Determination of ghrelin and desacyl ghrelin in human plasma and urine by means of LC-MS/MS for doping controls

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The hunger hormone ghrelin (G) is classified as prohibited substance in professional sport by the World Anti-Doping Agency (WADA), due to its known growth hormone releasing properties. The endogenous bioactive peptide consists of 28 amino acids with a caprylic acid attached to serine at position 3. Within this study, it was aimed to develop methods to determine G and desacyl ghrelin (DAG) in plasma and urine by means of LC-MS/MS. Two strategies were applied with a bottom-up approach for plasma and top-down analyses for urine. Both sample preparation procedures were based on solid-phase extraction for enrichment and sample clean-up. Method validation showed good results for plasma and urine with limits of detection (LODs) for G and DAG between 30 and 50 pg/ml, recoveries between 45-50%, and imprecisions (intra- and inter-day) between 3% and 24%. Plasma analysis was also valid for quantification with accuracies determined with ~100% for G and ~106% for DAG. The minimum required performance level for doping control laboratories is set to 2 ng/ml in urine, and the herein established method yielded acceptable results even at 5% of this level. As proof-of-concept, plasma levels (G and DAG) of healthy volunteers were determined and ranged between 30 and 100 pg/ml for G and 100-1200 pg/ml for DAG. In contrast to earlier reported studies using ligand binding assays for urinary G and DAG, in this mass spectrometry-based study, no endogenous urinary G and DAG were found, although the LODs should enable this.

Original languageEnglish
JournalDrug testing and analysis
Pages (from-to)1-9
Number of pages9
Publication statusElectronically/ online published ahead of print - 11.10.2021

Research areas and keywords

  • doping
  • high resolution mass spectrometry


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