Electrochemical production of H2O2 from O2 using simulated seawater provides a promising alternative to the energy-intensive industrial anthraquinone process. In this study, a flow cell system is built for electrocatalytic production of H2O2 under an air atmosphere in simulated seawater using cobalt single-atom catalysts (Co SACs). The Co SACs can achieve a high H2O2 production rate of 3.4 mol gcatalyst-1 h-1 under an air flow at a current density of 50 mA cmgeo-2 and long-term stability over 24 h in 0.5 M NaCl. It is found that Co-N5 rather than the Co-N4 structure in Co SACs is the main active site for H2O2 formation in the two-electron oxygen reduction reaction (ORR) pathway. It also shows high chloride-endurability without inhibition of the ORR process in simulated seawater. The fast production of H2O2 on Co-N5 sites in a flow cell provides a promising path of electrocatalytic oxygen reduction in simulated seawater, eventually converting ubiquitous air and seawater towards energy sustainability.