Low Mass Species Research Articles

Degradation strategies for structural characterization of insoluble synthetic polymers by mass spectrometry.

With the advent of soft ionization techniques such as electrospray (ESI) and matrix-assisted laser desorption/ionization (MALDI) to produce intact gas-phase ions from nonvolatile macromolecules, mass spectrometry has become an essential technique in the field of polymeric materials. However, (co)polymers of very high molecular weight or with reticulated architectures still escape ESI or MALDI, mainly due to solubility issues. Strategies developed to tackle such an analytical challenge all rely on sample degradation to produce low-mass species amenable to existing ionization methods. Yet, chain degradation needs to be partial and controlled to generate sufficiently large species that still contain topological or architectural information. The present article reviews the different analytical degradation strategies implemented to perform mass spectrometry of these challenging synthetic polymers, covering thermal degradation approaches in sources developed in the 2000s, off-line sample pre-treatments for controlled chemical degradation of polymeric substrates, and most recent achievements employing reactive ionization modes to perform chemolysis on-line with MS.

The environmental degradability of DEMNUM, a typical PFPE polymer

The three environmental degradation tests of hydrolysis, indirect photolysis and Zahn-Wellens microbial degradation were conducted according to the OECD and the US EPA guidelines on DEMNUM, a typical linear perfluoropolyether polymer. Low mass degradation products that formed in each test were structurally characterized and indirectly quantified by liquid chromatography mass spectrometry (LC/MS) using a reference compound and an internal standard of similar structure. The degradation of the polymer was assumed to directly correlate with the appearance of lower mass species. The hydrolysis experiment at 50 °C showed the appearance of less than a dozen low mass species with increasing pH but at the negligible total estimated amount of ∼2 ppm relative to polymer. A dozen low mass perfluoro acid entities also appeared following the indirect photolysis experiment in synthetic humic water. Their maximum total amount was at ∼150 ppm relative to polymer. The largest total amount of low mass species formed during the Zahn-Wellens biodegradation test amounted to only ∼80 ppm relative to polymer. The Zahn-Wellens conditions tended to produce larger low mass molecules than the ones formed under photolysis. The results from all three tests indicate that the polymer is stable and non-degradable in the environment.

Photocatalytic degradation of organic pollutants by Ag2O/AgSiOx

Silver silicate (AgSiOx) is a visible-driven photocatalyst. However, the efficiency of single component photocatalysts can be improved by coupling with other compounds. In this work, AgSiOx was coupled with different amounts of Ag2O using a co-precipitation method. Ratios of Ag2O:AgSiOx were controlled by varying the concentrations of Na2SiO3.9H2O and NaOH. All prepared products were characterized by X-ray diffractometry, transmission electron microscopy, X-ray photoelectron spectroscopy, surface analysis, and UV–vis diffuse reflectance spectroscopy. Under visible light irradiation, the Ag2O/AgSiOx composite could degrade rhodamine B, reactive orange, and bisphenol A solutions. The Ag2O/AgSiOx exhibited greater photocatalytic activity than AgSiOx and Ag2O because this composite improved visible light absorption and inhibited charge recombination. The Ag2O/AgSiOx prepared from 0.012 mol NaOH exhibited the highest photocatalytic activity. Photocatalytic degradation of all pollutant models to low mass species was studied by electrospray ionization mass spectrometry and the cytotoxicity of pollutant model solutions before and after photocatalytic processes was evaluated on L929 mouse fibroblast cells using the MTT assay.

Body mass and geographic distribution determined the evolution of the wing flight-feather molt strategy in the Neornithes lineage

The evolutionary history of many organisms is characterized by major changes in morphology and distribution. Specifically, alterations of body mass and geographic distribution may profoundly influence organismal life-history traits. Here, we reconstructed the evolutionary history of flight-feather molt strategy using data from 1,808 Neornithes species. Our analysis suggests that the ancestral molt strategy of first-year birds was partial or entirely absent, and that complete wing flight-feather molt in first-year birds first evolved in the late Eocene and Oligocene (25–40 Ma), at least 30 Myr after birds first evolved. Complete flight-feather molt occurred mainly at equatorial latitudes and in relatively low body mass species, following a diversification of body mass within the lineage. We conclude that both body mass and geographic distribution shaped the evolution of molt strategies and propose that the evolutionary transition towards complete juvenile molt in the Neornithes is a novel, relatively late adaptation.

Low-energy electron ionization mass spectrometer for efficient detection of low mass species

The design of a high-efficiency mass spectrometer is described, aimed at residual gas detection of low mass species using low-energy electron impact, with particular applications in helium atom microscopy and atomic or molecular scattering. The instrument consists of an extended ionization volume, where electrons emitted from a hot filament are confined using a solenoidal magnetic field to give a high ionization probability. Electron space charge is used to confine and extract the gas ions formed, which are then passed through a magnetic sector mass filter before reaching an ion counter. The design and implementation of each of the major components are described in turn, followed by the overall performance of the detector in terms of mass separation, detection efficiency, time response, and background count rates. The linearity of response with emission current and magnetic field is discussed. The detection efficiency for helium is very high, reaching as much as 0.5%, with a time constant of (198 ± 6) ms and a background signal equivalent to an incoming helium flux of (8.7 ± 0.2) × 106 s-1.

A quantitative light-isotope measurement system for climate and energy applications

We describe the design, operation, and performance of a new instrumental configuration capable of quantitative determinations with sub-picomole accuracy of dilute concentrations of low mass species, such as He4, He3, Ne20, and Ar40, in a balance of stable hydrogen (H2, DH, and D2) gas. This inexpensive system may realize important applications in fields ranging from climate studies to hydrogen fusion energy research, thereby providing an expanded availability of this diagnostic within emerging energy systems research and development. These spectra, calibration curves, and determinations were obtained by using a novel method for the purification and subsequent removal of the hydrogen matrix gas, and an extensively modified commercial Fourier Transform Ion Cyclotron Resonance (FT-ICR) mass spectrometer with an electron impact (EI) ionizer. These high-resolution FT-ICR mass spectrometers have routinely achieved a resolution, R = m/Δm better than 10,000 Da at mass-3, with a mass resolution that scales as 1/m. These devices have easily resolved D2 from He4, and DH from He3. The performance of this upgraded instrument has demonstrated the ability to detect impurities from tiny air leaks, such as Ar40 and Ne20, in the presence of the hydrogen matrix gas. While no concentration measurements of radioactive species have been attempted to date with this system, it is expected to easily resolve DT from D2H (a 0.0059 Da mass difference) and HT from all other mass-4 species.

Rayleigh-Brillouin scattering in binary mixtures of disparate-mass constituents: SF_{6}-He,SF_{6}-D_{2}, and SF_{6}-H_{2}.

The spectral distribution of light scattered by microscopic thermal fluctuations in binary mixture gases was investigated experimentally and theoretically. Measurements of Rayleigh-Brillouin spectral profiles were performed at a wavelength of 532nm and at room temperature, for mixtures of SF_{6}-He,SF_{6}-D_{2}, and SF_{6}-H_{2}. In these measurements, the pressure of the gases with heavy molecular mass (SF_{6}) is set at 1bar, while the pressure of the lighter collision partner was varied. In view of the large polarizability of SF_{6} and the very small polarizabilities of He, H_{2}, and D_{2}, under the chosen pressure conditions these low mass species act as spectators and do not contribute to the light scattering spectrum, while they influence the motion and relaxation of the heavy SF_{6} molecules. A generalized hydrodynamic model was developed that should be applicable for the particular case of molecules with heavy and light disparate masses, as is the case for the heavy SF_{6} molecule, and the lighter collision partners. Based on the kinetic theory of gases, our model replaces the classical Navier-Stokes-Fourier relations with constitutive equations having an exponential memory kernel. The energy exchange between translational and internal modes of motion is included and quantified with a single parameter z that characterizes the ratio between the mean elastic and inelastic molecular collision frequencies. The model is compared with the experimental Rayleigh-Brillouin scattering data, where the value of the parameter z is determined in a least-squares procedure. Where very good agreement is found between experiment and the generalized hydrodynamic model, the computations in the framework of classical hydrodynamics strongly deviate. Only in the hydrodynamic regime both models are shown to converge.

Addressing contamination in ECR charge breeders

The Electron Cyclotron Resonance (ECR) ion source was first utilized for charge breeding in 1995 [1]. Since that time the charge breeding technique has been refined. Single charge state efficiency has improved by a factor of ten, the efficiency discrepancy between solid and gaseous species has narrowed, and low-mass species efficiency has improved. But the limiting characteristic of the ECR charge breeder continues to be a high level of contamination which often obscures the beam of interest [2]. Multiple techniques have been employed to reduce this contamination with varying levels of success, and attempts are currently underway to improve upon the successes achieved to date. This paper will review those past techniques, current attempts, and possible future paths for reducing the contamination level in ECR charge breeders.

Numerical study of the influence of surface reaction probabilities on reactive species in an rf atmospheric pressure plasma containing humidity

The quantification and control of reactive species (RS) in atmospheric pressure plasmas (APPs) is of great interest for their technological applications, in particular in biomedicine. Of key importance in simulating the densities of these species are fundamental data on their production and destruction. In particular, data concerning particle-surface reaction probabilities in APPs are scarce, with most of these probabilities measured in low-pressure systems. In this work, the role of surface reaction probabilities, γ, of reactive neutral species (H, O and OH) on neutral particle densities in a He–H2O radio-frequency micro APP jet (COST-APPJ) are investigated using a global model. It is found that the choice of γ, particularly for low-mass species having large diffusivities, such as H, can change computed species densities significantly. The importance of γ even at elevated pressures offers potential for tailoring the RS composition of atmospheric pressure microplasmas by choosing different wall materials or plasma geometries.

Enhanced capabilities for imaging gangliosides in murine brain with matrix-assisted laser desorption/ionization and desorption electrospray ionization mass spectrometry coupled to ion mobility separation

The increased interest in lipidomics calls for improved yet simplified methods of lipid analysis. Over the past two decades, mass spectrometry imaging (MSI) has been established as a powerful technique for the analysis of molecular distribution of a variety of compounds across tissue surfaces. Matrix-assisted laser desorption/ionization (MALDI) MSI is widely used to study the spatial distribution of common lipids. However, a thorough sample preparation and necessity of vacuum for efficient ionization might hamper its use for high-throughput lipid analysis. Desorption electrospray ionization (DESI) is a relatively young MS technique. In DESI, ionization of molecules occurs under ambient conditions, which alleviates sample preparation. Moreover, DESI does not require the application of an external matrix, making the detection of low mass species more feasible due to the lack of chemical matrix background. However, irrespective of the ionization method, the final information obtained during an MSI experiment is very complex and its analysis becomes challenging. It was shown that coupling MSI to ion mobility separation (IMS) simplifies imaging data interpretation. Here we employed DESI and MALDI MSI for a lipidomic analysis of the murine brain using the same IMS-enabled instrument. We report for the first time on the DESI IMS-MSI of multiply sialylated ganglioside species, as well as their acetylated versions, which we detected directly from the murine brain tissue. We show that poly-sialylated gangliosides can be imaged as multiply charged ions using DESI, while they are clearly separated from the rest of the lipid classes based on their charge state using ion mobility. This represents a major improvement in MSI of intact fragile lipid species. We additionally show that complementary lipid information is reached under particular conditions when DESI is compared to MALDI MSI.

Ultrahigh pressure fast size exclusion chromatography for top-down proteomics

Top-down MS-based proteomics has gained a solid growth over the past few years but still faces significant challenges in the LC separation of intact proteins. In top-down proteomics, it is essential to separate the high mass proteins from the low mass species due to the exponential decay in S/N as a function of increasing molecular mass. SEC is a favored LC method for size-based separation of proteins but suffers from notoriously low resolution and detrimental dilution. Herein, we reported the use of ultrahigh pressure (UHP) SEC for rapid and high-resolution separation of intact proteins for top-down proteomics. Fast separation of intact proteins (6-669 kDa) was achieved in < 7 min with high resolution and high efficiency. More importantly, we have shown that this UHP-SEC provides high-resolution separation of intact proteins using a MS-friendly volatile solvent system, allowing the direct top-down MS analysis of SEC-eluted proteins without an additional desalting step. Taken together, we have demonstrated that UHP-SEC is an attractive LC strategy for the size separation of proteins with great potential for top-down proteomics.

Improved charge breeding efficiency of light ions with an electron cyclotron resonance ion source

The Californium Rare Isotope Breeder Upgrade is a new radioactive beam facility for the Argonne Tandem Linac Accelerator System (ATLAS). The facility utilizes a (252)Cf fission source coupled with an electron cyclotron resonance ion source to provide radioactive beam species for the ATLAS experimental program. The californium fission fragment distribution provides nuclei in the mid-mass range which are difficult to extract from production targets using the isotope separation on line technique and are not well populated by low-energy fission of uranium. To date the charge breeding program has focused on optimizing these mid-mass beams, achieving high charge breeding efficiencies of both gaseous and solid species including 14.7% for the radioactive species (143)Ba(27+). In an effort to better understand the charge breeding mechanism, we have recently focused on the low-mass species sodium and potassium which up to present have been difficult to charge breed efficiently. Unprecedented charge breeding efficiencies of 10.1% for (23)Na(7+) and 17.9% for (39)K(10+) were obtained injecting stable Na(+) and K(+) beams from a surface ionization source.

Modulation of in vitro activity of zymogenic and mature recombinant human β‐secretase by dietary plants

The in vitro activity of human recombinant β-secretase (BACE1) was studied using a fluorogenic substrate based on the cleavage site for the enzyme in the Swedish mutation of amyloid precursor protein. The enzyme was inhibited by a control peptide inhibitor with good repeatability. The enzyme preparation comprised a mixture of pro-enzyme or zymogen and mature enzyme whereby the pro-enzyme sequence forms a 'flap' that can obstruct the binding site. 'Open flap' forms of the zymogen and mature enzyme are active, but the 'closed flap' form of the zymogen is inactive. This mixture of enzyme populations permitted apparent stimulation of enzyme activity under particular conditions, presumably due to facilitating flap-opening of the zymogen. As reported for heparin, enzyme activation was stimulated in the presence of low concentrations of Tween 20 and dimethylsulfoxide before becoming inhibited at higher concentrations. Dietary plant extracts either consistently inhibited (e.g. clove, tea, cinammon) or consistently stimulated (e.g. mushroom, parsley, asparagus) BACE1. Common structural features identified by Fourier transform infrared spectroscopy revealed that BACE1 activity could be explained by differential interactions of either small molecule or polymeric species with mature versus zymogen forms of the enzyme, respectively. Further, enzyme activity could be reversed by mixtures of high and low mass species. These results may have implications for the regulation of β-secretase activity in vivo by either endogenous or possibly dietary factors and for a potential role of BACE1 in stimulation of the production of amyloid beta peptide in sporadic Alzheimer's disease.

MALDI-TOF MS stability study of model poly(p-phenylene terephthalamide)s

In the present study, we address the possibility of matrix-assisted laser desorption/ionization (MALDI)-time-of-flight MS analysis-induced chain fragmentation in poly(p-phenylene terephthalamide) (PPD-T) by considering two possible sources: (1) grinding-induced fragmentation resulting from the evaporation-grinding MALDI sample preparation method (E-G method) and (2) in-source/metastable fragmentation induced by the MALDI laser. An analysis of variance (ANOVA) statistical study found, with a high probability, that obtaining MALDI spectra with the effective laser area as large as possible (the "fanned-out" setting) did not cause any chain fragmentation due to the E-G MALDI sample preparation method, even when three additional grinding steps were used. However, the effect of laser fluence was less clear. A significant effect of laser fluence was observed for lower mass oligomers (<1,400 Da), but there was essentially no effect for higher mass species up to our limit of ANOVA measurement (approximately 2,300 Da). Plausible explanations are presented to explain these observations. The most likely scenario is that "unexpected" end-group modifications occur during PPD-T synthesis, producing small quantities of low mass species, which are amplified by the MALDI-EG extraction procedure.

Metabolite Imaging Using Matrix-Enhanced Surface-Assisted Laser Desorption/Ionization Mass Spectrometry (ME-SALDI-MS)

We describe here the use of a hybrid ionization approach, matrix-enhanced surface-assisted laser desorption/ionization mass spectrometry (ME-SALDI-MS) in bioimaging. ME-SALDI combines the strengths of traditional matrix-assisted laser desorption/ionization (MALDI) and SALDI and enables successful MS imaging of low-mass species with improved detection sensitivity. Using 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) as the MS standard, MS performances of MALDI, SALDI, and ME-SALDI are systematically compared. The analyte desorption and ionization mechanism in ME-SALDI is qualitatively speculated based on the observation of significantly reduced matrix background and improved survival yields of molecular ions. Improvements in detection sensitivity of low-mass species using ME-SALDI over MALDI in imaging are demonstrated with mouse heart and brain tissues.

Imaging mass spectrometry of proteins and peptides: 3D volume reconstruction

As large genomic and proteomic datasets are generated from homogenates of various tissues, the need for information on the spatial localization of their encoded products has become more pressing. Matrix-assisted laser desorption-ionization (MALDI) imaging mass spectrometry (IMS) offers investigators the means with which to unambiguously study peptides and proteins with molecular specificity, and to determine their distribution in two and three dimensions. In the past few years, several parameters have been optimized for IMS, including sample preparation, matrix application and instrumental acquisition parameters (Box 1). These developments have resulted in a high degree of reproducibility in mass accuracy and peak intensities (Supplementary Fig. 1 online). Recently, we have optimized our protocol to be able to increase the number of molecular species analyzed by collecting two sets of sections, covering one set of sections with sinapinic acid for optimal detection of proteins and adjacent sections with 2,5-dihydroxybenzoic acid (DHB) matrix for the optimal detection of low-mass species, including peptides. Approximately 1,000 peaks can be observed in each dataset (Fig. 1). Furthermore, the sections are collected at an equal distance, 200 mum instead of 400-500 mum used previously, thus enabling the use of virtual z-stacks and three-dimensional (3D) volume renderings to investigate differential localization patterns in much smaller brain structures such as the substantia nigra and the interpeduncular nucleus. Here we present our optimized step-by-step procedure based on previous work in our laboratory, describing how to make 3D volume reconstructions of MALDI IMS data, as applied to the rat brain.

Denaturant sensitive regions in creatine kinase identified by hydrogen/deuterium exchange

The GdmHCl-induced unfolding of creatine kinase (CK) has been studied by hydrogen/deuterium (H/D) exchange combined with mass spectrometry. MM-CK unfolded for various periods in different denaturant concentrations was pulsed-labeled with deuterium to identify different conformational intermediate states. For all denaturation times or GdmHCl concentrations, we observed variable proportions of only two species. The low-mass envelope of isotope peaks corresponds to a species that has gained about 10 deuteriums more than native CK, and the high-mass envelope to a completely deuterated species. To localize precisely the unfolded regions in the states highly populated during denaturation, the protein was digested with two proteases (pepsin and type XIII protease) after H/D exchange and rapid quenching of the reaction. The two sets of fragments obtained were analyzed by liquid chromatography coupled to mass spectrometry to determine the deuterium level in each fragment. Bimodal distributions of deuterium were found for most peptides, indicating that these regions were either folded or unfolded. This behavior is consistent with cooperative, localized unfolding. However, we observed a monomodal distribution of deuterium in two regions (1-12 and 162-186). We conclude that the increment of mass observed in the low-mass species of the intact protein (+10 Da) has its origin in these two segments. These regions, which are very sensitive to low GdmHCl concentrations, are involved in the monomer-monomer interface of CK and their perturbation is likely to weaken the dimeric structure. At higher denaturant concentration, this would induce dissociation of the dimer.

Analytical characterization of bioactive metal species in the cellular domain (metallomics) to simplify environmental and biological proteomics

The speciation approach for the characterization of low-mass metal species including sample treatment and storage is at present a well-established topic based on chromatography-atomic detector coupling. More recently, new endogenous and exogenous metal species from biological systems are attracting considerable interest. Bioactive molecules such as proteins, DNA restriction fragments, phytochelatins, metallothioneins and others are target species of a new generation of analytical tools (bioanalysis) which substitute the traditional atomic detectors based on the use of photons (AAS, FPD, ICP-AES, AFS) by mass detectors (MS and ICP-MS) for ion characterization. Several cases related to biological molecules involving proteins and multiprotein systems, in which frequently metals (metallomics) participate, are described, and a generic metallomic analytical approach is proposed for the identification and quantification of metalloproteins, and other metal molecules present in living systems. In this work, a multiplexed analytical approach (MAA) is described on the basis of three experimental components: (1) a separation technique—selectivity component; (2) a highly sensitive elemental detector—sensitivity component; and (3) a molecule-specific detector, generally based on mass spectrometry—structural component. This approach brings together both elemental and molecular detectors to simplify the identification of metal-tagged biomolecules in environmental, food, and health studies.

Reductive Assays forS-Nitrosothiols: Implications for Measurements in Biological Systems

Bioactive SNOs are found in many tissues. We speculated SNOs might be misidentified in conventional assays which reduce NO−3to NO.S-Nitrosothiols were exposed to saturated VCl3in HCl, 1% KI in acetic acid, photolysis, or CuCl and CSH in He; NO was measured by chemiluminescence.S-Nitrosothiols were readily detected in VCl3but not in KI. Reduction in CuCl/cysteine was linear (r2= 1.0,n= 6), sensitive to 10 pmol, and eliminated by HgCl2; it did not detect NO−2, NO−3, or 3-nitrotyrosine.S-Nitrosothiols represented ∼2.9% of NOxassayed by VCl3in human serum, of which <5% were low-mass species. In summary, (i) conventional assays may misidentify NO−3, but not NO−2, as SNOs; and (ii) chemiluminescence/reduction systems may be sensitive and specific as SNO assays. We suggest that assay of the SNO fraction in biological NOxmay be more relevant and feasible than is now appreciated.

A simulation of gas transport in polymeric membranes. Application to heterogeneous aging

A simplified mechanism of polymer aging has been proposed to simulate the effect of diffusion on degradation processes. Simple observations have been made on different events simulated using a modelization of the diffusion of low mass species: (1) the bulk oxidation rate, in the case of heterogeneous oxidation, is proportional to the square root of the initiation rate (formation of radicals). (2) The relative oxidation profile (relative to the surface oxidation) as a function of aging time is not constant. (3) The initial concentration of oxygen has, in some cases, a large influence on the apparent bulk oxidation rate during all the degradation processes. (4) The same observation is made for model samples stored in air and aged in inert atmosphere. (5) When heterogeneous oxidation occurs in samples containing antioxidant additives, the concentration of antioxidant has an effect on the induction period and on the bulk oxidation rate (this is not the case for homogeneous oxidation).(6) When heterogeneous oxidation occurs, the effect of antioxidant mobility can be practically neglected when D oxygen /D antioxidant (D = diffusion coefficient) varies from to 100 to 1.( (7) The time required to reach a pseudostationary state (corresponding to the egality of oxidation rate and oxygen diffusion rate) can be interpreted by a simple induction period (antioxidant consumption and/or hydroperoxide accumulation).