Simultaneous speciation analysis of glutathione peroxidase, selenoprotein P and selenoalbumin in human serum by tandem anion exchange-affinity HPLC and on-line isotope dilution ICP-quadrupole MS.
A method based on anion exchange (AE) and affinity (AF)-HPLC (AE-AF-HPLC) hyphenated to inductively coupled plasma-(quadrupole) mass spectrometry (ICP-QMS) was developed for the speciation analysis of selenoprotein P (SelP), glutathione peroxidase (GPx) and selenoalbumin (SeAlb) in human serum. AE-HPLC is proposed here for the on-line alleviation of Cl and Br spectral interferences on (77)Se ((40)Ar(37)Cl) and (82)Se ((81)Br(1)H). Separation of GPx, SelP and SeAlb by AE-AF-HPLC was obtained within a total chromatographic runtime of <20 min. On-line (post-column) isotope dilution (ON-ID) and on-line external calibration (ON-EC)-ICP-QMS were used for the quantification of Se in GPx, SelP and SeAlb. ON-EC using a Se-L: -cystine standard was shown to be a suitable approach for the routine simultaneous speciation analysis of serum GPx, SelP and SeAlb. The method validation was carried out by direct ICP-sector field MS determination of Se in GPx, SelP and SeAlb fractions collected after AE-AF-HPLC separation. In addition, the method accuracy for the determination of total protein-bound Se was assessed by analyzing a human serum reference material (BCR-637) certified for total Se content.
Jitaru
Petru
P
Institute for the Dynamics of Environmental Processes (IDPA-CNR), Dorsoduro 2137, 30123, Venice, Italy.
Cozzi
Giulio
G
Gambaro
Andrea
A
Cescon
Paolo
P
Barbante
Carlo
C
D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D002852:Chromatography, Ion Exchange / Q000379:methods*; D005979:Glutathione Peroxidase / Q000097:blood* / Q000378:metabolism; D006801:Humans; D007554:Isotopes / Q000737:chemistry; D012643:Selenium / Q000097:blood; D051149:Selenoprotein P / Q000097:blood* / Q000378:metabolism; D012709:Serum Albumin / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
D007554:Isotopes; D051149:Selenoprotein P; D012709:Serum Albumin; D005979:Glutathione Peroxidase; D012643:Selenium