The precise measurement of three-dimensional stress in the rotor is a key means to ensure the assembly accuracy and operational safety of aero-engines. The unclear coupling influence mechanism of different directions of stress on ultrasonic propagation makes it challenging to establish a three-dimensional stress measurement model. This paper proposes a three-dimensional assembly stress measurement method for aero-engine rotors based on ultrasonic propagation time difference decoupling. The three-dimensional assembly stress is converted into the stress component parallel and perpendicular to the ultrasonic wave propagation direction in this method. The acoustoelastic equation including three-dimensional stress parameters is established based on the function model of different direction stress and ultrasonic wave propagation velocity. An innovative three-transducer stress measurement scheme is designed, which integrates ultrasonic wave transmitting and receiving transducers and time difference feedback transducers. The acoustoelastic equations in three propagation directions are constructed by rotating the transducer group to change the propagation direction of ultrasonic waves. Finally, the acoustoelastic equations are solved simultaneously to decouple the ultrasonic propagation time difference, thus achieving a three-dimensional assembly stress measurement of the aero-engine rotor. The experimental results show that the maximum deviation between the measured values and the simulation results in the x ,y, and z directions is 15.885 MPa, 14.113 MPa, and 17.093 MPa, respectively. This study provides effective theoretical and technical support for precisely measuring three-dimensional assembly stress in high-end equipment represented by aero-engines.