Abstract: Nowadays, sustainability and circular economy are two principles to be pursued in all fields. In road pavement engineering, they can be put into practice through the partial substitution of bitumen with industrial residues and by-products deriving from renewable materials. Within this framework, this paper presents an extensive investigation of the chemical, morphological and rheological properties of bio-binders obtained by mixing a conventional 50/70 bitumen with different percentages by weight (0, 5%, 10% and 15%) of a renewable bio-oil, generated as a residue in the processing of wood into pulp and paper. Results show that overall the bio-oil provides a softening effect, which, in terms of performance, leads to an improvement of the low-temperature behaviour and fatigue resistance with respect to the control bitumen, in spite of an increased tendency to permanent deformation. Although no chemical reaction appears to occur after blending, the peculiarities of the bio-oil affect the chemistry of the resulting bio-binders, whereas no phase separation is observed from the microscopic analysis. In addition, a Newtonian behaviour, an unchanged temperature susceptibility and a good fitting of 1S2P1D model to the rheological data are found, regardless of the bio-oil percentage considered. These promising outcomes suggest that such bio-binders can be favourably employed for several applications in road pavements.