Adjustment techniques of heavy metals bioaccumulation in the mussel mytilus galloprovincialis for evaluation of contamination in the moroccan atlantic coast

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bioaccumulation in the mussel mytilus galloprovincialis for...

Article in Fresenius Environmental Bulletin · January 2009

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Ikram Brahim

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ADJUSTMENT TECHNIQUES OF

HEAVY METALS BIOACCUMULATION IN THE

MUSSEL Mytilus galloprovincialis FOR EVALUATION OF CONTAMINATION IN THE MOROCCAN ATLANTIC COAST

Ikram Brahim^1*, Younes Karhat² and Samir Benbrahim³

1 Department of Biology, Faculty of Science Ain Chock, University Hassan II, Km 8 Route d'El Jadida, Maarif, Casablanca, Morocco.

2Department of Chemistry, Laboratory of Catalysis and Environment,

Faculty of Science Ain Chock, University Hassan II, Km 8 Route d'El Jadida, Maarif, Casablanca, Morocco.

3 Laboratory of Marine Chemistry, Institut National de Recherche Halieutique (INRH), 2 rue de Tiznit, Casablanca, Morocco.

ABSTRACT

In this work, contamination levels in two Moroccan coastal zones (Ouled Ghanem near El Jadida and Dar Bouazza near Casablanca) through spatial and temporal monitoring of the levels of some metallic elements (Cd, Hg and Zn) in the mussel Mytilus galloprovincialis by calculating the Condition Index (CI: the ratio between dry flesh weight and dry shell weight) were examined. Heavy metal bioaccumulation measurements in the mussel Mytilus gal- loprovincialis collected in the studied zones show a sea- sonal pattern with a maximum in spring and a minimum in summer because of physiological variations including metabolism and reproduction (gametogenesis, egg laying) with- out neglecting the effect of geographical distribution. Com- putation of the Condition Index enables us to adjust the levels of heavy metals measured. Adjusted concentrations of heavy metals in the mussel Mytilus galloprovincialis, after taking into account their CI, are supposed to be represen- tative of the state of marine environmental contamination.

They also make it possible to compare the contamination levels in two studied zones. In our case, results show that Ouled Ghanem area is more contaminated by Cd than Dar Bouazza.

KEYWORDS: Bioaccumulation, adjusted concentration, heavy metals, Moroccan coast, Mytilus galloprovincials, Condition Index.

INTRODUCTION

The marine environment, particularly rich biotope, is characterized at the same time by a remarkable stability of its fundamental properties and a great variability of its mi-

cro-constituents. Though the contribution of metal contaminants via the industrial effluents and the atmosphere, the rivers and their estuaries, may modify the seawater com- position, and make it toxic for plants and animals [1].

Measurements of seawater contaminants are realized by sophisticated and expensive analysis techniques which are not easily applied to enough samples collected all over the Moroccan coast. In addition, the temporal variability over the coast confers only little significance on a specific measurement taken in the water column. This is why the mussel, pollution indicator species, is largely used in various monitoring and research programs aiming the estab- lishment of the temporal and spatial variability of contaminants over the coastal environment [2].

The purpose of this study is to assess the contamination level by heavy metals at Moroccan Atlantic coast. The Condition Index (CI) was calculated as the ratio between dry flesh and dry shell weight which allows to adjust the contamination levels of heavy metals measured in the mussel Mytilus galloprovincialis. This Condition Index repre- sents a biometric variable mostly correlated with the heavy metals concentrations [3].

MATERIAL AND METHODS

Sites of taking away

Two sites on the Moroccan Atlantic Ocean were se- lected for realization of this study:

Dar Bouazza is located 30 km south of Casablanca on the Atlantic coast of Morocco.

Ouled Ghanem is located 76 km south of El Jadida and 66 km north of Safi.

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FIGURE 1 – Locations of the two studied sites (Dar Bouazza and Ouled Ghanem) in the Moroccan Atlantic Ocean.

Sampling and pretreatment

• The sampling of the marine organisms is done from January to September 2007. In all sites, we carried out:

• Measurement of length, width and thickness of the mussel shells with a slide caliper giving a measure to 1/10 mm;

• Weight of each individual to 1/1000 g of “the total fresh weight”;

• Weight of flesh and shell “fresh weight”;

• Drying at low temperature (42 °C) in a stainless drying oven;

• Weight of flesh and shell “dry weight”;

• Crushing and homogenization of the dried flesh.

Analyses

Acid mineralization was carried out in a microwave oven (950 W) “Mars 5 CEM” programmed by computer with regulation system of pressure and temperature inside the mineralization bombs.

Cd was analyzed by graphite furnace atomic absorption spectrometry (Schimadzu AA 6800), and Zn by flame atomic absorption spectrometry (Varian). Mercury concentrations were determined by cold vapor atomic absorption spectrometry (Aula 254).

Results are expressed in µg of contaminants per gram of dry mussel flesh.

RESULTS AND DISCUSSION

Temporal variations of metal contents in the mussels Dar Bouazza

Fig. 2 shows the temporal variations of Hg, Cd and Zn concentrations in mussel tissues at Dar Bouazza.

The Hg recorded a high concentration for the period from April to June with a maximum in May (0.14 µg/g of dry weight). The minimal value was recorded in Septem- ber (0.04 µg/g of dry weight).

The monthly evolution of Cd and Zn contents showed a period of strong values in spring with a maximum in June (0.90 µg/g of dry weight for Cd and 248.69 µg/g of dry weight for Zn). The minimal concentrations for these two metals were recorded in August (0.28 µg/g of dry weight for Cd and 108.92 µg/g of dry weight for Zn).

The temporal variations of metal contents in the mussel show a seasonal rhythm for the three analyzed elements [4]. Indeed, the maximum concentrations of Cd, Zn and Hg in mussel tissues at Dar Bouazza were recorded in spring with a minimum in summer [5-7].

Ouled Ghanem

The temporal variations of Hg, Cd and Zn levels in mussel tissue at Ouled Ghanem are visualized in Fig. 3.

The temporal evolution of Hg showed an increase in its content for the period from February (0.09 µg/g of dry weight) to June (0.17 µg/g of dry weight).

Cadmium and zinc recorded a peak in April (1.87 µg/g of dry weight for Cd and 304.56 µg/g of dry weight for Zn),

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Mercury

0,02 0,06 0,10 0,14 0,18

January March May July September

months µg/g

Cadmium

0,2 0,4 0,6 0,8 1,0

Zinc

100 150 200 250 300

m onths µg/g

FIGURE 2 - Monthly variation of mercury, cadmium and zinc concentrations in mussel tissues at Dar Bouazza.

Mercury

0,06 0,10 0,14 0,18 0,22

February April

June Augus

t m onths µg/g

Cadm ium

0,8 1,1 1,4 1,7 2,0

February April

June August m onths

µg/g

Zinc

100 200 300 400

February April

June August months

µg/g

FIGURE 3 - Temporal evolution of mercury, cadmium and zinc concentrations in mussel tissues at Ouled ghanem.

followed by an important decrease in August (1.07 µg/g of dry weight for Cd and 122.10µg/g of dry weight for Zn).

These results present a seasonal rhythm of bioaccumulation of the metals analyzed in the mussel tissues with a maximum in spring and a minimum in summer [5, 7].

Various hypotheses can be advanced to explain this phenomenon:

• The seasonal change of metals` environmental sources which affect the metal accumulation in the mussel;

• The increase of biological activity of the mussel, de- pending on the availability of phytoplankton, during the spring season, can lead to an increase of the complexa- tion capacity of metals to the organic matter and, con- sequently, to a change of their metal bioavailability [8];

• The intense metabolic activity during the reproduction period obliges the animal to filter more quantities of

water and to be continuously open, which leads to more important accumulation;

• Variations of the mussel weights during different phases of life cycle (gametogenesis and egg laying) [6, 12].

The physiological processes related to the metabolism and, especially, to the reproduction (gametogenesis, egg laying) seem to play a key role in the seasonal variations of the metal concentrations in the mussel, jointly with the temporal changes of the weight of their flesh, in agree- ment with many authors [10, 11].

The seasonal increase of the studied metal concentrations in the mussels coincided with the multiplication and gametes maturation phases, during which the cellular metabolism of the mussel was very active and the gonads constituted a “trap” of metals, leading to a strong metal accumulation [9]. The emission of the sexual products (egg laying) occurred in summer, and was followed by a progressive decrease in metal contents in mussels until

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they reached a very low level at the end of this season [6, 7].

Adjustment model of the raw data

The Condition Index is strongly correlated with the contaminant concentrations. For heavy metals, tissue concentration is inversely proportional to the Condition Index:

[ML] = a * 1/CI+b [3, 13].

Dar Bouazza

Figure 4 presents the linear regression of contaminant concentration in the mussel to the inverse of Condition Index in the site of Dar Bouazza.

The zinc and cadmium levels in the mussel were significantly correlated to the inverse of Condition Index (R²= 0.69 for Zn and R²=0.75 for Cd), whereas mercury concentrations showed a very weak correlation to the inverse of Condition Index with R²=0.11.

Mercury

y = 0,0042x + 0,0613 R² = 0,1089

0,0 0,1 0,2 0,3

0 5 10 15 20

1/CI µg/g

Cadm ium

y = 0,0719x - 0,1112 R² = 0,7478

0,0 0,4 0,8 1,2 1,6

4 9 1/CI 14 19

µg/g

Zinc

y = 15,895x + 31,723 R² = 0,6881 0

100 200 300

0 10 20

1/CI µg/g

FIGURE 4 - Linear regression between the contaminant concentrations in the mussel and the inverse of the Condition Index (site of Dar Bouazza).

Ouled Ghanem

Figure 5 shows the linear regression between the heavy metals concentrations in the mussel to the inverse of the Condition Index in the site of Ouled Ghanem.

Similarly to the mussels of Dar Bouazza, the linear regression of Hg concentrations to the inverse of the Condi- tion Index showed that there was no significant linear correlation between these two parameters (R² =0.09), whereas the linear regression of Cd concentrations to the inverse of the Condition Index evidenced a moderate linear correlation (R²=0.41), but the one for zinc was significantly correlated to the inverse of Condition Index (R²=0.75).

Mercury

y = 0,0021x + 0,1115 R² = 0,0904 0,0

0,1 0,2 0,3

5 15 25 35

1/CI µg/g

Cadm ium

y = 0,0497x + 0,776 R² = 0,4116

0 1 2 3 4

5 15 25

1/CI µg/g

Zinc

y = 11,525x + 25,474 R² = 0,7527 50

150 250 350 450

5 15 1/CI 25 35

µg/g

FIGURE 5 - Linear regression of the contaminant concentrations in the mussel to the inverse of Condition Index in the site of Ouled Ghanem.

For mercury, the linear regression relating concentrations in the mussel tissue to the inverse of the Condition In- dex were not significantly because the amounts of this metal in the marine environment were very weak and at the detection limit of the measuring devices.

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We noticed that low values of Condition Index in the mussel corresponded to high concentrations of zinc and cadmium in the flesh of this species because:

• the increase of biomass during the reproduction cycles, in particular at the period of egg laying, resulted in a di- lution of bio-accumulated metals [12],

• the small individuals accumulated more metals because of the very high rate of their metabolism compared to low masses of their tissue [12].

Summary table of linear regression models

TABLE 1 - Linear regression models of the heavy metal concen- trations to the inverse of Condition Index in the two sites of study.

Area of study Contaminant Linear Regression Model R² (correlation coefficient) Cadmium [Cd] =

0.0719 * 1/CI – 0.1112 0.75 Dar Bouazza

Zinc [Zn] =

15.895 * 1/CI + 31.723 0.69 Cadmium [Cd] =

0.0497 * 1/CI + 0.776 0.41 Ouled Gha-

nem Zinc [Zn] =

11.525 * 1/CI + 25.474 0.75 Adjustment method of the raw results

For each analyzed metal, tissue concentration was inversely proportional to the condition index [ML] = a*1/CI+b. The use of these linear models (when being significant) makes it possible to adjust the concentrations measured to a unique reference Condition Index (CI r) by calculating the average or mean from the Condition Indi- ces of the samples applied to each model [14].

These adjusted concentrations are used as an indicator of the marine environment contamination along the year whatever the period is of reproduction or not [15].

Adjustment formula used here is similar to that given by Stanisière et al. in the year 2002 [16]:

[ML] aj = [ML] mes + [a * (1/CI r – 1/CI mes)]

Where

• [ML] = heavy metal concentration: aj = adjusted; mes

= measured

• CI mes = Condition Index measured;

• CI r = reference Condition Index;

• a = slope of the line.

Dar Bouazza

The monthly evolution of heavy metal concentrations in the mussel (Cd and Zn) measured and adjusted in the site of Dar Bouazza is presented in Fig. 6.

These results show that, over months, adjusted metal concentrations present more stability and weaker fluctuations compared to the measured concentrations. This sim- ply means that the remarkable highest values (in April and

June) and lowest values (in August and September) are at- tenuated.

Measured Cadmium

0,0 0,2 0,4 0,6 0,8 1,0

Januar y Febr

uary Mar

ch April

May June July August

September months

µg/g

Adjusted Cadmium

0,0 0,2 0,4 0,6 0,8 1,0

Januar y February

March April

May

June July August

Septembe r months

µg/g

Measured Zinc

0 100 200 300

January February

March April

May

June July August

µg/g

Adjusted Zinc

0 100 200 300

Januar y Febr

uary Mar

ch April

May June July August

µg/g

FIGURE 6 - Temporal evolution of the contents of Cd and Zn measured and adjusted in the site of Dar Bouazza.

Cadmium measured in the flesh of the mussel varied in time (between 0.28 and 0.90 µg/g of dry weight), but the adjusted concentrations showed a small variation in time (from 0.52 to 0.74µg/g of dry weight).

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Zinc measured in mussel varied in time (between 109 and 249 µg/g of dry weight), whereas the adjusted concentrations showed relatively small changes during the year (from 173 to 208 µg/g of dry weight).

Ouled Ghanem

The temporal evolution of the metal contents (Cd and Zn) measured and adjusted in the site of Ouled Ghanem are presented in Fig. 7.

Measured Cadmium

0,0 0,5 1,0 1,5 2,0

February April June August months

µg/g

0,0 0,5 1,0 1,5 2,0

February April June August

Measured Zinc

0 100 200 300 400

Adjusted Zinc

0 100 200 300 400

FIGURE 7 - Temporal evolution of the contents of Cd and Zn measured and adjusted in the site of Ouled Ghanem.

Similarly, one can make the same remark about stability (weaker fluctuations) of adjusted concentrations of zinc and cadmium, with respect to the ones measured in the mussels.

The notable fluctuations in the concentrations measured in the mussels in the two studied sites, seemed then to be due to the physiological variations. According to that, the adjusted concentrations are supposed to be more repre- sentative of heavy metal concentrations in the marine environment. The physiology of the mussel distorts somehow the metal contents in the marine environment [2].

Spatial variations of the metal contents in mussel tissue

The study of the mussels at Dar Bouazza and Ouled Ghanem sites enabled us to compare their contamination with the adjusted concentrations, by eliminating the fluctuations due to physiology of the mussel (egg laying, gametogenesis).

The spatial variations of Cd and Zn in mussels from both study sites are shown in Fig. 8.

The Cd contents in the mussels taken at the level of Ouled Ghanem were very high compared to those found in Dar Bouazza, while Zn levels were almost the same in the two studied sites.

0,0 0,4 0,8 1,2 1,6

Dar Bouazza Ouled Ghanem Adjusted Zinc

0 50 100 150 200 250

February April June August m onths

µg/g

Dar Bouazza Ouled Ghanem

FIGURE 8 - Spatial variation of the metal contents in the mussels from Dar Bouazza and Ouled Ghanem.

The important contents of Cd in the site of Ouled Ghanem (more than two times contaminated to those of Dar Bouazza) are related to the fact that Ouled Ghanem is located in an agricultural area of great importance charac-

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terized by the intensive use of artificial fertilizers and pes- ticides rich in Cd. Moreover, the localization of Ouled Ghanem near the Jorf Lasfar zone where there is an important production of phosphoric acid and derived manures (rates raised out of Cd) by Phosphates Industrial Trans- formation [17]. There, average Zn amounts showed a small difference concerning the two studied sites.

CONCLUSION

The heavy metal contents recorded in the mussels (Mytilus galloprovincialis), considered to be bio-indicators of marine environment, showed seasonal variations, essentially due to the metabolism and reproduction cycle of the mussels (gametogenesis, eggs laying), and agreed with that reported in previous works [5]. The adjusted concentrations of heavy metals showed more stable variations.

The Condition Index made it possible to take into account the factors related to the physiology of the species [14], and to make adjustment of the raw results. For the studied metals, linear regression models of measured concentrations in the mussels to the inverse of the Condition Index showed that Hg had no significant correlation, con- trary to Cd and Zn. Only Zn and Cd were retained in this study and their accumulation rates were adjusted.

The adjusted concentrations of Cd and Zn can be considered as indicators of local environment contamination.

On this basis, we have compared the contamination levels of both studied sites and deduced that Ouled Ghanem is clearly more Cd-contaminated than Dar Bouazza, and this contamination is essentially due to local agricultural activity, whereas Cd level of Jorf Lasfar may be attributed to transport of water masses to Ouled Ghanem.

Finally, the accumulation rates of Cd, Hg and Zn recorded in the mussel Mytilus galloprovincialis do not indi- cate any notable contamination of the sites having been subject of this study.

ACKNOWLEDGMENTS

This work was supported by the Institute of Marine Research (Institut National de Recherche Halieutique

“INRH”). Special thanks to researchers from INRH, namely, Azeddine Ramzi, Fatima Zohra Bouthir and Ra- chid Chfiri for their help.

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Received: January 30, 2009 Accepted: March 02, 2009

CORRESPONDING AUTHOR Ikram Brahim

369 Bd du 2 mars 2^ème étage Casablanca MOROCCO

Phone: + 212 662 113242 E-mail: [email protected]

FEB/ Vol 18/ No 7b/ 2009 – pages 1348 - 1355