A stereospecific HPLC method has been developed for the resolution of the enantiomers of salbutamol in human urine. After solid-phase extraction and derivatization with (S)-(-)-${alpha}$-methyl-benzyl isocyanate, the diastereomeric derivatives were resolved $(R_s=1.59)$ on $5{mu}M$ octadecylsilan column using 47% methanol as a mobile phase with fluorescence detection. The detection limit of each enantiomer was 10 ng/ml (S/N=3).