Baseline activity concentration of ²¹⁰Po and ²¹⁰Pb and dose assessment in bivalve molluscs at the Andalusian coast

Baseline activity concentration of 210Po and 210Pb and dose assessment in bivalve molluscs at the Andalusian coast. Authors: Santiago Hurtado-Bermúdez* (1), José Antonio Jurado-González (3), Juan Luis Santos (2), Carlos Francisco Díaz-Amigo (3), Irene Aparicio (2), José Luis Mas (1, 4), Esteban Alonso (1, 2) (1) Servicio de Radioisótopos, Centro de Investigación, Tecnología e Innovación, Universidad de Sevilla, Av. Reina Mercedes 4B, 41012 Sevilla, Spain (2) Departamento de Química Analítica, Escuela Politécnica Superior, Universidad de Sevilla, C/ Virgen de África 7, 41011 Sevilla, Spain (3) Laboratorio de Control de Calidad de los Recursos Pesqueros, Agencia de Gestión Agraria y Pesquera de Andalucía, Ctra. Punta Umbría-Cartaya, km 12., 21459, El Rompido (Huelva), Spain (4) Departamento de Física Aplicada I, Escuela Universitaria Politécnica, Universidad de Sevilla, Spain Highlights • The first comprehensive study related to radioactivity concentrations in the area. • Assessment of 210Pb and 210Po in different types of bivalve molluscs. • The results have been compared with previous studies from other countries. • The effective dose assessment to the adult population was performed. Abstract In this study, the activity concentrations of 210Po and 210Pb were determined in different types of bivalve molluscs sampled during the period of May 2014–June 2015 along the Andalusian littoral. Radioactivity concentrations of 210Po were determined through alphaparticle spectrometry using 209Po as an internal tracer. Radioactivity concentrations of 210Pb were determined through low-level gamma-ray spectrometry. The activity concentrations of 210Po and 210Pb varied between 40 ± 2 and 515 ± 9 Bq kg−1 dry weight (d.w.), and ND (lower than limit of detection) and 73 ± 10 Bq kg−1 d.w., respectively. The committed effective dose to humans was calculated to range from 39–477 μSv year−1. Radioactivity and dose levels were compared with previous studies from other countries. Keywords: Baseline assessment; bivalve molluscs; 210Po; 210Pb ; Andalusia; dose There are two main sources of radioactivity in the marine environment (Shannon and Cherry, 1967): 40K and natural decay series of 238U and 232Th; and artificial radionuclides such as 137Cs, (239+240)Pu or 90Sr. These radionuclides are chemical analog of metabolically essential elements, and are finally bioaccumulated in hard and soft tissues of the marine organisms (Carvalho, 2018; KIlIç and Çotuk, 2011). Consequently, the radionuclides get into the human food chain via consumption of marine food. An assessment of radionuclide levels in marine organisms and radiation doses is therefore essential to evaluate the intake of radionuclides by man (Feroz Khan et al., 2014). The release of artificial radionuclides into the environment is mainly due to nuclear power plant (NPP) accidents (Chernobyl in 1986, or Fukushima in 2011), and nuclear weapon tests (during the 1960s), and therefore the Earth becomes globally contaminated with radioactive debris (Fathivand and Amidi, 2007). Several studies revealed that the contribution of anthropogenic radionuclides to the radiation dose is lower than the naturally occurring ones (Aarkrog et al., 1997; Connan et al., 2007). Therefore, we focused on the determination of natural radionuclides levels in marine organisms. Among the natural radionuclides 210Pb, and 210Po are members of 238U decay chain. The sources of 210Po (T1/2 = 138 days) and 210Pb (T1/2 = 22.3 years) in the environment are the emanation of 222Rn from the earth’s crust to the atmosphere and their subsequent fallout through wet and dry deposition (Baskaran, 2011), the radioactive decay of 226Ra dissolved in seawater (Bacon et al., 1976; Carvalho and Fowler, 1993), and also the technologically enhanced natural occurring radioactive materials (TENORM) from phosphate, oil, and gas industries (Carvalho et al., 2010; Shakhashiro et al., 2011; Villa et al., 2009). 210Po is a hazardous element with high chemical and radiological toxicity connected with the fact that the path length of alpha particles of 5.3 MeV in tissue is about 40 mm, releasing very locally the radiation dose, and potentially originating a fatal damage to different organs and tissues (Stewart et al., 2008). Therefore the major contributor to radiation dose received by humans is from 210Po in sea food, such as fish, crustaceans and molluscs (Dahlgaard, 1996; Wildgust et al., 2000). Furthermore, 210Pb is a beta emitter and also it is the second highest radiotoxic radionuclide in 238U decay chain (Štrok and Smodiš, 2011). Bivalve molluscs have been commonly used as first-order biological indicators of the contaminant levels, thus they are helpful for assessing the contamination and ecotoxicology in the marine environment (Carvalho et al., 2010; Fonollosa et al., 2017; Francioni et al., 2007; Kiliç et al., 2014; Rožmarić et al., 2012; Thébault et al., 2008; Topcuoǧlu et al., 2003; Uǧur et al., 2011). With all of this in mind, the main objectives of this study are (1) to assess the activity concentration of 210Po and 210Pb on various bivalve species distributed and consumed in the Andalusian coast, and (2) to estimate the annual effective ingestion dose to which the public is exposed in terms of health and safety. The littoral of Andalusia extends along the Mediterranean Sea, the Gibraltar Strait, and the Atlantic Ocean (Fig. 1) and it corresponds to 17.5% of the 1,100 km of coasts in Spain. Five of the 10 large urban areas in Andalusia are located in coastal areas, in addition to other systems along the coast, such as thermal power plants, ports or chemical industries. Fig. 1. Map showing the study area and sampling stations. The bivalves molluscs samples were collected from 14 locations of the Andalusian coast in May 2014 and June 2015 (see Fig. 1). In particular, different bivalves that are usually consumed in this area were collected: cockles (Cerastoderma edule) with 2.3–3.1 cm shell length, mussels (Mytilus galloprovincialis), wedge clams (Donax trunculus), peppery furrow shells (Scrobicularia plana), striped venus clams (Chamelea gallina), warty venus clams (Venus verrucosa), and grooved razor shells (Solen marginatus). In the laboratory the bivalves molluscs samples were washed with distilled water in order to eliminate any sediment residues and other impurities. However, the digestive track was not depurated as it was considered as edible material. Then, their soft tissues, including interstitial fluid, were dissected to remove the flesh using stainless steel scalpel blades to prevent contamination of samples, weighted, frozen at -20 ºC and freeze-dried for 24 h, using a freeze-dryer module; finally the dehydrated sample was weighed, crushed and homogenized. In order to determine 210Po activity concentration, a known activity of 209Po was added to the lyophilized sample to calculate the chemical recovery. Then, the sample was digested with concentrated HNO3 and H2O2, and if foam was generated during this process, the solution was cooled to a lower temperature. The obtained solution was evaporated slowly to near dryness on a hot plate at 80-90 ºC, and the resulting residue was treated again 2 or 3 times with concentrated HNO3 and H2O2 until all the organic material was digested. Finally, the residue was dissolved in 100 ml of 1 M HCl and the solution was filtered using a 0.45 µm filter. About 0.1 g of ascorbic acid was added as a reducing agent to this HCl solution. The solution was stirred continuously for 4 h at 85-90 ºC and pH 1.5–2.0 in contact with a silver disc, and then 210Po was spontaneously plated onto the disc. The disc was removed and washed with distilled water and acetone, and dried under an infrared lamp. It was then counted using an alpha-spectrometry instrument (Alpha Analyst, Canberra) containing a Passivated Implanted Planar Silicon (PIPS) detector inside. High chemical yields (>70%) were achieved (Hurtado-Bermudez et al., 2017). To measure 210Pb activity concentration, non-destructive gamma-ray spectrometry technique was used. Lyophilized samples were weighted and put into 80 ml sample containers, sealed, and 210Pb activity was determined directly by its gamma emission at 46.5 keV. The samples were measured using a low-background Canberra HPGe reverse electrode coaxial detector, and the efficiency was determined using LabSOCS software (Hurtado and Villa, 2010). Each sample was counted for 1–3 days, and activity values were reported as the activity on the date of sampling. Uncertainties reflect one sigma counting statistics. Both methods were validated in terms of the quantification limit (LQ), defined as (Currie, 1968): 𝐿𝐿𝑄𝑄=50�1+�1+𝑛𝑛0/12.5� 𝜀𝜀∙𝑡𝑡0∙𝑚𝑚∙𝑅𝑅𝑐𝑐 (1) where n0 is the total background counts; t0 the background counting time (in seconds); ε is the counting efficiency; Rc the radiochemical yield (100% for gamma-ray spectrometry); and m is the mass of the sample. The result was 2.0 Bq kg-1 for 210Po assuming 1 g of sample, and 16.0 Bq kg-1 for 210Pb assuming 15 g of sample. Additionally, the proposed methods were validated by the analysis of Certified Reference Materials (CRM) provided by the International Atomic Energy Agency (IAEA). IAEA- 437 is a reference material designed for the determination of anthropogenic and natural radionuclides in mussel (Mytilus galloprovincialis species) including 210Pb/210Po in equilibrium (median value of 4.2 Bq kg-1). IAEA-414, a mixed fish species from eastern Irish Sea, is a reference material certified for 210Pb/210Po in equilibrium (median value of 2.1 Bq kg-1). Following the IUPAC (Thompson et al., 2006) and ISO (ISO/IEC, 2010) recommendations for assessment of performance of laboratories, a z-score methodology was used in the data evaluation. The z-score was calculated as following: 𝑧𝑧=𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝐿𝐿𝐿𝐿𝐿𝐿−𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑅𝑅𝑅𝑅𝑅𝑅 𝜎𝜎𝑅𝑅𝑅𝑅𝑅𝑅 (2) where ValueREF is the reference value, ValueLAB is our laboratory value, and σREF is the uncertainty of the sample published by IAEA. The performance in term of accuracy is evaluated as satisfactory if |z-score| ≤ 2, questionable for 2 ≤ |z-score| ≤ 3 and unsatisfactory for |z-score| ≥ 3. The value of the parameter u (trueness) includes the uncertainty of the assigned value (σLAB) for bias, and it was calculated from the following equation: 𝑢𝑢=|𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝐿𝐿𝐿𝐿𝐿𝐿−𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑅𝑅𝑅𝑅𝑅𝑅| �𝜎𝜎𝐿𝐿𝐿𝐿𝐿𝐿 2+𝜎𝜎𝑅𝑅𝑅𝑅𝑅𝑅 2 (3) The calculated u-test value is compared with the critical values listed in the t-statistic tables to determine if the reported result differs significantly from the expected value at a given level of probability. The advantage of u-test is that it takes into consideration the propagation of measurement uncertainties when defining the normalised error, this is especially useful when evaluating results, which may overlap with the reference interval. The limiting value for the u-test parameter was set to 2.58 for level of probability at 99% to determine if a result passes the test (u < 2.58). Finally, the results were evaluated against the acceptance criteria for trueness and precision and assigned the status “Acceptable”, “Warning” or “Not Acceptable” accordingly. The result is assigned “Acceptable” status for trueness if: |𝑉𝑉𝑉𝑉𝑉𝑉𝑢𝑢𝑉𝑉𝐿𝐿𝐿𝐿𝐿𝐿 −𝑉𝑉𝑉𝑉𝑉𝑉𝑢𝑢𝑉𝑉𝑅𝑅𝑅𝑅𝑅𝑅|≤2.58 ∙�𝜎𝜎𝐿𝐿𝐿𝐿𝐿𝐿 2+𝜎𝜎𝑅𝑅𝑅𝑅𝑅𝑅 2 (4) And additionally the result is assigned “Acceptable” status for precision if: 𝑃𝑃=��𝜎𝜎𝐿𝐿𝐿𝐿𝐿𝐿 𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝐿𝐿𝐿𝐿𝐿𝐿�2+�𝜎𝜎𝑅𝑅𝑅𝑅𝑅𝑅 𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑉𝑅𝑅𝑅𝑅𝑅𝑅�2𝑥𝑥100% (5) Applying the above reported equation the result is assigned “Acceptable” status if P is < 20% for alpha-particle spectrometry, and P is < 30% for gamma-ray spectrometry due to the high uncertainties associated to this method because of low-energy and lowprobability gamma-ray emission of 210Pb. A result must obtain “Acceptable” status in both criteria to be assigned final status of “Acceptable”. In view of the above, the statistical tests were applied to the results obtained for the determination of 210Pb and 210Po in the IAEA CRMs. Five aliquots of approximately 1 g and 55 g were analyzed for the determination of 210Po and 210Pb presented in each CRM respectively, and results are shown in Table 1. The z-score and u values obtained are inside the acceptable range for trueness of the proposed method. All the results obtained an “Acceptable” status referring to precision and trueness. The analysis of the IAEA-437 samples gives a median value of 4.5 Bq kg-1 for 210Po and 4.9 Bq kg-1 for 210Pb, very close to the reference material reported value and within its 95% confidence interval. Lastly for IAEA-414 samples the obtained median value is within the reported 95% confidence interval. The observed deviations are in the order of the precision values associated to environmental materials, and overall, we conclude that the proposed method is adequate to analyse 210Po and 210Pb in seafood samples. Table 1. Results obtained for two IAEA reference materials for five aliquots (N=5). Uncertainties (σ) are expressed at k=1. ID Radionuclide ALAB AREF z-score u Trueness Precision P (%) Bq kg-1 σ (k=2) Bq kg-1 σ (k=2) IAEA-437 (mussel) 210Po 4.5 0.3 4.2 0.7 0.4 0.4 Acceptable 18 IAEA-437 (mussel) 210Pb 4.9 1.1 4.2 0.7 1 0.5 Acceptable 28 IAEA-414 (fish) 210Po 2.4 0.3 2.1 0.3 1 0.7 Acceptable 11 IAEA-414 (fish) 210Pb 2.7 0.7 2.1 0.3 2 0.8 Acceptable 29 Average, minimum and maximum activity concentrations of 210Pb and 210Po in bivalve molluscs in the sampling locations are presented for 2014 in Table 2, and for 2015 in Table 3. Table 2. Activity concentrations of radionuclides (Bq/kg d.w.) and annual ingestion dose for bivalve molluscs samples in the study area during 2014. bivalve specie 210Po 210Pb ww/dw (%) E d (µSv/year) Bq/kg uncertainty Bq/kg uncertainty S1-Río Guadiana Cerastoderma edule 73 3 <16.0 16.6 69 S2-Isla Cristina Mytilus galloprovincialis 167 6 39 8 17.1 180 S3-Barra del Terrón Donax trunculus 112 4 <16.0 18.3 115 S4-Desembocadura Río Piedras Donax trunculus 46 2 <16.0 15.5 41 S5-Punta Umbría Donax trunculus 40 2 18 13 17.4 50 S6-Caño de Sancti Petri Scrobicularia plana 48 2 <16.0 18.6 52 S7-Río Barbate Scrobicularia plana 187 8 17 6 18.7 198 S8-Getares Mytilus galloprovincialis 41 1 75 8 16.5 77 S9-Bahía de Algeciras Chamelea gallina 149 5 <16.0 19.3 158 S10-Bahía de Algeciras Venus verrucosa 179 9 26 8 20.5 220 S11-La Línea Mytilus galloprovincialis 129 5 37 12 16.1 133 S12-Puerto de Marbella Mytilus galloprovincialis 382 14 92 10 16.0 382 S13-Puerto de Benalmádena Mytilus galloprovincialis 308 18 20 5 19.1 336 S14-Caleta de Vélez Mytilus galloprovincialis 314 12 24 7 17.7 319 The activity concentrations of 210Po in bivalve molluscs samples ranged between 40 ± 2 and 383 ± 14 Bq kg−1 d.w. for 2014, and 85 ± 3 and 506 ± 9 Bq kg−1 d.w. for 2015. The highest 210Po activity concentration was found in bivalves collected from ports, Puerto de Marbella (S12) and Puerto de Benalmádena (S13), during both years. The lowest activities were determined in the samples from Punta Umbría (S5) and Caño de Sancti Petri (S6) in the Atlantic coast. As the accumulation of 210Po in marine organisms is associated with the ingestion of food, it seems that bivalves at the stations located at the ports (S12 and S13) have on the one hand more phytoplankton available as food, and on the other hand more organic particulate matter from sewage discharges accumulates inside the ports. The activity concentrations of 210Po in bivalve molluscs samples during summer 2014 and summer 2015 are shown in Fig. 2. There is an overall average increment of the activity concentrations in summer 2015. Probably, there was an annual change of some environmental parameters, such as the amount of suspended particulate matter or the volume of the existing plankton, that implies higher values for activity concentrations of 210Po in bivalve molluscs during summer 2015. Table 3. Activity concentrations of radionuclides (Bq/kg d.w.) and annual ingestion dose for bivalve molluscs samples in the study area during 2015. bivalve specie 210Po 210Pb ww/dw (%) E d (µSv/year) Bq/kg uncertainty Bq/kg uncertainty S1-Río Guadiana Solen marginatus 354 17 <16.0 19.8 385 S2-Isla Cristina Mytilus galloprovincialis 224 10 <16.0 17.0 210 S3-Barra del Terrón Donax trunculus 172 9 <16.0 17.1 163 S4-Desembocadura Río Piedras Donax trunculus 186 10 <16.0 16.7 172 S5-Punta Umbría Donax trunculus 91 3 <16.0 16.8 86 S6-Caño de Sancti Petri Scrobicularia plana 85 3 <16.0 20.9 99 S7-Río Barbate Scrobicularia plana 256 12 31 13 20.0 301 S8-Getares Mytilus galloprovincialis 436 24 37 12 16.6 417 S9-Bahía de Algeciras Chamelea gallina 390 14 <16.0 18.2 390 S10-Bahía de Algeciras Venus verrucosa 187 9 <16.0 19.9 205 S11-La Línea Mytilus galloprovincialis 397 15 34 17 18.8 429 S12-Puerto de Marbella Mytilus galloprovincialis 451 21 28 4 15.5 389 S13-Puerto de Benalmádena Mytilus galloprovincialis 506 9 30 14 16.7 479 S14-Caleta de Vélez Mytilus galloprovincialis 320 15 <16.0 16.8 295 Fig. 2. Concentrations of 210Po (Bq kg−1 dw) in bivalve molluscs sampled during summer 2014 and summer 2015 from different locations at Andalusian coast. 0 100 200 300 400 500 600 S1 S2 S3 S4 S5 S6 S7 S8 S9 S10 S11 S12 S13 S14 210Po (Bq/kg) sampling station 2014 2015 The concentration of 210Pb in bivalves ranged between ND (not detected or below Minimum Detectable Activity) and 92 ± 10 Bq kg−1 d.w. in 2014, and ND and 37 ± 12 Bq kg−1 d.w. in 2015. 210Pb activity concentrations were relatively low probably due to the fact that the sampling stations are under the influence of Mediterranean climate with cool wet winters and hot and dry summers. Because a high concentrations of 210Pb in molluscs is usually linked to high atmospheric deposition of 210Pb during the wet season, a low activity concentration of 210Pb is expected for samples collected during summer (Uǧur et al., 2011). On the other hand, 210Pb activity concentrations are higher at Mediterranean coast than at the Atlantic coast of Andalusia (see Fig. 3). Fig. 3. Concentrations of 210Pb (Bq kg−1 d.w.) in bivalve molluscs sampled during summer 2014 and summer 2015 from different locations at Andalusian coast. 210Po and 210Pb activity concentrations measured in this study are comparable with other reported values in the literature (see Table 4). In this study, the 210Po/210Pb ratio was found to range from 4 to 16 with a mean value of 8, which shows that the 210Po majority in the bivalves is unsupported. The range of 210Po/210Pb ratio is similar to those found by several studies (see Table 4). Table 4. Comparison of 210Po and 210Pb activity concentrations, 210Po/210Pb ratio, and annual ingestion dose in bivalves collected from various studies. 0 20 40 60 80 100 120 S1 S2 S3 S4 S5 S6 S7 S8 S9 S10 S11 S12 S13 S14 210Pb (Bq/kg) sampling station 2014 2015 References Country 210Po (Bq kg -1 dw) 210Pb (Bq kg -1 dw) 210Po/210Pb E d (µSv year -1 ) (McDonald et al., 1996) UK 103–3124 - - - (Uğur et al., 2002)ide Turkey 52–1344 6–167 3-25 - (Desideri et al., 2011) Italy 75–223 2–25 4-62 96–466 (Carvalho et al., 2011) Portugal 102-759 2.6-45 18-51 - (Charmasson et al., 2011) France 203 20 10-15 - (Uǧur et al., 2011) Turkey 53-1960 6-135 4-137 - (Štrok and Smodiš, 2011) Slovenia 51-106 2.7-3.0 17-111 8.5 (Rožmarić et al., 2012) Croatia 22-207 2.8-9.3 6-31 53-497 (Kiliç et al., 2014) Turkey 26-280 1-23 8-24 0.2-3.3 (Kim et al., 2017) Korea 240 - - 19-189 The activity concentrations for each species of bivalve molluscs are shown in Table 3 and 4 for both years. 210Pb was not found or the values were too low (clams) in all samples, except for mussels (M. Galloprovincialis) with 92 Bq kg−1 d.w. as the maximum value. Significant variation of 210Po activity concentrations were observed within different species of bivalves, which may be due to difference in the size, age, metabolism, feeding habit and the environment of the specific bivalves species. The highest value in average corresponds to mussels (M. Galloprovincialis) with 506 Bq kg−1 d.w., and the lowest one to cockles (D. trunculus) with 40 Bq kg−1 d.w. and peppery furrow shells (S. plana) with 48 Bq kg−1 d.w. Table 5. Activity concentrations of radionuclides (Bq/kg d.w.) and annual ingestion dose for different species of bivalve molluscs. 210Po 210Pb ww/dw (%) Ed (µSv/year) min-max average min-max average Cockles (C. Edule) 70.0-76.0 73.0 <16.0 <16.0 16.6 66.0 Razor shells (S. Marginatus) 337-371 354 <16.0 <16.0 19.8 382 Mussels (M. Galloprovincialis) 40.0-515 306 15.0-102 41.2 17.0 302 Clams (D. Trunculus) 40.0-196 108 5.00-31.0 18.0 17.0 102 Peppery furrow shells (S. Plana) 48.0-268 144 11.0-44.0 24.0 19.6 160 Clams (C. Gallina) 144-404 270 <16.0 <16.1 18.8 271 Clams (V. Verrucosa) 170-196 183 18.0-34.0 26.0 20.2 210 Annual ingestion doses due to the consumption of bivalves molluscs were calculated for adults using the following equation (IAEA, 2014): 𝐸𝐸𝑑𝑑=𝐴𝐴 × 𝑚𝑚 × 𝐶𝐶𝑓𝑓 (6) where Ed (Sv) is the annual effective ingestion dose of each radionuclide, A is the activity concentration of each radionuclide (Bq kg-1 w.w.) in the bivalve molluscs samples, m (kg) is the estimation of annual intake of each bivalve mollusc specie, and Cf is the dose coefficient for adults (Sv Bq-1). The annual intake of molluscs was obtained via the database of food consumption maintained by the Spanish Ministry of Agriculture, Food and Environment (m = 3.5 kg of bivalves of wet tissue per year per capita). The values of the dose coefficients used for 210Po and 210Pb were 1.26·10-6 and 6.90·10-7 Sv Bq-1, respectively (IAEA, 2014). The annual effective doses resulting from the internal incorporation of 210Po and 210Pb through the consumption of all of the species analyzed in this study are reported in Table 5. The obtained values were found to range between 66-382 μSv year-1. The annual ingestion doses calculated for all of the bivalve molluscs analyzed are comparable with other studies reported in the literature (see Table 4), and are within the normal range of This work Spain (Andalusia) 40-506 17-92 4-16 41-479