Column-switching system for selenium speciation by coupling reversed-phase and ion-exchange high-performance liquid chromatography with microwave-assisted digestion–hydride generation–atomic fluorescence spectrometry

Speciation of selenocysteine (SeCys), selenomethionine (SeMet), selenoethionine (SeET), selenite (Se(IV)) and selenate (Se(VI)) has been accomplished using high-performance liquid chromatography, with the aid of an anion exchange column and a reversed-phase column, both connected through a six-port switching valve. On-line microwave-assisted digestion and hydride generation steps were performed prior to the atomic fluorescence detection. The elution of the seleno amino acids was accomplished in the reversed-phased column using water as mobile phase. Selenite and selenate were separated in the anion exchange column, using gradient elution with an acetate buffer. The separation of the five selenium compounds took place in 15 min. The detection limits obtained ranged between 0.6 and 0.9 μg l−1. Values of r>0.998 were obtained for linear fit graphs. A commercial available urine sample was analyzed, in which SeCys and Se(IV) were quantified.