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.     

Defensive Mechanisms in Cucurbits against Melon Fly (Bactrocera cucurbitae) Infestation through Excessive Production of Defensive Enzymes and Antioxidants

Melon fly (Bactrocera cucurbitae) is the most common pest of cucurbits, and it directly causes damage to cucurbit fruits in the early developmental stage. The infection of fruit tissues induces oxidative damage through increased generation of cellular reactive oxygen species. The effects of melon fly infestation on the production of defensive enzymes and antioxidant capabilities in five cucurbit species, namely, bottle gourd, chayote, cucumber, snake gourd, and bitter gourd, were investigated in this study. The total phenolic and flavonoid content was considerably higher in melon fly infestation tissues compared to healthy and apparently healthy tissues. The chayote and bottle gourd tissues expressed almost 1.5- to 2-fold higher phenolic and flavonoid contents compared to the tissues of bitter gourd, snake gourd, and cucumber upon infestation. Defensive enzymes, such as peroxidase (POD), superoxide dismutase (SOD), polyphenol oxidase (PPO), and catalase (CAT), were high in healthy and infected tissues of chayote and bottle gourd compared to bitter gourd, snake gourd, and cucumber. The activity of POD (60–80%), SOD (30–35%), PPO (70–75%), and CAT (40–50%) were high in infected chayote and bottle gourd tissue, representing resistance against infestation, while bitter gourd, snake gourd, and cucumber exhibited comparatively lower activity suggesting susceptibility to melon fly infection. The antioxidant properties were also high in the resistant cucurbits compared to the susceptible cucurbits. The current research has enlightened the importance of redox-regulatory pathways involving ROS neutralization through infection-induced antioxidative enzymes in host cucurbit resistance. The melon fly infestation depicts the possible induction of pathways that upregulate the production of defensive enzymes and antioxidants as a defensive strategy against melon fly infestation in resistant cucurbits.     

Effects of UV-B radiation on growth, photosynthesis, UV-B-absorbing compounds and NADP-malic enzyme in bean (Phaseolus vulgaris L.) grown under different nitrogen conditions.

The effects of UV-B radiation on growth, photosynthesis, UV-B-absorbing compounds and NADP-malic enzyme have been examined in different cultivars of Phaseolous vulgaris L. grown under 1 and 12 mM nitrogen. Low nitrogen nutrition reduces chlorophyll and soluble protein contents in the leaves and thus the photosynthesis rate and dry-matter accumulation. Chlorophyll, soluble protein and Rubisco contents and photosynthesis rate are not significantly altered by ambient levels of UV-B radiation (17 microW m-2, 290-320 nm, 4 h/day for one week). Comparative studies show that under high nitrogen, UV-B radiation slightly enhances leaf expansion and dry-matter accumulation in cultivar Pinto, but inhibits these parameters in Vilmorin. These results suggest that the UV-B effect on growth is mediated through leaf expansion, which is particularly sensitive to UV-B, and that Pinto is more tolerant than Vilmorin. The effect of UV-B radiation on UV-B-absorbing compounds and on NADP-malic enzyme (NADP-ME) activity is also examined. Both UV-B radiation and low-nitrogen nutrition enhance the content of UV-B-absorbing compounds, and among the three cultivars used, Pinto exhibits the highest increases and Arroz the lowest. The same trend is observed for the specific activity and content of NADP-ME. On a leaf-area basis, the amount of UV-B-absorbing compounds is highly correlated with the enzyme activity (r2 = 0.83), suggesting that NADP-ME plays a key role in biosynthesis of these compounds. Furthermore, the higher sensitivity of Vilmorin than Pinto to UV-B radiation appears to be related to the activity of NADP-ME and the capacity of the plants to accumulate UV-B-absorbing compounds.     

Effects of CO2 laser pretreatment on drought stress resistance in wheat

In order to determine the role of laser in drought stress resistance of spring wheat (Triticum aestivum L.), seed embryos were exposed to CO2 laser radiation for 0min, 1min, 3min and 5min, respectively, and when the seedlings were 12 days old they were treated with 10% (w/v) PEG6000 solution for 10 days. Changes in the concentration of malondialdehyde (MDA), hydrogen peroxide (H2O2), glutathione (GSH), ascorbate (AsA), oxidized glutathione (GSSG), carotenoid, zeaxanthin, the production rate of superoxide radical (O2(-)), the activities of ascorbate peroxidase (APX), peroxidase (POD), catalase (CAT), superoxide dismutase (SOD), glutathione reductase (GR), glutathione peroxidase (GP), glutathione-S-transferase (GST) and the growth parameters of seedlings (plant height, leaf area and dry weight) were measured to test the effects of laser pretreatment. The results showed that suitable laser pretreatment of embryos enhanced drought stress resistance in wheat seedlings by decreasing the concentration of MDA and H2O2, GSSG, the production rate of O2(-), leaf area and increasing the activities of APX, GST, GP and POD and AsA, carotenoid and zeaxanthin concentration. It is suggested that those changes in MDA, O2(-) H2O2, anti-oxidative enzymes and anti-oxidative compounds were responsible for the increase in drought stress resistance observed in the experiments. The results also showed that the laser had a long-term positive physiological effect on the growth of drought stress seedlings. This is the first investigation reporting the use of CO2 laser pretreatment to enhance drought stress resistance of spring wheat.     

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.     

Hydrogen Peroxide‐mediated Inactivation of Two Chloroplastic Peroxidases,Ascorbate Peroxidase and 2‐Cys Peroxiredoxin†

Reactive oxygen species (ROS), such as the superoxide anion and hydrogen peroxide, are generated by the photosystems because photoexcited electrons are often generated in excess of requirements for CO₂ fixation and used for reducing molecular oxygen, even under normal environmental conditions. Moreover, ROS generation is increased in chloroplasts if plants are subjected to stresses, such as drought, high salinity and chilling. Chloroplast‐localized isoforms of ascorbate peroxidase and possibly peroxiredoxins assume the principal role of scavenging hydrogen peroxide. However, in vitro studies revealed that both types of peroxidases are easily damaged by hydrogen peroxide and lose their catalytic activities. This is one contributing factor for cellular damage that occurs under severe oxidative stress. In this review, I describe mechanisms of hydrogen peroxide‐mediated inactivation of these two enzymes and discuss a reason why they became susceptible to damage by hydrogen peroxide.     

Identification and cloning of molecular markers for UV-B tolerant gene in wild sugarcane (Saccharum spontaneum L.)

Previously we have selected wild sugarcane (Saccharum spontaneum L.) sterile lines that are tolerant or susceptible to UV-B radiation based on response index (RI) in a field screening test. The RI was established according to plant height, tiller number, leaf index, total biomass and brix under enhanced ultraviolet-B (UV-B, 280-310 nm) radiation. In this experiment, molecular markers linked to the UV-B tolerant and susceptible genes were identified and cloned. RAPD (Randomly amplified polymorphic DNAs) assay using 100 arbitrary primers followed by clustering analysis separated the tolerant and susceptible lines into two groups at the genetic distance of 0.380. The UV-B tolerant and susceptible gene pools were constructed and compared using the Bulked Segregate Analysis (BSA) approach. Of the 100 arbitrary RAPD primers, primer OPR16 produced polymorphic DNA banding patterns from both gene pools. The OPR16-1200 bp DNA fragment was only amplified from the tolerant lines and the OPR16-800 bp from the susceptible ones. These two PCR fragments were cloned onto T-vector. DNA sequence alignment analysis determined that 42% homology existed between the reverse and forward sequences of the OPR16-1200 bp clone, and 36% homology between the forward sequences of the OPR16-800 bp and OPR16-1200 bp clones. The two DNA clones were determined to be linked to the UV-B tolerant and susceptible genes, and they can be used to develop molecular markers for the associated traits.     

Herba Epimedii: anti-oxidative properties and its medical implications

Herba Epimedii is a Chinese herbal medicine with proven efficacy in treating cardiovascular diseases and osteoporosis, and in improving sexual and neurological functions. This efficacy is found to be related to the potent anti-oxidative ability of Herba Epimedii and its flavonoid components, with icarrin as the main effective constituent, along with polysaccharides and vitamin C. These ingredients have been proven to be effective against oxidative-stress related pathologies (cardiovascular diseases, Alzheimer's disease and inflammation) in animal rodent models and in vitro studies. Their anti-oxidative properties are found to be related to an inductive effect on endogenous free-radical scavenging enzymes such as catalase and glutathione peroxidase and the inherent electron-donating ability of flavonoids.     

Barley cultivar discrimination: I. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and glycoprotein blotting.

Two different methods of detecting electroblotted glycoproteins after sodium dodecyl sulfate-polyacrylamide gel electrophoresis of Tris-buffer soluble barley seed proteins were examined for their applicability for barley cultivar discrimination. These are the highly specific, lectin-based concanavalin A/peroxidase method and the more general periodate/danyslhydrazine method. The results of the periodate/dansylhydrazine method enabled us to divide the 20 examined cultivars into three groups, whereas the more sensitive concanavalin A/peroxidase method revealed six different glycoprotein patterns. In comparison, sodium dodecyl sulfate-polyacrylamide gel electrophoresis and silver staining of the alcohol-soluble barley seed proteins (hordeins) gave nine different banding patterns. A combination of hordein electrophoresis together with glycoprotein staining by the concanavalin A/peroxidase method made it possible to classify the cultivars into twelve groups, the largest of which contained four cultivars. The qualitative expression of the glycoprotein patterns seemed to be independent of growth conditions, whereas the band intensities obviously were not. As a whole, glycoprotein blotting is a valuable supplement to sodium dodecyl sulfate-polyacrylamide gel electrophoresis of hordeins in barley cultivar discrimination.     

Analysis by two-dimensional electrophoresis of the effect of salt stress on the polypeptide patterns in roots of a salt-tolerant and a salt-sensitive cultivar of wheat

The effect of salt stress on the polypeptide levels in roots of two wheat (Triticum durum) cultivars with different sensitivity to NaCl (cv. Ben Bachir, sensitive; cv. Chili, tolerant), was examined by two-dimensional polyacrylamide gel electrophoresis. Blue-stained gels were analyzed by visual inspection to identify changes that resulted when seedlings were grown in the presence of 200 mM NaCl for four days. Although the protein patterns for control and salt-stressed seedlings were qualitatively similar, the net synthesis of a 26 kDa polypeptide was significantly changed. This observation was mainly noticeable in the more tolerant cultivar. With the intention of identifying its function, the NH2-terminal of this polypeptide was sequenced. A 20 amino acid sequence was obtained and compared to sequences available in different databases. Possible roles of this polypeptide, depending on the homologies of its amino acid sequence with known proteins, in salinity tolerance are discussed.     

The role of lignification in the resistance of plants to attack by pathogens

Reviews by Vance et al. (1980) and Friend (1981) contain details of and references to the earlier work on lignification in both cucumber and grasses that is briefly described below. The suggestion of Hijwegen in 1963 that active lignification could be a general resistance mechanism in plants arose from experiments in which it was found that lignin deposition was associated with the reaction of a resistant cultivar of cucumber to inoculation withCladosporium cucumerimum. More recently Hammerschmidt and Kuć (1982) have found that systemic resistance of cucumber, induced by pre-inoculation with eitherC. cucumerimum orColletotrichum lagenarium, was associated with the ability of the induced plants to produce lignin rapidly in response to a challenge inoculation with either fungus. They concluded that induced resistance may be similar to cultivar resistance. Hammerschmidt et al. (1982) also showed that induction inoculation gave enhanced peroxidase activity in protected plants, associated with the fast-moving anodic isoenzymes. Lignification, especially the formation of lignified papillae, seems to be a common reaction of grass plants to attempted infection by nonpathogens. This has been found in reed canary-grass,Phalaris arundinacea, and a range of grasses by Vance and Sherwood, and in wheat by Ride. In reed canary-grass, active lignification is associated with increases in the enzymes of phenolic acid biosynthesis, and in peroxidase with the appearance of one new isoenzyme. The inducible resistance response, together with the associated increases in enzyme levels, were inhibited by the protein synthesis inhibitor, cycloheximide. The difference in response of wounded wheat leaves to inoculation by the nonpathogenBotrytis cinerea and the pathogenSeptoria nodorum appears to be related to the speed of lignification. It was rapid in the former case, confining the fungus to the wounded area; in the latter, it was slower and the fungus was not restricted. Recent experiments indicate that the accumulation of phenolic compounds in cell walls is a major factor in the non-race-specific resistance of potato tuber discs toPhytophthora infestans and of the same cultivars toPhoma exigua var foveata (Ampomah, 1983; Ampomah and Friend, in preparation). Accumulation of these compounds, estimated either by the level of browning of discs or the intensity of blue-green staining with toluidine blue of the walls of the top layer of cells, was greater following inoculation of resistant than of susceptible cultivars. Treatment of discs with aminooxyacetic acid (AOA), which is an inhibitor of PAL activity in vivo, inhibited the deposition of the phenolic compounds and permitted the fungi to grow further through tuber tissue. After freezing the discs in liquid nitrogen and extracting with chloroform: methanol: water (CMW), the residue was extracted with phenol: acetic acid:water (PAW). The CMW extracts contained chlorogenic acid; its level was lower in inoculated than uninoculated tissue. δεi measurements indicated that phenolic compounds were present both in the dialyzed PAW extract and in the PAW residue. There was always more phenolic material in both extract and residue from the inoculated discs of both resistant and susceptible cultivars than from uninoculated discs; the levels were always higher in the resistant than in the susceptible discs. The presence of quinic acid in the PAW extracts indicates the possibility that they contain oxidized, polymerized chlorogenic acid. Such material is absent from the PAW residue that contains esterified hydroxy cinnamic acids. We are, as yet, unable to determine whether these acids are esterified to lignin, to carbohydrate, or to the hydroxy-aliphatic acids in suberin.     

Cytochemical labeling for fungal and host components in plant tissues inoculated with fungal wilt pathogens.

Antibodies to detect pectin in present investigations attached to distinct fibrils in vessel lumina. In carnation infected with an isolate of Fusarium oxysporum f.sp., labeling of pathogen cells also occurred; in a resistant cultivar (cv.), it was coincident with proximate pectin fibrils and linked to altered fungal walls, which was the opposite in the susceptible cv., indicating that hindrance of pathogen ability to degrade pectin may be related to resistance. Labeling of the fungus in culture was nil, except in media containing pectin, showing that pectin is not native to the pathogen. Labeling of fungal walls for cellulose in elm (inoculated with Ophiostoma novo-ulmi) and carnation also occurred, linked to adsorbed host wall components. The chitin probe often attached to dispersed matter, in vessel lumina, traceable to irregularly labeled fungal cells and host wall degradation products. With an anti-horseradish peroxidase probe, host and fungal walls were equally labeled, and with a glucosidase, differences of labeling between these walls were observed, depending on pH of the test solution. Fungal extracellular matter and filamentous structures, present in fungal walls, predominantly in another elm isolate (Phaeotheca dimorphospora), did not label with any of the probes used. However, in cultures of this fungus, extracellular material labeled, even at a distance from the colony margin, with an anti-fimbriae probe.     

Citrus essential oil of Nigeria. Part V: Volatile constituents of sweet orange leaf oil (Citrus sinensis)

The volatile oils extracted from leaves of eight cultivars of Citrus sinensis (L) Osbeck were comprehensively analysed by a combination of GC and GC-MS. Fifty four constituents accounting for 82.3-98.2% were identified. Sabinene (20.9-49.1%), delta-3-carene (0.3-14.3%), (E)-beta-ocimene (4.4-12.6%), linalool (3.7-11.1%) and terpinen-4-ol (1.7-12.5%) were the major constituents that are common to all the volatile oils. In addition, a cluster analysis was carried out and indicated at least four different chemotypes for the C. sinensis cultivars.     

Essential Oil Volatile Fingerprint Differentiates Croatian cv. Oblica from Other Olea europaea L. Cultivars

Olive leaves are a highly available by-product from table olive and olive oil production. They are nowadays strongly valuable for their major bioactive compounds and their beneficial effects. To determine the differences between two Croatian domestic (Lastovka, Oblica) and two introduced (Leccino, Frantoio) cultivars, physical and chemical analysis of olive leaves were performed: surface area, color variability, total phenolic amounts, and essential oil volatile profiles were analyzed at three harvest periods. All cultivars greatly differed in surface area, with cv. Lastovka being the smallest. Color variability resulted in an overall decrease in darkness and amounts of green and yellow that could be attributed to a decrease in photosynthetic demand and chlorophyll content. The highest amount of total phenolic content occurred in the summer months, followed by a reduction until October. Essential oils volatiles were determined by GC-MS and showed great diversity not only amongst cultivars but also between harvest periods, with overall 45 compounds identified. Principal component analysis distinguished domestic cultivar Oblica from the other observed cultivars, mainly due to its essential oil volatile fingerprint. Compounds that differentiated cv. Oblica were aldehydes ((E,Z)-2,4-heptadienal, (E,E)-2,4-heptadienal, decanal), ketones ((E)-β-damascone, dihydrodehydro-β-ionone), sesquiterpenes (cyclosativene, α-copaene, α-muurolene) and saturated hydrocarbons (tetradecane, hexadecane). Essential oil volatile fingerprint attributed the highest to the biodiversity of domestic cv. Oblica through all three harvest periods.     

Up-regulation of defense enzymes is responsible for low reactive oxygen species in malignant prostate cancer cells

Reactive oxygen species (ROS) are involved in a diversity of important phenomena in the process of tumor development. To investigate the alterations of oxidative stress and their related systems in tumor progression, a variety of components in the antioxidative stress defense system were examined in prostate cancer cell lines, PC3 and LNCaP. Cell surface molecules involved in metastasis were expressed highly in PC3 cells compared with LNCaP cells, and strong invasion ability was shown in PC3 cells only. ROS level in LNCaP cells was twice higher than that in PC3 cells, although nitric oxide (NO) level was similar between the two cell lines. The content of GSH increased up to about 2-fold in PC3 compared with LNCaP. Activities of glutathione reductase, thioredoxin reductase, and glutathione S-transferase except catalase are significantly higher in PC3 cells than in LNCaP cells. Furthermore, oxidative stress-inducing agents caused down-regulation of GSH and glutathione S-transferase much more significantly in LNCaP cells than in PC3 cells. These results imply that malignant tumor cells may maintain low ROS content by preserving relatively high anti-oxidative capacity, even in the presence of stressful agents.     

Effects of Sphingomyelin-Containing Milk Phospholipids on Skin Hydration in UVB-Exposed Hairless Mice

Reactive oxygen species (ROS) generated by ultraviolet (UV) exposure cause skin barrier dysfunction, which leads to dry skin. In this study, the skin moisturizing effect of sphingomyelin-containing milk phospholipids in UV-induced hairless mice was evaluated. Hairless mice were irradiated with UVB for eight weeks, and milk phospholipids (50, 100, and 150 mg/kg) were administered daily. Milk phospholipids suppressed UV-induced increase in erythema and skin thickness, decreased transepidermal water loss, and increased skin moisture. Milk phospholipids increased the expression of filaggrin, involucrin, and aquaporin3 (AQP3), which are skin moisture-related factors. Additionally, hyaluronic acid (HA) content in the skin tissue was maintained by regulating the expression of HA synthesis- and degradation-related enzymes. Milk phospholipids alleviated UV-induced decrease in the expression of the antioxidant enzymes superoxidase dismutase1 and 2, catalase, and glutathione peroxidase1. Moreover, ROS levels were reduced by regulating heme oxygenase-1 (HO-1), an ROS regulator, through milk phospholipid-mediated activation of nuclear factor erythroid-2-related factor 2 (Nrf2). Collectively, sphingomyelin-containing milk phospholipids contributed to moisturizing the skin by maintaining HA content and reducing ROS levels in UVB-irradiated hairless mice, thereby, minimizing damage to the skin barrier caused by photoaging.     

Multifunctional Antioxidants: Regenerable Radical‐Trapping and Hydroperoxide‐Decomposing Ebselenols

Regenerable, multifunctional ebselenol antioxidants were prepared that could quench peroxyl radicals more efficiently than α‐tocopherol. These compounds act as better mimics of the glutathione peroxidase enzymes than ebselen. Production of reactive oxygen species (ROS) and reactive nitrogen species (RNS) in human mononuclear cells was considerably decreased upon exposure to the organoselenium compounds. At a concentration of 25 μm , the ebselenol derivatives showed minimal toxicity in pre‐osteoblast MC3T3 cells.     

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.