The effects of free ammonia (FA) or free nitrous acid (FNA) on partial nitrification (PN) has been well investigated. Nevertheless, little information was known about the combined effects of nitrogen loading rate (NLR) and substrate inhibition, as well as the impact on the community structure of nitrifiers. In this work, real reject water was treated in a pre-denitrification reactor. PN was achieved by gradually increasing NLR, and it was successfully maintained when average FA and FNA were within 0.8-3.2 mg NH₃-N/L and 0.003-0.067 mg HNO₂-N/L, respectively. When NLR was reduced, PN was slightly affected due to the FA declination. As FNA inhibition was also eliminated by adding alkalinity into the influent, PN was completely destroyed quickly. FISH results indicated the deterioration of the PN was mainly attributed to the recovery of NOB when inhibition effects were limited. It concluded the increase of NLR benefited the partial nitrification. However, the stability of the nitrite path way was more dependent on selective substrate inhibition effects, especially the FNA inhibition. These findings would be important for further treatment of actual reject water.