The longer start-up period of the Anammox process is due to the very low cellular yield and growth rates of Anammox bacteria. Nitrite inhibition is considered to be the key factor in the instability of the Anammox process during the operation. However, little attention was paid to the inhibitory effect of pH and free ammonia. This paper presents start-up and inhibition analysis of an Anammox biofilm reactor seeded with anaerobic granular sludge. Results showed that the start-up period could be divided into the sludge lysis phase, lag phase, propagation phase, stationary phase and inhibition phase. Optimization control could be implemented correspondingly to accelerate the start-up of Anammox bioreactors. Effluent pH increased to 8.7-9.1 when the nitrogen removal rate was higher than 1,200 mg l(-1) day(-1). The free ammonia concentration was accompanied with a higher level of 64-73 mg l(-1). Inhibitory effects of high pH and free ammonia on Anammox bacteria contributed to the destabilization of the Anammox bioreactor during the first 125 days with influent KHCO(3) of 0.5 g l(-1). Increasing the suffering capacity in the inlet by dosing 1.25 g KHCO(3) l(-1) effectively reduced the pH variation, and the nitrogen removal performance of the reactor was further developed.