Long-term hydrogenotrophic denitrification of high-nitrate synthetic and spring water in dual-recirculation UASB reactors

Hydrogenotrophic denitrification (H 2 Den) using hydrogen gas (H 2 ) as an electron donor and anaerobic granular sludge represents an effective strategy for nitrate (NO 3 - ) removal from organic carbon-deficient waters. This study investigates a novel-designed up-flow anaerobic sludge blanket (UASB) reactor, which exploits a dual-recirculation system (for both liquid and gaseous phases) to enhance the H 2 Den process. Two reactors were operated in biological duplicate for 420 days under varying conditions, including different hydraulic retention times (HRTs) and H 2 supply amounts. Optimal conditions (i.e., 200% excess H 2 and 1 day HRT) resulted in minimal nitrite (NO 2 - ) accumulation (< 0.5 mg NO 2 - /L), and a NO 3 - removal efficiency up to 99% maintained over time, with a denitrification rate up to 166.6 ± 61.8 mg NO 3 - /L/d, also when validating the process in the presence of a low organic carbon-to-nitrogen spring water amended with NO 3 - . As regards greenhouse gas emissions, the nitrous oxide (N 2 O) emission rate under optimal conditions was 5.3·10 -3 μg N 2 O/min. Microbial community revealed an enrichment of the Comamonadaceae (i.e., from 1.7% to 19.3%) family, known to be actively involved in hydrogenotrophic denitrification, as well as of Spirochaetaceae (i.e., from 3.3% to 19.3%), frequently detected in anoxic environments but with a yet unexplored potential for nitrate removal.