects of B toxicity and salinity in the presence or absence of silicon on the shoot growth, concentrations of sodium (Na), chloride (Cl), boron (B) and silicon (Si), and stomatal resistance (SR), lipid peroxidation (MDA), proline accumulation, H₂O₂ accumulation and the activities of major antioxidant enzymes (superoxide dismutase, SOD; catalase, CAT and ascorbate peroxidase, APX) activity grapevine rootstocks of 41B (V. Vinifera × V. Berlandieri) and 1103P (V. Berlandieri × V. Rupestris). Applied Si counteracted the deleterious effects of salinity and boron toxicity on shoot growth by lowering the accumulation of Na in 1103P, and B and Cl in the both rootstocks. Stomatal resistance, MDA, and the concentrations of H₂O₂ and proline were higher in the plants grown in conditions of B toxicity, salinity and their combination while applied Si lowered these parameters. Lowering SOD and CAT but increasing APX, Si treatment significantly affected the enzyme activities of both rootstocks. Based on the present work, it can be concluded that Si alleviates the adverse effects of salinity, B toxicity and combined salinity-B toxicity on grapevine rootstocks by preventing both oxidative membrane damage and translocation of Na and B from root to shoots and/or soil to plant, and also lowering the phytotoxic effects of Na and B within plant tissues. When considering the antioxidative response and membrane systems, it was concluded that the rootstock 1103P was responsive to Si under stress conditions. To our knowledge, this is the first report that Si improves the combined salt and B tolerance of grapevine grown under saline, B toxic, and B toxic and saline conditions which describes membrane related parameters and antioxidant responses.     

Antioxidative activity associated with chilling injury tolerance of muskmelon (Cucumis melo L.) rind

‘Galia’ melon (Cucumis melo L. var. reticulatus) is a climacteric fruit with a short storage life. To prevent over-ripening, fruits are stored at 4–6 °C, which also results in chilling injury (CI) that appears as numerous brown spots on the surface of the fruit. It has been suggested that exposure of ‘Galia’ fruits to high growth temperatures in the field renders them sensitive to low storage temperatures. Activity profile of the antioxidative enzymes catalase (CAT), superoxide dismutase (SOD), peroxidase (POD), ascorbate peroxidase (APX) and glutathione reductase (GR) was monitored in the rind of ‘Galia’ fruits during low-temperature storage, to associate each enzyme's activity with CI incidence. Experiments were performed with low-temperature-sensitive and tolerant varieties, 5080^S and 1537^T respectively, and included a pre-storage treatment of hot-water wash of 5080^S fruits.     

Induction of resistance in Yali pear (Pyrus bretschneideri Rehd.) fruit against postharvest diseases by acibenzolar-S-methyl sprays on trees during fruit growth

The objective of this work was to evaluate how disease resistance in harvested fruit of Yali pears (Pyrus bretschneideri Rehd.) may be affected by acibenzolar-S-methyl (ASM) sprays on trees. Results showed that disease incidence and lesion diameter in mature pears from trees sprayed with ASM for three times during growth and inoculated with Penicillium expansum or Alternaria alternata after harvest were 27.9 and 42.7%, or 29.1 and 23.4% lower, respectively, than in control fruit 17 days after inoculation. Mature fruit from ASM-treated trees exhibited higher activities of defense enzymes including phenylalanine ammonia-lyase (PAL), chitinase and β-1,3-glucanase than in control at harvest. In young pears, activities of peroxidase, polyphenoloxidase, PAL, chitinase and β-1,3-glucanase were significantly enhanced by ASM after first spray on trees. The ASM spray also significantly increased H₂O₂ level and glutathione reductase activity, but reduced activities of catalase and ascorbate peroxidase in young pears. The study indicated that enhancement of disease resistance in harvested Yali pear fruit could be the result of multiple effects of several factors related to plant defenses induced by ASM sprays on trees during fruit growth. Application of ASM in the field holds great promise for controlling postharvest diseases of the fruit.     

BTH-mediated antioxidant system responses in apple leaf tissues

BTH (S-methylbenzo-1,2,3-thiadiazole-7-carbothiate), an active compound of the commercial preparation Bion, has been studied as an elicitor of resistance to fire blight (Erwinia amylovora) in apple. However, the biochemical mechanisms of its action are not fully elucidated. Our study indicated that BTH at the best time of its protection activity (2–14 days after application) induced changes in prooxidant–antioxidant balance in the leaves of apple trees, but in different ways in the enzymatic and nonenzymatic antioxidants. Glutathione as low molecular antioxidant as well as superoxide anion radical and lipid peroxides as oxidants exhibited changes at the early phase of BTH action. Glutathione-dependent enzymes were strongly affected by the elicitor used. On the 2nd day glutathione transferase (GST) and glutathione peroxidase (GSH-Px) activities increased by about 70% and 30% above the control, respectively. GST activity normalized about the 14th day but GSH-Px at the same time showed 27% of the control value. Among enzymes utilising hydrogen peroxide only catalase showed increase (37%) at the early phase of experiment. Compared with the control, BTH-treated plants did not show changes in ascorbate peroxidase and phenylalanine ammonia-lyase activities. Tocopherol (TOC) level diminished starting from the 7th day after BTH treatment and on the 14th day it was only 28% of the control. It is proposed that extinguishing of BTH-mediated signal resulted from TOC and glutathione action. The diminished ascorbate level at all examined times may play a crucial role in BTH-mediated cell growth regulation. The direct influence of BTH on lipid metabolism should be also taken into consideration.     

Changes in oxidative damage, antioxidant enzyme activities and polyamine contents in leaves of grafted and non-grafted eggplant seedlings under stress by excess of calcium nitrate

The effect of 80 mmol L⁻¹ stress by excess of calcium nitrate [Ca(NO₃)₂] on biomass production, oxidative damage, antioxidant enzymes activities and polyamine contents in leaves of grafted and non-grafted eggplant (Solanum melongena L.) seedlings were studied, in which grafted plants were grafted on a salinity tolerant rootstock (Solanum torvum Swartz). The results showed that on the 15th day of treatment, the biomass production reduction of non-grafted seedlings was significantly higher than that of grafted seedlings. Under stress by excess of Ca(NO₃)₂, superoxide anion radical (O₂⁻) producing rate, electrolyte leakage percentage, contents of hydrogen peroxide (H₂O₂) and malondialdehyde (MDA) of non-grafted seedlings were significantly higher than those of grafted seedlings, but activities of superoxide dismutase (SOD, EC 1.15.1.1), peroxidase (POD, EC 1.11.1.7), ascorbate peroxidase (APX, EC 1.11.1.11) and glutathione reductase (GR, EC 1.6.4.2) of grafted seedlings were significantly higher than those of non-grafted seedlings, moreover, contents of free, soluble conjugated and insoluble bound polyamines of grafted seedlings were significantly higher than those of non-grafted seedlings, and activities of diamine oxidase (DAO, EC 1.4.3.6) and polyamine oxidase (PAO, EC 1.5.3.3) of grafted seedlings were significantly lower than those of non-grafted seedlings. The possible roles of antioxidant enzymes and polyamines in protective mechanism of grafted eggplant seedlings to stress by excess of Ca(NO₃)₂ were discussed.     

Molecular cloning and characterization of a cucumber MAP kinase gene in response to excess NO3 and other abiotic stresses

Mitogen-activated protein kinases (MAPKs) play important roles in the transduction of extracellular signals to the intracellular targets in all eukaryotes. Here, a cucumber cDNA designated CsNMAPK, encoding a mitogen-activated protein kinase was isolated using RT-PCR, 3′ and 5′ RACE. The full-length cDNA sequence contains 1636 bp and an open reading frame (ORF) of 1113 bp, which encodes 370 amino acid residues. According to the phylogenetic analysis, CsNMAPK belongs to subgroup I MAPK in plants. Northern blot analysis revealed that CsNMAPK expressed differently in response to excess NO₃⁻. And the CsNMAPK expression kinetics between a salt-resistant cultivar (Xintaimici) and a salt-sensitive cultivar (Shennongchunwu) was slightly different under 182 mmol L⁻¹ NO₃⁻ treatment. The mRNA levels also increased after 24 h treatments with H₂O₂ and salicylic acid (SA), but decreased with abscisic acid (ABA) and low-temperature. However, there was no significant induction of CsNMAPK gene after 24 h drought and high-temperature treatments. Our results suggested that a MAP kinase cascade may function in excess NO₃⁻ and other abiotic stresses in cucumber.     

Effect of arbuscular mycorrhizal fungi on growth,mineral nutrition,antioxidant enzymes activity and fruit yield of tomato grown under salinity stress

The purpose of this study was to investigate the mechanisms underlying alleviation of salt stress by arbuscular mycorrhizal fungi Glomus mosseae. Tomato (Lycopersicon esculentum L. cv. Zhongzha105) plants were cultivated in soil with 0, 50 and 100 mM NaCl. Mycorrhization alleviated salt induced reduction of root colonization, growth, leaf area, chlorophyll content, fruit fresh weight and fruit yield. The concentrations of P and K were higher in AM compared with nonAM plants grown under nonsaline and saline conditions. Na concentration was lower in AM than nonAM plants grown under nonsaline and saline conditions. AMF colonization was accompanied by an enhancement of activity of superoxide dismutase (SOD), catalase (CAT), peroxidase (POD) and ascorbate peroxidase (APX) in leaves of both salt-affected and control plants. In addition, inoculation with AMF caused reduction in MDA content in comparison to salinized plants, indicating lower oxidative damage in the colonized plants.