Abstract An increase in the use of biomaterials has been noted in recent years because of several impacts caused by human activities, especially for engineering and paving industry benefits. Several renewable resources, such as a nonfood source of soybean oil, have been successfully tested on modification of asphalt binders. However, their impact on the pavement life cycle is still unknown. Thereby, the objective of this research was to investigate soybean oil as a new, green supply for the maintenance of flexible pavements. The experimental plan consisted of the construction of field test sections through the application of sub-epoxidized soybean oil (SESO) over a flexible pavement surface. Rates of 0.1 and 0.3 L/m2 were chosen, and the main tests intended for study were functional procedures including the British pendulum and sand patch tests on the pavement surface, which were complemented with the indirect tensile strength test for mechanical analysis. The change in the chemical composition of the recovered binder was analyzed by thin layer chromatography and Fourier-transform infrared spectroscopy. The rheological properties were analyzed by frequency sweep, linear amplitude sweep, and multiple stress creep and recovery, and the tests were carried out with a dynamic shear rheometer. The effects of the SESO bio-emulsion demonstrated a rejuvenating activity on the properties of the aged binder, with mechanical and rheological improvements for both analyzed rates. A further investigation is indicated to evaluate the influence of time on the modification studied in this work. Such testing could ensure the use of SESO bio-emulsion as an environmentally friendly alternative for the maintenance of flexible pavements.