Mercury isotope ratio measurements of methylmercury in fish
A method for mercury isotope ratio measurements for methylmercury in fish based on HPLC combined with MC-ICP-MS has been developed by UK-based scientists from LGC.
Species-specific Hg isotope ratio data has increasingly become an important tool in understanding biogeochemistry of mercury. Such species-specific isotope data provides information on natural sources of CH3Hg production, enables the tracing of species pathways in the environment, and supports the assessment of the potential impact on the aquatic ecosystems. In the past, large variations in the bulk isotopic composition of Hg were observed in fish. Due to the ability of CH3Hg to bioaccumulate and biomagnify in aquatic biota, species-specific Hg isotope ratio data in fish can hopefully facilitate discriminating source relating isotopic signatures from those formed during biotransformation processes including degradation.
Analysis of mercury species is most often performed by speciation analysis using a separation technique such as chromatography, electrophoresis and others combined with element selective detection techniques such as ICP-MS. For precise and accurate Hg isotope ratio determinations only multi-collector instruments can be used. While gas chromatography coupled to ICP-MS (GC-ICP-MS) is the most sensitive technique for Hg speciation analysis, it requires derivatization to convert the different Hg species into volatile ones. Also since the Hg isotope ratio should be used to trace reactions it is not a good idea to rely on reactions for derivatization for detection.
The new study:
Scientists from LGC (UK) now developed a method based on HPLC. Especially problematic is to obtain accurate isotope ratio data from transient signals created by on-line separation techniques. Many steps of optimization were necessary to obtain the required sensitivity and accuracy. It involved developing improved, quantitative Hg species extraction incurring minimal dilution, matrix matching between the extraction and separation procedures and high capacity HPLC separation of Hg species as well as instrumental protocols to account for the aforementioned features of Hg isotopic analysis involving the use of cold vapor generation (CVG) for effective sample introduction. It turned out that offline separation produced more reliable data than on-line separation.
The analytical steps of the whole procedure are shown in figure 1.
Figure 1: Workflow of the analytical procedure for the mercury isotope ratio determination of methylmercury in fish
The chromatographic separation of Hg species was carried out under isocratic conditions using a reversed phase C18 column avoiding changing conditions that could have an influence on sensitivity. Low detection limits were obtained by using a high sample injection volume of 500 µL. Also the collected analyte fraction were preconcentrated by freeze drying and digested with a small volume (0.5 mL) of nitric acid and 0.1 ml of hydrogen peroxide. The digestion of the fractions avoided any matrix effects produced by the binding partners of mercury.
The digested fractions were measured by multi-collector ICP-MS using CVG. The authors evaluated the uncertainties of the measurements by propagating individual uncertainty components according to the ISO/GUM guide. The isotope data obtained in this study show that fractionation can occur during dissociation of CH3Hg as well as Hg° volatilisation from solution containing reduced sulphur. Lighter Hg isotopes were preferentially partitioned into reaction products in both cases.
The authors concluded that the developed method could be helpful for certifying reference materials for mercury isotopes in biological materials.
The original study:
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