Burn injury is the main cause more than 70% scar formation for patients. In previous study has been reported fetal stem cells conditioned medium significantly promoted skin regeneration compared to mesenchymal stromal cells (MSCs). However, little is known about the functionality of key factors and mechanisms through the analysis of conditioned medium from fetal stem cells. In this study, we aimed to fabricate a biologically functional human fetal cartilage derived progenitor cells-conditioned medium (hFCPCs-CM) that can improve wound healing and minimize scar formation on a burn skin injury model in rats. The hFCPCs-CM was prepared by a series of processes including a 3D culture of hFCPCs, a 3.5 kDa cutoff dialysis membrane of conditioned medium and lyophilization of extracellular matrix. The final lyophilized hFCPCs-CM did not contain the cellular content and contained large amount of total protein, collagen, glycosaminoglycans and soluble proteins of IGFBP-2, IGFBP-6, HGF, VEGF, TGF β3 and M-CSF compared to human bone marrow-derived MSCs-conditioned medium (hBMSCs-CM). It also contained 2 specific proteins of collagen alpha-1(XIV) chain and collagen alpha-2(I) chain. The acceleration effect of hFCPCs-CM on wound healing was more prominent than hBMSCs-CM, in enhancing the viability, proliferation and migration of fibroblast and polarization of M2 macrophages in vitro. The therapeutic potential of the hFCPCs-CM was also observed better than that of hBMSCs-CM in a burn skin injury model in rats by efficiently accelerating wound healing and minimize scar formation. Our results show that the hFCPCs-CM could be a potential therapeutic approach to promote wound healing process.