The responses of the enzymes related with ascorbate–glutathione cycle during drought stress in apple leaves

To explore the significance of the ascorbate–glutathione cycle under drought stress, the leaves of 2-year-old potted apple (Malus domestica Borkh.) plants were used to investigate the changes of each component of the ascorbate–glutathione cycle as well as the gene expression of dehydroascorbate reductase (DHAR, EC 1.8.5.1), ascorbate peroxidase (APX, EC 1.11.1.11) and glutathione reductase (GR, EC 1.6.4.2) under drought stress. The results showed that the malondialdehyde (MDA) and H₂O₂ concentrations in apple leaves increased during drought stress and began to decrease after re-watering. The contents of total ascorbate, reduced ascorbic acid (AsA), total glutathione and glutathione (GSH) were obviously upregulated in apple leaves when the soil water content was 40–45%. With further increase of the drought level, the contents of the antioxidants and especially redox state of AsA and GSH declined. However, levels of them increased again after re-watering. Moreover, drought stress induced significant increase of the activities of enzymes such as APX, scavenging H₂O₂, and also of monodehydroascorbate reductase (MDHAR, EC 1.6.5.4), DHAR and GR used to regenerate AsA and GSH, especially when the soil water content was above 40–45%. During severe drought stress, activities of the enzymes were decreased and after re-watering increased again. Gene expression of cytoplasmic DHAR, cytoplasmic APX and cytoplasmic GR showed similar changes as the enzyme activities, respectively. The results suggest that the ascorbate–glutathione cycle is up-regulated in response to drought stress, but cannot be regulated at severe drought stress conditions.     

Time course of antioxidant responses of Capsicum annuum subjected to a progressive magnesium deficiency

The effect of magnesium (Mg²⁺)‐deficiency on the antioxidant responses of Capsicum annuum was investigated over a 60‐day period under controlled conditions. This Mg²⁺‐deficiency aimed to mimic the physiological conditions that plants may experience in the field. At each harvest time, five different leaf‐levels (L2 to L6) were distinguished. L2 and L6 correspond to the second and sixth youngest leaves, respectively. The following parameters were determined: Mg²⁺, chlorophyll and protein contents, total and redox pools of ascorbate and glutathione, and the activities of superoxide dismutase, ascorbate peroxidase, dehydroascorbate reductase, and glutathione reductase. Under Mg²⁺‐deficiency, leaf Mg²⁺ contents decreased over time in all leaf‐levels except in the second youngest leaves (L2), where they remained constant at about 0.25% (dry weight basis). Mg²⁺‐deficiency led to an increase in the antioxidant enzyme activities concomitant with an increase in the ascorbate and glutathione pools, whereas total chlorophyll and soluble protein contents decreased. The L2 leaves showed an increase in glutathione reductase activity and in the ascorbate redox state whereas no difference was observed for the other parameters. Superoxide dismutase activities increased in L5 leaves from day 15 and, afterwards, in L3 to L5 leaves, irrespective of Mg²⁺ content. At day 30, glutathione reductase activities increased in L2 to L4 leaves and dehydroascorbate reductase activities in L4 leaves. At day 45, we observed an increase in the ascorbate peroxidase activities in L3 to L5 leaves. At the same time, ascorbate and glutathione pools increased in intermediate leaves, whereas chlorophyll content decreased in L3 and L4 leaves, and protein content decreased in L4 leaves. Results suggest that pepper leaves enhance their defence capacities against oxidative stress by increasing ascorbate more than glutathione synthesis. However, cells showed higher regeneration rates for the glutathione redox state than for the ascorbate redox state.     

Kinetics of the Anti-oxidant Response to Salinity in the Halophyte Cakile maritima

The effects of NaCl stress on the activity of anti-oxidant enzymes （superoxide dismutase, catalase （CAT）, peroxidase （POD）, ascorbate peroxidase （APX）, monodehydroascorbate reductase, dehydroascorbate reductase （DHAR）, and glutathione reductase （GR））, anti-oxidant molecules （ascorbate and glutathione）, and parameters of oxidative stress （malondialdehyde （MDA）, electrolyte leakage, and H2O2 concentrations） were investigated in Cakile maritima, a halophyte frequent along the Tunisian seashore. Seedlings were grown in the presence of salt （100, 200, and 400 mmol/L NaCl）. Plants were harvested periodically over 20 days. Growth was maximal in the presence of 0-100 mmol/L NaCl. At 400 mmol/L NaCl, growth decreased significantly. The salt tolerance of C. maritima, at moderate salinities, was associated with the lowest values of the parameters indicative of oxidative stress, namely the highest activities of POD, CAT, APX, DHAR, and GR and high tissue content of ascorbate and glutathione. However, prolonged exposure to high salinity resulted in a decrease in anti-oxidant activities and high MDA content, electrolyte leakage, and H2O2 concentrations. These results suggest that anti-oxidant systems participate in the tolerance of C. maritima to moderate salinities.     

Kinetics of the Anti-oxidant Response to Salinity in the Halophyte Cakile maritima

The effects of NaCl stress on the activity of anti-oxidant enzymes (superoxide dismutase, catalase (CAT),peroxidase (POD),ascorbate peroxidase (APX), monodehydroascorbate reductase, dehydroascorbate reductase (DHAR), and glutathionereductase (GR)), anti-oxidant molecules (ascorbate and glutathione), and parameters of oxidative stress (malondialdehyde(MDA), electrolyte leakage, and H_2O_2 concentrations) were investigated in Cakile maritima, a halophyte frequent along theTunisian seashore. Seedlings were grown in the presence of salt (100, 200, and 400 mmol/L NaCI). Plants were harvestedperiodically over 20 days. Growth was maximal in the presence of 0-100 mmol/L NaCl. At 400 mmol/L NaCl, growthdecreased significantly. The salt tolerance of C. maritima, at moderate salinities, was associated with the lowest values ofthe parameters indicative of oxidative stress, namely the highest activities of POD, CAT, APX, DHAR, and GR and high tissuecontent of ascorbate and glutathione. However, prolonged exposure to high salinity resulted in a decrease in anti-oxidantactivities and high MDA content, electrolyte leakage, and H_2O_2 concentrations. These results suggest that anti-oxidantsystems participate in the tolerance of C. maritima to moderate salinities.     

Changes in activities of antioxidative system and monoterpene and photochemical efficiency during seasonal leaf senescence in Hevea brasiliensis trees

A variety of ecophysiological parameters were monitored in leaves of Hevea brasiliensis (rubber tree) during seasonal leaf senescence. Higher levels of hydrogen peroxide and malondialdehyde, and lower content of total protein and efficiency of photochemistry of photosystem II (PSII) were observed in the senescent leaves (SL) compared to the mature leaves (ML). A significant decrease in the contents of chlorophyll (Chl) and carotenoids (Car) in SL was also observed, but with increase in ratio of Car/Chl. Moreover, activities of superoxide dismutases, catalase, and glutathione reductase in SL were strongly suppressed. In contrast, the activities of guaiacol peroxidase (POD) and ascorbate peroxidase (APX), and the contents of reduced ascorbate, total ascorbate, reduced glutathione and total glutathione were considerably increased in SL compared to ML. In addition, α-pinene, β-pinene, sabinene and total monoterpene pool in SL were drastically decreased. Taken together, these results indicate that the enhanced activities of POD and APX, and further activation of ascorbate-glutathione cycle conferred an important photoprotection against oxidative stress in senescent leaves of rubber trees. The increased Car/Chl could give the protection against photoxidation as well.     

Influence of salt stress on growth,lipid peroxidation and antioxidative enzyme activity in borage (Borago officinalis L.)

Abstract

The effects of increasing salt concentrations on the growth, electrolyte leakage, lipid peroxidation, and major antioxidant enzyme activities (superoxide dismutase, catalase, ascorbate peroxidase and glutathione reductase) of borage plants were investigated. Plants were grown in half strength of Hoagland nutrient solution added with 0, 25, 50, and 75 mM of NaCl. Most measured parameters were affected by salinity. Increasing salt levels caused a significant reduction in leaf area, stem length, stem diameter, flower number, and dry masses of different organs. Growth of borage plants, in terms of dry weight, was affected. As a consequence of salinity stress, lipid peroxidation and membrane permeability was increased. Antioxidant activity showed an increase in the activity of superoxide dismutase, a non-induced activity of catalase and ascorbate peroxidase, and a slight increase in glutathione reductase activity. The results indicate that borage plants appear to be sensitive to salt stress, since enzymes related to antioxidant enzymatic defense system in treated leaves should be highly active.     

Developmental changes of antioxidative systems in tobacco leaves as affected by limited sucrose export in transgenic plants expressing yeast-invertase in the apoplastic space

It is generally believed that a restricted export of carbohydrates from source leaves causes oxidative stress because of an enhanced utilisation of O₂ instead of NADP⁺ as electron acceptor in photosynthesis. To test this hypothesis, developmental changes of antioxidative systems were investigated in wild-type and transgenic tobacco (Nicotiana tabacum L.) suffering from disturbed sink-source relations by expression of yeast invertase in the apoplastic space. Young expanding leaves of the wild type contained higher activities of Superoxide dismutase (EC 1.15.1.1), ascorbate peroxidase (EC 1.11.1.11), catalase (EC 1.11.1.6), dehydroascorbate reductase (EC 1.8.5.1), glutathione reductase (EC 1.6.4.2) and a higher glutathione content than mature source leaves. The activity of monodehydroascorbate-radical reductase (EC 1.1.5.4) and the ascorbate content remained unaffected by the developmental stage in the wild type. In young expanding leaves of the transgenic plants the capacity of the antioxidative systems was similar to or higher than in corresponding leaves from the wild type. Source leaves of transgenic tobacco with an increased carbohydrate content showed a small chlorophyll loss, an increased malondialdehyde content, a selective loss of the activities of Cu/Zn-superoxide dismutase isoenzymes and a fourfold decrease in ascorbate compared with the wild type. There was no evidence that the protection from H₂O₂ was insufficient since source leaves of transgenic tobacco contained increased activities of catalase, ascorbate peroxidase, and monodehydroascorbate-radical reductase and an increased ascorbate-to-dehydroascorbate ratio compared with source leaves of the wild type. In severely chlorotic leaf sections of the transgenic plants, most components of the antioxidative system were lower than in green leaf sections, but the ascorbate-to-dehydroascorbate ratio was increased. These results suggest that carbohydrate-accumulating cells have an increased availability of reductant, which can increase the degree of reduction of the ascorbate system via glutathione-related systems or via the activity of monodehydroascorbate-radical reductase. At the same time, transgenic tobacco leaves seem to suffer from an increased oxidative stress, presumably as a result of a decreased consumption of O ₂ ^.- by Cu/Zn-superoxide dismutases in the chloroplasts. There was no evidence that carbohydrate-accumulating leaves acclimated to enhanced O ₂ ^.- production rates in the chloroplasts.     

Changes in ascorbate, glutathione, and related enzyme activities during thidiazuron-induced bud break of apple

The changes of ascorbic acid, dehydroascorbic acid, and glutathione content and related enzyme activities were studied in apple buds during dormancy and thidiazuron-induced bud break. An increase in ascorbic acid, reduced form of glutathione (GSH), total glutathione, total non-protein thiol (NPSH) and non-glutathione thiol (RSH) occurred as a result of induction by thidiazuron during bud break, whereas dehydroascorbic acid and oxidized glutathione (GSSG) decreased during the same period. Thidiazuron also enhanced the ratio of GSH/GSSG, and activities of ascorbate free radical reductase (AFR; EC 1.6.5.4), ascorbate peroxidase (EC 1.11.1.11). dehydroascorbate reductase (DHAR; EC 1.8.5.1) and glutathione reductase (GR; EC 1.6.4.2). The ascorbic acid content and the activities of AFR, ascorbate peroxidase, and DHAR peaked when buds were in the side green or green tip stage just prior to the start of rapid expansion, and declined thereafter. The GSH, NPSH, RSH, ratio of GSH/GSSG, and activities of GR increased steadily during bud development.     

Salinity induced oxidative stress and antioxidant system in salt-tolerant and salt-sensitive cultivars of rice (Oryza sativa L.)

Indices of oxidative stress viz., superoxide radical and H₂O₂ content increased in leaves of all the cultivars with the rise in salinity level, the increase was more pronounced and significant in salt-sensitive varieties and non-significant in resistant cultivars. Except for glutathione reductase (GR), basal activities of all other antioxidative enzymes viz. superoxide dismutase (SOD), catalase (CAT), peroxidase (POX), ascorbate peroxidase (APX) and glutathione reductase (GR) were significantly higher in leaves of all the resistant cultivars as compared to the sensitive ones. A differential response of salinity was observed on various enzymatic and non-enzymatic components of antioxidant system in leaves of salt-tolerant and salt-sensitive cultivars of rice (Oryza sativa L.). Activities of superoxide dismutase and glutathione reductase enhanced in all the tolerant cultivar while declined in the sensitive cultivars with increasing salinity from 0 to 100 mM. Salt-stress induced the activities of catalase and peroxidase in all the cultivars but the magnitude of increase was more pronounced in the sensitive cultivars than in the tolerant cultivars. Contrarily, APX activity increased in the salt-sensitive cultivars but showed no significant change in the salt-tolerant cultivars. The amount of ascorbic acid content, reduced glutathione (GSH), reduced/oxidized glutathione (GSSG) ratio was higher in leaves of the tolerant cultivars than that of the sensitive cultivars under saline conditions. It is inferred that leaves of salt-tolerant cultivars tend to attain greater capacity to perform reactions of antioxidative pathway under saline conditions to combat salinity-induced oxidative stress.     

Effects of exogenous hydrogen sulfide on the ascorbate and glutathione metabolism in wheat seedlings leaves under water stress

This study investigated the effects of exogenous hydrogen sulfide on the ascorbate and glutathione metabolism in wheat seedlings leaves under water stress. The results showed that pretreatment with sodium hydrosulfide (NaHS), hydrogen sulfide donor, increased the activities of ascorbate peroxidase, glutathione reductase, dehydroascorbate reductase and gamma-glutamylcysteine synthetase, and the contents of reduced ascorbic acid, reduced glutathione, total ascorbate and total glutathione under water stress, compared to control and water stress without NaHS. Meanwhile, pretreatment with NaHS decreased the malondialdehyde content and electrolyte leakage induced by water stress in plants, compared to control and water stress without NaHS. Our results suggested that exogenous hydrogen sulfide alleviated oxidative damage by regulating the ascorbate and glutathione metabolism in wheat seedlings under water stress.     

Response of the cultivated tomato and its wild salt-tolerant relative Lycopersicon pennellii to salt-dependent oxidative stress: increased activities of antioxidant enzymes in root plastids

Root plastids of the cultivated tomato Lycopersicon esculentum (Lem) exhibited salt-induced oxidative stress as indicated by the increased H 2 O 2 and lipid peroxidation levels which were accompanied with increased contents of the oxidized forms of ascorbate and glutathione. In contrast, H 2 O 2 level decreased, lipid peroxidation level slightly decreased and the levels of the reduced forms of ascorbate and glutathione increased in plastids of L. pennellii (Lpa) species in response to salinity. This better protection of Lpa root plastids from salt-induced oxidative stress was correlated with increased activities of superoxide dismutase (SOD), ascorbate peroxidase (APX), guaiacol peroxidases (POD), monodehydroascorbate reductase (MDHAR), glutathione peroxidase (GPX), glutathione- S -transferase (GST) and phospholipid hydroperoxide glutathione peroxidase (PHGPX). In the plastids of both species, activities of SOD, APX, and POD could be resolved into several isozymes. In Lem plastids two Cu/ZnSOD isozymes were found whereas in Lpa an additional FeSOD type could also be detected. In response to salinity, activities of selected SOD, APX, and POD isozymes were increased in Lpa, while in Lem plastids the activities of most of SOD and POD isozymes decreased. Taken together, it is suggested that plastids play an important role in the adaptation of Lpa roots to salinity.     

Effects of exogenous hydrogen sulfide on the redox states of ascorbate and glutathione in maize leaves under salt stress

This study investigated the effects of exogenous hydrogen sulfide (H₂S) on the redox states of ascorbate (AsA) and glutathione (GSH) in maize leaves under NaCl (100 mM) stress. Salt stress increased the activities of ascorbate peroxidase (APX), glutathione reductase (GR), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR), Γ-glutamylcysteine synthetase (Γ-ECS), and L-galactono-1,4-lactone dehydrogenase (GalLDH), malondialdehyde content and electrolyte leakage, and reduced the ratios of reduced and oxidised forms of AsA (AsA/DHA) and GSH (GSH/GSSG) compared with control. Pretreatment with NaHS (H₂S donor) further enhanced the activities of the above enzymes except MDHAR and ameliorated the decrease in the ratios of AsA/DHA and GSH/GSSG compared with the salt stress alone. Pretreatment with NaHS significantly reduced the malondialdehyde content and electrolyte leakage induced by the salt stress. Pretreatment with NaHS alone did not affect any of the above mentioned parameters compared with the control. Our results suggest that exogenous H2S could maintain the redox states of ascorbate and glutathione by up-regulating the ascorbate and glutathione metabolism and thus play an important role for acquisition of salt stress tolerance in maize.     

Antioxidant defences of the apoplast

Summary The apoplast of barley and oat leaves contained superoxide dismutase (SOD), catalase, ascorbate peroxidase, dehydroascorbate reductase, monodehydroascorbate reductase, and glutathione reductase activities. The activities of these enzymes in the apoplastic extracts were greatly modified 24 h after inoculation with the biotrophic fungal pathogenBlumeria graminis. The quantum efficiency of photosystem II, which is related to photosynthetic electron transport flux, was comparable in inoculated and healthy leaves during this period. Apoplastic soluble acid invertase activity was also modified in inoculated leaves. Inoculation-dependent increases in apoplastic SOD activity were observed in all lines. Major bands of SOD activity, observed in apoplastic protein extracts by activity staining of gels following isoelectric focusing, were similar to those observed in whole leaves but two additional minor bands were found in the apoplastic fraction. The apoplastic extracts contained substantial amounts of dehydroascorbate (DHA) but little or no glutathione (GSH). Biotic stress decreased apoplastic ascorbate and DHA but increased apoplastic GSH in resistant lines. The antioxidant cycle enzymes may function to remove apoplastic H₂O₂ with ascorbate and GSH derived from the cytoplasm. DHA and oxidized glutathione may be reduced in the apoplast or returned to the cytosol for rereduction.Abbreviations AA reduced ascorbate - APX ascorbate peroxidase - DHA dehydroascorbate (oxidised ascorbate) - DHAR dehydroascorbate reductase - G6PDH glucose-6-phosphate dehydrogenase - GSH reduced glutathione - GSSG glutathione disulphide - GR glutathione reductase - MDHA monodehydroascorbate - MDHAR monodehydroascorbate reductase - SOD superoxide dismutase     

Biphasic effect of copper on the ascorbate-glutathione pathway in primary leaves of Phaseolus vulgaris seedlings during the early stages of metal assimilation

Copper‐imposed oxidative stress and antioxidative defence responses were investigated in the primary leaves of Phaseolus vulgaris L. plants grown on hydroponics containing 50 μM CuSO₄. Copper mainly accumulates in roots; therefore, an increase of the copper content in the leaves was only observed 48 h after the start of the copper supply. Nevertheless, an increase of the thiobarbituric acid reactive metabolites (TBArm) content, an indication of stress, occurred immediately following copper application. Because the ascorbate‐glutathione pathway is considered as a major antioxidative defence mechanism, the evolution of the enzymes and the related metabolites involved in this pathway were studied in the primary leaves as a function of plant copper assimilation. The capacities of monodehydroascorbate reductase (EC 1.6.5.4), dehydroascorbate reductase (EC 1.8.5.1), and glutathione reductase (EC 1.6.4.2) were increased before elevated amounts of copper could be detected in the leaves. The early enhancement of glutathione reductase was only temporary. After copper accumulation in the leaves, a second increase of the glutathione reductase capacity and also an increase of the ascorbate peroxidase capacity (EC 1.11.1.11) were observed. These changes in enzymatic capacity modified the level of the metabolites involved. Increase of the ascorbate pool and maintenance in its reduced form was observed immediately after the start of the treatment. In the beginning of the experiment, the glutathione disulphide/reduced glutathione ratio was higher in the treated plants as compared to the controls. However, towards the end of the experiment, the total glutathione pool, as well as the reduced glutathione content, increased, resulting in a lower ratio value for the treated plants. In conlusion, copper‐imposed oxidative stress, as well as the antioxidative defence response in the leaves, appears to be biphasic. An indirect preventive effect on the antioxidative defence system was observed during the first phase before the leaf copper content increased. A root‐to‐shoot signalling system appears to be involved. Direct oxidation by copper of reduced cell metabolites occurred during the second phase when the leaf copper content was enhanced.     

Cadmium-induced oxidative damage and antioxidant responses in Brassica juncea plants

Indian mustard (Brassica juncea L. cv. Vitasso) plants exposed to 10, 30, 50 and 100 μM of Cd for 5 d in hydroponic culture were analysed with reference to the distribution of Cd²⁺, the accumulation of biomass and antioxidants and antioxidative enzymes in leaves. Cd induced a decrease in plant biomass. The maximum accumulation of Cd occurred in roots followed by stems and leaves. Cd induced a decrease in catalase (CAT) and guiacol peroxidase (GPX) activities but an increase in ascorbate peroxidase (APX) and monodehydroascorbate reductase (MDHAR) activities. Enhancement in dehydroascorbate reductase (DHAR) activity was also at 10 μM Cd. Glutathione reductase (GR) activity showed pronounced stimulation after all treatments, but glutathione S-transferase (GST) and glutathione peroxidase (GPOX) activities decreased. The effectiveness of ascorbate-glutathione cycle (AGC) was determined by the ratio of ascorbate to H₂O₂. This ratio decreased in the Cd-treated leaves which indicated that the cycle was disordered.     

The effect of Botrytis cinerea infection on the antioxidant profile of mitochondria from tomato leaves

Infection of tomato leaves with the necrotrophic fungus Botrytis cinerea resulted in substantial changes in enzymatic and non-enzymatic components of the ascorbate-glutathione cycle as well as in superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), glutathione transferase (GST), and l-galactono-gamma-lactone dehydrogenase (GLDH) activities. In the initial phase of the 5 d experiment CuZn SOD was the most rapidly induced isoform (up to 209% of control), whereas later on its activity increase was not concomitant with the constant total SOD enhancement. Starting from the second day B. cinerea infection diminished the mitochondrial antioxidant capacity by decreasing activities of ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR) as well as declining ascorbate and glutathione contents. This was accompanied by dehydroascorbate (DHA) and oxidized glutathione (GSSG) accumulation that resulted in ascorbate and glutathione redox ratios decreases. The strongest redox ratio decline of 29% for ascorbate and of 34% for glutathione was found on the 3rd and 2nd days, respectively. Glutathione reductase (GR) induction (185% of control 2 d after inoculation) was insufficient to overcome the decreased antioxidant potential of glutathione. Changes in the ascorbate pool size were closely related to the activity of l-galactono-gamma-lactone dehydrogenase (GLDH). The activities of two glutathione-dependent enzymes: GSH-Px and GST were increased from day 1 to day 4. These results demonstrated that in B. cinerea-tomato interaction mitochondria could be one of the main targets for infection-induced oxidative stress.     

Nitric oxide is involved in the regulation of ascorbate and glutathione metabolism in Agropyron cristatum leaves under water stress

This study investigated the regulation of ascorbate and glutathione metabolism by nitric oxide in Agropyron cristatum leaves under water stress. The activities of ascorbate peroxidase (APX), glutathione reductase (GR), monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR), L-galactono-1,4-lactone dehydrogenase (GalLDH) and γ-glutamylcysteine synthetase (γ-ECS), and the contents of NO, reduced ascorbic acid (AsA), reduced glutathione (GSH), total ascorbate and total glutathione increased under water stress. These increases were suppressed by pretreatments with NO synthesis inhibitors N ^G-nitro-L-arginine methyl ester (L-NAME) and 4-carboxyphenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (cPTIO). However, application of L-NAME and cPTIO to plants sufficiently supplied with water did not affect the activities of above mentioned enzymes and the contents of NO and above mentioned antioxidants. Pretreatments with L-NAME and cPTIO increased the malondialdehyde (MDA) content and electrolyte leakage of plants under water stress. Our results suggested that water stress-induced NO is a signal that leads to the upregulation of ascorbate and glutathione metabolism and has important role for acquisition of water stress tolerance.     

Age-dependence of the antioxidative system in tobacco with enhanced glutathione reductase activity or senescence-induced production of cytokinins

Tobacco leaves of plants with enhanced glutathione reductase activity (GR46-27, Nicotiana tabacum L. cv. Samsun) or with autoregulated senescence-induced production of cytokinins (P_SAG12-IPT, N. tabacum L. cv. Wisconsin) were studied during the course of leaf development and senescence by measuring photosynthesis, chlorophyll and protein content, the antioxidants ascorbate, glutathione and α -tocopherol as well as the antioxidative enzymes ascorbate peroxidase (APX, EC 1.11.1.11), glutathione reductase (GR, EC 1.6.4.2) and superoxide dismutase (SOD, EC 1.15.1.1). The photosynthetic rate, as well as the chlorophyll and protein content, dropped with increasing leaf age after having reached a maximum at the end of the exponential growth phase. The concentrations of the water-soluble antioxidants ascorbate and glutathione fell continuously with age, whereas the concentration of the lipophilic α -tocopherol increased. The activities of the antioxidative enzymes APX, GR and SOD reached their maximum at the beginning of leaf development, but were reduced in senescing leaves. The age-dependent course of the measured leaf parameters in GR46-27 leaves was similar to the one in wild-type leaves, with the exception of an overall enhanced GR activity. In contrast, in old leaves of P_SAG12-IPT plants, which possess a much higher life span, the chlorophyll and protein content, the photosynthetic rate, the antioxidant concentrations of ascorbate and glutathione as well as the activities of the antioxidative enzymes were higher than in wild-type leaves. The results show that the capacity of the antioxidative system to scavenge radicals is sufficiently balanced with the plant metabolism, and its decline with increasing age is not the cause, but a consequence of senescence and ageing in plants.     

Synthesis and Secretion of Acid Phosphatase in Halotolerant Zygosaccharomyces rouxii

Changes in the contents of defensive substances against the active oxygen in water-stressed spinach plants were examined. The contents of ascorbate peroxidase (AP; EC 1.11.1.7), glutathione reductase (GR; EC 1.6.4.2) and α-tocopherol increased remarkably in water-stressed spinach leaves, while those of Superoxide dismutase (SOD; EC 1.15.1.1), dehydroascorbate reductase (EC 1.8.5.1), ascorbate and glutathione changed little. The content of α-tocopherol in chloroplast thylakoid membranes isolated from water-stressed leaves was higher than that from normal leaves. It is, therefore, conceivable that GR, AP and α-tocopherol might be related to the tolerance of plants to water deficiency.