Detection of seventy-two anabolic and androgenic steroids and/or their esters in horse hair using ultra-high performance liquid chromatography-high resolution mass spectrometry in multiplexed targeted MS2 mode and gas chromatography-tandem mass spectrometry.
Abstract: Anabolic and androgenic steroids (AAS) are banned substances in both human and equine sports. They are often administered intramuscularly to horses in esterified forms for the purpose of extending their time of action. The authors' laboratory has previously reported an UHPLC/HRMS method using quadrupole-Orbitrap mass spectrometer in full scan and parallel reaction monitoring (PRM) mode for the detection of 48 AAS and/or their esters in horse hair. However, two injections were required due to the long duty cycle time. In this paper, an UHPLC/HRMS method using multiplexed targeted MS mode was developed and validated to improve the coverage to 65 AAS and/or their esters in a single injection. In addition, a GC/MS/MS method in selected reaction monitoring (SRM) mode was developed to screen for another seven AAS and/or their esters not adequately covered by the UHPLC/HRMS method using the same sample extract after derivatisation with pentafluoropropionic anhydride. The UHPLC/HRMS and GC/MS/MS methods in combination allowed the detection of 72 AAS and/or their esters with estimated limits of detection down to sub to low ppb levels with good interday precision. Method applicability was demonstrated by the detection of boldione and 4-androstenedione in two out-of-competition hair samples and testosterone propionate in a referee hair sample.
Copyright © 2018 Elsevier B.V. All rights reserved.
Publication Date: 2018-06-20 PubMed ID: 29958683DOI: 10.1016/j.chroma.2018.06.049Google Scholar: Lookup
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Summary
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The research paper presents the development of an improved method for the detection of anabolic and androgenic steroids (AAS) in horse hair using ultra-high performance liquid chromatography-high resolution mass spectrometry and gas chromatography-tandem mass spectrometry. This developed technique is capable of detecting 72 different steroids in a single sample run.
Use of Anabolic and Androgenic Steroids in Equine Sports
- The paper begins with establishing the context, explaining how Anabolic and Androgenic Steroids (AAS) are forbidden substances in both human and equine sports. These substances, often administered intramuscularly in esterified forms, are designed to increase muscle strength and performance.
- Noting the need for effective detection methods to ensure the integrity of the sport and health of the animals, the authors highlight their previous work with an ultra-high performance liquid chromatography-high resolution mass spectrometry (UHPLC/HRMS) method which successfully detected 48 different types of AAS and/or their esters in horse hair samples.
Improvement in Steroid Detection: Single-Injection Multiplexed Targeted MS Mode
- The research paper presents an advanced version of the previous method where an UHPLC/HRMS process employs multiplexed targeted MS mode.
- This advanced method allows the detection of 65 different AAS in a single injection, which offers an improvement in efficiency and detection scope compared to the older two-injection method.
Gas Chromatography-Tandem Mass Spectrometry (GC/MS/MS) Method
- In addition to the UHPLC/HRMS method, the authors also establish a GC/MS/MS method to detect seven additional AAS substances that were not within the detection range of the UHPLC/HRMS method.
- This secondary method involves the use of selected reaction monitoring (SRM) mode and requires chemical modification of the sample with pentafluoropropionic anhydride before analysis.
Results and Implications
- As a result, the combination of these two methods allows the detection of a total of 72 different AAS and/or their esters from a single sample.
- The newly developed method delivers highly accurate and precise results with limits of detection down to the sub to low ppb levels.
- The authors verified the effectiveness of the method by testing hair samples from out-of-competition horses and confirming the presence of specific AAS substances.
Cite This Article
APA
Choi TLS, Kwok KY, Kwok WH, Tsoi YYK, Wong JKY, Wan TSM.
(2018).
Detection of seventy-two anabolic and androgenic steroids and/or their esters in horse hair using ultra-high performance liquid chromatography-high resolution mass spectrometry in multiplexed targeted MS2 mode and gas chromatography-tandem mass spectrometry.
J Chromatogr A, 1566, 51-63.
https://doi.org/10.1016/j.chroma.2018.06.049 Publication
Researcher Affiliations
- Racing Laboratory, The Hong Kong Jockey Club, Sha Tin Racecourse, Sha Tin, N.T., Hong Kong, China. Electronic address: timmy.ls.choi@hkjc.org.hk.
- Racing Laboratory, The Hong Kong Jockey Club, Sha Tin Racecourse, Sha Tin, N.T., Hong Kong, China.
- Racing Laboratory, The Hong Kong Jockey Club, Sha Tin Racecourse, Sha Tin, N.T., Hong Kong, China.
- Racing Laboratory, The Hong Kong Jockey Club, Sha Tin Racecourse, Sha Tin, N.T., Hong Kong, China.
- Racing Laboratory, The Hong Kong Jockey Club, Sha Tin Racecourse, Sha Tin, N.T., Hong Kong, China.
- Racing Laboratory, The Hong Kong Jockey Club, Sha Tin Racecourse, Sha Tin, N.T., Hong Kong, China. Electronic address: terence.sm.wan@hkjc.org.hk.
MeSH Terms
- Androstenedione / analysis
- Animals
- Chromatography, High Pressure Liquid
- Doping in Sports
- Esters / analysis
- Esters / chemistry
- Gas Chromatography-Mass Spectrometry
- Hair / chemistry
- Horses
- Steroids / analysis
- Steroids / chemistry
- Tandem Mass Spectrometry
- Testosterone Propionate / analysis
Citations
This article has been cited 4 times.- Cloteau C, Dervilly G, Loup B, Delcourt V, Kaabia Z, Bagilet F, Groseille G, Dauriac K, Fisher S, Popot MA, Garcia P, Le Bizec B, Bailly-Chouriberry L. Performance assessment of an equine metabolomics model for screening a range of anabolic agents.. Metabolomics 2023 Apr 7;19(4):38.
- Haynes AZ, Levine M. Detection of anabolic steroids via cyclodextrin-promoted fluorescence modulation.. RSC Adv 2020 Jun 29;10(42):25108-25115.
- Caprioli G, Genangeli M, Mustafa AM, Petrelli R, Ricciutelli M, Sagratini G, Sartori S, Laus F, Vittori S, Cortese M. Quantification of 17 Endogenous and Exogenous Steroidal Hormones in Equine and Bovine Blood for Doping Control with UHPLC-MS/MS.. Pharmaceuticals (Basel) 2021 Apr 21;14(5).
- Chan WS, Wong GF, Hung CW, Wong YN, Fung KM, Lee WK, Dao KL, Leung CW, Lo KM, Lee WM, Cheung BK. Interpol review of toxicology 2016-2019.. Forensic Sci Int Synerg 2020;2:563-607.
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