The First Assessment on Metal Contamination in the Critically Endangered Sawback Angelshark (Squatina Aculeata) From North-Eastern Mediterranean


 A Sawback angelshark (Squatina aculeata) classified as “Critically Endangered” Global Red List by the IUCN (2017) and CITES (2017) is one of the threated elasmobranch groups and the increased pollution in the marine environment negatively impact angelshark species. Despite this, limited information exists about the effects of toxic chemical contamination in angelsharks. Therefore, present study was conducted to elucidate heavy metal accumulation in Squatina aculeata captured from the Iskenderun Bay, North-Eastern Mediterranean by electrochemical technique. Arsenic (As), Mercury (Hg), Copper (Cu), Iron (Fe), Zinc (Zn) and Mangan (Mn) were found by electrochemical analyzer. These results showed that Fe and Zn were the highest concentrations in all tissues of S. aculeata and, the accumulations of Hg, Fe, Zn and Mn were significantly different in the gill, liver and muscle tissues (P < 0.05). As and Hg concentrations exceeded maximum limit legalized by FAO (2003) and UNEP (2008). This results confirmed that the metal contamination in North-Eastern Mediterranean area may pose a potential threat to the sustainability of S. aculeata in marine ecosystem


Introduction
The environment pollution by trace metals is an important global problem. Toxic metals reason damaging impacts to terrestrial and aquatic animals. Both natural and anthropogenic events generally generate heavy metals that contaminate the aquatic habitats through sewage canals, river loads and atmospheric depositions ).
Sharks are susceptible to heavy metals due to the slow elimination and large capacity to incorporate metals and, accumulate trace metals in their tissues because of their diet as they feed on many trophic organisms (Lopez et al. 2013). Heavy metal accumulation in each aquatic organism show variation according to its diet and habit. Hereafter, sharks as a top predator feeding on a large number of organisms would be having in a higher level of heavy metal accumulation into their body. Furthermore, their widespread distribution and signi cance to the ecosystem makes them ideal sentinel organisms for A notable number of elasmobranch species (about 25%) are currently regarded as threatened on the authority of International Union for Conservation of Nature (IUCN). Angel sharks (Squatinidae) are among the critically endangered sh species in marine ecosystem and actions goaling to these species conservation has been recommended in lately times (Martins et al. 2020). Populations of the three Critically Endangered angel sharks have been severely depleted in the Mediterranean Sea. Sawback angelshark (S. aculeata) is classi ed as "Critically Endangered" Global Red List by IUCN (2017) and CITES (2017). Sawback angelshark is an Atlanto-Mediterranean coastal shark live in continental shelf and tropical eastern Atlantic, and is extremely rare in the east and west parts of the Mediterranean (Ergenler et al. 2020). Altough the occurrence of the species in the Turkish marine waters has been reported before (Basusta, 2002;Filiz et al. 2005;Ergüden and Bayhan 2015), the new information on the rare occurrence of a male specimen of Sawback angelshark S. aculeata from the northeastern Mediterranean coast of Turkey was made by our laboratory team on February 19, 2019 (Ergenler et al. 2020). Hereby, the preliminary data on metal contamination in threatened sawback angelsharks from the northeastern Mediterranean will available with this research. This species inhabits coastal areas of the northeastern Mediterranean coast of Turkey regions under great anthropogenic impact. It has been often reported that the Mediterranean Sea is at high toxicological risk (Storelli et al. 2011), as a result of speci c hydrographical features and actual density of the anthropological effect. Especially Iskenderun bay is highly exposed to heavy metal accumulation impact due to agricultural, industrial and urban wastes. This bay has a major input of contaminants from maritime, domestic and industrial sources  This species was transported to the laboratory. Morphometric measurements were carried out to the nearest 0.1 mm by a caliper. The specimen was identi ed as S. aculeata with the diagnostic characteristics described by Compagno (1984) and Serena (2005). Measurements such as total body length, standard length and wet body weight of sampling shark (mean ± SD) were found as 1170 mm, Liver and gill tissues were also dissected with the aid of a scalpel with stainless steel blade, and samples were stored in plastic bags at − 18°C until analyses.

Trace Metal Analysis
Brie y, the trace metal determination in the muscle and liver tissue was performed on a wet weight basis through an acid digestion adapted from AOAC O cial Method 999. 10 (2002), where a mixture of 10 mL of nitric acid (HNO 3 ), 0.25 mL of hydrogen peroxide (H 2 O 2 ) and 1 g of the sample (taken with a scalpel from the central part of the muscle) kept in a bath-water at 60° C for one hour to perform acid digestion, after which the samples were allowed to cool at room temperature. Later, the resulting solution was ltered and increased to 100 mL with distilled and deionized water. To avoid contamination, all glassware Mean values for trace metals measured in the different tissues of Sawback angelshark are illustrated in Table 1. The mean concentrations of Fe and Zn were the highest concentrations in gill, liver and muscle tissues of S. aculeata (Fig. 2). The mean concentrations of Cu and Mn were the lowest concentrations in all tissues of S. aculeata.
The concentrations of all the heavy metals were signi cantly different in the gill, liver and tissues (P < 0.005) except As and Cu in S. aculeata ( In general, the concentration orders of heavy metals in the tissues were found to be gill > liver > muscle for the studied sawback angelshark, except for Hg ( Table 3). The concentrations order of Hg was found to be muscle > liver > gill. Among the gill and liver tissue samples of S. aculeata, Fe and Zn were detected as highest concentrations followed by As, Hg, Mn and Cu; Among the muscle tissue of S. aculeata, Fe and Zn were detected as highest followed by Hg, As, Mn and Cu (Table 1; Fig. 2).

Discussion
The present study provides rstly, the preliminary data on metal contamination in the Critically endangered Sawback angelshark from the northeastern Mediterranean by electrochemical technique.
Angel sharks often occupy coastal waters and thus exposed to different contaminants, the high trophic level of this environment could increase the accumulation of such pollutants (Ellis et al., 2020 As seen in Table 2, the restricted data is available on the levels of metal contamination in angel sharks. Especially, no study on metal accumulations of Critically endangered Sawback angelshark was found in the literature searches. reported that there is no report on the contaminants that have impacted on angel sharks, and further studies are required on the potential impacts of habitat degradation and water quality for inhabiting areas of angel shark, especially industrialized areas. Therefore, the present nding with exceeded legalized limit of Hg concentrations are very valuable for the conservation and sustainability of S. aculeata as management consideration.

Human Health Risk Assessment
Page 7/12 The regulatory limits of As, Hg, Cu, Fe, Zn and Mn concentrations are directly compared in Table 5.
Concentrations of Cu, Fe, Zn and Mn as found in this study did not constitute a risk factor for human health being below the permissible limits for human consumption reported by FAO (2003) and UNEP (2008). Average total arsenic contents in S. aculeata was 7.5416 ± 0.6548 µg g − 1 wet weight, which exceeded maximum limit legalized for any kind of food. Similarly, Total mercury average content in S. aculeata was 19.9942 ± 1.6116 µg g − 1 (wet weight), which exceeded the proposed limit value which is 0.2-2 µg g − 1 , con rming that the consumption of S. aculeata is a high risk threating the health of consumer (Table 3) Recently, much attention has been given on electrochemical detection methods which are inexpensive, highly sensitive and easily adaptable for in situ assessment with short analytical periods in the eld of

Conclusion
The present study revealed rst data on metal contamination in the critically endangered Sawback angelshark from the northeastern Mediterranean by electrochemical technique. As and Hg concentrations exceeded maximum limit legalized by FAO (2003) and UNEP (2008). This results con rmed that the metal contamination in North-Eastern Mediterranean area may pose a potential threat to the sustainability of S. aculeata in marine ecosystem and the concentrations detected is the above the safety limits for angelshark that should be taken into consideration regarding human consumption. Our data also reveal that the electrochemical technique can be used successfully in determination of trace metal concentrations in sawback angel shark.
Declarations 32. Zar JH (1996) Biostatistical analysis, 3rd edn. Prentice Hall Inc, Englewood Cliffs. Figure 1 Map showing capturing site (black dot) of Squatina aculeata in the Iskenderun Bay. Note: The designations employed and the presentation of the material on this map do not imply the expression of any opinion whatsoever on the part of Research Square concerning the legal status of any country, territory, city or area or of its authorities, or concerning the delimitation of its frontiers or boundaries. This map has been provided by the authors.