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

Determination of ghrelin and desacyl ghrelin in human plasma and urine by means of LC-MS/MS for doping controls / Andreas Thomas, Sophia Krombholz, Carina Wolf, Mario Thevis. - (Drug Testing and Analysis (2021) 11 October)

  • PMID: 34633773
  • DOI: 10.1002/dta.3176


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 - 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 document


Research / Study
11 October 2021
Krombholz, Sophia
Thevis, Mario
Thomas, Andreas
Wolf, Carina
Other organisations
Deutsche Sporthochschule Köln (DSHS) - German Sport University Cologne
European Monitoring Center for Emerging Doping Agents (EuMoCEDA)
Analytical aspects
Mass spectrometry analysis
Testing method development
Doping classes
S2. Peptide Hormones, Growth Factors
Lenomorelin (ghrelin)
Document category
Scientific article
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Pdf file
Date generated
18 October 2021
Date of last modification
25 November 2021
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