The aim of this paper is to investigate the catalytic activity, the mechanism on degrading tetracycline (TC) with co-doped biochar modifying with non-metallic heteroatoms and transition metal oxides in a persulfate (PS) system. To this end, we ingeniously prepared nitrogen and copper oxide co-doped biochar (CNB-500) and activated it with PS to develop a CNB-500-PS system that can efficiently degrade TC. Owing to constructing a multi-pathway electron transfer channel, the non-radical interaction-electron transfer in the system was enhanced; the surface charge distribution of the catalyst was broken, and its adsorption with dissolved oxygen in water was regulated towards induce the generation of singlet oxygen (1O2); the mechanism on radical-non-radical synergism was elucidated, and the possible degradation pathways for TC were speculated. Therefore, the CNB-500-PS system provides a new strategy for degrading antibiotics in water.