Study On The Role Of Extracellular Polymeric Substances During The Anaerobic Sludge Granulation And Application

Anaerobic granular sludge as a type of microbial aggregation formed throughextracellular polymeric substances (EPS), which having superior settleability, high volumetricloading capacity and strong resistance to shock loading etc., is extensively applied in thetreatment of high concentration organic wastewater. However, there is scarce data about thekey influence factors of EPS in the formation of granular sludge as well as the compositionand structure maintaining method for mature granular sludge during beer wastewatertreatment under low organic loading conditions. Thus, in this paper, the characteristics ofgranular sludge and influence factors during the cultivation process from aerobic sludge wascomprehensively investigated, and the response mechanism of mature granular sludge tosemi-starvation stress was further discussed. Moreover, the key composition of EPS, whichwould influence the granular sludge formation and structure stability, was analyzed usingadvanced three-dimensional excitation emission fluorescence matrix (3D-EEM) and fouriertransform infrared(FT-IR), etc. Finally, industrial-scale cultivation of granular sludge wascarried out on the base of successful lab-scale cultivation. This industrial-scale cultivationcould provide theoretical basis and practice guide for large-scale application of granularsludge. Main results are as follows:(1) Anaerobic granular sludge was cultivated by gradually increasing volume loading ina CLR anaerobic reactor, which was self-developed, inoculated with aerobic sludge and fedwith simulative beer wastewater. The anaerobic granular sludge was successfully cultivatedin the continuous-running CLR after125days, where the volume loading gradually increasedfrom0.5kg COD/(m3d) to18kg COD/(m3d). The cultivated granular sludge with15.6%ofcomponents’ diameter larger than2.8nm, had an appearance of black, smooth andnear-spherical. During the granulation of sludge, the PN contents in EPS was improved from21.4mg/g VSS to97.7mg/g VSS, while the PS contents showed no significant change.Meanwhile, the PN/PS value was positively correlated to both Zeta potential and cell surfacehydrophobicity. Results indicated that Zeta potential and cell hydrophobicity could beimproved by increasing PN concentration in EPS, thus prompting microbial aggregate togranular sludge. This means that PN played an important role in the granulation of sludge.The3D-EEM and FT-IR were employed to characterize the EPS composition. Threerepresentative aromatic protein-like fluorescent substance such as tyrosine (low-excitation,peak A and high-excitation, peak C), tryptophan(low-excitation, peak B)were identified by3D-EEM, and tyrosine and tryptophan were also detected at infrared wavelength (1520cm-1and1520cm-1) by FT-IR respectively. In addition, the fluorescence intensity from3D-EEMenhanced with the granular size. However, the fulvic-like substance (F) was only discoveredin seed sludge. The above results manefisted that aromatic protein-like substances, tyrosine and tryptophan, could boost the granular sludge formation.(2) Structure stability of mature granular sludge under semi-starvation condition (lowervolumetric loading) was investigated according to the characteristics of beer wastewatertreatment, and the key EPS compositions during the granulation process were furtherconfirmed. Two reactors were performed under semi-starvation conditions by continuous (R1)and intermittent (R2) operation, respectively. During the whole operation period of87days,the granular sludge disintegration in both reactors was found with average size reduction inthe first2months. With time going on (the3rdmonth), the sludge in reactor R2still continuedto break, while the broken sludge in R1was found to be re-aggregated to oval shapedgranular sludge. And COD removal rate in reactor R1was higher than that in reactor R2,illustrating that continuous operation of reactor R1was in favour of granular sludgemaintaining its structure stability in semi-starvation condition. Results from3D-EEM andFT-IR found that the broken sludge in reactor R2was simultaneously accompanied bydisappearance of aromatic protein-like tyrosine (low-excitation, peak A and high-excitation,peak C) in3D-EEM and appearance of fulvic-like suabstance and protein secondary structureAmide Ⅱ (1549cm-1). However, protein-like tyrosine was re-detected after sludgere-aggregation in reactor R1. In brief, it could be concluded that aromatic protein-liketyrosine were significant for granular sludge in maintaining its structure stability,whilefulvic-like substance (peak F)showed the negative effect.(3) The granular sludge was successfully cultivated in a industrial-scale reactor,inoculated with aerobic sludge and fed with beer wastewater. During cultivation period of193days and continous semi-starvation period of90days, volumetric loading was firstlyincreased from0.5kg COD/(m3d) to9.6kg COD/(m3d), and then declined to4.8kg COD/(m3d). The cultivated granular sludge with15.2%of components’ diameter largerthan2.8nm, had an appearance of near-spherical. It was found that the mature granularsludge would partly break under semi-starvation condition, and then re-aggregated to ovalshaped granule after two months when the reactor was operated continuously. Tyrosine andtryptophan were also measured along with granulation and increased with granular size,whichillustrated that protein-like tyrosine and tryptophan were the key components in granularsludge formation and structure stability maintenance. Thus, tyrosine and tryptophan in EPScould be adopted as an important index for guiding granular sludge cultivation.