Element content in cultured and wild sea bass ( Dicentrarchus labrax ) from the Adriatic Sea

The aim of this study was to determine the levels of potentially toxic elements in cultured and wild fish tissues and to assess their risk for human health. For this purpose, sea bass specimens (Dicentrarchus labrax) were sampled in selected fish farm and three other locations along the eastern Adriatic coast. Ranges of element concentrations in sea bass muscles were 1.60-4.46 ppm for As, 0.001-0.079 ppm for Cd, 0.14-49.10 ppm for Cr, 1.38-4.85 ppm for Cu, 0.11-1.31 ppm for Hg, 0.01-0.65 ppm for Pb and 21.9136.0 ppm for Zn. Mean Cd, Cr, Cu, Hg, Pb, and Zn concentrations in commercially interesting cultured fish samples were below the permissible levels, while mean As values slightly exceed those limits. In wild fishes mean Cd, Cu, Pb and Zn concentrations were below the recommended limits, for As, Cr and Hg the mean values were higher. The smallest cultured sea bass samples showed As, Cr, Pb, and Zn concentrations exceeding the recommended limits but values decreased with fish size. Therefore, the metal concentrations in commercial fishes showed no threat for human consumption.


Introduction
Among several different contaminants, metals are recognized as one of the greatest threats to organisms due to their persistence and possible bioaccumulation and biomagnification in the food chain (Demirak et al., 2006;Uysal et al., 2009).Some metals as mercury (Hg), lead (Pb), and cadmium (Cd) are considered to be very dangerous, while some other essential elements, such as copper (Cu), nickel (Ni), chromium (Cr), and zinc (Zn), can also cause toxic effects at excessively elevated values (Cid et al., 2001 and references therein).Food presents the main and a constant source of potentially toxic trace metals in different parts of human body and can cause damage in many of its basic systems (renal, cardiovascular, gastrointestinal, endocrine, nervous, etc. (Cid et al., 2001 and references therein).In Mediterranean countries, especially in coastal areas, human nutrition is highly represented by sea products including fishes.Therefore, the determination of trace metals in fish tissues is necessary.Numerous researches focused to investigation of potentially toxic metals in marine organisms as well as their potential sources (sea water, food) (Ersoy et al., 2006;Burger et al., 2007;Dural et al., 2007;Biladžić et al., 2011;Fallah et al., 2011;Mieiro et al., 2011;Percın et et al., 2011 and many others).
In our study, levels of trace metal contamination of cultured and wild sea bass were determined.The investigation focused (1) in assess the differences between cultured and wild specimens, (2) to ascertain the possible differences between different fish sizes and (3) to observe possible risk for human health (measured values were compared with guidelines proposed by various authorities).

Materials and Methods
Sea bass samples were collected in fish farm near Vrgada Island (Central Adriatic) and three coastal locations in the Adriatic Sea: Korčula Island, Pirovac Bay, and Savudrija (northern Istra Peninsula).The observed fish farm was located around 4 nautical miles from the mainland, south of Vrgada Island, which is located in Murter Sea between Zadar and Šibenik (Croatia).The farm produced about 550 tons of fish (sea bass and sea bream) annually.Production ran throughout the whole year, with new generations being born between April and July.During the winter the amount of feed given was obviously smaller than during the summer, when the breeding of fishes was highest.Korčula Island is located in the southern part of the Central Adriatic Sea between Split and Dubrovnik.Samples were collected at the western side of the island orientated towards the open sea.Pirovac Bay is a semienclosed bay located in the shore area of Central Adriatic between Zadar and Šibenik.Savudrija town is positioned at the shore of northern Istra.These three locations are anthropogenically affected by different sources i.e. untreated municipal and industrial sewage from cities, tourist facilities in the summer etc.
European sea bass (Dicentrarchus labrax) were sampled in 2009 with scuba diving and by catching in the farm.In order to determine the element between fish sizes, four various sea bass specimens, 6 to 25.2 cm in length and 15 to 312.2 g in wet weight, were collected at fish farms.After sampling, fishes were stored in a cool box and later frozen.In the laboratory, fishes were sectioned and freeze-dried for at least 72 h.Dried samples were homogenized and crushed to a fine powder by grinding in an agate mortar.
Fish tissues were analysed for certain trace metals in a certified Canadian commercial laboratory (ACTLABS: Activation Laboratories Ltd., Ancaster, Ontario, Canada) using inductively coupled plasma mass spectrometry (ICP/MS).Dry, ashed samples were dissolved in acid and analysed by High Resolution ICP/MS.Additionally, a special microwave digestion procedure (closed vessel digestion) was used.The analytical precision and accuracy were within ±10% for the analysed elements.This was indicated by the results of the duplicate measurements of both fish samples and standards (NIST 1575a and DORM-2).

Results and Discussion
Measured results show the ranges of element concentrations in sea bass muscles 1.60-4.46ppm for arsenic, 0.001-0.079ppm for cadmium, 0.14-49.10ppm for chromium, 1.38-4.85ppm for cooper, 0.11-1.31ppm for mercury, 0.01-0.65 ppm for lead and 21.9-136.0ppm for zinc.Generally, mean concentrations of As, Cd, Cr, Cu, Pb, and Zn were higher in cultured fishes, while only Hg values were higher in wild specimens.The higher general mean content of cultured fish was mainly the result of the concentrations in the smallest sea bass samples.By ignoring the latter samples, the mean values of cultures specimens were mostly below the wild concentrations.
Several studies show that sizes of marine animals play an important role in metal contents in their tissues.Previous investigations established a significant negative relationship between metal concentrations and fish size (Nussey et al., 2000;Canli & Atli, 2003;Henry et al., 2004).Equally, our results showed generally a negative relationship.The highest metal concentrations were observed in the smallest sea basses and the content of metals decreases with increases in the weight and length of specimens.Thus, for As, Pb, and Zn the lowest concentrations were found in the biggest sea bass.The lowest values of Cd, Cr, Cu, and Hg were measured in the medium size sea bass, while the concentrations in the biggest sea bass were again slightly elevated.This negative relationship in our study between metal content and fish size can be explained by the difference in metabolic activity between younger and older fishes as previously described in similar studies (Nussey et al., 2000;Canli & Atli, 2003).
In order to assess the possible risk for human health, measured values of fish tissues were compared with the limit values proposed by various authorities: the Ministry of Agriculture, Fisheries and Food (MAFF, 2000), the United Nations Food and Agriculture Organization (FAO), which published the "Compilation of Legal limits for Hazardous Substances and Fish and Fishery Products" (Nauen, 1983) and the European Commission (EC, 2001).Mean values of Cd, Cu, Hg, Pb, and Zn were below the permissible levels in cultured and wild fish specimens, with the exception of Hg in wild fish samples and Zn in cultured one, which slightly exceeded them.The mean concentrations of As and Cr obtained in cultured and wild fish specimens were higher than those established by the regulations.Arsenic concentrations were at least for little exceeding the permissible levels in practical all samples, chromium was elevated in wild samples and the smallest cultured fish.Among the wild samples, obviously elevated values of As, Cr, Hg, and Zn were measured in fishes collected at Korčula sampling site and also As and Cr concentrations were higher in Pirovac Bay and Savudrija locations.The elevated values of some metals (As, Cd, Cr, Zn) in cultured fishes were observed in the smallest sea basses.Moreover, the values of potentially commercial cultured fishes were practically below the recommended limits with the only little elevation of arsenic.

Conclusion
If we exclude the smallest cultured sea basses, the concentrations of cultured fish specimens were lower than the wild one, with the exception of cooper.These indicate several different potential natural and anthropogenic sources of elements, such as geologic weathering, fish feed, untreated municipal and industrial sewages, waste from sewage treatment plants, or agricultural waste sewages.
The results of cultured sea basses showed a generally negative relationship between fish size and metal level.Generally, the highest values were observed in smaller fishes and the lowest in the biggest, commercial one.
Mean values of Cd, Cu, Hg, Pb, and Zn were below the permissible levels in cultured and wild fish specimens, with the exception of Hg in wild fish samples and Zn in cultured one.Elevated values of As and Cr of cultured samples were high mainly due to the juvenile 11001-p.2non-commercial sea basses, which also showed elevated concentrations of some other metals (Pb and Zn).In larger, commercial cultured fish samples only As showed slightly higher concentrations regarding to recommended legal limits and were even lower than in wild one.Consequently investigated fishes from farm present no toxically danger for human consumption.
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