Benchmarking breath analysis using peppermint approach with gas chromatography ion mobility spectrometer coupled to micro thermal desorber.

The Peppermint Initiative, established within the International Association of Breath Research, introduced the peppermint protocol, a breath analysis benchmarking effort designed to address the lack of inter-comparability of outcomes across different breath sampling techniques and analytical platforms. Benchmarking with gas chromatography-ion mobility spectrometry (GC-IMS) using peppermint has been previously reported however, coupling micro-thermal desorption (µTD) to GC-IMS has not yet, been benchmarked for breath analysis. To benchmarkµTD-GC-IMS for breath analysis using the peppermint protocol. Ten healthy participants (4 males and 6 females, aged 20-73 years), were enrolled to give six breath samples into Nalophan bags via a modified peppermint protocol. Breath sampling after peppermint ingestion occurred over 6 h att= 60, 120, 200, 280, and 360 min. The breath samples (120 cm3) were pre-concentrated in theµTD before being transferred into the GC-IMS for detection. Data was processed using VOCal, including background subtractions, peak volume measurements, and room air assessment. During peppermint washout, eucalyptol showed the highest change in concentration levels, followed byα-pinene andß-pinene. The reproducibility of the technique for breath analysis was demonstrated by constructing logarithmic washout curves, with the average linearity coefficient ofR2= 0.99. The time to baseline (benchmark) value for the eucalyptol washout was 1111 min (95% CI: 529-1693 min), obtained by extrapolating the average logarithmic washout curve. The study demonstrated thatµTD-GC-IMS is reproducible and suitable technique for breath analysis, with benchmark values for eucalyptol comparable to the gold standard GC-MS.

Travelling Wave Ion Mobility-Derived Collision Cross Section for Mycotoxins: Investigating Interlaboratory and Interplatform Reproducibility.

Parent and modified mycotoxin analysis remains a challenge because of their chemical diversity, the presence of isomeric forms, and the lack of analytical standards. The creation and application of a collision cross section (CCS) database for mycotoxins may bring new opportunities to overcome these analytical challenges. However, it is still an open question whether common CCS databases can be used independently from the instrument type and ion mobility mass spectrometry (IM-MS) technologies, which utilize different methodologies for determining the gas-phase mobility. Here, we demonstrated the reproducibility of CCS measurements for mycotoxins in an interlaboratory study (average RSD 0.14% ± 0.079) and across different traveling wave IM-MS (TWIMS) systems commercially available (ΔCCS% < 2). The separation in the drift time dimension of critical pairs of isomers for modified mycotoxins was also achieved. In addition, the comparison of measured and predicted CCS values, including regulated and emerging mycotoxins, was addressed.

Using SLIM-Based IMS-IMS Together with Cryogenic Infrared Spectroscopy for Glycan Analysis.

The isomeric heterogeneity of glycans poses a great challenge for their analysis. While combining ion mobility spectrometry (IMS) with tandem mass spectrometry is a powerful means for identifying and characterizing glycans, it has difficulty distinguishing the subtlest differences between isomers. Cryogenic infrared spectroscopy provides an additional dimension for glycan identification that is extremely sensitive to their structure. Our approach to glycan analysis combines ultrahigh-resolution IMS-IMS using structures for lossless ion manipulation (SLIM) with cryogenic infrared spectroscopy. We present here the design of a SLIM board containing a series of on-board traps in which we perform collision-induced dissociation (CID) at pressures in the millibar range. We characterize the on-board CID process by comparing the fragments generated from a pentapeptide to those obtained on a commercial tandem mass spectrometer. We then apply our new technique to study the mobility and vibrational spectra of CID fragments from two human milk oligosaccharides. Comparison of both the fragment drift times and IR spectra with those of suitable reference compounds allows us to identify their specific isomeric form, including the anomericity of the glycosidic linkage, demonstrating the power of this tool for glycan analysis.

Sensing Precursors of Illegal Drugs-Rapid Detection of Acetic Anhydride Vapors at Trace Levels Using Photoionization Detection and Ion Mobility Spectrometry.

Sensitive real-time detection of vapors produced by the precursors, reagents and solvents used in the illegal drugs manufacture represents a priority nowadays. Acetic anhydride (AA) is the key chemical used as acetylation agent in producing the illegal drugs heroin and methaqualone. This study was directed towards quick detection and quantification of AA in air, using two fast and very sensitive analytical techniques: photoionization detection (PID) and ion mobility spectrometry (IMS). Results obtained indicated that both PID and IMS can sense AA at ultra-trace levels in air, but while PID produces a non-selective response, IMS offers richer information. Ion mobility spectrometric response in the positive ion mode presented one product ion, at reduced ion mobility K0 of 1.89 cm2 V-1 s-1 (almost overlapped with positive reactant ion peak), while in the negative ion mode two well separated product ions, with K0 of 1.90 and 1.71 cm2 V-1 s-1, were noticed. Our study showed that by using a portable, commercial IMS system (model Mini IMS, I.U.T. GmbH Berlin) AA can be easily measured at concentrations of 0.05 ppmv (0.2 mg m-3) in negative ion mode. Best selectivity and sensitivity of the IMS response were therefore achieved in the negative operation mode.

Mass Spectrometry Characterization of Higher Order Structural Changes Associated with the Fc-glycan Structure of the NISTmAb Reference Material, RM 8761.

As monoclonal antibodies (mAbs) rapidly emerge as a dominant class of therapeutics, so does the need for suitable analytical technologies to monitor for changes in protein higher order structure (HOS) of these biomolecules. Reference materials (RM) serve a key analytical purpose of benchmarking the suitability and robustness of both established and emerging analytical procedures for both drug producers and regulators. Here, two simple enzymatic protocols for generating Fc-glycan variants from the NISTmAb RM are described and both global and localized changes in HOS between the RM and these Fc-glycan variants are characterized using hydrogen deuterium exchange-mass spectrometry (HDX-MS) and ion mobility spectrometry-mass spectrometry (IMS-MS) measurements. An alternative statistical approach is described where measurement thresholds that differentiate between measurement variability and significant structural changes were established on the basis of experimental data. Measurements revealed decreases in structural stability correlating with the degree of Fc-glycan structure loss, especially at the CH2/CH3 domain interface. These data promote the use of this RM and these Fc-glycan variants for establishing the sensitivity of and validating analytical methods for the detection of HOS measurements of mAbs.

Quantitation of Cyclosporin A in Cell Culture Media by Differential Mobility Mass Spectrometry (DMS-MS/MS).

Cell permeability is an important factor in determining the bioavailability of therapeutics that is usually measured by cell culture testing. The concentration of pharmaceutical in a medium such as Hank's Balanced Salt Solution with HEPES organic buffer (HBSS-HEPES) is measured at a series of time points, making simplicity and high throughput of the analytical method important characteristics. We report an electrospray differential mobility spectrometry mass spectrometry method (nanoESI-DMS-MS) for the rapid determination of cyclosporin A (CsA, cyclosporine) concentration in such a buffer. DMS technology provides gas phase atmospheric pressure ion filtration for small-molecule bioanalytical methods that suppresses interfering ions and reduces chemical noise, without the use of chromatography. This allows simplified sample preparation, fast calibration curve development, and shortened analysis times. It has also been noted that the DMS prefilter can reduce contamination of the mass spectrometer by salts, thereby extending mass spectrometer system uptime.In the application described here, DMS-MS/MS is applied to cyclosporine A (CsA) in cell medium. Sample preparation is limited to dilution with an ammonium acetate-methanol-water mobile phase and the addition of CsA-d4 internal standard. The isotope ratio data are obtained in DMS-MS MRM mode observing NH3 loss from the ammonium adduct of the two species. A calibration curve with high linearity (R2 = 0.998) is rapidly obtained with nearly zero intercept, while it was found that a liquid chromatography LC-MS method required a preliminary SPE step to obtain a linear calibration curve. The time for data acquisition in the DMS-MS MRM method with flow injection (FIA) or infusion introduction at ESI flow of 400 nL/min is typically 30 s leading to a cycle time of less than 1 min.

DMS as an orthogonal separation to LC/ESI/MS/MS for quantifying isomeric cerebrosides in plasma and cerebrospinal fluid.

Cerebrosides, including glucosylceramides (GlcCers) and galactosylceramides (GalCers), are important membrane components of animal cells with deficiencies resulting in devastating lysosomal storage disorders. Their quantification is essential for disease diagnosis and a better understanding of disease mechanisms. The simultaneous quantification of GlcCer and GalCer isomers is, however, particularly challenging due to their virtually identical structures. To address this challenge, we developed a new LC/MS-based method using differential ion mobility spectrometry (DMS) capable of rapidly and reproducibly separating and quantifying isomeric cerebrosides in a single run. We show that this LC/ESI/DMS/MS/MS method exhibits robust quantitative performance within an analyte concentration range of 2.8-355 nM. We further report the simultaneous quantification of nine GlcCers (16:0, 18:0, 20:0, 22:0, 23:0, 24:1, 24:0, 25:0, and 26:0) and five GalCers (16:0, 22:0, 23:0, 24:1, and 24:0) molecular species in human plasma, as well as six GalCers (18:0, 22:0, 23:0, 24:1, 24:0 and 25:0) and two GlcCers (24:1 and 24:0) in human cerebrospinal fluid. Our method expands the potential of DMS technology in the field of glycosphingolipid analysis for both biomarker discovery and drug screening by enabling the unambiguous assignment and quantification of cerebroside lipid species in biological samples.

Aptamer-modified carbon nanomaterial based sorption coupled to paper spray ion mobility spectrometry for highly sensitive and selective determination of methamphetamine.

A cellulose paper was modified with an aptamer against methamphetamine on either carbon dots (CDs) or on multichannel carbon nanotubes (CNTs). The resulting sorbent was applied to the extraction of METH from blood or saliva. The METH-loaded paper than also was directly applied as a paper spray ionization source in ion mobility spectrometry. The carbon nanomaterial enhances sensitivity, and the aptamer enhances selectivity. The materials were covalently bound to the paper on one side, while the aptamer was immobilized on the other. After optimization of the extraction process and instrumental parameters, the limits of detection when using the aptamer-CNT modified paper are 0.6 ng·mL-1 for saliva, and 0.45 ng·mL-1 for plasma. The respective values when using aptamer-CD modified paper are 1.5 ng·mL-1 for saliva and 0.9 ng·mL-1 for plasma. Calibration plots are linear in the 2 to 150 ng·mL-1 METH concentration range for saliva, and in the 1.5 to 200 ng·mL-1 concentration ranges for blood when using the aptamer-CNT based method. When using the aptamer-CDs, the dynamic ranges extend from 5 to 200 ng·mL-1 and from 3 to 250 ng·mL-1, respectively. The method was applied to the determination of METH in real samples of saliva and blood, and the accuracy of the method was confirmed by comparison of the results with data analyzed by GC-MS. Graphical abstract ᅟ.

Evaluation of ion mobility for the separation of glycoconjugate isomers due to different types of sialic acid linkage, at the intact glycoprotein, glycopeptide and glycan level.

The study of protein glycosylation can be regarded as an intricate but very important task, making glycomics one of the most challenging and interesting, albeit under-researched, type of "omics" science. Complexity escalates remarkably when considering that carbohydrates can form severely branched structures with many different constituents, which often leads to the formation of multiple isomers. In this regard, ion mobility (IM) spectrometry has recently demonstrated its power for the separation of isomeric compounds. In the present work, the potential of traveling wave IM (TWIMS) for the separation of isomeric glycoconjugates was evaluated, using mouse transferrin (mTf) as model glycoprotein. Particularly, we aim to assess the performance of this platform for the separation of isomeric glycoconjugates due to the type of sialic acid linkage, at the intact glycoprotein, glycopeptide and glycan level. Straightforward separation of isomers was achieved with the analysis of released glycans, as opposed to the glycopeptides which showed a more complex pattern. Finally, the developed methodology was applied to serum samples of mice, to investigate its robustness when analyzing real complex samples. BIOLOGICAL SIGNIFICANCE: Ion mobility mass spectrometry is a promising analytical technique for the separation of glycoconjugate isomers due to type of sialic acid linkage. The impact of such a small modification in the glycan structure is more evident in smaller analytes, reason why the analysis of free glycans was easier compared to the intact protein or the glycopeptides. The established methodology could be regarded as starting point in the separation of highly decorated glycoconjugates. This is an important topic nowadays, as differences in the abundance of some glycan isomers could be the key for the early diagnosis, control or differentiation of certain diseases, such as inflammation or cancer.

Correlating Resolving Power, Resolution, and Collision Cross Section: Unifying Cross-Platform Assessment of Separation Efficiency in Ion Mobility Spectrometry.

Here we examine the relationship among resolving power (Rp), resolution (Rpp), and collision cross section (CCS) for compounds analyzed in previous ion mobility (IM) experiments representing a wide variety of instrument platforms and IM techniques. Our previous work indicated these three variables effectively describe and predict separation efficiency for drift tube ion mobility spectrometry experiments. In this work, we seek to determine if our previous findings are a general reflection of IM behavior that can be applied to various instrument platforms and mobility techniques. Results suggest IM distributions are well characterized by a Gaussian model and separation efficiency can be predicted on the basis of the empirical difference in the gas-phase CCS and a CCS-based resolving power definition (CCS/ΔCCS). Notably traveling wave (TWIMS) was found to operate at resolutions substantially higher than a single-peak resolving power suggested. When a CCS-based Rp definition was utilized, TWIMS was found to operate at a resolving power between 40 and 50, confirming the previous observations by Giles and co-workers. After the separation axis (and corresponding resolving power) is converted to cross section space, it is possible to effectively predict separation behavior for all mobility techniques evaluated (i.e., uniform field, trapped ion mobility, traveling wave, cyclic, and overtone instruments) using the equations described in this work. Finally, we are able to establish for the first time that the current state-of-the-art ion mobility separations benchmark at a CCS-based resolving power of >300 that is sufficient to differentiate analyte ions with CCS differences as small as 0.5%.