Monitoring the Effect of Water Pollution on Four Bioindicators of Aquatic Resources of Sindh Pakistan
This investigation was aimed to estimate the heavy metal
pollution in marine and fresh water and their acute toxicity and its toxicological
affects on survival, physiological and biochemical parameters of the widely
consumed fresh water and marine water fishes of Sindh. Four bioindicators,
two from marine water (Liza subviridus and Johnius belengerii)
and two from fresh water species (Cyprius carpio and Pomodasy
argyrew) were collected to study the species-site interaction. Water
samples from both stations collected to analyze the essential and non
essential metals and muscles of fish were sampled for metabolic parameters
and persistent metals pollutant. Total lipids, proteins, amino acids and
glycogen were estimated by Spectrophotometery whereas Atomic Absorption
Spectrophotometery was used for metals detection. Results showed that
interaction of metal pollutants vary specie to specie. This showed that
pollutants act by changing the structural or biological function of bioindicator.
High concentrations of contaminants were found in tissues of fishes collected
from marine water as compared to fresh water fishes.
Untreated wastes of industrial, technological and agricultural origin
containing various metallic compounds often contaminate natural waters.
Heavy metals due to their bio-accumulative and non-biodegradable properties
constitute a core group of aquatic pollutants. These metals particulates
enter the aquatic medium through effluents discharged from tanneries,
textiles, electroplating, metal finishing, mining, dyeing and printing
industries, ceramic and pharmaceutical industries etc. (Azmat and Talat,
2006). They concentrate in the tissues of aquatic biota and are known
to produce cumulative deleterious effects (Cosson, 1994).
Furthermore, there is an increased public awareness regarding pesticides,
fertilizers, agricultural products and metals that might endanger our
indigenous fish populations and aquatic ecosystems. This is mainly because
humans use these natural resources as food and water supplies, are therefore
also exposed to produce polluting these resources (Evans et al.,
2000). Of particular concern is the exposure of bio-organisms to metal
pollution, as it is known that metals act as mutagenic/genotoxic compounds,
interfere with xenbiotic metabolic pathways and may also affect glycolysis,
the Krebs cycle, oxidative phosphoorylation, protein, amino acid metabolism
as well as carbohydrate and lipid metabolism (Drastichová et
al., 2005; De la Torre et al., 2000). Knowledge of acute toxicity
of a xenobiotic often can be very helpful in predicting and preventing
acute damage to aquatic life in receiving waters as well as in regulating
toxic waste discharges. A perusal of the available literature reveals
that studies on the acute effects of toxic metals on the biochemical constituents
of fishes are scanty (El-Demeerdash and Elgamy, 1999; El-Naga et al.,
Heavy metals concentration in the tissues of fish enter into human beings
through food chain and due to there cumulative action causes potential
health hazards sometimes even lethal (El-Shehawi et al., 2007).
The toxic effects may result from the bio concentration of metals and
their consequence binding with biologically active constituents of the
body such as lipids, amino acids and proteins (Smedes and Thomson, 1996;
Thangam and Sivakumar, 2004; Vutukuru, 2005).
Chuddar et al. (2002) report that heavy metal, nickel effects
biochemical component like glycogen of gill, digestive gland and whole
body of freshwater bivalve, Parreysia cylindrical under studied.
The significant decrease in total glycogen content of gill, digestive
gland and whole body was observed due to pollution stress caused by nickel.
In this context, an attempt was made to investigate bioaccumulation of
heavy metals in marine and fresh water fishes and their acute effects
on some biochemical profiles to show an important link in the aquatic
food chain. Biochemical profiles in fish and other aquatic organisms under
heavy metal stress serve as important bioindicators in the monitoring
of aquatic environment.
MATERIALS AND METHODS
Two marine water species (Liza subviridus (L.) and Johnius
belengerii (J)) from Ireland Manora (Fig. 1). Karachi
and two fresh water fishes ((Cyprius carpio (C) and Pomodasy
argyrew (P)) from Halegyy Lake (Fig. 2). Sindh were
collected and selected for monitoring the integrated effect of water born
metal pollution. Water sample from above site were also collected for
determination of heavy metal pollution in both aquatic resources. Collection
of water was made during fishing by fisher man in the depth of 7 to 9
feet in both aquatic resources. Ten sample of each fish were analysed
for essential and non-essential metal and their biochemical profile were
estimated for nutritional quality.
||Manora Island Karachi where the collection of marine
water fish were made
||Haleji lake of Sindh where collection of fresh water
fish were made
Metal Analysis was done after collection of fishes by dry ashing. Equal
weight of four fishes was put into crucibles. The crucibles were placed
in the oven for 2 h at 135°C. After that the samples were mineralized
at 400°C in the chemical oven for 24 h then 2 mL of nitric acid was
added and sampled were dehydrated at 450°C. To each sample 10 mL of
hydrochloric acid was added and then make up to 50 mL with double distilled
water. Water sample were digested in the same manner for the detection
of pollutant toxic metals.
Macronutrient elements were determined by flame photometry in laboratory
and reported in mg kg-1. The content of heavy metals were determined
by atomic absorption Spectrophotometry (AAS) along with standards (As
a reference), supplied by the Agilent Technologies and results were given
in μg kg-1. Hg was analyzed on AAS by using mercury lamp,
along with standard reference. All data obtained, subjected to statistical
analysis on Excel 2000.
Glycogen content was measured by Anthrone reagent as described by
Carroll et al. (1956). Results were expressed in mg g-1.
Total lipids contents were extracted from tissues by using phosphovanilline
method and were determined according to Smedes and Thomson (1996) using
diagonastic kit. Protein and amino acids were estimated by Sawhney and
Singh (2005) methods.
RESULTS AND DISCUSSION
The toxicity tests are necessary in water pollution evolution because
chemical and physical measurements alone are not sufficient to asses potential
effects on aquatic biota.
Analysis of metals in water samples from marine and fresh water resources
under study showed that there is a significant difference in concentration
of pollutants that were higher in marine water as compared to fresh water
(Table 1, 2). In addition it is an
important step to detect the level of toxicants and their effects in the
marine organism. Such effects might lead to integrated effects on metabolic
functions such as behavioral, growth, reproduction and survival. This
can result in changes in fish health and reproduction that may alter fish
population and community structure. The values of pollutants were compared
with Current National Recommended Water Quality Criteria of priority pollutant
which indicates that except as all metals having higher concentration
in both aquatic resources.
||Macronutrients metal analysis of aquatic resources of
||Trace metal analysis of aquatic resources of Sindh Pakistan
|| Analysis of macronutrients of four bioindicators of
Sindh Pakistan (mg kg-1)
Table 3 showed the concentration of macronutrients
in marine and fresh water fish in ranged of 61-52 mg kg-1 for
Na, 38-40 mg kg-1 for K, for Ca 22-27 mg kg-1 and
Mg 16-19 mg kg-1 which is approximately same for four bioindicator
of Sindh region. This showed that although there is significant difference
in concentration in of these ions (Table 1) in both
aquatic resource but rate of accumulation these elements were same as
the Na, K, Ca and Mg are very important minerals elements and found insoluble
salts in the sacroplasm of the muscular cells, inter cellular fluid, blood
and plasma (Azmat et al., 2006). The values of macronutrients compared
with literature showed decline in concentration which may effects fish
health because these elements are also play an important role in physiological
processes involves in structure of several organic compounds (Oilvereau
et al., 1981). An increase in concentration of K, Na or Mg contents
in sea-water (Table 1) may alter the morpho-functional
changes in fishes. These changes include the increase in the height and
the diameter of the micli of pinealcytes, the increase being followed
by apocrynic secretion in the cells which may disturb the ionic balance
of internal miles (Deane and Woo, 2005).
Table 4 showed bioaccumulation potential of heavy toxic
metals in four species under investigation and compared with international
literature. Pb concentration in muscles of two biomarkers from fresh water
(Cyprius carpio (0.6 μg kg-1) and Pomodasy argyrew
(5.8 μg kg-1) and two from marine water (Liza subviridus
(7.4 μg kg-1) and Johnius belengerii (7.6 μg
kg-1) showed significant difference. Johnius belengerii
showed averaged highest value (7.6 μg kg-1) whereas Pomodasy
argyrew showed averaged lowered value (5.8 μg kg-1).
It indicated that interaction of heavy toxic metals with biomarkers vary
specie to specie and more prominent in marine water fish. Extensive clinical
and experimental evidence support the significance of Pb-Ca interaction
which is apparent in current study that Liza subviridus has lowest
concentration of Ca (22 mg kg-1) and that of Pb 7.4 μg
kg-1 and Johnius belengerii (7.6 μg kg-1)
while Ca 24 mg kg-1. These interactions occur at the cellular
and molecular level and are the abilities of Pb to displace Ca during
specific physiological process (Table 3, 4).
It is likely that Pb blocks Ca efflux from cells by substituting Ca in
Ca++/Na adenosine triphosphate (Simons, 1986).
Hg concentration in muscle of two fresh water species was higher as compared
to marine water fish. Pomodasy argyrew showed highest averaged
value (5 μg kg-1) whereas Liza subviridus showed
lowest (3.2 μg kg-1). In addition there was a significant
difference among the average concentration of Hg in two stations. This
data indicate that different species have various capabilities to accumulate
and store water contaminates independent of their level in water. Same
phenomena were observed by De la Torre et al. (2000). Hg also interact
with the metal binding protein metalothionine (MT), a low molecular weight
cytosolic protein protect the biological system by binding metal ions.
Higher concentration of amino acids reported in these fish also support
Hg amino acid interaction, which may control Hg toxicity.
|| Heavy metals level in four bioindicators of Sindh Pakistan
As concentration in muscles of the four species showed variation in two
fresh water and two marine water species Liza subviridus showed
highest (9 μg kg-1) accumulation of As while Cyprius
carpio showed lowest value (3 μg kg-1). In addition
to this Pomodasy argyrew (5 μg kg-1) and Johnius
belengerii (5 μg kg-1) showed the same pattern of
accumulation of As regarding less the marine or fresh water sites. As
interact with thiol and effects many functional proteins and its effect
is significant on protein contents (Table 5) of reported
There are many similarities between Cd and Zn both are member of group
11B metals having similar tendency to form complexes. Cd has an inhibitory
effect on the activity of Zn-containing enzymes. Indeed Cd replace Zn
in MT and low protein values in current investigation may result in increased
Cd uptake (Gulfaraz and Ahmed, 2001; Funk et al., 1987). The lower
concentration of Zn in fishes may be related with higher concentration
of heavy metal Cd (Table 4) which may be attributed
with replacement of Zn with Cd due to chemical similarity.
Cadmium derives its toxicological properties from its chemical similarity
to Zn, an essential micronutrient for plants, animals and human. Cd as
an ion affects on respiration and binders in exchange of gases (Gulfaraz
and Ahmed, 2001). Cosson (1994) reported that Zn ions of metallothionine
(MT) molecule were replaced by those of Cd when both metals were combined
in the organism. This metal also showed affinity to protein SH group.
This may be related with interesting pattern of interaction between metal
and biochemical constitutes of these species like protein, amino acids
glycogen and total lipids content of these biomarkers (Table
5). In fresh water fish Cyprius carpio the highest level of
protein was reported (2.36 wet wt. and 3.36 dry wt.) as compared to other
fishes, which showed Cyprius carpio was more tolerant to heavy
metal stress. During stress condition, fish needed more energy to detoxify
the toxicants and overcome stress. Since fish have a very little amount
of carbohydrate, therefore next alternative source of energy is protein
to meet increased energy demand. The observed depletion of protein fraction
may have been due to their degradation and possible utilization of degraded
products for metabolic purpose. Table 5 showed that
nutritional composition of marine fish protein was lower than that of
fresh water fish under studied which may be related with water body condition.
Results obtained from biochemical analysis of these common edible fish
can give a useful indication for proper use of biochemical response as
biomarkers in monitoring water born pollution by heavy metals.
Increment in free amino acids level may be the results of break down
of protein for energy requirement and impaired incorporation of amino
acids in protein synthesis (Table 5) which may be attributed
to lesser use of amino acids and their involvement of an acid-base balance.
Glyeogen content in muscle were investigated as a biological monitoring
tool for assessment of effect of heavy metals present in water. Table
5 showed that the value of glycogen content in Liza subviridus
was (0.017%) which was lowest as compared to the other species. This clearly
indicates the change in biological functions due to the highest accumulation
of metal in the Liza subviridus (Table 4). Glycogen
is localized in sarcoplasm under the sarcolemma, in sarcoplasmic reticulum
and throughout the sarcoplasmic core. They are defined as a change in
biological response that differs from molecular to organisamal level due
to metals toxicity. Carbohydrates are the primary and immediate source
of energy. In stress condition, carbohydrate reserve depleted to meet
energy demand. Depletion of glycogen may be due to direct utilization
for energy utilization (Table 5).
Lipid content in muscles were inversely related level of As, Hg and Pb.
Pomadasy agyrew (22.86) contain highest total lipid concentration
as compared to marine fish Liza subviridus (14.04) while it was
lower in marine water fishes that indicates that marine water fish use
more lipids contents due to heavy load of metals (Michael et al.,
1986; Ozmen et al., 2006).
Investigation showed that appreciable decline in the biochemical profiles
such as total glycogen, total lipids and total protein contents of the
fish in presence of toxins, results in decrease productivity of fish population.
However, the decrease in protein content was significant in marine water
fish. This study reflects the extent of the toxic effects of toxic metals
and the metal induced cumulative deleterious effects at various functional
levels in the widely consumed freshwater fish and marine fish. The toxicity
of heavy metal caused the glucose level to decrease with increase of pollutants
concentration and decrease the glycogen content in muscle as reported
by Scott et al. (2006).
Statistical data analysis showed significant difference in macro elements
of fresh water and marine water fish and order follows (CNa-PNa) >
(JNa-LNa) > (CMg-PMg) > (CCa-PCa) > (JMg-LMg) > (JCa-LCa)
> (JK-LK). Only one pair of metal i.e., (CK-PK) has no significant
difference with marine fish. Similarly for heavy metals order of variation
is (JPb-LPb) > (CHg-PHg) > (JHg-LHg) > (JZn-LZn) > (CPb-PPb)
> (JCd-LCd) > (CCd-PCd) > (CZn-PZn) > (CAs-PAs) > (JAs-LAs).
Whereas biochemical parameters of fish belongs to two different aquatic
resources showed variation in proteins, amino acids, glycogen and lipids.
On the basis of above investigation it may be concluded that concentration
of heavy metals in fish of Sindh region is a matter of serious fact because
ultimately its accumulate in human body and can cause damages in human
body therefore heavy metals in the tissues of aquatic animals should occasionally
monitored. As the heavy metal concentration in tissues reflects past exposure
via water or food. It can demonstrate the current situation of the animals
before toxicity affects the ecological balance of population in the aquatic
environment. Therefore it is suggested that Pakistan coastal metal management
smelting facility shell required assessing the potential toxicity of metals-contaminated
effluent at its point of discharge to avoid the determining effects of
toxic metals in high quality food. Otherwise changes in fish health due
to pollution may decline in fish population.
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