Simultaneous determination of ibuprofen and its metabolites in complex equine urine matrices by GC-EI-MS in excretion study in view of doping control

Emilia Waraksa; Marzena Wójtowicz-Zawadka; Dorota Kwiatkowska; Anna Jarek; Anna Małkowska; Robert Wrzesień; Jacek Namieśnik

doi:10.1016/j.jpba.2018.02.004

Simultaneous determination of ibuprofen and its metabolites in complex equine urine matrices by GC-EI-MS in excretion study in view of doping control

J Pharm Biomed Anal. 2018 Apr 15:152:279-288. doi: 10.1016/j.jpba.2018.02.004. Epub 2018 Feb 5.

Authors

Emilia Waraksa¹, Marzena Wójtowicz-Zawadka², Dorota Kwiatkowska³, Anna Jarek³, Anna Małkowska⁴, Robert Wrzesień⁵, Jacek Namieśnik⁶

Affiliations

¹ Gdańsk University of Technology, Faculty of Chemistry, Department of Analytical Chemistry, G. Narutowicz 11/12 Street, 80-233 Gdansk, Poland; Institute of Sport - National Research Institute, Trylogii 2/16 Street, 01-982 Warsaw, Poland.
² Institute of Sport - National Research Institute, Department of Anti-Doping Research, Trylogii 2/16 Street, 01-982 Warsaw, Poland. Electronic address: marzena.wojtowicz@insp.waw.pl.
³ Institute of Sport - National Research Institute, Department of Anti-Doping Research, Trylogii 2/16 Street, 01-982 Warsaw, Poland.
⁴ Medical University of Warsaw, Department of Toxicology, Banacha 1 Street, 02-097 Warsaw, Poland.
⁵ Medical University of Warsaw, Central Laboratory of Experimental Animal, Banacha 1B Street, 02-097 Warsaw, Poland.
⁶ Gdańsk University of Technology, Faculty of Chemistry, Department of Analytical Chemistry, G. Narutowicz 11/12 Street, 80-233 Gdansk, Poland.

PMID: 29453024
DOI: 10.1016/j.jpba.2018.02.004

Abstract

A novel assay for the simultaneous determination of ibuprofen (IBU) and its four probable metabolites, 1-hydroxyibuprofen (1-OH IBU), 2-hydroxyibuprofen (2-OH IBU), 3-hydroxyibuprofen (3-OH IBU) and carboxyibuprofen (CBX IBU) in equine urine samples with the application of Gas Chromatography-Electron Ionization-Mass Spectrometry (GC-EI-MS) has been developed and elaborated. The new approach for sample preparation including minimizing matrix effects by the application of weak cation exchange solid-phase extraction together with strong cation exchange solid-phase extraction has been applied. The GC-EI-MS method was validated to demonstrate specificity, matrix effect, linearity, limit of detection (LOD) and quantification (LOQ), precision, trueness, carry-over and stability by using the matrix-matched quality control samples. Additionally, extraction yield was evaluated. The assay achieved the LOQ of 1.75 μg mL^-1, 0.62 μg mL^-1, 4.15 μg mL^-1, 0.58 μg mL^-1 and 4.04 μg mL^-1 for IBU, 1-OH IBU, 2-OH IBU, 3-OH IBU and CBX IBU, respectively. The developed method has been successfully applied to the excretion study in horses, in which a single oral IBU dose was administered to twelve horses (mares and geldings) and equine urine samples were collected for 5 or 6 days after the drug administration. Data on the detection and determination of three IBU metabolites, 2-OH IBU, 3-OH IBU and CBX IBU in equine urine samples has been presented for the first time. The obtained results indicated the rapid excretion of IBU and its metabolites that were detectable only in the first day after the drug administration. IBU was mainly the most abundant compound detected in equine urine samples (with two exceptions in the case of samples collected from two horses, for which the highest instrumental responses were obtained for CBX IBU). The received results have indicated that two major IBU metabolites, CBX IBU and 2-OH IBU can be important markers for the IBU abuse in view of doping control in equestrian sports.

Keywords: Doping; Equine excretion study; GC-EI-MS analysis; Ibuprofen; Sport.

MeSH terms

Animals
Doping in Sports / methods
Gas Chromatography-Mass Spectrometry / methods
Horses
Ibuprofen / analogs & derivatives
Ibuprofen / metabolism*
Ibuprofen / urine*
Limit of Detection
Solid Phase Extraction / methods

Substances

hydroxyibuprofen
carboxyibuprofen
Ibuprofen