Using electrochemistry for metabolite simulation and synthesis in preventive doping research: application to the Rycal S107 and the PPARδ-agonist GW1516

Abstract

In view of the numerous newly emerging drug candidates, it is a constant challenge for doping control laboratories to keep sports drug testing procedures up-to-date, thus, allowing for comprehensive analyses. Therefore, it is of major importance to elucidate the biotransformation pathway of potentially performance-enhancing therapeutics as early as possible in order to determine analytical targets that can reliably identify drug abuse. Usually, in vivo and/or in vitro assays with rodents and/or liver cell preparations, respectively, serve this purpose. However, these models for metabolism simulation are time-consuming and uneconomical for the generation and isolation of larger amounts of metabolite reference material for thorough method development. In this work, electrochemistry (EC) in hyphenation with liquid chromatography (LC) and electrospray mass spectrometry (ESI-MS) was used to study the metabolic fate of the ryanodine receptor-calstabin-complex stabilizer (“Rycal”) S107 and the peroxisome proliferator-activated receptor (PPAR) δ agonist GW1516. High resolution/high accuracy MS and tandem MS were utilized and supported by experiments with stable isotope-labeling for S107, or human liver microsomes (HLM) and S9 fraction for GW1516. The results were compared with recently published in vitro data. For both potential doping agents, the purely instrumental approach succeeded in generating all of the known in vitro phase I metabolites. Mono-oxygenated products like N- and/or S-oxides were generated, next to demethylated and dehydrogenated species (S107) or bis-oxygenated products (e.g. the GW1516 sulfone). This demonstrates the value of EC as a fast and easy method for metabolism simulation while facilitating the generation of pure reference material for preventive doping research.