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Investigation of Hg Content by a Rapid Analytical Technique in Mediterranean Pelagic Fishes

Department of Biomedical and Dental Sciences and Morphofunctional Imaging, University of Messina, 98125 Messina, Italy
Author to whom correspondence should be addressed.
Separations 2018, 5(4), 51;
Received: 25 July 2018 / Revised: 1 October 2018 / Accepted: 9 October 2018 / Published: 24 October 2018
(This article belongs to the Special Issue Analytical Tools for Food Analysis)


Mercury (Hg) fish and seafood contamination is a global concern and needs worldwide sea investigations in order to protect consumers. The aim of this study was to investigate the Hg concentration by means of a rapid and simple analytical technique with a direct Mercury Analyzer (DMA-80) in pelagic fish species, Tetrapturus belone (spearfish), Thunnus thynnus (tuna) and Xiphias gladius (swordfish) caught in the Mediterranean Sea. Hg contents were evaluated also in Salmo salar (salmon) as pelagic fish not belonging to the Mediterranean area. The results obtained were variable, ranging between 0.015–2.562 mg kg−1 for T. thynnus species, 0.477–3.182 mg kg−1 for X. gladius, 0.434–1.730 mg kg−1 for T. belone and 0.004–0.019 mg kg−1 for S. salar, respectively. The total Hg tolerable weekly intake (TWI) and tolerable weekly intake % (TWI%) values according to the European Food Safety Authority (EFSA) were calculated. The results highlighted that the pelagic species caught in the Mediterranean Sea should be constantly monitored due to their high Hg contents as well as their TWI and TWI% with respect to S. salar samples.

Graphical Abstract

1. Introduction

The Mediterranean diet with its constituents is nowadays recognised to be one of the most healthy diets worldwide [1,2]. In addition to the several plant-based foods promoted by this diet, a moderate consumption of fish is important for its contribution of healthy nutrients such as Omega 3, or “n-3 long-chain polyunsaturated” fatty acids but with low levels of saturated fatty acids [3,4,5]. However, according to the European Food Safety Authority (EFSA), the consumption of seafood and fish is relevant for the assessment of dietary exposure to mercury from food. Fish meat is considered among the most important contributors of mercury intake for people, from children to adults [3].
Mercury (Hg) commonly known also as quicksilver, is a heavy, silvery-white liquid metal chemical element, highly toxic to the environment and living creatures. Indeed, contamination of fish species is a potential health hazard for humans as the last component of the food chain. In this context, since the Mediterranean Sea is a closed sea, with a limited exchange of water mainly coming from the Atlantic Ocean and with a minor contribution from the Black Sea, it results to be one of the geographic areas of concern for Hg concentrations due to the ever-increasing industrialization processes [6,7,8,9,10,11]. People may be exposed to inorganic Hg by their employment or pollution, and exposed to organic Hg (methylmercury, CH3Hg+) predominantly through the consumption of seafood [9,10]. The pelagic species represent a significant fishing resource and a widely consumed food in the human diet because of the traditional recipes of the Mediterranean diet and also as sushi and sashimi, typical of Japanese cuisine, have rapidly spread through Western countries during the last few decades [4,5,12]. In the light of all the above, the purpose of this study was to carry out an investigation using a rapid analytical technique on the Hg content in some of the most representative pelagic fish species, in particular, Tetrapturus belone, Thunnus thynnus and Xiphias gladius (spearfish, tuna and swordfish, respectively) caught in the Mediterranean Sea and Salmo salar (salmon) as a commercial pelagic fish not belonging to the Mediterranean area.
The Hg contents were also used to calculate the total Hg Tolerable Weekly Intake (TWI) and Tolerable Weekly Intake % (TWI%) values for each species according to the EFSA guidelines [3] to underline the potential health risk for consumers.

2. Materials and Methods

A total of 48 fishes from the Mediterranean Sea were kindly provided by the Italian Institute for Environmental Protection and Research. Thunnus thynnus, Xiphias gladius and Tetrapturus belone samples were collected from Cape d’Orlando, Cape Rasocolmo, Patti Gulf, Tyrrenian Sea and Strait of Messina (Italy). The commercial Salmo salar samples were produced in the North Sea (Norway and Scotland, UK). The specimens had lengths of 15 to 240 cm and weights of 150 g to 220 kg. From each sample, an approximate amount of about 50–100 g of muscle tissue from the region around abdominal cavity was taken. Once in the laboratory, samples were frozen at −20 °C and stored until analysis.
For Hg level determination, the analyser used was a Milestone DMA-80 Direct Mercury Analyser (Milestone GmbH, Leutkirch im Allgäu, Germany). It is an innovative analytical instrument that allows for the elimination of the use of reagents, and to simultaneously obtain validated results with no digestion and chemical pre-treatment steps.
The DMA-80 has been used according to the US EPA method 7473 (mercury in solids and solutions by thermal decomposition, amalgamation, and atomic absorption spectrophotometry). Furthermore, it was compliant with the ASTM method D-6722-01 (total mercury in coal and coal combustion residues) and ASTM method D-7623-10 (total mercury in crude oil).
About 0.1 ± 0.001 g of the samples were weighted, put onto nickel vessels and introduced to the direct analyser, dried at 200 °C for 3 min, then chemically and thermally decomposed at 650 °C for 2 min. Hg content of samples was determined by measuring absorbance at 253.7 nm.
The calibration curve was constructed using standards with known concentrations of mercury, associating a value of absorbance to each known Hg concentration. The standard solutions were prepared in glass flasks (5 concentration points from 0.050 to 10 mg kg−1) from the 1000 mg/L certified standard (CZECH Metrology Institute Analytika). The evaluation of the linearity was based on six injections of the standard solution. Good linearity was observed, achieving a correlation coefficient r = 0.9996. The detection limit (DL) and quantification limit (QL) were counted as 3 times and 6 times the standard deviation for blanks and were 0.25 μg kg−1 and 0.51 μg kg−1, respectively.
Hg content for a certified reference material TB149, a spearfish sample, is shown in Table 1. Analyses of the certified reference material TB 149 showed satisfactory recoveries of about 95% for Hg content both for total Hg and methylmercury (CH3Hg+). The data obtained were shown to agree with those of the certified value, thus the procedure was confirmed to be adequate for measuring Hg contents in the muscle sample collected.
The software of the instrumentation automatically calculates the total Hg levels of the sample by interpolating the absorbance values directly with the calibration curve [4].
The risk exposure to CH3Hg+ expressed as TWI (mg kg−1 b.w.) was calculated for an average serving portion of 200 g of fish for a 60 kg adult according to the following formula (Equation (1)).
Tolerable Weekly Intake Hg (TWI) = [Hg] × serving portion/body weight
the corresponding weekly risk exposure to CH3Hg+ was calculated as TWI% of 4 mg kg−1 b.w. recommended by EFSA (2012).

3. Results

Table 1 shows the reported Hg content obtained for T. thynnus, X. gladius, T. belone and S. salar samples expressed in mg kg−1.
The results were variable, ranging from 0.015–2.562 mg kg−1 for T. thynnus species, 0.477–3.182 mg kg−1 for X. gladius, and 0.434–1.730 mg kg−1 for T. belone. For S. salar the Hg content obtained ranged from 0.004–0.019 mg kg−1. The following decreasing order among the examined pelagic species could be defined: X. gladius (mean 1.829 mg kg−1) > T. thynnus (mean 1.288 mg kg−1) > T. belone (mean 1.082 mg kg−1) > S. salar (mean 0.012 mg kg−1). The significant p-level below 0.05 obtained by a Kruskal-Wallis test, confirmed that the samples of T. thynnus and of T. belone have a Hg level not significantly different between them (Figure 1).
In Table 2, the ranges of Hg contents and their relative TWI value calculated in this study are shown. The weekly consumption of a portion of 200 g of the most examined pelagic fish species by a 60 kg adult body weight, point out a remarkable health risk of exposure to the toxic action of CH3Hg+ for T. thynnus, X. gladius and T. belone, as the TWI value was exceeded in many samples caught in the Mediterranean Sea. Meanwhile, no risk were observed for S. salar samples from the North Sea.

4. Discussion

The use of the DMA-80 permitted fast and reliable results. In fact, its unique processing of the sample by thermal decomposition, amalgamation, and atomic absorption spectrometry allowed the direct analysis of the fish samples.
The Hg values showed a noteworthy variability among the examined samples. As expected, these differences could be due to different biological and ecological aspects of the species, including different territory characteristics between the collection sites around Sicily (Italy) and especially between the Mediterranean Sea and the North Sea. In regard to the collection sites, the samples caught in the Strait of Messina were highly rich in Hg with respect to the other Mediterranean Sea collection sites and the North Sea.
Furthermore, focusing on T. thynnus samples, it was possible to notice that the highest Hg contents were found in the samples collected from the Strait of Messina and the Patti Gulf, followed by the samples collected in the other sites. Fish size correlations with Hg contents were difficult to establish and further studies should investigate this aspect.
A comparison of the Hg contents with the literature data on pelagic fish species previously investigated, pointed out that the results obtained in this study were in agreement, with some minor differences, with Hg levels in muscle tissue previously reported in this geographic area and also with fishes from oceans around the world [4,5,11,13,14,15].
It is important to also compare the results obtained with the EC (European Commission) [16] maximum levels for certain contaminants in foodstuffs, including seafood. Unfortunately, it was seen that except for the S. salar species, all other samples fell outside the EC maximum level established. For the T. thynnus species, two samples exceeded the maximum limit, while for T. belone and X. gladius, most samples were above the limit values.
In light of the those results, it is interesting to note that an adult who consumes 200 g of T. thynnus, X. gladius and T. belone fish weekly, the limit of mercury exposure, which corresponds to one or two serving portions of those fishes, is highly exceed. There is also a minor risk for the weekly consumption of S. salar fish muscle.
The ingestion of Hg from the analysed fish samples could present health risks for the average consumer even at 200 g portions, which is a regular serving size according to the Mediterranean diet. Indeed, it should be kept in mind that regular or excessive consumption of such pelagic fish species might exceed the recommended weekly intake (TWI).
In light of all the above, pelagic fishes may contribute significantly to the intake of hazardous elements from the environment especially, in this case, the Mediterranean Sea.

5. Conclusions

The results established that the three pelagic species T. thynnus, X. gladius and T. belone caught in the Mediterranean Sea need to be constantly monitored due to their high Hg content. In most cases, the content level was higher or very close to the European legislative limit [3]; while, S. salar Hg concentrations were far from the maximum limits.
Further studies should address the monitoring of Hg levels in Mediterranean fish specimens, in order to ensure the population’s safety.

Author Contributions

G.D.B. and N.C. conceived and designed the experiments; R.T. performed the experiments; G.D.B. and N.C. analyzed the data; G.D.B and N.C. contributed reagents/materials/analysis tools; R.T. wrote the paper.


This research was funded by (Leo Club International District 108 ITALY) grant number (Leo Hunting Mercury Project grant).

Conflicts of Interest

The authors declare no conflict of interest and the founding sponsors had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, and in the decision to publish the results.


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Figure 1. Kruskall-Wallis test for analyzed samples.
Figure 1. Kruskall-Wallis test for analyzed samples.
Separations 05 00051 g001
Table 1. Sample information: Site of collection, length (cm) and weight (kg), concentration levels of Hg expressed as mg kg−1.
Table 1. Sample information: Site of collection, length (cm) and weight (kg), concentration levels of Hg expressed as mg kg−1.
SampleSiteLength (cm)Weight (kg)Hg (mg kg−1) a
TT0032Rasocolmo Cape26.800.320.0562 ± 0.0032
TT0033Rasocolmo Cape27.000.360.0833 ± 0.0035
TT005d’Orlando Cape20.000.220.0192 ± 0.0007
TT094Patti Gulf195.00150.000.5570 ± 0.0413
TT0951Patti Gulf115.0028.000.7057 ± 0.0275
TT0952Patti Gulf240.00180.000.9057 ± 0.0514
TT0953Patti Gulf90.0016.000.7697 ± 0.0265
TT096Southern Tyrrhenian sea15.400.150.0146 ± 0.0028
TT097Southern Tyrrhenian sea24.800.300.0348 ± 0.0140
TT158Strait of Messina128.0036.001.3689 ± 0.0631
TT159Strait of Messina165.0066.002.5618 ± 0.4609
XG321Strait of Messina173.0074.002.0623 ± 0.2434
XG330Strait of Messina52.005.000.4775 ± 0.0287
XG343Strait of Messina171.0058.000.8837 ± 0.0279
XG344Strait of Messina170.0061.002.1336 ± 0.0908
XG352Strait of Messina161.0055.002.2072 ± 0.0310
XG365Strait of Messina181.0048.002.5557 ± 0.2700
XG367Strait of Messina133.5035.002.0135 ± 0.1949
XG372Strait of Messina180.0070.001.7924 ± 0.0598
XG376Strait of Messina180.0065.001.6910 ± 0.0302
XG377Strait of Messina140.0030.002.1496 ± 0.1989
XG379Strait of Messina164.0054.002.9100 ± 0.0171
XG380Strait of Messina196.0083.001.5631 ± 0.1708
XG381Strait of Messina180.0075.001.2367 ± 0.2219
XG384Strait of Messina183.0049.003.1819 ± 0.1949
TB 103Strait of Messina155.0013.000.4344 ± 0.0335
TB 106Strait of Messina151.0011.000.7986 ± 0.0889
TB 108Strait of Messina156.0016.001.6499 ± 0.2199
TB 113Strait of Messina150.0011.000.6695 ± 0.0563
TB 117Strait of Messina170.0016.000.8687 ± 0.0230
TB 120Strait of Messina179.0022.001.7303 ± 0.0268
TB 121Strait of Messina151.006.000.6021 ± 0.0642
TB 122Strait of Messina178.0021.001.6086 ± 0.1290
TB 126Strait of Messina157.0016.000.5199 ± 0.0209
TB 131Strait of Messina155.0016.000.6017 ± 0.0433
TB 132Strait of Messina142.0013.000.5306 ± 0.0525
TB 144Strait of Messina151.0011.000.5693 ± 0.0534
TB 145Strait of Messina155.0014.001.0583 ± 0.2837
TB 149Strait of Messina148.0012.000.5559 ± 0.0228
SS1Scotlandn.d.n.d.0.0188 ± 0.0007
SS2Norwayn.d.n.d.0.0089 ± 0.0013
SS3Norwayn.d.n.d.0.0114 ± 0.0004
SS4Norwayn.d.n.d.0.0193 ± 0.0014
SS5Norwayn.d.n.d.0.0091 ± 0.0003
SS6Norwayn.d.n.d.0.0117 ± 0.0004
SS7Norwayn.d.n.d.0.0037 ± 0.0006
SS8Norwayn.d.n.d.0.0159 ± 0.0004
SS9Norwayn.d.n.d.0.0087 ± 0.0005
n.d.: Not determined. a Mean value (n = 3).
Table 2. Ranges and mean values of tolerable weekly intake (TWI) and relative % (TWI%) for Hg contents in analyzed fishes.
Table 2. Ranges and mean values of tolerable weekly intake (TWI) and relative % (TWI%) for Hg contents in analyzed fishes.
SpeciesN SamplesHg
Range Min–MaxMean ValueMean Value ± Dev.stRange Min–MaxMean Value
S. salar90.004–0.0020.0120.0003 ± 0.0015.10–11.286.97
T. belone140.434–1.7301.0820.0200 ± 0.011253.41–1009.35508.23
T. thynnus110.015–2.5621.2880.0150 ± 0.0188.52–1494.38375.29
X. gladius140.477–3.1821.8290.0400 ± 0.017278.53–1856.081119.08

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Di Bella, G.; Tardugno, R.; Cicero, N. Investigation of Hg Content by a Rapid Analytical Technique in Mediterranean Pelagic Fishes. Separations 2018, 5, 51.

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Di Bella G, Tardugno R, Cicero N. Investigation of Hg Content by a Rapid Analytical Technique in Mediterranean Pelagic Fishes. Separations. 2018; 5(4):51.

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Di Bella, Giuseppa, Roberta Tardugno, and Nicola Cicero. 2018. "Investigation of Hg Content by a Rapid Analytical Technique in Mediterranean Pelagic Fishes" Separations 5, no. 4: 51.

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