Automated Simultaneous Analysis of Monomethyl and Mercuric Hg in Biotic Samples by Hg-Thiourea Complex Liquid Chromatography Following Acidic Thiourea Leaching

A simple leaching procedure has been validated for quantitative isolation of both monomethyl (CH₃Hg⁺) and inorganic (Hg^II) mercury from fresh or dried biotic tissue for simultaneous analysis via separation and quantification with Hg-thiourea complex liquid chromatography cold vapor atomic fluorescence spectrometry (HgTU/LC-CVAFS). The leaching solution comprises thiourea, hydrochloric acid, and glacial acetic acid and works by protonating thiol binding sites and forming water-soluble cationic CH₃HgSC(NH₂)₂⁺ and Hg[SC(NH₂)₂]₂²⁺ complexes, which are easily separated from the solid matrix. The isolated complexes are preconcentrated online by either thiol resin trapping or a new iodide-complex polydivinylbenzene resin trapping (I-PDVB). The I-PDVB trapping involves only one reagent addition, requires no pH adjustments, and is quantitative over a large range of volumes and flow rates. The chromatography system can use either ion chromatography or a new ion-pairing reversed phase separation coupled to cold vapor generation and atomic fluorescence detection. The system allows quantitative sample introduction and yields absolute detection limits of 0.4 pg and 0.7 pg, for CH₃Hg⁺ and Hg^II respectively, enabling relative detection limits as low as 4 and 7 pg g⁻¹ with 100 mg samples, and yields %CV routinely less than 5% with well homogenized samples. Accuracy for both forms of mercury has been validated with multiple biotic reference materials and by comparison of the sum of CH₃Hg⁺ and Hg^II with total Hg on a variety of different biotic sample types (n = 49). The system can be calibrated with either aqueous standards or leached reference materials.