A potential hemostatic chitosan/gelatin cryogel impregnated with Verbascum thapsus leaf extract for noncompressible hemorrhage management

Abstract In this study, we prepared a series of chitosan/gelatin (CS/GEL) cryogels containing Verbascum thapsus ( V. thapsus ) leaf extract and identified a lead formulation for noncompressible hemorrhage (NCH). Cryogels with average pore diameters ranging from 225 to 478 µm were fabricated through cryogelation at various CS/GEL ratios. C15 was chosen as the base scaffold due to its homogeneous pore distribution, with a pore size coefficient of variation (CV) of approximately 0.22. Extract loading was 1%, 5%, 10%, and 20% w/v. Functional porosity was reported by the relative accessible void index (RAVI). In PBS, the values relative to neat C15 were 1.00, 0.27, 0.20, 0.13, and 0.09 for concentrations of 0%, 1%, 5%, 10%, and 20% w/v, respectively. In citrated blood, the series was 1.00, 0.29, 0.12, 0.14, and 0.09. After loading, equilibrium swelling decreased and the compressive modulus increased, consistent with partial pore filling in a fixed network. The cryogels maintained an interconnected macroporous network and showed swelling from 300% to 3600% in blood and PBS. Antibacterial activity reached 89% inhibition, and cell viability remained above 80%. Hemolysis was low and within acceptance limits. Clotting improved in whole blood as the blood clotting index decreased from 11.9 to 6.5, and the clotting time was approximately 6 min. The 5% w/v group provided the optimal balance of clotting, antibacterial effects, and biocompatibility. This study presents a novel hemostatic CS/GEL cryogel containing V. thapsus leaf extract that holds strong potential for future applications in NCH management.