Fig. 2.

Elution profile of Se in the assay mixture. The assay mixture consisted of 7 mM GSH, 1 mM SAM as the methyl group donor, and 1 μM iSe^IV in 20 mM sodium phosphate buffer in the presence (B) or absence (A) of 30 μg of rhAS3MT. The assay mixture was incubated at 37 °C for 4 h, and then heated at 95 °C for 5 min to terminate the reaction. H₂O₂ was added to the mixture at the final concentration of 3%, and further incubation was carried out for 1 h. Then, 50-fold diluted catalase was added to eliminate excess H₂O₂ from the mixture. A 50 μL aliquot of the filtrate was applied to GS-320HQ, a multi-mode size exclusion column. The eluate was directly introduced to an ICP–MS to detect Se at m/z 82. Se standards, such as iSe^IV, iSe^VI, and TMSe without treatment with H₂O₂, were also analyzed (C).