High resolution secondary ion mass spectrometry analysis of trace elements in cereal grain and roots

This thesis presents information on the subcellular localisation of two important trace elements, selenium and arsenic, in wheat, rice and rice roots for what is believed to be the first time. The general aim of this work was to illustrate the potential of using physical science techniques to solve biological problems. High resolution secondary ion mass spectrometry was undertaken using the CAMECA NanoSIMS50 with a sensitivity down to ppm concentrations and a lateral resolution of less than 100 nm. Selenium in wheat grain was found to be distributed across both the bran layer and the endosperm region with Se-rich hotspots found in the aleurone cells and a higher intensity of Se in the subaleurone region. Arsenic in rice grain was found in two key regions. In grains with high As and high dimethylarsinic acid (DMA) content, As was predominantly localised to the subaleurone region yet in lower concentration, hydroponically grown As(III)-treated grains the As was only localised to the aleurone layer near the ovular vascular trace (OVT). A combined NanoSIMS and S-XRF experiment revealed As hotspots near the OVT. A combination of high pressure freezing, high resolution secondary ion mass spectrometry and TEM was used to localise As in the roots of rice plants revealing a contrasting subcellular distribution of As and Si in the roots even though arsenite and silicic acid are transported across the plasma membranes by the same transporters. Fe plaque forms only on the root epidermis and was shown to be a strong sink for As. Colocalisation of S with As in the vacuoles of the endodermis, pericycle and xylem parenchyma supports the notion that As is stored as arsenite-phytochelatin complexes in the roots while Si is localised in the endodermis cell walls and is not strongly affected by the Lsi2 mutation. Some figures have been removed from this version of the thesis for copyright reasons