A convenient method for simultaneous quantification of multiple phytohormones and metabolites: application in study of rice-bacterium interaction

A method was developed for simultaneous quantification of multiple phytohormones, abscisic acid, indole-3-acetic acid (IAA), jasmonic acid (JA), and salicylic acid, hormone conjugates, IAA-aspartic acid, JA-isoleucine, and methyl JA, and phytoalexins, momilactone A, naringenin, and sakuranetin. This method combines a convenient procedure for preparing filtrated crude extracted samples and a sensitive quantification assay using ultra fast liquid chromatography-electrospray ionization tandem mass spectrometry (UFLC-ESI-MS). With this method, we determined the dynamic profiles of defense-related phytohormones, hormone metabolites, and phytoalexins in the interaction of rice with Xanthomonas oryzae pv. oryzae (Xoo), which causes bacterial blight, one of the most devastating diseases of rice worldwide.This UFLC-ESI-MS method is convenient, sensitive, reliable, and inexpensive for quantification of multiple phytohormones and metabolites compared to current methods. The results obtained by application of this method in studying rice-bacterial interaction provide a basis for understanding the molecular mechanisms of rice defense responses.Phytohormones are essential for the regulation of diverse physiologic processes of plants, including development, growth, reproduction, and responses to biotic and abiotic stresses. Plant-produced hormones include nonpeptide hormones, abscisic acid (ABA), auxin or indole-3-acetic acid (IAA, the major form of auxin in most plants), brassinosteroids, cytokinins, ethylene, gibberellins, jasmonic acid (JA), nitric oxide, salicylic acid (SA), and strigolactones, and peptide hormones [1,2]. The nonpeptide phytohormones are structurally unrelated small molecules. Phytohormones act as signal molecules in biological activities and frequently occur in low concentration. The homeostasis of these hormones is tightly controlled between the biosynthetic and metabolic pathways. The metabolism of nonpeptide phytohormones is generally categorized into thre