The mechanism of isotope fractionation during algal nitrate assimilation as illuminated by the ¹⁵N/¹⁴N of intracellular nitrate

The ¹⁵N/¹⁴N of nitrate in the external medium and intracellular pool of the cultured marine diatom Thalassiosira weissflogii (Grun.) Fryxell et Hasle was measured during nitrate assimilation under low light, a 12:12-h light:dark cycle, low temperature, or low iron conditions. The ¹⁵N/¹⁴N of the nitrate in the medium and the particulate matter both followed the predicted Rayleigh fractionation model, and the intracellular nitrate always had a higher ¹⁵N/¹⁴N than did the medium nitrate. When the experiments were compared, the results showed a negative correlation between the isotope fractionation factor and the difference in the ¹⁵N/¹⁴N between the two pools of nitrate. These observations imply that the variations in the isotope effect result from variations in the degree to which the fractionation by nitrate reductase is expressed outside the cell, which is, in turn, controlled by the rate of nitrate efflux relative to nitrate reduction. The low iron and low temperature experiments showed relatively small isotope effects but a large intracellular-medium difference in nitrate ¹⁵N/¹⁴N, consistent with a relative rate of efflux (compared with influx) that is small and similar to fast-growing cells. In contrast, large isotope effects and small intracellular-medium differences in nitrate ¹⁵N/¹⁴N were observed for low light and light:dark cycle grown cells and are explained by higher relative rates of nitrate efflux under these growth conditions.