Hyaluronic acid (HA) was chemically crosslinked with a water-soluble carbodiimide (WSC) to produce low-water-content films when brought into contact with water. The crosslinking reaction was performed in two different ways; one was by using HA films and the other by casting HA solutions. Both methods produced water-insoluble HA films. The lowest water content of the crosslinked HA films subjected to swelling with water was 60 wt % at 37 degrees C, which was lower than any reported values. Infrared spectra of the crosslinked films suggested that intermolecular formation of ester bonds between the hydroxyl and carboxyl groups belonging to different polysaccharide molecules led to crosslinking. For comparison, pectin which possesses hydroxyl and carboxyl groups in one molecule, similar to HA, was subjected to crosslinking with WSC. The finding on pectin also supported ester formation between different polysaccharide molecules. The crosslinking of HA film with WSC in the presence of L-lysine methyl ester prolonged the in vivo degradation of HA film, probably because of amide bond formation as the crosslink.