The objective of this study is to use synchrotron-based X-ray fluorescence imaging (XFI) and bulk analysis to investigate elements (Mn, Fe, Cu, Zn, P, S, K, Ca) distributions and relative concentrations in four cool-season oat varieties (CDC Arborg, CDC Nasser, CDC Haymaker, and Summit) obtained from the same growing location, soil conditions and harvest time at the University of Saskatchewan. XFI at the Canadian Light Source's BioXAS-Imaging beamline (5 μm resolution, 15 keV) revealed that P, K, Mn, and Zn were concentrated in the aleurone layer, scutellum, and embryo, while Ca was only localized in the aleurone layer and scutellum in the four oat varieties. Notably, S and Cu were distributed in all parts of the seed across four varieties, but the intensity was low in the endosperm. Bulk analysis results show that there were significant differences in the relative concentrations of K, Fe and Zn among four oat varieties harvested for three consecutive years (2018, 2019, 2020) at the completely mature stage. CDC Nasser oat had the lowest K and Zn, while CDC Haymaker had the highest Fe among the oat varieties. These findings highlight the impact of variety on nutritional quality and could help inform future biofortification strategies to enhance the micronutrient content for human and animal diets. This work is the first to map the oat mineral distributions across cool-season varieties using high-resolution XFI. Unlike rice, oats showed minimal mineral accumulation in the hull, ensuring nutritional retention post-milling. Overall, this study shows XFI's potential as a non-destructive tool for cereal grain analysis and supports breeding nutrient-dense oat varieties to address global micronutrient deficiencies.

Membranes and electrode ionomers in proton exchange membrane fuel cells are prone to cation contamination, leading to a reduction in performance. Despite the importance, the characteristics of cation mobility within membranes and ionomer thin films remain poorly understood. Here, we investigate Co2+ transport properties in membranes and electrode ionomers using synchrotron X-ray fluorescence imaging. Specifically, the samples are doped with a controlled Co2+ exchange and the samples are subsequently subject to hydrogen pump operation under fixed humidity and potential gradient. A 1-D model is developed based on the Nernst-Planck relation, which predicts the diffusion and mobility coefficients of Co2+. We also develop a characterization platform, termed Electrode Ionomer Network (EIN), for studying cation transport properties in electrode ionomers that are inherently tortuous and non-uniform. We observe that an increase in relatively humidity from 40 to 75% increases both the diffusion and mobility in Nafion membranes (by factors of 2.2 and 7.1, respectively), determined via fitting the Nernst-Planck relation to our experimental data. Despite the identical humidity conditions, Co2+ become less mobile in EINs relative to membrane (lower by 67% and 44% for diffusivity and mobility, respectively), which are attributed to confinement effects, and the tortuous and disconnected ionomer network in the electrode. Our results provide insights that can help predict cation concentration distributions across membrane-electrode assemblies for hydrogen fuel cell applications.

Bacteria are becoming increasingly resistant to antibiotics, therefore there is an urgent need for new classes of antibiotics to fight antibiotic resistance. Mammals do not express Nɑ -acetyl-L-ornithine deacetylase (ArgE), an enzyme that is critical for bacterial survival and growth, thus ArgE represents a promising new antibiotic drug target, as inhibitors would not suffer from mechanism-based toxicity. A new ninhydrin-based assay was designed and validated that included the synthesis of the substrate analog N5, N5-di-methyl Nα-acetyl-L-ornithine (kcat/Km = 7.32 ± 0.94 × 104 M−1s−1). This new assay enabled the screening of potential inhibitors that absorb in the UV region, and thus is superior to the established 214 nm assay. Using this new ninhydrin-based assay, captopril was confirmed as an ArgE inhibitor (IC50 = 58.7 μM; Ki = 37.1 ± 0.85 μM), and a number of phenylboronic acid derivatives were identified as inhibitors, including 4-(diethylamino)phenylboronic acid (IC50 = 50.1 μM). Selected inhibitors were also tested in a thermal shift assay with ArgE using SYPRO Orange dye against Escherichia coli ArgE to observe the stability of the enzyme in the presence of inhibitors (captopril Ki = 35.9 ± 5.1 μM). The active site structure of di-Zn EcArgE was confirmed using X-ray absorption spectroscopy, and we reported two X-ray crystal structures of E. coli ArgE. In summary, we describe the development of a new ninhydrin-based assay for ArgE, the identification of captopril and phenylboronic acids as ArgE inhibitors, thermal shift studies with ArgE + captopril, and the first two published crystal structures of ArgE (mono-Zn and di-Zn).

The efficiency of N2-fixation in legume–rhizobia symbiosis is a function of root nodule activity. Nodules consist of 2 functionally important tissues: (a) a central infected zone (CIZ), colonized by rhizobia bacteria, which serves as the site of N2-fixation, and (b) vascular bundles (VBs), serving as conduits for the transport of water, nutrients, and fixed nitrogen compounds between the nodules and plant. A quantitative evaluation of these tissues is essential to unravel their functional importance in N2-fixation. Employing synchrotron-based x-ray microcomputed tomography (SR-μCT) at submicron resolutions, we obtained high-quality tomograms of fresh soybean root nodules in a non-invasive manner. A semi-automated segmentation algorithm was employed to generate 3-dimensional (3D) models of the internal root nodule structure of the CIZ and VBs, and their volumes were quantified based on the reconstructed 3D structures. Furthermore, synchrotron x-ray fluorescence imaging revealed a distinctive localization of Fe within CIZ tissue and Zn within VBs, allowing for their visualization in 2 dimensions. This study represents a pioneer application of the SR-μCT technique for volumetric quantification of CIZ and VB tissues in fresh, intact soybean root nodules. The proposed methods enable the exploitation of root nodule’s anatomical features as novel traits in breeding, aiming to enhance N2-fixation through improved root nodule activity.

It is important to know the mineral distribution in cereal grains for nutritional improvement or genetic biofortification. Distributions and intensities of micro-elements (Mn, Fe, Cu, and Zn) and macro-elements (P, S, K and Ca) in Arborg oat were investigated using synchrotron-based on X-ray fluorescence imaging (XFI). Arborg oat provided by the Crop Development Center (CDC, Aaron Beattie) of the University of Saskatchewan for 2D X-ray fluorescence scans were measured at the BioXAS-Imaging beamline at the Canadian Light Source. The results show that the Ca and Mn were mainly localized in the aleurone layer and scutellum. P, K, Fe, Cu, and Zn were mainly accumulated in the aleurone layer and embryo. Particularly the intensities of P, K, Cu, and Zn in the scutellum were higher compared to other areas. S was also distributed in each tissue and its abundance in the sub-aleurone was the highest. In addition, the intensities of S and Cu were highest in the nucellar projection of the crease region. All these elements were also found in the pericarp but they were at lower levels than other tissues. Overall, the details of these experimental results can provide important information for micronutrient biofortification and processing strategies on oat through elemental mapping in Arborg oat.

Mercury is ubiquitous in the environment, with rising levels due to pollution and climate change being a current global concern. Many mercury compounds are notorious for their toxicity, with the potential of organometallic mercury compounds for devastating effects on the structures and functions of the central nervous system being of particular concern. Chronic exposure of human populations to low levels of methylmercury compounds occurs through consumption of fish and other seafood, although the health consequences, if any, from this exposure remain controversial. We have used high energy resolution fluorescence detected X-ray absorption spectroscopy to determine the speciation of mercury and selenium in human brain tissue. We show that the molecular fate of mercury differs dramatically between individuals who suffered acute organometallic mercury exposure (poisoning) and individuals with chronic low-level exposure from a diet rich in marine fish. For long-term low-level methylmercury exposure from fish consumption, mercury speciation in brain tissue shows methylmercury coordinated to an aliphatic thiolate, resembling the coordination environment observed in marine fish. In marked contrast, for short-term high-level exposure, we observe the presence of biologically less available mercuric selenide deposits, confirmed by X-ray fluorescence imaging, as well as mercury(II)-bis-thiolate complexes, which may be signatures of severe poisoning in humans. These differences between low-level and high-level exposures challenge the relevance of studies involving acute exposure as a proxy for low-level chronic exposure.

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Neuronal and sensory toxicity of mercury (Hg) compounds has been largely investigated in humans/mammals with a focus on public health, while research in fish is less prolific and dispersed by different species. Well-established premises for mammals have been governing fish research, but some contradictory findings suggest that knowledge translation between these animal groups needs prudence [e.g. the relative higher neurotoxicity of methylmercury (MeHg) vs. inorganic Hg (iHg)]. Biochemical/physiological differences between the groups (e.g. higher brain regeneration in fish) may determine distinct patterns. This review undertakes the challenge of identifying sensitive cellular targets, Hg-driven biochemical/physiological vulnerabilities in fish, while discriminating specificities for Hg forms.A functional neuroanatomical perspective was conceived, comprising: (i) Hg occurrence in the aquatic environment; (ii) toxicokinetics on central nervous system (CNS)/sensory organs; (iii) effects on neurotransmission; (iv) biochemical/physiological effects on CNS/sensory organs; (v) morpho-structural changes on CNS/sensory organs; (vi) behavioral effects. The literature was also analyzed to generate a multidimensional conceptualization translated into a Rubik's Cube where key factors/processes were proposed.Hg neurosensory toxicity was unequivocally demonstrated. Some correspondence with toxicity mechanisms described for mammals (mainly at biochemical level) was identified. Although the research has been dispersed by numerous fish species, 29 key factors/processes were pinpointed.Future trends were identified and translated into 25 factors/processes to be addressed. Unveiling the neurosensory toxicity of Hg in fish has a major motivation of protecting ichtyopopulations and ecosystems, but can also provide fundamental knowledge to the field of human neurodevelopment.

The newly-created Biological and Life Sciences Department at the Canadian Light Source (CLS) encompasses four sets of beamlines devoted to biological studies ranging in scope from the atomic scale to cells, tissues and whole organisms. The Canadian Macromolecular Crystallography Facility (CMCF) consists of two beamlines devoted primarily to crystallographic studies of proteins and other macromolecules. The Mid-Infrared Spectromicroscopy (Mid-IR) beamline focusses on using infrared energy to obtain biochemical, structural and dynamical information about biological systems. The Bio-Medical Imaging and Therapy (BMIT) facility consists of two beamlines devoted to advanced imaging and X-ray therapy techniques. The Biological X-ray Absorption Spectroscopy (BioXAS) facility is being commissioned and houses three beamlines devoted to X-ray absorption spectroscopy and multi-mode X-ray fluorescence imaging. Together, these beamlines provide CLS Users with a powerful array of techniques to study today’s most pressing biological questions. We describe these beamlines along with their current powerful features and envisioned future capabilities.

Fusarium head blight (FHB) is a serious disease of wheat worldwide. Cultivar resistance to FHB depends on biochemical factors that confine the pathogen spread in spikes. Breeding for cultivar resistance is considered the most practical way to manage this disease. In this study, different spectroscopy and microscopy techniques were applied to discriminate resistance in wheat genotypes against FHB. Synchrotron-based spectroscopy and imaging techniques, including focal plane array infrared and X-ray fluorescence (XRF) spectroscopy were used to understand changes in biochemical and nutrients in rachis following FHB infection. Sumai3 and Muchmore were used to represent resistant and susceptible cultivars to FHB, respectively, in this study. The histological comparison of rachis showed substantial differences in the cell wall thickness between the cultivars after infection. Synchrotron-based infrared imaging emphasized substantial difference in biochemical composition of rachis samples between the two cultivars prior to visible symptoms; in the resistant Sumai3, infrared bands representing lignin and hemicellulose were stronger and more persistent compared to the susceptible cultivar. These bands may be the candidates of biochemical markers for FHB resistance. Focal plane array infrared imaging (FPA) spectra from the rachis epidermis and vascular bundles revealed a new band (1710 cm(-1)) related to the oxidative stress on the susceptible cultivar only. XRF spectroscopy data revealed differences in nutrients composition between cultivars, and between controls and inoculated samples, with substantial increases observed for Ca, K, Mn, Fe, Zn, and Si in the resistant cultivar. These nutrients are related to cell wall stability, metabolic process, and plant defense mechanisms such as lignification pathway and callose deposition. The combination of cell wall composition and lignification plays a role in the mechanism of type II host resistance to FHB. Biochemical profiling using the synchrotron-based spectroscopy holds potential for screening wheat genotypes for FHB resistance.

In recent years larval stage zebrafish have been emerging as a standard vertebrate model in a number of fields, ranging from developmental biology to pharmacology and toxicology. The tyrosinase inhibitor 1-phenyl-2-thiourea (PTU) is used very widely with larval zebrafish to generate essentially transparent organisms through inhibition of melanogenesis, which has enabled many elegant studies in areas ranging from neurological development to cancer research. Here we show that PTU can have dramatic synergistic and antagonistic effects on the chemical toxicology of different mercury compounds. Our results indicate that extreme caution should be used when employing PTU in toxicological studies, particularly when studying toxic metal ions.

Melano-macrophage aggregates, collections of specialized cells of the innate immune system of fish, are considered a general biomarker for contaminant toxicity. To elucidate further the relationship between macrophage aggregates and metals exposure, yelloweye rockfish (Sebastes ruberrimus), a long-lived species, were sampled from the east and west coasts of Prince of Wales Island, Alaska. Metals concentrations in livers (inorganic Hg, methyl mercury, Se, Ni, Cd, Cu, Zn) and spleens (inorganic Hg and methyl mercury) were determined, as well as their correlations with melano-macrophage aggregate area. Sections of liver tissue were analyzed by laser ablation-inductively coupled plasma-mass spectrometry to determine how metals were spatially distributed between hepatocytes and macrophage aggregates. The concentration of inorganic Hg in whole tissue was the best predictor of macrophage area in yelloweye livers and spleens. Macrophage aggregates had higher relative concentrations than most metals compared with the surrounding hepatocytes. However, not all metals were accumulated to the same degree, as evidenced by differences in the ratios of metals in macrophages compared with hepatocytes. Laser ablation data were corroborated with the results of X-ray synchrotron fluorescence imaging of a yelloweye liver section. Hepatic macrophage aggregates in yelloweye rockfish may play an important role in the detoxification and storage of Hg and other metals. Environ Toxicol Chem 2015;34:1918-1925. (c) 2015 SETAC

The compounds of mercury can be more toxic than those of any other non-radioactive heavy element. Despite this, environmental mercury pollution and human exposure to mercury are widespread, and are increasing. While the unusual ability of selenium to cancel the toxicity of mercury compounds has been known for nearly five decades, only recently have some aspects of the molecular mechanisms begun to be understood. We report herein a study of the interaction of mercury and selenium in the larval stage zebrafish, a model vertebrate system, using X-ray fluorescence imaging. Exposure of larval zebrafish to inorganic mercury shows nano-scale structures containing co-localized mercury and selenium. No such co-localization is seen with methylmercury exposure under similar conditions. Micro X-ray absorption spectra support the hypothesis that the co-localized deposits are most likely comprised of highly insoluble mixed chalcogenide HgSxSe(1-x) where x is 0.4-0.9, probably with the cubic zincblende structure.

Methylmercury (MeHg) is a ubiquitous and persistent neurotoxin that poses a risk to human health. Although the mechanisms of MeHg toxicity are not fully understood, factors that contribute to susceptibility are even less well known. Studies of human gene polymorphisms have identified a potential role for the multidrug resistance-like protein (MRP/ABCC) family, ATP-dependent transporters, in MeHg susceptibility. MRP transporters have been shown to be important for MeHg excretion in adult mouse models, but their role in moderating MeHg toxicity during development has not been explored. We therefore investigated effects of manipulating expression levels of MRP using a Drosophila development assay. Drosophila MRP (dMRP) is homologous to human MRP1-4 (ABCC1-4), sharing 50% identity and 67% similarity with MRP1. A greater susceptibility to MeHg is seen in dMRP mutant flies, demonstrated by reduced rates of eclosion on MeHg-containing food. Furthermore, targeted knockdown of dMRP expression using GAL4>UAS RNAi methods demonstrates a tissue-specific function for dMRP in gut, Malpighian tubules, and the nervous system in moderating developmental susceptibility to MeHg. Using X-ray synchrotron fluorescence imaging, these same tissues were also identified as the highest Hg-accumulating tissues in fly larvae. Moreover, higher levels of Hg are seen in dMRP mutant larvae compared with a control strain fed an equivalent dose of MeHg. In sum, these data demonstrate that dMRP expression, both globally and within Hg-targeted organs, has a profound effect on susceptibility to MeHg in developing flies. Our findings point to a potentially novel and specific role for dMRP in neurons in the protection against MeHg. Finally, this experimental system provides a tractable model to evaluate human polymorphic variants of MRP and other gene variants relevant to genetic studies of mercury-exposed populations.

ADVERTISEMENT RETURN TO ISSUEPREVReviewNEXTElemental and Chemically Specific X-ray Fluorescence Imaging of Biological SystemsM. Jake Pushie†, Ingrid J. Pickering†‡§, Malgorzata Korbas∥⊥, Mark J. Hackett†, and Graham N. George*†‡§View Author Information† Molecular and Environmental Sciences Research Group, Department of Geological Sciences, University of Saskatchewan, Saskatoon, SK S7N 5E2, Canada‡ Toxicology Centre, University of Saskatchewan, Saskatoon, SK S7N 5B3, Canada§ Department of Chemistry, University of Saskatchewan, Saskatoon, SK S7N 5C9, Canada∥ Canadian Light Source Inc., 44 Innovation Boulevard, Saskatoon, SK S7N 2V3, Canada⊥ Department of Anatomy and Cell Biology, University of Saskatchewan, Saskatoon, SK S7N 5E5, Canada*E-mail: [email protected]Cite this: Chem. Rev. 2014, 114, 17, 8499–8541Publication Date (Web):August 7, 2014Publication History Received22 December 2013Published online7 August 2014Published inissue 10 September 2014https://doi.org/10.1021/cr4007297Copyright © 2014 American Chemical SocietyRIGHTS & PERMISSIONSACS AuthorChoiceArticle Views13612Altmetric-Citations206LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (27 MB) Get e-AlertsSUBJECTS:Elements,Energy,Fluorescence,Mercury,X-rays Get e-Alerts

Human populations experience widespread low level exposure to organometallic methylmercury compounds through consumption of fish and other seafood. At higher levels, methylmercury compounds specifically target nervous systems, and among the many effects of their exposure are visual disturbances, including blindness, which previously were thought to be due to methylmercury-induced damage to the visual cortex. Here, we employ high-resolution X-ray fluorescence imaging using beam sizes of 500 × 500 and 250 × 250 nm(2) to investigate the localization of mercury at unprecedented resolution in sections of zebrafish larvae ( Danio rerio ), a model developing vertebrate. We demonstrate that methylmercury specifically targets the outer segments of photoreceptor cells in both the retina and pineal gland. Methylmercury distribution in both tissues was correlated with that of sulfur, which, together with methylmercury's affinity for thiolate donors, suggests involvement of protein cysteine residues in methylmercury binding. In contrast, in the lens, the mercury distribution was different from that of sulfur, with methylmercury specifically accumulating in the secondary fiber cells immediately underlying the lens epithelial cells rather than in the lens epithelial cells themselves. Since methylmercury targets two main eye tissues (lens and photoreceptors) that are directly involved in visual perception, it now seems likely that the visual disruption associated with methylmercury exposure in higher animals including humans may arise from direct damage to photoreceptors, in addition to injury of the visual cortex.

Human exposure to potentially neurotoxic methylmercury species is a public-health concern for many populations worldwide. Both fish and whale are known to contain varying amounts of methylmercury species. However studies of populations that consume large quantities of fish or whale have provided no clear consensus as to the extent of the risk. The toxicological profile of an element depends strongly on its chemical form. We have used X-ray absorption spectroscopy to investigate the comparative chemical forms of mercury and selenium in fish and whale skeletal muscle. The predominant chemical form of mercury in whale is found to closely resemble that found in fish. In the samples of skeletal muscle studied, no involvement of selenium in coordination of mercury is indicated in either whale or fish, with no significant inorganic HgSe or HgS type phases being detected. The selenium speciation in fish and whale shows that similar chemical types are present in each, but in significantly different proportions. Our results suggest that for equal amounts of Hg in skeletal muscle, the direct detrimental effects arising from the mercury content from consuming skeletal muscle from whale and fish should be similar if the effects of interactions with other components in the meat are not considered.

Mercury, one of the most toxic elements, exists in various chemical forms each with different toxicities and health implications. Some methylated mercury forms, one of which exists in fish and other seafood products, pose a potential threat, especially during embryonic and early postnatal development. Despite global concerns, little is known about the mechanisms underlying transport and toxicity of different mercury species. To investigate the impact of different mercury chemical forms on vertebrate development, we have successfully combined the zebrafish, a well-established developmental biology model system, with synchrotron-based X-ray fluorescence imaging. Our work revealed substantial differences in tissue-specific accumulation patterns of mercury in zebrafish larvae exposed to four different mercury formulations in water. Methylmercury species not only resulted in overall higher mercury burdens but also targeted different cells and tissues than their inorganic counterparts, thus revealing a significant role of speciation in cellular and molecular targeting and mercury sequestration. For methylmercury species, the highest mercury concentrations were in the eye lens epithelial cells, independent of the formulation ligand (chloride versusl-cysteine). For inorganic mercury species, in absence of l-cysteine, the olfactory epithelium and kidney accumulated the greatest amounts of mercury. However, with l-cysteine present in the treatment solution, mercuric bis-l-cysteineate species dominated the treatment, significantly decreasing uptake. Our results clearly demonstrate that the common differentiation between organic and inorganic mercury is not sufficient to determine the toxicity of various mercury species.