Oxytetracycline hyper-production through targeted genome reduction of Streptomyces rimosus.

Most biosynthetic gene clusters (BGC) encoding the synthesis of important microbial secondary metabolites, such as antibiotics, are either silent or poorly expressed; therefore, to ensure a strong pipeline of novel antibiotics, there is a need to develop rapid and efficient strain development approaches. This study uses comparative genome analysis to instruct rational strain improvement, using Streptomyces rimosus, the producer of the important antibiotic oxytetracycline (OTC) as a model system. Sequencing of the genomes of two industrial strains M4018 and R6-500, developed independently from a common ancestor, identified large DNA rearrangements located at the chromosome end. We evaluated the effect of these genome deletions on the parental S. rimosus Type Strain (ATCC 10970) genome where introduction of a 145 kb deletion close to the OTC BGC in the Type Strain resulted in massive OTC overproduction, achieving titers that were equivalent to M4018 and R6-500. Transcriptome data supported the hypothesis that the reason for such an increase in OTC biosynthesis was due to enhanced transcription of the OTC BGC and not due to enhanced substrate supply. We also observed changes in the expression of other cryptic BGCs; some metabolites, undetectable in ATCC 10970, were now produced at high titers. This study demonstrated for the first time that the main force behind BGC overexpression is genome rearrangement. This new approach demonstrates great potential to activate cryptic gene clusters of yet unexplored natural products of medical and industrial value.IMPORTANCEThere is a critical need to develop novel antibiotics to combat antimicrobial resistance. Streptomyces species are very rich source of antibiotics, typically encoding 20-60 biosynthetic gene clusters (BGCs). However, under laboratory conditions, most are either silent or poorly expressed so that their products are only detectable at nanogram quantities, which hampers drug development efforts. To address this subject, we used comparative genome analysis of industrial Streptomyces rimosus strains producing high titers of a broad spectrum antibiotic oxytetracycline (OTC), developed during decades of industrial strain improvement. Interestingly, large-scale chromosomal deletions were observed. Based on this information, we carried out targeted genome deletions in the native strain S. rimosus ATCC 10970, and we show that a targeted deletion in the vicinity of the OTC BGC significantly induced expression of the OTC BGC, as well as some other silent BGCs, thus suggesting that this approach may be a useful way to identify new natural products.

Transient Ruddlesden-Popper-Type Defects and Their Influence on Grain Growth and Properties of Lithium Lanthanum Titanate Solid Electrolyte.

Lithium lanthanum titanate (LLTO) perovskite is one of the most promising electrolytes for all-solid-state batteries, but its performance is limited by the presence of grain boundaries (GBs). The fraction of GBs can be significantly reduced by the preparation of coarse-grained LLTO ceramics. In this work, we describe an alternative approach to the fabrication of ceramics with large LLTO grains based on self-seeded grain growth. In compositions with the starting stoichiometry for the Li0.20La0.60TiO3 phase and with a high excess addition of Li (Li:La:Ti = 11:15:25), microstructure development starts with the formation of the layered RP-type Li2La2Ti3O10 phase. Grains with many RP-type defects initially develop into large platelets with thicknesses of up to 10 µm and lengths over 100 µm. Microstructure development continues with the crystallization of LLTO perovskite, epitaxially on the platelets and as smaller grains with thinner in-grain RP-lamellae. Theoretical calculations confirmed that the formation of RP-type sequences is energetically favored and precedes the formation of the LLTO perovskite phase. At around 1250 °C, the RP-type sequences become thermally unstable and gradually recrystallize to LLTO via the ionic exchange between the Li-rich RP-layers and the neighboring Ti and La layers as shown by quantitative HAADF-STEM. At higher sintering temperatures, LLTO grains become free of RP-type defects and the small grains recrystallize onto the large platelike seed grains via Ostwald ripening. The final microstructure is coarse-grained LLTO with total ionic conductivity in the range of 1 × 10-4 S/cm.

Adjusting the Operational Potential Window as a Tool for Prolonging the Durability of Carbon-Supported Pt-Alloy Nanoparticles as Oxygen Reduction Reaction Electrocatalysts.

A current trend in the investigation of state-of-the-art Pt-alloys as proton exchange membrane fuel cell (PEMFC) electrocatalysts is to study their long-term stability as a bottleneck for their full commercialization. Although many parameters have been appropriately addressed, there are still certain issues that must be considered. Here, the stability of an experimental Pt-Co/C electrocatalyst is investigated by high-temperature accelerated degradation tests (HT-ADTs) in a high-temperature disk electrode (HT-DE) setup, allowing the imitation of close-to-real operational conditions in terms of temperature (60 °C). Although the US Department of Energy (DoE) protocol has been chosen as the basis of the study (30,000 trapezoidal wave cycling steps between 0.6 and 0.95 VRHE with a 3 s hold time at both the lower potential limit (LPL) and the upper potential limit (UPL)), this works demonstrates that limiting both the LPL and UPL (from 0.6-0.95 to 0.7-0.85 VRHE) can dramatically reduce the degradation rate of state-of-the-art Pt-alloy electrocatalysts. This has been additionally confirmed with the use of an electrochemical flow cell coupled to inductively coupled plasma mass spectrometry (EFC-ICP-MS), which enables real-time monitoring of the dissolution mechanisms of Pt and Co. In line with the HT-DE methodology observations, a dramatic decrease in the total dissolution of Pt and Co has once again been observed upon narrowing the potential window to 0.7-0.85 VRHE rather than 0.6-0.95 VRHE. Additionally, the effect of the potential hold time at both LPL and UPL on metal dissolution has also been investigated. The findings demonstrate that the dissolution rate of both metals is proportional to the hold time at UPL regardless of the applied potential window, whereas the hold time at the LPL does not appear to be as detrimental to the stability of metals.

Non-matrix-matched calibration in bulk multi-element laser ablation - Inductively coupled plasma - Mass spectrometry analysis of diverse materials.

Laser ablation inductively coupled plasma - mass spectrometry (LA-ICP-MS) is a frequently used microanalytical technique in elemental analysis of solid samples. In most instances the use of matrix-matched calibration standards is necessary for the accurate determination of elemental concentrations. However, the main drawback of this approach is the limited availability of certified reference materials. Here, we present a novel conceptual framework in LA-ICP-MS quantification without the use of matrix-matched calibration standards but instead employment of an ablation volume-normalization method (via measurement of post-ablation line scan volumes by optical profilometry) in combination with a matrix-adapted fluence (slightly above the ablation threshold). This method was validated by cross-matrix quantification of reference materials typically investigated by LA-ICP-MS, including geological and biological materials. This allows for more accurate and precise multi-element quantification, and enables quantification of previously unquantifiable elements/materials.

Toxicity of nitrophenolic pollutant 4-nitroguaiacol to terrestrial plants and comparison with its non-nitro analogue guaiacol (2-methoxyphenol).

Phenols, and especially their nitrated analogues, are ubiquitous pollutants and known carcinogens which have already been linked to forest decline. Although nitrophenols have been widely recognized as harmful to different aquatic and terrestrial organisms, we could not find any literature assessing their toxicity to terrestrial plants. Maize (monocot) and sunflower (dicot) were exposed to phenolic pollutants, guaiacol (GUA) and 4-nitroguaiacol (4NG), through a hydroponics system under controlled conditions in a growth chamber. Their acute physiological response was studied during a two-week root exposure to different concentrations of xenobiotics (0.1, 1.0, and 10 mM). The exposure visibly affected plant growth and the effect increased with increasing xenobiotic concentration. In general, 4NG affected plants more than GUA. Moreover, sunflower exhibited an adaptive response, especially to low and moderate GUA concentrations. The integrity of both plant species deteriorated during the exposure: biomass and photochemical pigment content were significantly reduced, which reflected in the poorer photochemical efficiency of photosystem II. Our results imply that 4NG is taken up by sunflower plants, where it could enter a lignin biosynthesis pathway.

Evaluation of two-phase sample transport upon ablation of gelatin as a proxy for soft biological matrices using nanosecond laser ablation - inductively coupled plasma - mass spectrometry.

BACKGROUND: Recent papers on LA-ICP-MS have reported that certain elements are transported in particulate form, others in gaseous form and still others in a combination of both upon ablation of C-based materials. These two phases display different transport behaviour characteristics, potentially causing smearing in elemental maps, and could be processed differently in the ICP which raises concerns as to accuracy of quantification and emphasizes the need for matrix-matching of external standards. This work aims at a better understanding of two-phase sample transport by evaluating the peak profile changes observed upon varying parameters such as laser energy density and wavelength. RESULTS: It is demonstrated that upon ablation of gelatin, elements are transported predominantly in particulate phase, but already at low laser energy density, a significant fraction of some elements is transported in the gaseous phase, which is even more expressed at higher energy density. This behaviour is element-specific since the ratio of the signal intensity for the analyte element transported in gas phase to the total signal intensity was 0 % for 23Na, 43 % for 66Zn and as high as 95 % for 13C using a 193 nm laser. The results also suggest an effect of the laser wavelength, as all elements show either the same or higher amount of gas phase formation upon ablating with 213 nm versus 193 nm. It was even established that elements that fully occur in particulate form upon ablation using 193 nm laser radiation are partly converted into gaseous phase when using 213 nm. SIGNIFICANCE: This work provides a thorough investigation of the underexposed phenomenon of two-phase sample transport upon ablation of biological samples upon via LA-ICP-MS. It is shown that for some elements a fraction of the ablated material is converted and transported in the gas phase, which can lead to significant smearing effects. As such, it is important to evaluate element-specific peak profiles on beforehand and, if necessary, adapt instrument settings and slow down data acquisition.

Utilizing ablation volume for calibration in LA-ICP-MS mapping to address variations in ablation rates within and between matrices.

Quantification in 2D LA-ICP-MS mapping generally requires matrix-matched standards to minimize issues related to elemental fractionation. In addition, internal standardization is commonly applied to correct for instrumental drift and fluctuation, whereas also differences in ablated mass can be rectified for samples that cannot be sectioned and subjected to total ablation. However, it is crucial that the internal standard element is homogeneously distributed in the sample and that the laser light absorptivity is uniform over the surface. As in practice these requirements are often not met, this work will focus on correction of ablation rate differences within/between samples and standards by normalizing the element maps using the associated ablation volume per pixel as measured by optical profilometry. Due to the volume correction approach the element concentrations are no longer defined as mass per mass concentrations (in µg g-1) but by mass per volume concentrations (in µg cm-3), which can be interconverted in case matrix densities are known. The findings show that ablation volume-aided calibration yields more accurate element concentrations in 2D LA-ICP-MS maps for a decorative glass with highly varying elemental concentrations (murrina). This research presents a warning that if there are variations in ablation rates between samples and standards within and across matrices, even when their sensitivities are the same, generic LA-ICP-MS calibration protocols may not accurately depict the actual element concentrations.

Kinetics and product identification of water-dissolved nitroguaiacol photolysis under artificial sunlight.

Nitroguaiacols are typical constituents of biomass-burning emissions, including absorbing aerosols which contribute to climate change. Although they are also harmful to humans and plants, their atmospheric fate and lifetimes are still very speculative. Therefore, in this work, the photolysis kinetics of aqueous-phase 4-nitroguaiacol (4NG) and 5-nitroguaiacol (5NG), and the resulting photo-formed products were investigated under artificial sunlight, observing also the effect of sunlight on the absorption properties of the solutions. We found the photolysis of 5NG slower than that of 4NG, whereas the absorbance in the visible range prevailed in the 5NG solutions at the end of experiments. Although we identified dinitroguaiacol as one of the 4NG photolysis products, which increased light absorption of 4NG-containing solutions, considerably more chromophores formed in the 5NG photolyzed solutions, implying its stronger potential for secondary BrC formation in the atmosphere. In general, denitration, carbon loss, hydroxylation, nitration, and carbon gain were characteristic of 4NG phototransformation, while carbon loss, hydroxylation, and carbon gain were observed in the case of 5NG. The photolysis kinetics was found of the first order at low precursor concentrations (<0.45 mM), resulting in their lifetimes in the order of days (125 and 167 h illumination for 4NG and 5NG, respectively), which suggests long-range transport of the investigated compounds in the atmosphere and proposes their use as biomass-burning aerosol tracer compounds.

Exploring the Benefits of Ablation Grid Adaptation in 2D/3D Laser Ablation Inductively Coupled Plasma Mass Spectrometry Mapping through Geometrical Modeling.

This study aims to investigate the potential benefits of adapting the ablating grid in two-dimensional (2D) and three-dimensional (3D) laser ablation inductively coupled plasma mass spectrometry in a single pulse mapping mode. The goals include enhancing the accuracy of surface sampling of element distributions, improving the control of depth-related sampling, smoothing the post-ablation surface for layer-by-layer sampling, and increasing the image quality. To emulate the capabilities of currently unavailable laser ablation stages, a computational approach using geometrical modeling was employed to compound square or round experimentally obtained 3D crater profiles on variable orthogonal or hexagonal ablation grids. These grids were optimized by minimizing surface roughness as a function of average ablation depth, followed by simulating the post-ablation surface and related image quality. An online application (https://laicpms-apps.ki.si/webapps/home/) is available for users to virtually experiment with contracting/expanding orthogonal and hexagonal ablation grids for generic 3D super-Gaussian laser crater profiles, allowing for exploration of the resulting post-ablation surface layer roughness and depth.

Mechanistic Study of Fast Performance Decay of PtCu Alloy-based Catalyst Layers for Polymer Electrolyte Fuel Cells through Electrochemical Impedance Spectroscopy.

In the past, platinum-copper catalysts have proven to be highly active for the oxygen reduction reaction (ORR), but transferring the high activities measured in thin-film rotating disk electrodes (TF-RDEs) to high-performing membrane electrode assemblies (MEAs) has proven difficult due to stability issues during operation. High initial performance can be achieved. However, fast performance decay on a timescale of 24 h is induced by repeated voltage load steps with H2/air supplied. This performance decay is accelerated if high relative humidity (>60% RH) is set for a prolonged time and low voltages are applied during polarization. The reasons and possible solutions for this issue have been investigated by means of electrochemical impedance spectroscopy and distribution of relaxation time analysis (EIS-DRT). The affected electrochemical sub-processes have been identified by comparing the PtCu electrocatalyst with commercial Pt/C benchmark materials in homemade catalyst-coated membranes (CCMs). The proton transport resistance (Rpt) increased by a factor of ~2 compared to the benchmark materials. These results provide important insight into the challenges encountered with the de-alloyed PtCu/KB electrocatalyst during cell break-in and operation. This provides a basis for improvements in the catalysts' design and break-in procedures for the highly attractive PtCu/KB catalyst system.

Semiquantitative Analysis for High-Speed Mapping Applications of Biological Samples Using LA-ICP-TOFMS.

Laser ablation (LA) in combination with inductively coupled plasma time-of-flight mass spectrometry (ICP-TOFMS) enables monitoring of elements from the entire mass range for every pixel, regardless of the isotopes of interest for a certain application. This provides nontargeted multi-element (bio-)imaging capabilities and the unique possibility to screen for elements that were initially not expected in the sample. Quantification of a large range of elements is limited as the preparation of highly multiplexed calibration standards for bioimaging applications by LA-ICP-(TOF)MS is challenging. In this study, we have developed a workflow for semiquantitative analysis by LA-ICP-TOFMS based on multi-element gelatin micro-droplet standards. The presented approach is intended for the mapping of biological samples due to the requirement of matrix-matched standards for accurate quantification in LA-ICPMS, a prerequisite that is given by the use of gelatin-based standards. A library of response factors was constructed based on 72 elements for the semiquantitative calculations. The presented method was evaluated in two stages: (i) on gelatin samples with known elemental concentrations and (ii) on real-world samples that included prime examples of bioimaging (mouse spleen and tumor tissue). The developed semiquantification approach was based on 10 elements as calibration standards and provided the determination of 136 nuclides of 63 elements, with errors below 25%, and for half of the nuclides, below 10%. A web application for quantification and semiquantification of LA-ICP(-TOF)MS data was developed, and a detailed description is presented to easily allow others to use the presented method.

Nanotubular TiO x N y -Supported Ir Single Atoms and Clusters as Thin-Film Electrocatalysts for Oxygen Evolution in Acid Media.

A versatile approach to the production of cluster- and single atom-based thin-film electrode composites is presented. The developed TiO x N y -Ir catalyst was prepared from sputtered Ti-Ir alloy constituted of 0.8 ± 0.2 at % Ir in α-Ti solid solution. The Ti-Ir solid solution on the Ti metal foil substrate was anodically oxidized to form amorphous TiO2-Ir and later subjected to heat treatment in air and in ammonia to prepare the final catalyst. Detailed morphological, structural, compositional, and electrochemical characterization revealed a nanoporous film with Ir single atoms and clusters that are present throughout the entire film thickness and concentrated at the Ti/TiO x N y -Ir interface as a result of the anodic oxidation mechanism. The developed TiO x N y -Ir catalyst exhibits very high oxygen evolution reaction activity in 0.1 M HClO4, reaching 1460 A g-1 Ir at 1.6 V vs reference hydrogen electrode. The new preparation concept of single atom- and cluster-based thin-film catalysts has wide potential applications in electrocatalysis and beyond. In the present paper, a detailed description of the new and unique method and a high-performance thin film catalyst are provided along with directions for the future development of high-performance cluster and single-atom catalysts prepared from solid solutions.

Alternative and facile production pathway towards obtaining high surface area PtCo/C intermetallic catalysts for improved PEM fuel cell performance.

The design of catalysts with stable and finely dispersed platinum or platinum alloy nanoparticles on the carbon support is key in controlling the performance of proton exchange membrane (PEM) fuel cells. In the present work, an intermetallic PtCo/C catalyst is synthesized via double-passivation galvanic displacement. TEM and XRD confirm a significantly narrowed particle size distribution for the catalyst particles compared to commercial benchmark catalysts (Umicore PtCo/C). Only about 10% of the mass fraction of PtCo particles show a diameter larger than 8 nm, whereas this is up to or even more than 35% for the reference systems. This directly results in a considerable increase in electrochemically active surface area (96 m2 g-1 vs. >70 m2 g-1), which confirms the more efficient usage of precious catalyst metal in the novel catalyst. Single-cell tests validate this finding by improved PEM fuel cell performance. Reducing the cathode catalyst loading from 0.4 mg cm-2 to 0.25 mg cm-2 resulted in a power density drop at an application-relevant 0.7 V of only 4% for the novel catalyst, compared to the 10% and 20% for the commercial benchmarks reference catalysts.

Direct quantification of PAHs and nitro-PAHs in atmospheric PM by thermal desorption gas chromatography with electron ionization mass spectroscopic detection.

In this work, we developed and optimized a method for the analysis of PAHs and nitro-PAHs in atmospheric particulate matter (PM) samples by using thermal desorption gas chromatography coupled with electron ionization single quadrupole mass spectrometry (TD-GC-(EI)-MS). The method uses thermal desorption from a PM on a filter sample as means of sample introduction to a column and obviates the need for complex extraction procedures, which are time-consuming and require environmentally unfriendly solvents. Moreover, the possibility of systematic errors is minimized and a significantly smaller amount of sample is required compared to traditional techniques requiring a pre-extraction step (approx. 10-times). Thirteen PAHs and three nitro-PAHs were used during method development. Although Tenax cartridges are typically used to capture volatile pollutants from the air, we found that glass-wool liner is the most suitable trap for the examined analytes after desorption from a quartz filter. Among the various instrument parameters which were tested and optimized, TD desorption flow and hold time, and temperature of the cooled injection system (CIS) proved to be most critical. We also found out that the matrix effect is especially pronounced in the case of high PM loadings, which should be kept in mind when planning the analysis. After the optimization, standard reference materials (ERM-CZ100 and NIST 1648a) were used for partial method validation and finally, real PM10 and PM2.5 samples from two Slovenian cities were successfully analyzed.

Characterization of fresh PM deposits on calcareous stone surfaces: Seasonality, source apportionment and soiling potential.

Particulate matter (PM) pollution is one of the major threats to cultural heritage outdoors. It has been recently implied that organic aerosols will prevail over inorganic carbon particulates in the future, changing the main mechanisms of damage caused by poor air quality to calcareous heritage in particular. We studied fresh particulate deposits on marble and limestone surfaces exposed to urban air in sheltered and unsheltered configurations. Due to different air pollution sources in different seasons, the amount and composition of surface deposits varied throughout the year. The main and most constant contributor to PM2.5 (particles smaller than 2.5 µm) were primary traffic emissions (30 %), followed by secondary formation of acidic inorganic aerosols, such as sulphate in summer and nitrate in winter (33 % altogether), and seasonal biomass-burning emissions (14 %). Although biomass burning is the major source of primary organic aerosols including the light-absorbing fraction that prevailed over black carbon (BC) in colder months (up to 60 % carbonaceous aerosol mass), we show that surface darkening causing the soiling effect is still governed by the minor BC fraction of atmospheric aerosols, which remained below 20 % of the carbonaceous aerosol mass throughout the year. This, however, can change in remote environments affected by biomass-burning emissions, such as winter resorts, or by rigorous BC mitigation measures in the future. In the short run, sheltered positions were less affected by different removal processes, but we show that surface deposits are not simply additive when considering longer periods of time. This must be taken into account when extrapolating surface accumulation to longer time scales.

Controlled growth of ZnO nanoparticles using ethanolic root extract of Japanese knotweed: photocatalytic and antimicrobial properties.

Synthesis of zinc oxide (ZnO) nanoparticles (NPs) was mediated by plant extracts to assist in the reduction of zinc atoms during the synthesis and act as a capping agent during annealing. The preparation used ethanolic extracts from the roots of Japanese knotweed (Fallopia japonica). Two major outcomes could be made. (i) A synergistic effect of multiple polyphenolic components in the extract is needed to achieve the capping effect of crystallite growth during thermal annealing at 450 °C characterized by an exponential growth factor (n) of 4.4 compared to n = 3 for bare ZnO. (ii) Synergism between the ZnO NPs and plant extracts resulted in superior antimicrobial activity against both Gram-positive bacteria, e.g., Staphylococcus aureus, and Gram-negative bacteria, e.g., Escherichia coli and Campylobacter jejuni. The materials were also tested for their antimicrobial activity against S. aureus under ultraviolet (UV) illumination. Also here, the photocatalyst prepared with plant extracts was found to be superior. The residues of the plant extract molecules on the surface of the catalyst were identified as the main cause of the observed differences, as proved by thermal gravimetry. Such a preparation using ethanolic extract of Fallopia japonica could serve as a more controlled synthesis of ZnO and potentially other metal oxides, with low environmental impact and high abundance in nature.

Quantification anomalies in single pulse LA-ICP-MS analysis associated with laser fluence and beam size.

Laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) has undergone major improvements in recent years which have led to reduction of the analysis time, higher spatial resolution, and better sensitivity. However, quantification and accurate analysis remain one of the bottlenecks in LA-ICP-MS analysis and so far satisfactory calibration solutions are restricted to well-documented matrices and suitable internal standards. Additional uncertainties associated with laser fluence and beam size via various ablation cells and interfaces make quantification even more challenging. This work is focused on the influence of fluence, beam size and aerosol transport on quantification in single pulse LA-ICP-MS analysis via approaches based on pulse intensity, LA spot volumes, noise characteristics, etc. for different elements (As, Gd, La, Ni, Te and Zn), concentrations (between 10 and 1000 µg g-1), and matrices (gelatin standards and NIST SRM 612). The findings indicate that selection of the appropriate laser fluence, just above the ablation threshold, and beam size, depending on the interface of LA and ICP-MS, are critical for reliable quantification and should be properly adjusted to avoid excessive Poisson and Flicker noise, achieve maximum sensitivity, and prevent the formation of double peaks in single pulses.

Importance of Chemical Activation and the Effect of Low Operation Voltage on the Performance of Pt-Alloy Fuel Cell Electrocatalysts.

Pt-alloy (Pt-M) nanoparticles (NPs) with less-expensive 3d transition metals (M = Ni, Cu, Co) supported on high-surface-area carbon supports are currently the state-of-the-art (SoA) solution to reach the production phase in proton exchange membrane fuel cells (PEMFCs). However, while Pt-M electrocatalysts show promise in terms of increased activity for oxygen reduction reaction (ORR) and, thus, cost reductions from the significantly lower use of expensive and rare Pt, key challenges in terms of synthesis, activation, and stability remain to unlock their true potential. This work systematically tackles them with a combination of electrocatalyst synthesis and characterization methodologies including thin-film rotating disc electrodes (TF-RDEs), an electrochemical flow cell linked to an inductively coupled plasma mass spectrometer (EFC-ICP-MS), and testing in 50 cm2 membrane electrode assemblies (MEAs). In the first part of the present work, we highlight the crucial importance of the chemical activation (dealloying) step on the performance of Pt-M electrocatalysts in the MEA at high current densities (HCDs). In addition, we provide the scientific community with a preliminary and facile method of distinguishing between a "poorly" and "adequately" dealloyed (activated) Pt-alloy electrocatalyst using a much simpler and affordable TF-RDE methodology using the well-known CO-stripping process. Since the transition-metal cations can also be introduced in a PEMFC due to the degradation of the Pt-M NPs, the second part of the work focuses on presenting clear evidence on the direct impact of the lower voltage limit (LVL) on the stability of Pt-M electrocatalysts. The data suggests that in addition to intrinsic improvements in stability, significant improvements in the PEMFC lifetime can also be obtained via the correct MEA design and applied limits of operation, namely, restricting not just the upper but equally important also the lower operation voltage.

Graphene-Derived Carbon Support Boosts Proton Exchange Membrane Fuel Cell Catalyst Stability.

The lack of efficient and durable proton exchange membrane fuel cell electrocatalysts for the oxygen reduction reaction is still restraining the present hydrogen technology. Graphene-based carbon materials have emerged as a potential solution to replace the existing carbon black (CB) supports; however, their potential was never fully exploited as a commercial solution because of their more demanding properties. Here, a unique and industrially scalable synthesis of platinum-based electrocatalysts on graphene derivative (GD) supports is presented. With an innovative approach, highly homogeneous as well as high metal loaded platinum-alloy (up to 60 wt %) intermetallic catalysts on GDs are achieved. Accelerated degradation tests show enhanced durability when compared to the CB-supported analogues including the commercial benchmark. Additionally, in combination with X-ray photoelectron spectroscopy Auger characterization and Raman spectroscopy, a clear connection between the sp 2 content and structural defects in carbon materials with the catalyst durability is observed. Advanced gas diffusion electrode results show that the GD-supported catalysts exhibit excellent mass activities and possess the properties necessary to reach high currents if utilized correctly. We show record-high peak power densities in comparison to the prior best literature on platinum-based GD-supported materials which is promising information for future application.

Quantification in bioimaging by LA-ICPMS - Evaluation of isotope dilution and standard addition enabled by micro-droplets.

This study explores quantitative bioimaging as enabled by laser ablation-inductively coupled plasma time-of-flight mass spectrometry (LA-ICP-TOFMS), designing standardization methods based on robotic micro-droplet dispensing. The potential of producing controlled and highly precise pL-volume droplets was exploited to establish on-tissue isotope dilution and standard addition. Both strategies eliminate matrix effects and offer high metrological order traceable to SI units. The absolute quantity was obtained for µm-sized regions of interest in tissue samples, as defined by the extension of the deposited pL-volume droplet. While the gold standard isotope dilution (ID) was restricted to the accurate quantification of a single element, i.d. platinum in different tissue samples (mouse liver, spleen and tumor tissue), multiplexed matrix-matched calibration was obtained by on-tissue standard addition by depositing a dilution series of certified multi-element standards. Here, the working range was determined by the heterogeneity of the tissue samples and the background levels of elements intrinsically present and/or artificially introduced during sample preparation. Both methods, ID and standard addition served as reference methods for validation of external calibration using gelatin-based micro-droplet standards. Given full ablation, these external standards revealed a high dynamic range together with an excellent repeatability. Where applicable, the cross-validation revealed consistent quantitative results for the three quantification approaches. The comparable sensitivity obtained for standard addition and external standardization, respectively expressed as slope of the calibration function, provided proof that gelatin-based micro-droplets could serve as matrix-matched calibrations. Therefore, gelatin micro-droplets offer a valid tool for multiplexed matrix-mimicking standardization at high-throughput.