Measurement of inorganic and methylmercury in fish tissues by enzymatic hydrolysis and HPLC-ICP-MS

Abstract

The measurement of mercury(II) and methylmercury(I) in fish muscle tissues after protease type XIV extraction by HPLC-ICP-MS is described. A SphereClone 5 μm ODS2 80A PEEK column (100 mm × 4.6 mm, Phenomenex, USA) and a 5% v/v CH₃OH–water mobile phase containing 0.06 M CH₃COONH₄ and 0.1% w/v cysteine, pH 6.8 (flow rate, 1.0 ml min⁻¹; temperature, 25 °C) was used for the separation of mercury species. The addition of cysteine during incubation with protease and HPLC-ICP-MS separation was essential to eliminate adsorption, peak tailing and memory problems. The recovery of mercury from two reference materials (NRCC DORM-2 Dogfish and NIST RM 50 Albacore tuna) and sample fish tissues was 92–107%. The calibration curves for mercury(II) and methylmercury standards were linear in the range of 0.5–20 µg L⁻¹ (r² = 0.9992, r² = 0.9999, respectively). The lowest measurable mercury was 0.5 µg l⁻¹ and this corresponds to 0.025 µg g⁻¹ in fish tissues. Methylmercury measured in NIST RM 50 (0.87 ± 0.03 µg g⁻¹) was similar to that reported in the literature (0.89–0.91 µg g⁻¹). The methylmercury concentration measured in DORM 2, when results are corrected for the mercury extracted during incubation with the enzyme, was similar (4.4 ± 0.1 µg g⁻¹) to the certified methylmercury concentration (4.47 ± 0.32 µg g⁻¹). Most of the mercury in fish tissues is present as methylmercury (88–100%).