Exploring Intracellular Se Speciation in Marine Phytoplankton by LC-ICP-MS/MS: from Se Enrichments to Extraction and Chromatographic Techniques

Marine phytoplankton play a key role in biogeochemical selenium (Se) cycling. By transforming inorganic Se into organic species, many of which remain uncharacterized, they influence Se availability for other organisms and may contribute to Se volatilization. Characterizing organic Se species in phytoplankton is challenging and no standard analytical workflow has been established. Here, we used the marine phytoplankton species Gephyrocapsa huxleyi and Effrenium voratum to systematically assess how cultivation (Se concentrations in the medium), cell harvesting, extraction, and chromatographic parameters influence Se speciation measurements quantitatively (extraction yields, column recoveries) and qualitatively (species distribution), when performed by HPLC-ICP-MS/MS. We show that medium Se concentrations and choice of chromatography had the largest effect on measured Se speciation. The two phytoplankton showed marked differences in intracellular Se speciation, likely reflecting a dissimilar selenometabolism. Resolving these speciation differences was only possible with select chromatographies (e.g. anion-exchange but not mixed-mode). Both phytoplankton also showed changes to intracellular Se speciation when grown in Se-enriched media in comparison with near-natural concentrations. The selection of Se concentrations in the medium is therefore crucial for studying Se metabolites, e.g. under environmentally relevant conditions. We recommend extraction–chromatography workflows tailored to specific experimental needs in phytoplankton Se research, providing a framework for future studies to uncover further effects these organisms have on biogeochemical Se cycling.