Bone regeneration may benefit from the local delivery of bone-promoting growth factors. In this view, we developed, optimized and evaluated in vivo a novel syringeable hydrogel based on chitosan, intended as a drug carrier. The effect of the carrier pH and nature on recombinant human Bone morphogenetic protein-2 (rhBMP-2) bioactivity was demonstrated. As for carrier pH, rhBMP-2 bone-forming ability decreased when shifting hydrogel pH from acidic to neutral conditions. Observed aggregation and conformational changes of rhBMP-2 in solution might explain the decrease of growth factor bioactivity. Although combining rhBMP-2 with a cross-linked hyaluronan hydrogel induced formation of higher amounts of bone, a more mature bone was observed with the chitosan hydrogel carrier. The addition of β-tricalcium phosphate to chitosan hydrogel increased rhBMP-2-induced bone formation. Finally, the sustained release of a platelet growth factor from chitosan hydrogel formulations demonstrated the potential of this approach as an alternative to exogenous growth factors.