Revealing the metabolic characteristics and nitrite accumulation mechanism of glycerol-driven partial denitrification process by batch tests and untargeted metabolomic analysis

Abstract

Glycerol has been proven as an effective carbon source for partial denitrification (PD), while the underlying mechanism of the nitrite accumulation remains unknown. In this study, the nitrite accumulation characteristics was firstly investigated by batch tests. Denitrifying bacteria exhibited high nitrate to nitrite ratio (NTR) (84.6%) and nitrite accumulation rate (NAR) (30.23 mg N h−1 gMLVSS−1) in a typical PD process with sufficient glycerol supply. But when glycerol was exhausted, denitrifying bacteria entered endogenous PD process driven by polyhydroxyalkanoates (PHA) and glycogen, and the NTR and NAR significantly decreased to 63.9% and 1.74 mg N h−1 gMLVSS−1, respectively. More analysis found that keeping the endogenous carbon source at a relatively high level before PD reaction could further improved the NTR up to 99.7%. Metabolomic analysis showed that most required amino acids were synthesized in nitrite accumulation phase with active RNA metabolism. While in the subsequent nitrite reduction phase with limited glycerol, microbes tended to synthesize those amino acids with lower energetic cost, and activities related to DNA metabolism was more active in this phase. Moreover, reactive oxygen species generated in the glycerol degradation were deduced to be the key factor that induced the efficient nitrite accumulation. This study provided a comprehensive insight into the mechanism of glycerol-driven PD process.