Batch experiments are conducted at various concentrations of initial total ammonia nitrogen (TAN) with acetate as an electron donor to examine the effects of free ammonia (NH 3 ) inhibition on electricity production in single-chambered microbial fuel cells (MFCs). This research demonstrates that initial TAN concentrations of over 500 mg N L -1 significantly inhibit electricity generation in MFCs. The maximum power density of 4240 mW m -3 at 500 mg N L -1 drastically decreases to 1700 mW m -3 as the initial TAN increases up to 4000 mg N L -1 . Nitrite and nitrate analysis confirms that nitrification after complete acetate removal consumes some TAN. Ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) are also inhibited by increasing the initial TAN concentrations. Another batch experiment verifies the strong inhibitory effect of TAN with only small differences between the half-maximum effective concentration (EC 50 ) for TAN (894 mg N L -1 equivalent to 10 mg N L -1 as NH 3 ) and optimum TAN conditions; it requires careful monitoring of the TAN for MFCs. In addition, abiotic control experiments reveal that granular activated carbon, which is used as an auxiliary anode material, adsorbs a significant amount of ammonia at each TAN concentration in batch MFCs.