Control of methylxanthines in the competition horse: pharmacokinetic/pharmacodynamic studies on caffeine, theobromine and theophylline for the assessment of irrelevant concentrations

Methylxanthines positives in competition samples have challenged doping control laboratories and racing jurisdictions since methylxanthines are naturally occurring prohibited substances and often constituents of feed. For theobromine an international threshold (renamed in International Residue Limit, IRL) of 2 µg/mL in urine has been established. On the basis of the here presented data a threshold or rather an IRL for theobromine in plasma of 0.3 µg/mL was proposed and was thereupon approved by the International Federation of Horseracing Authorities (IFHA). Official recommendations for reporting caffeine and theophylline is still lacking. The aim of the study was to investigate IRLs for theobromine in blood and for caffeine and theophylline in blood and urine. Therefore a set of 6 administrations were carried out including both single i.v. and single oral administrations of caffeine, theobromine and theophylline. Plasma and urine concentrations were determined using a validated liquid chromatography - tandem mass spectrometry (LC-MS/MS). Applying the Toutain model approach an effective plasma concentration (EPC) of caffeine was estimated at 3.05 µg/mL, irrelevant concentrations in blood (IPC) and urine (IUC) approached to 6 and 12 ng/mL, respectively. EPC of theobromine was calculated with 3.80 µg/mL, and irrelevant concentrations of theobromine were determined at 8 ng/mL in plasma and at 142 ng/mL in urine. Toutain modelling of the theophylline data produced an EPC, IPC and IUC of 3.20 µg/mL, 6 ng/mL and 75 ng/mL, respectively. The obtained irrelevant concentrations were used to postulate IRLs for theobromine in plasma and for caffeine and theophylline in plasma and urine. This article is protected by copyright. All rights reserved.