The effect of ¹⁵N to ¹⁴N ratio on nitrification, denitrification and dissimilatory nitrate reduction

RATIONALE Earlier experiments demonstrated that isotopic effects during nitrification, denitrification and dissimilatory nitrate reduction can be affected by high ¹⁵N contents. These findings call into question whether the reaction parameters (rate constants and Michaelis-Menten concentrations) are function of δ¹⁵N values, and if these can also lead to significant effects on the bulk reaction rate. METHODS Five experiments at initial δ¹⁵N-NO₃^- values ranging from 0° to 1700° were carried out in a recent study using elemental analyser, gas chromatography, and mass spectrometry techniques coupled at various levels. These data were combined here with kinetic equations of isotopologue speciation and fractionation. Our approach specifically addressed the combinatorial nature of reactions involving labeled atoms and explicitly described substrate competition and time-dependent isotopic effects. RESULTS With the method presented here, we determined with relatively high accuracy that the reaction rate constants increased linearly up to 270% and the Michaelis-Menten concentrations decreased linearly by about 30% over the tested δ¹⁵N-NO₃^- values. Because the parameters were found to depend on the ¹⁵N enrichment level, we could determine that increasing δ¹⁵N-NO₃^- values caused a decrease in bulk nitrification, denitrification and dissimilatory nitrate reduction rates by 50% to 60%. CONCLUSIONS We addressed a method that allowed us to quantify the effect of substrate isotopic enrichment on the reaction kinetics. Our results enable us to reject the assumption of constant reaction parameters. The implications of δ-dependent (variable) reaction parameters extend beyond the study-case analysed here to all instances in which high and variable isotopic enrichments occur.