Variations in extracellular polymeric substances (EPS) during adaptation of activated sludges to new feeding conditions Ayse Gul Geyik * , Ferhan Çeçen Institute of Environmental Sciences, Bogazici University, 34342 Bebek, Istanbul, Turkey article info Article history: Received 11 August 2015 Received in revised form 25 August 2015 Accepted 29 August 2015 Available online xxx Keywords: Adaptation Carbon to nitrogen EPS Fractionation Nitrifiers Very loosely bound EPS abstract The study examined the dynamics of EPS production in activated sludges. Semi-continuously fed batch activated sludge reactors were operated at different feeding conditions, namely at COD/TKN ratios of 10, 5 and 0, respectively. Parallel to this operation, EPS production and fractionation were evaluated. The study identified for the first time a very loosely bound fraction in EPS, the VLB-EPS. Bound EPS were divided into many fractions as Very Loosely Bound-EPS (VLB-EPS), Loosely Bound-EPS (LB-EPS) and Tightly Bound-EPS (TB-EPS) while the carbohydrate and protein content of each fraction was measured. Regardless of the COD/TKN ratio, tightly bound EPS and protein-EPS were dominant in all sludges. As determined by HPSEC analyses, in all EPS fractions, a great portion of protein-EPS had a molecular weight (MW) 12.5 kDa. Although all substrates were very effectively removed after start-up of reactors, a long operation time was required for stabilization of EPS. The only exception was the nitrifying sludge in which EPS were soon stabilized. © 2015 Elsevier Ltd. All rights reserved. 1. Introduction Extensive research has shown that extracellular polymeric substances (EPS) are a major component of the matrix found in activated sludge (Wingender et al., 1999; Liu and Fang, 2003). EPS are sticky solid materials secreted by cells. In a biological sludge, EPS serve in adhesion to surface, formation of matrix structure, control of microbial physiology, long-term stability of sludge (Liu and Fang, 2003) and sludge dewaterability (Ruiz-Hernando et al., 2015). EPS may also aid in accumulation of nutrients from envi- ronment (Flemming and Wingender, 2010) and provide resistance to cells in the case of toxic metals or inorganics. In activated sludge EPS are hypothesized to bridge two neigh- boring bacterial cells physically to each other, as well as with inert particulate matter (Sobeck and Higgins, 2002; Mcswain et al., 2005). This cohesive binding creates large and dense aggregates. EPS exhibit a double-layered structure. Loosely bound EPS (LB-EPS) diffuses from the tightly bound EPS (TB-EPS) that surrounds the cells (Sheng et al., 2006). The cells in the outer region of flocs are entangled by weak interactions through LB-EPS. Another fraction in EPS is the soluble microbial products (SMP). While bacteria carry out biomass synthesis, they also convert a fraction of organic sub- strate into SMP which account for the bulk of soluble organic car- bon in effluent (de Silva and Rittmann, 2000a,b). In a full-scale wastewater treatment system composition and strength of the influent wastewater may fluctuate with respect to time. In particular, changes may occur in the carbon to nitrogen (C/ N) ratio, also known as the COD/TKN ratio. Such changes might have a great influence on microbial ecology of sludge. Possibly, during adaptation of microrganisms to a feed, also variations are expected in fractionation and composition of EPS (Yang and Li, 2009; Miqueleto et al., 2010; Ye et al., 2011). A thorough examination of literature reveals that there is still some missing information on EPS. Studies examing the production of EPS in activated sludge systems mostly deal with the case of high heterotrophic activity in the presence of organic carbon. On the other hand, to date only few studies examined nitrifying systems that are operated in the absence of organic carbon (COD/TKN ¼ 0) (Choi and Hu, 2009; Çeçen et al., 2010a,b; Xie et al., 2012). Also few studies paid attention to EPS production in such systems (Liang et al., 2010; Zhang et al., 2011). Therefore, one of the aims of the present study was to evaluate the effect of the COD/TKN ratio. Often, most studies do not mention whether EPS results were achieved under steady- or transient conditions. Former study examined in detail the production and composition of EPS in * Corresponding author. E-mail addresses: gul.geyik@boun.edu.tr (A.G. Geyik), cecenf@boun.edu.tr (F. Çeçen). Contents lists available at ScienceDirect International Biodeterioration & Biodegradation journal homepage: www.elsevier.com/locate/ibiod http://dx.doi.org/10.1016/j.ibiod.2015.08.021 0964-8305/© 2015 Elsevier Ltd. All rights reserved. International Biodeterioration & Biodegradation 105 (2015) 137e145