Development of a new column switching method for simultaneous speciation of selenometabolites and selenoproteins in human serum

•Speciation of serum Se-proteins and Se-metabolites by 2D/SEC-AF-SUID-ICP-ORS-qMS.•Speciation of serum Se-proteins and Se-metabolites by 3D/SEC-AF-AEC-SUID-ICP-ORS-qMS.•The method is time-efficient and reliable using post-column SUID analysis.•A method for analysis of eGPx, selenite, selenate, SeP a...

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Published in:Journal of Chromatography A 2013-11, Vol.1318, p.171-179
Main Authors: García-Sevillano, M.A., García-Barrera, T., Gómez-Ariza, J.L.
Format: Article
Language:English
Subjects:
anion exchange
Biological and medical sciences
blood serum
Column-switching
detection limit
glutathione peroxidase
high performance liquid chromatography
Human serum
humans
Inductively coupled plasma-mass spectrometer
Investigative techniques, diagnostic techniques (general aspects)
isotope dilution technique
Isotopic dilution analysis
mass spectrometry
Medical sciences
Miscellaneous. Technology
molecular weight
Pathology. Cytology. Biochemistry. Spectrometry. Miscellaneous investigative techniques
quality control
selenium
Selenometabolites
Selenoproteins
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Summary:•Speciation of serum Se-proteins and Se-metabolites by 2D/SEC-AF-SUID-ICP-ORS-qMS.•Speciation of serum Se-proteins and Se-metabolites by 3D/SEC-AF-AEC-SUID-ICP-ORS-qMS.•The method is time-efficient and reliable using post-column SUID analysis.•A method for analysis of eGPx, selenite, selenate, SeP and SeAlb in serum is proposed. A method for the simultaneous speciation of selenoproteins and selenometabolites in human serum has been developed on the basis of in series three dimensional chromatography: size exclusion, affinity and anion exchange high performance liquid chromatography (3D/SE-AF-AEC-HPLC), using different columns of each type and hyphenation to inductively coupled plasma-(quadrupole) mass spectrometry (ICP-qMS). The method allows the quantitative simultaneous analysis of selenoprotein P (SeP), extracellular glutathione peroxidase (eGPx), selenoalbumin (SeAlb), selenite and selenate in human serum using species-unspecific isotope dilution (SUID). The 3D chromatographic separation is proposed to remove typical spectral interferences in this matrix from chloride and bromide on 77Se (40Ar37Cl), 80Se (79Br1H) and 82Se (81Br1H). In addition, a previous method based on 2D/SE-AF-HPLC is proposed as a simple alternative when low molecular mass selenium species are absent in the samples. The method is robust, reliable and fast with typical chromatographic runtime less than 35min. Detection limits are in the range of 0.2–1.3ng of Seg−1. Method accuracy for determination of total protein-bound to Se was assessed by analyzing an human serum reference material (BCR-637) certified for total Se content and method reliability checked in samples of human serum providing results in good agreement with the total selenium concentration. In addition, the application of the method to commercial human serum and plasma reference materials for quality control analysis, certified for total Se, has provided, for the first time, indicative levels of selenium containing proteins in these samples.
ISSN:0021-9673
DOI:10.1016/j.chroma.2013.10.012