Explore the vast range of titles available.

Colorectal cancer (CRC) ranks as the third most frequently diagnosed cancer and the second leading cause of cancer-related deaths. The current endoscopic-based or stool-based diagnostic techniques are either highly invasive or lack sufficient sensitivity. Thus, there is a need for less invasive and more sensitive screening approaches. We, therefore, conducted a study on 64 human serum samples representing three different groups (adenocarcinoma, adenoma, and control) using cutting-edge GC×GC–LR/HR-TOFMS (comprehensive two-dimensional gas chromatography coupled with low/high-resolution time-of-flight mass spectrometry). We analyzed samples with two different specifically tailored sample preparation approaches for lipidomics (fatty acids) (25 μL serum) and metabolomics (50 μL serum). In-depth chemometric screening with supervised and unsupervised approaches and metabolic pathway analysis were applied to both datasets. A lipidomics study revealed that specific PUFA (ω-3) molecules are inversely associated with increased odds of CRC, while some PUFA (ω-6) analytes show a positive correlation. The metabolomics approach revealed downregulation of amino acids (alanine, glutamate, methionine, threonine, tyrosine, and valine) and myo-inositol in CRC, while 3-hydroxybutyrate levels were increased. This unique study provides comprehensive insight into molecular-level changes associated with CRC and allows for a comparison of the efficiency of two different analytical approaches for CRC screening using same serum samples and single instrumentation.

Mass spectrometry-based metabolomics approaches can enable detection and quantification of many thousands of metabolite features simultaneously. However, compound identification and reliable quantification are greatly complicated owing to the chemical complexity and dynamic range of the metabolome. Simultaneous quantification of many metabolites within complex mixtures can additionally be complicated by ion suppression, fragmentation and the presence of isomers. Here we present guidelines covering sample preparation, replication and randomization, quantification, recovery and recombination, ion suppression and peak misidentification, as a means to enable high-quality reporting of liquid chromatography– and gas chromatography–mass spectrometry-based metabolomics-derived data.This Perspective, from a large group of metabolomics experts, provides best practices and simplified reporting guidelines for practitioners of liquid chromatography– and gas chromatography–mass spectrometry-based metabolomics.

Huangqi Guizhi Wuwu decoction (HGWD), as a classical formula, has been used for thousands of years in China. In this work, a comprehensive strategy was proposed for characterizing the chemical profile of HGWD based on online two-dimensional hydrophilic interaction and reversed-phase liquid chromatography coupled with hybrid linear ion trap-Orbitrap mass spectrometry (online HILIC × RP-ESI/HRMS/MS n ). The compounds in HGWD were first separated by the combined use of an XBridge amide column (150 × 4.6 mm, 3.5 μm) and Accucore C18 column (50 mm × 4.6 mm, 2.6 μm). Modulation with assistant technology, including trap columns and online dilution, was optimized and developed to decrease potential analyte loss and improve the resolution of the system. Subsequently, the accurate mass was determined by high-resolution Orbitrap and MS n fragment data by a hybrid linear ion trap (LTQ). In total, 170 chemical constituents were unambiguously identified or tentatively characterized in both positive and negative ion modes. Our study demonstrated that the proposed online HILIC × RP system coupled to the LTQ-Orbitrap MS platform is an efficient analytical technique for characterizing the chemical profile of multicomponent systems.

Untargeted mass spectrometry (MS) experiments produce complex, multidimensional data that are practically impossible to investigate manually. For this reason, computational pipelines are needed to extract relevant information from raw spectral data and convert it into a more comprehensible format. Depending on the sample type and/or goal of the study, a variety of MS platforms can be used for such analysis. MZmine is an open-source software for the processing of raw spectral data generated by different MS platforms. Examples include liquid chromatography–MS, gas chromatography–MS and MS–imaging. These data might typically be associated with various applications including metabolomics and lipidomics. Moreover, the third version of the software, described herein, supports the processing of ion mobility spectrometry (IMS) data. The present protocol provides three distinct procedures to perform feature detection and annotation of untargeted MS data produced by different instrumental setups: liquid chromatography–(IMS–)MS, gas chromatography–MS and (IMS–)MS imaging. For training purposes, example datasets are provided together with configuration batch files (i.e., list of processing steps and parameters) to allow new users to easily replicate the described workflows. Depending on the number of data files and available computing resources, we anticipate this to take between 2 and 24 h for new MZmine users and nonexperts. Within each procedure, we provide a detailed description for all processing parameters together with instructions/recommendations for their optimization. The main generated outputs are represented by aligned feature tables and fragmentation spectra lists that can be used by other third-party tools for further downstream analysis. Key points MZmine is a program designed to process data from untargeted mass spectrometry (MS) experiments acquired in data-dependent acquisition mode; specifically, collision-induced dissociation and higher-energy collisional dissociation. This protocol provides three distinct procedures to perform feature detection and annotation of untargeted MS data produced by instrumental setups: liquid chromatography–(ion mobility spectrometry–)MS, gas chromatography–MS and (ion mobility spectrometry–)MS imaging. Untargeted mass spectrometry (MS) produces complex, multidimensional data. The MZmine open-source project enables processing of spectral data from various MS platforms, e.g., liquid chromatography–MS, gas chromatography–MS, MS–imaging and ion mobility spectrometry–MS, and is specialized for metabolomics.

IntroductionUrsodeoxycholic acid (UDCA) is an intestinal bacterial metabolite with hepatoprotective effects. However, molecular mechanisms underlying its effects remain unclear.ObjectivesThe aim of this study was to investigate the mechanisms underlying the therapeutic effects of UDCA by using global metabolomics analyses in healthy subjects.MethodsHealthy Korean men were administered UDCA at dosage of 400, 800, or 1200 mg daily for 2 weeks. Serum samples were collected and used for liver function tests and to determine miR-122 expression levels. Urinary and plasma global metabolomics analyses were conducted using a liquid chromatography system coupled with quadrupole-time-of-flight mass spectrometry (LC/QTOFMS) and gas chromatography-TOFMS (GC/TOFMS). Unsupervised multivariate analysis (principal component analysis) was performed to identify discriminative markers before and after treatment.ResultsAlanine transaminase score and serum miR-122 levels decreased significantly after 2 weeks of treatment. Through LC- and GC-based metabolomic profiling, we identified 40 differential metabolites in plasma and urine samples.ConclusionsRegulation of liver function scores and metabolic alternations highlight the potential hepatoprotective action of UDCA, which were primarily associated with amino acid, flavonoid, and fatty acid metabolism in healthy men.

Microcystins (MCs) are cyclic peptides produced by cyanobacteria, which can be harmful to humans and animals when ingested. Differences in the coding of the non‑ribosomal peptide synthetase/polyketide synthase enzyme complex responsible for microcystin production have resulted in more than 100 microcystin variants being reported to date. The microcystin diversity of Microcystis CAWBG11 was investigated using matrix-assisted laser desorption/ionization-time of flight mass spectrometry and liquid chromatography-mass spectrometry. This revealed that CAWBG11 simultaneously produced 21 known microcystins and six new congeners: [Asp3] MC-RA, [Asp3] MC-RAba, [Asp3] MC-FA, [Asp3] MC-WA, MC-FAba and MC-FL. The new congeners were putatively characterized by tandem mass spectrometry and chemical derivatization. A survey of the microcystin congeners produced by 49 cyanobacterial strains documented in scientific literature showed that cyanobacteria generally produce four microcystin congeners, but strains which produce up to 47 microcystin congeners have been reported. Microcystis CAWBG11 (which produces at least 27 congeners) was positioned in the top ten percentile of the strains surveyed, and showed fluidity of the amino acids incorporated into both position two and position four.

Lipidomics and metabolomics communities comprise various informatics tools; however, software programs handling multimodal mass spectrometry (MS) data with structural annotations guided by the Lipidomics Standards Initiative are limited. Here, we provide MS-DIAL 5 for in-depth lipidome structural elucidation through electron-activated dissociation (EAD)-based tandem MS and determining their molecular localization through MS imaging (MSI) data using a species/tissue-specific lipidome database containing the predicted collision-cross section values. With the optimized EAD settings using 14 eV kinetic energy, the program correctly delineated lipid structures for 96.4% of authentic standards, among which 78.0% had the sn -, OH-, and/or C = C positions correctly assigned at concentrations exceeding 1 μM. We showcased our workflow by annotating the sn - and double-bond positions of eye-specific phosphatidylcholines containing very-long-chain polyunsaturated fatty acids (VLC-PUFAs), characterized as PC n-3-VLC-PUFA/FA. Using MSI data from the eye and n-3-VLC-PUFA-supplemented HeLa cells, we identified glycerol 3-phosphate acyltransferase as an enzyme candidate responsible for incorporating n-3 VLC-PUFAs into the sn1 position of phospholipids in mammalian cells, which was confirmed using EAD-MS/MS and recombinant proteins in a cell-free system. Therefore, the MS-DIAL 5 environment, combined with optimized MS data acquisition methods, facilitates a better understanding of lipid structures and their localization, offering insights into lipid biology. Lipids are complex molecules crucial for life. The authors developed MS-DIAL 5, a tool that enhances in-depth structure elucidations and the spatial mapping of lipidome. Using advanced techniques, they uncovered new insights into the biosynthetic pathway of eye-specific phosphatidylcholines.

In polymicrobial infections, microbes can interact with both the host immune system and one another through direct contact or the secretion of metabolites, affecting disease progression and treatment options. The thick mucus in the lungs of patients with cystic fibrosis is highly susceptible to polymicrobial infections by opportunistic pathogens, including the bacterium Pseudomonas aeruginosa and the fungus Aspergillus fumigatus. Unravelling the hidden molecular interactions within such polymicrobial communities and their metabolic exchange processes will require effective enabling technologies applied to model systems. In the present study, MALDI-TOF and MALDI-FT-ICR imaging mass spectrometry (MALDI-IMS) combined with MS/MS networking were used to provide insight into the interkingdom interaction between P. aeruginosa and A. fumigatus at the molecular level. The combination of these technologies enabled the visualization and identification of metabolites secreted by these microorganisms grown on agar. A complex molecular interplay was revealed involving suppression, increased production, and biotransformation of a range of metabolites. Of particular interest is the observation that P. aeruginosa phenazine metabolites were converted by A. fumigatus into other chemical entities with alternative properties, including enhanced toxicities and the ability to induce fungal siderophores. This work highlights the capabilities of MALDI-IMS and MS/MS network analysis to study interkingdom interactions and provides insight into the complex nature of polymicrobial metabolic exchange and biotransformations.

Ubiquitin-binding proteins play an important role in eukaryotes by translating differently linked polyubiquitin chains into proper cellular responses. Current knowledge about ubiquitin-binding proteins and ubiquitin linkage-selective interactions is mostly based on case-by-case studies. We have recently reported a method called ubiquitin interactor affinity enrichment-mass spectrometry (UbIA-MS), which enables comprehensive identification of ubiquitin interactors for all ubiquitin linkages from crude cell lysates. One major strength of UbIA-MS is the fact that ubiquitin interactors are enriched from crude cell lysates, in which proteins are present at endogenous levels, contain biologically relevant post-translational modifications (PTMs) and are assembled in native protein complexes. In addition, UbIA-MS uses chemically synthesized nonhydrolyzable diubiquitin, which mimics native diubiquitin and is inert to cleavage by endogenous deubiquitinases (DUBs). Here, we present a detailed protocol for UbIA-MS that proceeds in five stages: (i) chemical synthesis of ubiquitin precursors and click chemistry for the generation of biotinylated nonhydrolyzable diubiquitin baits, (ii) in vitro affinity purification of ubiquitin interactors, (iii) on-bead interactor digestion, (iv) liquid chromatography (LC)-MS/MS analysis and (v) data analysis to identify differentially enriched proteins. The computational analysis tools are freely available as an open-source R software package, including a graphical interface. Typically, UbIA-MS allows the identification of dozens to hundreds of ubiquitin interactors from any type of cell lysate, and can be used to study cell type or stimulus-dependent ubiquitin interactions. The nonhydrolyzable diubiquitin synthesis can be completed in 3 weeks, followed by ubiquitin interactor enrichment and identification, which can be completed within another 2 weeks.

Over several millennia, various native plant species in South America have been used for their healing and psychoactive properties. Chemical analysis of archaeological artifacts provides an opportunity to study the use of psychoactive plants in the past and to better understand ancient botanical knowledge systems. Liquid chromatography tandem mass spectrometry (LC-MS/MS) was used to analyze organic residues from a ritual bundle, radiocarbon dated to approximately 1,000 C.E., recovered from archaeological excavations in a rock shelter located in the Lípez Altiplano of southwestern Bolivia. The site is located at an elevation of ∼3,900 m above sea level and contains evidence of intermittent human occupations during the last 4,000 years. Chemical traces of bufotenine, dimethyltryptamine, harmine, and cocaine, including its degradation product benzoylecgonine, were identified, suggesting that at least three plants containing these compounds were part of the shamanic paraphernalia dating back 1,000 years ago, the largest number of compounds recovered from a single artifact from this area of the world, to date. This is also a documented case of a ritual bundle containing both harmine and dimethyltryptamine, the two primary ingredients of ayahuasca. The presence of multiple plants that come from disparate and distant ecological areas in South America suggests that hallucinogenic plants moved across significant distances and that an intricate botanical knowledge was intrinsic to pre-Columbian ritual practices.