Multiple Adducts Research Articles

DNA Adductomics: A Narrative Review of Its Development, Applications, and Future.

DNA adductomics is the global study of all DNA adducts and was first proposed in 2006 by the Matsuda group. Its development has been greatly credited to the advances in mass spectrometric techniques, particularly tandem and multiple-stage mass spectrometry. In fact, liquid chromatography-mass spectrometry (LC-MS)-based methods are virtually the sole technique with practicality for DNA adductomic studies to date. At present, DNA adductomics is primarily used as a tool to search for DNA adducts, known and unknown, providing evidence for exposure to exogenous genotoxins and/or for the molecular mechanisms of their genotoxicity. Some DNA adducts discovered in this way have the potential to predict cancer risks and/or to be associated with adverse health outcomes. DNA adductomics has been successfully used to identify and determine exogenous carcinogens that may contribute to the etiology of certain cancers, including bacterial genotoxins and an N-nitrosamine. Also using the DNA adductomic approach, multiple DNA adducts have been observed to show age dependence and may serve as aging biomarkers. These achievements highlight the capability and power of DNA adductomics in the studies of medicine, biological science, and environmental science. Nonetheless, DNA adductomics is still in its infancy, and great advances are expected in the future.

GlyCombo Enables Rapid, Complete Glycan Composition Identification across Diverse Glycomic Sample Types.

Glycans are sugar-based polymers found to modify biomolecules, including lipids and proteins, as well as occur unconjugated as free polysaccharides. Due to their ubiquitous cellular presentation, glycans mediate crucial biological processes and are frequently sought after as biomarkers for a wide range of diseases. Identification of glycans present in samples acquired with mass spectrometry (MS) is a cornerstone of glycomics research; thus, the ability to rapidly identify glycans in each acquisition is integral to glycomics analysis pipelines. Here we introduce GlyCombo (https://github.com/Protea-Glycosciences/GlyCombo), an open-source, freely available software tool designed to rapidly assign monosaccharide combinations to glycan precursor masses including those subjected to MS2 in LC-MS/MS experiments. GlyCombo was evaluated across six diverse data sets, demonstrating MS vendor, derivatization, and glycan-type neutrality. Compositional assignments using GlyCombo are shown to be faster than the current predominant approach, GlycoMod, a closed-source web application. Two unique features of GlyCombo, multiple adduct search and off-by-one error anticipation, reduced unassigned MS2 scans in a benchmark data set by 40%. Finally, the comprehensiveness of glycan feature identification is exhibited in Skyline, a software that requires predefined transitions that are derived from GlyCombo output files.

MARS: A Multipurpose Software for Untargeted LC-MS-Based Metabolomics and Exposomics.

Untargeted metabolomics is a growing field, in which recent advances in high-resolution mass spectrometry coupled with liquid chromatography (LC-MS) have facilitated untargeted approaches as a result of improvements in sensitivity, mass accuracy, and resolving power. However, a very large amount of data are generated. Consequently, using computational tools is now mandatory for the in-depth analysis of untargeted metabolomics data. This article describes MetAbolomics ReSearch (MARS), an all-in-one vendor-agnostic graphical user interface-based software applying LC-MS analysis to untargeted metabolomics. All of the analytical steps are described (from instrument data conversion and processing to statistical analysis, annotation/identification, quantification, and preliminary biological interpretation), and tools developed to improve annotation accuracy (e.g., multiple adducts and in-source fragmentation detection, trends across samples, and the MS/MS validator) are highlighted. In addition, MARS allows in-house building of reference databases, to bypass the limits of freely available MS/MS spectra collections. Focusing on the flexibility of the software and its user-friendliness, which are two important features in multipurpose software, MARS could provide new perspectives in untargeted metabolomics data analysis.

Rapid characterization and identification of non-volatile constituents in Pogostemonis Herba by UPLC-Q-TOF-MS combined with UNIFI and an in-house library

This study aimed to characterize and identify the non-volatile components in Pogostemonis Herba by using ultra-perfor-mance liquid chromatography-quadrupole-time of flight-mass spectrometry(UPLC-Q-TOF-MS) combined with UNIFI and an in-house library. The chemical components in 50% methanol extract of Pogostemonis Herba were detected by UPLC-Q-TOF-MS in both positive and negative MS~E continuum modes. Then, the MS data were processed in UNIFI combined with an in-house library to automatically characterize the metabolites. Based on the multiple adduct ions, exact mass, diagnostic fragment ions, and peak intensity of compounds and the fragmentation pathways and retention behaviors of reference substances, the structures identified by UNIFI were further verified and those of the unidentified compounds were tentatively elucidated. A total of 120 compound structures were identified or tentatively identified, including flavonoids, phenylpropanoids, phenolic acids, terpenes, fatty acids, alkaloids, and phenylethanoid glycosides. Sixteen of them were accurately identified by comparison with reference substances, and 53 compounds were reported the first time for Pogostemonis Herba. This study systematically characterized and identified the non-volatile compounds in Pogostemonis Herba for the first time. The findings provide a scientific basis for revealing the pharmacodynamic material basis, establishing a quality control system, and developing products of Pogostemonis Herba.

A comprehensive review on synthesis of dihydropyrimidione via Biginelli reaction catalyzed by reusable magnetic nanocatalyst (from 2020–till date)

The discovery of a new class of heterocyclic compounds by Pietro Biginelli in 1891 was regarded as a watershed point in the history of chemistry. Biginelli is a well‐known organic chemistry technique for generating dihydropyrimidones/thiones, which are important biological compounds. The chemistry of the Biginelli motifs has been studied in drug development because of the extensive use of multiple Biginelli adducts in various domains. The current review focuses on the magnetic reusable iron nanocatalysts, imparting their catalytic role in the Biginelli reaction, which can be impregnated with ionic liquids, Lewis's acids, metal (monometallic or bimetallic), or hybrid polymers as catalyst support. Our ongoing interest in developing environmentally friendly synthetic methodologies prompted us to investigate the utility of magnetically reusable catalysts where workup provides an eco‐friendly approach in comparison with traditional methods to provide a clear understanding of pathways leading to the formation of Biginelli adducts.

Quantitation by Liquid Chromatography-Nanoelectrospray Ionization-High-Resolution Tandem Mass Spectrometry of Multiple DNA Adducts Related to Cigarette Smoking in Oral Cells in the Shanghai Cohort Study.

We developed a liquid chromatography-nanoelectrospray ionization-high-resolution tandem mass spectrometry (LC-NSI-HRMS/MS) method for simultaneous quantitative analysis of 5 oral cell DNA adducts associated with cigarette smoking: (8R/S)-3-(2'-deoxyribos-1'-yl)-5,6,7,8-tetrahydro-8-hydroxypyrimido[1,2-a]purine-10(3H)-one (γ-OH-Acr-dGuo, 1) from acrolein; (6S,8S and 6R,8R)-3-(2'-deoxyribos-1'-yl)-5,6,7,8-tetrahydro-8-hydroxy-6-methylpyrimido[1,2-a]purine-10(3H)-one [(6S,8S)γ-OH-Cro-dGuo, 2; and (6R,8R)γ-OH-Cro-dGuo, 3] from crotonaldehyde; 1,N6-etheno-dAdo (4) from acrylonitrile, vinyl chloride, lipid peroxidation, and inflammation; and 8-oxo-dGuo (5) from oxidative damage. Oral cell DNA was isolated in the presence of glutathione to prevent artifact formation. Clear LC-NSI-HRMS/MS chromatograms were obtained allowing quantitation of each adduct using the appropriately labeled internal standards. The accuracy and precision of the method were validated, and the assay limit of quantitation was 5 fmol/μmol dGuo for adducts 1-4 and 20 fmol/μmol for adduct 5. The assay was applied to 80 buccal cell samples selected from those collected in the Shanghai Cohort Study: 40 from current smokers and 40 from never smokers. Significant differences were found in all adduct levels between smokers and nonsmokers. Levels of 8-oxo-dGuo (5) were at least 3000 times greater than those of the other adducts in both smokers and nonsmokers, and the difference between amounts of this adduct in smokers versus nonsmokers, while significant (P = 0.013), was not as great as the differences of the other DNA adducts between smokers and nonsmokers (P-values all less than 0.001). No significant relationship of adduct levels to risk of lung cancer incidence was found. This study provides a new LC-NSI-HRMS/MS methodology for the quantitation of diverse DNA adducts resulting from exposure to the α,β-unsaturated aldehydes acrolein and crotonaldehyde, inflammation, and oxidative damage which are all associated with carcinogenesis. We anticipate application of this assay in ongoing studies of the molecular epidemiology of cancers of the lung and oral cavity related to cigarette smoking.

Guidelines and considerations for building multidimensional libraries for untargeted MS-based metabolomics.

Feature annotation is crucial in untargeted metabolomics but remains a major challenge. The large pool of metabolites collected under various instrumental conditions is underrepresented in publicly available databases. Retention time (RT) and collision cross section (CCS) measurements from liquid chromatography ion mobility high-resolution mass spectrometers can be employed in addition to MS/MS spectra to improve the confidence of metabolite annotation. Recent advancements in machine learning focus on improving the accuracy of predictions for CCS and RT values. Therefore, high-quality experimental data are crucial to be used either as training datasets or as a reference for high-confidence matching. This manuscript provides an easy-to-use workflow for the creation of an in-house metabolite library, offers an overview of alternative solutions, and discusses the challenges and advantages of using open-source software. A total of 100 metabolite standards from various classes were analyzed and subjected to the described workflow for library generation. The outcome was an open-access available NIST format metabolite library (.msp) with multidimensional information. The library was used to evaluate CCS prediction tools, MS/MS spectra heterogeneities (e.g., multiple adducts, in-source fragmentation, radical fragment ions using collision-induced dissociation), and the reporting of RT.

Simultaneous quantification of multiple amino acid adducts from sulfur mustard-modified human serum albumin in plasma at trace exposure levels by ultra-high performance liquid chromatography–triple quadrupole mass spectrometry after propionyl derivatization

Sulfur mustard (HD) is a highly toxic vesicant and is prohibited by the Organisation for the Prohibition of Chemical Weapons (OPCW). HD can modify human serum albumin (HSA) to generate hydroxyethylthioethyl (HETE) adducts, which could be utilized as biomarkers for verifying HD exposure in forensic analysis. Here, five amino acid adducts generated from pronase digestion of HD-exposed human serum albumin (HD–HSA) in plasma were selected as biomarkers to retrospectively detect HD exposure. HD–HSA was precipitated from plasma with acetone, digested by pronase, derivatized with propionic anhydride (PA), and analysed with ultra-high performance liquid chromatography coupled with triple quadrupole mass spectrometry (UHPLC–TQ MS). The limits of detection (LODs) and limits of quantification (LOQs) of the HD exposure concentrations were evaluated as 1.00 ng/mL at S/N≥3 and 3.00 ng/mL at S/N≥10, respectively, which are approximately 60 times lower than those of the reported method. The approach shows good linearity (R2≥0.997) from 3.00 ng/mL to 10.0 µg/mL of HD-exposed human plasma with satisfactory precision and accuracy. The developed approach was applied to analysing samples from the 6th OPCW Biomedical Proficiency Test (BioPT). The study showed that the developed approach was also suitable for analysing human plasma samples that were exposed to six of HD analogues, which were common impurities in sulfur mustard mixtures. Moreover, the method was successfully applied to plasma from other species, including rabbits, rats and cattle. This study provides a reliable and sensitive tool for the retrospective detection of vesicants exposure based on multiple biomarkers.

The Effect of Benzannulation on the Structures, Reactivity and Molecular Dynamics of Indenes, Pentalenes, Azulenes and Related Molecules.

The stabilising effect of benzannulation on isoindenes formed in the course of sigmatropic shifts of (C5H5)Fe(CO)2 or of organo-silyl groups, and on exocyclic allyl intermediates in the course of haptotropic shifts of organometallic fragments over polycyclic skeletons (fluorene, cyclopenta[def]phenanthrene, syn and anti dibenzpentalenes) is exemplified. This approach led to the development of the first organometallic molecular brake. Benzyne cycloadditions to anthracenes to form triptycenes also led to unexpected or multiple adducts that were characterised by X-ray crystallography. Synthetic routes to the previously elusive benz[cd]azulene system are presented. Finally, the complete mechanism of the stepwise assembly of dispiro- and diindenyltetracenes from fluorenylallenes is presented, whereby every intermediate has been unambiguously structurally characterised.

Mass spectrometric profiling of DNA adducts in the human stomach associated with damage from environmental factors

BackgroundA comprehensive understanding of DNA adducts, one of the most plausible origins of cancer mutations, is still elusive, especially in human tissues in clinical settings. Recent technological developments have facilitated the identification of multiple DNA adducts in a single experiment. Only a few attempts toward this “DNA adductome approach” in human tissues have been reported. Geospatial information on DNA adducts in human organs has been scarce.AimMass spectrometry of human gastric mucosal DNA was performed to identify DNA adducts associated with environmental factors.Materials and methodsFrom 59 subjects who had received gastrectomy for gastric cancer, 306 samples of nontumor tissues and 15 samples of tumors (14 cases) were taken for DNA adductome analysis. Gastric nontumor tissue from autopsies of 7 subjects without gastric cancer (urothelial cancer, hepatocellular carcinoma, lung cancer each; the other four cases were without any cancers) was also investigated. Briefly, DNA was extracted from each sample with antioxidants, digested into nucleosides, separated by liquid chromatography, and then electrospray-ionized. Specific DNA adducts were identified by mass/charge number and column retention time compared to standards. Information on lifestyle factors such as tobacco smoking and alcohol drinking was taken from the clinical records of each subject.ResultsSeven DNA adducts, including modified bases, C5-methyl-2′-deoxycytidine, 2′-deoxyinosine, C5-hydroxymethyl-2′-deoxycytidine, N6-methyl-2′-deoxyadenosine, 1,N6-etheno-2′-deoxyadenosine, N6-hydroxymethyl-2′-deoxyadenosine, and C8-oxo-2′-deoxyguanosine, were identified in the human stomach and characterized. Intraindividual differences according to the multiple sites of these adducts were noted but were less substantial than interindividual differences. N6-hydroxymethyl-2′-deoxyadenosine was identified in the human stomach for the first time. The amount of C5-hydroxymethyl-2′-deoxycytidine was higher in the stomachs of subjects without gastric cancer than in the nontumor and tumor portions of the stomach in gastric cancer patients. Higher levels of 1,N6-etheno-2′-deoxyadenosine were detected in the subjects who reported both smoking and drinking than in those without these habits. These DNA adducts showed considerable correlations with each other.ConclusionsWe characterized 7 DNA adducts in the nontumor portion of the human stomach in both gastric cancer subjects and nongastric cancer subjects. A reduction in C5-hydroxymethyl-dC even in the nontumor mucosa of patients with gastric cancer was observed. Smoking and drinking habits significantly influenced the quantity of one of the lipid peroxidation-derived adducts, etheno-dA. A more expansive DNA adductome profile would provide a comprehensive understanding of the origin of human cancer in the future.

Nanoparticle‐Assisted Cation Adduction and Fragmentation of Small Metabolites

AbstractMass spectrometry (MS) promises small‐metabolite profiling as a tool of the future and calls for the comprehensive understanding of key procedures to enhance its capability. Herein, we studied cation adduction and fragmentation of small metabolites by a combination of theoretical and experimental approaches, via nanoparticle‐assisted laser desorption/ionization (LDI)‐MS and MS/MS. We calculated the energies of adduction conformers and atomic bond orders to establish the rules of cation–metabolite affinity and multiple cation adductions in charge transfer. Further, we demonstrated the reaction paths of adducted ions and mapped the potential energy surfaces to characterize the loss of given groups during fragmentation. Finally, we successfully controlled metabolite fragmentation by selected and multiple adductions to enhance the atomic/fragment coverage as defined for metabolite identification toward profiling. Considering the success of MS in the analysis of large biomolecules, our work may have an impact and guide to advanced analysis of small metabolites.

Nanoparticle-Assisted Cation Adduction and Fragmentation of Small Metabolites.

Mass spectrometry (MS) promises small-metabolite profiling as a tool of the future and calls for the comprehensive understanding of key procedures to enhance its capability. Herein, we studied cation adduction and fragmentation of small metabolites by a combination of theoretical and experimental approaches, via nanoparticle-assisted laser desorption/ionization (LDI)-MS and MS/MS. We calculated the energies of adduction conformers and atomic bond orders to establish the rules of cation-metabolite affinity and multiple cation adductions in charge transfer. Further, we demonstrated the reaction paths of adducted ions and mapped the potential energy surfaces to characterize the loss of given groups during fragmentation. Finally, we successfully controlled metabolite fragmentation by selected and multiple adductions to enhance the atomic/fragment coverage as defined for metabolite identification toward profiling. Considering the success of MS in the analysis of large biomolecules, our work may have an impact and guide to advanced analysis of small metabolites.

Paper spray ionization-high-resolution mass spectrometry (PSI-HRMS) of peroxide explosives in biological matrices.

Mitigation of the peroxide explosive threat, specifically triacetone triperoxide (TATP) and hexamethylene triperoxide diamine (HMTD), is a priority among the law enforcement community, as scientists and canine (K9) units are constantly working to improve detection. We propose the use of paper spray ionization-high-resolution mass spectrometry (PSI-HRMS) for detection of peroxide explosives in biological matrices. Occurrence of peroxide explosives and/or their metabolites in biological samples, obtained from urine or blood tests, give scientific evidence of peroxide explosives exposure. PSI-HRMS promote analysis of samples in situ by eliminating laborious sample preparation steps. However, it increases matrix background issues, which were overcome by the formation of multiple alkali metal adducts with the peroxide explosives. Multiple ion formation increases confidence when identifying these peroxide explosives in direct sample analysis. Our previous work examined aspects of TATP metabolism. Herein, we investigate the excretion of a TATP glucuronide conjugate in the urine of bomb-sniffing dogs and demonstrate its detection using PSI from the in vivo sample.

Removal of optimal cutting temperature (O.C.T.) compound from embedded tissue for MALDI imaging of lipids.

Matrix-assisted laser desorption/ionisation mass spectrometry imaging (MALDI-MSI) is a common molecular imaging modality used to characterise the abundance and spatial distribution of lipids in situ. There are several technical challenges predominantly involving sample pre-treatment and preparation which have complicated the analysis of clinical tissues by MALDI-MSI. Firstly, the common embedding of samples in optimal cutting temperature (O.C.T.), which contains high concentrations of polyethylene glycol (PEG) polymers, causes analyte signal suppression during mass spectrometry (MS) by competing for available ions during ionisation. This suppressive effect has constrained the application of MALDI-MSI for the molecular mapping of clinical tissues. Secondly, the complexity of the mass spectra is obtained by the formation of multiple adduct ions. The process of analyte ion formation during MALDI can generate multiple m/z peaks from a single lipid species due to the presence of alkali salts in tissues, resulting in the suppression of protonated adduct formation and the generation of multiple near isobaric ions which produce overlapping spatial distributions. Presented is a method to simultaneously remove O.C.T. and endogenous salts. This approach was applied to lipid imaging in order to prevent analyte suppression, simplify data interpretation, and improve sensitivity by promoting lipid protonation and reducing the formation of alkali adducts.

Fluorescent salen-type Zn(II) Complexes As Probes for Detecting Hydrogen Sulfide and Its Anion: Bioimaging Applications.

In this work, we investigate the mode of interaction of a family of fluorescent zinc complexes with HS– and H2S. Different experiments, performed by diverse spectroscopic techniques, provide evidence that HS– binds the zinc center of all the complexes under investigation. Treatment with neutral H2S exhibits a markedly different reactivity which indicates selectivity for HS– over H2S of the systems under investigation. Striking color changes, visible to the naked eye, occur when treating the systems with HS– or by an H2S flow. Accordingly, also the fluorescence is modulated by the presence of HS–, with the possible formation of multiple adducts. The results highlight the potential of the devised systems to be implemented as HS–/H2S colorimetric and fluorescent sensors. Bioimaging experiments indicate the potential of using this class of compounds as probes for the detection of H2S in living cells.

New horizons of DNA adductome for exploring environmental causes of cancer.

Chemical carcinogenesis is focused on the formation of DNA adducts, a form of DNA damage caused by covalent binding of a chemical moiety to DNA. The detection of carcinogen‐DNA adducts in human tissues, along with demonstration of mutagenicity/carcinogenicity in experimental systems, and validation of adducts as biomarkers of environmental exposure and indicators of cancer risk in molecular epidemiological studies suggests a pivotal role of DNA adducts in cancer development. However, accurate measurement of DNA adducts in varied biological samples is challenging. Advances in mass spectrometry have prompted the development of DNA adductome analysis, an emerging method that simultaneously screens for multiple DNA adducts and provides relevant structural information. In this review, we summarize the basic principle and applications of DNA adductome analysis that would contribute to the elucidation of the environmental causes of cancer. Based on parallel developments in several fields, including next‐generation sequencing, we describe a new approach used to explore cancer etiology, which integrates analyses of DNA adductome data and mutational signatures derived from whole‐genome/exome sequencing.

High-Throughput Amenable MALDI-MS Detection of RNA and DNA with On-Surface Analyte Enrichment Using Fluorous Partitioning.

High-throughput matrix-assisted laser desorption/ionization mass spectrometry (HT-MALDI-MS) has garnered considerable attention within the drug discovery industry as an information-rich alternative to assays using light-based detection methods. To date, these efforts have been primarily focused on assays using protein or peptide substrates. Methods for RNA or DNA analysis by HT-MALDI-MS have not been extensively reported due to the challenges associated with MALDI-MS of oligonucleotides, including the propensity to form multiple salt adducts, low ionization potential, and ease of fragmentation. The objective of this work was to develop a platform suitable for HT-MS analysis of RNA and DNA substrates that overcomes these hurdles by combining on-surface sample preparation with soft ionization. This has been accomplished through the selective immobilization of fluorous-tagged oligonucleotides on a fluorous-modified MS target plate, followed by on-surface enrichment, matrix addition, and direct laser desorption/ionization, a process dubbed fluorous HT-MS (F-HT-MS). The work has resulted in methods by which RNA and DNA substrates can be detected at nanomolar concentrations from a typical assay buffer system using procedures that are amenable to full automation. The protocols were applied to an miRNA biogenesis assay, demonstrating its potential for RNA processes and thereby filling a prominent gap in RNA drug discovery: the paucity of in vitro functional assays.

Abstract A24: Exploration of esophageal cancer etiology using comprehensive DNA adduct analysis (DNA adductome analysis)

Abstract Objective: China has among the highest incidence and mortality rates of esophageal cancer in the world. Cixian, one of the high-incidence areas, demonstrates a much higher incidence when compared with other rural or urban areas in China. To address the etiology of esophageal cancer in Cixian, we carried out a comprehensive DNA adduct analysis (DNA adductome) using surgical specimens that were collected from esophageal cancer patients living in Cixian and Shijiazhuang (low-incidence area for esophageal cancer). Methods: Both tumorous and nontumorous tissues were collected from patients who underwent surgical procedures at Cixian Cancer Hospital and the Fourth Hospital of Hebei Medical University. A peripheral blood sample was also collected in each hospital. DNA adductome analysis was performed with LC-QTOF mass spectrometer by using DNA derived from surgical specimen. Whole-exome sequencing analysis was also conducted using esophageal tumor/peripheral blood paired samples. Mutation analyses of candidate chemical substance, which was predicted by DNA adductome analysis, were performed with Ames assay and gpt delta transgenic rats. Results: Multiple DNA adducts were detected in samples from both areas in the DNA adductome analysis. The 2D-PCA scores plot of the DNA adducts showed a clear clustering of the high- and low-incidence areas, and the associated loadings plot demonstrated that several DNA adducts made a greater contribution to the high-incidence area. By referring to an in-house DNA adduct database, N2-(3,4,5,6-tetrahydro-2H-pyran-2-yl)deoxyguanosine (THP-dG), which was derived from N-nitrosopiperidine (NPIP), emerged as a major DNA adduct. In order to confirm the DNA adduct that was highly correlated to the high-incidence area, we synthesized authentic 15N5-THP-dG and analyzed it by quantitative LC-MS/MS apparatus. An elution peak, which appeared at the same position of authentic 15N5-THP-dG, was observed in the surgical specimens collected from the high-incidence area. Examination of THP-dG adduct in peripheral blood samples demonstrated that the adduct levels were significantly higher in the high-incidence area than in the low-incidence area. Moreover, NPIP exhibited a strong mutagenic activity under metabolic activation in the Ames test and a significant dose-dependent increase in mutation frequency during in vivo mutagenicity tests with gpt delta rats. The NPIP-induced mutation was dominated by A:T to C:G transversions, followed by G:C to A:T and A:T to G:C transitions, in both liver and esophagus of animal samples. Similar mutational patterns were observed in the mutational signatures of esophageal cancer patients, which demonstrated weak correlation with THP-dG levels. Conclusion: This is the first study that used DNA adductome analysis to reveal the etiology of esophageal cancer in Cixian. These findings suggested that NPIP exposure is partly involved in the development of esophageal cancer in Cixian residents. Citation Format: Hitoshi Nakagama, Yukari Totsuka, Yingsong Lin, Yutong He, Haruna Sato, Tomonari Matsuda, Yoshitaka Matsushima, Mamoru Kato, Asmaa Elzawahry, Yasushi Totoki, Tatsuhiro Shibata, Baoen Shan. Exploration of esophageal cancer etiology using comprehensive DNA adduct analysis (DNA adductome analysis) [abstract]. In: Proceedings of the AACR Special Conference on Environmental Carcinogenesis: Potential Pathway to Cancer Prevention; 2019 Jun 22-24; Charlotte, NC. Philadelphia (PA): AACR; Can Prev Res 2020;13(7 Suppl): Abstract nr A24.

Protocol for community-created public MS/MS reference spectra within the Global Natural Products Social Molecular Networking infrastructure.

A major hurdle in identifying chemicals in mass spectrometry experiments is the availability of tandem mass spectrometry (MS/MS) reference spectra in public databases. Currently, scientists purchase databases or use public databases such as Global Natural Products Social Molecular Networking (GNPS). The MSMS-Chooser workflow is an open-source protocol for the creation of MS/MS reference spectra directly in the GNPS infrastructure. An MSMS-Chooser Sample Template is provided and completed manually. The MSMS-Chooser Submission File and Sequence Table for data acquisition were programmatically generated. Standards from the Mass Spectrometry Metabolite Library (MSMLS) suspended in a methanol-water (1:1) solution were analyzed. Flow injection on an LC/MS/MS system was used to generate negative and positive mode data using data-dependent acquisition. The MS/MS spectra and Submission File were uploaded to MSMS-Chooser workflow in GNPS for automatic selection of MS/MS spectra. Data acquisition and processing required ~2 h and ~2 min, respectively, per 96-well plate using MSMS-Chooser. Analysis of the MSMLS, over 600 small molecules, using MSMS-Chooser added 889 spectra (including multiple adducts) to the public library in GNPS. Manual validation of one plate indicated accurate selection of MS/MS scans (true positive rate of 0.96 and a true negative rate of 0.99). The MSMS-Chooser output includes a table formatted for inclusion in the GNPS library as well as the ability to directly launch searches via MASST. MSMS-Chooser enables rapid data acquisition, data analysis (selection of MS/MS spectra), and a formatted table for inspection and upload to GNPS. Open file-format data (.mzML or.mzXML) from most mass spectrometry platforms containing MS/MS spectra can be processed using MSMS-Chooser. MSMS-Chooser democratizes the creation of MS/MS reference spectra in GNPS which will improve annotation and strengthen the tools which use the annotation information.

Reading patterns of proteome damage by glycation, oxidation and nitration: quantitation by stable isotopic dilution analysis LC-MS/MS.

Liquid chromatography-tandem mass spectrometry (LC-MS/MS) provides a high sensitivity, high specificity multiplexed method for concurrent detection of adducts formed by protein glycation, oxidation and nitration, also called AGEomics. Combined with stable isotopic dilution analysis, it provides for robust quantitation of protein glycation, oxidation and nitration adduct analytes. It is the reference method for such measurements. LC-MS/MS has been used to measure glycated, oxidized and nitrated amino acids - also called glycation, oxidation and nitration free adducts, with a concurrent quantitation of the amino acid metabolome in physiological fluids. Similar adduct residues in proteins may be quantitated with prior exhaustive enzymatic hydrolysis. It has also been applied to quantitation of other post-translation modifications, such as citrullination and formation of Nε-(γ-glutamyl)lysine crosslink by transglutaminases. Application to cellular and extracellular proteins gives estimates of the steady-state levels of protein modification by glycation, oxidation and nitration, and measurement of the accumulation of glycation, oxidation and nitration adducts in cell culture medium and urinary excretion gives an indication of flux of adduct formation. Measurement of glycation, oxidation and nitration free adducts in plasma and urine provides for estimates of renal clearance of free adducts. Diagnostic potential in clinical studies has been enhanced by the combination of estimates of multiple adducts in optimized diagnostic algorithms by machine learning. Recent applications have been in early-stage detection of metabolic, vascular and renal disease, and arthritis, metabolic control and risk of developing vascular complication in diabetes, and a blood test for autism.