Chitosan was selectively N-acylated with acetic, propionic and hexanoic anhydrides under homogeneous condition to prepare N-acetyl chitosan (NACS), N-propionyl chitosan (NPCS) and N-hexanoyl chitosan (NHCS), respectively. NACSs with different N-acetylation degrees were obtained by controlling the degree of N-acetylation. The chemical structures of N-acylated chitosans including degree of deacetylation (DD), weight-average molecular weight (M-w), radius of gyration ((S-2)(Z)(1/2)) and crystal structure were studied by FTIR, GPC-LLS and X-ray diffraction techniques. Aggregation behavior of N-acylated chitosan was investigated by rheometer. Intramolecular aggregation of NPCS and NACS was stronger with NPCS stronger than NACS. The effect of concentration of polymer, concentration of salt and temperature on self-aggregation of NACS and NPCS was investigated. Hydrophobic interaction of N-acylated chitosan substituted with longer acyl chains was stronger. With moderate DD, intramolecular aggregation occurs predominantly. In vitro antibacterial activity test of N-acylated chitosans was evaluated against two Gram-positive bacteria and two Gram-negative bacteria. Relative inhibition time (RIT) of NHCS with concentration of 1 mg/ml against Escherichia coli and Pseudomonas aeruginosa was more than 2-6 times longer than that of NACS and NPCS. N-acylated chitosan with lower DD had inhibitory effect on the growth of bacteria than that with moderate DD. The results showed that intermolecular aggregation characteristic of N-acetylated chitosans with low DD may help in forming bridge to interact with bacterial cell. (c) 2007 Elsevier Ltd. All rights reserved.