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Influence of environmental factors on nitrate utilization and nitrogen isotope fractionation by marine phytoplankton Needoba, Joseph Andrew


Nitrate utilization by marine phytoplankton is an important component of the marine nitrogen cycle. The ¹⁵N / ¹⁴N ratio that is created during nitrate assimilation can be propagated throughout marine food webs, throughout nitrogen pools in the water column, and recorded in the organic fraction of marine sediments. Variations in the ¹⁵N / ¹⁴N ratio are potentially useful for understanding processes of the nitrogen cycle on a variety of spatial and temporal scales. Our present understanding of the processes involved in nitrogen isotope fractionation does not allow us to confidently interpret the ¹⁵N / ¹⁴N signal in the various pools of nitrogen in the ocean. Using laboratory batch culture experiments, the uptake and assimilation characteristics of nitrate during growth in light, temperature, and iron limiting conditions were determined as a means to investigate the variability and possible physiological mechanisms of nitrogen isotope fractionation by marine phytoplankton. Irradiance level had the most significant impacts on nitrate assimilation and nitrogen isotope fractionation. The phytoplankton species that expressed an uncoupling between the uptake and assimilation processes under low light conditions had relatively large isotope fractionation factors. The marine diatom Thalassiosira weissflogii showed the largest isotope discrimination, with an observed epsilon value of 6.2 per mil in high light, and 15.2 per mil in low light. Under a 12h lightdark cycle, the isotope fractionation factor was dependent on the light versus dark nitrate assimilation rates. A coccolithophorid, Emiliania huxleyi, with no dark nitrate assimilation, had the same fractionation compared to growth in 24 hours of continuous light, whereas three marine diatoms displayed higher fractionation factors during nitrate uptake in the dark, and therefore had higher fractionation factors than when grown in 24 hours of continuous light. The growth limiting effects of temperature or iron had little influence on isotope fractionation in the two marine diatoms that were measured. Despite the lower growth rate, isotope fractionation was similar to conditions of higher temperature (18°C) and high iron concentrations in cultures of Thalassiosira weissflogii and Thalassiosira pseudonana. These results indicate that the mechanism of isotope fractionation is not altered by slow growth rates induced by temperature or iron limitation. Lastly, the isotope fractionation mechanism of Thalassiosira weissflogii was investigated by measuring the ¹⁵N / ¹⁴N ratio of nitrate inside and outside the cell during growth in the four growth conditions described above. The results show that the ¹⁵N / ¹⁴N ratio is higher in the internal pool of nitrate than in the nitrate in the medium, and therefore the fractionation step that produces the overall phytoplankton δ¹⁵N value is a result of assimilation processes inside the cell. The ¹⁵N / ¹⁴N values of the internal nitrate pool in the different growth conditions suggested that the overall isotope fractionation by marine phytoplankton is a result of the isotope discrimination step during nitrate reduction combined with the efflux of nitrate from the cytoplasm to the external medium. A linear correlation was observed between the magnitude of isotope fractionation and the relative efflux: influx ratio that suggests that fractionation associated with nitrate reductase was 23 per mil. The results and discussion in this thesis describe how isotope fractionation by marine phytoplankton varies as a result of species composition, nutrient, and light conditions. The variations in the magnitude of nitrogen isotope fractionation imply that an accurate interpretation of a specific nitrogen isotope signal will depend on knowledge of the growth parameters that control phytoplankton species composition and nitrate utilization.

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