Recent ¹⁵N tracing studies have highlighted the important role biotic assimilation plays in stream N retention, yet the fate of N following assimilation is not well understood. One potentially important fate is indirect denitrification, a process in which locally mineralized and nitrified N is denitrified before being exported to the water column. We conducted a series of in-situ chamber experiments in which patches of stream bottom were labeled with ¹⁵N to investigate the fate of assimilated N. We measured nitrate transformations, including uptake and denitrification, at the start of the experiment and continued to monitor dissolved N species (including N₂) for isotopic enrichment in the patches over the next month. Direct denitrification (denitrification of nitrate from the water column) was an order of magnitude higher in cobble habitats than in sand habitats, accounting for ~26% and ~2% of nitrate uptake in cobble and sand, respectively. Mean ¹⁵N turnover times of benthic biomass were 18 days at cobble sites and 10 days at the sand sites. Production of ¹⁵N ammonium and nitrate by stream biofilms we approximately equal, and accounted for a majority of the loss of assimilated N. We detected slight enrichment of N₂ gas in cobble and sand habitats, indication that indirect denitrification was taking place. However, indirect denitrification only accounted for <0.1% of the assimilated N, suggesting that assimilation is only a short-term N retention mechanism.