Antioxidant potential and indole alkaloid profile variations with water deficits along different parts of two varieties of Catharanthus roseus

The variations in antioxidant potentials and indole alkaloid content were studied in the present investigation, in two varieties (rosea and alba) of Catharanthus roseus, an important herb used in traditional as well as modern medicine, exposed to water deficit stress. The antioxidant and alkaloid profiles were estimated from root, stem, leaf, flowers and pods. The antioxidant potentials were examined in terms of non-enzymatic antioxidant molecules and activities of antioxidant enzymes. The non-enzymatic antioxidant molecules studied were ascorbic acid (AA), -tocopherol (-toc) and reduced glutathione (GSH). The estimated antioxidant enzymes were superoxide dismutase (SOD), ascorbate peroxidase (APX), catalase (CAT), peroxidase (POX) and polyphenol oxidase (PPO). The antioxidant concentrations and activities of antioxidant enzymes were high under water deficit stress in all parts of the plants. Indole alkaloid content was high in the roots of rosea variety in response to stress when compared to alba variety.     

Lipoic Acid Combined with Melatonin Mitigates Oxidative Stress and Promotes Root Formation and Growth in Salt-Stressed Canola Seedlings (Brassica napus L.)

Lipoic acid (LA) and melatonin (MT) are pleiotropic molecules participating in plant stress resistance by modulating cellular biochemical changes, ion homeostasis, and antioxidant enzyme activities. However, the combined role of these two molecules in counteracting the detrimental impacts of salinity stress is still unknown. In the present study, we determined the effects of exogenous LA (0.5 µM), MT (1 µM) and their combination (LA + MT) on growth performance and biomass accumulation, photosynthetic pigments, enzymatic and non-enzymatic antioxidant activities, and ions homeostatic in canola (Brassica napus L.) seedlings under salinity stress (0, 100 mM) for 40 days. The results indicate that exogenous application of LA + MT improved the phenotypic growth (by 25 to 45%), root thickness (by 68%), number of later lateral roots (by 52%), root viability (by 44%), and root length (by 50%) under salinity stress. Moreover, total soluble protein, chlorophyll pigments, the concentration of superoxide dismutase (SOD), catalase peroxidase (CAT), and ascorbic peroxidase (ASA) increased with the presence of salt concentration into the growth media and then decreased with the addition of LA + MT to saline solution. Leaf protein contents and the degradation of photosynthetic pigments were lower when LA + MT treatments were added into NaCl media. The proline and phenol contents decreased in the exogenous application of LA + MT treatments more than individual LA or MT treatments under the salinity stress. The incorporation of LA or MT or a combination of LA + MT to saline solution decreased salinity-induced malondialdehyde and electrolyte leakage. In conclusion, the alteration of metabolic pathways, redox modulation, and ions homeostasis in plant tissues by the combined LA and MT application are helpful towards the adaptation of Brassica napus L. seedlings in a saline environment. The results of this study provide, for the first time, conclusive evidence about the protective role of exogenous LA + MT in canola seedlings under salinity stress.     

Water deficit stress effects on reactive oxygen metabolism in Catharanthus roseus; impacts on ajmalicine accumulation

In the present work, we have analysed the changes in the reactive oxygen metabolism of Catharanthus roseus (L.) G. Don. plants in terms of H(2)O(2) content, lipid peroxidation and the free radical quenching systems (non-enzymatic and enzymatic antioxidants) under drought stress. In addition to this, the root alkaloid ajmalicine was extracted and quantified from both control and drought stressed plants. The H(2)O(2) content was analysed from both stressed and unstressed control plants. Lipid peroxidation was estimated as thiobarbituric acid reactive substances. The non-enzymatic antioxidants viz., ascorbic acid, alpha-tocopherol and reduced glutathione contents, antioxidant enzymes like superoxide dismutase, ascorbate peroxidase, and catalase were extracted and estimated from the samples. The alkaloid ajmalicine was extracted and quantified from shade dried root samples and found significantly increased over control. From the results of this investigation, it can be concluded that the water deficit areas may be well used for the cultivation of medicinal plants like C. roseus and the economically important alkaloid production can be enhanced in the plant level.     

Comparative effects of different triazole compounds on antioxidant metabolism of Solenostemon rotundifolius

The effect of different triazole compounds, viz., triadimefon (TDM) and hexaconazole (HEX) treatments on the antioxidant metabolism of Solenostemon rotundifolius Poir., Morton plants was investigated in the present study under pot culture. Plants were treated with TDM at 15 mg l⁻¹ and HEX at 10 mg l⁻¹ separately by soil drenching on 80, 110 and 140 days after planting (DAP). The plants were harvested randomly on 90, 120 and 150 DAP for determining the effect of both the triazoles on non-enzymatic antioxidant contents like ascorbic acid (AA), reduced glutathione (GSH) and -tocopherol (-toc), activities of antioxidant enzymes like superoxide dismutase (SOD) and ascorbate peroxidase (APX). All the analyses were made in leaf, stem and tubers of both control and treated plants. It was found that both these triazole compounds have profound effects on the antioxidant metabolism and caused an enhancement in both non-enzymatic and enzymatic antioxidant potentials under treatments. These results suggest that, the application of triazoles may be a useful tool to increase the antioxidant production in S. rotundifolius and thereby make it an economical food crop.     

Effect of paclobutrazol on water stress amelioration through antioxidants and free radical scavenging enzymes in Arachis hypogaea L

An investigation was carried out to find out the extent of changes occurred in groundnut (Arachis hypogaea L.) cultivars in response to paclobutrazol (PBZ) treatment under water deficit stress. Two groundnut cultivars namely ICG 221 and ICG 476 were used for the study. Individual treatment with PBZ and drought stress showed an increase in ascorbic acid, -tocopherol and reduced glutathione, superoxide dismutase (SOD), ascorbate peroxidase (APX) and catalase (CAT) activities. PBZ with drought stressed plants maintained higher levels of antioxidant and scavenging enzymes. Significant differences were observed between cultivars and treatments. These results suggests that the adverse effects of water stress can be minimized by the application of PBZ by increasing the antioxidant levels and activities of scavenging enzymes such as SOD, APX and CAT. The Cv. ICG 221 appears to be more tolerant to water stress than the ICG 476.     

Changes in antioxidant metabolism of Vigna unguiculata (L.) Walp. by propiconazole under water deficit stress

In the present study, a pot culture experiment was conducted to estimate the ameliorating effect of propiconazole (PCZ) on drought stress in cowpea (Vigna unguiculata (L.) Walp.) plants. From 30 days after sowing (DAS), the plants were subjected to 3, 6 and 9 days interval drought (DID) stress and drought stress with PCZ at 15 and 15 mg l(-1) PCZ alone and 1 day interval irrigation was kept as control. The plant samples were collected on 34 DAS (3 DID), 37 DAS (6 DID) and 40 DAS (9 DID). The plants were separated into root, stem and leaf for estimating the antioxidant contents and activities of antioxidant enzymes. Individual and combined drought stress and PCZ treatments increased ascorbic acid (AA), alpha-tocopherol (alpha-toc) contents, superoxide dismutase (SOD), ascorbate peroxidase (APX), catalase (CAT) and polyphenol oxidase (PPO) activities when compared to control. The PCZ treatment mitigated the adverse effects of drought stress by increasing the antioxidant potentials and thereby paved the way for overcoming drought stress in V. unguiculata plants.     

Changes of antioxidative enzymes and cell membrane osmosis in tomato colonized by arbuscular Mycorrhizae under NaCl stress

Salinity toxicity is a worldwide agricultural and eco-environmental problem. Many literatures show that arbuscular mycorrhizal fungi (AMF) can enhance salt tolerance of many plants and some physiological changes occurred in AM symbiosis under salt stress. However, the role of ROS-scavenging enzymes in AM tomato is still unknown in continuous salt stress. This study investigated the effect of Glomus mosseae on tomato growth, cell membrane osmosis and examined the antioxidants (superoxide-dismutase, SOD; catalase, CAT; ascorbate peroxidase, APX; peroxidase, POD) responses in roots of mycorrhizal tomato and control under different NaCl stress for 40 days in potted culture. NaCl solution (0, 0.5 and 1%) was added to organic soil in the irrigation water after 45 days inoculated by AMF (Glomus mosseae). (1) AMF inoculation improved tomato growth under salt or saltless condition and reduced cell membrane osmosis, MDA (malonaldehyde) content in salinity. So the salt tolerance of tomato was enhanced by AMF; (2) SOD, APX and POD activity in roots of AM symbiosis were significantly higher than corresponding non-AM plants in salinity or saltless condition. However, CAT activity was transiently induced by AMF and then suppressed to a level similar with non-AM seedlings; (3) higher salinity (1% level) and long stress time suppressed the effect of AMF on SOD, APX, POD and CAT activity; (4) this research suggested that the enhanced salt tolerance in AM symbiosis was mainly related with the elevated SOD, POD and APX activity by AMF which degraded more reactive oxygen species and so alleviated the cell membrane damages under salt stress. Whereas, the elevated SOD, POD and APX activity due to AMF depended on salinity environment.     

Oxidative stress and defence mechanisms of the freshwater cladoceran Daphnia longispina exposed to UV radiation

The aim of the present study is to evaluate the occurrence of oxidative stress in the cladoceran Daphnia longispina exposed to UV-A and UV-B radiation. The activity of antioxidant enzymes and lipid peroxidation markers is investigated and the protective action of ascorbic acid determined. Results show differences in the lethality radioinduced by UV-A and UV-B. Both UV-A and UV-B exposure cause an important increase in malonaldehyde (MDA) concentration and catalase activity. Ascorbic acid addition reduces the MDA concentration, indicating that the oxidative stress caused by either UV-A or UV-B radiation can be controlled by antioxidants. The increase of the antioxidant enzymes may be a response mechanism to oxidative stress.     

Induction of drought stress tolerance by ketoconazole in Catharanthus roseus is mediated by enhanced antioxidant potentials and secondary metabolite accumulation

A pot culture experiment was conducted to estimate the drought stress mitigating effect of ketoconazole (KCZ), a fungicide cum plant growth regulator, in Catharanthus roseus plants. The plants under pot culture were subjected to drought stress and drought stress with KCZ from 30 days after sowing (DAS) and regular irrigation was kept as control. Antioxidant contents and activities of antioxidant enzymes were estimated from root, stem and leaf of both control and treated plants. The alkaloid ajmalicine was extracted and estimated from the roots of control, drought stressed and KCZ treated plants. Individual and combined drought stress and KCZ treatments increased ascorbic acid, -tocopherol contents, superoxide dismutase, ascorbate peroxidase, catalase and polyphenol oxidase activities when compared to control. There was a significant enhancement in ajmalicine production under KCZ treated plants under drought stress when compared to well watered control as well as drought stressed plants. The KCZ treatment resulted in partial mitigation of drought stress by increasing the antioxidant potentials in C. roseus plants.     

Protection of Vigna unguiculata (L.) Walp. plants from salt stress by paclobutrazol

A pot culture experiment was conducted to estimate the stress ameliorating ability of paclobutrazol, a triazole fungicide in Vigna unguiculata (L.) Walp. plants. Treatments were given as 80 mM NaCl, 80 mM NaCl + 15 mg l⁻¹ paclobutrazol and 15 mg l⁻¹ paclobutrazol alone. The samples were collected on 60 and 80 days after sowing (DAS). NaCl stress inhibited the root and stem length, total leaf area, fresh weight (FW), dry weight (DW) and activities of antioxidant enzymes like superoxide dismutase (SOD), catalase (CAT) and peroxidase (POX). Plants treated with NaCl with paclobutrazol increased these parameters to a larger extent when compared to NaCl stressed plants. The results showed that the paclobutrazol significantly ameliorated the adverse effects of NaCl stress in V. unguiculata plants.     

Growth, Osmolyte Concentration and Antioxidant Enzymes in the Leaves of Sesuvium portulacastrum L. Under Salinity Stress

In this study, growth and osmolyte concentration in the leaves of halophyte, Sesuvium portulacastrum, were studied with respect to salinity. Therefore, the changes in shoot growth, leaf tissue water content, osmolyte concentration (proline content, glycine betaine) and antioxidant enzymes [polyphenol oxidase (PPO), superoxide dismutase (SOD) and catalase (CAT)] were investigated. The 30-day old S. portulacastrum plants were subjected to 100, 200, 300, 400, 500 and 600 mM NaCl for 28 days. The plant growth was steadily increased up to 500 mM NaCl stress at 28 days. TWC was higher in 300 mM NaCl treated leaves than that of 600 mM NaCl. Salinity stress induced the accumulation of osmolyte concentration when compared to control during the study period. The antioxidant enzymes PPO, CAT and SOD were increased under salinity.     

Drought Stress Effects on Growth,ROS Markers,Compatible Solutes,Phenolics, Flavonoids,and Antioxidant Activity in <Emphasis Type="Italic">Amaranthus tricolor</Emphasis>

Four selected Amaranthus tricolor cultivars were grown under four irrigation regimes (25, 50, 80, and 100% field capacity) to evaluate the mechanisms of growth and physiological and biochemical responses against drought stress in randomized complete block design with three replications. Drought stress led to decrease in total biomass, specific leaf area, relative water content (RWC), photosynthetic pigments (chlorophyll a, chlorophyll b, chlorophyll ab), and soluble protein and increase in MDA, H₂O₂, EL, proline, total carotenoid, ascorbic acid, polyphenols, flavonoids, and antioxidant activity. However, responses of these parameters were differential in respect to cultivars and the degree of drought stresses. No significant difference was observed in control and LDS for most of the traits. The cultivars VA14 and VA16 were identified as more tolerant to drought and could be used for further evaluations in future breeding programs and new cultivar release programs. Positively significant correlations among MDA, H₂O₂, compatible solutes, and non-enzymatic antioxidant (proline, TPC, TFC, and TAC) suggested that compatible solutes and non-enzymatic antioxidant played vital role in detoxifying of ROS in A. tricolor cultivar. The increased content of ascorbic acid indicated the crucial role of the ASC–GSH cycle for scavenging ROS in A. tricolor.     

Antioxidant Effect of Azadirachta Indica on High Fat Diet Induced Diabetic Charles Foster Rats

Oxidative stress plays a major role in the pathogenesis of both types of diabetes mellitus. Excessively high levels of free radicals cause damage to cellular proteins, membrane lipids and nucleic acids, and eventually cell death. The present study was designed to investigate the possible effect of Azadirachta indica leaf extract in high fat diet induced diabetic Charles Foster rats. The increased level of lipidperoxidation and altered levels of enzymatic (superoxide dismutase, glutathione peroxidase and catalase) and non-enzymatic (glutathione) antioxidants were seen in high fructose fed animals. The treatment with A. indica leaf extract significantly normalized the altered levels of lipid peroxidation and antioxidant status at 400?mg/kg b.w. dose. The A. indica leaf extract was also tested for in vitro inhibition of generation of superoxide anion and hydroxyl free radical in both enzymatic and non-enzymatic systems. The A. indica leaf extract was found to inhibit generation of superoxide anion and hydroxyl free radical significantly at 200???g/ml concentration. Data of present study demonstrated that the A. indica leaf extract has both antidiabetic and antioxidant properties.     

Oxidation of kojic acid catalyzed by manganese peroxidase from Ceriporiopsis subvermispora in the absence of hydrogen peroxide

We have previously reported the oxidation of kojic acid catalyzed by manganese peroxidase (MnP) from Ceriporiopsis subvermispora. This reaction is strictly dependent on Mn(II), although it does not require the addition of hydrogen peroxide. We have extended these studies because this reaction can be considered as a model system for the in situ generation of hydrogen peroxide in natural environments. We show here that oxidation of kojic acid with horseradish peroxidase (HRP) plus hydrogen peroxide or with manganic acetate rendered a product with identical chromatographic and spectral properties as the one obtained in the reaction catalyzed by MnP. The initial lag observed in the latter reaction decreased significantly upon UV irradiation of the substrate. On the other hand, ascorbic acid increased the lag and did not affect the yield of the reaction. The superoxide anion trapping agents glutathione, nitroblue tetrazolium, and superoxide dismutase markedly affected the reaction. In contrast, addition of the hydroxyl radical scavengers mannitol and salicylic acid had no effect. Based on these results, a mechanism for the MnP-catalyzed reaction is proposed.     

Methyl jasmonate and salicylic acid induced oxidative stress and accumulation of phenolics in Panax ginseng bioreactor root suspension cultures

To investigate the enzyme variations responsible for the synthesis of phenolics, 40 day-old adventitious roots of Panax ginseng were treated with 200 microM methyl jasmonate (MJ) or salicylic acid (SA) in a 5 L bioreactor suspension culture (working volume 4 L). Both treatments caused an increase in the carbonyl and hydrogen peroxide (H2O2) contents, although the levels were lower in SA treated roots. Total phenolic, flavonoid, ascorbic acid, non-protein thiol (NPSH) and cysteine contents and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical reducing activity were increased by MJ and SA. Fresh weight (FW) and dry weight (DW) decreased significantly after 9 days of exposure to SA and MJ. The highest total phenolics (62%), DPPH activity (40%), flavonoids (88%), ascorbic acid (55%), NPSH (33%), and cysteine (62%) contents compared to control were obtained after 9 days in SA treated roots. The activities of glucose 6-phosphate dehydrogenase, phenylalanine ammonia lyase, substrate specific peroxidases (caffeic acid peroxidase, quercetin peroxidase and ferulic acid peroxidase) were higher in MJ treated roots than the SA treated ones. Increased shikimate dehydrogenase, chlorogenic acid peroxidase and beta-glucosidase activities and proline content were observed in SA treated roots than in MJ ones. Cinnamyl alcohol dehydrogenase activity remained unaffected by both MJ and SA. These results strongly indicate that MJ and SA induce the accumulation of phenolic compounds in ginseng root by altering the phenolic synthesis enzymes.     

A hydroponic rice seedling culture model system for investigating proteome of salt stress in rice leaf

By using an in vivo hydroponic rice seedling culture system, we investigated the physiological and biochemical responses of a model rice japonica cultivar Nipponbare to salt stress using proteomics and classical biochemical methods. Yoshida's nutrient solution (YS) was used to grow rice seedlings. YS-grown 18-day-old seedlings manifested highly stable and reproducible symptoms, prominently the wilting and browning of the 3rd leaf, reduced photosynthetic activity, inhibition in overall seedling growth, and failure to develop new (5th) leaf, when subjected to salt stress by transferring them to YS containing 130 mM NaCl for 4 days. As leaf response to salt stress is least investigated in rice by proteomics, we used the 3rd leaf as source material. A comparison of 2-DE protein profiles between the untreated control and salt-stressed 3rd leaves revealed 55 differentially expressed CBB-stained spots, where 47 spots were increased over the control. Of these changed spots, the identity of 33 protein spots (27 increased and 5 decreased) was determined by nESI-LC-MS/MS. Most of these identified proteins belonged to major metabolic processes like photosynthetic carbon dioxide assimilation and photorespiration, suggesting a good correlation between salt stress-responsive proteins and leaf morphology. Moreover, 2-DE immunoblot and enzymatic activity analyses of 3rd leaves revealed remarkable changes in the key marker enzymes associated with oxidative damage to salt stress: ascorbate peroxidase and lipid peroxidation were induced, and catalase was suppressed. These results demonstrate that hydroponic culture system is best suited for proteomics of salt stress in rice seedling.     

Rosmarinic Acid and Antioxidant Enzyme Activities in <Emphasis Type="Italic">Lavandula vera</Emphasis> MM Cell Suspension Culture: A Comparative Study

The growth and intracellular protein content of lavender (Lavandula vera MM) cell suspension culture was followed along with some antioxidant defense system members—non-enzymatic (rosmarinic acid) and enzymatic [superoxide dismutase (EC 1.15.1.1) and catalase (EC 1.11.1.6)]. It was found that the media content and the cultivation mode strongly influenced the production of plant defense compounds as well as the ratio between non-enzymatic and enzymatic ones. The bioreactor culture contains about two times more rosmarinic acid, superoxide dismutase, and catalase compared to the shake-flask cultivation. These findings are discussed with respect to the relative stress levels and plant antioxidant orchestra system. It was concluded that investigated defense system components (enzymatic and non-enzymatic) were closely associated in a complex balance. The three isoenzyme forms of SOD (Cu/ZnSOD, FeSOD, and MnSOD) in the cells of Lavandula vera were revealed by polyacrylamide gel electrophoresis analysis, and the FeSOD isoform exhibited highest activity.     

Effect of sound wave stress on antioxidant enzyme activities and lipid peroxidation of Dendrobium candidum

The effect of sound wave stress on important medicinal plant, Dendrobium candidum Wall. ex Lindl, was investigated, including the responses on malondialdehyde (MDA) content, the activities change of superoxide dismutase (SOD), catalase (CAT), peroxidase (POD) and ascorbate peroxidase (APX). Results were found that the activities of SOD, CAT, POD and APX enhanced totally in different organs of D. candidum, as leaves, stems and roots, in response to the stress. Furthermore there happened similar shift of antioxidant enzymes activities, which increased in the initial stimulation and decreased afterwards. Data showed SOD, CAT, POD and APX activities ascended to max at day 9, 6, 9 and 12 in leaves, at day 9, 6, 12 and 9 in stems, and at day 12, 6, 9 and 9 in roots, respectively. As a lipid peroxidation parameter, MDA content in different organs increased in the beginning, dropped afterward, and increased again in the late. Anyway the total trend was the rise of MDA level compared to the control. It was interesting that the MDA content appeared the lowest levels almost when the antioxidant enzymes activities were up to the highest. Our results demonstrated the different organs of D. candidum might produce accumulation of active oxygen species (AOS) under initial treatment of sound wave stress. Later AOS might start to reduce due to the enhancement of antioxidant enzymes activities treated by the stress. The data revealed that the antioxidant metabolism was to be important in determining the ability of plants to survive in sound stress, and the up regulation of these enzymes activities would help to reduce the build up of AOS, which could protect plant cells from oxidative damage. Moreover, different cell compartments might activate different defensive system to reduce excessive amount of AOS. Finally the mechanism of this action was also discussed simply.     

Electrochemical Reduction of a Conjugated Cinnamic Acid Diazonium Salt as an Immobilization Matrix for Glucose Biosensor

Initial results on the synthesis of a new conjugated diazonium salt of trans‐4‐cinnamic acid diazonium fluoroborate, which is used for the chemical modification of the glassy carbon (GC) electrode with trans‐4‐cinnamic acid groups through electrochemical reduction, and direct covalent immobilization of glucose oxidase (GOD) on the cinnamic acid groups are presented. The chemically modified GC electrode exhibits a good selectivity relative to the bare GC electrode for the various possible interfering compounds in glucose analysis: namely ascorbic acid and 4‐acetamidophenol. Covalent immobilization of GOD on the chemically modified GC electrode produces a biosensor which responds to glucose concentration changes in the presence of a soluble redox mediator (ferrocenemethanol). The chemical modification of GC by cinnamic acid groups is potentially useful for the attachment of other enzymes and biochemical reagents.