The microstructure evolution and mechanical properties of hot-rolled (HR) 40CrNiMoA steel affected cyclic quenching treatment were investigated in this work. The results reveal that the average austenite grain size of 40CrNiMoA steel is refined and more uniformly distributed after cyclic quenching. With first pass of cyclic quenching (CQ1), the austenite grain size of 40CrNiMoA steel reduces from 9.14 μm(HR) to 4.51 μm(CQ1). Concurrently, the ultimate tensile strength and hardness experience a notable increase from 1081 MPa to 2314 MPa and 362HV to 620HV, respectively. With the decrease of grain size, the ultimate tensile strength and hardness of the sample increased significantly, and the elongation remained basically unchanged. The wear resistance ranking of the samples, from strong to weak, is CQ1 > CQ2 > CQ0 > HR. The 40CrNiMoA steel undergoes a single pass of cyclic quenching treatment, and considering economic factors, its mechanical properties and wear resistances undergo significant enhancement, thereby exerting a substantial impact on its overall performance.