Changes in Activities of Antioxidant Enzymes and Their Relationship to Genetic and Paclobutrazol-Induced Chilling Tolerance of Maize Seedlings

The potential role of antioxidant enzymes in protecting maize (Zea mays L.) seedlings from chilling injury was examined by analyzing enzyme activities and isozyme profiles of chilling-susceptible (CO 316) and chilling-tolerant (CO 328) inbreds. Leaf superoxide dismutase (SOD) activity in CO 316 was nearly one-half that of CO 328, in which the high activity was maintained during the chilling and postchilling periods. Activity of glutathione reductase (GR) was much higher in roots than in leaves. CO 328 also possessed a new GR isozyme that was absent in roots of CO 316. Ascorbate peroxidase (APX) activity was considerably lower in leaves of CO 328 than in CO 316, and nearly similar in roots. Paclobutrazol treatment of CO 316 induced several changes in the antioxidant enzyme profiles and enhanced their activities, especially those of SOD and APX, along with the induction of chilling tolerance. These results suggest that increased activities of SOD in leaves and GR in roots of CO 328, as well as SOD and APX in leaves and roots of paclobutrazol-treated CO 316, contribute to their enhanced chilling tolerance.     

梨树花芽休眠解除与活性氧代谢的关系

梨树(Pyrus bretschneideri Rehd.)自然休眠和休眠解除时,花芽的活性氧代谢发生变化.O2-·产生速率和H2O2的含量在休眠期间上升,在休眠后期下降.抗氧化系统中SOD活性在自然休眠期呈下降趋势,自然休眠结束活性上升.POD和CAT活性在自然休眠期上升.抗氧化物质AsA和GSH的含量随休眠进行而下降,休眠解除过程中重新升高.APX和GR的活性在休眠期间活性下降,休眠结束活性迅速上升.这些结果表明:花芽的休眠与活性氧的代谢有很大关系.     

Changes in antioxidative enzymes in cucumber cotyledons during natural senescence: comparison with those during dark-induced senescence

Changes in 7 antioxidative enzymes in naturally senescent cotyledons of cucumber ( Cucumis sativus ) were investigated. The activities of superoxide dismutase (SOD; EC 1.15.1.1), catalase (EC 1.11.1.6), dehydroascorbate reductase (EC 1.8.5.1) and glutathione reductase (GR; EC 1.6.4.2) gradually decreased during the progression of senescence, while those of ascorbate peroxidase (APX; EC 1.11.1.11) and guaiacol peroxidase (GPX; EC 1.11.1.7) gradually increased. The activity of monodehydroascorbate reductase (MDAR; EC 1.6.5.4) was not significantly changed. Western blot analysis showed that the protein level of mitochondrial SOD gradually declined. The protein level of catalase transiently decreased and then increased in the later stages of senescence, despite the decrease in its activity. The overall behavior was markedly different from that found in cotyledons of artificially senescing seedlings transferred into darkness; the activities of SOD, catalase, APX, GPX and GR gradually increased.     

Postharvest chilling induces oxidative stress response in the dwarf tomato cultivar Micro-Tom

Oxidative stress is involved in the response of Lycopersicon esculentum fruits (cultivar Micro-Tom) to chilling. Changes in activated oxygen scavenging enzymes, superoxide dismutase (SOD, EC 1.15.1.1), catalase (CAT, EC 1.11.1.6), ascorbate peroxidase (APX, EC 1.11.1.11), and glutathione reductase (GR, EC 1.6.4.2) were examined during ripening after postharvest chilling. Also, lipid peroxidation, respiration, and pigment contents were determined. These parameters were affected by chilling, especially the lycopene content and the respiration rate that showed a high value when the fruits were transferred to higher temperatures. CAT activity increased the day after the fruits were re-warmed, while the activity of GR was higher in the chilled than in the non-chilled green fruits. Lipid peroxidation was more evident at the 'pre-chilled' yellow and red fruits. APX and SOD were not affected by previous chilling in ripening fruits. These results indicate that oxidative stress is generated by conservation at 4°C. The antioxidant response of tomato fruit could be mediated by CAT and GR but not by SOD or APX. Moreover, CAT seemed to respond to the increase in the respiration rate.     

Effect of drought and low light on growth and enzymatic antioxidant system of Picea asperata seedlings

The combined effects of drought and low light on biomass partition, foliar nitrogen concentration, membrane stability and active oxygen species (AOS) and antioxidant system were investigated in dragon spruce (Picea asperata Mast.) seedlings grown at two watering regimes (well-watered, 100% of field capacity and drought, 30% of field capacity) and light availabilities (HL, 100% of full sunlight and low light, 15% of full sunlight). Under high light condition drought not only reduced foliar nitrogen concentration (Nmass) and membrane stability index (MSI) but also significantly increased biomass partitioning to roots, AOS, ascorbic acid (AsA) content and antioxidant enzyme activities including superoxide dismutase (SOD, EC 1.15.1.1), peroxidase (POD, EC 1.11.1.7), catalase (CAT, EC 1.11.1.6), ascorbate peroxidase (APX, EC 1.11.1.11) and glutathione reductase(GR, EC 1.6.4.2). However, no prominently drought-induced differences in biomass partitioning to root, SOD, GR activities, hydrogen peroxide (H₂O₂) and MSI were observed in low light seedlings. On the other hand, significant interaction of drought and low light was found on MSI, the antioxidant enzymes activities (SOD, POD, CAT, APX, GR), H₂O₂ and superoxide radical (O₂ ⁻). These results suggested that seedlings grown at the understory were more sensitive to drought than low light.     

Nickel elicits a fast antioxidant response in Coffea arabica cells.

The antioxidant responses of coffee (Coffea arabica L.) cell suspension cultures to nickel (Ni) were investigated. Ni was very rapidly accumulated in the cells and the accumulation could be directly correlated with the increase of NiCl(2) concentration in the medium. At 0.05 mM NiCl(2) growth was stimulated, but at 0.5 mM NiCl(2), the growth rate was reduced. An indication of alterations in the presence of reactive oxygen species was detected by an increase in lipid peroxidation at 0.5 mM NiCl(2). Catalase (CAT; EC 1.11.1.6), glutathione reductase (GR; EC 1.6.4.2), ascorbate peroxidase (APX; EC 1.11.1.11), guaiacol peroxidase (GOPX; EC 1.11.1.7) and superoxide dismutase (SOD; EC 1.15.1.1) activities were increased, particularly at earlier NiCl(2) exposure times and the activities were higher at 0.5 mM NiCl(2) for most of exposure times tested. Non-denaturing PAGE revealed one CAT isoenzyme, nine SOD isoenzymes and four GR isoenzymes. The SOD isoenzymes were differentially affected by NiCl(2) treatment and one GR isoenzyme was increased by NiCl(2). NiCl(2) at 0.05 mM did not induce lipid peroxidation and the main response appeared to be via the induction of SOD, CAT, GOPX and APX activities for the removal of the reactive oxygen species and through the induction of GR to ensure the availability of reduced glutathione.     

Changes in the antioxidative systems in mitochondria during ripening of pepper fruits

The presence of enzymes of the ascorbate–glutathione cycle was studied in mitochondria purified from green and red pepper (Capsicum annuum L.) fruits. All four enzymes, ascorbate peroxidase (APX; EC 1.11.1.11), monodehydroascorbate reductase (MDHAR; EC 1.6.5.4), dehydroascorbate reductase (DHAR; EC 1.8.5.1) and glutathione reductase (GR; EC 1.6.4.2) were present in the isolated mitochondria of both fruit ripening stages. The activity of the reductive ascorbate–glutathione cycle enzymes (MDHAR, GR and DHAR) was higher in mitochondria isolated from green than from red fruits, while APX and the antioxidative enzyme superoxide dismutase (SOD; EC 1.15.1.1) were higher in the red fruits. The levels of ascorbate and L-galactono-γ-lactone dehydrogenase (GLDH; EC 1.3.2.3) activity were found to be similar in the mitochondria of both fruits. The higher APX and Mn-SOD specific activities in mitochondria from red fruits might play a role in avoiding the accumulation of any activated oxygen species generated in these mitochondria, and suggests an active role for these enzymes during ripening.     

美洲黑杨与大青杨杂种无性系耐寒性的初步研究

以美洲黑杨与大青杨杂种无性系1年生扦插苗为材料,测定其低温胁迫中多个生理指标的变化。结果表明,6个杂种无性系SOD、POD和APX活性在低温胁迫中均表现出先上升后下降的趋势,而电解质渗漏率、活性氧含量、CAT和GR活性保持逐渐升高的趋势。无性系间差异显著,其中DU146和DU147电解质渗漏率和活性氧含量最低,而5种抗氧化酶活性均最高,表明其具有较强的耐寒性;而DU135和DU150电解质渗漏率和活性氧含量最高,而5种抗氧化酶活性均最低,表明其耐寒性差。     

Antioxidative defense system in an upland rice cultivar subjected to increasing intensity of water stress followed by recovery

Rice ( Oryza sativa L.) cv. Tulsi is recommended for Eastern India, for upland ecological cultivation systems where a crop experiences natural cycles of water deficit and water sufficiency, depending upon the monsoon rains. In this experiment, this cultivar was subjected to three cycles of water stress of increasing stress intensity. Each stress cycle was terminated by rewatering the plants for a 48-h period. The level of stress was measured by quantification of H₂O₂. The response of antioxidant metabolites such as ascorbate and glutathione, and enzymes such as superoxide dismutase (SOD, EC 1.15.1.1), catalase (CAT, EC 1.11.1.6), ascorbate peroxidase (APX, EC 1.11.1.11), glutathione reductase (GR, EC 1.6.4.2) and guaiacol peroxidase (POX, EC 1.11.1.7) was analysed in terms of activity and isozyme pattern for each cycle of stress and recovery. The differential response of the antioxidant enzymes with increasing stress intensity followed by recovery, highlight the different role of each in the drought acclimation process of upland rice. SOD and POX activity in stressed plants was higher than the controls in all the three cycles. The second level of stress saw an increase in all the enzymes with APX and GR showing its maximum activity and there was a better management of H₂O₂ levels. There was an induction of a new CAT isoform in stressed plants in the third cycle of water stress. The co-ordinated defense helped the plants to recover in terms of growth on rewatering after stress cycles.     

Antioxidative responses to mercury in the cell cultures of Sesbania drummondii.

The effect of mercury (Hg) on the growth and the response of antioxidative systems have been investigated in Sesbania cell cultures to determine the tolerance limits and the mechanisms of metal (Hg) tolerance in plant cells. Cell cultures of Sesbania were developed in different concentrations (0-50 microM) of mercury. Cultures tolerated Hg up to a concentration of 40 microM and showed an increase in the fresh weight growth by 620% in 3 weeks. The levels of antioxidants: glutathione (GSH) and non-protein thiols (NPSH) and the activities of antioxidative enzymes: superoxide dismutase (SOD, EC 1.15.1.1), ascorbate peroxidase (APX, EC 1.11.1.11) and glutathione reductase (GR, EC 1.6.4.2) were influenced by Hg treatments. The contents of GSH, NPSH and GSH/GSSG ratio increased up to a concentration of 40 muM Hg and then severely declined at 50 microM Hg. The activities of antioxidative enzymes, SOD, APX and GR followed the same trends as antioxidants, first increased up to a concentration of 40 muM Hg and then declined in the presence of 50 microM Hg.     

Genetic mapping of tomato cDNA clones encoding the chloroplastic and the cytosolic isozymes of superoxide dismutase

The isozyme pattern of superoxide dismutase (SOD) in tomato consists of two Cu,Zn isozymes located, respectively, in the chloroplast and in the cytosol, as well as additional isozymes of the Mn or Fe SOD type. We have shown that SOD-1 is the chloroplastic Cu,Zn SOD and is related to cDNA clone T10. Restriction fragment length polymorphism (RFLP) analysis was performed with two cDNA clones representing tomato Cu,Zn-superoxide dismutases. T10, coding for the chloroplast isozyme, was thus mapped to chromosome 11, between marker TG46 and TG108, while clone P31, coding for the cytosolic Cu,Zn SOD isozyme, was mapped to chromosome 1 between TG24 and TG81. SOD is associated with the response of plants to various environmental stresses; the mapping information presented here would permit the demonstration of this association by genetic analysis.     

The activity of antioxidant defence enzymes in the mussel Mytilus galloprovincialis from the Adriatic Sea

The activity of the antioxidant defence enzymes superoxide dismutase (SOD, EC 1.15.1.1), catalase (CAT, EC 1.11.1.6), glutathione peroxidase (GSH-Px, EC 1.11.1.9), glutathione reductase (GR, EC 1.6.4.2) and the phase II biotransformation enzyme glutathione-S-transferase (GST, EC 2.5.1.18) in whole mussels (Mytilus galloprovincialis) were studied. The mussels were collected in winter and in spring at two localities in the Adriatic Sea: Bar Port and Tivat Bay. Our results show that the activities of SOD, GSH-Px and GST were seasonally dependent with higher activities in winter. GR activity was also higher in winter, but only in mussels from Bar Port. In mussels from Tivat Bay, GR activity was lower in winter compared to spring. In addition, a decrease in CAT activity in mussels from Bar Port compared to those from Tivat Bay was found. It can be concluded that seasonal variations should be incorporated into interpretation of biomonitoring studies in mussels.     

Protecting cotton photosynthesis during moderate chilling at high light intensity by increasing chloroplastic antioxidant enzyme activity.

This study examined the effect of increasing chloroplastic superoxide dismutase (SOD), ascorbate peroxidase (APX), or glutathione reductase (GR) activity via plant transformation of cotton on the initial recovery of photosynthesis following exposures to 10 degrees C and high photon flux density (PFD). Growing wild-type or non-expressing segregate plants (controls) and transformants at two PFDs (600 micromol m(-2) s(-1) and full sun) resulted in a range of total antioxidant enzyme activities. Total SOD activities above that for control leaves grown in full sun did not substantially improve the recoveries of CO(2)-saturated photosynthesis, especially for stress treatments lasting more than 1 h, while elevated APX or GR activity did improve recoveries after 1-3 h of the chilling treatment. No synergistic effects were noted when the activities of more than one antioxidant enzyme were elevated in transgenic hybrids. Although these results suggest that the protection of photosynthesis can be realized by reducing either superoxide or H(2)O(2) levels, thereby reducing the possibility of hydroxyl radical formation, the situation is complicated, since elevated APX or GR activity can improve recoveries even when additional SOD activity has no effect. In conclusion, to enhance the protection of photosynthesis using stroma-targeted antioxidant enzymes, enhancing metabolism associated with H(2)O(2) is more effective than enhancing the capacity for superoxide scavenging. Although small, the improvement in the protection of photosynthetic capacity may be sufficient to improve cotton yield in temperate regions with large diurnal temperature fluctuations.     

The influence of low-temperature on the photochemical activity of chloroplasts and activity of antioxidant enzymes in maize leaves

The effects of low growth temperature on the activities of photosystems (PS) 1 and 2 and antioxidant enzymes superoxide dismutase (SOD), ascorbate peroxidase (APX), glutathione reductase (GR) and catalase (CAT) in leaves of various maize inbred and hybrid genotypes (parental lines, F1 hybrids, F2 and backcross generations) were investigated. Considerable decrease of the PS 2 activity (contrary to the activity of PS 1) due to low-temperature stress was observed in the majority of genotypes/generations examined. The GR, APX and SOD activities markedly increased due to chilling, whereas the activity of CAT showed lesser changes which depended on the genotype/generation analysed. The higher susceptibility of the inbred line 2013 to low temperature was transmitted to further generations in case this line was used as the maternal parent. The intraspecific variability in photosynthetic and antioxidant parameters was caused particularly by the dominance (negative or positive), however, the level of the expression of this effect often changed after low-temperature stress and was probably the cause of the increase in the positive F1 heterosis observed in this case. Other genetic effects (e.g. the additivity, and particularly the additive or dominant maternal effects) were also found to contribute to the intraspecific variability in parameters analyzed. The dominant maternal effects possibly played an important role in maintaining positive heterosis in F2 generation.     

Antioxidant defense in the leaves of C3 and C4 plants under salinity stress

The effect of salt stress (50, 100 and 150 m M of NaCl) on the activity of superoxide dismutase (SOD, EC. 1.15.1.1), ascorbate peroxidase (APX, EC. 1.11.1.11), glutathione reductase (GR, EC. 1.6.4.2) enzymes and also on the rate of lipid peroxidation in terms of thiobarbituric acid-reactive substances (TBARS) content and photosynthetic capacity in two wheat (C3 plants) and two maize (C4 plants) varieties was studied. In the non-salined control plants, the antioxidant enzymes activities were significantly higher for maize than for wheat. Adding salt to the nutrient solution increased the level of antioxidants in leaves of both maize and wheat. The first substantial response to salinity was found for SOD on the 2nd day, whereas changes occurred for APX on the 4th day and for GR on the 4th/5th day of salt treatment. Although SOD activity increased considerably more in wheat (C3), it never reached as high levels as in maize (C4) grown in the same treatment combinations. The total increase in APX activity was similar for wheat and maize, whereas GR activity was higher in leaves of maize. Lipid peroxidation analyses showed an increase in TBARS contents in both plants' species grown under salinity that corresponded to the damage that occurred in secondary oxidative stress. However, as a result of advanced antioxidant defense in maize, the TBARS quantities did not elevate to as high level as in wheat. Chlorophyll fluorescence measurements revealed a considerable decrease in the efficiency of PS II and electron-transport chain (ETC). Assimilation rate of CO₂ decreased in both plant groups; however, in C4 maize, we observed a much better capacity to preserve the photosynthetic apparatus against overproduction of ROS. Results suggest that efficient antioxidant defense plays an important role in maize, the C₄ plant, resistance to environmental stresses like salinity or drought.     

Photochemical responses and oxidative stress in two clones of Coffea canephora under water deficit conditions

The effects of water deficit on photochemical parameters and activities of superoxide dismutase (SOD), catalase (CAT) and ascorbate peroxidase (APX), as well as, cellular damages were investigated in two clones of Coffea canephora differing in drought tolerance. After 6 days without irrigation, predawn leaf water potential fell to −3.0 MPa that was accompanied by the suppression of net photosynthesis in both clones. The variable to maximum chlorophyll fluorescence ratio remained unchanged regardless of the imposed treatments. Both clones showed a similar decline (about 25%) in the photochemical quenching coefficient, but only the drought-sensitive clone exhibited an enhancement (31%) of thermal deactivation under water deficit conditions. The quantum yield of electron transport decreased similarly in both genotypes. Under drought conditions, activities of SOD, CAT and APX increased to a greater extent in the drought-tolerant clone than in the drought-sensitive one. This seemed to be matched with higher protection against oxidative stress, as judged from the lower levels of lipid peroxidation and electrolyte leakage in the drought-tolerant clone. Thus, the ability to increase the antioxidant system activity in order to limit cellular damages might be an important attribute linked to the drought tolerance in C. canephora.     

Antioxidative responses of Calendula officinalis under salinity conditions.

To gain a better insight into long-term salt-induced oxidative stress, some physiological parameters in marigold (Calendula officinalis L.) under 0, 50 and 100 mM NaCl were investigated. Salinity affected most of the considered parameters. High salinity caused reduction in growth parameters, lipid peroxidation and hydrogen peroxide accumulation. Under high salinity stress, a decrease in total glutathione and an increase in total ascorbate (AsA + DHA), accompanied with enhanced glutathione reductase (GR, EC 1.6.4.2) and ascorbate peroxidase (APX, EC 1.11.1.11) activities, were observed in leaves. In addition, salinity induced a decrease in superoxide dismutase (SOD, EC 1.15.1.1) and peroxidase (POX, EC 1.11.1.7) activities. The decrease in dehydroascorbate reductase (DHAR, EC 1.8.5.1) and monodehydroascorbate reductase (MDHAR, EC 1.6.5.4) activities suggests that other mechanisms play a major role in the regeneration of reduced ascorbate. The changes in catalase (CAT, EC 1.11.1.6) activities, both in roots and in leaves, may be important in H2O2 homeostasis.     

Exogenous treatment with salicylic acid leads to increased antioxidant capacity in leaves of barley plants exposed to paraquat

Our previous study suggests that salicylic acid mediates tolerance in barley plants to paraquat (Ananieva et al. 2002). To further define the role of SA in paraquat induced responses, we analysed the capacity of the antioxidative defence system by measuring the activities of several antioxidative enzymes: superoxide dismutase (SOD, EC 1.15.1.1), ascorbate peroxidase (APX, EC 1.11.1.11), glutathione reductase (GR, EC 1.6.4.2), dehydroascorbate reductase (DHAR, EC 1.8.5.1), catalase (CAT, EC 1.11.1.6), and guaiacol peroxidase (POX, EC 1.11.1.7). Twelve-day-old barley seedlings were supplied with 500 micromol/L SA or 10 micromol/L Pq via the transpiration stream and kept in the dark for 24 h. Then they were exposed to 100 micromol m(-2) s(-1) PAR and samples were taken 6 h after the light exposure. Treatment of seedlings with 10 micromol/L Pq reduced the activity of APX and GR, did not affect the activity of POX and DHAR but caused over a 40% increase in the activity of CAT. Pre-treatment with 500 micromol/L SA for 24 h in the dark before Pq application increased the activities of the studied enzymes in both the chloroplasts (SOD activity) and the other compartments of the cell (POX, CAT activity). The effect of SA pre-treatment was highly expressed on DHAR and POX activity. The data suggest that SA antagonizes Pq effects, via elicitation of an antioxidative response in barley plants.