A rapid and sensitive method for the quantitative analysis of ibuprofen and its metabolites in equine urine samples by gas chromatography with tandem mass spectrometry.

Ibuprofen is widely used in human and veterinary medicine for the treatment of chronic pain as well as rheumatic and musculoskeletal disorders. However, the analgesic and anti-inflammatory properties of Ibuprofen have contributed to frequent drug abuse in equestrian sports. A sensitive and rapid gas chromatography with tandem mass spectrometry based method with a simple liquid-liquid extraction and derivatization requiring 200 μL volume of sample and 2 mL of extraction solvent for the simultaneous determination of ibuprofen and its metabolites was developed. The proposed procedure was optimized and validated according to the principles for bioanalytical methods. The assay achieved satisfactory validation parameters, namely, recovery (92.2-105%), interday accuracy (92.5-106%), and precision (0.3-4.4%) for all investigated compounds as well as limits of quantification of 50 ng/mL for ibuprofen, 2-hydroxyibuprofen, and carboxyibuprofen, 25 ng/mL for 1-hydroxyibuprofen and 100 ng/mL for 3-hydroxyibuprofen. The applicability of the method was evaluated by the analysis of five real urine samples collected from different horses after drug administration. In view of the low limits of quantification, high selectivity, repeatability, and recovery, the procedure can be utilized for laboratory applications, including the control of ibuprofen abuse in equestrian sports for anti-doping purposes and drug/pharmaceutical mentality investigations.