Cassava alcohol stillage is high-strength organic wastewater generated during bioethanol production. Its high organic load and propensity for volatile fatty acid accumulation during anaerobic digestion (AD) can inhibit its degradation efficiency significantly. This study introduced porphyrin-mediated inoculum acclimation under light to investigate the synergy between photosynthetic and methanogenic communities in AD. In the present study, preliminary experiments were conducted by supplementing the AD system with varying concentrations of hemin (2.5–7.5 g/L), using cassava alcohol stillage as the substrate. Methane (CH4) production enhancement was limited, ranging from 4.66% to 17.49%. Under 5 g/L hemin combined with acclimated inoculum, methane production increased by > 120%, total organic carbon degradation rate improved by 20.94%, acidification lag phase was shortened by 80%, and the digestion cycle was reduced from 28 d to 14–16 d. Following introduction of the inoculum acclimated under light in the presence of hemin over a four-month period, the methanogenic community was enriched substantially at the end of fermentation. The relative abundances of Methanobacteriota—the core methanogenic archaeal phylum—reached 5.92%, 10.12%, and 4.11% in the 2.5 g/L, 5 g/L, and 7.5 g/L treatment groups, respectively, exceeding the control group value of 2.69%. Furthermore, the proportion of persistent microbial taxa increased from 69.57% to approximately 82%, indicating selective enrichment and stabilization of the core microbiome responsible for AD through the combined effects of light and hemin. The results confirm that pretreatment with light and hemin effectively enhances the AD of cassava alcohol stillage.
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