Development of a metabonomic approach based on LC-ESI-HRMS measurements for profiling of metabolic changes induced by recombinant equine growth hormone in horse urine

Despite the worldwide existing regulation banning the use of the recombinant equine growth hormone (reGH) as growth promoter, it is suspected to be used in horseracing to improve performances. Various analytical methods previously developed to screen for its misuse have encountered some limitations in terms of detection timeframes, in particular during the first days following reGH administration. A novel strategy involving the characterization of global metabolomic fingerprints in urine samples of non-treated and reGH-treated horses by liquid chromatography-electrospray-high-resolution mass spectrometry (LC-ESI-HRMS) is described and assessed in this paper in order to develop a new screening tool for growth hormone abuse in horseracing. The strategy involves a limited sample preparation of the urine samples and the use of appropriate software for data processing and analysis. As preliminary work, reproducibility of both sample preparation and mass spectrometry (MS) measurements was evaluated in order to demonstrate the reliability of the method. Application of the developed protocol on two horses demonstrated the suitability of the developed strategy and preliminary results showed significant modifications of the metabolome after treatment with reGH.