Detection of Over 110 Anabolic–Androgenic Steroids, Corticosteroids and/or Their Esters in Horse Hair Using Ultra‐High‐Performance Liquid Chromatography–High‐Resolution Mass Spectrometry and Gas Chromatography–Tandem Mass Spectrometry After Solid‐Supported Liquid Extraction

Abstract

Anabolic–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 duration of action. As such, the detection of intact esters of endogenous steroids in hair is particularly advantageous, as it can provide unequivocal proof of their external origin. Compared with urine and blood matrices, hair in general allows a longer detection window as long as the administered drug can be incorporated into hair and has not degraded. This can be useful in doping control of equine sports, where the timing of sample collection may be critical due to the transient nature of AAS in other biological fluids. This paper describes the detection of AAS and/or their esters, as well as corticosteroids, in horse hair using ultra‐high‐performance liquid chromatography–high‐resolution mass spectrometry (UPLC‐HRMS) and gas chromatography–tandem mass spectrometry (GC‐MS/MS) after solid‐supported liquid extraction (SLE) at basic pH. The developed UPLC‐HRMS and GC‐MS/MS methods in combination allowed the detection of 117 AAS, corticosteroids and/or their esters with estimated limits of detection down to sub‐ppb levels and with good inter‐day precision. Method applicability has been demonstrated through the detections of (i) boldenone undecylenate, nandrolone phenylpropionate and trenbolone acetate in three proficiency testing hair samples and (ii) nandrolone as a metabolite in a post‐administration hair sample collected from a castrated horse having been administered nandrolone decanoate.