Speciation of selenoamino acids, selenonium ions and inorganic selenium by ion exchange HPLC with mass spectrometric detection and its application to yeast and algae

Cation and anion exchange HPLC were used to separate a mixture of 12 selenium species comprising selenoamino acids, selenonium ions and inorganic selenium. The cationic species were separated from each other and from the co-injected anions using a cation exchange column with gradient elution by aqueous pyridinium formate at pH similar to 3 as the mobile phase. The anionic species were separated using an anion exchange column with isocratic elution by an aqueous salicylate-TRIS mobile phase at pH 8.5. The separated selenium species were detected as Se-80 by ICP-dynamic reaction cell (DRC)-MS. The analytical methods were applied to the analysis of yeast and algae enriched in selenium. The yeast was treated with beta -glucosidase followed by a protease mixture for dissolution of the cell walls and selenium-containing peptides, respectively. The second to largest HPLC peak after that corresponding to selenomethionine was ascribed to selenomethionine-Se-oxide (SeOMet) by retention time matching with a SeOMet standard, which was characterised by HPLC-electrospray (ES)-MS. Se-methylselenocysteine was detected based on co-chromatography with the standard substance spiked to the yeast hydrolysate. A trichloroacetic acid extract of Chlorella algae contained dimethylselenonium propionate (DMSeP), which was verified by HPLC-ES-MS. Se-allylselenocysteine and selenoethionine was detected at the low ng g(-1) concentration level based on co-chromatography with the standard substances spiked to the algal extract.