Current Research Journal of Biological Sciences 3(2): 124-128, 2011
ISSN: 2041-0778
© Maxwell Scientific Organization, 2011
Received: November 13, 2010
Accepted: December 30, 2010
Published: March 05, 2011
Effect of Triazophos on Protein Metabolism in the Fish, Channa punctatus (Bloch)
Abdul Naveed and C. Janaiah
Department of Zoology, Panchsheel College of Education, Nirmal - 504106,
Adilabad, A.P. India
Department of Zoology, Kakatiya University, Warangal -506 009, A.P. India
Abstract: On treatment with sublethal dose of triazophos exposure i.e., 24, 48, 72 and 96 h on protein
metabolism of fish Channa punctatus were observed. The harmful toxic effect of triazophos, is an Organo
Phosphorus (OP), is investigated by measuring key enzymes in protein metabolism, Viz. catalases, Super Oxide
Dismutase (SOD) and Xanthine Oxidase (XOD) and these antioxidant enzymes were altered. In the present
investigations the activity of catalases reduced under toxicity because that triazophos initiates the enhancement
of super oxide radicals. The inhibition of SOD activity in the fish reveals the impairment of antioxidant defense
mechanism and also reduction in molecular oxygen and normal metabolism step. The decreased activity of
XOD leads to increase in cellular damage through photolytic activity or non availability of iron under stress
condition.
Key words: Antioxidant enzymes, catalase, Channa punctatus, Triazophos, SOD, XOD
dismutation of the superoxide ion (O2G) to hydrogen
peroxide and oxygen molecule during oxidative energy
processes. The reaction diminishes the destructive
oxidative processes in cells. The levels of antioxidant
enzyme have been extensively used as an early warning
indicator of like pollution (Lin et al., 2001). Fish have
been proposed as indicators for monitoring land-based
pollution because they may concentrate indicative
pollutants in their tissue, directly from water through
respiration and also through their diet. Fish are frequently
subjected to proxidant effects of different pollutants often
present in the aquatic environment. In the present study on
attempt has been made to observe the effect of triazophos
on certain enzymes of protein metabolism in C. punctatus.
INTRODUCTION
Triazophos (O, O - diethyl, 0.1 phenyl, 1H, 1, 2, 4,
Triazol 3yl - phosphorothio) is an organophosphate,
widely used singly or in mixture as an insecticide to
control the pests on the paddy and on cotton fields.
Triazophos, as in other organophosphate compounds, is
known to produce toxic effects by inhibiting of the acetyl
cholinesterase activity. Organophosphate (OP) pesticides
are widely used because of their biodegradability
(BookHout and Monroe, 1977). Basically the most
prevalent xenobiotics arising out of agricultural and
industrial activities are pesticides and heavy metals where
the stress response mechanisms have been widely
addressed in vertebrates in general and fish in particular.
Oxygen availability is a limiting growth factor and
chronic hypoxia or hyperoxia may be an important
environmental stressor influencing fish growth
(Wilhel et al., 2005). However, biological systems protect
themselves against free radicals (and others derived from
oxygen) aggression converting these free radicals in
oxygen by oxidation phenomena or reduction through
antioxidant system. Univalently reduced O2- are reduced
to uncharged H2O2 either spontaneously or by Superoxide
dismutase (Halliwell and Guteridge, 1985). Superoxide
dismutase (SOD) and Catalase (CAT) have been detected
in a wide variety of mammalian cells. These enzymes play
an important role in protecting the cell against the
potentially toxic effects of environmental pollutants
(Kuthan et al., 1986). Superoxide dismutase catalyzes the
MATERIALS AND METHODS
The freshwater fish Channa punctatus were collected
from the lake of Nirmal, Adilabad district of Andhra
Pradesh. The fish were stored in cement tank (6x3x3 feet)
containing 60 L-dechlorinated tap water for 3 week for
acclimatization under laboratory conditions. Water was
changed every 24 h. Commercial fish food was supplied
to fish during acclimatization period. Dead fish (if any)
were removed from the cement tank as soon as possible to
avoid water fouling. Adult fish of nearly similar (weight
50±1.30 g) and (length 25.5±1.21 cm) were selected for
experiments. Stock solution of the pesticide was prepared
in acetone. Acclimatized fish were treated with 90% pure
technical grade triazophos. The IUPAC name of
Corresponding Author: Abdul Naveed, Department of Zoology, Panchsheel College of Education, Nirmal - 504106. Adilabad,
A.P. India
124
Curr. Res. J. Biol. Sci., 3(2): 124-128, 2011
equilibration and to establish blank rate if any. The
enzyme activity was expressed as micromoles H2O2
decomposed/mg protein/min.
Xanthine oxidase (XOD: CE. 1.17.3.2) activities
measured by the method of (Worthington, 2004). The
assay mixture contained 1.9 mL of phosphate buffer (pH
7.5), 1.0 mL of hypoxanthine and 0.1 mL of enzyme
source. The reaction was initiated by the addition of
enzyme source. Blank contains all the assay mixture
except 1.0 mL reagent grade water as a substitute to the
substrate. The rate is proportional to enzyme
concentration within limits of 0.01 to 0.02 units per test.
The activity was expressed as micro moles of Urate
formed/mg protein/min.
Data exhibited homogeneous variances, so
comparisons among different treatments were made by
one-way analysis of variance, followed by Manu-Whitney
U-test. All the tests were made with the software
statistical. Minimum significance level was 95% (p<0.05).
triazophos pesticide is (O, O-diethyl, 0.1 phenyl, 1H, 1,
2, 4, Triazol 3yl-phosphorothio). According to
(Bayne et al., 1977), LC50 (40% E.C) 48h value of
triazophos for fish C. panctatus is 0.019 ppm. Six tubs
were set up for each concentration and each tub contained
6 fish in 6L dichlorinated tap water. Water temperature
was kept at 22±1.0ºC during whole experimental period.
Qualities of experimental water PH = 7.2-7.3; dissolved
oxygen = 9.2 mg/L; free carbon dioxide =10.0 mg/L;
alkalinity = 69 mg/L were measured according to the
method of (APHA/ AWWA/WEF, 1998) Control groups
were kept in dechlorinated tap water without any
treatment. Fish were not fed before 24 h and during the
experiment. After 24, 48, 72 and 96 h of exposure, fish
were removed from tubs and washed with freshwater.
Fishes of both treated as will as control groups were killed
by a severe blow on the head.
Superoxide dismutase (SOD: EC. 1.15.1.1) activities
were measured by the method of (Beachamp and
Fridovich, 1971). The tissues were homogenized (20%
w/v) in potassium phosphate buffer pH (7.5) containing
1% poly Vinyl Pyrolydone, and centrifuged at 16000 rpm
for 15 min. Values are expressed as micro moles of
formazan formed / mg protein.
Catalases (CAT: EC. 1.11.1.6) activities were
measured by the method of (Beers and Sizer, 1952). The
(10% W/v) tissue homogenate was prepared in 50 moles
of phosphate buffer (pH 7.0) and centrifuged at 16000
rpm for 45 min and again the supernatant was centrifuged
at 10500 rpm for 1 h and resulting supernatant was used
as the enzyme source. The reaction mixture was incubated
in spectrophotometer for 45 min to achieve temperature
RESULTS
The activity of SOD observed in the tissue of liver,
brain and kidney tissue of Channa punctaurs during
sublethal concentration of triazophos for 24, 48, 72 and 96
h of exposure periods. The SOD activity significantly
decreased in all the tissues during the toxic exposure
periods.
The activity of Catalases (CAT) is observed in the
tissue of liver, brain and kidney tissue of Channa
punctatus during sublethal concentration of triazophos for
24, 48, 72 and 96 h of exposure periods. The Catalases
Table 1: Effect of (XOD, SOD, CAT) in Liver tissue of Channa punctatus (Bloch) intoxicated with Triazophos
Triazophos
-------------------------------------------------------------------------------------------------------------------------Parameters
Control
24 h
48 h
72 h
96 h
Superoxide dismutase 3.46±0.27
3.21±0.19*
2.97±0.38*
2.46±0.76
2.03±0.46
PC = -7.22
PC = -14.16
PC = -28.90
PC = -41.32
Catalases
0.172±0.014
0.164±0.017*
0.150±0.021*
1.46±0.26
0.139±0.015
PC = -4.65
PC = -12.79
PC = -15.11
PC = -19.18
Xanthine oxidase
0.41±0.036
0.350±0.02*
0.271±0.01
0.186±0.003
0.149±0.062
PC = -14.63
PC = -33.90
PC = -54.63
PC = -63.65
Each value is mean SD of 6(Six) observations; All values are statistically significant from control at 5% level (p<0.05); PC denotes percent change
over control; *: Not significant; Units as per Table 3
Table 2: Effect of (XOD, SOD, CAT) in Brain tissue of Channa punctatus (Bloch) intoxicated with Triazophos
Triazophos
-------------------------------------------------------------------------------------------------------------------------Parameters
Control
24 h
48 h
72 h
96 h
Superoxide dismutase 2.94±0.39
2.68±0.11*
2.43±0.26
2.07±0.21
1.93±0.41
PC = -8.84
PC = -17.34
PC = -29.59
PC = -34.35
Catalases
0.145±0.018
0.136±0.013*
0.131±0.16*
0.128±0.011*
0.124±0.15*
PC = -6.20
PC = -9.65
PC = -11.72
PC = -14.48
Xanthine oxidase
0.563±0.046
0.475±0.015
0.420±0.021
0.379±0.017
0.327±0.053
PC = -15.63
PC = -25.39
PC = -32.68
PC = -41.91
Each value is mean SD of 6(Six) observations; All values are statistically significant from control at 5% level (p<0.05); PC denotes percent change
over control; *: Not significant; Units as per Table 3
125
Curr. Res. J. Biol. Sci., 3(2): 124-128, 2011
3.5
3.0
2.5
Control
2.0
24 h
1.5
48 h
1.0
72 h
0.5
0.0
96 h
Liver
3.5
3.0
2.5
2.0
Control
1.5
24 h
48 h
1.0
72 h
0.5
96 h
0.0
5.0
4.5
4.0
3.5
3.0
2.5
Control
2.0
24 h
1.5
48 h
1.0
72 h
0.5
96 h
0.0
Micro moles of Urate formed/mg protien/min
Micro moles of Urate formed/mg protien/min
4.0
0.7
Micro moles of Urate formed/mg protien/min
Micro moles of formazan formed/mg protien/h
Micro moles of formazan formed/mg protien/h
Micro moles of formazan formed/mg protien/h
Table 3: Effect of (XOD, SOD, CAT) in Kidney tissue of Channa punctatus (Bloch) intoxicated with Triazophos
Triazophos
------------------------------------------------------------------------------------------------Parameters
Control
24 h
48 h
72 h
96 h
Superoxide dismutase
3.01±0.13
2.71±0.19*
2.63±0.36*
2.17±0.41
2.10±0.36
micro moles formazan formed/mg of protein/h
PC = -9.96
PC = -12.62
PC = -27.90
PC = -30.23
Catalases
0.136±0.015
0.130±0.019*
0.125±0.026*
0.120±0.036*
0.115±0.045
Values are expressed in micro moles formazan
PC = -4.41
PC = -8.08
PC = -11.76
PC = -15.44
formed/mg of protein/h
Xanthine oxidase
0.684±0.019
0.630±0.01*
0.572±0.071
0.519±0.016
0.492±0.061
Values are expressed in micro moles formazan
PC = -7.89
PC = -16.37
PC = -24.12
PC = -28.07
formed/mg of protein/h
Each value is mean SD of 6(Six) observations; All values are statistically significant from control at 5% level (p<0.05); PC denotes percent change
over control; *: Not significant
0.8
0.6
0.5
0.4
3.0
2.5
2.0
Control
1.5
24 h
1.0
48 h
72 h
0.5
0.0
96 h
Control
0.3
24 h
48 h
0.2
72 h
0.1
0.0
Brain
3.5
Liver
96 h
Brain
0.7
0.6
0.5
Control
0.4
24 h
0.3
48 h
0.2
72 h
0.1
0.0
96 h
Kidney
Kidney
Fig. 2: Activity of Xanthine Oxidase (XOD) in different
tissues of Channa punctatus under sublethal
concentration of triazophos during 24, 48,72 and 96 h.
Of exposure period
Fig. 1: Activity of Super Oxide Dismutase (SOD) in different
tissues of Channa punctatus under sublethal
concentration of triazophos during 24, 48,72 and 96 h.
Of exposure period
126
0.20
after prolonged exposure periods. All the results are
presented in the Table 1, 2, 3 and Fig. 1, 2, 3.
0.18
0.16
DISCUSSION
0.14
0.12
0.10
Control
0.08
24 h
0.06
48 h
0.04
72 h
0.02
96 h
Micro moles of H 2O 2 formed/mg protien/h
0.0
0.25
Micro moles of H2 O 2 formed/mg protien/h
Micro moles of H 2 O 2 formed/mg protien/h
Curr. Res. J. Biol. Sci., 3(2): 124-128, 2011
0.18
Insecticides are extensively used in agriculturally
developing countries to protect the crop from harmful
pests. SOD enzyme present in all eukaryotes contained
copper or Zinc which played an important role in the
catalyses of Superoxide radicals to form hydrogen
peroxide, further it is converted in to H2O by Catalases
(Nelson and Cox, 2005). Svadlenka et al. (1975) reported
that SOD is a link in the biological defense mechanism
through disposition of endogenous cytotoxic O2, which
are deleterious to structural proteins of plasma membrane.
The decreased activity of SOD in erythrocytes of calves
was observed by (Patra and Swarup, 2000). It is observed
that the pesticides produce oxidative stress by inhibiting
the activity of SOD. According to (Nelson and Cox, 2005;
Sathyanarayana, 2005) catalases play an important role in
protection of cell from the hydrogen peroxide toxicity.
Gaetani et al. (1994) reported that catalases consist of
four protein sub units containing haem group and it acts
as antioxidant enzyme. Mostly these catalases are found
in peroxisomes. In C. punctatus the activity of catalases
reduced under toxicity of pesticides, because pesticides
initiate the enhancement of superoxide radicals that can
inhibit the catalases activities (Yu, 1994). XOD is an
important enzyme that converts hypoxanthine to xanthine,
subsequent to uric acid. This enzyme contains FAD,
molybdenum and Iron are exclusively found in liver,
intestine and little amount in other tissues of animals
(Sathyanarayana, 2005; Hille and Nishino, 1995;
Hellesten, 1994) also reported XOD played a vital role in
conversion of toxic ammonia into nontoxic uric acid.
Xanthine oxidase produces hydrogen peroxide which is
very harmful to the animal, and then it converts into H2O
and O2. Further, the uric acid may act as an antioxidant
and free radical scavenger protects the cells from
oxidative damage. (Sheehan et al., 2001;
Guskov et al., 2002). The decrease in XOD activity leads
to increase in cellular damage and may be due to non
availability of Iron to the fish during toxic period. It is
proposed that measurement of antioxidant in fish tissues
may prove to be useful in biomonitering of exposure to
aquatic pollutants.
Liver
0.20
0.15
Control
0.10
24 h
0.05
72 h
48 h
96 h
0.0
Brain
0.16
0.14
0.12
0.10
Control
0.08
24 h
0.06
48 h
0.04
72 h
0.02
96 h
0.0
Kidney
Fig. 3: Activity of catalase in different tissues of Channa
punctatus under sublethal concentration of triazophos
during 24, 48,72 and 96 h. Of exposure period
activity significantly decreased in all the tissues during
the toxic exposure periods.
The activity of XOD is observed in the tissue of liver,
brain and kidney tissue of Channa punctatus during
sublethal concentration of triazophos for 24, 48, 72 and 96
h of exposure periods. The XOD activity significantly
decreased in all the tissues during the toxic exposure
periods.
The activity of SOD, XOD and Catalases in the
tissues of liver, brain and kidney tissues of Channa
punctatus during sublethal concentration of triazophos for
24, 48, 72 and 96 h of exposure periods. SOD, XOD and
CAT activities in all the tissues significantly decreased
CONCLUSION
It is observed that in the C. punctatus the decreased
SOD indicates the triazophos produces oxidative stress by
inhibiting activity of SOD. The decreased catalase activity
reduces peroxidative damage in the tissues in response to
the levels of antioxidant were modulated and decreased
XOD activity in C. punctatus reported that triazophos
127
Curr. Res. J. Biol. Sci., 3(2): 124-128, 2011
induces peroxidative damage in liver, kidney and brain
which led to modulation of antioxidant may prove to be
useful in biomonitoring of exposure to aquatic pollutants.
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ACKNOWLEDGEMENT
We thank Abdul Shukur; founder Panchsheel College
of Education, Nirmal for support and providing lab
facilities for the present study. Author’s also thankful to
the management of Maxwell Scientific Organization for
financing the publication.
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