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The development of new qualitative and quantitative metal speciation approaches for low molecular weight metals complexes in plants

Abstract : The main goal of this thesis was to develop and validate protocols of mass spectrometry methods, which could be applied for determination and quantitation of low molecular weight metal compounds in plant liquids. Unfortunately, the amount of plant liquid (eg. xylem), which usually can be sampled after several weeks of fastidious plant cultivation, is in low microliters range (or lower), which make the method development difficult task. Thus, as a compromise, coconut (Cocos nucifera) water was chosen as a model for method development studies of plant endosperm metallome, as the amount of sample available for experiments is much higher. Based on the information available in literature the most commonly identified iron complexes were Fe-citrate ones and mixed complexes of iron with citrate and malate. The detailed study of formation of different complexes was carried out to evaluate their stability and equilibrium between them. Additionally different chromatographic columns were tested and elution condition were optimized to be used later on to the real samples. ESI MS condition were adjusted (positive or negative mode ionization) depending on chromatographic conditions used. The principal part of the thesis concerns the development of methods for characterization of the model sample. Influence of different sample treatment steps (filtration, centrifugation) and storage condition was studied. Also, sample preparation before HILIC separation was evaluated, and the separation conditions optimized on model complexes was tested. The quest of a generic analytical strategy for the standardless quantification of different low-molecular weight metal complexes by molecular mass spectrometry is pursued using the Isotope Dilution Analysis (IDA) with the addition of an isotope spike 58Fe in ionic form and HILIC-ICP-MS. The protocol was carefully studied in order to keep the original metal complexes in non-degraded way and achieve the formation of metal complexes with the excess of ligands present in the sample. Finally, the developed procedures were applied for characterisation and quantification of iron compounds in xylem samples of several plants. The plants chosen (Pisum sativum, Paspalum urvilleli, and Setaria parviflora) were hypertolerant or overaccumulating ones.
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Ghaya Alchoubassi. The development of new qualitative and quantitative metal speciation approaches for low molecular weight metals complexes in plants. Analytical chemistry. Université de Pau et des Pays de l'Adour, 2019. English. ⟨NNT : 2019PAUU3031⟩. ⟨tel-02519778v1⟩

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