Preparation of Short Collagen Nanofibers for Injectable Hydrogels: Comparative Assessment of Fragmentation Methods, Physicomechanical Properties, and Biocompatibility

Abstract Collagen nanofibers can be employed in hydrogels to create injectable nanocomposite hydrogels, mimicking the fibrous architecture of the natural extracellular matrix (ECM). As long continuous electrospun collagen nanofibers are not applicable, fragmentation is inevitable to obtain injectable hydrogels with a fine viscosity. Here, four methods: hand grinding (HG), homogenizer (HM), mixer milling (MM), and ultrasonication (UH) are used to disintegrate and shorten collagen nanofiber mats before incorporation into an injectable hyaluronic acid hydrogel as a matrix. The Length‐to‐diameter (L/d) ratio and morphology of fragmented collagen are compared by SEM. The injection force, mechanical properties, and cell viability of the selected collagen‐incorporated hydrogels are also evaluated. UH emerges as the most effective method, yielding the highest L/d ratio of 46 and a notable compressive modulus of 8.7 ± 0.92 kPa. Assessment of the in vitro cell viability of the encapsulated chondrocytes in the collagen‐incorporated hydrogels demonstrates good biocompatibility, and hydrogels containing UH short nanofiber, in particular, show an increase in cell proliferation. This work indicates how collagen mats can be effectively broken down and combined with injectable hydrogels to enhance both their mechanical behavior and biocompatibility.