Nanostructure and Crystallinity Govern the Rheology and Printability of Cellulose-Based Bioinks

Despite growing interest in bioinks containing nanocellulose, the mechanistic contribution of cellulose crystallinity to rheology, microarchitecture, and printability remains unclear. Here, we directly compared cellulose nanocrystal (CNC)- and cellulose nanofiber (CNF)-based fibrin (FIB) bioinks to isolate crystallinity-associated effects. Cell-free formulations (CNC-FIB, CNF-FIB, and CNC-CNF-FIB) were characterized by attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR), X-ray diffraction (XRD), oscillatory/steady rheology, extrusion printability, and scanning electron microscopy (SEM)-based image analysis. XRD showed higher cellulose-region crystallinity for CNC than CNF (95.20 ± 0.88% vs 72.30 ± 7.04%, p ≤ 0.01), consistent with FTIR total crystallinity index values (2.16 ± 0.09 vs 1.04 ± 0.11, p ≤ 0.001). Power-law analysis revealed distinct flow regimes: CNC-FIB exhibited low consistency and favorable flow (K = 2.354 ± 0.179; n = 0.672 ± 0.029), CNC-CNF-FIB was intermediate (K = 5.887 ± 0.168; n = 0.545 ± 0.011), and CNF-FIB showed markedly high flow resistance (K = 191.685 ± 17.296; n = -0.063 ± 0.008). At 10 rad·s^-1, tanδ further separated formulations (3.59 ± 0.06, 1.63 ± 0.15, and 0.141 ± 0.003 for CNC-FIB, CNC-CNF-FIB, and CNF-FIB, respectively), matching printability (0.946 ± 0.011, 0.897 ± 0.0213, and 0.284 ± 0.227). SEM showed no significant differences in 2D porosity; however, anisotropy increased in cellulose-containing groups. Overall, the data indicate that bioink performance is governed by the interplay of crystalline order and network topology. CNC improves processability and shape fidelity, while CNF enhances stiffness but compromises flow. Combined CNC/CNF formulations provide a tunable compromise for mechanically demanding bioprinting applications, with potential relevance to bone- and cartilage-oriented constructs.