bims-metlip 2026-08-30 papers

bims-metlip

Biomed News

on Methods and protocols in metabolomics and lipidomics

Issue of 2026–08–30
35 papers selected by
Sofia Costa, Matterworks

Quantitative Profiling of Metabolites, Sugars, and Lipids in Pollen Tubes by Mass Spectrometry.
Enhanced selectivity for PFAS analysis in complex biological matrices by LC-MS/MS: Application to mouse feces and liver.
PyCompound: An Open-Source Python Package for Spectral-Library Matching in Mass Spectrometry-Based Metabolomics.
Comprehensive non-targeted LC×LC-HRMS lipidomics workflow with active solvent modulation for the characterization of endocrine disruptor effects in zebrafish eleutheroembryos.
Dual-functional SiO2@Cr2O3-C18 sorbent combining Lewis acid-base and hydrophobic interactions for the selective SPE-LC-MS/MS determination of perfluoroalkyl carboxylates in protein-rich biological matrices.
A Sustainable Air-Assisted Liquid-Liquid Microextraction Procedure Coupled to Liquid Chromatography-Tandem Mass Spectrometry for the Determination of 11-Nor-9-carboxy-Δ9-tetrahydrocannabinol and Hexahydrocannabinol Metabolites in Urine.
Quantitative MALDI-MSI Workflow for Lipids with a LC-MS-Based Quality Control Pipeline.
Protocol for preparing small, fragile samples from Schistosoma mansoni and Drosophila melanogaster brains for MALDI mass spectrometry imaging.
Tebuconazole and penconazole - Determination of TEB-OH, TEB-COOH, PEN-OH, and PEN-COOH in urine by LC-MS/MS: Biomonitoring Method - Translation of the German version from 2026.
Salivary cortisone quantification by paper-based ambient mass spectrometry with integrated sample cleanup and enrichment.
From Chromatographic Optimisation to Bioanalysis: HPLC System Comparison and HPLC-QTRAP-MS/MS Determination of Cariprazine and Lurasidone in Human Serum, Urine, and Saliva.
ExpoLib: a framework for an MS/MS exposome library of anthropogenic and natural toxicants and their biotransformation products.
Aptamer-based magnetic solid-phase extraction coupled with LC-MS/MS for the determination of recombinant FGF10 in rat plasma.
TIMSImaging: An open and interoperable workflow for trapped ion mobility mass spectrometry imaging data processing and visualization.
A novel composite based on cellulose and hyper-crosslinked polystyrene for spin column microextraction of fluoroquinolones from human serum prior to their LC-MS/MS determination.
Development and Validation of an LC-MS/MS Method for Simultaneous Quantification of Pazopanib, Cabozantinib, Sorafenib, Axitinib, Tivozanib, Sunitinib, and the Metabolites Sorafenib N-Oxide and Desethyl-Sunitinib in Human Plasma.
From "Undetectable" to "Sensitive Detection": Advances in Derivatization Techniques for LC-MS Analysis of Genotoxic Impurities.
Development and validation of a liquid chromatographic tandem mass spectrometric method (LC-MS/MS) for the determination of bisphenols and parabens in menstrual products after solid-phase extraction.
Bridging the Gap Between Chemiluminescence Immunoassay and Liquid Chromatography-Tandem Mass Spectrometry for Infliximab Monitoring: Method Validation and Clinical Comparability Study.
FERMO: A Dashboard for Biochemometric Prioritization of Molecular Features from Mass Spectral Data.
Simultaneous determination of belzutifan and cabozantinib, oral agents for renal cell carcinoma, in human plasma using high-performance liquid chromatography.
From known chemical space to unannotated metabolites: a cluster-guided retention-time driven framework for biologically informed annotation.
Resolving challenges in detection and quantification of D-2-hydroxyglutarate and L-2-hydroxyglutarate via LC/MS.
MALDI Mass Spectrometry Imaging in Alzheimer's Disease Lipidomics: Matrix Selection, Spatial Lipid Pathology and Emerging Analytical Strategies.
Chemical chemical derivatization-assisted LC-MS for food analysis: Analytical principles, performance enhancement, applications, and perspectives.
Advances in Single-Cell Metabolomics and Their Applications in Disease Biology.
Development and Validation of a Rapid UPLC-MS/MS Determination of Cereulide in ARA Oil and ARA Powder.
Application of Metabolomics in Defence Responses of Brassica Crops.
Truthful visualizations for mass spectrometry imaging enable high-spatial-resolution interactive m/z mapping and exploration.
Simultaneous RP-HPLC-DAD quantification of benzaldehyde, 4-methoxybenzaldehyde, and 3,4-dimethoxybenzaldehyde in fermented acid whey without derivatization.
Exercise and Sports Metabolomics: Analytical Platforms, Metabolite Annotation, Pathway-Level Interpretation, and Biomarker-Panel Readiness.
Dispersive liquid-liquid aerosol phase extraction for the determination of acetylated bromophenols in urine by gas chromatography - mass spectrometry.
Estimation of DNA Methylation in Wheat Using Reverse Phase Ultra-High-Performance Liquid Chromatography Coupled with Mass Spectrometry During Leaf Rust Pathogenesis.
Rapid screening method of glucocorticoid residues in sample-limited cell-cultured meat.
Gas Chromatography Method for Quantitation of Residual Solvent Impurities in Nanoformulations.

Methods Mol Biol. 2026 ;3042 93-109

Quantitative Profiling of Metabolites, Sugars, and Lipids in Pollen Tubes by Mass Spectrometry.

Maneesh Lingwan, Neeta Lohani, Erin Cullen.

  Pollen tube germination and elongation are energy-demanding, metabolically active processes essential for plant reproduction. Recent research combining metabolomics, imaging, and molecular genetics has highlighted the crucial roles of sucrose synthase activity, starch breakdown, and lipid signaling in maintaining pollen tube health and promoting precise growth, particularly under stress conditions. This chapter presents a mass spectrometry-based workflow for high-resolution profiling of metabolites, sugars, and lipids in pollen and pollen tubes. The workflow includes metabolite extraction, chemical derivatization, and chromatographic separation steps compatible with both gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS). These techniques enable simultaneous quantification of polar metabolites, lipids, and fatty acids, capturing the metabolic complexity that supports pollen tube germination, elongation, and fertilization. It also covers the application of internal standards and isotopic normalization to ensure precise quantification and reproducibility across experiments. These analytical approaches collectively provide a foundation for comparative metabolite profiling in reproductive biology and can be adapted to various plant species, lipid signaling, and membrane biogenesis, thereby offering a mechanistic understanding of how pollen tubes sustain polarized growth under both developmental and stress conditions.

Keywords:  Chromatography; Lipid; Mass spectrometry; Metabolites; Pollen tube; Sugar

DOI:  https://doi.org/10.1007/978-1-0716-5308-1_7
J Chromatogr B Analyt Technol Biomed Life Sci. 2026 Aug 22. pii: S1570-0232(26)00353-3. [Epub ahead of print]1283 125264

Enhanced selectivity for PFAS analysis in complex biological matrices by LC-MS/MS: Application to mouse feces and liver.

Gong Zhang, Amélie Blais, Azam F Tayabali, Guillaume Pelletier, Allison Loan, David Prescott, Kristin M Eccles, Rocio Aranda-Rodriguez.

  A sensitive and robust micro-solid phase extraction LC-tandem mass spectrometry (μSPE-LC-MS/MS) method was established for quantifying 23 per- and polyfluoroalkyl substances (PFAS) in mouse feces and liver. Method development identified interferences due to the complex fecal matrix, especially at unit-mass resolution detection, where co-eluting endogenous compounds caused transition-specific interferences. Conventional C18 stationary phases failed to adequately resolve these matrix components, particularly for perfluoroalkyl sulfonic acids (PFSAs), resulting in compromised selectivity. To reduce these matrix effects, alternative stationary phases, including Phenyl-Hexyl (Phe) and Pentafluorophenyl (PFP) columns, were systematically evaluated. The PFP column, operated with acetonitrile in the mobile phase, demonstrated superior chromatographic selectivity and effectively eliminated co-eluting interferences. Notably, transition-specific interferences previously observed for PFOS on C18 columns were resolved, allowing accurate quantification using both m/z 499 → 80 and 499 → 99 transitions. Using the improved selectivity of the PFP column, automated μSPE cleanup combined with isotope-labelled internal standards achieved high extraction efficiencies (96-104%), ensuring reliable accuracy and precision (recoveries 96-99%, RSD <5%) for most analytes despite matrix complexity. These findings demonstrate the importance of chromatographic selectivity, rigorous cleanup, and isotopic correction for quantifying PFAS in highly complex biological matrices using unit-resolution mass spectrometry. The developed workflow provides a reliable analytical platform for toxicokinetic studies and may be applicable to other challenging biological matrices.

Keywords:  LC–MS/MS; Mouse feces; PFOS interferences; Per- and polyfluoroalkyl substances (PFAS); micro solid-phase extraction (μSPE)

DOI:  https://doi.org/10.1016/j.jchromb.2026.125264
Metabolites. 2026 Aug 18. pii: 585. [Epub ahead of print]16(8):

PyCompound: An Open-Source Python Package for Spectral-Library Matching in Mass Spectrometry-Based Metabolomics.

Hunter Dlugas, Xiang Zhang, Xun Bao, Jing Li, Ikuko Kato, Seongho Kim.

  Spectral-library matching is a widely used approach for compound annotation in mass spectrometry (MS)-based metabolomics, yet annotation performance is influenced by spectrum preprocessing, parameter selection, and similarity-measure choice. We present PyCompound, an open-source Python package for spectral-library matching with flexible preprocessing, parameter optimization, and diverse similarity measures for both nominal-resolution and high-resolution mass spectrometry data. PyCompound implements six preprocessing procedures, nineteen similarity measures, including the newly developed Rényi entropy similarity in this work for spectral-library matching, user-defined composite similarity scores, and automated parameter optimization using grid search and differential evolution. The software supports common spectral formats and is accessible through a Python API, command-line interface, and interactive Shiny application. The software was validated using public GNPS LC-MS/MS and WebNIST GC-MS datasets, where the newly developed Rényi entropy similarity demonstrated annotation performance comparable to that of the established Shannon and Tsallis entropy similarity measures. PyCompound provides a flexible platform for practical compound annotation through configurable preprocessing workflows, diverse similarity measures, and automated parameter optimization.

Keywords:  chemoinformatics; compound annotation; entropy; mass spectrometry; metabolomics; similarity measures; spectral library matching

DOI:  https://doi.org/10.3390/metabo16080585
Anal Chim Acta. 2026 10 22. pii: S0003-2670(26)00911-6. [Epub ahead of print]1420 345961

Comprehensive non-targeted LC×LC-HRMS lipidomics workflow with active solvent modulation for the characterization of endocrine disruptor effects in zebrafish eleutheroembryos.

Miriam Pérez-Cova, Laia Navarro-Martin, Gabriel Mazzi Leme, Romà Tauler, Benjamin Piña, Joaquim Jaumot, Dwight R Stoll.

  Lipidomics is a powerful approach for investigating lipid alterations in complex biological systems. However, conventional liquid chromatography coupled to mass spectrometry (LC-MS) may be limited in non-targeted studies due to the high structural diversity and wide concentration range of lipid species. In this work, an optimized comprehensive two-dimensional liquid chromatography coupled to high-resolution mass spectrometry (LC×LC-HRMS) workflow was developed for non-targeted lipidomics. The method combines reversed-phase (RP) separation in the first chromatographic dimension with hydrophilic interaction liquid chromatography (HILIC) separation in the second dimension (RP×HILIC) and incorporates active solvent modulation (ASM) as an interface between the two chromatographic dimensions to improve analytical sensitivity and solvent compatibility across both dimensions. This approach improves sensitivity, solvent compatibility and lipidome coverage compared to conventional LC-MS workflows. The proposed workflow was applied to investigate lipidomic alterations in zebrafish (Danio rerio) eleutheroembryos exposed to the endocrine-disrupting chemical bisphenol A (BPA), using 17β-estradiol (E2) as an estrogenic control. The LC×LC-HRMS approach enabled enhanced lipidome coverage, resulting in the detection of 567 lipid features, of which 134 showed significant alterations associated with endocrine-disruptor exposure. Comparative analysis revealed lipidomic changes consistent with estrogenic responses for both BPA and E2 treatments, while additional lipid alterations suggested potential obesogenic effects under E2 exposure. In summary, the proposed LC×LC-HRMS workflow with ASM provides enhanced separation performance. The obtained results demonstrate the potential of comprehensive multidimensional chromatography combined with multivariate analysis tools for in-depth lipidomics studies in complex biological samples.

Keywords:  Bisphenol A; Chemometrics; Endocrine disrupting chemicals; LC × LC-HRMS; Lipidomics; Zebrafish

DOI:  https://doi.org/10.1016/j.aca.2026.345961
Anal Chim Acta. 2026 10 22. pii: S0003-2670(26)00909-8. [Epub ahead of print]1420 345959

Dual-functional SiO2@Cr2O3-C18 sorbent combining Lewis acid-base and hydrophobic interactions for the selective SPE-LC-MS/MS determination of perfluoroalkyl carboxylates in protein-rich biological matrices.

Mohamed Rashad, Abdullah Alhendal.

   BACKGROUND: The growing concern of perfluoroalkyl substances (PFAS), or "forever chemicals," with serious health implications across the globe necessitates the development of sophisticated analytical techniques for the sensitive analysis of these compounds in biological samples. However, the critical limitations of common SPE sorbents in terms of low selectivity, poor recoveries in biological samples containing high protein content, and high susceptibility to matrix effects in the form of ion suppression have hindered the analysis of these compounds. To overcome these problems, a new dual-functional sorbent was synthesized using a novel composite material consisting of chromia nanoparticles (Cr2O3) and hydrophobic octadecyl chains (C18) on a high-surface-area silica support.
RESULTS: Comprehensive characterization (FTIR, TEM, XPS, BET) confirmed uniform chromia dispersion, successful dense C18 grafting, and high structural stability. Under optimized SPE-LC-MS/MS conditions, the sorbent provided enrichment factors (194-198) for three representative perfluoroalkyl carboxylates (PFHpA, PFOA, PFDoA), yielding ultra-trace limits of detection (LOD) of 25 - 26 ng L-1. To rigorously evaluate its practical utility, the method was validated in standardized synthetic human plasma. The dual-retention mechanism successfully mitigated protein fouling and severe ion suppression, demonstrating an absolute matrix effect of <15% (representing an >85% reduction compared to the direct injection of untreated plasma). Furthermore, the sorbent exhibited robust operational reusability (>50 extraction cycles with <5% efficiency loss).
SIGNIFICANCE: The sorbent exhibited an excellent green analytical profile, achieving a Sample Preparation Metric of Sustainability (SPMS) score of 5.16. This scalable and environmentally friendly platform establishes a highly reliable analytical methodology for PFAS trace analysis in complex biological matrices, offering a robust solution for accurate human biomonitoring and exposure assessment.

Keywords:  Chromia nanoparticles; LC-MS/MS; Lewis acid-base interactions; Matrix effect mitigation; Perfluoroalkyl carboxylic acids (PFCAs); Solid-phase extraction

DOI:  https://doi.org/10.1016/j.aca.2026.345959
J Xenobiot. 2026 Aug 01. pii: 143. [Epub ahead of print]16(4):

A Sustainable Air-Assisted Liquid-Liquid Microextraction Procedure Coupled to Liquid Chromatography-Tandem Mass Spectrometry for the Determination of 11-Nor-9-carboxy-Δ9-tetrahydrocannabinol and Hexahydrocannabinol Metabolites in Urine.

Pamela Cabarcos-Fernández, Sara Odoardi, Serena Mestria, Valeria Valentini, Giulia Biosa, Ana María Bermejo-Barrera, Sabina Strano-Rossi.

  Hexahydrocannabinol (HHC) is a semi-synthetic cannabinoid that has recently emerged in the European market, raising concerns regarding its detection in forensic toxicology. Reliable analytical approaches are required for the determination of HHC metabolites together with conventional cannabis biomarkers in biological samples. This study aimed to develop and validate a sample preparation method based on air-assisted liquid-liquid microextraction (AALLME) coupled to liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the determination of 11-nor-9-carboxy-Δ9-tetrahydrocannabinol (THC-COOH) and HHC metabolites in urine. Urine samples underwent alkaline hydrolysis followed by AALLME using a cyclohexane/ethyl acetate mixture (9:1) prior to LC-MS/MS analysis. Experimental conditions affecting extraction performance were optimized. Method validation was performed according to international guidelines. The method achieved a limit of detection of 2.5 ng/mL and a lower limit of quantification of 5 ng/mL. Precision and accuracy fulfilled the established acceptance criteria across all concentration levels. The applicability of the method was evaluated using eleven authentic THC-COOH-positive routine casework urine samples. THC-COOH was successfully quantified, whereas no HHC metabolites were detected because no urine samples from confirmed HHC users were available. The proposed workflow provides a reduced-solvent sample preparation approach compared with conventional procedures and demonstrated adequate analytical performance for the simultaneous determination of THC-COOH and HHC metabolites in urine.

Keywords:  LC-MS/MS; air-assisted liquid–liquid microextraction; hexahydrocannabinol; metabolites of HHC; urine

DOI:  https://doi.org/10.3390/jox16040143
ACS Meas Sci Au. 2026 Aug 19. 6(4): 1194-1205

Quantitative MALDI-MSI Workflow for Lipids with a LC-MS-Based Quality Control Pipeline.

Felix-Levin Hormann, Sven Heiles.

  Mass spectrometry imaging (MSI) has established itself as a major analytical method in the spatial analysis of biological tissue. Besides qualitative mapping of analyte distributions, recent developments aim to achieve quantification of target analytes. After successful spatial quantification of pharmaceuticals, the field is now shifting toward endogenous biomolecules such as lipids and metabolites. Here, we present a quantitative MSI (qMSI) workflow for endogenous lipids employing matrix-assisted laser desorption/ionization (MALDI) and stable isotope labeled standards (SILS) integrated into a quality control pipeline utilizing quantitative reversed-phase liquid chromatography tandem mass spectrometry (RP-LC-MS/MS). We thoroughly characterized the standard deposition of 13 lipid SILS with a focus on homogeneity and detection errors on mouse heart septa with intraday coefficient of variations (CVs) of ≤15%. Unlike RP-LC-MS/MS for which CV values increase mainly for lowly abundant lipids, variations of lipids in MALDI-MSI are affected by concentration as well as lipid class. Additionally, we compared lipid annotations based on MALDI-MSI and RP-LC-MS/MS and correlated these annotations between the methods based on quantification results. This method comparison revealed that most polar lipids such as PCs or SMs are captured in both MALDI-MSI and RP-LC-MS/MS, whereas less polar compounds such as TGs exhibit limited overlap. These results highlight that a determination of the SILS amounts and lipid annotations via RP-LC-MS/MS is necessary to counter the increase variability during quantitative MALDI-MSI. To demonstrate the analytical capabilities of the quantitative MALDI-MSI pipeline combined with RP-LC-MS/MS quality control steps, we analyzed mouse kidney from a two-hit heart failure with preserved ejection fraction model and achieved good comparability between RP-LC-MS/MS and MALDI-MSI quantitation results. Our data suggest increased ether lipid concentrations in obese mice kidney, likely to cope with increased oxidative stress induced by the progressing disease state of the mice.

Keywords:  MALDI; lipid metabolism; lipidomics; mass spectrometry imaging; quantification

DOI:  https://doi.org/10.1021/acsmeasuresciau.6c00083
STAR Protoc. 2026 Aug 24. pii: S2666-1667(26)00442-9. [Epub ahead of print]7(3): 104789

Protocol for preparing small, fragile samples from Schistosoma mansoni and Drosophila melanogaster brains for MALDI mass spectrometry imaging.

Annika Sabine Mokosch, Max A Müller, Laura Garmendia Sanchez, Stefanie Gerbig, Bernhard Spengler.

  Mass spectrometry imaging is a routine tool for investigating biological samples with high spatial and mass resolution. However, sample preparation, especially when dealing with very small samples, can be challenging. Here, we describe a sample preparation protocol for small and fragile samples based on examples of Schistosoma mansoni parasites and dissected Drosophila melanogaster brains. We describe steps for cryosectioning, matrix application, measurement, and data analysis to enable the acquisition of high-quality spatial lipidomics and metabolomics data from small samples. For complete details on the use and execution of this protocol, please refer to Mokosch et al. and Rorsman et al., respectively.1,2.

Keywords:  Chemistry; Organoids; Protocols in Metabolomics and Lipidomics

DOI:  https://doi.org/10.1016/j.xpro.2026.104789
MAK Collect Occup Health Saf. 2026 ;11(1): Doc019

Tebuconazole and penconazole - Determination of TEB-OH, TEB-COOH, PEN-OH, and PEN-COOH in urine by LC-MS/MS: Biomonitoring Method - Translation of the German version from 2026.

MAK Commission

  The working group "Analyses in Biological Materials" of the German Senate Commission for the Investigation of Health Hazards of Chemical Compounds in the Work Area (MAK Commission) developed and verified this biomonitoring method for the measurement of the most important urinary metabolites of the triazole fungicides tebuconazole and penconazole. Specifically, this method determines (RS)-5-(4-chlorophenyl)-2,2-dimethyl-3-(1H-1,2,4-triazol-1-ylmethyl)-1,3-pentanediol (TEB-OH) and (RS)-5-(4-chlorophenyl)-2,2-dimethyl-3-(1H-1,2,4-triazol-1-ylmethyl)-3-olpentanoic acid (TEB-COOH) as well as 4-(2,4-dichlorophenyl)-5-[1,2,4]-triazol-1-ylpentanol (PEN-OH) and 4-(2,4-dichlorophenyl)-5-[1,2,4]-triazol-1-ylpentanoic acid (PEN-COOH) in urine. After adding isotope-labelled internal standards, the samples are enzymatically hydrolysed to release the analytes from the glucuronide and sulfate conjugates. After online purification, the analytes are separated by liquid chromatography and analysed using tandem mass spectrometry. Calibration is performed using calibration standards prepared in pooled urine and processed analogously to the samples to be analysed. The method provides reliable and accurate analytical results, as shown by the good precision data with standard deviations in the range of 0.1-10.7%. Good accuracy data were obtained with mean relative recoveries in the range of 100-107%. The method is selective, sensitive, and provides quantitation limits of 0.3 μg/l for TEB-OH and TEB-COOH and of 1.0 μg/l for PEN-OH and PEN-COOH.

Keywords:  LC-MS/MS; biomonitoring; penconazole; tebuconazole; urine

DOI:  https://doi.org/10.34865/bi10753496e11_1or
Anal Chim Acta. 2026 10 22. pii: S0003-2670(26)00873-1. [Epub ahead of print]1420 345923

Salivary cortisone quantification by paper-based ambient mass spectrometry with integrated sample cleanup and enrichment.

Tung-Ting Sham, Yufeng Zhou, Barry L Smith, Joanne C Blair, Simon Maher.

  Salivary cortisone correlates more closely with serum free cortisol than salivary cortisol itself, making it the most clinically informative non-invasive marker of adrenal status. Yet its trace endogenous concentration has, until now, largely confined accurate quantification to laboratory-based ultra-high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS). UHPLC-MS/MS demands substantial sample volumes and labour intensive offline preparation. These barriers disproportionately exclude patients who cannot reliably provide the required volumes, including paediatric, elderly and xerostomic populations. Here we report, to our knowledge, the first ambient mass spectrometry method to deliver UHPLC-MS/MS-equivalent quantification of an endogenous steroid biomarker from healthy human saliva across the clinically relevant range. Paper-arrow mass spectrometry (PA-MS), an ambient ionisation technique within the paper spray family, unifies extraction, analyte enrichment and ionisation on a single disposable paper substrate. Cortisone is quantified from 4 μL of saliva, a 50-fold reduction relative to UHPLC-MS/MS, within 10 min. Validated to ICH M10 bioanalytical criteria, the method achieves an LOQ of 0.5 ng/mL, precision ≤8.4% RSD, accuracy 96 to 105%, linearity over 0.5 to 50 ng/mL, and negligible matrix effects. Cross-validation against UHPLC-MS/MS in 28 specimens showed excellent agreement (mean difference 4.17 ± 6.98%). Cortisone, unlike cortisol, was free of co-eluting isobaric interferences, making it well suited to chromatography-free ambient analysis. PA-MS opens a practical route to decentralised endocrine diagnostics for populations underserved by current laboratory-based methods.

Keywords:  Ambient ionisation; Cortisol; Cortisone; Mass spectrometry; Paper spray; Saliva

DOI:  https://doi.org/10.1016/j.aca.2026.345923
Int J Mol Sci. 2026 Aug 07. pii: 7090. [Epub ahead of print]27(16):

From Chromatographic Optimisation to Bioanalysis: HPLC System Comparison and HPLC-QTRAP-MS/MS Determination of Cariprazine and Lurasidone in Human Serum, Urine, and Saliva.

Karol Wróblewski, Anna Petruczynik, Zuzanna Rząd, Hanna Karakuła-Juchnowicz.

  Cariprazine (CAR) and lurasidone (LUR) are antipsychotic drugs used to treat schizophrenia. These drugs are relatively new in clinical practice, and there is a need to optimise and develop analytical methods for their determination in various biological matrices for biomedical analysis. To date, the detection of these drugs has been performed in serum and urine, but there are no methods for determining these drugs in saliva. In the first part of this study, various chromatographic systems were compared using high-performance liquid chromatography with diode array detection (HPLC-DAD) or coupled with quadrupole-linear ion trap tandem mass spectrometry (HPLC-QTRAP-MS/MS), taking into account the retention of tested compounds, system efficiency and peak symmetry. Next, a simple, rapid, and sensitive HPLC-QTRAP-MS/MS method has been developed for the determination of CAR and LUR in human serum, urine, and, for the first time, in saliva samples. Solid-phase extraction (SPE) was used for sample pre-treatment. Quantifications were carried out using a Polar RP column with a mobile phase consisting of acetonitrile and a formate buffer at pH 4.0 in gradient mode. The method was successfully applied for the determination of CAR and LUR in biological samples obtained from psychiatric patients. The findings suggest that saliva may be a non-invasive alternative for the quantification of free levels of investigated drugs, although further studies are required to clarify its relationship with plasma concentrations.

Keywords:  HPLC-DAD; HPLC-QTRAP-MS/MS; biological samples; cariprazine; chromatographic system optimisation; drug determination; lurasidone

DOI:  https://doi.org/10.3390/ijms27167090
Metabolomics. 2026 Aug 26. pii: 145. [Epub ahead of print]22(5):

ExpoLib: a framework for an MS/MS exposome library of anthropogenic and natural toxicants and their biotransformation products.

Vinicius Verri Hernandes, Miguel A Aguilar Ramos, Rolf Breinbauer, Philipp Fruhmann, Hannes Mikula, Monika Ehling-Schulz, Ellen L Zechner, Emily P Balskus, Benedikt Warth.

   INTRODUCTION: Despite technological advancements over the last three decades in small-molecule omics, compound annotation remains a major bottleneck in untargeted metabolomics and non-targeted environmental analysis. This is especially true for exposomics applications, which remain significantly affected by the limited chemical space coverage.
OBJECTIVES: This work aims at describing the development of an MS/MS spectral library containing > 170 relevant xenobiotics from different classes of food, environmental, and microbial toxicants.
METHODS: LC-MS/MS data was acquired using collision-induced dissociation in data dependent acquisition mode under 13 different single collision energies with four additional collision energy spread experiments. A diverse set of compounds including natural toxins produced by bacteria, fungi (mycotoxins), and plants (phytotoxins), as well as anthropogenic chemicals such as bisphenols, phthalates, PFAS chemicals, drugs, consumer care products ingredients, and pesticides, and additional toxicologically relevant chemical classes were screened. Metabolic products for which commercially available reference standards and/or MS/MS spectra are not available in any public or commercial database have been included (e.g. colibactin-DNA-adduct, cereulide, deoxynivalenol-3-glucuronide). Library generation was performed in mzmine.
RESULTS: Open-format data based on representative spectra are provided.This new resource, available at https://zenodo.org/records/20715576 , is aimed at providing a ready-to-use tool for the annotation of key exogenous compounds which are frequently overlooked in clinical metabolomics but may exert potent biological effects. A detailed discussion from a user perspective is provided regarding the library generation workflow in mzmine, aiming at facilitating the work of fellow researchers in the creation of their own in-house libraries.
CONCLUSION: We intend to provide the metabolomics community with better tools for exposomics research and to reduce perceived barriers in developing specialized MS/MS libraries for widening chemical space coverage and increasing quality and confidence.

Keywords:  (Xeno-)metabolomics; Endocrine disrupting chemicals; Exposure and effect analysis; High-resolution mass spectrometry (HRMS); Microbiome-related genotoxins; Structural elucidation; Suspect and non-targeted screening

DOI:  https://doi.org/10.1007/s11306-026-02481-x
J Chromatogr A. 2026 Aug 20. pii: S0021-9673(26)00704-1. [Epub ahead of print]1786 467377

Aptamer-based magnetic solid-phase extraction coupled with LC-MS/MS for the determination of recombinant FGF10 in rat plasma.

Xiaojun Cai, Xu Xie, Yile Xie, Yang Yu, Zhou Li, Xiangzhun Chen, Zhiyuan Zhou, Yuepiao Cai, Jinmei Yang, Lei Jin.

  Establishing sensitive and reliable methods for the quantification of structurally intact recombinant protein therapeutics in biological matrices remains essential for pharmacokinetic studies but continues to be analytically challenging. In this study, we developed an aptamer-based magnetic solid-phase extraction coupled with liquid chromatography-tandem mass spectrometry (Apt-MSPE-LC-MS/MS) method for the quantitative determination of recombinant fibroblast growth factor 10 (rFGF10) in rat plasma. The aptamer-mediated extraction enabled selective enrichment of rFGF10 prior to LC-MS/MS analysis, resulting in a 7.2-fold increase in analytical sensitivity compared with direct LC-MS/MS analysis. The developed method exhibited satisfactory linearity over a concentration range of 1.0-50 μg⋅mL⁻¹ (R² = 0.998), with acceptable precision (6.3-13.6%), accuracy (%Bias, -5.8% to 8.0%), and a limit of detection (LOD) of 0.53 μg⋅mL⁻¹. Matrix effect evaluation demonstrated acceptable and reproducible matrix effects, with coefficient of variation (CV) values below 15%. The validated Apt-MSPE-LC-MS/MS method was successfully applied to the pharmacokinetic analysis of rFGF10 in rats. Overall, this workflow provides a selective and reliable strategy for the quantitative analysis of structurally intact recombinant proteins in complex biological matrices.

Keywords:  Aptamer; Liquid chromatography–tandem mass spectrometry; Magnetic solid-phase extraction; Pharmacokinetic analysis; Recombinant fibroblast growth factor 10

DOI:  https://doi.org/10.1016/j.chroma.2026.467377
Bioinformatics. 2026 Aug 25. pii: btag624. [Epub ahead of print]

TIMSImaging: An open and interoperable workflow for trapped ion mobility mass spectrometry imaging data processing and visualization.

Yinyue Zhu, Kylie Bemis, Sai Srikanth Lakkimsetty, Mujia Jenny Li, Larissa Chiara Meyer, Andreas Weber-Steinhilber, Melanie Christine Föll, Olga Vitek.

SUMMARY: Mass spectrometry imaging (MSI) enables high-throughput spatial mapping of molecules, but the lack of chromatographic separation limits its utility for complex biological samples. Ion mobility (IM) provides key orthogonal separation. However, open-source tools dedicated for IM-MSI data analysis remain scarce and the integration of ion mobility information into downstream analysis is underexplored. Here, we present TIMSImaging, an open-source workflow for processing and visualization of MALDI-TIMS-MS data from Bruker timsTOF instruments. TIMSImaging incorporates a graph-based two-dimensional feature extraction algorithm for separation of isobaric peaks by ion mobility, supports collision cross section (CCS) calculation, and exports results as imzML files with ion mobility for downstream analysis. We demonstrate its capabilities on three case studies spanning different sample types, analyte types, and downstream tasks.
AVAILABILITY AND IMPLEMENTATION: TIMSImaging is released as open-source software under the MIT License. The source code, installation instructions, documentation, and case study Vignettes are available at https://github.com/YinyueZhu/TIMSImaging.

DOI: https://doi.org/10.1093/bioinformatics/btag624
Talanta. 2026 Aug 20. pii: S0039-9140(26)01143-4. [Epub ahead of print]312(Pt B): 130487

A novel composite based on cellulose and hyper-crosslinked polystyrene for spin column microextraction of fluoroquinolones from human serum prior to their LC-MS/MS determination.

Yana A Kulinich, Alexander Yu Popov, Anton A Grigoriev, Veronika V Tolmacheva, Artem O Melekhin, Vladimir V Apyari, Stanislava G Dmitrienko.

  A composite based on cellulose and hyper-crosslinked polystyrene (cellulose-HCP) was developed as a novel sorbent for micro-solid-phase extraction (μSPE) using spin columns. The composite was prepared via a simple, low-cost, and environmentally friendly procedure based on cellulose dissolution in NaOH solution. The extraction conditions were systematically optimized with respect to sorbent amount, centrifugation parameters, solution pH, and desorption conditions. Quantitative extraction of fluoroquinolones (98%, RSD ≤ 2%) were achieved at pH 5.8 using spin columns packed with three composite disks and centrifugation at 1500 rpm for 1 min. Efficient desorption was obtained using an acetonitrile-ammonia mixture (1:1, v/v). The method was validated using liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the determination of fluoroquinolones in human serum. Good linearity was observed in the range of 2.0-400 μg L-1 (R2 ≥ 0.9916) with limits of detection of 0.3-0.7 μg L-1. Matrix effects were negligible (ME ≤ ±16%), and satisfactory reproducibility was achieved (RSD < 18%). The total sample preparation took 24 min and required only a centrifuge without protein precipitation. The proposed approach combines simplicity, rapidity, and low solvent consumption, meeting the principles of green sample preparation and demonstrating its suitability for the determination of fluoroquinolones in human serum.

Keywords:  Centrifugal-assisted; Green sample preparation; Simultaneous quantification; Therapeutic drug monitoring

DOI:  https://doi.org/10.1016/j.talanta.2026.130487
Pharmaceuticals (Basel). 2026 Jul 28. pii: 1180. [Epub ahead of print]19(8):

Development and Validation of an LC-MS/MS Method for Simultaneous Quantification of Pazopanib, Cabozantinib, Sorafenib, Axitinib, Tivozanib, Sunitinib, and the Metabolites Sorafenib N-Oxide and Desethyl-Sunitinib in Human Plasma.

Eva Greibe, Jakob N Henriksen, Niels Fristrup, Elke Hoffmann-Lücke.

  Background: Pazopanib, cabozantinib, sorafenib, axitinib, tivozanib, and sunitinib are widely used tyrosine kinase inhibitors (TKIs) targeting the vascular endothelial growth factor receptor signaling pathway. Considerable interindividual pharmacokinetic variability, together with established exposure-toxicity relationships, underlines the need for reliable analytical methods for therapeutic drug monitoring (TDM). The aim of this study was to develop and validate an LC-MS/MS method for the simultaneous determination of these six TKIs and the two metabolites, sorafenib N-oxide and desethyl-sunitinib, in human plasma and to investigate their stability under relevant preanalytical conditions. Methods: Plasma samples were prepared by protein precipitation and analyzed using LC-MS/MS with positive electrospray ionization and multiple reaction monitoring. Validation included assessments of linearity, accuracy, precision, lower limit of quantification, dilution integrity, carry-over, and stability under different storage conditions. As proof of concept, the method was applied to plasma samples from patients receiving TKI treatment. Results: All analytes demonstrated linear responses across the validated calibration ranges. Validation parameters met predefined acceptance criteria for all analytes, except for sorafenib N-oxide, which exhibited CV% values slightly above the acceptance limit (15%). All compounds remained stable during storage at -80 °C for at least five years. Furthermore, all analytes were stable at room temperature for 24 h, except sorafenib N-oxide, which showed instability under these conditions. Analysis of patient samples confirmed the suitability of the method for clinical application. Conclusions: We established and validated a fast, sensitive, and accurate LC-MS/MS method enabling simultaneous quantification of six TKIs and two metabolites in human plasma. The analytical method was successfully applied to plasma samples from patients in treatment with the six TKIs, demonstrating proof of concept for its application in TDM.

Keywords:  axitinib; cabozantinib; desethyl-sunitinib; pazopanib; renal cell carcinoma; sorafenib; sorafenib N-oxide; stability; sunitinib; tivozanib; tyrosine kinase inhibitor

DOI:  https://doi.org/10.3390/ph19081180
Molecules. 2026 Aug 19. pii: 2889. [Epub ahead of print]31(16):

From "Undetectable" to "Sensitive Detection": Advances in Derivatization Techniques for LC-MS Analysis of Genotoxic Impurities.

Xingchen Wang, Zhuzi Chen, Shunli Ji.

  Many genotoxic impurities (GTIs) remain "invisible" to conventional LC-MS due to poor ionization or chemical instability under electrospray ionization, yet their sub-ppm acceptable intake limits under ICH M7(R2) demand exceptional analytical sensitivity. Derivatization-the chemical introduction of ionizable moieties, stable tags, or MS/MS information carriers-offers a powerful strategy to overcome this limitation. This review provides a critical systematic overview of derivatization techniques for LC-MS analysis of GTIs over the past decade (2015-2025, based on a literature search across PubMed, Web of Science, and Scopus). We construct a functional-group-based strategic framework covering alkyl halides, nitroaromatics, sulfonyl chlorides, hydroxylamine, alcohols, aldehydes, carboxylic acids, and amines, while placing specific emphasis on typical impurities within these classes such as methyl iodide, methyl chloride, nitrobenzene, and benzenesulfonyl chloride, and discuss the evolution of reagents from simple "reaction tags" to "MS/MS information carriers" that provide characteristic neutral losses or product ions for enhanced selectivity. Quantitative analysis reveals that derivatization typically enhances ESI response by 2-3 orders of magnitude, consistently achieving LODs below 1 ppm-the ICH M7(R2) threshold. Key analytical trade-offs are critically evaluated, including the balance between derivatization efficiency and reaction time, by-product management, and the fundamental kinetic and chromatographic constraints that render post-column derivatization impractical for most GTIs. We conclude with perspectives on high-throughput automation, smart multifunctional reagents, online integration, and green chemistry, aiming to provide a practical roadmap for developing robust, sensitive, and regulatory-compliant LC-MS methods for GTI control.

Keywords:  ICH M7; LC-MS/MS; derivatization; genotoxic impurities; sample pretreatment

DOI:  https://doi.org/10.3390/molecules31162889
Anal Chim Acta. 2026 10 22. pii: S0003-2670(26)00891-3. [Epub ahead of print]1420 345941

Development and validation of a liquid chromatographic tandem mass spectrometric method (LC-MS/MS) for the determination of bisphenols and parabens in menstrual products after solid-phase extraction.

Pavlos Voutsas, Natasa Kalogiouri, Victoria Samanidou.

   BACKGROUND: Currently, thousands of personal care and hygiene products are widely produced and distributed worldwide. Even though in recent years legislation concerning these compounds has become stricter than ever before, limited information is provided on product labels.
RESULTS: In this research, a novel method was developed and validated for the determination of parabens and bisphenols in menstrual products, including menstrual pads, panty liners, and tampons. Sample preparation was carried out using solid-phase extraction (SPE) prior to analysis by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) equipped with an electrospray ionization (ESI) source. The optimized method showed high recoveries ranging from 84.0 to 112.0%, with relative standard deviation (RSDs) not exceeding 20.0% at the limit of quantification (LOQ) and 15.0% at the remaining concentration levels. The method was also applied to real samples, in which bisphenol A was the most prevalent compound detected, while parabens were also determined in several analysed samples.
SIGNIFICANCE AND NOVELTY: This work introduces a rapid LC method for the simultaneous extraction, purification, and quantification of bisphenols and parabens in menstrual products, with reduced solvent consumption and analysis time. The applicability, analytical performance, and environmental sustainability of the method were evaluated using established assessment tools. The results demonstrate that the proposed method aligns with the principles of green chemistry.

Keywords:  Bisphenols; LC-MS/MS; Menstrual products; Parabens; SPE

DOI:  https://doi.org/10.1016/j.aca.2026.345941
J Sep Sci. 2026 Sep;49(9): e70515

Bridging the Gap Between Chemiluminescence Immunoassay and Liquid Chromatography-Tandem Mass Spectrometry for Infliximab Monitoring: Method Validation and Clinical Comparability Study.

Ying Xia, Ming-Mei Zhu, Meng-Qing Xiao, Chen-Chun Zhong, Yu-Jun Zhou, Yi-Hao Xue, Shi-Wei Zheng, Li Li, Feng Chen.

  Infliximab (IFX) is a cornerstone biologic for pediatric inflammatory bowel disease, where therapeutic drug monitoring (TDM) is essential but complicated by inter-platform variability. Systematic comparison of automated immunoassays with reference methods is therefore warranted. This study aimed to validate an in-house chemiluminescence immunoassay (CLIA) for IFX quantification and evaluate its agreement with liquid chromatography-tandem mass spectrometry (LC-MS/MS). A CLIA kit was validated by assessing the limit of blank, specificity, linearity, accuracy, precision, matrix equivalence, and stability. IFX concentrations in 52 pediatric plasma samples were measured by CLIA and a reference LC-MS/MS method. Comparability was evaluated using linear regression, Passing-Bablok regression, and Bland-Altman analysis. CLIA showed excellent performance (lower limit of quantification: 0.640 µg/mL; specificity <10.0% interference; reportable range: 0.640-462 µg/mL). LC-MS/MS yielded higher concentrations than the CLIA (median: 6.25 vs. 2.87 µg/mL). Strong correlation was observed (r = 0.9101) with no significant deviation from linearity (Passing-Bablok, p > 0.05). However, Bland-Altman revealed substantial mean relative bias (67.9%) and poor categorical agreement (52% concordance) within the therapeutic range (3-7 µg/mL), indicating non-interchangeability. We validated a rapid, automated CLIA suitable for high-throughput TDM. Despite strong correlation, significant quantitative differences preclude direct result substitution, underscoring the need for method-specific therapeutic thresholds.

Keywords:  LC‐MS/MS; chemiluminescence immunoassay; children; infliximab; therapeutic drug monitoring

DOI:  https://doi.org/10.1002/jssc.70515
ACS Meas Sci Au. 2026 Aug 19. 6(4): 1011-1022

FERMO: A Dashboard for Biochemometric Prioritization of Molecular Features from Mass Spectral Data.

Mitja M Zdouc, Hannah E Augustijn, Nataliia V Machushynets, Lina M Bayona, Sylvia Soldatou, Niek F de Jonge, Sarah Casu, Marcel Jaspars, Gilles P van Wezel, Marnix H Medema, Justin J J van der Hooft.

  Many natural products can selectively modulate biological processes, making them prime candidates for drug discovery. However, the complexity of biological samples makes clear attribution of activity to molecules challenging, thereby hampering hypothesis-driven prioritization, with liquid chromatography-tandem mass spectrometry routinely detecting hundreds of molecules per sample. Existing biochemometric tools typically focus on facilitating data-driven exploration to support manual interpretation, rather than more objective, data-driven prioritization and hypothesis generation. Here, we introduce FERMO, a free online dashboard interface for biochemometrics-based prioritization of molecular features and samples. FERMO accepts qualitative and quantitative bioactivity assay data and further integrates group metadata and results from genome mining. FERMO performs automated data processing, organization, and annotation, supporting prioritization with the calculation of custom scores. FERMO supports both exploratory and targeted analysis through efficient interactive visualization, reproducible prioritization, and data filtering. We demonstrate FERMO's utility in benchmarking studies prioritizing bioactive natural products from complex biological matrices. FERMO is freely available at https://fermo.bioinformatics.nl/.

Keywords:  Antibiotics; Computational Metabolomics; Data Integration; Mass Spectrometry; Natural Products; Secondary/Specialized Metabolites

DOI:  https://doi.org/10.1021/acsmeasuresciau.6c00022
Drug Discov Ther. 2026 Aug 26.

Simultaneous determination of belzutifan and cabozantinib, oral agents for renal cell carcinoma, in human plasma using high-performance liquid chromatography.

Yoshito Gando, Takeo Yasu.

  Belzutifan and cabozantinib are oral drugs for advanced renal cell carcinoma, which display exposure-dependent toxicities. No validated method exists for simultaneously quantifying these drugs in human plasma. In this study, we developed and validated a high-performance liquid chromatography-ultraviolet method for simultaneously quantifying belzutifan and cabozantinib in human plasma. Plasma samples (50 μL) were processed via simple protein precipitation. Chromatographic separation on a C18 column was performed isocratically with 0.5% KH2PO4 (pH 4.5)/acetonitrile (47/53, v/v) at 1.0 mL/min, with UV detection at 230 nm. Performance characteristics (calibration, precision, accuracy, recovery, stability, selectivity, and interference by concomitant drugs) were systematically evaluated. Linear calibration ranges were 50-4,000 ng/mL for belzutifan and 25-4,000 ng/mL for cabozantinib with excellent linearity (r2 ≥ 0.9998). Intra- and interday coefficients of variation were ≤ 5.41% for belzutifan and ≤ 12.25% for cabozantinib. Both analytes were stable under bench-top, short-term, long-term, and freeze-thaw conditions. No interference from endogenous plasma components or six commonly co-administered medications was observed. The newly developed assay for the simultaneous quantification of belzutifan and cabozantinib in human plasma is rapid, requires minimal sample volumes, and demonstrates robust analytical performance.

Keywords:  belzutifan; cabozantinib; high-performance liquid chromatography–ultraviolet; human plasma concentration

DOI:  https://doi.org/10.5582/ddt.2026.01033
Metabolomics. 2026 Aug 26. pii: 146. [Epub ahead of print]22(5):

From known chemical space to unannotated metabolites: a cluster-guided retention-time driven framework for biologically informed annotation.

Dipendra Bhandari, Henry A Paz, Keith Henderson, Kiran Kumar Adepu, Ahmad Mani-Varnosfaderani, Hailemariam Abrha Assress, Brian D Piccolo, Renny S Lan, Elisabet Børsheim, Colin D Kay, Sree V Chintapalli.

   INTRODUCTION: Untargeted metabolomics often results in a significant portion of unannotated metabolites, or "metabolic dark matter," which hinders biological interpretation.
OBJECTIVES: A two-step analytical approach was developed to systematically prioritize and interpret unannotated metabolites using plasma LC-MS/MS data from pregnant women with obesity as a biologically relevant test dataset.
METHODS: The first step involved clustering 1,021 known metabolites into ten structurally coherent groups based on the Tanimoto similarity, thus defining the biologically relevant chemical space of the dataset. These metabolites were further characterized by Absorption, Distribution, Metabolism, and Excretion (ADME) profiling, protein target prediction, molecular docking and Kyoto Encyclopedia of Genes and Genomes pathway mapping analysis, to establish biological plausibility and functional perspective. Candidate structures for 1,836 unannotated features were retrieved from PubChem using molecular formula and molecular weight matching within a ±0.5 Da tolerance.
RESULTS: This search yielded 569,115 candidate structures, of which 368,197 unique structures were retained after curation. Tanimoto coefficient filtering reduced the candidate pool to 19,868 structurally plausible candidates, and retention time-based prioritization further refined this set to 418 high confidence candidate annotations, including 83 database-supported candidates identified through HMDB and LIPID MAPS structure database cross-referencing. RT-based prioritization effectively distinguished positional isomers sharing the same molecular formula by incorporating agreement between predicted and experimentally observed retention times.
CONCLUSION: This improved discrimination among structurally similar candidates, expanded metabolite annotation confidence, and provided a scalable framework for prioritizing dark matter metabolites in untargeted metabolomics.

Keywords:  Dark matter; Gene; In silico; Metabolomics; Protein; Retention time; Target prediction

DOI:  https://doi.org/10.1007/s11306-026-02520-7
Methods. 2026 Aug 22. pii: S1046-2023(26)00183-0. [Epub ahead of print]255 48-61

Resolving challenges in detection and quantification of D-2-hydroxyglutarate and L-2-hydroxyglutarate via LC/MS.

Tyrone Dowdy, Mioara Larion.

  D-2-hydroxyglutarate and L-2-hydroxyglutarate (D-2HG/L-2HG) are typically metabolites of non-specific enzymatic reactions that are kept in check by the housekeeping enzymes, D-2HG /L-2HG dehydrogenase (D2HGDH/L2HGDH). In certain disease states, such as D-2HG or L-2HG aciduria and cancers, the accumulation of these biomarkers interferes with the activity of oxoglutarate-dependent enzymes that regulate bioenergetic metabolism, histone methylation, post-translational modification, and protein expression. The complex role of D-2HG in tumorigenesis has driven metabolomics investigations in cancer research. Meanwhile, L-2HG is produced by non-specific action of malate dehydrogenase and lactate dehydrogenase under acidic or hypoxic environments. Characterization of the divergent effects of D-2HG/L-2HG on the activity of specific enzymes in diseased metabolism depends on their accurate quantification via mass spectrometry. Despite advancements in high-resolution mass spectrometry (HRMS), challenges are typically encountered when attempting to resolve isobaric and isomeric metabolites such as D-2HG/L-2HG for quantitative analysis. Herein, D-2HG/L-2HG derivatization and liquid chromatography (LC) MS quantification methods were examined. This outcome led to the development of a high-throughput LC-HRMS approach that permits concomitant quantification of the D-2HG and L-2HG enantiomers. This approach also provided the added benefit of quantifying other intermediates that are dysregulated within interconnecting pathways. A calibration curve was obtained over the linear range of 0.8-104 nmol/mL with R2 ≥ 0.995 for each enantiomer. The LC/MS-based assay had an overall precision with intra-day CV % ≤ 8.0 and inter-day CV % ≤ 6.3 across the quality control level for commercial standard and pooled biological samples; relative error % ≤ 2.7 for accuracy; and resolution, Rs = 1.6 between 2HG enantiomers (m/z 147.030), D-2HG and L-2HG (at retention time of 5.82 min and 4.75 min, respectively) following chiral derivatization with diacetyl-L-tartaric anhydride (DATAN). Our methodology was applied to disease-relevant samples to illustrate the implications of proper enantioselective quantification of both D-2HG and L-2HG. The method allows scaling to large cohorts of clinical samples in the future.

Keywords:  D-2-hydroxyglutarate and L-2-hydroxyglutarate, D-2HG/L-2HG; Diacetyl-L-tartaric anhydride, DATAN; LC/MS; Mutant IDH1 glioma; N-p-tosyl-L-phenylalanyl chloride, TSPC

DOI:  https://doi.org/10.1016/j.ymeth.2026.08.005
Int J Mol Sci. 2026 Aug 07. pii: 7074. [Epub ahead of print]27(16):

MALDI Mass Spectrometry Imaging in Alzheimer's Disease Lipidomics: Matrix Selection, Spatial Lipid Pathology and Emerging Analytical Strategies.

David Aebisher, Anna Krzysztofińska, Barbara Smolak, Patrycja Bernat, Wiktoria Czajka, Magdalena Kowal, Sylwia Krasoń, Klaudia Dynarowicz, Wiesław Guz, Dorota Bartusik-Aebisher.

  Alzheimer's disease (AD) involves not only amyloid-β and tau pathology but also extensive disturbances in lipid metabolism, membrane organization, neuroinflammatory signaling, and tissue homeostasis. Conventional lipidomics has identified changes in phospholipids, sphingolipids, sulfatides, ceramides, gangliosides, and cholesterol-related pathways, but tissue homogenization removes their anatomical context. The aim of this review is to critically assess how matrix selection, sample preparation, ionization polarity, and emerging analytical strategies influence the detection and interpretation of spatial lipid alterations specifically associated with AD neuropathology. Current evidence shows that AD-related lipid remodeling is region- and lesion-specific, with recurrent findings including ganglioside accumulation, sulfatide depletion, ceramide-related alterations, phospholipid remodeling, lysosomal lipid changes, and disturbed cholesterol homeostasis within or around amyloid plaques. Matrix chemistry strongly influences lipid-class coverage, ionization efficiency, spectral background, adduct formation, spatial resolution, and biological interpretation. Matrix-Assisted Laser Desorption/Ionization with Laser-Induced Post-Ionization (MALDI-2), ion mobility, reactive matrices, on-tissue derivatization, structural lipidomics, single-cell imaging, and spatial multiomics are expanding molecular coverage and annotation confidence. However, broader translation requires standardized workflows, structurally validated assignments, quantitative quality control, larger human cohorts, and improved interlaboratory reproducibility. Collectively, the available evidence indicates that the principal value of Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry Imaging (MALDI-MSI) in AD lies not merely in detecting altered lipid abundance, but in resolving lesion-specific lipid microenvironments whose interpretation depends directly on matrix chemistry, spatial resolution, and structural validation.

Keywords:  Alzheimer’s disease; MALDI mass spectrometry imaging; amyloid plaques; gangliosides; ion mobility; matrix selection; molecular neuropathology; on-tissue derivatization; spatial lipidomics; sulfatides

DOI:  https://doi.org/10.3390/ijms27167074
J Chromatogr A. 2026 Aug 22. pii: S0021-9673(26)00709-0. [Epub ahead of print]1786 467382

Chemical chemical derivatization-assisted LC-MS for food analysis: Analytical principles, performance enhancement, applications, and perspectives.

Xiaoyu Xie, Chunling Zhao, Ruonan Zhang, Shuhui Wang, Chao Li, Zhihong Yan.

  The increasing complexity of food matrices and the demand for comprehensive foodomics have exposed the limitations of conventional LC-MS workflows in chemical coverage, structural annotation, and quantitative analysis. Chemical derivatization provides an effective strategy to address these challenges through selective functional-group modification and integration with advanced analytical approaches, enabling improved chromatographic performance, ionization efficiency, and structural characterization. Recent advances integrating derivatization with high-resolution mass spectrometry, stable isotope encoding, retention time prediction, and computational annotation have further expanded its role from sensitivity enhancement to information-rich analysis. This review summarizes recent developments in derivatization-assisted LC-MS strategies for food analysis from 2013 to 2026. Derivatization approaches targeting amino, hydroxyl, carbonyl, carboxyl, and other functional groups are systematically discussed, with emphasis on reaction principles, analytical advantages, representative applications, and methodological limitations. Particular attention is given to the role of derivatization in improving molecular coverage, structural confidence, and quantitative reliability in complex food matrices. Remaining challenges include balancing reaction selectivity and applicability, improving workflow standardization, and enhancing automation compatibility. Future advances will rely on integrating selective derivatization chemistry with advanced MS technologies and computational approaches to achieve more comprehensive, reliable, and accurate characterization of chemically diverse compounds in complex food systems.

Keywords:  Chemical derivatization; Food analysis; Liquid chromatography-mass spectrometry

DOI:  https://doi.org/10.1016/j.chroma.2026.467382
OMICS. 2026 Aug 26. 15578100261480571

Advances in Single-Cell Metabolomics and Their Applications in Disease Biology.

Jie Du, Huihui Tao, Yujie Zhang, Mengyao Wu, Erjiao Hao, Tianyuan Li, Min Feng, Feng Zhu, Ruijie Yang, Ning Ni, Yong Dai.

  Single-cell metabolomics (SCM) provides a functional view of cellular heterogeneity by measuring metabolic states at cellular or subcellular resolution. Unlike bulk metabolomics, SCM can reveal rare metabolic cell states, spatially restricted metabolic niches, dynamic pathway activity, and treatment-associated metabolic adaptations. This review summarizes major SCM strategies, including spatial mass spectrometry imaging (MSI), isolated single-cell mass spectrometry (MS), isotope-assisted approaches, fluorogenic probes, and vibrational spectroscopy-based methods. Rather than treating these platforms as interchangeable technologies, we organize the review around a question-driven framework linking biological questions to platform selection, data structures, computational workflows, and interpretation boundaries. We also discuss major analytical challenges, including limited sample amount, missing values, ion suppression, batch effects, metabolite annotation uncertainty, and incomplete standardization. Disease-related studies suggest that SCM can identify recurrent metabolic programs, candidate biomarkers, and intervention-relevant metabolic nodes, but most applications remain at the discovery or early translational stage. Future progress will require standardized workflows, improved quantitative confidence, multimodal integration, and validation in clinically relevant models and cohorts.

Keywords:  biomarker discovery; mass spectrometry; metabolic heterogeneity; single-cell metabolomics; standardized workflows

DOI:  https://doi.org/10.1177/15578100261480571
Toxins (Basel). 2026 Jul 23. pii: 320. [Epub ahead of print]18(8):

Development and Validation of a Rapid UPLC-MS/MS Determination of Cereulide in ARA Oil and ARA Powder.

Xia Cui, Jing Zhang, Zixiao Zhou, Ziyi Wang, Jing Hou, Rong Zhao, Sai Fan.

  Arachidonic acid (ARA) is an essential long-chain polyunsaturated fatty acid for infant growth and development, commonly added to infant formula and supplementary foods as ARA oil or ARA powder. Since December 2025, infant formula products have been recalled in several countries due to cereulide contamination in ARA ingredients. Cereulide, a heat-resistant emetic toxin produced by Bacillus cereus, presents a serious threat to infant health. However, specific analytical methods for cereulide detection in ARA raw materials remain scarce, hindering effective quality control and source prevention of food safety hazards. This study developed and validated a rapid ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for the quantitative determination of cereulide in ARA oil and powder. Sample pretreatment was optimized according to matrix characteristics: for microencapsulated ARA powder, water dissolution was performed to release cereulide before acetonitrile extraction, whereas direct extraction was applied to ARA oil. Subsequent HLB-P pass-through solid-phase extraction effectively eliminated matrix interference in both matrices. The method exhibited excellent linearity (R2 > 0.999) from 0.1 to 20 μg·L-1, with an LOD of 0.03 μg·kg-1 and an LOQ of 0.1 μg·kg-1, respectively. Spike recoveries reached 90.7-107.0% for ARA oil (RSDs ≤ 6.5%) and 88.5-107.9% for ARA powder (RSDs ≤ 7.0%). This rapid and reliable method provides a critical analytical tool to monitor cereulide in ARA raw materials and mitigate infant safety hazards at the source.

Keywords:  ARA oil; ARA powder; HLB-P; UPLC-MS/MS; cereulide

DOI:  https://doi.org/10.3390/toxins18080320
Metabolites. 2026 Aug 10. pii: 563. [Epub ahead of print]16(8):

Application of Metabolomics in Defence Responses of Brassica Crops.

Yufei Li, Junxing Lu.

  Brassica crops, encompassing globally important vegetables and oilseeds, face severe threats from diverse biotic and abiotic stresses. Plant secondary metabolites constitute the chemical foundation of defence, and metabolomics has emerged as an effective systems biology tool for comprehensively dissecting stress-induced metabolic changes. Recent progress in applying metabolomics to elucidate defence mechanisms in Brassica crops is systematically synthesised here. Major stresses confronting Brassica crop production and the metabolic basis of plant defence are first outlined. Current analytical platforms, including liquid chromatography-mass spectrometry, gas chromatography-mass spectrometry, ion mobility spectrometry, and mass spectrometry imaging, are critically evaluated alongside data processing workflows and multi-omics integration strategies. Key defence-related metabolite classes identified in Brassica crops, notably glucosinolates (GSLs) and their hydrolysis products, phenolic compounds, and lipid-derived signalling molecules, are surveyed with emphasis on their respective functions in biotic and abiotic stress responses. Metabolomics has been instrumental in revealing distinct metabolic reprogramming patterns triggered by diverse stresses, including pathogen infection, insect herbivory, drought, salinity, temperature extremes, and heavy metal stress. Metabolomics-informed crop improvement strategies, including marker-assisted breeding, genetic and metabolic engineering, and precision agronomic practices, are discussed together with current technical bottlenecks and future directions involving artificial intelligence, metabolic modelling, and spatial metabolomics. The compiled knowledge provides a comprehensive reference for leveraging metabolomics to enhance stress resilience and sustainable production of Brassica crops.

Keywords:  Brassica crops; metabolomics; plant defence; secondary metabolites; stress responses

DOI:  https://doi.org/10.3390/metabo16080563
Sci Adv. 2026 Aug 28. 12(35): eaed3650

Truthful visualizations for mass spectrometry imaging enable high-spatial-resolution interactive m/z mapping and exploration.

Jacob Gildenblat, Jens Pahnke.

Mass spectrometry imaging (MSI) produces high-dimensional molecular data, but practical interpretation remains limited by visualizations that incompletely preserve global structure. We present MSI-VISUAL, an open-source framework for interactive MSI analysis that integrates truthful dimensionality reduction visualizations with region-of-interest selection, statistical comparison, and direct mass/charge ratio (m/z) mapping. MSI-VISUAL introduces four visualization strategies: SALO and SPEAR (optimization-based methods designed to improve global structure preservation across distance metrics) and TOP3 and PR3D (lightweight approaches for memory-efficient, rapid visualization of large datasets). Across benchmarks and lipidomics case studies, including mouse brain and kidney pathology examples, the proposed methods outperform commonly used alternatives in our benchmarks, improve detection of subtle tissue differences, and reveal fine molecular-anatomical patterns that support biological insights. These results establish MSI-VISUAL as a scalable framework for discovery-oriented and diagnostic MSI workflows.

DOI: https://doi.org/10.1126/sciadv.aed3650
J Chromatogr B Analyt Technol Biomed Life Sci. 2026 Aug 26. pii: S1570-0232(26)00355-7. [Epub ahead of print]1283 125266

Simultaneous RP-HPLC-DAD quantification of benzaldehyde, 4-methoxybenzaldehyde, and 3,4-dimethoxybenzaldehyde in fermented acid whey without derivatization.

Sabrina Stýblová, Sonja Berensmeier.

  A reversed-phase high-performance liquid chromatography method with a diode-array detector (RP-HPLC-DAD) was developed and validated for the simultaneous determination of 3,4-dimethoxybenzaldehyde (3,4-DMBA), benzaldehyde (BA), and 4-methoxybenzaldehyde (4-MBA) in acid whey fermented with Ischnoderma benzoinum. These structurally related aromatic aldehydes are valuable aroma compounds formed during basidiomycete-mediated fermentation of dairy by-products; however, no method enabling their simultaneous quantification in this complex matrix without prior derivatization has been reported to date. Chromatographic separation was achieved on an XBridge BEH C18 column (250 × 4.6 mm, 5 μm) using gradient elution with water and methanol as mobile phases, with a total runtime of 30 min. The method was validated following the relevant ICH Q2(R2) validation characteristics and showed linearity (R2 > 0.999) over ranges of 0.1-100 mg L-1 (BA, 4-MBA) and 0.1-75 mg L-1 (3,4-DMBA). Limits of detection and quantification ranged from 0.32 to 0.66 mg L-1 and 0.96-1.98 mg L-1, respectively. Accuracy (94.9-111.5%), intra- and inter-day precision (RSD ≤ 3.16%), and matrix recovery (90.7-105.5%) confirmed the reliability of the method for quantitative analysis in real fermentation samples. The developed method provides a practical, direct analytical tool for monitoring aromatic aldehyde production at mg L-1 level in complex aqueous fermentation matrices, rather than for trace-level determination.

Keywords:  Acid whey valorization; Aromatic aldehydes; Ischnoderma benzoinum fermentation; Matrix effect; Method validation

DOI:  https://doi.org/10.1016/j.jchromb.2026.125266
Metabolites. 2026 Jul 27. pii: 530. [Epub ahead of print]16(8):

Exercise and Sports Metabolomics: Analytical Platforms, Metabolite Annotation, Pathway-Level Interpretation, and Biomarker-Panel Readiness.

Donghai Lin, Yifen Chen, Caihua Huang.

  Exercise and sports metabolomics provide a systems-level approach to characterizing how acute exercise, training adaptation, nutrition, recovery, and environmental stress reshape human metabolism. By profiling metabolites related to substrate utilization, mitochondrial function, redox balance, inflammation, muscle stress, and recovery kinetics, these approaches can reveal pathway-level responses that conventional single biomarkers cannot capture. However, many exercise-responsive features remain difficult to interpret because of incomplete chemical identification, uncertain annotation confidence, limited quantitative reproducibility, variable pre-analytical control, inconsistent data processing, and insufficient biological validation. This narrative review examines recent advances in exercise and sports metabolomics, with emphasis on LC-MS, GC-MS, NMR spectroscopy, IMS-MS, and CE-MS workflows; platform selection; metabolite annotation and identification; pathway-level interpretation; and evidence requirements for candidate-panel development. Exercise-responsive metabolites should be interpreted as context-dependent pathway signals rather than isolated indicators of fatigue, recovery, adaptation, or performance. The review consolidates requirements for sampling, quality control, metadata capture, repeated-measures analysis, and external validation within an evidence-readiness roadmap. Wearable biochemical monitoring, AI-assisted analysis, and multi-omics integration may support future applications, but their value depends on analytical robustness, external validation, and physiological interpretability. Exercise and sports metabolomics should therefore progress from descriptive feature discovery toward reproducible, quantitatively reliable, and biologically validated pathway-level interpretation.

Keywords:  biomarker-panel readiness; exercise metabolomics; metabolite annotation; metabolite identification; pathway validation; quality control; sports metabolomics

DOI:  https://doi.org/10.3390/metabo16080530
Anal Chim Acta. 2026 10 22. pii: S0003-2670(26)00892-5. [Epub ahead of print]1420 345942

Dispersive liquid-liquid aerosol phase extraction for the determination of acetylated bromophenols in urine by gas chromatography - mass spectrometry.

Raquel Sánchez-Romero, Javier Peña, José Luis Todolí, Iria González-Mariño.

   BACKGROUND: Bromophenols are urinary biomarkers of human exposure to brominated flame retardants, but their determination remains challenging because they occur at ultra-trace levels in this complex biological matrix. In this work, dispersive liquid-liquid aerosol phase extraction (DLLAPE) was combined with in-situ aqueous acetylation and gas chromatography-mass spectrometry analysis.
RESULTS: Under optimized conditions, the acetylating reagent (acetic anhydride) and the extracting solvent (heptane) are nebulized directly into the buffered urine sample, generating an interfacial surface area that accelerates mass transfer. The extraction is completed in 80 s and yields a ready-to-inject organic extract through spontaneous phase separation, avoiding the time-consuming centrifugation steps typically required in conventional liquid-liquid extraction. The method demonstrated satisfactory trueness (recoveries of 74-130 %), good overall precision with RSD values generally ≤20 %, and limits of quantification ranging from 70 to 370 pg mL-1 in human urine. Finally, it was successfully applied to urine samples from five healthy adult volunteers with no known exposure to brominated flame retardants, allowing the quantification of several mono-, di-, and tri-bromophenols at sub-ng mL-1 levels.
SIGNIFICANCE: The study represents the first application of DLLAPE to the extraction of organic compounds from aqueous matrices, specifically from urine. It provides a rapid, solvent-minimized, and high-throughput alternative to conventional liquid-liquid extraction for the determination of bromophenols in urine.

Keywords:  Brominated flame retardants; Bromophenols; Dispersive liquid-liquid aerosol phase extraction; Gas chromatography-mass spectrometry; Urine

DOI:  https://doi.org/10.1016/j.aca.2026.345942
Methods Mol Biol. 2026 ;3017 185-194

Estimation of DNA Methylation in Wheat Using Reverse Phase Ultra-High-Performance Liquid Chromatography Coupled with Mass Spectrometry During Leaf Rust Pathogenesis.

Uzma Afreen, Aryan Verma, Kunal Mukhopadhyay, Manish Kumar.

  Reverse phase ultra-high-performance liquid chromatography is considered the most reliable and sensitive technique to determine and separate analytes with small differences in their molecular weights. Leveraging this technique coupled with mass spectrometry, we developed a method for simultaneous determination of 2'-deoxycytidine and its methylated analogue, 5-methyl-2'-deoxycytidine, in wheat. The nonpolar C18 column was selected that greatly shortened the detection time to approximately 2 min. Formic acid was added to the mobile phase to achieve the complete separation. Thus, avoiding the signal suppression and interference of 2'-deoxycytidine in the detection of 5-methyl-2'-deoxycytidine. The method utilized a least toxic solvent system consisting of water and methanol. Although, DNA methylation can be detected in numerous ways, bisulfite sequencing achieve single-base resolution and methylation-sensitive amplified polymorphism is effective in pattern analysis. Whole-genome and reduced representation bisulfite sequencing provide more detailed or more targeted methylation profiles, respectively. However, these methods are data and cost-intensive and provide incomplete information due to DNA degradation. Methylated DNA immunoprecipitation sequencing is biased toward highly methylated regions, and ELISA is more high-throughput but less precise. Therefore, the optimized method, is specific, reliable, and highly sensitive for the detection and quantification of cytidine methylation levels in wheat during leaf rust pathogenesis.

Keywords:  2′-deoxycytidine; 5-methyl 2′-deoxycytidine; Cytosine Methylation; Leaf rust; UHPLC-MS/MS; Wheat

DOI:  https://doi.org/10.1007/978-1-0716-5162-9_15
Food Res Int. 2026 Oct 31. pii: S0963-9969(26)01832-6. [Epub ahead of print]242(Pt 2): 120146

Rapid screening method of glucocorticoid residues in sample-limited cell-cultured meat.

Peiwen Yu, Simeng Li, Chen Zhang, Xiaohui Li, Hongping Wang, Fen Jin.

  The emergence of cell-cultured meat as a sustainable protein source necessitates robust safety assessment frameworks, particularly for novel inputs like glucocorticoids-potent media supplements used to enhance cell growth. This study addresses the critical analytical gap for monitoring these compounds in complex cultured meat matrices. A sensitive and selective isotope-dilution ultra-performance liquid chromatography-tandem mass spectrometry method was established for 40 glucocorticoids, overcoming key challenges including significant matrix effects and the separation of six critical isomer pairs. Through optimized miniaturized sample preparation (0.2 g) combined with chromatographic separation on a BEH Phenyl column, the method achieved quantification limits (MQLs) of 0.5-3 μg/kg in cell-cultured biomass and supernatant, with recoveries of 70-118%. The method was applied to analyze residual glucocorticoids during the cell-cultured pork production. Hydrocortisone was the only glucocorticoid detected, with a concentration of 3.15 ± 0.07 μg/kg in final biomass, well below the maximum residue limit for milk (10 μg/kg). Furthermore, a safety-assessed maximum intake (SAMI) limit was derived to guide future media formulation. This work provides the first systematic study of glucocorticoid residues in cell-cultured meat production, establishing a vital analytical method to support the safe development of the cell-cultured meat industry.

Keywords:  Cell-cultured meat; Glucocorticoids; Residue; Safety assessment; UPLC-MS/MS

DOI:  https://doi.org/10.1016/j.foodres.2026.120146
Methods Protoc. 2026 Jul 23. pii: 110. [Epub ahead of print]9(4):

Gas Chromatography Method for Quantitation of Residual Solvent Impurities in Nanoformulations.

Krishna Kattel, Rachael M Crist, Jeffrey D Clogston.

  The development and validation of a sensitive, rapid, and specific gas chromatography method for the evaluation of 19 common Class 2 and Class 3 solvents frequently used in nanomedicine formulation is described. Method validation was performed using PerkinElmer's headspace gas chromatograph system with flame ionization detection and an Elite 624 Crossbond 6% cyanopropylphenyl-94% dimethylpolysiloxane or DB-Fatwax-Ultra Inert column with helium as the carrier gas. Validation characteristics such as linearity, spike recovery, method precision, specificity, sensitivity, limit of detection/quantitation, and analyte stability were evaluated. The validated methods showed excellent linearity, with a correlation coefficient > 0.99, and good precision, with intra-day precision < 7.4% for all tested analytes. The percent recoveries ranged 83-104% within the method's quantitation range. In comparison to previously reported methods, the current method has a much shorter equilibration time, higher sensitivity, better separation for many solvents, and a wide concentration detection range. The current method is also perfectly suitable to analyze short chain fatty acids such as formic acid, acetic acid, butyric acid, and valeric acid without requiring additional extraction or derivatization steps. Notably, the method was found to be suitable for analysis of formic acid-a common solvent in certain nanoformulations and one in which there is no prior gas chromatography method available which does not require this additional sample manipulation-down to approximately 75 ppm. Herein, the method is demonstrated using various nanoformulations, including the commercial Doxil formulation as well as several research nanoformulations, including polymeric, cross-linked polymeric, and dendrimer platforms.

Keywords:  flame ionization detector; formic acid; headspace gas chromatography; liposome; nanomedicine; polymeric nanoparticles; residual solvents; short chain fatty acids; volatile organic impurities

DOI:  https://doi.org/10.3390/mps9040110