Trapped Ion Mobility Mass Spectrometry Research Articles

Exploring Isomerism in Isolated Cyclodextrin Oligomers through Trapped Ion Mobility Mass Spectrometry.

Cyclodextrin (CD) macrocycles are used to create a wide range of supramolecular architectures which are also of interest in applications such as selective gas adsorption, drug delivery, and catalysis. However, predicting their assemblies and identifying the possible isomers in CD oligomers have always remained challenging due to their dynamic nature. Herein, we interacted CDs (α, β, and γ) with a divalent metal ion, Cu2+, to create a series of Cu2+-linked CD oligomers, from dimers to pentamers. We characterized these oligomers using electrospray ionization mass spectrometry and probed isomerism in each of these isolated oligomers using high resolution trapped ion mobility spectrometry. Using this technique, we separated multiple isomers for each of the Cu2+-interlinked CD oligomers and estimated their relative population, which was not accessible previously using other characterization techniques. We further carried out structural analysis of the observed isomers by comparing the experimental collision cross sections (CCSs) to that of modeled structures. We infer that the isomeric heterogeneity reflects size-specific packing patterns of individual CDs (e.g., close-packed/linear). In some cases, we also reveal the existence of kinetically trapped structures in the gas phase and study their transformation to thermodynamically controlled forms by examining the influence of activation of the ions on isomer interconversion.

Four-dimensional trapped ion mobility spectrometry lipidomics for high throughput clinical profiling of human blood samples

Lipidomics encompassing automated lipid extraction, a four-dimensional (4D) feature selection strategy for confident lipid annotation as well as reproducible and cross-validated quantification can expedite clinical profiling. Here, we determine 4D descriptors (mass to charge, retention time, collision cross section, and fragmentation spectra) of 200 lipid standards and 493 lipids from reference plasma via trapped ion mobility mass spectrometry to enable the implementation of stringent criteria for lipid annotation. We use 4D lipidomics to confidently annotate 370 lipids in reference plasma samples and 364 lipids in serum samples, and reproducibly quantify 359 lipids using level-3 internal standards. We show the utility of our 4D lipidomics workflow for high-throughput applications by reliable profiling of intra-individual lipidome phenotypes in plasma, serum, whole blood, venous and finger-prick dried blood spots.

Monitoring Conformational Changes of Lysozyme–Polyelectrolyte Complexes Using Trapped Ion Mobility-Mass Spectrometry (IM-MS)

The conformational changes of lysozyme due to the formation of its complexes with polystyrene sulfonate (PSS) oligomers were monitored at different charge states using trapped ion mobility spectrometry-time-of-flight mass spectrometer (TIMS-TOF-MS). The mixtures of disulfide-intact lysozyme and PSS oligomers were directly infused into the ESI source of the TIMS-TOF-MS instrument. At charge states from 7+ to 12+ in the gas phase, mobilograms obtained from TIMS-TOF-MS analyses were extracted for the protein and each complex ion. Changes occurring in the protein conformation as a result of the binding of PSS oligomers of different chain lengths were evaluated by comparing the obtained integrated peak areas for compact and extended conformers in the extracted ion mobilograms. The increase in the abundance of compact conformers due to the complexation of lysozyme and PSS oligomers was observed more clearly in the 8+ charge state. The number and lengths of PSS chains had little or no effect on the complex conformation in the higher charge states that did not provide compaction in the entire structure. It was observed that the molecular dynamics (MD) simulations of the protein and complexes correlate well with the results obtained by TIMS-TOF-MS. The PSS chains interact with the positively charged sites of the protein and cause compaction in the conformation by changing the locations of the excess charge on the structure in the gas phase that was monitored using TIMS regarding the length and number of the attached polyelectrolyte chain.

Developments in Trapped Ion Mobility Mass Spectrometry to Probe the Early Stages of Peptide Aggregation.

Ion mobility mass spectrometry (IM-MS) has proven to be an excellent method to characterize the structure of amyloidogenic protein and peptide aggregates, which are formed in coincidence with the development of neurodegenerative diseases. However, it remains a challenge to obtain detailed structural information on all conformational intermediates, originating from the early onset of those pathologies, due to their complex and heterogeneous environment. One way to enhance the insights and the identification of these early stage oligomers is by employing high resolution ion mobility mass spectrometry experiments. This would allow us to enhance the mobility separation and MS characterization. Trapped ion mobility spectrometry (TIMS) is an ion mobility technique known for its inherently high resolution and has successfully been applied to the analysis of protein conformations among others. To obtain conformational information on fragile peptide aggregates, the instrumental parameters of the TIMS-Quadrupole-Time-of-Flight mass spectrometer (TIMS-qToF-MS) have to be optimized to allow the study of intact aggregates and ensure their transmission toward the detector. Here, we investigate the suitability and application of TIMS to probe the aggregation process, targeting the well-characterized M307-N319 peptide segment of the TDP-43 protein, which is involved in the development of amyotrophic lateral sclerosis. By studying the influence of key parameters over the full mass spectrometer, such as source temperature, applied voltages or RFs among others, we demonstrate that by using an optimized instrumental method TIMS can be used to probe peptide aggregation.

Quantitation of Enantiomeric Excess in an Achiral Environment Using Trapped Ion Mobility Mass Spectrometry.

We present a novel, straightforward method to determine the enantiomeric excess (ee) of tryptophan (Trp) and N-tert-butyloxycarbonyl-O-benzylserine (BBS) solutions without chiral additives. For this, lithium carbonate, sodium carbonate, or silver acetate was added to solutions of Trp or BBS. Singly negatively charged dimer and trimer clusters were then formed by electrospray ionization and analyzed using trapped ion mobility spectrometry (TIMS) and time-of-flight mass spectrometry. When a solution contains both enantiomers, homo- and heterochiral clusters are generated which can be separated in the TIMS-tunnel based on their different mobilities using a nitrogen buffer gas. The ratio of homochiral to heterochiral clusters shows a binomial distribution and can be calibrated with solutions of known ee to yield ee measurements of samples with better than 1% accuracy. Samples can be prepared rapidly, and measurements are completed in less than 5 min. Current instrumental limitations restrict this method to rigid molecules with large functional groups adjacent to the chiral centers. Nevertheless, we expect this method to be applicable to many pharmaceuticals and provide the example of 1-methyltryptophan to demonstrate this.

Probing topology of supramolecular complexes between cyclodextrins and alkali metals by ion mobility-mass spectrometry

In this study, the size and shape of supramolecular assemblies between cyclo-oligosaccharides and proton, ammonium or a series of alkali metals by electrospray coupled to trapped ion mobility-mass spectrometry (ESI-TIMS) have investigated. Native cyclodextrins (CD) were selected as models, and collision cross section (CCS) values were deducted for the main positive singly and doubly charged species. Experimental CCS values were in good agreement with those obtained from molecular modeling. Due to the high mobility resolving power and resolution, it was possible to highlight the presence of various conformers. Also, TIMS allowed to discriminate and estimate the content of various orientations from non-covalent nanotubes-based CD, involving secondary/secondary rim hydroxyl groups (head-to head), primary/secondary rim (head-to-tail) hydroxyl groups or primary/primary rim (tail-to-tail) hydroxyl groups interactions. Such results pave the way for a better knowledge of the topology of cyclo-oligosaccharides based supramolecular complexes, demonstrating that TIMS can be a particularly attractive molecular descriptor.

Yellow Fever Blood Proteomics Study by Trapped Ion Mobility Mass Spectrometry

Yellow Fever Blood Proteomics Study by Trapped Ion Mobility Mass Spectrometry

Structural amyloid plaque polymorphism is associated with distinct lipid accumulations revealed by trapped ion mobility mass spectrometry imaging.

Understanding of Alzheimer's disease (AD) pathophysiology requires molecular assessment of how key pathological factors, specifically amyloid β (Aβ) plaques, influence the surrounding microenvironment. Here, neuronal lipids have been implicated in Aβ plaque pathology, though the lipid microenvironment in direct proximity to Aβ plaques is still not fully resolved. A further challenge is the microenvironmental molecular heterogeneity, across structurally polymorphic Aβ features, such as diffuse, immature, and mature, fibrillary aggregates, whose resolution requires the integration of advanced, multimodal chemical imaging tools. Herein, we used matrix-assisted laser desorption/ionization trapped ion mobility spectrometry time-of-flight based mass spectrometry imaging (MALDI TIMS TOF MSI) in combination with hyperspectral confocal microscopy to probe the lipidomic microenvironment associated with structural polymorphism of Aβ plaques in transgenic Alzheimer's disease mice (tgAPPSWE ). Using on tissue and ex situ validation, TIMS MS/MS facilitated unambiguous identification of isobaric lipid species that showed plaque pathology-associated localizations. Integrated multivariate imaging data analysis revealed multiple, Aβ plaque-enriched lipid patterns for gangliosides (GM), phosphoinositols (PI), phosphoethanolamines (PE), and phosphatidic acids (PA). Conversely, sulfatides (ST), cardiolipins (CL), and polyunsaturated fatty acid (PUFA)-conjugated phosphoserines (PS), and PE were depleted at plaques. Hyperspectral amyloid imaging further delineated the unique distribution of PA and PE species to mature plaque core regions, while PI, LPI, GM2 and GM3lipids localized to immature Aβ aggregates present within the periphery of Aβ plaques. Finally, we followed AD pathology-associated lipid changes over time, identifying plaque- growth and maturation to be characterized by peripheral accumulation of PI (18:0/22:6). Together, these data demonstrate the potential of multimodal imaging approaches to overcome limitations associated with conventional advanced MS imaging applications. This allowed for the differentiation of both distinct lipid components in a complex micro-environment as well as their correlation to disease-relevant amyloid plaque polymorphs. Cover Image for this issue: https://doi.org/10.1111/jnc.15390.

Distinction of chiral penicillamine using metal-ion coupled cyclodextrin complex as chiral selector by trapped ion mobility-mass spectrometry and a structure investigation of the complexes

Penicillamine (Pen) is a common chiral drug that is obtained from penicillin. Between the two enantiomers of Pen, only D-Pen can be used to treat cystinuria and rheumatoid arthritis while L-Pen is toxic. Therefore, it requires great efforts for the research of the rigorous analysis and distinction of the two enantiomers. The non-covalent combination of chiral molecules and chiral selectors (CSs) has been proved as a unique strategy for chiral distinction by ion mobility spectrometry in coupling with -mss spectrometry (IM-MS). Here, we developed a simple method to distinguish D, L-Pen by using special CSs for IM-MS separation. The CSs utilized here include cyclodextrins (CD) and linear chain oligosaccharides plus metal ions. We found that non-covalent complexes [Pen+β-CD + Li]+ could be easily formed by electrospray ionization of the mixture of the solution, and the chirality of Pen could be effectively recognized by measuring their mobilities due to the different collision cross collision sections of [D-Pen+β-CD + Li]+ and [L-Pen+β-CD + Li]+. A detailed analysis of [Pen+β-CD + Li]+ was then conducted by the optical rotation measurements and NMR experiments to reveal their structural differences. Furthermore, DFT calculation showed the differences of molecular conformation between the complexes. The results provide a new powerful method for fast analysis and recognition of chirality of Pen compounds by IM-MS.

Complementing Matrix-Assisted Laser Desorption Ionization-Mass Spectrometry Imaging with Chromatography Data for Improved Assignment of Isobaric and Isomeric Phospholipids Utilizing Trapped Ion Mobility-Mass Spectrometry.

Lipids, such for example the multifaceted category of glycerophospholipids (GP), play a major role in many biological processes. High-resolution mass spectrometry is able to identify these highly diverse lipid species in combination with fragmentation experiments (MS/MS) on the basis of the accurate m/z and fragmentation pattern. However, for the differentiation of isomeric lipids or isobaric interferences, more elaborate separation methods are required. Especially for imaging techniques, such as matrix-assisted laser desorption/ionization (MALDI)-MS imaging, the identification is often exclusively based on the accurate m/z. Fragmentation via MS/MS increases the confidence in lipid annotation in imaging approaches. However, this is sometimes not feasible due to insufficient sensitivity and significantly prolonged analysis time. The use of a separation dimension such as trapped ion mobility spectrometry (TIMS) after ionization strengthens the confidence of the identification based on the collision cross section (CCS). Since CCS libraries are limited, a tissue-specific database was initially generated using hydrophilic interaction liquid chromatography-TIMS-MS. Using this database, the identification of isomeric lipid classes as well as isobaric interferences in a lipid class was performed using a mouse spleen sample in a workflow described in this study. Besides a CCS-based identification as an additional identification criterion for GP in general, the focus was on the distinction of the isomeric GP classes phosphatidylglycerol and bis(monoacylglycero)phosphate, as well as the differentiation of possible isobaric interferences based on the formation of adducts by MALDI-TIMS-MS imaging on a molecular level.

Automated Chiral Analysis of Amino Acids Based on Chiral Derivatization and Trapped Ion Mobility-Mass Spectrometry.

A fast and fully automated method for chiral analysis has been developed by combining a chiral derivatization approach with high-resolution trapped ion mobility separation. Although the presented approach can be potentially applied to diverse types of chiral compounds, several benchmark amino acids were used as model compounds, focusing on the smallest amino acid alanine. An autosampler with an integrated chromatography system was used for inline chiral derivatization with (S)-naproxen chloride and subsequent preseparation. Afterwards, derivatized amino acids were directly introduced into the electrospray interface of a trapped ion mobility-mass spectrometer for rapid diastereomer separation in the gas phase. This unique combination of preseparation and trapped ion mobility spectrometry separation in the negative ion mode enabled rapid chiral analysis within 3 min per sample. Furthermore, the diastereomer separation proved to be independent of alkali salts or other metal ions, offering robustness with regard to samples containing high amounts of salts. Highly sensitive detection of amino acid diastereomers was possible down to the lower nanomolar concentration range, and enantiomeric ratios could be readily determined from the recorded mobilograms with excellent reproducibility and precision. To demonstrate the general applicability of our method, alanine and other amino acids were analyzed from soy sauces and seasonings, which revealed extraordinarily high d-Ala contents of up to 99% in all samples.

Fast Online Separation and Identification of Electrochemically Generated Isomeric Oxidation Products by Trapped Ion Mobility-Mass Spectrometry.

Trapped ion mobility spectrometry (TIMS) is presented as a new and rapid method to distinguish between electrochemically generated isomeric oxidation products. Separation was performed online directly after generation and ionization of the analytes, thus providing the opportunity to detect even short-lived and reactive transformation products. The same setup enables structure elucidation based on TIMS aligned fragmentation experiments. Due to the high resolution, TIMS was able to distinguish between two isomeric transformation products of the model compound metoprolol, which only differ in the position of the hydroxylation taking place in the benzylic and aromatic positions, respectively. Using this method, the analysis time is at least five times shorter compared to conventional chromatography approaches. Consequently, TIMS may arise as a powerful tool in electrochemical metabolism studies.

Nanogymnastics: Visualization of Intercluster Reactions by High-Resolution Trapped Ion Mobility Mass Spectrometry

Although single-crystal X-ray diffraction is a proven technique to determine the structure of monolayer-protected coinage metal clusters in solid state, it is not readily applicable to the characte...

Exploring the Conformational Space of Growth-Hormone-Releasing Hormone Analogues Using Dopant Assisted Trapped Ion Mobility Spectrometry-Mass Spectrometry.

Recently, we proposed a high-throughput screening workflow for the elucidation of agonistic or antagonistic growth hormone-releasing hormone (GHRH) potencies based on structural motif descriptors as a function of the starting solution. In the present work, we revisited the influence of solution and gas-phase GHRH molecular microenvironment using trapped ion mobility-mass spectrometry (TIMS-MS). The effect of the starting solvent composition (10 mM ammonium acetate (NH4Ac), 50% methanol (MeOH), 50% acetonitrile (MeCN), and 50% acetone (Ac)) and gas-phase modifiers (N2, N2 + MeOH, N2 + MeCN, and N2 + Ac) on the conformational states of three GHRH analogues, GHRH (1-29), MR-406, and MIA-602, is described as a function of the trapping time (100-500 ms). Changes in the mobility profiles were observed showing the dependence of the conformational states of GHRH analogues according to the molecular microenvironment in solution, suggesting the presence of solution memory effects on the gas-phase observed structures. Modifying the bath gas composition resulted in smaller mobilities that are correlated with the size and mass of the organic modifier, and more importantly led to substantial changes in relative abundances of the IMS profiles. We attributed the observed changes in the mobility profiles by a clustering/declustering mechanism between the GHRH analogue ions and the gas modifiers, redefining the free energy landscape and leading to other local minima structures. Moreover, inspection of the mobility profiles as a function of the trapping time (100-500 ms) allowed for conformational interconversions toward more stable "gas-phase" structures. These experiments enabled us to outline a more detailed description of the structures and intermediates involved in the biological activity of GHRH, MR-406, and MIA-602.

Analysis of Conjugated Fatty Acid Isomers by the Paternò-Büchi Reaction and Trapped Ion Mobility Mass Spectrometry.

Fatty acids containing conjugated carbon-carbon double bonds (C═Cs), such as conjugated linoleic acids (CLAs), attract growing research interest due to their bioactivities against diabetes, cancer, and atherosclerosis. Analysis of conjugated fatty acid (CFA) is challenging for existing analytical techniques because it requires determination of geometry (cis ( Z) vs trans ( E)) and location of individual C═C. In this study, we developed a method to achieve confident, fast, and quantitative analysis of CFA isomers from mixtures. This method combines the strength of trapped ion mobility spectrometry (TIMS) for fast isomer separation and the Paternò-Büchi (PB) reaction followed by tandem mass spectrometry (MS/MS) for C═C location determination. Notably, the PB reaction of CFA is regioselective to terminal C═Cs, thus forming diagnostic fragment ions unique to conjugated C═Cs from PB-MS/MS. These fragment ions facilitate identification and quantitation of individual CLA isomers differing in C═C locations, affording limit of identification of 1 nM. Given that PB-MS/MS alone cannot identify the geometry of C═C, TIMS has been employed for characterizing C═C geometry. TIMS is capable to separate various C═C geometric isomers of CLAs, allowing visualization of C═C isomerization during the PB reaction. By coupling the PB-MS/MS with TIMS, two CLA isomers, CLA 18:2(9 Z,11 E) (46.9 ± 1.1%) and CLA 18:2(10 E,12 Z) (53.1 ± 1.1%), are quantified in a commercial CLA supplement.

Resolution of minor size differences in a family of heteroleptic coordination cages by trapped ion mobility ESI-MS.

We report a complex system of heteroleptic coordination cages based on the combination of four bis-monodentate ligands whose backbones only slightly differ in shape and length. cis-[Pd2L2L'2] assemblies cleanly form after addition of PdII cations to a 1 : 1 mixture of two shape-complementary ligands, each. When three or even all four ligands are used in combination, the unambiguous discrimination of all individual species in the product mixture becomes difficult by conventional NMR spectroscopic and mass spectrometric methods. Due to steric constraints, the system is restricted to the formation of ten different coordination cages in total, two of which are isomeric. We show that high-resolution trapped ion mobility mass spectrometry (TIMS) allows the clear differentiation of all ten species. Observed size trends could be readily reproduced by the calculation of theoretical values for collisional cross sections (CCS) from geometry-optimized models.

Altering the mobility-time continuum: nonlinear scan functions for targeted high resolution trapped ion mobility-mass spectrometry

Trapped ion mobility spectrometry (TIMS) is a versatile high resolution technique that provides the user with the flexibility to adjust the mobility range of interest, duty cycle (up to 100 %), and resolving power (up to ~300) according to the application requirements. Furthermore, TIMS offers the flexibility of operating as either a mobility-selective or conventional ion funnel, thus permitting ion mobility separations to be turned on or off. Here, we extend the flexibility of TIMS by introducing multilinear and nonlinear scanning methods that allow enhanced resolution in user-defined mobility regions. The performance of the new method is demonstrated using a variety of nonlinear scan functions that allow the resolving power to be continuously varied across the mobility spectrum. Further, we demonstrate that mobility analysis can be targeted over disparate regions using a multilinear scan function. In this example, high resolution mobility analysis is targeted on two analytes on opposite ends of a mobility range, while other ions that fall between the regions of interest remain unanalyzed. Using this approach, the resolving power for targeted species was increased by a factor of two over the conventional linear scanning approach (R ~60 versus ~120) without reducing the duty cycle of the TIMS measurement. Importantly, in such an analysis, ions in the untargeted regions are not mobility analyzed, however, they are also not discarded. Rather, these ions are ejected for downstream mass analysis. In this sense, TIMS bridges the gap between dispersive and scanning mobility techniques. That is, TIMS disperses ions according to their elution voltage, however, TIMS can also perform target mobility analyses without eliminating untargeted ions.