Diane Eichert

New insights into globoids of protein storage vacuoles in wheat aleurone using synchrotron soft X-ray microscopy

Mature developed seeds are physiologically and biochemically committed to store nutrients, principally as starch, protein, oils, and minerals. The composition and distribution of elements inside the aleurone cell layer reflect their biogenesis, structural characteristics, and physiological functions. It is therefore of primary importance to understand the mechanisms underlying metal ion accumulation, distribution, storage, and bioavailability in aleurone subcellular organelles for seed fortification purposes. Synchrotron radiation soft X-ray full-field imaging mode (FFIM) and low-energy X-ray fluorescence (LEXRF) spectromicroscopy were applied to characterize major structural features and the subcellular distribution of physiologically important elements (Zn, Fe, Na, Mg, Al, Si, and P). These direct imaging methods reveal the accumulation patterns between the apoplast and symplast, and highlight the importance of globoids with phytic acid mineral salts and walls as preferential storage structures. C, N, and O chemical topographies are directly linked to the structural backbone of plant substructures. Zn, Fe, Na, Mg, Al, and P were linked to globoid structures within protein storage vacuoles with variable levels of co-localization. Si distribution was atypical, being contained in the aleurone apoplast and symplast, supporting a physiological role for Si in addition to its structural function. These results reveal that the immobilization of metals within the observed endomembrane structures presents a structural and functional barrier and affects bioavailability. The combination of high spatial and chemical X-ray microscopy techniques highlights how in situ analysis can yield new insights into the complexity of the wheat aleurone layer, whose precise biochemical composition, morphology, and structural characteristics are still not unequivocally resolved.

Low-energy X-ray fluorescence microscopy opening new opportunities for bio-related research

Biological systems are unique matter with very complex morphology and highly heterogeneous chemical composition dominated by light elements. Discriminating qualitatively at the sub-micrometer level the lateral distribution of constituent elements, and correlating it to the sub-cellular biological structure, continues to be a challenge. The low-energy X-ray fluorescence microspectroscopy, recently implemented in TwinMic scanning transmission mode, has opened up new opportunities for mapping the distribution of the light elements, complemented by morphology information provided by simultaneous acquisition of absorption and phase contrast images. The important new information that can be obtained in bio-related research domains is demonstrated by two pilot experiments with specimens of interest for marine biology and food science. They demonstrate the potential to yield important insights into the structural and compositional enrichment, distribution and correlation of essential trace elements in the lorica of Tintinnopsis radix, and the lateral distribution of trace nutrients in the seeds of wheat Triticum aestivum.

Beeswax as dental filling on a Neolithic human tooth

Evidence of prehistoric dentistry has been limited to a few cases, the most ancient dating back to the Neolithic. Here we report a 6500-year-old human mandible from Slovenia whose left canine crown bears the traces of a filling with beeswax. The use of different analytical techniques, including synchrotron radiation computed micro-tomography (micro-CT), Accelerator Mass Spectrometry (AMS) radiocarbon dating, Infrared (IR) Spectroscopy and Scanning Electron Microscopy (SEM), has shown that the exposed area of dentine resulting from occlusal wear and the upper part of a vertical crack affecting enamel and dentin tissues were filled with beeswax shortly before or after the individual’s death. If the filling was done when the person was still alive, the intervention was likely aimed to relieve tooth sensitivity derived from either exposed dentine and/or the pain resulting from chewing on a cracked tooth: this would provide the earliest known direct evidence of therapeutic-palliative dental filling.

Relevance for food sciences of quantitative spatially resolved element profile investigations in wheat (Triticum aestivum) grain

Bulk element concentrations of whole grain and element spatial distributions at the tissue level were investigated in wheat (Triticum aestivum) grain grown in Zn-enriched soil. Inductively coupled plasma mass spectrometry and inductively coupled plasma optical emission spectrometry were used for bulk analysis, whereas micro-proton-induced X-ray emission was used to resolve the two-dimensional localization of the elements. Soil Zn application did not significantly affect the grain yield, but did significantly increase the grain Ca, Fe and Zn concentrations, and decrease the grain Na, P and Mo concentrations; bulk Mg, S, K, Mn, Cu, Cd and Pb concentrations remained unchanged. These changes observed in bulk element concentrations are the reflection of tissue-specific variations within the grain, revealing that Zn application to soil can lead to considerable alterations in the element distributions within the grain, which might ultimately influence the quality of the milling fractions. Spatially resolved investigations into the partitioning of the element concentrations identified the tissues with the highest element concentrations, which is of utmost importance for accurate prediction of element losses during the grain milling and polishing processes.

Optimisation of a compact optical system for the beamtransport at the x-ray fluorescence beamline at Elettra for experiments with small spots

This paper reports a simplified and compact mechanical scheme designed for providing alternatively high flux or high spectral resolution from a monochromator for the 2 - 14 keV X-ray range in a stationary spot at a sample. The example case treats a bending magnet source at the Elettra storage ring. The properties of the continuously tunable monochromatic beam are adapted for a variety of experimental techniques, which include also the need to operate the sample in the total reflection regime, i.e. at angles of grazing incidence smaller than the critical angle of the sample material. The positional stability of the monochromatised beam during tuning was the major concern in the design of the monochromator.

Biochemical characterization of cell types within leaves of metal-hyperaccumulating Noccaea praecox (Brassicaceae)

Background and aimsDistinct metal distribution patterns within leaves of metal hyperaccumulating plants are repeatedly observed however, the presumable role of key structural biochemical molecules in determining and regulating their allocation remains largely unknown. We aimed to characterise in a spatially resolved manner the distribution of the main biochemical components in leaves of field-collected Cd/Zn-hyperaccumulating Noccaea praecox in order to relate them to metal distribution patterns at tissue level.MethodsThe biomolecular composition of the leaves was spatially analysed using synchrotron radiation Fourier Transform Infrared (FTIR) and the distribution of Zn with synchrotron radiation Low-Energy X-Ray Fluorescence (LEXRF) microspectroscopy was determined on the same tissues of interest (epidermis, sub-epidermis, mesophyll).ResultsIn epidermal cells high proportion of free-carboxyl, nitro and phosphate groups standing for pectin, nitroaromatics, phytic and other organic acids were found. Adjacent mesophyll cells had higher proportions of proteins, carbohydrates and cellulosic compounds.ConclusionsPectin compounds were indicated as important components of Zn enriched epidermal cell walls. In addition, intense lignification of epidermal cell walls might limit leakage of the trapped metals back to the metabolically active and thus more sensitive mesophyll. Distribution of metal-binding compounds in particular cell types/tissues may therefore predispose metal distribution patterns and tolerance in leaves.

Fast pixelated quantum-well-based sensor for multi-wavelength photon detection

In order to cover a wide range of experimental applications, the opportunity to use InGaAs/InAlAs quantum well (QW) devices as fast pixelated photon detectors has been investigated. QW structures are planar objects in which electrons are confined in one dimension. Devices with several combinations of barrier and well materials can be fabricated by using compound semiconductors; in the case of InGaAs/InAlAs QWs this allows to tune the energy band gap down to 0.6 eV. Thanks to their direct, low-energy gap such devices operated at room temperature may be used as detectors for photon energies ranging from visible to hard X-ray. Internal charge amplification mechanism can be applied for very low signal levels, while the high carrier mobility allows the design of very fast photon detectors with sub-nanosecond response times. QW devices grown with molecular beam epitaxy have been pixelated by using standard photolithographic techniques. In order to fit commercially available readout chips, a pixelated sensor with pixel size of 172 × 172 μm2 is currently under development. A small-scale version of the pixelated QW sensor has been preliminarily tested with synchrotron radiation, conventional X-rays and UV laser light. The reported results indicate that these devices show fair charge sharing in the clearances between the pixels and feature very short response times to 100-fs-wide laser pulses.

On symmetric X-ray beam splitting with high efficiency by use of reflection gratings with rectangular profile in the extreme off-plane configuration

In order to be reflected or diffracted off a surface structure soft X-rays and hard X-rays need to impinge at grazing angles of incidence onto the surface. In case of a reflection grating of highly symmetric structure with rectangular groove profile these grooves can be oriented parallel to the beam trajectory. In such a symmetric situation the distribution of the diffracted intensity with respect to the plane of incidence is then expected to be symmetric. This is indeed observed with symmetrically oriented diffraction peaks. It can be predicted that for appropriate structure parameters the intensity can be contained mostly in two symmetrically oriented diffraction peaks. This will also be the case for hard X-rays. The diffraction efficiency will be particularly high, when the angle of grazing incidence is chosen in the total reflection regime below the critical angle of the grating coating. These predictions were experimentally verified in this work for hard X-rays with photon energies between 4 keV and 12.4 keV. In the experiment of the order of 30% of the incident intensity was diffracted into the two first orders. This is to be compared to reflectivities of the order of 50% measured at the same coating in an unruled area of the substrate. Consequently the relative structural diffraction efficiency for each first order was about 30%, while ideally it could have been 40%. The presented grating structure will thus be a rather efficient amplitude beam splitter for hard X-rays, e.g. in the coherent beam from a free electron laser. In addition such object could then be used as the first component in Michelson interferometers for the beam characterisation or for introducing a time delay between two coherent beams.

Position sensitive photon detectors using epitaxial InGaAs/InAlAs quantum wells

This work deals with the investigation of novel position-sensitive devices based on InGaAs/InAlAs quantum wells, which could be applied to several applications of either synchrotron or conventional light sources. Such devices may be used as fast and efficient detectors due to the direct, low-energy band gap and high electron mobility at room temperature. Metamorphic In0.75Ga0.25As/In0.75Al0.25As quantum wells containing a two-dimensional electron gas were grown by molecular beam epitaxy. Two devices with size of 5 × 5 mm2 were prepared by using optical lithography. In the first, the active layers were segmented into four electrically insulated quadrants. Indium ohmic contacts were realized on the corner of each quadrant (for readout) and on the back surface (for bias). In the second, the quantum well was left unsegmented and covered by 400 nm of Al providing a single bias electrode, while four readout electrodes were fabricated on the back side by depositing and segmenting a Ni/Ge/Au layer. Photo-generated carriers can be collected at the readout electrodes by biasing from either the QW side or the back side of the devices during beam exposure. Individual currents obtained from each electrode allow monitoring of both the position and the intensity of the impinging beam for photon energies ranging from visible to hard X-ray. Such detector prototypes were tested with synchrotron radiation. Moreover, the position of the beam can be estimated with a precision of 800 nm in the segmented QW. A lower precision of 10 μm was recorded in the unsegmented QW due to the charge diffusion through the 500-μm-thick wafer, with however a lower electronic noise due to the better uniformity of the contacts.

Surface impurities on giant gypsum crystals from “la Cueva de las Espadas” (Cave of Swords), Naica, Mexico

The Cave of Swords was discovered in 1910 at Naica, Chihuahua, México. During the last century, human presence has changed the microclimate conditions inside this cave, raising the question of whether anthropogenic action resulted in the deterioration of its gypsum single crystals and in the deposition of impurities on their surfaces. The present work provides a detailed characterization of representative samples of this cave and suggests an answer to the origin of the impurities on the surface of these nature-made large crystals. Laboratory and synchrotron characterization techniques are applied. For the first time, the samples single-crystal nature and the fragmentation effect of impurities are characterized. Synchrotron light diffraction measurements performed on a collection of characteristic samples consistently reveal sharp textures, with crystal reciprocal vectors [0,1,0] preferably perpendicular to the samples principal cleavage planes and orientation widths averaging 8°. X-ray diffraction identifies galena, sphalerite, hematite, goethite and cuprite on the crystals’ surface. Diffraction results indicate no correlation between the number of phases of impurities present and crystallinity. Micro X-ray fluorescence clarifies the elemental spatial distribution. The correlations between the elemental distributions confirm the phase identification obtained by diffraction. For Mn and Pb, the correlations point to the presence of amorphous oxides. Minor phases’ characterization suggests they have been deposited on iron oxy-hydroxide substrates. All the identified phases correspond to minerals that were abundantly present in Naica ore deposit before any anthropogenic activity. The impurities on the surface of gypsum crystals at the Cave of Swords were not produced by human presence.