Development and validation of an LC-MS/MS method for the quantification of ephedrines in urine

The objective of this study was to develop a simple and robust LC-MS/MS method for the quantification of ephedrine type substances in urine. Sample preparation consisted of a 10-fold dilution step of the samples into the internal standard solution (ephedrine-d(3), 4 mu g/mL in water). Baseline separation of the diastereoisomers norpseudoephedrine-norephedrine and ephedrine-pseudoephedrine was performed On a C8-column using isocratic conditions followed by positive electrospray ionisation and tandem mass spectrometric detection. The mobile phase consisted of 98/2 (H(2)O/ACN) containing 0.1% HAc and 0.01% TFA. Calibration curves were constructed between 2.5 and 10 mu g/mL for norephedrine and norpseudoephedrine and 5 and 20 mu g/mL for ephedrine, pseudoephedrine and methylephedrine. The bias ranged from -5.5 to 12% for norephedrine, -4.1 to 8.0% for norpseudoephedrine, 0.3 to 2.1% for ephedrine,1.6 to 2.6% for pseudoephedrine and 2.9 to 5.0% for methylephedrine. Precision of the method varied between 2.8 and 10.4% for all compounds and the matrix effect was less than 15%. The applicability of the method has been checked by the analysis of 40 urine samples. The results were compared with those obtained with the common GC-NPD method. Results show a good correlation between both methods with correlation coefficients higher than 0.95 for all analytes. (C) 2009 Elsevier B.V. All rights reserved.