THE INFLUENCE OF WATER HARDNESS, ON HEAVYMETALS TOXICITY TO FLATHEAD

GREY MULLET

Hussien M .Elsahfei,

General Authority For Fish Resources Development, Port Said, Egypt.

Abstract

The present investigation was designed to determine the effects of water hardness and heavy metals concentrations on a freshwater Mugilidae Mugil Cephalus, L (Flathead Grey Mullet). Very few data concerning the effect of water hardness on the acute toxicity of heavy metals are available on grey mullet, which has been proposed as a test organism for ecotoxicological studies. The effect of water hardness on the toxicity of heavy metals is discussed. The acute toxicity of selected heavy metals to a freshwater grey mullet was determined in very soft, soft, hard and very hard (12, 45, 170 and 300 mg CaCO3 L-1total hardness, respectively) water. Percentage mortality of grey mullet as influenced by heavy metals was studied in water of variable hardness. Water hardness had a significant effect on heavy metals toxicity. The concentrations of metals necessary to immobilize 50% of the test fish at 24, 48, 72, 96 hr were significantly different in soft and hard water. The 96 hr EC50 values for grey mullet were higher in hard water compared with soft and very soft water. Median effective concentrations (EC50) and their 95% confidence limits were determined for cadmium, cobalt, , copper, lead, mercury, nickel, and zinc during exposure for 24, 48, 72 and 96 hr in three different water hardness. Hardness has a much smaller effect upon the acute toxicity of mercury than the other heavy metals tested. The results indicate that Cu, Cd, Hg and Zn induced autotomy of the caudal region and mucus production. It is concluded that water hardness parameters should be considered in establishing appropriate water quality criteria and standards for the protection of aquatic fauna and flora, and ultimately human health.

Keywords: hardness, heavy metals, grey mullet, toxicity, mortality.

Introduction

Water quality is a vital element of the environment and is strongly determined by pollution. Water seems like a bountiful natural resource. However, if humans do not monitor pollution and take proper precautions to preserve good water quality then the environmental effects will progress to have a great influence. The contamination of one source of water does not only have an effect on that source of water. The contamination of streams affects rivers which then affects bays and then oceans. There is a chain of events that results in a vast majority of water being influenced by pollution. This chain of events also results in many ecosystems and organisms being affected by the water supply as well as the drinking water of humans. These organisms must learn to adapt to the water contaminants or less pollution must occur or the pollution refers to the presence of chemical, physical, or biological materials in a body of water that will affect its future use (Pipkin and Trent, 2001). After three decades of federal cleanup efforts, about 40 percent of the nation’s streams, rivers, lakes and coastal waters are still too dirty for people to fish or swim, in the Environmental Protection Agency reports (Cooper, 2000). Many fish, however, are found in hard water and show a liking for it. It may be necessary to investigate a wider range of water quality parameters in catchments that have industrial areas (Emily Bowling2003). The water may contain heavy metals or toxic organic compounds. Harmful effects can include death (lethal effects) or inability to reproduce.

Heavy metals are widely used in industry and are common water pollutants; thus knowledge of their toxicity to aquatic organisms is important. Numerous studies have documented the acute toxicity of trace metals to various invertebrate species under conditions of stable water quality. The toxicity of chemicals with particular reference to heavy metals can be altered in natural water by a number of environmental

factors (Rattner and Heath, 1995), which may alter the rate of metabolism of the organisms, and the state of toxicity of the chemical. Therefore, from the early days of toxicity testing, several attempts were made to study the effects of environmental factors on the toxicity of chemicals

to aquatic organisms. Acute tests comparing the toxicity of heavy metals to tubificid worms under conditions where water quality parameters were varied have also been conducted. Among all environmental factors, the effects of water hardness and temperature on the toxicity of chemicals are studied most often (Welsh et al, 2000). For a long time, it has been known that most heavy metals become less toxic in harder water. For example, toxicity of copper, to various fish species decreased with an increase in water hardness (Taylor et al., 2000).

Spear and Pierce (1979) discussed the lethal levels of copper for fish, as determined by several investigators, and find that Cu toxicity decreased with an increase in water hardness. Metals are generally more toxic at low pH where they occur in their most bioavailable, free ionic form; i.e., M2 (Pyle.2000).

. The toxicity of heavy metals will be affected by their bioavailability. Organisms uptake metals through the food they consume, through absorption onto gills and through the skin. Bioavailability in turn is affected by water hardness, pH and age of the organism (EPA ,1996).

Metals exert their toxic effects on microorganisms through one or more mechanisms. An excellent review is available that describes modes of metal toxicity and the mechanisms by which microorganisms resist such toxicity . Toxic metal cations may substitute for physiologically essential cations within an enzyme (e.g., Cd2+ may substitute for Zn2+), rendering the enzyme nonfunctional. Similarly, metal oxyanions such as arsenate, may be used in place of structurally similar, essential nonmetal oxy anions, such as phosphate. In addition, metals impose oxidative stress on microorganisms (Kachur et al. 1998). Metal toxicity is most commonly ascribed to the tight binding of metal ions to sulfhydryl (-SH) groups of enzymes essential for microbial metabolism. It follows, then, that the metal concentration of interest is that which is capable of binding to enzymes and interfering with microbial activity. It is this metal concentration that we define here as bioavailable metal. Although the concept of bioavailable metal is important, measurement of bioavailable metal is difficult because it varies depending on the environment and the type of organism exposed. Divalent cations, such as zinc, have been reported to mitigate metal toxicity. Higham et al. (1985) showed that addition of 60 µM total zinc reduced toxicity of 3 mM total cadmium to P. putida. Specifically, the lag phase was reduced, and the growth rate and cell yield were increased. Zinc had no effect on cells grown without cadmium. Similarly, magnesium reduced toxicity of nickel to bacteria and yeast to filamentous fungi, and to a filamentous alga. Calcium has been reported to reduce cadmium toxicity to an alga and to reduce zinc toxicity to a cyanobacterium and algae The protective effect of divalent cations such as zinc against metal toxicity is not limited to microorganisms. Zinc has been implicated in protection from cadmium-induced formation of tumors , sarcomas and lesion development in rats and mice . Despite the widespread demonstration of the protective effects of divalent cations such as zinc against metal toxicity, little is understood with regard to the mechanism of protection. However, cadmium uptake has been found to be very dependent on zinc concentration. In studies investigating uptake of 109Cd2+, zinc was a competitive inhibitor of cadmium uptake and exhibited a Ki of 4.6 µM. A more detailed understanding of the mode of protection by divalent cations might lead to the development of strategies to bioremediate co-contaminated sites in which a relatively nontoxic divalent cation (e.g., calcium) is added to a site to induce metal resistance and enhance organic biodegradation ( Rashmi et ai., 2003). Sandrin (2000)investigated the ability of seven divalent cations (calcium, cobalt, copper, iron, magnesium, manganese, zinc) to reduce inhibition of NAPH biodegradation caused by 10 and 37.5 mg solution-phase cadmium/L. Addition of 90 mg total zinc/L to treatments containing 37.5 mg solution-phase cadmium/L cadmium eliminated a 48-hr cadmium-induced lag phase. The remaining cations had inhibitory or no effects on NAPH biodegradation. elicit similar effects.

In this study, we evaluate the effects of water hardness on the acute toxicity of Seven heavy metals (viz. Cd, Hg, Ni, Pb, Cu, Co and Zn) to a freshwater Grey mullet(Mugil Cephalus,L) whish is the most economic and commercial fish. Grey mullet is a world widely distributed sea and freshwater, bottom feeder on the benthic fauna which accumulate the heavy metal results partly from chronic effects of metals contamination in the benthic invertebrates that are impoetant as food for young-of -the-year fish resulted in reduced growth, feeding activity and eleveted levels of metals in the whole fish, and as also increased in liver ,increased cell membrane damage(liped peroxidation), decreased digestive enzyme production (zymogen),and a sloughing of intestinal mucosal epithelial cells(Woodward et al., 1994). These fish are exposed to water pollutants, via sediment (during feeding), polluted water.

MATERIALS AND METHODS

Fish. Mature fingerlings Grey mullet( Mugil Cephalus, L) (7-10gm)were reared in 182-liter glass aquariums with plastic chips as gravel. Each tank was supplied with dechlorinated, circulated, and aerated local tap water at 27-29°C under a photoperiod of 12-14 h light. Fish were fed with commercial fish food. Five ( Mugil Cephalus, L )were introduced to each flux chamber after being rinsed in deionized water to remove extra ions from the body. All studies were carried out in environmental growth chambers, maintained at a constant temperature of 25±1.C under a photoperiod of 12L: 12D. (Mugil Cephalus,L )were acclimatized at to the test conditions for 48–96 hr.

A synthetic medium consisting of double-glass distilled water and salts in the concentration indicated in Table (1) were used for experiments involving water hardness.

Four levels of reconstituted test water (The Committee on Methods for Toxicity Tests with Aquatic Organisms, 1975) hardness were used in this experiments:

Verysoft, soft and hard water. The water was then bubbled with air for 2– 3 hr. At the initiation and termination of each bioassay pH was measured with a pH meter, water hardness, alkalinity and dissolved oxygen were by dissolved oxygen. (APHA et al., 1989). Seven heavy metals were used in the present study: cadmium (as CdCl2·2.5H2O), lead as Pb (NO3)2 , mercury (asHgCl2) nickel (as NiCl2·6H2O), cobalt (as COCl2·6H2O), copper (as CuSO4·5H2O), and zinc (as ZnSO4·7H2O). Stock solutions of reagent grade of heavy metallic salts were prepared in double-glass distilled water. The desired test concentrations were prepared by standard dilution techniques, using appropriate aliquots of the stock solution with test water one hour before the addition of grey mullet (Mugil Cephalus,L).Test concentrations were selected by logarithmic bisectioning. Static bioassay test procedure used in the present study were based upon recommendations of APHA et al. (1989).

Prior to test situations, the grey mullet were feed on a diet of fish food. During a test situation, however grey mullet (Mugil Cephalus,L) were not fed and the test water was not aerated or renewed for the duration of the experiment. grey mullet (Mugil Cephalus,L) mortality and behavior changes were recorded at 30 min intervals for the first 6 to 8 hr; thereafter, every 6 to 8 hr until the end of the experiment. Dead fish were removed after every 6–8 hr.

Control experiments without toxicant were run for 96 hr for each metal at each level of water hardness. Mortality in the control chamber of >10% indicated a problem with the test conditions, thus invalidating the test. Only tests where mortality in the control chamber was >10% were included in the calculations of the results. Two replicates of each test were conducted. EC50 values and their 95% confidence limits were calculated for 24, 48, 72 and 96 hr. Two replicate tests for each concentration and controls were run.

Results

Results of chemical analysis showed that exposure waters essentially achieved desired hardness and alkalinity for very soft, soft, and hard water as earlier suggested by The Committee on Methods for Toxicity Tests with Aquatic Organisms (1975) Table (1).

We studied the effect of hardness on the acute toxicity of seven heavy metals by measuring the mortality rate of grey mullet (Mugil Cephalus,L) on exposure for up to 96 hr in static bioassay chambers. Analysis of test solutions indicated that water hardness and alkalinity (Table 1) were within the limits suggested by The Committee on Methods

for Toxicity Tests with Aquatic Organisms (1975). Dissolved oxygen concentration were not less than 5.0 mg L-1 during the experiment. In the present study we found that in general, for lower concentrations of heavy metals percent mortality decreased with increasing water hardness (Table 2). Percent mortality resultingfrom heavy metals concentrations in 48 and 96 hr for grey mullet (Mugil Cephalus,L) at different hardness tests is shown in Table (2).

In the present study we found that as hardness of test water increases the percent mortality decreases. A summary of the 24, 48, 72, and 96 hr EC50values and their 95% confidence limits calculated from mortality data is shown in Table (3). It is clear from the table that toxicity of zinc, copper, cobalt, lead, manganese, nickel and cadmium decreased (increasing EC50 values) with increasing hardness of test water. In control tests of very soft, soft and hard water, grey mullet (Mugil Cephalus,L) remained active during the test period.

The results of the present study suggested that for grey mullet (Mugil Cephalus,L) , mercury and copper are the most toxic , manganese and cobalt the least toxic of the heavy metal ions.

The rank order toxicity of tested metals were as follows:

Very soft water: Hg > Cu > Cd > Pb > Zn > Ni > Co

Soft water: Hg > Cu > Cd > Zn > Ni > Pb > Co

Hard water: Hg > Cu > Zn > Cd > Ni > Pb > Co

The rank order toxicity Zn, Pb, Cd, varied in different water hardness tests. In the very soft and soft water, the acute toxicity of Cu, Zn, Pb, and Cd was very high due to the high solubility of these metals in the test solution. It is known that thes oluble form of heavy metals is more toxic than the insoluble form (Holcombe and Andrew, 1978).

The percent mortality of grey mullet (Mugil Cephalus,L) for the various hardness tests at 48, and 96 hrof exposure to various concentrations of heavy metals are presented in Table ( 2.) In very soft water with concentrations of 1.0, 0.64, 0.1 Cu mg L-1, the mortality at 48 hr of exposure was 100, 100, and 20%, respectively, while at 96 hr death was 100, 100, 45%, respectively. However, in hard water with the same period of exposure and metal concentrations, the percent death was 55, 20, and 0 at 48 hr and 45, 30 and 0 at 96 hr of exposure. These observations indicated that acute copper toxicity decreases with the increase in water hardness. Similar trends in acute toxicity of Cd, Co, Pb, and Ni metals were observed. The EC50 values and their 95% confidence limits from 24 to 96 hr of exposure are shown in Table (3).

The acute toxicity of all the heavy metals tested increased with exposure time.

For example, in very soft water the 24 hr EC50 values were 0.239 mg L-1 and 4.74 mg L-1 Cu and Pb, respectively, whereas at 96 hr the values were 0.1 Cu mg L-1 and 3.50 Pb mg L-1. Similar trends were also observed in other experiments tested in different hardness. There was a rapid decline of EC50values between 24 and 48 hr of exposure. However, between 72 and 96 hr there was only a small difference in EC50 values for exposure to Zn, Pb, Cd and Hg. Therefore, mortality was more

likely to occur in the first 48 hr of the experiment, after which the rate of mortality declined; suggesting that most of the fish mortality appeared in the initial 24 and 48 hr of exposure and at 72 and 96 hr, the mortality rate considerably decline. Furthermore, the EC50 values indicate that in very soft and soft waters most of the tested heavy metals were more toxic than in hard water. Similar trends were also observed for other tested heavy metals except for Hg where water hardness has no effect on acute toxicity

In control tests of very soft, soft, and hard water, grey mullet (Mugil Cephalus,L) remained active during the test period. They were clustered at the bottom of the test container and showed typical movement patterns. In the higher concentrations of heavy metals, test fingerling fish remains separated at the beginning of the experiment and showed a rapid movement. The later phase of toxicity was reduced movement of the fish and finally death appeared. At the lethal concentrations of many heavy metals after 24 hr of exposure, the hemoglobin content disappeared from gill and the body become white due to secretion of mucus in Cu, Zn, Hg and Cd solutions. In chromium and manganese test solutions the body became elongated and enlarged in size. There was no loss of hemoglobin and fish remains separated at medium to higher test concentrations of Mn and Co.

Discussion

Heavy metals are ubiquitous pollutants and occur in water from natural sources as well. Several important factors are known to make heavy metals biologically less active and therefore less toxic. Inherent water quality characteristics such as pH, temperature, hardness, and alkalinity also change the biological activity of heavy metals. The work described in this paper further elucidates the subtleties these later effects. Such information should make the task specifying acceptable limits easier for a variety of natural waters. The 96 hr EC50 values and their 95% confidence limits for a fresh water grey mullet (Mugil Cephalus,L) were reported for sven heavy metals viz., Cu, Cd, Co, Ni, Pb, Hg and Zn in very soft, soft, and hard water. Based on the results of present study it is concluded that the toxicity of several heavy metals decreased with increasing water hardness. Acute toxicity tests are important steps in establishing appropriate water-quality criteria and standards. If such tests are to be ecologically relevant, the important sensitive test animals should be used. However, except for daphnids and certain species of fish, little effort has made to do so.

Short-term toxicity of heavy metals decreases when pH and/or hardness increases (Lucan-Bouche et al., 1999). All these parameters such as pH, temperature hardness, mode of exposure, origin of the fish -may greatly vary between the present and other previous studies and therefore, comparisons ofLC50 or EC50 values are difficult indeed quite impossible. Previously published heavy metals LC50 values for grey mullet (Mugil Cephalus,L) exposed in hard water closely agree, with the 96 hr EC50 and their 95% confidence limits for grey mullet (Mugil Cephalus,L) in the present study.

Pyle.(2000) demonstrated that the increasing water hardness was reduced Ni toxicity to larval fathead minnows. These results suggest three separate mechanisms affecting Ni toxicity to larval fathead minnows:

(1) competitive cationic binding at the gill surface (effect of hardness); (2) competitive cationic binding to suspended solids (effect of TSS); and (3) Ni-carbonate formation with increasing pH (effect of pH). Water hardness is well known to attenuate metal toxicity to fish (Erickson et al., 1998). A previous study using adult fathead minnows exposed to Ni in soft (20 mg L.1 as CaCO3) and hard (360 mg L.1 as CaCO3) water gave 96-h LC50s of 4.58–5 and 42.4–44.5 mg Ni Ll, respectively (Pickering and Henderson, 1966).

Schubauer-Berigan et al.(1993) reported the 96-h LC50 to larval fathead minnows as 3.4 mg Ni L.1 (1.9-4.0 mg Ni L.1 95% CI) in very hard water (300–320 mg L.1 as CaCO3). This result is similar to the 96-h LC50 reported here for fathead minnows exposed to Ni under hard water conditions (i.e., 2.27 mg L.1, 1.99–2.59 mg L.1 95% CI), given the difference in hardness between the two studies.

Calcium maintains gill membrane structural integrity by cross binding gill surface anions . In very soft water, fish are close to their ionoregulatory threshold and gill structural integrity is jeopardized owing to the lack of available Ca2+ (McDonald and Rogano, 1986). Ambient metals displace Ca2+from the negatively charged gill surface causing structural damage and a reduction in osmotic integrity (Mueller et al., 1991). This alteration of gill tissue is apparent by the observed high toxicity of metals in soft water. On the other hand, high concentrations of Ca2+in the water (i.e., hard water) reduce metal toxicity through competitive inhibition of metal binding to gill surfaces by Ca2+ (Hollis et al., 1997).

In the present study the 96 hr EC50 values for cadmium were 0.134 mg L-1 in very soft water (hardness = 10–13 mg L-1 as CaCO3). Pickering & Henderson (1966) calculated the LC50 values for five species of warm-water fish, some of which were tested in both soft and hard water. There was surprisingly little difference in fathead minnows (Pimephales promelas) between the 24-h, 48-h, and 96-h LC50 values, which in soft water, were 1.09, 1.09, and 1.05 mg/litre, respectively. This species was very much more resistant to cadmium in hard water with 24-h, 48-h, and 96-h LC50 values of 78.1, 72.6, and 72.6 mg/litre, respectively. The hardness for the two types of water was 20 and 360 mg CaCO3 per litre. However, the pH was 7.5 for soft water and 8.2 for hard water, this being an area of the pH range where speciation of cadmium undergoes major change. Bluegill sunfish, goldfish, and guppies showed a decrease in LC50 with increase in exposure duration from 24 to 96 h , but these were tested only in soft water. The green sunfish Lepomis cyanellus showed LC50 values in soft water of 7.84, 3.68, and 2.84 mg/litre with exposure durations of 24 h, 48 h, and 96 h, respectively. This species was also tested in hard water, where, like the fathead minnow, it showed considerably less cadmium toxicity. The 24-h LC50 rose from 7.84 in soft water to 88.6 mg/litre in hard water. A maximum acceptable toxicant concentration (MATC) for the fathead minnow Pimephales promelas of between 37 and 57 µg cadmium/litre .The protective effects of various constituents of hard water on the toxicity of cadmium to the brook trout ( Salvelinus fontinalis) and concluded that calcium, added as either the sulfate or carbonate, was the most significant source of protection. This protective effect was observed in the absence of significant cadmium precipitation. Magnesium, sulfate, and sodium ions and the carbonate system provided little or no protection.

Holcombe and Andrew(1978) reported a protective effect of water hardness against the effects of cadmium on the eggs of Oryzias latipes.

In the hard water cadmium toxicity was decreased due to formation of precipitation. Copper, zinc and nickel are intermediate in their interaction with hardness. The situation is more complex in case of copper because water pH generally affects copper speciation . Calcium concentration of the water has been known to have protective effects on heavy metals at low pH. Aquatic reservoirs are most sensitive to acid pollution because they are poorly buffered and, therefore, always have low water hardness. Several lakes in northern Norway which are devoid of fish and fish food organisms due to acid and heavy metals pollution (Rattner and Heath, 1995).

The decrease Cu and Zn toxicity with increased of water hardness was found in the present study , those was agreed with previous reports for freshwater invertebrates, including T. tubifex, and several fish species (Taylor et al., 2000). The results of acute toxicity tests may also be affected by the chemical composition of test water hardness.

Of the seven heavy metals tested in the present study, four were found to vary several fold in toxicity to grey mullet fingelings (Mugil Cephalus,L) between soft and hard water conditions (Table 3).

Hardness of water is mainly due to the presence of calcium and magnesium salts, usually the bicarbonate and sulfate or both. The calcium cations are known to involved in a variety of biological functions related to ionic and osmotic regulation and maintenance of background levels of calcium in the body of aquatic organisms. Water hardness substantially affects metal toxicity, but has little effect on the toxicity of organic contaminants. This is the conclusion of several aquatic toxicologists who have reviewed the fairly extensive literature. The almost universal observation is that metal are less toxic in hard water, provided pH is kept constant (Rashmi, 2003).

Inorganic mercury is toxic to fish at low concentrations. The 96-h LC50 s mercury compounds are more toxic. Toxicity is affected by temperature, salinity, dissolved oxygen, and water hardness. A wide variety of physiological and biochemical abnormalities have been reported after exposure of fish to sublethal concentrations of mercury.

An increase in the water hardness from 23 to 120 mg CaCO3/litre also decreased the toxicity Spear and Price (1979) found that the mean survival time for the minnow Phoxinus phoxinus in mercuric chloride rose from 15 min for 10-3mol/litre to 230 min at 5 × 10-6mol/litre. The addition of enough sodium chloride to convert the whole of the mercuric chloride into a double-chloride sodium mercuric chloride, and even the addition of ten times this amount, did not affect the toxicity of the solution. The addition of a considerable excess of sodium chloride caused a marked prolongation of the survival time, the maximum effect being attained when the solution was approximately isotonic.

The present study revealed that Cu, Cd, Zn and Hg induced segmentation, degeneration, autotomy and excess mucus production in the grey mullet. This phenomenon was also earlier observed with Cu, Pb, and Cd exposure.

Fragmentation by heavy metals exposure was also observed in another freshwater annelids Lumbriculus variegates (Bailey and Liu, 1980).

Bouche et al. (2000) suggested that the autotomy could constitute a valid endpoint of sublethal toxicity for heavy metals, since this phenomenon is easily monitored because it is time and concentration dependent, as also revealed in the present study. The excessive mucus secretion was observed with Cd, Cu, Pb and Zn metals. Mucus secretion may be adaptive response of tubificid worm to the physiological resistance phase. Mucus from complexes with heavy metals, therefore, represents a detoxification mechanism .

In conclusion, increasing water hardness, reduce metal toxicity to Grey mullet( Mugil Cephalus,L) . Increasing pH reduces toxicity by progressive formation of less bioavailable metal species, like metal-carbonates. Water hardness reduces toxicity by out-competing metal for gill-surface binding sites. Total suspended solids, represented by commercially available clay, removes free metal from the water column reducing metal bioavailability to fish. However, the protection fish receive from TSS against metal toxicity might be offset by physical irritation to fish gills caused by high concentrations of TSS.

REFERANCES

APHA. (1989): Standard Methods for the Examination of Water and Wastewaters, 17th ed., American Public Health Association, Washington, 17th D.C.

Bailey, H. C. and Liu, D. H.W.( 1980): in J. C. Eaton, P. R. Parrish and A. C. Hendricks (eds), Aquatic Toxicology, ASTM STP 707, ASTM, Philadelphia, pp. 205–215,

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Emily Bowling.(2003):Water Chemistry: An Analysis of Hardness, Alkalinity, Iron, and Nitrate in Susquehanna River Water Sample University Selinsgrove, PA 1787.

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Higham D.P, Sadler P.J, Scawen M.D. (1985): Cadmium resistance in Pseudomonas putida: growth and uptake of cadmium. J Gen Microbiol 131:2539-2544.

Holcombe, G. W. and Andrew, R.W.(1978) : The Acute Toxicity of Zinc to Rainbow and Brook Trout: Comparison in Hard and Soft Water, U.S. EPA, Environmental Research Laboratory, EPA-600/3-78-094, Duluth, Mn., 17 pp.

Hollis, L., Muench, L. and Playle, R. C.( 1997): ‘Influence of Dissolved Organic Matter on Copper Binding, and Calcium on Cadmium Binding, by Gills of Rainbow Trout’, J. Fish Biol. 50, 703–

720.

Kachur A.V, Koch C.J, Biaglow J.E. (1998): Mechanism of copper-catalyzed oxidation of glutathione. Free Radic Res 28:259-26

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McDonald, D. G. and Rogano, M. S.(1986): ‘Ion Regulation by the Rainbow Trout, Salmo gairdneri,in Ion-Poor Water’, Physiol. Zool. 59, 318–331.

Mueller, M. E., Sanchez, D. A., Bergman, H. L., McDonald, D. G., Rhem, R. G. and Wood, C. M.( 1991): ‘Nature and Time Course of Acclimation to Aluminum in Juvenile Brook Trout (Salvelinus fontinalis): II Gill Histology’, Can. J. Fish. Aquat. Sci. 48, 2016–2027.

Pickering, Q. H. and Henderson, C.(1966), ‘The Acute Toxicity of Some Heavy Metals to Different Species of Warmwater Fishes’, Int. J. Air Wat. Pollut. 10, 453–463.

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Pyle G.G., Swanson S.M and Lehmkuhld D.M. (2000): The influence of water hardness, PH,and suspended solids on Nickel toxicity to larval fathead minnows (PIMEPHALES PROMELAS),water air and soil pollution,2002.

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Table (1): Composition and water quality characteristics of reconstituted fresh water

fingerlings grey mullet(Mugil Cephalus,L) toxicity test.

Water type	Salt required mg/l				Water quality –Mean and range
Water type	CaCO3.2H2O	NaHCO3	KCl	MgSO4	PH	Hardness CaCO3mg/l	Alkalinty CaCO3 mg/l
Very soft	15	12	1.0	12	6.5 (6.3-6.7)	13(11-15)	11.5 (10-23)
Soft	45	55	5.0	40	7.2 (7.1-7.5)	47(40-54)	32 (28-36)
hard	160	200	12	160	7.8 (7.6-8.2)	174 (160-184)	117.5 (110-125)

Table (2) Percentage mortality at 48 and 96 hr of exposure in fingerlings grey mullet (Mugil Cephalus,L) at different hardness.

Conc.mg /l

Percent mortality at different hardness

Very soft Soft Hard

Cadmium

4.2

0.42

0.1

Lead

640

150

Mercury

2.0

0.64

0.064

0.034

0.1

Nickel

640

360

150

Zinc

6.4

0.64

Copper

6.4

2.0

1.0

0.1

Cobalt

1800

1000

560

100

48hr 96hr

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Dr.Hussien Mohammed Elshafei

THE INFLUENCE OF WATER HARDNESS, ON HEAVYMETALS TOXICITY TO FLATHEAD GREY MULLET

Hussien M .Elsahfei,

General Authority For Fish Resources Development, Port Said, Egypt.

Abstract

Introduction

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