CHAPTER 7: Focus on Determination of Selenocysteine and Selenomethionine in Foodstuffs of Animal Origin by 2D Size-Exclusion Reversed-Phase HPLC-ICP-MS

The chapter, after summarizing the state-of-the art for selenium analysis and selenium status of different aliments, describes an analytical procedure that was developed for the quantitative recovery of selenomethionine (SeMet) and selenocysteine (SeCys) from foodstuffs of animal origin. It points out all the crucial steps of raw sample treatment and gives the details of all the steps that had to be optimized to achieve efficient protein extraction (protein unfolding/denaturation by urea) and ensure lack of losses (carbamidomethylation of the aminoacid residues using iodoacetamide) due to Se-compound degradation. The seleno-aminoacids were specifically determined by ion-paring reversed-phase HPLC-ICP MS and quantified by the method of standard addition after proteolysis followed by their isolation from the postreaction mixture by size-exclusion LC. In addition, formal identification by ESI MS/MS was carried out for compounds for which standards are not commercially available. Determination and quantification of separate selenoamino acids gives us information about specific or nonspecific incorporation of selenium into proteins.