Self-crosslinked gelatin-oxidized hyaluronic acid/hydroxyapatite(GEL-OHA/HAp) composite bone substitute was successfully prepared with a novel in situ precipitation method without using any toxic chemical cross-linkers,and its characterizations,including elemental composition,surface morphology,crystallinity,and structure of crystalline phase,were carried out. In order to evaluate its corresponding performances,comparisons with glutaraldehyde crosslinked gelatin/hydroxyapatite(GEL-Glu/HAp) composite were made in detail. The results indicate that nano-HAp crystallites are homogeneously dispersed in both GEL-OHA/HAp and GEL-Glu/HAp composites,and the HAp crystallites in the former take larger particle size than those in the latter. Mechanical property tests show the acceptable mechanical strength at high strain of GEL-OHA/HAp composite. Study of in vitro degradation and swelling demonstrate that the two composites have similar degradation rate and water absorption capability.By in vitro cell culture,it has been found out that the cells on the GEL-OHA/HAp composite show higher proliferative potential than the cells on the GEL-Glu/HAp composite. Compared with GEL-Glu/HAp composite,GEL-OHA/HAp composite provides an excellent strategy for preparation of non-toxic bone substitute with acceptable corresponding properties. Self-crosslinked gelatin-oxidized hyaluronic acid / hydroxyapatite (GEL-OHA / HAp) composite bone substitute was successfully prepared with a novel in situ precipitation method without using any toxic chemical cross-linkers, and its characterizations, including elemental composition, surface morphology, crystallinity, and structure of crystalline phase, were carried out. In order to evaluate its corresponding performances, comparisons with glutaraldehyde crosslinked gelatin / hydroxyapatite (GEL-Glu / HAp) composite were made in detail. The results indicate that nano-HAp crystallites are homogeneously dispersed in both GEL-OHA / HAp and GEL-Glu / HAp composites, and the HAp crystallites in the former take larger particle size than those in the latter. Mechanical property tests show the acceptable mechanical strength at high strain of GEL-OHA / HAp composite. Study of in vitro degradation and swelling demonstrated that the two composites have similar degradation rate and water absorption capability.By in vitro cell culture, it has been found out that the cells on the GEL-OHA / HAp composite show higher proliferative potential than the cells on the GEL-Glu / HAp composite. Compared with GEL-Glu / HAp composite, GEL-OHA / HAp composite provides an excellent strategy for preparation of non-toxic bone substitute with acceptable corresponding properties.