Search

Details of interfacing a high-pressure gas chromatograph to the internal ion source of a Fourier transform ion cyclotron resonance mass spectrometer (FT-ICR MS) are described. We present our preliminary results and potential analytical applications of GC/FT-ICR for analyzing complex biological and environmental sample matrixes, such as petroleum mixtures. Based on GC/FT-ICR data, rapid characterization of various automobile gasoline samples is possible. Comparison between acquired data from the GC/FT-ICR MS (in broadband mode) and a commercial GC quadrupole mass spectrometer (QMS) (over a wide mass range) indicates that sensitivity of the GC/FT-ICR MS is an order of magnitude lower. High mass resolution and mass measurement accuracy of FT-ICR MS can be utilized for unambiguous molecular formula identification of unknown analytes.

Researchers from the University of Maine utilized a gas-phase hydrogen-deuterium exchange reactions and ion storage capabilities of a resonance mass spectrometer to investigate the molecular complexation of aluminum in fulvic acid samples.

A gas-phase hydrogen/deuterium (H/D) exchange reaction technique to determine the number of active hydrogens (NOAH) in fulvic acid ions using electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry is described. First, fulvic acid precursor ions are isolated by stored waveform inverse Fourier transform dipolar excitation. Second, ion-molecule reactions of the selected fulvic acid ions with neutral H/D exchange reagent gases are monitored. The number of incorporated deuterium isotopes in the product ion provides the NOAH for the precursor ion. Previously characterized northern hardwood (NHFA) and red spruce (NCFA) fulvic acid samples were analyzed in this study. Selected ions of both fulvic acid samples undergo H/D exchange with D(sub 2)O, ND(sub 3), and CD(sub 3)OD reagent gases. The extent of H/D exchange of the fulvic acid ions increases with reagent gas basicity, D(sub 2)O < CD(sub 3)OD < ND(sub 3). For the first time, we were able to count the NOAH of selected fulvic acid molecules. The average maximum NOAH for NCFA and NHFA ions at m/z region 700-1000 Th is approximately 7-9. For example, the singly charged NCFA positive ions at m/z 800 Th contain eight active hydrogens. There is no significant difference between NOAH for NCFA and NHFA. The proton affinities of fulvic acid ions at m/z range of 600-1000 Th do not vary significantly.

Endogenously occurring salts and nonvolatile matrix components in untreated biological surfaces can suppress protein ionization and promote adduct formation, challenging protein identification. Characterization of labile proteins within biological specimens is particularly demanding because additional purification or sample treatment steps can be time-intensive and can disrupt noncovalent interactions. It is demonstrated that the combined use of collision-induced unfolding, tandem mass spectrometry, and bottom-up proteomics improves protein characterization in native surface mass spectrometry (NSMS). This multiprong analysis is achieved by acquiring NSMS, MS/MS, ion mobility (IM), and bottom-up proteomics data from a single surface extracted sample. The validity of this multiprong approach was confirmed by the successful characterization of nine surface-deposited proteins, with molecular weights ranging from 8 to 147 kDa, in two separate mixtures. Bottom-up proteomics provided a list of proteins to match against observed proteins in NSMS and their detected subunits in tandem MS. The method was applied to characterize endogenous proteins from untreated chicken liver samples. The subcapsular liver sampling for NSMS analysis allowed for the detection of endogenous proteins with molecular weights of up to ∼220 kDa. Moreover, using IM-MS, collision cross sections and collision-induced unfolding pathways of enzymatic proteins and protein complexes of up to 145 kDa were obtained.

Infrared laser ablation sample transfer (LAST) was used to collect samples from solid surfaces for mass spectrometry under native spray conditions. Native mass spectrometry was utilized to probe the charge states and collision-induced unfolding (CIU) characteristics of bovine serum albumin (BSA), bovine hemoglobin (BHb), and jack-bean concanavalin A (ConA) via direct injection electrospray, after liquid extraction surface sampling, and after LAST. Each protein was deposited from solution on solid surfaces and laser-ablated for off-line analysis or sampled for online analysis. It was found that the protein ion gas-phase charge-state distributions were comparable for direct infusion, liquid extraction, and laser ablation experiments. Moreover, calculated average collision cross section (CCS) values from direct injection, liquid extraction, and laser ablation experiments were consistent with previously reported literature values. Additionally, an equivalent number of mobility features and conformational turnovers were identified from unfolding pathways from all three methods for all charge states of each protein analyzed in this work. The presented work suggests that laser ablation yields intact proteins (BSA, BHb, and ConA), is compatible with native mass spectrometry, and could be suitable for spatially resolved interrogation of unfolding pathways of proteins.

Because of their diverse functionalities in cells, lipids are of primary importance when characterizing molecular profiles of physiological and disease states. Imaging mass spectrometry (IMS) provides the spatial distributions of lipid populations in tissues. Referenced Kendrick mass defect (RKMD) analysis is an effective mass spectrometry (MS) data analysis tool for classification and annotation of lipids. Herein, we extend the capabilities of RKMD analysis and demonstrate an integrated method for lipid annotation and chemical structure-based filtering for IMS datasets. Annotation of lipid features with lipid molecular class, radyl carbon chain length, and degree of unsaturation allows image reconstruction and visualization based on each structural characteristic. We show a proof-of-concept application of the method to a computationally generated IMS dataset and validate that the RKMD method is highly specific for lipid components in the presence of confounding background ions. Moreover, we demonstrate an application of the RKMD-based annotation and filtering to matrix-assisted laser desorption/ionization (MALDI) IMS lipidomic data from human kidney tissue analysis.

A Schwarzschild reflective objective with a numerical aperture of 0.3 and working distance of 10 cm was used for laser ablation sampling of tissue for off-line mass spectrometry. The objective focused the laser to a diameter of 5 μm and produced 10 μm ablation spots on thin ink films and tissue sections. Rat brain tissue sections 50 μm thick were ablated in transmission geometry, and the ablated material was captured in a microcentrifuge tube containing solvent. Proteins from ablated tissue sections were quantified with a Bradford assay, which indicated that approximately 300 ng of protein was captured from a 1 mm 2 area of ablated tissue. Areas of tissue ranging from 0.01 to 1 mm 2 were ablated and captured for bottom-up proteomics. Proteins were extracted from the captured tissue and digested for liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis for peptide and protein identification.

Firefighters are exposed to many different contaminants during structural fires. Moreover, if their protective gear is not successfully decontaminated, firefighters are at risk of being repeatedly exposed to contaminants from previous fires. Thus, the successful removal of contaminants from firefighter turnout gear is necessary to prevent or reduce repeated exposure risks. Laundering methods can reduce the probability of re-exposure to contaminants, such as heavy metals, thus reducing repeated exposure risks. In this study, the efficiencies of heavy metal removal from the firefighter turnout gear outer textile by Decon7 cleaning solution and a standard reference detergent were compared. Nitric acid digests were used to extract metals from textile samples, which were cut from small sections of firefighter jackets, before and after their laundering with either cleaning solution. Inductively coupled plasma mass spectrometry (ICP-MS) was utilized to determine metal contents, including arsenic (As), antimony (Sb), cadmium (Cd), chromium (Cr), and lead (Pb) concentrations. Results from multiplicate samples indicated that, on average, Decon7 was significantly more efficient than a standard detergent in decreasing the concentrations of the five metals studied herein.

Deep-ultraviolet laser ablation with a pulsed 193 nm ArF excimer laser was used to remove localized regions from tissue sections from which proteins were extracted for spatially resolved proteomic analysis by liquid chromatography tandem mass spectrometry (LC-MS/MS). The ability to capture intact proteins by ablation at 193 nm wavelength was verified by matrix-assisted laser desorption ionization (MALDI) of the protein standard bovine serum albumin (BSA), which showed that BSA was ablated and captured without fragmentation. A Bradford assay of the ablated and captured proteins indicated 90% efficiency for transfer of the intact protein at a laser fluence of 3 kJ/m 2 . Rat brain tissue sections mounted on quartz microscope slides and ablated in transmission mode yielded 2 μg protein per mm 2 as quantified by the Bradford assay. Tissue areas ranging from 0.06 mm 2 to 1 mm 2 were ablated and the ejected material was collected for proteomic analysis. Extracted proteins were digested and the resulting peptides were analyzed by LC-MS/MS. The proteins extracted from the ablated areas were identified and the average number of identified proteins ranged from 85 in the 0.06 mm 2 area to 2400 in the 1 mm 2 area of a 50 μm thick tissue. In comparison to infrared laser ablation of equivalent sampled areas, both the protein mass and number of proteins identified using DUV laser ablation sampling were approximately four times larger., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Repulsive electrostatic forces between prion-like proteins are a barrier against aggregation. In neuropharmacology, however, a prion's net charge (Z) is not a targeted parameter. Compounds that selectively boost prion Z remain unreported. Here, we synthesized compounds that amplified the negative charge of misfolded superoxide dismutase-1 (SOD1) by acetylating lysine-NH 3 + in amyloid-SOD1, without acetylating native-SOD1. Compounds resembled a "ball and chain" mace: a rigid amyloid-binding "handle" (benzothiazole, stilbene, or styrylpyridine); an aryl ester "ball"; and a triethylene glycol chain connecting ball to handle. At stoichiometric excess, compounds acetylated up to 9 of 11 lysine per misfolded subunit (ΔZ fibril =-8100 per 10 3 subunits). Acetylated amyloid-SOD1 seeded aggregation more slowly than unacetylated amyloid-SOD1 in vitro and organotypic spinal cord (these effects were partially due to compound binding). Compounds exhibited reactivity with other amyloid and non-amyloid proteins (e.g., fibrillar α-synuclein was peracetylated; serum albumin was partially acetylated; carbonic anhydrase was largely unacetylated)., (© 2021 Wiley-VCH GmbH.)

Hepatic encephalopathy (HE), a neurological disease resulting from liver failure, is difficult to manage and its causes are unclear. Bile acids have been postulated to be involved in the provenance and progression of various diseases including HE. Hence, the characterization of bile acid profiles in the brains of subjects with and without liver failure can provide important clues for the potential treatment of HE. Nanoflow ultra-performance liquid chromatography electrospray ionization ion mobility mass spectrometry (UPLC-ESI-IM-MS) is a highly sensitive method for detection of specific molecules, such as bile acids in brain samples, at biologically relevant concentrations. We used UPLC-ESI-IM-MS to characterize bile acid profiles in brain samples from seven "healthy" control rodents and 22 "diseased" rodents with liver failure (i.e., induced HE). An isomer of trihydroxycholanoyl-taurine was detected in brain tissue samples from both rats and mice with induced HE; however, this isomer was not detected in the brains of healthy rats and mice. Our findings were confirmed by comparing IM arrival times (AT), exact mass measurements (m/z), and mass spectral fragmentation patterns of the experimentally observed suspected species to standards of trihydroxycholanoyl-taurine isomers. Moreover, In Silico Fractionation was employed to provide an additional analytical dimension to verify bile acid identifications., (© 2021 John Wiley & Sons Ltd. This article has been contributed to by US Government employees and their work is in the public domain in the USA.)

Untargeted mass spectrometry (MS) workflows are more suitable than targeted workflows for high throughput characterization of complex biological samples. However, analysis workflows for untargeted methods are inadequate for characterization of complex samples that contain multiple classes of compounds as each chemical class might require a different type of data processing approach. To increase the feasibility of analyzing MS data for multi-class/component complex mixtures (i.e., mixtures containing more than one major class of biomolecules), we developed a neural network-based approach for classification of MS data. In our in silico fractionation (iSF) approach, we utilize a neural decision tree to sequentially classify biomolecules based on their MS-detected isotopic patterns. In the presented demonstration, the neural decision tree consisted of two supervised binary classifiers to positively classify polypeptides and lipids, respectively, and a third supervised network was trained to classify lipids into the eight main sub-categories of lipids. The two binary classifiers assigned polypeptide and lipid experimental components with 100% sensitivity and 100% specificity; however, the 8-target classifier assigned lipids into their respective subclasses with 95% sensitivity and 99% specificity. Here, we discuss important relationships between class-specific chemical properties and MS isotopic envelopes that enable analyte classification. Moreover, we evaluate the performance characteristics of the utilized networks., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

A multimodal workflow for mass spectrometry imaging was developed that combines MALDI imaging with protein identification and quantification by liquid chromatography tandem mass spectrometry (LC-MS/MS). Thin tissue sections were analyzed by MALDI imaging, and the regions of interest (ROI) were identified using a smoothing and edge detection procedure. A midinfrared laser at 3-μm wavelength was used to remove the ROI from the brain tissue section after MALDI mass spectrometry imaging (MALDI MSI). The captured material was processed using a single-pot solid-phase-enhanced sample preparation (SP3) method and analyzed by LC-MS/MS using ion mobility (IM) enhanced data independent acquisition (DIA) to identify and quantify proteins; more than 600 proteins were identified. Using a modified database that included isoform and the post-translational modifications chain, loss of the initial methionine, and acetylation, 14 MALDI MSI peaks were identified. Comparison of the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways of the identified proteins was achieved through an evolutionary relationships classification system., (© 2019 John Wiley & Sons, Ltd.)

The conformations of glycans are crucial for their biological functions. In-electrospray ionization (ESI) hydrogen/deuterium exchange-mass spectrometry (HDX-MS) is a promising technique for studying carbohydrate conformations since rapidly exchanging functional groups, e.g., hydroxyls, can be labeled on the timeframe of ESI. However, regular application of in-ESI HDX to characterize carbohydrates requires further analysis of the in-ESI HDX methodology. For instance, in this method, HDX occurs concurrently to the analyte transitioning from solution to gas-phase ions. Therefore, there is a possibility of sampling both gas-phase and solution-phase conformations of the analyte. Herein, we differentiate in-ESI HDX of metal-adducted carbohydrates from gas-phase HDX and illustrate that this method analyzes solvated species. We also systematically examine the effects of ESI parameters, including spray solvent composition, auxiliary gas flow rate, sheath gas flow rate, sample infusion rate, sample concentration, and spray voltage, and discuss their effects on in-ESI HDX. Further, we model the structural changes of a trisaccharide, melezitose, and its intramolecular and intermolecular hydrogen bonding in solvents with different compositions of methanol and water. These molecular dynamic simulations support our experimental results and illustrate how an individual ESI parameter can alter the conformations we sample by in-ESI HDX. In total, this work illustrates how the fundamental processes of ESI alter the magnitude of HDX for carbohydrates and suggest parameters that should be considered and/or optimized prior to performing experiments with this in-ESI HDX technique. Graphical Abstract ᅟ.

Infrared laser ablation microsampling was used with data-dependent acquisition (DDA) and ion mobility-enhanced data-independent acquisition (HDMS E ) for mass spectrometry based bottom-up proteomics analysis of rat brain tissue. Results from HDMS E and DDA analyses of the 12 laser ablation sampled tissue sections showed that HDMS E consistently identified approximately seven times more peptides and four times more proteins than DDA. To evaluate the impact of ultra-performance liquid chromatography (UPLC) peak congestion on HDMS E and DDA analysis, whole tissue digests from rat brain were analyzed at six different UPLC separation times. Analogous to results from laser ablated samples, HDMS E analyses of whole tissue digests yielded about four times more proteins identified than DDA for all six UPLC separation times., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Background: Glioblastoma multiforme (GBM) is a fatal disease without effective therapy. Identification of new biomarkers for prognosis would enable more rational selections of strategies to cure patients with GBM and prevent disease relapse., Methods: Seven datasets derived from GBM patients using microarray or next generation sequencing in R2 online database (http://r2.amc.nl) were extracted and then analyzed using JMP software. The survival distribution was calculated according to the Kaplan-Meier method and the significance was determined using log-rank statistics. The sensitivity of a panel of GBM cell lines in response to temozolomide (TMZ), salinomycin, celastrol, and triptolide treatments was evaluated using MTS and tumor-sphere formation assay., Findings: We identified that CD44, ATP binding cassette subfamily C member 3 (ABCC3), and tumor necrosis factor receptor subfamily member 1A (TNFRSF1A) as highly expressed genes in GBMs are associated with patients' poor outcomes and therapy resistance. Furthermore, these three markers combined with MGMT, a conventional GBM marker, can classify GBM patients into five new subtypes with different overall survival time in response to treatment. The four-gene signature and the therapy response of GBMs to a panel of therapeutic compounds were confirmed in a panel of GBM cell lines., Interpretation: The data indicate that the four-gene panel can be used as a therapy response index for GBM patients and potential therapeutic targets. These results provide important new insights into the early diagnosis and the prognosis for GBM patients and introduce potential targets for GBM therapeutics. FUND: Baylor Scott & White Health Startup Fund (E.W.); Collaborative Faculty Research Investment Program (CFRIP) of Baylor University, Baylor Scott & White Health, and Baylor College of Medicine (E.W., T.S., J.H.H.); NIH R01 NS067435 (J.H.H.); Scott & White Plummer Foundation Grant (J.H.H.); National Natural Science Foundation of China 816280007 (J.H.H. and Fu.W.)., (Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.)

Chemical identification often relies on matching measured chemical properties and/or spectral "fingerprints" of unknowns against their precompiled libraries. Chromatography, absorption spectroscopy, and mass spectrometry are all among analytical approaches that provide chemical measurement databases amenable to library searching. Occasionally, using conventional single-library or single-domain searches can lead to misidentification of unknowns. To improve chemical identification, we present a tandem gas chromatography/vacuum ultraviolet-mass spectrometry (GC/VUV-MS) chemical identification approach that utilizes databases from GC, VUV spectroscopy, and mass spectrometry analyses for a "multidomain" library search. Using standard chemical mixtures as well as aroma compounds as test cases, we demonstrate that multidatabase library searches utilizing GC, VUV, and MS data results in fully correct identification of chemical mixtures examined here that could only be identified with a 69.2% or an 88.5% success rate with MS or VUV library searches alone, respectively. Additionally, we introduce a library- and data domain-independent metric for evaluating the confidence of library search results. Using multidomain library searches improves both the chemical assignment accuracy and confidence.

Several recent reports suggest that HNO may be produced endogenously by reaction of H 2 S and S-nitrosoglutathione (GSNO). This hypothesis was tested using deoxymyoglobin (MbFe II ) to trap the expected HNO released from the target reaction, which should generate the stable HNO adduct, HNO-Mb, under anaerobic conditions. Under numerous experimental conditions, the sole globin product was NO-Mb, as characterized by absorbance, EPR, and NMR spectroscopies. Analogous reactions of GSNO with other biological reductants such as ascorbic acid, dithiothreitol, glutathione, and dithionite also yielded NO-Mb as the sole globin product; however, whereas analogous reduction of GSNO using NaBH 4 generates HNO-Mb in high yield. Quantitative GC/MS analyses of reactions of GS 15 NO with H 2 S showed that the main reaction product was 15 NO, with 15 N 2 produced at a comparable level to 15 N 2 O. Overall yield of N 2 O is unchanged by the presence of MbFe II , discounting the intermediacy of either NO or HNO in its formation. Taken together, these results argue against the generation of free HNO as a major pathway in the reactions of GSNO with H 2 S, and instead imply some as yet uncharacterized intermediates generate the nitrogenic gases., (Copyright © 2018 Elsevier Inc. All rights reserved.)

High resolving power ion mobility (IM) allows for accurate characterization of complex mixtures in high-throughput IM mass spectrometry (IM-MS) experiments. We previously demonstrated that pure component IM-MS data can be extracted from IM unresolved post-IM/collision-induced dissociation (CID) MS data using automated ion mobility deconvolution (AIMD) software [Matthew Brantley, Behrooz Zekavat, Brett Harper, Rachel Mason, and Touradj Solouki, J. Am. Soc. Mass Spectrom., 2014, 25, 1810-1819]. In our previous reports, we utilized a quadrupole ion filter for m/z-isolation of IM unresolved monoisotopic species prior to post-IM/CID MS. Here, we utilize a broadband IM-MS deconvolution strategy to remove the m/z-isolation requirement for successful deconvolution of IM unresolved peaks. Broadband data collection has throughput and multiplexing advantages; hence, elimination of the ion isolation step reduces experimental run times and thus expands the applicability of AIMD to high-throughput bottom-up proteomics. We demonstrate broadband IM-MS deconvolution of two separate and unrelated pairs of IM unresolved isomers (viz., a pair of isomeric hexapeptides and a pair of isomeric trisaccharides) in a simulated complex mixture. Moreover, we show that broadband IM-MS deconvolution improves high-throughput bottom-up characterization of a proteolytic digest of rat brain tissue. To our knowledge, this manuscript is the first to report successful deconvolution of pure component IM and MS data from an IM-assisted data-independent analysis (DIA) or HDMSE dataset.

For wide class characterizations of volatile organic compounds (VOCs), conventional gas chromatography mass spectrometry (GC-MS)-based techniques are utilized. These GC-MS-based chemical identification approaches typically rely on library searches against ion fragmentation patterns of known compounds. Although MS library searches can often provide correct chemical identities, erroneous chemical assignments of structurally similar unknown compounds are also possible. Other detection systems, such as absorption spectrometers, have been used for VOC analysis and can provide complementary absorption data. Here, we demonstrate the analytical advantages of coupling vacuum ultraviolet (VUV) absorption spectroscopy and MS in tandem for the improved characterization of structurally similar VOCs. We also discuss technical considerations and limitations of coupling a VUV spectrometer to a quadrupole mass spectrometer. Moreover, we show that combining the isomer selectivity of VUV spectroscopy, as a nondestructive analyte detection approach, with the mass selectivity of MS in a VUV-MS detection system improves characterization of GC-eluting compounds. Utilizing GC/VUV-MS data, we demonstrate that orthogonal VUV and MS library searches improve identification of VOCs present in complex mixtures such as a mixed standard sample, a commercial perfume product, and an essential oil sample.

Peak broadening in ion mobility (IM) is a relatively predictable process and abnormally broad peaks can be indicative of the presence of unresolved species. Here, we introduce a new ion mobility peak fitting (IM&#95;FIT) software package for automated and systematic determination of traveling wave ion mobility (TWIM) unresolved species. To identify IM unresolved species, the IM&#95;FIT software generates a trend line by plotting ions' mobility peak widths as a function of their arrival times. Utilizing user-defined thresholds, IM&#95;FIT allows for automated and rapid detection of ions that deviate from the peak width trend line. To demonstrate the advantages of IM&#95;FIT for automated detection of IM unresolved species, IM-mass spectrometry (IM-MS) data from a sample mixture containing polypropylene glycol and multiple peptides were analyzed. A total of 14 out of the 34 observed singly-charged IM peaks above 5% relative abundance (i.e., signal-to-noise ratios above ∼200) were tagged as potentially co-eluting ions by IM&#95;FIT. Subsequently, the 14 IM peaks tagged as potentially unresolved (presumably, peaks corresponding to co-eluting compounds), were further analyzed by automated IM deconvolution (AIMD), liquid chromatography-IM-MS (LC-IM-MS), and/or ultra-high resolution mass spectrometry. Using the aforementioned techniques, more than 85% of the tagged IM peaks (12 out of 14) were confirmed to contain co-eluting ions. As an additional new finding, IM&#95;FIT facilitated the discovery of an unexpected sequence-scrambled y-type fragment ion., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Ion mobility (IM) is an important analytical technique for determining ion collision cross section (CCS) values in the gas-phase and gaining insight into molecular structures and conformations. However, limited instrument resolving powers for IM may restrict adequate characterization of conformationally similar ions, such as structural isomers, and reduce the accuracy of IM-based CCS calculations. Recently, we introduced an automated technique for extracting "pure" IM and collision-induced dissociation (CID) mass spectra of IM overlapping species using chemometric deconvolution of post-IM/CID mass spectrometry (MS) data [J. Am. Soc. Mass Spectrom., 2014, 25, 1810-1819]. Here we extend those capabilities to demonstrate how extracted IM profiles can be used to calculate accurate CCS values of peptide isomer ions which are not fully resolved by IM. We show that CCS values obtained from deconvoluted IM spectra match with CCS values measured from the individually analyzed corresponding peptides on uniform field IM instrumentation. We introduce an approach that utilizes experimentally determined IM arrival time (AT) "shift factors" to compensate for ion acceleration variations during post-IM/CID and significantly improve the accuracy of the calculated CCS values. Also, we discuss details of this IM deconvolution approach and compare empirical CCS values from traveling wave (TW)IM-MS and drift tube (DT)IM-MS with theoretically calculated CCS values using the projected superposition approximation (PSA). For example, experimentally measured deconvoluted TWIM-MS mean CCS values for doubly-protonated RYGGFM, RMFGYG, MFRYGG, and FRMYGG peptide isomers were 288.8 Å(2), 295.1 Å(2), 296.8 Å(2), and 300.1 Å(2); all four of these CCS values were within 1.5% of independently measured DTIM-MS values., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Radio-frequency ionization (RFI) is a novel ionization method coupled to mass spectrometry (MS) for analysis of semi-volatile and volatile organic compounds (VOCs). Despite the demonstrated capabilities of RFI MS for VOC analysis in both positive- and negative-ion modes, mechanism of RFI is not completely understood. Improved understanding of the ion generation process in RFI should expand its utility in MS. Here, we studied the possibility of electron emission in RFI using both direct charged particle current measurements and indirect electron detection in a 9.4-T Fourier transform-ion cyclotron resonance (FT-ICR) mass spectrometer. We show that RF-generated electrons can be trapped in the ICR cell and, subsequently, reacted with neutral hexafluorobenzene (C6 F6 ) molecules to generate C6 F6 (●-) . Intensity of observed C6 F6 (●-) species correlated with the number of trapped electrons and decreased as a function of electron quenching period. We also measured the electron attachment rate constant of hexafluorobenzene using a post-RF electron trapping experiment. Measured electron attachment rate constant of hexafluorobenzene (1.19 (±0.53) × 10(-9)  cm(3)  molecule(-1)  s(-1) ) for post-RF FT-ICR MS agreed with the previously reported value (1.60 (±0.30) × 10(-9)  cm(3)  molecule(-1)  s(-1) ) from low-pressure ICR MS measurements. Experimental results from direct and indirect electron measurements suggest that RFI process involves RF-generated electrons under ultrahigh vacuum conditions., (Copyright © 2015 John Wiley & Sons, Ltd.)

A combination of density functional theory calculations, hydrogen/deuterium exchange (HDX) reactions, ion mobility-mass spectrometry, and isotope labeling tandem mass spectrometry was used to study gas-phase "host-guest" type interactions of a benzyloxycarbonyl (Z)-capped proline (P) glycine (G) model dipeptide (i.e., Z-PG) and its various structural analogues with ND3. It is shown that in a solvent-free environment, structural differences between protonated and alkali metal ion (Na(+), K(+), or Cs(+))-complexed species of Z-PG affect ND3 adduct formation. Specifically, [Z-PG + H](+) and [Z-PG-OCH3 + H](+) formed gas-phase ND3 adducts ([Z-PG (or Z-PG-OCH3) + H + ND3](+)) but no ND3 adducts were observed for [Z-PG + alkali metal](+) or [Z-PG + H - CO2](+). Experimentally measured and theoretically calculated collision cross sections (CCSs) of protonated and alkali metal ion-complexed Z-PG species showed similar trends that agreed with the observed structural differences from molecular modeling results. Moreover, results from theoretical ND3 affinity calculations were consistent with experimental HDX observations, indicating a more stable ND3 adduct for [Z-PG + H](+) compared to [Z-PG + alkali metal](+) species. Molecular modeling and experimental MS results for [Z-PG + H](+) and [Z-PG + alkali metal](+) suggest that optimized cation-π and hydrogen bonding interactions of carbonyl groups in final products are important for ND3 adduct formation. Graphical Abstract ᅟ.

Existing instrumental resolving power limitations in ion mobility spectrometry (IMS) often restrict adequate characterization of unresolved or co-eluting chemical isomers. Recently, we introduced a novel chemometric deconvolution approach that utilized post-IM collision-induced dissociation (CID) mass spectrometry (MS) data to extract "pure" IM profiles and construct CID mass spectra of individual components from a mixture containing two IM-overlapped components [J. Am. Soc. Mass Spectrom., 2012, 23, 1873-1884]. In this manuscript we extend the capabilities of the IM-MS deconvolution methodology and demonstrate the utility of energy resolved IM deconvolution for successful characterization of ternary and quaternary isomer mixtures with overlapping IM profiles. Furthermore, we show that the success of IM-MS deconvolution is a collision-energy dependent process where different isomers can be identified at various ion fragmentation collision-energies. Details on how to identify a single collision-energy or suitable collision-energy ranges for successful characterization of isomer mixtures are discussed. To confirm the validity of the proposed approach, deconvoluted IM and MS spectra from IM overlapped analyte mixtures are compared to IM and MS data from individually run mixture components. Criteria for "successful" deconvolution of overlapping IM profiles and extraction of their corresponding pure mass spectra are discussed.

Collision-induced dissociation (CID) of m/z-isolated w type fragment ions and an intact 5' phosphorylated DNA oligonucleotide generated rearranged product ions. Of the 21 studied w ions of various nucleotide sequences, fragment ion sizes, and charge states, 18 (~86%) generated rearranged product ions upon CID in a Synapt G2-S HDMS (Waters Corporation, Manchester, England, UK) ion mobility-mass spectrometer. Mass spectrometry (MS), ion mobility spectrometry (IMS), and theoretical modeling data suggest that purine bases can attack the free 5' phosphate group in w type ions and 5' phosphorylated DNA to generate sequence permuted [phosphopurine](-) fragment ions. We propose and discuss a potential mechanism for generation of rearranged [phosphopurine](-) and complementary y-B type product ions.

Presence of unresolved ion mobility (IM) profiles limits the efficient utilization of IM mass spectrometry (IM-MS) systems for isomer differentiation. Here, we introduce an automated ion mobility deconvolution (AIMD) computer software for streamlined deconvolution of overlapped IM-MS profiles. AIMD is based on a previously reported post-IM/collision-induced dissociation (CID) deconvolution approach [J. Am. Soc. Mass Spectrom. 23, 1873 (2012)] and, unlike the previously reported manual approach, it does not require resampling of post-IM/CID data. A novel data preprocessing approach is utilized to improve the accuracy and efficiency of the deconvolution process. Results from AIMD analysis of overlapped IM profiles of data from (1) Waters Synapt G1 for a binary mixture of isomeric peptides (amino acid sequences: GRGDS and SDGRG) and (2) Waters Synapt G2-S for a binary mixture of isomeric trisaccharides (raffinose and isomaltotriose) are presented.

It is shown that y-type ions, after losing C-terminal H2O or NH3, can lose an internal backbone carbonyl (CO) from different peptide positions and yield structurally different product fragment ions upon collision-induced dissociation (CID). Such CO losses from internal peptide backbones of y-fragment ions are not unique to a single peptide and were observed in four of five model peptides studied herein. Experimental details on examples of CO losses from y-type fragment ions for an isotopically labeled AAAAHAA-NH2 heptapeptide and des-acetylated-α-melanocyte-stimulating hormone (dα-MSH) (SYSMEHFRWGKPV-NH2) are reported. Results from isotope labeling, tandem mass spectrometry (MS(n)), and ion mobility-mass spectrometry (IM-MS) confirm that CO losses from different amino acids of m/z-isolated y-type ions yield structurally different ions. It is shown that losses of internal backbone carbonyls (as CID products of m/z-isolated y-type ions) are among intermediate steps towards formation of rearranged or permutated product fragment ions. Possible mechanisms for generation of the observed sequence-scrambled a-"like" ions, as intermediates in sequence-scrambling pathways of y-type ions, are proposed and discussed.

To date, only a limited number of reports are available on structural variants of multiply-charged b-fragment ions. We report on observed bimodal gas-phase hydrogen/deuterium exchange (HDX) reaction kinetics and patterns for substance P b10(2+) that point to presence of isomeric structures. We also compare HDX reactions, post-ion mobility/collision-induced dissociation (post-IM/CID), and sustained off-resonance irradiation-collision induced dissociation (SORI-CID) of substance P b10(2+) and a cyclic peptide with an identical amino acid (AA) sequence order to substance P b10. The observed HDX patterns and reaction kinetics and SORI-CID pattern for the doubly charged head-to-tail cyclized peptide were different from either of the presumed isomers of substance P b10(2+), suggesting that b10(2+) may not exist exclusively as a head-to-tail cyclized structure. Ultra-high mass measurement accuracy was used to assign identities of the observed SORI-CID fragment ions of substance P b10(2+); over 30% of the observed SORI-CID fragment ions from substance P b10(2+) had rearranged (scrambled) AA sequences. Moreover, post-IM/CID experiments revealed the presence of two conformer types for substance P b10(2+), whereas only one conformer type was observed for the head-to-tail cyclized peptide. We also show that AA sequence scrambling from CID of doubly-charged b-fragment ions is not unique to substance P b10(2+).

Rationale: Low-mass cut-off restrictions for injecting ions from external ion sources into high magnetic fields impose limitations for wide mass range analyses with Fourier transform ion cyclotron resonance (FTICR) instruments. Radio-frequency (RF)-only quadrupole ion guides (QIGs) with higher frequencies can be used to overcome low-mass cut-off in FTICR instruments., Methods: RF signals (1.0 MHz to 10.0 MHz) were applied to QIGs to transfer externally generated ions from either electron ionization (EI) or electrospray ionization (ESI) sources into ICR cells of 9.4 T FTICR mass spectrometers. Efficiencies of QIGs were evaluated using externally generated ions from: EI of acetone, air, and perfluorotributylamine mixture, EI of gas chromatography (GC)-separated components of a standard sample mixture, and ESI of complex mixtures such as petroleum and fulvic acid samples., Results: We were able to transfer ions with m/z as low as 26 from an external EI source into the ICR cell of a 9.4 T FTICR mass spectrometer and extend the operational low-mass range for ESI-FTICR analyses. High mass resolving power and mass measurement accuracy of GC/FTICR mass spectrometry were utilized to discriminate between oxygenated and non-oxygenated compounds in a 'Grob' sample. Ion losses based on SIMION ion trajectory predictions were consistent with experimental findings., Conclusions: We demonstrated that the use of high-frequency QIGs can extend the operational lower m/z range for both external EI- and ESI-FTICR mass spectrometers. By considering both ICR and Mathieu equations of motions to describe ion trajectories, theoretical ion ejection thresholds (consistent with our experimental findings) could be predicted., (Copyright © 2013 John Wiley & Sons, Ltd.)

Sequence scrambling from y-type fragment ions has not been previously reported. In a study designed to probe structural variations among b-type fragment ions, it was noted that y fragment ions might also yield sequence-scrambled ions. In this study, we examined the possibility and extent of sequence-scrambled fragment ion generation from collision-induced dissociation (CID) of y-type ions from four peptides (all containing basic residues near the C-terminus) including: AAAAHAA-NH2 (where "A" denotes carbon thirteen ((13)C1) isotope on the alanine carbonyl group), des-acetylated-α-melanocyte (SYSMEHFRWGKPV-NH2), angiotensin II antipeptide (EGVYVHPV), and glu-fibrinopeptide b (EGVNDNEEGFFSAR). We investigated fragmentation patterns of 32 y-type fragment ions, including y fragment ions with different charge states (+1 to +3) and sizes (3 to 12 amino acids). Sequence-scrambled fragment ions were observed from ~50 % (16 out of 32) of the studied y-type ions. However, observed sequence-scrambled ions had low relative intensities from ~0.1 % to a maximum of ~12 %. We present and discuss potential mechanisms for generation of sequence-scrambled fragment ions. To the best of our knowledge, results on y fragment dissociation presented here provide the first experimental evidence for generation of sequence-scrambled fragments from CID of y ions through intermediate cyclic "b-type" ions.

Although structural isomers may yield indistinguishable ion mobility (IM) arrival times and similar fragment ions in tandem mass spectrometry (MS), it is demonstrated that post-IM/collision-induced dissociation MS (post-IM/CID MS) combined with chemometrics can enable independent study of the IM-overlapped isomers. The new approach allowed us to investigate the propensity of selected b type fragment ions from AlaAlaAlaHisAlaAlaAla-NH2 (AAA(His)AAA) heptapeptide to form different isomers. Principle component analysis (PCA) of the unresolved post-IM/CID profiles indicated the presence of two different isomer types for b4(+), b5(+), and b6(+) and a single isomer type for b7(+) fragments of AAA(His)AAA. We employed a simple-to-use interactive self-modeling mixture analysis (SIMPLISMA) to calculate the total IM profiles and CID mass spectra of b fragment isomers. The deconvoluted CID mass spectra showed discernible fragmentation patterns for the two isomers of b4(+), b5(+), and b6(+) fragments. Under our experimental conditions, calculated percentages of the "cyclic" isomers (at the 95% confidence level for n = 3) for b4(+), b5(+), and b6(+) were 61 (± 5)%, 36 (± 5)%, and 48 (± 2)%, respectively. Results from the SIMPLISMA deconvolution of b5(+) species resembled the CID MS patterns of fully resolved IM profiles for the two b5(+) isomers. The "cyclic" isomers for each of the two-component b fragment ions were less susceptible to ion fragmentation than their "linear" counterparts.

A new ionization technique: A radio-frequency signal was used to ionize neutral organic molecules in the ultrahigh-vacuum region of a Fourier transform ion cyclotron resonance mass spectrometer. Radio-frequency ionization (RFI) yielded signal/noise (S/N) ratios roughly six times higher than those generated by conventional 70 eV electron impact ionization (EI)., (Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

We present the details of a data analysis approach for deconvolution of the ion mobility (IM) overlapped or unresolved species. This approach takes advantage of the ion fragmentation variations as a function of the IM arrival time. The data analysis involves the use of an in-house developed data preprocessing platform for the conversion of the original post-IM/collision-induced dissociation mass spectrometry (post-IM/CID MS) data to a Matlab compatible format for chemometric analysis. We show that principle component analysis (PCA) can be used to examine the post-IM/CID MS profiles for the presence of mobility-overlapped species. Subsequently, using an interactive self-modeling mixture analysis technique, we show how to calculate the total IM spectrum (TIMS) and CID mass spectrum for each component of the IM overlapped mixtures. Moreover, we show that PCA and IM deconvolution techniques provide complementary results to evaluate the validity of the calculated TIMS profiles. We use two binary mixtures with overlapping IM profiles, including (1) a mixture of two non-isobaric peptides (neurotensin (RRPYIL) and a hexapeptide (WHWLQL)), and (2) an isobaric sugar isomer mixture of raffinose and maltotriose, to demonstrate the applicability of the IM deconvolution.

Electrospray ionization (ESI) Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) combined with H/D exchange reactions was utilized to explore the existence of different b(5)(+) and b(4)(+) fragment ion conformers/isomers of hexapeptide WHWLQL in the gas phase. Distinct H/D exchange trends for protonated WHWLQL ([M + H](+)) and its b(5)(+) and b(4)(+) fragment ions (with ND(3)) were observed. Isolated (12)C(all) isotopomers of both b(5)(+) and b(4)(+) fragment ions yielded bimodal distributions of H/D exchanged product ions. The H/D exchange reaction kinetics also confirmed that b(5)(+) and b(4)(+) fragment ions exist as combination of slow-exchanging ("s") and fast-exchanging ("f") species. The calculated rate constant for the first labile hydrogen exchange of [M + H](+) (k([M + H](+)) = 3.80 +/- 0.7 x 10(-10) cm(3) mol(-1) s(-1)) was approximately 30 and approximately 5 times greater than those for the "s" and "f" species of b(5)(+), respectively. Data from H/D exchange of isolated "s" species at longer ND(3) reaction times confirmed the existence of different conformers or isomers for b(5)(+) fragment ions. The sustained off-resonance irradiation collision-activated dissociation (SORI-CAD) of WHWLQL combined with the H/D exchange reactions indicate that "s" and "f" species of b(5)(+) and b(4)(+) fragment ions can be produced in the ICR cell as well as the ESI source. The significance of these observations for detailed understanding of protein sequencing and ion fragmentation pathways is discussed., (Copyright 2010. Published by Elsevier Inc.)

The contributions of molecular and fragment ions toward the disparate self-chemical ionization (SCI) of alpha-pinene and camphene isomers were investigated. A kinetic model was constructed to predict the SCI outcomes for these two C(10)H(16) isomers. A major portion of the camphene molecular ions (isolated 500 ms after the 10 ms EI event at 24 eV) unimolecularly dissociated within 200 s of the ionization event. Conversely, under similar experimental conditions, the alpha-pinene molecular ions as well as the major fragment ions of alpha-pinene and camphene showed no unimolecular dissociation. The alpha-pinene and camphene molecular ions yielded product ions through two different reaction mechanisms (direct charge-transfer {CT} and indirect proton transfer {PT}). The isolated terpene fragment ions at m/z 93 and 121 reacted with their respective neutrals to produce [M + H](+). Proton affinity (PA) bracketing experiments, PA additivity schemes, and alkene PA versus adiabatic ionization energy (IE) linear correlation indicated that the PAs of camphene and alpha-pinene were comparable ( approximately 210 +/- 2 kcal x mol(-1)). The observed [M + H](+) SCI terpene ions were mainly the products of various fragment ion reactions.

Water samples from a local water treatment plant were analyzed, using gas chromatography Fourier transform ion cyclotron resonance mass spectrometry (GC/FT-ICR MS), to identify potential disinfection byproducts (DBPs). Both liquid-liquid extraction (LLE) and solid-phase microextraction (SPME) techniques were used for sample preparation prior to GC/MS analyses. Based on the averaged mass measurement accuracy (MMA) of better than five parts-per-million (<5 ppm), multiple solvent artifacts were identified. It is shown that solventless SPME can be utilized to reduce potential interferences from solvent stabilizers. Six DBPs were detected and their molecular compositions were assigned at a high level of confidence. At the ppb concentration ranges and in the broadband mass spectral detection mode, internally calibrated mass spectra provided concurrent high resolution (resolving power M/deltaM50% > 30,000 at m/z values -110) and MMA of better than one part-per-million (MMA < 1 ppm). The use of thermochemical data, such as proton affinities, as a complementary tool to enhance analytical resolution is also demonstrated.

Coupling of a cryogenic preconcentrator (PC) to a gas chromatograph/Fourier transform ion cyclotron resonance mass spectrometer (GC/FT-ICR MS) is reported. To demonstrate the analytical capabilities of the PC/GC/FT-ICR MS, headspace samples containing volatile organic compounds (VOCs) emitted from detached pine tree twigs were analyzed. Sub-ppm mass measurement accuracy (MMA) for highly resolved (m/Deltam(50%) > 150 k) terpene ions was achieved. Direct PC/GC/FT-ICR MS analyses revealed that detached twigs from pine trees emit acetone, camphor, and four detectable hydrocarbon isomers with C(10)H(16) empirical formula. The unknown analytes were identified based on accurate mass measurement and their mass spectral appearances. Authentic samples were used to confirm initially unknown identifications. Self-chemical-ionization (SCI) reactions furnished an additional dimension for rapid isomer differentiation of GC eluents in real time.

Mass spectra of fragments of permethylated oligosaccharides are analyzed by Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry. Sustained off-resonance irradiation (SORI) collision-induced dissociation (CID), quadrupolar axialization, multiple stages of isolation and dissociation (MSn), and ion remeasurement are exploited for carbohydrate structural analyses. That SORI CID internal energies are adequate for linkage analysis of a permethylated glucose oligomer is demonstrated by identifying ring-opened fragment ions from MALDI-generated mass-isolated and collisionally activated ions. Ion remeasurement and axialization techniques enhance the sensitivity of ion fragmentation analysis. Multiple stages of isolation and dissociation of ion fragments (MSn) provide for structural analysis of an electrospray-ionized permethylated lacto-N-fucopentaose isomer (LNFP II). Compared to MS2 spectra taken with a triple quadrupole, FT-ICR MSn (n > 2) provides more extensive characterization of the parent molecular structure than is available from a single stage of ion isolation and dissociation (MS2).