Full-scale Cultivation Of Anaerobic Granular Sludge And Its Underlying Mechanisms

In full-scale anaerobic wastewater treatment, the operational stability and highperformance of an anaerobic reactor depends largely on the efficient settling properties andhigh methanogenic activity of anaerobic granular sludge. If loose sludge floc exists in theanaerobic reactor, the reactor becomes prone to sludge washout, low organic loading rate, andpoor performance. Mature granular sludge merchandise is scarce and expensive, grows slowly,involves difficult training, and has high transportation cost. Therefore, a study on full-scalecultivation of anaerobic granular sludge can solve engineering problems as well as providepractical knowledge and theoretical value.In this dissertation, a full-scale anaerobic reactor was inoculated with residual activatedsludge to investigate the following: sludge granulation process, factors influencing anaerobicgranulation by grey relational analysis, microbial population dynamics by molecularbiotechnology, enhanced granulation of anaerobic granular sludge by Ca²⁺andpolyacrylamide （PAM）, and comparison of the kinetic modeling of identical full-scale reactorswith residual activated sludge and anaerobic granular sludge for brewery wastewatertreatment. These concepts demonstrated the full-scale cultivation process of anaerobicgranular sludge and its mechanisms. The main results were described as follows:（1） Anaerobic sludge granulation with residual activated sludge could accomplishfull-scale cultivation for brewery wastewater treatment by a continuously average organicloading rate （OLR） of8.8kg chemical oxygen demand （COD）·m^-3·d^-1, upflow velocity of5.4m·h^-1, and hydraulic retention time （HRT） of5.9h. After granulation, the bioparticles largerthan0.5mm accounted for57%of all bioparticles. Mature particle size in the internalcirculation reactor was between2.0mm and2.5mm in the first reaction, whereas that in thesecond reaction chamber was between0.8mm and1.5mm. When the diameter size was1.74mm, the specific methanogenic activity could reach the maximum level which was1.95gCH4-COD·g volatile suspended solids （VSS）^-1·d^-1. The specific gravity was between1.02and1.06and the mechanical strength was between1600Pa and3400Pa in the diameter size of1.0mm to2.5mm. The tightly bound-extracellular polymeric substances （TB-EPS） andloosely bound-EPS （LB-EPS） were30.2mg total organic carbon （TOC）·g SS^-1and15.2mgTOC·g suspended solids （SS）^-1in the diameter size of2.5mm. The results showed that sludgegranulation mechanism could be attributed to EPS and selection pressure in full-scalecultivation of anaerobic granular sludge. TB-EPS affected the particle size and LB-EPSaffected the particles capability and structural strength. Selection pressure affected thebiomass concentration, particle size distribution, and the bioparticle rate, but also affected theproportion of the specific gravity, mechanical strength, sedimentation and other properties.（2） The grey relational analysis results showed that the factors that influenced anaerobicgranulation with significant effects on anaerobic granulation were as follows: liquid upflowvelocity> hydraulic retention time （HRT）> organic loading rate （OLR）> influent COD. The sludge in the effluent and in the reactor influenced by liquid upflow velocity was examinedusing a scanning electron microscope, energy dispersive spectrometer, and X-ray diffractionanalysis. The results revealed that the liquid upflow velocity affected the surface roughness,sludge shape, and integrity of the granular sludge through the collision friction andhydrodynamic shear force. The characteristics of the organic substrate could influence theformation, composition, and structure of anaerobic granules. The complexity of the substratecould also exert selection pressure on microbial diversity in anaerobic granules, which wouldconsequently influence the formation and microstructure of granules.（3） The main archaeal species in the sludge samples at different OLRs variedsignificantly compared with bacteria species using the polymerase chain reaction withdenaturing gradient gel electrophoresis （DGGE）. The Shannon–Wiener index （H’）, Margalefrichness index, and dominance index S of bacteria and archaea calculated by DGGE profilesshowed that a higher diversity index resulted in less ecological dominance and reducedrichness. The dominant archaeal species in the treated sludge at low OLRs wasMethanosarcina, whereas that at high OLRs was Methanosaeta.（4） Compared with the control group, the contents of EPS and proteins could reach themaximal values and respectively increased by31.8%and53.1%when the Ca²⁺concentrationwas300mg L^-1. However, polysaccharides content decreased by11.0%, the ratio of proteinand polysaccharide increased from2.0to3.5, and Zeta potential increased from-45mV to-30mV. The above variations enhanced granular sludge particle size from2.5mm to2.75mm,and methanogenic rate increased by5.0%. Compared with the control group, thedehydrogenase, BAA-protein hydrolase, β-glucosidase, alkaline phosphatase, and coenzymeF420activities increased by36.4%,28%,11.2%,29.8%, and28.0%respectively. Comparedwith the control group, the contents of EPS, proteins, polysaccharides, and Zeta potentialcould reach the maximal values and respectively increased by54.0%,82.0%,3.9%, and41.3%when the PAM concentration was20mg L^-1. The ratio of protein and polysaccharideincreased from2.0to4.2, and Zeta potential increased from-46mV to-27mV. The abovevariations enhanced granular sludge particle size from2.5mm to3.0mm, and methanogenicrate increased by15.1%. The above enzyme activities increased by120%,40%,29.4%,40.4%, and166.7.0%, respectively, compared with the control group. Anaerobic sludgegranulation enhanced by Ca2+and PAM.（5） Two identical full-scale biogas-lift reactors treating brewery wastewater wereinoculated with different types of sludge. One reactor （R1） started up with anaerobic granularsludge in12weeks and obtained a continuously average OLR of7.4kg COD·m^-3·d^-1, effluentCOD of approximately450mg·L^-1, and COD removal efficiency of80%. The other reactor（R2） started up with residual activated sludge in30weeks and granulation was accomplishedwhen the reactor reached an average OLR of8.8kg COD·m^-3·d^-1, effluent COD ofapproximately240mg·L^-1, and COD removal efficiency of90%. The difference between thevolatile fatty acids （VFA） concentrations of R1and R2were significant, which were approximately200mg·L^-1and100mg·L^-1as acetic acid, respectively. The VFA/alkalinityratios of R1and R2were approximately0.3and0.15, respectively. Methane contents in thebiogas of R1and R2were about77.2%and78.6%, respectively. The result suggested that thedominant methanogens in R2were able to maintain a higher level of methanogenesis becauseof achieved anaerobic granulation sludge. EPS production increased slightly from36.4mg·gSS^-1to36.8mg·g SS^-1in R1, whereas that increased from22.5mg·g SS^-1to46.1mg·g SS^-1inR2. The increased amount of EPS indicated that EPS could help in granule formation. A singleparticle size of R1was between1.5mm and2.5mm, whereas different particle sizedistribution of R2was0.0⁰.3mm,0.3⁰.5mm,0.5¹.5mm, and1.5².5mm. Differences insludge characteristics, methane content, VFA, and VFA/alkalinity ratio may be accounted forthe superior efficiency of the treatment performance of R2over R1. Grau second-order andModified Stover–Kincannon models were successfully used to develop kinetic parameters ofthe experimental data with high correlation coefficients. These findings demonstrated that theresidual activated sludge could be used effectively instead of anaerobic granular sludge toachieve higher performance and sludge granulation. These results indicated that the highactivity of anaerobic granular sludge full-scale cultivation system could be established.