The invention provides a regulating method of the reverse-running electricity generation of a water pump of a pump station, which relates to the technical field of pump stations. The regulating method comprises the following steps of: (1) starting the water pump to run in a no-load mode by a frequency conversion device at 40Hz frequency corresponding rotating speed and absorbing electric energy from an electric network by a water-pump motor through the frequency conversion device at the moment; (2) starting a water discharging gate, running a water pump and a water turbine in the same direction to enter an electricity generation state from an electric state and stabilizing electricity generation rotating speed in the 40Hz frequency corresponding rotating speed, at the moment, the water pump works in an electricity generator state under a water head, and energy flows into the electric network through the frequency conversion device; and (3) searching the optimal running electricity generation rotating speed of the water turbine, running the electricity generator at the optimal electricity generation rotating speed and obtaining optimal electricity generation efficiency. The invention can improve electricity generation quantity of 50 percent and ensure that the reverse-running electricity generation possibility of the water pump which can not run under an electricity generation work condition originally is increased by regulating the electricity generation running frequency.