Current Research Journal of Biological Sciences 4(4): 462-470, 2012
ISSN: 2041-0778
© Maxwell Scientific Organization, 2012
Submitted: March 14, 2012
Accepted: March 30, 2012
Published: July 10, 2012
Effects of Zeolite and Selenium Application on Some Physiological Traits and Oil
Yield of Medicinal Pumpkin (Cucurbita pepo L.) under Drought Stress
Kourosh Eskandari Zanjani, Amir Hossein Shirani Rad, Masoumeh Naeemi, Amin
Moradi Aghdam and Tofigh Taherkhani
Department of Agronomy and Plant Breeding, Takestan Branch, Islamic Azad University,
Takestan, Iran
Abstract: There is no doubt that certain plants offer valuable medicinal properties. But there are many
challenges that local communities face in the harvesting and cultivation of these plants. Medicinal pumpkin
(Cucurbita pepo L. var. Styriaca) is an economically important medicinal plant cultivated for oil production.
Water source limitation is one of the important problems in agriculture worldwide. Natural zeolits have some
properties such as water absorption and emission and nitrate leaching inhibition, which is useful for soil
amendment. This study was carried out to investigate the effects of zeolite and selenium application on some
physiological traits and oil yield of medicinal pumpkin (Cucurbita pepo L.) under drought stress. A field
experiment was carried out in a factorial based on a randomized complete block design with three replications.
The factors included: Irrigation (control, drought stress imposed during at flowering phase and drought stress
imposed during fruit formation), Zeolite (non-application and application of 10 ton/ha) and Selenium (nonapplication and 30 g/Lit) from sodium selenate. All physiological parameters were affected by drought stress
and zeolite application. Drought stress decreased total chlorophyll content, stomatal conductance, RWC, leaf
soluble protein, oil content and oil yield and zeolite application in drought stress increased mentioned traits
except oil content. Proline accumulation, WSD and CAT antioxidant enzyme activity were reduced in presence
of zeolite in drought stress conditions than under control. Selenium spraying with zeolite application in drought
conditions increased RWC, CAT antioxidant enzyme activity and oil yield. Drought stress at fruit formation
phase reduced oil content and oil yield more severe than that on flowering stage. Finally, it seems that zeolite
and selenium application in dry lands are exposed to drought stress be helpful for physiological traits and oil
yield improvement and prevention of decreased oil yield.
Keywords: Chlorophyll, drought, medicinal pumpkin, proline, stomatal conductance
management practices have been tried to reduce the
drought effects (Manivannan et al., 2007), but the
application of zeolite to discharged plants attracted little
attention. One possible approach to reducing the effect of
drought on plant productivity is through the addition of
zeolite to soil. Zeolite is a group of naturally occurring
minerals with physical and physicochemical properties
that can be used in such diverse areas as construction and
agriculture that can absorb and hold potentially harmful or
toxic substances. It also is capable of absorbing part of the
excess nutrients and also water, resulting in more
balanced macronutrient cation ratios in the root
environment and also can keep water in root zone (Savvas
et al., 2004). There are several zeolite types, one of them
being clinoptilolite, a hydrated aluminosilicate of alkali
and alkaline earth metals (Na+, K+, Ca2+, Mg2+, Ba2+)
having an infinite three-dimensional crystal structure, a
polyhedric shape and a great open cavity (Dakoviƒ
et al., 2007). Zeolites can be successfully used in
INTRODUCTION
Water is one of the most important ecological factors
determining crop growth and development; drought plays
a very important role in inhibiting the yields of crops
(Jaleel et al., 2007a). In recent years, many studies about
the effects of supplemental irrigation on yield
performance and Water Use Efficiency (WUE) have
shown that proper supplemental irrigation can increase
crop yield by significantly improving soil water
conditions and their WUE (Deng et al., 2002). Improving
the efficiency of water use in agriculture is associated
with increasing the fraction of the available water
resources that is transpired, because of the unavoidable
association between yield and water use (Jaleel et al.,
2007b). In regions where water scarcity is the principal
limiting factor for cultivation, farmers are interested in
using some methods to deduce injurious effects of water
deficiency. Chemical treatment and agronomical crop
Corresponding Author: Kourosh Eskandari Zanjani, Department of Agronomy and Plant Breeding, Takestan Branch, Islamic Azad
University, Takestan, Iran, Tel.: 989122607652
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Curr. Res. J. Biol. Sci., 4(4): 462-470, 2012
cultivating different crops such as cereals, forage crops,
vegetables, vine and fruit crops (Torii, 1978) due to their
exceptionally high ion-exchange capacity (Butorac et al.,
2002). Their porous structure assures a permanent water
reservoir in the root zone, improving the horizontal spread
of water after irrigation (Polat et al., 2004).
These abiotic stresses such as drought can result in
the accumulation of Reactive Oxygen Species (ROS) and
other toxic compounds (Xiong et al., 2002). Production of
ROS during environmental stress is one of the main
causes for decreases in productivity, injury and death that
accompany these stresses in plants (Upadhyaya and
Panda, 2004). Oxidative stress can prevent photosynthetic
activity, respiration process and plant growth (Sajedi
et al., 2011). Plants have developed the scavenging
mechanism of ROS categorized as enzymatic and nonenzymatic (Shalata et al., 2001). When ROS increases,
chain reactions start, in which Superoxide Dismutase
(SOD) and catalyzes the dismutation of O-2 radicals to
molecular O2 and H2O2. The H2O2 is then detoxified in the
ascorbate-glutathione cycle (Ouchi et al., 1990). Bailly
et al. (2000) reported that the content of Superoxide
Dismutase (SOD), Catalase (CAT) and glutathione
reductase (GR) in sunflower seeds will increase under
drought stress condition. In response to drought stress,
plants are able to control transpiration water loss by
reducing the leaf expansion rate to prevent dehydration of
leaf tissues (Liu and Stutzel, 2002).
Se is an essential trace element for animals and
humans (Tapiero et al., 2003) but its role in plants is still
unclear. Most cereal crops and fodder plant are relatively
weakly able to absorb Se, even when grown on soils with
higher Se content (Hegedus et al., 2001). Therefore, the
source potential of this intermediate element in plant can
be increased by increasing its application up to a normal
level (Nejat et al., 2009). It has been reported that
Selenium (Se) has an antioxidant effect and can increase
the antioxidative capacity and stress tolerance of plants
(Seppänen et al., 2003; Xu et al., 2001). There is
increasing evidence that Se has a positive effect on crop
growth and stress tolerance at low concentrations
(Rani et al., 2005; Turakainen et al., 2004). The specific
physiological mechanisms that underlie the positive
effects of Se in plants have not been clearly elucidated but
for senescing plants like maize and the antioxidative
effect of Se was closely associated with the increased
activity of glutathione peroxidase (GSH-Px) (Xu et al.,
2001). The Results showed that plant growth promoted by
Se is due to the increased starch accumulation in
chloroplasts (Pennanen and Hartikainen, 2002). Recently
it has been shown that Se can regulate the water status of
plants under conditions of water deficiency and thereby
performs its protective effect (Kuznetsov et al., 2003).
Medicinal pumpkin (Cucurbita pepo convar. pepo
var. styriaca) is an important annual plant that belongs to
the Cucurbitaceae family. Pumpkin seed oil has strong
antioxidant properties (Stevenson et al., 2007) and has
been recognized for several health benefits such as
prevention of the growth and reduction of the size of
prostate, retardation of the progression of hypertension,
mitigation of hypercholesterolemia and arthritis, reduction
of bladder and urethral pressure and improving bladder
compliance, alleviation of diabetes by promoting
hypoglycemic activity and lowering levels of gastric,
breast, lung and colorectal cancer (Stevenson et al.,
2007).
In order to meet the ever increasing demand of
medicinal plants, for the indigenous systems of medicine
as well as for the pharmaceutical industry, some
medicinal plants need to be cultivated commercially, but
the soil drought pose serious threats to plant production
(Jaleel et al., 2007b). Based on our knowledge,
information about the effects of drought stress on
physiological and agronomical traits changes in this
species are scarce. Therefore, the primary objective of the
present study was to find out the effect of drought stress
on physiological (leaf Relative Water Content (RWC),
leaf Water Saturation Deficient (WSD), stomatal
conductance, proline, chlorophyll, leaf soluble protein,
catalase enzyme activity) and agronomical (oil content
and oil yield) in medicinal pumpkin. The research was
aimed also whether zeolite soil application and selenium
foliar spraying might be strategies for increasing the
drought tolerance of plant.
MATERIAL AND METHODS
This experiments was carried out in the Agricultural
Research Station of Islamic Azad University, Takestan
Branch, Iran, during the growing season of 2010. This site
is located at 49º42’N latitude, 36º3 E longitude, with the
altitude of 1283 m above sea level in Qazvin Province in
the west of Iran. This region has a semi-arid climate on
the base of metrological data in Takestan, Iran in 2010.
Experiment was conducted in factorial based on
randomized complete block design with three replications.
Irrigation strategy had three levels: (1) 60% of
evaporation as control, (2) withheld irrigation at the
flowering stage and (3) withheld irrigation at fruit
formation stage. Soil application of zeolite factor had two
levels: (1) Non-application of zeolite as control and (2)
Zeolite application of 10 tons per hectare. Selenium was
sprayed in two concentrations: (1) zero and (2) 30 gram
per liter). Before seed sowing, multiple soil samples were
collected for determination of their physical and chemical
properties (Table 1). Seeds planted on four rows in each
plots, the rows distance was 2.5 m and the plant distance
on each row was 50 cm, it had used 150 kg/ha pure
nitrogen fertilizer and 60 kg/ha phosphorus and
potassium, for maximum exact, beginning and end of each
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Curr. Res. J. Biol. Sci., 4(4): 462-470, 2012
Table 1: Soil properties (0-30 cm) before plant sowing
OC
EC
Sand
Silt
Clay
Year (%)
pH
(dS/m) (%)
(%)
(%)
2010 0.73
7.1
1.20
62.1
22.0
16.3
Soil
texture
Sandy loam
P
K
(ppm) (ppm)
21.8
363
Fe
(ppm)
6.6
Cu
Zn
Mn
Total
(ppm) (ppm) (ppm) N(%)
1.8
3.7
24.3
0.07
plots closed, with regarding area of each plots, irrigation
intervals and water amounts used in each irrigation,
volume of water for each plot determined. In this research
for uniform Se spraying of each plot at first sprayer
machine calibrated with water and selenium selenit
sprayed by sodium selenit (Na2O3Se.5H2O) form in time
of 50% flowering and fruit formation stage, separately.
First and last rows of each plot considered as edge effect
and sampling for traits evaluation did from 2 central
Rows.
made on six leaf disks, 1 cm in diameter, from the same
leaves. The disks were weighed and floated on water for
6 h at a temperature of 46ºC in darkness. The disks were
blotted dry and turgid weight was determined and the dry
weight was measured after drying the disks for 24 h in
80ºC. RWC was calculated by the formula:
(Chartzoulalis et al., 2002):
Sampling: After drought stress treatment, three leaves of
each plant were removed. The samples were washed and
then frozen in liquid N2 and then stored at -80º C pending
biochemical analysis. Also, fruits of plants were harvested
from 5 m2 of central rows with ignoring border effect.
Seeds after separating from fruits, dried and then
weighted.
Stomatal conductance: Measurements of gs were made
using an AP4 Porometer (Delta-T Devices Ltd). The
measurements were taken between 12:00 and 13:00 h.
using five leaves from each treatment.
RWC = [(fresh weight - dry weight)/
(turgid weight - dry weight)] ×100
Chlorophyll content: Chlorophyll content were estimated
by extracting the leaf material (0.05 g) in 10-cm3
dimethylsulfoxide (DMSO) (Hiscox and Israelstam,
1979). The samples were heated at 65ºC in water bath and
for 4 hours and then absorbance of extract was recorded
at 665, 645 and 470 nm. Chlorophyll contents were
calculated as per standard method (Arnon, 1949).
Preparation of extracts: 0.500 g plant leaf tissue was
homogenized in a mortar and pestle with 3 mL ice-cold
extraction buffer (50 mM potassium phosphate, pH 7.0)
and 0.5% insoluble polyvynil pyrrilidone (the tissue/
buffer ratio of 1:6, w/v). The homogenate was centrifuged
at 15000 g for 30 min at 4ºC and then supernatant was
filtered through paper. The supernatant fraction was used
as a crude extract for the assay of enzyme activity. All
operations were carried out at 4ºC.
Oil content and oil yield: Seed samples per replicate plot
were ground with blender and three grams of dried
medicinal pumpkin seeds were extracted with petroleum
ether for 6 h in a Soxhlet system (B.chi Universal
Extraction System B-811, Germany) according to the
AOCS method (AOCS, 1993). The oil extract was
evaporated by distillation at a reduced pressure in a rotary
evaporator at 40ºC until the solvent was totally removed.
Oil yield was obtained multiplying the oil content and
seed yield.
Assay of antioxidant enzyme and protein: Catalase
(CAT, EC 1.11.1.6) activity was estimated by the method
of Aebi (1984). The reaction mixture contained 20 :L
crude enzyme extract, 250 :L 70 mM H2O2 and 250 :L
50 mM potassium phosphate buffer. The decrease in the
absorbance at 240 nm was recorded for 3 min by
spectrophotometer (Varian- UV- carry300, USA). CAT
activity of the extract was expressed as CAT units per
milligram of protein. Protein content in the supernatant
was quantified using bovine serum albumin as standard
(Bradford, 1976).
Data Analysis: All data were analyzed using SAS version
9.1. (SAS Inst., Cary, Nc) software. Each treatment was
analyzed in three replications. When analysis of variance
(ANOVA) showed significant treatment effects, Duncan’s
Multiple Range Test was applied to compare the means at
p<0.05.
Measurement of proline content: Determination of the
free proline levels was done according to Bates et al.
(1973). Proline was extracted in 3% (w/v) aqueous
sulfosalicylic acid. Extracts (2 mL) were held for 1 h in
boiling water by adding 2 mL ninhydrin and 2 mL glacial
acetic acid, after which cold toluene (4 mL) was added.
Proline content was measured at 520 nm and calculated as
mmol/g FW against standard proline.
RESULTS AND DISCUSSION
RWC and WSD: A significant decrease in the RWC and
increase in WSD observed in both drought stress
treatments as compare to normal irrigation (Table 2), that
this results agreement with Sepehri and Golparvar (2011).
According to results of Bayoumi et al. (2008), RWC may
be attributed to differences in the ability of the variation
to absorb more water from the soil and/or the ability to
control water loss through stomata. The greatest RWC
Relative Water Content (RWC) measurement:
Measurements of Relative Water Content (RWC) were
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Curr. Res. J. Biol. Sci., 4(4): 462-470, 2012
Table 2: Comparison of main and interaction effects on studied traits of medicinal pumpkin
RWC
WSD
Chlorophyll
Proline
Protein
Catalase activity Gs
Oil content Oil yield
(kg/ha)
Treatments
(%)
(%)
(mg/g fw)
(mmol/g fw)
(mg/g fw)
(u/mg.protein)
(mmol/m2/s) (%)
Irrigation
86.59a
13.41c
16.96a
11.37b
1.38a
5.71c
123.31a
43.92a
559.01a
I1
69.40b
30.59b
12.57b
25.99a
0.84b
23.12b
99.06 b
43.12ab
490.99b
I2
67.76c
32.13a
11.94c
25.63a
0.78c
27.55a
102.12b
42.33b
369.57c
I3
Zeolite (ton/ha)
0
69.92b
30.08a
12.81b
23.68a
0.87b
27.39a
100.34b
43.17a
445.16b
(Z0)
10
79.32a
20.68b
14.84a
18.31b
1.13a
10.20b
115.99a
43.08a
501.22a
(Z1)
Selenium (g/Lit)
0
72.28b
27.72a
13.87a
21.32a
0.95b
15.01b
107.55a
43.42a
459.69b
(S0)
30
76.33a
23.03a
13.67a
20.67b
1.05a
22.58a
108.77a
42.83a
486.68a
(S1)
86.04a
13.96d
17.18a
11.55c
1.35b
6.12d
124.75a
43.93a
550.35ab
I1 Z0
87.14a
12.85d
16.75a
11.20c
1.41a
5.30d
121.87a
43.92a
567.78a
I1 Z1
62.21d
37.79a
11.17c
30.06a
0.66d
34.62b
88.95d
42.47ab
456.47c
I2 Z0
76.58b
23.40c
13.97b
21.91b
1.02c
11.61c
109.16c
43.77a
525.51b
I2 Z1
61.52d
38.48a
10.08d
29.43a
0.59e
41.41a
87.32d
43.10ab
328.67e
I3 Z0
74.21c
25.79b
13.79b
21.83b
0.97c
13.68c
116.93b
41.56b
410.48d
I3 Z1
85.44b
14.56c
17.11a
11.00c
1.35b
4.38d
122.67a
43.65ab
545.35a
I1 S0
87.75a
12.25d
16.83a
11.75c
1.42a
7.05d
123.96a
44.20a
572.66a
I1 S1
66.46d
33.54a
12.73b
26.95a
0.78ed
18.87c
97.92b
43.85ab
487.64b
I2 S0
72.35c
27.65b
12.42bc
25.03b
0.91c
27.37b
100.21b
42.40bc
494.35b
I2 S1
64.93d
35.07a
11.78d
26.01a
0.73 e
21.78c
102.09b
42.78abc
346.10d
I3 S0
70.80c
29.20b
12.10dc
25.25b
0.83d
33.31a
102.17b
41.89c
393.05c
I3 S1
I1: Normal irrigation, I2: Withheld irrigation at flowering stage, I3: Withheld irrigation at fruit formation stage. Means at each column for each character,
followed by similar letters are not significantly different using Duncan's multiple range tests (p#0.05).
indicated that leaf RWC and/or proline content are useful
indicators for early detecting plant water-deficit stress
(Bates et al., 1973; Paknejad et al., 2009). Proline
accumulation during drought stress is an adaptive
response that enhances survival and tissue water status
(Chu et al., 1974). The accumulation of osmolyte
compounds like sugars and amino acids, especially
proline in the cells, as a result of water stress is often
associated with a possible mechanism to tolerate the
harmful effect of water shortage (Pirzad et al., 2011).
Zeolite application decreased the adverse drought
stress effects and caused to reduce the proline
accumulation in drought stressed medicinal pumpkin
plants by 22.6% (Table 2). Whereas, Se application
caused to decrease the proline accumulation (Table 2).
Means of comparison result showed that highest proline
content was obtained from Non-application of zeolite in
withheld irrigation at the flowering (30.06 mmol.g-1 fw).
Moreover, there was no significant difference between
Non-application of zeolite in withheld irrigation at fruit
formation and Non-application of zeolite in withheld
irrigation at the flowering, for proline content (Table 2).
Under normal irrigation conditions, Non-application or
application of zeolite was no significant effect on proline
content whereas, zeolite application caused to
considerable decrease the proline content, at drought
stress treatments at both flowering and fruit formation
phases. On the other hand, result of means comparison
indicated that proline content at Non-application Se in
withheld irrigation at the flowering was higher to Nonapplication or application Se in normal irrigation
conditions, considerably (Table 2). Also, Non-application
obtained from normal irrigation treatment (86.6%) and
drought stress caused to decrease the RWC by 20% at
withheld irrigation at the flowering stage and 22% at
withheld irrigation at fruit formation, respectively
(Table 2).
However, Zeolite application and Se spraying had
positive and desirable effect on RWC and increased it, but
zeolite application significantly decreased WSD trait.
Relatively higher RWC was noted in drought stress with
zeolite application than stress conditions without zeolite
indicating that zeolite have the ability to reserve and
maintain water in itself and so increases the water
availability to plant under stress conditions (Zahedi et al.,
2009).
Se spraying at drought stress conditions had desirable
effect on RWC and increased it. It was shown that Se has
the ability to regulate the water status of plants under
conditions of drought (Kuznetsov et al., 2003) and that
the protective effect of Se under drought stress conditions
was achieved by increasing the water uptake capacity of
the root system (Kuznetsov et al., 2003). Also in drought
stress treatments the highest amount of RWC and the
lowest of WSD were obtained in the presence of zeolite
and Se together (Table 3).
Proline: Proline contents in fresh leaves of irrigations
treatments are shown in Table 2. A minimum Proline
content was obtained from normal irrigation condition and
drought stress significantly enhanced accumulation of
proline in the leaf of drought-stressed medicinal pumpkin
plants by 56.2 and 55.6% at withheld irrigation at the
flowering stage and withheld irrigation at fruit formation,
respectively (Table 2). Studies on other crops have
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Curr. Res. J. Biol. Sci., 4(4): 462-470, 2012
Table 3: Irrigation, Zeolite and selenium three way interaction effects on studied traits of medicinal pumpkin
Treatments
----------------------------------------------------------------------RWC
WSD
Catalase activity
Oil yield
Irrigation
Zeolite (ton/ha) Selenium (g/Lit)
(%)
(%)
(u/mg.protein)
(kg/ha)
Normal irrigation
0
0
84.92b
15.08f
4.73h
544.68b
30
87.16ab
12.84fg
7.51gh
556.02ab
10
0
86.96ab
14.05fg
4.02h
546.03b
30
88.34a
11.66g
6.58gh
589.30a
Withheld irrigation
0
0
58.66g
41.34a
27.25d
442.10c
at flowering stage
30
65.76f
34.24b
41.99b
470.84c
10
0
74.25d
25.75d
10.48fg
533.17b
30
78.94c
21.06e
12.72ef
517.85b
Withheld irrigation
0
0
59.28g
40.72a
32.35c
318.70e
at fruit formation stage
30
63.75 f
36.24b
50.46a
338.65ed
10
0
70.58e
29.42c
11.20fg
373.51d
30
77.84c
22.16e
16.17e
447.46c
Means at each column for each character, followed by similar letters are not significantly different using Duncan's multiple range tests(p # 0.05).
Decreased or unchanged chlorophyll level during drought
stress has been reported in other species, depending on the
duration and severity of drought (Kpyoarissis et al.,
1995). Manivannan et al. (2007) reported that drought
stress caused a large decline in the chlorophyll a content,
the chlorophyll b content and the total chlorophyll content
in sunflower. A decrease of total chlorophyll with drought
stress implies a lowered capacity for light harvesting.
Since the production of reactive oxygen species is mainly
driven by excess energy absorption in the photosynthetic
apparatus, this might be avoided by degrading the
absorbing pigments (Herbinger et al., 2002). Also
decrease in chlorophyll under drought stress is mainly the
result of damage to chloroplasts caused by active oxygen
species (Smirnoff, 1995). Parida et al. (2007) reported
that the decrease in Chlorophyll contents in droughtstressed plants might possibly be due to changes in the
lipid protein ratio of pigment–protein complexes or
increased chlorophyllase activity (Parida et al., 2007).
The total Chlorophyll content was increased from nonapplication of zeolite as 12.81 mg/g fw to 14.84 mg/g fw,
at zeolite application treatment. Also, in this study zeolite
application under drought stress treatments caused to
increase the total Chlorophyll content by 20.2 and 26.9%,
respectively (Table 2).
or application of Se under Non-application zeolite was
produce higher proline content as compared to Nonapplication or application Se under application zeolite
(23.6 and 23.7 vs. 19.0 and 17.6 mmol g fw).
Stomatal conductance: Withheld irrigation at the
flowering and fruit formation stages caused to decrease
the stomatal conductance by 19.7 and 17.0% respectively,
as compared to normal irrigation conditions (Table 2).
Moreover, stomatal conductance at zeolite application
treatment was increase by 13.5% as compared to nonapplication of zeolite (Table 2). It is well known that
water-deficit stress alters a variety of physiological
processes such as stomatal conductance (Gardner et al.,
1984). This physiological trait are directly or indirectly
associated with crop growth and yields (Silva et al., 2007;
Zhang et al., 2001). Plants grown under drought condition
have a lower stomatal conductance in order to conserve
water. Consequently, CO2 fixation is reduced and
photosynthetic rate decreases, resulting in less assimilate
production for growth and yield of plants (Mafakheri
et al., 2010). On the other hand, means of comparison
results indicated that zeolite application at both imposed
drought stress conditions caused to significant increase
the stomatal conductance (109.16 vs. 88.95 mmol/m2.s
and 116.93 vs 87.32 mmol/m2.s). In this study, zeolite
application may be attributed to save as higher moisture
and absorb more water from soil and therefore, it was
caused to increasing water status of plant and leaf
stomatal conductance, Probability.
Leaf soluble protein content: Leaf soluble protein
contents in fresh leaves of irrigations treatments are
shown in Table 2. Results showed that drought stress
decreased protein content in leafs. A maximum amount of
soluble protein was obtained from normal irrigation and
water stress significantly decreased soluble protein
concentration in plant leafs. Imposed drought stress at
both flowering and fruit formation stages decreases
protein by 39.13 and 43.48%, respectively. A drought
induced decrease in total soluble protein has also been
reported in safflower (Carthamus mareoticus L.) by
Abdel-Nasser and Abdel-Aal (2002). Changes observed
in total protein, free amino acid and proline contents of
Total Chlorophyll: In present study water stress impose
decreased total chlorophyll content both at the flowering
and fruit formation stages. Maximum amount of total
Chlorophyll was obtained from normal irrigation (16.96
mg/g fw) and drought stress caused to reduce the total
Chlorophyll content to 12.57 and 11.94 mg/g fw,
respectively (Table 2). The results are agreement with
those of Pirzad et al. (2011) and Mafakheri et al. (2010).
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Curr. Res. J. Biol. Sci., 4(4): 462-470, 2012
spraying. Application of zeolite to stressed plants
decreased the CAT enzyme activity in the leaves. In our
study, se foliar spraying enhanced enzyme activity in
stressed plants as compared to treatments of not using Se
(Table 2). These results are similar to those reported by
other researchers (Sajedi et al., 2011; Timothy, 2001). Se
role in plants under water stress could increase
antioxidant enzymes activities by reducing oxidative
conditions and free radicals which have a determinate
effect on plant cells. Earlier studies have indicated that Se
maintains antioxidative defence systems and enhances
sugar and starch accumulation (Cakmak, 2000).
In the present study, drought stress and lack of zeolite
led to a significant increase in the CAT activity compared
to the respective controls, These results may point out that
the lack of zeolite in stress conditions provokes
antioxidant enzyme responses. The effects of three way
factors interactions on CAT enzyme activity was
significant. In drought stress condition in each two stages,
the highest CAT enzyme activity was observed from
treatments of without zeolite and with Se spraying
(Table 3). This might indicate plant sensitivity due to no
protection factor under water stress. According to these
results it can be suggested that usage of zeolite and se can
reduce the harmful effects of ROS and improves plant
resistance.
several drought-stressed plant species have been
attributed to a reduction in the rates of protein synthesis
and an increase in proteolytic activity, both of which tend
to cause an increase in the total soluble nitrogen (Shen et
al., 1990). The decrease in protein contents might be due
to increased proteolytic activity. Proteins are hyrolysed by
proteases to release amino acids for storage and/or
transport and for osmotic adjustment (like as proline)
during drought stress in plant. Osmotic adjustment,
protection of cellular macromolecules, storage form of
nitrogen, maintaining cellular pH, detoxification of the
cells and scavenging of free radicals are proposed
functions of free amino acid accumulation (Parida et al.,
2007). Zelolite application and Se foliar spraying had
positive and enhancing effects on protein. Also, zeolite
application in all irrigation treatments had desirable
effects on protein, so that, Zeolite application decreased
the adverse drought stress effects and caused to prevent
loss of protein content in drought stressed medicinal
pumpkin plants by 23.01% (Table 2). In summary, our
results showed that, in medicinal pumpkin, highest protein
content was obtained from application of zeolite in normal
irrigation (1.41 mg/g fw) and lowest of it was obtained
from Non-application of zeolite in withheld irrigation at
the fruit formation stage (0.59 mg/g fw). Also, Se foliar
spraying increased the leaf soluble protein contents
(Table 2).
Oil content: In present study, water stress only at fruit
formation stage had significant effect on seed oil content
and decreased it by 4%, However oil content not affected
by impose water stress at flowering phase (Table 2). Lack
of water during all stages of growth and development is
one of the limiting factors for seed growth and can
influence the composition of its oil. In addition drought
stress can reduces the seed-filling period and oil content
(Flagella et al., 2002). Pasban Eslam et al. (2010)
reported that safflower seed oil content was not affected
by drought stress but Ali et al. (2009) Concluded that oil
content will be affected by irrigation regimes, With the
increase in oil content and water also increases. Trials
have shown that unfavorable conditions, especially
drought, might alter the seed composition and related
qualities (Anwar et al., 2006). According to a number of
reports, it is evident that the stress tolerance of most plant
species varies at different growth stages (Nam et al.,
2001). Zahedi et al. (2009) reported that oil percentage
was decreased by drought stress, most likely because of
a reduction by photosynthesis and assimilate
remobilization. This trait had not affected by zeolite or Se
application.
Catalase enzyme activity: Environmental stresses such
as drought, lead to disturbance in plant metabolism and
cause oxidative injuries by enhancing the production of
Reactive Oxygen Species (ROS). Production of ROS
during drought stress is one of the main causes for
decreases in productivity, injury and death that
accompany this stress in plants. (Upadhyaya and Panda,
2004). Oxidative stress can prevent photosynthetic
activity, respiration process and plant growth (Sajedi et
al., 2011). Plant resistance to stress factors was associated
with their antioxidant capacity and the increased levels of
the antioxidant constituents may prevent stress damage
(Monk et al., 1989). CAT is one of the major antioxidant
enzymes associated with scavenging ROS (Shen et al.,
1990). Catalase, which is present in peroxisome,
dismutates H2O2 into water and molecular O2 whereas
peroxidase decomposes H2O2 by oxidation of substrate
such as phenolic compounds and/or antioxidants (Pan et
al., 2006). In our study, CAT activities was stimulated
under drought stress in comparison with the control and
the highest enzyme activity under drought stress were
found at fruit formation stage (Table 2). Bailly et al.
(2000) and Tohidi-Moghadam et al. (2009) reported that
the content of Superoxide Dismutase (SOD), Catalase
(CAT) and Glutathione Reductase (GR) in sunflower will
increase under drought stress condition. However, this
enzyme activity was significantly increased by Se
Oil yield: The oil yield response to drought stress of
medicinal pumpkin is given in 2. The oil yield was
affected by water levels. Plants in normal irrigation gave
a significantly higher oil yield (559.01 kg/ha) than plants
stressed during flowering (490.99 kg/ha) or during the
467
Curr. Res. J. Biol. Sci., 4(4): 462-470, 2012
deficiency, produce plant water need and increases
medicinal pumpkin drought tolerance under water
deficiency.
fruit formation stage (369.57 kh/ha). This results were
agreement with Pasban Eslam et al. (2010) and Zahedi
et al. (2009). Oil yield under water stress at fruit
formation stage showed 25% less than under drought
stress at flowering stage (Table 2). Oil yield is dependent
on seed yield and oil content and it seems that impose
drought stress at flowering and fruit formation stages
decreases seed yield (data not shown) and oil yield,
mainly through their components (Pasban Eslam et al.,
2010).
Zeolite application and Se spraying had significant
and positive effects on oil yield trait, as compared to Nonapplication of this treatments (Table 2). Although the
application of zeolite in all irrigation levels had desirable
effects on oil yield, however, this effects were significant
only at water stress treatments and maximum amounts of
oil yield were produced in plots that containing of zeolite
at stress conditions.
However, interactions between irrigation and zeolite
and Se treatments were significant. Results showed that
zeolite application with Se spraying in water stress
conditions specially in irrigation cease at fruit formation
stage significantly had highest oil yield as compared to
stress condition without mentioned treatments (Table 3).
Zahedi et al. (2009) reported that zeolite and selenium
application in drought stress conditions increased seed
yield and oil yield of canola cultivars. An increase in seed
yield and oil yield can be related to increments of water
and nutrition availability due to zeolite soil application as
a soil amendment. Finally they proposed that zeolite and
selenium application in dry land that exposed to drought
stress could helpful to yield improvement and prevention
of decreased yield.
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