Visualising the ionome in resistant and susceptible plant-pathogen interactions.

At the morphological and anatomical levels, the ionome, or the elemental composition of an organism, is an understudied area of plant biology. In particular, the ionomic responses of plant-pathogen interactions are scarcely described, and there are no studies on immune reactions. In this study we explored two X-ray fluorescence (XRF)-based ionome visualisation methods (benchtop- and synchrotron-based micro-XRF [µXRF]), as well as the quantitative inductively coupled plasma optical emission spectroscopy (ICP-OES) method, to investigate the changes that occur in the ionome of compatible and incompatible plant-pathogen interactions. We utilised the agronomically important and comprehensively studied interaction between potato (Solanum tuberosum) and the late blight oomycete pathogen Phytophthora infestans as an example. We used one late blight-susceptible potato cultivar and two resistant transgenic plant lines (only differing from the susceptible cultivar in one or three resistance genes) both in control and P. infestans-inoculated conditions. In the lesions from the compatible interaction, we observed rearrangements of several elements, including a decrease of the mobile macronutrient potassium (K) and an increase in iron (Fe) and manganese (Mn), compared with the tissue outside the lesion. Interestingly, we observed distinctly different distribution patterns of accumulation at the site of inoculation in the resistant lines for calcium (Ca), magnesium (Mg), Mn and silicon (Si) compared to the susceptible cultivar. The results reveal different ionomes in diseased plants compared to resistant plants. Our results demonstrate a technical advance and pave the way for deeper studies of the plant-pathogen ionome in the future.

Identifying anthropogenic features at Seoke (Botswana) using pXRF: Expanding the record of southern African Stone Walled Sites.

Numerous and extensive 'Stone Walled Sites' have been identified in southern African Iron Age landscapes. Appearing from around 1200 CE, and showing considerable variability in size and form, these settlements are named after the dry-stone wall structures that characterize them. Stone Walled Sites were occupied by various Bantu-speaking agropastoral communities. In this paper we test the use of pXRF (portable X-ray fluorescence analysis) to generate a 'supplementary' archaeological record where evident stratigraphy is lacking, survey conditions may be uneven, and excavations limited, due to the overall site size. We propose herein the application of portable X-ray fluorescence analysis (pXRF) coupled with multivariate exploratory analysis and geostatistical modelling at Seoke, a southern African SWS of historical age (18th century CE). The aim of the paper is twofold: to explore the potential of the application of a low cost, quick, and minimally invasive technique to detect chemical markers in anthropogenic sediments from a Stone Walled Site, and to propose a way to analyse the results in order to improve our understanding of the use of space at non-generalized scales in such sites.

X-ray fluorescence analysis of metal distributions in cryogenic biological samples using large-acceptance-angle SDD detection and continuous scanning at the Hard X-ray Micro/Nano-Probe beamline P06 at PETRA III.

A new Rococo 2 X-ray fluorescence detector was implemented into the cryogenic sample environment at the Hard X-ray Micro/Nano-Probe beamline P06 at PETRA III, DESY, Hamburg, Germany. A four sensor-field cloverleaf design is optimized for the investigation of planar samples and operates in a backscattering geometry resulting in a large solid angle of up to 1.1 steradian. The detector, coupled with the Xspress 3 pulse processor, enables measurements at high count rates of up to 106 counts per second per sensor. The measured energy resolution of ∼129 eV (Mn Kα at 10000 counts s-1) is only minimally impaired at the highest count rates. The resulting high detection sensitivity allows for an accurate determination of trace element distributions such as in thin frozen hydrated biological specimens. First proof-of-principle measurements using continuous-movement 2D scans of frozen hydrated HeLa cells as a model system are reported to demonstrate the potential of the new detection system.

Large-scale micron-order 3D surface correlative chemical imaging of ancient Roman concrete.

There has been significant progress in recent years aimed at the development of new analytical techniques for investigating structure-function relationships in hierarchically ordered materials. Inspired by these technological advances and the potential for applying these approaches to the study of construction materials from antiquity, we present a new set of high throughput characterization tools for investigating ancient Roman concrete, which like many ancient construction materials, exhibits compositional heterogeneity and structural complexity across multiple length scales. The detailed characterization of ancient Roman concrete at each of these scales is important for understanding its mechanics, resilience, degradation pathways, and for making informed decisions regarding its preservation. In this multi-scale characterization investigation of ancient Roman concrete samples collected from the ancient city of Privernum (Priverno, Italy), cm-scale maps with micron-scale features were collected using multi-detector energy dispersive spectroscopy (EDS) and confocal Raman microscopy on both polished cross-sections and topographically complex fracture surfaces to extract both bulk and surface information. Raman spectroscopy was used for chemical profiling and phase characterization, and data collected using EDS was used to construct ternary diagrams to supplement our understanding of the different phases. We also present a methodology for correlating data collected using different techniques on the same sample at different orientations, which shows remarkable potential in using complementary characterization approaches in the study of heterogeneous materials with complex surface topographies.

Biologically Synthesized Silver Nanoparticles for Enhancing Tetracycline Activity Against Staphylococcus aureus and Klebsiella pneumoniae

Abstract Phytochemical content of plant extracts can be used effectively to reduce the metal ions to nanoparticles in one-step green synthesis process. In this study, six plant extracts were used for the synthesis of silver nanoparticles (AgNPs). Biologically synthesized AgNPs was characterized using UV-Vis Spectrophotometer, Field Emission Scanning Electron Microscope (FE-SEM), X-ray diffraction (XRD), Energy Dispersive X-ray spectroscopy (EDX) and Fourier Transform Infrared (FTIR) spectroscopy. The individual and combined effects of AgNPs and tetracycline against S. aureus and K. pneumoniae were assessed. Ginger, onion and sidr extracts supported AgNPs formation while arak, garlic and mint extracts failed to convert the silver ions to AgNPs. The present findings revealed significant differences between the tested plant extracts in supporting AgNPs synthesis. AgNPs synthesized by ginger showed the highest individual and combined activity against tested strains followed by AgNPs prepared by sidr then that synthesized by onion. AgNPs significantly enhanced tetracycline activity (p≤0.05) against S. aureus and K. pneumoniae. The results of this study demonstrated that the combination of tetracycline and biologically synthesized AgNPs presented a useful therapeutically method for the treatment of bacterial infection and counterattacking bacterial resistance.

Portable X-ray fluorescence system to measure Th and U concentrations.

This study reports the results obtained in the analysis of waste material samples generated by the industries of phosphate fertilizers, in particular, the use of specific filters in a portable X-ray fluorescence system, a simple equipment allowing the characterization, identification and quantification of low concentrations of Th and U (ppm). The industrial byproduct is classified as a Technologically-Enhanced, Naturally-Occurring Radioactive Material - TENORM, and therefore requires monitoring for its radio-toxic activity due to the presence of radioactive thorium and uranium families. From the results obtained, it is concluded that this technique is able to determine the contents of these elements to concentrations of tens of ppm in measurements of about 300 s, and a small sample amount (∼0.1 g).

Assessing Statistically Significant Heavy-Metal Concentrations in Abandoned Mine Areas via Hot Spot Analysis of Portable XRF Data.

To develop appropriate measures to prevent soil contamination in abandoned mining areas, an understanding of the spatial variation of the potentially toxic trace elements (PTEs) in the soil is necessary. For the purpose of effective soil sampling, this study uses hot spot analysis, which calculates a z-score based on the Getis-Ord Gi* statistic to identify a statistically significant hot spot sample. To constitute a statistically significant hot spot, a feature with a high value should also be surrounded by other features with high values. Using relatively cost- and time-effective portable X-ray fluorescence (PXRF) analysis, sufficient input data are acquired from the Busan abandoned mine and used for hot spot analysis. To calibrate the PXRF data, which have a relatively low accuracy, the PXRF analysis data are transformed using the inductively coupled plasma atomic emission spectrometry (ICP-AES) data. The transformed PXRF data of the Busan abandoned mine are classified into four groups according to their normalized content and z-scores: high content with a high z-score (HH), high content with a low z-score (HL), low content with a high z-score (LH), and low content with a low z-score (LL). The HL and LH cases may be due to measurement errors. Additional or complementary surveys are required for the areas surrounding these suspect samples or for significant hot spot areas. The soil sampling is conducted according to a four-phase procedure in which the hot spot analysis and proposed group classification method are employed to support the development of a sampling plan for the following phase. Overall, 30, 50, 80, and 100 samples are investigated and analyzed in phases 1-4, respectively. The method implemented in this case study may be utilized in the field for the assessment of statistically significant soil contamination and the identification of areas for which an additional survey is required.

Assessing arsenic and selenium in a single nail clipping using portable X-ray fluorescence.

The feasibility of measuring arsenic and selenium contents in a single nail clipping was investigated using a small-focus portable X-ray fluorescence (XRF) instrument with monochromatic excitation beams. Nail clipping phantoms supplemented with arsenic and selenium to produce materials with 0, 5, 10, 15, and 20µg/g were used for calibration purposes. In total, 10 different clippings were analyzed at two different measurement positions. Energy spectra were fit with detection peaks for arsenic Kα, selenium Kα, arsenic Kß, selenium Kß, and bromine Kα characteristic X-rays. Data analysis was performed under two distinct conditions of fitting constraint. Calibration lines were established from the amplitude of each of the arsenic and selenium peaks as a function of the elemental contents in the clippings. The slopes of the four calibration lines were consistent between the two conditions of analysis. The calculated minimum detection limit (MDL) of the method, when considering the Kα peak only, ranged from 0.210±0.002µg/g selenium under one condition of analysis to 0.777±0.009µg/g selenium under another. Compared with previous portable XRF nail clipping studies, MDLs were substantially improved for both arsenic and selenium. The new measurement technique had the additional benefits of being short in duration (~3min) and requiring only a single nail clipping. The mass of the individual clipping used did not appear to play a major role in signal strength, but positioning of the clipping is important.

Detection of terrestrial radionuclides with X-ray fluorescence analysis.

This paper provides an overview of analytical methods frequently used to identify terrestrial radionuclides in samples. While radioactivity is normally measured through the ionising radiation produced during the spontaneous decay of unstable atoms, selected radionuclides or their chemical elements can be quantified with instrumental techniques based on stimulated emission or counting of atoms. The advantages and disadvantages of these analytical methods are discussed. Particular attention is paid to X-ray fluorescence analysis of materials containing uranium and thorium. It is also possible to determine the area distributions of these chemical elements in samples with the use of scanning X-ray fluorescence systems.

Tracking sulfur and phosphorus within single starch granules using synchrotron X-ray microfluorescence mapping.

BACKGROUND: Native starch accumulates as granules containing two glucose polymers: amylose and amylopectin. Phosphate (0.2-0.5%) and proteins (0.1-0.7%) are also present in some starches. Phosphate groups play a major role in starch metabolism while granule-bound starch synthase 1 (GBSS1) which represents up to 95% of the proteins bound to the granule is responsible for amylose biosynthesis. METHODS: Synchrotron micro-X-ray fluorescence (µXRF) was used for the first time for high-resolution mapping of GBSS1 and phosphate groups based on the XRF signal of sulfur (S) and phosphorus (P), respectively. Wild-type starches were studied as well as their related mutants lacking GBSS1 or starch-phosphorylating enzyme. RESULTS: Wild-type potato and maize starch exhibited high level of phosphorylation and high content of sulfur respectively when compared to mutant potato starch lacking glucan water dikinase (GWD) and mutant maize starch lacking GBSS1. Phosphate groups are mostly present at the periphery of wild-type potato starch granules, and spread all over the granule in the amylose-free mutant. P and S XRF were also measured within single small starch granules from Arabidopsis or Chlamydomonas not exceeding 3-5µm in diameter. CONCLUSIONS: Imaging GBSS1 (by S mapping) in potato starch sections showed that the antisense technique suppresses the expression of GBSS1 during biosynthesis. P mapping confirmed that amylose is mostly present in the center of the granule, which had been suggested before. GENERAL SIGNIFICANCE: µXRF is a potentially powerful technique to analyze the minor constituents of starch and understand starch structure/properties or biosynthesis by the use of selected genetic backgrounds.

Rapid determination of silver in nanobased liquid dietary supplements using a portable X-ray fluorescence analyzer.

This paper reports a rapid and straightforward method for the quantitation of total Ag content in nanobased commercially available liquid dietary supplements using a portable X-ray fluorescence (pXRF) analyzer. Figures of merits were evaluated by analyzing a series of AgNO3 standards. This method was shown to have a detection limit of 3 ppm, a quantitation limit of 10 ppm, and a broad linear range from the detection limit to 10000 ppm (1%). Accurate detection and quantitation of Ag content in well-characterized Ag nanoparticle samples and in nanobased liquid dietary supplements were achieved with good correlation (i.e., percentage difference average values under 15%) between the total Ag concentration obtained by the pXRF analyzer and by inductively coupled plasma mass spectrometry (ICP-MS). Furthermore, accurate quantitation of Ag in the presence of high concentrations of potential spectral interferences was also demonstrated.

Evaluation of a prototype point-of-care instrument based on monochromatic x-ray fluorescence spectrometry: potential for monitoring trace element status of subjects with neurodegenerative disease.

Assessment of trace elements such as Cu, Zn, and Se in patients with neurodegenerative disease, such as Alzheimer's (AD) and Parkinson's disease (PD), may be useful in etiologic studies and in assessing the risk of developing these conditions. A prototype point-of-care (POC) instrument based on monochromatic x-ray fluorescence (M-XRF) was assembled and evaluated for the determination of Cu, Zn, and Se in whole blood, plasma, and urine. The prototype instrument was validated using certified reference materials for Cu and Zn in serum/plasma, and the reported bias and relative imprecision were <10%. The M-XRF prototype performance was further assessed using human specimens collected from AD and PD subjects, and was found to be satisfactory (<20% bias) for monitoring Cu and Zn levels in plasma and whole blood. However, the prototype M-XRF sensitivity was not sufficient for quantifying Cu, Zn, or Se in urine. Nonetheless, while validating the prototype instrument, body fluids (whole blood, plasma, and urine) were collected from 19 AD patients, 23 PD patients, and 24 controls specifically for trace element analysis using well-validated methods based on inductively coupled plasma mass spectrometry (ICP-MS). This limited biomonitoring study provided robust data for up to 16 elements including Sb, As, Ba, Cd, Cs, Co, Cr, Cu, Hg, Pb, Mo, Se, Tl, Sn, Zn, and U in plasma, whole blood, and urine. The results did not indicate any significant differences in most trace elements studied between AD or PD patients compared to controls, although the sample size is limited. A statistically significant increase in plasma Se was identified for PD patients relative to AD patients, but this could be due to age differences.

Matrix effects in the energy dispersive X-ray analysis of CaO-Al(2)O(3)-MgO inclusions in steel.

Energy dispersive X-ray microanalysis of micron-sized inclusions in steel is of considerable industrial importance. Measured spectra and Monte Carlo simulations show a significant effect of the steel matrix on analysis of CaO-Al(2)O(3)-MgO inclusions: the steel matrix filters the softer (Al and Mg) characteristic X-rays, increasing the relative height of the Ca peak. Bulk matrix correction methods would not result in correct inclusion compositions, but operating at a lower acceleration voltage shifts the effect to smaller inclusion sizes.

Use of field-portable XRF analyzers for rapid screening of toxic elements in FDA-regulated products.

Analytical instrumentation continues its amazing evolution, especially in regard to generating ever more sensitive, faster, and reliable measurements. Perhaps the most difficult challenges are making these instruments small enough to use in the field, equipping them with well-designed software that facilitates and simplifies their use by nonexperts while preserving enough of their analytical capabilities to render them useful for a wide variety of applications. Perhaps the most impressive and underappreciated example of instruments that meet these criteria are field-portable X-ray fluorescence (XRF) analyzers. In the past, these analyzers have been routinely used for environmental applications (lead in paint and soil, metal particulates in air samples collected onto filters), geology studies (ore and soil analysis, precious metal identification), and recycling industries (alloy identification). However, their use in the analysis of toxic elements in food, food ingredients, dietary supplements, and medicinal and herbal products, especially within the FDA and regulatory environments, has been surprisingly limited to date. Although XRF will not replace atomic spectrometry techniques such as ICP-MS for sub-parts per million level analyses, it offers a number of significant advantages including minimal sample preparation, high sample throughputs, rapid and definitive identification of many toxic elements, and accurate quantitative results. As should be obvious from many recent news reports on elevated levels of toxic elements in children's lunchboxes, toys, and supplements, field-portable XRF analyzers can fill a very important niche and are becoming increasingly popular for a wide variety of elemental analysis applications. This perspective begins with a brief review of the theory of XRF to highlight the underlying principle, instrumentation, and spectra. It includes a discussion of various analytical figures of merit of XRF to illustrate its strengths and limitations compared to existing methods such as ICP-MS. It concludes with a discussion of a number of different FDA applications and case studies in which XRF has been used to screen, identify, and in some cases quantify toxic elements in various products. This work clearly demonstrates that XRF analyzers are an exceedingly valuable tool for routine and nonroutine elemental analysis investigations, both in the laboratory and in the field. In the future, it is hoped that both field-portable and laboratory-grade XRF analyzers will see more widespread use for investigational and forensic-type applications of food and other regulated consumer products.

Topographic trace-elemental analysis in the brain of Wistar rats by X-ray microfluorescence with synchrotron radiation.

Knowledge about the spatial distribution and the local concentration of trace elements in tissues is of great importance, since trace elements are involved in many biological functions of living organisms. However, there are few methods available to measure the spatial (two (three)-dimensional) elemental distribution in animal brain. X-ray microfluorescence with synchrotron radiation is a multielemental mapping technique, which was used in this work to determine the topographic of iron, zinc and copper in coronal sections of female Wistar rats of different ages. Young (14 days old) and middle-aged (20 months old) rats (n = 8) were analyzed. The measurements were carried out at the XRF beam line at the Synchrotron Light National Laboratory (Campinas, Brazil). Two-dimensional scanning was performed in order to study the tendency of elemental concentration variation. The acquisition time for each pixel was 10 s/step and the step size was 300 microm/step in both directions. It was observed that the iron distribution was more conspicuous in the cortical area, thalamus and bellow the thalamus. On the other hand, the zinc distribution was more pronounced in the hippocampus. The iron, copper and zinc levels increased with advancing age. Therefore, this study reinforces the idea that these elements are involved in the chemical mechanisms of the brain that induce some neurological diseases, since they are also present in high levels in specific areas of the brain, such as the hippocampus and the substantia nigra of patients with these disorders.

Differential phase contrast with a segmented detector in a scanning X-ray microprobe.

Scanning X-ray microprobes are unique tools for the nanoscale investigation of specimens from the life, environmental, materials and other fields of sciences. Typically they utilize absorption and fluorescence as contrast mechanisms. Phase contrast is a complementary technique that can provide strong contrast with reduced radiation dose for weakly absorbing structures in the multi-keV range. In this paper the development of a segmented charge-integrating silicon detector which provides simultaneous absorption and differential phase contrast is reported. The detector can be used together with a fluorescence detector for the simultaneous acquisition of transmission and fluorescence data. It can be used over a wide range of photon energies, photon rates and exposure times at third-generation synchrotron radiation sources, and is currently operating at two beamlines at the Advanced Photon Source. Images obtained at around 2 keV and 10 keV demonstrate the superiority of phase contrast over absorption for specimens composed of light elements.

A short working distance multiple crystal x-ray spectrometer.

For x-ray spot sizes of a few tens of microns or smaller, a millimeter-sized flat analyzer crystal placed approximately 1 cm from the sample will exhibit high energy resolution while subtending a collection solid angle comparable to that of a typical spherically bent crystal analyzer (SBCA) at much larger working distances. Based on this observation and a nonfocusing geometry for the analyzer optic, we have constructed and tested a short working distance (SWD) multicrystal x-ray spectrometer. This prototype instrument has a maximum effective collection solid angle of 0.14 sr, comparable to that of 17 SBCA at 1 m working distance. We find good agreement with prior work for measurements of the Mn Kbeta x-ray emission and resonant inelastic x-ray scattering for MnO, and also for measurements of the x-ray absorption near-edge structure for Dy metal using Lalpha(2) partial-fluorescence yield detection. We discuss future applications at third- and fourth-generation light sources. For concentrated samples, the extremely large collection angle of SWD spectrometers will permit collection of high-resolution x-ray emission spectra with a single pulse of the Linac Coherent Light Source. The range of applications of SWD spectrometers and traditional multi-SBCA instruments has some overlap, but also is significantly complementary.

Inferring the geometry of fourth-period metallic elements in arabidopsis thaliana seeds using synchrotron-based multi-angle X-ray fluorescence mapping.

BACKGROUND: Improving our knowledge of plant metal metabolism is facilitated by the use of analytical techniques to map the distribution of elements in tissues. One such technique is X-ray fluorescence (XRF), which has been used previously to map metal distribution in both two and three dimensions. One of the difficulties of mapping metal distribution in two dimensions is that it can be difficult to normalize for tissue thickness. When mapping metal distribution in three dimensions, the time required to collect the data can become a major constraint. In this article a compromise is suggested between two- and three-dimensional mapping using multi-angle XRF imaging. METHODS: A synchrotron-based XRF microprobe was used to map the distribution of K, Ca, Mn, Fe, Ni, Cu and Zn in whole Arabidopsis thaliana seeds. Relative concentrations of each element were determined by measuring fluorescence emitted from a 10 microm excitation beam at 13 keV. XRF spectra were collected from an array of points with 25 or 30 microm steps. Maps were recorded at 0 and 90 degrees , or at 0, 60 and 120 degrees for each seed. Using these data, circular or ellipsoidal cross-sections were modelled, and from these an apparent pathlength for the excitation beam was calculated to normalize the data. Elemental distribution was mapped in seeds from ecotype Columbia-4 plants, as well as the metal accumulation mutants manganese accumulator 1 (man1) and nicotianamine synthetase (nasx). CONCLUSIONS: Multi-angle XRF imaging will be useful for mapping elemental distribution in plant tissues. It offers a compromise between two- and three-dimensional XRF mapping, as far as collection times, image resolution and ease of visualization. It is also complementary to other metal-mapping techniques. Mn, Fe and Cu had tissue-specific accumulation patterns. Metal accumulation patterns were different between seeds of the Col-4, man1 and nasx genotypes.

X-ray spectroscopy and X-ray diffraction at wavelengths near the K-absorption edge of phosphorus.

Phosphorus is an abundant element in living organisms. It is traceable by its X-ray absorption spectrum which shows a strong white line at its K-edge, comparable with that observed for the L(III) edges of rare earth ions. With purple membrane, the variation of the imaginary part of the anomalous dispersion of phosphorus is found to be close to 20 anomalous electron units. Anomalous diffraction experiments at wavelengths near the K-absorption edge of phosphorus confirm this result. The spatial distribution of lipids derived from anomalous diffraction agrees with earlier results from neutron diffraction. Test experiments on single crystals of the carrier protein using 5.76 A photons gave a first low-resolution diffraction pattern. Various techniques of crystal mounting were attempted. In addition, fluorescence measurements on a solution of threonine synthase appear to hint at a change of the phosphate environment of the cofactor upon activator binding.

Microchemical element imaging of yeast and human cells using synchrotron X-ray microprobe with Kirkpatrick-Baez optics.

Trace element imaging and speciation analysis in cells and subcellular compartments is a challenging and important objective for modern analytical chemistry in order to better understand the biological chemistry of essential and toxic elements. A focusing system based on Kirkpatrick-Baez design optics mounted on a synchrotron radiation scanning X-ray microscope has been developed at the ESRF and was used for trace element quantitative imaging in single cells. The focused microbeam (1.3 x 3.2 microm(2)) obtained in that way led to a photon flux as bright as 1.5 x 10(11) photons/s at 14 keV. The absolute detection limit of this analytical probe, as measured on standard reference materials, was shown to be 2 x 10(-)(17) g for most elements. Chemical maps of human carcinoma and of Saccharomyces cerevisiae cells were obtained for minor (P, S, Cl, K) and trace elements (Fe, Zn). Within human cancer cells, chemical elements are homogeneously distributed at the current spatial resolution and correlated with the sample's mass, except Fe, which shows micrometer-sized structures around the cell nucleus, and Zn, which slightly concentrates in the nucleus, while chemical maps of S. cerevisiae show homogeneous pattern distribution at the cellular level.