Distribution of Lead, Cadmium and Chromium in Sediment and their Availability to Various Organs of a Freshwater Teleost, Labeo rohita (Hamilton)
The present investigation deals with the distributions of three heavy metals, lead (Pb), cadmium (Cd) and chromium (Cr) in sediment and their availability in tissues of Labeo rohita (Hamilton) fingerlings. This study showed that Cd was available in exchangeable and carbonate fractions of the sediment, where as Pb and Cr was mainly existed in organic and residual organic fractions. This study revealed that Pb and Cr were more in gill tissues, while Cd was mainly concentrated in viscera. Significant relationships (p<0.05) between metals in different chemical fractions (exchangeable, carbonate, Fe-Mn oxide, organic and residual) of sediment and metals in fish organs (Pb in all organs, Cd in muscle, skin and viscera and Cr in muscle and gill) were found.
During the last two decades, sediment analysis has acquired a new dimension, being used as a tool to trace non natural pollution influences in inland and coastal waters. It is well known that natural processes for the formation of aquatic bottom sediments are altered by human activities. Most hydrophobic organic contaminants, metal compounds and nutrients entering water bodies become associated with particulate matter, which after settling accumulated in bottom sediments. Contaminants in bottom sediments may be released to overlying water or accumulate in the food chain as a result of physical, chemical or biological process.
Consequently, bottom sediments are a source as well as sink of contaminant
in an aquatic environment as stated by Johnson and Nicholls (1988). For heavy
metal pollution, several studies have stressed the relevance of geochemical
partitioning on lagoonal/estuarine sediments to understand metal mobilization,
associations, availability to biota and diagenesis (Luoma, 1983; Desouza et
al., 1986; Lacerda et al., 1992). The analysis of the total metal
concentrations only provides information of metal enrichment of the sediment,
but not direct information on the biological effects of metal. This is due to
the fact that different chemical forms of a metal in the sediment will determine
its behaviors in the environment. On the contrary, sequential extraction which
provides information on the speciation of metals can give detailed information
concerning the mode of occurrence, metal mobility and bioavailability (Teisser
et al., 1979). Usually, the fractions considered are: exchangeable,
weakly absorbed, hydrous-oxide, organic linked and the lattice material components
(referred as residual). More than ten different sequential extraction procedures
have been developed to study the geochemical phase of metals in aquatic sediments,
road dust and sludge (Teisser et al., 1979; Miller and McFee, 1983).
Among the various sequential procedures presented, the most widely used is that
proposed by Teisser et al. (1979) which was originally developed for
The aim of the present research is to study the different forms of metals in the sediment using sequential extraction to fractionate the metals contained in sediments into six groups: water soluble, exchangeable, carbonate, Fe-Mn oxide, organic and residual fractions; to determine the role of resulting metal distribution in toxicity to the fish which live in contaminated sediment.
Materials and Methods
Sediment sample was collected from Kolleru lake area, Andhra Pradesh, India, where intensive fish culture is being practiced using industrial effluent and brought to the laboratory. It was dried, powdered and mixed properly. Chemical properties of sediment sample and water were analyzed before experimentation. Sediment sample was transferred to 100 L glass jar with 4" sediment base. Ninty five liter tap water (chloride free) was added to jar and kept for 15 days to attain equilibrium condition. The 1/10th of 96 h LC50 of rohu fingerlings of lead (1.0 mg L-1), cadmium (0.5 mg L-1) and chromium (1.5 mg L-1) were added to respective jars. For each metal three replications were there along with control (without addition of heavy metal). In each jar 10 numbers of Labeo rohita fingerlings average weight of 20±0.45 g and length of 12.8±0.55 cm were released and surface water was aerated continuously with air diffuser stones to achieve desired oxygen concentration. The chemical parameters of water were analyzed by the methodologies of APHA (1992) every 7 days interval and Pb, Cd and Cr concentration of water were also measured by the standard procedures of Black (1965). Experiment was continued for 90 days. To ensure reliable tissue metal concentration estimates, food was not supplied to fish from 24 h before sampling of total fish. After the termination of the experimental period, fish samples were collected from each jar and dissected to muscle, skin, gill, eye and whole viscera. Tissues were dried at 60oC overnight. Sediment samples were collected from each jar and dried, powdered and used for chemical fractionation of metals in sediments. The analysis was triplicate for each sediment samples. Characteristics of the processed sediments (<2.0 mm size) were determined.
Chemical Fractionation of Metals in Sediments
To evaluate the heavy metal geochemistry and to assess elemental bioavailability
of the sediment, the samples were subjected to sequential chemical extraction
techniques proposed by Tessier et al. (1979). The flow chart of the fractionation
of sediment is as follows:
After each successive extraction, the solution was centrifuged at 6000 rpm
for 20 min and then filtered through Whatman 42 filter paper and stored in acid
rinsed polyethylene bottles at 4°C prior to analysis. The heavy metal concentrations
were determined using Atomic Absorption Spectrophotometer (Perkin Elmer, 3011).
Analysis of Total Metal Contents in Sediment and Fish Tissues
One gram finely powered sediment and fish tissues were transferred to porcelain
basin and put to a Heraeus Thermicon P muffle furnace at a temperature of about
550°C for 4 to 5 h when all the carbon was destroyed, it was taken to 125
mL Erlenmeyer flask. Samples were digested with tri acid mixture (HNO3:HClO4:
H2SO4 = 10:4:1) at a rate of 5 mL per 0.5 g of sample
and was placed on hot plate at 100°C temperature. Digestion was continued
until the liquor was clear (AOAC, 1990). All the digested liquors were filtered
through Whatmann 42 filter paper and diluted to 25 mL with distilled water.
Measurement of Heavy Metals by Atomic Absorption Spectrophotometer
All heavy metals were measured with a Perkin-Elmer Atomic Absorption Spectrophotometer
(Model No. 3110) by specific cathode lamp using wavelength and potential detection
limit for respective heavy metals as follows:
Heavy metal concentration was calculated as follows:
Student t-test were used to determine the significance differences at (p<0.05)
in terms of metal concentrations in fish tissues between control and treatment.
Correlation of coefficients were determined at 0.05 and 0.01 probability levels
using two- tailed test to show relationship of metals between various tissues
of fish and different chemical fractions of sediment.
|| Physico-chemical characteristics of test water treated with
lead, cadmium and chromium with sediment base
|Temp = Temperature; TA = Total Alkalinity; TH = Total Hardness;
EC= Electrical Conductivity and pH. TA and TH are in mg L-1;
temp. in °C, EC-dS m-1, heavy metals (μg L-1)
|| Chemical characteristics of sediment treated with lead, cadmium
and chromium with sediment base
||Metal contents in different chemical fraction of sediment
(mg/kg dry wt.) treated with lead (1.0 ppm), cadmium (0.5 ppm) and chromium
(1.5 ppm) with sediment base for 90 days exposure. Values are mean±SE
|t = trace; a = significant at 5% level; NS = not significant
The physico-chemical characteristics of overlying water and sediment are given
in Table 1 and 2. The chemical characteristics
of overlying water and sediment in various treatments did not vary significantly
(p>0.05) from those of the control during 90 days exposure period.
||Pearson correlation matrix showing relationship of metals
between different tissues of Labeo rohita and each chemical fraction
|* and ** indicate the correlation coefficients were significant
at 0.05 and 0.01 probability levels, using two-tailed test, n = 5.-'
During experimental period, no mortality of fish was observed in both control
and treatment but fish weight was increased in all treatment along with control.
Heavy metal (Pb, Cd and Cr) concentration in water was almost traced in amount
in both control and treatment during experimental period. There was no significant
difference of total Pb concentration in the sediment between control and treatment.
Lead was mainly associated with residual, organic and Fe-Mn oxide fractions
whereas exchangeable and carbonate fractions were in very less amount (Table
3). Significant differences of Pb were observed between control and treated
groups in respect of Fe-Mn oxide, organic and residual fractions. No significant
difference of total Cd concentration in sediment between control and treatment
group was observed. Cadmium was more associated with exchangeable and carbonate
fractions and to some extent Fe-Mn oxide fraction also (Table
3). Organic fraction of cadmium was negligible in amount. Significant differences
of Cd were observed between control and treated groups in respect of exchangeable,
carbonate and Fe-Mn oxide fractions. Significant differences of total Cr concentration
in the sediment were observed between control and treatment. Chromium contents
were not detected in exchangeable and carbonate fractions. It was mainly associated
with residual fraction (86.41%) and to some extent in organic fraction (11.61%)
and Fe-Mn oxide was in negligible amount (Table 3).
Heavy metal (Pb, Cd and Cr) contents in whole body of Labeo rohita treated
with Pb, Cd and Cr were significantly (p<0.05) different from control. The
distribution of Pb was significant in all tissues except muscle and eye tissues,
whereas for Cd, skin and eye tissues and for Cr, skin, eye and viscera tissues,
the accumulation was not significant. Significantly (p<0.05) higher concentrations
of Pb and Cr were found in gill tissue and Cd was concentrated mainly in viscera
and gill tissues (Fig. 1A- C). The order
of distributions of heavy metals in various tissues of Labeo rohita as
follows: gill >viscera> skin> muscle>eye for Pb and Cr and viscera>
gill> skin> muscle> eye for Cd.
||Variations in metal content in different tissues of Labeo
rohita between control and treatment in sediment base heavy metals experiment.
(A) Pb, (B) Cd and (C) Cr. Vertical line at each point represents standard
error. *Significant at 5% level (p<0.05) Legend: WB-whole body, M-muscle,
S-skin , G- gill, V- viscera, E-eye
Significant correlations between different chemical fractions of Pb, Cd and
Cr (exchangeable, carbonate, Fe-Mn oxide, organic and residual) in sediments
and their accumulation in fish tissues were found (Table 4).
Mobility and Bioavailability of Metals in Sediment Base Heavy Metals Treatment
The water soluble, exchangeable and carbonate fractions are considered to
be moderately available in the environment (Ma and Rao, 1997). Levels in the
residual fraction should be considered as the background value for the elements
in the sediments (Tessier et al., 1979). The present study showed that
Pb was mainly associated with residual fraction as well as to some extent of
Fe-Mn oxide and organic fraction whereas exchangeable and carbonate fractions
was very less. The organic matter-bound Pb was more in the sediment indicates
the formation of stable Pb organic complexes due to the combination of Pb with
organic matter. Similarly, the strong affinity of Pb with humic acid has also
been reported (Takenaga and Aso, 1975). The decreasing status with the increase
in depth and higher amounts of Pb bound to organic matter had also been reported
The strong adsorption of Pb by minerals present in the soil can increase the amount of Pb bound to Fe-Mn oxide in the soil (MaClaren et al., 1981). The amount of exchangeable Pb was comparatively small due to low solubility and mobility of Pb in the soil (Xian, 1989). The carbonate Pb also may have lesser mobility due to lesser solubility as the pKsp value of PbCO3 is quite low (12.8) as compared to other Pb compounds. This observation is analogous to Santillian-Medrano and Jurinak (1975). Fernandes (1997) also reported that reducible phase was the most important sink for Pb among the mobile phases in sediments. Lead also formed stable complexes with Fe-Mn oxide and these compounds were more stable (Ramos et al., 1994). According to Zhou et al. (1998) Pb was existed in Fe-Mn oxides and organically bound form whereas Maiz et al. (1997) reported that residual fraction was the predominant form (62-87%) for Pb.
The study indicated that Cd was more associated with exchangeable and carbonate fraction compared to Fe-Mn oxide and residual fractions. Large amounts of Cd in exchangeable and carbonate fractions suggest that an appreciable percentage of particulate Cd would be available through solubilization with lower in pHs. Tessier et al. (1980) reported that large amounts of Cd were present in the carbonate and exchangeable fractions and significant lower values occurred in the residual and Fe-Mn oxide fractions, whereas no interaction of Cd was observed with organic matter. Cadmium was not found in the organic fraction for low adsorption constant and labile complexion with organic matter (Baron et al., 1990). Similar findings were also observed by Zhou et al. (1998) and reported that Cd was mainly existed in exchangeable and carbonate fractions whereas, Maiz et al. (1997) also reported that Cd was associated with carbonates, exchangeable and Fe-Mn oxides.
Chromium was mainly associated with residual and organic fractions whereas exchangeable and carbonate fractions were negligible in amounts. Similar findings were made by De Souza et al. (1986) in which significant Cr concentrations in the oxidizable phase of an organic rich sediment from a polluted river was reported. The predominant lithogenous origin of Cr may be inferred by the high percentage of the residual phase as well as detectable concentrations of the oxidizable phase (Fernandes, 1997). Zhou et al. (1998) noted that residual fraction of Cr was the predominant form (79-93%) in the sediment while according to Maiz et al. (1997) Cr was mainly associated with Fe-Mn oxide and organically bound forms.
Distribution of Heavy Metals (Pb, Cd and Cr) in Different Tissues of Labeo
Rohita in Sediment Base Heavy Metals Treatment
The present study revealed that Cr concentration in the whole fish and also
in gill and viscera was significantly (p<0.05) lower than that of Pb and
Cd. The accumulation of lead and chromium was more in gill tissue while cadmium
was mainly concentrated in viscera. In alkaline pH of water, availability of
metals was less due to the precipitation of hydroxide which accumulates in sediments.
Bioconcentration of Cd in fish occurred not only from the water but also from
the sediment as Cd was also available in exchangeable and carbonate fractions
of the sediment whereas Cr and Pb were in mainly existed in residual and organic
fractions. Fish were not able to accumulate Pb and Cr directly from the sediment.
Such findings suggest that residues may release from sediment into the overlying
water column, under natural conditions, viz., water solubility of the compound
and organic matter content of the sediment. Results of this study show that
waterborne metal is more available than sediment associated metal for uptake
by fish. Rohu exposed to waterborne Pb, Cd and Cr (without sediment) accumulated
significantly (p<0.01) much more metal than did rohu exposed to metal in
the presence of sediment. This difference in bioavailability may be largely
due to the sorption of heavy metal to inorganic or organic materials in the
sediment (Campbell et al., 1988; Fu and Cao, 1992). Cadmium absorbed
to materials in the sediment was probably not readily available for direct uptake
by fish (Rodgers et al., 1987). Similar findings with Cd was used by
Sherman et al. (1987), who found that Cd additions to a pond environment
did not produce the expected toxicity based on Cd2+ concentrations
in laboratory experiments, because high pond pH caused precipitation as CdCO3.
Cope et al. (1994) also showed that waterborne Cd was more available
than sediment associated Cd for uptake by bluegill. This study showed that most
of the Pb and Cr were accumulated in gill tissue as gill is the first target
for pollutants in water. Absorption across the gill (Sorensen, 1991; Zhou, 1998)
seemed to be the major pathway of Pb, Cd entering tilapia. The relatively high
contents of heavy metals found in viscera may be due to the fact that most of
the heavy metals are accumulated in the liver and kidney after ingestion (Badsa
and Gold Spink, 1982).
Relationship of Metals in Each Fraction of Sediment with Metals Contents
in Different Tissues of Fish in Sediment Base Heavy Metals Treatment
Relationships of metals in each fraction (except water-soluble fraction)
of sediment with metals contents in different organs of fish were studied using
Pearson correlation matrix. The accumulation of Pb in whole fish and each organ
were closely related (p<0.05) to the concentrations of this metal in exchangeable
and organic fraction whereas gill and viscera in residual portion and viscera
in Fe-Mn oxide fraction (Table 4). Cadmium concentrations
in whole fish and in each organ were closely related (p<0.05) to exchangeable
and carbonate and skin in Fe-Mn oxide fraction whereas skin and viscera in residual
portion. As regards Cr, significant correlations were noted in Cr contents in
whole fish, muscle and gill tissues with Cr contents in organic and residual
portion (Table 4). The effect of heavy metals on aquatic organisms
is controlled by the concentrations and chemical forms of the metals in water
and sediment. It is observed that complexion of metals by organic substances
reduces metal bioavailability and its potential toxicity. It is assumed that
bioavailability is related to solubility and six extraction fractions followed
the decreased solubility order of water soluble>exchangeable>carbonate
bound>Fe-Mn oxide bound>organic bound>residual (Ma and Rao, 1997).
Thus metals bioavailability decreased in the same order.
Authors are grateful to Indian Council of Agricultural Research, New Delhi for financial assistance to carryout the present study.
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