Sample preparation for identification of selenocompounds in urine by electrospray-MS/MS

Abstract

The speciation of selenium in urine stands as a challenge to analytical chemists for two main reasons: the low physiological concentration of selenium in urine and the lack of commercially available Se-species standards for comparisons. Therefore, techniques giving direct access to structural information of the selenocompounds formed must be used to identify the chemical nature of urine Se species and sample preparation is critical for that purpose. Two sample preparation procedures to analyse selenocompounds in urine by electrospray-MS/MS have been developed here. The proposed sample treatments allow the purification of selenocompounds and consist of removal of the saline interfering matrix by different approaches followed by preparative size exclusion chromatography and final reversed-phase HPLC separation. Preconcentration of selenocompounds was achieved by several freeze-drying steps. One of the developed procedures was successfully tested in human urine spiked with selenocompounds but it was found to be unsuitable for basal samples. An alternative treatment is described that allows the detection of the main Se metabolite reported in urine samples, Se-methyl-N-acetylselenohexosamine, in rats under supplementation with selenomethionine.