The invention discloses a thiamphenicol molecularly imprinted electrochemical sensor and a preparation method and application thereof. A porous graphene-molybdenum disulfide nano flower-shaped compound is prepared through a hydrothermal one-step method, and the compound is dropped on the surface of an L-type glassy carbon electrode; aminated multi-walled carbon nanotubes and porous Pt-Pd nanoparticles are dispersed so that the imprinting surface area of the electrode can be increased and stability of a modification interface can be enhanced, and accordingly a three-dimensional porous imprinting substrate is obtained; thiamphenicol serves as template molecules, 1,2-diaminobenzene serves as functional monomers, and an imprinted membrane is prepared through cyclic voltammetry; ascorbic acid serves as a photoelectric chemical probe, and the electrochemical sensor for detecting thiamphenicol is built. The sensor has good thiamphenicol responsiveness, the linear range of the sensor is 1.0*10<-9>-3.5*<-6> mol L<-1>, and the detection lower limit is 5.0*10<-9> mol L<-1>. The sensor can be used for detecting thiamphenicol in meat samples and feed samples.