Universität zu Köln

Knoop, Andre, Thomas, Andreas ORCID: 0000-0003-1199-0743, Fichant, Eric, Delahaut, Philippe, Schaenzer, Wilhelm and Thevis, Mario (2016). Qualitative identification of growth hormone-releasing hormones in human plasma by means of immunoaffinity purification and LC-HRMS/MS. Anal. Bioanal. Chem., 408 (12). S. 3145 - 3154. HEIDELBERG: SPRINGER HEIDELBERG. ISSN 1618-2650

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Abstract

The use of growth hormone-releasing hormones (GHRHs) is prohibited in sports according to the regulations of the World Anti-Doping Agency (WADA). The aim of the present study was to develop a method for the simultaneous detection of four different GHRHs and respective metabolites from human plasma by means of immunoaffinity purification and subsequent nano-ultrahigh performance liquid chromatography-high resolution/high accuracy (tandem) mass spectrometry. The target analytes included Geref (Sermorelin), CJC-1293, CJC-1295, and Egrifta (Tesamorelin) as well as two metabolites of Geref and CJC-1293, which were captured from plasma samples using a polyclonal GHRH antibody in concert with protein A/G monolithic MSIA (TM) D.A.R.T.'SA (R) (Disposable Automation Research Tips) prior to separation and detection. The method was fully validated and found to be fit for purpose considering the parameters specificity, linearity, recovery (19-37 %), lower limit of detection (< 50 pg/mL), imprecision (< 20 %), and ion suppression/enhancement effects. The analytes' stability and metabolism were elucidated using in vitro and in vivo approaches. EDTA blood samples were collected from rats 2, 4, and 8 h after intravenous administration of GHRH (one compound per test animal). All intact substances were detected for at least 4 h but no anticipated metabolite was confirmed in laboratory rodents' samples; conversely, a Geref metabolite (GHRH(3-29)) was found in a human plasma sample collected after subcutaneous injection of the drug to a healthy male volunteer. The obtained results demonstrate that GHRHs are successfully detected in plasma using an immunoaffinity-mass spectrometry-based method, which can be applied to sports drug testing samples. Further studies are however required and warranted to account for potential species-related differences in metabolism and elimination of the target analytes.

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 Fichant, Eric UNSPECIFIED UNSPECIFIED UNSPECIFIED Delahaut, Philippe UNSPECIFIED UNSPECIFIED UNSPECIFIED Schaenzer, Wilhelm UNSPECIFIED UNSPECIFIED UNSPECIFIED Thevis, Mario UNSPECIFIED UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-277322
DOI:	10.1007/s00216-016-9377-3
Journal or Publication Title:	Anal. Bioanal. Chem.
Volume:	408
Number:	12
Page Range:	S. 3145 - 3154
Date:	2016
Publisher:	SPRINGER HEIDELBERG
Place of Publication:	HEIDELBERG
ISSN:	1618-2650
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language CHROMATOGRAPHY-MASS SPECTROMETRY; CJC-1295; ANALOG; PEPTIDE; IMMUNOASSAY; STIMULATION; STRATEGIES; SECRETION; SERUM; GH Multiple languages Biochemical Research Methods; Chemistry, Analytical Multiple languages
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/27732