Speciation of selenoamino acids, selenonium ions and inorganic selenium byion exchange HPLC with mass spectrometric detection and its application toyeast and algae

Cation and anion exchange HPLC were used to separate a mixture of 12 seleni um species comprising selenoamino acids, selenonium ions and inorganic sele nium. The cationic species were separated from each other and from the co-i njected anions using a cation exchange column with gradient elution by aque ous pyridinium formate at pH similar to 3 as the mobile phase. The anionic species were separated using an anion exchange column with isocratic elutio n by an aqueous salicylate-TRIS mobile phase at pH 8.5. The separated selen ium species were detected as Se-80 by ICP-dynamic reaction cell (DRC)-MS. T he analytical methods were applied to the analysis of yeast and algae enric hed 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 corresp onding to selenomethionine was ascribed to selenomethionine-Se-oxide (SeOMe t) by retention time matching with a SeOMet standard, which was characteris ed by HPLC-electrospray (ES)-MS. Se-methylselenocysteine was detected based on co-chromatography with the standard substance spiked to the yeast hydro lysate. A trichloroacetic acid extract of Chlorella algae contained dimethy lselenonium propionate (DMSeP), which was verified by HPLC-ES-MS. Se-allyls elenocysteine and selenoethionine was detected at the low ng g(-1) concentr ation level based on co-chromatography with the standard substances spiked to the algal extract.