Direct separation and quantitative determination of clenbuterol enantiomers by high performance liquid chromatography...

Enantiomers of clenbuterol were directly separated by a new high performance chromatographic method on Chirex 3005 column. Several parameters such as mobile phase composition, column temperature and flow rate were studied. Baseline enantioseparation was achieved, using the optimized mobile phase of n-hexane-1,2-dicholoethane-methanol (54:38:8, v/v/v) at 17 degrees C and 1.0 ml/min, with the separation factor (alpha) 1.43 and the resolution factor (R(S)) 1.81. The mechanism of separation was also discussed. Standard linear calibration cures were established for the R- and S-enantiomers, over the range of 26.1-1,045.8 and 5.7-229.6 nmol/ml, with the correlation coefficient of 0.9999 for both. The limits of detection were 0.47 and 1.04 nmol/ml for R- and S-enantiomers, respectively. Recovery and precision of the method were also evaluated, which had been successfully used to monitor and identify quantitatively the profile of the clenbuterol enantiomers in human serum.