Nitrate and nitrite inhibition of methanogenesis during denitrification in granular biofilms and digested domestic sludges

Anaerobic bioreactors that can support simultaneous microbial processes of denitrification and methanogenesis are of interest to nutrient nitrogen removal. However, an important concern is the potential toxicity of nitrate (NO₃^-) and nitrite (NO₂^-) to methanogenesis. The methanogenic toxicity of the NO_x^- compounds to anaerobic granular biofilms and municipal anaerobic digested sludge with two types of substrates, acetate and hydrogen, was studied. The inhibition was the severest when the NO_x^- compounds were still present in the media (exposure period). During this period, 95% or greater inhibition of methanogenesis was evident at the lowest concentrations of added NO₂^- tested (7.6-10.2 mg NO₂^--N l^-1) or 8.3-121 mg NO₃^--N l^-1 of added NO₃^-, depending on substrate and inoculum source. The inhibition imparted by NO₃^- was not due directly to NO₃^- itself, but instead due to reduced intermediates (e.g., NO₂^-) formed during the denitrification process. The toxicity of NO_x^- was found to be reversible after the exposure period. The recovery of activity was nearly complete at low added NO_x^- concentrations; whereas the recovery was only partial at high added NO_x^- concentrations. The recovery is attributed to the metabolism of the NO_x^- compounds. The assay substrate had a large impact on the rate of NO₂^- metabolism. Hydrogen reduced NO₂^- slowly such that NO₂^- accumulated more and as a result, the toxicity was greater compared to acetate as a substrate. The final methane yield was inversely proportional to the amount of NO_x^- compounds added indicating that they were the preferred electron acceptors compared to methanogenesis.