Inhibitory effects of nitrate, nitrite, NO and N 2O on methanogenesis by Methanosarcina barkeri and Methanobacterium bryantii

Abstract

In order to elucidate the mechanism of the inhibitory effect of nitrate and its denitrification intermediates nitrite, NO and N 2O on methanogenesis in anoxic environments, we tested possible toxic effects of these N-compounds on the methanogenic bacteria Methanosarcina barkeri and Methanobacterium bryantii which are ubiquitous in methanogenic rice field soils. The different N-compounds inhibited H 2-dependent methanogenesis by these bacteria to different extents. Nitrate showed the weakest inhibition of methanogenesis in both bacteria, followed by N 2O and nitrite for Ms. barkeri, and nitrite and N 2O for Mb. bryantii, respectively. In both bacteria, the strongest inhibition was caused by NO. Concentrations of 30 mM nitrate still enabled a CH 4 production rate of 25–40% of that before the addition of the N-compound, whereas NO completely inhibited methanogenesis at concentrations ≥0.8–1.7 μM (equivalent to 50–100 Pa NO partial pressure). Removal of NO by replacing the atmosphere with H 2/CO 2 (8:2) resulted in resumption of methanogenesis only if the bacteria had been treated with NO concentrations ≤0.8 μM (50 Pa). Removal of N 2O from the cultures resulted in resumption of methanogenesis if Mb. bryantii had been treated with ≤95 μM N 2O (500 Pa) or Ms. barkeri with ≤950 μM N 2O (5 kPa). These results show that the denitrification products of nitrate can inhibit CH 4 production both reversibly and irreversibly depending on the type of methanogenic bacterium and the applied concentration of the N-compound. In a separate experiment with methanogenic rice field slurries addition of nitrate resulted in immediate inhibition of CH 4 production. Nitrate was consumed resulting in the sequential accumulation of nitrite, NO and N 2O which were subsequently utilized. Nitrite and N 2O reached maximum concentrations that would have been inhibitory in the methanogenic bacterial cultures examined.