AbstractAmmonia inhibition of unacclimated anaerobic mesophilic mixed cultures was studied in batch mode. Bacterial growth rate and specific acetate‐uptake rate were affected by the concentration of un‐ionised ammonia (UAN) in a three‐stage pattern: initial inhibition, plateau and final inhibition stage. The un‐competitive inhibition model provided the most reasonable data fit. The stage pattern of inhibition suggested that more than one mechanism of UAN inhibition are acting on the mixed culture. The effects of UAN on the length of initial lag‐time also followed a three‐staged curve. High concentration of UAN resulted in acetate release. The biological basis and origin of the acetate release were not determined.