Three bio-based compounds have been investigated as emulsifiers and stabilisers for bitumen emulsion preparation. The emulsifiers used are biopolymer, native nanoparticles and nanoparticles modified to be more hydrophobic. The comparison was made with a usual cationic surfactant for bitumen emulsion, used to manufacture the emulsion. Bitumen-in-water emulsions with a bitumen content of 60% were further obtained with biopolymers and nanoparticles. These emulsions were characterised and compared to a classical bitumen emulsion with amine-type surfactant. Cold mix asphalt specimens were successfully manufactured, moulded and used to evaluate the water recovery and stripping resistance. Different behaviours have been observed and interpreted. They are strongly linked to the emulsion formulation.