Enatiomeric determination of tramadol and O-desmethyltramadol in human urine by gas chromatography–mass spectrometry

A GC–MS assay for stereoselective determination of tramadol and its pharmacologically active phase I metabolite O-desmethyltramadol in human urine was developed. Nefopam was used as internal standard. The method involves a simple solid phase extraction with chiral analysis by gas chromatography–electron ionization mass spectrometry using m/z 263; 58, 249; 58, and 179; 58 for the determination of concentration of tramadol, O-desmethyltramadol and internal standard, respectively. Chromatography was performed on a Rt-βDEXcst column containing alkylated beta-cyclodextrins as a chiral selector. The calibration curves were linear in the concentration range 0.1–20 μg/mL (R2 ≥ 0.998). Intra-day accuracies ranged between 97.2–104.9%, 96.1–103.2%, and 97.3–102.8% at the lower, intermediate, and high concentration for all analytes, respectively. Inter-day accuracies ranged between 95.2–105.7%, 99.1–105.2%, and 96.5–101.2% at the lower, intermediate, and high concentration for all analytes, respectively. This method was successfully used to determine the concentration of enantiomers of T and ODT in a pharmacogenetic study.