Effect of Habitat and Feeding Pattern on the Chemical Composition and Accumulation of Heavy Elements in Some Marine Fish Species in the Syrian Coast

With the increase in scientific knowledge, the importance of the chemical composition of fish for both the producer and the consumer, nutritionists, and environmentalists became clear, as interest began to study the chemical composition of fish and link it to the fish’s environment and its environmental and nutritional behavior [1]. The body of a fish is mainly composed of chemical substances represented by proteins, fats, water, some vitamins, especially fat-soluble ones (A, D, E, K), hormones, various mineral elements, pigments, and others. Knowing the chemical composition of fish and the factors that affect this composition allows us to evaluate the health of the fish and determine the efficiency of transferring nutrients and minerals from the environment to the fish [1]. Marine organisms, like other living organisms, are exposed to pollution by organic and inorganic pollutants, which negatively affect them and the surrounding environment, especially humans if they consume them. Among the inorganic pollutants are heavy elements found in very low concentrations in any ecosystem, which is natural. However, increasing the concentrations of these elements above their natural levels leads to physiological damage or death of various organisms, as they pose a direct threat to the lives of most aquatic organisms and are transmitted through food chains to cause harm to humans alike [2]. Differences in the chemical composition and extent of accumulation of heavy elements can exist between species and geographic location, and even at the level of individual fish according to many internal and external factors. External factors are environmental and nutritional influences and include water temperature, salinity, dissolved oxygen, mineral and heavy elements and diet composition [3], while internal factors include genetic factors related to the nature of the species composition, life stage, size, sex and sexual maturity, habitat and distribution of the fish (bottom or floating) and anatomical location in fish [3]. As for their location within the water column, migratory fish species have high levels of fat in their muscles, and this is closely related to the nature of the high activity of these species, as they are migratory fish species that are constantly moving and travelling, which requires large amounts of energy [4]. As for the bottom species, the presence of these fish at great depths and temperatures less than 10 degrees Celsius makes these fish slow-metabolizing and moving creatures, and even feeding and reproduction processes are low compared to pelagic fish, which leads to the accumulation of fats within their tissues, which in turn contributes to the adaptation of these species to low temperatures of 3-5 degrees Celsius [4]. This was confirmed by many previous studies that compared the chemical composition of fish according to their location within the water column, which found that bottom species are superior in terms of their nutrient content compared to pelagic species [5]. The results of a study conducted by Ashraf et al. [6]. To assess the relationship between the type of food for a group of fish and their chemical composition showed that the herbivorous species had the highest percentage of muscle fat, but at the same time, the results showed that the percentage of proteins was the highest in the muscles of the carnivorous species compared to the rest of the other species. In a study by Hussain et al. [7], on the effect of several factors and their relationship to the chemical composition of fish, their study concluded that carnivorous species have the highest moisture content. Their results also concluded that the highest percentage of proteins was in carnivorous species, as well as fats, which were the highest in omnivorous species. A study by Jovanovic et al. [8], showed that carnivorous and herbivorous species had the highest accumulation of heavy metals, followed by omnivorous and then herbivorous species. Another study by Hashim [9], showed similar results, with concentrations of most of the studied heavy metals being high in carnivorous and herbivorous fish species, followed by carnivorous and then herbivorous species. The present study aims to prove the extent to which the habitat of some economically important fish species within the marine environment affects their approximate chemical composition and the accumulation of heavy elements in their muscle tissues.

Study Area 

The study was carried out in the entire marine waters of the Latakia Governorate - Syria (Figure 1).

 Figure 1: Study site, marine waters of the Lattakia Governorate

Studied Species 

• Marbled Spinefoot (Siganus Rivulatus): Herbivorous fish, prefer shallow bottoms of compacted sand or rocks and coral [10] (Figure 2).

Figure 2: Marbled spinefoot Siganus rivulatus.

• Round Sardinella (Sardinella Aurita): Omnivorous fish, move in groups, and are highly migratory [11] (Figure 3).

 Figure 3: Round sardinella Sardinella aurita. 

• Red Sea Goatfish (Parupeneus Forsskali): Carnivorous fish, are found on sandy and rocky bottoms [12] (Figure 4).

 Figure 4: Red Sea goatfish Parupeneus forsskali.

• Golden Grey Mullet (Chelon Auratus): Omnivorous fish, live near beaches, in gulfs and in estuaries [13] (Figure 5).

 Figure 5: Golden grey mullet Chelon auratus.

Sample Preparation 

For fish samples, fish were brought regardless of size, age, and sex (random selection) during the seasons of the year, with 15-20 fish samples each season transported to the laboratory on the same day. Then the fish samples were weighed to the nearest gram, and the weights ranged (45.1-152.1)g, and the total and standard lengths were measured to the nearest cm, and the total lengths ranged (15.5-27.3)cm. After that, the scales and skin were removed. Then the meat was separated from the bone and kept in the freezer at a temperature (less than 0°C) until laboratory analyses were performed.

The Chemical Analysis 

All chemical analyses were expressed in (g/100g) of fresh fish muscle, according to AOAC [14].

• Moisture: At first, the initial weight of the samples was taken. Then samples were dried in an oven at about 1050°C for about 8 to 10 hours until a constant weight was reached and cooled in a desiccator and weight again. Then the samples were minced in an electric grinder. The percentage of moisture content was calculated by the following equation: 

Percentage (%) of moisture = (Weight loses/Original weight of sample) × 100

• Fat: For the estimation of fat content, the dried samples left after moisture determination were finely grinded and the fat was extracted with a nonpolar solvent, ethyl ether. After extraction, the solvent was evaporated and the extracted materials were weighed. The percentage of fat content was calculated as: 

Percentage (%) of fat = (Weight of extract/Weight of sample) × 100

Protein: The protein content of the fish was determined by micro-kjeldahl method. It involves conversion of organic nitrogen to ammonium sulphate by digestion with concentrated sulphuric acid in a microkjeldahl flask. The digest was diluted, made alkaline with sodium hydroxide and distilled. The liberated ammonia was collected in a boric acid solution and was determined titrametrically. The percentage of protien in the sample was calculated by the following equation:

Percentage (%) of protein = (c-b) × 14 × d × 6.25/a × 1000 × 100

Where:

a = sample weight (g)

b = volume of NaOH required for back titration and neutralize 25ml of 0.1N H2SO4 (for sample)

c = volume of NaOH required for back titration and neutralize 25ml of 0.1N H2SO4 (for blank)

d = normality of NaOH used for titration

6.25 = conversion factor of N to protein

14 = atomic weight of N

Ash: The ash content of a sample is the residue left after ashing in a muffle furnace (Gerhardt) at about 550-600 C till the residue become white. The percent of ash was calculated as follows:

Percentage (%) of ash = (Weight of ash / Weight of Sample) × 100

Heavy Element Measurement 

About 3 grams of fish muscle sample was taken. The wet digestion method was used to analyze heavy elements. Samples were transferred into digestion flasks and treated with 5ml HNO₃ (ultrapure, Merck) on the hot plate until the color turns into light yellow, nearly white. After this process, the samples were transferred to 25ml flanks and added double distilled water until 25ml. The solution was filtered by filter papers [15]. 

At each step of the digestion process, acid blanks (laboratory blanks) were prepared using the identical procedure to ensure that the samples and chemicals used were not contaminated from any of the mentioned possible sources. They contain the same digestion reagents as the real samples with the same acid ratios but without the fish sample. They were analyzed by Atomic Absorption Spectrophotometer (ShimadzoAA6800) before the real samples, to check if it will give the exact values of heavy metals in real samples [15].

Statistical Analysis 

The tested results were analyzed by ANOVA and the results were compared using the Least Significant Difference (LSD) method at the 0.05% level. using the programs: Microsoft Excel 2016, and SPSS 2011.

In the following, and through table 1, we note that the migratory species (rounded sardines) had the highest fat content (9%) and the lowest moisture content (72.1%), compared to the (Marbled spinefoot), which prefers shallow bottoms and had the lowest fat content (3%) and the highest moisture content (75.4%), while there were no significant differences between the studied species with regard to protein and ash content.

Table 1: Chemical composition of the studied species, their location in the marine environment, and their food pattern.

Table 1 shows that the rounded sardine fish species has the highest fat content among all the studied species. The high-fat content in rounded sardines may be due to the high-activity nature of this species, as it is a migratory fish species that is constantly moving and shifting, which requires large amounts of energy [16]. The Red Sea goatfish follows in terms of fat content, which is a benthic species. The presence of these fish at great depths and temperatures of less than 10°C makes these fish slow-metabolizing and slow-moving creatures, and even the feeding and reproduction processes are low compared to pelagic fish, which leads to the accumulation of fat within their tissues, which in turn contributes to the adaptation of these species to low temperatures of 3-5°C [17]. As for moisture, it is known that the moisture content of fish muscles is inversely proportional to their fat content, and this is what our results showed, as the moisture content of pelagic fish was higher than that of benthic fish, but this contradicts the study of Nybakken [18], which confirms that the water content of bottom fish exceeds that of surface fish due to the increase in water salinity with depth, and as a response by the fish to this, it leads to an increase in their water content. 

Table 1 shows the chemical composition of the studied fish species with their type of food. We notice that the Marbled spinefoot (herbivorous) had the highest moisture content (75.4%), while the Round sardinella (omnivorous) had the lowest (72.1%). As for the ash values, there were no significant differences between the studied species, as the omnivorous species (sardines and mullet) were the highest (1.9%), while the carnivorous and herbivorous species were the lowest (1.8%). Protein values did not differ significantly between the studied species (P>0.05), as the Red Sea goatfish (carnivorous) species was the highest (21%), while the herbivorous species (Marbled spinefoot) was the lowest (18.4%). As for fats, the Round sardinella (omnivorous) significantly outperformed all studied species (9%), while the Marbled spinefoot (herbivorous) was the lowest (3%). 

Comparing our results with the results of a study conducted by Ashraf et al. [6], to study the relationship between the quality of food for a group of fish with their chemical composition, we find that the results of our study contradict this study in that the herbivorous species they reached, where the highest in terms of muscle fat percentage. While the omnivorous species, specifically the rounded sardines in our study, had the highest percentage of fat, their results were consistent with our results in terms of the percentage of proteins, the carnivorous species had the highest percentage of protein in the muscles. 

In comparison with the study of Hussain et al. [7], on the effect of several factors and their relationship to the chemical composition of fish, their study concluded that carnivorous species have the highest moisture content, which is contrary to our study, in which we found that the highest muscle moisture rate was in the herbivorous species. They also concluded that the highest percentage of proteins was in carnivorous species, which is consistent with our study, as well as fats, which were the highest in omnivorous species. 

Table 2 shows the concentrations of heavy elements in the muscle tissues of the studied fish and their location in the marine environment. It can be concluded from this table that the rounded sardine had the highest concentrations of all heavy elements significantly (p<0.05) and its values were (0.113, 1.307, 1.912, and 0.221 mg/kg wet weight) for cadmium, nickel, copper, and lead respectively, followed by the Red Sea goatfish for cadmium and lead (0.083 and 0.105 mg/kg wet weight respectively) and the Golden grey mullet for nickel and copper (1.138 and 1.637mg/kg wet weight respectively). Finally, the Marbled spinefoot had the lowest concentrations of all the studied heavy elements, as its values were (0.040, 0.782, 0.944, and 0.070mg/kg wet weight Wet) for cadmium, nickel, copper and lead respectively. 

Table 2: concentrations of heavy elements in the muscle tissues of the studied fish, their location in the marine environment, and their food pattern. 

From the above, we note that the rounded sardine fish species was the highest in terms of accumulation of heavy elements in its muscle tissues. Since it is a migratory species and therefore moves continuously, it is exposed to many different marine environments with varying degrees of pollution, which makes it more susceptible to receiving heavy elements from different surrounding environments and thus accumulating them within its tissues to a greater extent than species that are stable in a specific environment or in a small number of marine environments. The rounded sardine species is characterized by being an active species that is always moving. Therefore a physiologically active species, and it has high metabolic rates, which increases its ability to accumulate heavy elements in its various tissues, including muscles [16]. Finally, as we found previously, the nature of its nomadic and constantly moving life makes it one of the fish species that require large amounts of energy sources, and thus the percentage of fat is high in its muscle tissues, which also leads to an increase in the ability of those tissues to accumulate heavy elements in them. 

As for the high concentrations of cadmium and lead in the muscle tissues of the Red Sea goatfish compared to the two species, the Golden grey mullet and Marbled spinefoot, this is consistent with many previous studies [19], which confirmed that heavy elements accumulate in the muscle and non-muscle tissues of benthic species more than pelagic species, especially cadmium and lead. The superiority of benthic species in accumulating heavy elements within their tissues compared to pelagic species can be explained by the fact that benthic species are in direct contact with bottom sediments rich in heavy elements, especially those with diameters of 63μm or less, especially in polluted sites, which enter the mouth and gills directly with breathing water and during feeding processes [20]. 

As for the high concentrations of nickel and copper in the muscle tissues of the Golden grey mullet, it may be due to the nature of the species that prefers river estuaries, which are rich in surface erosion sediments from agricultural lands and agricultural drainage water which is rich in pesticide and fertilizer residues, which are considered an important source of both copper and nickel [21]. 

From table 2, we note that omnivorous fish have the highest concentrations of heavy elements in the muscles of the studied fish. This may be because these fish feed on a wide spectrum of plant and animal nutrients, and therefore the sources of heavy elements entering their bodies are many and varied, which increases the chance of these elements accumulating in their muscle tissues, in addition to other factors related to their high physiological activity and the high percentage of fat within their muscle tissues. This is consistent with the study by Jovanovic et al. [17], where omnivorous species had the highest accumulation of heavy elements, followed by carnivorous species and then herbivorous species. Our study is also consistent with another study in 2014 by Hashim [9], where the concentrations of most of the studied heavy elements were high in omnivorous fish species (C. auratus and S. aurita), followed by carnivorous species (P. forsskali), and then herbivorous species (S. rivulatus). We also note from the previous results that the carnivorous fish species, (P. forsskali), has a high accumulation of heavy elements in its muscle tissues. This may be because carnivorous species generally exist at high levels within the food pyramid, in addition to the fact that most of them are benthic species and thus heavy elements accumulate in them to a large extent, as we found previously. 

For the herbivorous fish species, (S. rivulatus), the concentrations of heavy elements within its muscle tissues were somewhat low compared to the rest of the studied species and under our study conditions. This can be explained by the fact that herbivorous species generally have a narrow food spectrum and a low level within the food pyramid, and therefore the bioaccumulation of heavy elements within their tissues is very low [22], in addition to the low percentage of fat within their muscle tissues, which reduces the accumulation of heavy elements within their muscle tissues, as we found previously. 

The following is table 3, showing the average concentrations of the studied heavy elements in the muscle tissue of studied fish species and comparing them with the international limit values. 

Table 3: Average concentrations of heavy elements in muscle tissues of the studied fish compared to the permissible limits. 

We note from table 2, that the concentrations of heavy elements did not exceed the permissible limits and are therefore suitable for human consumption without risks. However, the fact that the concentrations of heavy metals in the muscles do not exceed the permissible limits does not necessarily mean that the fish are not contaminated or affected by pollution, because the heavy metals accumulate in varying proportions. Within the fish’s body, it varies according to the type of tissue, its physiological metabolic activity, and the percentage of fat in it. Therefore, heavy elements tend to accumulate in physiologically active tissues and organs such as the liver, kidneys, gonads, and gills, and to a lesser extent in the muscles [16].

From all of the above, we conclude that the chemical composition of fish and accumulation of heavy elements in their tissue is related to many internal and external factors, which work individually and together to influence the formation of the chemical composition of fish. We also found from the above that the muscles of migratory and benthic fish species were characterized by a high nutritional content compared to other fish species. In addition, the nutritional content in the muscles of omnivorous and carnivorous fish species was high compared to the muscles of herbivorous fish species. Migratory and benthic fish species have the highest capacity to accumulate heavy elements in their muscle tissues compared to other fish species, and omnivorous and carnivorous fish species have the highest capacity to accumulate heavy elements in their muscles compared to herbivorous species.

The authors thank Tishreen University and Manara University for providing logistical facilities and laboratory equipment’s, and ensuring appropriate conditions and conditions for carrying out the research. We also thank the fishermen who were diligent in collecting field data.

No conflicts of interest.

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