Nitrite acts as the key “switch” governing the free nitrous acid pretreatment in anaerobic digestion: A comprehensive mechanism of abiotic, bioinformatics and bioenergetics effects

Free nitrous acid (FNA) pretreatment has been an effective and environmentally-friendly strategy to improve anaerobic digestion (AD) performance, and the mechanism was generally ascribed to the abiotic role of enhancing substrate solubilization. However, a comprehensive understanding of FNA is lack considering the different biotic roles on various steps, microbes and functional genes, as well as the variation of thermodynamic conditions. Herein, FNA pretreatment significantly enhanced the AD performance of food waste (FW), and the maximum increment of methane yield (36.45 %) was observed for FNA level of 1.42 mg HNO₂ − N/L. Nevertheless, FNA exhibited an inhibition-to-enhancement effects as the AD process proceeds, and the introduced nitrite was the key “switch”. Initially, the abiotic effects of FNA reduced the molecular weight of components in FW, benefiting the solubilization and formation of available organics (glucose and amino acid). The individual steps experiment confirmed that residual nitrite severely inhibited methanogenesis rather than hydrolysis and acidogenesis, which resulted in the decrement of methane yield and accumulation of VFAs. Meanwhile, the methanogens (Methanothrix and Methanobacterium) and genes involved in methanogenesis metabolisms (aceticlastic and hydrogenotrophic pathways) were downregulated. However, with the elimination of nitrite, the microbial activities and gene expressions restored and were even enhanced, and Gibbs-energy-based analysis suggested that accumulated VFAs provided favorable thermodynamic conditions for subsequent methanogenesis. This study firstly proved that the abiotic breakdown of macromolecule components, selective and reversible inhibition on microbes and functional genes induced by nitrite and varying thermodynamic conditions together governed the FNA affecting the AD process of FW.