Abstract

This study reports the development and validation of a new analytical method for simultaneous speciation analysis of Se and Hg in fish muscle. For this purpose, four Se species (selenite/Se(IV), selenate/Se(VI), selenomethionine/SeMet, and selenocysteine/SeCys) and two Hg species (inorganic mercury/iHg and methylmercury/MeHg) were extracted simultaneously by microwave-assisted enzymatic hydrolysis and then separated by HPLC in less than 15min by using a column with both anion and cation exchange mechanisms and a mobile phase consisting of a mixture of methanol 5% (v/v), 45mM HNO3, 0.015% 2-mercaptoethanol, and 1.5mM sodium 3-mercapto-1-propanesulfonate. The separated species of Hg and Se were detected online by inductively coupled plasma-mass spectrometry (ICP-MS). The speciation analysis method was validated by means of the accuracy profile approach by carrying out three series of measurements in duplicate on three different days over a time-span of 3weeks. The limits of quantification (LOQ) are in the range of 0.010-0.013mg/kg wet weight (ww) for all selenium species, except for Se(IV) (0.15mg/kg ww), while the coefficient of variation in terms of intermediate reproducibility (CVR) was < 7%. The LOQ for MeHg was 0.006mg/kg ww, while the CVR was 3%. The method was successfully applied to the analysis of muscle samples from four different fish species: rainbow trout, tuna, swordfish, and dogfish.

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