The primary focus of this study is to assess the stripping resistance of bitumen when partially replaced with rice-straw biomass-based lignin from bio-ethanol industry residue. Moreover, the study examines the relationship between the chemical and physicochemical (surface free energy or SFE) properties of lignin-substituted bitumen blends and their stripping potential. Two distinct lignin materials were examined: fermentation residue (FRL), a by-product directly obtained from the bio-ethanol industry, and Isolated Lignin (IL), which was extracted from FRL through an alkali-acid precipitation process. These materials were partially substituted at dosages of 5%, 10%, and 15% by weight of bitumen. Test results indicated that the addition of lignin to bitumen led to a change in SFE, with an increase in the polar component and a reduction in the dispersive component. The rolling bottle test revealed that bitumen blends containing IL and FRL displayed superior stripping resistance compared to the base bitumen, retaining more coating after testing. This enhancement in performance was attributed to the increased heteroatom content [sum of nitrogen (N), oxygen (O), and sulfur (S)] and higher IS═O and IC═O indices upon adding the lignin materials. While the energy ratio obtained from the SFE alone did not entirely explain the stripping resistance of lignin-substituted bitumen, the presence of heteroatoms like N, O, and S, and the presence of silicon (Si) and associated functional groups played a substantial role in the chemical adhesion of these bitumen blends to aggregates. In conclusion, the incorporation of IL and FRL lignin materials holds promise for enhancing the stripping resistance of base bitumen. [ABSTRACT FROM AUTHOR]