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SYNAPT G2-Si Mass Spectrometer


Background In recent years, mass spectrometry (MS) has been applied to clinical microbial biotyping, exploiting the speed of matrix-assisted laser desorption/ionization (MALDI) in recording microbe-specific MS profiles. More recently, liquid atmospheric pressure (AP) MALDI has been shown to produce extremely stable ion flux from homogenous samples and 'electrospray ionization (ESI)-like' multiply charged ions for larger biomolecules, whilst maintaining the benefits of traditional MALDI including high tolerance to contaminants, low analyte consumption and rapid analysis. These and other advantages of liquid AP-MALDI MS have been explored in this study to investigate its potential in microbial biotyping. Methods Genetically diverse bacterial strains were analyzed using liquid AP-MALDI MS, including clinically relevant species such as Escherichia coli, Staphylococcus aureus and Klebsiella pneumoniae. Bacterial cultures were subjected to a simple and fast extraction protocol using ethanol and formic acid. Extracts were spotted with a liquid support matrix (LSM) and analyzed using a Synapt G2-Si mass spectrometer with an in-house built AP-MALDI source. Results Each species produces a unique lipid profile in the m/z range of 400-1100, allowing species discrimination. Traditional (solid) MALDI MS produced spectra containing a high abundance of matrix-related clusters and an absence of lipid peaks. The MS profiles of the bacterial species tested form distinct clusters using principle component analysis (PCA) with a classification accuracy of 98.63% using a PCA-based prediction model. Conclusions Liquid AP-MALDI MS profiles can be sufficient to distinguish clinically relevant bacterial pathogens and other bacteria, based on their unique lipid profiles. The analysis of the lipid MS profiles is typically excluded from commercial instruments approved for clinical diagnostics.

Waters Corporation (USA) recently unveiled the new Waters SYNAPT G2-Si Mass Spectrometer at the 61st conference of the American Society of Mass Spectrometry (ASMS). The company is also previewing its UNIFI CCS Research Edition and the new TransOmics Version 2.0 informatics products for the new SYNAPT G2-Si mass spectrometer and demonstrating their benefits for a variety of small molecule applications and cutting-edge Omics-focused research.

The SYNAPT G2-Si System integrates a third dimension of resolution and separation power into a new suite of untargeted and targeted LC/MS/MS workflows. This powerful new tool gives researchers a means with which to gain a deeper understanding of molecular biology and disease mechanisms, to develop the next generation of healthcare treatments or chemical materials, or to screen food products or environmental samples for contaminants. The SYNAPT G2-Si mass spectrometer now combines the unique power of Travelling Wave (T-Wave) Ion Mobility Separations with new data acquisition and informatics technologies, and collision cross section (CCS) measurements to bring to the toughest analytical applications, unparalleled information and confidence at a level not possible by mass and chromatographic separation alone.

The SYNAPT G2-Si mass spectrometer is the first MS system to elevate CCS alongside retention time and mass to charge ratio (m/z) as a robust, reliable identification parameter in library-based screening.

As the newest member of the SYNAPT family of mass spectrometers, the SYNAPT G2-Si system can separate individual isomers, conformers, and isobaric compounds; help determine sites of biotransformation; study structure at physiological concentrations; comprehensively profile complex mixtures and minimize false positives and negatives in screening experiments.

A timed ion mobility quadrupole time-of-flight mass spectrometer enabling exceptional detection and sensitivity for quantitative proteomics workflows. Single-shot applications using fast (high-throughput) or traditional (deep coverage) nanoflow gradients. Ideal for stable isotope label (e.g. SILAC) or label-free (peptide peak area) quantitative proteomics workflows. Parallel search engine in real time (PaSER) provides comprehensive qualitative results for immediate feedback on experimental results, sample integrity, or sytems suitability.

A quadrupole-Orbitrap-linear ion trap tribrid mass spectrometer employing HCD, CID and ETD fragmentation options. Instrument is equiped with a FAIMS PRO source for atmospheric ion mobility separations providing improved analyte selectivity and expanded proteome coverage. Analysis of complex protein digests including identification of post-translational modifications, covalently bound protein-drug adducts, and chemically cross-linked peptides. Ideal for stable isotope label (e.g. TMT, SILAC) or label-free (peptide peak area) quantitative proteomics workflows.

A linear ion trap-Orbitrap hybrid mass spectrometer employing CID, HCD, and ETD fragmentation. Untargeted metabolomic and lipidomic profiling and targeted assays using HCD fragmentation using conventional flow UHPLC. Nanoflow UPLC for high-sensitivity applications including neuroactive steroids, modified nucleotides, and peptides.

Modern mass spectrometry methods provide a huge benefit to saponin structural characterization, especially when combined with collision-induced dissociation experiments to obtain a partial description of the saponin (ion) structure. However, the complete description of the structures of these ubiquitous secondary metabolites remain challenging, especially since isomeric saponins presenting small differences are often present in a single extract. As a typical example, the horse chestnut triterpene glycosides, the so-called escins, comprise isomeric saponins containing subtle differences such as cis-trans ethylenic configuration (stereoisomers) of a side chain or distinct positions of an acetyl group (regioisomers) on the aglycone. In the present paper, the coupling of liquid chromatography and ion mobility mass spectrometry has been used to distinguish regioisomeric and stereoisomeric saponins. Ion mobility arrival time distributions (ATDs) were recorded for the stereoisomeric and regioisomeric saponin ions demonstrating that isomeric saponins can be partially separated using ion mobility on a commercially available traveling wave ion mobility (TWIMS) mass spectrometer. Small differences in the ATD can only be monitored when the isomeric saponins are separated with liquid chromatography prior to the IM-MS analysis. However, gas phase separation between stereoisomeric and regioisomeric saponin ions can be successfully realized, without any LC separation, on a cyclic ion mobility-enabled quadrupole time-of-flight (Q-cIM-oaToF) mass spectrometer. The main outcome of the present paper is that the structural analysis of regioisomeric and stereoisomeric natural compounds that represents a real challenge can take huge advantages of ion mobility experiments but only if increased ion mobility resolution is attainable.

Further studies were performed on a cyclic ion mobility (cIM) enabled quadrupole time-of-flight (Q-cIM-oaToF) mass spectrometer (Waters, UK) [28, 29]. This system has a similar geometry to the SYNAPT G2-Si, with a trap cell proceeding the cIM device, followed by a transfer cell and an oaToF operating at resolutions of up to 100,000 FWHM (full width at half maximum).

The HC extract is analyzed by LC-IMS-MS on a Waters SYNAPT G2-Si mass spectrometer and the ATDs of all the saponin ions, [M+Na]+, are recorded in the positive ion mode. The saponin molecules presented in Table 1 are detected in the saponin extract and are resolved using liquid chromatography as shown in Figure 1a for the typical case of escin 1 isomers. The structural assignment of the LC peaks is achieved based upon the mass spectrometry data together with nuclear magnetic spectrometry (NMR) experiments on isolated saponins (see SI for more details). Table 1 also gathers the arrival times (tA) of the [M+Na]+ ions. As reported in a previous investigation concerning saponin ion mobility [20], the tA are quite similar for isomeric ions presenting only subtle structural differences. For instance, the tA of the [M+Na]+ ions generated from the four isomeric escin 1 saponins are measured at 9.83, 9.75, 9.97, and 9.83 ms respectively for escin 1a, escin 1b, isoescin 1a, and isoescin 1b (Table 1 and Figure 1b). The Tig-containing saponins appear less compact than the Ang-containing counterparts, 9.83 ms vs 9.75 ms (escin 1a vs escin 1b) and 9.97 vs 9.83 ms (isoescin 1a vs isoescin 1b) for the [M+Na]+ ions. The regioisomer separation of the [M+Na]+ ions of escin 1a and escin 1b are characterized by shorter tA than their isoescin 1a and isoescin 1b regioisomers. However, escin 1a and isoescin 1b that are both regioisomers and stereoisomers cannot be separated using ion mobility (Figure 1b and Table 1). The same discussion holds for the different sets of isomers as presented in Table 1.

Aesculus hippocastanum, commonly known as the horse chestnut tree or conker tree, is a well-known large tree, abundantly cultivated in streets and parks in temperate countries, especially in Western Europe. Its seeds are well known to contain saponin congeners presenting different compositions and different structures. The HC saponin extract is particularly challenging from an analytical point of view since regioisomeric and stereoisomeric saponins are present. In the present paper, it has been shown that ion mobility experiments together with liquid chromatography separation can be utilized for the structural characterization of stereoisomeric and regioisomeric saponins. Saponins presenting isomeric side chains, such as tiglic and angelic acid residues, can be distinguished by recording the ATDs, provided that high-resolution ion mobility separation is used. This was demonstrated by comparing the capabilities of the Waters SYNAPT G2-Si mass spectrometer equipped with a conventional TWIMS device to an experimental cyclic ion mobility system (cIM) setup. Based on higher ion mobility resolution due to the multi-pass IMSn experiments and versatile fragmentation/spectrum clean-up/ion manipulation capabilities, we succeeded in discriminating stereoisomeric and regioisomeric natural molecules. The present work suggests that natural product analysis will benefit enormously from improvements in ion mobility techniques, in particular increased resolution. 59ce067264

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