Ultratrace Metal Speciation Analysis by Coupling of Sector-Field ICP-MS to High-Resolution Size Exclusion and Reversed-Phase Liquid ChromatographyClick to copy article linkArticle link copied!
- Hendrik Küpper*Hendrik Küpper*E-mail: [email protected]Department of Plant Biophysics and Biochemistry, Institute of Plant Molecular Biology, Biology Center of the Czech Academy of Sciences, Branišovská 31/1160, 370 05 České Budějovice, Czech RepublicDepartment of Experimental Plant Biology, University of South Bohemia, Branišovská 31/1160, 370 05 České Budějovice, Czech RepublicMore by Hendrik Küpper
- Syed Nadeem Hussain BokhariSyed Nadeem Hussain BokhariDepartment of Plant Biophysics and Biochemistry, Institute of Plant Molecular Biology, Biology Center of the Czech Academy of Sciences, Branišovská 31/1160, 370 05 České Budějovice, Czech RepublicMore by Syed Nadeem Hussain Bokhari
- Noelia Jaime-PérezNoelia Jaime-PérezDepartment of Plant Biophysics and Biochemistry, Institute of Plant Molecular Biology, Biology Center of the Czech Academy of Sciences, Branišovská 31/1160, 370 05 České Budějovice, Czech RepublicMore by Noelia Jaime-Pérez
- Lyudmila LyubenovaLyudmila LyubenovaDepartment of Plant Biophysics and Biochemistry, Institute of Plant Molecular Biology, Biology Center of the Czech Academy of Sciences, Branišovská 31/1160, 370 05 České Budějovice, Czech RepublicMore by Lyudmila Lyubenova
- Nermeen AshrafNermeen AshrafDepartment of Plant Biophysics and Biochemistry, Institute of Plant Molecular Biology, Biology Center of the Czech Academy of Sciences, Branišovská 31/1160, 370 05 České Budějovice, Czech RepublicDepartment of Experimental Plant Biology, University of South Bohemia, Branišovská 31/1160, 370 05 České Budějovice, Czech RepublicMore by Nermeen Ashraf
- Elisa AndresenElisa AndresenDepartment of Plant Biophysics and Biochemistry, Institute of Plant Molecular Biology, Biology Center of the Czech Academy of Sciences, Branišovská 31/1160, 370 05 České Budějovice, Czech RepublicMore by Elisa Andresen
Abstract
Techniques for metal speciation analysis with subnanomolar (ppt) detection limits in complex matrices, with simultaneous quantification of matrix elements, have become a necessity for investigating targets of trace metal binding to macromolecules and pigments at environmentally relevant concentrations. In this work we optimized the analysis of such metal binding in a custom-built HPLC-ICP-MS system. Key elements of the optimization were the choice of components for the metal-free HPLC-DAD system and sector-field ICP-MS detection (ICF-sfMS) with desolvating injection and optimization of sample handling. Protein analysis was done using ammonium bicarbonate buffer and size exclusion chromatography (SEC-ICP-sfMS), with possible addition of anion exchange chromatography. Detection of metal exchange in pigments (chlorophylls and bacteriochlorophylls) was based on reversed-phase chromatography with a methanol-acetone gradient and coupling to the ICP-sfMS via a dedicated organic matrix interface (RPC-ICP-sfMS). The resulting HPLC-DAD-ICP-sfMS system has detection limits in the picomolar range in protein buffer, limited by the maximal achievable purity of buffers/solvents and not by system sensitivity. Tests for method optimization showed that sonication, meant to increase protein solubilization, leads to artifacts of metal loss from metalloproteins. Examples for Cd binding to soybean proteins and chlorophyll, Cr binding to Arabidopsis thaliana proteins, La binding to Desmodesmus quadricauda proteins, and Cu binding to Rhodospirillum rubrum proteins and pigments are shown. These application examples demonstrate that the system is sensitive enough to detect binding of metals to proteins and pigments at background concentration levels of typical nutrient solutions made from analytical grade chemicals, equivalent to ultratrace metal concentrations in nonpolluted environments.
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