Selective androgen receptor modulators:in vitroandin vivometabolism and analysis

Abstract

For future targeted screening in National Residue Control Programmes, the metabolism of seven SARMs, from the arylpropionamide and the quinolinone classes, was studied in vitro using S9 bovine liver enzymes. Metabolites were detected and identified with ultra-performance liquid chromatography (UPLC) coupled to time-of-flight mass spectrometry (ToF-MS) and triple quadrupole mass spectrometry (QqQ-MS). Several metabolites were identified and results were compared with literature data on metabolism using a human cell line. Monohydroxylation, nitro-reduction, dephenylation and demethylation were the main S9 in vitro metabolic routes established. Next, an in vivo study was performed by oral administration of the arylpropionamide ostarine to a male calf and urine samples were analysed with UPLC-QToF-MS. Apart from two metabolites resulting from hydroxylation and dephenylation that were also observed in the in vitro study, the bovine in vivo metabolites of ostarine resulted in glucuronidation, sulfation and carboxylation, combined with either a hydroxylation or a dephenylation step. As the intact mother compounds of all SARMs tested are the main compounds present after in vitro incubations, and ostarine is still clearly present in the urine after the in vivo metabolism study in veal calves, the intact mother molecules were selected as the indicator to reveal treatment. The analytical UPLC-QqQ-MS/MS procedure was validated for three commercially available arylpropionamides according to European Union criteria (Commission Decision 2002/657/EC), and resulted in decision limits ranging from 0.025 to 0.05 µg l⁻¹ and a detection capability of 0.025 µg l⁻¹ in all cases. Adequate precision and intra-laboratory reproducibility (relative standard deviation below 20%) were obtained for all SARMs and the linearity was 0.999 for all compounds. This newly developed method is sensitive and robust, and therefore useful for confirmation and quantification of SARMs in bovine urine samples for residue control programmes and research purposes.

Specific characterization of non‐steroidal selective androgen peceptor modulators using supercritical fluid chromatography coupled to ion‐mobility mass spectrometry: application to the detection of enobosarm in bovine urine

“…Patterns of fragmentation and fragments identification have already been proposed by Thevis et al [31] and De Rijke et al [15] for andarine and enobosarm. They have therefore been considered as a basis to explain observed fragmentation using MS/MS, IM-MS/MS or IM-MS E (Figure 3).…”

“…One animal was orally administered a unique dose of enobosarm (GTx-024, GTx Inc., Memphis, TN, USA) (200 mg) in ethanolic solution diluted in apple juice, adapted from a protocol described by De Rijke et al [15] The five others formed the control group and were orally administrated the same ethanol/apple juice mix without enobosarm. All were aged three months (56 ± 5 days) (solid feed and hay) and weighed approximately 100 ± 5 kg at the beginning of the experiment.…”

Selective androgen receptor modulators: comparative excretion study of bicalutamide in bovine urine and faeces

“…[15][16][17] SARMs may also exhibit a high potential for misuse in animal husbandry as growth promoters to increase weight gain, improve carcass quality, and reduce production costs. More studies are necessary to develop the comprehensiveness of SARMs metabolism through in vivo approaches, [20,21] as already available for humans and horses. Efficient analytical strategies to detect the use of SARMs have already been proposed and while main developments have been realized in the human anti-doping arena, few applications are now available in food producing animals.…”

“…[18,19] Since these compounds have recently emerged, they have not been investigated a lot. [21] Bicalutamide exhibits an anti-androgen effect and is used for the treatment of human non-metastatic prostate cancer. We therefore propose in this study to investigate further SARMs elimination and metabolism in bovine.…”

Investigation of the selective androgen receptor modulators S1, S4 and S22 and their metabolites in equine plasma using high‐resolution mass spectrometry

“…[7] Furthermore, the identification of metabolites also corroborates that a substance has passed through a metabolic system and that an adverse finding is not caused by contamination of the sample. The metabolism of different SARMs of the arylpropionamide class has been studied in vitro in a variety of systems such as human liver microsomes, [8] equine liver microsomes, [9] bovine liver microsomes [10] and fungal incubates, [11] as well as in vivo in both human [12,13] and equine [14] urine. The three aforementioned in vivo studies showed that the SARMs were metabolized rapidly and extensively.…”