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Microbial fuel cells are one of the new methods of energy production that microorganisms generate power from biomass by performing oxidation and reduction processes. In this research, a single-chamber reactor with a volume of 157 ml was constructed and investigated. This reactor had three brush anodes and an air cathode. The maximum voltage and power density were 500 mv and 33 mW/m2, respectively. The COD removal efficiency was 53, 66, 85 and 89% at 8, 12, 24 and 48 hours, respectively, which is a high efficiency for an anaerobic system. The Coulombic efficiency after 25 and 80 hours was 13.64 and 20.9, respectively. The higher the concentration of COD, the higher the efficiency of electricity production. Also, the maximum power density for concentrations of 4500, 2000, 1000, 500, 300 and 150 were obtained as 393, 330, 285, 252, 230 and 140 mW/m2, respectively. An increase in temperature first leads to an increase in efficiency (power production) and then a decrease. An increase in pH also leads to an increase in efficiency (power production) and then to its decrease, and neutral pH is optimal. Also, three different substrates, including acetate, glucose, and domestic wastewater, were evaluated. The type of substrate affects more than the maximum power value; it affects the three increasing, static and decreasing phases. Acetate and domestic wastewater also act very similarly.