Mercury and methylmercury concentrations in Mediterranean seafood and surface sediments, intake evaluation and risk for consumers

International Journal of Hygiene and Environmental Health 215 (2012) 418–426

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Mercury and methylmercury concentrations in Mediterranean seafood and surface sediments, intake evaluation and risk for consumers Lucia Spada ∗ , Cristina Annicchiarico, Nicola Cardellicchio, Santina Giandomenico, Antonella Di Leo C.N.R. – Institute for Coastal Marine Environment, Taranto Section, via Roma 3, 74100 Taranto, Italy

a r t i c l e

i n f o

Article history: Received 14 March 2011 Received in revised form 26 August 2011 Accepted 8 September 2011 Keywords: Mercury Methylmercury Sediment Edible organisms Health hazard

a b s t r a c t Total mercury and methylmercury concentrations were measured in sediments and marine organisms from the Taranto Gulf to understand their distribution and partitioning. Sediment concentrations ranged from 0.036 to 7.730 mg/kg (mean: 2.777 mg/kg d.w.) and from 1 to 40 ␮g/kg (mean: 11 ␮g/kg d.w.) for total mercury (THg) and methylmercury (Me–Hg), respectively. In mollusks THg ranged from n.d. to 1870 ␮g/kg d.w. while in fish from 324 to 1740 ␮g/kg d.w. Me–Hg concentrations in fish ranged from 190 to 1040 ␮g/kg d.w. and from n.d. to 1321 ␮g/kg d.w. in mollusks. THg exceeded the maximum level fixed by the European Commission (0.5 mg/kg w.w.) only in gastropod Hexaplex t. The calculated weekly intake was in many cases over the Provisional Tolerable Weekly Intake established by EFSA for all edible species. These results seem to indicate that dietary consumption of this seafood implicates an appreciable risk for human health. © 2011 Elsevier GmbH. All rights reserved.

Introduction In the marine environment, levels of contaminants have been increasing over the last decades, as a consequence of anthropogenic activities and pollutants are potentially accumulated in organisms and sediments, and subsequently transferred to man through the food chain. Coastal areas, particularly near large urban centres, are of concern, as they receive the largest exposure to chemical contamination, due to source proximity. Contamination of marine organisms with toxic chemicals, such as mercury and its compounds, has been intensively studied in recent years, due to the fact that these contaminants are persistent, toxic, tend to bioaccumulate, and pose human and ecosystem risks. Mercury may occur naturally in the environment or from anthropogenic sources like mining, fossil fuels combustion, incineration, emission from smelters, fungicides and catalyst activities. Most mercury is volatilized and returned to the atmosphere, but the greater part of this metal introduced into the coastal sea precipitates, because of the very low solubility products of its compounds. It also accumulates in the sediment, which represents the principal sink. Most of inorganic and organic mercury in aquatic environment appears to be bound to particles, colloids and high molecular weight organic matter (Horvat, 1997; Schiff, 2000). In sediments, due to bacterial activity, inorganic mercury may also be converted into methylmercury, the most toxic chemical species which may

∗ Corresponding author. Tel.: +39 099 4542206; fax: +39 099 4542215. E-mail address: [email protected] (L. Spada). 1438-4639/$ – see front matter © 2011 Elsevier GmbH. All rights reserved. doi:10.1016/j.ijheh.2011.09.003

cause the permanent harm to the central nervous system (Harada et al., 1998), such as behavioural disorders and deficiencies in the immune system and development (Harada et al., 1998; UNEP, 2002). In this form methylmercury dissolves in the water column, becoming readily bioavailable; then it bioaccumulates and biomagnifies up into marine food chains leading to elevated concentrations especially in predatory organisms. Therefore, the consumption of marine products represents a non-negligible exposure pathway to Hg and, thereby, a risk for human health. The aim of this work was to determine the concentrations of total mercury (THg) and methylmercury (Me–Hg) in the sediments, bivalve molluscs (Mytilus galloprovincialis, Chlamys varia), gastropod molluscs (Hexaplex trunculus) and fish (Symphodus melops) collected at 5 sites from Taranto Gulf, in order to investigate contamination level and public health risks, associated with consuming fish and seafood harvested from these areas. Moreover the goal of this study was also to estimate the weekly intake and compare it with the Provisional Tolerable Weekly Intake (PTWI) recommended by the European Food Safety Authority (EFSA, 2004). Materials and methods Study area Taranto seas, Mar Piccolo and Mar Grande basins (Fig. 1), represent a coastal marine ecosystem example, whose biological balances have been modified step by step, in relation to the anthropic development and, in particular, to the industry settlement (iron and steel factory, petroleum refinery and shipyard). For

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Fig. 1. Location of the sampling stations.

these intense anthropogenic impacts Taranto city has been identified as “High Risk of Environmental Crisis Area” (Law n. 349/86). Later on, the Law n. 426/98 classified it as “Site of National Interest”, including the site in the “National Project of Environmental Restoration”. The Mar Piccolo basin is located in the Northern area of the Taranto town, Italy. It is an inner, semi-enclosed basin (surface area of 20.72 km2 ), with lagoon features, divided into two inlets, called first and second inlet, which have a maximum depth of 13 and 8 m, respectively. It is connected with the Mar Grande through two channels, the “Navigabile” the “Porta Napoli” channels. Tidal range does not exceed 30–40 cm. The scarce hydrodynamism and the low water exchange with the nearby Mar Grande determine (mainly in summer), a high water stratification. Mar Piccolo basin is the most important area for mussel culture in Italy; it is influenced by urbanization, by harbour activities, by aquaculture and commercial fishing. The main problems of environmental impact are: nine pipes discharge sewages, the ship-yard of the Italian Navy with its dry-docks (located in the first inlet), the largest mussel farm distributed in both the inlets, the fishing-boat fleet localized in the first inlet and small rivers and freshwater springs which drain the surrounding agricultural soils in the basin. Mar Grande basin is a wide roadstead, which lies to the NorthEastern area of the Taranto Gulf. Twenty-eight urban and five industrial discharges drain directly into the basin. Taranto seas are a noteworthy economic resource, being the site of intensive mussel farming. Mussel farming in Taranto has a long history that dates back to the sixteenth century, so that the typical chestnut stakes, which stick out of the sea, have always been part of the city view. This industry has grown from the idea of an enterprising local to become a big export earner. Actually, the annual output amounts to 30,000 tonnes of mussels. Only a part of the locally harvested seafood is used for home consumption, while most of it is exported to European Economic Community countries, in particular to Spain. Sampling Surface marine sediments and marine organisms were collected from four stations of the Mar Piccolo and from one station of the Mar Grande (Fig. 1). Sampling stations were selected by considering

distribution of contamination sources, in order to obtain a good evaluation of the monitored area. In particular, station one was near the Navigable Channel, station two was located in proximity to the former shipyard “Tosi”, station three was near the Navy Arsenal. Station four was located at the North side of the second inlet of the Mar Piccolo while station five was in proximity of the touristic and commercial port, located in Mar Grande basin. Surface sediment samples (0–3 cm) were collected with a Van Veen grab in three replicates per station. After sampling, redox potential was determined by a platinum redox electrode Crison (Crison Instruments, Spain). Sediments were stored into a plastic vessel and frozen at −20 ◦ C until analysis. In the laboratory, sediment samples were defrosted at room temperature, dried at 30 ◦ C up to a constant weight, ground and homogenized to a fine powder in a mortar. Fish species, corkwing wrasse (Symphodus melops) was caught with gill nets. Bivalve molluscs (Mytilus galloprovincialis, Chlamys varia) and gastropod mollusc (Hexaplex trunculus) were collected by scuba divers. Sampling, handling and storage of the specimen were carried out according to the FAO (1976) recommendations. Samples were stored below −20 ◦ C until analysis. Composite samples of fish species, lower than 100 g, were prepared blending together muscle tissue of three specimens of almost similar size (about 15 cm). For analysis of molluscs, individuals of commercial size were partially thawed, the shell removed and a composite sample of at least 20 individuals was made (mean wet weight 4 g/individual). Dry weight calculation on organism tissues was carried out by oven drying at 105 ◦ C until constant weight. Total mercury determination THg concentrations were determined (both for sediments and organisms) with an Advanced Mercury Analyser (AMA-254, LECO) that uses catalytic combustion of the sample, pre-concentration by gold amalgamation, thermal desorption and UV determination at 254 nm. About 100 mg of dry sediments and wet organism’s tissue were precisely weighed in a nickel boat, placed in the instrument and automatically introduced into the AMA. The entire analytical procedure was validated by analysing CRM DOLT-2 (dogfish tissue) and IAEA 356 (marine sediment) samples at the beginning

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Table 1 Analytical results of standard reference materials DOLT-2 (dogfish tissue) and IAEA-356 (marine sediment). DOLT-2

IAEA-356

Total Hg (mg/kg d.w.)

MeHg (as Hg) (mg/kg d.w.)

Total Hg (mg/kg d.w.)

MeHg (as Hg) (␮g/kg d.w.)

0.672 ± 0.065

7.61 ± 0.16

5.29 ± 0.28

0.693 ± 0.053

7.62 ± 0.62

5.46 ± 0.38

Measureda 1.87 ± 0.15 Certified 1.99 ± 0.10 d.w.: dry weight. a Number of replicates is 5.

and end of each set of (usually 10) tissue samples, thereby ensuring that the instrument remained calibrated throughout the study. Results were in agreement with certified values and the standard deviations were low, proving good repeatability of the method (Table 1). The detection limit (0.2 ng/g d.w.) was calculated based on blank measurements (Buccolieri et al., 2006). All chemicals used in sample treatments were ultrapure grade (Merck, Daemastadt, Germany). Working standard solutions were prepared by serial dilution of stock mercury standard solutions containing 1000 mg/L (BDH, Poole, UK).

International Atomic Energy Agency (Marine Sediment IAEA 356). Results were in agreement with certified values and the standard deviations were low, proving good repeatability of the method (Table 1). The detection limits (LOD) of 0.6 ng/g d.w. in sediments and 0.4 ng/g d.w. in organisms were obtained with three times the SD of the blank. The solvent used was pesticide analysis grade (Merck, Daemastadt, Germany). The acid (ultrapure grade) was sulphuric acid (96%) (Merck, Daemastadt, Germany). The reagents used were: KOH, CuSO4 , KBr, cysteine, (BDH). Aqueous solutions were prepared daily by dissolving an appropriate amount in Milli-Q® water.

Methylmercury determination Me–Hg was determined using a method described by Caricchia et al. (1997) and modified by us. An aliquot of dry sediment (2 g) and wet pooled organism tissue (1 g) was weighed and extracted with 5 ml KOH/CH3 OH (25%) in ultrasonic bath (45 min). After cooling, 5 ml of H2 SO4 (4 M, saturated with CuSO4 ), 5 ml of 4 M KBr and 4 ml of toluene were carefully added and the sample was manually shaken for about 3 min. After centrifugation (2200 rpm for 10 min), the supernatant organic phase was collected. The solvent extraction was repeated three times by 2 ml of toluene. The collected organic extract (10 ml) was subjected twice to a back-extraction by 1 ml of a cysteine solution (1%). The two cysteine extracts (1 + 1 ml) were collected in the same vial and extracted by a mixture of toluene (0.5 ml), CuSO4 saturated solution (0.5 ml), and 4 M KBr (1 ml). After manual shaking, the organic phase was separated from the aqueous phase and 2 ␮l were injected into GC-ECD system. Precision and accuracy of the analytical methods were determined using the following certified material from the National Research Council—Canada (Dogfish DOLT – 2) and from the

Statistical analysis Multivariate analysis methods such as two-way analysis of variance (ANOVA), principal component analysis (PCA), and correlation analysis (bivariate correlations) have been used to extract information from the chemical analysis, in order to find the relationships between THg and Me–Hg in organisms and sediments. As regard one-way ANOVA the F test was used (p < 0.01). Principal component analysis (PCA) was executed on the analytical data, in order to obtain a visual representation of the main characteristics of relationship among THg and Me–Hg concentrations in the studied organisms and sediments. The combined plot of scores and loadings allowed us to recognize groups of samples with similar behaviour and the existing correlation among the original variables. The correlation analysis was performed by Pearson correlation. All statistical analyses were performed with the software package STATISTICA® (StatSoft Inc., Tulsa, OK, USA).

Table 2 THg, Me–Hg concentrations (␮g/kg d.w.), standard deviation and percentage of Me–Hg respect to THg in sediments and organisms from Mar Piccolo and Mar Grande. Location THg Mytilus g. Symphodus m. Hexaplex t. Chalmys v. Sediment Me–Hg Mytilus g. Symphodus m. Hexaplex t. Chalmys v. Sediment % Me–Hg as THg Mytilus g. Symphodus m. Hexaplex t. Chalmys v. Sediment

1

2

3

4

5

368 ± 29 887 ± 76 474 ± 30 1443 ± 98 2620 ± 202

1038 ± 95 1740 ± 187 1431 ± 157 703 ± 34 2200 ± 33

1403 ± 219 1368 ± 177 1867 ± 197 1676 ± 184 7730 ± 685

21 ± 2 324 ± 40 224 ± 21 n.d. 36 ± 3

300 ± 32 945 ± 82 1099 ± 121 370 ± 34 1300 ± 178

129 ± 16 456 ± 34 199 ± 28 559 ± 52 4 ± 0.3 35.0 51.4 42.0 38.7 0.2

Each value corresponds to the mean of five determinations.

301 ± 36 1040 ± 146 1321 ± 164 315 ± 29 2 ± 0.2 29.0 59.7 92.3 44.7 0.1

377 ± 48 821 ± 87 800 ± 74 381 ± 48 7 ± 0.6 26.9 60.0 42.9 22.8 0.1

n.d. 190 ± 25 37 ± 4 n.d. 1 ± 0.1 – 58.7 16.7 – 2.8

75 ± 6 591 ± 87 355 ± 40 118 ± 15 40 ± 3 25.0 62.5 32.3 31.9 3.1

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Table 3 Total mercury (mg/kg d.w.) and methylmercury (␮g/kg d.w.) levels found in sediments from different Mediterranean areas. Study area

THg (mg/kg)

Range (mg/kg)

Me–Hg (␮g/kg)

Range (␮g/kg)

Bibliography

Gulf of Trieste – Adriatic sea Kastek Bay, Croatia – Adriatic Sea Lagoon of Bizerte – Tunisia Mar Piccolo – Ionian Sea Venice Lagoon – Adriatic Sea Gulf of Taranto – Ionian Sea Gulf of Taranto – Ionian Sea

5.24 21.60 0.13 1.62

0.10–23.30 14.28–30.40 0.01–0.65 1.07–5.72 0.21–1.15 0.03–11.60 0.36–7.73

16.90 19.70 0.53

0.20–60.10 6.10–36.70 nd–3.20

Covelli et al. (2001) Kwokal et al. (2002) Mzoughi et al. (2002) Cardellicchio et al. (2006) Seunghee et al. (2007) Cardellicchio et al. (2009) Present study

1.36 2.77

Results and discussion Mercury in sediments The sediment redox potential showed similar and negative values in all sampled stations, from −457 mV (station 4) to −348 mV (station 5). Table 2 showed mean values of THg and Me–Hg concentrations, together with the percentages of methylmercury to total mercury in analyzed sediments. Concentrations in sediments ranged from 0.04 to 7.7 mg/kg (mean: 2.8 mg/kg d.w.) for THg and from 1.0 to 40.0 ␮g/kg (mean: 11.0 ␮g/kg d.w.) for Me–Hg. The highest levels of THg and Me–Hg were found in sediments from stations 3 (located near to Military Navy Arsenal in the first inlet of the Mar Piccolo) and station 5 (located near touristic port “San Eligio” from Mar Grande), respectively. Instead, the lowest levels, both THg and Me–Hg, were found in sediments from station 4 located in the Northeast side from second inlet of the Mar Piccolo basin. Craig (1986) reported concentration ranges of 0.2–0.4 mg/kg THg for uncontaminated sediments, whereas sediments in urban, industrial or mining areas can contain up to 100.0 ␮g/g of THg. Buccolieri et al. (2006) reported background levels of 0.07 mg/kg in sediments from Ionian Sea (Mediterranean Sea). Therefore, the THg concentrations in sediments collected in this study (ranged from 1.3 to 7.7 mg/kg d.w.) with the exception of the station 4 were up of 19–110 times higher than background levels. Thus, these sediments could be considered as heavily contaminated. Me–Hg concentrations are characteristic of anoxic polluted sediments (Kwokal et al., 2002). Moreover the percentage of methylmercury in respect of total mercury concentrations in sediments ranged from 0.1 to 3.1% which falls within the normal values reported in the literature for coastal marine environments (Cossa et al., 1996; Mason et al., 1993). Table 3 showed THg and Me–Hg levels in sediments, found in this work, compared to those found in literature from different Mediterranean areas. This survey showed that the mean

0.30–1.70 10.80

1.00–40.00

concentration and range of mercury, determined in sediments from the Taranto Gulf, were higher than those reported by other studies (Cardellicchio et al., 2006, 2009; Mzoughi et al., 2002; Seunghee et al., 2007) with the exception of the level indicated by Kwokal et al. (2002) in the Kastella Bay (Croatia), heavily contaminated by mercury, derived from a PCV chlor-alkali plant, which operated during the period 1950–1990, and Covelli et al. (2001), in the Gulf of Trieste (Italy), influenced by the contaminated river Soca/Isonzo, for centuries draining the cinnabar-rich deposits of the Idrija mining district, in the Northwestern part of Slovenia. However, concentrations of Me–Hg in Taranto Gulf are comparable than those observed by Kwokal et al. (2002) and Covelli et al. (2001). Therefore in comparison with published data from other coastal areas, the THg concentrations reported here for Taranto Gulf were indicative of a polluted environment.

Mercury in organisms THg and Me–Hg concentrations and Me–Hg percentage in dry tissue of the marine organisms found in this study are summarized in Table 2. THg concentrations in soft tissue of bivalve mollusks (Mytilus g.), ranged from 21 to 1403 ␮g/kg d.w., while the levels of Me–Hg ranged from not detectable to 377 ␮g/kg d.w. The highest levels both THg and Me–Hg were found in sediments from station 3 (localized near to the Military Navy Arsenal, in the first inlet of the Mar Piccolo). Concentrations of THg in bivalve mollusks Chlamys v. ranged from not detectable to 1676 ␮g/kg d.w., while Me–Hg ranged from not detectable to 559 ␮g/kg d.w. In the sediments from stations 3 and 1 (located near to the Navigable Channel) were observed highest values of THg and Me–Hg, respectively. THg and Me–Hg concentrations in gastropod mollusks Hexaplex t. were in the range 224–1867 ␮g/kg d.w. and 37–1321 ␮g/kg d.w., respectively.

Table 4 Total mercury and methylmercury levels (␮g/kg) found in mussel Mytilus galloprovincialis and in gastropod mollusks Hexaplex trunculus from different Mediterranean areas. Study area Mytilus galloprovincialis Krka estuary – Croatia French coasts – Mediterranea Sea Gulf of Naples – Tirrenian Sea Sardinia Coasts – Tirrenian Sea Kastela Bay – Adriatic Sea Mar Piccolo – Ionian Sea Gulf of Taranto – Ionian Sea Hexaplex triinculus Croatian Coasts– Adriatic Sea La Spezia – Tirrenian Sea Gulf of Naples – Tirrenian Sea Gulf of Manfredonia – Adriatic Sea Gulf of Taranto – Ionian Sea d.w. = dry weight. w.w. = wet weight.

THg

146.9 d.w. 204.0 w.w. 181.0 d.w. 200.0 d.w. 626.0 d.w. 293 d.w. 558.6 d.w. 266.2 d.w. 266.4 d.w. 1019.0 d.w.

Range 14.5–30.2 w.w. 99.0–220.0 d.w. 5.0–366.0 w.w. 40.0–830.0 d.w. 138.0–325.0 d.w. 110.0–290.0 d.w. 21.0–1403.0 d.w.

Me–Hg

Range

Bibliography

67.1 d.w.

4.1–8.5 w.w. 43.0–88.0 d.w.

65.0 d.w.

17.0–116.0 d.w. 30.3–59.3 d.w.

176.4 d.w.

n.d.–377.0 d.w.

Mikac et al. (1996) Claisse et al. (2001) Amodio-Cocchieri et al. (2003) Ipolyi et al. (2004) ´ Kljakovic-Gaˇ spic´ et al. (2006) Cardellicchio et al. (2010) Present study

37.0–1321.0 d.w.

Buzina et al. (1989) Giordano et al. (1991) Giordano et al. (1991) Giordano et al. (1991) Present study

231 d.w.

224.0–1867.0 d.w.

524.4 d.w

422

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Fig. 2. Loading of the variables on the first two principal components.

In fish Symphodus m. THg concentrations ranged from 324 to 1740 ␮g/kg d.w., while the levels of Me–Hg ranged from 190 to 1040 ␮g/kg d.w. Maximum THg and Me–Hg levels in fish were found in station 2. It is important to note that the lowest levels both for THg and Me–Hg in all organisms and sediments analyzed were found in the station 4 localized in the Northeast side from second inlet of the Mar Piccolo far from direct sources of contamination. The percentage of Me–Hg as THg observed in bivalve mollusks Mytilus g. and Chlamys v. were about 23% and 27%, respectively. These values were in according to other literature data concerning Me–Hg percentage in bivalve mollusks (Berzas Nevado et al., 2003; Mikac et al., 1996). In the gastropod mollusks Hexaplex t., the percentage of Me–Hg as THg showed high variability ranging from 17 to 92%. The maximum value, observed in the station 2, was in agreement with that cited by Buzina et al. (1989), in the Croatian Coasts (Adriatic Sea). In fish Symphodus m. the percentage of Me–Hg as THg was similar in all sampled organisms, with a mean value of about 60%, according to different bioaccumulation studies of Me–Hg in fish (Agah et al., 2007). Other authors report a percentage of Me–Hg as THg in fish from 70 to 100% (Claisse et al., 2001; Storelli et al., 2005). However, factors such as age, sex, and feeding habitat of fish may influence such ratios (Lasorsa and Allen-Gil, 1995). In general, the mussels have Me–Hg/THg ratios which are lower compared to the ratios found in fish. The difference in ratio between mussels and fish may be due to the trophic level occupied by each organism, their physiology and metabolism and distribution in various tissues of proteins that are capable to complex metals and its compounds. There are very few published data about the contemporary evaluation of THg and Me–Hg concentrations in the marine organisms collected from Taranto Gulf, with the exception of Mytilus galloprovincialis and, in lower measure, Hexaplex trunculus which are widespread species. Table 4 showed THg and Me–Hg levels in mussels and gastropod mollusks found in this work, compared to those found in literature from different Mediterranean areas. Comparing these different data, one must pay attention as they are often reported in wet weight. While the content of water is not so important for fish, it is very variable for mollusks, being affected by factors such as: habitat, physiological conditions, and sample storage conditions (Cardellicchio et al., 2008). However, in order to

have a clear comparison among values, the average water content of the samples (85%) was used in the calculations (Ipolyi et al., 2004). This survey showed that the mean concentration and range of THg found in the Mytilus g. from Taranto Gulf were higher than those reported by other studies (Cardellicchio et al., 2010; Claisse ´ et al., 2001; Ipolyi et al., 2004; Kljakovic-Gaˇ spic´ et al., 2006; Mikac et al., 1996) with the exception of the level indicated by AmodioCocchieri et al. (2003) in the Gulf of Naples, which is considered a very polluted area subject by civil and industrial wastes. Concerning Me–Hg levels found in Mytilus g. in this work, the mean concentration and range were higher than those reported ´ by other studies (Claisse et al., 2001; Ipolyi et al., 2004; KljakovicGaˇspic´ et al., 2006; Mikac et al., 1996). Regarding the gastropod Hexaplex t., THg and Me–Hg levels in this study were much more higher than those reported by other literature studies (Buzina et al., 1989; Giordano et al., 1991). Therefore in comparison with published data from other coastal areas, the THg and Me–Hg concentrations reported here for the Gulf of Taranto are indicative of a polluted environment. Statistical analysis Correlation analyses of Hg and Me–Hg contents of organisms and sediment, performed according to Pearson Product-Moment, showed that THg was not correlated with Me–Hg concentrations in sediments (p > 0.05). Under anaerobic conditions, Hg2+ has a high affinity for sulphide, resulting in the formation of insoluble HgS, which is deposited in the sediment. Once deposited as HgS, mercury is presumably not available for methylation (Andersson et al., 1990). Physical perturbation or bioturbation can oxidize HgS and thus remobilize a small percentage of HgS (Stein et al., 1996). In contrast to sediments, THg concentrations in Mytilus g., Chlamys v. and Symphodus m. were correlated with Me–Hg (p < 0.05) in agreement with earlier studies (Kannan et al., 1998; Kim, 1995). Concerning THg and Me–Hg concentrations no correlations were found among sediments and all analyzed organisms. PCA was applied using as variables the THg and Me–Hg mean concentrations in the bivalve mollusks (Mytilus g. and Chlamys v.), gastropod mollusks (Hexaplex t.) fish (Symphodus m.) and sediments, in order to verify possible bioaccumulation patterns and detect possible different contamination levels among sites in the study area. Three principal components were extracted by covering 94.7% of the cumulative variance. Results are reported in

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423

Fig. 3. Scatter plot of the scores for the first two principal components.

Figs. 2 and 3. As shown by the factor loadings reported in Fig. 2, the stations 1, 2, 3 and 5 were correlated and station 4 was independent of the other stations. Factor scores of PCA (Fig. 3) for THg, Me–Hg concentrations in organisms and sediments showed that the stations 1, 2, 3 and 5 seemed to be associated with high THg levels, in sediments, in respect of THg and Me–Hg concentrations, in organisms and also Me–Hg in sediments. Moreover, the station 4 seemed to be different from the others, with low concentrations of THg and Me–Hg in sediments and organisms. Indeed, ANOVAs showed a statistically significant difference in the observed levels of THg and Me–Hg in the organisms and sediments between the stations (p < 0.01). Furthermore, a statistically significant difference was observed between THg and Me–Hg levels in organisms inside the stations (p < 0.01).

Risk for human health For most people the main route of exposure to mercury and methylmercury is through the diet. Consequently, information about dietary intake is necessary, in order to evaluate the potential health risk for the individual. Apart from comparing the concentrations of mercury with the maximum levels fixed by European Legislation, a more significant assessment of potential health hazards for consumers can be obtained by calculating the weekly intake of mercury and methylmercury, deriving from the consumption of fish and seafood and comparing it with the Provisional Tolerable Weekly Intake (PTWI) set by international organizations. The European Legislation (EC, 2006) establishes the maximum permissible concentration of mercury in edible marine species at

Table 5 Mean THg and Me–Hg concentrations and estimated weekly intake (␮g/kg body weight) in adults and children of Taranto, Italy. Location

THg (␮g/kg) w.w.

Mytilus galloprovindalis 1 40.0 2 100.0 3 160.0 4 2.0 5 32.0 Syntphodus ntelops 1 210.0 2 390.0 3 300.0 4 92.0 5 200.0 Hexaplex trunculus 1 150.0 2 390.0 3 560.0 4 60.0 5 310.0 Chlamys varia 1 230.0 2 123.0 3 290.0 4 n.d. 5 72.0

ETHg–w adultsa

ETHg–w childrenb

EMe–Hgw adultsa

EMe–Hgw childrenb

0.3 0.8 1.3 0.02 0.3

0.6 1.4 2.3 0.03 0.5

14.0 29.0 43.0 n.d. 8.0

0.1 0.2 0.4 n.d. 0.1

0.2 0.4 0.6 n.d. 0.1

1.7 3.2 2.5 0.8 1.6

3.0 5.5 4.3 1.3 2.8

108.0 233.0 180.0 54.0 125.0

0.9 1.9 1.5 0.4 1.0

1.5 3.3 2.6 0.8 1.8

1.2 3.2 4.6 0.5 2.5

2.1 5.5 8.0 0.9 4.4

63.0 360.0 240.0 10.0 100.0

0.5 2.9 2.0 0.1 0.8

0.9 5.1 3.4 0.1 1.4

1.9 1.0 2.4 n.d. 0.6

3.3 1.7 4.1 n.d. 1.0

89.0 55.0 66.0 n.d. 23.0

0.7 0.4 0.5 n.d. 0.2

1.3 0.8 0.9 n.d. 0.3

ETHg–w and EMe–Hg–w were expressed as ␮g/kgbw week. a Average adults body weight 60 kg (Monteduro et al., 2007). b Average children body weight 34.5 kg (7–12 years) (Yusà et al., 2010).

Me–Hg (␮g/kg) w.w.

424

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Fig. 4. The estimated weekly intake of THg and Me–Hg from Mytilus galloprovincialis, Symphodus melops, Hexaplex trunculus and Chlamys varia according to the PTWI EFSA limit (1.6 ␮g/kg body weight).

0.5 mg/kg of fresh weight. Considering this standard, concentrations exceeding it were observed solely in Hexaplex t. samples from the stations 3, located in proximity of the naval base of the Italian Marine Military (Mar Piccolo, first inlet). European Food Safety Authority (EFSA) has established regulatory guidelines regarding dietary THg and Me–Hg intake, recommending a Provisional Tolerable Weekly Intake (PTWI) of 1.6 ␮g/kg body weight (kgbw ), based on epidemiological studies that investigated the relationship between maternal exposure to

mercury and impaired neurodevelopment in their children (EFSA, 2004). PTWI of 1.6 ␮g/kg body weight represent safe values, which can be accumulated by the human population over lifetime. However, different considerations are recommended for pregnant women, nursing mothers and young children; due to Hg toxic effects on the developing nervous system, these categories should avoid the consumption of fish species which normally have higher mercury content.

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The risk to human health as a result of eating fish and seafood were evaluated by dietary THg and Me–Hg exposure from fish and seafood calculating daily mercury exposure Em (US-EPA, 1994) as follows: Em =

Cm · IR BW

Cm represents THg or Me–Hg concentration in fish or seafood (␮g/g w.w.); IR the ingestion rate (g/d) of fish or seafood and BW represents the body weight (kgbw ). Dietary THg and Me–Hg exposure was expressed as ␮g/kgbw day. In order to calculate the total daily exposure, average seafood consumption of 70 g/d was considered. This value is 10 g more than the value suggested by ISTAT (2000) for Italy, because the population inTaranto can be considered a high level fish and seafood consumer, as its diet is highly dependent on fish and seafood items. In this study it was considered a Cm value equal to 70 g/d, as a consequence seafood mean weekly consumption of 490 g. Moreover, daily intake was estimated for two age groups: children and adults. Considering the average children’s body weight of 34.5 kg (Yusà et al., 2010), and the average body weight of 60 kg (Monteduro et al., 2007) for adults. Therefore weekly THg and Me–Hg exposure (ETHg–w , EMe–Hg–w ), obtained by multiplying Em by 7 days are shown in Table 5. Concerning the Mytilus g., weekly THg and Me–Hg exposure (ETHg–w , EMe–Hg–w ), obtained were always below the established PTWI by ESFA, except to the children ETHg–w value (2.3 ␮g/kgbw week), found in the station 3. Regarding bivalve molluscs Chlamys v., weekly THg exposure (ETHg–w ) for adults, exceeded the PTWI in the station 1 and 3 (1.9 and 2.4 ␮g/kgbw week, respectively) while for children, also in the station 2. In the same organisms, in both adults and children, the EMe–Hg–w was always below the established PTWI. ETHg–w of gastropod mollusks Hexaplex t., calculated for children, exceeded the PTWI in all sampled station with the exception of station 4. Similarly in the adults the ETHg–w exceeded the PTWI in the stations 2, 3 and 5. EMe–Hg–w exceeded the limit in the stations 3 and 2 both in adults and in children (2.0 and 2.9; 3.2 and 5.1 ␮g/kgbw week, respectively). For mobile benthic carnivorous fish Symphodus m., ETHg–w , calculated both for adults and children, was always over the established PTWI, especially in the stations 3 and 2 (2.5 and 3.2 ␮g/kgbw week for adults; 4.3 and 5.5 ␮g/kgbw week for children, respectively) with the exception of the station 4. Concerning EMe–Hg–w in the adults, calculated values exceeded the PTWI only in the station 2 (1.9 ␮g/kgbw week), while in the children the EMe–Hg–w was over the mentioned limit in the station 2, 3 and 5 (3.3, 2.6, and 1.8 ␮g/kgbw week, respectively). In general, in relation to the average body weight, intake by children of THg and Me–Hg was higher than that found for adults (Fig. 4), especially for THg in fish, gastropod mollusks and bivalve mollusks Chlamys v. Therefore, these results indicate that THg and Me–Hg can pose a risk, especially to children who eat these organisms that are sampled in the stations 3 and 2. In fact, the sediments of these stations were the most contaminated. However, the highest value found in fish might be related to its feeding behaviour, trophic position in the food chain and fat content. Moreover, Hexaplex t. and Chlamys v. represent benthic organisms which live in contact with the polluted sediments. For this reason, mercury is taken up by adsorption of freely dissolved chemicals from the overlying and pore water, and also by direct contact and ingestion of sediment particles (Landrum et al., 1992). In addition, it must be borne in mind that the estimated intake does not include the contribution of other foods that may constitute further contamination sources to which population is also subjected. Besides, it should be considered

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that the estimation of daily mercury intake from seafood increase considering certain community members, such as fishermen and their families, which consume greater amounts of seafood. Thus, high health risks due to the dietary THg and Me–Hg intake for children especially for fishermen’s sons in Taranto population cannot be excluded. Furthermore, pregnant and lactating women should pay particular attention to the amount of seafood and fish introduced in their diets, due to the toxic effects of Hg on the developing nervous system.

Conclusions The obtained results demonstrate high contamination levels for THg in sediments of Taranto coastal area. Extreme concentrations of THg were found in the stations located close to the Military Navy Arsenal, Harbour and shipbuilding activities. Me–Hg concentrations, obtained in this study, are characteristic of anoxic polluted sediments. In fact in anoxic sediments, conditions do not appear to promote in situ methylation (Gilmour and Henry, 1991). However, it is important that the rates of methylation processes and pools of mercury which may potentially be methylated are continuously monitored. THg concentrations in edible species were below limit established by the European Community (0.5 mg/kg w.w.) with the exception to gastropod mollusks Hexaplex t., sampled in the station 3 localized near to the Military Navy Arsenal (Mar Piccolo). From obtained results, the risk for human health from fish and seafood consumption appears to be a real issue, especially for children. In fact, the estimated weekly intake for THg and Me–Hg was mostly over the Provisional Tolerable Weekly Intake (PTWI) established by European Food Safety Authority (EFSA) for all edible species, and in lower measure for Mytilus g. In general, the risk of excessive mercury intake is high based on the fish and seafood consumption pattern in Taranto population. In particular, fishermen and their families, or seafood-lovers who consume more seafood have higher risk of mercury intake so, to those people, the reduction of fish and seafood consumption is recommended. In this perspective, it is imperative that continuous monitoring of mercury and methylmercury levels in all foods be done, with crucial attention to fish and seafood, considered a primary vector of these substances for man.

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