Reduced isotope fractionation by denitrification under conditions relevant to the ocean

Experiments with two well-studied denitrifiers and one recently isolated marine suboxic zone denitrifier show that the cellular-level denitrification N isotope effect ( ¹⁵ε) is typically lower than the canonical value of ∼25‰ under many conditions prevalent in the ocean. Across all three strains, ¹⁵ε is 10-15‰ at cellular nitrate reduction rates that are more representative of the environment than the very high rates under which we and previous investigators measure ¹⁵ε to be 20-30‰. A sharp decrease in ¹⁵ε is also observed in individual nitrate drawdown assays as the extracellular nitrate concentrations approach 2-35μM and nitrate uptake becomes the rate-limiting step. On an apparently strain-specific basis, lower values of ¹⁵ε are observed under diverse conditions common in the natural environment: less reduced carbon sources, small inputs of oxygen, nutrient availability, agitation, and age of starter culture (i.e., initiation of assays with cells that had recently depleted a large previous nitrate amendment or were more recently in the exponential growth (" bloom" ) phase). A conserved oxygen-to-nitrogen isotope relationship across the experiments for all three denitrifiers ( ¹⁸ε/ ¹⁵ε=0.93±0.06 (1SD)) supports the interpretation that fractionation is imparted solely by the internal respiratory nitrate reductase, with the amplitude of ¹⁵ε varying with the proportional importance of cellular nitrate efflux relative to uptake. Aspects of the ¹⁵ε variation are unexpected; nevertheless, the occurrence of lower ¹⁵ε is robust. It is uncertain if our lower ¹⁵ε estimates apply to oceanic water column denitrification because field studies have generally yielded ¹⁵ε _wc between 20-30‰, more similar to previous culture estimates and our estimates at high cell specific nitrate reduction rates. If denitrification in the ocean's major suboxic zones does have an ¹⁵ε of ∼10-15‰, it would remove an apparent imbalance between global ocean N inputs and outputs previously suggested by fixed N isotope budgeting.