Full-Scale Thermal Hydrolysis Optimization: Impact on Refractory Organics and Anaerobic Digestion Robustness

Introduction Thermal hydrolysis (THP) is widely deployed to improve sludge treatment performance by enhancing anaerobic digestion (AD), increasing biogas production, boosting dewaterability, and achieving hygienization. However, THP also generates biologically recalcitrant compounds-particularly refractory chemical oxygen demand (rCOD) and refractory dissolved organic nitrogen (rDON)-that can resist downstream biological treatment and contribute to effluent residuals. This challenge is increasingly important as utilities face tightening discharge standards for COD and nitrogen and rising interest in high load solids strategies. This full scale study at the Dijon EauVitale WRRF (France) investigates how THP operating temperature (140-165  °C) influences the quantity and typology of refractory organics, their fate through digestion, and microbial community resilience. We combine high resolution chemometric fingerprints (UPLC SEC with UV/fluorescence detection) and artificial neural network (ANN) feature selection with parsimonious multiple linear regressions (MLRs) to build predictive models for rCOD and rDON using interpretable descriptors. We further assess PCDD/Fs concentrations and Toxic Equivalents (TEQ) around THP and characterize bacterial/archaeal diversity in digestate to evaluate operational robustness. A set of mass balances at full scale complements bench scale degradability tests to quantify how much refractory material is formed by THP and retained/released through AD and dewatering. Materials & Methods The study was conducted at the Dijon EauVitale WRRF equipped with thermal hydrolysis (140-165  °C) and mesophilic anaerobic digestion. Samples were collected at key process stages (pre-THP, post-THP, digestate, effluent) across multiple campaigns. Refractory COD and DON were quantified using standard colorimetric assays, while molecular fingerprints were characterized by UPLC-SEC with UV/fluorescence detection. Predictive models were developed using artificial neural networks and multiple linear regressions based on selected spectral descriptors. Microbial community structure was assessed via 16S rRNA sequencing and diversity metrics. Results & Discussion 1) Temperature vs Quantity and Typology of Refractory Organics Linear scaling of refractory formation. THP increased rCOD proportion linearly from 140 to 165  °C: ≈2.9 fold at 140  °C and ≈4 fold at 165  °C relative to PWAS. Despite quantity scaling, the rDON/rCOD ratio remained stable at ~4.5% (mean; 95% CI ≈3.8-5.1%), indicating a constant nitrogen share in refractory pools across the studied range. This stability suggests that temperature mainly scales the refractory quantity, not the nitrogenization of the refractory fraction. Chemometric fingerprints: typology conserved. PCA/HCA of the 144 spectral ratios revealed three robust clusters: (i) raw PWAS, (ii) THP/digestate streams, and (iii) final effluent. Analytical descriptors most discriminant (ANOVA, p < 0.05) were dominated by 20-40 kDa signals combining tryptophan like and humic /melanoidin like features (e.g., TRY1/MEL, TRY1/HA, TRY2/AF2). Across temperatures, these fingerprints remained consistent, supporting that THP temperature does not materially alter molecular typology, even though it amplifies the amount of refractory material. ANN MLR predictive modeling. ANNs prioritized a subset of descriptors, from which compact MLRs reproduced measured rCOD and rDON with R² = 0.972 and 0.964 using 4-5 interpretable ratios per model. Coefficient signs provided mechanistic insight: low MW protein-melanoidin and humic normalized protein/fulvic contrasts acted as negative predictors of recalcitrance, while 20-40 kDa protein/humic enrichments were positive predictors, consistent with the cluster discrimination. Practically, these descriptors can be embedded into monitoring to track refractory risk and guide temperature setpoints. Operational implication. For design/operations, increasing THP temperature predictably raises refractory load without altering its spectral signature, enabling utilities to use ANN MLR monitors as early indicators and control levers (e.g., temperature windows, blend strategies) rather than expecting typology shifts to ease downstream treatment. 2) Fate Through Anaerobic Digestion and Return Pathways AD mitigation (~31%). Full scale mass balances combined with degradability assays indicated that AD reduces refractory loads by ~30.6 ± 2.3% (95% CI ≈20-40%). Evidence points to physical mechanisms-adsorption/copolymerization/association with biomass-more than biodegradation, consistent with literature on hydrophobic recalcitrants binding to flocs. This mitigation means not all THP derived refractory material returns to headworks; a significant share remains associated with digested sludge and exits with dewatered cake. Centrate contribution and effluent context. At Dijon, THP derived refractory accounted for ~23-34% of effluent refractory COD during campaigns, with the remainder dominated by external industrial leachates. Even so, the AD mediated reduction increases the liquids line's headroom, limiting residual build up during high load operation. For centralized solids hubs, this synergy suggests greater acceptance capacity for external sludges than expected if considering THP alone. Operational implication. Coupling THP with robust AD provides a two stage strategy: push hydrolysis for digestion gains while buffering refractory recirculation through physical capture in digesters and solids handling. Monitoring centrates and adjusting cake handling strategies (e.g., capture, conditioning) becomes essential when effluent residuals are rate limiting for compliance. 3) PCDD/Fs Risk Profile at THP Temperatures Measured concentrations and TEQ. Around THP, PCDD/Fs totals showed negligible change at 140  °C (PWAS→HWAS; within method uncertainty) but increased at 165  °C, with TEQ rising to ~9.2 ng TEQ/kg TS. Although higher than at 140  °C (~2.9 ng TEQ/kg TS), values remained below applicable agronomic thresholds currently discussed in the EU context (e.g., France's Socle Decree limit: 20 ng TEQ/kg TS for land application). The total mass increase at 165  °C was accompanied by toxicity weighted increase, suggesting formation/enrichment of more dioxin like congeners at the upper end of the THP window. Operational implication. Within 140-165  °C, TEQ remains manageable, but facilities should adopt periodic screening when operating ≥160-165  °C, especially if feedstocks or legacy contamination could bias congener profiles. Pressure effects and catalyst/ash interactions warrant further study under THP conditions. 4) Microbial Diversity and Process Robustness Stable alpha diversity. Across THP temperatures, both bacterial and archaeal communities exhibited high and stable diversity (Shannon/Simpson), indicating that THP setpoints do not compromise the functional breadth of digestion. Archaeal communities remained dominated by hydrogenotrophic methanogens (e.g., Methanoculleus) typical of high ammonia sludge digestion, while bacteria showed moderate compositional shifts (e.g., Fastidiosipila vs. Proteiniphilum/Brevefilum) consistent with changes in substrate complexity rather than loss of core hydrolytic/acidogenic functions. Beta diversity insight. Bray-Curtis ordination highlighted greater sensitivity in bacterial assemblages to temperature than archaea, aligning with their broader roles in hydrolysis/acidogenesis. Functionally, the core trophic network remained intact, supporting operational resilience across the THP window. Operational implication. Utilities can tune THP temperature for digestion/performance gains without destabilizing microbial function, provided ammonia management and organic loading remain within design envelopes. Conclusions and Practical Guidance - Temperature effect: Increasing THP temperature (140→165  °C) linearly amplifies refractory organics without changing their molecular fingerprint; rDON shares remain proportionally constant. - Predictive control: A small, interpretable set of chemometric descriptors predicts rCOD/rDON with high fidelity, enabling ANN MLR monitoring to proactively manage refractory risk and temperature setpoints. - AD synergy: Anaerobic digestion removes ~31% of THP derived refractory material-primarily via physical association-meaning not all recalcitrants recirculate to the liquids line. - PCDD/Fs vigilance: TEQ remains below agronomic thresholds at 165  °C but rises versus 140  °C; adopt periodic screening when operating at the upper THP range. - Microbial resilience: High and stable diversity and conserved core functions confirm process robustness across the studied THP temperatures.