Implementation of Ion Mobility Spectrometry-Based Separations in Structures for Lossless Ion Manipulations (SLIM).

Structures for Lossless Ion Manipulations (SLIM) is a powerful variant of traveling wave ion mobility spectrometry (TW-IMS) that uses a serpentine pattern of microelectrodes deposited onto printed circuit boards to achieve ultralong ion path lengths (13.5 m). Ions are propelled through SLIM platforms via arrays of TW electrodes while RF and DC electrodes provide radial confinement, establishing near lossless transmission.

Measurement and Theory of Gas-Phase Ion Mobility Shifts Resulting from Isotopomer Mass Distribution Changes.

The unanticipated discovery of recent ultra-high-resolution ion mobility spectrometry (IMS) measurements revealing that isotopomers─compounds that differ only in the isotopic substitution sites─can be separated has raised questions as to the physical basis for their separation. A study comparing IMS separations for two isotopomer sets in conjunction with theory and simulations accounting for ion rotational effects provides the first-ever prediction of rotation-mediated shifts. The simulations produce observable mobility shifts due to differences in gas-ion collision frequency and translational-to-rotational energy transfer.

Diketopiperazine Formation from FPGK ( = 1-9) Peptides: Rates of Structural Rearrangements and Mechanisms.

Peptides with penultimate proline residues undergo isomerization of the Phe-Pro peptide bond followed by spontaneous bond cleavage at the Pro-Xxx bond (where Xxx is another amino acid residue), leading to cleavage of the Pro-Xxx bond and formation of a diketopiperazine (DKP). In this paper, ion mobility spectrometry and mass spectrometry techniques were used to study the dissociation kinetics of nine peptides [Phe-Pro-Gly-Lys ( = 1-9)] in ethanol. Shorter ( = 1-3) peptides are found to be more stable than longer ( = 4-9) peptides.

AutoCCS: automated collision cross-section calculation software for ion mobility spectrometry-mass spectrometry.

Motivation: Ion mobility spectrometry (IMS) separations are increasingly used in conjunction with mass spectrometry (MS) for separation and characterization of ionized molecular species. Information obtained from IMS measurements includes the ion's collision cross section (CCS), which reflects its size and structure and constitutes a descriptor for distinguishing similar species in mixtures that cannot be separated using conventional approaches. Incorporating CCS into MS-based workflows can improve the specificity and confidence of molecular identification.

Dynamic Time-Warping Correction for Shifts in Ultrahigh Resolving Power Ion Mobility Spectrometry and Structures for Lossless Ion Manipulations.

Detection of arrival time shifts between ion mobility spectrometry (IMS) separations can limit achievable resolving power (Rp), particularly when multiple separations are summed or averaged, as commonly practiced in IMS. Such variations can be apparent in higher Rp measurements and are particularly evident in long path length traveling wave structures for lossless ion manipulations (SLIM) IMS due to their typically much longer separation times. Here, we explore data processing approaches employing single value alignment (SVA) and nonlinear dynamic time warping (DTW) to correct for variations between IMS separations, such as due to pressure fluctuations, to enable more effective spectrum summation for improving Rp and detection of low-intensity species.

Assessing Collision Cross Section Calibration Strategies for Traveling Wave-Based Ion Mobility Separations in Structures for Lossless Ion Manipulations.

The collision cross section (CCS) is an important property that aids in the structural characterization of molecules. Here, we investigated the CCS calibration accuracy with traveling wave ion mobility spectrometry (TWIMS) separations in structures for lossless ion manipulations (SLIM) using three sets of calibrants. A series of singly negatively charged phospholipids and bile acids were calibrated in nitrogen buffer gas using two different TW waveform profiles (square and sine) and amplitudes (20, 25, and 30 V).

Evaluation of Waveform Profiles for Traveling Wave Ion Mobility Separations in Structures for Lossless Ion Manipulations.

Structures for lossless ion manipulations (SLIM) have recently enabled a powerful implementation of traveling wave ion mobility spectrometry (TWIMS) for ultrahigh resolution separations; however, experimental parameters have not been optimized, and potential significant gains may be feasible. Most TWIMS separations have utilized square-shaped waveforms applied by time-dependent voltage stepping across repeating sets of electrodes, but alternative waveforms may provide further improvements to resolution. Here, we characterize five waveforms (including square and sine) in terms of their transmission efficiency, IMS resolution, and resolving power, and explore the effects of TW amplitude and speed on the performance of each.

Ion Mobility Spectrometry with High Ion Utilization Efficiency Using Traveling Wave-Based Structures for Lossless Ion Manipulations.

Ion packets introduced from gates, ion funnel traps, and other conventional ion injection mechanisms produce ion pulse widths typically around a few microseconds or less for ion mobility spectrometry (IMS)-based separations on the order of 100 milliseconds. When such ion injection techniques are coupled with ultralong path length traveling wave (TW)-based IMS separations (i.e.

Ultra-High-Resolution Ion Mobility Separations Over Extended Path Lengths and Mobility Ranges Achieved using a Multilevel Structures for Lossless Ion Manipulations Module.

Over the past few years, structures for lossless ion manipulations (SLIM) have used traveling waves (TWs) to move ions over long serpentine paths that can be further lengthened by routing the ions through multiple passages of the same path. Such SLIM "multipass" separations provide unprecedentedly high ion mobility resolving powers but are ultimately limited in their ion mobility range because of the range of mobilities spanned in a single pass; that is, higher mobility ions ultimately "overtake" and "lap" lower mobility ions that have experienced fewer passes, convoluting their arrival time distribution at the detector. To achieve ultrahigh resolution separations over broader mobility ranges, we have developed a new multilevel SLIM possessing multiple stacked serpentine paths.

Rapid and Simultaneous Characterization of Drug Conjugation in Heavy and Light Chains of a Monoclonal Antibody Revealed by High-Resolution Ion Mobility Separations in SLIM.

Antibody-drug conjugates (ADCs) have recently gained traction in the biomedical community due to their promise for human therapeutics and an alternative to chemotherapy for cancer. Crucial metrics for ADC efficacy, safety, and selectivity are their drug-antibody ratios (DARs). However, DAR characterization (i.

Substance P in Solution: Trans-to-Cis Configurational Changes of Penultimate Prolines Initiate Non-enzymatic Peptide Bond Cleavages.

We report ion mobility spectrometry and mass spectrometry studies of the non-enzymatic step-by-step degradation of substance P (subP), an 11-residue neuropeptide, with the sequence Arg-Pro-Lys-Pro-Gln-Gln-Phe-Phe-Gly-Leu-Met-NH, in ethanol. At elevated solution temperatures (55 to 75 °C), several reactions are observed, including a protonation event, i.e.

Substance P in the Gas Phase: Conformational Changes and Dissociations Induced by Collisional Activation in a Drift Tube.

The work presented below is related to our companion paper in this issue, entitled: Substance P in solution: trans-to-cis configurational changes of penultimate prolines initiate non-enzymatic peptide bond cleavages. Two-dimensional ion mobility spectrometry (IMS-IMS) and mass spectrometry techniques are used to investigate structural transitions for [M+3H] ions of substance P (subP) upon collisional activation (CA) in the gas phase. In this approach, different conformations of ions having a specified mobility are selected after an initial IMS separation, collisionally activated to produce new conformers, and these product structures are separated again using a second IMS region.

Monitoring the stabilities of a mixture of peptides by mass-spectrometry-based techniques.

Biomolecular degradation plays a key role in proteostasis. Typically, proteolytic enzymes degrade proteins into smaller peptides by breaking amino acid bonds between specific residues. Cleavage around proline residues is often missed and requires highly specific enzymes for peptide processing due to the cyclic proline side-chain.

Conformationally Regulated Peptide Bond Cleavage in Bradykinin.

Ion mobility and mass spectrometry techniques are used to investigate the stabilities of different conformations of bradykinin (BK, Arg-Pro-Pro-Gly-Phe-Ser-Pro-Phe-Arg). At elevated solution temperatures, we observe a slow protonation reaction, i.e.

Melting proteins confined in nanodroplets with 10.6 μm light provides clues about early steps of denaturation.

Ubiquitin confined within nanodroplets was irradiated with a variable-power CO2 laser. Mass spectrometry analysis shows evidence for a protein "melting"-like transition within droplets prior to solvent evaporation and ion formation. Ion mobility spectrometry reveals that structures associated with early steps of denaturation are trapped because of short droplet lifetimes.

Selected overtone mobility spectrometry.

A new means of acquiring overtone mobility spectrometry (OMS) data sets that allows distributions of ions for a prescribed overtone number is described. In this approach, the drift fields applied to specific OMS drift regions are varied to make it possible to select different ions from a specific overtone that is resonant over a range of applied frequencies. This is accomplished by applying different fields for fixed ratios of time while scanning the applied frequency.