Consequences Of Excess Urea Application On Carbon Fixation And Nitrogen Metabolism Of Robinia Pseudoacacia

Nitrogen as one of the essential mineral nutrients for plant growth and development,plays an important role in plant growth.Large amounts of nitrogen fertilizer are added to agriculture and horticulture to achieve and maintain high yields of food and biomass.Urea accounts for more than 50%of the global nitrogen fertilizer application.Nearly half of urea cannot be used by plants after application,but is released into the environment and accumulates continuously,causing a variety of problems such as nitrogen deposition.It was found that China has become the third largest nitrogen deposition area only after Europe and America.With the development of our country’s industry and agriculture,the atmospheric nitrogen deposition will be increased.Large dose of nitrogen deposition is certainly to affect the natural environment where plants are exposed to high concentration nitrogen,resulting in a series of potential negative effects.The roots of leguminous plants can symbiosis and nodulate with the rhizobia in soil to produce nitrogen for plant use.Robinia pseudoacacia L.is a typical nitrogen-fixing woody leguminous plant,has a wide distribution,developed root system and strong resistance to stess.It is a pioneer tree species of sand fixation to protect soil and afforestation barren mountain in our country.Therefore,we selected Robinia seedlings from Northeast China and tested their photosynthetic and physiological responses by inoculating rhizobia and applying different concentrations of urea to reveal the physiological response mechanism of Robinia to high nitrogen stress under symbiotic nitrogen fixation conditions.Three-month-old Robinia seedlings（inoculated with Mesorhizobium huakuii QD9）that were cultured normally in a greenhouse were selected as experimental materials for nitrogen stress treatment.Nitrogen stress treatment levels included normal nitrogen concentration(7 mmol·L^-1),high nitrogen concentration(57 mmol·L^-1)and extremely high nitrogen concentration(107 mmol·L^-1).The treatment method was to apply 50 mmol·L^-1 urea to the seedlings in the normal nitrogen treatment group once every 5 days for two consecutive times.Take the sampling from the day before the application of high concentration urea as the control group CK.Samples were collected 1,3,and 5 days after each urea application.After sampling,the biomass,leaf photosynthetic parameters,nitrogen content and distribution in leaves and roots,nitrogen metabolism and antioxidant metabolism related enzyme activities in leaves and roots of Robinia were determined and compared.The main research results are as follows:（1）Different urea application rates and exposure time had significant effects on the growth and photosynthesis of Robinia leaves and roots.Compared with normal nitrogen Robinia,the photosynthetic efficiency of Robinia leaves was significantly enhanced on the 1st and 3rd days after each urea application,and recovered to CK level on the 5th day.The root and nodule biomass increased significantly after the first urea application,but the total leaf and plant biomass did not change significantly.The leaves,roots,nodules and total biomass of Robinia increased after the second urea application.（2）Nitrogen content and distribution in leaves and roots of Robinia were significantly affected by different urea application rates and exposure time.Compared with normal nitrogen application,the total nitrogen content of root showed a dynamic increase after urea application of different concentrations.The first application of urea briefly increased the total nitrogen content in the roots of Robinia,but this effect gradually disappeared with the passage of time,and appeared again after the second application of urea.However,urea application had no significant effect on the total nitrogen content in leaves,and the root nitrogen content was significantly lower than that in leaves.The first application of urea significantly increased the content of soluble protein nitrogen in leaves.This effect was short-lived and reappeared after the second application of urea.The content of total amino acid nitrogen in leaves did not change significantly after the first application of urea,but only increased briefly after the second application of urea.Urea application also significantly reduced the nitrate nitrogen content in leaves,but the ammonium nitrogen and structural nitrogen contents had no significant changes.Urea application firstly increased soluble protein nitrogen content in root system,and then increased total amino acid nitrogen content in root system.The first application of urea significantly increased the content of total nitrogen and structural nitrogen in roots.This effect was short-lived and was repeated with the second application of urea.（3）Urea application and exposure time had significant effects on nitrogen metabolism and antioxidant metabolism of Robinia leaves and roots.Compared with normal nitrogen Robinia,NR activity in Robinia leaves and roots increased instantaneously after two urea applications.Different from NR,the activities of GS and NADH-GOGAT in leaves did not change significantly under different urea application rate and exposure time,and FD-GOGAT activities in leaves decreased with the increase of urea application rate and extension of treatment time.However,the activity of GS in roots increased only briefly after the first application of urea,but did not change significantly after the second application of urea.The activity of NADH-GOGAT in roots increased briefly after both urea application,but this effect disappeared over time.GR activity in leaves and roots was increased after the first urea application,and remained unchanged after the second urea application.However,the DHAR activity in leaves and roots did not increase after urea application,but showed a temporary decrease trend with the extension of exposure time.In conclusion,excessive urea application can significantly promote photosynthesis of Robinia.Urea application can promote the growth of Robinia,but its effect on overground biomass lags behind that of underground.Nitrogen from leaves and roots of Robinia under excessive urea application will be preferentially used to synthesize nitrogen forms that can effectively resist high nitrogen stress,so as to improve the stress resistance of Robinia.Different urea application rates can promote nitrogen metabolism in Robinia leaves and roots.In addition,excessive urea application to a certain extent can enhance antioxidant capacity of Robinia.The results can provide a theoretical basis for establishing the physiological response mechanism of robinia-rhizobia microbial interaction resistance system in forest trees,and provide scientific guidance for coping with the possible high nitrogen stress caused by nitrogen deposition.