2.1. Sample collection and preparation: Fresh samples of three species included Rui (Labeo rohita),Pangas (Pangasius pangasius)and Tilapia (Oreochromis mossambi-cus)(Table S1) were collected from different markets occupying the 30 agro-ecological zones (Fig. 2andTable S2) of Bangladesh.These three fish species are highly consumed by the Bangladeshi population. Total fish individuals (muscle and head) were analyzed for cadmium (Cd), arsenic (As), lead (Pb), chromium (Cr), nickel(Ni), zinc (Zn), selenium (Se), copper (Cu), molybdenum (Mo), manganese (Mn), antimony (Sb), barium (Ba), vanadium (V), and silver (Ag). A composite sample for each species was prepared and homogenized in a food processor and 50 g test portions were stored at 20°C. Metal contents were expressed as mg kg1wetwt. of fresh fish.2.2. Analytical methods2.2.1. Reagents: All solutions were prepared with analytical reagent-grade chemicals and ultrapure water. SUPRAPURÒnitric acid (HNO3, 67% (v/v)) was purchased from Kanto Chemical Co, Japan and H2O2 were purchased from Wako Chemical Co, Japan. Standard stock solutions containing 10lgL1of each element (Cd, As, Pb, Cr, Ni, Zn, Se, Cu, Mo, Mn, Sb, Ba, V and Ag and internal standard solutions containing 1.0 mg L1of indium (In), yttrium (Y), beryllium (Be), telium(Te), cobalt (Co) and titanium (Ti) were purchased from SPEXertifi-cateÒand were used to prepare calibration and internal standards.Working standards were prepared daily in 5% (v/v) HNO3at 67%and were used without further purification. A solution of10.0 mg L1multielement solution (Merck, Darmstadt, Germany)was used to prepare a tuning solution with several elements such as indium, uranium, barium and lithium, capable of covering a wide range of masses. Ultra-pure grade carrier (Argon (Ar)) was supplied from Air Liquide (Japan).2.2.2. Sample digestion procedure: A microwave (Berghoff Microwave MWS-2, Germany) was used in the preparation of samples to analyze various metals with ICP–MS(Agilent 7700, USA). The microwave digestion system has been designed to performed chemical digestion procedures under extreme pressure and temperature conditions in chemical laboratories. Digestion reagents that were used included 5 ml HNO3acid (Kanto Chemical Co, Japan) and 2 ml H2O2(Wako Chemical Co,Japan). The weighed samples of 0.2 g were then placed into the digestion reagent in a Teflon vessel. DAP-60K type pressure vessels (Berghoff, Germany) which are made entirely of tetrafluoro-methoxylene (TFM) were used in this study. Three step-digestion procedures were followed: (1) temperature and power were maintained at 180°C and 85% respectively for 15 min; (2) temperature was kept steady at 200°C for 15 min together with 90% of the pow-er and (3) reduced temperature (100°C) and power (40%) were used for 10 min to cool down the Teflon vessels. After that, all Teflon vessels were kept in cold water to reduce the residual pressure inside the Teflon vessel. Samples were then transferred into a Teflon beaker and total volume was made up to 25 ml for water and 50 ml for sediments with MilliQ water (Elix UV5 and MilliQ, Millipore, USA). The digest solution then filtered (DISMICÒ–25HP PTTF syringe filter (pore size = 0.45 mm) Toyo Roshi Kaisha,Ltd., Japan), and stored in a crew cap plastic tube. The samples were then subjected to analysis for various trace metals using ICP–MS followed by three times digestion for each sample. Afterward, the vessels were cleaned by MilliQ water and dried with air.Finally, two blank digestions with 5 ml HNO3following the said digestion procedures were carried out to clean up the digestion vessels (Berghoff’s product user manual).2.2.3. Quality assurance: All test batches were evaluated using an internal quality approach and validated if they satisfied the defined Internal Quality Controls (IQCs). For trace metals, samples were analyzed using an inductively coupled plasma mass spectrometer (Agilent 7700 series). Multielement standard XSTC-13 (Spex CertiPrep, USA) solutions were used to prepare the calibration curve (ASEANFOODS,2011). The calibration curves withR2> 0.999 were accepted for concentration calculation. Before starting the sequence, relative SD ([RSD] <5%) was checked by using a tuning solution purchased from Agilent. Internal calibration standard solutions containing1.0 mg L1of indium, yttrium, beryllium, tellurium, cobalt, and thallium were purchased from Spex CertiPrep, USA. Multielement solution (Agilent), 1.0 mg L1, was used as a tuning solution covering a wide range of masses of elements. All test batches were evaluated using an internal quality approach and validated if they satisfied the defined internal quality controls. For each experiment, a run included blank, certified reference materials (CRM), and samples, all of which were analyzed in duplicate to eliminate any batch-specific errors. The CRM (DORM-2–dogfish muscle from the National Research Council, Canada) was analyzed to confirm analytical the performance and good precision ([RSD] <20%) of the applied method.