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Growth of Tomato (Lycopersicon esculentum)

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 Journal of Plant Interactions

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Growth of tomato (Lycopersicon esculentum) in

response to salicylic acid under water stress

Shamsul Hayat , Syed Aiman Hasan , Qazi Fariduddin & Aqil Ahmad

To cite this article: Shamsul Hayat , Syed Aiman Hasan , Qazi Fariduddin & Aqil Ahmad (2008)
Growth of tomato (Lycopersicon esculentum) in response to salicylic acid under water stress,
Journal of Plant Interactions, 3:4, 297-304

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Journal of Plant Interactions
Vol. 3, No. 4, December 2008, 297304

ORIGINAL ARTICLE
Growth of tomato (Lycopersicon esculentum) in response to salicylic acid under water stress
Shamsul Hayata*, Syed Aiman Hasana, Qazi Fariduddina and Aqil Ahmadb

a
Plant Physiology Section, Department of Botany, Aligarh Muslim University, Aligarh, India; bDepartment of Applied Sciences,
Higher College of Technology, Al-Khuwair, Sultanate of Oman
(Received 28 May 2008; final version received 3 July 2008)

Plants of Lycopersicon esculentum L. cv. K-25 were subjected to water stress by withholding water for 10 days at
20 (WS I) and 30 (WS II) days after sowing (DAS). Seedlings were sprayed with double distilled water (DDW) or
10
5M salicylic acid (SA) at 45 DAS. The water stress at earlier stage of growth (20 day stage) was more
inhibitory as compared to the later stage (30 day stage). The plants exposed to water stress exhibited a significant
(p B0.05) decline in photosynthetic parameters, membrane stability index (MSI), leaf water potential, activity of
nitrate reductase (NR), carbonic anhydrase (CA), chlorophyll and relative water content (RWC). A follow-up
treatment with SA protected against the stress generated by water and significantly improved the above
parameters. However, proline content and antioxidant enzymes increased under drought as well as under SA
treatments.
Keywords: antioxidants; electrolyte leakage; lipid peroxidation; membrane stability index; photosynthesis;
water potential

Introduction nificant increase in response to water deficit in

Salicylic acid (SA) is a phenolic compound that had
been recognized as a plant hormone (Hayat and
Ahmad 2007). It play a diverse physiological role in
plants which include plant growth (Khan et al. 2003)
thermogenesis, flower induction, nutrient uptake,
ethylene biosynthesis (Hayat and Ahmad 2007),
stomatal movement (Larque-Saavedra 1979) and
photosynthesis (Fariduddin et al. 2003). The disease
resistance and abiotic stress tolerance are the other
roles assigned to SA (Janda et al. 2007). Among
abiotic stresses, SA has been reported to counter
heavy metal stress (Choudhary and Panda 2004), low
temperature (Tasgin et al. 2003), high temperature
(He et al. 2005) and salinity (Yusuf et al. 2008).
Plants experience various type of environmental
stresses such as salt stress (Sairam et al. 2003),
heavy metal stress (Hasan et al. 2008), UV-B-
radiation (Casati et al. 2002), chilling (Van Heerden
and Kruger 2002) drought stress (Porcel et al. 2003).
Among them drought stress or water stress is a
major abiotic factor that limits photosynthesis by
causing damage to the chloroplast and inactivating
electron transfer reaction which in turn result in the
formation of active oxygen species (AOS) that can
lead to photo-oxidative damage (Asada 1999).
Moreover, water deficit generate oxidative stress
that results from a disturbance of pro-oxidant
antioxidant balance in the cell (Reddy et al. 2005).
The activity of antioxidant enzyme, namely, catalase
(CAT), peroxidase (POX), superoxide dismutase
(SOD), glutathione reductase (GR) exhibits a sig-
mulberry cultivars (Reddy et al. 2005) and wheat
seedlings (Dalmia and Sawhney 2004). Moreover it
also causes damage to the membrane stability index
(Nayyar and Walia 2003) which in turn causes
many physiological and biochemical alterations in
plant.
Rapid and effective measures of plant treatment
are necessary so that deterioration of crops due to
stresses can be countered successfully. The present
research was designed with an objective to evaluate
the changes in antioxidants under the influence of
salicylic acid in the Lycopersicon esculentum L. plants,
exposed to water stress and to establish a relationship
between antioxidant and the degree of tolerance. The
hypothesis tested is that salicylic acid will ameliorate
the adverse effects of water stress.
Material and methods
Plant material and growth conditions
The seeds of Lycopersicon esculentum L. cv. K-25
were purchased from Local Seed Store. Healthy seeds
were surface sterilized with 5% sodium hypochlorite
followed by repeated washing with distilled water.
The seeds were sown in earthen pots (6 inch
diameter), filled with sandy loam and farmyard
manure (ratio 6:1) to create a nursery. At the 15-
day stage these seedlings were uprooted and trans-
planted to the maintained pots under similar condi-
tions, as in the case of nursery pots. The plants were
grown in a net house under normal environmental

*Corresponding author. Email: shayat@lycos.com

ISSN 1742-9145 print/ISSN 1742-9153 online
# 2008 Taylor & Francis
DOI: 10.1080/17429140802320797
http://www.informaworld.com
298 S. Hayat et al.
conditions. The average temperature, humidity, and
day/night photoperiods were 26928C, 6595% and
10 h, respectively. Stress was imposed by withholding
water. At the 20-day stage plants were divided into
different sets. In the first set, water stress was given to
20-day old seedling by withholding water for
10 days (WS I). In the second set water stress was
given to 30-day-old seedling by withholding water for
10 days and in the third set, control plants was
maintained by supplying water at regular intervals.
The plants were allow to grow and at day 45, the
foliage had 0 or 10
5 M SA supplied (Fariduddin
et al. 2003). Stress plants were divided into two
groups. In the first it was sprayed with water and in
the second it was not sprayed. The results of both the
groups are the same, therefore, only the stress plants
sprayed with water were presented in the tables and
figures. Plant samples were collected 24 h after spray,
i.e. at day 46 to assess the parameters discussed
below.
Chlorophyll content and net photosynthetic rate
The chlorophyll in the fresh leaf samples was
measured by using Minolta chlorophyll meter
(SPAD-502, Konica Minolta Sensing Inc. Japan),
whereas the water use efficiency, internal CO2 con-
centration, stomatal conductance, transpiration rate
and photosynthetic rate were measured by LI-COR
6400 portable photosynthesis system (LI-COR, Lin-
coln, NE, USA).
Membrane stability index (MSI)
MSI was estimated by taking 200 mg leaf material in
10 cm3 of DDW in two sets. One set was heated at
408C for 30 min in a water bath and the electrical
conductivity bridge C1 was measured using a con-
ductivity meter. The second set was boiled at 1008C in
a water bath for 10 min and its conductivity was also
measured as conductivity bridge C2. MSI was calcu-
lated using the formula (Sairam 1994).
MSI[1
(C1 =C2 )]100:
Electrolyte leakage
The total inorganic ions leaked out in the leaves were
estimated by the method of Sullivan and Ross (1979).
Twenty leaf discs were taken in a boiling tube
containing 10 ml of deionized water for 10 min and
filter with Whatman filter paper and the filtrate was
measured for their electrical conductivity (EC) and
denoted as ECa. The filtrate was again heated at 558C
for 30 min in a water bath and electrical conductivity
was again measured and denoted as ECb. Later the
filtrate was again boiled at 1008C for 10 min and
electrical conductivity was again recorded and de-
noted as ECc. The electrolyte leakage was calculated
by using the formula:
ECb
ECa
Electrolyte leakage (%) 100:
ECc
Estimation of carbonic anhydrase (CA) and nitrate
reductase (NR) activities
The activity of carbonic anhydrase (CA) was
determined following the procedure described by
Dwivedi and Randhawa (1974). The leaf samples
were cut into small pieces and suspended in cystein
hydrochloride solution. The sample was incubated at
48C for 20 min. The pieces were blotted on the filter
paper and transferred to the test tubes, containing
phosphate buffer (pH 6.8) followed by the addition
of alkaline bicarbonate solution and bromothymol
blue indicator. The test tube was incubated at 58C
for 20 min. The reaction mixture was titrated
against 0.05N HCl after addition of 0.2 ml of
methyl red indicator.
The activity of nitrate reductase (NR) was mea-
sured following the method of Jaworski (1971). The
fresh leaf samples were cut into small pieces and
transferred to plastic vials containing phosphate
buffer (pH 7.5) followed by the addition of potassium
nitrate and isopropanol solutions. The reaction
mixture was incubated at 308C, for 2 h followed
with the addition of N-1-naphthyletylenediamine
dihydrochloride and sulphanilamide. The absorbance
of the colour was read at 540 nm.
Relative water content (RWC)
The relative water content (RWC) was determined in
fresh leaf discs of 2 cm diameter, excluding midrib.
Discs were weighed quickly and immediately floated
on double distilled water, in Petri dishes to saturate
them with water for the next 24 h, in dark. The
adhering water of the discs was blotted and turgor
weight was taken. Dry mass of these discs was
obtained, after dehydrating them at 708C for 48 h.
Relative water content was calculated as described
earlier (Jones and Turner 1978).
Leaf water potential
Leaf water potential of the fresh leaf sample was
measured by PSYPRO water potential system (WES-
COR, Inc, Logan, USA).
Lipid peroxidation
Lipid peroxidation rates were estimated by measuring
the malondialdehyde equivalents according to
Hodges et al. (1999). Some 0.5 g of the leaf was
homogenized in a mortar with 80% ethanol. The
homogenate was centrifuged at 3000 g for 10 min at
48C. The pallet was extracted twice with the same
solvent. The supernatants were pooled and 1 ml of
this sample was added to a test tube with an equal
volume of the solution comprised of 20% trichlor-
oacetic acid, 0.01% butylated hydroxy toluene and
 Journal of Plant Interactions
0.65% thiobarbutyric acid. Samples were heated at
958C for 25 min and cooled to room temperature.
Absorbance of the samples was recorded at 440, 532
and 600 nm. Lipid peroxidation rates (n mol mal-
ondialdehyde ml
1) were calculated by using the
formula given by Hodges et al. (1999).
Assay of antioxidant enzymes
Leaf tissue (0.5 g) was homogenized in 5 ml of 50 m
mol phosphate buffer (pH 7.0) containing 1% poly-
vinylpyrolidone. The homogenate was centrifuged at
15,000 rpm for 10 min and the supernatant was used
as the source of enzyme. The extraction was carried
out at 48C. Peroxidase and catalase was assayed
following the procedure described by Chance and
Maehly (1955). Catalase was estimated by titrating
the reaction mixture, consisting of phosphate buffer
(pH 6.8), 0.1M H2O2, enzyme extract and 2% H2SO4
against potassium permanganate. The reaction mix-
ture for peroxidase consisted of pyrogallol phosphate
buffer (pH 6.8), 1% H2O2 and enzyme extract.
Change in absorbance, due to catalytic conversion
of pyrogallol to perpurogallin, was noted at an
interval of 20 s for 2 min at 420 nm. A control set
was prepared by using distilled water instead of
enzyme extract.
The activity of superoxide dismutase was assayed
by measuring its ability to inhibit the photochemical
reduction of nitroblue tetrazolium (NBT) using the
method of Beauchamp and Fridovich (1971). The
reaction mixture containing 50 m mol phosphate
buffer (pH 7.8) 13 m mol methionine, 74 m mol
NBT, 2 m mol riboflavin, 0.1 m mol EDTA and 0.5
ml enzyme extract and was placed under a 15 W
fluorescent lamp. The reaction was started by switch-
ing on the light and was allowed to run for 10 min.
The reaction was stopped by switching off the light;
50% inhibition by light was considered as one
enzyme unit.
Estimation of proline content
The proline content in fresh leaf was determined by
adopting the method of Bates et al. (1973). A sample
was extracted in sulphosalicylic acid. To the extract,
an equal volume of glacial acetic acid and ninhydrin
solutions were added. The sample was heated at
1008C, to which 5 ml of toluene was added. The
absorbance of toluene layer was read at 528 nm, on a
spectrophotometer.
Statistical analysis
The experiment was conducted according to simple
randomized block design. A total of 10 replicates for
each treatment were taken. Treatment means were
compared by analysis of variance using SPSS (SPSS,
Chicago, IL, USA). Least significance difference
(LSD) was calculated at the 5% level of probability.
299
The treatments were separated by different letters.
Standard error of the replicate was also calculated.
Results
Photosynthesis and related parameters
Water stress significantly (p B0.05) decreased all the
photosynthetic parameters, i.e. net photosynthetic
rate (PN) stomatal conductance (gs), internal carbon
dioxide concentration (Ci), water use efficiency
(WUE) and transpiration rate. Plants that are sub-
jected to water stress at day 20 possessed 28.7%,
50.9%, 14.8%, 41.8% and 17.6% lower values over
the control and at 30 day it decreased the values by
21.6%, 45.1%, 10.9%, 34.3%, and 14.2%, respec-
tively, over the non-stressed plants. The SA spray
neutralized the damage caused by the 30-day stage of
the growth to a greater extent and evoked the values
26.4%, 77.1%, 11.1%, 35.5%, and 13.4% over the
non-sprayed stressed plants (Figure 1; Table 1).
Chlorophyll content
Plants receiving SA alone possessed the maximum
value for SPAD chlorophyll and measured 27.5%
significantly (pB0.05) higher than that of the control
(Figure 2a). However, application of water stress to
the plants decreased the value of SPAD. The follow-
up treatment with SA partly overcame the negative
effects generated by water stress (20-day stage) and
almost completely by the water stress at 30-day stage.
Membrane Stability Index (MSI)
Plants subjected to water stress at 20- and 30-day
stages of growth possessed 4.3% and 13.0% lower
values for MSI as compared to the control and were
statistically significant (p B0.05). The treatment of
the plants with SA resulted in a significant (pB0.05)
increase in the MSI. However non-stressed plant
receiving SA spray alone possesses maximum values
(Figure 2c).
Relative water content (RWC) and leaf water
potential
Plants treated with SA alone possessed maximum
values for leaf water potential and was about 20.9%
higher than that of the control (Table 1). However,
application of water stress lowered leaf water poten-
tial. The follow-up treatment with SA partially over-
came the effects generated by water stress (20- or 30-
day stage), resulting in increased values of 23.02%
and 58.2% respectively, over the control. A similar
pattern was also recorded for RWC (Table 1).
Carbonic anhydrase and nitrate reductase activities
The activity of CA decreased in the plants that are
subjected to water stress and it was about 12.6%
lower when compared to the control at the 20-day
300 S. Hayat et al.

(a) 7 LSD at 5 % = 0.68

a
6
b bc
PM (M CO2 m-2 s-1)

5 cd
e e
4
3
2
1
0

(b) 0.14 LSD at 5 % = 0.015

0.12 a
ab
ab
0.1 b
gs (mol m s )
2 -1

0.08
c c
0.06
0.04
0.02
0

(c) 400 LSD at 5 % = 13

a
300 b b
d cd c
Ci (ppm)

200

100

0
WS I +
WS II

WS II
WS I
Control

+ SA
SA

SA

Figure 2. Effect of salicylic acid (SA) on the leaf (a) SPAD

Treatments value, (b) electrolyte leakage (%) and (c) membrane
stability index (%), of Lycopersicon esculentum at the 46-
Figure 1. Effect of salicylic acid (SA) on the (a) photo- day stage subjected to water stress WS1 (20-day stage),
synthetic rate (PN), (b) stomatal conductance (gs) and (c) WS2 (30-day stage). Control means sprayed with water
internal CO2 (Ci), in the leaf of Lycopersicon esculentum at only. Vertical bar shows9SE and treatments are separated
the 46-day stage subjected to water stress WS1 (at 20-day by different letters.
stage), WS2 (30-day stage). Control means sprayed with
water only. Vertical bar shows9SE and treatments are ill effect of water stress partially at both the stages
separated by different letters. (Figure 3a). Under ambient conditions treatment of
the plants with SA enhanced activity of NR (Figure
stage stress and the values of CA for 30-day stress 3b). However, application of water stress resulted in a
was statistically equal to the values of 20-day stage significant decrease in enzyme activity. Moreover, the
stress. The follow up treatment of SA neutralized the follow-up treatment with SA resulted in significant
Table 1. Effect of 10
5M of Salicylic acid (SA) on the leaf proline (mg g
1 F.M.), relative water content (RWC) (%), water
use efciency (WUE), transpiration rate (mol m
2 s
1), water potential (mpa), of Lycopersicon esculentum at the 46-day stage
subjected to differential water stress WS1 (at the 20-day stage) WS2 (30-day stage), control means sprayed with water only,
9standard error; LSDleast signicant difference.

Proline RWC WUE Transpiration Water potential

Control 14.5991.06e 75.6090.328b 0.13790.019b 1.4790.238b
1.0390.10de

SA 16.15 91.07d 80.9490.355a 0.19990.023a 2.2990.317a
0.9090.080e
WS I 18.6091.39bc 65.7990.428c 0.08090.010d 1.2190.380e
2.1790.23a
WS II 17.8791.00c 60.5090.235d 0.09090.020d 1.2690.417de
1.9590.25b
WS ISA 20.1091.01a 74.8190.395b 0.10690.050c 1.3590.416cd
1.0890.26cd
WS IISA 19.2191.02b 67.7290.241c 0.12290.011b 1.4390.350c
1.2190.13c
LSD at 5% 0.75 3.8 0.015 0.11 0.16
 Journal of Plant Interactions
(p B0.05) improvement in enzyme activity in plants
that were subjected to water stress.
Electrolyte leakage and lipid peroxidation
These two parameters show completely different
responses to stress as compared to other parameters.
Plants that are subjected to water stress at the 30-
day stage of growth induced more electrolyte
leakage (42.1%) and lipid peroxidation (9.2%) as
compared to the plant experiencing water stress
at the 20-day stage, where the values were, 26.3%
(electrolyte leakage) and 6.1% (lipid peroxidation)
higher, over the control. However, the follow-up
treatment of these stressed plants with SA caused a
remarkable decrease in both these parameters
(Figures 2b, 3c).
Figure 3. Effect of salicylic acid (SA) on the leaf (a)
carbonic anhydrase (CA), (b) nitrate reductase (NR) and
(c) lipid peroxidation, of Lycopersicon esculentum at day 46
subjected to water stress WS1 (at day 20), WS2 (day 30).
Control means sprayed with water only. Vertical bar
shows9SE and treatments are separated by different letters.
301
Antioxidant enzymes and proline content
The activities of antioxidant enzymes (Peroxidase,
catalase, superoxide-dismutase) were significantly
(p B0.05) enhanced when the plants were exposed
to either of water stress and SA. The maximum
activities of these enzymes were recorded from the
plants that were exposed to water stress at 30 day
stage and also receiving SA. Control plants had the
minimum values for these enzymes (Figure 4).
The plants incubated under controlled conditions
possessed the lowest level of proline (Table 1).
However, proline level increased, both in response
to water stress as well as to SA treatment. Interaction
of SA with water stress significantly (pB0.05) raised
proline levels. Plants receiving SA in association with
water stress at day 30 showed maximum levels of
proline.
Figure 4. Effect of salicylic acid (SA) on the activities of (a)
peroxidase, (b) catalase and (c) SOD, in the leaves of
Lycopersicon esculentum at day 46 subjected to water stress
WS1 (at day 20), WS2 (day 30). Control means sprayed
with water only. Vertical bar shows9SE and treatments are
separated by different letters.
302 S. Hayat et al.
Discussion
Plant growth and development is a complex phenom-
enon and determined by several endogenous and
exogenous factors. Among the internal factors, hor-
mones play a vital role in regulating growth and
development. The role of SA in plant growth regula-
tion was the first physiological process to be reported
(Dekock et al. 1974). Thereafter, both exogenous and
endogenous levels of SA have been shown to impart
various effects (Raskin 1992).
When the plants are subjected to drought stress
a variety of physiological and biochemical changes
occur that can be used as markers against stress. In
the present study it was observed that water stress
reduced SPAD chlorophyll value, photosynthetic
attributes, NR, CA activities and leaf water poten-
tial (Figures 13, Table 1) and hence may ultimately
lead to an overall decrease in crop yield. Decrease in
photosynthetic rate mediated by water stress was
the consequence of the closure of stomata, thereby
decreasing CO2 supply as well as internal CO2
concentration (Tiwari et al. 2005) and consequently
a decrease in the activity of CA (Figure 3a), because
its activity, to a large extent, is regulated by the
CO2 concentration (Tiwari et al. 2005). Moreover
water stress also inhibits the activity of Rubisco
(Yardanov et al. 2003) and that of other enzymes of
photosynthesis (Lawlor 2002; Reddy et al. 2005).
However, the application of SA alone enhanced the
chlorophyll content and photosynthetic rate and
also neutralized the negative effect of the stress
(Figure 1a, 2a). The increase in the photosynthetic
rate by SA may be the result of enhanced activity of
Rubisco and PEP carboxylase under stress (Popova
et al. 2003; Singh and Usha 2003) and that of CA
(Figure 3a). The SPAD chlorophyll in the plants
treated with SA also increased (Figure 2a) which
may be another reason for the increase in photo-
synthesis. Similar results were also reported by
Singh and Usha (2003).
The decreased in the NR activity in the stressed
plant may be considered a biochemical adaptation to
conserve energy by stopping nitrate assimilation. The
reason that seems most appropriate to explain the SA
mediated elevation in the activity of nitrate reductase
(Figure 3b) is that it corrects the stress mediated
damage to the plasma membrane, as evident from an
increase in the membrane stability index (Figure
2c).The membrane correction/stabilization could
have facilitated the increased uptake of nutrients
including that of nitrate, which act as an inducer of
NR (Campbell 1999). The increase in the uptake of
various nutrients including NO3 and activation of
NR, under normal condition is well established
(Hayat et al. 2005) which strongly support the present
findings.
When the plants are subjected to drought stress, a
variety of active oxygen species (AOS) are generated,
such as superoxide, H2O2 and hydroxyl radicals,
which cause oxidative damage to the plants. They
are toxic to living organisms and unless removed
rapidly, they destroy or inactivate various cellular
components (Foyer and Noctor 2002). Plants have
antioxidant enzymes to scavenge AOS; these enzymes
are postulated to be involved in stress tolerance of
plants. Senaratna et al. (2000) have suggested a
similar mechanism to be responsible for SA induced
multiple stress tolerance in bean and tomato plants.
Results of the present work on peroxidase, catalase
and super oxide dismutase enzymes involved in
antioxidant defense mechanism supports the above
reasoning (Figure 4).
Proline is another important component of the
defense system of the plants to counter stress.
According to Hong et al. (2000) proline is synthe-
sized by the enzyme D?-pyrroline-5-carboxylates
synthetase (D?-P5CS) and D?-pyrroline-5-carboxylate
reductase (D?-P5CR) and is subsequently metabo-
lized by the enzyme proline dehydrogenase (Pro
DH). The activities of the enzyme D?-P5CR are
reported to increase and that of Pro DH decrease
in cowpea grown under water stress (Sumithra and
Reddy 2004). Moreover, the genes responsible for
the transcription of these proteins were also over-
expressed in transgenic plants under stress (Kishore
et al. 1995). It was found that the level of proline
in the plants exposed to water stress was higher
than those of control (Table 1) and also in the SA
treated plants. It may be possible that
SA activated the enzymes of proline metabolism
and consequently enhanced the accumulation of
proline.
Similarly the leaf water potential show more
negative values as the intensity of the stress was
increased (Table 1). Similar observation was made in
different plant species subjected to drought, Medi-
cago sativa (Irigoyen et al. 1992) and wheat (Dalmia
and Sawhney 2004). However the SA spray improved
these values in the stressed plant at both the stages of
stress. The possible mechanism behind this may be
that SA spray reduced electrolyte leakage by improv-
ing membrane stability, as reflected from our results
(Figure 2b, 2c) as well as Agarwal et al. (2005), this in
turn may improve relative water content and leaf
water potential.
Conclusion
SA enhanced the level of antioxidant system (Perox-
idase, catalase, superoxide-dismutase and proline),
both under stress and stress-free conditions. The
influence of SA on antioxidant system was more
pronounced under stress situation, suggesting that
the elevated level of antioxidant system, at least in
part, increased the tolerance of tomato plants to
water stress, thus protected the photosynthetic ma-
chinery and may be the plant growth.
 Journal of Plant Interactions
Acknowledgements
The authors are thankful to the anonymous reviewers
for their valuable suggestions. We are also greatful to Dr
Zaki A. Siddiqui, Visiting Professor at Kyoto University,
Kyoto, Japan, for his valuable input for the statistical
analysis.
References
Agarwal S, Sairam RK, Srivastava GC, Tyagi A, Meena
RC. 2005. Role of ABA, salicylic acid, calcium and
hydrogen peroxide on antioxidant enzymes induction
in wheat seedlings. Plant Sci. 169:559570.
Asada K. 1999. The water-water cycle in chloroplasts:
scavenging of active oxygens and dissipation of excess
photons. Ann Rev Plant Physiol Plant Mol Biol.
50:601639.
Bates LS, Waldren RP, Teare ID. 1973. Rapid determina-
tion of free proline for water stress studies. Plant Soil.
39:205207.
Beauchamp CO, Fridovich I. 1971. Superoxide dismutase:
improved assays and assay applicable to acrylamide
gels. Anal Biochem. 44:276287.
Campbell HW. 1999. Nitrate reductase structure, function
and regulation. Bridging the gap between biochemistry
and physiology. Ann Rev Plant Physiol Plant Mol
Biol. 50:277303.
Casati P, Lara MV, Andreo CS. 2002. Regulation of
enzymes involved in C4 photosynthesis and antiox-
idant metabolism by UV-B radiation in Egeria denso, a
submerged aquatic species. Photosyn Res. 71:251264.
Chance B, Maehly AC. 1955. Assay of catalase and
peroxidase. Methods Enzymol. 2:764775.
Choudhury S, Panda SK. 2004. Role of salicylic acid in
regulating cadmium induced oxidative stress in Oryza
sativa L. roots. Bulg J Plant Physiol. 30:95110.
Dalmia A, Sawhney V. 2004. Antioxidant defense mechan-
ism under drought stress in wheat seedlings. Physiol
Mol Biol Plants 10:109114.
Dekock PC, Grabowska FB, Innes AM. 1974. The effect of
salicylic acid on the growth of Lemma gibba. Ann Bot.
38:903908.
Dwivedi RS, Randhawa NS. 1974. Evaluation of a rapid
test for the hidden hunger of zinc in plants. Plant Soil
40:445451.
Fariduddin Q, Hayat S, Ahmad A. 2003. Salicylic acid
inuences net photosynthetic rate, carboxylation ef-
ciency, nitrate reductase activity and seed yield in
Brssica juncea. Photosynthetica 41:281284.
Foyer CH, Noctor G. 2002. Oxygen processing in photo-
synthesis: regulation and signaling. New Phytol.
146:359388.
Hasan SA, Hayat S, Ali B, Ahmad A. 2008. 28-homo-
brassinolid protects chickpea (Cicer arietinum) from
cadmium toxicity by stimulating antioxidant. Environ
Poll. 151:6066.
Hayat S, Ahmad A. 2007. Salicylic acid  a plant hormone.
The Netherlands: Springer.
Hayat S, Fariduddin Q, Ali B, Ahmad A. 2005. Effect of
salicylic acid on growth and enzyme activities of wheat
seedlings. Acta Agron Hung. 53:433437.
303
He Y, Liu Y, Cao W, Huai M, Xu B, Huang B. 2005. Effect
of salicylic acid on heat tolerance associated with
antioxidant metabolism in Kentucky Blue grass.
Crop Sci. 45:988995.
Hodges MD, DeLong JM, Forney CF, Prange RK. 1999.
Improving the thiobarbituric acid-reactive substances
assay for estimating lipid peroxidation in plant tissues
containing anthocyanin and other interfering com-
pounds. Planta. 207:604611.
Hong Z, Kakkenent K, Zhang Z, Verma DP. 2000.
Removal of feed back inhibition of D?-pyrroline-S-
carboxylate synthetase results in increased accumula-
tion and protection of plants from osmotic stress. Plant
Physiol. 122:11291136.
Irigoyen JJ, Emerich DW, Sanchez-Diaz M. 1992. Water
stress induced concentration of proline and total
soluble sugars in nodulated alfalfa (Medicago sativa)
plants. Physiol. Plant. 84:5560.
Janda T, Horvath E, Szalai G, Paldi E. 2007. Role of
salicylic acid in the induction of abiotic stress toler-
ance. In: Hayat S, Ahmad A, editors. Salicylic acid: a
plant hormone. The Netherlands: Springer.
Jaworski EG. 1971. Nitrate reductase assay in intact plant
tissue. Biochem Biophys Res Commun. 43:12741279.
Jones MM, Turner NC. 1978. Osmotic adjustment in leaves
of sorghum in response to water decits. Plant Physiol.
61:122126.
Khan W, Prithiviraj B, Smith D. 2003. Photosynthetic
responses of corn and soybean to foliar application of
salicylates. J Plant Physiol. 160:485492.
Kishore RBK, Hong Z, Miao GH, Hu CAA, Verma DP.
1995. Over-expression of D?-pyrroline-5-carboxylate
synthetase increase proline production and confers
osmotolerance in transgenic plants. Plant Physiol.
108:13871394.
Larque-Saavedra A. 1979. Stomatal closure in response to
acetalsalicylic acid treatment. Z Panzenphysiol.
93:371375.
Lawlor DW. 2002. Limitation to photosynthesis in water-
stressed leaves: stomatal metabolism and the role of
ATP. Annals Bot. 89:871885.
Nayyar H, Walia DP. 2003. Water stress induced proline
accumulation in contrasting wheat genotypes as af-
fected by calcium and abscisic acid. Biol Plant. 46:
275279.
Popova L, Ananiewa E, Hristova V, Christov K, Georgieva
K, Alexieva V, Stoinova ZH. 2003. Salicylic acid and
methyl jasmonate-induced protection on photosynth-
esis to paraquat oxidative stress. Bulg J Plant Physiol
(Special Issue):133152.
Porcel R, Barea JM, Rui-Lozano JM. 2003. Antioxidant
activities in mycorrhizal soybean plants under drought
stress and their possible relationship to the process of
nodule senescence. New Phytol. 157:135143.
Raskin I. 1992. Role of salicylic acid in plants. Ann Rev
Plant Physiol Plant Mol Biol. 43:439463.
Reddy AR, Chaitanya KV, Jutur PP, Granam A. 2005.
Photosynthesis and oxidative stress responses to water
decit in ve different mulberry (Morus alba L.)
cultivars. Physiol Mol Biol Plants 11:291298.
Sairam RK. 1994. Effect of homobrassinolide application
on plant metabolism and grain yield under irrigated
and moisture stress condition on two wheat varieties.
Plant Growth Regul. 14:173181.
 304 S. Hayat et al.
Sairam RK, Singh DV, Srivastava GC. 2003. Changes in
activities of antioxidant enzymes in sunower leaves of
different age. Biol Plant. 47:6166.
Senaratna T, Touchell O, Bunn E, Dixon K. 2000. Acetyl
salicylic acid (Aspirin) and salicylic acid induce multi-
ple stress tolerance in bean and tomato plants. Plant
Growth Regul. 30:157161.
Singh B, Usha K. 2003. Salicylic acid induced physiological
and biochemical changes in wheat seedlings under
water stress. Plant Growth Regul. 39:137141.
Sullivan CY, Ross WM. 1979. Selecting the drought and
heat resistance in grain sorghum. In: Mussel H, Stapes
RC, editors. Stress physiology in crop plants. New
York: John Wiley and Sons. p. 263281.
Sumithra K, Reddy AR. 2004. Change in proline metabo-
lism of cowpea seedlings under water decit. J Plant
Biol. 31:201204.
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Tasgin E, Atici O, Nalbantoghu B. 2003. Effect of salicylic
acid and cold on freezing tolerance in wheat leaves.
Plant Growth Regul. 41:231236.
Tiwari A, Kumar P, Singh S, Ansari SA. 2005. Carbonic
anhydrase in relation to higher plants. Photosynthe-
tica. 43:111.
Van Heerden PDR, Kruger GHJ. 2002. Separately and
simultaneously induced dark, chilling and drought
stress effect on photosynthesis, proline accumulation
and antioxidant metabolism in soybean. J Plant
Physiol. 159:10771086.
Yardanov I, Velikova V, Tsonev T. 2003. Plant responses
to drought and stress tolerance. Bull J Plant Physiol
(Special Issue):187206.
Yusuf M, Hasan SA, Ali B, Hayat S, Fariduddin Q, Ahmad
A. 2008. Effect of salicylic acid on salinity induced
changes in Brassica juncea. J Integrative Plant Biol.
50(8):14.