Role of Paclobutrazol and ABA in Drought Stress Amelioration in Sesamum indicum L.

Transcription

1 Global Journal of Molecular Sciences 4 (2): 56-62, 29 ISSN IOSI Publications, 29 Role of Paclobutrazol and in rought Stress Amelioration in Sesamum indicum L ,3 1 R. Somasundaram, Cheruth Abdul Jaleel, Sindhu S. Abraham, M.M. Azooz and R. Panneerselvam 1 Stress Physiology Lab, epartment of Botany, Annamalai University, Annamalainagar 68 2, Tamilnadu, India 2 epartment of Botany, Faculty of Science, South Valley University, Qena, Egypt 3 epartment of Biology, Faculty of Science, King Faisal University, P.O. Box: 38, Al-Hassa 31982, Saudi Arabia Abstract: In this study, paclobutrazol, a triazole growth regulator and abscisic acid (), a traditional growth regulator were used to analyse their role in water stress amelioration in drought stressed Sesamum indicum L. plants. The main aspects studied were the antioxidant enzyme profile changes in different parts of treated, drought stressed as well as control plants. The enzymes superoxide dismutase (SO), ascorbate peroxidase (APX) and peroxidase (POX) were analysed. It is found that, these growth regulators can be used as stress ameliorating agents in this crop plant, as they increase the antioxidant enzyme activities and there by providing stress tolerance ability to the plant. Key words: Superoxide dismutase Ascorbate peroxidase Peroxidase rought stress Sesamum indicum INTROUCTION to drought is essential for a holistic perception of plant resistance mechanism to water limited condition in higher Sesame (Sesamum indicum L.) is one of the most plants [81]. For the last few decades, several scales of important oilseed crops and has been cultivated in physiological works have been conducted under drought Korea since ancient times for use as a traditional stress in crop plants [46]. health food. The sesame crop is usually grown under Triazole compounds are the chemicals belongs to a rain-fed conditions, where because of low and irregular class of compounds known as ergosterol biosynthesis precipitation is regularly subjected to mild to severe inhibitors and are used as fungicides as wells as plant water-deficit stress [1]. Although sesame has good growth regulators [179]. Triazoles have been called drought tolerance compared with many other crops, it is plant multiprotectants because of their ability to induce particularly susceptible to drought damage during both tolerance in plants to environmental and chemical stresses the seedling and early flowering stage [2]. [2-23]. The inhibition of gibberellin biosynthesis is the Water is one of the most important ecological factors main reason behind the PGR properties of triazoles. determining crop growth and development; water deficit Growth substance like ethylene, cytokinin and are plays a very important role in inhibiting the yields of also affected by the triazoles [16-2]. Protection of plants crops [3-5]. rought stress occurs when the available from apparently unrelated stress by triazole is also water in the soil is reduced and atmospheric conditions mediated by a reduction in free radical damage and cause continuous loss of water by transpiration or increase in the antioxidant potential and have an efficient evaporation [6]. Water deficit (commonly known as free-radical scavenging system that enables them to drought) can be defined as the absence of adequate detoxify active oxygen [18-24]. moisture necessary for a plant to grow normally and Exogenous application of was able to complete its life cycle [7-9]. One way to ensure future food increase plant adaptive response to various needs of the increasing world populations should involve environmental conditions. The resurrection plant, a better use of water by the development of crop varieties Craterostigma plantaginem can tolerate extreme which needs less amount of water and more tolerant to dehydration. However, in vitro propagated callus derived drought [12]. The biochemical and molecular responses from this plant has a strict requirement for exogenously Corresponding Author: r. R. Somasundaram, Stress Physiology Lab, epartment of Botany, Annamalai University, Annamalainagar 68 2, Tamilnadu, India 56

2 Global J. Mol. Sci., 4 (2): 56-62, 29 applied in order to survive severe dehydration [25]. saved and made up to 1 ml with extraction buffer and The plant hormone abscisic acid as a stress signal used for estimation of the SO enzyme activity. The increased as a result of water stress and play important enzyme protein was determined by Bradford [27] method. role in the regulation of plant responses from the whole plant level and the cellular level. Estimation: Superoxide dismutase activity was assayed as The objectives of the present study were to described by Beauchamp and Fridovich [28]. The reaction understand the effect of paclobutrazol and in medium was prepared and to 3 ml reaction medium, 1 ml of drought stress amelioration in Sesamum indicum L. enzyme extract was added. The reaction mixture contained through their effects on the plant s Antioxidant enzymes M riboflavin,.1 M methionine, 2 1 potassium -5 cyanide and M nitroblue tetrasodium salt (NBT), MATERIALS AN METHOS dissolved in.5 M sodium phosphate buffer (ph 7.8). The mixture was illuminated in glass test tubes by two The seeds of Sesamum indicum L. were surface sets of Philips 4 W fluorescent tubes. Illumination sterilized with.2% Mercuric chloride solution for five started to initiate the reaction at 3 C for one hour. Those minutes with frequent shaking and thoroughly washed without illumination saved as blank and kept in dark. The with tap water. The experiments were carried out in absorbance was read at 56 nm in the spectrophotometer polythene bags (27x16 cm). The pots were filled with 3 kg against blank. Superoxide dismutase activity was uniform soil mixture containing red soil: sand: farm yard expressed in units. One unit is defined as the amount of manure (FYM) in 1:1:1 ratio. The experiment was laid out change in the absorbance by.1 per hour per milligram in a Completely Randomized Block esign (CRB). protein under the assay condition [29]. In the preliminary experiments, 2, 5, 1, 15 and 2 mg L paclobutrazol and 5, 1 and 15 µm Ascorbate Peroxidase (APX, EC: ): Ascorbate were used for treatment to determine the optimum peroxidase was extracted and estimated by the method of concentration. Among the treatments, 5 mg L Nakano Asada [3]. paclobutrazol and 1 µm concentration increased the growth and dry weight significantly and higher Extraction: Five hundred milligrams of fresh plant tissue concentration slightly decreased the growth and dry was ground in a pestle and mortar under liquid nitrogen weight when compared to drought stressed plants. In the and 1 ml of 5mM potassium phosphate buffer (ph 7.) lower concentrations, there was no change in weight and containing 1 mm ETA, 1 per cent PVP and 1 mm growth. Hence 5 mg L paclobutrazol concentration was ascorbic acid. The homogenate was filtered through a used to study the effect of paclobutrazol and 1 µm double-layered cheesecloth and centrifuged at 15, rpm on the drought stress amelioration of Sesamum indicum. for 2 min at 4 C. The supernatant was used as source of rought Treatment intervals were from 3 AS 2, 4 enzymes. and 6 days interval drought (I). The treatments were given as foliar spray for and soil drenching for Estimation: One ml of reaction mixture contained 5 mm on 32, 34 and 36 days after sowing (AS). potassium phosphate buffer (ph 7.),.5 mm ascorbic The plants were taken randomly on 33 (2 I), acid,.1 mm H2O 2 and 2 µl of enzyme extract. The 35 (4 I) and 37 (6 I) AS and separated into roots, absorbance was read as decrease at 29 nm against stems and leaves and used for determining antioxidant the blank, correction was done for the low, non-enzymatic potentials. oxidation of ascorbic acid by H2O 2(extinction coefficient 2.9 mm cm ). The enzyme activity was expressed in µg Superoxide ismutase (SO, EC: ): Crude per gram dry weight. enzyme extract was prepared, for the assay of Superoxide dismutase by the method of Hwang et al. [26]. Peroxidase (POX, EC ): Peroxidase was assayed by the method of Kumar and Khan [31]. Assay mixture of Extraction: One gram of fresh tissue was homogenized Peroxidase contained 2 ml of.1 M phosphate buffer with 1 ml of ice-cold 5 mm sodium phosphate buffer (ph 6.8), 1 ml of.1 M pyrogallol, 1 ml of.5 M H2O2 containing 1 mm PMSF. The extract was filtered through and.5 ml of enzyme extract. The solution was incubated a double-layered cheesecloth. The extract was centrifuged for 5 min at 25 C after which the reaction was terminated at 12,5 rpm for 2 minutes at 4 C. The supernatant was by adding 1 ml of 2.5 N H SO. The amount of

3 Global J. Mol. Sci., 4 (2): 56-62, 29 purpurogallin formed was determined by measuring Leaf: SO activity increased in the leaves of the absorbance at 42 nm against a blank prepared by Sesamum indicum under drought condition. adding the extract after the addition of 2.5 N H2SO 4 at Paclobutrazol also resulted in increased SO zero time. The activity was expressed in unit mg protein. activity in leaves of Sesamum indicum. Paclobutrazol One unit is defined as the change in the absorbance by in combination with drought caused again.1 min mg protein. enhancement in SO activity when compared to stress and well-watered control plants. It was nearly per RESULTS cent over control on 6 I. treatment alone increased (163.3 per cent over control) the SO activity Superoxide ismutase (Fig. 1) significantly in leaves when compared to all other Root: The activity of SO increased in the root by water treatments. deficit in Sesamum indicum. It was nearly per cent over control on 6 I. Treatment with paclobutrazol and Ascorbate Peroxidase (Fig. 2) resulted in an enhancement of SO activity under Root: APX activity increased in roots of drought stress, which was 25.7 and percent over Sesamum indicum under drought condition. control respectively on 6 I. Paclobutrazol also resulted in increased APX activity in leaves of Sesamum indicum. Paclobutrazol in Stem: SO activity was found increased under water combination with drought caused again enhancement deficit condition in stem of Sesamum indicum. The SO in APX activity when compared to stress and activity increased greatly in paclobutrazol treated stem, well-watered control plants. It was nearly per cent which was and percent over control in over control on 6 I. treatment alone increased individual and drought stressed respectively. ( per cent over control) the APX activity caused an increased in SO activity and it was significantly in roots when compared to all other percent over control under drought stress on 6 I. treatments. Root Stem I 4 I 6 I I 4 I 6 I + + Leaf I 4 I 6 I + + Fig. 1: Individual and combined effects of drought,, on SO activity o f Sesamum indicum. [-rought, - Paclobutrazol, - Abscisic acid] 58

4 Global J. Mol. Sci., 4 (2): 56-62, 29 Root Stem I 4 I 6 I I 4 I 6 I + + Leaf I 4 I 6 I + + Fig. 2: Individual and combined effects of drought,, on APX activity of Sesamum indicum. [-rought, - Paclobutrazol, - Abscisic acid] Root Stem I 4 I 6 I I 4 I 6 I + + Leaf I 4 I 6 I + + Fig. 3: Individual and combined effects of drought,, on POX activity o f Sesamum indicum. [-rought, - Paclobutrazol, - Abscisic acid] 59

5 Global J. Mol. Sci., 4 (2): 56-62, 29 Stem: APX activity increased in stem of drought stressed and resulted in an enhancement of SO activity Sesamum indicum plants. The APX activity increased under drought stress. The SO activity increase under greatly in paclobutrazol treated stem, which was drought in Catharanthus roseus [6,9]. An increase in SO and percent over control in individual and drought activity was reported in Helianthus annuus plants under stressed respectively. caused an increased in APX water deficit stress and triazole application [13]. Water activity and it was percent over control in stem stress increased the SO activity in Abelmoschus under drought stress on 6 I. esculentus plants [12]. SO activity increased under drought stressed higher plants [5,6]. Antioxidant Leaf: APX activity increased in the leaves of Sesamum potentials increased in ioscorea rotundata Poir. indicum under drought condition. Paclobutrazol also following paclobutrazol drenching [23]. resulted in increased APX activity in leaves of Sesamum APX activity increased in Sesamum indicum indicum. Paclobutrazol in combination with drought under drought condition. Paclobutrazol also resulted in caused again enhancement in APX activity when increased APX activity in leaves of Sesamum indicum. compared to stress and well-watered control plants. It was Paclobutrazol in combination with drought caused again nearly per cent over control on 6 I. enhancement in APX activity when compared to stress treatment alone increased ( per cent over control) and well-watered control plants. Exogenous application of the APX activity significantly in leaves when compared to triadimefon affects the antioxidant defense system of all other treatments. Withania somnifera unal [32] and in Catharanthus roseus [33]. rought stress induced generation of Peroxidase (Fig. 3) active oxygen species is well recognized at the cellular Root: The peroxidase activity was showed an increase level and is tightly controlled at both the production and the root of Sesamum indicum under drought conditions. consumption levels through increased antioxidant Treatment with paclobutrazol and increased the systems [4-6]. Water stress and salt stress increased the peroxidase activity and it was per cent and APX activity in Catharanthus plants [8]. SO activity per cent over control respectively on 37 AP. increased under drought stressed Arachis hypogaea Paclobutrazol and in combination with plants [5]. Paclobutrazol treatment [34] and propiconazole drought increased the peroxidase activity and it was [35] increased the APX activity in Catharanthus plants per cent and 15 per cent over control respectively under salt stress. on 37 AP. The peroxidase activity was showed an increase the root of Sesamum indicum under drought conditions. Stem: Peroxidase activity increased in the stem of Treatment with paclobutrazol and increased the Sesamum indicum under drought condition. Paclobutrazol peroxidase activity. Wheat plants under water deficit also resulted in increased peroxidase activity in leaves of stress showed an enhancement in POX activity Sesamum indicum. Paclobutrazol in combination with irrespective of different genotypes [16]. Induction of drought caused again enhancement in peroxidase activity drought stress tolerance by ketoconazole in when compared to stress and well-watered control plants. Catharanthus roseus is mediated by enhanced It was nearly 29.2 per cent over control on 6 I. antioxidant potentials and secondary metabolite treatment alone increased ( per cent over control) accumulation [2]. Water deficit stress increased the the peroxidase activity significantly in stem when POX activity in Catharanthus roseus [1]. compared to all other treatments. REFERENCES Leaf: The peroxidase activity was increased in leaves of Sesamum indicum under drought condition. The 1. Ucan, K., F. Killi, C. Gencoglan and H. Merdun, peroxidase activity increased greatly in paclobutrazol 27. Effect of irrigation frequency and amount on treated leaf, which was and percent over water use efficiency andyield of sesame (Sesamum control in individual and drought stressed respectively. indicum L.) under field conditions. Field Crops Res., ISCUSSION 2. 11: Orruno, E., M.R.A. 27. Morgan Purification and characterisation of the 7S globulin storage protein The activity of SO increased in the root by water from sesame (Sesamum indicum L.). Food Chem., deficit in Sesamum indicum. Treatment with paclobutrazol 1:

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