Pre-ozonation is effective in improving the membrane filterability of the mixed liquor in membrane bioreactors. However, the amelioration mechanism is still not clear. The present study systematically investigated the influence of zonation on the properties of activated sludge, soluble microbial products (SMP), and extracellular polymeric substances (EPS). Chemical compositions of the SMP and EPS were quantified in terms of total organic carbon, polysaccharides, protein, and Ca2+/Mg2+. Functional groups were characterized using Fourier transform infrared spectroscopy and three-dimensional excitation-emission matrix fluorescence spectroscopy. On the basis of the analyses, the rationale behind the filterability improvement was elucidated. The appropriate zonation increased the sludge hydrophobicity and introduced more acidic functional groups to the SMP and [PS, thus facilitating a sludge reflocculation process in the subsequent aeration operation by virtue of cation-bridging and hydrophobic interactions. Ultimately, the increased sludge particle size and decreased SMP concentration jointly contributed to the amelioration of membrane filterability. (C) 2013 Elsevier B.V. All rights reserved