The effect of circadian rhythm on gentamicin pharmacokinetics was studied in rabbits who took a single intravenous 2 mg/kg dose of gentamicin at 09:00 in the morning (a.m.) and 22:00 in the evening (p.m.). A significant circadian rhythm of pharmacokinetic parameters as a function of time of day was noted in rabbits, showing lower total body clearance $CL_t$ and higher serum area under the curve (AUC) when given in the evening. The half-life $t_{1/2}$ was shorter in the morning $(3.88pm0.62h)$ when compared to the evening $(4.76pm0.75;h)$. The AUC was greater in the evening $(25.92pm3.49;{mu}g/ml{cdot}hr)$ than that in the morning $(22.42pm3.42;{mu}g/ml{cdot}hr)$, most likely because the CLt was significantly higher when gentamicin was given in the morning $(0.18pm0.28;ml/hr)$ versus in the evening $(0.15pm0.26;ml/hr)$. The $t_{1/2}$ of gentamicin in the evening was increased significantly(p<0.05) compared to those of gentamicin in the morning. It is reasonable to consider individual circadian rhythm for effective dosage regimen of gentamicin in clinical chronotherapeutics.