The effect of nitrate on the performance of a single chamber air cathode MFC system and the denitrification activity in the system were investigated. The maximum voltage output was not affected by 8.0mM nitrate in the medium solution at higher external resistance (270-1000Omega), but affected at lower resistance (150Omega) possibly due to the low organic carbon availability. The Coulombic efficiency was greatly affected by the nitrate concentration possibly due to the competition between the electricity generation and denitrification processes. Over 84-90% of nitrate (0.8-8.0mM) was removed from the single chamber MFCs in less than 8h in the first batch. After 4-month operation, over 85% of nitrate (8.0mM) was removed in 1h after the MFC was continuously fed with a medium solution containing nitrate. Only a small amount of nitrite (<0.01mM) was detected during the denitrification process. The similar denitrification activity observed at different external resistances (1000 and 270Omega) and open circuit mode indicates that the denitrification was not significantly affected by the electricity generation process. No electricity was generated when the MFC fed with 8.0mM nitrate was moved to a glove box (no oxygen), indicating that the bacteria on the cathode did not involve in accepting electrons from the circuit to reduce the nitrate. Denaturing Gradient Gel Electrophoresis (DGGE) profiles demonstrate a similar bacterial community composition on the electrodes and in the solution but with different dominant species.