The invention discloses a determination method of a bridge influence line when a vehicle passes at a non-uniform speed. The determination method of a bridge influence line when a vehicle passes at a non-uniform speed includes the steps: 1) acquiring response vectors of discrete measuring points of a determination influence line as needed; 2) collecting the real-time axle positional information when the vehicle passes a bridge at a non-uniform speed; 3) determining the space between the discrete measuring points of the determination influence line as needed; 4) establishing a position solutionequation of the corresponding set axle every time when collecting response, obtaining the position vector of the corrected set axle, and constructing an axle position matrix; 5) based on the axle position matrix in the step 4), establishing a vehicle load matrix; and 6) based on the response vector and the vehicle load matrix, establishing an equation, and solving the equation to obtain the bridgeinfluence line vector. The determination method of a bridge influence line when a vehicle passes at a non-uniform speed solves the problem that a traditional determination method cannot guarantee that the vehicle can pass the bridge at a uniform speed in the actual test, and solves the ill-posed problem of the bridge influence line solution equation, thus obtaining a more accurate bridge influence line.