Nitrogen Isotopic Fractionation During The Growth Of Freshwater Algae Under Different Nitrogen Sources

The algae bloom in rivers/reservoirs and lakes is caused by eutrophication,and the related changes in the elements cycle are very important environmental issues.Nitrogen stable isotope technology is widely used to analyz the transformation of nitrogen element and trace the sources of water environmental pollution.However,the isotopic composition of nitrogen is not only related to its source,but also affected by biological effects?such as assimilation?during nitrogen transformation rate in aqutic system.The study of nitrogen isotope fractionation effects during freshwater algae assimilation of different nitrogen sources is a non-negligible part of the nitrogen biogeochemical process in lake/reservoir systems,which is of great significance for accurately tracing the source of nitrogen and improving the theory of nitrogen biogeochemical cycles,as well as for the water quality management of lake/reservoir environmental systems.In this work,freshwater green algae-Chlamydomonas reinhardtii was used as research object,and different nitrogen sources systems were set,including two inorganic nitrogen sources?NO₃^-and NH₄⁺?,one organic nitrogen source?urea,Org-N?and two mixed nitrogen sources?NO₃^-&NH₄⁺and NO₃^-&Org-N?,aiming to understand the utilization of different nitrogen sources and isotopic fractionation effects during the growth of Chlamydomonas reinhardtii through laboratory culture.The cultural growth cycle of Chlamydomonas reinhardtii includes the growth phase,the stationary phase and the decline phase.The experiment resultes of single nitrogen sources showed that the consumption rate of NH₄⁺was the fastest,followed by urea and NO₃^-during the growth of Chlamydomonas reinhardtii,which was related to the selective absorption of different forms of nitrogen by algae cells;in the mixed nitrogen sources culture system,compared with NO₃^-,Chlamydomonas reinhardtii prefers to absorb NH₄⁺;while algae cells had higher consumption rate of NO₃^-than urea,which may be related to the number of specific transport proteins of different forms of nitrogen in Chlamydomonas reinhardtii cells.In all culture treatments,organic carbon was released during the growth of Chlamydomonas reinhardtii,including a small amount of organic carbon released by metabolism during the growth phase and a large amount of organic carbon released by algae cells death.Analyzing the nitrogen isotopic compositions of the remaining nitrogen sources in culture media and particulate nitrogen of algal in different time intervals,it showed that the?¹⁵N value in the growth phase of Chlamydomonas reinhardtii increased linearly with the decrease of nitrogen concentration.The isotopic enrichment factor?¹⁵N and?¹⁸O of the remaining nitrogen sources in the NO₃^-system were 2.8‰and5.1‰,respectively,and with a ratio 1.8:1 for?¹⁸O:?¹⁵N;while the nitrogen isotopic enrichment factors of the remaining nitrogen sources in the NH₄⁺and Org-N systems were 14.2‰and 1.7‰,respectively.The?¹⁵N_p value of the particulate nitrogen decreased rapidly in the early growth stage of Chlamydomonas reinhardtii,while the?¹⁵N_p value gradually increased in the later growth stage,and finally reached the isotope value similar to the initial nitrogen sources.The particulate nitrogen isotope enrichment factors of three singel nitrogen sources?NO₃^-,NH₄⁺and Org-N?were4.5‰,13.0‰and 2.6‰,respectively.The results indicated that the sequence of isotope enrichment factors of water-soluble nitrogen and particulate nitrogen are same,which was?_NH4+>?_NO3->?_Org-N.In summary,The above experimental results showed that freshwater algae preferentially absorb ammonium nitrogen,then nitrate nitrogen,and finally organic nitrogen.Therefore,controlling the discharge of point-sourced ammonium nitrogen in the catchment is an important measure to improve the quality of water environment.