To enhance the adhesion of bio-asphalt to aggregates and improve its practical application in engineering, this study investigates the effects of diatomite modification on bio-asphalt. The physical properties of diatomite-modified bio-asphalt are evaluated using standard index tests and rotational viscosity experiments. Adhesion to limestone is evaluated using an improved boiling method, Based on surface free energy, contact angles between the diatomite-modified bio-asphalt and test liquids are measured by the lying drop method, enabling the calculation of surface energy, cohesion work, adhesion work and spalling work. The results show that diatomite improves the physical properties of bio-asphalt, increases the softening point, reducces needle penetration, initially decreases ductility, and then slightly increases. Viscosity increases with diatomite content but stabilizes when the content exceeds 17%. When diatomite content is 17%, the asphalt quality loss during the boiling test is minimized, showing a decrease of 36.44% compared with unmodified bio-asphalt. Additionally, diatomite significantly improves the surface energy, adhesion and cohesion of bio-asphalt, and reduces spalling work. The dimensionless energy parameter (ER) reaches its peak at a diatomite content of 17%.