Abstract:[Background] The metabolic pathway of electrochemical anaerobic digestion (EAD) system is dominated by microorganisms with different functions, and the metabolic flux is related to the abundance, activity, and community structure of functional microorganisms. [Objective] To explore the relationship between methane flux and microorganisms in the EAD system. [Methods] We conducted the metabolic flux analysis by using pH as a disturbance factor. [Results] At pH 7.5, the maximum methane flux was 0.398 4±0.029 3, which was 25% and 65% higher than that 0.297 4±0.012 7 in the control group (pH 6.9) and that 0.136 5±0.012 0 in the disturbance group (pH 6.3), respectively. In addition, an average of 33.8%±3.1% of hydrogen (flux) was used to reduce carbon dioxide to produce methane and acetic acid, and an average of 21.0%±2.6% of acetic acid (flux) was converted into methane. The methane flux was positively correlated with the abundance of Mariniphaga, Methanosaeta, and Desulfomicrobium, while negatively correlated with that of Sedimentibacter. [Conclusion] In the EAD system with the coexistence of methane and acid-producing bacteria, a pH value slightly greater than 7.0 is conducive to the production of methane, and changing the pH value of the EAD system can regulate the metabolite flux by altering microbial abundance.