We reported a new approach for development of lignin-based carbon nanofibers (LCNFs) from two lignin-derived bio-oil (LB) obtained from catalytic depolymerization reaction combined with the organic solvent gradient separation process (including ethanol-soluble LB (EL) and tetrahydrofuran-soluble LB (TL)). This approach was particularly unique and translatable as it uses small molecule TL with high reactivity and low heterogeneity obtained via stalk lignin depolymerization to produce good morphologies and mechanical performances of LCNFs mixed with polyacrylonitrile under 85% high replacement rates. We first detailed characterization studies such as 2D-HSQC NMR, P-31 NMR, FTIR, GPC, TGA and DSC were conducted to understand the properties of EL and IL. Then, the goal of the influence of EL and TL on the LCNFs morphologies and mechanical properties were accomplished by using SEM, XRD, Raman and material testing machine. As compared with EL, the TL had excellent linear structure with relatively less total phenolic-OH groups and heterogeneity, which could improve lignin structure with better alignment along PAN, and thus increased the lignin replacement rates and mechanical performances of LCNFs. (C) 2020 Elsevier B.V. All rights reserved.