Robust microwave-assisted extraction protocol for determination of total mercury and methylmercury in fish tissues

A rapid and efficient closed vessel microwave-assisted extraction (MAE) method based on acidic leaching was developed and optimized for the extraction of total mercury (Hg), inorganic mercury (Hg2+) and methylmercury (CH3Hg+) from fish tissues. The quantitative extraction of total Hg and mercury species from biological samples was achieved by using 5 mol L−1 HCl and 0.25 mol L−1 NaCl during 10 min at 60 °C. Total Hg content was determined using inductively coupled plasma mass spectrometry (ICP–MS). Mercury species were measured by liquid chromatography hyphenated with inductively coupled plasma mass spectrometry (LC–ICP–MS). The method was validated using biological certified reference materials ERM-CE464, DOLT-3, and NIST SRM-1946. The analytical results were in good agreement with the certified reference values of total Hg and CH3Hg+ at a 95% confidence level. Further, accuracy validation using speciated isotope-dilution mass spectrometry (SIDMS, as described in the EPA Method 6800) was carried out. SIDMS was also applied to study and correct for unwanted species transformation reactions during and/or after sample preparation steps. For the studied reference materials, no statistically significant transformation between mercury species was observed during the extraction and determination procedures. The proposed method was successfully applied to fish tissues with good agreement between SIDMS results and external calibration (EC) results. Interspecies transformations in fish tissues were slightly higher than certified reference materials due to differences in matrix composition. Depending on the type of fish tissue, up to 10.24% of Hg2+ was methylated and up to 1.75% of CH3Hg+ was demethylated to Hg2+.