Metabonomics Of Arabidopsis Thaliana In Response To Salt Stress

Salt stress is one of the important abiotic stress factors affecting crop yields worldwide.Metabolomics is used to analyze metabolic network changes caused by factors such as external stimuli,pathophysiological disturbances,and changes from genetic information.Since the dynamic changes of metabolites in plants can be visually understood,it is widely used in the study of stress response and is a good way to study plant resistance.In this study,the Arabidopsis thaliana was used as the experimental material,and the A.thaliana plants were subjected to salt stress with NaCl solution.Neutral desorption-extractive electrospray ionization mass spectrometry(ND-EESI-MS),ultra performance liquid chromatography coupled with quadrupole-time of flight mass spectrometry(UPLC-QTOF-MS)and high performance liquid chromatography(HPLC)were used to analyze the metabolites of A.thaliana under salt stress.Key research findings are as followed:1.A.thaliana was treated with 50,100,200 and 300 mM NaCl solution for 1,4,and d,respectively.In situ nondestructive analysis of A.thaliana rosette leaves was performed by using ND-EESI-MS.Fingerprints of ND-EESI-MS metabolites from A.thaliana leaves obtained under different stress conditions were determined by tandem mass spectrometry combined with related databases to identify 12 metabolites,including serine,arginine,glutamic acid,cinnamic acid,caffeic acid,ferulic acid,myricetin,protocatechuic acid,epicatechin,morin,apigravin and ?-cotonefuran.In response to salt stress,the contents of serine,arginine,glutamic acid and cinnamic acid,caffeic acid,ferulic acid and epicatechin involved in the metabolic pathway of shikimate-phenylpropanoid increased under 50,100,200 mM NaCl salt stress,and the change was the largest at 200 mM,but decreased under the maximum concentration of 300 mM NaCl.Both Principal Component Analysis(PCA)and Partial least squares discrimination analysis(PLS-DA)showed that the higher concentrations(200 mM and 300 mM)of the salt stress treatment group could be well distinguished from the experimental control group and the lower concentration(50 mM and 100 mM)salt stress treatment group.ND-EESI-MS technology can realize in situ in vivo detection of samples without sample pretreatment,and will not cause obvious damage to samples.Plants can continue to grow after the test,and the positioning and continuous observation of the same plant can be realized.The detection time of each sample is only 30 s,which can realize high-throughput and rapid detection.ND-EESI-MS analysis technology has certain prospects in the study of plant metabolomics.2.The UPLC-QTOF-MS non-targeted detection method was used to analyze the ethanol extract of A.thaliana under the stress of 200 mM NaCl solution,which identified 34 compounds such as phenylalanine,abscisic acid,malic acid,ferulic acid,cinnamic acid,rutin,galactose,uridine,and glucoerucin.In response to salt stress,the content of abscisic acid accumulated in A.thaliana.Phenylalanine content decreased significantly(p <0.05).Malic acid,protocatechuic acid,gallic acid,kaempferol and rutin were significantly reduced(p <0.05).Cinnamic acid,p-coumaric acid,caffeic acid and ferulic acid involved in the metabolic pathway of shikimate-phenylpropanoid were also reduced.The contents of cellobiose in glucose metabolism increased significantly(p <0.05).While the changes of glycosides were different,the contents of uridine,cyclic-GMP and glucoraphanin decreased significantly(p <0.05),the contents of glucobrassicin increased significantly(p <0.05).These metabolites are mainly involved in metabolic pathways such as amino acids,shikimate-phenylpropanoids and sugars,and A.thaliana responds to salt stress through a series of changes in these pathways.3.The phenolic substances in the leaves of A.thaliana leaves under the stress of 200 mM NaCl solution were detected by HPLC combined with standard reagents.Ten phenolic substances were detected.Compared to the experimental control group,the contents of protocatechuic acid,caffeic acid,rutin,cinnamic acid and morin in the salt stress treatment group decreased significantly(p <0.05),and the cinnamic acid content was reduced by more than 3 times;the contents of epicatechin and catechin were significantly increased(p <0.05),and the content of catechin was increased by nearly 3 times.The seven phenolic substances with significant difference were related to the metabolism of shikimic acid-phenylpropanoid in plants,and played a key role in the regulation of salt stress in A.thaliana.In this paper,ND-EESI-MS of the direct mass spectrometry was used to achieve rapid nondestructive detection of A.thaliana non-targeted metabolomics under salt stress.Based on ND-EESI-MS results,200 mM NaCl was further determined as the treatment concentration.UPLC-QTOF-MS and HPLC were used to study the metabolites of A.thaliana under salt stress.The results reveal the effects of salt stress on metabolites in A.thaliana at the level of metabolomics,which provides a theoretical basis for further revealing the mechanism of plant metabolism in response to salt stress.