No abstract

CE-ESI-MS and CE-ICP-MS were implemented for studying three phytosiderophores (mugineic acid, epi-mugineic acid and deoxymugineic acid) and their metal complexes. Free ligands and ferric complexes were analyzed using the first methodology, while six free metals (Co(II), Cu(II), Fe(III), Mn(II), Ni(II) and Zn(II)) together with the corresponding complexes were investigated by the latter technique. CE separation was realized at a voltage of +25 kV employing a BGE containing 20 mM ammonium bicarbonate at pH 7.2. Both techniques revealed limits of detection in the high nM to low microM range. Standard additions to hydroponic samples of H. distichon, cv. Bodega (spring barley) showed regression coefficients for the metal-ligand complexes ranging from 0.984 to 0.999. Additionally, results of a competitivity study allowed the determination of relative metal-phytosiderophore complex stability constants of deoxymugineic/mugineic acid.

Analysis of underivatized amino acids is challenging regarding the separation as well as the detection of these small polar analytes. Hydrophilic interaction LC using a 2.1x150 mm ZIC (ZIC, zwitterionic)-hydrophilic interaction LC from SeQuant as stationary phase with 1% v/v formic acid in water and ACN as eluents was combined with MS/MS in multiple reaction monitoring mode for the separation and the detection of 16 underivatized amino acids. Regression coefficients of eight or seven point calibrations varied from 0.9454 to 0.9993. Absolute LODs and LOQ (on column) were in the fmol range (0.1-12 and 0.4-41 fmol, respectively). A fast screening method of 19 min total runtime has been developed offering applicability to samples from rhizosphere studies--characterized by low analyte concentrations and complex matrices. A successful application to the analysis of tyrosine in samples from soil adsorption experiments is presented as well as an evaluated enrichment procedure for amino acids derived from plant culture in nutrient solution.

Phytosiderophores (PS) form stable complexes with various transition metals. These ligands are exuded by the roots of graminacous plants as a mechanism for mobilizing and acquiring soil iron. To investigate iron mobilization and transport, a novel LC method in combination with ESI-MS/MS for the determination of three Fe(III)-complexes with mugineic acid (MA), 2'-epi-MA and 2'-deoxymugineic acid (DMA) has been developed. Liquid chromatographic separation was realized using a silica-based mixed-mode reversed phase/weak-anion exchange type stationary phase and a 50 mM ammonium acetate buffer, pH 6.5. Baseline separation of the two complex diastereomers Fe(III)-MA and Fe(III)-epi-MA could be achieved. ESI-MS/MS detection allowed for simultaneous quantification of the complexes and the free ligands. Limits of detection were determined to be 0.001 and 0.05 μM for DMA and Fe(III)-DMA, respectively. The analytical figures of merit of the novel method were evaluated and compared with a CE-ESI-MS method that we had published earlier. The LC-ESI-MS/MS method has been successfully applied to real samples derived from preliminary extraction experiments.

Analysis of underivatized amino acids is challenging regarding both separation and detection of this small, polar, and largely UV-inactive compounds. Additives for reversed phase chromatography such as ion-pairing reagents can hamper mass spectrometric detection. Zwitterionic hydrophilic interaction chromatography using MS compatible eluents together with tandem mass spectrometry in multiple reaction monitoring mode for selective detection is an attractive approach to overcome the above mentioned issues.