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A method for the analysis of Pt–protein complexes in biological samples, previously subjected to cisplatin treatment, has been developed. Proteins were separated by gel electrophoresis, and those bound to Pt were detected with high sensitivity by LA-ICP-(SF)-MS. Pt-containing spots were in-gel digested with trypsin, and the peptides produced identified using nHPLC-ESI-LTQ-FT-MS/MS. The influence of protein separation conditions, staining and gel processing prior to laser ablation on Ptprotein bonds preservation have been evaluated using standard proteins incubated with cisplatin. 2-DE separation under non-reducing conditions followed by either Coomassie blue brilliant or silver staining is appropriate for Pt–protein complexes, achieving a good separating resolution of the proteins in biological samples. Direct LA-ICP-MS analysis of glycerol-treated dried gels for Pt–protein monitoring resulted in better sensitivity, more reliable relative Pt signals and a simpler and less time-consuming approach compared to the analysis of blotted membranes. Ablation of gels allowed tackling protein identification of Pt-spots in the remaining non-ablated material in the gel, making it unnecessary to run several gels in parallel for separate Pt detection and protein identification. By using this approach, Pt coordinated to proteins, such as α-2-macroglobulin, transferrin, albumin or hemoglobin, was detected in the serum from a rat treated in vivo with cisplatin after nrSDS-PAGE separation. Furthermore, the first complete LA-ICP-MS metalloprotein contour map in a 2-DE gel has been produced, in this case for the detection of Pt–protein complexes in renal proximal tubule epithelial cells (RPTECs) incubated with cisplatin. Several proteins were identified in those spots containing Pt, which may have a connection with the drug-induced nephrotoxicity mainly affecting this cell type in the kidney.
The experimental density data of the binary System nitrogen−hydrogen available at the time of the development of the equation of state for natural gases and related mixtures, GERG-2008, were limited to hydrogen contents higher than 0.15 (amount-of-substance fraction) and temperatures above 270 K. This work provides accurate experimental (p, ρ, T) data for three binary mixtures of nitrogen and hydrogen: (0.95 N2 + 0.05 H2), (0.90 N2 + 0.10 H2), and (0.50 N2 + 0.50 H2) at temperatures of 240, 250, 260, 275, 300, 325, and 350 K, thus extending the range of available experimental data to low hydrogen contents and low temperatures. The density measurements were performed by using a single-sinker densimeter with magnetic suspension coupling at pressures up to 20 MPa. Experimental data were compared with the corresponding densities calculated from the GERG-2008 and the AGA8-DC92 equations of state. The relative deviations of the experimental data from both equations of state were within the estimated uncertainty value of the equations. Therefore, the experimental data agree very well with the values estimated from the equations. The virial coefficients B(T,x), C(T,x), and D(T,x) as well as the second interaction virial coefficient B12(T) for the nitrogen−hydrogen binary system were also calculated from the experimental data set at temperatures from 240 to 350 K. The resulting values agree with those from literature.
A pilot study for the thickness measurement of HfO2 films was performed by the Surface Analysis Working Group (SAWG) of the Consultative Committee for Amount of Substance (CCQM). The aim of this pilot study was to ensure the equivalency in the measurement capability of national metrology institutes for the thickness measurement of HfO2 films. In this pilot study, the thicknesses of six HfO2 films with nominal thickness from 1 nm to 4 nm were measured by X-ray Photoelectron Spectroscopy (XPS), X-ray Reflectometry(XRR), X-ray Fluorescence Analysis (XRF), Transmission Electron Spectroscopy (TEM), Spectroscopic Ellipsometry (SE) and Rutherford Backscattering Spectrometry (RBS). The reference thicknesses were determined by mutual calibration of a zero-offset method (Medium Energy Ion Scattering Spectroscopy (MEIS) of KRISS) and a method traceable to the length unit (the average thicknesses of three XRR data except the thinnest film). These reference thicknesses are traceable to the length unit because they are based on the traceability of XRR. For the thickness measurement by XPS, the effective attenuation length of Hf 4f electrons was determined. In the cases of XRR and TEM, the offset values were determined from a linear fitting between the reference thicknesses and the individual data by XRR and TEM. The amount of substance of HfO2, expressed as thickness of HfO2 films (in both linear and areal density units), was found to be a good subject for a CCQM key comparison.
To reach the main text of this paper, click on Final Report.
The final report has been peer-reviewed and approved for publication by the CCQM.
This key comparison aims to assess the core capabilities of the participants in gas analysis. Such competences include, among others, the capabilities to prepare Primary Standard gas Mixtures (PSMs), perform the necessary purity analysis on the materials used in the gas mixture preparation, the verification of the composition of newly prepared PSMs against existing ones, and the capability of calibrating the composition of a gas mixture. According to the Strategy for Key Comparisons of the Gas Analysis Working Group, this key comparison is classified as an RMO track A key comparison.
The artefacts were binary mixtures of propane in nitrogen at a nominal amount-of-substance fraction level of 1000 μmol/mol. The values and uncertainties from the gravimetric gas mixture preparation were used as key comparison reference values (KCRVs). Each transfer standard had its own KCRV. The results are generally good. All results are within ± 1 % of the KCRV.
Numerous membrane importers rely on accessory water-soluble proteins to capture their substrates. These substrate-binding proteins (SBP) have a strong affinity for their ligands; yet, substrate release onto the low-affinity membrane transporter must occur for uptake to proceed. It is generally accepted that release is facilitated by the association of SBP and transporter, upon which the SBP adopts a conformation similar to the unliganded state, whose affinity is sufficiently reduced. Despite the appeal of this mechanism, however, direct supporting evidence is lacking. Here, we use experimental and theoretical methods to demonstrate that an allosteric mechanism of enhanced substrate release is indeed plausible. First, we report the atomic-resolution structure of apo TeaA, the SBP of the Na+-coupled ectoine TRAP transporter TeaBC from Halomonas elongata DSM2581T, and compare it with the substrate-bound structure previously reported. Conformational free-energy landscape calculations based upon molecular dynamics simulations are then used to dissect the mechanism that couples ectoine binding to structural change in TeaA. These insights allow us to design a triple mutation that biases TeaA toward apo-like conformations without directly perturbing the binding cleft, thus mimicking the influence of the membrane transporter. Calorimetric measurements demonstrate that the ectoine affinity of the conformationally biased triple mutant is 100-fold weaker than that of the wild type. By contrast, a control mutant predicted to be conformationally unbiased displays wild-type affinity. This work thus demonstrates that substrate release from SBPs onto their membrane transporters can be facilitated by the latter through a mechanism of allosteric modulation of the former.
A multiproxy approach to evaluate biocidal treatments on biodeteriorated majolica glazed tiles
(2016)
The Fishing House located on the grounds of the Marquis of Pombal Palace, Oeiras, Portugal, was built in the 18th century. During this epoch, Portuguese gardens, such as the one surrounding the Fishing House, were commonly ornamented with glazed wall tile claddings. Currently, some of these outdoor tile panels are covered with dark colored biofilms, contributing to undesirable aesthetic changes and eventually inducing chemical and physical damage to the tile surfaces. Phylogenetic analyses revealed that the investigated biofilms are mainly composed of green algae, cyanobacteria and dematiaceous fungi. With the aim of mitigating biodeterioration, four different biocides (TiO2 nanoparticles, Biotin® T, Preventol® RI 80 and Albilex Biostat®) were applied in situ to the glazed wall tiles. Their efficacy was monitored by visual examination, epifluorescence microscopy and DNA-based analysis. Significant changes in the microbial community composition were observed 4 months after treatment with Preventol® RI 80 and Biotin® T. Although the original community was inactivated after these treatments, an early stage of re-colonization was detected 6 months after the biocide application. TiO2 nanoparticles showed promising results due to their self-cleaning effect, causing the detachment of the biofilm from the tile surface, which remained clean 6 and even 24 months after biocide application.
Numerous membrane importers rely on accessory water-soluble proteins to capture their substrates. These substrate-binding Proteins (SBP) have a strong affinity for their ligands; yet, Substrate release onto the low-affinity membrane transporter must occur for uptake to proceed. It is generally accepted that release is facilitated by the association of SBP and transporter, upon which the SBP adopts a conformation similar to the unliganded state, whose affinity is sufficiently reduced. Despite the appeal of this mechanism, however, direct supporting evidence is lacking. Here, we use experimental and theoretical methods to demonstrate that an allosteric mechanism of enhanced substrate release is indeed plausible.
First, we report the atomic-resolution structure of APO TeaA, the SBP of the Na!-coupled ectoine TRAP transporter TeaBC from Halomonas elongata DSM2581T, and compare it with the substrate-bound structure previously reported. Conformational freeenergy landscape calculations based upon molecular Dynamics simulations are then used to dissect the mechanism that couples ectoine binding to structural change in TeaA. These insights allow us to design a triple mutation that biases TeaA toward apo-like conformations without directly perturbing the binding cleft, thus mimicking the influence of the membrane transporter. Calorimetric measurements demonstrate that the ectoine affinity of the conformationally biased triple mutant is 100-fold weaker than that of the wild type. By contrast, a control mutant predicted to be conformationally unbiased displays wild-type affinity. This work thus demonstrates that substrate release from SBPs onto their Membrane transporters can be facilitated by the latter through a mechanism of allosteric modulation of the former.
Experimental characterization of the thermodynamic behavior of multicomponent gas mixtures of accurately known composition is of great importance for testing the current reference equation of state for natural gases and related mixtures, GERG-2008. Natural gas composition may change considerably due to the diversity of origins, or the possibility to incorporate hydrogen, biogas or other non-conventional energy gases.
Thus, the Consultative Committee for Amount of Substance: Metrology in Chemistry and Biology (CCQM) of the Bureau International des Poids et Mesures (BIPM) launched a key comparison on primary standard mixtures for natural gases of this particular characteristics. One mixture is a hydrogen-enriched natural gas mixture (13 components, x(H2) = 0.03) with low calorific value to facilitate support to power-to-gas applications, which often involve blending hydrogen with natural gas. The other mixture resembles a typical liquefied natural gas (“LNG”, 9 components) with high calorific value.
The GERG-2008 and AGA8-DC92 equations of state enable the calculation of thermal and caloric properties for natural gases and other related mixtures consisting of up to 21 pure components. Experimental characterization of the thermodynamic behavior of gas binary mixtures is of great importance because most of the binary systems were taken into account by using adjusted reducing functions for density and temperature. Only for those binary mixtures for which sufficient accurate experimental data were available, specific departure functions were developed. For the binary mixture methane-helium no departure function was developed so far.
This work studied the thermodynamic behavior of two gravimetrically prepared methane-helium binary mixtures through accurate (p, ρ, T) experimental data obtained by using a single-sinker densimeter with magnetic suspension coupling.
This work provides accurate experimental (p, rho, T) values for three binary mixtures of methane with helium: (0.95 (amount-of-substance fraction) CH₄+ 0.05 He) and (0.90 CH₄+ 0.10 He) at temperatures of (240, 250, and 260) K and (0.50 CH₄+ 0.50 He) from (240 to 400) K. This work is a continuation of a previous one which reported accurate experimental (p, rho, T) values for the (0.95 CH₄+ 0.05 He) and the (0.90 CH₄+ 0.10 He) binary mixtures over the temperature range from (250 to 400) K. All density measurements were performed by using a single-sinker densimeter with magnetic suspension coupling at pressures up to 20 MPa. Experimental values were compared with the corresponding densities calculated from the GERG-2008 and the AGA8-DC92 equations of state, respectively. Deviations from the GERG-2008 are much larger than from the AGA8-DC92 (up to -6.5%). These deviations increase with decreasing temperature, with increasing pressure, and with increasing helium fraction. In contrast, deviations from the AGA8-DC92 are within the 0.5% band. The experimental values were also used to calculate the second and the third virial coefficients, B(T,x) and C(T,x), as well as the second interaction virial coefficient B12(T) for this mixture.
Experimental characterization of the thermodynamic behavior of multicomponent gas mixtures of accurately known composition with traceability to National Standard is of great importance for testing the current reference equation of state (EoS) for natural gases and related mixtures, GERG-2008. Natural gas composition may change considerably due to the diversity of origins, or the possibility to incorporate hydrogen, biomethane or other non-conventional energy gases.
In this work, density measurements for three synthetic mixtures, representing two biomethane mixtures and a typical natural gas composition, are presented. Results are compared to the current reference EoS for natural gases and related mixtures, i.e., GERG-2008 and AGA8-DC92.
The basis for the development and evaluation of equations of state for mixtures is experimental data for several thermodynamic properties. The quality and the availability of experimental data limit the achievable accuracy of the equation. Referring to the fundamentals of GERG-2008 wide-range equation of state, no suitable data were available for many mixtures containing secondary natural gas components. This work provides accurate experimental (p,rho,T) data for two binary mixtures of methane with Helium (0.95 (amount-of-substance fraction) CH4 + 0.05 He and 0.90 CH4 + 0.10 He). Density measurements were performed at temperatures between (250 and 400) K and pressures up to 20 MPa by using a single-sinker densimeter with magnetic suspension coupling. Experimental data were compared with the corresponding densities calculated from the GERG-2008 and the AGA8-DC92 equations of state. Deviations from GERG-2008 were found within a 2% band for the (0.95 CH4 + 0.05 He) mixture but exceeded the 3% Limit for the (0.95 CH4 + 0.05 He) mixture. The highest deviations were observed at T = 250 K and pressures between (17 and 19) MPa. Values calculated from AGA8-DC92, however, deviated from the experimental data by only 0.1% at high pressures and exceeded the 0.2% limit only at temperatures of 300 K and above, for the (0.90 CH4 + 0.10 He) mixture.
The lead authors failed to name two collaborators as co-authors. The authors listed should include:
Miss Claudia L. Compean-Gonzalez (ORCID:
0000-0002-2367-8450) and Dr. Giacomo Ceccone (ORCID:
0000-0003-4637-0771).
These co-authors participated in VAMAS project A27, provided data that were analyzed and presented in this publication (and supporting information), and reviewed the manuscript before submission.
Most of the experimental density data of the methane−hydrogen binary system available at the time of the development of the equation of state for natural gases and related mixtures, GERG-2008, at temperatures above 270 K were limited to hydrogen contents higher than 0.20 (amount-of-substance fraction). On the contrary, for mixtures with hydrogen contents lower than 0.20, experimental data were limited to temperatures below 270 K. This work intends to close the gap and provides accurate experimental (p, ρ, and T) data for three binary mixtures of methane and hydrogen, (0.95 CH4 + 0.05 H2), (0.90 CH4 + 0.10 H2), and (0.50 CH4 + 0.50 H2), at temperatures of 240, 250, 260, 275, 300, 325, and 350 K, thus extending the range of available experimental data to higher temperatures for mixtures with hydrogen contents lower than 0.20 and, accordingly, to lower temperatures for mixtures with hydrogen contents higher than 0.20. The density measurements were performed by using a single-sinker densimeter with magnetic suspension coupling at pressures up to 20 MPa. Experimental data were compared to the corresponding densities calculated from the GERG-2008 and the AGA8-DC92 equations of state, respectively. The experimental data are within the uncertainty of both equations of state, except at the lower temperatures of 240 and 250 K and pressures over 14 MPa for the mixtures with a hydrogen content of 0.05 and 0.10, respectively. The virial coefficients B(T, x) and C(T, x), as well as the second interaction virial coefficient B12(T) for the methane−hydrogen binary system, were also calculated from the experimental data set at temperatures from 240 to 350 K using the virial equation of state.
The GERG-2008 equation of state is the approved ISO standard (ISO 20765-2) for the calculation of thermophysical properties of natural gas mixtures. The composition of natural gas can vary considerably due to the diversity of origin. Further diversification was generated by adding hydrogen, biogas, or other non-conventional energy gases. In this work, high-precision experimental (p, rho, T) data for two gravimetrically prepared synthetic natural gas mixtures are reported. One mixture resembled a conventional natural gas of 11 components (11 M) with a nominal mixture composition (amount-of-substance fraction) of 0.8845 for methane as the matrix compound with the other compounds being 0.005 for oxygen, 0.04 for nitrogen, 0.015 for carbon dioxide, 0.04 for ethane, 0.01 for propane, 0.002 each for n- and isobutane, and ultimately 0.0005 each for isopentane, n-pentane, and nhexane. The other mixture was a 13-component hydrogen-enriched natural gas with a low calorific value featuring a nominal composition of 0.7885 for methane, 0.03 for hydrogen, 0.005 for helium, 0.12 for nitrogen, 0.04 for carbon dioxide, 0.0075 for ethane, 0.003 for propane, 0.002 each for n- and isobutane, and 0.0005 each for neopentane, isopentane, npentane, and n-hexane. Density measurements were performed in an isothermal operational mode at temperatures between 260 and 350 K and at pressures up to 20 MPa by using a single-sinker densimeter with magnetic suspension coupling. The data were compared with the corresponding densities calculated from both GERG-2008 and AGA8-DC92 equations of state to test their performance on real mixtures. The average absolute Deviation from GERG-2008 (AGA8-DC92) is 0.027% (0.078%) for 11 M and 0.095% (0.062%) for the 13-component H2-enriched mixture, respectively. The corresponding maximum relative Deviation from GERG-2008 (AGA8-DC92) amounts to 0.095% (0.127%) for 11 M and 0.291% (0.193%) for the H2-enriched mixture.
The first international round-robin exercise for the measurement of the long-lived radionuclide 10Be has been conducted. Ten participating accelerator mass spectrometry (AMS) facilities have each measured three samples at the 10-12 to 10-14 10Be/9Be level. All results have been made traceable to the NIST SRM 4325 standard to avoid additional discrepancies that arise when different facilities use different calibration materials. Hence, the data concentrates on pure measurement distinctions. Multivariate statistical investigations have been performed to reveal a bias between facilities, i.e. two distinguished groups could be identified. Maximum discrepancies between two single facilities are in the range of 6–31% depending on the absolute 10Be/9Be value. These findings should be considered when comparing 10Be data produced at one AMS facility with that produced at another facility, which is e.g. often the case for in situ 10Be dating studies. Round-robin exercises are a very helpful tool as part of an overall quality assurance scheme to improve the accuracy, and not only the precision, of AMS data.
We report the results of a Versailles Project on Advanced Materials and Standards interlaboratory study on the intensity scale calibration of x-ray photoelectron spectrometers using low-density polyethylene (LDPE) as an alternative material to gold, silver, and copper. An improved set of LDPE reference spectra, corrected for different instrument geometries using a quartz-monochromated Al Kα x-ray source, was developed using data provided by participants in this study. Using
these new reference spectra, a transmission function was calculated for each dataset that participants provided. When compared to a similar calibration procedure using the NPL reference spectra for gold, the LDPE intensity calibration method achieves an absolute offset of ∼3.0% and a systematic deviation of ±6.5% on average across all participants. For spectra recorded at high pass energies (≥90 eV), values of absolute offset and systematic deviation are ∼5.8% and ±5.7%, respectively, whereas for spectra collected at lower pass energies (<90 eV), values of absolute offset and systematic deviation are ∼4.9% and ±8.8%, respectively; low pass energy spectra perform worse than the global average, in terms of systematic deviations, due to diminished count rates and signal-to-noise ratio. Differences in absolute offset are attributed to the surface roughness of the LDPE induced by sample preparation. We further assess the usability of LDPE as a secondary reference material and comment on its performance in the presence of issues such as variable dark noise, x-ray warm up times, inaccuracy at low count rates, and underlying spectrometer problems. In response to participant feedback and the results of the study, we provide an updated LDPE intensity calibration protocol to address the issues highlighted in the interlaboratory study. We also comment on the lack of implementation of a consistent and traceable intensity calibration method across the community of x-ray photoelectron spectroscopy (XPS) users and, therefore, propose a route to achieving this with the assistance of instrument manufacturers, metrology laboratories, and experts leading to an international standard for XPS intensity scale calibration.
In the last few years, coal mine methane (CMM) has gained significance as a potential non-conventional gas fuel. The progressive depletion of common fossil fuels reserves and, on the other hand, the positive estimates of CMM resources as a by-product of mining promote this fuel gas as a promising alternative fuel. The increasing importance of its exploitation makes it necessary to check the capability of the present-day models and equations of state for natural gas to predict the thermophysical properties of gases with a considerably different composition, like CMM. In this work, accurate density measurements of a synthetic CMM mixture are reported in the temperature range from (250 to 400) K and pressures up to 15 MPa, as part of the research project EMRP ENG01 of the European Metrology Research Program for the characterization of non-conventional energy gases. Experimental data were compared with the densities calculated with the GERG-2008 equation of state. Relative deviations between experimental and estimated densities were within a 0.2% band at temperatures above 275 K, while data at 250 K as well as at 275 K and pressures above 10 MPa showed higher deviations.