Optimization of micro‐HPLC peak focusing for the detection and quantification of low hepcidin concentrations |
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Authors: | François Mansion Patrice Chiap Virginie Houbart Jacques Crommen Anne‐Catherine Servais Marianne Fillet |
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Affiliation: | 1. Laboratory of Analytical Pharmaceutical Chemistry, Department of Pharmacy, CIRM, University of Liege, Liege, Belgium;2. Advanced Technology Corporation (ATC), Liege, Belgium |
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Abstract: | Micro‐high‐performance liquid chromatography is a miniaturized, economic and ecological chromatographic system allowing the use of reduced size chromatographic columns. Coupled with electrospray ionization tandem mass spectrometry, this technique can be used to detect and quantify low concentrations of peptides. In this study, hepcidin was used as the model compound and analysed using octadecylsilica stationary phase by means of a gradient elution mode at a flow rate of 4 μL/min. Several parameters were studied to optimize peak focusing. Using the methodology of experimental design, the mobile‐phase gradient conditions and the sample composition were optimized in order to maximize the sensitivity and minimize retention time. Stability of the target peptide in solution was also demonstrated. |
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Keywords: | Hepcidin Micro‐HPLC‐MS MS Peak focusing |
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