There has been an on-going effort and long term goal to accurately assess selenium status and selenium nutritional requirement in humans through the quantification of human plasma selenoprotein P (SelP) and glutathione peroxidase 3 (GPx3). To achieve this goal, this study explored the potential of mass spectrometry techniques for selenoprotein identification and determination in a Human Plasma candidate Standard Reference Material SRM 1950 with a total selenium concentration of 105.5 ± 2.3 µg kg-1. Since classical proteomic shotgun approach of depleted human plasma is not specific and sensitive enough to identify selenoproteins, a laser ablation ICP-MS method was first developed to target selenoprotein. The combination of better sample preparation, by concentrating selenoproteins onto membranes, and better sensitivity of Se detection, by humidifying ICP with an organic solution, resulted in a laser ablation ICP-MS method 80 times more sensitive compared to classical LA ICP-MS methods. The method was successfully applied in this study to the detection of human plasma selenoproteins at physiological concentrations. Accurate identification of SelP and GPx3 was then carried out by tandem mass spectrometry. More than seven peptides were identified for both proteins with good quality MS/MS spectra. For future quantification by MS/MS, these peptides were studied to select proteotypic candidates and LPTDSELAPR for SelP and NCSPPTSELLGTSDR for GPx3 were the best candidates. This study highlights the importance of an approach combining ICP-MS and tandem mass spectrometry for accurate selenoprotein characterization before further quantification by standard addition or isotope dilution mass spectrometry. This procedure may be easily performed in others laboratories to study selenoproteins or other metalloproteins in humans or other biological samples using a conventional laser device.
Citation: Journal of Analytical Atomic Spectrometry
Pub Type: Journals
glutathione peroxidase 3, human plasma, selenoprotein P, laser ablation ICP-MS, tandem mass spectrometry, Standard Reference Material