Development of an efficient hemostatic material based on cuttlefish ink nanoparticles loaded in cuttlebone biocomposite

Traumatic hemorrhage is one of the main causes of mortality in civilian and military accidents. This study aimed to evaluate the effectiveness of cuttlefish bone (cuttlebone, CB) and CB loaded with cuttlefish ink (CB-CFI) nanoparticles for hemorrhage control. CB and CB-CFI were prepared and characterized using different methods. The hemostasis behavior of constructed biocomposites was investigated in vitro and in vivo using a rat model. Results showed that CFI nanoparticles (NPs) are uniformly dispersed throughout the CB surface. CB-CFI₁₀ (10 mg CFI in 1.0 g of CB) showed the best blood clotting performance in both in vitro and in vivo tests. In vitro findings revealed that the blood clotting time of CB, CFI, and CB-CFI₁₀ was found to be 275.4 ± 12.4 s, 229.9 ± 19.9 s, and 144.0 ± 17.5 s, respectively. The bleeding time in rat liver injury treated with CB, CFI, and CB-CFI₁₀ was 158.1 ± 9.2 s, 114.0 ± 5.7 s, and 46.8 ± 2.7 s, respectively. CB-CFI₁₀ composite resulted in more reduction of aPTT (11.31 ± 1.51 s) in comparison with CB (17.34 ± 2.12 s) and CFI (16.79 ± 1.46 s) (p < 0.05). Furthermore, CB and CB-CFI₁₀ exhibited excellent hemocompatibility. The CB and CB-CFI did not show any cytotoxicity on human foreskin fibroblast (HFF) cells. The CB-CFI has a negative surface charge and may activate coagulation factors through direct contact with their components, including CaCO₃, chitin, and CFI-NPs with blood. Thus, the superior hemostatic potential, low cost, abundant, simple, and time-saving preparation process make CB-CFI a very favorable hemostatic material for traumatic bleeding control in clinical applications.