bims-metlip Biomed News
on Methods and protocols in metabolomics and lipidomics
Issue of 2020‒04‒05
forty-two papers selected by
Sofia Costa
Cold Spring Harbor Laboratory


  1. J Chromatogr B Analyt Technol Biomed Life Sci. 2020 Mar 21. pii: S1570-0232(20)30165-3. [Epub ahead of print]1144 122078
    Røst LM, Shafaei A, Fuchino K, Bruheim P.
      The pyridine nucleotides nicotineamide adenine dinucleotide (NAD) and nicotineamide adenine dinucleotide phosphate (NADP) are conserved coenzymes across all domains of life, and are involved in more than 200 different hydride transfer reactions supporting essential catabolic and anabolic functions. The intracellular levels of these metabolites, and the ratio of their oxidized to reduced forms regulate an extensive network of reactions ranging beyond metabolism. Hence, monitoring their intracellular levels provides information about, but not limited to, the metabolic state of a cell or tissue. Interconversion between oxidized and reduced forms, varying pH liability and varying intracellular concentrations of the different species leaves absolute quantification of the pyridine nucleotides analytically challenging. These polar metabolites are poorly retained on conventional reverseed-phase stationary phases without ion-pair reagents that contaminates the LC-system. Herein we demonstrate that zwitterionic HILIC-tandem mass spectroemtry can be applied to successfully resolve the pyridine nucleotides in biological extracts in a fast, robust and highly sensitive way. The presented method applies isotope dilution to compensate potential loss of these labile metabolites and is validated for low, medium and high biomass samples of two popular biological model systems; Escherichia coli and the human cell line JJN-3. High stability and rapid sample preparation without solvent removal allows for long sequence runs, making this method ideal for high-throughput analysis of biological extracts.
    Keywords:  (13)C-isotope dilution; NAD(P); Pyridine nucleotides; Tandem mass spectrometry; Zwitterionic HILIC
    DOI:  https://doi.org/10.1016/j.jchromb.2020.122078
  2. Anal Chem. 2020 Mar 30.
    Lv W, Wang L, Xuan Q, Zhao X, Liu X, Shi X, Xu G.
      Metabolite and lipid profilings usually need two liquid chromatography-mass spectrometry (LC-MS) methods due to great polarity difference. Pseudotargeted metabolomics method has emerged as a novel approach integrating the advantages of nontargeted and targeted methods. Here, we aim to establish a comprehensive method for metabolome and lipidome by using parallel column-based two-dimensional LC (PC-2DLC)-MS and pseudotargeted approach. To simultaneously extract as many polar metabolites and nonpolar lipids as possible, we systematically optimized the sample pretreatment process, and isopropanol/methanol (3:1, v/v) and isopropanol/water (7:3, v/v) were selected as the extraction and reconstitution solvents, respectively. The detected triglycerides have significantly increased after the sample pretreatment optimization. Then, PC-2DLC coupled with Triple TOF MS was applied to analyze a mixed sample from serum, urine and liver tissue matrices. The multiple reaction monitoring (MRM) transitions of metabolome and lipidome were defined according to the "MRM-Ion Pair Finder" software and lipidomics MRM-transition database, respectively. After verified by QTRAP MS in the scheduled MRM mode, 1609 potential metabolites and lipids corresponding to 1294 MRM transitions, and 847 potential metabolites and lipids corresponding to 687 MRM transitions were detected in positive and negative ion modes, respectively. They range about 30 orders of magnitude in octanol/water partition coefficient. Pseudotargeted 2DLC-MS method was validated to have good analytical characteristics. As a proof of applicability, sera from Type 2 diabetic patients were investigated by the established method. The results indicated that the pseudotargeted 2DLC-MS method is reliable, repeatable and can be used in the metabolomics study.
    DOI:  https://doi.org/10.1021/acs.analchem.0c00372
  3. J Chromatogr B Analyt Technol Biomed Life Sci. 2020 Apr 15. pii: S1570-0232(19)31713-1. [Epub ahead of print]1143 122072
    Le A, Mak J, Cowan TM.
      Metabolic profiling is commonly achieved by mass spectrometry (MS) following reversed-phase (RP) and hydrophilic interaction chromatography (HILIC) either performed independently, leading to overlapping datasets, or in a coupled configuration, requiring multiple liquid chromatography (LC) systems. To overcome these limitations, we developed a single, 20-minute chromatographic method using an in-line RP-ion-exchange (IEX) column arrangement and a single LC system. This configuration separates clinically significant polar and non-polar compounds without derivatization or ion-pairing reagents, allowing ionization in both polarities. An in-house library was created with 397 authentic standards, including acylcarnitines, amino acids, bile acids, nucleosides, organic acids, steroid hormones, and vitamins. Analysis of pooled plasma and urine samples revealed 5445 and 4111 ion features, leading to 88 and 82 confirmed metabolite identifications, respectively. Metabolites were detected at clinically relevant concentrations with good precision, and good chromatographic separation was demonstrated for clinically significant isomers including methylmalonic acid and succinic acid, as well as alloisoleucine and isoleucine/leucine. Evaluation of the samples by unsupervised principal component analysis showed excellent analytical quality.
    Keywords:  Ion-exchange; Liquid chromatography; Mass spectrometry; Metabolomics; Reversed-phase
    DOI:  https://doi.org/10.1016/j.jchromb.2020.122072
  4. Anal Chem. 2020 Apr 03.
    Xue J, Domingo-Almenara X, Guijas C, Palermo A, Rinschen M, Isbell J, Benton HP, Siuzdak G.
      Electrospray ionization (ESI) in-source fragmentation (ISF) has traditionally been minimized to promote precursor molecular ion formation, and therefore its value in molecular identification underappreciated. In-source annotation algorithms have been shown to increase confidence in putative identifications by using ubiquitous in-source fragments. However, these in-source annotation algorithms are limited by ESI sources that are generally designed to minimize ISF. In this study, enhanced in-source fragmentation annotation (eISA) was created by tuning the ISF conditions to generate in-source fragmentation patterns comparable with higher energy fragments generated at higher collision energies as deposited in the METLIN MS/MS library, without compromising the intensity of precursor ions (median loss ≤ 10% in both positive and negative ionization modes). The analysis of 50 molecules was used to validate the approach in comparison to MS/MS spectra produced via data dependent acquisition (DDA) and data independent acquisition (DIA) mode with quadrupole time-of-flight mass spectrometry (QTOF-MS). Enhanced ISF as compared to QTOF DDA, enables for higher peak intensities for the precursor ions (median: 18 times in negative mode and 210 times in positive mode), with the eISA fragmentation patterns consistent with METLIN for over 90% of the molecules with respect to fragment relative intensity and m/z. eISA also provides higher peak intensity as opposed to QTOF DIA for over 60% of the precursor ions in negative mode (median increase: 20%) and for 88% of the precursor ions in positive mode (median increase: 80%). Molecular identification with eISA was also successfully validated from the analysis of a metabolic extract from macrophages. An interesting side benefit of enhanced ISF is that it significantly improved molecular identification confidence with low resolution single quadrupole mass spectrometry-based untargeted LC/MS experiments. Overall, enhanced ISF allowed for eISA to be used as a more sensitive alternative to other QTOF DIA and DDA approaches, and further, it enables the acquisition of ESI TOF and ESI single quadrupole mass spectrometry instrumentation spectra with improved molecular identification confidence.
    DOI:  https://doi.org/10.1021/acs.analchem.0c00409
  5. Anal Bioanal Chem. 2020 Apr 02.
    Lange M, Fedorova M.
      Lipidomics analysis for large-scale studies aiming at the identification and quantification of natural lipidomes is often performed using LC-MS-based data acquisition. However, the choice of suitable LC-MS method for accurate lipid quantification remains a matter of debate. Here, we performed the systematic comparison between two HRAM-MS-based quantification workflows based on HILIC and RPLC MS by quantifying 191 lipids from five lipid classes in human blood plasma using deuterated standards in the "one ISTD-per-lipid class" approach. Lipid quantification was performed considering all necessary isotopic corrections, and obtained correction factors are illustrated. Concentrations of lipids in NIST® SRM® 1950 human blood plasma determined by the two methods were comparable for most of the studied lipid species except for highly unsaturated phosphatidylcholines (PC). A comparison of lipid concentrations to consensus values determined in a previously published multi-laboratory study illustrated possible "overestimation" of concentrations for these highly unsaturated lipids by HILIC MS. We evaluated the influence of lipid loading amounts as well as the difference between quantified lipid and internal standard concentrations on the HILIC MS quantification results. We conclude that both HILIC and RPLC HRAM-MS workflows can be equally used for accurate lysophosphatidylcholine (LPC), lysophosphatidylethanolamine (LPE), phosphatidylcholine (PC), phosphatidylethanolamine (PE), and sphingomyelin (SM) lipid quantification, despite significant differences in the concentration of highly unsaturated PC lipids which need to be addressed by establishing response factors to account for the differences in degree of lipid unsaturation. Graphical.
    Keywords:  HILIC; Human blood plasma; Lipidomics; Mass spectrometry; Quantification; RPLC; UHPLC
    DOI:  https://doi.org/10.1007/s00216-020-02576-x
  6. Clin Chem Lab Med. 2020 Mar 30. pii: /j/cclm.ahead-of-print/cclm-2020-0259/cclm-2020-0259.xml. [Epub ahead of print]
    Heaney LM.
      
    DOI:  https://doi.org/10.1515/cclm-2020-0259
  7. Mass Spectrom Rev. 2020 Mar 31.
    Züllig T, Köfeler HC.
      The boost of research output in lipidomics during the last decade is tightly linked to improved instrumentation in mass spectrometry. Associated with this trend is the shift from low resolution-toward high-resolution lipidomics platforms. This review article summarizes the state of the art in the lipidomics field with a particular focus on the merits of high mass resolution. Following some theoretical considerations on the benefits of high mass resolution in lipidomics, it starts with a historical perspective on lipid analysis by sector instruments and moves further to today's instrumental approaches, including shotgun lipidomics, liquid chromatography-mass spectrometry, matrix-assisted laser desorption ionization-time-of-flight, and imaging lipidomics. Subsequently, several data processing and data analysis software packages are critically evaluated with all their pros and cons. Finally, this article emphasizes the importance and necessity of quality standards as the field evolves from its pioneering phase into a mature and robust omics technology and lists various initiatives for improving the applicability of lipidomics. © 2020 The Authors. Mass Spectrometry Reviews published by John Wiley & Sons Ltd.
    Keywords:  chromatography; high mass resolution; lipidomics; mass spectrometry
    DOI:  https://doi.org/10.1002/mas.21627
  8. Metabolomics. 2020 Apr 03. 16(4): 46
    Cheng K, Brunius C, Fristedt R, Landberg R.
      INTRODUCTION: Consensus in sample preparation for untargeted human fecal metabolomics is lacking.OBJECTIVES: To obtain sample preparation with broad metabolite coverage for high-throughput LC-MS.
    METHODS: Extraction solvent, solvent ratio and fresh frozen-vs-lyophilized samples were evaluated by metabolite feature quality.
    RESULTS: Methanol at 5 mL per g wet feces provided a wide metabolite coverage with optimal balance between signal intensity and saturation for both fresh frozen and lyophilized samples. Lyophilization did not affect SCFA and is recommended because of convenience in normalizing to dry matter.
    CONCLUSION: The suggested sample preparation is simple, efficient and suitable for large-scale human fecal metabolomics.
    Keywords:  Fecal sample; Freeze-drying; Method optimization; Short-chain fatty acids; Untargeted fecal metabolomics; XCMS
    DOI:  https://doi.org/10.1007/s11306-020-01669-z
  9. Clin Chem Lab Med. 2020 Mar 25. pii: /j/cclm.ahead-of-print/cclm-2020-0177/cclm-2020-0177.xml. [Epub ahead of print]
    Habler K, Paal M, Vogeser M.
      Background Appropriate monitoring of tobacco smoking is extremely important in several areas of medicine, e.g. management of chronic obstructive pulmonary disease (COPD), epidemiological surveys, and allocation of heart or lung transplants. The major metabolite of nicotine is cotinine that is increasingly used as a laboratory parameter for assessing tobacco smoke exposure. Methods Creatinine and cotinine were analyzed simultaneously in urine by ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) in one run within 3 min using a biphenyl column. For quantification, the respective stable-isotope-labeled standards were used. Results Detuning and measuring a natural isotope of creatinine as precursor and product ion allowed a simultaneous quantification of creatinine and cotinine. The method revealed robust validation results. For both analytes, inaccuracy and imprecision of the quality control and external quality assessment (EQA) samples were ≤-11.1%. Conclusions One essential novelty of the method presented here is the simultaneous quantification of creatinine and cotinine covered by one analytical method. Despite the very different natural concentrations of creatinine and cotinine, this allows the immediate reporting of the cotinine-to-creatinine ratio without the need for a separate creatinine analysis.
    Keywords:  UHPLC-MS/MS; cotinine; creatinine; tobacco smoking exposure; urine
    DOI:  https://doi.org/10.1515/cclm-2020-0177
  10. Metabolites. 2020 Mar 26. pii: E126. [Epub ahead of print]10(4):
    Ten-Doménech I, Martínez-Sena T, Moreno-Torres M, Sanjuan-Herráez JD, Castell JV, Parra-Llorca A, Vento M, Quintás G, Kuligowski J.
      One of the most widely used strategies for metabolite annotation in untargeted LCMS is based on the analysis of MSn spectra acquired using data-dependent acquisition (DDA), where precursor ions are sequentially selected from MS scans based on user-selected criteria. However, the number of MSn spectra that can be acquired during a chromatogram is limited and a trade-off between analytical speed, sensitivity and coverage must be ensured. In this research, we compare four different strategies for automated MS2 DDA, which can be easily implemented in the frame of standard QA/QC workflows for untargeted LC-MS. These strategies consist of (i) DDA in the MS working range; (ii) iterated DDA split into several m/z intervals; (iii) dynamic iterated DDA of (pre)selected potentially informative features; and (iv) dynamic iterated DDA of (pre)annotated metabolic features using a reference database. Their performance was assessed using the analysis of human milk samples as model example by comparing the percentage of LC-MS features selected as the precursor ion for MS2, the number, and class of annotated features, the speed and confidence of feature annotation, and the number of LC runs required.
    Keywords:  data dependent acquisition; human milk; liquid chromatography–mass spectrometry; peak annotation
    DOI:  https://doi.org/10.3390/metabo10040126
  11. J Pharm Biomed Anal. 2020 Mar 17. pii: S0731-7085(19)33086-9. [Epub ahead of print]185 113261
    Di Gregorio E, Miolo G, Steffan A, Corona G.
      Few time-consuming bioanalytical methods are currently available for trabectedin quantification in clinical investigations. Here we present a novel, fast and sensitive method for trabectedin determination in human plasma based on hydrophilic interaction liquid chromatography and tandem mass spectrometry (HILIC-MS/MS). Plasma samples are treated with acetonitrile-0.1 % formic acid and the solvent extract is directly injected into an Acquity BEH Amide column (2.1 × 100 mm, 1.7 μm) operating in HILIC mode at 0.2 mL/min with 80:20 acetonitrile-0.1 % formic acid in water. The analyte is separated by an organic solvent gradient and quantified by an Agilent Ultivo triple quadrupole mass spectrometer operating in multiple reaction monitoring (MRM) mode. The quantitative MRM transitions were m/z 762→234 and m/z 765→234 for trabectedin and its d3-labeled derivative, respectively. The lower limit of quantification (LLOQ) was 0.01 ng/mL and the assay was linear up to 2.5 ng/mL. The intra- and inter-day relative error ranged from 1.19 % to 8.52 %, while the relative standard deviation was less than 12.35 %. The method was used to determine the pharmacokinetic profiles of trabectedin in 26 patients with soft tissue sarcoma, showing that this new HILIC-MS/MS method is suitable for use in clinical research.
    Keywords:  Cancer; HILIC; LC-MS/MS; Pharmacokinetics; Trabectedin
    DOI:  https://doi.org/10.1016/j.jpba.2020.113261
  12. Anal Chim Acta. 2020 Apr 29. pii: S0003-2670(20)30253-1. [Epub ahead of print]1108 79-88
    De Spiegeleer M, De Graeve M, Huysman S, Vanderbeke A, Van Meulebroek L, Vanhaecke L.
      Faecal metabolomics markedly emerged in clinical as well as analytical chemistry through the unveiling of aberrations in metabolic signatures as reflection of variance in gut (patho)physiology and beyond. Logistic hurdles, however, hinder the analysis of stool samples immediately following collection, inferring the need of biobanking. Yet, the optimum way of storing stool material remains to be determined, in order to conserve an accurate snapshot of the metabolome and circumvent artifacts regarding the disease and parameter(s) under observation. To address this problem, this study scrutinised the impact of freeze-thaw cycling, storage duration, temperature and aerobicity, thereby using ultra-high performance liquid chromatography-high-resolution mass spectrometry (UPLC-HRMS)-based polar metabolomics and lipidomics methodologies for faecal metabolomics. Both targeted (n > 400) and untargeted approaches were implemented to assess storage effects on individual chemical classes of metabolites as well as the faecal fingerprint. In general, recommendations are that intact stool samples should be divided into aliquots, lyophilised and stored at -80 °C for a period no longer than 18 weeks, and avoiding any freeze-thawing. The first preservation week exerted the most decisive impact regarding storage temperature, i.e. 12.1% and 6.4% of the polar metabolome experienced a shift at -20 °C and at -80 °C, respectively, whereas 8.6% and 7.9% was observed to be changed significantly for the lipidome. In addition, aside from the negligible impact of aerobicity, the polar metabolome appeared to be more dependent on the storage conditions applied compared to the lipidome, which emerged as the more stable fraction when assessing the storage duration for 25 weeks. If the interest would greatly align with particular chemical classes, such as branched-chain amino acids or short-chain fatty acids, specific storage duration recommendations are reported. The provided insights on the stability of the faecal metabolome may contribute to a more reasoned design of experiments in biomarker detection or pathway elucidation within the field of faecal metabolomics.
    Keywords:  Faecal fingerprinting; Lipidomics; Metabolomics; Sample storage; Ultra-high performance liquid chromatography-high-resolution mass spectrometry
    DOI:  https://doi.org/10.1016/j.aca.2020.02.046
  13. J Chromatogr B Analyt Technol Biomed Life Sci. 2020 Mar 19. pii: S1570-0232(20)30044-1. [Epub ahead of print]1144 122071
    Denghel H, Göen T.
      2-(2H-Benzotriazol-2-yl)-4,6-di-tert-pentylphenol (UV 328) is an ultraviolet light (UV) absorber which prolongs the stability of plastics and other organic compounds towards UV radiation. Therefore, it is frequently used as a preserving additive and may result in an exposure of consumers. Thus, an analytical method using dispersive liquid-liquid microextraction and subsequent gas chromatography tandem mass spectrometry analysis with advanced electron ionization was developed for a human biomonitoring of UV 328 exposure, which enabled the determination of UV 328 and six of its metabolites in human urine. Sample preparation and derivatization were optimized. Baseline separation of the analytes was assured by the application of a suitable temperature program. The validation resulted in limits of detection of 0.1 µg/L for all analytes, variation coefficients from 1 to 12% for precision in series and from 5 to 12% for interday precision. Furthermore, relative recovery rates between 80 and 120% were determined. Moreover, the procedure was successfully applied to urine samples of a volunteer exposed to UV 328. The method showed excellent sensitivity, repeatability and robustness for all parameters and may be applicable for studies to elucidate the metabolism and kinetics of the investigated UV absorber and for monitoring of individuals with specific exposure to UV 328.
    Keywords:  Dispersive liquid-liquid microextraction; Human biomonitoring; Metabolites; UV 328; UV absorber; Urine analysis
    DOI:  https://doi.org/10.1016/j.jchromb.2020.122071
  14. Clin Chem Lab Med. 2020 Mar 25. pii: /j/cclm.ahead-of-print/cclm-2019-0868/cclm-2019-0868.xml. [Epub ahead of print]
    Israr MZ, Bernieh D, Salzano A, Cassambai S, Yazaki Y, Suzuki T.
      Background Matrix-assisted laser desorption ionisation (MALDI) mass spectrometry (MS) has been used for more than 30 years. Compared with other analytical techniques, it offers ease of use, high throughput, robustness, cost-effectiveness, rapid analysis and sensitivity. As advantages, current clinical techniques (e.g. immunoassays) are unable to directly measure the biomarker; rather, they measure secondary signals. MALDI-MS has been extensively researched for clinical applications, and it is set for a breakthrough as a routine tool for clinical diagnostics. Content This review reports on the principles of MALDI-MS and discusses current clinical applications and the future clinical prospects for MALDI-MS. Furthermore, the review assesses the limitations currently experienced in clinical assays, the advantages and the impact of MALDI-MS to transform clinical laboratories. Summary MALDI-MS is widely used in clinical microbiology for the screening of microbial isolates; however, there is scope to apply MALDI-MS in the diagnosis, prognosis, therapeutic drug monitoring and biopsy imaging in many diseases. Outlook There is considerable potential for MALDI-MS in clinic as a tool for screening, profiling and imaging because of its high sensitivity and specificity over alternative techniques.
    Keywords:  MALDI; biomarker; clinical; disease; mass spectrometry
    DOI:  https://doi.org/10.1515/cclm-2019-0868
  15. Metabolites. 2020 Mar 31. pii: E135. [Epub ahead of print]10(4):
    Klåvus A, Kokla M, Noerman S, Koistinen VM, Tuomainen M, Zarei I, Meuronen T, Häkkinen MR, Rummukainen S, Farizah Babu A, Sallinen T, Kärkkäinen O, Paananen J, Broadhurst D, Brunius C, Hanhineva K.
      Metabolomics analysis generates vast arrays of data, necessitating comprehensive workflows involving expertise in analytics, biochemistry and bioinformatics in order to provide coherent and high-quality data that enable discovery of robust and biologically significant metabolic findings. In this protocol article, we introduce notame, an analytical workflow for non-targeted metabolic profiling approaches, utilizing liquid chromatography-mass spectrometry analysis. We provide an overview of lab protocols and statistical methods that we commonly practice for the analysis of nutritional metabolomics data. The paper is divided into three main sections: the first and second sections introducing the background and the study designs available for metabolomics research and the third section describing in detail the steps of the main methods and protocols used to produce, preprocess and statistically analyze metabolomics data and, finally, to identify and interpret the compounds that have emerged as interesting.
    Keywords:  LC–MS; computational statistical; mass spectrometry; metabolic profiling; metabolomics; pathway analysis; supervised learning; unsupervised learning
    DOI:  https://doi.org/10.3390/metabo10040135
  16. Metabolites. 2020 Mar 21. pii: E118. [Epub ahead of print]10(3):
    Jin H, Moseley HNB.
      Stable isotope resolved metabolomics (SIRM) experiments use stable isotope tracers to provide superior metabolomics datasets for metabolic flux analysis and metabolic modeling. Since assumptions of model correctness can seriously compromise interpretation of metabolic flux results, we have developed a metabolic modeling software package specifically designed for moiety model comparison and selection based on the metabolomics data provided. Here, we tested the effectiveness of model selection with two time-series mass spectrometry (MS) isotopologue datasets for uridine diphosphate N-acetyl-d-glucosamine (UDP-GlcNAc) generated from different platforms utilizing direct infusion nanoelectrospray and liquid chromatography. Analysis results demonstrate the robustness of our model selection methods by the successful selection of the optimal model from over 40 models provided. Moreover, the effects of specific optimization methods, degree of optimization, selection criteria, and specific objective functions on model selection are illustrated. Overall, these results indicate that over-optimization can lead to model selection failure, but combining multiple datasets can help control this overfitting effect. The implication is that SIRM datasets in public repositories of reasonable quality can be combined with newly acquired datasets to improve model selection. Furthermore, curation efforts of public metabolomics repositories to maintain high data quality could have a huge impact on future metabolic modeling efforts.
    Keywords:  isotopologue deconvolution; model selection; moiety modeling; nonlinear inverse problem; overfitting; stable isotope resolved metabolomics (SIRM)
    DOI:  https://doi.org/10.3390/metabo10030118
  17. Anal Chim Acta. 2020 Apr 29. pii: S0003-2670(20)30197-5. [Epub ahead of print]1108 142-151
    Khakimov B, Mobaraki N, Trimigno A, Aru V, Engelsen SB.
      Proton Nuclear Magnetic Resonance (NMR) spectroscopic analysis of urine generates rich but complex spectra. Retrieving metabolite information from such spectra is challenging due to signal overlapping, chemical shift changes, and large concentration variations of complex urine metabolome. This study demonstrates a new method, Signature Mapping (SigMa), for the rapid and efficient conversion of raw urine NMR spectra into an informative metabolite table. The principle behind SigMa relies on a division of the urine NMR spectra into Signature Signals (SS), Signals of Unknown spin Systems (SUS) and bins of complex unresolved regions (BINS). The method allows simultaneous detection of urinary metabolites in large NMR metabolomics studies using a SigMa chemical shift library and a new automatic peak picking algorithm. For quantification of SS and SUS SigMa uses multivariate curve resolution, while the unresolved inter-SS spectral regions are binned (BINS). SigMa is tested on three human urine 1H-NMR datasets including spiking experiments, and has proven to be extraordinarily efficient, quantitatively reliable and robust.
    Keywords:  MCR; Metabolomics; NMR; Signature Mapping; Urine
    DOI:  https://doi.org/10.1016/j.aca.2020.02.025
  18. Anal Bioanal Chem. 2020 Mar 31.
    Sun C, Wang F, Wang X, Geng Y, Song S, Yu Z, Liu W.
      The application of mass spectrometry imaging (MSI) for the study of spatiotemporal alterations of the metabolites in tumors has brought a number of significant biological results. At present, metabolite profiling based on MSI is typically performed on frozen tissue sections; however, the majority of clinical specimens need to be fixed in tissue fixative to avoid autolysis and to preserve antigenicity. In this study, we present the global impacts of different fixatives on the MS imaging of gastric cancer tissue metabolites. The MSI performances of 17 kinds of metabolites, such as amino acids, polyamines, cholines, organic acids, polypeptides, nucleotides, nucleosides, nitrogen bases, cholesterols, fatty acids, and phospholipids, in untreated, 10% formalin-, 4% paraformaldehyde-, acetone-, and 95% ethanol-fixed gastric cancer tissues were thoroughly explored for the first time. Furthermore, we also investigated the spatial expressions of 6 metabolic enzymes, namely, GLS, FASN, CHKA, PLD2, cPLA2, and EGFR, closely related to tumor-associated metabolites. Immunohistochemical staining carried out on the same tissue sections' which have undergone MSI analysis' suggests that enzymatic characterization is feasible after metabolite imaging. Combining the spatial signatures of metabolites and pathway-related metabolic enzymes in heterogeneous tumor tissues offers an insight to understand the complex tumor metabolism. Graphical abstract.
    Keywords:  Mass spectrometry imaging; Metabolic enzyme; Metabolites; Tissue fixative; Tumor metabolic reprogramming
    DOI:  https://doi.org/10.1007/s00216-020-02562-3
  19. J Proteome Res. 2020 Mar 31.
    Anderson JR, Phelan MM, Rubio-Martinez LM, Fitzgerald MM, Jones SW, Clegg PD, Peffers MJ.
      Synovial fluid (SF) is of great interest for the investigation of orthopaedic pathologies as it is in close proximity to various tissues which are primarily altered during these disease processes and can be collected using minimally invasive protocols. Multi 'omic' approaches are commonplace although little consideration is often given for multiple analysis techniques at sample collection. Nuclear magnetic resonance (NMR) metabolomics and liquid chromatography tandem mass spectrometry (LC-MS/MS) proteomics are two complementary techniques particularly suited to the study of SF. However, currently there are no agreed standard protocols which are published for SF collection and processing for use with NMR metabolomic analysis. Furthermore, the large protein concentration dynamic range present within SF can mask the detection of lower abundance proteins in proteomics. Whilst combinational ligand libraries (ProteoMinerTM columns) have been developed to reduce this dynamic range, their reproducibility when used in conjunction with SF, or on-bead protein digestion protocols, have yet to be investigated. Here we employ optimised protocols for the collection, processing and storage of SF for NMR metabolite analysis and LC-MS/MS proteome analysis, including a Lys-C endopeptidase digestion step prior to tryptic digestion which increased the number of protein identifications and improved reproducibility for on-bead ProteoMinerTM digestion.
    DOI:  https://doi.org/10.1021/acs.jproteome.0c00035
  20. J Sep Sci. 2020 Apr 02.
    Arena K, Cacciola F, Rigano F, Dugo P, Mondello L.
      Olea europaea, meaning "European olive", is a small tree belonging to the family Oleaceae, occurring in the Mediterranean Basin. Olive oil is an essential component of an equilibrated diet because of its nutritional value. Among micronutrients, phenolic compounds did show important beneficial effects for human health. The majority of the research studies on the phenol content are carried out by liquid chromatography combined to photodiode array and/or mass spectrometry detection; however, because of matrix complexity, one-dimensional liquid chromatography cannot be sometimes sufficient to obtain rewarding separations, requiring more advanced analytical techniques. In this work, comprehensive two-dimensional liquid chromatography, incorporating RP-Amide and C18 stationary phases, in the first and second dimension, respectively, both under reversed phase conditions, was investigated for the determination of the phenolic fraction in extra virgin olive oil samples. As far as detection is concerned, triple quadrupole mass spectrometry was employed under multi reaction monitoring mode offering superior selectivity and sensitivity. The reduction of matrix effects, when using LC × LC with respect to conventional 1D-LC, was assessed by comparing the slopes of calibration curves built from standard solutions and spiked olive oil samples. This article is protected by copyright. All rights reserved.
    Keywords:  comprehensive two-dimensional liquid chromatography; extra virgin olive oils; mass spectrometry; matrix effect; phenolic compounds
    DOI:  https://doi.org/10.1002/jssc.202000169
  21. Anal Chem. 2020 Apr 01.
    Kawai Y, Miyake Y, Hondo T, Lehmann JL, Terada K, Toyoda M.
      Simultaneous ion counting and waveform averaging implemented on a field-programmable gate array compiled with a high-speed digitizer was applied to ultra-performance liquid chromatography-time-of-flight mass spectrometric analysis of sulfa drugs. Ion counting was carried out by a "Peak Detection" (PKD) function that works together with signal averaging (AVG). Sulfadimidine (SDD) and sulfadimethoxine (SDMX) were measured in human serum (HS) model sample matrix. By using simultaneous PKD and AVG acquisition, we observed a unified calibration curve for more than three orders of magnitude of sample amounts (0.010 to 100.0 pmol). The ion count rate for the "practical" sample amounts, such as less than one pmol, was below 30%, which is suitable for PKD-based ion counting for quantitative accuracy and excellent peak identification performance. Samples containing 200 fmol or less could not be identified from the AVG waveform. Adding HS treated with acetonitrile severely suppressed the SDMX ion to less than half (58.1 %). However, a linear response was observed for chromatographic peak area for analytes calculated from PKD waveforms. Also, the mass resolving power calculated from the peak on PKD waveform was 24 % better than the corresponding AVG waveform, which also improves performance for analyte identification.
    DOI:  https://doi.org/10.1021/acs.analchem.0c00301
  22. Thorac Cancer. 2020 Mar 31.
    Tang X, Chen H, Chen G, Duan C, Fan Q, Li H, Wang Y, Li Z, Shi W, Liu Y.
      BACKGROUND: Sphingosine 1-phosphate (S1P), a bioactive lipid, has been shown to mediate cancer processes. Therefore, accurate qualitative and quantitative determination is essential. The current assay method is still cumbersome to be of practical use worldwide and the aim of this study was therefore to develop a fast, accurate, precise and efficient LC-MS/MS method for targeted analyses of S1P in serum samples.METHODS: Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is an established method used for monitoring and analyzing S1P levels in serum. We determined the level of serum S1P in 256 patients with lung cancer and 36 healthy donors, and used Spearman';s rank correlation analysis to evaluate the difference in serum S1P levels between radiotherapy and nonradiotherapy patients.
    RESULTS: Standard curves were linear over ranges of 25-600 ng/mL for S1P with correlation coefficient (r2 ) greater than 0.9996. The lower limit of quantifications (LLOQs) was 25 ng/mL. The intra- and interbatch precisions and accuracy was less than 10% for S1P. The recoveries of the method were found to be 80%-98%. Serum S1P levels in healthy donors were different from those in patients (P < 0.001). Of 256 lung cancer patients, 124 (48.4%) received radiotherapy and were identified to have concomitant low serum S1P levels (222.13 ± 48.63), whereas 132 (51.6%) who had not received radiotherapy were identified to have high levels (315.16 ± 51.06). The serum S1P levels were therefore associated with radiotherapy (Spearman's Rho = -0.653, P < 0.001).
    CONCLUSIONS: Our results indicated that this new LC-MS/MS method is rapid, sensitive, specific and reliable for the quantification of S1P levels in serum samples. The level of S1P in serum samples of patients with lung cancer who received radiotherapy was significantly lower than that in patients who did not receive radiotherapy.
    KEY POINTS: An improved method was established to quantify S1P levels in human serum by LC-MS/MS, which enabled the change in serum S1P levels in lung cancer patients to be monitored, in combination with radiotherapy, and their clinical significance to be analyzed.
    Keywords:  Correlation; LC-MS/MS; S1P; radiotherapy
    DOI:  https://doi.org/10.1111/1759-7714.13409
  23. Magn Reson Chem. 2020 05;58(5): 390-403
    Martineau E, Dumez JN, Giraudeau P.
      Nuclear magnetic resonance (NMR) is a well-known analytical technique for the analysis of complex mixtures. Its quantitative capability makes it ideally suited to metabolomics or lipidomics studies involving large sample collections of complex biological samples. To overcome the ubiquitous limitation of spectral overcrowding when recording 1D NMR spectra on such samples, the acquisition of 2D NMR spectra allows a better separation between overlapped resonances while yielding accurate quantitative data when appropriate analytical protocols are implemented. Moreover, the experiment duration can be considerably reduced by applying fast acquisition methods. Here, we describe the general workflow to acquire fast quantitative 2D NMR spectra in the "omics" context. It is illustrated on three representative and complementary experiments: UF COSY, ZF-TOCSY with nonuniform sampling, and HSQC with nonuniform sampling. After giving some details and recommendations on how to apply this protocol, its implementation in the case of targeted and untargeted metabolomics/lipidomics studies is described.
    Keywords:  2D NMR spectroscopy; fast methods; lipidomics; metabolomics; quantitative analysis; targeted; untargeted
    DOI:  https://doi.org/10.1002/mrc.4899
  24. Anticancer Res. 2020 Apr;40(4): 2406
      
  25. J Sep Sci. 2020 Mar 31.
    Jakubec P, Douša M, Nováková L.
      Nine different chiral columns based on covalently immobilized or coated tris(3,5-dimethylphenylcarbamate) cellulose and amylose have been explored. We evaluated their respective enantioselective potential including the quality of enantioseparation and qualitative characteristics of peaks. The generic screening conditions were using gradient elution from 5% to 40% organic modifier/CO2 during 3 min with about forty enantiomer pairs. Primary screening was carried out using ten different mobile phases varying in type of additives while using one representative amylose- and one cellulose-based column. The complete evaluation of all nine columns was then carried out using three best performing organic modifiers: (i) methanol + 0.1% trifluoroacetic acid + 0.1% diethylamine, (ii) isopropanol + 0.1% trifluoroacetic acid + 0.1% diethylamine, and (iii) methanol + 0.1% ammonium hydroxide. Equivalency of different columns with the same chiral selector was not confirmed. When comparing coated stationary phases, the similarity corresponded to 62 % and 63 %, for cellulose-based and 67 % and amylose-based columns. For immobilized columns the similarity was 69 % and 59 % for celluloses and amyloses, respectively. The best performing column based on success rate of enantioseparation was Chiralcel OD-3 when using methanol + 0.1% trifluoroacetic acid and 0.1% diethylamine combined additive. This article is protected by copyright. All rights reserved.
    Keywords:  chiral screening; column evaluation; enantioseparation; polysaccharide stationary phases; supercritical fluid chromatography
    DOI:  https://doi.org/10.1002/jssc.202000085
  26. Molecules. 2020 Mar 26. pii: E1514. [Epub ahead of print]25(7):
    Bąchor R, Konieczny A, Szewczuk Z.
      Kidneys play a crucial role in maintaining metabolic homeostasis in a body. Serum creatinine concentration is a simple test used as an indicator of renal function. One of the known ways of quantifying creatinine concentration is the liquid chromatography-mass spectrometry (LC-MS) method, using an isotopically labeled analog of creatinine as an internal standard. Unfortunately, such isotope-labeled analogs are expensive and their synthesis is complex. Here we demonstrate a facile preparation of deuterated analogues of creatinine, via the H/D exchange of hydrogens located at the α-carbon (α-C) of the N-methylated amino acid part, under basic conditions. The stability of retrieved isotopologues was analyzed under both neutral or acidic conditions, and the results revealed that the introduced deuterons do not undergo back-exchange. In addition, the coelution of deuterated and non-deuterated forms under acidic and neutral conditions was observed. The prepared isotopologues were successfully applied in the quantitative LC-MS analysis of urine samples, and the results demonstrated that the presented strategy is novel and inexpensive, and that the quantification correlates with the commonly used Jaffe test method.
    Keywords:  creatinine; hydrogen/deuterium exchange; isotope dilution; liquid chromatography-mass spectrometry
    DOI:  https://doi.org/10.3390/molecules25071514
  27. Molecules. 2020 Mar 31. pii: E1600. [Epub ahead of print]25(7):
    Ezzeldin E, Iqbal M, Asiri YA, Ali AA, Alam P, El-Nahhas T.
      Baricitinib, is a selective and reversible Janus kinase inhibitor, is commonly used to treat adult patients with moderately to severely active rheumatoid arthritis (RA). A fast, reproducible and sensitive method of liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the quantification of baricitinib in rat plasma has been developed. Irbersartan was used as the internal standard (IS). Baracitinib and IS were extracted from plasma by liquid-liquid extraction using a mixture of n-hexane and dichloromethane (1:1) as extracting agent. Chromatographic separation was performed using Acquity UPLC HILIC BEH 1.7 µm 2.1 × 50 mm column with the mobile phase consisting of 0.1% formic acid in acetonitrile and 20 mM ammonium acetate (pH 3) (97:3). The electrospray ionization in the positive-mode was used for sample ionization in the multiple reaction monitoring mode. Baricitinib and the IS were quantified using precursor-to-production transitions of m/z 372.15 > 251.24 and 429.69 > 207.35 for baricitinib and IS, respectively. The method was validated according to the recent FDA and EMA guidelines for bioanalytical method validation. The lower limit of quantification was 0.2 ng/mL, whereas the intra-day and inter-day accuracies of quality control (QCs) samples were ranged between 85.31% to 89.97% and 87.50% to 88.33%, respectively. Linearity, recovery, precision, and stability parameters were found to be within the acceptable range. The method was applied successfully applied in pilot pharmacokinetic studies.
    Keywords:  UPLC-MS/MS; baricitinib; irbersartan; pharmacokinetic study
    DOI:  https://doi.org/10.3390/molecules25071600
  28. J Am Soc Mass Spectrom. 2020 Apr 02.
    Chen LC.
      A new high-pressure ESI source that can be readily used for commercial API mass spectrometers in a plug-and-play manner without any modification on the ion sampling interface is introduced. The emitter can be operated at ground potential, and the positive mode electrospray is generated by applying a negative high potential to the counter electrode. A shielding electrode effectively shields the opposing electric field and improves the ion transmission. This feature facilitates the direct connection of ESI emitter to the electrically grounded components. Application of the present ion source to the high-temperature (>100 °C) capillary liquid chromatography for high-speed separation of peptide and proteins is demonstrated using a monolithic polymeric column.
    DOI:  https://doi.org/10.1021/jasms.0c00052
  29. Metabolites. 2020 Mar 28. pii: E130. [Epub ahead of print]10(4):
    Witting M.
      Genome scale metabolic models (GSMs) are a representation of the current knowledge on the metabolism of a given organism or superorganism. They group metabolites, genes, enzymes and reactions together to form a mathematical model and representation that can be used to analyze metabolic networks in silico or used for analysis of omics data. Beside correct mass and charge balance, correct structural annotation of metabolites represents an important factor for analysis of these metabolic networks. However, several metabolites in different GSMs have no or only partial structural information associated with them. Here, a new systematic nomenclature for acyl-based metabolites such as fatty acids, acyl-carnitines, acyl-coenzymes A or acyl-carrier proteins is presented. This nomenclature enables one to encode structural details in the metabolite identifiers and improves human readability of reactions. As proof of principle, it was applied to the fatty acid biosynthesis and degradation in the Caenorhabditis elegans consensus model WormJam.
    Keywords:  Caenorhabditis elegans; Genome scale metabolic networks; standardization
    DOI:  https://doi.org/10.3390/metabo10040130
  30. Magn Reson Chem. 2020 05;58(5): 427-444
    Liaghati Mobarhan Y, Soong R, Lane D, Simpson AJ.
      Traditionally, due to different hardware requirements, nuclear magnetic resonance (NMR) has developed as two separate fields: one dealing with solids, and one with solutions. Comprehensive multiphase (CMP) NMR combines all electronics and hardware (magic angle spinning [MAS], gradients, high power Radio Frequency (RF) handling, lock, susceptibility matching) into a universal probe that permits a comprehensive study of all phases (i.e., liquid, gel-like, semisolid, and solid), in intact samples. When applied in vivo, it provides unique insight into the wide array of bonds in a living system from the most mobile liquids (blood, fluids) through gels (muscle, tissues) to the most rigid (exoskeleton, shell). In this tutorial, the practical aspects of in vivo CMP NMR are discussed including: handling the organisms, rotor preparation, sample spinning, water suppression, editing experiments, and finishes with a brief look at the potential of other heteronuclei (2 H, 15 N, 19 F, 31 P) for in vivo research. The tutorial is aimed as a general resource for researchers interested in developing and applying MAS-based approaches to living organisms. Although the focus here is CMP NMR, many of the approaches can be adapted (or directly applied) using conventional high-resolution magic angle spinning, and in some cases, even standard solid-state NMR probes.
    Keywords:  Comprehensive Multiphase NMR; Daphnia magna; Hyalella azteca; In-vivo; environmental organisms; magic angle spinning; metabolomics
    DOI:  https://doi.org/10.1002/mrc.4832
  31. Expert Opin Drug Discov. 2020 Mar 31. 1-5
    Jadhav GP, Prathipati PK, Chauhan H.
      
    Keywords:  Drug discovery; Orbitrap; Raman; SPR; characterization; drug development; mass spec; new compounds; surface plasmon resonance
    DOI:  https://doi.org/10.1080/17460441.2020.1745771
  32. Plants (Basel). 2020 Mar 20. pii: E388. [Epub ahead of print]9(3):
    Saleem H, Htar TT, Naidu R, Anwar S, Zengin G, Locatelli M, Ahemad N.
      The plants of the Bougainvillea genus are widely explored regarding nutritive and medicinal purposes. In this study, dichloromethane (DCM) and methanol (MeOH) extracts of Bougainvillea glabra (Choisy.) aerial and flower parts were analyzed for high-performance liquid chromatography with photodiode array detection (HPLC-PDA), ultra-high-performance liquid chromatography-mass spectrometry (UHPLC-MS) phytochemical composition, and enzyme inhibition potential against key enzymes involved in diabetes (α-amylase), skin problems (tyrosinase), and inflammatory disorders (lipoxygenase (LOX)). HPLC-PDA quantification revealed the identification of nine different polyphenolics, amongst which both flower extracts were richest. The flower MeOH extract contained the highest amount of catechin (6.31 μg/g), gallic acid (2.39 μg/g), and rutin (1.26 μg/g). However, none of the quantified compounds were detected in the aerial DCM extract. UHPLC-MS analysis of DCM extracts revealed the tentative identification of 27 secondary metabolites, where the most common belonged to terpenoid, alkaloid, and phenolic derivatives. Similarly, for enzyme inhibition, all the extracts presented moderate activity against tyrosinase and α-amylases, whereas, for LOX, both methanolic extracts showed higher percentage inhibition compared with DCM extracts. Based on our findings, B. glabra could be regarded as a perspective starting material for designing novel pharmaceuticals.
    Keywords:  B. glabra; amylase; lipoxygenase; phenolic compounds; tyrosinase
    DOI:  https://doi.org/10.3390/plants9030388
  33. Br J Surg. 2020 Apr 04.
    Iwano T, Yoshimura K, Inoue S, Odate T, Ogata K, Funatsu S, Tanihata H, Kondo T, Ichikawa D, Takeda S.
      Probe electrospray ionization mass spectrometry (PESI-MS) is an ambient ionization-based mass spectrometry method that surpasses the original electrospray ionization technique in features such as the rapidity of analysis, simplicity of the equipment and procedure, and lower cost. This study found that the PESI-MS system with machine learning has the potential to establish a lipid-based diagnosis of breast cancer with higher accuracy, using a simpler approach. Rapid MS for breast cancer.
    DOI:  https://doi.org/10.1002/bjs.11613
  34. J Anal Toxicol. 2020 Mar 30. pii: bkaa028. [Epub ahead of print]
    Cüpper M, Dalsgaard PW, Linnet K.
      The unpredictable pharmacological and toxicological effects associated with the recreational use of new psychoactive substances (NPS) represent a threat to the public health. Analysts are constantly facing a challenge to identify these designer drugs. In this article, five seized samples were submitted for analysis using ultra-high-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-MS). To tentatively identify the NPS in the samples, the potential usage of an online mass spectral database (HighResNPS.com) was explored by searching the exact mass of the precursor ion and evaluating the fragmentation profile. This approach successfully identified a suspected candidate compound present in three of the five samples. However, conclusive identification of the remaining two was not possible, due to indistinguishable fragmentation profiles of positional isomers. Therefore, complementary analytical methodologies are of paramount importance. In light of the above, HighResNPS.com is a useful tool in presumptively identifying an NPS without a reference standard.
    Keywords:  HighResNPS.com; UHPLC–QTOF-MS; fragmentation profile; new psychoactive substances; targeted screening method
    DOI:  https://doi.org/10.1093/jat/bkaa028
  35. Chem Res Toxicol. 2020 Mar 30.
    Guo J, Turesky RJ, Tarifa A, DeCaprio AP, Cooke MS, Walmsley SJ, Villalta PW.
      Mass spectrometry-based DNA adductomics is an emerging approach for the human biomonitoring of hazardous chemicals. A mass spectral database of DNA adducts will be created for the scientific community to investigate the associations between chemical exposures, DNA damage, and disease risk.
    DOI:  https://doi.org/10.1021/acs.chemrestox.0c00031
  36. Proteomes. 2020 Mar 31. pii: E6. [Epub ahead of print]8(2):
    Freitas Leal JK, Lasonder E, Sharma V, Schiller J, Fanelli G, Rinalducci S, Brock R, Bosman G.
      Microvesicle generation is an integral part of the aging process of red blood cells in vivo and in vitro. Extensive vesiculation impairs function and survival of red blood cells after transfusion, and microvesicles contribute to transfusion reactions. The triggers and mechanisms of microvesicle generation are largely unknown. In this study, we combined morphological, immunochemical, proteomic, lipidomic, and metabolomic analyses to obtain an integrated understanding of the mechanisms underlying microvesicle generation during the storage of red blood cell concentrates. Our data indicate that changes in membrane organization, triggered by altered protein conformation, constitute the main mechanism of vesiculation, and precede changes in lipid organization. The resulting selective accumulation of membrane components in microvesicles is accompanied by the recruitment of plasma proteins involved in inflammation and coagulation. Our data may serve as a basis for further dissection of the fundamental mechanisms of red blood cell aging and vesiculation, for identifying the cause-effect relationship between blood bank storage and transfusion complications, and for assessing the role of microvesicles in pathologies affecting red blood cells.
    Keywords:  aging; lipidomics; metabolomics; proteomics; red blood cell; storage; vesicles
    DOI:  https://doi.org/10.3390/proteomes8020006
  37. ACS Nano. 2020 Apr 02.
    Thomen A, Najafinobar N, Penen F, Kay E, Upadhyay PP, Li X, Phan NTN, Malmberg P, Klarqvist M, Andersson S, Kurczy ME, Ewing AG.
      Mass spectrometry imaging is a field that promises to become a mainstream bioanalysis technology by allowing the combination of single cell imaging and subcellular quantitative analysis. The frontier of single cell imaging has advanced to the point where it is now possible to compare the chemical contents of individual organelles in terms of raw or normalized ion signal. However, to realize the full potential of this technology, it is necessary to move beyond this concept of relative quantification. Here we present a NanoSIMS imaging method which directly measures the absolute concentration of an organelle-associated, isotopically labeled, pro-drug directly from a mass spectrometry image. This is validated with a recently developed nanoelectrochemistry method for single organelles. We establish a limit of detection based on the number of isotopic labels used and the volume of the organelle of interest, also offering this calculation as a web application. This approach allows sub-cellular quantification of drugs and metabolites, an overarching and previously unmet goal in cell science and pharmaceutical development.
    DOI:  https://doi.org/10.1021/acsnano.9b09804
  38. Anal Chem. 2020 Apr 01.
    Hauck BC, Harden CS, McHugh VM.
      Ion mobility spectrometry (IMS)-based instruments have historically been accurate to, at best, ±2% of the reduced ion mobility (K0) value of the chemical of interest. Fielded IMS-based detectors that are in use for hazardous and illicit substance detection are subject to false-positive alarms because of this inaccuracy and the resulting wide alarm windows, which are required to maintain a high rate of true-positive alarms. To reduce false-positive alarm rates and improve the accuracy of any IMS-based instrument, accurate K0 values of an ion mobility reference standard need to be used for ion mobility scale calibration. However, a suitable calibrant has yet to be accurately analyzed and agreed upon by the IMS community. In this study, we have chosen five potential IMS calibrants on the basis of their rating against seven criteria for suitable standards and analyzed them as a function of drift gas temperature and humidity using an accurate ion mobility instrument. Recommendations are made herein for each potential calibrant's suitability as a standard for the wider IMS community.
    DOI:  https://doi.org/10.1021/acs.analchem.0c00859
  39. BMC Bioinformatics. 2020 Apr 03. 21(1): 130
    Buchweitz LF, Yurkovich JT, Blessing C, Kohler V, Schwarzkopf F, King ZA, Yang L, Jóhannsson F, Sigurjónsson ÓE, Rolfsson Ó, Heinrich J, Dräger A.
      BACKGROUND: New technologies have given rise to an abundance of -omics data, particularly metabolomic data. The scale of these data introduces new challenges for the interpretation and extraction of knowledge, requiring the development of innovative computational visualization methodologies. Here, we present GEM-Vis, an original method for the visualization of time-course metabolomic data within the context of metabolic network maps. We demonstrate the utility of the GEM-Vis method by examining previously published data for two cellular systems-the human platelet and erythrocyte under cold storage for use in transfusion medicine.RESULTS: The results comprise two animated videos that allow for new insights into the metabolic state of both cell types. In the case study of the platelet metabolome during storage, the new visualization technique elucidates a nicotinamide accumulation that mirrors that of hypoxanthine and might, therefore, reflect similar pathway usage. This visual analysis provides a possible explanation for why the salvage reactions in purine metabolism exhibit lower activity during the first few days of the storage period. The second case study displays drastic changes in specific erythrocyte metabolite pools at different times during storage at different temperatures.
    CONCLUSIONS: The new visualization technique GEM-Vis introduced in this article constitutes a well-suitable approach for large-scale network exploration and advances hypothesis generation. This method can be applied to any system with data and a metabolic map to promote visualization and understand physiology at the network level. More broadly, we hope that our approach will provide the blueprints for new visualizations of other longitudinal -omics data types. The supplement includes a comprehensive user's guide and links to a series of tutorial videos that explain how to prepare model and data files, and how to use the software SBMLsimulator in combination with further tools to create similar animations as highlighted in the case studies.
    Keywords:  Data visualization; Metabolism; Metabolomics; Platelet; Red blood cell
    DOI:  https://doi.org/10.1186/s12859-020-3415-z
  40. Anal Chim Acta. 2020 Apr 29. pii: S0003-2670(20)30065-9. [Epub ahead of print]1108 129-141
    Münster-Müller S, Scheid N, Zimmermann R, Pütz M.
      In this study, we utilized elemental analyser (EA) and gas-chromatography (GC) isotope ratio mass spectrometry (IRMS) and ultra-high-performance liquid chromatography coupled to mass spectrometry (UHPLC-MS) in a comprehensive profiling approach assessing the chromatographic impurity signatures and δ13C and δ15N isotope ratios of synthetic cannabinoids from police seizures and internet test purchases. Main target of this study was the highly prevalent synthetic cannabinoid MDMB-CHMICA (methyl (2S)-2-([1-(cyclohexylmethyl)-1H-indol-3-yl]formamido)-3,3-dimethylbutaoate). Overall, 61 powder and 118 herbal blend (also called "Spice-Products") samples were analysed using both analytical techniques and evaluated in a joint model to link samples from a common source. As a key finding, three agglomerates of Spice-product samples with similar dates of purchase were identified in the IRMS data, possibly representing larger shipments of MDMB-CHMICA, each produced with the same precursor material, successively delivered to the European market. The three agglomerates were refined into multiple sub-clusters based on the impurity profiling data, each representing an individual synthesis batch. One of the agglomerates identified in the IRMS data was found to consist two groups of four sub-clusters, respectively, with majorly different impurity profiles, demonstrating the necessity for both analytical techniques to extract the maximum amount of information from a limited sample pool. Additionally, 31 samples containing the recently surfaced synthetic cannabinoid Cumyl-PeGaClone (5-pentyl-2-(2-phenylpropan-2-yl)-2,5-dihydro-1H-pyrido[4,3-b]indol-1-one) were analysed for their and δ13C and δ15N isotope ratios to put the isotopic data recorded for MDMB-CHMNICA in a more global perspective. Three building blocks of precursor chemicals (indole, tert-leucine, cumylamine) potentially used for the synthesis of the two named synthetic cannabinoids were acquired from different global vendors and measured for their δ13C and δ15N isotope ratios to better understand variations in the isotopic composition of the synthetic cannabinoids and to trace their origin.
    Keywords:  Cumyl-PeGaClone; Impurity profiling; Isotope ratio mass spectrometry; MDMB-CHMICA; New psychoactive substances
    DOI:  https://doi.org/10.1016/j.aca.2020.01.029
  41. Gigascience. 2020 Apr 01. pii: giaa025. [Epub ahead of print]9(4):
    McGowan T, Johnson JE, Kumar P, Sajulga R, Mehta S, Jagtap PD, Griffin TJ.
      BACKGROUND: Proteogenomics integrates genomics, transcriptomics, and mass spectrometry (MS)-based proteomics data to identify novel protein sequences arising from gene and transcript sequence variants. Proteogenomic data analysis requires integration of disparate 'omic software tools, as well as customized tools to view and interpret results. The flexible Galaxy platform has proven valuable for proteogenomic data analysis. Here, we describe a novel Multi-omics Visualization Platform (MVP) for organizing, visualizing, and exploring proteogenomic results, adding a critically needed tool for data exploration and interpretation.FINDINGS: MVP is built as an HTML Galaxy plug-in, primarily based on JavaScript. Via the Galaxy API, MVP uses SQLite databases as input-a custom data type (mzSQLite) containing MS-based peptide identification information, a variant annotation table, and a coding sequence table. Users can interactively filter identified peptides based on sequence and data quality metrics, view annotated peptide MS data, and visualize protein-level information, along with genomic coordinates. Peptides that pass the user-defined thresholds can be sent back to Galaxy via the API for further analysis; processed data and visualizations can also be saved and shared. MVP leverages the Integrated Genomics Viewer JavaScript framework, enabling interactive visualization of peptides and corresponding transcript and genomic coding information within the MVP interface.
    CONCLUSIONS: MVP provides a powerful, extensible platform for automated, interactive visualization of proteogenomic results within the Galaxy environment, adding a unique and critically needed tool for empowering exploration and interpretation of results. The platform is extensible, providing a basis for further development of new functionalities for proteogenomic data visualization.
    Keywords:  Galaxy; Integrated Genomics Viewer; RNA-Seq; mass spectrometry; proteogenomics; proteomics; transcriptomics; visualization
    DOI:  https://doi.org/10.1093/gigascience/giaa025
  42. Anal Chem. 2020 Apr 01.
    Fülöp A, Bausbacher T, Rizzo S, Zhou Q, Gillandt H, Hopf C, Rittner M.
      Several diseases are associated with disturbed redox signaling and altered metabolism of sulfur-containing metabolites and proteins. Importantly, oxidative degradation of fresh-frozen tissues begins within the normal timescale of MALDI MSI sample preparation. As a result, analytical methods that preserve the redox state of the tissue are urgently needed for refined studies of the underlying mechanisms. Nevertheless, no derivatization strategy for free sulfhydryl groups in tissue is known for MALDI MSI. Here, we report the first derivatization reagent, (E)-2-cyano-N-(2-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)ethyl)-3-(4-hydroxyphenyl)acrylamide (CHC-Mal), for selective detection of free thiols using MALDI MSI. We per-formed in-situ derivatization of free thiol groups from thiol-containing metabolites such as glutathione and cysteine and reduced proteins such as insulin and imaged their spatial distribution in porcine and mouse xenograft tissue. Derivatization of thiol-containing metabolites with CHC-Mal for MALDI MSI was also possible when using aged tissue in the presence of excess reducing agents. Importantly, CHC-Mal-derivatized low mass-metabolites could be detected without the use of a conventional MALDI matrix.
    DOI:  https://doi.org/10.1021/acs.analchem.9b05630