Hemostasis plays a critical role in the early stage of wound healing, especially in acute wounds which can significantly improve the survival of patients. Based on the excellent biocompatibility of natural biomaterials, in this study, we prepared a series of novel hemostatic sponges by using tilapia skin, a marine biological resource, and extracting tilapia skin-derived gelatin, collagen, and acellular dermal matrix through five different methods. Using in vitro sheep blood and in vivo rat liver hemorrhage models, we found that tilapia skin sponges had excellent coagulation and hemostatic abilities. Among them, the collagen sponge exhibited optimal hemostasis performance because it could accelerate clotting by binding to the specific sites of blood cells and platelets. Furthermore, the sponges' porous structure enhanced the capability to absorb blood, thus effectively promoting hemostasis. In summary, we reported an efficient and convenient method to prepare marine biological resources into sponges, which provided a novel class of alternatives for hemostasis in acute wounds with broad application prospects.