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Antioxidative Enzymes in Sunflower Subjected to Drought Stress

Dawoud Habibi1, Masoud Mashdi Akbar Boojar2, Ali Mahmoudi1, Mohammad Reza Ardakani1 and Daryoush Taleghani3

Islamic Azad University of Karaj , www.tylakoid.com Email d_habibi2004@Yahoo.com
2
University of Tarbiat Moalem
3
Sugarbeet Seed Institute , Email Taleghani@sbsi.ir

Abstract

In order to study the role of antioxidant enzymes superoxide dismutase (SOD), catalze (CAT) and glutathion peroxidase (GPX) in the drought resistance of five sunflower varieties an experiment was carried out under drought and control conditions. Results showed that the activity of these enzymes were significantly different between control and stress treatments. Results also showed there were no significant differences among varieties in the level of these enzymes. There was no relationship between drought resistance as measured by grain yield and the content of glutathion peroxidase, catalaze and superoxide dismutase among varieties. There was also no significant relationship among varieties between seed germination susceptibility to stress imposed by mannitol and antioxidant enzymes measured in the field. Therefore, selection for drought resistance by evaluation of SOD, GPX and CAT in these varieties was impossible.

Media summary

Oil sunflower varieties, in order to protect their cells from oxidative damage, increase their antioxidant enzymes superoxide dismutase, catalaze and glutathion peroxidase when subjected to drought stress.

Keywords :

Yield Susceptibility, Catalaze, Superoxide Dismutase, Glutathion Peroxidase, Germination

Introduction

H202 can be harmful because of its oxidative and destructive effects on the metabolism of plants. In organisms, H202 can be destroyed by catalase and glutathion peroxidace. Catalase protects cells from the effects of H202. Under normal condition catalase for some cells has an important role in increasing the resistance to oxidative stress. The reaction of free and semifree radicals of oxygen can be seen in destructive functions such as senecence (Ames et al, 1993). Glutathion peroxidase (GPX) has a residue of selenium of selenosistein on four unit branches that its very important for enzyme activity. GPX catalyzes the reduction of H202 by GSH (reduced glutathion), thereby protecting the cells from oxidative damage. Metabolism of glutathion is one of the protective mechanisms for antioxidants (Esterbauer et al, 1992). Increasing the glutathion and reductase activity in cotton and wheat under drought stress showed that, at the same time as H202 production, the increase can absorb ferrodoxin electrons by NADP, so superoxide production will be reduced. Halliwel et al. (1990) reported that in oilseed crops such as sunflower, the content of free radicals such as superoxide and peroxide in tissue will increase under stress conditions. These radicals cause destruction of lipids in sunflower and will affect their natural production. Peroxidase and catalaze have been seen in some physiological cycles under biotic and abiotic stresses. Under environmental stresses some products such an 02, H202 and OH will increase. Superoxide dismutase activity in plants will increase by the use of herbicides (Paraquat), SO2 concentration in the atmosphere, drought stress or high levels of Mg and Zn.

Material and Methods

This research was done in 2003 at the experimental farm of the Islamic Azad University of Karaj in Iran using a split plot completely randomized block design with four replications. Irrigation was the main factor at two levels (normal and stress conditions) and five oil sunflower varieties (Record, Progress, Azargol, Colshid, Gabour) as sub-plots within the main plots. The level of stress was measured by a soil electrical conductivity meter. Additionally, the percent germination of seeds of the five varieties was measured in mannitol at an osmotic potential of -0.8 MPa (moderate stress) and in distilled water (Habibi et al., 1995).

Sample preparation for enzyme assay and protein measurement:

Leaves from each plant were washed with distilled water and homogenized in 0.16M Tris buffer ( pH = 7.5) at 4C. Then, 0.5 mL of total homogenized solution was used for protein determination by the Lowery et al. (1951) method. Based on the amount of protein per volume of homogenized solution, the following enzymes were assayed in the volume containing a known protein concentration in order to calculate the specific activities of the enzymes.

Glutathion peroxidase (GPX) activity:

The activity was measured by the Paglia and valentine (1987) method in which 0.56M (pH=7) phosphate buffer, , 0.5 M EDTA, 1mM NaN3, 0.2mM NADPH were added to the extracted solution. GPX catalyses the oxidation of glutathion (GSH) by cumene hydroperoxide. In the presence of glutathion reductase and NADPH, the oxidized glutathion is immediately converted to the reduced form with the concomitant oxidation of NADPH to NADP. The decrease in absorbance at 340 nm was measured with a spectrophotometer.

Superoxide dismutase (SOD) activity:

The activity was measured based on Misra and Fridovich (1972), in which the activity was measured on the basis of its ability to inhibit free radical chain oxidation in which O.-2 was a chain propagating radical and the autooxidation of epinephrine (0.25mM) was inducedA SOD standard was used for calibration of activity.

Catalaze (CAT) activity:

Catalaze activity was measured at 25C as previously described by Paglia and Valentine (1987), that used hydrogen peroxide as substrate and 1 k of catalaze activity was defined as the rate constant of the first order reaction.

Results and conclusions

The results showed that there were significant differences (P<0.01) between activity levels of superoxide dismutase, catalaze and glutathion peroxidase in the irrigated and drought stress treatments. The activity of all antioxidants enzymes were increased under drought stress in all the varieties, but there was no relationship between yield stability under drought stress and the content of catalze, glutathion peroxidase and superoxide dismutase in the varieties (Table 1). Among the varieties, Golshid had highest susceptibility to drought stress and the lowest SOD content, and Record had lowest susceptibility and high SOD content (Table 1). In Record, the susceptibility of germination to the drought stress (mannitol) treatment was low and SOD was very high, and this variety also produced more grain yield in the field. We can conclude those varieties that have more uniform germination in laboratory when exposed to an osmoticum may have more seed yield stability in the field and higher drought resistance. There was no significant relationship between antioxidant enzymes activity, and the stability of seed yield and seed germination under drought conditions among these varieties, so selection for drought resistance by using antioxidant enzymes is not possible.

Table 1. Antioxidative enzyme activity, seed yield and germination susceptibility of sunflower varieties subjected to drought stress.

(1) Fischer and Maurer (1976)
(2) Habibi et al. (1995)

References

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Esterbauer H, Grebicki J, Puhl H, and Jugens G (1992). The role of lipid peroxidation and antioxidants in oxidation modification of LDL. Free Radic Biol Med : 100, 341-90 .

Fischer RA and Maurer R (1976). Drought resistance in spring wheat cultivar. I. Grain yield responses. Aust. J. Agric. Res. 29 : 897-912.

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Habibi D, Khodabandeh N, Heravan E M, Gohari J, Chegini M A (1995). Selection of the tolerant sugarbeet genotypes under drought stress. 3th Iranian Congress of Crop Production and Plant Breeding. 31 Aug . – 4 Sep.

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Misra HP and Fridovich I (1972). The generation of superoxide radical during auto oxidation . J. Biol. Chem. 247,6960-6966.

Paglia DE, and Valentine WN (1987). Studies on the quantitative and qualitative characterization of glutathion proxidase. J. Lab. Med. 70 : 158-165.

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