Introduction: Phosphodiesterase 4 (PDE4) inhibitors represent an emerging class of pharmacological agents known for their bronchodilatory and anti-inflammatory effects. These therapeutic benefits have positioned PDE4 inhibitors, as effective treatments for respiratory and inflammatory conditions. The detection and characterization of metabolites of these compounds are therefore critical for both clinical monitoring and the enforcement of anti-doping regulations. This study aims to investigate the metabolic fate of two PDE4 inhibitors roflumilast and ibudilast following oral administration in thoroughbred horses, as well as through in vitro experiments using equine liver microsomes and the fungal model Cunninghamella elegans.
Methods: Following oral administration of roflumilast and ibudilast in horses, blood and urine samples were collected and analyzed. Parallel in vitro experiments employed equine liver microsomes and Cunninghamella elegans to simulate metabolic pathways. High-performance liquid chromatography coupled with high-resolution mass spectrometry was applied to identify and characterize metabolite structures.
Results: For roflumilast, ten metabolites were detected in vivo, consisting of six phase I and four phase II metabolites. In vitro studies revealed eleven metabolites, with eight belonging to phase I and three to phase II. For ibudilast, a total of twenty metabolites were detected in vivo, including twelve phase I and eight phase II metabolites. Hydroxylation (mono, di, and tri-hydroxylation) was the predominant pathway, along with demethylation. Phase II metabolism produced glucuronide and sulfonic acid conjugates.
Conclusion: This study highlights the complex metabolic pathways of PDE4 inhibitors in equine systems. The identification of multiple phase I and phase II metabolites, both in vivo and in vitro, provides valuable insights into the biotransformation of roflumilast and ibudilast. These findings contribute to the development of analytical methods for detecting their unauthorized use in equestrian sports and enhance anti-doping surveillance strategies.
Keywords
PDE4 inhibitors, Roflumilast, Ibudilast, Drug biotransformation, Equine anti-doping