Crosslinking of hyaluronic acid with glutaraldehyde

Hyaluronic acid (HA) was chemically crosslinked with glutaraldehyde (GA) to produce water-insoluble films having low water contents when brought into contact with water. The crosslinking reaction was performed using uncrosslinked HA films in acetone–water mixtures. This method could produce water-insoluble HA films with water contents as low as 60 wt % when subjected to swelling with phosphate-buffered saline of pH 7.4 at 37°C. This 60 wt % water content was lower than any values for HA ever reported. There was an optimal HCl concentration around 0.01N for the HA crosslinking with GA in acetone—water mixtures. To get information on the crosslinking mechanism, alginic acid, which possesses hydroxyl and carboxyl groups in one molecule, similar to HA, and poly(vinyl alcohol) (PVA) and amylopectin, which possess only hydroxyl groups, were subjected to crosslinking with GA. PVA and amylopectin were also found to become water-insoluble after reaction with GA. On the basis of the infrared spectra of these crosslinked films, it was concluded that intermolecular formation of hemiacetal bonds with GA between the hydroxyl groups belonging to different HA molecules led to crosslinking.