Hydrolysis, where the complex insoluble organic matter is hydrolyzed by hydrolytic enzymes, is regarded as rate-limiting step for anaerobic digestion of waste activated sludge (WAS). The cascade system, which consisted of small-reactors in series, was developed in this study to enhance the hydrolysis rate. Degradation efficiency of sludge was strongly related to the sludge retention time (SRT) and recirculation ratio (RR). Therefore, this study investigated the effect of SRT and RR on process performance and hydrolysis enzymatic activity of WAS treatment in the cascade system and conventional single continuous stirred tank reactor (CSTR) system. The cascade system’s chemical oxygen demand (COD), volatile solid (VS), protein and carbohydrates reduction, and methane production could be maintained despite the decrease of SRT, which was associated with an increase in organic loading rate (OLR) (15 d = 3.33 gCOD/L/d, 12 d = 4.16 gCOD/L/d). The performance of reference CSTR system deteriorated at shorter SRT. Hydrolysis, acidogenesis and methanogenesis analysis indicated that shorter SRT accelerated the hydrolysis rate of cascade system from 1.29±0.23 gCOD/L/d (SRT=15 d) to 1.59±0.19 gCOD/L/d (SRT=12 d), leading to the increase of overall degradation of WAS. The enhancement of hydrolysis rate was supported by the increased protease and cellulase activity detected at shorter SRT. Recirculating the effluent from reactor3 (R3) to reactor 1 (R1) was also essential to maintain the performance. The results not only showed deteriorating performance of cascade system regarding lower COD, VS, protein, carbohydrates reduction and methane production, but also presented lower hydrolysis enzymatic activity at 2% RR compared to 10% RR. 10% recirculation of solid phase, which contained biomass and sludge-attached enzymes, was beneficial to recover the performance of cascade system but the performance was not as good as that with 10% recirculation of both solid and liquid phase of R3 effluent. The cations analysis revealed that the required quantities of Cobalt (Co), Nickel (Ni) and Zinc (Zn) was highest in R1 while the concentration of soluble Co, Ni and Zn was lowest in R1. The concentration of Co, Zn and Ni was increased along AD of cascade system, providing the opportunities of offering increment of trace elements to the growth of microbial community and enzymes formation in R1. In conclusion, cascade system had better performance at shorter SRT compared to single CSTR. The recirculation played an important role to maintain the superior performance of cascade system under high OLR.

Large amounts of residual waste activated sludge are produced as by-products during biological wastewater treatment processes. Anaerobic digestion is a widely accepted stabilisation method for waste activated sludge (WAS) treatment. However, the application of anaerobic digestion is limited by the long retention time and low degradation efficiency of compounds. Structural extra cellular polymeric substances (EPS) are metabolic products released by microorganisms that play an important role in the disintegration of sludge structure. The limitations in anaerobic digestion mentioned above pertain to the hydrolysis step in anaerobic digestion. The cascade reactor (cascade AD) system i.e. continuous stirred tank reactor (CSTR) in series, is a robust reactor system that is expected to enhance the hydrolysis step and to show a superior performance at low solid retention time compared to a conventional CSTR. This research is aimed at observing the difference in performance of cascade AD and a conventional CSTR at shortened retention time. Various indicators were used to understand the performance enhancement of a cascade AD in comparison to a conventional CSTR. Moreover, the degradation of structural EPS by selected enzyme groups such as protease, cellulase and polygalacturonase were also studied. The cascade AD showed better performance than a conventional CSTR at a retention time of 22 and 15 days. This improved performance was enabled by the smaller reactors in cascade AD that provided higher hydrolysis rate. Higher removal efficiency of protein, carbohydrate and structural EPS was observed in cascade AD. Improved ammonium and phosphate release were also indications of better performance of cascade AD. Although the mass balance in nitrogen was maintained in the reactor system, phosphorous mass balance indicated possibilities of precipitation. The batch tests performed with the enzyme protease, cellulase and polygalacturonase were aimed at understanding the degradation of SEPS. It was inferred from the test that volatile suspended solids proved to be a better indicator for solubilisation compared to COD and ammonium concentration. Protease showed higher solubilisation compared to cellulase and polygalacturonase. Particle size distribution did not indicate a significant difference upon the addition of all three enzyme groups; indicating that a significant change in structure was not caused by the enzymes. Hydrolysis kinetics and SEPS degradation could not be derived from the test because of the variations in results based on the substrates used. Nevertheless, the tests proved to be useful in improving methodology for deriving hydrolysis kinetics of WAS. In conclusion, the novel cascade AD showed better performance at shortened retention time. The system also showed stable performance despite the shortened retention time compared to a conventional CSTR. Thus, the stable performance suggests the opportunities to further lower the retention time in cascade AD.

The hydrolysis of sludge solids especially for difficultly degradable sludges such as WAS is not fully understood, yet. The first-order hydrolysis rate was shown to function well for most easily degradable sludges and soluble substrates. This description for substrate hydrolysis in the context of anaerobic digestion has the benefit of being very simple and therefore applicable for many engineering applications where little data is available. On the other hand, in the last decades many studies reported that the first-order hydrolysis would need a modification to better describe the degradation of difficulty degradable solids. Guo et al. (2021) developed a cascade system for anaerobic digestion of WAS that does not seem to follow first-order hydrolysis kinetics when lowering the applied SRTs from 22 to 15 and 12 days, respectively. Based on observations by Guo et al. (2021) and a statistical analysis of the cascade system performed in the study at hand it seems that the first order hydrolysis rate constant is in fact a coefficient and that the first-order hydrolysis rate is not solely dependent on sludge characteristics and substrate concentrations. This hypothesis is tested in the thesis at hand. In Guo et al.’s study the cascade system was always compared to a reference system. To test this hypothesis and understand the kinetics of the cascade system in more detail a statistical analysis was performed for both systems from which an empirical hydrolysis model was derived. This model was implemented in ADM1 to replace the existing hydrolysis rate expression and was tested for the mentioned cascade system and the reference system. The empirical model was compared to the results of the standard ADM1 which uses a first-order hydrolysis expression. The empirical model assumed a dependency of the hydrolysis rate based on load and residence time along the cascade system to achieve a change in hydrolysis rate coefficients along the cascade system. The models were compared based on visual inspection and quantitative analysis of the simulated results. Both models showed low R² values which is likely due to the high level of detail implemented in ADM1 that does not fit to the resolution of the experimental data. However, calculated RMSE values agreed with the standard deviations of the experimental results. Therefore, the overall predictive capability for both models is given. The ADM1 managed to model the reference system with reasonable agreement to the experimental data. The performance of the empirical model for the reference was comparable. For the cascade system however the ADM1 could not fully describe the experimental at the applied low SRTs of 15 and 12 days. The empirical model in this case showed better predictive capabilities. This is an indication that a hydrolysis rate which is made dependent on system characteristics such as load and residence time might indeed have its justification and be better applicable to anaerobic digestion systems that show a concentration profile along the reactor as it is in the case with plug-flow and cascade systems.

A comparative batch experiment as well as a biochemical methane potential(BMP) test were conducted over 44d for aerobic granular sludge (WAGS) and waste flocculent activated sludge(WAS) on sludge anaerobic digestion (AD), to give an insight into the chemical and mechanical properties change of the EPS in different sludge source during AD process, via the analysis of polysaccharides (PS), protein (PN), loosely bound extracellular polymeric substance (LB-EPS), tightly bound extracellular polymeric substance (TB-EPS) change and its gel-forming property with multi-valent ions. The EPS, who play a vital role in forming the compact gel-like sludge structure, was extracted from WAGS and AS and analyzed for several test. During AD process, WAGS demonstrated a lower hydrolysis rate than AS over the first 6 days and reached a comparable overall methane production potential afterwards, suggesting the necessity of pre-treatment of WAGS to enhance sludge hydrolysis. The isolated SEP was slowly degraded over the AD process with a biodegradability of 31% for WAGS and 39% for WAS and this degradation of EPS mainly contributed from the PS and PN it contained. This EPS degradation led to the loss of gel-like properties for both sludge after AD process. Although the results from fourier transformed infrared (FT-IR) spectra exhibited the similar typical brands within the range of 1000~3000 cm-1 in all samples, the second-derivative spectra manifested the occurrence of polysaccharides degradation, especially for homopolymannuronic acid blocks (MM) and homopolyguluronic acid blocks during sludge digestion. Meanwhile, the dewaterability of both sludge was observed to deteriorate with AD ongoing due to the internal transmission from the compact TB-EPS to porous LB-EPS as well as the increase in PN/PS ratio in EPS. Moreover, the mechanical property variation of different sludge was examined by the crosslink of EPS solution with ion to form hydro-bead. Reduction on mechanical strength of EPS formed hydro-beads was seen for both WAGS and WAS but mechanical structure loss of WAGS was less than that of WAS after digestion which can be attributed to higher EPS residues in WAGS. However, no matter for WAGS or WAS, only a limited amount of EPS was degraded during AD process and this provided a possibility of recovery EPS as a coating material from digested sludge.