KHANH Trang Bay

Hoa Province

Tran Thi

Mai Phuong
email: ttmaiphuong@yahoo.com

Nguyen Ky

Nicolas Marmier
email: marmier@unice.fr

Hoa Khanh

VIET Nam Coastal

Thi Mai Tran

Phuong
email: ttmaphuong@yahoo.com

Charlotte Hurel

Nguyen Ky Phung

HEAVY METAL CONTAMINATION IN SEDIMENTS OF NHA

Keywords: Coastal pollution, sediment, mollusk, heavy metals, bioaccumulation, Nha Trang Bay Heavy metal, contamination, enrichment factor, sediment, Nha Trang bay Heavy metal, mollusk, bioaccumulation, Khanh Hoa coastal

Over the last three decades, the pollution of heavy metals in Nha Trang coast and the environs comes from many human activity sources, in which 90% of wastewater discharged directly into the rivers then coastal zone without treatment. Other major sources from industrial, agriculture, aquaculture activities, oil drilling, tourism and port activities may also cause direct contamination of heavy metal in this zone. The metals body burden in mollusks may reflect the concentrations of metals in surrounding water and sediment, and may thus be an indication of quality of the surrounding environment.

The study on the potential bioaccumulation of mollusks as bioindicator is an important effort that contributes to the findings of method in monitoring pollution in an environment of tropical regions. This thesis reports the results of our investigation of heavy metals in mollusks and sediment samples collected from four coastal sites of different environmentally background in Nha Trang bay and adjacent areas, Khanh Hoa province, Viet Nam during 2 years from 2012 to 2013. The study used suitable analyzing methods update to find the bioaccumulation capacity of trace metals in marine ecosystems, evaluation of the bioavailability of potentially toxic substances on human health risks and available for predicting the environmental fat and effects of pollutions. Results from this study demonstrated that the 5 metals As, Cr, Cd, Cu and Zn concentrations of sediments were acceptable or moderate biological effects. These studied mollusk species including Laternula anatine, Glauconome virens, Katelysia hiantina and Geloina coaxans have high potential factors in term of metal BSAF in their tissues. They have served as good bioindicator organisms, in which K.hiantina might be the best indicator of metal pollution as it is high presence, abudance and wide geographical distribution in the study area.

The obtained results of the research not only provided the environmental status in term of heavy metal pollution and the bioaccumulation in Nha Trang bay, they are also scientific bases for further studies regarding bio-monitoring for marine environmental protection and natural conservation in Nha Trang bay and southern coast of Viet Nam.

ACKNOWLEDGEMENT

Foremost, I would like to express my deepest gratitude to my supervisors, Prof.

Nicolas Marmier and Ass.Prof Nguyen Ky Phung for the continuous support of my Ph.D study and their excellent guidance, caring, and providing me with an excellent atmosphere for doing research and writing of this thesis.

My sincere thanks also go to for VIED offering me the scholarship and Ministry of Education and Formation of Viet Nam for financially supported my study.

Many thanks to Dr. Nguyen Thi Thanh Thuy, Dr. Vu Tuan Anh, Hua Thai Tuyen, Nguyen Chi Cong and other workers in the Institute of Oceanography of Nha Trang for helping me collect sediments and mollusk samples from the field. My research would not have been possible without their helps.

I thank my fellow labmates in ECOMERS Group: Dr. Charlotte Hurel, Dr. Claire Lomenech, Ines Mnif, Mehwish Taneez, Brice Campredon, Andrea Sabau, Salome Ansanay, and Yassine Bentahar for the stimulating discussions and for all the fun we have had in the last three years. Also I thank my collegues in Ho Chi Minh University of Science: Le Ngoc Tuan, Tran Bich Chau, Duong Thuy Nga, Nguyen Ngoc Mai, Nguyen Thi Thuy Luyen, Tran Ngoc Diem My. They were always supporting me and encouraging me with their best wishes. In particular, I am grateful to Prof. Ha Quang Hai for enlightening me the first glance of research.

Last but not the least, I would like to thank my family: my parents Tran The Son and Vu Thi Minh Nguyet, for giving birth to me at the first place; my husband Vu Van Thang and two my daughters Vu Mai Cam Quynh, Vu Mai Quynh Thu for supporting me spiritually throughout my life.

RÉSUME

Au cours des trois dernières décennies, la baie de Nha Trang et ses environs sont pollues par diverses activités humaines. 90% des eaux usées non traitées sont déverses directement dans les rivières sur la zone côtière. L'industrie, l'agriculture, l'aquaculture, le forage pétrolier, le tourisme ainsi que les activités portuaires peuvent également provoquer une contamination directe par des métaux lourds dans cette zone. La charge corporelle des métaux dans les mollusques reflète les concentrations de métaux dans l'eau et dans les sédiments environnants, telle est l'indication de la qualité de l'environnement.

L'étude sur le potentiel de bioaccumulation des mollusques comme bioindicateur est un effort important a appliquer rigoureusement dans les régions tropicales. Cette thèse présente les résultats de notre enquête pendant deux ans (2012-2013) dans 4 sites côtiers de la baie de Nha Trang de la province de Khanh Hoa (Viet Nam). L'étude a utilise des méthodes d'analyse appropriées pour déterminer la pollution en métaux lourds des sédiments et interpréter la capacité de bioaccumulation ces métaux lourds dans plusieurs types de mollusques pour la proposer des bioindicateur et évaluer les risques lies a la consommation de ces mollusques provient des substances toxiques. Les résultats de cette étude ont démontré que la concentration de métaux lourds comme Cr, Cd, Cu et Zn ne présent pas les effets biologiques. Un traitement des résultats sur le choix d'une espece comme bioindicateurs, y compris Laternula anatine, Glauconome virens, Katelysia hiantina et Geloina coaxans en terme de bioaccumulation BSAF de métal dans leurs tissus.

Les facteurs de bioaccumulation conduisant au choix d'un espèce particulière, la quelle K.hiantina pourrait être le meilleur indicateur de la pollution par les métaux.

Enfin, la recherche présente des résultats scientifiques sur la contamination des sédiments, à la bioaccumulation des métaux lourds, puis au choix de bioindicateur que pour la surveillance biologique et la protection de l'environnement marin en baie de Nha Trang et sur la côte sud-est du Viet Nam.

Mots clés: pollution des zones côtières, les sédiments, les mollusques, les métaux lourds, la bioaccumulation, baie de Nha Trang.

vi

AIM AND STEPS OF THE STUDY

This study is to identify heavy metals in coastal sediments and in mollusks from four principal areas inside the Nha Trang bay and adjacent areas, Khanh Hoa province, Viet Nam during 2 year: 2012-2013 and would provide important evident of bioaccumulation potential of heavy metals as well as a whole of ecosystem ecological risk impacts of selected trace elements in the health of the marine ecosystem in this area.

To achieve these objectives, the study conducted on the 3 main steps of research:

Step 1: Definition the ranges and state of contamination of these seven metals of sediments as As, Cd, Cr, Cu, Zn, Fe and Al. This step including collected and analyzed coastal surface sediments to determine the physico-chemical properties and heavy metal contents of the surface sediment; comparison with results from other geographical regions and to assess relationship between heavy metal contents and physicochemical characteristics of the sediments.

Step 2: Assessments important evident of bioaccumulation potential of heavy metals in this area: To investigate the relation of body size to total body burdens of metals in organisms as well as there parameters such as TOC, pH and particle size distribution possibly controlling the degree of contaminations are also discussed

Step 3: Investigate the effects of some heavy metals from human activities on the marine ecosystems. This integrated approach allows a better understanding of the fate of trace elements in different components of the coastal ecosystem, and an evaluation of the bioavailability of potentially toxic substances and of human health risks. quality and increased pressures on marine resources. There have, however, been significant changes in perspective, and new concerns have emerged [START_REF] Moste | State of the environment of VietNam[END_REF]. Elevated concentrations of trace metals in aquatic bodies as a result of human activities have been recorded since ancient times. However, excessive releases of toxic trace metals into the urban environment and the associated health implications only became apparent in the 1960s when anthropogenic metal contamination of the environment was denoted. From an environmental and health perspective, this profound geographical development will have a critical influence on our immediate environment and its quality for human health. On a daily basis, numerous human activities including municipal, industrial, commercial and agricultural operations release a variety of toxic and potentially toxic pollutants into the environment [START_REF] Cheung | Assessment of metal and nutrient concentrations in river water and sediment collected from the cities in the Pearl River Delta, South China[END_REF].

All these elements reach the ocean floor through coastal region, which connects pollutions to the marine ecosystem. Though coastal region are highly productive, dynamic and much diversified regions, the entry of these elements in the biotic system is much easier. From the marine biotic communities the bottom dwelling mollusks have a tremendous capacity of bioaccumulation. Since the coasts are more prone to the accumulation of all toxic elements and chemicals there is a higher chance of accumulation in the body of mollusks. In the dynamic lotic riverine ecosystem deposition and accumulation of such elements are rare because of its fluvial dynamics. But due to the continuous and alternate tidal actions, the retention times for such elements are high in the coastal region, which results in the better chance for their entry in to the biotic communities. The fast growing and highly edible green mussel in the coastal region are more susceptible to heavy metal accumulation and act as a route of toxicity to human population.

It is widely accepted that anthropogenic activity makes a significant contribution to the total aquatic burden of toxic metals by both point source and non point source contamination can occur. Non point source contamination usually arises from agricultural, industrial, and urban effluents that reach the coast by way of waterways, surface runoff, and precipitation. Both benthic and pelagic species may thus become contaminated by direct uptake and or through biomagnifications. Nevertheless, a permanent control of water quality is indispensable. To reveal the presence of pollutants and to measure their toxic effect biological indicators can be used, which are suitable for prediction of the expectable toxic influence of known or unknown substances. [START_REF] Sanjay | Evaluation of mollusc as sensitive indicator of heavy metal pollution in aquatic system: a review -IIOAB[END_REF] The pollution of heavy metals in Nha Trang coastal comes from many human activity sources. About 90 percents of wastewater from Nha trang city discharged directly into the rivers without treatment then make their way to estuaries and Nha trang bay. Other major sources from industrial, agriculture activities, oil drilling, tourism and port activities may also cause the direct contamination of heavy metals in this zone. Aquaculture fish in the sea with nearly 7,000 carges caused more serious affecting to the bay nowadays. In fact, approximately 10 tons per day of solid waster were discharged into the sea by 5.000 people living in islands. In the other hand, Nha Trang bay is a tourism city, however, they are more than 40 tourist boats transports every day. [START_REF] Phung | Environmental risk assessment of coastal region of Khanh Hoa Province -Department of Environment and Resources of Khanh Hoa province[END_REF].

In the recent year, contamination of heavy metals has been great problems to the natural environment, especially to marine ecosystems in coastal Viet Nam. Since 1996, in Viet Nam, the increase of metal concentration in the sediment had been observed in Quang Ninh, Hai Phong, Da Nang and Khanh Hoa (Phuong et al, 2012), but there have never been any published reports on the background of heavy metals in such a mollusk species, especially in Khanh Hoa coastal zone. In Nha Trang bay, the concentrations of heavy metal in surface sediments in period 1996-2011 were ranged between 4.58-43.2; 5.51-13.6; 0.32-34.1; 10.23-2,7; 0,15-0,33, 0,09-0,43 (µg/g DW) for Zn, Cu, Pb, As, Cd and Hg respectively. The fine fraction of sediment in Nha Trang bay goes from moderately to strongly contamination with respect to the analysis of 4 heavy metals (Zn, As, Cu and Pb) (Phuong et al, 2012) and receives attention from local managers.

The heavy metals can be either adsorbed into sediments or accumulated in benthic organism, sometimes to toxic levels. Studies on heavy metal pollution especially in coastal zones increased over the last few decades at global scale. Therefore, the mobility, bioavailability and subsequent toxicity of metals have been a major research area [START_REF] Ghabbour | Thermodynamics of metal cation binding by a solid soil-derived humic acid. Binding of Fe (III), Pb (II), and Cu (II)[END_REF]. Generally, the presence of contamination by metals have been considered only in important harbours such as New York [START_REF] Feng | Distribution of heavy metal and PCB contaminants in the sediments of an urban estuary: the Hudson River[END_REF], Boston (Manheim F.T and others, 1998), the Atlantic French harbours [START_REF] Fichet | Concentration and mobilisation of Cd, Cu, Pb and Zn by meiofauna populations living in harbour sediment: their role in the heavy metal flux from sediment to food web[END_REF] and, more recently, in Baltimore [START_REF] Mason | Metal accumulation in Baltimore Harbor: current and past inputs[END_REF], Montevideo [START_REF] Muniz | Assessment of contamination by heavy metals and petroleum hydrocarbons in sediments of Montevideo Harbour (Uruguay)[END_REF] and

Naples [START_REF] Adamo | Distribution and partition of heavy metals in surface and sub-surface sediments of Naples city port[END_REF]. However, very few studies deal with the effect of levels of toxic elements on the health of mollusks in tropical and subtropical regions such as

Vietnam and other Southeast Asian countries, particularly in Khanh Hoa province, where, in addition to human activities such as harbor activities within the estuary, industrial, agricultural and residential activities around the coastal can release heavy metals to the environment.

HEAVY METAL POLLUTIONS

Metals in environment

Metals are considered as important toxic pollutants and there is extensive literature concerning their accumulation in ecosystems. Some metals enter the sea from the atmosphere, by volcanoes, natural weathering of rocks, e.g. natural inputs of metals, such as Aluminum in wind-blowing dust of rocks and shales, but also by numerous anthropogenic activities, such as mining, combustion of fuels, industrial and urban sewage and agricultural practices. On a global scale there is now abundant evidence that anthropogenic activities have polluted the environment with heavy metals from the poles to the tropics and from the mountains to the depths of the oceans. Some metals are deposited by gas exchange at the sea surface, by fallout of particles (dry deposition) or are scavenged from the air column by precipitation (rain) which is called wet deposition. For example, Lead inputs in the atmosphere from industrial and vehicular exhaust are much greater than natural inputs. The natural levels of heavy metals in the environment had never been a threat to health but in the recent years increased industrial activities leading to air born emissions, auto exhausts, effluents from industries as well as solid waste dumping have become the sources of large quantities of heavy metals into the environment (Mhatre G.N, 1991).

Metals enter the environment by means of natural processes or are derived from human activities. For some metals, natural and anthropogenic inputs may be of the same order (Zn), whereas for others (Pb) inputs due to human activities dwarf natural inputs (Clark R.B, 2001). Much of those human activities are located in the fluvial watersheds and in the margins of estuaries (Salomons and others, 1984), being important areas for the concentration of contaminants, due to coastal industrial activity and human settlement.

There, trade is growing rapidly and much of it depends on shipping, most of import and export travel by sea, and the marine harbour environment is degraded. The increase of metal concentrations in the sediment is a symptom of this process as has been observed in the Pacific harbours [START_REF] Wolanski | The Environment en Asia Pacific Harbours[END_REF]. Rivers make a major contribution of metals in the marine environment. The nature of metals depends on bearing deposits in the catchments area and the discharge of human waste and discharges when the river passes through urban areas. Dredging of shipping channels produces large quantities of metal pollution. Much smaller quantities of metals are added to the sea by direct discharges of industrial and other waste and the dumping of sewage sludge. [START_REF] Clark | Marine Pollution, 4 th edit[END_REF][START_REF] Depledge | Heavy metals[END_REF].

Atmospheric and river inputs, dredging soil, direct discharges, industrial dumping and sewage sludge are some of the important contributors to metal pollution, which lead to the release of metals to the marine environment [START_REF] Valavanidis | Metal Pollution in Ecosystems. Ecotoxicology -Studies and Risk Assessment in the Marine Environment. Science advances on Environment, Toxicology & Ecotoxicology issues. www.chem-tox-ecotox[END_REF].

The oceans provide a vital sink for many heavy metals and their compounds. There is a growing concern that the natural cycling rates of many metals are being disturbed by anthropogenic activities, especially the release from industrial, domestic and urban effluents of increasing amounts of Pb, Zn, Cd, Hg and Cu. (Schindler P.W,1991).

Atmospheric metal pollution is responsible from most of the dissolved Cd, Cu, Fe, Zn, Ni and As in the oceans. [START_REF] Gesamp | The State of the Marine Environment[END_REF]. The world wide emissions of metals to the atmosphere (thousands of tons per year) by natural sources is estimated as: Ni: 26, Pb: 19, Cu: 19, As: 7.8, Zn: 4, Cd: 1.0, Se: 0.4 (tons x10 3 /yr). Whereas, from anthropogenic sources: Pb: 450, Zn: 320, Ni: 47, Cu: 56, As: 24, Cd: 7.5, Se: 1.1 (tons x10 3 /yr). [START_REF] Clark | Marine Pollution, 4 th edit[END_REF]. It is obvious from these numbers that Pb, Zn, As, Cd and Cu are the most important metal pollutants from human activities.

Metals of major interest in bioavailability studies, as listed by the U.S.

Environmental Protection Agency, are Al, As, Be, Cd, Cr, Cu, Hg, Ni, Pb, Se, and Sb (EPA, 1978). Other metals that are presently of lesser interest are Ag, Ba, Co, Mn, Mo, Na, Ti, V, and Zn. [START_REF] Mckinney | Metal bioavailability[END_REF]. These metals were selected because of their highly toxic properties, their effects on the environment and the living organisms, their potential for human exposure and increased health risk. Some highlights concerning the bioavailability of As, Cd, Cr, Cu, Mo, Ni, Pb, and Zn in sediment are discussed in this

study. An additional data are available in the references listed for some major such heavy metals are:

Arsenic (As)

Arsenic mobility, bioavailability, and toxicity are dependent on speciation: arsenite -3 ) forms are much more toxic to biological species and are more mobile than arsenate (AsO 4 -3 ) forms [START_REF] Kersten | Geochemistry of priority pollutants in anoxic sludges: Cadmium, arsenic, methyl mercury, and chlorinated organic, Chemistry and biology of solid waste[END_REF]. Arsenic is chemically similar to phosphorous.

(AsO 3
Arsenate interferes with phosphate metabolism that is widespread in the biosphere.

Metallo-organic forms of arsenic also may be much more bioavailable than inorganic forms; however, organic-bound arsenic is excreted by most species and does not appear to be highly toxic (Luoma S.N, 1983). Adsorption and desorption on iron and aluminum oxide minerals is the main factor controlling arsenic behavior in soil and sediment.

Maximal adsorption occurs at different pH for As {III} (pH 9.2) and As {V} (pH 5.5) as a function of the adsorbing mineral; As +3 mobility is enhanced under oxic conditions.

Arsenic is apparently highly mobile in anoxic sediment-water systems. Development of acidic and oxidizing conditions tends to release large amounts of arsenic into solution due to decreased sorption capacity of both forms of arsenic [START_REF] Léonard | Arsenic[END_REF]. Arsenic can be found in some film chemistry, but is not very common.

Cadmium (Cd)

The redox potential of sediment-water systems exerts controlling regulation on the chemical association of particulate cadmium, whereas pH and salinity affect the stability of its various forms [START_REF] Kersten | Geochemistry of priority pollutants in anoxic sludges: Cadmium, arsenic, methyl mercury, and chlorinated organic, Chemistry and biology of solid waste[END_REF]. Elevated chloride contents tend to enhance chloride complex formation, which decreases the adsorption of cadmium on sediment, thereby increasing cadmium mobility (Bourg A.C, 1988) and decreasing the concentration of dissolved Cd +2 and bioavailability (Luoma S.N, 1983). In anoxic environments, nearly all particulate cadmium is complexed by insoluble organic matter or bound to sulfide minerals. Greenockite (CdS) has extremely low solubility under reducing conditions thereby decreasing cadmium bioavailability. Oxidation of reduced sediment or exposure to an acidic environment results in transformation of insoluble sulfide-bound cadmium into more mobile and potentially bioavailable hydroxide, carbonate, and exchangeable forms [START_REF] Kersten | Geochemistry of priority pollutants in anoxic sludges: Cadmium, arsenic, methyl mercury, and chlorinated organic, Chemistry and biology of solid waste[END_REF]. Studies of lake and fluvial sediment indicate that most cadmium is bound to exchangeable site, carbonate fraction, and iron-manganese oxide minerals, which can be exposed to chemical changes at the sediment-water interface, and are susceptible to remobilization in water [START_REF] Schintu | Heavy metal distribution and mobilization in sediments from a drinking water reservoir near a mining area: Water[END_REF]. Cadmium is a common metal found in anthropogenically contaminated aquatic environments and is toxic to aquatic biota at elevated levels. Cadmium can be found in some pigments, especially orange, red, and yellow colors. In oxidized, near neutral water, CdCO 3 limits the solubility of Cd 2+ [START_REF] Kersten | Geochemistry of priority pollutants in anoxic sludges: Cadmium, arsenic, methyl mercury, and chlorinated organic, Chemistry and biology of solid waste[END_REF]. In a river polluted by base-metal mining, cadmium was the most mobile and potentially bioavailable metal and was primarily scavenged by non-detrital carbonate minerals, organic matter, and iron-manganese oxide minerals [START_REF] Prusty | Metal contamination due to mining and milling activities at the Zawar zinc mine, Rajasthan, India. 1[END_REF].

Mollusks accumulate large concentration of calcium ranging from 1900 -2000 ppm dry weight [START_REF]Marine Pollution[END_REF]. Highest concentration in cadmium causes several health problems in human. Cadmium and its compounds along with mercury and some other dangerous metals are, however, included in the blacklist. It is being used routinely in different industrial processes and its potential hazard to life form is predominant. Eating food or drinking water with very high cadmium levels severely irritates the stomach, leading to vomiting and diarrhea, and sometimes death. Eating lower levels of cadmium over a long period of time can lead to a build-up of cadmium in the kidneys. If the levels reach a high enough level, the cadmium in the kidney will cause kidney damage, and also causes bones to become fragile and break easily. As a conservative approach, and based on the limited human data and the studies in rats, the United States Department of Health and Human Services (DHHS, 1999) has determined that cadmium and cadmium compounds may reasonably be anticipated to be carcinogens.

Chromium (Cr)

The major source of chromium emission in to the environment from the chemical manufacturing industry, combustion of fossil fuel, cement producing plants, waste from

Copper (Cu)

Copper is most efficiently scavenged by carbonate minerals and iron-manganese oxide minerals and coatings and is less mobile than cadmium, lead, and zinc (Prusty et al,1994); in most other situations lead is less mobile than copper. Elevated chloride contents decrease adsorption of copper on sediment, due to chloride complexation, which results in greater solubility and mobility (Bourg A.C, 1988); [START_REF] Gambrell | The effects of pH, redox, and salinity on metal release from contaminated sediment: Water[END_REF]. In systems with high total copper contents, precipitation of malachite controls dissolved copper contents at low pH (Bourg A.C, 1988;[START_REF] Salomons | Environmental impact of metals derived from mining activities: Processes, predictions, prevention[END_REF]. Sometimes, elemental substitution is more complex; for example, copper toxicity is related to low abundances of zinc, iron, molybdenum, and (or) sulfate (Chaney R.L, 1988).

The natural input of copper to the marine environment from erosion of mineralized rocks. Anthropogenic inputs of copper are from the production of electrical equipments, as chemical catalysts, as antifouling agents in paints, as algicides, in alloys, and as wood preservatives. Copper dissolved in seawater is chiefly in the form of CuCO 3 or in reduced salinity as CuOH + . It also forms complexes with organic molecules. Mollusks have a tremendous capacity to accumulate copper from contaminated waters. Reports are saying the copper concentration factor for oysters growing in contaminated waters is 7500 and they may accumulate 2000 ppm of copper in their blood [START_REF]Marine Pollution[END_REF].

Lead (Pb)

The main sources of lead in the aquatic environment are leaded gasoline and mining (Prosi F, 1989). Leaded gasoline results in introduction of organometallic lead compounds, which eventually reach surface water, into the atmosphere. Mining releases inorganic lead compounds. Both organic and inorganic forms of lead pose serious health risks to all forms of life [START_REF] Ewers | Lead[END_REF]. Inorganic lead compounds (sulfide, carbonate, and sulfate minerals) are commonly abundant in sediment but have low solubilities in natural water. Naturally occurring lead in mineral deposits is not very mobile under normal environmental conditions, but becomes slightly more soluble under moderately acidic conditions. Soluble lead is little affected by redox potential [START_REF] Gambrell | The effects of pH, redox, and salinity on metal release from contaminated sediment: Water[END_REF]. Lead is tightly bound under strongly reducing conditions by sulfide mineral precipitation and complexion with insoluble organic matter, and is very effectively immobilized by precipitated iron oxide minerals under well oxidized conditions [START_REF] Gambrell | The effects of pH, redox, and salinity on metal release from contaminated sediment: Water[END_REF]. In the aquatic environment, total dissolved lead abundances in water and pore water control primary uptake by organisms. Lead bioaccumulation is primarily dependent on the amount of active lead compounds (predominantly aqueous species) in the environment and the capacity of animal species to store lead (Prosi F, 1989). Particulate lead may contribute to bioaccumulation in organisms. For humans, particles that are inhaled but not exhaled are especially important. Variations in physiological and ecological characteristics of individual species lead to different enrichment factors and tolerances for each organism. A study of bottom dwelling organisms suggests that iron rich sediment inhibits lead bioavailability (Luoma S.N, 1989). In a study of lake and fluvial sediment, most lead was bound to a carbonate fraction or to iron-manganese oxide minerals, both of which respond to chemical changes at the sediment-water interface, and are susceptible to remobilization in water [START_REF] Schintu | Heavy metal distribution and mobilization in sediments from a drinking water reservoir near a mining area: Water[END_REF]. In a polluted river environment, lead is most efficiently scavenged by non-detrital carbonate and iron-manganese oxide minerals and is less mobile than cadmium [START_REF] Prusty | Metal contamination due to mining and milling activities at the Zawar zinc mine, Rajasthan, India. 1[END_REF]. Lead that can be found in some inks and coatings pigments (although not common), electrodes, solder, battery plates (if maintenance is performed on batteries), and paper but is reportable only if 0.5 lbs or more of lead is released in the form of dust.

Lead is a highly toxic substance, exposure to which can produce a wide range of adverse health effects. Both adults and children can suffer from the effects of lead poisoning, but childhood lead poisoning is much more frequent. Even today, at minimum more than four hundred thousand children under the age of six who have too much lead in their blood. Lead (Pb) is typical example of anthropogenic metal pollution. Beginning with very low levels at about 2.700 years ago, Pb concentration increased during the industrial age and has risen rapidly since Pb was added to gasoline fuel of vehicles. Pb levels in Greenland ice have risen 200-fold from the natural level. (Harrison et al, 1991).

There are many ways in which humans are exposed to lead: through deteriorating paint, household dust, bare soil, air, drinking water, food, ceramics, home remedies, hair dyes and other cosmetics. Much of this lead is of microscopic size, invisible to the naked eye. More often than not, children with elevated blood lead levels are exposed to lead in their own home. Young children under the age of six are especially vulnerable to lead's harmful health effects, because their brains and central nervous system are still being formed. For them, even very low levels of exposure can result in reduced IQ, learning disabilities, attention deficit disorders, behavioral problems, stunted growth, impaired hearing, and kidney damage. At high levels of exposure, a child may become mentally retarded, fall into a coma, and even die from lead poisoning. In adults, lead can increase blood pressure and cause fertility problems, nerve disorders, muscle and joint pain, irritability, and memory or concentration problems. When a pregnant woman has an elevated blood lead level, that lead can easily be transferred to the fetus, as lead crosses the placenta. In fact, pregnancy itself can cause lead to be released from the bone, where lead is stored often for decades after it first enters the blood stream.

Nickel (Ni)

Nickel is a silver-white metal with siderophilic properties that facilitate the

Molybdenum (Mo)

Molybdenum is an essential element for many animals and plants as it is required in their enzyme system. Molybdenum can be present in molybdate anions, MoO 4 -2 , in soil where it can be mobile and bioavailable, because it is geochemically similar to sulfate.

Molybdate ion is often associated with iron oxyhydroxide minerals, where it competes with phosphate and organic matter. Molybdenosis in animals is associated with soil that contains large amounts of available molybdenum, especially in forage plants with low sulfur and copper contents [START_REF] Neuman | Copper and molybdenum[END_REF].

Zinc (Zn)

In slightly basic, anoxic marsh sediment environments, zinc is effectively immobilized and not bioavailable [START_REF] Gambrell | The effects of pH, redox, and salinity on metal release from contaminated sediment: Water[END_REF]. Substantial amounts of zinc are released to solution if this sediment is oxidized or exposed to an acidic environment. Very high abundances of soluble zinc are present under well oxidized conditions and at pH 5.0 to 6.5, whereas low abundances of soluble zinc are present at pH 8 under all redox conditions and at pH 5.0 to 6.5 under moderately and strongly reducing conditions [START_REF] Gambrell | The effects of pH, redox, and salinity on metal release from contaminated sediment: Water[END_REF]. In polluted river environments, most zinc is scavenged by nondetrital carbonate minerals, organic matter and oxide minerals and is less mobile than cadmium (and perhaps less mobile than lead) [START_REF] Prusty | Metal contamination due to mining and milling activities at the Zawar zinc mine, Rajasthan, India. 1[END_REF]. Elevated chloride contents decrease adsorption of zinc on sediment (Bourg A.C, 1988).

Zn, as other transition metals, is essential for the health and growth of most organisms. It is a cofactor of nearly 300 enzymes. In phytoplankton, carbonic anhydrate, zinc-based metalloenzyme, is involved in the inorganic carbon acquisition from seawater.

The activity of this enzyme has been shown to be dependent on the level of CO 2 and on the availability of Zn, thus conferring on Zn a key role in oceanic carbon cycling.

The Zn enrichment in the marine environment stimulates the synthetic capacity of the mollusk hence the calcification of them. In natural seawater, concentrations of dissolved zinc are often very low (ca. 21.7 nmol.l -1 ). However, while zinc is an important at trace concentrations, it can also be very toxic when in excess, forming dangerous free radicals. A mollusk has an opportunity to be exposed to high metal concentrations as a result of human activities or natural disaster.

Heavy metals are among the most common environmental pollutants and their occurrence in waters and biota indicate the presence of natural or anthropogenic sources.

Numerous studies have demonstrated that the concentrations of heavy metals in suspended and bed sediments can be sensitive indicators of contaminants in hydrological systems [START_REF] Diagomanolin | Heavy metals (Ni, Cr, Cu) in the Karoon waterway river, Iran[END_REF][START_REF] Idris | Assessment of heavy metals pollution in Sudanese harbours along the Red Sea Coast[END_REF]Jain C.K, 2004) Zinc is an essential element to human being. Zinc is widely seen in nature. The natural concentration of zinc in soil estimated to be 10-30 mg/kg. Zinc is used in coating of other metals, in alloys and many common goods. Besides this zinc is used for wood preservation, catalyst, ceramic, fertilizers, batteries, paints, explosives household and medical appliances. According to WHO, 1996 the dietary requirement of zinc up to 22mg/day, which is equivalent to 0.3mg/kg bw/day. Gastrointestinal absorption of zinc varied substantially from 8-80%. The absorption decreases after ingestion with calcium and phosphorus. This is due to the precipitation of zinc in the intestine. Dermal absorption of zinc also noticed. There is little information about the toxicity of zinc exposure. Chronic exposure of zinc leads to anemia.

Pollution of heavy metal in marine sediments

Because sediments can contain significantly higher concentrations of some chemicals than the overlying water, it is important to evaluate the potential for such chemicals to accumulate in aquatic organisms [START_REF] Renier | Relationships between ambient geochemistry, watershed land-use and trace metal concentrations in aquatic invertebrates living in storm water treatment ponds[END_REF]. Among aquatic pollutants, heavy metals are the most appropriate indicator of pollution, because of their stability in sediments and scarcity in natural environments (Saeki et al, 1993). Heavy metals introduced to aquatic environments by industrial, domestic and mining activities are ultimately absorbed by deposits and incorporated into sediments. Hence sediments are the most concentrated physical pool of metals in aquatic systems.

Heavy metal concentrations in sediment are many times greater than the same metals in the water column. Sediments can act as a scavenger agent for heavy metal and an adsorptive sink in aquatic environment. It is therefore considered to be an appropriate indicator of heavy metal pollution (Idris et al, 2004). Surface sediments are a feeding source for biological life, a transporting agent for pollutants and an ultimate sink for organic and inorganic matter settling. In heavily polluted sediments, the anthropogenically introduced components by far exceed the natural components and pose a risk to the marine ecosystem (Algan et al, 1999;Waldichuk et al, 1985). Metals in contaminated sediments may persist and impact upon ecosystems for decades, or remain largely dormant until desorption via a resuspension event releases the toxicants to seawater, greatly increasing their potential impact. In general, environmental pollutants such as metals pose serious risks to many aquatic organisms.

Trace metals, when entering into natural water become part of the water-sediment system and their distribution processes are controlled by a dynamic set of physicalchemical interactions and equilibrium. Sediments are basic components of the environment as it provides nutrients for living organisms and serves as sink for deleterious chemical species, reflect the history of the pollution [START_REF] Singh | Geogenic distribution and baseline concentration of heavy metals in sediments of the Ganges River, India[END_REF]. Heavy metals are mainly distributed between the aqueous phase and the suspended sediments during their transport.

Riverine suspended load and sediments have important function of buffering heavy metal concentrations particularly by adsorption or precipitation. More than 97% of the mass transport of heavy metals to the oceans is associated with river sediments (Jain C.K, 2004).

The presence of heavy metals in sediments is affected by the particle size, composition of the sediments and other organic substances. [START_REF] Ghabbour | Thermodynamics of metal cation binding by a solid soil-derived humic acid. Binding of Fe (III), Pb (II), and Cu (II)[END_REF].

Physical mixing of fluvial and marine particulates leads to a continuous decrease in the trace metal content of the suspended matter with increasing salinity. [START_REF] Cheung | Assessment of metal and nutrient concentrations in river water and sediment collected from the cities in the Pearl River Delta, South China[END_REF].

Metals are natural constituents of the sediments in coastal zones and they may be oligoelements, such as zinc for example, which is a natural element essential for living organisms when present in small amounts, or do not have any known biological role, as is the case for lead. However, both can become toxic for living organisms at high concentrations [START_REF] Ewers | Lead[END_REF][START_REF] Ohnesorge | Zinc[END_REF].

Sediments may contribute significantly to concentrations of metals in benthic invertebrates, either by absorption/adsorption from interstitial water or by direct ingestion within food chains and the potential risk of human exposure, makes it necessary to monitor the levels of these contaminants in marine organisms. This information is needed to help predict potentially adverse effects on fish, shellfish, and other aquatic prey animals, or on wildlife or humans that consume them [START_REF] Odeq | Guidance for Assessing Bioaccumulative Chemicals of Concern in Sediment[END_REF]. In the Atlantic and Mediterranean harbours these changes in the sediment reservoir have been occasionally studied and some researchs were maked as information show in Table 1.1 universal in accurately predicting the bioavailability of metals accumulated by animals living in contaminated sediments although some methods show promise (Luoma et al, 1983).

Unlike organic pollutants, natural processes of decomposition do not remove heavy metals. On the contrary, they may be enriched by organisms and can be converted to organic complexes, which may be even more toxic. The metal solubility is principally controlled by pH, concentration and type of ligands and chelating agents, oxidation-state of the mineral components and the redox environment of the system. Since each form may have different bioavailability and toxicity, the environmentalists are rightly concerned about the exact forms of metal present in the aquatic environment. The toxicity and fate of the water borne metal is dependent on its chemical form and therefore quantification of the different forms of metal is more meaningful than the estimation of its total metal concentrations. Critical assessment of the endpoints of determination for potentially and actually available and accessible metal fractions in the environmental matrices of water, soil, and sediment become the basis for a need specific monitoring strategy.

Some research for mechanisms of metal toxicity on organisms is indicated that:

The mechanisms by which metals exert their toxicity in living organisms is very diverse, especially their involvement in oxidative biochemical reactions through the formation of reactive oxygen species (ROS). (Goyer R.A, 1991). Molecular mechanisms of heavy metal cytotoxicity include damage to plasma membranes, following binding to proteins and phospholipids, inhibition of Na, K dependent ATPases, inhibition of transmebrane amino acid transport, enzyme inhibition, lipid peroxidation and oxidative DNA damage, depletion of antioxidant enzymes (such as glutathione) through the generation of ROS. (Stohs S, 1995;Sigel A et al, 1992). Metal ions can penetrate inside the cell, interrupting cellular metabolism and in some cases can enter the nucleus. Metals cations can bind to DNA through ionic and coordinated bonds in a reversible way, but cannot produce all the lesions observed in chromatin of cells. Hence, not only the direct, but mostly indirect effects of metals on nuclear chromatin must be considered more important in DNA damage. In the last decade it has been proved that metal carcinogenicity is mediated by the generation of reactive free radicals (especially hydroxyl radicals, HO•) and ROS. (Kasprzak K, 1995).

The entrance of certain metals into the nucleus can enhance the synthesis of RNA that codes for metallothioneins. Metallothieneins (MT) are peptides found mainly in the cytosol, lysosomes and in the nucleus, low molecular weight peptides, high in the amino acid cysteine which contains a thiol group (-SH). The thiol group enables MTs to bind heavy metals. Metallothioneins can be induced by essential and non-essential metals in aquatic organisms (mollusks, crustaceans). The MT induction is leading to changes in several biochemical processes that have the potential to be used as biomarkers of exposure and evaluation of pollution in the marine environment (Hamer D.H, 1986). In bivalves, metallothioneins may be trapped in whole soft tissues. Induction of metallothioneins binding cadmium in various soft tissues, gills, labial pulps, digestive gland as well as in remaining tissues, have been studied by several researchers [START_REF] Biliaff | US mussel watch data from 1986 to 1994, Temporal trend detection at large spatial scales[END_REF][START_REF] Bebianno | Cadmium metabolism in the clam Ruditapes decussata: the role of metallothioneins[END_REF].

Trace metal exposure may induce specific metal-binding ligands. Other ligands such as sulfide are important for Ag biokinetic changes in bivalves. Metals also interact strongly in their accumulation by aquatic animals. Generally, dissolved Hg uptake is reduced following exposure to other metals such as Ag, Cd, Cu, and Zn in mollusks and invertebrates. The tissue body burden and the detoxificatory fate of metals in animals seem to be more important in affecting metal accumulation than the nature of the exposure routes (aqueous vs. dietary) or of the exposure regimes. Trace metal accumulation may also be variable in different natural populations of bivalves as a result of different physicochemical environments and histories of exposure (Jain C.K, 2004). Large amounts of dissolved organic complexes and particulate matter with heavy metals are transported great distances to end up in the sediments of the estuaries. Some metals, such as Cd, can be released from their organic complexes by increasing Cl (chlorine) concentrations, which form chloride complexes. [START_REF] Elbay | Dissolved Cd behaviour in some selected French and Chinese estuaries. Consequences on Cd supply to the ocean[END_REF].

Some metals are available for uptake into organisms from solution only as free ions, whereas others are transported over biological membranes as inorganic complexes. In experiments with Cu and Cd their toxicity (and their bioavailability) is correlated with the concentration of free metal concentration (Zamuda C, 1982;[START_REF] Sunda | Effect of chemical speciation on toxicity of cadmium to grass shrimp Palaemonetes pugio: importance of free cadmium ion[END_REF].

Effect of metal on mollusks

Mollusks have the ability to efficiently bioaccumulate in the high concentrations of dissolved metals and its effects on enzymatic activity determining them growth. They are a major seafood source of protein and nutrition, therefore, becomes a potential carrier of contaminants from aquatic environment to man (Malek et al, 2004).

Many metals are essential to living organisms but some of them are highly toxic or become toxic at high concentrations. Fe (hemoglobin), Cu (respiratory pigments), Zn (enzymes), Co (Vitamin B12), Mo and Mn (enzyme). Light metals Sodium (Na), Potassium (K) and Calcium (Ca) which plays important biological roles. Transition metals Fe, Cu, Co and Mn which are essential but may be toxic at high concentrations. Metals such as arsenic (As), mercury (Hg), cadmium (Cd), copper (Cu), chromium (Cr), lead (Pb), iron (Fe), manganese (Mn), zinc (Zn) etc. which are generally not required for metabolic activity and are toxic to living organisms at quite low concentrations. [START_REF] Forster | Metal Pollution in the Aquatic Environment[END_REF]Meria E,1991).

Metals such as Hg, Pb, Sn, Ni, Se, Cr and As do not degrade in general; therefore, they accumulate throughout the trophic chain. Marine organisms are able to accumulate certain amount of toxic elements naturally through continuous exposure to pollutants present in seawater and food (Malek et al, 2004). Accumulation in living organisms leads to concentrations several orders of magnitude higher than those of the surrounding water [START_REF] Casas | Relation between metal concentration in water and metal content of marine mussels (Mytilus galloprovincialis): impact of physiology[END_REF]. Despite this the relationship between the concentration of a metal in the environment and in an organism is far from straight forward as the accumulation ratio depends on many factors; some of them have an environmental origin (temperature, pH, salinity, etc.), whereas others are related to biological factors like age, sex, sexual maturity stage, etc. (Mubiana et al, 2000). Toxic elements accumulation by marine organisms is a complex dynamic process, determined by both environmental and physiological factors i.e.

water chemistry, size, contamination of feedstuffs, feeding intensity, position in the food chain etc. (Kryshev et al, 2000;Smith et al, 2002). However, marine organisms eventually lose much of their activity in the contaminated area then heavy metal toxicity in aquatic organisms, in association with the long residence time.

In the last few decades increasing attention has been paid to the relationship between the conformation of heavy metals and their impact on aquatic organisms. coronatus, suggesting that the bioavailability of these metals is disproportionably increased with a degree of enrichment of the sediments in Fe, Cd and Ni, respectively. A slope constant approximating to unity (1.14) for Cu in A. haddoni relative to its concentration in sediment extract implies that bioavailability of this metal proportionally increased with growing concentrations of its labile forms in the associated sediment (Szefer et al, 1999).

Relationship of

Content of heavy metals in mollusks

Metals can be accumulated by biota from the water column, sediment or diet, and transferred through the food chain to eventually impact on human health (Forstner et al, 1983). Uptake pathways in mollusks may include exposure through diet (prey items living among contaminated sediments), water (direct exposure of metals via the gills) and sediments (ingestion of sediments). Mollusks are filter feeders, which feed on algae, zooplanktons and excreta of all aquatic vertebrates mainly present in the bottom sediments of the water bodies. Mollusks are the major bottom feeders in the marine ecosystem, which also have tremendous capacity to accumulate all the microelements present in their food.

Mollusks are considered as the main bioaccumulators of toxic chemicals as pesticides, heavy metals… Heavy metals are the class of highly toxic elements, causing great health problem to human life through bioaccumulation from the edible bivalves.

Sediments act as both carriers and sinks for contaminants in aquatic environments.

Sediments directly stress marine ecosystem by reducing available reproduction or light energy, impeding recruitment and smothering, which also leads to more organism's disease. Metals in contaminated sediments are persistent and have the potential to impact upon coastal ecosystems for decades. They may remain largely dormant until desorption during a coastal hazard releases the toxicants to seawater, greatly increasing their impact.

In general, metals pose a significant risk to mollusk ecosystems. Unfortunately, little information is presently available on the toxic impact of contaminant metals on organisms.

Numerous studies have demonstrated that the concentrations of the majority of heavy metal emissions from anthropogenic activities accumulates in river and ocean sediments, where they are absorbed onto clays and other fine-grained materials. However, the sediments can release heavy metals to the overlying water via natural or anthropogenic processes, causing potential adverse effects to the ecosystems. Moreover, benthic organisms can take up metals directly from the sediments, which in turn enhance the potential of some metals entering into the food chain [START_REF] Adamo | Distribution and partition of heavy metals in surface and sub-surface sediments of Naples city port[END_REF][START_REF] Chen | Distribution and accumulation of heavy metals in the sediments of Kaohsiung Harbor, Taiwan[END_REF].

The pollution levels of the marine environment by heavy metals can be estimated by analyzing sediments and marine organisms. The levels of heavy metals in mollusks and other invertebrates are often considerably higher than in other constituents of the marine environment. Compared to sediments, mollusks exhibit greater spatial sensitivity and therefore, are the most reliable tool for identifying sources of biologically available heavy metal contamination [START_REF] Szefer | Some metals in benthic invertebrates in Gdansk Bay[END_REF]. Heavy metals occur naturally and from anthropogenic sources in the ecosystems with large variations in concentrations. They can be bioaccumulated through the food chain posing a toxic risk to species higher in the food chain and to humans (Stankovic et al, 2013) In sediments, very little of the total metal content is present in free ionic or chelated form in the interstitial solution. Most of the metal content occurs in complex form with insoluble inorganic and organic ligands. Particularly, heavy metals can be absorbed on negatively charged surfaces of clay minerals, organic matter or iron and manganese hydrous oxides from which they can be removed by ion-exchange with hydrogen ions and other cation species. This labile or exchangeable fraction of heavy metals is largely bioavailable. Heavy metals are also occluded in precipitated iron and manganese hydrous oxides or present as carbonates or sulfides. In addition, metal atoms may also be present in the lattices of primary and secondary minerals. In the latter forms, metals are largely unavailable to organisms, therefore bioavailability is low [START_REF] Thornton | Metal concentrations in sediment, water and clams from the Indian River Lagoon, Florida[END_REF].

High sediment load is known to negatively impact organisms but the effect of toxic elements attached to these sediments on mollusk has not been determined. The impact of heavy metals on mollusks is largely unexplored, but potential long-term damage includes:

reduced population, increased susceptibility to disease, reduced fertility/reproduction and reduced survival of larvae/juveniles. The accumulation of trace elements in aquatic consumers is of interest to environmental scientists concerned with the fate and effect of contaminants, as well as to ecologists interested in food web dynamics and trace metal biogeochemical cycles to assess the toxic impact or distribution of contaminants (Nguyen H.L et al, 2005).

So far, the individual effects of these metal contaminants have not been defined. In recent years, researchers have focused their attention on the identification of other possible bioindicators for trace metal pollution, such as the mollusks. The available data are of particular interest concerning filter-feeding bivalves which are well known for their ability to reflect environmental levels of trace metal contaminants in marine ecosystems (Chassard B, 1989). Several biomonitoring studies for the heavy metals pollution in marine

Definition

Bioaccumulation is a general term applied when there is a net accumulation of a chemical by an organism as a result of uptake from all routes of exposure.

Bioaccumulation of contaminants from sediments to benthic organisms and their subsequent transfer through the food web provides an exposure pathway to higher-level organisms [START_REF] Odeq | Guidance for Assessing Bioaccumulative Chemicals of Concern in Sediment[END_REF] The term bioaccumulation includes bioconcentration, which is the net accumulation of a chemical directly from living environment by an organism, and biomagnifications, which refers to the process by which chemicals tend to accumulate to higher concentrations at higher levels in the food web due to dietary accumulation [START_REF] Odeq | Guidance for Assessing Bioaccumulative Chemicals of Concern in Sediment[END_REF] Biomagnification is generally defined as the process of bioaccumulation along food chains or more precisely: within food webs, following various pathways on different tropical levels. However, this process must not necessarily end up in a magnification, leading to a stepwise increase with highest concentrations in organisms being in terminal positions of food webs e.g. whales, crocodiles, humans (Bernd Beek et al, 2000) The terms bioconcentration and bioconcentration factor (BCF) as defined by OECD guidelines should be limited to laboratory test systems (OECD, 2000) where the uptake of a chemical is nearly exclusively restricted to the soluble fraction and any other uptake routes can be neglected e.g. by minimizing the particle bound fraction of suspended matter.

But even under controlled laboratory conditions a certain uptake of adsorbed fractions onto food may occur. Generally all fractions must be considered potentially bioavailable regardless of the route of uptake.

The extent of bioaccumulation of metals is dependent on the total amount, the biovailability of each metal in the environmental medium and the route of uptake, storage and excretion mechanisms. The requirements of different organisms for essential metals vary substantially but optimal concentration ranges are narrow and frequently under careful homeostatic control. Excess metal concentration in an organism must be actively excreted, compartmentalized in cells or tissues, or metabolically immobilized. Some metal escape all these actions causing toxic and other adverse effects. [START_REF] Chapman | Evaluation of bioaccumulation factors in regulating metals[END_REF]Rand et al, 1985) Metal in the aquatic environment are bioaccumulated by organisms either passively from water or by facilitated uptake. Essential metals are maintained by binding to organic molecules at a variety of biochemical sites where they function mainly as catalysts to induce or enhance enzymatic activity [START_REF] Regan | The design of metal-binding in proteins[END_REF] Essential metals at high concentrations can have sublethal toxicity effects to some organisms or lethal consequences to others. Also, metals at deficient concentrations can have again adverse health effects. Thus essential metals can have a double "toxic" threshold (Rainbow P.S, 2007) Living organisms exposed environmentally to high metal concentrations follow various mechanisms to counter potential toxicity. They reduce intake, enhance excretion and/or sequestration of the metals within tissues to render them non toxic. Metals can be sequestered through storage by metal binding proteins, such as metallothioneins, in cellular vesicles and granules. Some storage mechanisms can be related in some organisms of providing essential metals for future needs (Malins D.C, 1993) Futhermore, once heavy metals are in the sediment, it can easily up take and accumulate in living tissues of benthic organisms as well as mollusk community. And when humans eat these organisms, that metals can be built up large concentration of them in human and finally can put human health at risk, because they cannot be metabolized.

The subsequent remobilization of these metals from sediments and their transfer to food web can impact local communities, sources of food supply as well as their sources of livelihood. A need to assess the impacts of contamination of heavy metals on the health and function of the vulnerable components of coastal ecosystems in Viet Nam is crucial.

This knowledge is a key to manage coastal environments for sustainable and wise use, and to assure the public health and safety of the population consuming seafood in Viet Nam.

Bioaccumulation factor (BAF)

Bioaccumulation is quantitatively expressed by the bioaccumulation factor (BAF), the ratio of the concentration reached in the organism under steady state condition and the concentration of the surrounding medium. This factor can be related to the whole organism or tissues and organs there of on a wet, dry or lipid weight basis depending on the context.

Usage of BAF offers unique approach for investigating and understanding the behaveior and fate of these toxic substances in coastal ecosystems. Transfer factors, also referred to as bioaccumulation or concentration factors, indicate the most likely metals concentration in an organism due to its exposure. Transfer factors for a given element may differ by several orders of magnitude in freshwater compared to marine biota. Since critical pathway for contaminant to enter our body systems are generally from food chains, particular emphasis must be given to study the transfer factors and dose assessment in marine biota which later can be used to estimate health risk from consumption of contaminated seafood. Therefore, this study will establish the transfer factors and dose response of marine biota found in the region and later be used to estimate health risk from consumption of contaminated seafood.

Bioaccumulation of heavy metal in mollusks

Bioaccumulation in mollusks and most of the other aquatic animals involves the uptake of chemical from the water and sediments. Bioaccumulation is the process, which causes an increased chemical concentration in organisms compared to that in environment, due to uptake by all exposure routes including dietary absorption, transport across respiratory surfaces and dermal absorption. Bioaccumulation can thus be viewed as a combination of bioconcentration and food uptake. Biomagnification can be regarded as a special case of bioaccumulation in which the chemical concentration in the organism exceeds that in the organism's diet due to dietary absorption. The extent of bioaccumulation thus can play key role in determining water and sediment quality criteria.

The assessment of the levels of heavy metals pollution in aquatic mollusks which are used as bioaccumulation indicators has become an important task in preventing risks to public health. It is to be pointed out that for a living species to be used as bioaccumulator some essential characteristics are necessary [START_REF] Cheung | Assessment of metal and nutrient concentrations in river water and sediment collected from the cities in the Pearl River Delta, South China[END_REF][START_REF] Bebianno | Cadmium metabolism in the clam Ruditapes decussata: the role of metallothioneins[END_REF].

-It must be typical of the ecosystem studied (eg non-migratory), ubiquitous and abundant, -Its size, biotype and behavior must be such as to make sapling easy.

-It must bioaconcentrate xenobiotics substances to a level sufficient to perform a direct analysis without pre-concentrations.

-It must be able to stand high concentrations of different toxic substances so as to survive the pollutant studied.

-It lives in a sessile style, thus definitely representing the local pollution.

-Its life long enough for the comparisons between various ages.

-It occupies the important position in food chain.

-Dose effect relationship can be observed in it

Assessing bioaccumulation is also a component of international efforts to identify and control chemicals of environmental concern. It is now generally accepted that substances, which are persistent, bioaccumulative, and toxic and are subject to long range transport are of particular concern. There is a need to establish reliable procedures for estimating bioaccumulation potential from knowledge of molecular structure or from readily measurable properties of the substance. There is a further inventive to adopt a tiered assessment system in order that those substances which are not bioaccumulative can be rejected from the assessment process at an early stage with minimal expense and effort.

Even toxic effects on ecosystems start with these chemical reactions in individuals. In past few years monitoring programs, conducted to evaluate water quality, usually include chemical and common biological parameters, the use of biochemical markers for surveys is less frequent, but recently more efforts have been given to propose these biomarkers of exposure and effect, in toxicity testing aiming an application in pollution monitoring [START_REF] Christensen | Human exposure to toxic metals. Factors influencing interpretation of biomonitoring results[END_REF] Both essential and non-essential trace elements are known to be highly accumulated by invertebrates, in particular by a variety of mollusks species. Aquatic mollusks seem to reflect ambient metal contamination and are therefore widely used as bioindicator organisms. Undoubtedly, aquatic mollusks are amongst the most thoroughly investigated bioindicator organisms. The translocation of sentinel species, mainly mussels from a reference site to the study areas has been demonstrated as a useful strategy for the assessment of water quality in coastal and estuarine environments, either through bioaccumulation or biomarkers analysis [START_REF] Gundacker | Comparison of heavy metal bioaccumulation in freshwater mollusks of urban river habitats in Vienna[END_REF][START_REF] Lee | Metal contamination in urban, suburban and country park soils of Hong Kong. A study based on GIS and multivariate statistics[END_REF][START_REF] Nakhle | Brachidontes variabilis and Patella sp. as quantitative biological indicators for cadmium, lead and mercury in the Lebanese coastal waters[END_REF].

In an earlier chemical and biological study of a marine bay in Washington State [START_REF] Epa | Chemical and Biological Survey of Liberty Bay[END_REF], Pacific oysters Crassostrea gigas and Ostrea edulis from the upper bay contained the highest concentrations of cadmium (1.1 mg/kg DW), copper (7.6 mg/kg DW), and zinc (150 mg/kg DW), while exhibiting the lowest levels of arsenic (0.20 mg/kg DW) and mercury (0.008 mg/kg DW).

Last some years, the concentrations of some heavy metals in the tissues of mussels, oysters and cockles have been observed in some samples collected along the coastal water of Khanh Hoa province. Concerning seafood safety, the average metal contents (Zn, Cu, Cr) in the samples from Van Previous studies have indicated that lead, zinc, cadmium, and copper were major heavy metal contaminants in the Nha Phu region (Vinh L.T, 2003). For this time, they are not research with combined analysis for the coastal sediment and mollusks species to recognize relationship between them and their bioaccumulation in this coastal region. The first aspect of this study was therefore to utilize mollusks to assess the range and distribution of the heavy metal contaminants at four strategic marine sites noted for different potential environmental contamination in the Nha Trang bay. The second aspect was to investigate whether the extent of contaminations in mollusks in each location were associated with differences observed in population densities and banding pattern attributes which were previously linked to nutritional factors in the Khanh Hoa region.

Factors affecting bioaccumulation of heavy metal in mollusks

Knowledge on the factors affecting bioaccumulation of heavy metal in mollusks may help us to understand the processes involved in the uptake of metals by the organisms. The greater metabolic rate of small organisms may partially account for the higher concentration of the essential elements as Cu and Zn [START_REF] Williamson | Variables affecting body burdens of lead, zinc and cadmium in a roadside population of snail Cepaca hortinsis[END_REF]. Uptake of metals and subsequent bioavailability are not only highly dependent on geochemical but also biological factors.

Geochemical factors

Geochemical factors that influence bioaccumulation are organic carbon, water hardness, temperature, pH, dissolved oxygen, sediment grain size, and hydrologic features of the system [START_REF] Elder | Freshwater molluscs as indicators of bioavailability and toxicity of metals in surface water systems[END_REF].

Uptake and accumulation in deposit feeders would be expected to correlate to metal concentrations in sediments, whereas accumulation in filter feeders would most likely reflect metal concentrations in water [START_REF] Newman | Influence of lead in components of a fresh water ecosystem on molluscan tissue lead concentrations[END_REF]and high environmental phosphate concentration facilitates the uptake of cadmium by organisms [START_REF] Romeo | Heavy metals in various molluscs from the Mauritanian coast[END_REF]. Several of these factors are described above and others are discussed in the following paragraphs.

pH

Hydrogen ion activity (pH) is probably the most important factor governing metal speciation, solubility from mineral surfaces, transport, and eventual bioavailability of metals in aqueous solutions. pH affects both solubility of metal hydroxide minerals and adsorption-desorption processes. Most metal hydroxide minerals have very low solubilities under pH conditions in natural water. Because hydroxide ion activity is directly related to pH, the solubility of metal hydroxide minerals increases with decreasing pH, and more dissolved metals become potentially available for incorporation in biological processes as pH decreases. Ionic metal species also are commonly the most toxic form to aquatic organisms [START_REF] Salomons | Environmental impact of metals derived from mining activities: Processes, predictions, prevention[END_REF].

Adsorption, which occurs when dissolved metals are attached to surfaces of particulate matter (notably iron, manganese, and aluminum oxide minerals, clay, and organic matter), is also strongly dependent on pH and, of course, the availability of particulate surfaces and total dissolved metal content [START_REF] Bourg | Metal in aquatic and terrestrial systems: Sorption, speciation, and mobilization[END_REF]Elder, 1989). Metals tend to be adsorbed at different pH values, and sorption capacity of oxide surfaces generally varies from near 0 percent to near 100 percent over a range of about 2 pH units.

The adsorption edge, the pH range over which the rapid change in sorption capacity occurs, varies among metals, which results in precipitation of different metals over a large range of pH units. Consequently, mixing metal-rich, acidic water with higher pH, metalpoor water may result in dispersion and separation of metals as different metals are adsorbed onto various media over a range of pH values. Cadmium and zinc tend to have adsorption edges at higher pH than iron and copper, and consequently they are likely to be more mobile and more widely dispersed. Adsorption edges also vary with concentration of the complexing agent; thus, increasing concentrations of complexing agent increases pH of the adsorption edge [START_REF] Bourg | Metal in aquatic and terrestrial systems: Sorption, speciation, and mobilization[END_REF]. Major cations such as Mg +2 and Ca +2 also compete for adsorption sites with metals and can reduce the amount of metal adsorption [START_REF] Salomons | Environmental impact of metals derived from mining activities: Processes, predictions, prevention[END_REF]. Davies C. and others, 1982 found adsorption of Cd to seawater sediment increased with increasing pH levels between 6.5 and 9.0.

Metals

The efficiency of bioaccumulation via sediment ingestion is dependent on geochemical characteristics of the sediment. Luoma (1989) describes variation of cadmium uptake from sediment by clams as a function of sediment iron content; cadmium uptake from iron oxide-rich sediment was not detected, but a significant proportion of total cadmium uptake was a consequence of iron poor sediment ingestion by clams.

Interaction among metals can result both in stimulation and antagonism, as described by Luoma (1989). Stimulation occurs when uptake of one metal induces synthesis of binding sites that affect accumulation of both metals. Stimulation of cadmium uptake by lead exposure and of zinc by cadmium exposure has been observed in rats.

Antagonism is commonly observed in simultaneous exposure to several metals.

Antagonism to zinc uptake occurs during exposure to cobalt in clams and to copper in phytoplankton and microalgae. Antagonism between zinc and cadmium occurs in phytoplankton and macroalgae. Antagonism between copper and lead and between cadmium and mercury has also been reported in some types of organisms. In addition, more widespread elements, such as calcium and magnesium, can inhibit some types of metal-organic interactions. Zinc uptake by fish is influenced by this phenomenon but uptake of copper and cadmium are not affected.

Bioavailability studies indicate that aquatic organisms uptake free metal ions (metal hydroxides) from solution quite efficiently; similarly, terrestrial animals uptake metal from solutions more efficiently than via direct particulate matter ingestion (Luoma, 1983).

The data suggest that, in low and moderate heavy metal medium gradients, Fe ions, (CBA). Heavy metal tissue enrichment was calculated with respect to Fe, which is the most vitally important element for metabolic processes; the coefficient of enrichment (CE) was calculated with modifications for the aquatic medium.

which

Particulate size

Particulate size and resulting total surface area available for adsorption are both important factors in adsorption processes and can affect metal bioavailability (Luoma, 1989). Small particles with large surface area to mass ratios allow more adsorption than an equivalent mass of large particles with small surface area to mass ratios.

Reduced adsorption can increase metal bioavailability by increasing concentrations of dissolved metals in associated water. The size of particles released during mining depends on mining and beneficiation methods. Finely milled ore may release much smaller particles that can both be more widely dispersed by water and wind, and which can also serve as sites of enhanced adsorption. Consequently, mine tailings released into fine grained sediment such as silty clays found in many playas can have much lower environmental impact than those released into sand or coarse grained sediment with lower surface area and adsorption. Consequently, geologic and (or) environmental conditions that enhance dissolved metal abundances (for example, lower pH) result in greater metal bioavailability. Indirect controls, such as larger particle or sediment size, also can result in greater bioavailability of metals by reducing adsorption and increasing dissolved metal contents. Metal assimilation from ingested particulate matter is also important, however, because metals are highly concentrated in this form (Luoma, 1989).

Moreover, several sediment types possess an adsorption affinity for zinc (Zn) ions, thereby decreasing Cd availability to benthic invertebrates [START_REF] Jackim | Effects of environmental factors on radiocadmium uptake by four species of marine bivalves[END_REF].

Organic carbon

In recent organic carbon-rich sediments, trapped interstitial fluids can commonly form a strongly reducing (anoxic) environment. Low redox potential in this environment can promote sulfate reduction and sulfide mineral deposition. During diagenesis, much of the non silicate bound fraction of potentially toxic metals such as arsenic, cadmium, copper, mercury, lead, and zinc, can be coprecipitated with pyrite, form insoluble sulfides, and become unavailable to biota (Morse, 1994). Seasonal variation in flow rates or storms that induce an influx of oxygenated sea water can result in rapid reaction of this anoxic sediment and thereby release significant proportions of these metals. Pyritization and depyritization of trace metals probably can be an important process in controlling bioavailability of many trace metals, especially in the marine environment (Morse, 1994).

Temperature

Accumulation by marine bivalves can increase with a rise in temperature [START_REF] Phillips | Biomonitoring of Trace Aquatic Contaminants[END_REF].As noted above, temperature can strongly influence the rate of biological processes; rates double for every 10 o C temperature increment. The increase in biological process rates does not necessarily result in increased bioaccumulation of metals, because both influx and efflux rates of metals may increase.

In Green land, blue mussels Mytilus edulis have been employed as indicators in areas influenced by mining activities. It is known that metal levels in mussels not only reflect the heavy metal concentration in the surrounding water, but element accumulation is also influenced by sampling season, location within the intertidal zone, and mussel size [START_REF] Riget | Influence of length on element concentrations in blue mussels (Mytilus edulis)[END_REF]. Cadmium accumulation increases with temperature because of its effect on the metabolic activity of the host [START_REF] Ray | Bioaccumulation of cadmium in marine organisms[END_REF].

Salinity

Accumulation by marine bivalves can decrease with increasing salinity [START_REF] Gibb | The application of biomonitors for the assessment of mine-derived pollution on the West Coast of the Isle of man[END_REF][START_REF] Eisler | Trace Metal Concentrations in Marine Organisms[END_REF]. Moreover, some studies have shown that increasing salinity will tend to decrease Cd uptake (Davies C. et al., 1982;[START_REF] Luoma | Temporally intensive study of trace metals in sediments and bivalves from a large river-estuarine system: Suisun Bay/delta in San Fransisco Bay[END_REF]. Salinity influences availability of contaminants whereas temperature changes can alter bivalve spawning, which will affect the soft tissue mass and lipid composition of the mollusks [START_REF] Biliaff | US mussel watch data from 1986 to 1994, Temporal trend detection at large spatial scales[END_REF].

Biological factors

Among biological factors, there are major differences in bioaccumulation between mollusks species or metal concentration may differ according to the species. Body size and weight also play an important role in bioaccumulation of metals. Within a single species accumulation can be a function of age, size, sex, genotype, phenotype, feeding activity and reproductive state (Boening D.W, 1997).

Sex plays a significant role in metals accumulation in molluscs. Significantly lower concentrations of cadmium and zinc concentrations were found in males as compared to females in Perna perna species and high concentration of cadmium was reported in Pecten maximus [START_REF] Romeo | Heavy metals in various molluscs from the Mauritanian coast[END_REF]Boyden C.R, 1974).

Body size, condition index and tidal height also affects the concentrations of As, Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb and Zn in Mytilus edulis and findings suggested that the body weight was inversely related to metal concentrations and for Cd, Mn, Pb and Zn the regression was affected by tidal height. Except for As, Fe and Mn metal concentrations were inversely related to physiological status though no differences between essential and non-essential metals were recorded. Thus it is recommended that stringent measures during sampling for biomonitoring or metal concentrations at each location must be normalized to a common body size, condition index and tidal height [START_REF] Mubiana | The influence of body size, condition index and tidal exposure on the variability in metal bioaccumulation in Mytilus edulis[END_REF].

However, the magnitude to this accumulation can vary widely between species. [START_REF] Talbot | Naturally occurring heavy metal binding proteins in invertebrates[END_REF] measured Cd levels in oysters and mussels from Philip Bay and Corio Bay near Melbourne, Australia. Both shellfishes were highly contaminated, with oysters containing more Cd than mussels (35.5-174.3mg/kg DW and 2.8-17.0mg/kg DW, respectively). Mussels collected from piers contained less Cd than mussels living on sediment, implicating sediments as the 'sink' for the metal, and therefore the matrix contributing most to uptake. Oysters, abalone and mussels are capable of accumulating extremely high levels of Cd in edible portions and therefore represent a greater hazard to human consumers than other marine organisms [START_REF] Epa | Metal Bioaccumulation in Fishes and Aquatic Invertebrates[END_REF].

Influence of length and weight

Aside from the species, the body size of oysters is one of the most important factors that affect the metal accumulation rates in biological tissues [START_REF] Meyer | Mercury in a northeastern Brazilian mangrove area, a case study: potential of the mangrove oyster Crassostrea rhizophorae as bioindicator for mercury[END_REF]; however, the soft tissue weight was higher in samples from the dry season, when the condition index is greater. Condition Index is normal if CI =90; high if CI = 120 and low if CI = 70 -80.

During the present study, consistent with as light increase in length and weight of mollusks from dry to rainy periods, metal concentrations also showed an increase from dry to rainy periods suggesting that metals accumulated in mollusks are not totally excreted to baseline concentrations (Ju et al., 2011). This is further supported by significant correlations between all the metals and both length and weight of mollusks. Borchardt and others, 1988 suggested that an increase in bivalves' biomass in the unpolluted estuarine areas of the North Sea (Atlantic Ocean) from monsoon to non-monsoon periods lead to an increase in metal concentrations and, when expressed graphically, metal concentrations in bivalves always follow a sinusoidal curve with respect to biomass.

Metal concentrations in mollusks during the dry and rainy seasons were plotted against their respective lengths and irregular patterns were observed, indicating the bioavailability of metals and an efficient metabolism. In certain cases, a short shell length or long shell length does not correlate with a low metal concentration or high metal concentration, thus disrupting a sinusoidal curve pattern and suggesting that levels of metals present in mollusks are strongly dependent on the anthropogenic input into the estuary.

Based on the results of Spearman's correlation coefficients between the biological parameters of the specimens and the mean levels of the elements in the prismatic layer, significant positive relationships were found for Mg with weight and length and also for Al with width and height, but significant negative correlations were observed for Fe and Mn with height. In terms of nacreous layer, significant positive correlations were detected for Al with width and height, while the negative relationships were found for Mn with weight and length and for Ba and Li with hinge length.

Organ of organism

Hard clams Mercenaria mercenaria were exposed to 0.1µg Cd/ml seawater for either a period of 31 days, or 3 days, each followed by depuration in clean seawater for 2 to 64 days. Cadmium accumulated at increasing rates in the kidney, but at decreasing rates in all other organs during the exposure period. The overall rate of accumulation was significantly greater for the kidney than for the gill, mantle, digestive gland, and remaining viscera. No loss of Cd or other detectable metals occurred even after 64 days depuration.

The degree to which Cd accumulated in the kidney, gill, and digestive gland did not correlate with the concentrations of other metals present in these organs [START_REF] Robinson | Metal interactions within the kidney, gill and digestive gland of the hard clam, Mercenari mercenaria, following laboratory exposure to cadmium[END_REF].

Mechanisms of bioaccumulation

These mechanisms may also vary with physiological and environmental factors (Bryan, 1973) or even with the sexual state of the animal (Alexander et al., 1976). So the observed variation in metal levels in mollusks at different stations may be related to one or both of the following mechanisms:

The availability of different metals to the animal varies with different stations, and the animal involves different uptake and retention mechanisms for the same metal at different stations. Since these mollusks inhabit such different environs, one cannot expect the same level of exposure to a metal of potential concern. The littleneck clam P. staminea and butter clam S. gigantus differ in their rate of bioaccumulation, based on their metabolic rate, living environment, and feeding habits (Wayne D, 1994). Butter clams have a higher metabolic rate, and a more accelerated growth rate than the littleneck. The feeding behavior of the butter clam relies on deeper solids. It is often found near the lower littoral zone deeply buried in sand and in bays or off rocky coasts in water depths up to 50 meters [START_REF] Ricketts | Between Pacific Tides[END_REF]. For instance, littlenecks might be more sensitive to compounds in surface deposits and those suspended in fluids. In contrast, butter clams and other

Protobranchia would be more sensitive to the bulk chemistry of the subsurface sediment matrix. In contrast, littleneck clams are more sessile. The littleneck is a poor digger and never lives in shifting sand where rapid digging is essential. It inhabits bays and estuaries in coarse, sandy mud. This species is also widely harvested for food, and is most often found in the upper littoral zone [START_REF] Audubon | Pacific Coast Nature Guide[END_REF]. Therefore, the order of metal accumulation in the animal was Fe>Zn>Mn>Pb>Cu>Ni>Cr>Cd, where, Fe was the highest followed by Zn, Mn and Pb then Cd, while Cu, Ni and Cr were changeable in their order at different stations. This order may be due to variation in the levels of discharged metals at different stations and also the chemical changes of metal before being taken up by the tested limpet.

Ayling, 1974 suggested that different mechanisms exist for the up take of Cu, Cr, Zn and Cd within the oyster Crassostre agigas. Cadmium uptake is believed to be a passive mechanism [START_REF] Spacie | Bioaccumulation and bioavailability in multiphase systems[END_REF][START_REF] Daskalakis | Variability of metal concentrations in oyster tissue and implications to biomonitoring[END_REF]. Crassostrea virginica exposed to Cd rapidly accumulated high levels of the metal in soft tissues. Maximum concentrations of Cd were independent of exposure concentrations [START_REF] Daskalakis | Variability of metal concentrations in oyster tissue and implications to biomonitoring[END_REF]. The target organelle of Cr accumulation was shown to be the lysosome where the metal was associated with phosphorus and sulfur and trapped in an insoluble form. In M. edulis, the metabolism and transport of Cr was compared with other metals, and led to the conclusion that Cr is metabolized and distributed differently from most toxic metals [START_REF] Usero | Trace metals in the bivalve mollusc Chamelea gallina from the Atlantic Coast of Southern Spain[END_REF][START_REF] Phillips | Biomonitoring of Trace Aquatic Contaminants[END_REF]. Cadmium and lead concentrations in sediment and tissues tend to be of similar magnitude, suggesting that they are not regulated by the organism [START_REF] Phillips | Biomonitoring of Trace Aquatic Contaminants[END_REF]. These metals have not been demonstrated to be utilized in metabolism [START_REF] Eisler | Trace Metal Concentrations in Marine Organisms[END_REF]. Tissue Cu and Zn concentrations were higher than the sediments, indicating that these metals are probably taken up actively possibly reflecting their use in metabolism [START_REF] Genest | Heavy metals in Mercenaria and sediments form the New Bedford harbor region of Buzzard's Bay, Massachusetts[END_REF]. Some investigators have reported significant depuration of Cd by some bivalves but other organisms show very effective retention of Cd. [START_REF] Genest | Heavy metals in Mercenaria and sediments form the New Bedford harbor region of Buzzard's Bay, Massachusetts[END_REF] found that Zn accelerated Cd transport from gills to other organs. Prior exposure to Zn retarded Cd uptake in the excised gill of A. anatina. Much of what is known about Cd bioaccumulation by marine organisms has come from laboratory studies and therefore extrapolation to certain field conditions may be inherently uncertain [START_REF] Ray | Bioaccumulation of cadmium in marine organisms[END_REF].

The second mechanism could operate for one or two reasons, either the different animal populations have various abilities to accumulate metals at the different stations or the uptake of other metals. The second alternative has been extensively studied with other molluscs (Romeril, 1971;Martin et al., 1975;Howard, 1975;Coughtrey et al., 1977). It is also suggested by the results of the present data that a significant correlation exists between Cu, Cr and Ni at all stations. Coughtrey et al., 1977) showed that the correlation between Cd, Pb, Zn and Cu for Helix aspersa was significant at the less contaminant site and non significant and marginally significant at the more contaminant site.

Considering that many factors could affect the trace metals bioaccumulation in mollusks, e.g. physiological state, size and season (Romeo et al., 1994) our study

investigated the metallic concentrations according to the season, animal size and distribution in organs. It was realized in two different sites of the Nha Trang bay.

BIOMONITORING

Biomonitoring

Chemical analysis of the environment matrix such as water, sediment is the most direct approach to reveal the heavy metal pollution status in the environment, while it cannot afford the powerful evidence on the integrated influence and possible toxicity of such pollution on the organisms and ecosystem. Biomonitoring is a scientific technique for assessing environment including human exposures to natural and synthetic chemicals, based on sampling and analysis of an individual organism's tissues and fluids. The results of these measurements provide information about the amounts of natural and manmade chemicals that have entered and remained in the organisms and the corresponding effects induced. Due to consistency between the selected organisms and the corresponding living space, biomonitoring can directly offer the data on the potential effects and actual integrated toxicities of pollutants, reflecting the corresponding deleterious degree in the environment [START_REF] Zhoua | Biomonitoring: An appealing tool for assessment of metal pollution in the aquatic ecosystem[END_REF] Biomonitoring of heavy metals and effect studies on natural populations of organisms must take into account the pollution induced community tolerance that is expected from communities that are exposed to particular pollutants for a long time (several generations). [START_REF] Blanck | Pollution-induced community tolerance-A new ecotoxicological tool[END_REF][START_REF] Chapman | Ecotoxicology of metals in aquatic sediments: binding and release, bioavailability, risk assessment, and remedation[END_REF]. By virtue of their wide spread geographic distribution, reasonable size and availability throughout the year, bivalves (mussels, clam and oysters) have been used as sentinel bioindicators of trace metal pollution monitoring in coastal regions of many countries.

In biological indication passive and active monitoring are accepted as a general approach, at different levels of organization. In passive monitoring degradation of the ecosystem, elimination of sensitive species and reduction of biodiversity can be revealed as adverse consequences of pollution at the level of populations, while at the level of individual's accumulation of toxic substances in specimen, in organs and tissues indicative of pollution in the environment can be traced. In active monitoring the response of artificial or modified populations, behavioral patterns of specimen, specific function of organs like movement, feeding, respiration, reproduction and the neural regulation as well as cellular and sub cellular events are studied under the effect of toxic substances [START_REF] Solanki | Molluscs in biological monitoring of water quality[END_REF]. The main purpose of biomonitoring heavy metal concentrations in biota has been to determine the toxicological threat posed to organisms and also for health risks to humans from the ingestion of edible species. (Valavanidis et al., 2000) An important approach to assessment of risk from environmental and occupational exposures is biomonitoring which provides an estimate of the total dose absorbed and gives indirect access to determination of target site concentrations. To reveal the presence of pollutants and to measure their toxic effect biological indicators can be used. At the level of individual's accumulation of toxic substances in specimen, in organs and tissues indicative of pollution in the environment can be traced. [START_REF] Sanjay | Evaluation of mollusc as sensitive indicator of heavy metal pollution in aquatic system: a review -IIOAB[END_REF]. In biomonitoring surveys, the toxic elements arsenic, cadmium, chromium, cobalt, lead and nickel etc are used as examples to illustrate the disturbing factors in the interpretation of biomonitoring results (Christensen J.M, 1995) The use of aquatic organisms as bioindicators for trace metal pollution is very common these days. Molluscs are among the organisms most used for this purposes. It is widely observed that various species of molluscs are the effective sentinel organisms and can achieve high concentrations of metals and metalloids relative to concentrations gradient of these substances in the surrounding environment (Rainbow et al., 2000). In contrast, biomonitoring using specific organisms offers an appealing tool for the assessment of trace metal bioavailabilities over space and time in coastal water bodies by providing integrated measures of the ecotoxicologically significant fraction of trace metals partitioned in water, suspended particulates, and sediments (Rainbow 1995(Rainbow , 2006;;Zhou et al., 2008).

Marine mollusks, including clams, mussels and oysters, can be employed as bioindicators to assess the marine environment, since they can accumulate (concentrate) trace elements and other substances [START_REF] Labrecque | The concentrations of selected trace elements in clams Trivela mactroidea along the Venezuelan coast in the state of Miranda[END_REF]. In addition to using bivalve molluscs as bio-indicator organisms of coastal contamination, it is also recognized that they can be important links for contaminants between sediments and higher organisms, including man, that consume them. Hence, information on the contaminant concentrations in their tissues is potentially useful in considering toxicological and public health implications of estuarine contamination [START_REF] Cheggour | Metal concentrations in sediments and clams in four Moroccan estuaries[END_REF]

Bioindicator

Bioindicators are species that accumulate bioavailable chemical forms of contaminants. Among aquatic organisms, gastropods and bivalves mollusks have been recognized as a useful tool for monitoring of the environment they live in because of their ability to accumulate chemical elements and/or compounds in their tissues proportionally to their bioavailability and thus can be used as indicators of aquatic metallic pollution.

They are filter feeders, herbivores or carnivores and have the potential to bioconcentrate contaminants, which would normally be present in the water or within sediments at concentrations too low for detection by routine monitoring techniques. They are also ideal species for environmental monitoring, because their sedentary nature does not require consideration of complex migratory factors in the interpretation of the bioaccumulation data. They are sedentary organisms filtrating large amounts of water allowing them to accumulate the substances from the environment. They also satisfy the other conditions to be bioindicators hence very appropriate for monitoring because of their abundance and wide geo-graphical distribution, relative longer life span, suitable dimensions, size, weight, easy identification and collection, abundance in an ecosystem and accumulate the elements to a degree suitable to measure for hazard and risk assessment [START_REF] Chase | Gulf watch: monitoring spatial and temporal patterns of trace metal and organic contaminants in the Gulf of Maine [1991-1997] with the Blue Mussel Mytilus edulis L[END_REF].

Mollusks are benthic fauna in the food chain ecosystem, they live and move slowly to feed on the surface of sediments and thus accumulate higher concentrations of heavy metals than other organisms (Eisler, 1980). Many benthic organisms accumulate trace metals to the levels reflecting those in the environment. Tissue metal concentrations can reflect contamination, and molluscs in particular may therefore be sensitive bioindicators of anthropogenic metal inputs [START_REF] Hendozko | Heavy metals in Macoma balthica and extractable metals in sediments from the southern Baltic Sea[END_REF]. To achieve a better estimate of bioavailable metal exposure, it is recommended that the tissues of the organisms be analyses for trace metals (Luoma S.N, 1983).

Benthic mollusks play an important role as bioindicators for trace metal pollution and appear more and more often in global monitoring programs (Rainbow et al., 2000).

Among aquatic organisms suitable for biological monitoring mollusks occupy a prominent place and they are often used both for passive and active biomonitoring and in hazard and risk assessment [START_REF] Salanki | In situ bioassays with caged bivalves to estimate exposure and bioeffects in a comprehensive monitoring program[END_REF]. As even closely related species may exhibit different accumulation strategies for trace elements, there is a need to identify widespread cosmopolitan bioindicators to allow intra-specific comparison of accumulated metals concentrations over large geographical areas (Rainbow et al., 2000). Mollusks can accumulate and integrate concentrations of several metals in seawater for relatively long intervals. They also assimilate trace metals from their food and from the ingestion of inorganic particulate material (Philips D.J, 1977).

The potential ecological effects of rising levels of heavy metals concentrations in the environment are of great concern due to their highly bioaccumulative nature, persistent behavior and higher toxicity. These chemicals biomagnify in the food chain and impose various toxic effects in aquatic organisms. Mollusks reflect the higher degree of environmental contamination by heavy metals and are the most useful bioindicator tools.

Several studies and research work have been cited to establish and evaluate the relationship between metal contents of water column, sediment fractions, suspended matter and mollusk tissue concentrations.

Suggested characteristics for bioindicator organisms (ODEQ, 2007)

-Tolerance to a wide range of metal exposures, ability to accumulate metals without suffering mortality -Sedentary habits, slow and limited range of movement -Sufficient life span to allow for sampling of more than a one year class -High presence, abudance and wide geographical distribution in the study area -Sufficient size to allow chemical analysis of tissue samples -Hardiness, ability to remain healthy during sampling and laboratory incubation -Size, weight, relative ease of sampling and identification -High metal accumulation rate -Responsiveness to changes in metal exposure

The potential ecological effects of rising levels of heavy metals concentrations in the environment are of great concern due to their highly bioaccumulative nature, persistent behavior and higher toxicity. These chemicals biomagnify in the food chain and impose various toxic effects in aquatic organisms. Mollusks reflect the higher degree of environmental contamination by heavy metals and are the most useful bioindicator tools. (Cevik F et al., 2011)

Marine moluscs as biomonitors for heavy metals

Benthic mollusks are the organisms most often used for the biomonitoring of metal contamination. Bivalve mollusks have an ability to accumulate heavy metals to various orders of magnitude with respect to the levels found in their environment [START_REF] Usero | Heavy metal concentrations in molluscs from the Atlantic coast of southern Spain[END_REF]. The levels of heavy metals accumulated by marine mollusks are a function of water quality, but also of seasonal factors, temperature, salinity, diet, spawning and individual variation, among other factors.

Moreover, the levels of metals accumulated in some marine organisms may be many orders of magnitude above background concentrations, thus demonstrating the potential of certain species as bioindicators of heavy metal pollution (Hamed et al., 2006).

Mollusk, especially the species from bivalve class is frequently used as bioindicator in environmental monitoring. Bivalve is one of the organisms that have the criteria of potential bioindicator due to its ability to accumulate pollutants from its ambient. Usually, the level of pollutant accumulated in such an organisms tissue is used for assessing the level of pollution in its habitat (Al-Madfa et al., 1998;[START_REF] Abdallah | Accumulation of Lead and Cadmium in the Marine Prosobranch Nerita Saxtilis, Chemical Analysis, Light and Electron microscopy[END_REF].

Blue mussels Mytilus edulis are commonly used to monitor heavy metal pollution (Riget and others, 1996). One study found the blue mussel to be a particularly effective sentinel organism because soft tissue residues closely parelleled the heavy metal concentration gradient of their surroundings [START_REF] Julshamn | Distribution of trace elements from industrial discharges in the Hardangerfjord, Norway: A multivariate data analysis of Saithe, Flounder and Blue Mussel as sentinel organisms[END_REF]. The mussels Mytilus edulis and Mytilus californicus are indigenous and abundant in bays and estuaries around much of the United States coastline. Mussels feed continuously throughout the year, making them a convenient species for physiological study. Furthermore, mussels are excellent for transplanting because healthy adult animals are plentiful and can be distributed easily into field cages at various sampling locales [START_REF] Epa | Biomonitoring for Control of Toxicity in Effluent Discharges to the Marine Environment[END_REF][START_REF] Phelps | A report on the coastal environmental assessment stations (CEAS) program[END_REF].

American oysters Crassostrea virginica have shown an ability to concentrate cadmium (Cd) to a great degree in edible tissues [START_REF] Eisler | Cadmium uptake by marine organisms[END_REF]. Cadmium concentrations, exceeding levels considered dangerous for human consumption (13.0 mg/kg), were attained by oysters exposed to Cd concentrations in water considered safe for drinking (10.0µg/l).

In another study [START_REF] Epa | Chemical and Biological Survey of Liberty Bay[END_REF] In general, marine shellfishes have a low bioaccumulation tendency for chromium, but a rather high tendency to accumulate arsenic, cadmium, copper, lead, mercury, silver and zinc (Trocine et al., 1996;[START_REF] Saiz-Salinas | Heavy metal levels in intertidal sediments and biota from the Bidasoa Estuary[END_REF]. Shellfishes, particularly oysters, passively accumulate many metals much more readily than fishes: this suggests a priority for monitoring in metal contaminated areas [START_REF] Epa | Metal Bioaccumulation in Fishes and Aquatic Invertebrates[END_REF][START_REF] Julshamn | Distribution of trace elements from industrial discharges in the Hardangerfjord, Norway: A multivariate data analysis of Saithe, Flounder and Blue Mussel as sentinel organisms[END_REF]. Moreover, sediments are an important sink for most metals in aquatic environments. Further, information concerning the biological and physicochemical factors affecting metals mobilization in sediments, would be valuable.

Various metal accumulating bivalve and gastropod species show a high presence and abundance in marine ecosystems therefore they are suitable for different monitoring projects for example:

• Perumytilus purpuratus, Semelle solida and Tagellus dombeii [START_REF] Gregori | Variability of cadmium, copper and zinc levels in molluscs and associated sediments from chile[END_REF] • Bembicium nanum [START_REF] Gay | Natural variation of copper, zinc, cadmium and selenium concentrations in Bembicium nanum and their potential use as a biomonitor of trace metals[END_REF] • Pyganodon gandis [START_REF] Bonneris | Sub-cellular partitioning of metals (Cd, Cu, Zn) in the gills of a freshwater bivalve, Pyganodon grandis. Role of calcium concretions in metal sequestration[END_REF] • Crassostrea angulata, Scrobicularia plana, Palameon longirostris, Uca tangeri, Melicertus kerathurus [START_REF] Blasco | Heavy metals in organisms of the River Guadalquivir estuary. Possible incidence of the Aznalc´ollar disaster[END_REF] • Crassostrea virginica [START_REF] Apeti | A Model for Bioaccumulation of Metals in Crassostrea virginica from Apalachicola Bay, Florida[END_REF] • Radix ovata, Vivitarus spp [START_REF] Gundacker | Comparison of heavy metal bioaccumulation in freshwater mollusks of urban river habitats in Vienna[END_REF] • Rapana venosa, Neverita didyma (Lee C et al., 2006) Some mollusks species propose to use as bioindicator in specific regional as:

Adriatic Sea, Black Sea: Mytilus galloprovincialis, Perna perna, Crassostrea angulata, Crassostrea virginica [START_REF] Blasco | Heavy metals in organisms of the River Guadalquivir estuary. Possible incidence of the Aznalc´ollar disaster[END_REF][START_REF] Apeti | A Model for Bioaccumulation of Metals in Crassostrea virginica from Apalachicola Bay, Florida[END_REF] Mediterranean Area: Monodonta turbinata, Pattela caerulea, Mytilopsis sallei, Mytilus californicus (EPA,1989) Chinese Bohai Sea: Mytilus edulis, Crassostrea talienwha nensis, Ruditapes philippinarum, Rapana venosa [START_REF] Chassard | Chromium uptake, distribution and loss in the mussel Mytilus edulis: a structural, ultrastructural and microanalytical study[END_REF][START_REF] Lee | Metal contamination in urban, suburban and country park soils of Hong Kong. A study based on GIS and multivariate statistics[END_REF] Gulf of Aden, Yemen: Turbo coronatus, Acanthopleura haddoni, Ostrea cucullata, Pitar spp A preliminary investigation conducted to assess the pollution in the coastal part of Vietnam with focus on trace elements including heavy metals at significant site specific differences were reported in the Table 1.3 as bellow: The bay of Nha Trang is the protected beach in Viet Nam, yes its fauna with the world's richest diversity, which is very important in term of biodiversity conservation. A total of 694 species of molluscs in 2 classes Gastropoda (435 species) and Bivalvia (259 species) were recorded in Khanh Hoa coast (Nghi B.Q, 2001). Compared with some other areas, species composition of mollusks in Nha Trang Bay was more abundant than Cat Ba -Ha Long Bay (372 species) (Duc N.X, 2001) and Tonkin Gulf (470 species, Cai Yingya et al., 1988). Some of there dominated bivalve species in Nha Trang bay that are showed in Table 1.4 T et al., 2002).

Upwelling off Vietnam occurs during SW monsoon when a cyclonic circulation cell is the northern and an anti-cyclonic give in the southern are initiated by spatial asymmetry in the wind field (Wu et al., 1998) The bay of Nha Trang is the marine protected area (MPA) in Viet Nam; yes its fauna with the world's richest diversity, which is very important in term of biodiversity conservation. These mollusks are also indispensable for local communities because they provide livelihood and economy as fishery and tourism bases and protect land as a natural breakwater. However, they are at high risk of destruction due to coastal development accompanied with the rapid population growth in the neighboring coastal area. In this context this paper tries to evaluate the current state of knowledge trace metal impact on marine mollusks.

Human pressure

There are a number of environmental issues in the coastal zone of Khanh Hoa province. Amongst these the most significant are the pollution problems (mainly along Nha Trang coast) and the occurrence of tropical cyclones. The pollution issues along Nha

Trang coast that have arisen due mainly to the indiscriminate discharge of effluent from industrial and agricultural sources and disposal of untreated liquid and solid wastes generated from domestic sources into the coastal environment. The sewage waste generation in Nha Trang is some more than million cubic meters /day out of which 40% is domestic waste and 60% is industrial waste. This waste is dumped into the Nha Trang sea via Cai river, Be river, Nha Trang harbour area, and small waste drains. The other coastal areas having industrial pollution problems are ship breaking industries based in area. The heavy metals, persistent organic pollutants, air pollution and oil pollution are more significant.

Population Pressure

The coastal city of Nha Trang has an estimated population of 500 thousands, and is the biggest tourist center of Viet Nam with more than 1 million tourists per year. In 1995, the total population of the coastal province was estimated to be 675,000 (6,5% of the total national population). Population densities vary from 7.07 people/km in with an average density of 37 people/km 2 . Average growth rates in 1993 range from 2.7 to 4.6% in region.

Coastal Development

The Government's goals for Khanh Hoa provinces and municipality originally planned for economic development in Viet Nam through the rebuilding of urban growth tourism port expansion and industry.

Urbanization

Investment in coastal development will lead to increased urban growth as population from the rural areas move to tourism and associated services for increased economic opportunities. The Urban environment infrastructure is current insufficient to meet the requirement of even the current urban population. Without appropriate investments the environmental quality of this town will degrade.

Tourism development

Coastal area is a matter of great in importance to the future of both in term of economical and environmental considerations. This development is expected to lead the way for Coast to develop as commercial centre which would substantially increase its population. The impact from this development may be affected to coastal inland resources coastal water resources. The number of tourists in this coastal province has been estimated at 1.206.000/year in 2010. An approximately 9% growth in tourism arrivals in Nha Trang is Russian and Chinese. This indicated that the positive growth in the potential tourism development sector in the country.

Port development

Coastal ports expansions developments can make a contribution to the economy as a main hub for growth of maritime transport which should in turn attract manufacturing entries but may also have an adverse impact on the surrounding environment. These effects of new expansion port can be focused upon location port construction and port operation.

These lead to impacts on water quality coastal hydrology bottom contamination marine ecology air noise waste management and visual quality.

Industrial development

The industrial development zone was established in Khanh Hoa called Cam Ranh Industrial Zone. This industrial zone include petrochemical production to exploit recently confirmed oil and gas reserves in the seafood processing based on the local fisheries in the area timber processing and are manufacturing. However these industries pose potential damage to the environment outlets generally heavily contaminated with municipal sewage, industrial effluent and sediments contribute a significant quantity of pollution load to the coast. Regarding heavy metal presence in the biota, natural sources account for a background exposure. Among biota, zooplankton may contribute to the transfer of trace metals to higher trophic levels are the recommended groups for the base line studies of trace metals in the marine environment (Rejomon et al., 2008). Zooplankton accumulates metals directly from water by absorption, also by assimilation through food substances.

The bioavailable fraction can be assessed only by determining the amount of metals incorporate into organisms which is the main goal in biomonitoring

Reversing environmental degradation trends in the Nha Trang bay

In Nha Trang Bay, the Cai and Be rivers discharge into the sea, and the catchments of these two transboundary rivers cover parts of coastal region, and their water and sediment discharges greatly influence the coastal seas. The impact of human activities include changes in the quality of the coastal and marine environments due to the increased use and accumulation of pollutants and the loss of habitats. These impacts have resulted in increasing unpredictability and severity of coastal problems such as floods, erosion, sedimentation, and saltwater intrusion; environmental pollution; and the degradation of ecosystems, with accompanying decrease in biodiversity and fishery productivity (Thanh et al., 2004).

Environmental issues of Nha Trang coast have become more prominent recently mainly due to increasing population and intensifying industry after year of 1997. Among the various contaminants, trace metals are of particular concern due to their environmental persistence, available through biogeochemical recycling and encountering ecological risks.

Hence, studies on trace metals influenced by seasonal variability and their concentrations in the coastal area are much needed.

Environmental quality

Water quality

According to the results of monitoring program of University of Fisheries Nha

Trang in 2003, the salinity was ranged 25.2-34.0 mg/l, pH: 7.9-8. The size structure:

Perna viridis is a large mussel, 80-100 mm in length, occasionally reaching 165 mm (Rajagopal and others, 1998). The shell has a smooth exterior surface characterized by concentric growth lines and slightly concave ventral margin with a bright green colour in young and fading to brown with green edges as it matures.

Characteristic:

The sexes of the species are separate and fertilization is external. Spawning generally occurs twice in the year (Chatterji and others, 1984). Fertilized egg develops in to larvae and remains in the water column for two weeks. During this planktonic period, larvae will be widely dispersed by physical processes, but may aggregate periodically at certain depths through a variety of biological processes, most notably vertical migration (Hayes and others, 2005). The larvae completely metamorphose in eight to twelve days of growth. Sexual maturity typically occurs at 2-3 months of age with a length of about 15-30mm (Benson and others, 2001). Perna viridis has the greatest growth rate of the mussels studied, which have a life span of about 2-3 years (Shafee, 1979). Maximum growth of the green mussel occurs 2m below the surface because of the increased productivity of the water at that depth and a narrow area of temperature and salinity fluctuation (Sivalingam, 1977).

Perna viridis is commonly available, popular and most consumed seafood in Khanh Hoa region. It is available throughout the year and consumes by making various preparations.

Solen regularis (Dunker, 1862)

Classification:

Phylum: Mollusca

SELECTION OF SAMPLING SITES

The 4 sampling locations were selected based on differences in physicochemical conditions, contamination status, varied anthropogenic pressure and feasibility of sampling. These sites still sustain rich biodiversity and were selected to be far from direct discharges. The characteristics of the 4 studied locations were:

Location 1 (Tan Dao -TD): is characterized by dense mangroves, is located between Nha Trang and Nha Phu Bay. During the past 30 years, a considerable decline in mangrove stands occurred, threatening their survival in this area. This area far removed from obvious sources of metal pollution, with no appreciable human activities. Hence, this location was chosen as a reference site to provide information on natural metal levels in non contaminated macrobenthic. Map of the sampling sites was showed in Figure 1. Their accurate positions were determined by GPS localities instrument with the accuracy of ±5m as is shown in Table 3.1. Common physicochemical parameters like pH, temperature, moisture, grain size, TOC; essential trace metals like zinc, copper, aluminum, iron, and non-essential arsenic, lead, cadmium, and chromium were estimated to know their levels present in the surface sediment and mollusk tissues.

ANALYZING METHODS

The quality and quantity of sampling size, site selection and sample analysis methods for each of these parameters study is based on an International Standard Analytical Methods.

Sediment samples

Method of collection sediment samples

The uppermost 15cm of surface sediment was sampled using a grab sampler. Subsamples were taken from the central part of the grab to avoid contamination. Further more, the state of the sediment surface was inspected to make sure it was relatively undisturbed (i.e., lack of channeling or sample wash out) and that the desired penetration depth was achieved. All sampling equipment (e.g., scoops, containers) was made of non contaminating material cleaned before and after each sample. If all the sampling criteria were met, the surface sediment was subsampled by a flat scoop. This device will allow a methylene chloride prior to initial use and between uses at each station. Finally, the sediment samples were placed carefully into acid-rinsed glass vials via a stainless spatula and then stored at -4 o C before analysis to insure undisturbed samples and eliminate cross contamination.

Preparation and storage of sediment samples

Using plastic container for storage sediment samples and frozen at -4 o C to be transported to the laboratory of Environmental faculty of HCMUS. Here, the sediment samples were analyzed some physicochemical parameters as pH, Total organic carbon, moisture and grain size. There sediment samples were dried at room temperature then pulverized to fine powder using an agate mortar and pestle and then sieved through a 2mm mesh sieve to remove large particles. The samples were storaged at -20 o C until they were transported to the ECOMERS laboratory (UNS-France) for further analyses.

Analyzing physico -chemical characteristic

Grain size analysis

The grain size of the sediment samples was determined by standard sieving method for grain size >0.063mm and by pipetting for particle <0.063mm according to Casagrande method (Strmac, 1952). Fifty grams from dried sediment were prepared and then sieved to produce six sediment particle size fractions, starting with particles smaller than 2mm in diameter. The percentage of a given particle size in the sample, on a mass basis, was then calculated the size distribution of sediment and classification according to the Shepard classification [START_REF] Shepard | Nomenclature based on sand-silt-clay ratios[END_REF]: fraction of sand, silt, clay or silt clay in table 3.

= ------------------------------------------x 100%
Weigh of dry soil

Bulk density determination Db

Soil bulk density is a measure of how dense and tightly packed a sample of soil is.

It is determined by measuring the mass of dry soil per unit of volume (g/ml or g/cm 3 ). Soils made of minerals will have a different bulk density than soils made of organic material. The carbonate content was determined as weight loss after the treatment with 4 M HCl (Loring et al, 1992).

Sediment digestion method

Dried sediment samples were sieved through a nylon mesh to obtain particles smaller than 0.2mm in diameter for determination of heavy metal concentration.Schema of sediment digestion methos show in figure 3.2.

Figure 3.2 Schema of sediment digestion method

The samples were digested following the EPA guideline for siliceous sediments (EPA 3050b, 1996) for digestion of sediments using an Automatic Microwave Digestion System (Microwave 3000). A 0.1 gram of dried and sieved sediment samples were weighed out in a Teflon (R) reaction vessel then added with mixed of 9ml concentrated acid HNO 3 (69%) and 3ml of HCl (36,46%). Vessels then were placed in the rotor in the microwave. The vessels were heated to at least 240 o C over 35 minutes and then held at 210 o C for at least 15 minute. The samples were cooled and then filtered to remove insoluble materials, before dilution to volume to 50 mL with ultra-pure water (Figure 3.2).

The solution in preparation of being analyzed the concentration of heavy metals measured using Perkin Elmer inductively coupled plasma optical mission spectrometer ICP-OES (Picture 3.1).

Dried sediment 1g

Place in acid washed digestion tubes 9ml of HNO The accuracy of the analytical procedures employed for the analysis of metals in sediment samples was checked using the marine sediment IAEA-433 as certified reference material (Appendix 4.1)

Mollusk samples

All the mollusks examined in this study are commercial species which have sedentary life style, filter-feeder invertebrates and have the ability to accumulate metals from the surrounding environment. Mollusk samples were identified to species level by Biological department of HCMUS.

Preparation of tissue samples

In the laboratory, all samples were removed from the plastic containers and washed with running distilled water. The whole soft tissues were removed from their shells and prepared for analysis. Mollusks were placed on a critically cleaned glass cutting board, and

Sample measurement

Moisture analysis

Moisture analyses were conducted on all samples in duplicate at laboratory HCMUS. Three individuals from each mollusks species sample were placed into each of three aluminum weighing pans. Moisture was determined by measuring the difference between sample mass before and after drying in an oven at 60 degrees for more than 24 hours. Percentage moisture can be used to compute metal concentrations in the tissue on a dry weight basis.

Body size measuring

Each individual sample was classified, weighed and the weight was recorded on the sample composite form. Samples were measured using caliper 0.1mm for lengh, wide of shell and digital balance 0.01g for weight measurement

Digestion method of mollusks

Processing of the samples for analysis began in 1 month after collection.

Determination of heavy metal concentration in mussels tissues according to Digestion method of organically based matrices (EPA 3052).

Before beginning the processing of the mollusks samples, in preparation for metals analysis, the equipment was cleaned using the method described in EPA 3052. The twelve individual mollusk samples from each sampling site were composited into a single sample by combining portions of each species into a common sample used for analysis of metals.

The dried tissues were powdered, and aliquots of 0,5g were digested for 30min at 95°C with 9ml HNO 3 (65%, Merck) in closed beaker. Additional nitric acid was added if the samples were evaporated to near dryness. After cooled, 3ml HCl (Merck) then 0.5ml of H 2 O 2 were added and heated for few minute to make the solution clear. The digests were cooled and diluted to 50ml with ultra pure water and metals were analyzed (Figure 3.4).

Digested tissues were analyzed for the same heavy metals as in sediment using the same procedure using an ICP-OES (Perkin Elmer). All metal concentrations in the tissues

Standard solutions

Standard solutions of known concentrations were prepared from purchased certified solutions. A standard curve was prepared each day of analysis using the standard solutions.

Metal standards were made as sets of five concentrations plus a reagent blank with one set run at the beginning of the analysis and another full set analyzed with each set of forty samples. Three absorbance readings are taken for each sample by the instrument, with the reported concentration being an average of those readings to ensure accuracy of standards and calibration curve.

Lab control

Quality of analysis was monitored by several methods during this study. Analysis of reagent blanks determined if reagents contained appreciable quantities of metals or if contamination occurred during the sample preparation. Analysis of blanks showed significant amounts of iron and aluminum compared to the reference material and would account for the high percentage agreements for these metals.

Certified reference materials

Accuracy is measured by analyzing certified reference standards IAEA -433 for marine sediment and mussel tissue with MAB3 reference material (Appendix 4.1)

Quality assurance for sediment analysis

Reference material IAEA-433 was purchased to use as a reference standard for determining method accuracy. The marine sediment was prepared from the International Atomic Energy agency, Analytical Quality Control Services, A-1400 Vienna, Austria.

Metal concentrations measured in reference sediments following sample digestion procedures were in general agreement with expected values with a low mean agreement of 77.95% for magnesium and a high agreement of 319, 57% for manganese. The acceptable recoveries for sediment sample ranged from 88.52 % for arsenic to 114.19 % for lead (Appendix 4.1).

The Standard Reference Material IAEA-433 certified concentrations of arsenic, cadmium, chromium, copper, zinc, and lead in sediment for method analysing. Certified values were not provided for nickel, molybdenum, magnesium and manganese which the method using.

Quality assurance for mollusks analysis

Mussel tissue was also purchased to use as a reference standard for determining method accuracy. The mussel tissue was purchased from the International Atomic Energy In conclusion, the used method certified a good quality assurance for measure concentrations of arsenic, copper, iron, lead, zinc and aluminum in tissue. Mn and magnesium had recovery percentages ranging from 148.77 to 473.45 % which are both unreasonable and should be ignored. Certified values were not recommended for chrome, cadmium, nickel, molybdenum and aluminum.

Detection limit of the method

Data are reported in one of three categories. Some samples sediment concentrations below the Detection Limit (DL) of the method. When this happens, the concentration for that sample is reported as a "less than" numerical value. Some data were measured above the detection limit (DL), but are less than ten-thirds of the detection limit and are marked as data between the DL and the Limit of Quantitation (LOQ). There is a lower confidence with values between the DL and LOQ than those above the LOQ. The third categories of data are values above the LOQ. Proposed general limit of heavy metals analysis method as show in Appendix 5

The sequence of metal concentration varies with the distance between stations and seasons. An overall view of the result indicated that the heavy metals Fe, Al, As, Zn, Ni, Pb, Cr, Cu, Cd and Mo concentrations were above the permissible limit. The highest metal levels estimated along the shoreline of the Be river, reflecting long-term exposure to various activities. Fe is the most abundant metal, whereas arsenic and chromium present in minimum concentration when compared to other metal's concentrations. The range of variation for each metal is unique; their concentration is not restricted only by anthropogenic input. The results suggest that complex combination of factors act on surface sediment thereby increase the metal concentration. If this valid point consider in future for bay of Nha Trang possible influences on metal accumulation in surface sediment may be worked out and follow up monitoring programmer may be initiated.

DATA ANALYSIS METHODS

Method assessment of heavy metal contamination in sediment

Metal assessment indices

Degree of contamination Cd

Contamination indices Cd, which compare the contaminants with clean and/or polluted stations, measured in the study area or simplyaggregate the metal concentrations.

Enrichment factor EF

Application of enrichment factor to the interpretations of result from the biomonitoring studies. EF is ratio of chemical concentration of an element in a soil or sediment to that in its fresh parent material (Battelle 2003) [START_REF] Hakanson | An Ecological risk index for aquatic pollution control. A sediment tological approach[END_REF].

Calculating enrichment factor EF as

([As]/[M]) sample (sediment) EF = ------------------------------------ ([As]/[M]) background (crust

Sediment Quality Guidelines (SQGs)

As the mean to predict adverse effect derived from contaminated sediments numerous sediment quality guidelines (SQGs) were developed by overseas researchers.

Basically, the purpose of SQGs development is to protect the aquatic and benthic organisms from the adverse effects of sediment contaminations. SQGs are also being used for prioritizing contaminated areas, evaluating contamination spatial patterns as well as for designing a monitoring programmer. Biological affects are not able to be predicted from metals concentrations in sediment. As an alternative, adverse ecological effects are identified and being used as the reference in the SQGs. The adverse ecological effects are attributable to sediment-borne metals concentrations. Among the most widely applied SQGs are as following:

-Effect-range guidelines [START_REF] Long | Incidence of adserve biological effect within ranges of chemical concentrations in marine and estuarine sediment[END_REF] -National Oceanic and Atmospheric Administration (NOAA), US -Provincial Sediment Quality Guidelines for Metals and Nutrients (Persaud et. al., 1992) -Ontario Ministry of the Environment, Canada -New York Sediment Screening criteria (1999) -Department of Environmental Conservation, New York -Interim Sediment Quality Values (ISQVs) [START_REF] Chapman | Development of Sediment Quality Values for Hong Kong Special Administrative Region: A Possible Model for Other Jurisdictions[END_REF]. Hong Kong Special

Administrative Region

These guidelines are summarized in Table 3.8 according to their terms and application.

Effect-Range Guidelines

Effect-range guidelines is one of the most widely applied SQGs for estuarine and marine location. These SQGs was developed by National Oceanic and Atmospheric Administration (NOAA) in the United States of America (Longet al, 1995). This set of guidelines adopted the terms of effect-range-low (ER-L) and effect-range-median (ER-M).

Effect

Evaluation of bioaccumulation

The methods used in this study evaluate potential bioaccumulation by:

-Using the metal pollution index with mollusk tissues data -Comparing the estimated or measured concentration of matals in tissue to acceptable tissue levels (ATLs) for humans and relevant classes of wildlife and/or to critical tissue levels (CTLs).

-Calculating bioaccumulation factor BSAF with site-specific benthic invertebrate tissue data.

Calculation of metal indices

Metal pollution index (MPI)

MPI is a mathematical model which enables the presentation of all results from the trace metals concentrations in the muscle tissue of bivalve as one value to classify the biological variation from an unpolluted zone to a highly polluted zone. The overall metal content of bivalves at various sites investigated in this study was compared using the metal pollution index (MPI) calculated with the formula [START_REF] Usero | Trace metals in the bivalve mollusc Chamelea gallina from the Atlantic Coast of Southern Spain[END_REF] MPI = (Al x Fe x Zn x Cu x Cr x As x Cd) 1/7 The MPI value is used for screening and estimating the spatial and temporal variation of trace metal pollution based on the metal content with in muscle tissues of bivalves. The available data are of particular interest concerning filter-feeding bivalves which are well known for their ability to reflect environmental levels of trace metal contaminants in estuarine ecosystems. According to [START_REF] Usero | Heavy metal concentrations in molluscs from the Atlantic coast of southern Spain[END_REF] The metal/shell weight index (micrograms of metal per gram dry shell weight or the metal content per unit of shell weight) was calculated according to [START_REF] Fischer | Cadmium body burden/shell weight of mussels: aprecise index for environmental monitoring[END_REF] According to Sotoet al., 1995, the metal/shell weight index provides a reliable tool for the assessment of metal bioavailability to sentinel mollusks in coastal waters. It is useful in eliminating part of the variability due to the inherent biology of sentinel mussels (Bartoloméet al., 2010). Analysis tissues of cookle or mussels (popular seafood) to evaluate the bioaccumulation factor of toxic element by marine biota.

Metal bioavailability was calculated as metal concentrations in soft tissues (mg/kg dry weigh metal content per unit shell weight (mg/kg dry shell weight). Additionally, the MSWI was calculated according to Fischer, 1984 as below:

Metal conc. in soft body (µg/g) x Dry wt of soft tissues (g) Metal/shell weight index = ---------------------------------------------------------------------Dry shell weight (g)

The metal/shell-weight index is independent of conditions such as nutritional stage, spawning, or tidal exposure (Fischer, 1983) and of environmental factors such as temperature, salinity and oxygen levels (Fischer, 1986). The metal/shell-weight index is also independent of changes in organism condition associated with the presence of pollutants, including metals (Marigomezet al., 1990) and has been significantly correlated with the biological effects of metals quantified at the cell and organism levels (Marigomez et al., 1990). According to Sotoet al., 1995 the metal/shell weight index does not exhibit seasonal fluctuations in non-polluted sites, as could be expected by the absence of local inputs. On the contrary, at a polluted site, the metal/shell weight index changed seasonally in relation to the intensity of the industrial activity, indicating fluctuations in the metal bioavailability.

Consequently, the metal/shell weight indices provide a method to detect differences and changes in the over all availability of cumulative metals, as Fischer, 1988 concluded from laboratory studies and Regoli and Orlando, 1993 as applied to the particular case of metal exposed mussels.

Condition index (CI)

According Romeo and Gnassia B., 1988, many factors, such as body size, could affect the concentration of trace metals in mollusks. [START_REF] Boyden | Trace element content and body size in molluscs[END_REF] reported that the total trace metal content per individual (µg/g) could be related to body weight (W) as the power function condition index CI.

A condition index (CI) was calculated (Crosby et al., 1990) for the majority of the cockles used for trace metal analysis:

Soft tissue dry weight (g)

CI = -------------------------------------- Shell cavity volume (litre)
The shell cavity volume is calculated by multiplying the length, width and depth measurements of the shell. However site had a greater affect on CI than season with seasonal differences being significant only a tone of the most coastal sites (MPI).

To investigate any relationship between trace metal concentrations and CI, the results from individual mollusks were combined for all sites and sampling times.

Spearmanrank correlation coefficients comparing the adjusted mean trace metal concentrations for all sites and the mean condition index (CI)

Assessment bioaccumulation

Screening level value (SLVs)

Compare the concentration of each metals in sediment at each location to its generic bioaccumulation screening level value (SLVs) or critical tissue levels (CTL). If the concentration is lower, no further action is required with respect to bioaccumulation for that metal. If the metal concentration is greater than its generic SLVs or CTLs, consider an area-wide statistical evaluation of the exposure point concentration taking into account the appropriate range of relevant species. Using some guidance for assessing bioacumulative potential with WDOE or DEQ's general screening approach for SLVs (mg/kg dry weigh)

and WQC -USEPA for CTL (mg/kgdry weigh). The SLVs value of some potentially heavy metals in soil/sediment was developed by WDOE, 1994 and BCE, 1999 is seeing in appendix 6.4.

Compare the concentration of each heavy metal in mollusk at each location to its Maximum permissible levels MPL. If the concentration is lower, no further action is required with respect to bioaccumulation for that metal. If the metal concentration is greater than its MPL, consider an area-wide statistical evaluation of the exposure point concentration taking into account the appropriate range of relevant species. However, for benthic organisms that are stationary or range over small distances, a comparison with the maximum concentration is appropriate. Some MPL values depose by different institution is combined in the appendix 6.5.

Bioaccumulation factor

Biaccumulation or bioconcentration screening following constituent as likely to bioaccumulate or bioconcentrate for soil or sediment. The some of factors represent for accumulation of chemical in organisms are listed in Table 3.9 as below: The most potentially bioaccumulative chemicals are: As, Cd, Pb, Hg,

Ecological risk analysis (ERA)

They are 3 guidance documents for interprete the ecological risk of metals:

-DEQ,2000: Guidance for assessing risk to human health -DEQ,2001b: Guidance for ecological risk assessment -DEQ, 2001a: Program quality assurance policy

Potential ecological risk index (PERI)

The potential ecological risk index is used to assess the potential ecological risks from metals in the soil (Medicietal. 2011; Abdallah 2011). It is defined by where C sample is the content of the metal measured in the studied soil sample,C reference material is the background value of the element (as mentioned above), and Ti is the response coefficient for the toxicity of the single element i. The values of Ti have been reported to be: As = 10, Cu = Pb = Ni = 5, Zn = 1, Cr = 2 and Cd = 30 [START_REF] Hakanson | An Ecological risk index for aquatic pollution control. A sediment tological approach[END_REF]Madiseh and others, 2009). No Ti value appears to be given for Mn. Comparing its toxicity with that of Cu, Cr, and Zn, a Ti value of Mn = 1 was adopted. RI<150 means a low ecological risk, 150 < RI<300 a moderate ecological risk 300 < RI<600 a higher ecological risk RI > 600 a severe ecological risk

Determination of Estimated daily intake (EDI)

Determination of EID for the metals in each shellfish species based on tissue concentration of metals and the average daily shellfish consumption rate (Santos and

The target hazard quotient (THQ)

The health risks from consumption of shellfish by local inhabitant also assess by THQ. The THQ is a ratio of determined dose of a pollutant to a reference dose level. If the ratio is less than 1, the exposed population is unlikely to experience obvious adverse effects. The method of estimating risk using THQ provides in the USEPA and it is described by the following equation: BW: is the average body weigh 60kg for VN AT: is the average time for non-carcinogen in day (365 day/year x number of exposure years)

EFr x ED x FI x MC THQ = -----------------------------x 0,
An important aspect in assessing risk to human health from potentially harmuful chemicals in food is the knowledge of the dietary intake of such substances that must remain within determined safety margins.

The target cancer risk (TR)

The method for estimating TR values is provided by the US. We compared these levels to the recommended dietary intake levels published by the World Health Organization (Demirzen et al., 2006;WHO 1993).

The USEPA, 2006 has recommended the estimation method of the potential health risk for the intake of inorganic As from various seafood species, such as the target cancer risk TR. TR is the incremental individual lifetime cancer risk fromthe ingestion of shellfish. The carcinogenic risk from inorganic As is expressed as the excess the probability of contracting cancer over a life time of 70years. The TR id difined as follows:

EFr x ED x IR x Cinorg x CPSo TR = -----------------------------------------------x 0,001

BW x ATc

Where EFr is exposure frequency (365 days/year) ED is the exposure duration (70year)

IR is the ingestion rate of shell fish (g/day)

Cinorg is As concentration in shellfish (mg/kg fresh weight)

CPSo is the oral carcinogenic potency slope obtained from the IRIS Integrated Risk Information system (1,5mg/kg/day USEPA, 2006) BW: is the average body weigh 60kg for VN AT: is the average time for non-carcinogen in day (365 day/year x number of exposure years) 0,001: conversion factor

Acceptable Tissue Levels for Humans

For human consumption, acceptable tissue levels for carcinogens (ATLhC) and noncarcinogens (ATLhN) are back-calculated from acceptable risk levels in accordance with federal guidance for establishing shellfish consumption limits and for conducting human health risk assessments (USEPA, 1989(USEPA, , 1997)). Separate levels were not calculated for men and women, as differences in consumption rates relative to body weight are minor.

Acceptable fish tissue levels for humans are calculated using the following equations. At 4 stations, sediments were found to be hightly sorted, indicating, therefore, a same origin of settled particles (terrigenous, anthropogenus, and biogenous). Grain size analysis and composition of sediment samples signified that they were mainly sand which range from 80. in CL. Coarse clay fraction prevails in most of samples, after what follows silt, which is also obvious from Shepard's classification of material (Shepard,1954).

In TD and ND, the mean size of muddy particles is in the range from coarse clay to fine silt, while median of particles is in narrow range from very fine silt to fine silt were represented in BT and CL. Sediment type is determined according to [START_REF] Shepard | Nomenclature based on sand-silt-clay ratios[END_REF] and at BT and CL locations is classified as clayeysilt. Other two samples TD and ND are classified as silty clay, due to some how increased percentage of clay. At all locations, percentage of gravel is very high, and only in the CL location, it is 0.54-6.8%, what could be due to close sea. Particles size of the sediments in rainy and dry season will compare and describe in the next part.

Study of changement between seasons versus grain size fraction in the 4 sampled sites is an evident of this study. Figure 4.5;4.6;4.7 and 4.8 summarises the result of particles size of every sediment samples in two seasons. Percentage of sand is rather high on all locations in dry season, which could be also due to anthropogenic influence. On rainy season, percentage of sand is some how decreased. Fine particles were found on the whole sediment samples in dry season but not in the rainy season. The fine-grained fractions (grain size <0.0063mm) content of more than 76% was found in quiet areas samples as CL and ND, mean while only 23% of very fine mud was found at estuaries zone sediment BT. In addition, the fraction of medium coarse sand (grain size 0.25-0.5mm) in sediments was strongly varies between sampling locations.

pH of sediment

Acidity is another main factor influencing the form of metal. Under low pH condition (<pH 5), metals such as Fe and Al can become soluble from most minerals.

However, some elements may become insoluble under low pH and redox conditions.

Acidic condition in the digestive tract of organisms can cause particulate pollutants become soluble as well as a consequence, matals are more available to the organism and lead to the occurrence of bioaccumulations. The pH values at the four sites ranged from 7.2-7.6 indicate that sediments were neutral medium, especially pH were higher in the dry season 7.5-7.6, which contrasted with those lower of the rainy season 7.2-7.4 (Figure 4.9)

pH of sediment samples

Distribution of organic carbon in sediment

Adsorption is the most reactive form of metal. Metals are also tending to form bonding with the organic materials, has very high specific area. As a consequence, it has high adsorption capacity. Therefore, concentration of metal may also be proportional to the amount of organic particulates. Generally there is inverse relationship between bulk density and organic carbon contents, bulk density increases is increased as organic carbon content are decreased. The bulk density and organic carbon content of the sediment samples remained nearly constant with an average of 0.60-0.99 (g/cm 3 ) with lowest bulk density was found 0.36 (g/cm 3 ) in ND coast sediment and highest value was 1.26 (g/cm 3 ) in CL lagoon area. The sediments were classified as lighter organic matter according to Avnimelech Y, 2001 classification. Significant variations in AVS in sediment samples were observed, with variation between 0,168 (TD) and 0,416 (ND), that justifies the low AVS levels in this coastal area according to the another region. The most significant AVS values were found in the ND sediments (0,416) and very distinct concentrations were obtained for the BT and CL (0,243 and 0,266). The highest bicarbonat content was obtained for the ND: 0,786mgdl/100g. The percentage of bicarbonates varies from about 2,72 to 4,97% and is in all samples lower than percentage of organic matter.

Considering the physicochemical factor and accessibility, sampling locations were first being classified by referring to the structures. According to the physicochemical characteristics, the sediments type were classified as silt clay to clayely silt with pH neutral, lighter organic matter, low AVS and differences of moisture content from low to high, depends of location of sampling. Classification of sediment samples is shown in table 4.6. The concentrations of trace metals were showed a variation between the metals and between sites. The maximum average concentration values for Cu, Cr and Cd (mg/kg DW)

were observed in BT estuary site. Site TD and CL showed similar concentrations of trace metals, where as mangrove site TD showed comparatively low concentrations due to poor sediment accumulation, and CL located in high energy system showed the least concentrations. In the present study, the elevated concentrations of Zn and Cu from in the all four sites could be attributed to the increase in natural weathering of soil materials and inputs from rapid industrial development and urbanization. In surface sediment, an increase in concentrations of chrome and copper at BT and CL sites showed exchange able fraction and gradual increase due to additional metal precipitation around there two boundaries.

It may be concluded that the increase in trace metals in the sediments mainly come from extensive fishing activity, public boat services, domestic sewage release from near by areas, and other industrial waste discharge. Concentrations of heavy metals in sediments from four collecting stations within the affected area are illustrated in Figures 4.13;4.14;4.15 and 4.16. When the analyses of a metal for seasonal, there rainy sediments contained higher concentrations than dry sediments of all most metal. Some significants that sediment in dry season contained higher concentrations of heavy metal as chromium at BT and CL sediment samples or arsenic in TD and copper in CL.

The average concentration of Zn was highest in both seasons compared with other metals. Furthermore, the highest concentrations of Zn were detected in rainy season (80.66mg/kg DW) and 64.67mg/kg DW in dry season. Nevertheless, the concentrations of arsenic were not detected in dry seasons (arsenic concentration in CL = 0.15 mg/kg DW).

The highest average concentration of Cu was in rainy compared by dry seasons (24.67 and 18.21 mg/kg DW, respectively). The concentrations of Cr were higher on rainy season than dry season, while concentrations of Cd were detected with an average of 1.85 mg/kg DW in rainy season but were lower of detected limit in dry season in ND sample. The elements that are essential for mollusk growth (copper, iron and zinc) are the most abundant elements measured in the sediment on the rainy basis.

In general, the area distribution of metals in the ND estuarine reflects the inputs from anthropogenic origins, with higher concentrations at locations close to the metal Zinc has a similar trend to copper with local differences: the highest concentrations in the surface sediment occur in the estuary zone of the BT (112 mg/kg DW) and the area of the ND (99.90 mg/kgDW). The influence of the Zn in TD and CL are low, however it is obvious in the TD (50.99 mg/kg DW) and in CL (59.60 mg/kg DW). Another difference is that the zinc levels in the CL show small signs of anthropogenic enrichment. The zinc contamination in the sediment of preindustrial (MOSTE, 1996) was very low found (6.6mg/kg DW).

Once the copper and zinc levels in the surficial sediments are established, the contamination state could be ascertained by comparison with typical concentrations of these two metals in the unpolluted sediments and/or their natural presence in the lithosphere. The measured values in the four areas are higher than average concentration in the preindustrial sediment and also exceed the range for unpolluted sediments. However, it is more appropriate to use local background equations to define the degree of contamination.

In the mouth of the estuary, copper and lead tend to precipitate, probably with iron oxides. Fe, Ni and Cr are found in varying concentrations along the bay of Nha trang. The chemical forms of these precipitations vary from one hydrological zone to the next. Nickel, most often associated with organic matter, can precipitate with manganese oxides and ferrohumic complexes in contact with salinated waters. These two metals are the most labile and ubiquitous. Concentrations of Zn, Pb, Cr and Cu reach their peak in the dry period, indicating the importance of the evaporation.

Sediment Quality Guidelines (SQGs)

As the mean to predict adverse effect derived from contaminated sediments, numerous sediment quality guidelines (SQGs) were developed by overseas researchers.

Among the most widely applied SQGs are as following, both SQGs produced by NOAA and New York Sediment Screening Criteria were chosen as the most appropriate values, which is being used as main reference in this study. Especial, cadmium concentration in sediment collected in both dry and rainy season had all values above the LEL value. Those concentrations are interpreted as a very high level which compare the values of cadmium were found in preindustrial sediments from near by port of Nha trang (Phuong et al., 2012) and in other northern coastal sediments.

Additionally, that may conclude that Cd concentrations are anthropogenically influenced and that they are about 10 times higher than in unpolluted predidustrial sediments, but also that there are many spots in the Pacific Sea with much higher concentrations.

From the assessments in this study, most metals have acceptable (<LEL) or moderate (≥ LEL and < SEL) biological effects, except Cd. Percentage of samples amongst range of SQGs for dry and rainy sampling is showed in Figure 4.17 and Figure 4.18

Interactions between metals

To study the inter-elemental associations, Pearson correlation coefficients of the trace metals were computed for both two seasons. One can observe a strong correlation between the amount of Fe and Al in both two season (Table 4. 10 and 4.11). This can be mainly attributed to synergic co-precipitation in the form of hydrated aluminum oxide or hydrated ferric oxides. In sediment, a significant correlation exists between Cr and Al, Fe, To study the interactions between metals and physicochemical characteristics of sediments, Pearson correlation coefficients of the trace metals were computed for four locations separately (Table 4. 12, 4.13, 4.14, 4.15 and 4.16). In all the sites studied, trace metals and their inter relationship showed strong correlation with Cr and Al, Fe and in significant correlation with organic matter and moisture content. Due to relatively low concentration of organic matter in sediments, it does not act as a carrier of metals, and therefore shows asynchronous distribution pattern with trace metals. Strong inter-element relationships suggest that one common process determines the sediment composition for all trace metals.

The matrix correlating the geophysical parameters (grain size, TOC, pH and moisture) and pollutant concentrations is presented in Table 4.12. A high correlation was observed between metal levels in the sediment and silt-clay content while a moderately negative correlation resulted between Cd in sediment and other metals in the sediment.

Decreases in TOC and pH in the dry and rainy season appear to be linked to a accumulation of metals in the sediments.

In all the time sampling, trace metals and their inter relationship showed strong correlation with Al and in significant correlation with OC as well as moisture contents. Rank-order correlation coefficients for metal ion interconnections are showed in Table 4.12. The strong positive correlation between all the metals in sediment collected from TD that these metals are carried by industrial discharges, fishing activities, and constant movement of fishing and commercial boats into the near by environment. In TD, the poor association of As with other metals suggests that Fe oxide may only be aminorhostphase for these metals in the mangrove environment. In TD, Cd showed no significant correlation with other metals possibly due to different processes like biological effects and external inputs operating in the mangrove and adjacent estuarine sediments.

The differences in correlation among the above metals reflect differential behaviors, which require further examination. Significant correlations of Cr, Cu and Zn with Fe and Zn in TD indicate the industrial contribution of these metals into the system. >0.063mm -0,6894 0,0749 0,2266 1 <0.063mm 0,5889 -0,0735 -0,2223 -0,9824 1

As -0,3075 -0,1367 -0,0414 0,0738 -0,0053 1 Cd -0,1199 -0,0364 0,0436 0,6853 -0,7644 -0,5395 1 Cr 0,1897 0,9389 0,9460 0,1294 -0,1633 -0,1450 0,1122 1 Cu 0,0799 0,3139 0,3957 0,2000 -0,2936 0,1399 0,2407 0,6037 1 Zn -0,0085 0,6096 0,6537 0,1478 -0,2020 0,1051 0,0481 0,8072 0,9004 1 Fe 0,3258 0,9047 0,8642 -0,0175 -0,0369 -0,4250 0,1635 0,9481 0,4847 0,6999 1 Al 0,2222 0,7324 0,7520 0,3396 -0,4225 -0,4026 0,5347 0,8665 0,6178 0,6945 0,8677 1 Ni 0,2339 0,9564 0,9247 -0,0729 0,0658 -0,1115 -0,1275 0,9612 0,4628 0,7435 0,9291 0,7366 1 Mo -0,3863 -0,5822 -0,5002 0,4597 -0,5030 -0,187 0,6250 -0,3435 0,3050 0,0500 -0,3309 0,0028 -0,5070 >0.063mm -0,9524 -0,5596 0,0895 1,0000 <0.063mm 0,8360 0,3175 -0,3527 -0,9635 1,0000

As -0,9515 -0,9379 -0,5081 0,8124 -0,6267 1,0000 Cd 0,7573 0,9990 0,8028 -0,5221 0,2749 -0,9215 1,0000 Cr 0,5937 0,9643 0,9150 -0,3200 0,0549 -0,8124 0,9751 1,0000 Cu 0,1822 0,7514 0,9993 0,1263 -0,3871 -0,4758 0,7801 0,8994 1,0000 Zn 0,1533 0,7317 0,9978 0,1554 -0,4140 -0,4498 0,7614 0,8862 0,9996 1,0000 Fe 0,9600 0,9275 0,4831 -0,8289 0,6490 -0,9996 0,9099 0,7953 0,4503 0,4238 1,0000 Al 0,8192 0,9984 0,7387 -0,6052 0,3702 -0,9559 0,9949 0,9479 0,7132 0,6923 0,9470 showed that, for the metals, concentration of metals in shell and tissue were significantly correlated. These correlations may be partially attributed to metabolic rate of each species. In general, concentrations of Al and Fe in mollusk samples from Nha Trang bay were higher in shell than in soft tissue, but for other heavy metals, concentration of metals in shell and tissue were significantly correlated.

These results highlight that there not necessity to analyse metals in shell when comparing trace metal concentrations amongst mollusk samples.

Contents of heavy metals in soft tissues

Eleven species of mollusks collected from four locations were analyzed for metal For comparisons of data between two seasons the concentration of Cu and Zn in mollusks G.coaxans collected form TD during rainy season was much higher than that found in dry seasons. It was found that concentration of Zn was higher than Cu and Cr in mollusks in two seasons.

In addition, concentrations of As, Cd and Cr in G.coaxans collected in ND were higher than that in TD at rainy season (6.10, 2.10 and 1.28 mg/kg DW in ND and 0.01, 0.83 and 1.22mg/kg DW in TD, respectively) but concentration of Cu and Zn in G.coaxans collected from TD were higher than that found in ND. The local distribution of Cu in tissue of A.antiquate at TD showed higher concentration than the other stations. This may be due to the anti-fouling paints. The seasonal concentration of accumulate Zn showed higher level at TD (59.04 and 37.85mg/kgDW ) during rainy and dry season, respectively, while the level recorded 42.76mg/kg DW during dry season at station CL. This may be due to the high amount of suspended organic matter coming from different industrial wastes that precipitate to the bottom and also due to the decrease uptake of zinc by phytoplankton at lower temperature. The present concentration of Cu in tissue of the S. regularis was higher than other sites. The maximum value of Cu (52.26mg/kg DW) was observed at BT in dry season probably due to sewage discharge and industrial effluents at the Nha trang city. This can be explained by the fact that their hydroxides are precipitated together in the bottom and the common geochemistry of both. 15.53 mg/kg DW. Among the five metals tested in all tissue of mollusk, zinc Zn concentration was the highest in all tissue of mollusks. This could be due to zinc is essential element which is important in the metabolic activities of mollusk. Copper concentration was the second elements found to be highest in the mollusk tissue. This could be explained due to the high concentration of this metal in the water body and the role of this metal as essential elements for aquatic organism.

Contents of m etals in L.unguis at TD rainy season

As

Contents of m etals in S. regularis at CL rainy season

As

Contents of m etals in T.literatus at ND dry season

As

Metal pollution index

Metal pollution index MPI values provides a representative picture of contaminant impacts on bivalves due to their ability to concentrate and reflect the environmental levels of trace metal contaminants in coastal ecosystems. Table 4.17 and Table 4 The highest MPI values observed at BT is found to be coincided with the high overall percentage contribution of the rivers Be to the flow rate in rainy season (5,87),

where as the lowest MPI valves observed at ND is found to be coincided with the low percentage contribution of the low flow rate. Results from this study demonstrated that As were detected at high concentration for comparison with WHO standard. The mean range concentrations of As from 1.7 to 9.89 mg/kg DW. This is due to different species have different mechanism for metal binding and regulation in their body. Lower concentration of As in the A.antiquate tissue might be due to its toxicity and this metals is non essential metal to this clam, but As concentration in K.hiantina and G.coaxans were found high exceeded the permissible limit WHO (5 mg/kg DW). They are similar to concentrations reported for the bivalve R.philippinarum collected in Jiaozhou Bay (Lui et al, 1983). They are also similar to the Cd concentrations in the S.plana (Cheggouret al, 2005), where the average ranges was between 0.2 and 1.6 for irregular samples collected from the Moroccan estuarines. The Cd concentrations in the recent study were however lower than some other bivalves including V.gigas collected form Gulf of California [START_REF] Ruelas | Heavy-metal accumulation in the hydrothermal vent clam Vesicomya gigas from Guaymas basin, Gulf of California[END_REF]. The elevated Cd concentrations at two species A.antiquate and S.regularis in rainy season are very high compared with values reported for P.viridis from the Nha Phu bay, Khanh Hoa province where average Cd concentrations were between 0.03 and 0.31 µg/g (Ha D.et al., 2007).

Cd 0 2 4 K . h i a n t i n a L . A n a t i n a L . U n g u is G . C o a x a n s A . a n t i q u a t a S .
Cadmium contents to different species have a big different for metal binding.

Higher concentration of Cd was foud in the mollusk tissue of A.antiquate in both dry and rainy season. Their cadmium concentrations in were found exceeded the FDA permissible limit (3.95 and 4.84 mg/kg DW). However, concentrations of Chromium were still below the permissible limit when compared with FDA, 2001 (13mg/kg DW) exept Cr concentration found in L.unguis, which concentration were exceed the permissible limit (16.46 mg/kg DW) For chromium, which is considered a non-essential trace metal for bivalves, the tissue concentrations in the mollusks were lower values over rainy. These values are similar to those reported for mussels from Chile and pearl oysters from the Arabian Gulf where average values were below 3.8 mg/kg DW (Al-Sayedet al., 1994); Manley et al., 1996). From this study the concentration of copper was relatively lower when compared FDA (permissible limit 100 mg/kg DW) but Cu concentrations in almost mollusks are also most toxic metals when exceeded the certain limit of WHO (permissible limit 10mg/kgDW).

There have been fewer studies reporting copper concentrations in bivalve tissues (Table 1.2) but the average value of 15 mg/kg DW determined in the present study for the Nha Trang Bay mollusks is similar to the value reported for S.plana from Moroccan estuarines [START_REF] Cheggour | Metal concentrations in sediments and clams in four Moroccan estuaries[END_REF]. Similarly, Zn concentrations in the Nha Trang bay mollusks were comparable to those reported for T.mactroidea collected from the Edo Miranda Venezuela [START_REF] Labrecque | The concentrations of selected trace elements in clams Trivela mactroidea along the Venezuelan coast in the state of Miranda[END_REF] from sites with low levels of trace metal contamination. This figure showed that Zn were concentrated at low concentration when compared WHO limit, then Zn concentration in G.virens was found exceeded the permissible limit WHO (100 mg/kg DW).

Results from this study demonstrated that Cr and Zn were detected at low concentration. This is due to different species have different mechanism for metal binding and regulation in their body. The low fluctuation level of Cr and Zn in the tissues of mollusks clearly showed that they need of same level of these metals to maintain the normal bodies function of marine organisms or might be due to its toxicity and those metals are non essential metal to mollusks.

On the other side, these observed concentrations of As, Cd and Cu in the samples were above permissible level WHO, but from this study the concentration of As and Cu were relatively lower when compared FDA (permissible limit 86 and 100 mg/kg DW respectively) but it's concentration in all mollusks was found exceeded the permissible limit WHO (5 and 10 mg/kg DW respectively). In conclusion, metals as As, Cu and Cd are also most toxic metals when exceeded the certain limit where they habited may pose hazard to human in terms of health risk.

Based on those results, it showed that the magnitude of the heavy metal accumulation in mollusks tissues depend on the type of the heavy metals, the species of the mollusks, seasonal factors and also showed that mollusks living in the area accumulate heavy metals in their body tissues. Toxic heavy metals are dangerous since heavy metal tend to accumulate in the tissue of mollusk and could be transferred to human through the consumption of these mollusks.

Heavy metal concentration and biological parameters

Biometric parameters

Moisture of mollusks tissues

Tissue moisture was measured in all of the mollusks at the time of metal analysis as show in Table 4.19. There is a significant difference in moisture values between mollusks.

The mean moisture concentration for mollusks was 86.24 ± 1.22 % for 10 samples with a range of moisture concentrations of 81.99 to 90.49 %.

Body size

In order to minimize the influence of the body weight of animals on metal concentrations, comparison was made between sites with animals where no significant differences in weight were registered. CI values obtained from seven mollusks of four sites is showed in Table 4.20. Biometrics, which consisted of determining the length of the organisms and their fresh weight, were determined individually as Condition Index = 90 (normal); > 120 (high); < 70 -80 (low) according to [START_REF] Mann | A comparison of morphometric, biochemi-cal and physiological indexes of condition in marine bivalve molluscs[END_REF], Hetzel et al., 1983[START_REF] Lucas | The use of physiological condition indexes in marine bivalve aquaculture[END_REF]. Table 4.20 shows the average values of condition index. The mean mollusk condition index ranged from 14.38 (min value) to 116.67 (max value) and was significant variation among the species. The remaining biometric parameters were similar during all sample collections, and their weights and indexes were acceptable from the allometric point of view.

Metals determination and condition index

The matrix correlating the biometric parameters (length, weight, and condition index) and pollutant concentrations is presented in Table 4.21. Metal concentrations in tissues also depicted statistically significant correlations, from moderate to high, related to the condition index.

To investigate the relationship between trace metal concentrations and CI, the results from individual mollusks were combined for all sites and sampling times. The mean mollusk condition index ( For Cr, Cu, Zn, Fe and Al, tissue concentration was dependent of As concentration.

For Cd, the concentration appeared to decline with increasing As tissue concentration.

Condition index was negative correlated with all metal concentrations in the clam, only correlated well with Cd concentrations in the G.virens (r=0.446). The clam may become the best indicator for bioavailability than others mussel for which the CI was independent of all of metal concentrations (negative correlation). Cu and Fe metals in L.anatina showed highly correlations with condition index (r=0.795 and 0.624). The clam may also be a better indicator for bioavailability than others mussel for As which the CI was independent of As concentration. For the mussel L.unguis, there was a significant effect of CI on the trace metal concentrations for Cd and Fe. However in contrast, tissue arsenic and chrome concentrations were negative dependent of body size for mussels sampled in this study.

The mussel may also be a indicator for bioavailability than others mussel for which the CI was independent of As and Cr concentration. Mean tissue concentrations for Fe, Zn and Cu were positively correlated (Table 4.25) as were As and Cr concentrations. For these trace metals there was negative relationship between tissue concentration and the CI with contamination. Cd concentrations were negatively correlated with Cu and Zn. Condition index also correlated negatively with Cu, Zn and Fe (r=-0.866, -0.731 and -0.681 respectively). The condition index also correlated low with Al concentrations in the bivalve (r=0.116). The clam may also be a better indicator for bioavailability for which the CI was independent of all trace metal concentration. (Except Cd) For the mussel A.antiquate collected from 4 sites, there was a significant effect of CI on the trace metal concentrations for As and Zn. However in contrast, tissue chromium concentrations were negative dependent of body size for mussels sampled at this site. Cu and Zn was high correlated with Fe and Al concentrations in the bivalve (r=0.822-0.955).

The clam may also be a better indicator for bioavailability for Cd and Cu for which the CI was independent of Cd and Cu concentration. between As and Cd in tissue clam (-0.984). The clam may also be a indicator for As and Cr pollutions for which the CI was independent of As and Cr concentration.

There was negative correlated with As for all mollusk species, but not in A.antiquate (Coefficient = 0.996) the condition index appeared to decline within creasing As tissue concentration. For Cd, tissue concentration was highly positive dependent of CI.

For Cu, tissue concentration was not significant depend on the CI, except A.antiquate and K.hiantian. Coefficients of correlation were 0,795 and 0,775 respectively. For Cr, tissue concentration was independent of condition. For Al and Fe, where the range of tissue values examined was some what limited because of missing condition values for samples with highest Al and Fe concentrations. There was no clear significant effectt of any of these trace metal concentrations on mussel CI (regression analysis).

In order to establish this relation for 7 mollusks from the Nha Trang bay, we have considered all samples collected during the rainy and dry season.

Equation parameters for these 7 species were reported in Table 4.22 to 4.27. These results showed that, for the 7 metals, concentration and body weight were significantly correlated. The negative values of CI indicated that the higher metal concentrations were in small individuals. There negative correlations, more commonly observed than other correlations in mollusks, may be partially attributed to greater metabolic rate per biomass of smaller individuals or by a slight adsorption of metal by larger individuals (Blueweiss et al.,1978).

These results highlight the necessity to adjust for weight differences when comparing trace metal concentrations amongst mollusk samples.

EVALUATION OF BIOACCUMULATION

The biota-sediment accumulation factor (BSAF) is an index of the ability of the species to accumulate a particular metal with respect to its concentration in the sediment. It was calculated as the ratio between the metal concentration in the organism and that in the sediment [START_REF] Usero | Heavy metal concentrations in molluscs from the Atlantic coast of southern Spain[END_REF]

Comparison of BSAF in soft tissue and shell

Metal concentrations in mollusks varied along the 7 species collected first time on dry season (March 2012) in Nha Trang were calculated for BSAF in shell (BSAF-S) and tissues (BSAF-T) is showed in Table 4.28 The shell and tissue showed similar pattern in the accumulation of heavy metals analyzed and the tissue always had higher concentration than the shell. There was clear pattern of lowest bioaccumulation factor among the all metals and analyzed two body parts (shell and tissue) of the K.hiantina, A.antiquate and L.anatine. However, within the other species, Cd and Zn were recorded to have highest concentration in two body parts (shell and soft tissue).

The mollusks L.anatine, L.unguis, G.virens, C.rivularis and G.coaxans have high potential factor BSAF in tissue for As (BSAF > 2) but not for shell, exhibiting that this element possibly essential in its tissue. Comparatively, oyster C.rivularis recorded higher factor value for Cd, As, Cr, Cu and Zn in tissue and low factor value for these metals in shell. Such a condition indicated that all species are capable of accumulating that metal Cd in relatively larger quantity, which indeed shows that it is potential bioindicator for Cd pollution in sediement. On the other hand, it has low potential factor for the accumulation of Cr and Pb in both shell and tissue.

By comparing the accumulation of heavy metals in two body part of mollusks, this study found that the concentrations of metals were higher in tissue of mollusks and there was significant difference in metals accumulation between shell and tissue. Thus, the data from the study showed that mollusks living in the effluent from the study area high accumulate heavy metals in their body tissue.

Bioaccumulation of heavy metal in the mollusk tissues

Metal concentrations of Al and Fe in the tissue of the four sites were similar and did not vary appreciably and presented very lower compared to concentrations of these metals in the sediments. Concentrations of Al and Fe in the sediments were 20-100 times higher than the concentration in the mollusks.

By comparing BSAF values, we can compare the ability of those mollusks in taking up metal from sediment. The highest or lowest BSAF values also varied for each different metals ans species during 2012-2013 was showed in Table 4.29 Table 4.29 Values of bioaccumulation factor of heavy metal in mollusk species In general comparision the biocumulative factor of heavy metals from sediments within mollusks found in the Nha Trang bay, the BSAF indicated a low potential bioaccumulation for Pb, Mo and Ni (BSAF<1) and a high potential of accumulation for As, Cd, Cu, Cr and Zn. Arsenic had the high BSAF ratio for 5 species as: Crassostrea rivularis, Glauconoma virens, Katelysia hiantina, Lingula unguis and Laternula anatine.

Copper had the highest BSAF ratio for 5 mollusk species Anadara antiquate, Crassostrea rivularis, Glauconoma virens, Gelonia coaxans and Katelysia hiantina, while

Cd had the highest ratio for the remaining two species Anadara antiquata and Crassostrea rivularis. The BSAF values for Pb obtained by all of the mollusks were below 1.0, indicating higher content of this metal were found in the sediments compared to the tissues of the mollusks. Such a condition possibly reflected that the mollusk were rather selective in accumulating Pb metal into their tissues. Comparatively, an interesting observation indicated that mollusk species from the study area showed highest BSAF for As, Cd and Zn.

The trends in the BSAF for metals in seven mollusks species were in the descending order of Cu > Zn > As > Cd > Cr. The trend in BSAF for metal in G.coaxans were in the decending order Zn > Cu > As > Cd > Cr. Finally, if we compare the seven species studied, we can conclude that G.virens by far had the highest BSAF for As followed by K.hiantina. It showed that the organisms living in the area were high accumulated heavy metals in their body tissues.

Based on the bioaccumulation factor (BSAF), all mollusks showed different degrees of the accumulation of heavy metal in their tissues which is depending upon the type of heavy metals and the species of the mollusks. G.coaxans recorded high factor value of Cr, Cu and Ni. This indicates that G.coaxans is capable in accumulating Cr, Cu and Ni in relatively great quantity. Comparatively, C.rivularis, G.virens and K.hiantina have high potential factor for As, Cu and Zn, demonstrated that this element is probably essential element to they body. Therefore, these species also have the potential to be us as bioindicator for metals in aquatic environment. The study showed that the accumulation of heavy metals were different between species of mollusk. In general, higher metal concentration bioaccumulation was found in the samples from the vicinity of industrial complexes and urbanized area which lied between TD and CL.

CHOISE K.HIANTINA AS BIOINDICATEUR

The purpose of this study was to obtain information on the concentration of trace metals in mollusks from coastal area of Khanh Hoa Province and choising mollusk specie as a biomonitor for the area. The results reported here will provide valuable information and assessment the potential of using the K.hiantina as a biomonitors of heavy metal pollution. Another that the mineral composition of K.hiantina clam was to study their information on microstructure is one of the most significant relative datasets for clarification of evolutionary trends of shell structure. Thus, the findings may suggest the possibility of using the K.hiantina as an alternative biomaterial for bone substitute in managing bone defects.

Seasonal and locational variation

Although comparison between different stations is open to criticism, the mean concentrations of all studied metals in K.hiantina were of the same range with exception for the extreme values. For example, concentrations of heavy metals in K.hiantina from BT, in general, were significantly higher than that in samples from others locations as TD, ND and CL (as seen in Figure 4.41 to Figure 4.47). Only Cr and Cu concentrations in tissue samples from CL and ND were significantly higher than samples from BT.

For Al as well as Fe, although their concentrations were also higher in samples from BT, statistically they were not significantly different. Concentrations of metals in soft tissue of samples from BT were higher than that of samples from, but only Cd and Cu were statistically significant different. The highest mean value of Cr (11.58 mg/kg DW) in CL was higher 15.44 times than the lowest mean value (0.75 mg/kg DW) in BT, while other values, which varied between 3-15 insignificant ranges. Moreover, Cd contents were higher about 2.08-5.47 times in rainy season than in dry season.

The high concentrations of Zn in BT and other metals in stations from Nha Trang bay were assumed to be related to the human activities along the coastal waters of these stations. Two large rivers are draining their load to these waters, one of which is passing the densely populated area in the Nha trang city center. However, the industrial and anthropogenic activities in Nha Trang are still much less when compared to those in the southern coast of Viet Nam

Higher concentrations of metals in samples from BT, in general, might be related to the more anthropogenic activities in this area. MOSTE, 2002 reported that the southern coast of Viet Nam is more developed than the north coast. Many activities such as petrochemical plants, land reclamation, urbanization, shipping and other industrial activities and some major ports are concentrated in this location.

As accumulation in tissue of K.hiantina showed for those metals: the higher levels were observed in rainy season. For example, in from the 4 site, the mean values were 0.87 ± 0.46, 27.72 ± 16.78 and 64.53 ± 20.41 mg/kgDW respectively for Cd, Cu and Zn in the rainy season against 0,24 ± 0. The intense correlation of Zn and Cu shell/weight indices in K.hiantina species with their corresponding concentrations in the weakly bound fraction of sediment confirms the impact of sediment to the specific organism living within the sediment, under these a bottom surface, since in its filter-feeding mode it is exposed to the dissolved metals of its burrow water, which is a mixture of hypoxic/anoxic pore water and overlying (oxygenated)

water (Griscom et al., 2002).

Metal/shell-weight indices were used to characterize each site and season depending on the metal bioavailability. Metal bioavailability was highest in TD in both seasons than at any other site.

In dry season, TD showed higher metal bioavailability than other sites with values over those recorded in the rainy season. In rainy period, a general decline in metal bioavailability was observed, the highest bioavailability being restricted to the ND location. In this season, the metal bioavailability found in TD was close to that exhibited by ND. Afterwards, metal bioavailability decreased at all sampling points, mostly in BT then CL. As, Cr, Al bioavailability were higher in dry than in rainy season and Cd, Cu, Zn, Fe bioavailability occasionally increased in rainy season.

In the present field investigation, shell dry weights did not change significantly over the year for clams K.hiantina measuring 2.5 to 3.5 cm in shell length. At a polluted site, such as CL, the Cd concentration, for instance, changed seasonally but, conversely, the Cd/shell-weight index did not exhibit seasonal fluctuations in relation to the intensity of the industrial activity, indicating fluctuations in metal bioavailability. At a non polluted site, such as TD, both Cd concentration and Cd/shell-weight index changed seasonally as could be expected by absence of local inputs. Consequently, the present field study supports the proposition that metal/shell-weight indices provide a method to detect differences and changes in the bioavailability of cumulative metals (Fischer, 1988). Since the term metal/shell-weight index (Fischer, 1983[START_REF] Fischer | Cadmium body burden/shell weight of mussels: aprecise index for environmental monitoring[END_REF](Fischer, , 1986(Fischer, , 1988) might be misinterpreted as a measure of shell metal content, we suggest using Fischer's bioavailability index as a routine name for the metal burden in flesh related to shell weight in sentinel mussels. Such an index was applied in this study to investigate the metal bioavailability in the Nha trang bay.

Different metal bioavailabilities have been found, however, at different sites in relation to the influence of the waste waters out coming from the highly industrialized Nha Phu. Thus, metal bioavailability was generally higher on the north part of the bay than on the southern part and the outer sampling sites. The north receives the direct influence of the industrialized Nha Phu whereas the south is mainly influenced by water coming from the close sea, as a resul of wind and water currents. Interestingly, according to the metal concentration values a high metal bioavailability was envisaged in TD where flesh weight was heavily depressed due to the continuous stress provoked by very strong wave turbulence. When metal/shell-weight indices were applied, however, TD resulted to be a non polluted site, as expected from the scarce influence of industrial inputs. Finally, although the metal/shell-weight index appears to be kept unchanged by seasonal factors other than metal bioavailability, the present results indicate the existence of seasonal trends in metal/shell-weight indices. Thus, these fluctuations might be attributed to different metal inputs at different times of the year.

Bioaccumulation of heavy metal in clam K.hiantina

Metal concentrations of Al and Fe in the tissue of the four sites were similar and did not vary appreciably and presented very lower compared to concentrations of these metals in the sediments. Concentrations of Al and Fe in the sediments were 20-100 times higher than the concentration in the mollusks. By comparing BSAF, we can compare the ability of clam K.hiantina in taking up metal from sediment. Table 4.31 shows the calculated BSAF values in the different sampling sites. ,1993), it can be calculated that a 60kg human would reach there commended level of intake with a daily consumption of about 1kg of mussel flesh from this area. Additional sources of metals are thus required to cause adverse effects on human health. In addition, other factors should be considered.

The need for regular monitoring of trace metal pollution is emphasized to acquire data for environmental protection and conservation measures. Because shellfish consumption rates can vary greatly between specific sections of acommunity, the values reported here should only be used for screening purposes. Clearly, further assessment is necessary before more definitive conclusions can be drawn.

The target hazard quotient THQ

The target hazard quotient THQ has been recognized as a useful parameter for evaluation of risk associated with the consumption of metal contaminated food.

Metal contents in sediments

For remaining heavy metals in sediments in both two seasons, metal concentrations in the sediments ranked in the following order: Aluminium> iron > zinc > copper > chromium > asennic > cadmium. The most of metals concentrations were acceptable (<LEL) or moderate (≥ LEL and < SEL) biological effects, except Cd.

Correlation of metal concentration and environmental factors

A high correlation was observed between metal levels in the sediment and silt-clay content while a moderately negative correlation resulted between Cd in sediment and other metals in the sediment. Decreases in TOC and pH in the dry and rainy season appear to be linked to an accumulation of metals in the sediments.

One can observe a strong correlation between the amount of Fe and Al in both two seasons. On the other side, a significant correlation exists between Cr and Al, Fe, Zn and Ni in both dry and rainy season and between Cu-Zn and Fe, Al, Ni in dry season.

Bioaccumulation of heavy metals in mollusks

Metal contents in mollusks

In general, concentrations of Al and Fe in mollusk samples from Nha Trang bay were higher in shell than in soft tissue, but for other heavy metals, concentration of metals in shell and tissue were significantly correlated.

Among the five heavy metals tested in all tissue of mollusk, Zn concentration was the highest in all tissue of mollusks and Cu concentration was the second elements found to be highest in the mollusk tissue. But, these observed contents of Cu and Zn were not excceded permissible level WHO. These concentrations of As, Cd and Cr were detected at low concentration, but As, Cd and Cr in the samples were above permissible level WHO, where they habited may pose hazard to human in terms of health risk.

Bioaccumulation of heavy metals

The study also showed that the magnitude of the heavy metal accumulation in mollusks tissues depend on the type of the heavy metals, the species of the mollusks, seasonal factors.

There was well obvious overall pattern in the seasonal dependence of the metal concentrations measured for the mollusks in this study although the concentrations of some metals were generally high in rainy season, which is the period of somatic growth and gametogenesis. The majority of this research supports accelerated accumulation during the rainy season.

Physicochemical factors such as pH, TOC, together with the inorganic components of the sediments, can affect the uptake of metals and the subsequent accumulation of metals in the tissues. While in most mollusks the increased CI reflects directly the accumulation of metals, in some clams increases can occur without apparent weight gain.

This was also found in the present study where the variation in mollusks CI amongst species was greater than it was between sampling locations. These results highlight the necessity to adjust for weight differences when comparing trace metal concentrations amongst mollusk samples.

Mollusk species have served as good bioindicator organisms. Thus, the data showed that mollusks living in the effluent from the study area high accumulate heavy metals in their body tissue. The mollusks L.anatine, L.unguis, G.virens, K.hiantina and G.coaxans have high potential factor in term of metal BSAF in their tissues. They have served as good bioindicator organisms,in which K.hiantina might be the best indicator of metal pollution. Trace metal concentrations in this clam may be low compared with some other species, but it is high presence, abudance and wide geographical distribution also in potentially contaminated sites.

Ecological risks

A moderate trace level consumption of bivalve from the Nha Trang bay is not harmful to humans for Cd, Cu and Zn, but due to high level accumulations of metals in soft tissues, heavy consumption of mollusks from the Nha Trang bay may pose a health risk to consumers.

LIMITATIONS AND SUGGESTIONS

However this study also consider for a local programs. There was disturbance of the monitoring area (i.e., dredging, harvesting, recreational activities), as this was suspend contaminated material and possibly desorbs the sediment bound metals, as well as compromise the diversity of the benthic Sediment analysis should include salinity, DO, pyrite FeSO 4 , LOI, total sulfite volatil, as well as determination of CaCO 3 and Mn in sediment. These physicochemical parameters are important for explan of bioavailability and uptake.

The number or organisms should be sufficient so that a level of statistical significance can be assigned to the data. At least 10 surviving mollusks or better still, 20-30 bivalves per monitoring site should be sufficient.

These results highlight that there not necessity to analyse metals in shell when comparing trace metal concentrations amongst mollusk samples.The differences in correlation among the above metals reflect differential behaviors, which require further study. Although seasonal fluctuations in metals accumulation by mollusks has been reported, tissue sampling and analysis should occur four times per year.

Because shellfish consumption rates can vary greatly between specific sections of a community, the value ecological risks reported here should only be used for screening purposes. Clearly, further assessment is necessary before more definitive conclusions can be drawn.

FURTHER RESEARCH

When undertaking monitoring studies using mollusks, it is important to account for differences in biomonitoring considerations. The need for regular monitoring of bioaccumulation of trace metal pollution is emphasized to acquire data for environmental protection and conservation measures in Nha Trang bay in particular or .coastal of Viet nam in general.

Clam K.hiantina as good bioindicators, that found that clams had an ability to survive very well under natural conditions. A field experiment of the clam will be transplant, either suspended in seawater or in sand over the 6 month duration of the study.

I. INTRODUCTION

Heavy metals like Pb, Cd, Cu, As, Hg and Zn, coming from natural and/or anthropogenic sources, are significant input components of the sediment in marine zones, which are being important areas for concentration of contamination. The most significant sources of pollution include oil from ships, sewage and other domestic wastes, and industrial effluents [4]. Since 1996, in Viet Nam, the increase of metal concentration in the sediment had been observed in Quang Ninh [1], Hai Phong [START_REF] Thị | Accumulation of lead and copper in some bivalvia mollusks species from coastal water of Da Nang. Tạp chí Khoa học và Công nghệ, Đại học Đà nẵng[END_REF], Da Nang [3] and Khanh Hoa [3], and receives attention from local managers.

The objective of this study was to evaluate the ranges and state of contamination by 6 metals in the period of time going from 1996 to 2011 in Nha Trang bay, Khanh Hoa province by making comparison with standard criteria for sediment. Concentrations of these heavy metals for two different seasons have been monitored: the dry and the rainy seasons, and calculations of geoaccumulation using the Igeo index, which compare the metal contaminants concentrations for the pre-industrial period (1996) with values measured for sediments collected from 1998 to 2011, supposed to be affected by human activities.

II. MATERIAL AND METHODS

Survey area

The station of monitoring with latitude 12 0 12'45"N and longitude 109 0 13'12"E, is located in Nha trang bay -Central southern Vietnam, which encompasses some 507km 2 of offshore waters and island coasts. The Nha Trang bay is situated immediately adjacent to the coastal city of Nha Trang and its port at Cau Da. Two rivers, the Cai River in the north and the Be River in the south, flow into Nha Trang Bay, with the potential to influence water quality in the Bay. There are 19 islands included in the Nha Trang Bay, situated from ~ 1km to ~ 15 km offshore, with an average depth of 5 -20m, providing a wide range of coastal and marine habitat types in relation to prevailing oceanographic conditions and gradients in mainland -oceanic influences. The river catchments and riparian vegetation have been extensively modified for agriculture and aquaculture in recent decades.

The surface sediment samples were collected every year in the Nha Trang bay from 1996 to 2011 during both dry and rainy seasons, by Institute of Oceanography of Nha trang.

Field sampling

These sediments were taken using an Ekman grab from a boat. Samples of the top surface centimeter were collected with a polyethylene spool and placed in a plastic bag, which were stored in an ice box at 4 0 C, to be transported in the laboratory of Faculty of Environment (HCMUS) Ho Chi Minh City. In the laboratory, some of sediment characteristics were determined as pH, moisture content, organic matter and particlessize.

Sediment samples were dried at room temperature, and then were sieved through a nylon mesh to obtain 4 different particle-size fractions, starting with particles smaller than 0,2mm in diameter for determination of heavy metal concentration. For each particle-size fraction, 1.0 gram was taken for heavy metal analysis.

Analysis

Determination of sediment characteristics

Moisture was determined in two steps: sediments were dried in air at room temperature then oven drying at 105 o C in 16h (Method ASTM 2974-87). Moisture contents were measured by weight loss.

Organic matter was determined by ash dried sediments in the furnace to 750 o C and hold until the samples were completely ashed. (Method ASTM, 2974) For the bulk density determination, 10ml of post were added in sediment samples and allowed to dry to a constant weight at 70 o C. Bulk density was calculated as dry weight divided by wet volume.

For grain size analysis, a standard sieving method has been used for particles bigger than 0.1mm, and by pipetting method for particles smaller than 0.1mm.

Heavy metal analysis

The samples were digested, following the guideline EPA 3050 for digestion sediments using a hot plate. About 1,0 gram of dried and sieved shells were transferred in 250 ml pyrex beaker and added with 4 ml of acid (1+1) HNO 3 and 10 ml of (1+4) HCl for decomposition of organic matter. Then the beakers were put on a hot place for reflux extraction at 95 o C, for 30min. Then, the samples were cooled and the extract quantitatively transferred in a 100 ml volumetric flask, then final solutions diluted and mixed to volume to 100 mL with ultra-poor water. The obtained solution was then centrifuged and the concentration of heavy metals measured using AAS -Atomic Absorption Spectrophotometry.

III. RESULTS AND DISCUSSION

Sediment characteristics

The results have showed that the pH value in the station corresponding to the samples varies from 7.15 to 7.32. The sediment sample was then in a neutral environment. The moisture content of sediment sample was about 7.4%. The total organic matter of sediments is equal to 17.97%, and the bulk density gave an average value of 0.26g/cm 3 . Fine particles were found on the sediment sample. Meanwhile the average fraction of very fine sand and mud (grain size 0.01-0.125mm) in sediment was 26.77%, compared with an amount of about 73.02-74.71% of silty clay. The distribution of the coastal sand was not found in this zone.

The grain size distributions of sample showed a highest muddy fractions, while there is only 0,66% medium coarse sand (fraction whose practice-size >0.1).

Heavy metal concentrations

The concentrations of contaminated heavy metal in period 1996-2011 are summarized and compared with the New York guidelines criteria [6] for the effect of sediment on benthic organisms. Assessment of quality of sediment was focused on the LEL (Lowest effect level) and SEL (Severe effect level) as proposed by the Department of Environmental Conservation of New York, 1999. Definition of LEL and SEL [6] is:

Metal values<LEL: the effect in the sediment is considered to be acceptable LEL< Metal values<SEL: contaminated, moderate impact to benthic life Metal value >SEL: contaminated and significant harm to benthic aquatic life Table 1 shows the based concentration of metal in sediment samples from 1996 to 2011 and gives the LEL and SEL for each metal. In general, the results demonstrated that concentrations of heavy metal in Nha Trang were lower than LEL*. That means that most of the elements have no biological effect, except Hg. Hg is the most critical element that can have an effect on the organism living in sediment, as 100% of the collected sediments from 1998 to 2011 were contaminated with values exceeded LEL. In general, the concentrations of heavy metal in dry season were higher than in rainy season for the ten years.

Classification of contamination level

The geoaccumulation Igeo index [2] was used to calculate the metal contamination level for the period 1996-2011. Based on the classification of metal pollution proposed by the calculated Igeo factor, both Cu and As were in the range below uncontaminated or moderate contaminated level, when Zn was classified as moderately contaminated level. Only Pb shows a moderately to strongly contaminated level in this zone.

For Cd and Hg, low concentrations in the samples do not allow calculations.

IV. CONCLUSION

With regard to an overall review of contamination of heavy metal in Nha Trang bay, the heavy metal concentration was lower than LEL, meaning that most of the measured elements have no biological effect, except Hg. The concentrations of heavy metal in the dry season were higher than in the rainy season during the all ten years period of time.

The impact of contamination by heavy metal in Nha Trang bay in period from 1996-2011 was evaluated using Geoaccumalation Igeo indices. The Igeo suggests that fine fraction of sediment in Nha Trang bay goes from moderately to strongly contamination with respect to the analysis of 4 heavy metals (Zn, As, Cu and Pb). The Igeo index was not able to applied to the Hg and Cd concentrations for the assessment of their pollution impact.

INTRODUCTION

In the recent year, contamination of heavy metals has been negative impact to the natural environment, especially to marine ecosystems in coastal areas. The effects of heavy metal contamination can be severe and are a significant concern in the Arctic [1]. Once heavy metals are in the sediment, they can be easily be uptaken by organisms and accumulate in living tissues of bivalves as well as mussels community. The bioaccumulation of heavy metals by marine mollusks may reach many orders of magnitude above background concentration of certain locality [2]. Thus, collecting and analyzing the concentrations of heavy metals may help us to understand the bioaccumulation capacity for heavy metals by different species of mollusks, which habitats locates along the coastal of Khanh Hoa province. This research conducted the heavy metal analysis in the sediment and in the bivalves from the same location could be used to estimate bioaccumulation capacity in marine ecosystem and will help to propose the best bivalve indicators for an environmental biomonitoring.

MATERIAL AND METHODS

Research area

Nha Trang bay locates in Khanh Hoa province -central-southern Viet Nam, which is an important ecological zone and providing a wide range of coastal and marine ecosystems in relation to prevailing oceanographic conditions and gradients in mainland -oceanic influence. The sites of collection samples from the different geographic locations and substrate types (mainland-oceanic gradient). Sediments and mollusks samples were collected from eight sites in March and April 2012. The locations of sampling sites are showing in Table 1.

Sample collection and preparation

Mollusks collection

About 5-10 numbers of different sizes organisms from each sampled site were picked by hand and then put in Ice box to be transported in the laboratory. Different tissues are separated by knif and dried at 60 o C during two days to constant weight. These dried tissues were then stored in polyethylene bag and sent to Ecomer's laboratory (UNS-France) for the determination of heavy metals concentrations.

Sediment collection

These sediments were taken using an Ekman grab from a boat. Samples of the top surface centimeter were collected with a polyethylene spool and placed in a plastic bag, which were stored in an ice box at 4 0 C and sent to the laboratory. Sediment samples were dried at room temperature, and then were sieved through a nylon mesh to obtain particles smaller than 0,2mm in diameter for determination of heavy metal concentration. For each sediment sample, 1.0 gram was taken for heavy metal analysis.

Sample digestion and analysis

Mollusks digestion

Digestion method of organically based matrices was based on the guideline EPA 3052. Each biological sample (tissues or shells) was dried, pulverized and sieved for digestion and determination of metals concentrations. A weight of about 0, 5 gram of shell or 0,1gram of tissue was then transferred in a 100 ml Pyrex beaker and added with 4 ml of acid HNO 3 and 10 ml of HCl. Then the beakers were placed on a hot place for reflux extraction at 95 o C, for 30min. The digested samples were diluted and filled up with de-ionized water to a total volume of 50ml before metal analyzing by ICP-OES (Perkin Elmer). The accuracy of the analytical procedures employed for the analysis of metals in organic samples was checked using the Fish tissue reference IAEA MA-B-3/TM as certified reference material, and showed good agreement with the certified values.

Sediment digestion

The samples were digested, according to the guideline from EPA 3050 for digestion of sediments using a hot plate. About 1,0 gram of dried and sieved sediment were transferred in 100 ml Pyrex beaker and added with mixed of concentrated acid HNO 3 and HCl. Then the samples were put on a hot place for reflux extraction at 2 etapes: 95 o C, for 30min then 2h at 140 o C. The samples were cooled and mixed to volume to 100 mL with ultra-pure water. The obtained solution was then centrifuged and the concentrations of heavy metals were measured using ICP-OES (Perkin Elmer). The accuracy of the analytical procedures employed for the analysis of metals in sediment samples was checked using the marine sediment IAEA-433 as certified reference material.

Method of analyzing data

A mollusks capacity to accumulate metals from living medium into its tissue can be estimated using BSAF factor. BSAF were calculated for the selected metal in the mollusks studied according to formula BSAF = Cx/Cs where Cx and Cs were the metal concentrations in the organisms and in associated sediments [3].

By comparing BSAF, we can compare the ability of those mollusks in taking up metal from sediment. The categories of BSAF are presented as: High potential of accumulation if BSAF > 2, Moderate potential if 1<BSAF<2 and Low potential if BSAF <1.

RESULTS AND DISCUSSION

Heavy metal distribution in mollusk tissues

The overall heavy metal concentrations in the 7 analyzed species are included in Figure 1 and presented in µg/gram of dry weight. For most of the metals, when mean values were calculated, values were close to the maximum permissible level FDA (USA, 2001), except for Pb. It indicated that the coastal has been polluted by heavy metal which eventually leads to bioaccumulation of that pollutant in food chain of the marine environment. On the other side, the observed concentration of As, Cd, Cr and Cu in the samples were within permissible level, where the bivalves are safe for human consumption and where the ecosystems where they habited do not pose any hazard to human in terms of health risk. The low fluctuation level of Cd, Cu and Cr in the tissues of mollusks clearly showed that they need of same level of these metals to maintain the normal bodies function of marine organisms.

Based on there results, it showed that the magnitude of the heavy metal accumulation in mollusks tissues depends on the type of the heavy metal and the specie of the mollusks and also showed that mollusks living in the area accumulate heavy metals in their body tissue.

Bioaccumulation and concentrations of heavy metal in the different mollusks collected from Khanh Hoa coastal, Viet Nam

Heavy metal concentration in sediments

The concentrations of heavy metal in eight collected sediments are summarized in Figure 2 and compared with the New York guidelines criteria [4] for the effect of sediment on benthic organisms. Assessment of quality of sediment was focused on the LEL (Lowest effect level) and SEL (Severe effect level) as proposed by the Department of Environmental Conservation of New York, 1999. These variations between 8 sites may be due to their different geographic locations. The concentrations of heavy metal in sediments collected were ranged between 6.26-19,61; 0,45-1,82; 10,94-63,15; 11,03-381,30; 20,23-52,65, 40,99-465,65 (µg/g dry wt) for As, Cd, Cr, Cu, Pb and Zn respectively. The highest concentrations of Cu, Pb and Zn were in sediments sampled at NT8 and the value measured were 381, 30; 52, 65 and 465, 65 µg/g respectively. The highest concentration of As was observed in the samples from NT2, whereas the highest concentration of Cr was observed in those from NT1. However, the lowest value of Pb was measured in NT4. The mean concentrations of the heavy metals As, Cd and Pb in eight collected sediments were higher than the lowest effect level (LEL) but were lower than the severe effect level (SEL). In classification, these sediments are considered as contaminated, with a moderate impact on benthic life, except sample NT8. Indeed, the Cu and Zn concentrations in sample NT8 was exceeded the SEL, that means the NT8 was contaminated and can cause significant harm to benthic aquatic life.

As concentrations

In conclusion, the results showed that concentrations of heavy metal in Nha Trang are between LEL and SEL values. That means that most of the elements have a moderate impact to benthic life. At NT8 site, some heavy metals can cause harm to benthic life. In terms of geographical location the highest values appeared to be associated with river estuary areas and this may be due to the discharge of untreated water wastes.

Bioaccumulation of heavy metals

The BSAF were calculated for the selected metal in the mollusks studied and showed in the table 2. The data from table 2 also suggested that there are differences in accumulation of heavy metals among the different mollusk species. As indicated in the table 4, Bufonaria rana, Isonomom perna, Modiolus vaginus and Crosostrea rivularus were found to have a large capacity for As and Cd intake. That could be explained by the role of those metals as essential elements for them. Comparatively, the lower potentials of accumulation of metals as Pb and Cr in the tissue are possibly due to non essential metals for these mollusks.

It was also found a very high heavy metals level in the oyster Crasostrea rivularis collected in NT6, but this site of coast is an unpolluted area in terms of industrial impact. In the other hand, the heavy metal concentration in harbor of Nha trang NT1 were also found very low level in organisms, where this site has sources of anthropogenic inputs.

The living condition has a major impact on the accumulation rate of heavy metals in those mollusks. Anadara antiquata and Geloina coaxans are the species of mollusk that live on the bottom, where they burrow in the sand or mud. They are very much exposed to the heavy metal accumulated in the sediment. The BSAF values of these organisms were lower than 2, thus show that they have a low potential of accumulation of heavy metal from sediments. On the other hand, Crasostrea rivularis, Bufonaria rana and Modiolus vaginus which have a high potential of accumulation (BSAF values were higher than 2), are mollusks that live by stickling to any solid material such as rocks in the sea. As a result, they are more exposed to water and accumulate heavy metal by trapping food from water.

In conclusion, the accumulations of heavy metals were different between species and may exhibit different accumulation capacity for heavy metals. The protection of coastal pollution from damage due to these metals and other chemical requires an understanding of the sensitivity of mollusks to these substances and their ecological requirement. Monitoring programs for coastal pollutions may help to obtained relevant information.

CONCLUSION

As a conclusion, the bivalve mollusks can be useful as bioindicator for a coastal contaminant, especially soft tissues of mollusks which are suitable to be used as a toll for heavy metal contamination, due to its natural capacity to regulate and accumulate concentration of various metals. It is clear that the biomonitoring plan is not only site specific, but that it considers the use of species whenever possible. This research provides the evidences a general review of several bivalves and mollusks in Nha Trang Bay impacted by heavy metals, and give suggestion that the mollusks can be an established and essential biomonitor of heavy metal contamination particularly in the coastal Khanh Hoa area. 8-262.9, 0.80-6.24, 0.12-87.6, 8.28-23.7, and 4.31-3525 (µg/g dry wt) for As, Cd, Cr, Cu and Zn, respectively. Mean concentrations of As, Cu, Cr, and Cd in the selected mollusk species were below the maximum permissible level (MPL) when compared with the FDA guidance document.

SỰ TÍCH LŨY KIM

Key words: Bioaccumulation, Heavy metal, Mollusk species, Khanh Hoa.

I. INTRODUCTION

The pollution of aquatic ecosystems by trace metals is a significant problem, as trace metals constitute some of the most hazardous substances that can bioaccumulation Nguyen [START_REF] Van | Study concentration of heavy metals cadmium (Cd) and lead (Pb) in clam Corbicula sp. from estuarine, in Da Nang city -Tạp chí Khoa học và Công nghệ[END_REF]. The bioaccumulation of heavy metals by marine mollusc and other marine organisms may reach many orders of magnitude above background concentrations of certain locality. This phenomenon may demonstrate the potential of these species as a biomonitor of heavy metal pollution Chan (1989), Hamed & Emara (2006). Many studies have shown that intertidal molluscs can be good biomonitoring organisms [START_REF] Ismail | The use of intertidal molluscs in the monitoring of heavy metals and organotin compounds in the west coast of Peninsular, Malaysia[END_REF]. Due to heavy metals contamination are very localised and closed to discharge point sources [START_REF] Ismail | The background levels of heavy metal concentrations in sediments of the west coast of Peninsular, Malaysia[END_REF]) and molluscs inhabit in different microhabitat of intertidal areas, a detail studies on different species and tissues are important [START_REF] Edward Berandah | Bioaccumulation and Distribution of Heavy Metals (Cd, Cu, Fe, Ni, Pb and Zn) in the Different Tissues of Chicoreus capucinus Lamarck (Mollusca: Muricidae) collected from Sungai Janggut, Kuala Langat, Malaysia[END_REF].

A lot of studies on mollusks associated with heavy metal pollution have been done by many researchers [START_REF] Abdullah | Heavy Metals (Cd, Cu, Cr, Pb and Zn) in Meretrix meretrix Roding, Water and Sediments from Estuaries in Sabah[END_REF], but studies of this kind have been rare in Vietnam due to local difficulties in organizing major studies and legal restrictions regarding the export to fin digenous biologic material [START_REF] Wagner | Biomonitoring of trace elements in Vietnamese fresh water mussels[END_REF] For example, the concentrations of some heavy metals in the tissues of bivalve molluscs have been observed in the samples collected along the coastal of Viet Nam (Le Thi Mui, 2008; Nguyen [START_REF] Van | Study concentration of heavy metals cadmium (Cd) and lead (Pb) in clam Corbicula sp. from estuarine, in Da Nang city -Tạp chí Khoa học và Công nghệ[END_REF][START_REF] Tu | Concentrations of trace elements in Meretrix spp. along the coasts of Vietnam[END_REF]. However, none of those studies specifically investigated the bioacumulative factor of such a species.

The objective of this study is to provide important information of the metal distribution in the sediments and different mollusk tissues. Thus, helping in proposing the potential bioaccumulation by determining its Biota-Sediment Accumulation Factor (BSAF) (Dallinger et al. 1993) and suggesting bio-monitor for heavy metals in Khanh Hoa coastal.

The work is based on study of samples of sediments and 4 mollusks species (Glauconome virens (Linnaeus, 1767), Laternula anatina (Linaeus, 1758), Katelysia hiantina (Lamarck, 1818) and Anadara antiquata (Linaeus, 1758) taken in September 2012 in 4 locations within Nha Phu lagoon (Ninh Hoa) -Nha Trang bay and Thuy Trieu lagoon (Cam Lam). There locations provided wide intertidal zone across the Khanh Hoa coastal in terms of the sandy-muddy areas, represented high community of the bivalve mollusks.

II. MATERIAL AND METHOD

Study sites

Khanh Hoa provincecentral-southern Viet Nam, which is an important ecological zone and providing a wide range of coastal and marine ecosystems in relation to prevailing oceanographic conditions and gradients in mainlandoceanic influence. Compared with some other areas, species composition of molluscs in Nha Trang Bay (490 species) was more abundant than Cat Ba -Ha Long Bay (372 species) and Tonkin Gulf (470 species) Vo Si Tuat et al. (2002).

Sample collection and preparation

Organisms collection

About 10-20 numbers of the same sizes of each mollusk species (Glauconome virens (Linnaeus, 1767), Laternula anatina (Linaeus, 1758), Katelysia hiantina (Lamarck, 1818) and Anadara antiquata (Linaeus, 1758) were picked by hand from four locations and then put in ice box at 4 0 C to be transported to the laboratory. The tissues were separated by a plastic knife and dried at 60 o C during two days to constant dry weight. These dried tissues were then stored in a polyethylene bag to be sent in ECOMERS laboratory (UNS-France) for the determination of heavy metals concentrations.

Sediment collection

The sediment samples of the top 20cm surface centimeter were collected with a polyethylene spool or Ekman grab, then placed in a plastic bag, which were stored in an ice box at 4 0 C, to be transported to the laboratory. There sediment samples were dried at room temperature, and then were sieved through a nylon mesh to obtain particles smaller than 0.2mm in diameter for determination of heavy metal concentration. For each sediment sample, 0.1gram was taken for heavy metal analysis.

Sample digestion and analysis

Sediment digestion

The samples were digested following the guideline from EPA 3050b US EPA (1996) for digestion of sediments using a Microwave 3000. A 0.1 gram of dried and sieved sediment samples were weighed out in the reaction vessel then added with mixed of 9ml concentrated acid HNO 3 (69%) and 3ml of HCl (36,46%). Vessels then were placed in the rotor in the microwave. The vessels were heated to at least 240 o C over 35 minutes and then held at 210 o C for at least 15 minute. The samples were cooled and mixed to volume to 50 mL with ultra-pure water. The obtained solution was then centrifuged and the concentration of heavy metals measured using ICP-OES (Perkin Elmer). The accuracy of the analytical procedures employed for the analysis of metals in sediment samples was checked using the marine sediment IAEA SP-M1 as certified reference material.

Tissues digestion

The digestion method of organically based matrices used is the one described in the guideline EPA 3052 US EPA (2008). The tissue sample was dried, pulverized and sieved for digestion and determination of metals concentrations. A weight of about 0.5 gram of dry tissue was then transferred in a 100 ml Pyrex beaker and added with 9 ml of acid HNO 3 (69%) and 1ml of HCl (36,46%). Then the beakers were placed on a hot place for reflux extraction at 95 o C, for 30min. The digested samples were diluted and filled up with de-ionized water to a total volume of 50ml before metal analyzing. Heavy metals were analyzed by ICP-OES (Perkin Elmer). The accuracy of the analytical procedures employed for the analysis of metals in organic samples was checked using the Fish tissue reference IAEA MA-B-3/TM as certified reference material, and showed good agreement with the certified values.

Data analysis

A mollusk's capacity to accumulate metals from living medium into its tissue can be estimated using Biota-Sediment Accumulation Factor (BSAF). BSAF were calculated for the selected metal in the studied mollusks according to the formula BSAF = Cx/Cs where Cx and Cs were the metal concentrations in the organisms and in associated sediments, respectively (Phillips 1997). By comparing BSAF values, we can compare the ability of those mollusks in taking up metal from sediment. The categories of BSAF are presented as: High potential of accumulation if BSAF>2, Moderate potential if 1<BSAF<2 and Low potential of accumulation if BSAF<1 [START_REF] Wich | Technical guidance for screening contaminated sediment[END_REF].

III. RESULTS AND DISCUSSION

Sediments quality

The sediment qualities from the four sites are given in Figure 2. Concentrations of As, Cd, Cr and Cu in collected sediemnts were between LEL (Lowest Effect Level) and SEL (Severe Effect Level) values event the concentrations of Zn and Pb were lower than LEL. It indicated that the coast has been polluted by most of the heavy metal. Indeed, the effect of these sediments is considered to be contaminated, moderate impact to benthic life [START_REF] Fletcher | Guidelines for identifying, assessing and managing contaminated sediment in Ontario: An integrated approach[END_REF].

As concentrations in sediments

Table 1. Metal concentrations (mg/kg DW) in tissues of four collected organisms.

Metal concentrations of Al and Fe in the tissue of the four sites were similar and did not vary significantly and presented very lower compared to concentrations of these metals in the sediments. Concentrations of Al and Fe in the sediments were 20-100 times higher than the concentration in the mollusks. For most of the heavy metals, when mean values were calculated, values were close to FDA maximum permissible level (Rainbow & Phillips, 1993;Rainbow et al., 2000), except for Pb. The Pb concentration in all of samples was more accumulated and was higher than the FDA maximum permissible level (Pb<1.7µg/g dry wt), which eventually may pose hazard to human in term of health risk. The observed concentration of heavy metals in the mollusk samples showed that the magnitude of the heavy metal accumulation in the tissues depend on the type of the heavy metals and the species. There values of BSAF in the different mullusk species are shown in Table 2. On the other side, when BSAF were calculated, it indicated that different species showed different bioaccumulation of metals levels depending on the geographical location. In general, for all of the studied metals, the BSAF values for As were highest than those for Pb were lowest. The BSAF indicated a low potential bioaccumulation for Pb and Cr (BSAF<1) and a very high potential of accumulation for As, Cd, Cu and Zn.

From the results, it was found that clam Katelysia hiatina (Lamarck, 1818) highly accumulate As, Cd, Cu and Zn (BSAF>2), and then could be chosen as sensitive indicators for monitoring heavy metal pollutions in Khanh Hoa coastal, Viet Nam.

The heavy metal contents in the tissues of Katelysia hiantina and sediments

The clam Katelysia hiantina (J.B.Lamarck, 1818) also known as other names Marcia hiantina and local names "ngao den" or "ngao gia", is an economical important bivalve and can be also represented in Khanh Hoa coast -Nha Trang bay as one of the local abundant mollusca. This clam belongs to the class Bivalve, order Veneroidea in which shell is medium, oval and thick, that live on the loose bottom of water bodies. They are filter feeder and thus very potential to accumulate toxic substances from water and sediment.

The concentrations of metals (mg/kg dry wt) in whole soft tissue of the clam K.hiantina and surface sediments from the four sites are given in Figure 3.

It was observed that clam K.hiatiana accumulate 8 metals at different concentrations and at different potential of bioaccumulation. The distribution of metals in the tissue of K.hiatina, which inhabit in different habitat of intertidal areas, shows that metals contamination are very localized and closed to the anthropogenic influences. The study has concluded that clam K.hiatina could be a good bio-indicator of the particular environment through bio-monitoring process.

The higher bioavailability of heavy metals to TD and CL could be explained by the significantly lower organic matter contents, which could potentially affect metal availability, or affect several physiological processes that influence the accumulation of heavy metals by K.hiatina.

IV. CONCLUSSION

The coast has been polluted by most of the heavy metal as As, Cd, Cu, Cr. Indeed, the effect of these sediments is considered to be moderate impact to benthic life. Concentrations of almost heavy metals in 4 species mollusk collected were closed to FDA maximum permissible level except for Pb.

The BSAF indicated a low potential accumulation for Pb and Cr and a high bioaccumulation for As, Cd, Cu and Zn. Clam K.hiantina is represented in study zone as the most local abundant mollusca. This species high accumulate heavy metals and then could be chosen as sensitive indicators for monitoring heavy metal pollutions in Khanh Hoa coastal, Viet Nam. This preliminary sampling should merit further studies using the BSAF values for assess contamintion in coastal areas by demonstrate the potential of bioaccumulation of mollusk species

Metals

APPENDIX 1 .

 1 PUBLICATION ...................................................................................200 APPRENDIX 2. SOME PICTURE OF COLLECTING SAMPLESError! Bookmark not defined. APPENDIX 3. METHOD OF ANALYSIS .................... Error! Bookmark not defined. OF ANALYSIS ...................... Error! Bookmark not defined. xii LIST OF TABLES

Figure 3 . 1 Figure 3 . 2 Figure 3 . 3 Figure 3 . 4 Figure 4 . 1 Figure 4 . 2 Figure 4 . 3 Figure 4 . 4 Figure 4 . 5 Figure 4 . 6 Figure 4 . 7 Figure 4 . 8 Figure 4 . 9 Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .Figure 4 .

 3132333441424344454647484944444444444444444444444444444444444444 Figure 3.1 Sampling and evaluation scheme of sediment ...................................................70

  are antagonistic of Cu and Zn ions, inhibit their assimilation and stimulate the accumulation of Cr, Ni, Co, Cd, and Pb, which have strong positive association with Fe ions. Extremely high concentrations of Cr, Ni, Co, Cd, and Pb in the individuals displaying the signs of pathology and mutagenesis are indicative of "excess" or toxic accumulation of these elements. In high heavy metal medium gradients, similar signs may be related to the toxic concentration of Zn, Cu, Cd, and Pb at a reduced Fe and predominant Mn accumulation. Synergistic interaction of Zn and Cu ions inhibits the accumulation of Ni, Cr, and Co, as their antagonists. No mollusks with signs of pathology were found in the low heavy metal medium gradients. The deficient or optimal heavy metal accumulation can be assessed from the concentrations of the above metal indicative of an intense or relatively low accumulation, as estimated by the coefficient of biological accumulation

  Size / Weight / Age: Max length: 6.3 cm TL male/unsexed; common length: 4.0 cm TL male/unsexed; Environment: Benthic; depth range 0 -25 m Climate / Range: Tropical Distribution: A.antiquata is known from islands of the Pacific, Indo-Pacific: Eastern Africa to Japan, Australia, eastern Polynesia and Hawaii. (Poutiers, J.M. 1998) Picture 1.2 Anadara antiquata (Linnaeus, 1758) pipes It is primarily found in estuarine habitat with salinities ranging from 18-33 ppt and temperatures from 11-32 0 C. Perna viridis

  Heterodonta Infraclass: Euheterodonta Size / Weight / Age: Max length: 10.8 cm SHL male/unsexed; common length: 8.5 cm SHL male/unsexed; Environment: Benthic; depth range 0 -20 m Climate / Range: Tropical Distribution: Indo-West Pacific: from East and Southeast Africa, to Melanesia; north to southern Japan and south to Queensland and New Caledonia. (Poutiers, J.M. 1998) Picture 2.9 Tapes literatus (Linnaeus, 1758)

Picture 2 .

 2 10 Tan Dao -TD sampling site Location 2 (Ngoc Diem -ND): Is located 15km from the North of Nha Trang bay, which receives agricultural effluents from the hinterlands predominantly which is an embayment influenced by strong tidal currents, a rare phenomenon in the Nha Trang bay. Picture 2.11 Ngoc Diem -ND sampling site Location 3 (Binh Tan -BT): The Be estuary is situated on the Southern coast of Nha Trang City (BT), which dump waste directly into the ocean. This is a estuary which is an almost enclosed 'polluted' area near Nha trang port receive disposal effluents of domestic sewage water as well as waste disposal through boats and ships and is affected by several activities of the Nha Trang City. TIMES AND SAMPLING SITES One of the objectives of this study was to obtain a dataset of trace metal concentrations in bay of Nha Trang. Sampling density was adapted to assumed points of anthropogenic inputs and characteristic of the Nha Trang bay. In order to achieve the objective, surface sediment and mollusk samples were collected from 4 stations: Tan Dao (Ninh Loc, Ninh Hoa) -TD, Ngoc Diem (Ninh Ich, Ninh Hoa) -ND; Binh Tan (Nha Trang) -BT and Cam Hai Tay (Cam Lam) -CL.

Figure 3 .

 3 Figure 3.1 shows the sampling techniques and processing of sediment samples for different analyses.

Figure 3 . 1

 31 Figure 3.1 Sampling and evaluation scheme of sediment

  relatively large sub sample to be taken accurately to a depth of 2cm. This procedure avoided any sample pollution during collection. All objects coming in contact with the sample were made of glass or stainless steel. To avoid pollution, all sampling equipment had been cleaned in sequence with local seawater, chromatographically pure acetone and Sampling Fe, Al, Cr, As, Cd, Zn, Cu Weight loss 16h-105 o C

pH

  The measuring the pH (H 2 O) of sediment following to ASTM Test Method. Use a 1:5 dilution of sediment: water, and then take a pH measurement on the resulting solution with a laboratory meter. Use the 1:5 dilutions, but instead of water they use a dilute Kalium Chloride (KCl) solution and then take a pH (KCl) measurement on the resulting solution with a laboratory meter.The pH measured in KCl is almost always lower than pH of the same soil measured in water due to the higher concentration of H + . The procedure gives a value similar to that for natural sediment solution because the sediment solution also contains dissolved K 2+ and other ionsMoisture content determinationStandard procedures (Method ASTM 2974-87) were adopted for determination of moisture contents. Moisture was determined by two steps: One gram of the sediments was dried in air at room temperature then oven drying them at 110 o C in 16h to constant weight. Moisture contents were measured by the weight loss as a percentage of the dry soil weight. weigh wet soil -wet of oven dry soil 110 o C MC% = ----------------------------------------------------------x 100% eigh of wet soil TOC Total organic carbon Organic matter determination Organic matter was determined by ash dried sediments in the furnace to 750 o C and hold until the samples completely ashed. (Method ASTM, 2974). The loss of weight after treatment was assumed to be due to the organic matter content weigh dry soil of oven 750 o C MC%

For

  dry density determination, 10 pots were filled with sediment samples and were to constant weight at 70 o C. Dry bulk density was calculated as dry weight divided by wet volume. Soil Bulk Density is the dry mass of a soil divided by its volume. Bulk density value for soil is expressed as follows: Db= (Oven dry soilg/Total soil volume) g/m 3 Bulk density related to OC content (Y Avnimelech 2001) and show structure of mineral aquatic bottom sediment as mineral particles by organic micelles.

  1 ICP -OE Spectrometer for digestion samples Metals as arsenic (As), cadmium (Cd), chromium (Cr), copper (Cu), lead (Pb), nikel (Ni), molipden (Mo), and zinc (Zn) as well as iron (Fe) and aluminum (Al) are all considered to be elements of concern to study, verified by literature reports.

Figure 3 .Figure 3 . 3

 333 Figure 3.3 Sampling and evaluation scheme of mollusks

a

  scalpel was used to open the shell and remove the soft tissue from the shell (Picture 3.2). Tissue of mollusks were analyzed as a composite of 5-10 animals because tissue mass was small. Once all individuals that comprised a composite had been cleaned and weighed, they were placed in a stainless steel blender cup and dry at 60 o C to constant weight. The dry sample was then placed in a labeled plastic bag, which was then capped and frozen until transferred to the Ecomers Laboratory UNS for analysis. Picture 3.2 Measurement of mollusk samples

Figure 3

 3 Figure 3.4 Schema of mollusks digestion method

Picture 3 . 4

 34 ECOMERS laboratoryReagent blanks were processed with each digestion set by completing the digestion and analysis procedure on samples containing only reagents. This was done to determine if reagents contribute measurable quantities of the metals in question or if contamination is added during the digestion. Arsenic, cadmium, chromium, copper, lead and zinc were all below the Limit of Detection (LOD) or within the Limit of Quantitation (LOQ) in the reagent blanks. LOQ is defined as 10/3 of the LOD. Both reagent blanks for iron and aluminum had values above the LOQ.

5 :

 5 Factor of enrichment due to lithological variation C n Concentration of element in sediment B n Concentration of element in background value

  -range-low (ER-L) illustrates 10th percentile of chemical concentrations corresponding to the adverse biological effect; while effect-range-median (ER-M) for 50 th percentiles of chemical concentrations are corresponded. Three ranges in chemical concentration are evaluated using these terms: Adverse effects were rarely observed < ER-L Adverse effects were occasionally observed ≥ ER-L and < ER-M Interim Sediment Quality Values ISQVs from Hong Kong are meant for assisting in decision-making of dredged seabed sediment disposal by contamination level, they are adapted in this study as an extra reference. ISQVs displayed high similarity with other SQGs mentioned in above in terms of characteristics. However, there are differences in terms of contamination effect values. The contaminants range values in ISQVs are expressed as ISQV-low and ISQV-high. The contamination values below ISQV-low indicate adverse biological effects are unlikely, while values above ISQV-high indicate that severe adverse biological effect are very likely as a consequence of sediment contamination.

  of risk index RI of all metals byMadiseh and others, 2009.

  concentration of metal in shellfish species (mg/kg on fresh weight basis) W food: represent the daily average consumption of mollusk in region BW: the body weight Then the health risks from consumption of shellfish by local inhabitants assess based on the compassion with acceptable daily intake limit given by National research council of the National academy of sciences of the USA (for Cu 2000-3000µg), Environmental health criteria 221 (for Zn 5600-15000µg), Guidance document for Lead in shellfish, FAO/WHO 2003 for Cd: 60µg and USFDA, 1993 for Pb: 25-75µg (See Appendix 6.6)

4 Figure 4 . 1 Figure 4 . 2 Figure 4 . 3 Figure 4 . 4

 441424344 Figure 4.1 Percentage of particles size in Tan Dao during 2012-2013

Figure 4 . 5 Figure 4 . 6 Figure 4 . 7 Figure 4 . 8

 45464748 Figure 4.5 Particles size of sediment samples in Tan Dao at two seasons

Figure 4 . 9

 49 Figure 4.9 Trend of pH in 4 sampling locations from 2012 to 2013

  . The sediment collected from TD mangroves showed a decreasing trend of organic carbon due to restricted microbial activity in this anoxic or sub-oxic region and also due to partial decomposition of leaf litters, root exudates, and their preservation as burial in the sediments. Over all a high distribution of organic cacbon was observed in both ND and BT due to solid waste coupled with boat activities and physical mixing processes. Trend of total organic carbon content in sediment samples during 2012-2013 is shown in figure4.10.

Figure 4 . 10 4 . 1 . 4

 410414 Figure 4.10 Trend of total organic carbon content in sediment samples

Figure 4 .Figure 4 . 11 4 . 1 . 5

 4411415 Figure 4.11 bellow shows a range of bulk density of sediment samples during 2012-2013:

Figure 4 . 13 HeavyFigure 4 . 14 Figure 4 . 15 HeavyFigure 4 . 16

 413414415416 Figure 4.13 Heavy metals concentration of sediment collected from TD 2012-2013

  sources. So, Cr levels in the sediment were in the range of 21.81 to 41.96 mg/kg DW. The strong concentration of As and Cu were observed in BT estuary. When this local area is not considered, the increase of metal concentrations in the surface sediments are associated with the harbour areas: up to 53.94 mg/kg DW of Cu in rainy season to 43.24 mg/kg DW in dry season and to 14.1 mg/kg DW As in rainy season. Thus, the pattern of distribution of this metal shows the affect of the domestic activities and secondly the shipyard such as is observed in the middle of the Nha Trang habourd. These results validate the industrial effects already pointed out for As.The strong gradient in the As was observed around the CL, that shown a big difference which is in the TD because in this area the As concentration changes strongly from the rainy season to dry season although maximum values are found 14.10 mg/kg DW in dry season. The anthropogenic origin is clear in the sedimentary register, however the arsenic is dispersed throughout the dry season at TD and is not concentrated in the this area.

Figure 4 . 17

 417 Figure 4.17 Percentage of samples collected in dry season amongst range of SQGs

  Zn and Ni in both dry and rainy season and between Cu-Zn and Fe, Al, Ni in dry season. The same holds true for sediment concentrations of Fe has also shown that the high correlation of Al and Ni (r = 0.99). Zn is high affected by a Cu concentration (r=0.94). Negative correlations are obtained between As and Cd metal (r = -0.82) and between Cd and Cu and Zn (r = -0.35 and -0.38) in rainy season. This can be attributed to the different longitudinal variations of these metals in the coastal. Major seasonal variations in metal concentrations occur in the four sites.

  Due to relatively low concentration of OC in sediments, it does not act as a carrier of metals, and therefore shows asynchronous distribution pattern with trace metals. Strong inter-element relationships suggest that one common process determines the sediment composition for all trace metals. The strong positive correlation between Cr and other metals as Cu, Zn, Fe and Al (r = 0.80 -0.96) reveals that this metal are carried by pollution of Cr, which come from industrial discharges, agricultural activities into the near by environment. In moreover, the poor association of As with other metals suggests that As possibly due to different processes like biological effects and external inputs operating in the mangrove and adjacent estuarine sediments. Significant correlations of Cr, Cu, Zn with Fe, Al indicate the industrial contribution of these metals into the system.

Figure 4 .

 4 19 to Figure 4.25. These results

Figure 4 .

 4 Figure 4.25 Concentrations of metals in shell and tissue of A.antiquate

Figure 4 .

 4 Figure 4.28 Contents of metal in soft tissue of Geloina coaxans

Figure 4 .Figure 4 .Figure 4 .

 444 Figure 4.29 Contents of metal in soft tissue of Lingula unguis

Figure 4 .

 4 Figure 4.32 Contents of metal in soft tissue of Solens regularis

  Figures 4.36, 4.37, 4.38 and 4.39 present metal concentrations in seven mollusks

Figure 4 .

 4 Figures 4.36, 4.37, 4.38 and 4.39 present metal concentrations in seven mollusks collected in Nha Trang bay in comparision with FDA and WHO standards. On the contrary, higher than WHO standard concentrations of As, Cd, Cr and Cu were noted in mollusks collected in the 4 site. The high concentrations of these matals in mollusks from the site could be linked to the state of pollution in Nha Trang bay.

Figure 4 .

 4 Figure 4.37 Cd contents (mg/kg DW) in soft tissue in comparing with standard values

Figure 4 .

 4 Figure 4.38 Cr contents in soft tissue of mollusks in comparing with standard values

Figure 4 .

 4 Figure 4.39 Cu contents (mg/kg DW) in soft tissue in comparing with standard values

Figure 4 .

 4 Figure 4.40 Zn contents (mg/kg DW) in soft tissue in comparing with standard values

  ND: not detected, Red Bold: High potential of accumulation, Bold Italic: Moderate potential of accumulation and Regular: Low potential of accumulation.

Figure 4 .Figure 4 .Figure 4 .

 444 Figure 4.41 Arsenic accumulation in soft tissue of K.hiantina

Figure 4 .

 4 Figure 4.47 Aluminium accumulation in soft tissue of K.hiantina

  The trace metal concentrations determined in the present study for 4 sediments and soft tissues of the 11 mollusk species collected in period 2012-2013 in Nha Trang bay, Khanh Hoa, province, Viet Nam are reported for each of ones. During each sampling event, both organisms and sediment were collected from the 4 areas and analyzed for the metals of concern. The variation in trace metal concentrations between individual mollusks was predictable to some extent based on environmental factors, their body weight/shell size, patterns of essential trace metals depending on the group and appears to be affected by both seasonal and locational variations. Results from this study demonstrated that:Acording to data analysis of physicochemical characteristics, the sediments collected in Nha Trang bay were classified as silt clay to clayely silt with pH neutral, lighter organic matter, low AVS and differences of moisture content from low to high, depends of location of sampling

Figure 1 :

 1 Figure 1: The grain size distributions of sample

Figure 2 :

 2 Figure 2: Heavy metal concentrations of Nha Trang Bay in period 1996-2011

Figure 2 .

 2 Figure 2. Heavy metal concentrations in sediment samples (µg/gram dry weight)

1 .

 1 Môi trường, Đại học Khoa học Tự nhiên, Đại học Quốc gia TP.HCM 2 . EA 4228 ECOMERS, Khoa Khoa học, Đại học Nice Sophia Antipolis, Pháp Tóm tắt: Nghiên cứu bao gồm việc xác định thành phần một số kim loại không ảnh hưởng đến sinh vật như sắt (Fe), nhôm (Al) cũng như hàm lượng các kim loại nặng độc hại khác như arsen (As), đồng (Cu), cadmium (Cd), crôm (Cr), chì (Pb) và kẽm (Zn) trong các mẫu trầm tích cũng như thịt của một số . Mẫu các cũng như mẫu lấy tại 4 điểm thuộc vùng ven biển từ đầm Nha Phu (thuộc huyện Ninh Hòa) tới T u ( huyện Cam Lâm) vào tháng 9/2012. Mẫu 4 phi cái (Glauconome virens (Linnaeus, 1767), phi đực Laternula anatina (Linaeus, 1758), ngao đen Katelysia hiantina (Lamarck, 1818) và sò lông Anadara antiquata (Linaeus, 1758) được mang về phòng thí nghiệm, sấy khô trong 24 giờ 60 o C, nhiệt phân hủy với hỗn hợp axit HNO 3 (69%) và HCL (36,46%) và phân tích các kim loại nặng bằng máy khối phổ plasma cảm ứng ICP-OES. Hàm lượng nhôm (Al) và sắt (Fe) không có sự khác biệt nhiều trong thịt các khác nhau nhưng rất thấp so với hàm lượng kim loại trong . Hàm lượng kim loại nặng As, Cu, Cr, Cd trong các mẫu sinh vật nằm dưới ngưỡng cho phép (MPL) theo tiêu chuẩn của Cục an toàn thực phẩm Mỹ FDA nhưng hàm lượng chì (Pb) và kẽm (Zn) cao hơn 4 lần ngưỡng cho phép của FDA. Từ khóa: Tích tụ sinh học Kim loại nặng, Nhuyễn thể, Khánh Hòa. BIOACCUMULATION OF HEAVY METAL IN SOME MOLLUSK SPECIES AT THE COAST OF KHANH HOA PROVINCE, VIET NAM Tran Thi Mai Phuong 1, 2 ,* Nicolas Marmier 2 , Charlotte Hurel 2 , Nguyen Ky Phung 1 Department of Environment, University of Sciences of Ho Chi Minh City 2 . EA 4228 ECOMERS, Faculty of Science, University of Nice Sophia Antipolis the tissues ranged between 33.

  Sediments and mollusks samples were collected from four sites: Tan Dao (Ninh Loc, Ninh Hoa) -TD (12°02'17"N; 109°08'14"E), Ngoc Diem (Ninh Ich, Ninh Hoa) -ND (12°23'42"N; 109°11'57"E); Binh Tan (Nha Trang) -BT (12 0 12'22"; 109 0 11'17"E) and Cam Hai Tay (Cam Lam) -CL (12 0 04'45"N; 109 0 10'42"E). The locations of sampling sites are showing in Figure 1.

Figure 1 :

 1 Figure 1: Map and locations of sampling site

  collected sediments (mg/kg dry wt) Cd contents in collected sediments (mg/kg dry wt) collected sediments (mg/kg dry wt) Cu contents in collected sediments (mg/kg dry wt) collected sediments (mg/kg dry wt) Cr contents in collected sediments (mg/kg dry wt)

Figure 2 :

 2 Figure 2: Concentrations of heavy metals in collected sediments.

Figure 3 :

 3 Figure 3: The contents of metals in soft tissue of the clam and sediments

Table 1 .

 1 1 Trace metals (µg/kg DW) in marine sediments with different world areas ........14

Table 1 .

 1 2 Trace metal cotents in dried soft tissue of clams from different environment ....21

Table 1 .

 1 3 Mollusk species are used as biomonitor in marine of Vietnam...........................43 Table 1.4 Dominated bivalve species in Nha Trang bay .....................................................43 Table 2.5 Water quality of Nha Trang Bay .........................................................................50 Table 2.6 Water concentration of some heavy metal in Nha Trang Bay.............................50

Table 2 .

 2 7 Chemical characteristics of sediment from Nha Trang Bay................................51

Table 2 .

 2 8 Heavy metal concentrations of sediment from Nha Trang samples ....................52

Table 2 .

 2

9 Concentration of metal (µg/g DW) in surface sediment at period 1996-2011 ....52 Table 3.1 Localities of sampling sites .................................................................................68 Table 3.2 Sampling dates.....................................................................................................69 Table 3.3 Grain size classification .......................................................................................71 Table 3.4 Mollusk species and collected numbers of them ................................................77 Table 3.5 Gradation of Contmination indices Cd ................................................................84 Table 3.6 EF categories .......................................................................................................85 Table 3.7 The 7 grades of classes Igeo ................................................................................86

Table 3 .

 3 8 Sediment quality criteria......................................................................................87

Table 3 .

 3 9 The factors represent for accumulation of chemical in organisms ......................92

Table 3 .

 3 10 Classification of sites by BSAF.........................................................................93

Table 3 .

 3 11 Classification of COPC by bioaccumulation index ...........................................94

Table 4 .

 4 1 pH measurement of sediment samples ..............................................................104

Table 4 .

 4

2 Percentage of total organic carbon TOC (%) in sediment samples ...................105 Table 4.3 Bulk density (g/cm 3 ) of sediment samples ........................................................106 xiii

Table 4 .

 4 4 Percent moisture contents in sediment samples................................................107

Table 4 .

 4 5 The AVS and Bi-carbonat content in the sampled sediments. .........................108

Table 4 .

 4 6 Sampling site classification ...............................................................................109

Table 4 .

 4 7 Sequential orders of heavy metals in sampling sites .........................................110

Table 4 .

 4 8 Values for metals in surface sediment collected in dry season .........................115 Table 4.9 Values for metals tested in surface sediment collected in rainy season ............115

Table 4 .

 4 10 Interactions between metals in rainy season....................................................118

Table 4 .

 4 11 Interactions between metals in dry season.......................................................119

Table 4 .

 4 12 The interactions between metals and physicochemical characteristics of sediments collected from Nha Trang bay and adjacent areas ............................................121

Table 4 .

 4 13 Metals in correlation with the physico-chemical factors in Tan Dao station ..122

Table 4 .

 4

14 Metals in correlation with the physicochemical factors in ND station............123 Table 4.15 Metals in correlation with the physico-chemical factors in Binh Tan station .124 Table 4.16 Metals in correlation with the physico-chemical factors in CL station ...........125 Table 4.17 MPI during rainy season for 4 locations of sampling......................................138 Table 4.18 MPI during dry season for 4 locations of sampling.........................................139 Table 4.19 Moisture contents in mollusk tissues...............................................................145 Table 4.20 The calculated condition index for seven species............................................146

Table 4 .

 4 21 Correlation coefs between biometric and metal concentrations in K.hiantina 147

Table 4 .

 4 22 Correl coefficients between biometric and metal concentrations in G.virens .147

Table 4 .

 4 23 Correl coefficients between biometric and metal concentrations in L.anatine148

Table 4 .

 4 24 Correl coefficients between biometric and metal concentrations in L.unguis.149

Table 4 .

 4 25 Corre coefficients between biometric and metal concentration in G.coavans 149

Table 4 .

 4 26 Corr coefficients between biometric and metal concentrations inA.antiquate150

Table 4

 4

.27 Corr coefficients between biometric and metal concentrations in S.reguraris150 xiv

Table 4 .

 4 28 Comparision BSAF in shell and tissue of different mollusk species...............152

Table 4 .

 4 29 Values of bioaccumulation factor of heavy metal in seven mollusk species ..153

Table 4 .

 4 30 Calculation of MSWI for K.hiantina ...............................................................160

Table 4 .

 4 31 Bioaccumulation of heavy metal in clam K.hiantina ......................................163

Table 4 .

 4 32 Daily trace metal intake EDI through K.hiantina consumption ......................165

Table 4 .

 4 33 Daily trace metal intake EDI through G.virens consumption..........................165

Table 4 .

 4 34 Daily trace metal intake EDI through L.anatina consumption........................165

Table 4 .

 4 35 Daily trace metal intake EDI through L.unguis consumption .........................166

Table 4 .

 4 36 Daily trace metal intake EDI through G.coaxans consumption ......................166

Table 4 .

 4 37 Daily trace metal intake EDI through A.antiquate consumption.....................166

Table 4

 4

.38 Daily trace metal intake EDI through S.regularis consumption......................167 Table 4.39 Daily trace metal intake EDI through C.revularis consumption .....................167 Table 4.40 The target hazard quotient for K.hiantina........................................................168 Table 4.41 The target hazard quotient for G.virens ...........................................................168 Table 4.42 The target hazard quotient for L.anatine .........................................................168 Table 4.43 The target hazard quotient for L.unguis..........................................................169 Table 4.44 The target hazard quotient for G

.coaxans........................................................

169 Table 4.45 The target hazard quotient for A.antiquate ......................................................170 Table 4.46 The target hazard quotient for S.regularis.......................................................170 Table 4.47 The target hazard quotient for C.rivularis .......................................................170 xv LIST OF FIGURES

  electroplating, leather tanning, textile industry and consumer products such as inks, paints, papers, toner powder used in copying machine… Chromium is the naturally occurring compound found in soil, rocks and plants. It is normally exists in oxidation states ranging

	from chromium (II) to Chromium (VI). However, two major forms trivalent (III) and
	hexavalent (VI) forms have biological significance.
	Physiologically chromium is considered as a trace element and it is required for the
	optimum function of insulin in mammalian tissues and the maintenance of normal
	metabolism of glucose, cholesterol and fat. The normal level of blood chromium
	concentration in human beings is between 20-30µg/l. It is found that the intake of
	chromium is about 50-200 µg/day is regarded to be safe and adequate.
	Hexavalent chromium is an extremely toxic metal, which exist as an anion (CrO 4	2-

) and most readily absorbed from the gastrointestinal tract, skin and lungs. Most reports describe the toxicity of chromium (VI) in the form of chromate of dichromate. It can cause chronic ulceration of skin surface, denaturation of tissue proteins, asthma, kidney failure, discoloration of teeth and inflammation of skin. Acute poisoning results in symptoms such as dizziness, intense thirst, abdominal pain, vomiting and shock and sometimes death may occur due to the presence of urea in blood.

  the average American diet containing about 300 µg Ni/d. Nickel is highly mobile in soil, particularly in acid soils. There is little evidence that nickel compounds accumulate in the food chain. Nickel is not a cumulative toxin in animals or in humans. The initial effects involve irritation of the respiratory tract and nonspecific symptoms. Patients with severe poisoning develop intense pulmonary and gastrointestinal toxicity. Diffuse interstitial pneumonitis and cerebral edema are the main cause of death.Nickel is a common sensitizing agent with a high prevalence of allergic contact dermatitis. Nickel and nickel compounds are well recognized carcinogens. However, the identity of the nickel compound or compounds, which cause the increased risk of cancer, remains unclear. Currently, there are little epidemiological data to indicate that exposure to metallic nickel increases the risk of cancer, or that exposure to the carcinogenic forms of nickel causes cancer outside the lung and the nasal cavity.

formation of nickel-iron alloys. In contrast to the soluble nickel salts (chloride, nitrate, and sulfate), metallic nickel, nickel sulfides, and nickel oxides are poorly water-soluble. Nickel carbonyl is a volatile liquid at room temperature that decomposes rapidly into carbon monoxide and nickel. Drinking water and food are the main sources of exposure for the general population with

Table 1 .

 1 1Trace metal concentrations (µg/kg DW) in marine sediments with different world areas

	Location	Cu	Zn	Pb	Cd	Cr	References
	Gulf of Aden	3.1-27	13-51	5.3-23	0.3-2.6	5.7-25	Mostaf et al, 2009
	Marmara Sea	12-30	34-50	21-31	0.02-0.5	27-61	Topcuozlu et al, 2004
	Suez gulf	2.7-12	25-58	12-42	1.2-3.9	2.5-12	Hamed et al,2006
	Moroccan estuarines	24-66	113-217		0.9-3.3		Cheggour et al, 2005
	Naples harbor	12-5743	17-7234	19-30	0.01-3	7-1798	Sprovieri et al, 2007
	Southern black Sea	15-119	24-141	12-69	-	13-238	Yucesoy et al,1992
	Jiaozhou Bay	17-34	80-110	24-49	0.1-0.3	41-88	Xyaoyu et al, 2010
	Hangzhou Bay	24.1		22.6	0.09		Che et al, 2003
	Changjiang Estuary	21.2		21.7	0.12		Che et al,2003
	Gulluk Bay	25	81	20	0.56		Dalman et al, 2005
	Texas Estuarines	1-20	40-295	8-150	0.2-45	3.5-1332 Sharma et al,1999
	Scheldt estuarines	1-2600	9-1500	4-455	0.1-20.2	7-202	Zwolsman et al, 1996
	Izmir bay	13-49	45-114	23-52			Balci et al, 1993
	Boston harbor	67	118	86		131	Bothner et al,1998

BIOACCUMULATION OF METALS IN LIVING ORGANISMS 1.3.1 Heavy metals on benthic organisms 1.3.1.1 Metabolism and biokinetic of metal in organisms

	Metals, being elements, cannot be broken down, although their chemical state may
	be altered and form compounds of varying toxicity to aquatic organisms depending upon
	the environmental conditions. Understanding the mechanisms that control bioavailability to
	benthic organisms has important implications for the cycling of metals through different
	compartments of coastal ecosystems. So far no one principle or method has proved

Table 1

 1

		.3Mollusk species are used as biomonitor in marine of Vietnam
	N	Especies	Region	References
	1	Saccostrea cucullata	Nha Phu, Khanh Hoa	Vinh L.T, 2005
	2	Paphia undulata	Son Tra, Da Nang	Mui L.T, 2008
	3	Anadara subcrenata	Nam O, Xuan Trieu	Mui L.T, 2008
	4	Meritrix spp	Central coastal zone	Tuyen B.C 2006
	5	Pletholophus Swinhoei	North Viet Nam	Wagner A, 2004
	6	Conus textile	Nha Trang	Nghi B.Q, 2011

Table 1 .

 1 : 4 Dominated bivalve species in Nha Trang bay(Nghi B.Q, 2001) Ocean currents of low/moderate velocity (<1knot) flow between the island driven by coastal winds, tides and regional oceanography. Sea surface temperatures usually peak is 30 O C (SW) falling to 24 O C during NW monsoon (An V.

	N	Species

rivers: the Cai river in the north and the Be river in the south, flow into Nha Trang bay, with the potential to influence water quality in the bay. The river catchments and riparian vegetation have been extensively modified for agriculture and aquaculture in recent decades. Apart from the inlands, which are located on the shelf in front of the bay, two large lagoons: Nha Phu lagoon is found just north of Nha Trang bay and a large lagoon Thuy Trieu is found just to the south with significant aquaculture and industrial activities.

Nha Trang bay covers about 50.700 hectares and contains 19 islands, which are located about from 1km to 15 km offshore, with an average deepth of 5-20m. They provide the topographic basis for a wide range of coastal and marine habitat types in relation to prevailing oceanographic conditions and gradients in mainland -oceanic influences. The diverse array of tropical habitats includes coral reefs, soft bottom communities, and sea grass beds, small stands of mangroves, sandy beaches and rocky shores. Picture 2.1 Map of Nha Trang Bay, Khanh Hoa province, Viet Nam

2.2 ENVIRONMENTAL STATE

The coastal province is under seasonal influence of the SW and NW monsoons.

They have effect of tropical weather on the oceannography, particularly water temperatures, circulation patter and mosoon winds. The climate is classed as humid tropical and is characterized by a relatively 6 months of dry season followed by a 6 months of rainy season with irregular rainfall. Rainfall generally occurs between November and April and the dry season is from May through October. height) generated by the NW monsoon and typhoons in the South sea impact on the sublitoral communities most years. Wave energy during SW monsoon is usually more moderate <1m high.

Table 2 .

 2 8 Heavy metal concentrations of sediment from Nha Trang samples(Son P, 2007

	2, DO: 6.40-7.04 mg/l,

increase in sediment samples collected in

Van Phong coastal (Khanh Hoa province)

. The concentration of Cu was 4.2-5.4mg/kg DW in the year of 2001 (Vinh L.T, 2002) but it's was measured about 844.3mg/kg DW in 2010 (Du H.T, 2010). In coastal sediment, there was a considerable rise of the concentration of Pb from 5.7mg/kg DW in 2001 to 25.6mg/kg DW in 2010. In contract, the concentration of As increased sharply, more than twice from 3.9 mg/kg DW in 2001 to 8.6 mg/kg DW in 2010 (Du H.T, 2010). Chemical characteristics of sediment from Nha Trang Bay are showed in table 2.7, 2.8 and 2.9.

Table 2 .

 2

	Times	Season	Zn (µg/g)	Cu (µg/g)	Pb (µg/g)	As (µg/g)	Cd (µg/g)
	Jun1996	D	6,33	8,68	0,19	18,04*	ND
	Oct1996	R	6,44	6,19	0,7	5,57	0,2
	Sep1996	R	0,97	1,68	0,07	7,1	0,1
	Mar1998	D	50,8	26	12,6	3,9	<0.05
	Feb1999	D	61,73	12,26	28,11	6,26*	0,38
	Jun1999	R	97,24	14,62	29,55	8,33*	0,04
	Feb2000	D	99,5	18,3*	36,7*	8,5*	4,8*
	Aug2000	R	57,53	10,23	26,09	2,32	Trace
	Feb2001	D	52,3	10,6	33,4*	4,05	0,21
	Aug2001	R	55,99	12,93	27,14	16,07*	0,104
	Feb2002	D	24,8	5,37	15,63	1,53	0

9 Concentration of metal (µg/g DW) in surface sediment at period 1996-2011 (MOSTE, 2010) **Higher value than SEL: Severe effect level (ppm) 2.

2.3 Biodiversity of bivalve molluscs in the bay of Nha Trang A

  total of 490 species of molluscs in 164 genera from 62 families in 2 classes gastropoda (429 species) and bivalvia (68 species) were recorded in Nha Trang Bay. This represents 61% of the total species recorded in Khanh Hoa province(Nghi B.Q, 2011).Saurin, 1959 published many species of Pyramidellidae sp. in Khanh Hoa province, of which 197 species were recorded in Nha Trang Bay. Many species in this family are ectoparasites, feeding on the body fluids of various invertebrates, mainly polychaete worms and other molluscs. The majority of pyramidellids live in subtidal to deep waters and prefer muddy or sandy substrates where annelid worms are abundant(Loi T.N, 1965).

Compared with some other areas, species composition of molluscs in Nha Trang Bay was more abundant than Cat Ba -Ha Long Bay (372 species) (Duc N.X, 2001) and Tonkin

Gulf (470 species, Cai Yingya et al., 1988)

.

Differences in geographical location and substrate types (mainland-oceanic gradient) are the major factors contributing to differences in distribution of molluscs. In the littoral, oysters were most numerous on rocks, developing into an oyster belt in the intertidal. Some species of families Littorinidae, Patellidae and Turbinidae were most strongly distributed in the lower littoral zone, particularly on soft substrates, with 27 species recorded in Cua Be estuary in 1999. In the deeper sea, species composition is more abundant and more diverse. In the coral reefs, molluscs have evolved many modes of life, free-living, boring in rocks or corals, in sandy bottom nearby reef. Some specific regional especies listed as below:

Anadara antiquata (Linnaeus, 1758)

	Genus: Anadara
	Vietnam's name: Sò lông
	The size structure:
	Shell inequivalve, solid, inequilateral, obliquely ovate and elongate in outline, with
	an extended posteroventral part. Umbones much inflated, situated rather forwards, cardinal
	area narrow and elongate. About 40 radial ribs (35 to 44) at each valve; ribs usually with a
	narrow median groove on top, most visible towards the anterior ventral margin of valves in
	mature specimens. Periostracum coarse and velvety often eroded on umbones. Internal
	margins with strong crenulations corresponding with the external radial ribs. No byssal
	Classification:
	Phylum: Mollusca
	Class: Bivalvia
	Subclass: Pteriomorphia
	Order: Arcoida
	Superfamily: Arcoidea
	Family: Arcidae

gape. Colour: outside of shell greyish white, often stained darker grey on umbonal and posterior areas; periostracum dark brown. Inner side is white, some times light yellow in the umbonal cavity.

Table 3 .

 3 The bivalve mollusk and sediments were collected twice a year during the dry season and rainy season of 2 years 2012 -2013 from 4 locations of the bay in low tide time of the day as shown in Table3.2.

	1 Localities of sampling sites

Table 3 .

 3 4 Mollusk species and collected numbers of them during 2012/2013

	N	Species	TD		ND	BT	CL
			D	R	D	R	D	R	D	R
			2012/2013	2012/2013	2012/2013	2012/2013	2012/2013	2012/2013	2012/2013	2012/2013
		Anadara antiquate	10/15	6/10	11/10	15/17			9/8
		Crassostrea rivularis	15/10	10/5			8/10	2/5
		Glauconoma virens			3/5	6/10	10/15	15/10	15/20	15/12
		Geloina coavans	3/10	13/15	9/10	6/10			5/10	10/16
		Katelysia hiantina	9/15	15/13	17/20	10/7	20/18	20/14	18/20	12/15
		Lingula unguis	15/10	15/10			15/20	10/8	20/23	15/10
		Laternula anatine					2/5	5/2	5/10	5/3
		Perna viridis	20/15	10/5
		Solen rivularis					10/15	2/5	15/11	15/13
		Tapes literatus			15/11	10/8

  B3 sample was analyzed as a quality assurance measure for metal in tissue. The sample has a recommended concentration of2.11 (1.42-2.51) µg As/g of tissue.Agreements with the analyzed concentrations were 2.43 ± 0.78 for 3 analyses. Again, measured values were in general agreement with certified values with a good agreement for Cu, Pb, Zn and Fe where in reference sample, recovery ranged from 95.87 % for zinc to 116.23 % for copper (Appendix 4.2).

	The MA-
	agency, AQCS, Laboratory Seibersdort, A-1400 Vienna. The Standard Reference Material
	MA-B-3 was prepared from Lyophilised Tissue from the coast of Monaco. The values
	obtained for the reference material were in comparison with the certified values (Appendix
	4.2).

Table 3 .

 3 [START_REF] Thị | Accumulation of lead and copper in some bivalvia mollusks species from coastal water of Da Nang. Tạp chí Khoa học và Công nghệ, Đại học Đà nẵng[END_REF] shows classification of Igeo according toMuller, 1996)

Table 3 .

 3 7 The 7 grades of classes Igeo

		Igeo value	I geo class Designation of sediment quality
	1	>5	6	Extremely contaminated
	2	4-5	5	Strongly to extremely contaminated
	3	3-4	4	Strongly contaminated
	4	2-3	3	Moderaly to strongly contaminated
	5	1-2	2	Moderaly contaminated
	6	0-1	1	Uncontaminated to moderaly contaminated
	7	0	0	Uncontaminated
		If Igeo = 6: 100 fold enrichment about background value

Table 3

 3

	.8 Sediment quality criteria

  reported that MPI values (based on Cu, Pb, Zn, Ni, Cr and Cd) in the bivalve Donax trunculus ranged from

	approximately 3.4 to 3.8 and from approximately 6.8 to 7.8 for unpolluted and polluted
	areas, respectively, from the southern Spanish Atlantic coast.
	The MPI values for mollusks reported in this study broadly agree with the MPI
	value (based on metals Fe, Mn, Zn, Cr, Cu, Pb, Ni, Cd and Ag) range of 10.5-25.1
	reported for the mussel Mytilus edulis from the coast
	Metal/shell weight indices (MSWI)

Table 3 .

 3

			9 The factors represent for accumulation of chemical in organisms
	N	BF	Definition
	1	BAF	Bioaccumulation factor: threshold for soil or sediment
	2	BCF	Bioconcentration factor: for inorganic in surface water
	3	BSAF	Biota to soil on sediment accumulation factor
	4	BMF	Biomagnification factor
	5	TMF	Trophic Magnification factor
	6	R BAC	Bioaccumulation index for assessing contaminats of interest
		Biota sediment accumulation factor
		Acording to Szeferet al., 1999, Abdallahet al., 2008, BSAF is calculated as
	equation like:

SLV. Chemical could be bioaccumulate threat to humant that consume aquatic organism.

Table 4 .

 4

			1 pH measurement of sediment samples
	pH KCl	TD	ND	BT	CL
	Mar 2012	7.43	7.43	7.43	7.4
	Sep 2012	7.26	7.2	7.14	7.3
	Mar 2013	7.47	7.44	7.58	7.53
	Sep 2013	7.24	7.09	7.04	7.17
	pH H 2 O	TD	ND	BT	CL
	Mar 2012	7.56	7.55	7.48	7.52
	Sep 2012	7.27	7.36	7.21	7.4
	Mar 2013	7.63	7.58	7.68	7.54
	Sep 2013	7.34	7.21	7.46	7.34

Table 4

 4

		.2 Percentage of total organic carbon TOC (%) in sediment samples
	Locations	TD	ND	BT	CL
	Mar2012	9.98	18.47	3.08	3.03
	Sep2012	9.06	27.06	10.4	3.14
	Mar2013	6.84	10.01	3.16	1.71
	Sep2013	6.45	19.64	8.26	3.17

Average values of sediment organic carbon (%dry weight) are showed in table 4.2, and the observed concentration of CL

(1.71-3.17)

was lowest when compared to that observed in other regions (TD, ND, and BT) that ranges from 3.08 to 27.06%. The values of organic cacbon obtained in this study area are comparatively higher than those reported (2.78%) for various tidal and estuarine regions of southeast coast of India

(Achyuthan and others, 2002)

. Relatively lowest concentration of organic cacbon in sediments of CL is due to and mixing process in sediment-water interface and also due to high microbial activity in this region, which use the organic matter as nutrient source and removes it from the systemas CO 2 . All the sediement collected showed relatively higher organic carbon values in the rainy season due to the nature of sedimentation in this season

Table 4 .

 4 3.

		Table 4.3 Bulk density (g/cm 3 ) of sediment samples
	Locations	TD	ND	BT	CL
	Mar2012	0.57	0.25	1.19	1.20
	Sep2012	0.62	0.05	0.55	1.18
	Mar2013	1.23	0.57	0.68	1.50
	Sep2013	0.80	0.59	1.16	1.17

Table 4 .

 4 4 Percent moisture contents in sediment samples Moisture variation in sediment samples was noted as low for all of samples and most likely due to variations in location at the 4 times of sampling. Figure4.12 is showed a trend of moisture content in sediment samples during2012-2013.

	Locations	TD	ND	BT	CL
	Mar2012	9.98	18.47	3.08	3.03
	Sep2012	9.06	27.06	10.4	3.14
	Mar2013	11.62	16.6	4.52	4.05
	Sep2013	13.77	23.56	11.62	4.4
	In general, rainy seasonal sediment contained the higher moisture of the dry
	sediment with a range of 9.06 to13.77% in TD; 23.56-27.06% in ND; 10.4-11.62% in BT
	and 3.14-4.4% in CL in rainy season. A range of 3.08-4.52% in BT and CL with moisture

measurements in dry season were very low moisture contents. Sediments in ND and BT had a grand difference of moisture between two seasons, but not for TD and CL sediments. Sep2013

Figure 4.12 Trend of moisture content in sediment samples during 2012-2013 4.1.6 Acid volatile sulfide (AVS) Only one measurement was obtained for the AVS and Bicarbonat contents in September 2013. The acid volatile sulfide (AVS) concentrations (mg/kg) and Bicarbonat contents in the sampled sediments are listed in

Table 4 . 5 Table 4 .

 454 5 The AVS and Bi-carbonat content in the sampled sediments.

	Parametres	TD	ND	BT	CL
	AVS (mg/kg)	0,168	0,416	0,243	0,266
	Bicarbonat (mgdl/100g)	0,446	0,786	0,488	0,457
	Bicarbonat (%)	2,79	4,79	2,97	2,72

Table 4

 4

			.6 Sediment type classification
		TD	ND	BT	CL
	Sediment	Silt clay	Silt clay	Clayely silt	Clayely silt
	type
	pH	Neutral medium Neutral medium Neutral medium Neutral medium
	OC	Lighter organic	Lighter organic	Lighter organic	Lighter organic
		matter	matter	matter	matter
	Moisture	Medium	High	Low	Low
	AVS	Low	Low	Low	Low
	4.2

METAL CONCENTRATIONS IN SEDIMENT SAMPLES 4.2.1 Ranges of heavy metal concentrations

	For remaining heavy metals in sediments in both two seasons, metal concentrations
	in the sediments ranked in the following order: Aluminium> iron > zinc > copper >
	chromium > asennic > cadmium. In particulier, the average concentrations of heavy metals
	in sediments obtained from 4 locations were compared in sequential order as show in Table
	4.7.

Table 4 .

 4 7 Sequential orders of heavy metals in sampling sites

	Sites	Seasons	Metals in decreasing orders
	TD	Rainy	Zn > Cr > Cu > As > Cd
		Dry	Zn > As > Cr > Cu > Cd
	ND	Rainy	Zn > Cr > Cu > As > Cd
		Dry	Zn > Cr > Cu > As > Cd
	BT	Rainy	Zn > Cu > Cr > As > Cd
		Dry	Zn > Cu > Cr > As > Cd
	CL	Rainy	Zn > As > Cu > Cr > Cd
		Dry	Zn > Cu > Cr > Cd > As

Table 4 .

 4 8 and 4.9 gives the values for elements tested in surface sediment collected in dry and rainy season for each parameter (where applicable). The element concentration was study with referred to the values of effect range (ERL and ERM) and effect levels (LEL and SEL). In the sediment, the recorded metal values did not reach biological effect levels (ERL), concentrations proposed by[START_REF] Long | Incidence of adserve biological effect within ranges of chemical concentrations in marine and estuarine sediment[END_REF]

Table 4 .

 4 8 Values for metals (mg/kg DW)in surface sediment collected in dry season

	Metals	TD	ND	BT	CL
	As	2,49-13,09*	4,39-16,95*	2,09-5,82	0,15-5,54
	Cd	0-1,15*	0-0,94*	0-2,20*	0-2,41*
	Cr	2,33-6,83	5,94-21,81	4,92-23,43	10,84-10,97
	Cu	2,95-7,95	15,31-21,93	0-43,24*	5,49-11,34
	Zn	36,93-71,43	53,96-81,51	70,61-90,94	40,99-49,30

*: Values above LEL (mg/kg DW)

The sediment samples that collected in dry season from TD had arsenic concentrations 13,09mg/kg DW in between the LEL-SEL values. Similar situations were with copper concentrations in sediments collected at BT (43,24mg/kg DW), that were more than two times higher when compared to LEL value. Only zinc concentration in sediment collected in dry season had all values below the LEL value with the mean value of 36,93-90,94mg/kg DW.

Table 4 .

 4 [START_REF] Van | Study concentration of heavy metals cadmium (Cd) and lead (Pb) in clam Corbicula sp. from estuarine, in Da Nang city -Tạp chí Khoa học và Công nghệ[END_REF] Values for metals (mg/kg DW) in surface sediment collected in rainy season

	Metals	TD	ND	BT	CL
	As	4,65-6,73	5,71-14,36	14,10-15,09*	9,09-14,87*
	Cd	0-2,07*	1,14-1,93*	0-1,72*	9,02-1,69*
	Cr	7,61-13,49	16,75-41,96*	5,81-23,14	7,73-19,83
	Cu	0-7,36	28,70-91,22*	19,04-53,94*	8,00-99,21*
	Zn	50,66-68,36	70,62-99,90	112,31-126,03	59,76-65,65
	*: Values above LEL
	The sediment samples that collected in rainy season from BT and CL had arsenic
	concentrations in between the LEL-SEL values (6 -33mg/kg DW), with mean
	concentration values were more than two times higher when compared to LEL value
	(14,10 and 9,09mg/kg DW respectively). Similar situations were with copper

Table 4 .

 4

				10 Interactions between metals in rainy season
	N	As	Cd	Cr	Cu	Zn	Fe	Al	Ni
	As	1,0000
	Cd -0,8223 1,0000
	Cr	-0,1312 0,2356	1,0000
	Cu 0,7467	-0,3853 0,5039	1,0000
	Zn	0,5915	-0,3541 0,7201	0,9443	1,0000
	Fe	-0,3489 0,3957	0,9746	0,3024	0,5464	1,0000
	Al	-0,1806 0,1776	0,9836	0,4060	0,6633	0,9739	1,0000
	Ni	-0,1552 0,2013	0,9951	0,4550	0,6944	0,9774	0,9966	1,0000

Table 4 .

 4 11 Interactions between metals in dry season

		As	Cd	Cr	Cu	Zn	Fe	Al	Ni
	As	1,0000
	Cd -0,7048 1,0000
	Cr	-0,5329 0,0372	1,0000
	Cu -0,5388 0,4726	0,8063	1,0000
	Zn	-0,5379 0,0836	0,9976	0,8452	1,0000
	Fe	-0,6656 0,1341	0,9839	0,7778	0,9783	1,0000
	Al	-0,8300 0,7642	0,6693	0,8941	0,7056	0,7205	1,0000
	Ni	-0,2678 -0,4031 0,8775	0,4361	0,8448	0,8507	0,2511	1,0000
	Mo -0,8418 0,9554	0,0823	0,3741	0,1114	0,2177	0,7403	-0,2850

Table 4 .

 4 [START_REF] Dallinger | Strategies of metal detoxification in terrestrial invertabrates[END_REF] The interactions between metals and physicochemical characteristics of sediments collected from Nha Trang bay and

	adjacent areas

Table 4 .

 4 [START_REF] Tu | Concentrations of trace elements in Meretrix spp. along the coasts of Vietnam[END_REF] Metals in correlation with the physico-chemical factors in Tan Dao station

		pH	Moisture TOC	>0.063mm <0.063mm As	Cd	Cr	Cu	Zn	Fe
	pH	1
	Moisture	-0,6699	1,0000
	TOC %	0,9930	-0,5772	1,0000
	>0.063mm 0,9993	-0,6976	0,9877	1,0000
	<0.063mm -0,9993	0,6976	-0,9877	-1,0000	1,0000
	As	-0,3201	0,9178	-0,2056	-0,3559	0,3559	1,0000
	Cd	0,1148	-0,8145	-0,0037	0,1525	-0,1525	-0,977	1,0000
	Cr	0,5907	0,2034	0,6822	0,5596	-0,5596	0,5753	-0,733	1,0000
	Cu	0,7909	-0,0755	0,8578	0,7671	-0,7671	0,3266	-0,517	0,9610	1,0000
	Zn	0,8016	-0,0930	0,8668	0,7783	-0,7783	0,3099	-0,501	0,9559	0,9998	1,0000
	Fe	0,7878	-0,0704	0,8552	0,7638	-0,7638	0,3313	-0,521	0,9623	1,0000	0,9997	1,0000
	Al	0,9940	-0,5844	1,0000	0,9891	-0,9891	-0,214	0,0052	0,6756	0,8532	0,8623	0,8506

Table 4 .

 4 14 Metals in correlation with the physico-chemical factors in Ngoc Diem station

		pH	Moisture TOC	>0.063	<0.063	As	Cd	Cr	Cu	Zn	Fe
					mm	mm
	pH	1,0000
	Moisture	0,7593	1,0000
	TOC	-0,0735	0,5931	1,0000
	>0.063mm 0,3380	0,8690	0,9138	1,0000
	<0.063mm -0,3380	-0,8690	-0,9138	-1,0000	1,0000
	As	0,9802	0,8733	0,1256	0,5178	-0,5178	1,0000
	Cd	0,1086	0,7293	0,9834	0,9723	-0,9723	0,3035	1,0000
	Cr	0,1052	0,7270	0,9840	0,9715	-0,9715	0,3003	1,0000	1,0000
	Cu	0,4260	0,9122	0,8709	0,9955	-0,9955	0,5969	0,9456	0,9445	1,0000
	Zn	-0,8247	-0,2581	0,6247	0,2536	-0,2536	-0,6962	0,4728	0,4757	0,1604	1,0000
	Fe	0,2413	0,8147	0,9501	0,9949	-0,9949	0,4288	0,9909	0,9905	0,9808	0,3500	1,0000
	Al	-0,6779	-0,0364	0,7830	0,4628	-0,4628	-0,5187	0,6572	0,6597	0,3763	0,9749	0,5499

Table 4 .

 4 15 Metals in correlation with the physico-chemical factors in Binh Tan station

		pH	Moisture TOC	>0.063mm <0.063mm As	Cd	Cr	Cu	Zn	Fe
	pH	1,0000
	Moisture	0,7857 1,0000
	TOC	0,2186 0,7754	1,0000

Table 4 .

 4 16 Metals in correlation with the physico-chemical factors in Cam Lam station

		pH	Moisture TOC	>0.063mm <0.063mm As	Cd	Cr	Cu	Zn	Fe
	pH	1,0000
	Moisture % -0,7485	1,0000
	TOC %	-0,2641	-0,4419	1,0000
	>0.063mm -0,9936	0,6688	0,3713	1,0000
	<0.063mm 0,9936	-0,6688	-0,3713	-1,0000	1,0000
	As	-0,9509	0,5066	0,5496	0,9798	-0,9798	1,0000
	Cd	0,7806	-0,9988	0,3967	-0,7050	0,7050	-0,5489	1,0000
	Cr	0,4772	0,2255	-0,9736	-0,5734	0,5734	-0,7257	-0,1768	1,0000
	Cu	0,5728	0,1148	-0,9419	-0,6617	0,6617	-0,7983	-0,0652	0,9937	1,0000
	Zn	-0,0655	-0,6126	0,9797	0,1778	-0,1778	0,3711	0,5725	-0,9082	-0,8555	1,0000
	Fe	0,9283	-0,4484	-0,6037	-0,9644	0,9644	-0,9978	0,4923	0,7697	0,8365	-0,4318	1,0000
	Al	0,9052	-0,9594	0,1708	-0,8514	0,8514	-0,7293	0,9722	0,0585	0,1701	0,3647	0,6823

HEAVY METALS IN MOLLUSK TISSUES 4.3.1 Concentrations of heavy metals between shell and tissue

	Concentrations of metals in shell and tissues of these 7 species, which collected on
	the first time (March 2012) were showed in

Concentration of metals in shell and tissue of G.coavans

		Concentration of metals in shell and tissue of G.virens
		mg/kg DW mg/kg DW
	1000
			Tissue Tissue
			Shell Shell
	100
	10
	Concentration of metals in shell and tissue of L.anatine
	mg/kg DW 1
	1000	As As	Cd Cd	Cr Cr	Cu Cu	Pb Pb	Zn Zn	Fe Fe	Al Al
		Tissue
	0	Shell						Metals Metals
	100
	10 Figure 4.22 Concentrations of metals in shell and tissue of G.coavans Figure 4.20 Concentrations of metals in shell and tissue of G.virens
		Concentration of metals in shell and tissue of L.unguis
	1	mg/kg DW
	1000	As	Cd	Cr	Cu	Pb	Zn	Fe	Al
			Tissue
	0		Shell						Metals
	100
	Figure 4.19 Concentrations of metals in shell and tissue of L.anatine
	10
	1
		As	Cd	Cr	Cu	Pb	Zn	Fe	Al
	0								Metals
	Figure 4.21 Concentrations of metals in shell and tissue of L.unguis

Concentration of metals in shell and tissue of C.rivularis

	Concentration of metals in shell and tissue of K.hiantina
	mg/kg DW
	100
	Tissue
	Shell
	10
	1
	As	Cd	Cr	Cu	Pb	Zn	Fe	Al
	0							Metals
	Figure 4.24 Concentrations of metals in shell and tissue of K.hiantina
	Concentration of metals in shell and tissue of A.antiquate
	mg/kg DW mg/kg DW
	1000
		Tissue Tissue
		Shell Shell
	100
	10
	1
	As As	Cd Cd	Cr Cr	Cu Cu	Pb Pb	Zn Zn	Fe Fe	Al Al
	0							Metals Metals
	Figure 4.23 Concentrations of metals in shell and tissue of C.rivlaris

  contents. Results from the analyses of 5 metals As, Cd, Cr, Cu and Zn in these species are given inFigures 4.26 as below. varied during dry and rainy seasons. The maximum value of As, Cd were found at ND and Cu, Zn at BT during rainy season, while the highest value of Cd, Cr, Cu, Zn were recorded at CL during dry season. The minimum values of Cd, Cr and Zn were found at station CL in rainy season, while Cu, Cd, Cr and Zn were found at BT during rainy season.

	Contents of m etals in L.anatina at BT		Contents of m etals in L.anatina at CL
	rainy season		rainy season
	Contents of m etals in G.virens at CL		Contents of m etals in G.virens at BT
	rainy season		rainy season
		As		As
		Cd		Cd
		Cr		Cr
		As Cu		As Cu
		Cd Zn		Cd Zn
		Cr		Cr
		Cu		Cu
		Zn		Zn
	Contents of m etals in L.anatina at BT		Contents of m etals in L.anatina at CL
	dry season		dry season
	Contents of m etals in G.virens at CL		Contents of m etals in G.virens at BT
	dry season	As	dry season	As
		Cd		Cd
		Cr		Cr
		As Cu		As Cu
		Cd Zn		Cd Zn
		Cr		Cr
		Cu		Cu
		Zn		Zn
	Figure 4.27 Contents of metal in soft tissue of Laternula anatine
	On a seasonal scale, Zn and Cu showed their highest values during rainy season at
	Figure 4.26 Contents of metal in soft tissue of G.virens both locations BT and CL (61.29 and 90.60mg/kgDW for Zn; 17.72 and 9.22mg/kg DW
	Generally, the highest values of heavy metals were observed during rainy season for Cu, respectively) where the highest values of As were exhibited during dry season.
	and the lowest during dry season, except Cu in CL. Mean concentrations of Zn found in (6.58 and 6.84mg/kg DW ).The levels of Cr varied from 2.22 and 2.64 during rainy season
	mollusks were high in both seasons. Moreover, the concentrations of heavy metals in the to 1.62 to 13.42mg/kg DW during dry season, respectively. The maximum values of Cr

G.virens of the ND could be arranged in the following sequence: As>Cd>Zn>Cr>Cu.

From these results, the local distribution of metals in mollusk tissue of G.virens at different stations also recorded 13.42 during rainy season at CL location and their minimum values 1.62 during dry season at BT.

of m etals in K.hiantina at BT rainy season

  Furthermore, the concentrations of Zn in B.rana were higher than in that in P.viridis during dry period. The comparative study showed that the accumulation of heavy metals in 2 mollusk tissues were significantly higher than in sediment samples.

			Contents of metals in T.literatus at ND
			rainy season
	Contents of m etals in K.hiantina at TD		Contents of m etals in K.hiantina at TD
	rainy season		dry season
				As
		Cd		Cd
		Cr		Cr
		Cu As		Cu As
		Zn Cd		Cd
		Cr		Cr
		Cu		Cu
		Zn		Zn
	Contents of m etals in K.hiantina at ND		Contents of m etals in K.hiantina at ND
	rainy season		dry season
		As		As
		Cd		Cd
		Cr		Cr
		Cu		Cu
	Contents of metals in P.viridis at TD	Zn	Contents of m etals in B.rana at ND	Zn
	dry season		dry season
		As		As
		Cd		Cd
		Cr		Cr
	Cu Zn Contents As	Contents	Cu Zn
		Cd
		Cr
	Cu Figure 4.34 Contents of metal in soft tissue of B.rana and P.viridis Zn
	High differences of concentration of heavy metals in 2 species of Gastropode
	B.rana and Bivalve P.viridis were detected in rainy season for comparision. In gastropode

Zn

Figure 4.33 Contents of metal in soft tissue of Tapes literatus In the present study, As content of tissue of T. literatus ranged from 17.98mg/kgDW in rainy season to 6.85mg/kg DW at ND and Zn concentration were decreased from 4.08.10 to 35.01mg/kg DW from rainy to dry season. The high concentration of Cd content in rainy season may be due to the precipitation of industrial leaching from north part of the bay.

B.rana tissues, concentrations of Zn and As were highest compared to Cu and others, but in bivalve P.viridis, these concentraions of As, Cr, Cu and Zn were similar.

of m etals in K.hiantina at BT dry season

	Contents of m etals in K.hiantina at CL	Contents of m etals in K.hiantina at CL
	rainy season	dry season
	As	As
	Cd	Cd
	Cr	Cr
	Cu	Cu
	Zn	Zn
	Figure 4.35 Contents of metal in soft tissue of K. hiantina
	Samples contained arsenic concentrations that ranged from 0.01 to 6.7 mg/kg DW
	in rainy season and lower than concentrations of this metal in tissue at dry season 5.85-
		As
		Cd
		Cr
		Cu
		Zn

Table 4 .

 4 .[START_REF] Us Epa | Microwave assisted acid digestion of siliceous and organically based matrices[END_REF] show on the whole, the highest, moderate and lowest MPI ranges for 7 metals: As, Cd, Cr, Cu, Zn, Fe and Al. The averages occurred during the dry and rainy season for 4 locations are respectively. 17 MPI during rainy season for 4 locations of sampling

		Rainy season	TD	ND	BT	CL
	1	Katelysia hiantina	3,72	2,62	7,44	4,46
	2	Glauconoma virens		1,17	5,05	1,61

Table 4 .

 4 18 MPI during dry season for 4 locations of sampling According to classification of Usero et al, 2005 for MPI and in comparing these values from those in Table 4.17 and 4.18, it is clear that all of the sediment of Nha Trang bay are not contaminated by these trace metals of various magnitudes at 4 locations. During rainy season, BT is severely contaminated with highest MPI values (5.87), where as ND is lastly contaminated with moderate MPI values 2.35 and CL is moderately contaminated with low MPI values 3.78 for these metals. The highest MPI values found in BT were plausibly related to discharge of effluents from domestic.

	N Dry season	TD	ND	BT	CL
	1	Katelysia hiantina	3,83	4,14	4,09	4,16
	2	Glauconoma virens			3,57	8,52
	3	Laternula anatine			1,76	2,49
	4	Lingula unguis			2,75	0,74
	5	Geloina coaxans	8,71	3,35
	6	Crassostrea rivularis	11,11		12,04
	7	Anatina antiquate	6,70			8,89
	8	Tapes literatus		4,09
	9	Solens regularis			3,37	3,01

Table 4 .

 4 19 Moisture contents in mollusk tissues

	N	Species	Min	Max
	1	Katelysia hiantina	92,59%	93,91%
	2	Glauconoma virens	80,67%	84,99%
	3	Laternula anatine	56,25%	75,47%
	4	Lingula unguis	83,53%	91,63%
	5	Geloina coaxans	89,59%	96,19%
	6	Anatina antiquate	96,13%	98,58%
	7	Tapes literatus	73,99%	94,55%
	8	Solens regularis	62,23%	74,01%
	9	Perna viridis	92,68%	97,97%
	10	Crassostrea rivularis	92,21%	97,60%

Table 4 .

 4 20 The calculated condition index for seven species

	Species	CI min	CI max	Average
	K.hiantina	27.18	54.52*	38.58±9.0
	G.virens	44.94	105.52***	74.58±22.49
	G.coavans	23.85	43.13*	33.06±8.90
	L.anatine	78.12	116.67***	73.6±16.98
	L.unguis	18.66	63.22*	44.64±19.87
	A.antiquate	14.38	26.99*	22.64±1.75
	S.regularis	93.49	99.26**	95.87±3.01
	Note: *Condition index low
		*** Condition index high

Table 4 .

 4 21; 4.22; 4.23; 4.24) ranged from 22.64 (A.antiquate) to 95.67 (S.regularis) and was significant correlated with the metals in mollusk tissues. The matrix correlating the biometric parameters CI and pollutant concentrations is presented in tables bellow:

Table 4 .

 4 21 Correlation coefficients between biometric and metal content inK.hiantina

		CI	As	Cd	Cr	Cu	Zn	Fe	Al
	CI	1,000
	As	-0,274	1,000
	Cd	0,836	-0,526	1,000
	Cr	-0,142	-0,168	-0,268	1,000
	Cu	0,770	-0,563	0,668	-0,173	1,000
	Zn	0,700	-0,308	0,838	-0,315	0,372	1,000
	Fe	0,273	-0,426	0,376	-0,263	0,399	-0,048	1,000
	Al	0,008	-0,357	0,249	-0,335	0,086	-0,059	0,925	1,000
		A high correlation was observed between Cd and Cu, Zn levels in the clam
	K.hiantina (0.668 and 0.838). While a moderately negative correlation resulted between As
	and Cd, Cu metals in the clam (-0.526 and -0.563). Metal Cd, Cu and Zn concentrations in
	tissues also depicted statistically significant high correlations related to the condition index
	(r =0.836, 0.770 and 0.700 respectively). Condition index was negative correlated with As
	and Cr concentrations in the clam (r=-0.274 and -0.142). Condition index correlated
	positively low with Fe and Al (r=0.273 and 0.008, respectively). The clam may be a
	indicator for bioavailability for which the CI was independent of As and Cr concentration.

Table 4 .

 4 22 Correlation coefficients between biometric and metal contents in G.virens

		CI	As	Cd	Cr	Cu	Zn	Fe	Al
	CI	1
	As	-0,181	1,000
	Cd	0,446	-0,479	1,000
	Cr	-0,817	0,534	-0,378	1,000

Table 4 .

 4 23 Correlation coefficients between biometric and metal contents in L.anatine

		CI	As	Cd	Cr	Cu	Zn	Fe	Al
	CI	1,000
	As	-0,729	1,000
	Cd	0,269	-0,786	1,000
	Cr	0,093	0,520	-0,500	1,000
	Cu	0,795	-0,976	0,637	-0,513	1,000
	Zn	0,118	-0,690	0,988	-0,517	0,524	1,000
	Fe	0,624	-0,928	0,920	-0,351	0,831	0,850	1,000
	Al	0,551	0,019	-0,140	0,864	-0,019	-0,219	0,126	1,000

Table 4 .

 4 23 shows metal relationships with other metals in soft tissue of L.anatine by using t-test and probability differences. All metals measured in L.anatine gave highly correlations as Cu, Cd and Zn with Fe (r = 0.831, 0.920 and 0.850 respectively), Cr and Al (r=0.864), while As with Cd, Cu, Zn and Fe were negative correlation. On the other hand,

Table 4 .

 4 24 Correlation coefficients between biometric and metal contents in L.unguis

		CI	As	Cd	Cr	Cu	Zn	Fe	Al
	CI	1,000
	As	-0,728	1,000
	Cd	0,886	-0,436	1,000
	Cr	-0,167	-0,496	-0,563	1,000
	Cu	0,206	0,470	0,327	-0,699	1,000
	Zn	0,275	0,397	0,652	-0,994	0,690	1,000
	Fe	0,793	-0,498	0,947	-0,476	0,043	0,563	1,000
	Al	-0,107	-0,353	-0,556	0,892	-0,300	-0,890	-0,618	1,000

Table 4 .

 4 25 Correlation coefficients between biometric and metal contents in G.coavans

		CI	As	Cd	Cr	Cu	Zn	Fe	Al
	CI	1,000
	As	-0,657	1,000
	Cd	0,820	-0,236	1,000
	Cr	-0,677	0,560	-0,200	1,000
	Cu	-0,866	0,192	-0,918	0,497	1,000
	Zn	-0,731	0,348	-0,939	-0,007	0,732	1,000
	Fe	-0,681	0,165	-0,453	0,856	0,767	0,149	1,000
	Al	0,116	0,377	0,659	0,583	-0,415	-0,712	0,192	1,000

Table 4 .

 4 26 Correlation coefficients between biometric and metal contents inA.antiquate

		CI	As	Cd	Cr	Cu	Zn	Fe	Al
	CI	1,000
	As	0,996	1,000
	Cd	-0,775	-0,719	1,000
	Cr	0,333	0,411	0,339	1,000
	Cu	-0,128	-0,211	-0,528	-0,978	1,000
	Zn	0,702	0,640	-0,994	-0,438	0,616	1,000
	Fe	0,171	0,088	-0,756	-0,872	0,955	0,822	1,000
	Al	0,376	0,297	-0,878	-0,748	0,871	0,924	0,977	1,000

Table 4 .

 4 27 Correlation coefficients between biometric and metal contents in S.reguraris

		CI	As	Cd	Cr	Cu	Zn	Fe	Al
	CI	1,000

Table 4 .

 4 28 Comparision BSAF in shell and tissue of different mollusk species

	Species		As	Cd	Cr	Cu	Zn
	L.anatina	BSAF-S	0,55	4,74	0,01	1,10	5,56
		BSAF-T	5,45	3,99	0,01	3,28	1,51
	L.unguis	BSAF-S	0,72	1,70	0,02	0,38	2,26
		BSAF-T	13,41	5,89	0,02	1,29	65,46
	G.virens	BSAF-S	0,54	2,54	0,02	0,27	0,22
		BSAF-T	2,36	5,25	0,34	2,49	1,83
	G.coaxans	BSAF-S	0,83	4,12	0,19	0,47	0,64
		BSAF-T	1,17	1,76	0,04	0,57	3,81
	C. rivularis	BSAF-S	0,46	0,25	0,10	0,40	0,69
		BSAF-T	10,57	4,15	7,51	14,71	46,68
	K. hiantina	BSAF-S	0,19	0,23	0,02	0,12	0,08
		BSAF-T	0,56	0,76	0,01	0,39	0,64
	A. antiquate BSAF-S	0,42	0,38	0,02	0,07	0,08
		BSAF-T	0,29	1,95	0,01	0,15	0,58

Table 4 .

 4 11, 7.33±2.41 and 34.38 ± 6.54 mg/kg DW in the dry season. Dry seasonal depending on As, Cr, Al pollutions as well as rainy season depending on Cd, Cu, Zn, Fe pollutions. The low concentrations in dry season could be explained by the metal dilution in the organism which caused by weight increase resulted and a metal reconcentration in clam.Nevertheless higher concentrations of Al and Fe in the K.hiantia population can be found in areas of estuary BT at dry season; also a higher accumulation of As goes in the 4 station in dry has shown that physico-chemical factors and a stable metal concentration modify the concentration in species of the K.hiantina. Physiological processes linked to metabolic functions, especially reproduction -gametogenesis and emission of eggs, described byCheggour, 1988 seem to play a key role in determining seasonal variations in the level of metals present in the tissue of the organisms. This laying period is followed by rainfall mating characterized by high metal levels. At the beginning of dry period metal concentrations clearly peak. 30 Calculation of MSWI for K.hiantina It is evident that the bioaccumulated Zn in K.hiantina from the TD and BT and Cu from the ND, during both dry and rainy, demonstrated the highest values among the total samples analyzed. Data of the study showed higher mean Cd, Cu and Zn shell/weight indices in mollusks in rainy season in comparison to dry season, excluding As and Cr. Ratios of MSWI rainy season/rainy of As and Cr were 0.83/3.39 and 0.7/1.4 respectively. In clams, higher mean Pb shell/weight indices were found in rainy season in all the examined areas (p<0.001), where as no significant seasonal variation was observed regarding Pb in dry season. Concerning Fe and Al, no specific seasonal trends were recorded. The metal/shell weight indices of the mollusks were found to correlate with the metal concentrations in the sediment. Regarding the clam K.hiantina, its Zn/shell weight indices are associated with Zn concentrations in the fraction of sediment.

	4.5.2 Metal/shell weight indices (MSWI)
	Metal bioavailability was first calculated as metal concentrations in soft tissues
	(mg/kg DW) and shell weigh of organisms. Biometric data of samples for analysis was
	presented in Table 3.22. Mean shell height of clams K.hiantina varied from 5.5 to 6.2 cm,
	total weight from 64.0 to 81.6 g/clam and body mass from 5.3-9.9 g/clam. Calculation of
	MSWI for K.hiantina is showed in table 4.30
			Dry season			Rainy season
	Sites	TD	ND	BT	CL	TD	ND	BT	CL
	As	0,50	0,95	1,34	0,60	0,03	0,00	0,51	0,29
	Cd	0,03	0,02	0,02	0,01	0,06	0,09	0,10	0,02
	Cr	0,23	0,11	0,06	0,99	0,21	0,27	0,11	0,11
	Cu	0,45	0,77	0,84	0,45	0,93	3,84	1,56	2,01
	Zn	2,42	2,88	3,76	2,71	5,00	4,17	6,95	3,35
	Fe	78,93	57,61	49,73	18,09	72,06	91,69	22,34	23,20
	Al	54,27	37,66	29,78	11,14	50,07	41,64	16,48	14,78
	MSWI	136,83	100	85,53	33,99	128,36	141,7	48,05	43,76
	Metal/shell weight indices were used to characterize each site and season
	depending on metal bioavailability.

  In general terms, metal shell-weight indices also indicate that metal bioavailability was very low in the CL sea close between dry and rainy season. MSWI values were 33.19 and 43.76, respectively. As an exception, chrome Cr bioavailability was high in CL in dry season. Occasionally, highest Fe, Cd and Cu bioavailability was found in ND at rainy season and Al in TD in dry season. Low Cd, Cu, Fe and Al bioavailability were usually found in CL in dry season and Cu and Zn in TD.

Table 4 .

 4 31 Bioaccumulation of heavy metal in clam K.hiantina Moderate potential of accumulation and Regular: Low potential of accumulation.BSAF is defined as the ratio of metal concentration in the mollusk to that in sediment. In this study, it was found that K.hiantina from ND, BT and CL sites showed higher BSAF values for As. The BSAF values were2.53, 7.42 and 46.16 respectively in dry season indicating that the mollusk contained much more metals than the sediment. It should also be noted that the concentration of As metals in the sediment and K.hiantina from there sites were lower in rainy season than dry season, as shown in Fugures 4.41.Similar pattern was also found for Zn in all 4 sites, although the BSAF values were higher than 2.0 in rainy season. This investigation also indicated that the K.hiantina were able to accumulate higher quantity of Cu and Cr than Cd, Mo and Ni. The BSAF values for Cd, Mo and Ni were lower than 1.0, which means limited ability of these heavy metals to be accumulated by the mollusk.The BSAF indicated a low potential bioaccumulation for Cd, Mo and Ni (BSAF<1) and a high potential of accumulation for As in dry season and for Zn in rainy season. Based on those results, it show that the magnitude of the heavy metal accumulations in tissues depend on the type of the heavy metals and seasonal.In conclusion, the clam K.hiantina has characteristics as sampling efforts, locational pollution classification according to MSWI, seasonal depending on metal pollution and could choice as biomonitor in Nha Trang bay, Khanh Hoa coastal.

	Metals		Dry season			Rainy season
		TD	ND	BT	CL	TD	ND	BT	CL
	As	0,45	2,53	7,42	46,16	0,09	0,00	0,48	0,42
	Cd	0,31	0,28	0,10	0,04	0,36	0,62	0,73	0,16
	Cr	0,39	0,06	0,03	1,07	0,21	0,09	0,06	0,19
	Cu	1,78	0,59	0,23	0,46	1,67	1,79	0,38	3,32
	Zn	0,77	0,42	0,48	0,64	18,29	6,52	4,03	7,51
	Mo	0,40	0,93	0,39	0,13	ND	ND	ND	ND
	Ni	0,31	0,06	0,12	0,03	ND	ND	ND	ND
	Fe	0,11	0,03	0,03	0,01	0,06	0,04	0,02	0,03
	Al	0,15	0,02	0,01	0,00	0,03	0,01	0,01	0,01
		Note: ND: not detected, Bold: High potential of accumulation, Bold Italic:

Table 4 .

 4 32 Daily trace metals intake EDI through K.hiantina consumption

		Max	Min	Average
	As	1,062	0,001	0,431
	Cd	0,086	0,006	0,038
	Cr	0,792	0,051	0,219
	Cu	3,507	0,359	1,198
	Zn	6,293	1,936	3,380

Table 4 .

 4 33 Daily trace metals intake EDI through G.virens consumption

		Max	Min	Average
	As	1,468	0,084	0,589
	Cd	0,230	0,001	0,086
	Cr	0,890	0,089	0,362
	Cu	3,093	0,066	1,546
	Zn	17,978	0,130	8,465

Table 4 .

 4 34 Daily trace metals intake EDI through L.anatina consumption

		Max	Min	Average
	As	1,168	0,240	0,805
	Cd	0,137	0,002	0,059
	Cr	2,293	0,277	0,850
	Cu	3,027	0,821	1,618
	Zn	15,477	3,672	8,502

Table 4 .

 4 35 Daily trace metals intake EDI through L.unguis consumption

		Max	Min	Average
	As	0,500	0,001	0,182
	Cd	0,063	0,000	0,025
	Cr	1,535	0,075	0,459
	Cu	1,236	0,136	0,628
	Zn	2,984	0,758	2,308

Table 4 .

 4 36 Daily trace metals intake EDI through G.coaxans consumption

		Max	Min	Average
	As	0,992	0,001	0,616
	Cd	0,210	0,008	0,091
	Cr	1,997	0,091	0,585
	Cu	0,898	0,148	0,642
	Zn	8,473	0,870	4,376

Table 4 .

 4 37 Daily trace metals intake EDI through A.antiquata consumption

		Max	Min	Average
	As	0,647	0,000	0,263
	Cd	0,053	0,004	0,023
	Cr	0,483	0,031	0,133
	Cu	2,138	0,219	0,730
	Zn	3,836	1,180	2,060

Table 4 .

 4 38 Daily trace metals intake EDI through S.regularis consumption

		Max	Min	Average
	As	0,876	0,296	0,646
	Cd	0,045	0,000	0,015
	Cr	2,014	0,238	0,834
	Cu	6,097	0,634	3,456
	Zn	6,517	4,674	5,849

Table 4 .

 4 39 Daily trace metals intake EDI through C.revularis consumption

		Max	Min	Average
	As	2,588	0,002	1,052
	Cd	0,211	0,015	0,092
	Cr	1,930	0,125	0,533
	Cu	8,552	0,875	2,921
	Zn	15,343	4,722	8,242
		Thus, moderate trace level consumption of bivalve from the Nha Trang bay is not
	harmful to humans for As, Cr, Cd, Cu and Zn, but due to high-level accumulations of
	metals in bivalve muscle, heavy consumption of bivalves from the Nha Trang bay may
	pose a health risk to consumers (human beings). Assuming a tolerable daily intake of
	metals per kg of body weight (USFDA

Table 4 .

 4 40to Table4.47 are showed the provisional THQ of As, Cr, Cd, Cu, and Zn recommended by the WHO/FAO and the USA for a reference weight of 60kg. It is evident from the table that target hazard quotient As, Cr, Cd, Cu and Zn from mollusks did not exceed the provisional THQ recommended by the WHO/FAO and the USA. However, the estimated Fe and Al THQ from the 5 species exceed the provisional THQ recommended by the WHO/FAO and the USA.

Table 4 .

 4 40 The target hazard quotient for K.hiantina

		Max	Min	Average
	As	0,02	0,00	0,01
	Cd	0,00	0,00	0,00
	Cr	0,01	0,00	0,00
	Cu	0,06	0,01	0,02
	Zn	0,11	0,04	0,06
	Fe	1,52	0,26	0,80
	Al	0,82	0,16	0,50

Table 4 .

 4 41 The target hazard quotient for G.virens

		Max	Min	Average
	As	0,033	0,002	0,013
	Cd	0,005	0,000	0,002
	Cr	0,020	0,002	0,008
	Cu	0,070	0,001	0,035
	Zn	0,406	0,003	0,191
	Fe	2,892	1,335	2,151
	Al	2,624	0,522	1,548

Table 4 .

 4 42 The target hazard quotient for L.anatina

		Max	Min	Average
	As	0,031	0,006	0,021
	Cd	0,004	0,000	0,002
	Cr	0,060	0,007	0,022
	Cu	0,080	0,022	0,043
	Zn	0,407	0,096	0,223
	Fe	8,795	3,920	6,450
	Al	5,530	2,135	3,724

Table 4 .

 4 43 The target hazard quotient for L.unguis

		Max	Min	Average
	As	0,017	0,000	0,006
	Cd	0,002	0,000	0,001
	Cr	0,053	0,003	0,016
	Cu	0,043	0,005	0,022
	Zn	0,103	0,026	0,080
	Fe	3,936	1,291	2,160
	Al	1,097	0,395	0,687

Table 4 .

 4 44 The target hazard quotient for G.coaxans

		Max	Min	Average
	As	0,017	0,000	0,011
	Cd	0,004	0,000	0,002
	Cr	0,034	0,002	0,010
	Cu	0,016	0,003	0,011
	Zn	0,146	0,015	0,076
	Fe	1,961	0,759	1,290
	Al	1,363	0,345	0,897

Table 4 .

 4 45 The target hazard quotient forA.antiquata

		Max	Min	Average
	As	0,003	0,002	0,003
	Cd	0,003	0,001	0,002
	Cr	0,012	0,001	0,006
	Cu	0,006	0,003	0,005
	Zn	0,041	0,026	0,032
	Fe	0,746	0,299	0,505
	Al	0,337	0,176	0,239

Table 4 .

 4 46 The target hazard quotient for S.regularis

		Max	Min	Average
	As	0,054	0,000	0,022
	Cd	0,004	0,000	0,002
	Cr	0,040	0,003	0,011
	Cu	0,177	0,018	0,061
	Zn	0,318	0,098	0,171
	Fe	4,229	0,729	2,232
	Al	2,289	0,449	1,375

Table 4 .

 4 47 The target hazard quotient for C.rivularis As shown in Table4.40 to 4.47, there were no THQ values for all metals greater than 1 for any metal, suggesting that the local inhabitants will not be exposed to a potential health risk from dietary As, Cd, Cr, Cu and Zn. However, the TTHQ values for Al and Fe caused by consuming mollusks were quite high, and likely would have been even higher had other metals and metalloids been included in the study. These species may pose a potential risk to the local inhabitants through their consumption in the diet. Consequently, some effective measures may be necessary to reduce Al and Fe metal contamination in sediments in this region. Further in depth studies are urgently needed, and should be identified to help safeguard human health.

		Max	Min	Average
	As	0,011	0,003	0,007
	Cd	0,004	0,000	0,002
	Cr	0,008	0,004	0,006
	Cu	0,162	0,049	0,106
	Zn	0,777	0,206	0,492
	Fe	0,595	0,395	0,495
	Al	0,406	0,310	0,358

Table 1 :

 1 Based concentration of metal in surface sediment at period 1996-2011

		Season	Zn	Cu	Pb	As	Cd	Hg
	Time		(µg/g)	(µg/g)	(µg/g)	(µg/g)	(µg/g)	(µg/g)
	Jun1996	D	6,33	8,68	0,19	18,04	ND	ND
	Oct1996	R	6,44	6,19	0,7	5,57	0,2	<0.1
	Sep1996	R	0,97	1,68	0,07	7,1	0,1	0,09
	Mar1998	D	50,8	26	12,6	3,9	<0.05	<0.02
	Feb1999	D	61,73	12,26	28,11	6,26	0,38	0,62
	Jun1999	R	97,24	14,62	29,55	8,33	0,04	0,78
	Feb2000	D	99,5	18,3	36,7	8,5	4,8	1,6
	Aug2000	R	57,53	10,23	26,09	2,32	Trace	2,76
	Feb 2001	D	52,3	10,6	33,4	4,05	0,21	1,99
	Aug2001	R	55,99	12,93	27,14	16,07	0,104	0,684
	Feb2002	D	24,8	5,37	15,63	1,53	0	0,28
	Feb2003	D	56,38	11,19	14,62	4,02	0,07	0,74
	Aug2003	R	57,4	13,3	4,7	3,19	0,5	0,53
	Feb2004	D	56,9	12,7	26	3,2	0,07	0,31
	Aug2004	R	58,64	11,21	36,1	3,6	0,05	0,33
	Feb2005	D	66,2	13,6	30,3	2,9	0,05	0,35
	Aug2005	R	49,5	15,4	26	3,4	0,29	0,28
	Feb2006	D	48,7	9,2	28,9	3,2	0,04	0,22
	Aug2006	R	62,8	5,7	26,3	2,6	0,12	0,26
	Feb2007	D	49,3	11,6	26,7	3	0,25	0,26
	Aug2007	R	47,5	9,9	25	3,2	0,2	0,39
	Feb2008	D	56,3	11,9	26,7	5,1	0,13	0,72
	Aug2008	R	53	7,8	23,7	4,9	0,07	0,8
	Feb2009	D	45,4	6,7	11,6	0,08	4,5	0,5
	Aug2009	R	21,9	4,3	9,2	0,06	2,6	0,25
	Mar2010	D	43	11,9	33,9	3,8	0,7	0,33
	Sep2010	R	39,8	11,2	30,7	3,8	0,45	0,41
	Mar2011	D	44,4	12,1	30,8	3,4	0,3	0,31
	Aug2011	R	43,2	13,6	34,1	2,7	0,33	0,43
	LEL*		120	16.0	31.0	6.0	0.6	0,15
	SEL*		270	110.0	110.0	33.0	9.0	1,3
	Note:	D: Dry season	R: Rainy season	ND: Not detected
	LEL*: Lowest effect level (ppm)		SEL*: Severe effect level (ppm)

  Table 5 describes classes for increasing Igeo values. The geoaccumulation index is calculated as follow

		Igeo = log 2 Cn/Bn
	Where:	Cn: concentrations of elements in samples (from 1998-2011)
		Bn: concentration values of the element in pre-industrial (1996)

Table 2 :

 2 Descriptive classes for increasing Igeo values[2] Detail of the results of calculating Igeo values for individual element is showed in table3. The negative Igeo presented in the table are the results of relatively low concentration for metal in sample.

	N	Igeo values Igeo class Designation of sediment quality
	1	0	0	Uncontaminated
	2	0-1	1	Uncontaminated to moderately contaminated
	3	1-2	2	Moderately contaminated
	4	2-3	3	Moderately contaminated to strongly contaminated
	5	3-4	4	Strongly contaminated
	6	4-5	5	Strongly contaminated to extremely contaminated
	7	>5	6	Extremely contaminated

Table 3 :

 3 Igeo values and classing of level contamination of each metal

	Time	Igeo	Igeo	Igeo	Igeo	Total
	Year	Zn	Cu	As	Pb	Igeo
	Mar 1998	1,02	0,85	0,19	2,45	0-3
	Feb 1999	1,07	0,66	0,26	3,22	0-4
	Jun 1999	1,19	0,70	0,30	3,27	0-4
	Feb 2000	1,19	0,76	0,30	3,48	0-4
	Aug 2000	1,05	0,61	0,12	3,15	0-4
	Feb 2001	1,03	0,62	0,20	3,39	0-4
	Aug 2001	1,04	0,67	0,39	3,19	0-4
	Feb 2002	0,83	0,44	0,06	2,65	0-3
	Feb 2003	1,05	0,63	0,20	2,59	0-3
	Aug 2003	1,05	0,68	0,16	1,49	0-2
	Feb 2004	1,05	0,67	0,16	3,14	0-4
	Aug 2004	1,06	0,63	0,18	3,46	0-4
	Feb 2005	1,09	0,68	0,15	3,29	0-4
	Aug 2005	1,01	0,72	0,17	3,14	0-4
	Feb 2006	1,01	0,58	0,16	3,25	0-4
	Aug 2006	1,07	0,46	0,13	3,16	0-4
	Feb 2007	1,01	0,64	0,15	3,17	0-4

Table 1 .

 1 Sampling location

	Samples	Sites	Longitude	Latitude
	NT1	Nha Trang port	12 0 12'45"N	109 0 13'12"E
	NT2	Be estuary	12 0 12'22"N	109 0 11'17"E
	NT3	Lon estuary	12°15'44"N	109°11'58"E
	NT4	Vinh Luong	12 0 20'10"N	109 0 12'15"E
	NT5	Ninh Loc	12°26'17"N	109°8'14"E
	NT6	Ninh Ich	12° 23'42"N	109°11'57''E
	NT7	Dinh estuary	12 0 27'00"N	109 0 09'38"E
	NT8	Cam Lam	12 0 04'45"N	109 0 10'42"E

As concentrations (ug/gr dry w t)

																	Cd concentrations (ug/gr dry w t)
	16													3
	12
														2
	8
	4													1
	0													0
	B . r a n a	I . p e r n a	M . v a g in u s	G . c o a x a n s	C . r iv u la r is	K . h ia n t in a	A . a n t iq u a ta	G . c o a x a n s	B . r a n a	I . p e r n a M . v a g in u s G . c o a x a n s	C . r iv u la r is	K . h ia n t in a A . a n t iq u a ta G . c o a x a n s
			Cu concentrations (ug/gr dry wt)						Pb concentrations (ug/gr dry wt)
	18													6
	12													4
	6													2
	0													0
	B . r a n a	I . p e r n a	M . v a g in u s	G . c o a x a n s	C . r iv u la r is	K . h ia n t in a	A . a n t iq u a ta	G . c o a x a n s	B . r a n a	I . p e r n a	M . v a g in u s	G . c o a x a n s	C . r iv u la r is	K . h ia n t in a A . a n t iq u a ta G . c o a x a n s
		Zn concentrations (ug/gr dry wt)					Cr concentrations (ug/gr dry wt)
	250													2,5
	200													2,0
	150													1,5
	100													1,0
	50													0,5
	0													0,0
	B . r a n a	I . p e r n a	M . v a g in u s	G . c o a x a n s		C . r iv u la r is	K . h ia n t in a	A . a n t iq u a ta	G . c o a x a n s	B . r a n a	I . p e r n a	M . v a g in u s	G . c o a x a n s	C . r iv u la r is	K . h ia n t in a	A . a n t iq u a ta	G . c o a x a n s
							Figure 1. Heavy metal concentrations in mollusk tissues

Table 2 .

 2 BSAF values for each mollusks for heavy metals

	Mollusk names	As	Cd	Cr	Cu	Pb	Zn
	Bufonaria rana	5,45	3,99	0,01	3,28	0,06	1,51
	Isonomon perna	13,41	5,89	0,02	1,29	0,13	65,46
	Modiolus vaginus	2,36	5,25	0,34	2,49	0,68	1,83
	Geloina coaxans	1,17	1,76	0,04	0,57	0,15	3,81
	Crasostrea rivularis	10,57	4,15	7,51	14,71	0,69	46,68
	Katelysia hiantina	0,56	0,76	0,01	0,39	0,05	0,64
	Anadara antiquata	0,29	1,95	0,01	0,15	0,06	0,58
	Glauconome virens	0,78	0,44	0,02	0,03	0,04	0,38

Table 2 .

 2 The BSAF values in the different mollusk species.

Notes:

Acknowledgement:

The authors wish to acknowledge and thank staff of Institute of Oceanography Nha Trang, especially to Mr.Vu Tuan Anh for their helps and helpful comments in this study. In addition, we wish to thank to Ms. Nguyen Thi Thuy Luyen for help and assistance geochemical analysis.

Acknowledgement:

The authors wish to acknowledge and thank staff of Institute of Oceanography Nha Trang, especially to Mr.Vu Tuan Anh and Ms.Nguyen Thi Thanh Thuy for their helps and helpful comments in this study. In addition, we wish to thank to Mr. Hua Thai Tuyen for help and assistance classification of mollusks.

Acknowledgement:

The authors wish to acknowledge and thank staff of Institute of Oceanography Nha Trang, especially to Mr.Vu Tuan Anh and Ms.Nguyen Thi Thanh Thuy for their helpful comments in this study. In addition, we wish to thank to Mr. Hua Thai Tuyen, Ms. Nguyen Thi Thuy Luyen for their help.

SOME PICTURE OF COLLECTING SAMPLES

Picture A2.1 Collecting mollusk samples at Binh Tan Picture A2.2 Collecting mollusk samples at Tan Dao site Picture A2.3 Collecting samples at Cam Lam site Picture A2.4 Collected mollusk samples

LIST OF ACRONYMS

Al

The bivalve Perna viridis is an economically important mussel and a native to Asia-Pacific region where it is widely distributed. The P.viridis in found in the coastal waters of the Indo-pacific region (Benson and others, 2002). It is harvested fro food but is also known to harbor toxins and cause damage to submerged structures such as drainage The guidelines were derived for 25 chemicals (9 trace elements, 13 PAHs, 2 OCs and total PCB). Appendix 6.1 is a summary of ER-L, ER-M and Overall Apparent Effects Thresholds concentrations for selected chemicals in sediment (dry weight) (Long et al., 1990). These effect-range guidelines can be applied to both fresh and salt water sediments.

Provincial Sediment Quality Guidelines for Metals and Nutrients

These The ATL values represent the maximum concentration of a given chemical in shellfish tissue that will not generate a risk greater than the maximum acceptable risk level (ARL) used for carcinogens (See appendix 6.7)

Therefore, the ATLh values permit a specific population of humans to consume safely any combination of fish and/or shellfish for an extended period, provided that the combined daily consumption rate remains below the value of IR used to calculate ATLh.

CTL values are concentrations of bioaccumulative chemicals in tissue that will not cause significant adverse effects on the health of aquatic organisms containing those