The precise role of extracellular polymeric substances (EPS) in relation to the formation and physicochemical properties of microbial floc in wastewater treatment systems is not well known. Studies were undertaken to provide more comprehensive descriptions of EPS and properties of microbial floc. Acidic polysaccharides and DNA were relatively labile components of the EPS when biomass was stored at 4°C or at −20°C, and significant losses of these components were observed within 24 hours. The composition and properties of activated sludge were found to vary between different full-scale treatment systems reflecting the importance of wastewater composition and operation conditions on microbial communities and the response to environmental conditions. The COD:N:P ratio was found to influence hydrophobicity, surface charge and the EPS composition of microbial flocs in well-controlled bench-scale sequencing batch reactors. Phosphorus depleted and P-limited conditions resulted in a decrease in surface charge but increases in acidic polysaccharides which corresponded to a strong carboxyl stretch at 1740 cm−1 when the biomass was analysed by FTIR-spectroscopy. Electron dense particles, identified by energy-dispersive spectroscopy as containing iron, phosphorus and sulfur, were observed in the fibrils of the floc matrix by transmission electron microscopy.

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