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Determination of selenocysteine and selenomethionine in edible animal tissues by 2D size-exclusion reversed-phase HPLC-ICP MS following carbamidomethylation and proteolytic extraction

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A method was developed for the simultaneous determination of selenomethionine (SeMet) and selenocysteine (SeCys) in meat (chicken and lamb muscles) and different offal tissues (heart, liver, kidney). The analytical procedure was based on the protein extraction with urea under reducing conditions (dithiothreitol), derivatization of SeCys and SeMet by carbamidomethylation with iodoacetamide (IAM) followed by quantitative proteolysis. The mixture of the derivatized Se-amino acids was purified by size-exclusion liquid chromatography (LC) and analysed by ion-paring reversed-phase HPLC–inductively coupled plasma mass spectroscopy (ICP MS). The quantification of SeCys and SeMet was carried out by the method of standard additions. 77SeMet was used to control the SeMet derivatization efficiency and recovery. The method was validated by the determination of the Se mass balance. The Se-amino acids accounted for 91 ± 8% of the total selenium (mean of 95 samples of seven tissues analysed over a period of 18 months). The method was applied to the discrimination of the contribution of selenoproteins (containing SeCys) and other Se-containing proteins (containing SeMet) in tissues of animals during supplementation studies (dose–effect and tolerance).

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M.D. acknowledges a Marie Curie fellowship (MEIF-CT-2003-501297). The participation of E. Czerska, an M.Sc. student from the Warsaw University of Technology and J. Yates, an M.Sc. student from the University of Aberdeen, in the preliminary phase of this study is acknowledged. The samples used in this study for the method development came from a research funded by Alltech EU Regulatory Department (trial ref SEL/LAM/DOS/01/0405/UK and SEL/LAM/TOL/01/0205/UK, SEL/BRO/TOL/01/1002/UK) carried out at Reading University, UK.

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Correspondence to Ryszard Lobinski.

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Bierla, K., Dernovics, M., Vacchina, V. et al. Determination of selenocysteine and selenomethionine in edible animal tissues by 2D size-exclusion reversed-phase HPLC-ICP MS following carbamidomethylation and proteolytic extraction. Anal Bioanal Chem 390, 1789–1798 (2008).

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