Speciation of different forms of Se in Bovine blood by enzymatic extraction and HPLC/photo-reduction/HG-ICP-OES

Abstract

Selenium is essential for the vita! functions of the human and animal organism, such as growth, reproduction, prevention, and the protection of its muscular integrity. Although this dependence is well characterized, aspects related to selenium speci- ation still need to be studied further. This work describes the development of an alternative method for selenium spcciation in bovine blood samples using a hyphenated analytical system (HPLC-UY-HG-ICP-OES). The influence of the main parameters related to the detection of Se species, such as separation efficiency, photo-reduction and hydride generation, were studied. Enzymatic reaction with proteinase-K (1 day at 37°C) was employed for sample preparation. An on-line UV reduction system was adopted to convert the different species of Se to Se(IV). For the chromatographic system, equipped with an anionic exchange column, a phosphate- buffer of 40 mmol L-1 in the pH of 6.0 was used as the mobile phase (flow rate of 1 mL min-1) for separation of the different Se species. To increase the sensitivity, hydride generation was used, followed by inductively coupled plasma optical emission spectrometry (ICPOES) analysis. The developed method for Se speciation in bovine blood samples was performed completely on-line. The limits of detection (LODs) were 9.5, 7.2, 5.8, 3.1, 4.4 μg L-1 and the limits of quantification (LOQs) were 31.4, 23.9, 20.1, 10.5, and 14.5 μg L-1 for selenomcthylselenocysteine, seleno-DL-methionine, Se(TV), and Se(VI), respectively. The recoveries obtained were 96% for Se(IV), 98% for Se(Vl), 91% for seleno- cystamine, 89% for selenocysteinc, and 84% for seleno-mcthionine.