Exploring Molecular Dynamics and Structure Change of Dissolved Organic Nitrogen under Nitrate and Phosphate Pulses in a Cascade Upflow Biofiltration System

Dissolved organic nitrogen (DON) remains a large, reactive, and under-managed fraction of stormwater nitrogen. We investigated how influent N:P stoichiometry and porous media chemistry jointly shape DON fate in a cascade upflow bioreactor packed with two specialty adsorbents, including BIPGEM (sand-clay-ZVI-perlite-biochar) and ZIPGEM (sand-clay-ZVI-perlite). Reactors received short pulses of nitrate alone (AC1) or nitrate plus phosphate simultaneously (AC2). Paired influent-effluent samples were analyzed using a 21 T Fourier transform ion cyclotron resonance mass spectrometer. Molecular formulas containing nitrogen (CHON formulas) were then classified as removed, produced, or retained. AC2 reprograms produced DON chemistry in a media-specific manner. BIPGEM shifted toward heavier, unsaturation rich products with lower inferred lability, whereas ZIPGEM shifted toward more oxidized, aliphatic products with higher inferred lability and preferential removal of unsaturated formulas. Bulk measurements showed moderate nitrate removal in both ZIPGEM and BIPGEM. However, phosphate removal was strongest in ZIPGEM under AC2, while BIPGEM released phosphate under AC1. These findings highlight influent N:P and media selection/ordering as levers to control nutrient removal and residual DON quality, with implications for downstream nitrogen cycling and stormwater treatment performance.