Fourier transform ion-cyclotron resonance mass spectrometry (FT-ICR MS) is the only mass analyzer that can resolve the molecular complexity of natural organic matter at the level of elemental ...composition assignment. Here, we leverage the high dynamic range, resolving power, resistance to peak coalescence, and maximum ion number and ion trapping duration in a custom built, 21 tesla hybrid linear ion trap /FT-ICR mass spectrometer for a dissolved organic matter standard (Suwanne River Fulvic Acid). We compare the effect of peak-picking threshold (3σ, 4σ, 5σ, and 6σ) on number of elemental composition assignments, mass measurement accuracy, and dynamic range for a 6.3 s transient across the mass range of m/z 200–1200 that comprises the highest achieved resolving power broadband FT-ICR mass spectrum collected to date. More than 36 000 species are assigned with signal magnitude greater than 3σ at root-mean-square mass error of 36 ppb, the most species identified reported to date for dissolved organic matter. We identify 18O and 17O isotopologues and resolve isobaric overlaps on the order of a few electrons across a wide mass range (up to m/z 1000) leveraging mass resolving powers (3 000 000 at m/z 200) only achievable by 21 T FT-ICR MS and increased by ∼30% through absorption mode data processing. Elemental compositions unique to the 3σ span a wide compositional range of aromaticity not detected at higher peak-picking thresholds. Furthermore, we leverage the high dynamic range at 21 T FT-ICR MS to provide a molecular catalogue of a widely utilized reference standard (SRFA) to the analytical community collected on the highest performing mass analyzer for complex mixture analysis to date. This instrument is available free of charge to scientists worldwide.
We describe the design and initial performance of the first 21 tesla Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometer. The 21 tesla magnet is the highest field superconducting ...magnet ever used for FT-ICR and features high spatial homogeneity, high temporal stability, and negligible liquid helium consumption. The instrument includes a commercial dual linear quadrupole trap front end that features high sensitivity, precise control of trapped ion number, and collisional and electron transfer dissociation. A third linear quadrupole trap offers high ion capacity and ejection efficiency, and rf quadrupole ion injection optics deliver ions to a novel dynamically harmonized ICR cell. Mass resolving power of 150,000 (
m/Δm
50%
) is achieved for bovine serum albumin (66 kDa) for a 0.38 s detection period, and greater than 2,000,000 resolving power is achieved for a 12 s detection period. Externally calibrated broadband mass measurement accuracy is typically less than 150 ppb rms, with resolving power greater than 300,000 at
m/z
400 for a 0.76 s detection period. Combined analysis of electron transfer and collisional dissociation spectra results in 68% sequence coverage for carbonic anhydrase. The instrument is part of the NSF High-Field FT-ICR User Facility and is available free of charge to qualified users.
Graphical Abstract
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As disinfection byproducts (DBPs) are ubiquitous sources of chemical exposure in disinfected drinking water, identifying unknown DBPs, especially unknown drivers of toxicity, is one of the major ...challenges in the safe supply of drinking water. While >700 low-molecular-weight DBPs have been identified, the molecular composition of high-molecular-weight DBPs remains poorly understood. Moreover, due to the absence of chemical standards for most DBPs, it is difficult to assess toxicity contributions for new DBPs identified. Based on effect-directed analysis, this study combined predictive cytotoxicity and quantitative genotoxicity analyses and Fourier transform ion cyclotron resonance mass spectrometry (21 T FT-ICR-MS) identification to resolve molecular weight fractions that induce toxicity in chloraminated and chlorinated drinking waters, along with the molecular composition of these DBP drivers. Fractionation using ultrafiltration membranes allowed the investigation of <1 kD, 1-3 kD, 3-5 kD, and >5 kD molecular weight fractions. Thiol reactivity based predictive cytotoxicity and single-cell gel electrophoresis based genotoxicity assays revealed that the <1 kD fraction for both chloraminated and chlorinated waters exhibited the highest levels of predictive cytotoxicity and direct genotoxicity. The <1 kD target fraction was used for subsequent molecular composition identification. Ultrahigh-resolution MS identified singly charged species (as evidenced by the 1 Da spacing in
C isotopologues), including 3599 chlorine-containing DBPs in the <1 kD fraction with the empirical formulas CHOCl, CHOCl
, and CHOCl
, with a relative abundance order of CHOCl > CHOCl
≫ CHOCl
. Interestingly, more high-molecular-weight CHOCl
DBPs were identified in the chloraminated vs chlorinated waters. This may be due to slower reactions of NH
Cl. Most of the DBPs formed in chloraminated waters were composed of high-molecular-weight Cl-DBPs (up to 1 kD) rather than known low-molecular-weight DBPs. Moreover, with the increase of chlorine number in the high-molecular-weight DBPs detected, the O/C ratio exhibited an increasing trend, while the modified aromaticity index (AI
) showed an opposite trend. In drinking water treatment processes, the removal of natural organic matter fractions with high O/C ratio and high AI
value should be strengthened to minimize the formation of known and unknown DBPs.
Asphaltenes challenge nearly all analytical techniques because of their immense polydispersity in molecular composition and structure. This operationally defined fraction of crude oil, insoluble in ...n-alkanes but soluble in aromatic solvents, is known to concentrate in vacuum residues and islinked to fouling and deposition issues. However, presence and subsequent characterization of asphaltenes are seldom discussed in conventional/unconventional distillate fractions. Here, we isolate asphaltenes from conventional (<593 °C/1099 F) and unconventional (>593 °C) distillation cuts and provide molecular-level characterization by electrospray ionization and atmospheric pressure photoionization Fourier transform ion cyclotron resonance mass spectrometry as a function of boiling point. Our results indicate that asphaltene molecular composition starts in the vacuum gas oil range and extends into vacuum residues. Moreover, we report that distillable asphaltene composition exists as both highly polar (heteroatom rich), aliphatic (atypical asphaltenes) species as well as condensed aromatic structures (classical asphaltenes). As a function of distillation temperature, asphaltene compounds exhibit structural trends consistent with thermal cracking that starts between 510 and 538 °C, increases between 538 and 593 °C, and is readily observed at temperatures up to 700 °C. Above 600 °C, low molecular weight compounds (expected to boil at much lower temperatures) that are n-heptane insoluble are detected across all heteroatom classes. Results herein suggest that these compounds are formed through structural rearrangement of archipelago asphaltenes because of thermal cracking reactions that occur during distillation and precipitate as asphaltenes. We report the isolation and mass spectral characterization of asphaltenes isolated from distillation cuts and propose that quantification of asphaltenes in distillates is critical to predict and prevent problems related to catalyst deactivation.
We apply two widely used extraction techniques, liquid/liquid extraction and solid-phase extraction with styrene–divinylbenzene polymer with a proprietary nonpolar surface priority pollutant (PPL) to ...water-soluble compounds generated through photodegradation and biodegradation of petroleum. We compare the molecular composition of bio- and photodegraded water-soluble organic (WSO) acids by 21 T negative-ion electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS). We highlight the compositional differences between the two extraction techniques for abiotic and biotic degradation processes and identify known toxic species (naphthenic acids) produced through hydrocarbon biodegradation identified by liquid/liquid extraction (LLE) that are not detected with solid-phase extraction (SPE) of the same sample. Photodegraded WSO compounds extracted by SPE-PPL correspond to species with higher O/C ratio and carbon number compared to LLE extracted compounds. Naphthenic acids, a recalcitrant class of nonaromatic carboxylic acids and known acute toxicants formed through biodegradation of oil, are detected in LLE extracts (up to C30 and double-bond equivalents, DBE < 3) but are not detected in SPE-PPL extracts. This suggests that LLE and SPE-PPL retain different water-soluble oil species based on the dominant type of oil weathering process.
In pathogenic Gram-negative bacteria, interactions among membrane proteins are key mediators of host cell attachment, invasion, pathogenesis, and antibiotic resistance. Membrane protein interactions ...are highly dependent upon local properties and environment, warranting direct measurements on native protein complex structures as they exist in cells. Here we apply in vivo chemical cross-linking mass spectrometry, to reveal the first large-scale protein interaction network in Pseudomonas aeruginosa, an opportunistic human pathogen, by covalently linking interacting protein partners, thereby fixing protein complexes in vivo. A total of 626 cross-linked peptide pairs, including previously unknown interactions of many membrane proteins, are reported. These pairs not only define the existence of these interactions in cells but also provide linkage constraints for complex structure predictions. Structures of three membrane proteins, namely, SecD-SecF, OprF, and OprI are predicted using in vivo cross-linked sites. These findings improve understanding of membrane protein interactions and structures in cells.
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•260 binary interactions of P. aeruginosa proteins were detected in live cells•In vivo cross-linking enabled identification of membrane protein interactions•Cross-link sites enabled complex structure predictions
Navare et al. apply in vivo chemical cross-linking mass spectrometry to reveal the first large-scale protein interaction network in Pseudomonas aeruginosa with over 600 cross-linked peptide pairs. Cross-linked sites provide constraints useful for complex structure prediction, even for membrane protein interactions.
Introduction of oil and gas extraction wastewaters (OGWs) to surface water leads to elevated halide levels from geogenic bromide and iodide, as well as enhanced formation of brominated and iodinated ...disinfection byproducts (DBPs) when treated. OGWs contain high levels of chemical additives used to optimize extraction activities, such as surfactants, which have the potential to serve as organic DBP precursors in OGW-impacted water sources. We report the first identification of olefin sulfonate surfactant-derived DBPs from laboratory-disinfected gas extraction wastewater. Over 300 sulfur-containing DBPs, with 43 unique molecular formulas, were found by high-resolution mass spectrometry, following bench-scale chlor(am)ination. DBPs consisted of mostly brominated species, including bromohydrin sulfonates, dihalo-bromosulfonates, and bromosultone sulfonates, with chlorinated/iodinated analogues formed to a lesser extent. Disinfection of a commercial C12-olefin sulfonate surfactant mixture revealed dodecene sulfonate as a likely precursor for most detected DBPs; disulfur-containing DBPs, like bromosultone sulfonate and bromohydrin disulfonate, originated from olefin disulfonate species, present as side-products of olefin sulfonate production. Disinfection of wastewaters increased mammalian cytotoxicity several orders of magnitude, with chloraminated water being more toxic. This finding is important to OGW-impacted source waters because drinking water plants with high-bromide source waters may switch to chloramination to meet DBP regulations.
The nosocomial pathogen Acinetobacter baumannii is a frequent cause of hospital-acquired infections worldwide and is a challenge for treatment due to its evolved resistance to antibiotics, including ...carbapenems. Here, to gain insight on A. baumannii antibiotic resistance mechanisms, we analyse the protein interaction network of a multidrug-resistant A. baumannii clinical strain (AB5075). Using in vivo chemical cross-linking and mass spectrometry, we identify 2,068 non-redundant cross-linked peptide pairs containing 245 intra- and 398 inter-molecular interactions. Outer membrane proteins OmpA and YiaD, and carbapenemase Oxa-23 are hubs of the identified interaction network. Eighteen novel interactors of Oxa-23 are identified. Interactions of Oxa-23 with outer membrane porins OmpA and CarO are verified with co-immunoprecipitation analysis. Furthermore, transposon mutagenesis of oxa-23 or interactors of Oxa-23 demonstrates changes in meropenem or imipenem sensitivity in strain AB5075. These results provide a view of porin-localized antibiotic inactivation and increase understanding of bacterial antibiotic resistance mechanisms.
Recent advances in instrumentation for high-field Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) have enabled access to ∼70 000 unique molecular formulas in broadband mass ...spectral characterization of unfractionated/whole asphaltenes. The results accumulated over a decade highlight the need for an asphaltene molecular model that acknowledges the coexistence of (1) monofunctional and polyfunctional species; (2) island and archipelago structural motifs; and (3) heteroatom-depleted/highly aromatic compounds, as well as atypical species with low aromaticity but increased heteroatom content. Collectively, results from FT-ICR MS, preparatory-scale separations (extrography/interfacial material), gel permeation chromatography, precipitation behavior in heptane:toluene, thermal decomposition, and aggregate microstructure by atomic force microscopy (among other techniques), suggest that the strong aggregation of asphaltenes results from the synergy between several intermolecular forces: π-stacking, hydrogen bonding, London forces, and acid/base interactions. This review presents general features of asphaltene molecular composition reported over the past five decades. We focus on mass spectrometry characterization and expose the reasons why early results supported the dominance of single-core motifs. Then, the discussion shifts to recent advances in instrumentation for high-field FT-ICR MS, which have enabled the detection of thousands of species in asphaltene samples, whose molecular composition and fragmentation behavior in ultrahigh vacuum agree with the coexistence of single-core and multicore structural motifs. Furthermore, evidence that highlights the limitations of commercially available/custom-built ion sources and selective ionization effects is presented. Consequently, the limitations require separations (e.g., chromatography, extrography) to gain more-comprehensive molecular-level insights into the composition of these complex organic mixtures. The final sections present evidence for the role of aggregation in selective ionization and suggest that advanced characterization by both thermal desorption/decomposition and liquid chromatography with online FT-ICR MS detection can be employed to mitigate the effects of aggregation and provide unique insights in molecular composition/structure.
Abstract
Investigations of abiotic and biotic contributions to dissolved organic carbon (DOC) are required to constrain microbial habitability in continental subsurface fluids. Here we investigate a ...large (101–283 mg C/L) DOC pool in an ancient (>1Ga), high temperature (45–55 °C), low biomass (10
2
−10
4
cells/mL), and deep (3.2 km) brine from an uranium-enriched South African gold mine. Excitation-emission matrices (EEMs), negative electrospray ionization (–ESI) 21 tesla Fourier-transform ion cyclotron resonance mass spectrometry (FT-ICR MS), and amino acid analyses suggest the brine DOC is primarily radiolytically oxidized kerogen-rich shales or reefs, methane and ethane, with trace amounts of C
3
–C
6
hydrocarbons and organic sulfides. δ
2
H and δ
13
C of C
1
–C
3
hydrocarbons are consistent with abiotic origins. These findings suggest water-rock processes control redox and C cycling, helping support a meagre, slow biosphere over geologic time. A radiolytic-driven, habitable brine may signal similar settings are good targets in the search for life beyond Earth.
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