To investigate the influences of free ammonia (FA) on the nitrogen removal performance, the contents of extracellular polymeric substances (EPS) and the chemical composition (proteins (PN), polysaccharides (PS), and DNA), four laboratory-scale sequencing batch reactors (SBRs) fed with synthetic wastewater were operated at FA concentrations of 0.5, 5, 10 and 15 mg/L (R0.5, R5, R10 and R15, respectively). Results showed that high nitrogen removal efficiencies (97.6-99.4%) were achieved under four FA concentrations. FA had a significant impact on the three kinds of EPS (loosely bound EPS (LB-EPS), tightly bound EPS (TB-EPS) and total EPS, respectively) and their composition. Results showed that with the increased of initial FA concentrations from 0.5 to 10 mg/L, the production of three kinds of EPS and their components were significantly increased. Further increased of FA concentrations to 15 mg/L caused a decreased trend of them. Moreover, PN was the main component of LB-EPS, while PS dominated in TB-EPS and total EPS under different FA concentrations. It was found that the contents of three kinds of EPS and their components illustrated a similar variation tendency to the NO- x-N contents during the whole cycle under four FA concentrations.