The biocompatibility and hemostatic efficacy of silk fibroin nanofibrils fabricated by Shear-Induced phase separation process

The objectives of this study were to investigate the biocompatibility and hemostatic efficacy of silk fibroin nanofibrils (SFNFs). Unlike a conventional dialysis/freeze-drying process, SFNFs were prepared using a novel shear-induced phase-separation method. The morphology, particle size distribution, protein secondary structures, and crystalline structures of the resulting SFNFs were respectively investigated by scanning electron microscopy (SEM), transmission electron microscopy (TEM), laser diffraction (LD), Fourier-transform infrared (FTIR) spectroscopy, and x-ray diffraction (XRD) deconvolution analyses. Polar and non-polar extractants of SFNFs under a bath ratio of 0.2 g/mL were collected after individually soaking them into 0.9% normal saline and cottonseed oil. In vitro hemolysis ratios, biocompatibility, and sensitization tests were carried out. Then, the hemostatic efficacy was assayed using a rat liver punch biopsy model. Compared to dialyzed/freeze-dried SF, the harvested SFNFs revealed high surface area in morphology. The median particle size distribution, D50, was 294 μm for SFNFs. Shear-induced hydrogen bonding, crystallization, and phase separation tended to give the SFNFs a high crystalline index (CI) and degree of crystallinity (DC). The hemolysis ratio of SFNFs was 0.86 ± 0.40%, and it was classified as being non-hemolytic (0%∼2%). The test results of toxicity and skin sensitization were both negative. The hemostasis time for SFNFs (2.8 ± 0.6 min) was statistically less than that of oxidized cellulose (4.3 ± 0.9 min) (p < 0.05). The high biocompatibility and hemostatic efficacy of SFNFs revealed their potential as an alternative hemostatic agent.