A streamlined workflow for high throughput metaproteomic analysis of the rumen microbiome

Publication date: Available online 4 August 2026 Source: Journal of Proteomics Author(s): Ane Laura Pedersen, Loïc Dayon, Michael Affolter, Jana Seifert

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Interactive Software for Interpreting and Curating High‐Resolution Electron Ionization Mass Spectra

Journal of Mass Spectrometry, Volume 61, Issue 8, August 2026.

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Mapping the Human Ghost Proteome: Classification and Experimental Detection Biases in the Identification of Alternative Microproteins

The discovery of alternative proteins (AltProts), translated from non-canonical ORFs, has expanded the human proteome and revealed a hidden layer known as the ghost proteome. Despite increasing evidence, AltProts detection remains challenging due to their small size, physicochemical heterogeneity, and lack of annotation. Here, we developed an integrated bioinformatic and proteomic workflow to benchmark the detection of reference proteins (RefProts), isoforms, and alternative microproteins (MicroAltProts) in colorectal cancer cells using four extraction protocols -HCl, RIPA buffer, RIPA with chloroform, and RIPA followed by 30 kDa filtration- combined with high-resolution data-independent acquisition mass spectrometry. We identified and quantified using the Orbitrap Astral mass spectrometer a total of 66,438 peptides corresponding to 12,584 different protein groups across methods, with RIPA-based extraction approaches providing the most comprehensive coverage. To reduce redundancy in the OpenProt database and focus on MicroAltProts, we curated the dataset by removing known isoforms and long proteins, yielding a non-redundant set of 183,937 MicroAltProts. K-means clustering based on eight ProtParam-derived features grouped MicroAltProts into four physicochemical clusters. Among them, 43 MicroAltProts (

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Resolving drug uptake heterogeneity in 3D organ on chip models via live single-cell microsampling and ion mobility mass spectrometry

Publication date: Available online 3 August 2026 Source: Analytica Chimica Acta Author(s): Vincent Verschoor, Laura Ann Castaneda, Arnaud Nicolas, Bas Trietsch, Thomas Hankemeier, Ahmed Ali

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Single laboratory validation of multi-residue extraction and cleanup method for analysis of 50 pesticides, 14 veterinary drugs, and 7 Polychlorinated biphenyls in bovine milk by tandem mass spectrometry

Publication date: Available online 3 August 2026 Source: Journal of Chromatography A Author(s): Nisala Gunasekera, Ruchika Fernando, Ilmi Hewajulige, G.V.V. Liyanaarachchi, D.A.T.W.K Dissanayake, Dilini Madhushani, Laksara Maheshika

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Mass Spectrometry‐Based Extracellular Vesicle Proteomics for Biomarker Discovery

ABSTRACT Extracellular vesicle (EV) proteomics has emerged as a powerful platform for decoding intercellular communication and advancing biomarker discovery across human diseases. EVs carry proteins that reflect their cells of origin, offering a minimally invasive window into physiological and pathological processes. Mass spectrometry (MS) now enables deep, high-resolution EV proteome profiling, aided by improved isolation and rigorous characterization that ensure sample purity and integrity. Advanced computational pipelines integrating quantitative modeling, spectral-library prediction, machine learning and multi-omics analysis extract meaningful biological signals, revealing subtle disease-associated EV signatures and establishing EV proteomics as a strong platform for biomarker discovery and precision medicine. This review provides an integrated framework linking EV isolation principles, characterization strategies, mass-spectrometric workflows, and computational analysis to the biological and clinical insights they generate. We also highlight key challenges and future directions, including the need for standardized reference materials, unified pre-analytical workflows, and EV proteome reference atlases. Together, these innovations are transforming EV proteomics into a next-generation tool for precision medicine.

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Rapid and sensitive multi-residue method for the determination of 235 organic microcontaminants in environmental water samples using direct injection coupled to ultra-high performance liquid chromatography-tandem mass spectrometry

Publication date: 1 February 2027 Source: Talanta, Volume 312, Part A Author(s): Eva Jambrina-Hernández, Patricia Plaza-Bolaños, Samira Nahim-Granados, Ana Agüera

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Structural dynamics underlying agonist activation of a GLP-1R-Gs precoupled complex

G-protein-coupled receptors (GPCRs) act as allosteric transmembrane signalling machines, generating distinct cellular responses depending on the conformational states induced by ligand binding. The glucagon-like peptide-1 receptor (GLP-1R), a class B GPCR central to insulin secretion and body-weight regulation, is a key therapeutic target for obesity-associated metabolic disease. Here, we used hydrogen-deuterium exchange mass spectrometry to characterize ligand-evoked structural dynamics within a pre-coupled GLP-1R-Gs protein complex. Non-peptide agonists Chu-128 and danuglipron elicited overlapping dynamic perturbation profiles, with distinct drug-specific effects within the transmembrane bundle. In contrast, the natural GLP-1 hormone produced a weaker stabilizing effect on receptor backbone dynamics, while its inactive metabolite exerted opposing localised destabilization. Notably, both peptides uniquely modulated the highly flexible G-protein switch III loop, a key mediator of downstream signalling. These findings pinpoint areas where structural dynamics shape agonist efficacy and facilitate functional dynamics-integrated drug discovery of non-peptide agonists.

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Decoding the Plasma Proteomic Landscape of Clear Cell Renal Cell Carcinoma Reveals Diagnostic and Prognostic Liquid Biopsy Biomarkers

Clear cell renal cell carcinoma (ccRCC) remains the most lethal urological malignancy, with high metastatic rates, both at initial diagnosis and during disease progression, contributing to poor survival outcomes. Current diagnostic and prognostic approaches rely primarily on histopathology, limiting early detection of localized disease and relevant intervention for metastatic patients. Here, we performed the most extensive to-date mass spectrometry-based discovery profiling of longitudinal plasma samples collected across multiple clinical follow-up points spanning up to five years post-diagnosis., to characterize the circulating plasma proteome and identify biomarkers for localized and metastatic disease. Network analysis identified protein modules enriched in pathways involved in matrix remodeling and metabolic deregulation, perpetuating the ccRCC phenotype. A five-protein signature, comprising PRL, THBS1, ANGPT1, IGFBP1, and SRGN, demonstrated high diagnostic performance for localized ccRCC. Notably, PRL appeared as a promising stand-alone biomarker (AUC = 0.812), with independent validation confirming its utility as a diagnostic biomarker. Importantly, a six-protein signature (AMBP, C1S, C2, IGFBP3, RASGRP2, TFRC) stringently distinguished metastatic from high-grade non-metastatic ccRCC cases. Further validation of these signatures could inform clinical decision-making, enabling early detection of metastasis and minimal residual disease and real-time longitudinal monitoring for ccRCC patients.

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Emerging tools to investigate the contribution of selenium to ferroptosis and beyond - Nature Chemical Biology

This Review examines the chemical complexity of selenium biology and the tools available to study it. Defining current limitations in detecting selenium species outlines pathways toward a more precise, mechanistic understanding.

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Fluorescence polarization-based fragment screen identifies inhibitors of APOBEC3A and APOBEC3B

APOBEC3A and APOBEC3B are antiviral cytidine deaminases found to drive cancer-associated mutagenesis, contributing to tumor evolution and therapeutic resistance across multiple cancer types. Inhibiting these enzymes holds promise for prolonging response to a wide range of cancer therapies by delaying development of resistance. However, APOBEC3A and APOBEC3B remain challenging drug targets, with no potent and selective small molecule inhibitors reported. Here, we use a fluorescence polarization-based assay to identify small molecules inhibitors of the APOBEC3A-single-stranded DNA interaction. From a library of 2,400 disulfide compounds, we identified 64 hits (mean polarization +/- 3 sigma, hit rate of 2.7%). Intact protein mass spectrometry revealed that a subset of compounds covalently engages A3A at cysteine 64, including Compounds 1 and 2. Compounds 1 and 2 disrupt APOBEC3A/APOBEC3B-single-stranded DNA interactions and inhibit APOBEC3A/APOBEC3B deaminase activity in a dose-dependent manner, with micromolar IC50. Surprisingly, inhibition of APOBEC3A/APOBEC3B by Compounds 1 and 2 is independent of covalent tethering to cysteine, suggesting a predominantly non-covalent mode of binding. Together, these studies establish an integrated workflow for APOBEC ligand discovery and identify Compounds 1 and 2 as starting points for developing chemical probes to investigate APOBEC-driven mutagenesis and therapeutic resistance.

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Distinct mechanisms of antibody-mediated HCV neutralization revealed by nanobody-guided epitope mapping

Hepatitis C virus (HCV) remains a major global health challenge despite the availability of highly effective antiviral therapies, underscoring the need for a broadly protective vaccine. The envelope glycoprotein E2 is the principal target of neutralizing antibodies, yet the full repertoire of vulnerable epitopes and mechanisms of antibody mediated neutralization remains incompletely understood. Here, we exploited the unique binding properties of camelid nanobodies to probe the antigenic landscape of HCV E2 beyond the immunodominant human antibody response. We isolated a diverse panel of E2-specific nanobodies, including broadly neutralizing antibodies with high-affinity cross reactivity toward genetically diverse HCV isolates. By combining cross-neutralization assays, competition binding experiments, and high resolution hydrogen deuterium exchange mass spectrometry (HDX-MS), we identified three mechanistically distinct classes of neutralizing epitopes. While one class targets the canonical E2 neutralization face, a second class recognizes antigenic region 1 (AR1), independently validating and extending recent evidence that this region represents a functional site of viral vulnerability. These findings demonstrate that broadly neutralizing antibody responses extend beyond the canonical neutralization face and establish a broader framework for understanding HCV neutralization. More broadly, our study illustrates how alternative antibody repertoires can reveal functionally important antigenic surfaces that are underrepresented in conventional human antibody responses, providing new opportunities for the rational design of next-generation HCV vaccine immunogens.

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A prospective multi-cohort study identifies reproducible molecular responses to musical stimulation in the human tear proteome

Music elicits complex emotional, cognitive, and physiological responses, yet the molecular mechanisms underlying these effects remain poorly understood. Tear fluid is an accessible biofluid that reflects both systemic and local physiology, providing a unique opportunity to investigate the biological effects of musical stimulation. Quantitative mass spectrometry-based proteomics was performed on paired tear samples (n = 336) collected before and after exposure to a standardized musical stimulus in two independent cohorts of healthy participants, including both music-stimulated and music-unstimulated individuals. Integrative analyses, including differential expression, functional enrichment, co-expression network, correlation, and machine-learning approaches, identified reproducible molecular responses across cohorts. Fourteen proteins, including PRTN3, LEG3, CATD, IGHG2, and ZA2G, remained significant after multiple-testing correction and showed concordant regulation in the stimulated cohorts but not in controls. Functional enrichment converged on innate immune, inflammatory, host-defense, and extracellular signaling pathways, and consensus network analysis identified reproducible immune-related modules. The 14-protein signature robustly discriminated pre- and post-stimulation samples, with no effect detected in the unstimulated control groups. These findings support tear proteomics as a promising non-invasive approach for investigating the molecular basis of auditory and emotional experiences.

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Multiparameter Simulation for the Optimization of Ion Transmission Efficiency Through Mass Spectrometry Inlet Capillary

ABSTRACT Rationale According to the mass spectrometry (MS) design principles, the ions generated in an atmospheric pressure ion source must pass through a narrow-bore capillary in order to maintain the needed system vacuum pressure. The ion transmission efficiency through the inlet capillary is one of the key factors directly influencing the analytical performance of MS. Its design and operating parameters, including inner diameter, length, temperature, and those related to the ion source operation, can alter ion diffusion losses, gas flow dynamics, and space-charge effects both around and inside the inlet capillary, thereby influencing ion sampling efficiency and transmission efficiency to the mass analyzer. Methods A comprehensive simulation of ion transmission efficiency through an MS inlet capillary has been performed in this study by adjusting the important design and operating parameters. Results The simulation results show that the ion transmission efficiency through the inlet capillary is positively correlated with the inner diameter of the capillary and negatively correlated with the length of the capillary. Specifically, under the conditions of a capillary diameter of 0.8 mm and a temperature of 573.15 K, the ion transmission efficiency through a 10-cm-long capillary can reach 90%. The efficiency decreases with a reduction in the inner diameter or an increase in the length. When the temperature exceeds 573.15 K, the flow field inside the capillary transits from laminar to turbulent flow, leading to a sharp decline in ion transmission efficiency. For every 100-K increase in temperature beyond this point, the ion transmission efficiency decreases by 10%–20%. Conclusions The main results from this study have provided crucial design guidelines, emphasizing the use of larger diameters, moderate lengths, and strict temperature control below the turbulent transition threshold to maximize ion transmission and overall instrument sensitivity for MS instruments.

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MassGAT: a graph-based collective learning approach for untargeted detection and annotation of LC-MS data

Motivation: The untargeted processing of Liquid Chromatography coupled to High-Resolution Mass Spectrometry (LC-HRMS) data is a major challenge for the comprehensive and robust characterisation of metabolites. In particular, peak detection and annotation are two challenging tasks due to the size and complexity of the data, that are currently addressed independently in existing pipelines, without taking into account the redundancy of information between the ion species of the same compound. Results: We developed an innovative and efficient approach that combines detection and annotation, by 1) representing signals putatively originating from the same molecule as a graph, and 2) inferring the validity of the peaks and their connections within each component using a Graph Attention Network (GAT). We demonstrate on real data sets that the resulting MassGAT model outperforms current approaches in terms of both detection and annotation. Availability and implementation: The MassGAT open-source Python module is publicly available at https://github.com/odisce/MassGAT.

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Measurement of Stable Isotope Ratios of Halogenated Organic Compounds by Liquid Chromatographic, High-Temperature Combustion Isotope Ratio Mass Spectrometry

Publication date: Available online 2 August 2026 Source: Analytica Chimica Acta Author(s): Sota Maehara, Hiroto Kawashima, Sachi Taniyasu

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ENANTIOSELECTIVE LOW-PRESSURE GAS CHROMATOGRAPHY COMBINED WITH TRIPLE QUADRUPOLE MASS SPECTROMETRY FOR THE ANALYSIS OF 107 CHIRAL AND ACHIRAL PESTICIDE RESIDUES IN WINE

Publication date: Available online 1 August 2026 Source: Analytica Chimica Acta Author(s): Giorgia Rinaldi, Mariosimone Zoccali, Peter Q. Tranchida, Luigi Mondello

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Implementation of Nonisobaric TMT Analogs for Accurate Precursor‐Level Quantification by plexDIA

ABSTRACT Rationale Multiplexed quantitative proteomics enables simultaneous analysis of multiple biological samples, increasing throughput while reducing instrument time and sample requirements. However, integrating sample multiplexing with data-independent acquisition (DIA) remains challenging. We present a TMTpro plexDIA strategy leveraging MS1-level mass differences between nonisobaric TMTpro reagent variants to enable multiplexed quantification without compromising DIA sensitivity. Methods Three Saccharomyces cerevisiae deletion strains (Δmet6, Δpfk2, and Δura2) were labeled with TMTproZero (light), TMTpro16 (standard), and super-heavy TMTpro (heavy), mixed in three permutations, and analyzed by narrow-window DIA (2 m/z isolation, 300 scan events) on an Orbitrap Astral mass spectrometer. Database searching was performed using FragPipe/MSFragger with plex-DIA quantification enabled. Results Over 2000 protein groups and over 20 000 precursors were identified per channel per mixture, with closely matched identification rates across all three channels. All nine expected deletion patterns were correctly identified, with channel-specific depletion reproduced consistently across precursor charge states (2+, 3+, and 4+). Conclusions TMTpro plex-DIA enables accurate, multiplexed quantitative proteomics through MS1-level mass separation of the nonisobaric TMTpro isotopologs. The characteristic deletion patterns observed for each knockout strain serve as intrinsic molecular barcodes, validating sample identity and demonstrating the broad utility of plex-DIA for high-throughput, multiplexed proteomics applications.

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Molecular‐Level Characterization of Native Lignin Oligomers From Flow‐Through Solvolysis of Biomass by High‐Performance Liquid Chromatography–Tandem Mass Spectrometry

ABSTRACT Rationale Comprehensive characterization of lignin-derived oligomers remains a major analytical challenge because of their high structural diversity and the lack of methods capable of resolving native interunit linkages in complex biomass-derived mixtures. Molecular-level understanding of these oligomers is essential for elucidating lignin depolymerization mechanisms and guiding biomass valorization strategies. Methods Lignin oligomers generated from methanol-based flow-through solvolysis of birch sawdust were analyzed by high-performance liquid chromatography coupled with ultraviolet detection and tandem mass spectrometry (HPLC-UVD/MSn). Chromatographic separation was performed on tandem reversed-phase C18 columns, and structural characterization was achieved using electrospray ionization Orbitrap mass spectrometry with higher-energy collisional dissociation (HCD). Results Distinct chromatographic behaviors were observed for lignin-derived products with different degrees of polymerization, enabling differentiation of monomeric and oligomeric species. Tandem MS analysis of representative oligomers revealed characteristic fragmentation pathways that enabled structural elucidation of oligomers. β-O-4 linkages were identified as the predominant interunit structures, and native β-β motifs were also characterized. Conclusions Untargeted molecular-level characterization of lignin-derived products generated from methanol-based flow-through solvolysis of birch biomass provides direct insights into the composition and interunit linkages of native-like lignin oligomers. These findings reveal the structural diversity of biomass-derived oligomers and bridge the gap between model-compound studies and real biomass-derived lignin systems.

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Design of a New FAIMS Device With Replaceable Accessories and Analysis of the Impact of Spacing

ABSTRACT Rationale The plate spacing has a significant impact on the resolution and sensitivity of High-Field Asymmetric Waveform Ion Mobility Spectrometry (FAIMS). Theoretical and experimental analysis of this parameter is of great significance for improving the performance of FAIMS. Methods A self-designed modular FAIMS device with replaceable upper and lower PCB plates was developed to provide a reliable experimental platform for investigating plate spacing as a single variable. In this device, the fixed mechanical positioning and conductive-contact connection help restore consistent electrode positioning. Theoretical analysis and SIMION simulations were first performed to predict the influence of plate spacing, followed by experimental validation. Plate spacings of 0.3, 0.4, 0.5, and 0.6 mm were set, and acetone samples were detected under the same electric field conditions. Results The simulation and experimental results show that with the increase of plate spacing, the FAIMS signal intensity (peak height) increases significantly: when the plate spacing increases from 0.3 to 0.6 mm, the signal intensity of the main peak increases by approximately 100%, and that of the secondary peak increases 4.6-fold. Moreover, the relative standard deviation (RSD) of all spacings is below 5%. Increasing the plate spacing significantly improves the sensitivity of the FAIMS system while the resolution does not change significantly, and the system maintains good separation capability. Conclusions This study provides a theoretical basis and practical guidance for the optimal design of FAIMS instruments.

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Orbitrap HRMS‐Based Comprehensive Metabolite Characterization of Duvelisib in Cross‐Species Hepatocytes

ABSTRACT Rationale Duvelisib (DVB), a selective PI3K-δ/γ inhibitor approved for chronic lymphocytic leukemia, requires detailed metabolic characterization to support early drug development and selection of appropriate toxicology species. Because drug metabolism varies across species, an integrated system such as primary hepatocytes containing both Phase I and Phase II enzymes is critical for accurately defining metabolic pathways and identifying potential reactive metabolites. Methods DVB metabolism was investigated using primary hepatocytes from human, monkey, dog, rat, and mouse. Incubated samples were analyzed using high-resolution mass spectrometry (HRMS) to detect and characterize metabolites. Structural elucidation was performed based on MS/MS fragmentation patterns. Mechanistic studies were conducted using a selective aldehyde oxidase (AO) inhibitor to assess the role of AO in DVB metabolism. Results DVB underwent diverse metabolic transformations, including oxidation, N-dealkylation, N-glucuronidation and glutathione (GSH) conjugation, leading to the identification of 18 putative metabolites. Thirteen metabolites, including several glucuronide and GSH conjugates were newly identified. AO-mediated oxidation on the purine ring emerged as a major pathway and was significantly reduced in the presence of an AO inhibitor, confirming its involvement. Two previously unreported GSH conjugates were also characterized, suggesting potential sites of reactive metabolite formation and associated toxicity risk. Comparative analysis revealed distinct species-dependent metabolic profiles, with both shared and species-specific metabolites identified among human, mouse, rat, dog, and monkey hepatocytes. Conclusions The present study provides a comprehensive cross-species metabolic profile of DVB, highlighting key pathways and novel metabolites, including those linked to potential toxicity. Mouse hepatocytes are recommended for future toxicological studies.

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A Dual GC–MS and LC–MS/MS Analytical Strategy for Evaluating Free Chlorine Toxicity in Aqueous Samples

ABSTRACT Rationale Chlorine gas is a highly toxic industrial chemical that poses a significant risk in accidental releases and deliberate terrorist attacks. Reliable analytical methods for the determination of chlorine in aqueous samples are therefore required for forensic and environmental investigations. This study aimed to develop a rapid and selective mass spectrometry-based method for chlorine determination following derivatization. Methods Chlorine in aqueous samples was derivatized with methyl 2-(3,4,5-trimethoxyphenyl)acetate (MTMPA) at 50°C for 15 min, yielding methyl 2-(2-chloro-3,4,5-trimethoxyphenyl)acetate (Cl-MTMPA). The derivatization product was analyzed by gas chromatography–mass spectrometry (GC–MS) and liquid chromatography–tandem mass spectrometry (LC–MS/MS). While LC–MS/MS analysis was performed directly, GC–MS analysis required an additional liquid–liquid extraction step. Reaction conditions were optimized to maximize derivatization efficiency. Results The derivatization reaction provided a high yield of Cl-MTMPA under the optimized conditions. GC–MS and LC–MS/MS analyses produced a common major fragment ion at m/z 215.0. Further structural confirmation by liquid chromatography–high-resolution tandem mass spectrometry (LC-HRMS/MS) revealed a characteristic product ion at m/z 215.0473 generated from the precursor ion at m/z 275.0681. Calibration curves covering acute toxicologically relevant chlorine concentrations (5–200 mg/L) showed excellent linearity, with correlation coefficients (R 2) exceeding 0.99. Conclusions A rapid and selective derivatization-based method was developed in which the resulting chlorine derivative is readily analyzed by both GC–MS and LC–MS/MS. The method affords reliable analyte identification through characteristic fragmentation and excellent quantitative performance, making it suitable for forensic toxicology and environmental investigations involving chlorine exposure.

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LC‐HRMS‐Based Metabolic Profiling of the REV‐ERB Agonist SR9009 in Camel Liver Homogenate and Cunninghamella elegans for Anti‐Doping Analysis

ABSTRACT Rationale SR9009 is a synthetic REV-ERB agonist with potential performance-enhancing properties and is included on the World Anti-Doping Agency (WADA) Prohibited List for human sport. Its potential misuse has also raised concerns in animal sports, particularly equine and camel racing. Although SR9009 metabolism has been investigated in human and equine models, its metabolic fate in racing camels (Camelus dromedarius) remains unclear. Characterizing its biotransformation pathways and identifying suitable biomarkers are essential for developing reliable anti-doping screening strategies for camel racing. Methods An in vitro metabolic study was conducted using camel liver homogenate and the fungal model Cunninghamella elegans. Metabolites were characterized using liquid chromatography-high-resolution mass spectrometry (LC-HRMS). Putative structural elucidation was achieved through accurate mass measurements of precursor and product ions. Diagnostic fragmentation pathways were established using collision-induced dissociation (CID) tandem mass spectrometry (MS/MS), enabling the systematic identification of metabolic modifications based on characteristic fragment ions and neutral losses. Results Seventeen Phase I metabolites of SR9009 were identified. Major biotransformation pathways included N-dealkylation, mono-hydroxylation, di-hydroxylation, and combinations thereof. The N-dealkylated metabolite was the predominant metabolite detected in both camel liver homogenate and Cunninghamella elegans, indicating its suitability as a primary analytical target for anti-doping screening. No Phase II metabolites were detected under the experimental conditions employed. Putative structural assignments were supported by characteristic high-resolution tandem mass spectral fragmentation patterns. Conclusions This study provides the first comprehensive metabolic profile of SR9009 in a camel-specific in vitro model. The identification of 17 Phase I metabolites, particularly the predominant N-dealkylated derivative, provides valuable analytical targets for doping control. These findings establish a metabolic foundation for future in vivo studies to determine pharmacokinetics and detection windows in racing camels.

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Integrated 1D‐LC‐HRMS and Heart‐Cutting 2D‐LC–MS for Impurity Profiling and Chiral Separation of Mitiglinide Drug Substance

ABSTRACT Rationale The comprehensive quality control of chiral pharmaceuticals like mitiglinide necessitates simultaneous assessment of chemical impurities and enantiomeric purity, yet conventional workflows address these separately, leading to inefficiency. This study develops a comprehensive analytical strategy to overcome this challenge for the anti-diabetic drug mitiglinide. Methods For chiral analysis, an online heart-cutting two-dimensional liquid chromatography–high-resolution mass spectrometry (2D-LC-HRMS) method was developed. First, impurity profiling of mitiglinide was accomplished using a one-dimensional reversed-phase LC-HRMS (1D-LC-HRMS) method. Subsequently, the 2D-LC-HRMS system achieved enantiomer separation by online coupling of a C18 column (first dimension) with a polysaccharide-based chiral column (second dimension), with the separated analytes detected by an Orbitrap mass spectrometer. Results 1D-LC-HRMS identified five major impurities, structurally characterizing four, with the main component accounting for only 49.12% of the total integrated peak area (relative abundance by EIC peak area normalization, not absolute purity). The 2D-LC-HRMS method achieved effective enantiomer separation. A consistent third minor chromatographic peak was observed across six replicate analyses, which is tentatively assigned as a potential diastereomeric impurity based on stereochemical interpretation of the chromatographic behavior; confirmatory evidence is required for definitive identification. Conclusions This work successfully establishes a comprehensive strategy that efficiently consolidates impurity profiling and chiral purity assessment for mitiglinide. It provides a reliable, more informative approach for the quality control of complex chiral pharmaceuticals.

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Enhancing targeted LC-Q-TOF analysis of emerging contaminants through multivariate mass spectrometry optimization

Publication date: Available online 31 July 2026 Source: Journal of Chromatography A Author(s): Erica Ceccardi, Emanuela Rosi, Caterina Durante, Samuele Pellacani, Marina Di Carro, Emanuele Magi, Barbara Benedetti

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Trace Analysis of Genotoxic Impurity 3‐Amino‐N‐Nitrosopiperidine in Alogliptin Benzoate by Ultra‐Sensitive Liquid Chromatography Tandem Mass Spectrometry

ABSTRACT For many years, it is known as Nitrosamines are responsible for carcinogenicity in human. These nitroso-impurities were found detected in plastics, food items, cigarette, insect killer, water and alcoholic beverages. In 2018 EMA and USFDA claimed the presence of N-nitrosodimethylamine in valsartan used for the treatment of hypertension. Consequently, to assess the adverse effects of nitrosamine generation during storage, in the course of production, or through contaminated supply chains, regulators have issued many guidelines. Hence, there is a crucial need for accurate evaluation technique to measure nitroso-impurities in pharmaceutical products. According to the Carcinogenic Potency Categorization Approach, the allowable intake is 400 ng/day, which translates to an allowable limit of 16 ppm for a 25 mg daily dose. A study was then performed at 1.5 ppm relative to sample concentration, which is less than 10% of this limit. The main aim of this development to optimize and prove suitability of a new liquid chromatography–tandem mass spectrometry method for quantitatively analyzing 3-amino-N-nitrosopiperidine (NTPA) in Alogliptin Benzoate (AGP), achieving detectable response of 0.03 ppm and quantifiable response value of 0.1 ppm relative to sample concentration. Current method was found to be linear with the coefficient of regression of 0.9979. Also, spiking study was performed by the calculating percentage spiked NTPA in the drug, which ranged from 96.67% to 100.14%. The results confirmed that the methodology was reliable, precise, robust and reproducible to quantify NTPA at 1.5 ppm in samples of 2.75 mg/mL concentration.

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Ru/RuO2 Heterostructures for Stable and Active Acidic Water Oxidation via Interfacial Charge Redistribution and Lattice‐Oxygen Participation

A dopant-free Ru/RuO2 metal/oxide heterojunction is constructed for acidic oxygen evolution. Interfacial charge redistribution and work-function modulation promote lattice-oxygen participation while suppressing Ru overoxidation and dissolution, thereby enabling simultaneously enhanced OER activity and durability under acidic conditions. ABSTRACT The rational design and synthesis of Ir-free acidic oxygen evolution catalysts combining high activity with long-term durability remains a formidable challenge. This work demonstrates an addressable and dopant-free strategy through the construction of Ru/RuO2 heterostructures in a solely Ru-based system, wherein interfacial charge redistribution and lattice-oxygen participation jointly promote catalytic enhancement. The catalyst delivers an overpotential of 182 mV at 10 mA·cm−2 and retains operational durability for over 270 h at 100 mA·cm−2 in 0.5 M H2SO4. Spectroscopic characterizations, including X-ray absorption spectroscopy (XAS), Raman spectroscopy, and X-ray photoelectron spectroscopy (XPS), coupled with theoretical calculations, elucidate that interfacial electron transfer from metallic-Ru phase to RuO2 phase, assisted by the formation of a built-in electric field, results in increased work functions across the heterointerface and lowered interfacial Ru oxidation states, suppressing Ru-site overoxidation into soluble RuO4 and accounting for the exceptional durability. Lattice-oxygen participation, which corresponds to excellent activity, was verified by operando investigations, including differential electrochemical mass spectrometry (DEMS), attenuated total reflection surface-enhanced infrared absorption spectroscopy (ATR-SEIRAS), and tetramethylammonium (TMA+) chemical probing. The overall findings establish heterointerface engineering as a powerful tool for the simultaneous enhancement of the coupled activity-stability in Ru-based acidic OER catalysts.

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Mass Spectrometric Characterization of Native Insulin Hexamers and Non‐Native Heptamers: Formation and Stability Across Insulin Analogs

Journal of Mass Spectrometry, Volume 61, Issue 8, August 2026.

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Single-organ proteomics in Drosophila melanogaster larva

Publication date: Available online 30 July 2026 Source: Journal of Proteomics Author(s): Ibon Iloro, Coralia Perez, Jabi Beaskoetxea, Mikel Azkargorta, Iraide Escobes, David Martín, James D. Sutherland, Rosa Barrio, Felix Elortza

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Candidate biomarkers for early Giardia duodenalis infection revealed by time-resolved secretome proteomics

Publication date: Available online 30 July 2026 Source: Journal of Proteomics Author(s): Bo Li, Xi Chen, Linfan Zhang, Yayun Wu, Xiaoying Li, Guanghui Zhao, Junqiang Li, Baole Ma, Jinfeng Zhao, Longxian Zhang

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