Mouldy feed: A possible explanation for the excretion of anabolic‐androgenic steroids in horses

Abstract

To ensure fair competition and to protect the horse's welfare, horses have to compete on their own merits, without any unfair advantage that might follow the use of drugs. Therefore, regulatory authorities list all substances that are not allowed in competition, including most anabolic-androgenic steroids. As zero-tolerance is retained, the question arose whether the consumption of mouldy feed could lead to the excretion of steroids, due to the biotransformation of plant phytosterols to steroids. A rapid ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) analytical method, previously validated according to AORC (Association of Official Racing Chemists) and EC (European Commission) guidelines, was used to measure steroids in different sample types. Multiple mouldy feed samples were tested for the presence of steroids. The effect of digestion was tested by in vitro simulation of the horse's hindgut in batch incubations. In most feed samples no steroids were detected, even when the products were mouldy. Mouldy corn however showed to contain up to 3.0 ± 0.4 μg/kg AED (4-androstenedione), the main testosterone precursor. This concentration increased when mouldy corn (with added phytosterols) was digested in vitro. An herbal phytosupplement also showed to contain α-testosterone. These results demonstrate that it is important to caution against the consumption of any feed or (herbal) supplement of which the detailed ingredients and quantitative analysis are unknown. The consumption of mouldy corn should especially be avoided, not only from a horse health and welfare point of view, but also to avoid possible inadvertent positive doping results.

A review of designer anabolic steroids in equine sports

“…The limitations of this approach are reflected in the sample preparation of the whole liver extracts, as well as potential variations in the metabolic profile due to the use of individual tissue donors. Other approaches to model equine metabolism involve microbial systems which are typically based on equine faecal bacteria, and although these are currently largely unimportant for modelling equine steroid metabolism in an anti‐doping context, they may be important in other areas …”

MetIDfyR: An Open-Source R Package to Decipher Small-Molecule Drug Metabolism through High-Resolution Mass Spectrometry

“…This is a crucial step involved in multiple stages of a compound's life such as drug discovery and development, [1][2][3] preliminary in-vitro and in-vivo experiments, pharmaco-kinetics/dynamics studies 4,5 and forensics, [6][7][8][9][10] including drug-testing of human and animal athletes. [11][12][13][14][15][16][17] Over the past few decades, multiple key techniques and findings conducted the scientific community to expand knowledge on these complex molecular pathways. This includes the use of radioisotopes, 18 immunoassays, the identification of enzymes responsible of various and possibly complex biotransformations, 19 leading to the definition of most prevalent biochemical reactions for detoxification 20 and key actors of these mechanisms such as the P450 cytochrome enzyme family.…”

MetIDfyR, an Open-Source R Package to Decipher Small-Molecule Drugs Metabolism Through High Resolution Mass Spectrometry

“…This is a crucial step involved in multiple stages of a compound's life such as drug discovery and development, [1][2][3] preliminary in-vitro and in-vivo experiments, pharmaco-kinetics/dynamics studies 4,5 and forensics, [6][7][8][9][10] including drug-testing of human and animal athletes. [11][12][13][14][15][16][17] Over the past few decades, multiple key techniques and findings conducted the scientific community to expand knowledge on these complex molecular pathways. This includes the use of radioisotopes, 18 immunoassays, the identification of enzymes responsible of various and possibly complex biotransformations, 19 leading to the definition of most prevalent biochemical reactions for detoxification 20 and key actors of these mechanisms such as the P450 cytochrome enzyme family.…”