Salinity-induced solubilization and betaine protection synergistically boost anaerobic acidogenesis of waste activated sludge: Osmoprotection mechanisms and life cycle assessment

ABSTRACT Anaerobic fermentation is crucial for sludge resource recovery, but its efficiency is limited by slow hydrolysis. Although salinity enhances hydrolysis and organic matter release, it severely inhibits microbial activity. This study exploits the synergistic interplay between salinity-induced solubilization and betaine-mediated salt-stress alleviation to enhance the synthesis of volatile fatty acids (VFAs). NaCl promoted massive organic matter release from sludge, with the SCOD yield reaching 664 mg COD/g VSS at 54 g/L NaCl. Betaine elevated the activities of hydrolases and antioxidant enzymes and VFA production, with a yield of 308 mg COD/g VSS at 36 g/L NaCl and 6 mM betaine, representing a 122% increase compared with the salinity control and an 863% increase relative to the blank control. Metagenomic and pathway analyses further revealed that under salt stress, the relative abundances of genes involved in maintaining ionic homeostasis and synthesizing extracellular polymeric substances (EPS) were lower, and that exogenous betaine may enable cells to reallocate more energy toward acidogenesis while maintaining relatively high microbial community diversity and acidogenic bacteria abundance. Betaine upregulated pyruvate, acetyl-CoA and acetate synthetic genes, jointly driving targeted acetate accumulation and acidification. According to life cycle assessment (LCA) and life cycle cost (LCC), the betaine groups reduced environmental burdens and total LCC by 30%–55% versus the betaine-free groups, with internal costs dominating overall expenses. These findings provide mechanistic insights into betaine-mediated salt stress alleviation and offer a feasible strategy for enhancing acid production efficiency under high-salt treatments.