Sensitive determination of ephedrine and norephedrine in human plasma samples using derivatization with 9-fluorenylmethyl chloroformate and liquid chromatography

A high-performance liquid chromatography procedure for the determination of ephedrine and norephedrine using fluorimetric detection in plasma samples is described. A double liquid-liquid extraction was performed, followed by derivatization with 9-fluorenylmethyl chloroformate. The extracts were chro matographed with a 5-mu m C-18 (150 x 4.6 mm I.D.) column using a mobile ph ase composed of acetonitrile and water (52:48; v/v). The excitation and emi ssion wavelengths were respectively 264 nm and 313 nm. Calibration curves w ere linear over the range 0 to 300 ng/ml for each analyte. The specificity of the method was demonstrated with several FMOC-reacting drugs. The limits of quantification are similar to those obtained with the reference method: 2 ng/ml for ephedrine and 5 ng/ml for norephedrine. This method has been s uccessfully applied to the determination of ephedrine and norephedrine plas ma levels after administration of low doses of ephedrine to healthy subject s. (C) 2000 Elsevier Science B.V. All rights reserved.