Energy-dispersive X-ray fluorescence analysis of zinc and iron concentration in rice and pearl millet grain

Background and aims

Rice (Oryza sativa L.) and pearl millet (Pennisetum glaucum L.) biofortification breeding programs require accurate and convenient methods to identify nutrient dense genotypes. The aim of this study was to investigate energy-dispersive X-ray fluorescence spectrometry (EDXRF) for the measurement of zinc (Zn) and iron (Fe) concentration in whole grain rice and pearl millet.

Methods

Grain samples were obtained from existing biofortification breeding programs. Reference Zn and Fe concentrations obtained by inductively-coupled plasma-optical emission spectroscopy (ICP-OES) were used to calibrate the EDXRF instrument. Calibration was performed with 24 samples and separate calibrations were developed for rice and pearl millet. To validate calibrations, EDXRF analyses were conducted on an additional 40 samples of each species.

Results

EDXRF results were highly correlated with ICP-OES values for both Zn and Fe in both species (r²?=?0.79 to 0.98). EDXRF predicted Zn and Fe in rice to within 1.9 and 1.6?mg?kg^?1 of ICP-OES values, and Zn and Fe in pearl millet to within 7.6 and 12.5?mg?kg^?1 of ICP-OES values, at a 95% confidence level.

Conclusion

EDXRF offers a convenient, economical tool for screening Zn and Fe concentration in rice and pearl millet biofortification breeding programs.     

Energy-dispersive X-ray fluorescence spectrometry as a tool for zinc, iron and selenium analysis in whole grain wheat

Background and aims

Crop biofortification programs require fast, accurate and inexpensive methods of identifying nutrient dense genotypes. This study investigated energy-dispersive X-ray fluorescence spectrometry (EDXRF) for the measurement of zinc (Zn), iron (Fe) and selenium (Se) concentrations in whole grain wheat.

Methods

Grain samples were obtained from existing biofortification programs. Reference Zn, Fe and Se concentrations were obtained using inductively coupled plasma optical emission spectrometry (ICP-OES) and/or inductively coupled plasma mass spectrometry (ICP-MS). One set of 25 samples was used to calibrate for Zn (19–60?mg?kg^–1) and Fe (26–41?mg?kg^–1), with 25 further samples used to calibrate for Se (2–31?mg?kg^–1 ). Calibrations were validated using an additional 40–50 wheat samples.

Results

EDXRF limits of quantification (LOQ) were estimated as 7, 3 and 2?mg?kg^–1 for Zn, Fe, and Se, respectively. EDXRF results were highly correlated with ICP-OES or -MS values. Standard errors of EDXRF predictions were ±2.2?mg Zn kg^–1, ±2.6?mg Fe kg^–1, and ±1.5?mg Se kg^–1.

Conclusion

EDXRF offers a fast and economical method for the assessment of Zn, Fe and Se concentration in wheat biofortification programs.     

Portable X-ray fluorescence of zinc applied to human toenail clippings

Zinc is an essential trace element in humans. Zinc deficiency can result in a range of serious medical conditions which include effects on growth and development, the immune system, the central nervous system, and the gastrointestinal system. Diagnosis of zinc deficiency is often precluded by the lack of a noninvasive and reliable biomarker. Zinc concentration in nail is considered an emerging biomarker of zinc status in humans. Whether zinc in nail accurately reflects zinc status is beyond the scope of the current study, but is an important research question. The development of a portable method to quickly assess zinc concentration from a single nail clipping could be a useful advance. In this study, single toenail clippings from 60 individuals living in Atlantic Canada were measured for zinc using a portable X-ray fluorescence (XRF) technique. These samples were obtained from the Atlantic PATH cohort, part of the largest chronic disease study ever performed in Canada. Each toenail clipping was measured using three 300 s trials with a mono-energetic portable XRF system. Results were then assessed using two different approaches to the XRF analysis: (1) factory-calibrated zinc concentrations were output from each trial, and (2) energy spectra were analyzed for the characteristic X-rays resulting from zinc. Following the measurement of zinc using the non-destructive portable XRF method, the same clippings were measured for zinc concentration using the “gold standard” technique of inductively coupled plasma-mass spectrometry (ICP-MS). A linear equation of best fit was determined for the relationship between average XRF output zinc concentration and ICP-MS zinc concentration, with a correlation coefficient r = 0.60. Similarly, a linear equation of best fit was found for the relationship between a normalized XRF energy spectrum zinc signal and ICP-MS zinc concentration, with a correlation coefficient r = 0.68. Individual ICP-MS zinc concentrations ranged from 32 μg/g to 140 μg/g, with a population average of 85 μg/g. The results of this study indicate that portable XRF is a sensitive method for the measurement of zinc in a single nail clipping, and provides a reasonable estimation of zinc concentration. Further method development is required before portable XRF be considered a routine alternative to ICP-MS for the assessment of zinc in nail clippings.     

Energy-dispersive X-ray analysis of the extracellular cadmium sulfide crystallites of Klebsiella aerogenes

Klebsiella aerogenes forms electron-dense partieles on the cell surface in response to the presence of cadmium ions in the growth medium. These particles ranged from 20 to 200 nm in size, and quantitative energy dispersive X-ray analysis established that they comprise cadmium and sulfur in a 1:1 ratio. This observation leads to the conclusion that the particles are cadmium sulfide crystallites. A combination of atomic absorption spectroscopy, inductively coupled plasma mass spectrometry, and acid-labile sulfide analysis revealed that the total intracellular and bound extracellular cadmium:sulfur ratio is also 1:1, which suggests that the bulk of the cadmium is fixed as extracellular cadmium sulfide. The tolerance of K. acrogenes to cadmium ions and the formation of the cadmium sulfide crystallites were dependent on the buffer composition of the growth medium. The addition of cadmium ions to phosphate-buffered media resulted in cadmium phosphate precipitates that remove the potentially toxic cadmium ions from the growth medium. Electrondense particles formed on the surfaces of bacteria grown under these conditions were a combination of cadmium sulfide and cadmium phosphates. The specific bacterial growth rate in the exponential phase of batch cultures was not affected by up to 2mM cadmium in Tricine-buffered medium, but formation of cadmium sulfide crystallites was maximal during the stationary phase of batch culture. Cadmium tolerance was much lower (10 to 150 M) in growth media buffered with Tris, Bistris propane, Bes, Tes, or Hepes. These results illustrate the importance of considering medium composition when comparing levels of bacterial cadmium tolerance.Abbreviations EDXA Energy dispersive X-ray analysis - AAS Atomic absorption spectroscopy - TEM Transmission electron microscopy - SEM Scanning electron microscopy - ICP-MS Inductively coupled plasma mass spectrometry - ALSA Acid-labile sulfide analysis     

Energy-dispersive X-ray microanalysis of membrane-associated inclusions in hydrogenosomes isolated from Trichomonas vaginalis

Energy-dispersive X-ray microprobe analysis of electron-dense inclusions in hydrogenosomes isolated from the aerotolerant anaerobic protozoal human parasite Trichomonas vaginalis, Bushby strain, indicated the presence of high levels of Mg, P and Ca. This suggested that divalent cation (e.g. Ca2+ or Mg2+) accumulation by hydrogenosomes may be important in the regulation of intracellular ion concentrations.     

Non-linear least-squares methods applied to the analysis of fluorescence energy transfer measurements.

We describe a new method for calculating the efficiency of fluorescence energy transfer on labeled macromolecules using steady-state measurements. A single estimation of the efficiency value is obtained by a global analysis of all the measurement data sets (absorption, emission and excitation spectra) using non-linear least-squares. The method was tested on simulated and experimental data obtained from three simple model compounds: an equimolar mixture of tryptophan-tyrosine and two peptides, Trp-Tyr and Trp-Gly-Gly-Tyr, in which transfer efficiencies are respectively nearly 100% and 50%. The method was found to be reliable and provides methodological and quantitative advantages in regard to the sequential methods currently used.     

Selective fluorescence detection of histamine based on ligand exchange mechanism and its application to biomonitoring

We report on a novel histamine monitoring method by using a fluorescent probe, a complex between Ni²⁺ and calcein, based on a ligand exchange mechanism. The fluorescence intensity of this probe, which has been reduced due to effective quenching by Ni²⁺ ion, increases drastically by an addition of histamine. Furthermore, the probe shows high selectivity toward histamine among the various neurotransmitters in 0.1 M phosphate buffer solution (pH 7.4). Biomonitoring studies to detect histamine released from RAW264 cells are successfully represented.     

Digital dynamic range expansion applied to X-ray densitometric analysis of total hip replacement

Due to the presence of the stiff prosthetic stem fitted in the medullary canal during total his replacement, the surrounding cortex of the femur changes its density over time. This bone remodelling takes place with every type of total hip prosthesis; however, its intensity may vary between prostheses and patients. In the worst cases this process can lead to the late failure of the implant. To monitor such bone density evolution, we are developing a tailored Computer-aided Densitometric Image Analysis system (the major part of this our system uses an 8-bit commercial hardware with 256 levels of grey). The equivalent dynamic range of an X-ray picture is about 10 bits. In this paper we present a method to overcome these hardware limitations by improving the software. Using a double-exposure acquisition it is possible to build a 9-bit image that is good enough for most applications involving bone density measurement.     

High energy X-ray scattering tomography applied to bone

High energy synchrotron X-ray scattering was developed for reconstruction of specimen cross-sections. The technique was applied to a model specimen of cortical bone containing a capillary tube of silicon, and reconstructions were produced with either full diffraction rings or texture-related subsets of a given ring. The carbonated apatite (cAp) 00.2 and 22.2 reconstructions and the Si 311 reconstructions agreed with absorption-based reconstructions from the measured X-ray transmissivity recorded during diffraction pattern acquisition and from reconstructions produced subsequently of the same specimen using a commercial microCT (microComputed Tomography) scanner.     

X-ray fluorescence analysis using synchrotron radiation at the photon factory

X-ray fluorescence experiments at the Photon Factory in Japan are described. An energy-dispersive X-ray fluorescence system was combined with various excitation modes, i.e., a continuum and a monochromatic excitation, which consist of a crystal monochromator or a wide band pass monochromator. These excitation modes provide a wide range of band width and photon flux of excitation beams. Minimum detection limits obtained for a thin sample were less than 0.1 ppm and 0.1 pg when there was no line interference. Advantages of using monochromatic excitation are discussed, with emphasis on the possibility of chemical state analysis. Grazing incidence X-ray fluorescence is a technique very appropriate to synchrotron radiation characteristics. Near-surface analysis and trace analysis of solution samples placed on a total reflection support were made. Future plans are discussed, including microbeam analysis, tomography, X-ray excited optical fluorescence, and applications of insertion devices (undulator and wiggler).     

Time-domain fluorescence lifetime imaging applied to biological tissue.

Fluorescence lifetime imaging (FLIM) is a functional imaging methodology that can provide information, not only concerning the localisation of specific fluorophores, but also about the local fluorophore environment. It may be implemented in scanning confocal or multi-photon microscopes, or in wide-field microscopes and endoscopes. When applied to tissue autofluorescence, it reveals intrinsic excellent contrast between different types and states of tissue. This article aims to review our recent progress in developing time-domain FLIM technology for microscopy and endoscopy and applying it to biological tissue.     

Comparison of SR-excited X-ray fluorescence analysis with neutron activation analysis for hair and fiber

Human scalp hair and some kinds of vegetable and animal fibers were analyzed by means of the SR excited X-ray fluorescence method (SRXFA) and the neutron activation method (NAA). Human hair samples collected from five males and five females were washed by the IAEA method prior to analysis. In the SRXFA analysis, samples were excited by monochromated X-rays. Fluorescence X-rays were measured by an Si(Li) detector. The elements detected in all hair samples were S, Ca, Cu, Fe, Zn, Br, and Sr. The elements K, Ti, Cr, Mn, Ni, Se, Hg, and Pb were also detected in several samples. After SRXFA analysis these same samples were analyzed by the NAA method. Elements such as Cu, Zn, and Br were detected by both methods, and their relative concentrations show a good agreement of variation between individuals. However, Pb was only detected by SRXFA, and Na, Au, and Sb were only detected by NAA. Therefore, these two methods are complementary to each other for trace element analysis.     

Application of X-ray absorption and X-ray fluorescence techniques to the study of metallodrug action

X-ray absorption spectroscopy and X-ray fluorescence microscopy are two synchrotron-based techniques frequently deployed either individually or in tandem to investigate the fates of metallodrugs and their biotransformation products in physiologically relevant sample material. These X-ray methods confer advantages over other analytical techniques in that they are nondestructive and require minimal chemical or physical manipulation of the sample before analysis, conserving both chemical and spatial information of the element(s) under investigation. In this review, we present selected examples of the use of X-ray absorption spectroscopy and X-ray fluorescence microscopy in studies of metallodrug speciation and localisation in vivo, in cell spheroids and in intact tissues and organs, and offer recent highlights in the advances of these techniques as they pertain to research on metallodrug action.     

Comparison of X-ray fluorescence and optical fluorescence measured behind scattering layers as a basis for X-ray fluorescence endoscopy.

Recent developments in the technology of capillary-fiber optics suitable for X-rays in the range of approximately 4-10keV point to the possible realization of endoscopes applicable in X-ray fluorescence analysis. A general problem is the determination of scattering and absorption processes with consideration to tissue optics, X-ray scattering and X-ray absorption in a diagnostic partial volume. Therefore comparative investigations were performed in order to answer these questions. Zinc-oxide nanoparticles configured as single particles and ZnO clusters provided the fluorescence source in cell layers. An artificial scattering material was employed, which closely approximated the tissue optical conditions and the X-ray optical application conditions in possible diagnostic situations. As a result imaging of spatially resolved X-ray contrasts was better than adequate optical fluorescence imaging by approximately one magnitude. Hence a very important precondition for realizing X-ray fluorescence endoscopy is fulfilled.     

Vegetation on avalanche paths in the alps

Vegetation pattern on avalanche paths in the Nordkette Mountains near Innsbruck (Tyrol, Austria) was studied using transect sampling. The plant communities represent a gradient from alpine grassland (Seslerio-Caricetum sempervirentis, via grassland with Trisetum distichophyllum) and nutrient-poor grassland (Carlino-Caricetum sempervirentis) to an avalanche grassland proper. Along this gradient disturbance decreases from higher to lower elevations. The topography of the avalanche path suggests a gradient from the central zone to the edge of the path, although a floristic zonation or grouping of species across the path cannot be detected. Disturbance and local topography are the dominant factors for the vegetation pattern on the avalanche paths investigated.Abbreviations DCA= detrended correspondence analysis     

Multicolor fluorescence in situ hybridization (FISH) applied to FISH-banding

During the last decade not only multicolor fluorescence in situ hybridization (FISH) using whole chromosome paints as probes, but also numerous chromosome banding techniques based on FISH have been developed for the human and for the murine genome. This review focuses on such FISH-banding techniques, which were recently defined as 'any kind of FISH technique, which provide the possibility to characterize simultaneously several chromosomal subregions smaller than a chromosome arm. FISH-banding methods fitting that definition may have quite different characteristics, but share the ability to produce a DNA-specific chromosomal banding'. While the standard chromosome banding techniques like GTG lead to a protein-related black and white banding pattern, FISH-banding techniques are DNA-specific, more colorful and, thus, more informative. For some, even high-resolution FISH-banding techniques the development is complete and they can be used for whole genome hybridizations in one step. Other FISH-banding methods are only available for selected chromosomes and/or are still under development. FISH-banding methods have successfully been applied in research in evolution- and radiation-biology, as well as in studies on the nuclear architecture. Moreover, their suitability for diagnostic purposes has been proven in prenatal, postnatal and tumor cytogenetics, indicating that they are an important tool with the potential to partly replace the conventional banding techniques in the future.     

Canopy chlorophyll fluorescence applied to stress detection using an easy-to-build micro-lidar

Photosynthesis Research - The dark-to-light transitions enable energization of the thylakoid membrane (TM), which is reflected in fast and slow (OJIPSMT or OABCDE) stages of fluorescence induction...     

Biomedical applications of photon-induced X-ray fluorescence

An X-ray fluorescence method for in vitro analysis of trace elements is presented. The method is characterized by the use of an X-ray tube with secondary targets as a quasimonoenergetic radiation source, and by “infinitely thin” specimens. Different aspects have been examined in order to optimize the sensitivity of the method. It is extremely important to use secondary targets as pure as possible and collimators internally covered by the same element as the secondary target. It is also important to reduce the contribution at the XRF spectrum of photons scattered by the sample, by the sample support, and by the air. Preconcentration techniques can conveniently also be used to this purpose. In this work, biological samples are preconcentrated by reducing them to ash. Typical sensitivities obtained are in the order of 1–5 ng/cm² in a counting time of 10³ s for elements with atomic number ranging from 24 (chromium) to 40 (zirconium).     

Overall-response parameter analysis applied to butyrophenones

If the first (and second) principal component function of psycho pharmacologically defined parameters rationalize most of the information, the component may be regressed against physicochemical features in order to study quantitative relationships between structure and function. An example is the analysis of butyrophenones.     

Energy-dispersive X-ray microanalysis of the methanogenMethanosarcina barkeri ‘Fusaro’ grown on methanol and in the presence of heavy metals

The general distribution of metals in cell compartments ofMethanosarcina barkeri ‘Fusaro’ was studied by energy-dispersive X-ray microanalysis. Cells were cultivated on methanol, acetate, or on a mixture of H₂+CO₂ as substrate. At decreasing degree (polyphosphate bodies ? clusters of small granula>cell walls, inside a cell clump> cytoplasma), the elemental pattern of Ca, P, and Fe could be seen in all cell compartments. Cells grown on methanol were further investigated for their ability to incorporate added heavy metal ions. After completed growth and methane production, Pb or Cd was found to a considerable extent in all cell compartments, whereas Zn or Cu was incorporated only to a minor degree. Evidence for an uptake of Hg could not be obtained.