Synchrotron Radiation Induced X-ray Emission Research Articles

Analysis of synchrotron radiation induced X-ray emission spectra with R environment

Life sciences have seen a huge increase in the amount and complexity of data being collected with every experiment. Scientists today are faced with increasingly difficult task to extract vital information from the vast amount of numbers. Software used for this purpose should be sufficiently powerful and flexible to handle large and complex data sets. On the other hand it should allow the user to exactly follow what is being calculated, black-box type of software should be avoided. R platform (R Development Core Team, 2011), open-source environment for statistical analysis nicely fits these requirements. With its rapidly expanding user community it is quickly becoming the most important tool in statistical analysis of data in broad range of applications. The most important feature of R is the package system, allowing users to address specific problems with dedicated package and even for more advanced users to contribute software for their own fields. In this paper multivariate analysis and data treatment of spectral data by using R environment is presented.

Synchrotron radiation induced X-ray emission studies of the antioxidant mechanism of the organoselenium drug ebselen

Synchrotron radiation induced X-ray emission (SRIXE) spectroscopy was used to map the cellular uptake of the organoselenium-based antioxidant drug ebselen using differentiated ND15 cells as a neuronal model. The cellular SRIXE spectra, acquired using a hard X-ray microprobe beam (12.8-keV), showed a large enhancement of fluorescence at the K(α) line for Se (11.2-keV) following treatment with ebselen (10 μM) at time periods from 60 to 240 min. Drug uptake was quantified and ebselen was shown to induce time-dependent changes in cellular elemental content that were characteristic of oxidative stress with the efflux of K, Cl, and Ca species. The SRIXE cellular Se distribution map revealed that ebselen was predominantly localized to a discreet region of the cell which, by comparison with the K and P elemental maps, is postulated to correspond to the endoplasmic reticulum. On the basis of these findings, it is hypothesized that a major outcome of ebselen redox catalysis is the induction of cellular stress. A mechanism of action of ebselen is proposed that involves the cell responding to drug-induced stress by increasing the expression of antioxidant genes. This hypothesis is supported by the observation that ebselen also regulated the homeostasis of the transition metals Mn, Cu, Fe, and Zn, with increases in transition metal uptake paralleling known induction times for the expression of antioxidant metalloenzymes.

Iron valence states in organic samples and tissues investigated by XANES and Mössbauer spectroscopy

AbstractThe results of XANES and Mössbauer spectroscopy studies of organic samples of [Fe(Hka)3] (Hka = kojic acid) and [Fe(etmal)3] (etmal = ethyl maltol) are presented and discussed in terms of their application in characterization of the local electronic state of Fe3+. In case of [Fe(etmal)3], a tiny increase in covalency of iron bonds was found. The local iron states in tissues have been concluded from x‐ray absorption near‐edge structures (XANES) spectra, compared with spectra obtained for the organic samples. Significant increase of iron concentration in pathological cells of the prostate was shown by the synchrotron radiation‐induced x‐ray emission (SRIXE) method. XANES analysis of healthy and cancerous tissue samples suggested higher iron oxidation state in pathological cells. This result may be a hint for understanding the role of iron in carcinogenesisand opens up the same possibility of using XANES spectroscopy as a diagnostic tool in oncology. The correlation between results obtained by XANES and Mössbauer spectroscopy may be helpful in overcoming the limitations characteristic of both of the methods in the determination of the iron valence and spin states in tissues. Copyright © 2008 John Wiley & Sons, Ltd.

Application of Linear Discriminant Analysis in Prostate Cancer Research by Synchrotron Radiation-Induced X-Ray Emission

The ability to visualize an object of interest is one of the cornerstones of advancement in science. For this reason, synchrotron radiation-induced X-ray emission (micro-SRIXE) holds special promise as a imaging technique in structural biology, biochemistry, and medicine. It gives the possibility to image concentration of most of the elements in a sample at high space resolution. Statistical analysis of data obtained for samples of prostate tissues in an experiment at L-beam line HASYLAB (Hamburg, Germany) is presented in this paper. The regions for the measurements were selected according to the histological view of the sample. By histological examination, samples were divided into five groups (from healthy to Gleason4, most advanced stage of cancerogenesis). Data obtained in micro-SRIXE experiments on prostate cancer samples provide information about concentrations of certain elements in these groups. The rising problem is to find out concentrations of which elements allow the researcher to discriminate between different (early mentioned) groups. Linear discriminant analysis, a basic technique for feature extraction, was used in statistical analysis of the data. Our results indicate that the use of synchrotron radiation and discriminant analysis in the study of prostate cancer tissues provide information that can be key to better understanding of biomolecular functions.

Transport kinetics of four- and six-coordinate platinum compounds in the multicell layer tumour model

Four-coordinate (Pt(II)) platinum-based anticancer drugs are widely used in primary or palliative chemotherapy and produce considerable efficacy in certain clinical applications, for example testicular cancer. However, in many cancers the Pt(II) drugs are beset by poor efficacy mainly due to suboptimal pharmacokinetic properties. Consequently, the six-coordinate (Pt(IV)) class of Pt drugs were developed to improve platinum efficacy by (i) increasing stability, (ii) reducing reactivity, (iii) increasing lipophilicity, and (iv) nuclear targeting. However, comparatively little information is available on the pharmacokinetic properties of these compounds within solid tumour tissue. In the present study, the distribution and fluxes of [14C]-labelled [PtCl2(en)] (where en stands for ethane-1,2-diamine) and cis,trans-[PtCl2(OH)2(en)] drugs were determined in the multicell layer (MCL) tumour model comprising colon cancer cells. Flux data were analysed by mathematical modelling of drug diffusion and cellular uptake in the transport system. The flux of the Pt(IV) compound through the MCL was not significantly different to that of the Pt(II) drug nor were the diffusion coefficient or tissue uptake; the latter confirmed with elemental imaging analysis by synchrotron radiation induced X-ray emission. However, the flux of the Pt(IV) through the MCL was increased by hydrostatic pressure, thereby demonstrating the potential to target cancer cells further away from the vessels with six-coordinate platinum drugs.

The application of synchrotron radiation induced X-ray emission in the measurement of zinc and lead in Wistar rat ameloblasts

The development of analytical techniques for the measurement of trace elements in cellular compartments of developing teeth remains an important methodological issue in dental research. Recent advances in third generation synchrotron facilities have provided high brilliance X-ray sources that can be effectively used to study trace element distributions in small spatial regions with low detection limits. The present study describes for the first time the application of synchrotron radiation induced X-ray emission (SRIXE) in measuring the distribution of zinc and lead in the ameloblasts of developing Wistar rat teeth. Wistar rats were fed a standard rat diet, containing the normal dietary requirements of zinc, ad libitum and exposed to 100 ppm of lead in drinking water. Resin embedded sections of first mandibular molars were analysed using a 13.3 keV incident monochromatic X-ray beam focussed to a 0.2 μm spot. Characteristic X-rays arising from the entire thickness of the sample were measured using an energy dispersive detector for quantitative analysis of elemental concentrations. The results showed that intranuclear concentrations of zinc were greater than levels in the cytoplasm. Furthermore, nuclear and cytoplasmic concentrations of zinc in the maturation stage (742 ± 27 and 424 ± 25 ppm, respectively) were significantly higher than the zinc levels observed in the nucleus and cytoplasm of presecretory stage ameloblasts (132 ± 10 and 109 ± 10 ppm, respectively) ( p < 0.05). A clear lead signal above the background was not detected in the ameloblasts and lead concentrations could only be reliably measured in the developing enamel. Overall, SRIXE was an effective method of studying the spatial distribution of zinc in the cells of developing teeth and offered a unique combination of sub-micron spatial resolution and parts-per-million detection limits (0.8–1 and 0.6–1 ppm for zinc and lead, respectively).

Risk assessment of TiO 2 photocatalyst by individual micrometer-size particle analysis with on-site combination of SEM-EDX and SR-XANES microscope

Applications of synchrotron radiation X-ray fluorescence (SR-XRF) microscopy combined with scanning electron microscopy (SEM) are reported. Electron beam excited and synchrotron radiation induced X-ray emission spectra of the same yellow sand single particles are reported and compared. The Ti-K edge absorption fine structure of single microparticles of TiO 2 (rutile, anatase, and a photocatalyst aerosol) are recorded by using monochromatic synchrotron radiation of tunable energy. It is shown that the discrimination between rutile and anatase is possible. Based on the single particle speciation, the toxicity of photocatalyst aerosol powder is discussed.

Preliminary Study on Chemical Speciation of Sulphur in Cancerous Tissues

Sulphur is vital for almost all living organisms by participating in a wide variety of metabolic processes. Nevertheless its biochemistry is only partially understood, due to it a few tools such as X-ray absorption near edge structure may be used to determine its chemical speciation in biological system. In this paper we focused on analysis of composition and elemental distribution in tissue structures of biological samples. We examined cancerous and noncancerous prostate tissues. Experiments were carried out on the L-beam line at the HASYLAB, DESY (Hamburg, Germany) and DAΦNE-Light Facility DRX1 at the Laboratori Nationali di Frascati (Frascati, Italy). Synchrotron radiation induced X-ray emission measurements reveal the significant differences in concentration of sulphur between cancerous and non-cancerous tissues. X-ray absorption near edge structure spectra show two main types of sulphur that are represented by X-ray peaks at 2476 eV and 2484 eV. This suggests that different sulphur components are present in the measured samples.

Elemental Mapping of Prostate Tissue by Micro-SRIXE

Synchrotron radiation-induced X-ray emission is now a proven analytical method for the determination of trace elements. Better insight of the role of trace elements in tissue samples can be gained by acquiring complete distribution maps of these elements. Our approach is based on the acquisition of digital maps. In this contribution we present two difierent methods for obtaining normalization matrix: flrst | based on argon intensity in the single spectrum and second | based on changes of beam current intensity. The region for the scan was carefully chosen according to the histological view of the sample. A total area was scanned with a step size of 15 „m in each direction and measuring time of 30 s per pixel. The thickness of the samples was 15 „m. A polycapillary was used for beam focusing. At the energy of 18 keV the beam size on the sample was approximately 15 „m which is of the order of a cell diameter.

Cancerous tissues analyzed by SRIXE

Synchrotron radiation induced X-ray emission (SRIXE) analysis of trace element (TE) concentrations in prostate tissues shows the significant differences between elemental concentrations in cancer and healthy parts of tissues. The analysis enables obtaining additional information about the prostate cancer, almost the most dangerous disease of the recent days. The cancer tissue sections were obtained after radical removal of the organ from the patient's body. The samples were histologically examined and classified as well as the Gleason score was determined. After histological examination, the samples were irradiated with monochromatic X-ray beam at the beam line L at HASYLAB in DESY. Our investigation proved that the elements such as Mn, Fe, Co, Ni which are involved in cancerogenesis appeared on elevated concentration levels in cancerous part of the tissues in comparison to non-cancerous parts. This study may also indicate that the advancement of prostate cancer is connected with the content of copper, zinc and iron.

Iron and other elements studies in cancerous and non-cancerous prostate tissues

To understand the role of trace elements in the oxidative stress, which may be an important factor in the pathogenesis of prostate cancer, we investigated the distribution and chemical state of Fe using synchrotron radiation. Therefore, both synchrotron radiation induced X-ray emission (SRIXE) and X-ray absorption near-edge structure (XANES) techniques have been applied to study cancerous and non-cancerous prostate tissues. The results obtained in this work prove that SRIXE and XANES techniques are complementary ones for analysis of trace elements in biomedical samples. The absorption K-edge position of iron determined in cancerous tissue is shifted by 2.9 eV towards higher energies in comparison to iron absorption K-edge determined in non-cancerous tissue. The information obtained in this way may help to understand the role of oxidation state of iron in the oxidative stress underlying the oncogenesis.

Time-dependent uptake, distribution and biotransformation of chromium(VI) in individual and bulk human lung cells: application of synchrotron radiation techniques

Chromium(VI) is a human carcinogen, primarily affecting the respiratory tract probably via active transport into cells, followed by the reduction to Cr(III) with the formation of DNA-damaging intermediates. Distribution of Cr and endogenous elements within A549 human lung adenocarcinoma epithelial cells, following treatment with Cr(VI) (100 microM, 20 min or 4 h) were studied by synchrotron-radiation-induced X-ray emission (SRIXE) of single freeze-dried cells. After the 20-min treatment, Cr was confined to a small area of the cytoplasm and strongly co-localized with S, Cl, K, and Ca. After the 4-h treatment, Cr was distributed throughout the cell, with higher concentrations in the nucleus and the cytoplasmic membrane. This time-dependence corresponded to approximately 100% or 0% clonogenic survival of the cells following the 20-min or 4-h treatments, respectively, and could potentially be explained by a new cellular protective mechanism. Such processes may also be important in reducing the potential hazards of Cr(III) dietary supplements, for which there is emerging evidence that they exert their anti-diabetic effects via biological oxidation to Cr(VI). The predominance of Cr(III) was confirmed by micro-XANES spectroscopy of intracellular Cr hotspots. X-ray absorption spectroscopy (XANES and EXAFS, using freeze-dried cells after the 0-4-h treatments) was used to gain insight into the chemical structures of Cr(III) complexes formed during the intracellular reduction of Cr(VI). The polynuclear nature of such complexes (probably with a combination of carboxylato and hydroxo bridging groups and O-donor atoms of small peptides or proteins) was established by XAFS data analyses.

Application of SRIXE and XANES to the determination of the oxidation state of iron in prostate tissue sections

Normal prostate and cancerous prostate tissue sections were analyzed using synchrotron radiation-induced X-ray emission and X-ray absorption near-edge structure (XANES) at the X-26A microprobe beam line located at the National Synchrotron Light Source, Brookhaven National Laboratory, USA. In both measurements, a monochromatic beam of size of 16×14 μm 2 was applied. Two-dimensional scans of selected areas of non-cancerous and cancerous tissue sections were performed in order to obtain the distributions of various trace element concentrations. The XANES spectra were recorded for selected points containing high concentrations of iron. These measurements were performed with the aim of determining the oxidation state of iron. It was found that the iron concentration was much higher in the cancerous sections of the analyzed tissues than in the non-cancerous sections. From the results of XANES spectra analysis, iron in cancerous prostate tissue sections mostly occurs in the +3 oxidation state.

The cellular distribution and oxidation state of platinum(II) and platinum(IV) antitumour complexes in cancer cells.

The cellular distribution of platinum in A2780 ovarian cancer cells treated with cisplatin and platinum(IV) complexes with a range of reduction potentials has been examined using elemental analysis (synchrotron radiation-induced X-ray emission). The cellular distribution of platinum(IV) drugs after 24 h is similar to that of cisplatin, consistent with the majority of administered platinum(IV) drugs being reduced. Micro-X-ray absorption near-edge spectra of cells treated with cisplatin and platinum(IV) complexes confirmed the reduction of platinum(IV) to platinum(II). In cells treated, the most difficult to reduce complex, cis, trans, cis-[PtCl(2)(OH)(2)(NH(3))(2)], platinum(IV) was detected in the cells along with platinum(II). The observations are in accordance with the relative ease of reduction of the platinum(IV) complexes used and support the requirement of reduction for activation of platinum(IV) complexes.

Literature trends in x‐ray emission spectrometry in the period 1990–2000–a review

AbstractA quantitative overview is given of the role that x‐ray emission analysis methods, in their various forms, play in the literature for the period from January 1990 to the end of December 2000. The major sources of information were computerized searches through Chemical Abstracts and Web of Science and, specifically for 1998, a manual search through Analytical Abstracts. Areas that are covered in this review pertain to the recent trends in x‐ray emission spectrometry in general and in some specific techniques such as x‐ray fluorescence analysis, synchrotron radiation‐induced and particle‐induced x‐ray emission, total‐reflection and micro‐x‐ray fluorescence analysis. A brief outline of recent developments in the specific fields is given, with emphasis on the various excitation and detection modes, on different application areas and on relative contributions of different countries and languages to the x‐ray emission spectrometry literature. It appears that environmental monitoring and research continue to be particularly important publication fields for x‐ray spectrometry. Copyright © 2003 John Wiley &amp; Sons, Ltd.

Trace elements distribution in renal cell carcinoma depending on stage of disease.

The aim of this study was to identify those trace elements which can be used to distinguish between normal and malignant tissue in renal cell cancer (RCC) kidney and to assess changes in trace elements concentration in tissue with progressing malignant disease. In case control study, 36 cases of RCC were analyzed by Synchrotron Radiation Induced X-ray Emission (SRIXE) in order to establish the concentration of 19 elements. Patients with RCC were examined to obtain staging of disease after radical nephrectomy, which was performed in each case. Results were compared with 15 control kidney cortex tissue obtained during autopsy in which cause of death was trauma. The most relevant decrease was detected in Cd content: from 81 +/- 39.2 ppm in normal control samples to 16.6 +/- 22.2 ppm concentration in RCC. We found that the concentrations of Ti, Pb and Rb were also lower in RCC tissue. On the other hand, the RCC tissue was rich in iron and zirconium. With the progress of malignant disease, assessed by TNM (UICC 1997) scale, lower concentration of S and higher concentration of Ca in both RCC and neoplastic kidney cortex can be seen. The same tendency is observed in Zn and Se concentrations. Cadmium shows raising concentration with progress of RCC only in cortex of neoplastic kidney. In all cases it was shown that the relatively high tissue concentration of iron in both investigated tissues is decreasing with the progress of disease. The zirconium has shown raising tissue concentration in advanced disease. Trace elements concentration is different in malignant tissue and surrounding macroscopically unchanged kidney cortex. Progress of the disease is connected with changes in trace elements concentration. This may reflect different biology of compared tissue with potential practical implication.

Calcium compartments in brain.

Excellent progress has been made toward understanding the physiology and pharmacology of specific calcium-related cellular processes of the brain, but few studies have provided an integrated view of brain calcium kinetics. To further the knowledge of the size and binding properties of brain calcium compartments, the authors have conducted a series of experiments in hippocampal brain slices exposed to high and low extracellular calcium. Slices were incubated in buffers containing 0.001 to 4.5 mmol/L calcium for up to 75 minutes. Slice calcium content was analyzed by three methods: exchange equilibrium with 45Ca, synchrotron-radiation-induced x-ray emission, and inductively coupled plasma. Data were analyzed using a model based on a Langmuir isotherm for two independent sites, with additional extracellular and bound compartments. In parallel experiments, altered low calcium had no effect on slice histology and only mild effects on slice adenylates. When combined with prior 45Ca and fluorescent probe binding experiments, these results suggest that there are at least five kinetically distinct calcium compartments: (1) free extracellular (approximately 10%); (2) loosely associated extracellular plasma membrane (approximately 55%); (3) intracellular compartment with moderate avidity (approximately 17%); (4) tightly bound, nonexchangeable intracellular compartment ( approximately 15%); and (5) free cytoplasmic (<0.01%). If only the third compartment is considered a potential calcium buffer, then the buffering ratio is calculated to be approximately 2,700:1, but if the second compartment is also included, then the buffering ratio would be approximately 13,000:1. This may explain the wide range of estimates observed by fluorescent probe studies.

Hard X-ray microprobe studies of chromium(VI)-treated V79 Chinese hamster lung cells: intracellular mapping of the biotransformation products of a chromium carcinogen

The uptake of carcinogenic and mutagenic Cr compounds and the intracellular distribution of their biotransformation products in V79 Chinese hamster lung cells were studied by synchrotron-radiation-induced X-ray emission (SRIXE). SRIXE analysis was performed on whole cells that had been treated with either Cr(III) or Cr(V) 1,10-phenanthroline complexes, or Cr(VI). The high spatial resolution (0.3 microm) and elemental sensitivity (~10(-15) g Cr/cell) of the technique provided detailed maps of Cr and other cellular elements in thin sections prepared from Cr(VI)-treated cells. The Cr carcinogen concentrated in P-rich regions corresponding to the nucleus, as well as other areas of the cell that are likely to correspond to organelles. This is the first study that has enabled the determination of the localization of the biotransformation products of Cr(VI) carcinogens in a target lung cell.

Synchrotron Radiation-Induced X-Ray Emission to Identify Metal Ions in Preparations of Purified Protein

The suitability of synchrotron radiation-induced X-ray emission (SRIXE) for the detection and identification of metal ions in preparations of purified, soluble proteins was tested. Glutathione S-transferase fused to the proximal zinc finger motif of human transcription factor IIIA or to the cysteine-rich motif of poliovirus protein 2C was expressed in bacteria and purified by affinity chromatography. Aqueous samples containing the purified proteins were analyzed with SRIXE, and trace amounts of zinc and iron were detected. Mutation of the zinc-coordinating residues in the cysteine-rich motif of poliovirus protein 2C resulted in the loss of the zinc-binding ability. Relative quantities of metal in the protein preparations as determined by SRIXE corresponded well with the metal:protein ratios calculated by using a 4-(2-pyridylazo)resorcinol-based assay. We conclude that SRIXE is an accurate, sensitive, and simple method for the detection and identification of protein-bound metal ions in small amounts of sample. Thus, SRIXE may have wide use as a particularly effective method for rapidly determining trace metals in microarray samples.

Preliminary study on the distribution of selected elements in cancerous and non-cancerous kidney tissues

In this study special interest was given to trace elements recognized as to be carcinogenic to humans. The kidney tissue sections were analyzed in order to determine the concentrations of elements present in the sample. The Synchrotron Radiation Induced X-ray Emission (SRIXE) technique was applied using a white photon microbeam. The results from cancerous parts of the kidney tissues were compared to non-cancerous parts and to the control group. In addition the iron concentration level was determined in the serum of those patients. Two-dimensional scans are presented to illustrate the differences between perfused and not-perfused tissues. According to this study there is no significant difference in the Mn concentration between cancerous and non-cancerous parts of the kidney, but the concentrations of Cd, Cr, Ti, V, Cu, Se, and Zn are at a lower concentration level in the cancerous parts than in the non-cancerous parts. A converse observation has been made for Fe. This may be associated with different metabolism and dynamics of the cancer process and both higher vascularization and need of higher blood supply in the cancerous tissue. The two-dimensional scanning of thin kidney sections showed differences in the trace element distributions depending on the analyzed samples: perfused and non-perfused. Perfusion removed blood mostly from the peritubular capillaries while in the glomerulus some capillaries had a relatively high Fe content. A low Fe concentration was observed in nephron tubules while a converse observation has been made for Cd. This may indicate that Cd is localized in the cells but not in the blood.