A comparison of different approaches for measuring denitrification rates in a nitrate removing bioreactor

Denitrifying woodchip bioreactors (denitrification beds) are increasingly used to remove excess nitrate (NO3−) from point-sources such as wastewater effluent or subsurface drains from agricultural fields. NO3− removal in these beds is assumed to be due to microbial denitrification but direct measurements of denitrification are lacking. Our objective was to test four different approaches for measuring denitrification rates in a denitrification bed that treated effluent discharged from a glasshouse. We compared these denitrification rates with the rate of NO3− removal along the length of the bed. The NO3− removal rate was 8.73 ± 1.45 g m−3 d−1. In vitro acetylene inhibition assays resulted in highly variable denitrification rates (DRAI) along the length of the bed and generally 5 times greater than the measured (NO3−—N removal rate. An in situ   push–pull test, where enriched N15—NO3− was injected into 2 locations along the bed, resulted in rates of 23.2 ± 1.43 g N m−3 d−1 and 8.06 ± 1.64 g N m−3 d−1. The denitrification rate calculated from the increase in dissolved N2 and N2O concentrations (DRN2) along the length of the denitrification bed was 6.7 ± 1.61 g N m−3 d−1. Lastly, denitrification rates calculated from changes in natural abundance measurements of δ15N–N2 and δ15N—NO3− along the length of the bed yielded a denitrification rate (DRNA) of 6.39 ± 2.07 g m−3 d−1. Based on our experience, DRN2DRN2 measurements were the easiest and most efficient approach for determining the denitrification rate and N2O production of a denitrification bed. However, the other approaches were useful for testing other hypotheses such as factors limiting denitrification or may be applied to determine denitrification rates in environmental systems different to our study site. DRN2DRN2 does require very careful sampling to avoid atmospheric N2 contamination but could be used to rapidly determine denitrification rates in a variety of aquatic systems with high N2 production and even water flows. These measurements demonstrated that the majority of NO3− removal was due to heterotrophic denitrification.

► The majority of NO3− removal was due to heterotrophic denitrification in the denitrifying bioreactor. ► Measurement of dissolved N2 and N2O along the length of the bioreactor is an appropriate technique to measure denitrification rates. ► In situ   push pull technique using enriched N15—NO3− is useful for measuring denitrification rates at specific locations. ► In lab acetylene inhibition largely overestimated the denitrification rate in denitrifying bioreactors. ► Changes in δ15N—NO3− and δ15N–N2 determined the proportion of NO3− removed by denitrification in denitrification beds.