DeepION: A Deep Learning-Based Low-Dimensional Representation Model of Ion Images for Mass Spectrometry Imaging.

Mass spectrometry imaging (MSI) is a high-throughput imaging technique capable of the qualitative and quantitative in situ detection of thousands of ions in biological samples. Ion image representation is a technique that produces a low-dimensional vector embedded with significant spectral and spatial information on an ion image, which further facilitates the distance-based similarity measurement for the identification of colocalized ions. However, given the low signal-to-noise ratios inherent in MSI data coupled with the scarcity of annotated data sets, achieving an effective ion image representation for each ion image remains a challenge. In this study, we propose DeepION, a novel deep learning-based method designed specifically for ion image representation, which is applied to the identification of colocalized ions and isotope ions. In DeepION, contrastive learning is introduced to ensure that the model can generate the ion image representation in a self-supervised manner without manual annotation. Since data augmentation is a crucial step in contrastive learning, a unique data augmentation strategy is designed by considering the characteristics of MSI data, such as the Poisson distribution of ion abundance and a random pattern of missing values, to generate plentiful ion image pairs for DeepION model training. Experimental results of rat brain tissue MSI show that DeepION outperforms other methods for both colocalized ion and isotope ion identification, demonstrating the effectiveness of ion image representation. The proposed model could serve as a crucial tool in the biomarker discovery and drug development of the MSI technique.

3-Acetylpyridine On-Tissue Paternò-Büchi Derivatization Enabling High Coverage Lipid C═C Location-Resolved MS Imaging in Biological Tissues.

Unsaturated lipids containing single or more carbon-carbon double bonds (C═C) within tissues are closely associated with various types of diseases. Mass spectrometry imaging (MSI) has been used to study the spatial distribution of lipid C═C location isomers in tissue sections. However, comprehensive characterization of lipid C═C location isomers using MSI remains challenging. Herein, we established an on-tissue charge-switching Paternò-Büchi (PB) derivatization method using 3-acetylpyridine (3-AP) as a reaction reagent, which can be used to detect and assign C═C location of glycerophospholipids (GPLs) as well as neutral lipids, such as fatty acids (FAs), under the same experimental workflow using matrix-assisted laser desorption/ionization (MALDI)-MSI. High coverage of mono- and poly-unsaturated C═C location isomers among various lipid classes including FA, phosphatidylcholine (PC), and sulfatide (SHexCer) in distinct regions of the mouse brain and kidney was visualized using MALDI-MS/MS imaging. This method has also been applied to map the spatial distribution of lipid C═C location isomers in the Alzheimer's disease (AD) mice model for the first time, which provides a new tool to study the relationships between the distribution of lipid structural diversity and neurodegenerative diseases.

Effective Chiral Discrimination of Amino Acids through Oligosaccharide Incorporation by Trapped Ion Mobility Spectrometry.

Chiral analysis is critical to many research fields due to different biological functions of enantiomers in living systems. Although the use of ion mobility spectrometry (IMS) has become an alternative technology in the area of chiral measurements, there is still a lack of a general chiral selector for IMS-based chiral recognition, especially for small chiral molecules. Here, a new method using oligosaccharides as the chiral selector has been developed to discriminate chiral amino acids (AAs) by trapped ion mobility spectrometry-mass spectrometry (TIMS-MS). We analyzed 21 chiral amino acids, including small molecules (e.g., alanine and cysteine). Our data showed that the use of nonreducing tetrasaccharides was effective for the separation of chiral AAs, which differentiated 21 chiral AAs without using metal ions. By further incorporating a copper ion, the separation resolution could be improved to 1.64 on average, which accounts for an additional 52% improvement on top of the already achieved separation in metal-free analysis. These results indicate that the use of tetrasaccharides is an effective strategy for the separation of AA enantiomers by TIMS. The method developed in this study may open up a new strategy for effective IMS-based chiral analysis.

Resolution enhancement of overlapping peaks of ion mobility spectrometry based on improved particle swarm optimization algorithm.

RATIONALE: Ion mobility spectrometry (IMS) is a powerful analytical tool that has been widely applied in many fields. However, the limited structural resolution of IMS results in peak overlapping in the analysis of samples with similar structures. We propose a novel method, improved particle swarm optimization (IPSO), for the separation of IMS overlapping peaks. METHODS: This method, which prevents local optimization, is used to identify the peak model coefficients of IMS. Moreover, we use the half-peak width characteristics of IMS to determine the particle position range, which eliminates impossible combinations of single peaks and reduces the difficulty of identification of coefficients. RESULTS: During a comparison in performance between IPSO and the genetic algorithm (GA), the results show that the maximum separation error of IPSO is only 1.45%, while the error of the GA is up to 17.43%. Moreover, the time consumed by IPSO is 95% less than that of the GA, and IPSO has a greater robustness under the same separation error conditions. CONCLUSIONS: The method proposed provides accurate analytical results in separating overlapping IMS peaks even in cases of severe overlaps, which greatly enhances the structural resolution of IMS.

A TIMS-TOF mass spectrometry study of disaccharides from in situ ESI derivatization with 3-pyridinylboronate.

3-Pyridinylboronate, a zwitterionic boronic acid, displayed effective in situ ESI for reversible covalent tagging of saccharides in both cation and anion modes. The ion mobilities of thus-generated ions were examined with the Bruker timsTOF fleX instrument. Nine disaccharides were examined using this method. They have identical mass-to-charge ratios, differing only in monomer compositions, regio-linkages, and anomeric configurations (α or ß). The IMS separations of the disaccharides from this method were compared with those from sodium adducts reported in the literature. The differentiation effects of this method on the disaccharide isomers were increased on average by an order of magnitude. Using this method, all the pairs of disaccharides selected from nine isomers were completely identified by comparing the mobility spectra of single-tagged and double-tagged ions.

FGF-21 biomarker detection at the sub-nanogram per mL level in human serum using normal-flow liquid chromatography/tandem mass spectrometry.

RATIONALE: Quantitative detection of the FGF-21 biomarker at the sub-nanogram per mL level in human serum has generally been achieved using nanoflow liquid chromatography/tandem mass spectrometry (LC/MS/MS) due to its high sensitivity. However, a nano-LC/MS/MS-based assay can suffer from limited reproducibility and MS signal instability making it challenging to employ it as a robust analytical method for routine clinical applications. METHODS: To tackle these limitations, parallel reaction monitoring (PRM)-based targeted protein quantification using normal-flow liquid chromatography coupled with high-resolution, accurate mass instrumentation was evaluated as a possible alternative. Different from the conventional selected reaction monitoring (SRM) using triple quadrupole MS, the proposed strategy used high-resolution orbitrap MS coupled with conventional normal-flow liquid chromatography. The primary goal of this assay development effort is to significantly improve the robustness of the LC/MS/MS-based assay while maintaining high sensitivity by the use of high-resolution MS and a large sample loading volume. RESULTS: The performance of the normal-flow LC/MS/MS assay was evaluated by using it to quantify the FGF-21 protein, a potential biomarker for non-alcoholic fatty liver disease, in serum samples. Multiple replicated PRM sample quantification results demonstrated the excellent reproducibility and operational robustness of the assay. A limit of quantification of less than 0.4 ng/mL for FGF-21 in a complex serum matrix could be achieved by using the heavy-isotope-labeled peptide technique, a result which is comparable with the sensitivity obtained using the nano-LC/SRM MS-based assay. CONCLUSIONS: The strategy offered an effective alternative to nano-LC/SRM MS for the quantification of protein biomarkers in a complex biomatrix with much improved reproducibility and operational robustness.

Janus and Amphiphilic MoS2 2D Sheets for Surface-Directed Orientational Assemblies toward Ex Vivo Dual Substrate Release.

The two-dimensional (2-D) Janus and amphiphilic molybdenum disulfide (MoS2) nanosheet with opposite optical activities on each side (amphichiral) is synthesized by modifying sandwich-like bulk MoS2 with tannic acid and cholesterol through biphasic emulsion method. This new type of amphichiral Janus MoS2 nanosheet consists of a hydrophilic and positive optical activity tannic acid side as well as a hydrophobic and negative optical activity cholesterol side thereby characterized by circular dichroism. Surface-directed orientational differentiation assemblies are performed for the as-synthesized 2D material and are characterized by contact angle, infrared spectroscopy, X-ray photoelectron, and circular dichroism spectroscopies. The amphiphilic nature of the materials is demonstrated by the pre-organization of the nanosheets on either hydrophobic or hydrophilic surfaces, providing unprecedented properties of circular dichroism signal enhancement and wettability. Selective detachment of the surface organic groups (cholesterol and tannic acid fragments) is realized by matrix-assisted laser desorption/ionisation - time-of-flight (MALDI-TOF) mass spectrometry, and the dual substrate release in tissue is detected by ex vivo mass spectrometry imaging.

Mass-to-Charge Ratio Set Matching: A Novel and Efficient Compound Identification Method.

BACKGROUND: GC-MS is a powerful tool for component analysis of unknown compounds, especially in the fields of analytical chemistry and detection of biological samples. To effectively identify compounds in GC-MS, one of the most important ways is to use a matching algorithm to compare the similarity between the reference spectrum and the query spectrum. OBJECTIVE: To propose a novel way to improve compound identification accuracy. METHODS: This article proposes a method based on an m/z set match. First of all, select the maximum m/z and the m/z corresponding to the highest peak intensity in a pre-search. Next, employ the space vector model to carry on a refined search in the remaining spectra after the pre-search. Then, distinguish stereoisomers according to the order of the G value. RESULTS: Compared with the 10 peaks and the method based on an m/z number matching pre-search, the method based on m/z set matching showed higher accuracy and fewer remaining (missing) spectra. Furthermore, the refined search which is based on the m/z set matching method possesses shorter calculation time compared with no pre-search. CONCLUSIONS: The method can reduce the remaining spectra and speed up the identification of compounds. HIGHLIGHTS: The accuracy is higher, the number of remaining spectra is less, and the computational time is shorter.

Spatially revealed perfluorooctane sulfonate-induced nephrotoxicity in mouse kidney using atmospheric pressure MALDI mass spectrometry imaging.

Perfluorooctane sulfonate (PFOS), an emerging environmental persistent pollutant, has attracted extensive attention due to its potential nephrotoxicity. However, little is known about the spatial variations of lipid metabolism associated with PFOS exposure. In this study, atmospheric pressure matrix-assisted laser desorption/ionization mass spectrometry imaging (AP-MALDI MSI) was used to reveal the spatial distributions of PFOS and its adverse effect on lipid metabolism directly in mouse kidney sections. We have observed that PFOS accumulated in the renal pelvis and outer cortex regions, with some found in the medulla and inner cortex regions. Hematoxylin and eosin (H&E) staining results also demonstrated that the accumulation of PFOS caused damage to the mouse kidney, which was consistent with AP-MALDI MSI results. Furthermore, a total of 42 lipids were shown to be significantly different in the spatial distribution patterns and variations between control and PFOS exposure mice groups, including the significant down-regulation of lyso-glycerophospholipids (Lyso-GPs), phosphatidic acids (PA), phosphatidylcholines (PC), phosphatidylethanolamines (PE), phosphatidylserines (PS) sphingomyelins (SM) and sulfatides (ST) in renal medulla or cortex region of mouse kidney sections, and remarkable up-regulation of cholesterol and phosphatidylinositols (PI) in the cortex regions of mouse kidney sections. The AP-MALDI MSI provides a new tool to explore spatial distributions and variations of the endogenous metabolites for the risk assessment of environmental pollutants.

Nickel-catalyzed enantioselective α-heteroarylation of ketones via C-F bond activation to construct all-carbon quaternary stereocenters.

Nickel-catalyzed asymmetric α-heteroarylation of ketones with fluorinated heteroarenes is reported via C-F bond activation. A series of ketones and 2-fluoropyridine derivatives with different functional groups proceed well to provide the corresponding products containing all-carbon quaternary stereocenters in good yields (up to 99% yield) and high ee values (up to 99% ee). In addition, drug molecule donepezil could also be compatible under the reaction conditions to afford late-stage diversification of pharmaceuticals.

Chiral derivatization-enabled discrimination and on-tissue detection of proteinogenic amino acids by ion mobility mass spectrometry.

The importance of chiral amino acids (AAs) in living organisms has been widely recognized since the discovery of endogenous d-AAs as potential biomarkers in several metabolic disorders. Chiral analysis by ion mobility spectrometry-mass spectrometry (IMS-MS) has the advantages of high speed and sensitivity but is still in its infancy. Here, an N α-(2,4-dinitro-5-fluorophenyl)-l-alaninamide (FDAA) derivatization is combined with trapped ion mobility spectrometry-mass spectrometry (TIMS-MS) for chiral AA analysis. For the first time, we demonstrate the simultaneous separation of 19 pairs of chiral proteinogenic AAs in a single fixed condition TIMS-MS run. The utility of this approach is presented for mouse brain extracts by direct-infusion TIMS-MS. The robust separation ability in complex biological samples was proven in matrix-assisted laser desorption/ionization (MALDI) TIMS mass spectrometry imaging (MSI) as well by directly depositing 19 pairs of chiral AAs on a tissue slide following on-tissue derivatization. In addition, endogenous chiral amino acids were also detected and distinguished. The developed methods show compelling application prospects in biomarker discovery and biological research.

Effective separation of carbohydrate isomers using metal cation and halogen anion complexes in trapped ion mobility spectrometry.

Comprehensive analysis of carbohydrates, one of the critical steps towards the fundamental understanding of biological systems, has been hindered by the identifications of their complex isomeric structures differing in monosaccharide constituents, linkage positions, and configurations. While ion mobility spectrometry-mass spectrometry (IMS-MS) based methods have shown their utilities in performing glycomics measurements, there is still a lack of analytical methods allowing effective isomeric carbohydrate separation. Here, we systematically investigated the effect of alkaline earth cations (Ca2+ and Ba2+) and halogen anions (Cl-, Br- and I-) addition on the separation of 21 pairs of isomeric oligosaccharides using trapped ion mobility spectrometry-mass spectrometry (TIMS-MS). [M + Ca + I]+ complex (M is an oligosaccharide) has shown experimentally to be the optimum complex for the separation of all 21 pairs of isomeric oligosaccharides used in this study with a calculated average resolution (r) ~ 1.82 and resolving power (R) ~ 100-150. The separation capability of this complex was further demonstrated by using mixtures of 15 oligosaccharides. The optimum [M + Ca + I]+ complex can be potentially used as an effective complex for enhancing the IMS separation resolution of isomeric carbohydrates.

Structural resolution of disaccharides through halogen anion complexation using negative trapped ion mobility spectrometry.

Carbohydrates are an indispensable part of early life evolution. The determination of their structures is a key step to analyze their critical roles in biological systems. A variation of composition, glycosidic linkage, and (or) configuration between carbohydrate isomers induces structure diversity and brings challenges for their structural determination. Ion mobility spectrometry (IMS), an emerging gas-phase ion separation technology, has been considered as a promising tool for performing carbohydrate structure elucidation. In this work, eight disaccharides were analyzed by trapped ion mobility spectrometry-mass spectrometry (TIMS-MS) in the negative ion mode as the complexed form of [M + X]-, where M = disaccharide, and X = Cl, Br, and I. As compared to the positive ion analysis of the selected disaccharide in a sodiated form, a reversal charge state provided the ability to eliminate or even reverse the collision cross section (CCS) difference between disaccharide isomers. By the combination of TIMS analysis and the calculation of density functional theory, the only observed two conformers of ions [lactulose + I]- may result from different adduction sites for an iodide anion. Based on the comparison of different halogen adducts, the [M + I]- ion form exhibited more powerful ability for isomeric disaccharide differentiation with an average resolution (RP-P) of 1.17, which results in a 34.5% improvement as compared to the corresponding chloride adducts. This result indicates that the use of negative charge states, especially the complexation of an iodide anion, could be a supplemental strategy to commonly used positive ion analysis for carbohydrate separation.

Improving glycan isomeric separation via metal ion incorporation for drift tube ion mobility-mass spectrometry.

Glycosylated proteins are an essential class of molecules playing critical roles in complex biological systems. Understanding their biological functions remains extremely difficult due to the extremely broad compositions and structure variations of glycans. Although the combination of ion mobility spectrometry and mass spectrometry (IMS-MS) has become a promising technique in glycan structure characterization and composition identification, the insufficient resolving power of most IMS-MS instruments has limited its utility in performing the comprehensive structure characterization of glycans. To mitigate the low IMS resolving power, metal ion incorporation has been employed to enhance the separation of isomeric glycans. Here, we present a systematic investigation of many different glycan-metal ion complexes in an attempt to optimize the IMS separation of different isomeric glycans. By selecting optimum glycan-metal ion complexes, partial IMS separation was realized for all the 21 isomeric glycan pairs used in the experimental study. Baseline IMS separation was achieved for 76% of these isomeric glycan pairs. The best IMS separation of isomeric glycans was achieved in some cases by incorporating multiple ions with a glycan, such as the complex [glycan + Ca + Cl]+. In addition, the well-known IMS-MS measurement trendlines, often used to identify specific compound classes, were preserved for glycans even for all the 270 glycan-metal ion complexes observed in IMS-MS spectra.

A Novel Approach of Matching Mass-to-Charge Ratio for Compound Identification in Gas Chromatography-Mass Spectrometry.

Background: Gas chromatography-mass spectrometry (GC-MS) is one of the most widely used analytical techniques for analyzing chemical or biological samples in many fields. One of the most important approaches for the identification of compound in GC-MS is to compare an experimental mass spectrum with a compound recorded in a reference spectral library through a spectrum-matching algorithm. Objective: To develop a novel method to speed up compound identification. Method: In this study, a method based on m/z matching is proposed. We selected the highest m/z values and m/z values corresponding to the largest peak intensities of a mass spectrum and stepwise modified the matching threshold (MTh) based on the principle of local optimum in the pre-search process. The performance of the approach is evaluated using the mass spectral library maintained by the National Institute of Standards and Technology as a reference library and repetitive mass spectra as query spectra. Results: Compared with two-step spectral library pre-search and "ten peaks," the method based on m/z matching has higher accuracy, smaller number of remaining (missing) spectra, and shorter computational time. Conclusions: Therefore, the method can effectively speed up compound identification. Highlights: A method based on m/z matching is proposed. The accuracy is higher, the number of remaining spectra is less, and the computational time is shorter.