A novel protocol for preparation of highly magnetic chitosan particles, with a coercive force as high as 3500 Oe, has been developed. The surface of the particles was functionalized with aldehyde groups to facilitate the attachment of affinity ligands. The optimum conditions for the preparation of highly magnetic chitosan particles and immobilization of trypsin on magnetic particles were obtained; these particles were then used for affinity purification of aprotinin, and the conditions of affinity purification are discussed in detail. The proposed method was successfully applied to the affinity purification of aprotinin. (C) 2003 Society of Chemical Industry.