| Jasmonic acid (JA), a linolenic acid-derived cyclopentanone, is a new kind of plant hormone. It plays regulatory roles in plant development and responses to environmental stresses, such as mechanical wounding, fungal infection, insects and microbial pathogen attack, drought and osmotic stress. In addition, there have been numerous reports on the physiological effects of jasmonates, including the inhibition of seed and pollen germination, stem growth, cell expansion and multiplication and the promotion of senescence abscission and tuber formation. Moreover, recent studies showed that JA could regulate the expression of many genes involved in plant development or in stress tolerance.In general, the concentration of JA in plant tissues is very low, as those of the other plant hormones. The quantification of endogenous JA level in plant extracts by means of a sensitive and reliable method is of interest because of the significance of these compounds as putative signaling molecules. Knowledge about establishing a higher sensitive analysis method is of great importance for understanding the physiological functions of JA.At present, several methods for determining the exist and concentration of JA have been reported, such as mass spectrometry (MS), gas-liquid chromatography—mass spectrometry (GC-MS), enzyme linked immunosorbent assay, high performance liquid chromatography (HPLC), radioimmunoassay (RIA). However, the procedures of both MS and GC-MS method, which required long chromatographic separations at elevated temperatures, could cause thermal decomposition of the analytes. As a result, JA could not be discriminated perfectly from other plant components, which rendered the accurate quantification of small amounts of JA. Meanwhile, GC-MS and RIA required a complicated purification process before analysis. Quantification by RIA and ELISA based on specific antibodies has been used successfully, but they were suffered from cross-reactivity with structurally related compounds. HPLC, equipped with a fluorescence detector, has also been used successfully to quantify JA, but a complex purification procedure had to be used to separate compounds of interest from an enormous amount of other fluorescent compounds derived from the plant sample. Therefore, a more practical and accurate method for the detection of JA is needed. Here, we established a new method, high-performance capillary electrophoresis (HPCE), to determine the contents of the plant hormone JA.HPCE is an analytical method which separates compounds based on their size and electric charge. In recent years, major progress in the technology of HPCE has taken place. Because of a small volume (nL) injected, high sensitive detection, high plate efficiencies (up to a million) and short analysis time for biomacromolecule, HPCE shows strong potential to be widely used in biological science. The main challenge in developing HPCE method for phytochemical analyses is the presence of a complex mixture of various chemically different compounds in plant samples. Furthermore, the normal determination of JA by CE-UV detection reported in a few papers is still not high sensitive. Fluorescence detection, particularly the laser-induced fluorometry (LIF), also has become popular in CE, mainly because of its extremely high sensitivity.Despite their widespread applicability to biological and clinical analyses, CE coupled with LIF detection has not been applied to the analysis of JA at present. In this study, we used the fluorescent derivatization reagent,8-aminopyrene-l,3,6-trisu (APTS), which is commonly utilized in sugar derivatization analysis, as fluorescent labeled compound, and developed a novel method to determine JA extracted from fresh Arabidopsis thaliana leaf, based on capillary electrophoresis coupled to laser-induced fluorescence detection (CE-LIF). |
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