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Temporal distribution of heavy metal concentrations in oysters Crassostrea rhizophorae from the central Venezuelan coast
Marine Pollution Bulletin 73 (2013) 394–398
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Marine Pollution Bulletin journal homepage: www.elsevier.com/locate/marpolbul
Baseline
Temporal distribution of heavy metal concentrations in oysters Crassostrea rhizophorae from the central Venezuelan coast Juan A. Alfonso a,⇑, Helga Handt a, Abrahan Mora a, Yaneth Vásquez a, José Azocar a, Eunice Marcano b a b
Centro de Oceanología y Estudios Antárticos, Instituto Venezolano de Investigaciones Científicas (IVIC), Apartado 20632, Caracas 1020A, Venezuela Centro de Química, Instituto Venezolano de Investigaciones Científicas (IVIC), Apartado 20632, Caracas 1020A, Venezuela
a r t i c l e
i n f o
Keywords: Oysters Bivalves Bioindicators Heavy metals
a b s t r a c t The oyster Crassostrea rhizophorae is a bivalve abundant in Venezuelan estuaries and consumed by local populations. No known values have been reported on trace metals in oysters from the central Venezuelan coast. We report the concentrations of Al, Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, Sr, V and Zn in the soft parts of C. rhizophorae, which were collected bimonthly between March 2008 and March 2009, at two sampling areas from the Central Venezuelan Coast: Buche estuary and Mochima estuary. Our results show that for each metal there is a similar temporal variation pattern. The concentrations of the heavy metals reported in this work are useful as reliable baselines and can be used for comparison in future environment studies. Concentrations in C. rhizophorae from the Buche estuary can be interpreted to be high on a global scale for Cd, Cu, Ni and Mn, indicating atypically raised bioavailabilities. Ó 2013 Elsevier Ltd. All rights reserved.
Bivalves have been extensively used as bioindicators to assess coastal aquatic environments, since they can accumulate trace metals and other substances. Although no official monitoring program is in progress, various studies by different groups have provided relevant data with respect to changing metals levels in clams Tivela mactroides from the central Venezuelan coast (Jaffé et al., 1995; LaBrecque et al., 2004a; Alfonso et al., 2005). However, the need to consider other species and taxa has been emphasized for regional, national and international monitoring programs (Ke and Wang, 2001; Baudrimont et al., 2005; Senthil Kumar et al., 2008; Bendell and Feng, 2009; Apeti et al., 2009). Oysters have been used as a companion bivalve species with mussels in the biomonitoring of coastal contamination in National Status and Trend Programs (e.g. the National Status and Trend program in the US, ´ Connor et al., 1994; the Mussel Watch program in France, Cantillo O et al., 1998). The oyster Crassostrea rhizophorae is a bivalve abundant in Venezuelan estuaries and widely consumed by local populations. Only a few studies have been reported on trace metals in oysters from Venezuela. However, such studies have not dealt with temporal variations; they have only considered a limited number of elements, and have been undertaken in restricted areas (See Fig. 1). The marine environment in the central Venezuelan coast is subjected to contamination by heavy metals introduced to this coastal environment by the Tuy river (Jaffé et al., 1995; LaBrecque et al., 2004a; Alfonso et al., 2005), which has a plume known to move ⇑ Corresponding author. Tel.: +58 212 504 1401; fax: +58 212 504 1935. E-mail address: [email protected] (J.A. Alfonso). 0025-326X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.marpolbul.2013.05.010
in a northwesterly direction and receives waste water effluents from the metropolitan area of Caracas, a city of approximately 7 million inhabitants, via the Guaire river. Most of this waste water undergoes only minor treatment. In addition, the Tuy river drainage basin, of approximately 6600 km2, covers an active industrial and agricultural area, which also affects water quality (Jaffé et al., 1995). The use of a bioindicator is a good integrative tool of the ecosystem (Fraser et al., 2011). Indeed, bioindicators, such as oysters, integrate many aspects of their ecosystem including the contamination and external factors which can influence this contamination, such as ocean currents, storms, seasons and water temperature (Markert et al., 1999). The objective of this study is to establish a baseline of the temporal distribution of heavy metals in two areas of the central Venezuelan coast by using the oyster C. rhizophorae as a bioindicator. It is well known that the accumulation of heavy metals in bivalves is affected by age, size and weight of the bivalves (Mubiana et al., 2006; Fialkowski et al., 2009; Spann et al., 2011; McKinley et al., 2012). As these factors were not the objective of our study, they were kept as constant as possible, in order to minimize their influence on the measured concentrations. Each sample consisted of a pool of at least 10 oysters (C. rhizophorae) of 6–7 cm in size. Samples of C. rhizophorae were collected bimonthly between March 2008 and March 2009, using a stainless steel knife at six sampling sites, three located in Buche estuary, Carenero, state of Miranda, and three located in Mochima estuary, state of Sucre. Spacing between sampling sites varied between 400 and 500 m, within each sampling area. Triplicate samples were collected at
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Fig. 1. A map of the Venezuelan coast showing the sampling areas.
each sampling site (10–15 m2). All samples were rinsed with seawater after collection to remove marine sediments, shells and other debris, placed in plastic bags and transported to the laboratory on ice. In the laboratory, the complete soft tissue of oysters was removed from the shell and washed with distilled deionized water to remove impurities. The bivalves tissues were transferred to clean one-liter beakers and washed several times with distilled deionized water until the solution was clear. After discarding the excess liquid, the beakers were placed in a drying oven at about 65 °C until most of the liquid was evaporated, then the temperature was increased to 80 °C for 24 h, after cooled in desiccators. The dried tissue samples were ground into a fine powder with an agate mortar and pestle and well mixed to prepare homogenous samples. One gram portions of the dried tissues were dissolved in 8 ml of SuprapureÒ nitric acid (Merck) by heating on a hot plate at about 70 °C for about 8 h. When the solution cooled, 2 ml of hydrogen peroxide (30%) (Merck) was added and heated again at 70 °C for another 4 h. Finally, the resulting solution was diluted to 50 ml with distilled deionized water. Duplicate digestions were performed for each sample. Method blanks were always low in trace metals indicating minimal contamination during sample
Table 1 Comparison of the determined values (n = 6) in standard reference materials NIST 2977 in this study with the certified (CV), recommended (RV) and information (IV) values. Element NIST 2977
Al Cd Co Cr Cu Fe Mn Ni Pb Sr V Zn
processing, and these concentrations were subtracted from sample readings to give final values of metal concentration. Table 2 Mean, minimum maximum and standard deviation values of the trace element concentrations in dried soft oyster tissue from the different sampling areas during this temporal study (n = 9) in estuaries Buche and Mochima. The p-values in boldface represent significant concentration differences (p<0.05) between the sampling sites. Buche
Mochima
p
Al (lg/g)
63.4 2.8–197.8 26.7
49.05 5.3–131.6 53.4
0.673
Cd (lg/g)
2.5 1.5–4.2 0.9
2.7 1.7–3.5 0.7
0.657
Co (lg/g)
1.4 <1.0–2.5 1.1
0.7 <1.0–1.8 0.5
0.209
Cr (lg/g)
1.2 1.1–1.8 0.3
1.2 1.0–1.2 0.1
0.891
Cu (lg/g)
50.4 27.5–83.0 22.9
24.9 16.8–31.6 5.9
0.023
Fe (lg/g)
233.8 41.6–124.2 92.8
190.9 131.0–341.7 81.5
0.399
Mn (lg/g)
27.4 13.6–62.2 17.0
11.1 6.6–18.4 4.0
0.044
NIST 1566b
Reference material values (lg/g)
This study (lg/g)
Reference material values (lg/g)
This study (lg/g)
Ni (lg/g)
7.8 1.9–16.9 4.7
4.5 2.3–7.7 2.2
0.140
400 (IV) 0.18 ± 0.03 (CV) 0.48 ± 0.13 (RV) 3.91 ± 0.47 (RV) 9.42 ± 0.52 (CV) 274.0 ± 18.00 (RV) 23.93 ± 0.29 (CV) 6.06 ± 0.24 (CV) 2.27 ± 0.13 (CV) 69.30 ± 4.20 (CV) 1.1 (IV) 135.0 ± 5.00 (RV)
393.65 ± 0.85
197.20 ± 6.00 (VC) 2.48 ± 0.08 (CV) 0.37 ± 0.01 (CV) – 71.60 ± 1.60 (CV) 205.80 ± 6.80 (CV)
197.41 ± 0.45 2.49 ± 0.005
Pb (lg/g)
2.6 2.5–3.0 0.2
3.1 2.3–3.5 0.6
0.066
Sr (lg/g)
71.3 48.0–117.0 23.8
74.3 51.5–86.1 12.9
0.790
24.23 ± 0.01 6.05 ± 0.0001 2.23 ± 0.0002 68.08 ± 0.27 1.12 ± 0.0004 133.44 ± 0.03
18.50 ± 0.20 (CV) 1.04 ± 0.09 (CV) 0.31 ± 0.01 (CV) 6.80 ± 0.20 (RV) 0.58 ± 0.52 (CV) 1424.00 ± 46.00(CV)
17.92 ± 0.02 1.05 ± 0.0001
V (lg/g)
3.9 3.2–4.9 0.9
4.7 1.9–5.5 1.4
0.237
Zn (lg/g)
563.3 330.5–876.5 181.7
621.6 571.4–719.3 53.8
0.466
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The samples were analyzed by Inductively Coupled Plasma Optical Emission Spectroscopy (ICP-OES) using a Perkin Elmer, Optima 3000 spectrometer. It was operated with a radio frequency generator at 40 MHz and 1400 W. Determination of Al, Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, Sr, V and Zn was carried out in the automatic mode, with background correction employing the WinLab 32 software package. The following wavelengths were used: 394.401 nm for Al, 226.502 nm for Cd, 228.616 nm for Co, 205.552 nm for Cr, 324.754 nm for Cu, 259.939 nm for Fe, 257.610 nm for Mn, 231.604 nm for Ni, 220.353 nm for Pb, 421.552 nm for Sr, 292.402 nm for V and 202.548 nm for Zn. A comparison of the trace metals determination using the above described methodology in tissue reference materials NIST-1566B (oyster) and NIST-2977 (mussel) with the certified, recommended and information values is presented in Table 1. It can be seen that the determined values were very similar to the certified and recommended values, thus the ICP-OES method (LaBrecque et al., 2004b) employed can be considered accurate. Standard deviations (1r) of the element
determinations (n = 6) are small, indicating good precision. The detection limits were determined as three times the standard deviation of the background under the emission lines of the selected trace elements for the blank solutions. The detection limits of Al, Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, Sr, V and Zn were 0.15, 0.15, 1.0, 0.275, 0.245, 0.25, 0.02, 0.3, 0.5, 0.025, 0.125 and 0.125 lg/g, respectively. A one way ANOVA was performed in order to evaluate the statistical significance between the sites, for each metal considered. Tukey a posteriori tests were performed whenever the null hypotheses were rejected. Statistical analyses were carried out using Statistical 6.0 software (StatSoft, USA). In each sampling area (Buche estuary and Mochima estuary), the difference in the mean metal concentrations between the three sampling sites was not significant (p > 0.05), so a combination of the three sampling sites data will be considered. Mean, minimum, maximum and standard deviation (n = 9) values of the heavy metal concentrations in soft oyster tissues from the two sampling areas are presented in Table 2.
Fig. 2. Annual variation of Al, Cd, Zn, Cu, Mn, Ni, Sr and Fe concentrations in C. rhizophorae from estuaries Buche and Mochima, between March 2008 and March 2009.
Vasquez et al. (1993) 200–1315 nr nr
Presley et al. (1990) 72–10000 nr
0.28– 31.0 nr 161– 662
170–2650
2.96– 45.0 21–63 74.1–1300 6.8–559
nr
0.06– 3.71 nr nr
nr C. virginica (Campeche, Mexico)
nr = Not reported.
nr C. virginica (Gulf of Mexico, USA)
0.54– 16.8 1.2–7.8
nr nr nr C. virginica (USA coast)
1.2–9.1
1.0–2.2 nr
nr
nr
Lauenstein et al. (1990) 520–6000 nr nr
0.18– 1.8 0.02– 12.5 2.9– 24.2 1.3–4.5 nr
80–189
7.6– 17.8 nr
0.3–4.1
nr
nr
nr
1440– 5440 416–994 nr nr 0.1–0.5 1.0–2.0 nr nr
160– 180 6.5– 64.5 30–530 1.01– 2.6 nr 0.8–2.1 nr
C. (as Saccostrea) commercialis (Hawkesbury River, Australia) C. iridescens (Nayarit, Mexico)
nr
nr nr nr 0.8 nr 124 100 nr nr 3.2 nr
C. virginica (selected estuaries, USA) C. rhizophorae (Potengi, Brasil)
C. commercialis (Thailand)
nr 20–38 nr 249–875 nr 33–234 nr nr
<1.5– 8.0 nr 0.6–5.1 <1.5– 2.9 1.1–9.9 0.8–1.9 C. virginica (Savannah estuary, USA)
C. rhizophorae (Mochima estuary, Venezuela)
Phillips and Muttarasin (1985) Hardiman and Pearson (1995) Paez-Osuna et al. (1995)
Apeti et al. (2009) Silva et al. (2001)
nr 1550– 3949 571 nr nr
<1.5– 4.0 nr 1.8–7.8 <1.5– 2.5 nr 0.9–3.2
2.3–3.5 1.0–1.2 1.7–3.5
nr nr
Senthil et al. (2008)
This study
This study
Reference Zn
330.5– 876.5 571.4– 719.3 980–2400 3.2– 4.9 1.9– 5.5 nr
V Sr
48.0– 117.0 51.5– 86.1 nr 2.5–3.0
Pb Ni
1.9– 16.9 2.3–7.7
13.6– 62.2 6.6– 18.4 17–54
Mn Fe
41.6– 124.2 131.0– 341.7 230–1400 27.5– 83.0 16.8– 31.6 67–120
Cu Cr
1.1–1.8
<0.5– 2.5 <0.5– 1.8 nr
Co Cd
2.8– 197.8 5.3– 131.6 400– 1500 nr nr C. rhizophorae (Buche estuary, Venezuela)
1.5–4.2
Al Oyster (location)
Table 3 Comparison .of soft tissue heavy metal concentrations (range of means, lg/g dry weight) in C. rhizophorae from Buche and Mochima estuaries, Venezuela, with concentrations in oysters of the genus Crassostrea (means or ranges of means) from elsewhere.
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397
In the last column of Table 2, the p-values from the ANOVA-test are listed. Significance levels were fixed at p < 0.05 on a 95% confidence level, p-values of elements with significant concentration differences between the sampling sites are presented in boldface. The mean concentrations are higher in Buche estuary for all determined elements with significant differences between sampling areas. This was not unexpected, considering the fact that the Buche estuary is located west of the mouth of the Tuy river, which has a plume known to move in a northwesterly direction and transports a variety of pollutants derived from anthropogenic activities (Jaffé et al., 1995). According with our results, Alfonso et al. (2005) found that the mean concentrations of heavy metals in clams T. mactroides along the Venezuelan coast are generally higher in sites located west of the mouth of the Tuy river, in the state of Miranda. No known values have been reported on trace metals in oysters from the central Venezuelan coast. The relatively large variations occurring within one population, expressed by high standard deviations and large variations between minimum and maximum values in Table 2 are not unexpected, because of the complex relationships between environmental concentrations and bioaccumulation. Cr, Co, Pb and V concentrations in oysters displayed very little variation during the year. Fig. 2 shows the temporal variation of Al, Cd, Cu, Fe, Mn, Ni, Sr and Zn concentrations in oysters from Buche and Mochima. The results show that for each metal there is a similar temporal variation pattern and that the higher concentrations of Cu and Mn in oyster samples from Buche were obtained in March– May and September–November, respectively. The maximum concentrations observed in this study were as follows: for Al (197.83 lg/g), Ni (16.9 lg/g), Fe (413.6 lg/g) and Sr (117.0 lg/g) in September, for Cd (4.2 lg/g) in July, for Co (2.5 lg/g) during March–May, for Cu (83.0 lg/g) and Zn (876.54 lg/g) in May, for Pb (3.5 lg/g) and V(5.5 lg/g) during September–November, and for Mn (62.2 lg/g) in November. Exogenous factors, such as the changes in salinity in those coastal areas during the year depending on rainfall and river freshwater input contributes to seasonal variations (Wagner and Boman, 2004). It was also reported that, one of the most important endogenous factors is the effect of the reproductive cycle on metal accumulation (Coimbra et al., 1991). Unfortunately, the previous reported studies with C. rhizophorae have not included information on the variability in metal concentration during the year, impeding comparative studies, and there is no precise information of reproductive cycles. However, our results suggest that the endogenous factors have a higher impact on annual variation than the exogenous factors in the sampling sites. Further investigation is necessary to understand the dynamics of bioaccumulation and to interpret the annual variations observed. The levels of heavy metals in soft C. rhizophorae tissue found in isla Buche are similar or even lower than those previously reported for T. mactroides at the same sampling region (LaBrecque et al., 2004a; Alfonso et al., 2005), with the exception of the elements Cd, Cu and Zn. It is well know that the metal concentrations vary very markedly among different species of marine bivalves. One example is the difference between mussels and oysters, both of which have been employed as biomonitors of coastal metal contamination over the past few decades. Oysters accumulate high concentrations of Zn in detoxified granules, while mussels excrete much accumulated Zn in granules from the kidney. Thus oysters are strong accumulators of Zn, whereas mussels are weak net accumulators or partial regulators of Zn (Amiard et al., 2008). The class B or borderline metals such as Ag, Cd, Zn, Cu and Hg are generally bioaccumulated to much higher concentrations in oysters as compared to mussels (Wang and Rainbow, 2008). Interspecies differences in metal concentrations are
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well-documented and in a number of cases, the biochemical bases responsible for these differences have been demonstrated. The Word Health Organization (WHO) has established a provisional tolerable weekly intake (PTWI) for Cd at 7 lg/kg of human body weight. This PTWI weekly value corresponds to daily tolerable intake level of 70 lg of Cd for the average 70-kg man, and 60 lg of Cd per day for the average 60-kg woman; the tolerable daily intake of Pb was 25 lg/kg body wt. (Senthil et al., 2008). Clearly, the daily intake of both, Cd (1.5–4.2 lg/g dw) and Pb (2.5–3.5 lg/g dw) from oysters analyzed in this study is below the guidelines established by WHO. The weekly consumption of 612–1633 g of oyster may sufficient to reach the Cd PTWI. However apparent exposure may not reflect a consistent weekly intake of this magnitude, as individuals who consume oysters consume them sporadically. The United States Food and Drug Administration (USFDA, 1993) set an estimated safe and allowable daily consumption level for Cr at 200 lg-person/day, and for Ni at 1200 lg/person/day. Concentrations of Cr in oyster tissues were 1–1.8 lg/g on dry weight basis. Consumption of oyster from any of studied areas does not pose any significant health problems to humans. Concentrations of Ni in oysters were 1.9–16.9 lg dw. Even a 500 g/day intake of oyster from the Buche estuary in September with Ni 16.9 lg may not produce any significant detrimental effects. The results attained in this study should be of considerable value in measuring the impact of future industrial development in the areas. Table 3 compares the data collected in this study with similar data for species of the genus Crassostrea from elsewhere in the world. Concentrations in C. rhizophorae from the Buche estuary can be interpreted to be high on a global scale for Cd, Cu, Mn and Ni, indicating atypically raised bioavailabilities. Thus this area is metal-contaminated. Concentrations of Cr, Fe, Pb and Zn are not apparently above a range that might be considered normal or background. Such comparisons are inevitably somewhat general due to the importance of considering potential influence of size, age, sex reproductive stage, and physiological condition on the concentrations of heavy metals in oysters (Presley et al., 1990). Acknowledgements This project was supported by the FONACIT-Venezuela. We thank Carlos Bastidas, Morelvis Acevedo, José Cáceres and Clara Gomez for their assistance. References Alfonso, J.A., Azocar, J.A., LaBrecque, J.J., Benzo, Z., Marcano, E., Gómez, C.V., Quintal, M., 2005. Temporal and spatial variation of trace metals in clams Tivela mactroidea along the Venezuelan coast. Mar. Pollut. Bull. 50, 1713–1744. Amiard, J.C., Amiard-Triquet, C., Charbonnier, L., Mesnil, A., Rainbow, P.S., Wang, W.X., 2008. Bioaccessibility of essential and non-essential metals in commercial shellfish from Western Europe and Asia. Food Chem. Toxicol. 46, 2010–2022. Apeti, D.A., Lauenstein, G.G., Riedel, G.F., 2009. Cadmium distribution in coastal sediments and mollusks of the US. Mar. Pollut. Bull. 58, 1016–1024. Baudrimont, M., Schafer, J., Marie, V., Maury-Brachet, R., Bossy, C., Boudou, A., Blanc, G., 2005. Geochemical survey and metal bioaccumulation of three bivalve species (Crassostrea gigas, Cerastoderma edule and Ruditapes philippinarum) in the Nord Médoc salt marshes (Gironde estuary, France). Sci. Total Environ. 337, 265–280. Bendell, L.I., Feng, C., 2009. Spatial and temporal variations in cadmium concentrations and burdens in the Pacific oyster (Crassostrea gigas) sampled from the Pacific north-west. Mar. Pollut. Bull. 58, 1137–1143.
Cantillo, A.Y., Lauenstein, G.G., O’Connor, T.P., 1998. Status and trends of contaminant levels in biota and sediment of the Chesapeake Bay. Regional Report Series 1. NOS CCMA, Silver Spring, MD. Coimbra, J., Carraca, S., Ferreira, A., 1991. Metals in Mytilus edulis from the Northern Coast of Portugal. Mar. Pollut. Bull. 22, 249–253. Fialkowski, W., Calosi, P., Dahlke, S., Dietrich, A., Moore, P.G., Olenin, S., Persson, L.E., Smith, B.D., Spegys, M., Rainbow, P.S., 2009. The sandhopper Talitrus saltator (Crustacea: Amphipoda) as a biomonitor of trace metal bioavailabilities in European coastal waters. Mar. Pollut. Bull. 58, 39–44. Fraser, M., Surette, C., Vaillancourt, C., 2011. Spatial and temporal distribution of heavy metal concentrations in mussels (Mytilus edulis) from the Baie des Chaleurs, New Brunswick, Canada. Mar. Pollut. Bull. 62, 1345–1351. Hardiman, S., Pearson, B., 1995. 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The influence of body size, condition index and tidal exposure on the variability in metal bioaccumulation in Mytilus edulis. Environ. Pollut. 144, 272–279. ´ Connor, T.P., Cantillo, A.Y., Lauenstein, G.G., 1994. Monitoring of temporal trends O in chemical contamination by the NOAA National Status and trends Mussel Watch Project. In: Kramer, K.J.M. (Ed.), Biomonitoring of Coastal Waters and Estuarine. CRC Press, Boca Raton, FL, pp. 29–45. Paez-Osuna, F., Frias-Espericueta, M.G., Osuna-Lopez, J.I., 1995. Trace metal concentrations in relation to season and gonadal maturation in the oyster Crassostrea iridescens. Mar. Environ. Res. 40, 19–31. Phillips, D.J.H., Muttarasin, K., 1985. Trace metals in bivalve mollusks in Thailand. Mar. Environ. Res. 16, 281–300. Presley, B.J., Taylor, R.J., Boothe, P.N., 1990. Trace metals in Gulf of Mexico oysters. Sci. Total Environ. 97 (98), 551–593. Spann, N., Aldridge, D.C., Griffin, J.L., Jones, O.A.H., 2011. Size-dependent effects of low level cadmium and zinc exposure on the metabolome of the Asian clam, Corbicula fluminea. Aquat. Toxicol. 105, 589–599. Senthil Kumar, K., Sajwan, K.S., Richardson, J.P., Kannan, K., 2008. Contamination profiles of heavy metals, organochlorine pesticides, polycyclic aromatic hydrocarbons and alkylphenols in sediment and oyster collected from marsh/ estuarine Savannah GA, USA. Mar. Pollut. Bull. 56, 136–162. Silva, C.A.R., Rainbow, P.S., Smith, B.D., Santos, Z.L., 2001. Biomonitoring of trace metal contamination in the Potengi estuary, Natal (Brasil), using the oyster Crassostrea Rhizophorae, a local food source. Water Res. 17, 4072–4078. USFDA, 1993. Guidance Document for Arsenic, Chromium, Nickel in Shellfish. Center for Food Safety and Applied Nutrition. United States food and Drug Administration, Washington, DC. Vasquez, G.F., Sanchez, G.M., Virender, K.S., 1993. Trace metals in the oyster Crassotrea virginica of the Terminos Laggon, Campeche, Mexico. 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Marine Pollution Bulletin journal homepage: www.elsevier.com/locate/marpolbul
Baseline
Temporal distribution of heavy metal concentrations in oysters Crassostrea rhizophorae from the central Venezuelan coast Juan A. Alfonso a,⇑, Helga Handt a, Abrahan Mora a, Yaneth Vásquez a, José Azocar a, Eunice Marcano b a b
Centro de Oceanología y Estudios Antárticos, Instituto Venezolano de Investigaciones Científicas (IVIC), Apartado 20632, Caracas 1020A, Venezuela Centro de Química, Instituto Venezolano de Investigaciones Científicas (IVIC), Apartado 20632, Caracas 1020A, Venezuela
a r t i c l e
i n f o
Keywords: Oysters Bivalves Bioindicators Heavy metals
a b s t r a c t The oyster Crassostrea rhizophorae is a bivalve abundant in Venezuelan estuaries and consumed by local populations. No known values have been reported on trace metals in oysters from the central Venezuelan coast. We report the concentrations of Al, Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, Sr, V and Zn in the soft parts of C. rhizophorae, which were collected bimonthly between March 2008 and March 2009, at two sampling areas from the Central Venezuelan Coast: Buche estuary and Mochima estuary. Our results show that for each metal there is a similar temporal variation pattern. The concentrations of the heavy metals reported in this work are useful as reliable baselines and can be used for comparison in future environment studies. Concentrations in C. rhizophorae from the Buche estuary can be interpreted to be high on a global scale for Cd, Cu, Ni and Mn, indicating atypically raised bioavailabilities. Ó 2013 Elsevier Ltd. All rights reserved.
Bivalves have been extensively used as bioindicators to assess coastal aquatic environments, since they can accumulate trace metals and other substances. Although no official monitoring program is in progress, various studies by different groups have provided relevant data with respect to changing metals levels in clams Tivela mactroides from the central Venezuelan coast (Jaffé et al., 1995; LaBrecque et al., 2004a; Alfonso et al., 2005). However, the need to consider other species and taxa has been emphasized for regional, national and international monitoring programs (Ke and Wang, 2001; Baudrimont et al., 2005; Senthil Kumar et al., 2008; Bendell and Feng, 2009; Apeti et al., 2009). Oysters have been used as a companion bivalve species with mussels in the biomonitoring of coastal contamination in National Status and Trend Programs (e.g. the National Status and Trend program in the US, ´ Connor et al., 1994; the Mussel Watch program in France, Cantillo O et al., 1998). The oyster Crassostrea rhizophorae is a bivalve abundant in Venezuelan estuaries and widely consumed by local populations. Only a few studies have been reported on trace metals in oysters from Venezuela. However, such studies have not dealt with temporal variations; they have only considered a limited number of elements, and have been undertaken in restricted areas (See Fig. 1). The marine environment in the central Venezuelan coast is subjected to contamination by heavy metals introduced to this coastal environment by the Tuy river (Jaffé et al., 1995; LaBrecque et al., 2004a; Alfonso et al., 2005), which has a plume known to move ⇑ Corresponding author. Tel.: +58 212 504 1401; fax: +58 212 504 1935. E-mail address: [email protected] (J.A. Alfonso). 0025-326X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.marpolbul.2013.05.010
in a northwesterly direction and receives waste water effluents from the metropolitan area of Caracas, a city of approximately 7 million inhabitants, via the Guaire river. Most of this waste water undergoes only minor treatment. In addition, the Tuy river drainage basin, of approximately 6600 km2, covers an active industrial and agricultural area, which also affects water quality (Jaffé et al., 1995). The use of a bioindicator is a good integrative tool of the ecosystem (Fraser et al., 2011). Indeed, bioindicators, such as oysters, integrate many aspects of their ecosystem including the contamination and external factors which can influence this contamination, such as ocean currents, storms, seasons and water temperature (Markert et al., 1999). The objective of this study is to establish a baseline of the temporal distribution of heavy metals in two areas of the central Venezuelan coast by using the oyster C. rhizophorae as a bioindicator. It is well known that the accumulation of heavy metals in bivalves is affected by age, size and weight of the bivalves (Mubiana et al., 2006; Fialkowski et al., 2009; Spann et al., 2011; McKinley et al., 2012). As these factors were not the objective of our study, they were kept as constant as possible, in order to minimize their influence on the measured concentrations. Each sample consisted of a pool of at least 10 oysters (C. rhizophorae) of 6–7 cm in size. Samples of C. rhizophorae were collected bimonthly between March 2008 and March 2009, using a stainless steel knife at six sampling sites, three located in Buche estuary, Carenero, state of Miranda, and three located in Mochima estuary, state of Sucre. Spacing between sampling sites varied between 400 and 500 m, within each sampling area. Triplicate samples were collected at
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Fig. 1. A map of the Venezuelan coast showing the sampling areas.
each sampling site (10–15 m2). All samples were rinsed with seawater after collection to remove marine sediments, shells and other debris, placed in plastic bags and transported to the laboratory on ice. In the laboratory, the complete soft tissue of oysters was removed from the shell and washed with distilled deionized water to remove impurities. The bivalves tissues were transferred to clean one-liter beakers and washed several times with distilled deionized water until the solution was clear. After discarding the excess liquid, the beakers were placed in a drying oven at about 65 °C until most of the liquid was evaporated, then the temperature was increased to 80 °C for 24 h, after cooled in desiccators. The dried tissue samples were ground into a fine powder with an agate mortar and pestle and well mixed to prepare homogenous samples. One gram portions of the dried tissues were dissolved in 8 ml of SuprapureÒ nitric acid (Merck) by heating on a hot plate at about 70 °C for about 8 h. When the solution cooled, 2 ml of hydrogen peroxide (30%) (Merck) was added and heated again at 70 °C for another 4 h. Finally, the resulting solution was diluted to 50 ml with distilled deionized water. Duplicate digestions were performed for each sample. Method blanks were always low in trace metals indicating minimal contamination during sample
Table 1 Comparison of the determined values (n = 6) in standard reference materials NIST 2977 in this study with the certified (CV), recommended (RV) and information (IV) values. Element NIST 2977
Al Cd Co Cr Cu Fe Mn Ni Pb Sr V Zn
processing, and these concentrations were subtracted from sample readings to give final values of metal concentration. Table 2 Mean, minimum maximum and standard deviation values of the trace element concentrations in dried soft oyster tissue from the different sampling areas during this temporal study (n = 9) in estuaries Buche and Mochima. The p-values in boldface represent significant concentration differences (p<0.05) between the sampling sites. Buche
Mochima
p
Al (lg/g)
63.4 2.8–197.8 26.7
49.05 5.3–131.6 53.4
0.673
Cd (lg/g)
2.5 1.5–4.2 0.9
2.7 1.7–3.5 0.7
0.657
Co (lg/g)
1.4 <1.0–2.5 1.1
0.7 <1.0–1.8 0.5
0.209
Cr (lg/g)
1.2 1.1–1.8 0.3
1.2 1.0–1.2 0.1
0.891
Cu (lg/g)
50.4 27.5–83.0 22.9
24.9 16.8–31.6 5.9
0.023
Fe (lg/g)
233.8 41.6–124.2 92.8
190.9 131.0–341.7 81.5
0.399
Mn (lg/g)
27.4 13.6–62.2 17.0
11.1 6.6–18.4 4.0
0.044
NIST 1566b
Reference material values (lg/g)
This study (lg/g)
Reference material values (lg/g)
This study (lg/g)
Ni (lg/g)
7.8 1.9–16.9 4.7
4.5 2.3–7.7 2.2
0.140
400 (IV) 0.18 ± 0.03 (CV) 0.48 ± 0.13 (RV) 3.91 ± 0.47 (RV) 9.42 ± 0.52 (CV) 274.0 ± 18.00 (RV) 23.93 ± 0.29 (CV) 6.06 ± 0.24 (CV) 2.27 ± 0.13 (CV) 69.30 ± 4.20 (CV) 1.1 (IV) 135.0 ± 5.00 (RV)
393.65 ± 0.85
197.20 ± 6.00 (VC) 2.48 ± 0.08 (CV) 0.37 ± 0.01 (CV) – 71.60 ± 1.60 (CV) 205.80 ± 6.80 (CV)
197.41 ± 0.45 2.49 ± 0.005
Pb (lg/g)
2.6 2.5–3.0 0.2
3.1 2.3–3.5 0.6
0.066
Sr (lg/g)
71.3 48.0–117.0 23.8
74.3 51.5–86.1 12.9
0.790
24.23 ± 0.01 6.05 ± 0.0001 2.23 ± 0.0002 68.08 ± 0.27 1.12 ± 0.0004 133.44 ± 0.03
18.50 ± 0.20 (CV) 1.04 ± 0.09 (CV) 0.31 ± 0.01 (CV) 6.80 ± 0.20 (RV) 0.58 ± 0.52 (CV) 1424.00 ± 46.00(CV)
17.92 ± 0.02 1.05 ± 0.0001
V (lg/g)
3.9 3.2–4.9 0.9
4.7 1.9–5.5 1.4
0.237
Zn (lg/g)
563.3 330.5–876.5 181.7
621.6 571.4–719.3 53.8
0.466
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The samples were analyzed by Inductively Coupled Plasma Optical Emission Spectroscopy (ICP-OES) using a Perkin Elmer, Optima 3000 spectrometer. It was operated with a radio frequency generator at 40 MHz and 1400 W. Determination of Al, Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, Sr, V and Zn was carried out in the automatic mode, with background correction employing the WinLab 32 software package. The following wavelengths were used: 394.401 nm for Al, 226.502 nm for Cd, 228.616 nm for Co, 205.552 nm for Cr, 324.754 nm for Cu, 259.939 nm for Fe, 257.610 nm for Mn, 231.604 nm for Ni, 220.353 nm for Pb, 421.552 nm for Sr, 292.402 nm for V and 202.548 nm for Zn. A comparison of the trace metals determination using the above described methodology in tissue reference materials NIST-1566B (oyster) and NIST-2977 (mussel) with the certified, recommended and information values is presented in Table 1. It can be seen that the determined values were very similar to the certified and recommended values, thus the ICP-OES method (LaBrecque et al., 2004b) employed can be considered accurate. Standard deviations (1r) of the element
determinations (n = 6) are small, indicating good precision. The detection limits were determined as three times the standard deviation of the background under the emission lines of the selected trace elements for the blank solutions. The detection limits of Al, Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, Sr, V and Zn were 0.15, 0.15, 1.0, 0.275, 0.245, 0.25, 0.02, 0.3, 0.5, 0.025, 0.125 and 0.125 lg/g, respectively. A one way ANOVA was performed in order to evaluate the statistical significance between the sites, for each metal considered. Tukey a posteriori tests were performed whenever the null hypotheses were rejected. Statistical analyses were carried out using Statistical 6.0 software (StatSoft, USA). In each sampling area (Buche estuary and Mochima estuary), the difference in the mean metal concentrations between the three sampling sites was not significant (p > 0.05), so a combination of the three sampling sites data will be considered. Mean, minimum, maximum and standard deviation (n = 9) values of the heavy metal concentrations in soft oyster tissues from the two sampling areas are presented in Table 2.
Fig. 2. Annual variation of Al, Cd, Zn, Cu, Mn, Ni, Sr and Fe concentrations in C. rhizophorae from estuaries Buche and Mochima, between March 2008 and March 2009.
Vasquez et al. (1993) 200–1315 nr nr
Presley et al. (1990) 72–10000 nr
0.28– 31.0 nr 161– 662
170–2650
2.96– 45.0 21–63 74.1–1300 6.8–559
nr
0.06– 3.71 nr nr
nr C. virginica (Campeche, Mexico)
nr = Not reported.
nr C. virginica (Gulf of Mexico, USA)
0.54– 16.8 1.2–7.8
nr nr nr C. virginica (USA coast)
1.2–9.1
1.0–2.2 nr
nr
nr
Lauenstein et al. (1990) 520–6000 nr nr
0.18– 1.8 0.02– 12.5 2.9– 24.2 1.3–4.5 nr
80–189
7.6– 17.8 nr
0.3–4.1
nr
nr
nr
1440– 5440 416–994 nr nr 0.1–0.5 1.0–2.0 nr nr
160– 180 6.5– 64.5 30–530 1.01– 2.6 nr 0.8–2.1 nr
C. (as Saccostrea) commercialis (Hawkesbury River, Australia) C. iridescens (Nayarit, Mexico)
nr
nr nr nr 0.8 nr 124 100 nr nr 3.2 nr
C. virginica (selected estuaries, USA) C. rhizophorae (Potengi, Brasil)
C. commercialis (Thailand)
nr 20–38 nr 249–875 nr 33–234 nr nr
<1.5– 8.0 nr 0.6–5.1 <1.5– 2.9 1.1–9.9 0.8–1.9 C. virginica (Savannah estuary, USA)
C. rhizophorae (Mochima estuary, Venezuela)
Phillips and Muttarasin (1985) Hardiman and Pearson (1995) Paez-Osuna et al. (1995)
Apeti et al. (2009) Silva et al. (2001)
nr 1550– 3949 571 nr nr
<1.5– 4.0 nr 1.8–7.8 <1.5– 2.5 nr 0.9–3.2
2.3–3.5 1.0–1.2 1.7–3.5
nr nr
Senthil et al. (2008)
This study
This study
Reference Zn
330.5– 876.5 571.4– 719.3 980–2400 3.2– 4.9 1.9– 5.5 nr
V Sr
48.0– 117.0 51.5– 86.1 nr 2.5–3.0
Pb Ni
1.9– 16.9 2.3–7.7
13.6– 62.2 6.6– 18.4 17–54
Mn Fe
41.6– 124.2 131.0– 341.7 230–1400 27.5– 83.0 16.8– 31.6 67–120
Cu Cr
1.1–1.8
<0.5– 2.5 <0.5– 1.8 nr
Co Cd
2.8– 197.8 5.3– 131.6 400– 1500 nr nr C. rhizophorae (Buche estuary, Venezuela)
1.5–4.2
Al Oyster (location)
Table 3 Comparison .of soft tissue heavy metal concentrations (range of means, lg/g dry weight) in C. rhizophorae from Buche and Mochima estuaries, Venezuela, with concentrations in oysters of the genus Crassostrea (means or ranges of means) from elsewhere.
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In the last column of Table 2, the p-values from the ANOVA-test are listed. Significance levels were fixed at p < 0.05 on a 95% confidence level, p-values of elements with significant concentration differences between the sampling sites are presented in boldface. The mean concentrations are higher in Buche estuary for all determined elements with significant differences between sampling areas. This was not unexpected, considering the fact that the Buche estuary is located west of the mouth of the Tuy river, which has a plume known to move in a northwesterly direction and transports a variety of pollutants derived from anthropogenic activities (Jaffé et al., 1995). According with our results, Alfonso et al. (2005) found that the mean concentrations of heavy metals in clams T. mactroides along the Venezuelan coast are generally higher in sites located west of the mouth of the Tuy river, in the state of Miranda. No known values have been reported on trace metals in oysters from the central Venezuelan coast. The relatively large variations occurring within one population, expressed by high standard deviations and large variations between minimum and maximum values in Table 2 are not unexpected, because of the complex relationships between environmental concentrations and bioaccumulation. Cr, Co, Pb and V concentrations in oysters displayed very little variation during the year. Fig. 2 shows the temporal variation of Al, Cd, Cu, Fe, Mn, Ni, Sr and Zn concentrations in oysters from Buche and Mochima. The results show that for each metal there is a similar temporal variation pattern and that the higher concentrations of Cu and Mn in oyster samples from Buche were obtained in March– May and September–November, respectively. The maximum concentrations observed in this study were as follows: for Al (197.83 lg/g), Ni (16.9 lg/g), Fe (413.6 lg/g) and Sr (117.0 lg/g) in September, for Cd (4.2 lg/g) in July, for Co (2.5 lg/g) during March–May, for Cu (83.0 lg/g) and Zn (876.54 lg/g) in May, for Pb (3.5 lg/g) and V(5.5 lg/g) during September–November, and for Mn (62.2 lg/g) in November. Exogenous factors, such as the changes in salinity in those coastal areas during the year depending on rainfall and river freshwater input contributes to seasonal variations (Wagner and Boman, 2004). It was also reported that, one of the most important endogenous factors is the effect of the reproductive cycle on metal accumulation (Coimbra et al., 1991). Unfortunately, the previous reported studies with C. rhizophorae have not included information on the variability in metal concentration during the year, impeding comparative studies, and there is no precise information of reproductive cycles. However, our results suggest that the endogenous factors have a higher impact on annual variation than the exogenous factors in the sampling sites. Further investigation is necessary to understand the dynamics of bioaccumulation and to interpret the annual variations observed. The levels of heavy metals in soft C. rhizophorae tissue found in isla Buche are similar or even lower than those previously reported for T. mactroides at the same sampling region (LaBrecque et al., 2004a; Alfonso et al., 2005), with the exception of the elements Cd, Cu and Zn. It is well know that the metal concentrations vary very markedly among different species of marine bivalves. One example is the difference between mussels and oysters, both of which have been employed as biomonitors of coastal metal contamination over the past few decades. Oysters accumulate high concentrations of Zn in detoxified granules, while mussels excrete much accumulated Zn in granules from the kidney. Thus oysters are strong accumulators of Zn, whereas mussels are weak net accumulators or partial regulators of Zn (Amiard et al., 2008). The class B or borderline metals such as Ag, Cd, Zn, Cu and Hg are generally bioaccumulated to much higher concentrations in oysters as compared to mussels (Wang and Rainbow, 2008). Interspecies differences in metal concentrations are
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well-documented and in a number of cases, the biochemical bases responsible for these differences have been demonstrated. The Word Health Organization (WHO) has established a provisional tolerable weekly intake (PTWI) for Cd at 7 lg/kg of human body weight. This PTWI weekly value corresponds to daily tolerable intake level of 70 lg of Cd for the average 70-kg man, and 60 lg of Cd per day for the average 60-kg woman; the tolerable daily intake of Pb was 25 lg/kg body wt. (Senthil et al., 2008). Clearly, the daily intake of both, Cd (1.5–4.2 lg/g dw) and Pb (2.5–3.5 lg/g dw) from oysters analyzed in this study is below the guidelines established by WHO. The weekly consumption of 612–1633 g of oyster may sufficient to reach the Cd PTWI. However apparent exposure may not reflect a consistent weekly intake of this magnitude, as individuals who consume oysters consume them sporadically. The United States Food and Drug Administration (USFDA, 1993) set an estimated safe and allowable daily consumption level for Cr at 200 lg-person/day, and for Ni at 1200 lg/person/day. Concentrations of Cr in oyster tissues were 1–1.8 lg/g on dry weight basis. Consumption of oyster from any of studied areas does not pose any significant health problems to humans. Concentrations of Ni in oysters were 1.9–16.9 lg dw. Even a 500 g/day intake of oyster from the Buche estuary in September with Ni 16.9 lg may not produce any significant detrimental effects. The results attained in this study should be of considerable value in measuring the impact of future industrial development in the areas. Table 3 compares the data collected in this study with similar data for species of the genus Crassostrea from elsewhere in the world. Concentrations in C. rhizophorae from the Buche estuary can be interpreted to be high on a global scale for Cd, Cu, Mn and Ni, indicating atypically raised bioavailabilities. Thus this area is metal-contaminated. Concentrations of Cr, Fe, Pb and Zn are not apparently above a range that might be considered normal or background. Such comparisons are inevitably somewhat general due to the importance of considering potential influence of size, age, sex reproductive stage, and physiological condition on the concentrations of heavy metals in oysters (Presley et al., 1990). Acknowledgements This project was supported by the FONACIT-Venezuela. We thank Carlos Bastidas, Morelvis Acevedo, José Cáceres and Clara Gomez for their assistance. References Alfonso, J.A., Azocar, J.A., LaBrecque, J.J., Benzo, Z., Marcano, E., Gómez, C.V., Quintal, M., 2005. Temporal and spatial variation of trace metals in clams Tivela mactroidea along the Venezuelan coast. Mar. Pollut. Bull. 50, 1713–1744. Amiard, J.C., Amiard-Triquet, C., Charbonnier, L., Mesnil, A., Rainbow, P.S., Wang, W.X., 2008. Bioaccessibility of essential and non-essential metals in commercial shellfish from Western Europe and Asia. Food Chem. Toxicol. 46, 2010–2022. Apeti, D.A., Lauenstein, G.G., Riedel, G.F., 2009. Cadmium distribution in coastal sediments and mollusks of the US. Mar. Pollut. Bull. 58, 1016–1024. Baudrimont, M., Schafer, J., Marie, V., Maury-Brachet, R., Bossy, C., Boudou, A., Blanc, G., 2005. Geochemical survey and metal bioaccumulation of three bivalve species (Crassostrea gigas, Cerastoderma edule and Ruditapes philippinarum) in the Nord Médoc salt marshes (Gironde estuary, France). Sci. Total Environ. 337, 265–280. Bendell, L.I., Feng, C., 2009. Spatial and temporal variations in cadmium concentrations and burdens in the Pacific oyster (Crassostrea gigas) sampled from the Pacific north-west. Mar. Pollut. Bull. 58, 1137–1143.
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