Optimizations of the analysis conditions and validations of the confirmation and detection method for small peptides by LC-MS/MS

Identifier:  TFM:310

Handle: https://hdl.handle.net/20.500.11797/TFM310

Authors:  Sauras Pastor, Victoria.

Abstract:
An analytical method for the detection of small peptides in human urine has been developed and validated. Peptides that are analysed are families of GHRPs, GHS, GnRH, diuretic agents, growth factors and growth factor modulators which are consumed by some athletes because of their ergogenic effect on the body. The method consist in an solid phase extraction by OASIS WCX cartridges and a separation by high-perfonmance liquid chromatography-electrospray tandem mass spectrometry (LC-ESI-MS/MS), with C18 column (1.7 μm particle size), a mobile phase containing acetonitrile with 0.1% formic acid of solvent A and deionized water with 0.1% formic acid of solvent B. The gradient programm starts with 5% solvent A and increases until 35% in 8 minuts, until 80% in the minut 10 and then decreases to 5% in the next minute and mantein this conditions until minut 15. The detection was carried out with electrospray ionization in positive mode monitorizing the two best precurssor ion – product ion specific transitions for each analyte. The methodology was validated in terms of extraction recovery, intra assay precision (repeatability) and limits of detection (LOD) with LODs from 1 to 2 ng/ml.