The objective of this work was to increase the nasal absorption of acyclovir by using absorption enhancers. Acyclovir was selected as a model drug. A rat in situ nasal perfusion technique was utilized in the investigation to examine the rate and extent of absorption of acyclovir. In vitro enzymatic drug degradation study was carried out with rat nasal washings. Various experimental conditions such as nasal perfusion rate, pH of the perfusion medium and concentrations of absorption enhancers such as sodium deoxycholate, hydroxypropyl β-cyclodextrin, sodium caprate, sodium tauroglycocholate and EDTA were optimized. Nasal absorption of acyclovir was pH dependent. Initial absorption rate constants were determined by the plot of log% remaining amount of drug in perfusate vs time. It was found maximum at pH 7.4 and decreased at lower and higher pH conditions. In in vitro enzymatic degradation study, no measurable degradation was observed during first week. The extent of drug absorption was increased via absorption enhancers. In vivo studies were carried out for the optimized formulation in rabbits and the pharmacokinetics parameters of nasal solution were compared with oral solution. Hydroxypropyl β-cyclodextrin appeared to be more effective for enhancing the nasal absorption of acyclovir than the other absorption enhancers. The order of increasing absorption of acyclovir caused by the enhancers was hydroxypropyl β-cyclodextrin>sodium deoxycholate>sodium caprate>sodium tauroglycocholate>EDTA. © 2004 Elsevier B.V. All rights reserved.