Single-run chemo- and enantio-selective high-performance liquid chromatography separation of tramadol and its principal metabolite, O-desmethyltramadol, using a chlorinated immobilized amylose-based chiral stationary phase under multimodal elution conditions

A direct enantio- and chemo-selective high-performance liquid chromatography method was developed for separating the enantiomers of the pain medicine tramadol and its O-desmethyl active metabolite in a single run. The simultaneous separation was achieved on the immobilized-type amylose tris(3-chloro-5-methylphenylcarbamate) chiral stationary phase. The method was optimized in normal-phase and reversed-phase using ethanol as an organic modifier. It was demonstrated that ethanol is a valid alternative to the predominant high-performance liquid chromatography solvents in use for the preparation of reversed-phase eluents such as methanol and acetonitrile. With the green mobile phase ethanol-water-diethylamine, 80:20:0.1 (v/v/v) the limits of quantification for the (+)/(-)-enantiomers of tramadol and its O-desmethyl active metabolite were 1.14/1.16 and 1.33/1.40 μg mL–1, respectively.