Stenella Coeruleoalba) from the Mediterranean Sea (Southern Italy)

Environmental Pollution 116 (2002) 265–271 www.elsevier.com/locate/envpol

Accumulation and tissue distribution of mercury and selenium in striped dolphins (Stenella coeruleoalba) from the Mediterranean Sea (southern Italy)

N. Cardellicchio *, A. Decataldo, A. Di Leo, A. Misino CNR — Istituto Sperimentale Talassograﬁco, via Roma 3, I-74100 Taranto, Italy

Received 13 October 2000; accepted 6 March 2001

Abstract Tissues and organs from Stenella coeruleoalba stranded along the Apulian coasts (southern Italy) during the period April–July 1991 were analyzed for their mercury and selenium content. Analysis showed considerable variations in the mercury concentration in the examined organs and tissues. The highest concentrations of mercury were found in the liver (from 2.27 to 374.50 mgg1 wet wt.). After the liver, lung, kidney, muscle and brain were the most contaminated, while the lowest mercury contamination was found in the melon. As mercury, the liver also showed the highest selenium levels. Liver samples were also analyzed for their methyl mercury contents. The role of selenium in detoxiﬁcation process of methyl mercury has been discussed. Mercury concentrations related to geographic variations and pollution of the marine environment have been examined. The possible implications between mercury accumulation and dolphin death have also been discussed. # 2001 Elsevier Science Ltd. All rights reserved. Keywords: Mercury; Selenium; Mediterranean; Dolphin; Accumulation

1. Introduction monitoring researches on stranded animals both in the anatomo-pathological and in chemico-toxicological The study of mercury accumulation in dolphins is field have been carried out (Andre´ et al., 1991; Leonzio particularly interesting from the ecotoxicological point et al., 1992; Augier et al., 1993b; Cardellicchio, 1995; of view because of the position of these organisms at the Monaci et al., 1998; Capelli et al., 2000; Cardellicchio et end of the trophic networks. Due to its persistence and al., 2000; Frodello et al., 2000). The correlations between high mobility in the marine ecosystem, mercury shows a contaminant accumulation and observed pathologies are high level of biomagnification in the upper levels of the actually a very important topic of researches. food chain. The dolphins of the Mediterranean have In this paper, a study on mercury and selenium dis- much higher levels of mercury and selenium than those tribution and accumulation in tissues and organs of 10 of the Pacific and Atlantic (Andre´ et al., 1991; Augier et specimens of Stenella coeruleoalba stranded along the al., 1993a). For mercury, this can be explained by the Apulian coasts (south-eastern Italy) during the period presence of cinnabar deposits in the Mediterranean Sea: April–July 1991 is presented. As mercury in muscles of the use of mercury in industrial activities may also con- cetaceans is found as methyl mercury (Itano et al., 1984 tribute to increase mercury levels in the marine envir- a, b) and then detoxified by demethylation in the liver onment (Bacci, 1989; Andre´ et al., 1991). and stored in this organ as mercury selenide (Koeman et Dolphins stranded along the coasts are nowadays a al., 1973; Martoja and Berry, 1980; Nigro, 1994), liver noteworthy source of information about physiology and samples have been analyzed for both mercury and biology of these organisms: for this reason, various methyl mercury contents. The role of selenium in the detoxification process of methyl mercury has been dis- * Corresponding author. Tel.: +39-99-4542-208; fax: +39-99- cussed. Finally mercury and selenium levels have been 4542-215. compared with those found in striped dolphins from E-mail address: [email protected] (N. Cardellicchio). other marine areas.

2. Materials and methods Baker) H2SO4-HNO3 mixture (1:1) for 4 h at 160 C. Mercury concentrations were determined by cold Fig. 1 shows the coastal area where the cetaceans were vapour atomic absorption spectrophotometry using a found. Table 1 reports the main morphological char- Perkin Elmer mod. 1100 B spectrophotometer. acteristics of 10 specimens analyzed, together with the After acid digestion of the homogenized liver with 10 date and the location of stranding. Classification of the ml of Ultrex-grade (J.T. Baker) HNO3 (4 h at 160 C), individuals in three classes (calves, young, adults) was selenium was determined by graphite furnace atomic based on the morphological data of body length, since it absorption spectrophotometry (GF-AAS) using a Per- was not possible to obtain teeth for aging. The rela- kin Elmer 3030 Z spectrophotometer. tionship between the age and total length of dolphins The validity of analytical methods was confirmed with was discussed by Miyazaki, (1977), Miyazaki et al., certified Standard Reference Materials (DOLT-2: dog- (1981), and Andre´ et al., (1990, 1991). According to the fish liver) obtained from the National Research Council authors, there is a considerable growth in dolphins dur- of Canada. Results of quality controls are reported in ing the first 4–5 years, and the total length of the indi- Table 2 and show a good agreement with certified data. viduals at that age corresponds to more than 80% of the Methyl mercury (HgMet) was determined by high average size measured after 20 years. Males and females pressure liquid cromathography coupled with cold reach sexual maturity at a length of 190–210 cm and vapour atomic absorption spectrophotometry (HPLC- 187–206 cm, respectively (Calzada et al., 1996, 1997). In CV-AAS) after an acid hydrolysis step at room tem- the Mediterranean Stenella coeruleoalba is usually 10% perature (RT) proposed by Palmisano et al. (1993). H2O shorter than the specimens from the Pacific (Andre´ et (5.5 ml), 1 g of NaCl and 1.5 ml of concentrated HCl al., 1991): therefore, it is not possible to obtain a general were added to 1 g (wet wt.) of homogenized liver. The equation between age and length for all species of dol- sample was sonicated for 30 min and centrifuged at phins from different marine areas. In this work, indivi- 5500g for 10 min. An aliquot of the supernatant was duals with lengths of approximately 120 cm were filtered through a 0.45-mm membrane, diluted as neces- considered as calves; the male specimen n. 4, whose sary with mobile phase (40 mmol cysteine in 0.1 mol/l body length was 168 cm, was considered as young; acetic acid at pH 2.9) and analyzed by HPLC-CV-AAS. females longer than 183 cm and males longer than 190 With this method a fraction of inorganic mercury cm were considered as adults. Organ and tissue samples (liver, brain, kidney, lung, Table 1 muscle, blubber, and melon) were collected during Main characteristics of striped dolphins (Stenella coeruleoalba) analy- autopsy. After collection, samples were transferred to zeda PTFE-containers and frozen at 20C. Before the analysis, samples were homogenized in a teflon Ultra-Turrax Specimen Sex Length Age Location of Date of (cm) stranding sampling T25 homogenizer (Janke & Kunkel, Staufen, Germany). For total mercury determination, samples were digested 1 M 195 A La Specchiolla 18 June 1991 under pressure with a 10 ml of a Ultrex-grade (J.T. 2 M 111 C Minervino 20 June 1991 3 M 208 A Gallipoli 29 June 1991 4 M 168 Y Porto Cesareo 18 July 1991 5 F 137 C Castrignano 21 June 1991 6 F 208 A Gallipoli 6 July 1991 7 F 195 A S. Caterina- Nardo` 20 July 1991 8 F 183 A S. Giovanni- Ugento 24 July 1991 9 F 190 A Chiatona 10 April 1991 10 F 208 A Castellaneta 25 April 1991

a C, calf; Y, young; A, adult.

Table 2 Precision and accuracy of analytical methods obtained using a certiﬁed dogﬁsh liver (DOLT-2)a

Metals DOLT-2

Certiﬁed Foundb

Total mercury 1.99 0.10 1.85 0.20 HgMet (as Hg) 0.693 0.053 0.750 0.100 Selenium 6.06 0.49 6.27 0.75

Fig. 1. Coastal area in the Mediterranean Sea where dolphins (Ste- a The concentrations are given in mgg1 dry weight. nella coeruleoalba) were found. b Number of replicates is ﬁve. N. Cardellicchio et al. / Environmental Pollution 116 (2002) 265–271 267

Table 3 Table 4 Average concentrations (mg/g wet wt.), standard deviation (in par- Mercury and selenium compounds in the liver of striped dolphins (mg entheses) and range of mercury and selenium in Stenella coeruleoalba g1 wet wt.)

Tissue Hg Se Specimensa Hg HgMet %HgMet HgPro Hg* Se Se/Hg*

Liver 170.76 (138.90) 2.27–374.50 63.18 (50.52) 1.90–141.00 10 374.50 6.80 1.82 8.7 359.00 141.00 1.00 Muscle 8.61 (9.31) 0.44–28.00 4.43 (3.39) 0.92–11.00 7 351.86 10.00 2.84 23 318.86 127.00 1.01 Kidney 8.99 (6.84) 1.49–23.78 7.68 (3.06) 3.21–12.92 3 263.00 8.00 3.04 5.5 249.50 92.10 0.94 Brain 8.04 (8.11) 0.22–26.26 5.84 (3.63) 1.63–12.74 6 242.00 7.00 2.89 9.6 225.40 81.00 0.91 Blubber 0.78 (0.87) 0.04–2.40 2.61 (2.18) 0.14–6.07 9 183.06 7.90 4.32 15.7 159.46 71.10 1.13 Melon 0.22 (0.23) 0.05–0.70 N.D.a 1 168.00 4.60 2.74 6.8 156.60 72.30 1.17 Lung 14.52 (12.40) 0.41–36.16 5.47 (3.04) 2.29–11.13 8 107.35 6.30 5.87 8.8 92.25 39.00 1.07 4 12.33 1.60 12.98 1.1 9.63 4.50 1.19 a N.D., no detectable. 5 3.22 0.70 21.74 0.8 1.72 1.90 2.81 2 2.27 0.80 35.24 0.7 0.77 1.90 6.27 (Hg2+), probably derived by mercury weakly bound to a proteins or to other compounds like thiols, can be deter- Specimens are listed in order of decreasing mercury concentration. HgMet, methyl mercury; HgPro, mercury weakly bound to proteins; mined after acid hydrolysis step at room temperature. Hg*, [Hgtot(HgMet+HgPro)]. According to Palmisano et al., (1993), we have called this mercury fraction as HgPro. HgPro and HgMet were The limit of tolerance for Hg in mammalian hepatic simultaneously detected by a mercury-specific detector. tissue seems to be within the range 100–400 mgg1 wet This method has an absolute detection limit of 0.8 ng weight (Wagemann and Muir, 1984); Hg concentrations (as Hg) for HgMet; no organic solvent extraction is reported in Table 3 fall within this range. However, in required, the liver hydrolysate being directly injected this range of concentrations Rawson et al. (1993) have into the chromatographic system. Quantitation of dif- found in dolphins (Tursiops truncatus) liver abnormal- ferent species was performed by peak-area measurement ities associated with chronic mercury accumulation. using daily constructed calibration curves. Mercury accumulated in the liver cells and inhibited the activity of lysosomal digestive enzymes; this would sug- gest that reduced degradation of proteins also might 3. Results and discussion lead to excessive accumulation of lipofuscin within cells. Mercury absorption in dolphins can occur via pul- Mercury and selenium concentrations in different tis- monary, cutaneous and digestive barriers (Andre´ et al., sues and organs examined are shown in Table 3. The 1990). Many authors consider the third contamination decreasing order of mercury concentrations was liver route, via consumed prey, to be the most significant in >> lung > kidney=muscle=brain. The level of mer- relation to the bioaccumulation of mercury. Marine cury contamination in the liver is clearly the highest mammals are top predators in the food chain and are one, indeed it is about 12 times greater than the average long-lived, and tend to have much higher Hg con- concentration measured in the organ with the second centrations than other marine organisms. Inorganic highest level (the lung). The lowest mercury concentra- compounds and metallic form Hg are absorbed to only tion was found in the melon. The liver showed also the a small degree, compared with organic forms, in parti- highest selenium content. cular methyl mercury. For S. coeruleoalba, Itano and In Table 4, mercury compounds and selenium con- Kawai (1981) estimated a methyl mercury biological centrations in the liver of the dolphins are reported. half-life of about 1000 days. In agreement with other Compared with the total mercury, the percentage of observations (Viale, 1978), HgMet only accounts for a methyl mercury in adult specimens ranged from 1.8 to fraction of total Hg present in the liver, being less than 5.9%. In the young specimens, the methyl mercury per- 10% of total mercury in adult specimens. In this work, centage was higher (from 13 to 35%). the results indicate that in the liver at the lowest total Table 5 shows a comparison among mercury and Hg concentrations correspond the highest percentages selenium concentrations found in dolphins from differ- of HgMet. Fig. 2 shows the comparison of total Hg ent marine areas. Data expressed in dry weight were concentration and HgMet percentage in the liver of converted to wet weight using the factor (ww/dw) of dolphins examined: it clearly appears that above a con- 0.25 established for dolphins (Becker et al., 1995). The centration of approximately 100 mg/g Hgtot, the pro- data should be considered with care as there are many portion of HgMet becomes relatively independent of the potential sources of variations due to condition, age, sex total Hg amount. When the level of mercury becomes of dolphins, as well as to sampling methods and analy- relatively high, biological demethylation of HgMet tical techniques used. Anyway, it is clear that the highest takes place. The mechanism of demethylation process in values of mercury were found by different authors in the the liver, has not been explained yet. Mercury demethyla- liver of dolphins. tion is probably activated when HgMet concentration 268 N. Cardellicchio et al. / Environmental Pollution 116 (2002) 265–271

1 Fig. 2. Comparison between total Hg (mgg1 wet wt.) and %HgMet Fig. 4. Comparison between molar ratio Se/Hg* and total Hg (mgg in the liver of striped dolphins. wet wt.) in the liver of striped dolphins.

Fig. 3. Comparison between total Hg and Se in the liver of striped Fig. 5. Correlation between Hg* and Se concentrations in the liver of dolphins (mgg1 wet wt.). striped dolphins (mgg1 wet wt.).

exceeds a threshold value of 100 mg/g Hgtot (Palmisano HgSe (tiemannite), an insoluble and little toxic com- et al., 1995). It has been proposed that at high HgMet pound. The presence of HgSe in the cytoplasm of dol- concentrations the liver accumulates selenium, which is phin hepatic cells has been confirmed by X-ray somehow involved in the detoxication process. The dis- spectrography in Ziphius cavirostris (Martoja and Berry, covery of the protective effect of selenium on mercury 1980) and in S. coeruleoalba (Nigro, 1994). In the young toxicity is attributed to Parizek and Ostadalova (1967). specimens (Table 4) the HgMet percentage is high (from Selenium plays an important role in the enzymatic 13 to 35%) and Se/Hg* ratio is in the range 1.19–6.27. activities, e.g. glutathione peroxidase. The relation In these specimens, the HgMet demethylation has between mercury and selenium accumulation in marine probably not been activated yet. mammals was found by (Koeman et al., 1973; Martoja Hg and Se regression equation by least square treat- and Viale, 1977; Wagemann et al., 1983, 1988; Itano et ment is shown in Fig. 5. The slope value of the equation al. 1984a, b, 1985; Martoja and Martoja, 1985; Muir et is 2.58 according to other authors who found slope al., 1988; Capelli et al. 1989; Augier et al. 1993b). The values from 2.59 to 3.72 in the liver of S. coeruleoalba results shown in Fig. 3 indicate that increasing Hg con- (Thibaud, 1978; Itano et al., 1984b). Least square treat- centrations are associated with increasing Se levels. ment of koeman’s data (Koeman et al., 1973) gives the According to Koeman et al. (1973), the molar ratio Se/ slope value of 2.70. 0 0 Hg (Hg =HgtotHgMet) in the liver of dolphins The detoxification process of mercury in dolphin liver (Delphinus delphis) was approximately 1. In this work, could be due not only to HgSe formation but also to we have calculated the fraction of inorganic mercury complexation with metallothioneins (MT) or thiols like (Hg*) probably linked to selenium by difference glutathione (GSH) (Andre´ et al., 1990; Caurant et al., between total Hg and the sum of HgMet and HgPro 1996). Different metallothioneins able to bind Cd, Zn, (mercury weakly bound to proteins or thiols). Fig. 4 Hg and Cu have been identified in marine mammals shows the variation in the molar ratio Se/Hg* versus the (Olafson and Tompson, 1974; Lee et al., 1977; Ridling- total Hg concentration in dolphin liver samples. The ton et al., 1981; Mochizuki et al., 1985; Tohyama et al., average value of this ratio was 1.03 for adult specimens, 1986). However, Gerson and Shaikh (1982) have repor- in which the threshold of 100 mg/g Hgtot has been ted that a maximum of about 10% of intracellular Hg exceeded. Such observations led to the hypothesis that was found associated with thioneins in hepatocytes of the final compound of HgMet demethylation could be rats exposed to 30 mmol-Hg. They also reported that Hg N. Cardellicchio et al. / Environmental Pollution 116 (2002) 265–271 269

Table 5 Mercury and selenium concentrations (mgg1 wet wt.) in dolphins (compiled from the sources indicated)

Species Locality Tissues Hg Se Reference

Stenella coeruleoalba Japanese coasts Liver 20.5 Honda et al., 1983 Muscle 7 Kidney 8.7

S. coeruleoalba Japanese coasts Liver 205 Itano et al., 1984b Kidney 14.7

S. coeruleoalba French Atlantic coasts Liver 1.2–87 Andre´ et al., 1991

S. coeruleoalba French Mediterranean Liver 668.4 Andre´ et al., 1991 coasts Kidney 87.2

S. coeruleoalba French Mediterranean Liver 17–568a Augier et al., 1993b coasts Muscle 1.8–38.8a Kidney 3.5–85.3a Brain 1–20.3a Lung 0.8–99a Heart 1–25a

S. coeruleoalba Mediterranean coasts Liver 260.8a 25.2a Monaci et al., 1998 of Spain Muscle 6.9a 1.8a Kidney 15.7a 12.8a Brain 2.3a 3.5a Skin 2.7a 20.25a

S. coeruleoalba Mediterranean coasts Liver 148.2a 66.5a Monaci et al., 1998 of Italy Muscle 13.2a 2.9a Kidney 10.9a 6.5a Brain 0.7a 1.3a Skin 2.5a 26.9a

S. coeruleoalba Italian Mediterranean Liver 156.2–216.7 65.8–92.4 Cardellicchio et al., 2000 coasts (Adriatic and Muscle 8.1–14.4 Ionian coasts) Kidney 7.6–13.1 Brain 6.9–22.0 Blubber 0.6–2.9 Melon 1.3–4.1 Lung 15.5–51.9

S. coeruleoalba Corsican coasts Liver 115a Frodello et al., 2000 Muscle 5.3a kidney 12.3a Lung 9a Skin 4.8a Bone 0.5a

S. coeruleoalba Italian Mediterranean Liver 3.3–748.3a 1.8–276.3a Capelli et al., 2000 coasts (Ligurian Sea) Muscle 1.3–65a 0.5–25a Kidney 1.3–40.8a 1.3–22.8a Brain 0.8–49a 0.5–18.3a Lung 0.5–112a 1–33.3a Heart 1.3–30a 0.5–6.3a Spleen 1–329a 2.3–107.8a

a Value expressed as mgg1 wet weight after conversion from dry weight. weakly stimulated the induction of thioneins, and did yet been studied in marine mammals. It has been not preferentially bind to them. Therefore, in mammals, demonstrated in other mammals that these thiols con- metallothioneins seem not to have an important role in jugated methyl mercury in the liver and excreted the the subcellular process of detoxiﬁcation from mercury. complex in the bile (Hirata and Takahashi, 1981; Bal- The role of GSH in the mercury detoxiﬁcation has not latori and Clarkson, 1984a, b; Di Simplicio et al., 1990). 270 N. Cardellicchio et al. / Environmental Pollution 116 (2002) 265–271

Table 5 shows that geographical area influences metal surveys on metal accumulation in stranded dolphins accumulation in dolphins: in fact, mercury levels in may contribute to know the replies of cetaceans to the dolphins from the Mediterranean are generally higher contamination of marine ecosystem. than those found in the same species from the Atlantic (Andre´ et al., 1991). This has been explained by the existence of natural mercury sources in the Mediterra- References nean Sea and mercury rich inputs via rivers flowing through the Mt. Amiata area (one of the richest natural Andre´ , J.M., Ribeyre, F., Boudou, A., 1990. Mercury contamination levels and distribution in tissues and organs of Delphinids (Stenella reserves of cinnabar; Bacci, 1989). In fact, very high attenuata) from the eastern tropical Pacific in relation to biological mercury concentrations have been found in fish tissues and ecological factors. Mar. Environ. Res. 30, 43–72. from the Tyrrhenian Sea (NW Mediterranean; Bern- Andre´ , J., Boudou, A., Ribeyre, F., Bernhard, M., 1991. Comparative hard, 1988). Mercury levels in dolphins stranded on the study of mercury accumulation in dolphins (Stenella coeruleoalba) south eastern Mediterranean coasts of Italy (Cardellic- from French Atlantic and Mediterranean coasts. Sci. Tot. Environ. 104, 191–209. chio et al., 2000) are generally lower than those found in Augier, H., Park, W.K., Ronneau, C., 1993a. Mercury contamination dolphins from the French Mediterranean coasts (Andre´ of the striped dolphin Stenella coeruleoalba Meyen from French et al., 1991; Augier et al., 1993a) and from the northern Mediterranean coasts. Mar. Poll. Bull. 26 (6), 306–311. Mediterranean coasts of Italy (Ligurian Sea; Capelli et Augier, H., Benkoel, L., Brisse, J., Chamlian, A., Park, W.K., 1993b. al., 2000). However, mercury concentrations found in Necroscopic localization of mercury–selenium interaction products in liver, kidney, lung and brain of Mediterranean striped dolphins this work, are comparable to those found in S. coer- (Stenella coeruleoalba) by silver enhancement kit. Cell. and Molec. uleoalba from the Japanese coasts (Honda et al., 1983; Biology 39, 765–772. Itano et al., 1984b), even though in this case animals Bacci, E., 1989. Mercury in the Mediterranean. Mar. Poll. Bull. 20 (2), had been captured. 59–63. The results of this work show that sex has no sig- Ballatori, N., Clarkson, T.W., 1984a. Dependence of biliary secretion of inorganic mercury on the biliary transport of glutathione. Bio- nificant influence on the total mercury concentrations in chem. Pharmacol. 33, 1093–1098. tissues and organs analysed. Various authors (Gaskin et Ballatori, N., Clarkson, T.W., 1984b. Inorganic mercury secretion into al., 1979; Honda et al., 1983) agree that no influence of bile as a low molecular weight complex. Biochem. Pharmacol. 33, sex on mercury accumulation is observed in marine 1087–1092. mammals. On the other hand, Andre´ et al. (1990) found Becker, P.R., Mackey, E.A., Demiralp, R., Suydam, R., Early, G., Koster, B.J., Wise, S.A., 1995. Relationship of silver with selenium differences in mercury concentration between males and and mercury in the liver of two species of toothed whales (odonto- females of 44 dolphins from the eastern tropical Pacific. cetes). Mar. Poll. Bull. 30, 262–271. Pathological, microbiological and parasitological Bernhard, M., 1988. Mercury in the Mediterranean. UNEP Reg. Seas studies were performed on the dolphins analyzed. The Rep. Studies, No. 98. animals were found to be affected by a wide range of Calzada, N., Aguilar, A., Lockyer, C., Grau, E., 1997. Patterns of growth and physical maturity in the western Mediterranean striped conditions. The majority of histopathological lesions dolphin Stenella coeruleoalba (Cetacea: Odontoceti). Can. J. Zool. were caused by parasitic infections of the respiratory 75, 632–637. tract, the alimentary system or the hepatic system. Calzada, N., Aguilar, A., Sorensen, T.B., Lockyer, C., 1996. Repro- Three dolphins had endocarditis, two other specimens ductive biology of female striped dolphin Stenella coeruleoalba from were affected by hemorrhagic gastritis. Specimen No. 2 the western Mediterranean. J. Zool. 240, 581–591. Capelli, R., De Pellegrini, R., Minganti, V., 1989. Preliminary results showed the lobes of the caudal fin removed, and con- on the presence of inorganic, organic mercury and selenium in dol- sequential traumatic lesions; probably the dolphin was phins (Stenella coeruleoalba) from the Ligurian sea. Third An. Conf. entangled in the nets and was released by the same fish- Eur. Cetacean Soc., La Rochelle, France 24-26 fe´ v., 19–24. ermen through a clean cut. Capelli, R., Drava, G., De Pellegrini, R., Minganti, V., Poggi, R., It is impossible to determine from a study such as this 2000. Study of trace elements in organs and tissues of striped dolphins (Stenella coeruleoalba) found dead along the Ligurian coasts whether even the highest concentrations of mercury (Italy). Adv. Environ. Res. 4, 31–43. found increased the susceptibility of the animals to Cardellicchio, N., 1995. Persistent contaminants in dolphins: an indi- infectious diseases. For the specimens examined, infec- cation of chemical pollution in the Mediterranean sea. Water tive diseases or parasitic infections cannot be considered Science and Technology 32 (9), 331–340. as causes of death on the basis of necroscopic and Cardellicchio, N., Giandomenico, S., Ragone, P., Di Leo, A., 2000. Tissue distribution of metals in striped dolphins (Stenella coer- microbiologic surveys. Although dolphins may be able uleoalba) from the Apulian coasts, southern Italy. Mar. Environ. to tolerate high metal concentrations, there is no infor- Res. 49, 55–66. mation about long-term or sublethal impact of Hg Caurant, F., Navarro, M., Amiard, J.C., 1996. Mercury in pilot bioaccumulation. Even if concentrations of mercury in whales: possible limits to the detoxification process. Sci. Total the livers of these animals were below the limit of toler- Environ. 186, 95–104. Di Simplicio, P., Gorelli, M., Ciuffreda, P., Leonzio, C., 1990. The ance for Hg in mammalian hepatic tissue, it may be relatioinship between gamma-glutamyl transpeptidase and Hg levels reasonably supposed that high mercury contents can in Se/Hg antagonism in mouse liver and kidney. Pharmacol. Res. disturb the normal physiology of the animals. Anyway, 22, 515– 526. N. Cardellicchio et al. / Environmental Pollution 116 (2002) 265–271 271

Frodello, J.P., Rome´ o, M., Viale, D., 2000. Distribution of mercury in Miyazaki, N., Fujise, Y., Fujiyama, T., 1981. Body and organ weights the organs and tissues of five toothed-whale species of the Medi- of striped and spotted dolphins off the Pacific coast of Japan. Sci. terranean. Environ. Poll. 108, 447–452. Rep. Whales Res. Inst. (Tokyo) 33, 27–67. Gaskin, D.E., Stonefiel, K.I., Suda, P., Frank, R., 1979. Changes in Mochizuki, Y., Suzuki, K.T., Sunaga, H., Kobayashi, T., Doi, R., 1985. mercury levels in harbour porpoises from the bay of Fundy, Canada Separation and characterization of metallothionein in two species of and adjacent waters during 1969-77. Arch. Environ. Con. Tox. 8, seals by high performance liquid chromatography-atomic absorption 733–762. spectrophotometry. Comp. Biochem. Physiol. 82C, 248–254. Gerson, R.J., Shaikh, Z.A., 1982. Uptake and binding of cadmium Monaci, F., Borrel, A., Leonzio, C., Marsili, L., Calzada, N., 1998. and mercury to metallothionein in rat hepatocyto primary cultures. Trace elements in striped dolphins (Stenella coeruleoalba) from the Biochem. J. 208, 465–472. western Mediterranean. Environ. Poll. 99, 61–68. Hirata, E., Takahashi, H., 1981. Degradation of methyl mercury glu- Muir, D.C.G., Wageman, R., Grift, N.P., Notstrom, R.J., Simon, M., tathione by the pancreatic enzymes in bile. Toxicol. Appl. Pharma- Lien, J., 1988. Organochlorine chemical and heavy metal con- col. 58, 483–491. taminants in white-beaked dolphins (Lagenorhynchus albirostris) Honda, K., Tatsukava, R., Itano, K., Miyazaki, N., Fujiyama, T., and pilot whales (Globicephala melaena) from the coast of New- 1983. Heavy metals concentration in muscle, liver and kidney tissue foundland, Canada. Arch. Environ. Contam. Toxicol 17, 613–629. of striped dolphin Stenella coeruleoalba and their variation with Nigro, M., 1994. Mercury and selenium localization in macrophages body length, weight, age and sex. Agric. Biol. Chem. 47 (6), 1219– of the striped dolphin, Stenella coeruleoalba. J. Mar. Biol. Ass. 74, 1228. 975–978. Itano, K., Kawai, S., 1981. Changes of mercury contents and biologi- Olafson, R.W., Thompson, J.A.J., 1974. Isolation of heavy metal cal half-life of mercury in the striped dolphins. In: Fujiyama, N. binding proteins from marine vertebrates. Mar. Biol. 28, 83–86. (Ed). Studies on the Levels and Organochorine Compounds and Palmisano, F., Cardellicchio, N., Zambonin, P.G., 1993. Speciation Heavy Metals in the Marine Organisms. University of the Ryukyus, and simultaneous determination of mercury species in dolphin liver Nishihara, Okinawa, pp. 49–73. by liquid chromatography with on-line cold vapor atomic absorp- Itano, K., Kawai, S., Tatsukawa, R., 1985. Properties of mercury and tion spectrometry. Fresenius’ J. Anal. Chem. 346, 648–652. selenium in salt-insoluble fraction of muscle in striped dolphins. Palmisano, F., Cardellicchio, N., Zambonin, P.G., 1995. Speciation of Bull. Jap. Soc. Sci. Fish 51, 1129–1131. mercury in dolphin liver. A two-stage mechanism for the demethy- Itano, K., Kawai, S., Miyazaki, N., Tatsukawa, R., Fujiyama, T., lation accumulation process and role of selenium. Mar. Environ. 1984a. Body burdens and distribution of mercury and selenium in Res. 40, 109–121. striped dolphins. Agric. Biol. Chem. 48 (5), 1117–1121. Parizek, J., Ostadalova, I., 1967. The protective effect of small amounts Itano, K., Kawai, S., Miyazaki, N., Tatsukawa, R., Fujiyama, T., of selenite in sublimate intoxication. Experientia 23, 142–143. 1984b. Mercury and selenium levels in striped dolphins caught Rawson, A.J., Patton, G.W., Hofmann, S., Petra, G.G., Johns, L., on the Pacific coast of Japan. Agric. Biol. Chem. 48 (5), 1109– 1993. Liver abnormalities associate with chronic mercury accumu- 1116. lation in stranded atlantic bottlenose dolphins. Ecotox. and Koeman, J.H., Peeters, W.H.M., Koudstaal-Hol, C.H.M., Tjioe, P.S., Environ. Safety 25, 41–47. de Goeij, J.J.M., 1973. Mercury-selenium correlations in marine Ridlington, J.W., Chapman, D.C., Goeger, D.E., Whanger, P.D., mammals. Nature 245, 385–386. 1981. Metallothionein and Cu-chelatin: characterization of metal- Lee, S.S., Mate, B.R., von der Trench, K.T., Rimerman, R.A., Buhler, binding proteins from tissues of four marine mammals. Comp. Bio- D.R., 1977. Metallothionein and subcellular localization of mercury chem. Physiol. 70B, 93–104. and cadmium in the Californian sea lion. Comp. Biochem. Physiol. Thibaud, Y., 1978. Pre´ sence simultane´ e de mercure et de se´ lenium chez 57, 45–53. le dauphin et le thon rouge. IVa J. Et. Pollut. Comm. Inter. Et. Sci. Leonzio, C., Focardi, S., Fossi, C., 1992. Heavy metals and selenium Mer Me´ dit., Antalya, Turquie, 193–196. in stranded dolphins of the Northern Tyrrhenian (NW Mediterra- Tohyama, C., Himeno, S.J., Watanabe, C., Suzuki, T., Morita, M., nean). Sci.Tot. Environ. 119, 77–84. 1986. The relationship of the increased level of metallothionein with Martoja, R., Berry, J.P., 1980. Identification of tiemannite as a prob- heavy metal levels in the tissue of the harbor seal (Phoca vitulina). able product of demethylation of mercury by selenium in cetaceans. Ecotox. Environ. Safety 12, 85–94. A complement to the scheme of the biological cycle of mercury. Vie Viale, D., 1978. Evidence of metal pollution in Cetacea of the western Milieu 30 (1), 7–10. Mediterranean. Ann. Inst. Oce´ anogr. Paris 54 (1), 5–16. Martoja, M., Martoja, R., 1985. La bioaccumulation des me´ taux: Wagemann, R., Muir, D.C.G., 1984. Concentrations of heavy metals processus physiologique normal et conse´ quence de la pollution. Le and organochlorines in marine mammals of northern waters: over- Courrier du CNRS 54, 32–37. view and evaluation. Can. Tech. Rep. Fish Aqu. Sci. 1279. Martoja, R., Viale, D., 1977. Accumulation de granules de se´ le´ niure Wagemann, R., Snow, N.B., Lutz, A., Scott, D.P., 1983. Heavy metals mercurique dans le foie d’Otontoce` tes (Mammife` res Ce´ tace´ s): un in tissues and organs of the narwhal Monodon monoceros. Can., J. me´ canisme possible de de´ toxication du me´ thyl — mercure par le Fish aquat. Sci. 40, 206–214. se´ le´ nium. C.r. Acad. Sci. 285, 109–112. Wagemann, R., Stewart, R.E.A., Lockhart, W.L., Stewart, B.E., 1988. Miyazaki, N., 1977. Growth and reproduction of Stenella coeruleoalba Trace metals aand methyl mercury: associations and trasfrer in harp off the Pacific coast of Japan. Sci. Rep. Whales Res. Inst. (Tokyo) seal (Phoca groenlandica) mothers and their pups. Mar. Mammal 29, 21–48. Sci. 4, 339–355.