• Chitin nanofibers reduced the blood-clotting time and increased clot firmness. • The morphology of chitin nanofibers impacts their hemostatic properties. • The positively-charged surface of chitin stimulates contact activation pathways. Recently, negatively-charged nanofibers made of macro-scale carbohydrate polymers, such as cellulose, have been investigated for hemostatic applications. It was demonstrated that nanostructural features including specific surface area and aspect ratio can be optimized to increase the interaction of blood components with nanofibers. However, the influence of nanofiber’s surface charge is unknown. In this paper, positively-charged nanofibers were produced from shrimp chitin using a ball-milling technique. In-vitro thromboelastometry studies revealed that nanofibers morphology has much stronger influence than the surface charge on their hemostatic efficacy. Consequently, with optimizing the morphology, clotting time was reduced by 70 ± 2%, and clot firmness was increased by 12 ± 1% comparable to commercial hemostats.