Two-phase anaerobic digestion of long chain fatty acids under sulfate regulation: Effects of volatile fatty acids production, microbial community structure, and metabolic pathways

This study explored the effect of sulfate ( SO 4 2 − ) on long-chain fatty acids (LCFAs) transformation in a two-phase anaerobic digestion (AD) system. After 35 days of long-term acclimation of low concentration SO 4 2 − (400 mg/L) and sodium oleate (1 g/L), the total volatile fatty acids (TVFAs) production of the system was significantly increased. The sulfate-supplemented reactor (Ra LS ) achieved a maximum TVFAs yield of (2083 ± 184) mg/L, which was 54% higher than that of the sodium-oleate-only reactor (Ra L ), while the methanogenic phase (Rm LS ) was inhibited. As per microbial and functional analyses, sulfate addition not only enhanced acidogenesis even under high LCFA loading (10 g/L) but enriched unclassified_f_Enterobacteriaceae alongside sulfate-reducing bacteria (SRB). Consistently, PICRUSt2 metabolic predictions revealed that SO 4 2 − not only strengthened propionate biosynthesis but also stimulated sulfate reduction via dissimilatory sulfite reductase pathway. These findings clarify sulfate-driven VFAs formation mechanisms and offer strategies for efficient carbon recovery from LCFA-rich wastewater. • Prolonged exposure to 400 mg/L sulfate and 1 g/L oleate increased TVFAs by 54% • High oleate (10 g/L) inhibited acidogenesis, while sulfate alleviated it • Sulfate enriched Enterobacteriaceae and SRB, thereby reshaping communities • The DSR pathway dominated sulfate reduction in Ra LS