Revealing the roles of biochar in anaerobic digestion of waste activated sludge with the overloading accumulation of humic acids

Anaerobic digestion (AD) can produce renewable energy of methane from waste activated sludge (WAS), thereby achieving sludge reduction and resource utilization. However, the abundant recalcitrant humic acids (HAs) in WAS may accumulate to inhibitory levels and suppress anaerobic methanogenesis by binding hydrolases, limiting organic matter conversion, and competing with methanogens for electrons. Herein, the roles of biochar (BC) in AD of WAS under different HAs shock conditions were evaluated, i.e., the overloading accumulation cases of HAs in the substrate of WAS or in the AD phase. Results showed that for the substrate of WAS with overloading of HAs, BC increased the cumulative methane production to 140.4 mL/g volatile suspended solids (VSS), which was 37% and 23% over that in HA and control groups. For the overloading accumulation of HAs during AD period, the cumulative methane productions were 25.6% and 17.5% over that in HA and control groups. Mechanistic analysis indicated that BC alleviated HAs-induced inhibition mainly by adsorbing HAs, reducing their binding potential to hydrolases, and enriching fermentative bacteria such as Firmicutes, Chloroflexi, and Bacteroidota, thereby facilitating organic matter transformation. In addition, compared with the HA group, the synergistic effects of BC and HAs increased coenzyme F420 activity by 34.2% and promoted the enrichment of electroactive microorganisms such as Methanosarcina and Clostridium_sensu_stricto_13 , thereby supporting methanogenesis under HAs shock. These findings advance the understanding of the roles and mechanisms of BC in sustaining methanogenesis during AD of WAS under overloading accumulation of HAs, supporting sustainable sludge management and energy recovery in wastewater treatment.