Introduction

Heavy metals, the collective name for a group of metallic elements (e.g., mercury, lead, cadmium, chromium, etc.) known for their high density, atomic weight, and/or atomic number, are persistent pollutants released into aquatic and terrestrial environments through industrial discharges, agricultural runoff, and urban wastes. Due to the increase in anthropogenic activities and industrialization, soil, water, and the atmosphere are exposed to heavy metal pollution, which creates a major environmental problem that affects human health and food quality (Wang et al. 2022). As these pollutants disperse in water, aquatic organisms absorb them and build up bioaccumulation in their tissues over time (Ali et al. 2019). Continuous exposure to heavy metals threatens not only the health and survival of individual organisms but also the integrity of entire aquatic ecosystems (Taiwo et al. 2014). Understanding how these heavy metals accumulate in organisms helps to assess their impact on the body (Rainbow 2018; Basaran 2022). Even at trace levels, heavy metals can be hazardous to most organisms because of the irreversible inhibition of certain enzymes (Henczová et al. 2008). Polluted water frequently contains combinations of multiple metals, significantly affecting aquatic organisms (Zeng et al. 2019; Fettweis et al. 2021). These heavy metals' interaction can influence their toxicity and accumulation (de Medeiros et al. 2020). Hence, studying how trace elements interact within tissues is essential, especially since their concentrations have risen in nature (Briffa et al. 2020).

The absorption of potentially toxic elements from the environment occurs in aquatic animals through their skin, respiration, and nutrition. Because of their inability to break down naturally, metals tend to accumulate in the environment, leading to potential hazards for organisms as they enter the aquatic food chain (Al-Yousuf et al. 2000; Tao et al. 2012; Ali et al. 2019). Additionally, investigating the amount of metal in the tissues of consumed organisms is vital for evaluating their nutritional quality (Mani et al. 2022). Amphibians serve as vital bioindicators for monitoring water pollution (Simon et al. 2010; Şişman et al. 2015, 2021; Dönmez and Şişman 2021). Thanks to their permeable skin, which enables them to absorb substances from the environment rapidly (Hecnar 1995; Tatlı et al. 2022a), frogs can absorb heavy metals from their aquatic habitats, effectively accumulating these harmful substances in their bodies. Analyzing natural frog populations can offer valuable insights as reliable bioindicators (Lower and Kendall 1990; Vogiatzis and Loumbourdis 1998), shedding light on the persistent trend of metal buildup and pollution dynamics in aquatic ecosystems across an extended duration. This accumulation can lead to potential health risks for humans if the contaminated amphibians are ingested. Indeed, all amphibian species within the Ranidae family are suitable for human consumption, except for the frog species, which might be hazardous for humans (Neveu 2009). Frogs are recognized for their nutritional value, being low in fat and rich in protein and minerals (Özogul et al. 2008). Meat is one of the most basic foodstuffs for humans, and it is recommended to consume it from different sources to get the full benefit of its nutrients. However, due to the high consumption of red meat and its potential health implications when overindulged, researchers are now investigating alternative food sources (Şimşek et al. 2022). Frogs are emerging as a promising option, more affordable and safer for consumption than red meat (Stuart et al. 2004). The edible portions of frogs mainly consist of their legs, which offer a taste similar to chicken but with a slightly firmer muscle texture.

Research on the potential for heavy metal exposure from the consumption of frog legs has gained momentum in recent years (Borković-Mitić et al. 2016; Mani et al. 2022; Tatlı et al. 2022b; Zhelev et al. 2020). There are three edible frog species (Pelophylax ridibundus, Pelophylax bedriagae, and Pelophylax caralitanus) in Türkiye, and risk estimation for environmental pollution and human health by evaluating them together has not been explored in the literature up to now. Hence, this study aimed to analyze the concentration of heavy metals in the tissues (muscle and liver) and aquatic habitats (water and sediment) of three species of frogs belonging to the genus Pelophylax, with bioindicator properties, and also to evaluate if there are any health risks associated with the consumption of frog legs.

Materials and method

Study area and sampling

The animal capture was approved with the permission of the local ethics committee of Recep Tayyip Erdogan University (Decision number: 2021/15). In this research, adult Anatolian water frogs (Pelophylax ssp.) were utilized, along with sediment and water samples from their natural habitats. Sampling was carried out between May and September 2022 at 11 different locations, namely Istanbul, Bursa, Manisa, Antalya, Burdur, Isparta, Adana, Kayseri, Şanlıurfa, Erzincan, and Samsun (Fig. 1), and the habitats' images were presented in Fig. 2. Also basic characteristics of the sampling sites were given on Table S1. Seven of these sampling stations were inhabited by Pelophylax ridibundus (locations 1, 2, 7, 8, 9, 10 and 11), two by Pelophylax bedriagae (locations 3 and 4) and two by Pelophylax caralitanus (locations 5 and 6).

Fig. 1
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Sampling locations in this study

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Fig. 2
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Different habitat types in the sampling stations

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A total of 110 frogs were sampled from 11 populations within the distribution range of Pelophylax ssp. Frogs living in lakes, ponds, puddles, or farmland/ditch (Fig. 2) were captured manually or with dip nets and put in plastic carrier bags. Samples of both water and sediment were taken from the same habitats. A polyethylene sampling bottle was used to collect 500 ml water samples. The sediment surface (0–3 cm depth) was excavated using a steel shovel to gather approximately 1 kg of sediments, which were then placed in plastic carrier bags. All samples (water, sediment, and frogs) were placed in insulated containers and conveyed to the lab, where muscle and liver tissue and sediment samples were kept in a freezer set to -20 °C for later experiments. Fifty milliliters of water were filtered through 0.45 mm PTFE filters of particle size, and five drops of HNO3 (Suprapur, Merck) were added to reduce the pH to a level of less than two prior to cooling for further heavy metal analyses (US EPA 1994).

Sample digestion

The Anatolian frog samples were thawed and washed with ultrapure water at room temperature (RT). The sediment samples were initially kept at RT and later dried in the oven. Each frog's snout-vent length (SVL) and weight were recorded (Table S2). Subsequently, with a stainless-steel dissection set, the frogs' livers and hind leg muscles were extracted with precision, and each frog was dissected individually. Two grams of fresh weight from the leg and liver tissues were allocated to separate digesting vessels. Digestion of the sediment was conducted using 0.5 g of the dry sample. 5 ml of HNO3 (Suprapur, Merck) were added to each to tubes containing tissues and sediment samples. The tubes were enclosed with polypropylene lids and were kept overnight. A block heater was used to incubate the tubes for 2.5 h at 95 °C. After being brought down to RT, the tubes underwent an additional 2 h of incubation at 95 °C with 2.5 mL of H2O2 (Suprapur, Merck). The lids were taken off, and the tubes were left inside the block heater with the fluid content was reduced to approximately 2 mL. Afterward, the solutions were diluted with ultrapure H2O, strained with PTFE syringe filters (0.45 mm pore size), and placed in a + 4 °C fridge until the measurements were completed (US EPA 1996).

Heavy metal analysis and quality assurance

To assess and confirm the procedures' effectiveness at digesting the reference material (ERM-CE278k Mussel Tissue), the same approach employed on the samples was likewise performed there (Table S3). ICP-MS (Agilent, 7800) was used to determine the concentration of TE. The internal standards and blank samples were also examined to look for potential interferences with the ICP-MS readings or sample preparation processes (Öztürk et al. 2021).

Estimation and evaluation of health risk

By utilizing the heavy metal concentrations in frog legs, the Estimated Daily Intake (EDI) was assessed and contrasted with the provisional tolerable daily intake (PTDI) according to the FAO/WHO Joint Expert Committee on Food Additives (JECFA 1982, 2011a, b). Moreover, a Risk Coefficient (RK) method was employed to estimate the possible risk to human health from metals from consuming of frog legs (USEPA 2015). The THQs, a quantification of heavy metal concentration compared to the dosage coefficient, illustrate the probability of persistent non-carcinogenic exposures (USEPA 2015). If the THQ is greater than 1, the studied heavy metal has deleterious impacts. EDI and THQ calculations were made as follows:

$$EDI=\frac{\mathrm{C }\times \mathrm{ FCR}}{BW}$$
$${THQ}_{s}=\frac{\mathrm{C }\times \mathrm{ EF }\times \mathrm{ ED }\times \mathrm{ FCR}}{\mathrm{RfD }\times \mathrm{ BW }\times \mathrm{ EF }\times \mathrm{ ED}} \times {10}^{-3}$$

C: average metal concentration in the frog leg, FCR: Frog consumption (g/person/day), Average serving size for adults (227 g: US EPA 2000), ED: Duration of exposure (76 years/mean life expectancy in Türkiye) (WHO 2015), EF: frequency of exposure (365 days/year), BW: Mean body weight (72.5 kg Basara et al. 2016), RfD: Reference dose (mg/kg/day), 10–3: Conversion factor coefficient.

The non-carcinogenic health risk of all elements contained in frog legs (Newman and Unger 2002) was evaluated using the Health Index (HI). This index is formulated by the sum of the Total Hazard Quotients (THQs), and is represented as:

$$\mathrm{HI}={\mathrm{THQ}}_{\mathrm{Cu}}+{\mathrm{THQ}}_{\mathrm{Pb}}+....+{\mathrm{THQ}}_{\mathrm{x}}$$

Data analysis

The Shapiro–Wilk and Levene tests were used to verify that the data conformed to a normal distribution and equal variance, respectively. When the data did not align with the normal distribution, it underwent the log10 transformation process. Detection and removal of outlier data were performed with the Boxplot technique. To investigate the variations in heavy metal contents among species and different stations, a one-way analysis of variance (ANOVA) was utilized, accompanied by a post hoc Tukey test. Additionally, to ascertain if any discrepancies in heavy metal amounts existed between the sexes, the independent sample t-test was used. The Pearson correlation method was applied to ascertain the links between elements. Furthermore, a linear regression analysis was implemented to analyze the potential connection between metal concentrations in Anatolian water frogs' liver and muscle tissues, considering their body lengths and weights. Using the SPPS 22 (IBM, USA) package program, statistical analyses were carried out and 5% (p < 0.05) was chosen as the significance level.

Results and discussion

Heavy metal concentrations in frog muscle and liver tissues were calculated in µg kg−1, and the results are given in Figs. 3 and 4, respectively. The concentrations of heavy metals in the muscle tissue of frogs obtained from multiple sites were as detailed: As, 7.08–114.90 µg kg−1; Cd, under detection limits (UDL)–13.52 µg kg−1; Cr, 41.75–479.35 µg kg−1; Cu, 358.45–896.05 µg kg−1; Pb, 34.74–1390.10 µg kg−1, and Zn 4744–22306.76 µg kg−1 (Table S4; Fig. 3). The average concentrations (µg kg−1) of heavy metals in muscle tissue were determined in the following order: Zn (9785.11) > Pb (52.58) > Cu (25.01) > Cr (15.36) > As (3.92) > Cd (0.56). The heavy metal content in liver tissue was quantified as: Cd, UDL –526.31 µg kg−1; Cr, 168.91–22268.04 µg kg−1; Cu, UDL –69406.77 µg kg−1; Pb, 29.23–5447.76 µg kg−1; and Zn 6883.33–43955.22 µg kg−1 (Table S4; Fig. 4). Following the mean concentration, the elements in the liver were sequenced from highest to lowest as Cu (25787.12) > Zn (15507.32) > Cr (5043.05 > Pb (938.02) > Cd (95.77).

Fig. 3
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Heavy metal concentrations in the muscle tissue of Anatolian water frogs collected from 11 different stations across Türkiye. Circles represent the results for each frog. Red squares and error bars indicate mean and std error, respectively

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Fig. 4
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Heavy metal concentrations in the liver tissue of Anatolian water frogs collected from 11 different stations across Türkiye. Circles represent the results for each frog. Red squares and error bars indicate mean and std error, respectively

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Concentrations of metals in water samples varied (min–max) within following limits (mg L−1): As, 0.18–9.39 µg kg−1; Cd, 0.01–0.05 µg kg−1; Cr, 0.15–1.26 µg kg−1; Cu, 0.99–3.13 µg kg−1; Pb, 0.01–5.88 µg kg−1, and Zn 1.48–246.92 µg kg−1 (Fig. 5). Heavy metal concentrations in sediments are listed as follows (µg kg−1): As, 478.74–8251.53; Cd, 78.01–549.90; Cr, 3420.81–189504.74; Cu, 1852.78–45617.42; Pb, 6182.68–27713.32; Zn, 7198.64–49863.42 (Fig. 6). Based on average concentration, heavy metals in water and sediment were ranked as follows (µg L−1 and µg kg−1): Zn (33.77) > As (2.58) > Cu (1.92) > Pb (1.82) > Cr (0.39) > Cd (0.02) and Cr (68867) > Zn (34544) > Cu (22821) > Pb (15545) > As (3700) > Cd (279), respectively.

Fig. 5
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Heavy metal concentrations in water samples collected from 11 different stations across Türkiye

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Fig. 6
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Heavy metal concentrations in sediment samples collected from 11 different stations across Türkiye

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Data analysis reveals that the aquatic environment has the lowest levels of heavy metals. Despite the low elemental concentration in water, it is common to find relatively significant quantities of several heavy metals in the tissues of frogs (Stolyar et al. 2008; Borković-Mitić et al. 2016; Prokić et al. 2016; Mani et al. 2022). Our findings align with previous research in this field. An analysis of water samples indicated heavy metal concentrations that were up to 15,000 times smaller than the amount measured in edible tissues of frogs, with readings in liver tissue being up to 30,000 times lower. Accordingly, Stolyar et al. (2008) conducted a study that compared the concentrations of metals in two P. ridibundus populations in western Ukraine, with the distinction between rural and urban sites. Despite the low metal amounts in the water samples, they detected higher metal concentrations in the frog tissues they examined (Stolyar et al. 2008). The variation in metal levels between water and frog tissues could be ascribed to several factors. One possible reason is that frogs may accumulate heavy metals from anthropogenic or natural sources, or have experienced intermittent exposure to cumulative metal contamination in the past (Papadimitriou and Loumbourdis 2002; Othman et al. 2009). Apart from directly absorbing heavy metals through their skin from water, frogs can also accumulate these metals in their bodies by consuming contaminated food or accidentally ingesting polluted sediment (Papadimitriou and Loumbourdis 2002). For this reason, metals accumulate much more in frog tissues over time than in the environment in which frogs live. Furthermore, sediment samples in this study were also found to have higher metal concentrations than water and frog samples. The findings of the current survey revealed increased metal concentrations in the sediment samples in comparison to both the water and frog samples. Because it is known that sediments act as a reservoir for pollutants, this is an expected result. Similar conclusions were reached in a recent study conducted in Nigeria, which discovered that sediment samples contained higher quantities of heavy metals than frog samples (Ediagbonya et al. 2022). However, Nummelin et al. (2007) suggested that frogs tend to accumulate metals in their tissues to much higher degrees than in the sediment, mainly through the consumption of aquatic plants or other creatures in the wetlands' food web.

In the genus Pelophylax, different frog species exhibit significant variations in mean Cr, As, and Cd concentrations in their muscle tissues, as determined by a One-Way ANOVA (Cr: F2.30 = 4.317, p < 0.05; As: F2.30 = 5.529, p < 0.01; Cd: F2.30 = 6.771, p < 0.01). Conversely, the mean Cu, Zn, and Pb concentrations among the frog species were found to be similar (One-Way ANOVA; Cu: F2.30 = 1.654, p = 0.208; Zn: F2.30 = 2.401, p = 0.108; Pb: F2.30 = 0.076, p = 0.927). The results of the statistical analysis to determine whether there was any difference in the mean heavy metal concentrations of male and female muscle tissue showed that Cu (t-test; t = 2.690, df = 31, p < 0.05) and Cd (t-test; t = 2.462, df = 31, p < 0.05) were higher in males and As (t-test; t = -2.443, df = 31, p < 0.05) was higher in females. On the other hand, in the liver tissue, no difference was found between the heavy metal concentrations of male and female individuals, except for Pb (t-test; t = 3.131, df = 23, p < 0.01). In addition, it was found that the mean Cr, As, and Cd (One Way Anova; Cr: F2,30 = 4.317, p < 0.05; As: F2,30 = 5.529, p < 0.01; Cd: F2,30 = 6.771, p < 0.01) concentrations in the muscle tissue of the species in the same genus were different from each other, and the mean Cu, Zn, and Pb (One Way Anova; Cu: F2,30 = 1.654, p = 0.208; Zn: F2,30 = 2.401, p = 0.108; Pb: F2,30 = 0.076, p = 0.927) values were similar. Metal concentrations of the three species were found to be similar in liver tissue, except for Pb (One Way Anova; F2,22 = 11.654, p < 0.001). The findings of the study conducted in Nigeria indicated that Zn and Cd contents were higher in male frogs, while Cr, Cu, and Pb values were higher in female frogs (Ediagbonya et al. 2022). The precise reason for this contrast remains unknown; however, researchers have proposed that it might be linked to genetic differences between the sexes.

The inability of organisms to effectively eliminate toxic elements from their bodies leads to their buildup in tissues, leading to bioaccumulation. This process continues up the food chain, resulting in biomagnification (Burger and Snodgrass 2001; Stolyar et al. 2008; Mani et al. 2022). The concentrations of Cu and Zn in the sediment, water, and frog tissues were notably higher than other metals, likely due to their abundance in the earth's crust and extensive industrial use (Hoffman et al. 2003).

Amphibians occupy both aquatic and terrestrial ecosystems, with water playing a vital role in supporting their lives for the majority of their lifespan (Altunışık 2018; Altunışık and Kara 2021; Tatlı et al. 2022a, b). Their dependence on water during a significant part of their life makes them susceptible to potential toxic element concentrations in aquatic environments (Wells 2007; Mani et al. 2022). The statistical analysis indicates noteworthy discrepancies in muscles’ heavy metal levels among sampling locations (ANOVA, post hoc Tukey; Fig. 3). It is possible that the deviations in metal concentrations at the sampling sites are due to the extent of contamination in the development habitats. Typically, the metal concentrations in frog tissues collected from different sites mirror the pollution levels in the area (Hodkinson and Jackson 2005; Vershinin 2007). In light of this information, the variations in heavy metal levels in water and sediment samples, as shown in Fig. 5 and 6, appear to be of significant concern. Considering frogs' close interaction with their surroundings and their role as reliable bioindicators of pollution (Othman et al. 2009; Şişman et al. 2021), variations in metal concentrations in their tissues can serve as important indicators of overall environmental quality.

Another influencing factor is the variation in frog sizes across different sampling locations, which depends on certain variables. To investigate this, a linear regression analysis was performed to explore the relationship between frog length/weight and metal levels in the liver and hind limb muscles (Fig. 7). The data indicated negative correlations between frog length/weight and Cd, Cr, Cu, Pb, and Zn concentrations in hind limb muscles, with a positive correlation observed between length–weight and As concentrations (Cd: r = -0.725, p < 0.001; Cr: r = -0.359, p < 0.05; Cu: r = -0.490, p < 0.01; Pb: r = -0.520, p < 0.01; Zn: r = -0.526, p < 0.01; As: r = 0.269, p = 0.13). Furthermore, negative correlations were found between length/weight and Cr and Pb levels in liver tissue. However, positive correlations were observed between length/weight and Cu and Zn concentrations (Fig. 7). Nevertheless, these correlations lacked statistical significance (p > 0.05), except for Cr (r = -0.527, p < 0.01). Similar to our findings, a previous study reported a negative correlation between frog length–weight and As, Cd and Pb concentrations in muscle tissues, and a positive correlation between length–weight and Cr, Cu and Zn concentrations (Mani et al. 2022). In addition, the study found that there was a negative correlation between As, Cd, Pb, Cr, and Zn concentrations and the length–weight of frog’s liver tissue (Mani et al. 2022).

Fig. 7
figure 7

The relationship between body size and mass and the heavy metal concentrations in the tissues of Anatolian water frogs collected from 11 different stations across Türkiye. Results were obtained using linear regression analysis. Each dot shows the result of different samples. X-axis shows length or weight of frog; y-axis shows concentration (µg kg−1)

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Tables 1 and 2 show the interrelationship between metals in liver and muscle tissues, respectively. In muscle tissue, there was a positive significant correlation between Cr-Zn and Cu–Zn (Table 1). As has a non-significant positive relationship with Cd, Pb and Zn, and a non-significant negative relationship with Cr and Cu. Cd has a non-significant positive correlation with Cr, Cu, Pb and Zn. Cr has an insignificant positive relationship with Cu and Pb. Cu has an insignificant positive correlation with Pb. Pb Cu has a non-significant positive correlation with Zn (Table 1). On the other hand in liver tissue, Cu has a significant positive relationship with Cr and a significant negative relationship with Pb (Table 2). Relationship between Cr-Pb, Cr-Zn, Cu–Zn and Pb–Zn was insignificant (Table 2). Similarly, a significant positive correlation was reported between Cu-Cr and Cu-Pb in the frog Pyxicephalus edulis (Ediagbonya et al. 2022). Metals with strong correlations point to comparable sources of contamination.

Table 1 The relationship among heavy metal concentrations detected in frog muscle tissue
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Table 2 The relationship among heavy metal concentrations detected in frog liver tissue
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Concentrations of heavy metals in the frog's muscle and liver tissues have been documented in various research studies, as presented in Table S4. An investigation in Northern Greece (Loumbourdis and Wray 1998) reported higher levels of Cd, Cr, Cu, Pb, and Zn in both liver and muscle tissues compared to our study. A further study conducted in the vicinity of an e- waste dump has found that the content of As, Cu, Pb and Zn in the leg muscle tissues of three diverse species (Hoplobatrachus rugulosus, Fejervarya limnocharis and Occidozyga lima) was higher than in this study (Thanomsangad et al. 2020). In addition, X. laevis and R. esculentus in Nigeria (Tyokumbur and Okorie 2011) and R. tigrina and E. cyanophlyctis species in Pakistan (Qureshi et al. 2015) had relatively high content of Cd, Cu, Cr, Zn, and Pb in liver and muscle tissues. On the other hand, the results of our study were similar in terms of many metals such as As, Cd, Cu, Pb, and Zn when compared with studies conducted in Bulgaria (Zhelev et al. 2020) and northeastern Türkiye (Mani et al. 2022).

Turkey plays an important role as a supplier and trader of frogs (Çiçek et al. 2020). For over four decades, Anatolian water frogs (Pelophylax spp.) have been gathered for consumption either from nature or through aquaculture (Akin and Bilgin 2010, Kürüm 2015). According to the data of the Turkish Statistical Institute, in 2022, 210 tons of frog legs were obtained by hunting and 25 tons by aquaculture (TUIK 2023). Various agencies have established guidelines to ensure that food organisms do not contain excessive levels of certain trace elements to ensure consumer safety. Some trace elements, including Cu, Co, Cr, Zn, V, Mn, and Ni play crucial roles in the organisms' metabolism, but they can become toxic at high doses. Additionally, Pb, Cd, and As can have damaging impacts on humans even when present in minuscule concentrations, earning them a spot in the top 10 of the "Substance Priority List" released by ASTDR (2019). Several global entities have also established the uppermost tolerable concentrations for Pb, Cd, and As. For instance, the European Commission (EC) has issued guidelines stating that the concentrations of Cd and Pb in fishery products should not exceed 1 mg kg−1 and 1.5 mg kg−1, respectively (EC 2006). Our data, compared to these limits (Table 3), showed that the Cd and Pb values remained within the specified thresholds. To determine the Estimated Daily Intake (EDI) levels for average serving size (227 g), we used the measured values of Pb, Cd, Cu, Zn, and As. Additionally, we assessed the EDI levels based on the Joint FAO/WHO Expert Committee on Food Additives (JECFA) Provisional Tolerable Daily Intake (PTDI) norms (Table 3). Remarkably, the calculated EDI rates for adults were below the JECFA thresholds (Table 3).

Table 3 Estimated Daily Intake (EDIs), Target Hazard Quotients (THQs), and Hazard Index (HI) through frog consumption from Türkiye
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To determine the potential non-carcinogenic health hazard associated with the consumption of the hind leg muscles of frogs gathered from 11 different sites, THQ values were calculated and presented in Table 3. These THQ values represent the risk associated with each potentially toxic element in the frogs. The results indicate that the regular portion size of frog consumption is not expected to be detrimental to adult consumers since all THQ values for the elements were less than 1. Additionally, we examined the HI levels, which represent the cumulative non-carcinogenic risk from multiple elements. All HI values lower than 1 (the limit value) were observed to range from 0.019 to 0.169, depending upon the mean portion size. This suggests that the consumption of frogs is not expected to impact human health negatively.

Conclusion

In this study, heavy metal bioaccumulation in the tissues and habitats of three commercially collected and exported edible frog species (Pelophylax ridibundus, Pelophylax bedriagae, Pelophylax caralitanus) in Türkiye was meticulously investigated. Additionally, the research aimed to assess the potential non-carcinogenic health risk associated with consuming the hind leg muscles of these frog species. The results revealed considerable variations in heavy metal concentrations among frog tissues, influenced by the sampling sites and species. Among the three edible species, P.ridibundus and its habitats have relatively higher heavy metal concentrations than other species. However, a reassuring outcome was the discovery that the concentrations of Cd and Pb in the edible tissues were well below the permissible limits set by the European Commission (EC). The Estimated Daily Intake (EDI) values, calculated based on the average serving size, were also lower than the Provisional Tolerable Daily Intake (PTDI) levels for adult consumers. Furthermore, the study computed the Target Hazard Quotient (THQ) values for heavy metals, all of which were below the critical value of 1, indicating that consuming the hind leg muscles from these frog species would not pose an adverse health risk for humans.