Effects Of Humic Acid On Cucumber (Cucumis Sativus L.) Growth And Physiological Metabolism Under Nitrogen Stress

Humic acid?HA?could improve soil water holding capacity,reduce soil compaction and make soil structure better,increase soil fertility,stimulate plant growth and increase plant stress resistance.In recent years,many experts in the world focused on the research of HA as fertilizer and pesticide additives.In China,though there were many researches about the application of HA as fertilizer additives,the unpurified humic acid and its complicated components limited the mechanism study.There was little research reported on the direct effects of HA on plant growth improvement,especially on its stimulatory mechanism.Cucumber?Cucumis sativus L.?is one of the largest areas of cultivation vegetable crops.It is sensitive to nitrogen supply and is prone to salt damage in the cultivation process.The excess NO₃^-in facility soil limited the vegetables growth and development.It is significant to clarify the mechanism of HA on cucumber seedlings physiological characteristics under nitrogen stress with theoretical and practical meaning for increasing the crops stress resistance and fertility efficiency,and comprehensively prevented greenhouse soil secondary salinization.In order to clarify physiological mechanism,the dissertation studied the effects of purified humic acid?PHA?on cucumber growth and nutrients uptake under nitrogen stress,as well as on carbon and nitrogen metabolism,ROS metabolism,photosynthesis and fluorescence characteristics,signaling molecular and endogenous hormones,and further studied PHA soaking on cucumber seed germination and physiological characteristics.The dissertation aimed at clarifying PHA physiological mechanism under nitrogen stress and providing a theoretical basis for suitble application of humic acid in facilities vegetables cultivation.The main results are as follows:1.Effects of PHA on growth and nutrients uptake of cucumber seedlings under nitrogen stressTaking ‘Xintaimici' and ‘Jinyou 1' as experimental cucumber cultivars,this section studied the effects of watering PHA different level on cucumber growth and nutrients uptake under low nitrogen(1.0 mmol·L^-1 NO₃^-)stress in sand cultivation pots.The results showed that PHA could promote ‘Xintaimici' cucumber seedlings height growth,stem diameter and root growth and increase root surface area,root volume and the number of tips.The maximum leaf area obtained from 100 mg·L^-1 PHA treatment.The plant height,stem diameter of ‘Jinyou 1' cucumber showed insensitive to PHA treatments.The leaf area,root/shoot,total root length,root surface area and root volume obtained from 150 mg·L^-1 PHA treatment are significantly higher than other treatments.The proline and soluble sugar content in leaves of two cultivars were significantly increased by PHA application.The total absorption of N in P,K,Ca,Mg,Fe content in cucumber seedlings roots,stems,leaves were promoted by PHA under low nitrogen stress.2.Effects of PHA on growth,carbon and nitrogen metabolism of cucumber seedlings under nitrogen stressTaking ‘Xintaimici' as experimental cucumber cultivar,this section studied the effects of PHA on cucumber growth and carbon and nitrogen metabolism under low nitrogen(1.0 mmol·L^-1 NO₃^-)stress and high nitrogen(101 mmol·L^-1 NO₃^-)stress in hydroponics compared with general level of nitrogen(11 mmol·L^-1 NO₃^-)which was CK.The results showed that nitrogen stress treatments inhibited the growth of cucumber seedlings.The plant height,stem diameter,leaf area and dry matter accumulation was decreased.PHA significantly increased dry matter accumulation of the general level and low nitrogen stress treatments,but there was no significant difference within cucumbers grown under high nitrogen stress.PHA could affect the absorption of NO₃^-in cucumber seedlings,increasing nitrate content under low nitrogen stress and reducing it under high nitrogen stress.PHA significantly reduced ammonium content in roots and leaves under nitrogen stress.Compared with general level of nitrogen?CK?,NR,GS,GOGAT,GDH activity in roots and leaves and NiR activity in roots decreased significantly under nitrogen stress.PHA increased NR,NiR,GS,GOGAT,GDH activity with varying degrees.PHA increased amino acid and proline content in cucumber roots and leaves.The P5 CS,P5CR and ProDH activity were no significantly different between different treatments.PHA increased soluble protein content of cucumber seedling roots and leaves and it was related to protease activity.In carbon metabolism,PHA improve SPS activity under nitrogen stress and improve SS activity in roots,and increased soluble sugar content of cucumber roots and leaves.3.Effects of PHA on ROS metabolism of cucumber seedlings under nitrogen stressCompared with general level nitrogen(11 mmol·L^-1 NO₃^-),in cucumber roots and leaves,O2·-production rates and H2O2 content in leaves were slight increased under low nitrogen,but significantly reduced under high nitrogen stress.SOD activity in roots and leaves were increased under nitrogen stress,and were also significantly increased in PHA treatments.PHA significantly enhanced POD activity under general level and high nitrogen stress.Root CAT activity under nitrogen stress was increased,while the leaves CAT activity was decreased.PHA significantly increased CAT activity in leaves.Roots and leaves AsA,DHA and?AsA+DHA?content decreased under low nitrogen stress and increased under high nitrogen stress,but AsA/DHA was the opposite trend.PHA could promote the content of ascorbic acid generally.Compared with CK,GSH,GSSG and?GSH+GSSG?content in roots were significantly increased under nitrogen stress.GSH,GSSG,GSH+GSSG in leaves decreased under low nitrogen stress and increased under high nitrogen stress.PHA increased the root and leaves?GSH+GSSG?content and increased the GSH/GSSG in roots.PHA changed AsA,GSH contents by regulating related enzymes activity in AsA-GSH cycle.Compared with CK,cucumber roots APX activity increased significantly under nitrogen stress,and GR activity significantly decreased.PHA increased APX and GR activity.Cucumber roots MDHAR activity and DHAR activity decreased under low nitrogen stress and increased under high nitrogen stress,while in the leaves it was the opposite trend.PHA improved MDHAR and DHAR activity.PHA increased roots and leaves of the H+-ATPase activity under high nitrogen stress,and increased Ca2+-ATPase activity in roots and leaves.4.Effects of PHA on photosynthesis and fluorescence characteristics of cucumber seedlings under nitrogen stressCompared with general level of nitrogen(11 mmol·L^-1 NO₃^-),the photosynthesis pigments significantly decreased under low nitrogen stress and there was no significance in high nitrogen stress with CK.PHA increased pigments content in leaves.RuBPCase activity was increased under low nitrogen stress and CK.Pn,Gs,Tr were significantly decreased under nitrogen stress.PHA increased Pn and Gs under CK and low nitrogen stress treatment,but there was no significance under high nitrogen stress in Pn and Gs,but Tr was decreased.PHA reduced Ci index.PHA increased Ls for each treatment,and improved WUE.Fv/Fm,?PSII,qP,qN and ETR were decreased under nitrogen stress,while Fv'/ Fm' increased.PHA could improve Fv/Fm,?PSII,q P,ETR,Fv'/ Fm'.PHA in creased Pc,and reduced Hd,Ex,improving the light use efficiency.Fast Chlorophyll fluorescence induction danamics showed that under nitrogen stress,fluorescence is higher than CK,and PHA decreased fluorescence dissipation.PHA increased PIABS and DFABS.Under nitrogen stress,?Ro and ?o decreased,while ?Eo and ?Do increased.PHA showed no significant effects to ?Ro,?o,?Eo,?Do.PHA increased quinone acceptor side capacity significantly under high nitrogen stress.Fk-j and Fi-p was significantly reduced under nitrogen stress,and PHA could increase Fk-j and Fi-p under nitrogen stress.5.Effects of PHA on endogenous signaling molecule and endogenous hormone of cucumber seedlings under nitrogen stressCompared with general level of nitrogens(11 mmol·L^-1 NO₃^-),NO content in roots under nitrogen stress increased significantly,while opposite in leaves.When added PHA,NO content in roots and leaves increased under low nitrogen and decreased under high nitrogen stress.NOS-like activity in roots decreased under low nitrogen stress and increased under high nitrogen stress.When adding PHA,roots NOS-like activity increased in all treatments and leaves NOS-like activity decreased with no significant difference under high nitrogen stress.IAA concentration in roots increased under low nitrogen stress while decreased by adding PHA.IAA concentration in leaves decreased under low nitrogen stress and increased under high nitrogen stress.PHA increased leaves IAA concentration under low nitrogen stress.Compared with general level,ABA concentration was significantly reduced under nitrogen stress in roots and leaves.PHA increased the ABA concentration in roots.Compared with CK,IAA/ABA was increased under nitrogen stress,and IAA/ABA was decreased by PHA.6.Effects of PHA on cucumber seed germination and physiological characteristics under nitrogen stressTaking ‘Xintaimici' and ‘Jinyou 1' as experimental cucumber cultivars,this section studied the effects of soaking PHA different level on cucumber seed germination and physiological characteristics under high nitrogen(75 mmol·L^-1 NO₃^-)stress in Petri dishes.The results showed that seed germination,radicle length,lateral root length,lateral root number,hypocotyl length and diameter,and seedling fresh weight were decreased under NO₃^-stress.The above indices of adding PHA treatments were increased obviously and showed that PHA could partly alleviate NO₃^-stress.The seed cotyledon ?-amylase activity was decreased but seedling electrolyte leakage rate and radicle dehydrogenase activity was increased under nitrogen stress.MDA and soluble protein content,SOD,POD,CAT activity were increased.PHA significantly reduced the seedling electrolyte leakage rate,but with the increasing concentration of PHA the decreases difference became smaller.Cotyledon ?-amylase activity and radicle dehydrogenase activity were increased and showed downward trend when added high PHA concentration.Radicle and hypocotyl MDA content decreased when treated with PHA but increased with high PHA concentration.Soluble protein content were increased but decreased with high PHA concentration.SOD,POD,CAT activity showed a downward trend treated with PHA compared with NO₃^-stress.