Detection of Growth Hormone Releasing Peptides in Serum by a Competitive Receptor Binding Assay

Ferro, P.; Krotov, Grigory; Zvereva, Irina; Mana, Pérez; Ja, Mateus; Segura, Jordi

doi:10.4172/2157-7064.1000351

Cited by 3 publications

(4 citation statements)

References 37 publications

(56 reference statements)

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“…After intravenous (i.v.) administration of GHRP-2, detection times appeared shorter in serum than in urine samples, but for GHRP-6, detection windows were found to be comparable in both matrices [21,22]. Nevertheless, for urine analysis, knowledge about the metabolic fate of peptide drugs is desirable as the presence of metabolites alongside the intact and unmodified drug (candidate) was shown in the past [3,9,23].…”

Section: Introductionmentioning

confidence: 99%

Fully automated dried blood spot sample preparation enables the detection of lower molecular mass peptide and non-peptide doping agents by means of LC-HRMS

et al. 2020

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The added value of dried blood spot (DBS) samples complementing the information obtained from commonly routine doping control matrices is continuously increasing in sports drug testing. In this project, a robotic-assisted non-destructive hematocrit measurement from dried blood spots by near-infrared spectroscopy followed by a fully automated sample preparation including strong cation exchange solid-phase extraction and evaporation enabled the detection of 46 lower molecular mass (< 2 kDa) peptide and non-peptide drugs and drug candidates by means of LC-HRMS. The target analytes included, amongst others, agonists of the gonadotropin-releasing hormone receptor, the ghrelin receptor, the human growth hormone receptor, and the antidiuretic hormone receptor. Furthermore, several glycine derivatives of growth hormone-releasing peptides (GHRPs), arguably designed to undermine current anti-doping testing approaches, were implemented to the presented detection method. The initial testing assay was validated according to the World Anti-Doping Agency guidelines with estimated LODs between 0.5 and 20 ng/mL. As a proof of concept, authentic post-administration specimens containing GHRP-2 and GHRP-6 were successfully analyzed. Furthermore, DBS obtained from a sampling device operating with microneedles for blood collection from the upper arm were analyzed and the matrix was cross-validated for selected parameters. The introduction of the hematocrit measurement method can be of great value for doping analysis as it allows for quantitative DBS applications by managing the well-recognized "hematocrit effect."

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Section: Introductionmentioning

confidence: 99%

Fully automated dried blood spot sample preparation enables the detection of lower molecular mass peptide and non-peptide doping agents by means of LC-HRMS

et al. 2020

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“…11 Sensitivity was good enough to detect GHRP-2 presence up to 3-4 h in the samples from the administration study. This is advantageous over a previous publication, in which the displacement effect of GHRP-2 over radioactive ghrelin in serum 30 was observed for shorter times after administration. The window of detection reached here with LC-MS/MS slightly improves the prospects on GHRPs detection.…”

Section: Discussionmentioning

confidence: 80%

On the road of dried blood spot sampling for antidoping tests: Detection of GHRP‐2 abuse

Reverter-Branchat

Segura

Pozo

2020

Drug Testing and Analysis

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Dried blood spots (DBSs) sampling is gaining support by the antidoping community because of simplicity and cost‐effective characteristics, especially in collection, transport, and storage. Nevertheless, DBS applicability demands specific studies for each of the analytes proposed for testing. Here, GHRP‐2 has been selected as a representing member of the growth hormone‐releasing peptides (GHRPs) family to provide further evidence of DBS suitability for GHRPs abuse detection in sport testing. An analytical procedure to extract GHRP‐2 and its main metabolite (AA‐3) from DBS and to detect them by liquid chromatography–tandem mass spectrometry (LC–MS/MS) has been developed. The method has been validated for the detection of GHRP‐2. Specificity and identification capabilities have been assessed in agreement with antidoping guidelines. The low AA‐3 levels found in DBS samples prevented its effective application for the determination of this metabolite. The limit of detection (LoD) for GHRP‐2 has been established at 50 pg/ml. Long‐term stability (>2 years) has been confirmed. The procedure has been successfully applied to actual DBS samples from an administration study with a single intravenous dose of GHRP‐2 (100 μg) being detected up to 4 h after drug injection. GHRP‐2 concentrations have been higher in venous blood DBS than in capillary blood DBS. Despite the observed differences, a similar detection window has been achieved independently of the type of blood used. In summary, this study provides specific evidence supporting DBS usefulness to detect GHRP‐2, and potentially other GHRPs family members, for antidoping tests.

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“…For example, measurement of GnRH analogs has relied upon the use of such assays [31], however, the lack of specificity for degradation products limited its applicability for doping control analysis. The feasibility of using receptor binding assays (GHS-R1a) for the extraction of GHSs from urine [32][33][34] and serum [35] has also been demonstrated. This approach could be applied not only to parent drugs showing affinity of the GHS-R1a receptor but also to catabolites [34], which can make it a promising strategy to complete non-LBA-based strategies for the measurement of GHSs.…”

Section: Trends In Sample Clean-up Of Prohibited Spsmentioning

confidence: 99%

“…Besides the numerous applications based on urinary matrices, there are a few applications in plasma [55,36,46] or serum [46,35] as well. This matrix is rarely used not only because of the lower concentrations and shorter detection windows that are generally observed compared to urine, but also because of the more invasive nature of blood collection.…”

Section: Trends In Sample Clean-up Of Prohibited Spsmentioning

confidence: 99%

Doping control analysis of small peptides: A decade of progress

Judák

Esposito

Coppieters

et al. 2021

Journal of Chromatography B

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Detection of Growth Hormone Releasing Peptides in Serum by a Competitive Receptor Binding Assay

Cited by 3 publications

References 37 publications

Fully automated dried blood spot sample preparation enables the detection of lower molecular mass peptide and non-peptide doping agents by means of LC-HRMS

Fully automated dried blood spot sample preparation enables the detection of lower molecular mass peptide and non-peptide doping agents by means of LC-HRMS

On the road of dried blood spot sampling for antidoping tests: Detection of GHRP‐2 abuse

Doping control analysis of small peptides: A decade of progress

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