The antioxidant potential: factor of abiotic stress tolerance in - - PDF document

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11th Meeting of the Inter-Regional Cooperative Research Network on Cotton for the Mediterranean and Middle East Regions Antalya, Turkey, November 05-07, 2012

The antioxidant potential: factor of abiotic stress tolerance in cotton

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• A. Gurel,

Ege University, Engineering Faculty, Bioengineering Department, Izmir, Turkey

• L. Yildiz-Aktas

Ege University, Science Faculty, Biology Department, Izmir, Turkey

• B. Yagmur
• H. Hakerlerler

Ege University, Agriculture Faculty, Soil and Plant Nutrition Department, Izmir, Turkey

• B. Izci

Çanakkale Onsekiz Mart University, Agriculture Faculty, Field Crops Department, Çanakkale, Turkey

• Y. Oren

Tariş Figs, Raisins, Cotton and Oil Seeds Agricultural Sales Cooperatives Unions Izmir, Turkey

• H. Akdemir

Ege University, Ödemiş Vocational Training School, Ödemiş-Izmir, Turkey

• A. Edreva
• V. Velikova
• T. Tsonev
• E. Gesheva

Institute of Plant Physiology and Genetics, Bulgarian Academy of Sciences, Sofia, Bulgaria

• S. Dagnon

P.Hilendarski Plovdiv University, Plovdiv, Bulgaria

• D. Stoyanova-Koleva
• St. Kl.Ohridsky Sofia University, Sofia, Bulgaria

It is commonly accepted that the primary event induced by various stress factors in plants is

the burst of reactive oxygen species

ROS i.e. a state of oxidative stress that can have deleterious effect on cell function and structure. Plants elaborated a diversified network of antioxidants (ROS scavengers) to regulate the oxidative stress. The antioxidant potential is deployed as a response to stress inflict.

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Pathogens Temperature extremes Water deficit Nutrient imbalance High light UV Metals

Stress factors

ROS

ROS

Antioxidants

The antioxidant defense of plants involves compounds

• f diverse chemical types

Enzymes

Peroxidase Catalase Superoxidedismutase Glutathione reductase Etc.

Non-enzymatic compounds

Carotenoids Tocopherols Polyphenols Proline Polyamnines Etc.

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In our long-term research on cotton stress physiology we examined

two cases of cotton- stress factor interactions:

Nutrient (K/Na) imbalance (leaf reddening) Water deficit

In both cases abiotic constraints induce

a state of oxidative stress

Cotton reddening

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Red cotton leaves

• Three locations in Aegean region:

Söke, Menemen, Bergama

• Cotton plants:

Nazilli 84 cv. ♦ Leaves of green plants (controls) Leaves with symptoms of reddening: ♦ Light symptoms ♦ Severe symptoms

• Soils:

♦Under green plants ♦Under reddening plants

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We have established that reddening of cotton leaves is provoked by

K deficiency in the soil and K/Na imbalance

leading to overaccumulation of Na in the leaves

K and Na content (ppm) of soils on which green and reddening plants are grown

ppm

500 1000 1500 2000

green plants reddening plants

K Na

Soils under:

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Plants make better use of Na in case of K deficiency K and Na content (%) in leaves

• f green and reddening plants

%

0.0 0.5 1.0 1.5 2.0

green plants reddening plants

K Na

Excess of Na induces over-accumulation of toxic OH• free radicals in plants, i.e. a state of oxidative stress (Alia et al. 1993).

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Biochemical changes related to cotton reddening

Increase of non-enzymatic (proline) and enzymatic (peroxidase) antioxidants in reddening leaves

50 100 150 200 250 300

Proline

% of controls

50 100 150 200 250 300 350 400 450

Peroxidase

% of controls

Green Light red Strong red

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100 200 300 400 500 600 700

Anthocyanins

% of controls

50 100 150 200 250 300 350 400 450 500

Total phenols

% of controls

Dramatic increase of anthocyanins and total phenols in the reddening leaves

Green Light red Strong red

Green leaves Reddening leaves

Retention time (min)

HPLC of anthocyanins (C6-C3-C6+) in green and reddening cotton leaves Peaks 4 and 5 are identified as cyanidin glycosides

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The shift from malvidin to cyanidin in reddening leaves determines a stronger protective potential against oxidative stress.

OCH3 OH OH OH HO OCH3 O

+

OH OH OH OH HO H O

+

Malvidin

• Low antioxidant/antiradical activity
• Predominant aglycone in green leaves

Cyanidin

• High antioxidant activity

(due to the o-OH grouping in the B-ring)

• Predominant aglycone in reddening leaves

Green leaves Reddening leaves

B B A A

The efficacy of the antioxidant defense in reddening leaves is evidenced by the low damage of membrane integrity as shown by the Malonyl dialdehyde (MDA) test Transmission electron microscopy

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20 40 60 80 100 120

Malonyl dialdehyde % of controls

MDA test Low membrane damage in reddening leaves

MDA

Transmission electron microscopy

Preserved membrane integrity in reddening leaves Reddening leaves Green leaves

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Antioxidant protection in reddening cotton leaves:

general proposed scheme

Shortage of K Accumulation

• f Na

Burst of ROS Accumulation

• f proline

Increase of peroxidase activity Accumulation

• f

anthocyanins Shift of aglycons

Green leaves Red leaves

Scavenging of ROS ROS

Drought tolerance

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Experimental design

Nazilli 84-S Drought sensitive (S) Şahin 2000 Drought tolerant (T)

Irrigation regimes

♦ Field capacity (normal water supply) ♦ 1/3 field capacity (drought stress)

Locality

♦ Söke, Aegean region of Turkey

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• Non-enzymatic antioxidants
• Polyphenols
• Proline
• Carotenoids
• Markers of membrane damage
• Malonyldialdehyde (MDA)

Parameters

Physiological

• Photosynthesis
• Water use efficiency (WUE)
• Max photochemical activity of PSII
• Relative water content (RWC)

Proline

Non-enzymatic antioxidants

Polyphenols Carotenoids

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HPLC pattern of polyphenols in the leaves of cotton genotype Nazilli 84-S.

1, 2, 3 – isomers of chlorogenic acid: 1 – 5-O-caffeoyl quinic acid 2 – 3-O-caffeoyl quinic acid 3 – 4-O-caffeoyl quinic acid 4 – 10 – flavonoids: 4 – isoquercitrin glycoside 6 – rutin 8 – quercitrin 9 – kaempferol-3-rutinoside 10 – quercetin

A B

OH O O OH OH Quercetin

Main polyphenols in cotton leaves

Flavonoids: quercetine derivatives Cinnamic acid derivatives

OH

Chlorogenic acid

OH OH OH HO CH H2 H2 O H H H CO COOH CH Both types have high antioxidant activity due to the presence of o-dihydroxy grouping

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Quercetin derivatives (rutin, isoquercitrin) are the major flavonoids in cotton leaves

Rutin

0,5 1 1,5 2 2,5 3

Normal water supply Drought stress

mg g -1 FM

Nazilli 84 (S) Sahin 2000 (T)

Isoquercitrin

0,5 1 1,5 2 Normal water supply Drought stress

mg g -1 FM

Nazilli 84 (S) Sahin 2000 (T)

Kaempferol -3-rutinoside

0,1 0,2 0,3 0,4 0,5

Normal water supply Drought stress

mg g -1 FM

Nazilli 84 (S) Sahin 2000 (T)

Total flavonoids

0,5 1 1,5 2 2,5 3 3,5 4 4,5 5 Normal water supply Drough tstress

mg g -1 FM

Nazilli 84 (S) Sahin 2000 (T)

Higher content of flavonoids in the drought-tolerant ( ) than in the sensitive ( ) genotype:

• at normal water supply
• at drought

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3-0-caffeoyl quinic acid

0,1 0,2 0,3 0,4 0,5 Normal water supply Drought stress

mg g -1 FM

Nazilli 84 (S) Sahin 2000 (T)

4-0-caffeoyl quinic acid

0,05 0,1 0,15 0,2 Normal water supply Drought stress

mg g -1 FM

Nazilli 84 (S) Sahin 2000 (T)

5-0-caffeoyl quinic acid

0,5 1 1,5 Normal water sullpy Drought stress

mg g -1 FM

Nazilli 84 (S) Sahin 2000 (T)

Total chlorogenic acid

0,5 1 1,5 2

Normal water supply Drought stress

mg g -1 FM

Nazilli 84 (S) Sahin 2000 (T)

Higher content of chlorogenic acid isomers in the drought-tolerant ( ) than in the sensitive ( ) genotype:

• at normal water supply
• at drought

β-carotene

Non-enzymatic antioxidants Proline Carotenoids

H N CH COOH H2C CH2 H2C

Proline

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Proline

0,4 0,8 1,2 1,6 Normal water content Drought stress

μM g-1 FM

Nazilli 84 (S) Sahin 2000 (T)

Carotenoids

1 2 3 4 5

Normal water supply Drought stress

mg g

• 1 FM

Nazilli 84 (S) Sahin 2000 (T)

Higher content of proline and carotenoids in the drought-tolerant ( ) than in the sensitive ( ) genotype:

• at normal water supply
• at drought

The efficacy of the antioxidant defense in the drought-tolerant genotype is evidenced by: lower membrane damage better physiological performance as compared to the sensitive genotype

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MDA

0,01 0,02 0,03 0,04 0,05 0,06

Normal water supply Drought stress

mol g-1 FM

Nazilli 84 (S) Sahin 2000 (T)

Lower membrane damage (malonyl dialdehyde content, MDA) in the drought-tolerant ( ) than in the sensitive ( ) genotype at:

• normal water supply
• drought

Photosynthetic parameters in drought-subjected plants as % of the plants grown at normal water supply

Better photosynthetic performance in the drought-tolerant ( ) than in the sensitive ( ) genotype

%

20 40 60 80 100 Nazilli 84 (S) Sahin 2000 (T) Photosynthesis Max PS2 activity WUE

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RWC

60 65 70 75 80 85 Normal water supply Drought stress

%

Nazilli 84 (S) Sahin 2000 (T)

Higher relative water content (RWC) in the drought-tolerant ( ) than in the sensitive ( ) genotype at:

• normal water supply
• drought

Conclusion

Cotton plants employ an effective versatile network of antioxidant compounds for defense against various abiotic stress constraints.

The data obtained contribute to the understanding of

the biochemical bases of abiotic stress tolerance in plants.

They can serve as a rationale in modeling and

engineering abiotic stress tolerant cotton crops.

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Acknowledgements

The financial support of Tübitak, Turkey, and the Bulgarian Academy of Sciences is acknowledged.