Enhancing caproate production in lactate-containing effluents by Megasphaera hexanoica through acetate modulation

The anaerobic bacterium Megasphaera hexanoica has great potential for caproate production through reverse β-oxidation using lactate, acetate, and butyrate as substrates. This study assessed the effect of acetate concentrations (10-50 mM) on caproate biosynthesis, lactate oxidation, and cell synthesis in M. hexanoica. At 30 mM acetate, caproate production reached 42.83 mM, with an electron efficiency of 67.03 % and a specific productivity of 4.47 g_CA·h^-1·g_DCW^-1. Subsequent fed-batch experiments with lactate, acetate, and butyrate maintained continuous caproate production, achieving 65.21 mM. Bioreactor assays further validated the strategy, yielding 65.25 mM caproate over 180 h. Mechanistic analysis demonstrated that 30 mM acetate optimized acetyl-CoA flux and enhanced caproyl-CoA transferase activity (3.5 U‧mg^-1), supporting caproate synthesis. Kinetic modeling demonstrated the Logistic model fit lactate consumption (R² = 0.991), while the Fitzhugh model captured caproate production (R² = 0.991, NRMSE = 1.137). The findings offer practical insights for industrial-scale caproate production.