Universität zu Köln

Knoop, Andre, Thomas, Andreas ORCID: 0000-0003-1199-0743 and Thevis, Mario (2019). Development of a mass spectrometry based detection method for the mitochondrion-derived peptide MOTS-c in plasma samples for doping control purposes. Rapid Commun. Mass Spectrom., 33 (4). S. 371 - 381. HOBOKEN: WILEY. ISSN 1097-0231

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Abstract

Rationale The mitochondrial open reading frame of 12S rRNA type-c (MOTS-c) peptide was recently discovered and described to control metabolic homeostasis through AMPK activation along with AICAR accumulation. Consequently, it appears advisable to monitor the potential use of synthetic MOTS-c in sports, and a detection method suitable for sports drug testing purposes is necessary. Methods For the detection of MOTS-c in doping control plasma samples, a test method employing liquid chromatography and mass spectrometry (LC/MS) was developed. Following optimization, the assay was comprehensively validated and additional parameters such as the (long-term) stability and in vitro metabolism of the peptide were evaluated. In order to determine endogenous MOTS-c reference limits, the results generated by LC/MS-based detection were compared with those obtained with a commercially available enzyme-linked immunosorbent assay (ELISA). Results The LC/MS-based test method was fully validated for quantitative results interpretation according to the World Anti-Doping Agency's International Standard for Laboratories (WADA's ISL). It was found to be specific and sensitive, enabling a lower limit of detection (LLOD) for hMOTS-c in plasma at 100 pg/mL. Following optimization, animal MOTS-c analogues and four metabolites as well as two oxidation products were implemented. However, endogenous levels of a reference population of 20 healthy subjects studied by ELISA experiments (45.9-218.5 ng/mL) could not be confirmed by LC/MS. Conclusions A mass spectrometric detection assay for MOTS-c in human plasma samples was developed and successfully validated according to WADA's ISL, providing an additional tool for future doping control purposes. Besides MOTS-c, the assay also includes four in vitro derived metabolites and two oxidation products, which might further improve the traceability of the drug. The analytical approach was compared with a commercially available ELISA, and considerable differences in measured MOTS-c levels were observed.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Knoop, Andre UNSPECIFIED UNSPECIFIED UNSPECIFIED Thomas, Andreas UNSPECIFIED orcid.org/0000-0003-1199-0743 UNSPECIFIED Thevis, Mario UNSPECIFIED UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-156236
DOI:	10.1002/rcm.8337
Journal or Publication Title:	Rapid Commun. Mass Spectrom.
Volume:	33
Number:	4
Page Range:	S. 371 - 381
Date:	2019
Publisher:	WILEY
Place of Publication:	HOBOKEN
ISSN:	1097-0231
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language SKELETAL-MUSCLE; SENSITIVITY; IONIZATION; OBESITY Multiple languages Biochemical Research Methods; Chemistry, Analytical; Spectroscopy Multiple languages
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/15623