A sensitive and simple method of determining the plasma levofloxacin (LFX, CAS 100986-85-4) concentrations in human volunteers by liquid-liquid extraction were developed and validated by using a high-performance liquid chromatography/diode array detector. The method was also applied to pharmacokinetic study of LFX. LFX was orally administered to 8 healthy male Korean volunteers at single lowest dose of 200 mg, compared to the published reports in which more than 500 mg of LFX was orally administered. LFX in human plasma was determined. The detection limit of LFX was $0.05;{mu}g/mL$. $C_{max}$ value was $2.48{pm}0.67;{mu}g/mL$. $AUC_{0{ o}24;hr$} and $AUC_{0{ o}{infty}}$ were $14.52{pm}3.35;{mu}g/mL$ and $16.00{pm}3.66;{mu}g{cdot}hr/mL$, respectively. The terminal half-life was $6.87{pm}0.46;hr$. Our pharmacokinetic parameters were very consistent with that previously reported, showing good correlation between LFX doses and AUC ($r^2=0.995$). This method can be useful for the pharmacokinetics and bioequivalence study with relatively low dose for reducing the main side effects of LFX.