H-NMR-based metabolomic profiling and proteomic analysis of soybean ( L.) in response to dicarboxylic acids (photon) application as a stress priming agent.

Soybean ( L.) serves not only as food for humans, animals, and industrial purposes, but is also a plant that can be used to comprehend molecular mechanisms occurring in stress response to various development techniques. To reveal the effect of applying dicarboxylic acids as stress priming agents on a metabolic level in soybean leaf extracts, the chemical profile of methanolic extracts were collected at different time points (1 h, 2 h, 12 h, 24 h, 1 week, 2 weeks and 3 weeks) after spraying were analyzed using H-NMR based metabolomics by way of PCA and OPLS-DA. The OPLS-DA revealed several metabolites, including organic acids (fumarate, citrate and malate) and amino acids (asparagine, alanine and GABA), which accumulated in higher amounts, with fumarate accumulating the highest in L. leaf extracts compared to untreated leaves. Denaturing 1DE gels were prepared for MS-based protein analysis and the presence of fatty acids (linolenic, oleic and α-linolenic acid) were confirmed by gas chromatography coupled with mass spectrometry (GC-MS), which served as jasmonic acid precursors. The MS-based profiling of proteins on the denaturing 1DE gels revealed several proteins that were differentiated between the treated and untreated leaf extracts. These proteins included ferritins, CaM, ferredoxin-thioredoxin reductase and chalcone-flavanone isomerase 1A. Following the treatment, fumarate was significantly elevated at 12 h to 3 weeks, compared to other compounds. It is, therefore, proposed that elevated quantities of fumarate could be related to the KEAP1-NRF2 metabolic pathway. This study represents the initial investigation of the effect of dicarboxylic acid application as a stress priming agent on L. using H-NMR metabolomic analysis, GC-MS and proteomic analysis.