Objective To study the role of azithromycin niosomes (AM-NS) in promoting the intestinal absorption of AM in rats. Methods Six healthy male rates were randomly divided into two groups. Single-pass intestinal perfusion was employed to investigate the intestinal absorption of the duodenum, jejunum, ileum and colon when the perfusion rate was 0.2 mL/min and period was 1 h. The concentrations of the drugs were determined by HPLC; the drug absorption rate constant (K_a) and the intestinal apparent permeation coefficients(P_eff) were calculated. Results The absorption K_a of AM-NS in the duodenum, jejunum, ileum and colon were 2.41, 2.35, 2.54 and 2.95 folds that of the free AM, and the P_eff were 2.34, 1.47, 2.20, and 1.40 folds that of the free AM, respectively, with significant differences found between the two groups for both K_a and P_eff (P<0.05). Conclusion AM-NS can significantly improve the absorption of azithromycin in the intestine of rats.