3.2. Au cellular localisation and distribution
Proton induced x-ray emission spectroscopy (-PIXE) was
performed at the Australian Nuclear Science and Technology
Organisation (ANSTO) using the 10MV tandem accelerator [44].
Samples were analysed using a 3MeV proton beam with a typical
spot size between 3 and 5m. At this spot size, beam currents
between 0.1 and 0.5 nA can be achieved, which is sufficient for -
PIXE analyses. A high-purity Ge detector was used with a 100mm2
active area, located 33mm from the sample. A 100m Mylar foil
was used to reduce low energy X-rays and thus pile-up in the
-PIXE spectrum. This set-up allowed the detection of the accumulated
trace metals such as Ni and Cu with as a high sensitivity
as for Au.
Listmode data files were collected using the MICRODAS data
acquisition system supplied by the University Of Melbourne. Data
analysis was performed using the GeoPIXE II software package [45].
Real time quantitative elemental maps were generated using a
dynamic analysis matrix transform method built into GeoPIXE II
[46]. These are overlap resolved and include background subtraction.
A typical micro-PIXE spectrum from BJ root tissues is shown in
Fig. 5. The peaks in the spectra clearly indicate the presence of Au,
as well as the essential trace elements, Calcium (Ca), Potassium (K),
Chlorine (Cl), Iron (Fe) and Copper (Cu).
For morphological and anatomical analysis, light micrographs
of stem and roots cross sections were analysed. The transverse section
of BJ root (Fig. 6 a) clearly shows the three tissue systems, i.e.
dermal, ground and vascular. A single epidermal layer, followed
by multi-layers of parenchymatous ground tissue and intercellular
spaces, characteristic of the root system are visible. In the centre
is the vascular bundle; strands of phloem are observed to alternate
with the xylem ridges. Four xylem ridges (a tetrarch) is clearly
indicated in the cross-section.
-PIXE analysis of BJ andMSstem and root show the presence of
Auin all three tissue systems.Aseries of typical in situ,Auelemental
maps for BJ root after 24 h exposure at 1000ppm and MS stem after
24 h of exposure at 40ppm are given in Fig. 6. Fig. 6 also indicates
the elemental maps achieved by varying the exposure time for MS
root at 80ppm and BJ root at 1000 ppm.
These maps show the highest accumulation of Au occurs in the
centre, indicating xylem loading, followed by the epidermis and
cortex cells. BJ root sections show that the highest metal localisation
is in the vascular bundles (VB) and xylem cells mainly because
they are the means of metal translocation. The presence of Au
was most notable in the vascular bundles (xylem cells), followed
by epidermal cells in all the root and stem sections mapped for
both BJ and MS. The results from -PIXE analysis of MS stems
after 24 h exposure at lower Au concentration of 40ppm (Fig. 6)
indicate the accumulation of gold on the outer part of the stem section
which constitutes epidermal cells and outer cortical region,
whereas at higher concentrations of 1000ppm the metal was
present everywhere in the stem section, but highest in the vascular
bundles.
Elemental maps for BJ root at 80ppm after 24 and 48 h exposure
clearly indicate the presence of Au in the outer epidermal cells
and outer cortical cells (Fig. 6). The maps also suggests the presence
of higher concentrations of Au in the epidermal and cortical
cells with longer exposure time of 48 h as compared to 24 h. Similar
results were observed at higher concentrations (1000 ppm) in MS
root, although the metal was present everywhere in the section,
higher concentrations were observed after longer exposure time.
The most striking difference between BJ and MS root at 1000ppm
after 24 h is the high concentration of metal in the central stele in BJ
root, where as in MS root the metal is dispersed equally throughout
the section. However, at 48 h the metal seem to be concentrated in
the central stele (Fig. 6).