bims-metlip 2026-10-04 papers

bims-metlip

Biomed News

on Methods and protocols in metabolomics and lipidomics

Issue of 2026–10–04
nineteen papers selected by
Sofia Costa, Matterworks

A Novel Polarity Switching LC-MS/MS Method for Simultaneous Quantification of Drospirenone and Estetrol: Application to Pharmacokinetic Studies in Human Plasma.
Advanced sample preparation methods for LC-MS/MS bioanalysis of naturally derived narcotics in clinical and forensic contexts.
Sub-3 μm porous graphitic carbon for highly polar metabolomics: orthogonal selectivity and comparison with HILIC and reversed-phase chromatography.
Simultaneous quantification of venetoclax, selinexor, voriconazole and fluconazole in human plasma by HPLC-MS/MS and its application in hematological malignancies.
A DOE-optimized supercritical fluid chromatography-mass spectrometry workflow complementing liquid chromatography for comprehensive profiling of Trichoderma reesei specialized metabolism.
Comparison of Extraction Methods for the Quantification of Phytohormones From Tomato Fruits and Leaves by LC-MS/MS.
SpotOn: A Tool to Enhance Echo Acoustic Standard Deposition for Quantitative Mass Spectrometry Imaging.
A rapid sequential salting-out/isoelectric precipitation LC-MS/MS method for quantifying cytokeratin 18 in human ascitic fluid.
TOF-SIMS Imaging of Intact Lipids in Biological Tissue with Enhanced Ion Intensity by Ionic Liquid.
One-step SPE-UHPLC-MS/MS for simultaneous detection of fenbendazole and metabolites in animal-derived foods.
TC-TopoRT: Tautomer-consistent and topology-aware retention time prediction for LC-MS metabolite annotation.
Application of an intracellular LC-MS/MS assay for the determination of carbovir-triphosphate and remdesivir-triphosphate in peripheral blood mononuclear cells and dried blood spots.
Fabrication of thermoplastic microfluidic probes for nanospray desorption electrospray ionization mass spectrometry.
DES-impregnated electrospun nanofiber microextraction for simultaneous determination of carvedilol and verapamil in human plasma by HPLC-fluorescence detection.
X-Align: annotation-independent cross-platform alignment of untargeted metabolomics features.
Ballpoint tip as an integrated tool for extraction and ionization: a dual-workflow for rapid screening and quantification of multiple mycotoxins in edible oils.
Determination of perfluorobutane sulfonyl fluoride in environmental water and soil using a novel quaternary ammonium derivatization LC-MS/MS method.
Times are changing but order matters: transferable prediction of small-molecule liquid chromatography retention times.
Rapid quantification of environmental pollutant metals in dried blood spots via direct microextraction coupled to ICP-MS (μEx-ICP-MS).

Biomed Chromatogr. 2026 Nov;40(11): e70633

A Novel Polarity Switching LC-MS/MS Method for Simultaneous Quantification of Drospirenone and Estetrol: Application to Pharmacokinetic Studies in Human Plasma.

Pravesh Singh, Sajad Khaliq Dar, Faisal Hamid Khan, Asjad Nabi Khan Sherwani, Sanjay Jagannath Gurule, Onkar Singh, Sudershan Kumar, Arshad H Khuroo.

  A sensitive, selective, and rapid liquid chromatography-tandem mass spectrometry (LC-MS/MS) method using a polarity switch technique was developed and validated for the simultaneous determination of drospirenone (DSP) and estetrol (E4) in human plasma for the first time. Solid phase extraction (SPE) with Bond Elut PLEXA cartridges was performed. Chromatographic separation was obtained on Eclipse XDB-C8 (150 mm × 4.6 mm, 3.5 μm) column using 2-mM ammonium bicarbonate solution:methanol (30:70, v/v) as the mobile phase at a flow rate of 0.750 mL/min. A polarity switching method was performed in a triple quadrupole mass spectrometer using multiple reaction monitoring (MRM). MRM was performed in positive mode for DSP transitions m/z 367.2 → 171.1 and DSP-D4 transitions m/z 371.2 → 171.1, and in negative mode for E4 transitions m/z 303.1 → 241.2, 273.0 and E4-d3 transitions m/z 306.2 → 243.2, 275.0. Calibration curves were linear over the concentration ranges of 0.351-100.446 and 0.351-50.134 ng/mL for DSP and E4, respectively. The accuracy measured for both analytes ranged from -2.74% to 4.21%. Both the analytes showed consistent recoveries ranged from 77.25% to 81.34% with no significant matrix effect. The method was successfully applied in a pharmacokinetic study of DSP and E4 tablets (3 mg + 14.2 mg).

Keywords:  SPE; drospirenone; estetrol; matrix effect; polarity switching; quality control; selectivity

DOI:  https://doi.org/10.1002/bmc.70633
J Chromatogr A. 2026 Sep 24. pii: S0021-9673(26)00816-2. [Epub ahead of print]1788 467490

Advanced sample preparation methods for LC-MS/MS bioanalysis of naturally derived narcotics in clinical and forensic contexts.

Melda Aldemir, Yasin Darı, Demet Dincel.

  Naturally occurring narcotics such as morphine and codeine require careful monitoring due to the risks of tolerance, addiction, and overdose. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is considered a gold standard method for the quantitative and qualitative analysis of these narcotics in biological samples. However, because of the complex structure of biological matrices, sample preparation directly affects analytical performance. Inadequate preparation can lead to matrix effects, poor recovery, and analytical uncertainties, making optimized strategies necessary for reliable results. This review presents a systematic classification of sample preparation techniques used in the LC-MS/MS-based bioanalysis of morphine and codeine. Protein precipitation, liquid-liquid extraction, solid-phase extraction, and next-generation microextraction approaches are comprehensively evaluated regarding matrix-effect reduction, ion suppression control, and analyte recovery. Current methodological limitations and literature gaps are discussed, alongside proposed innovative strategies for future research. Rather than viewing sample preparation as a simple preliminary step, this study emphasizes its role as a key component of the analytical workflow. Integrating optimized extraction with LC-MS/MS is essential for achieving reliable, sensitive, and reproducible narcotic analysis.

Keywords:  Analytical challenges of narcotics’ bioanalysis; LC–MS/MS; Naturally derived narcotics; Sample preparation techniques; Therapeutic drug monitoring

DOI:  https://doi.org/10.1016/j.chroma.2026.467490
Anal Bioanal Chem. 2026 Oct 02.

Sub-3 μm porous graphitic carbon for highly polar metabolomics: orthogonal selectivity and comparison with HILIC and reversed-phase chromatography.

Danila La Gioia, Emanuela Salviati, Fabrizio Merciai, Vicky Caponigro, Vincenzo Vestuto, Carlo Crescenzi, Pietro Campiglia, Eduardo Maria Sommella.

  The analysis of the polar metabolome remains a major challenge in LC-MS. Beyond established hydrophilic interaction liquid chromatography (HILIC) and reversed-phase (RP) workflows, alternative and orthogonal chromatographic approaches are required to improve the detection and quantification of compounds with low logP values and structurally related isomers. In this study, we evaluated the performance of sub-3 µm porous graphitic carbon (PGC) stationary phase and benchmarked it against state-of-the-art HILIC and RP approaches. Flow rate, column temperature, organic modifier, mobile-phase composition, and pH were optimized using a representative panel of highly polar endogenous metabolites. The three chromatographic modes exhibited markedly complementary selectivity profiles. HILIC provided the broadest coverage of nucleotides and related metabolites, whereas PGC showed enhanced performance for sugar phosphates, polyamines, and, particularly, tricarboxylic acid (TCA) cycle intermediates. Focused optimization for TCA cycle metabolites improved peak shape and signal-to-noise ratio and enabled the separation of citrate and isocitrate, outperforming both HILIC and reversed-phase chromatography for the detection of this metabolite class under the investigated conditions. In mouse liver extracts, PGC detected a complementary subset of metabolites not observed using HILIC or reversed-phase chromatography, confirming its distinctive selectivity in a complex biological matrix. As a proof of concept, the optimized PGC method was applied to lipopolysaccharide-stimulated macrophages using a hybrid workflow combining multiplexed parallel reaction monitoring for the absolute quantification of TCA cycle intermediates with data-dependent acquisition for global metabolic profiling. The workflow captured the characteristic metabolic reprogramming associated with macrophage inflammatory activation. Overall, sub-3 µm PGC represents a valuable orthogonal chromatographic platform for extending analytical coverage and improving the characterization of highly polar metabolites.

Keywords:  HILIC; Metabolomics; Polar metabolites; Porous graphitic carbon; Tricarboxylic acids

DOI:  https://doi.org/10.1007/s00216-026-06846-y
Front Pharmacol. 2026 ;17 1827916

Simultaneous quantification of venetoclax, selinexor, voriconazole and fluconazole in human plasma by HPLC-MS/MS and its application in hematological malignancies.

Feifei Han, Yinying Wang, Yuan Jian, Ting Hu, Ying Zhu, Xian Ding, Jinghui Zhang, Rao Wei, Honghu Zhu, Zhuoling An.

   Background: Venetoclax and selinexor are essential targeted agents for hematological malignancies, but their clinical application is challenged by pronounced ethnic pharmacokinetic differences-especially in Chinese patients-and CYP3A-mediated drug-drug interactions (DDIs) with azole antifungals (voriconazole, fluconazole). No existing LC-MS/MS method simultaneously quantifies all four drugs, creating an unmet need for therapeutic drug monitoring (TDM). We aimed to develop and validate a rapid, sensitive HPLC-MS/MS method for simultaneous quantification of venetoclax, selinexor, voriconazole, and fluconazole in human plasma.
Methods: Plasma samples underwent protein precipitation pretreatment. Chromatographic separation utilized a C18 column with acetonitrile and 0.1% formic acid gradient elution. Mass detection employed positive-ion multiple reaction monitoring (MRM). The method was validated for specificity, linearity, precision, accuracy, recovery, matrix effect, and stability, then applied to AML patients receiving these medications.
Results: Calibration ranges were 40-8000 ng/mL (venetoclax), 5-1000 ng/mL (selinexor), 100-10000 ng/mL (voriconazole), and 320-32000 ng/mL (fluconazole), all with r 2 > 0.9976. Intra-day and inter-day precision and accuracy met acceptance criteria. No significant matrix effect or carry-over was observed. Selinexor exhibited limited stability (<85% remaining after 4 h at room temperature or 24 h at 4 °C); other analytes were stable throughout. The median venetoclax steady-state trough concentration (Css,trough) in Chinese patients was 1025.08 ng/mL. Co-administration with voriconazole increased venetoclax exposure 1.75-14-fold, whereas fluconazole produced a consistent 2.4-2.9-fold increase. Venetoclax 100 mg/d combined with azole antifungals achieved exposures comparable to or exceeding 400 mg/d monotherapy.
Conclusion: This is the first HPLC-MS/MS method enabling simultaneous quantification of venetoclax, selinexor, voriconazole, and fluconazole, successfully deployed in routine TDM. Voriconazole's marked DDI variability underscores the necessity of TDM-guided dosing, while fluconazole offers a more predictable antifungal prophylaxis alternative. These results deliver population-specific pharmacokinetic data supporting rational co-administration of venetoclax with azole antifungals in Chinese hematologic malignancy patients.

Keywords:  HPLC-MS/MS; fluconazole; hematologic malignancies; selinexor; venetoclax; voriconazole

DOI:  https://doi.org/10.3389/fphar.2026.1827916
J Chromatogr A. 2026 Sep 17. pii: S0021-9673(26)00775-2. [Epub ahead of print]1788 467449

A DOE-optimized supercritical fluid chromatography-mass spectrometry workflow complementing liquid chromatography for comprehensive profiling of Trichoderma reesei specialized metabolism.

Ilias El Ouar, Agnès Le Masle, Frédérique Bidard, Alexandra Berlioz-Barbier, David Touboul.

  Fungal secretomes are prolific sources of structurally diverse bioactive natural products, including peptaibols, lipopeptides, and polar metabolites. Their chemical complexity makes comprehensive untargeted profiling a persistent analytical challenge, particularly for highly polar compounds that are poorly retained in conventional reversed-phase liquid chromatography (RPLC). This study presents the development and optimization of a supercritical fluid chromatography (SFC) method coupled to high-resolution tandem mass spectrometry (HRMS/MS) for the untargeted metabolite profiling of a Trichoderma reesei secretome extract. The SFC method was systematically compared to an established RPLCHRMS/MS workflow to evaluate their respective analytical performances and chemical space coverage. RPLC demonstrated superior overall metabolite detectability, with 3726 features detected compared to 2301 in SFC, and excelled at resolving structurally related isomers, particularly within the peptaibol class where up to six isomers were baseline-separated. SFC, by contrast, provided complementary class-specific retention behavior and uniquely revealed highly polar amino sugar-like compounds alongside a series of 14-residue harzianine-like peptaibols that remained undetected under RPLC conditions. Molecular networking analysis confirmed that 43% of SFC-detected features were absent from RPLC datasets, demonstrating substantial orthogonality between the two separation modes. Neither approach alone was sufficient to capture the full chemical diversity of the secretome. These findings establish that integrating SFC and RPLC workflows substantially expands the accessible chemical space in fungal metabolomics and provides a robust framework for the discovery of bioactive natural products.

Keywords:  DOE; High-resolution mass spectrometry; Molecular network; Natural products; Reversed-phase liquid chromatography; Supercritical fluid chromatography

DOI:  https://doi.org/10.1016/j.chroma.2026.467449
J Mass Spectrom. 2026 Oct;61(10): e70117

Comparison of Extraction Methods for the Quantification of Phytohormones From Tomato Fruits and Leaves by LC-MS/MS.

Carlos A Juarez Guzman, Linxing Yao, Corey D Broeckling, Cristiana T Argueso.

  Purification of crude plant extracts by solid-phase extraction (SPE) is assumed to reduce matrix effect in phytohormone quantification by LC-MS/MS, but its efficacy is rarely assessed. We used postextraction spiking of isotope-labeled phytohormones to estimate matrix effect, recovery, and processes efficiency in extracts prepared according to protocols that implement SPE with tC18 and HLB sorbents, respectively, and according to a protocol that does not include SPE. Across six structurally diverse phytohormone classes and tomato fruit and leaf matrices, we did not find significant matrix effects associated with extracts prepared with no SPE. For recovery efficiency, process efficiency, and variation, we rarely found conclusive evidence that the methods that implemented SPE were optimal relative to the one that does not. A notable exception was fruit extracts prepared using HLB sorbents, which had minimal variation for indole-3-acetic acid, trans-zeatin, and trans-zeatin riboside, all of which were detected with positive electrospray ionization.

Keywords:  LC–MS/MS; matrix effect; phytohormone quantification; phytohormones; plant hormones; solid phase extraction

DOI:  https://doi.org/10.1002/jms.70117
Chem Biomed Imaging. 2026 Sep 28. 4(9): 2119-2125

SpotOn: A Tool to Enhance Echo Acoustic Standard Deposition for Quantitative Mass Spectrometry Imaging.

Emily R Sekera, Nathaniel Twarog, Rebekah DeVries, P Jake Slavish, John J Bowling, Anang A Shelat.

  The quantitative evaluation of therapeutics in the drug discovery pipeline is essential for elucidating their pharmacokinetic and pharmacodynamic profiles within complex biological systems. Traditionally, autoradiography and liquid chromatography-mass spectrometry (LC-MS) have been employed for such analyses. However, matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI) is emerging as a promising alternative, offering a more rapid untargeted workflow. Quantitative MALDI-MSI (qMSI) relies on accurate calibration to achieve robust and reproducible quantification of analytes. Here, we present an open-source software tool, SpotOn, designed to facilitate the discrete deposition of low-volume calibration standards onto glass slides or tissue sections. By enabling precise placement and significantly reducing the volume of standards required, our tool affords the generation of more calibration points across a wider dynamic range, thereby improving the robustness and accuracy of quantitative analyses. Software was validated using JQ1 on both glass slides and murine liver tissue. The resulting calibration curves demonstrated strong linearity, with coefficients of determination (R 2) exceeding 0.92. This tool streamlines the qMSI workflow, conserves valuable reagents, and enhances analytical reliability, making it a helpful resource.

Keywords:  MALDI; acoustic deposition; image analysis; mass spectrometry imaging; quantitation; software

DOI:  https://doi.org/10.1021/cbmi.6c00010
Clin Chim Acta. 2026 Oct 01. pii: S0009-8981(26)02051-6. [Epub ahead of print] 122869

A rapid sequential salting-out/isoelectric precipitation LC-MS/MS method for quantifying cytokeratin 18 in human ascitic fluid.

Zhenni Liu, Xiaohua Xiao, Bingchu Li, Nijiao Qin, Ziyang Li, Xing Lyu, Zhongyuan Xiang, Min Hu, Qichen Long.

  Cytokeratin 18 (CK18), a predominant hepatocyte intermediate filament protein released during hepatocyte death, has been investigated as a hepatic injury biomarker. Conventional immunoassays for CK18 suffer from non-specific cross-reactivity and insufficient standardization. Though liquid chromatography-tandem mass spectrometry (LC-MS/MS) could overcome such immunoanalytical drawbacks, accurate CK18 quantification in ascitic fluid remains analytically challenging because of the complex matrix, low target abundance, and variability introduced by protein enrichment and digestion. Herein, we developed a rapid and practical LC-MS/MS method for CK18 quantification in ascitic fluid. A two-step non-immunoaffinity enrichment strategy, integrating sequential ammonium sulfate salting-out and isoelectric point-driven acidic precipitation, was optimized to balance enrichment yield, pretreatment turnaround, and operational cost. Subsequent pretreatment workflow comprising urea denaturation, enzymatic hydrolysis, and solid phase extraction purification can be completed within 6 h, providing an economical, instrument-compatible procedure for LC-MS/MS analysis. Systematic analytical performance evaluation showed satisfactory linearity spanning 0.10-10.36 nmol/L, a limit of quantification of 0.10 nmol/L, a limit of detection of 0.02 nmol/L, total imprecision <10%, and recoveries ranging from 93.07% to 108.92%. The internal standard effectively compensated for LC-MS/MS instrumental variability and ionization-related matrix effects. Method comparison showed acceptable agreement with a published multiplexed cytokeratin MS method. Clinical analysis demonstrated significantly elevated ascitic-fluid CK18 concentration in cirrhotic patients versus non-cirrhotic controls (P = 0.001). This novel enrichment strategy preserves the simplicity and cost-efficiency of conventional precipitation methods while avoiding costly antibody capture reagents. Collectively, the validated LC-MS/MS method is appropriate for preliminary exploration of the clinical value of ascitic-fluid CK18 and can facilitate translational research in cirrhotic populations.

Keywords:  Cirrhosis; Cytokeratin 18; Liquid chromatography-mass spectrometry; Protein precipitation; Quantification; Salting-out

DOI:  https://doi.org/10.1016/j.cca.2026.122869
ACS Omega. 2026 Sep 01. 11(34): 51034-51041

TOF-SIMS Imaging of Intact Lipids in Biological Tissue with Enhanced Ion Intensity by Ionic Liquid.

Itsuko Ishizaki, Noriaki Sanada.

Mass spectrometry imaging (MSI) is a powerful technique for visualizing the spatial distribution of chemical species in biological samples. Time-of-flight secondary ion mass spectrometry (TOF-SIMS) enables high-spatial-resolution imaging and the detection of a broad range of molecular ions, including intact lipids. However, the observation of high-mass components remains challenging due to limitations in ionization efficiency and sample degradation. In this study, we present a novel sample preparation method for TOF-SIMS using an ionic liquid (IL) pretreatment to enhance the ionization efficiency of high-mass lipid ions. This approach increases the ionization efficiency of lipid molecules by achieving an appropriate distribution of ionic liquid to the surface of the sample. By reducing fragmentation, this method exceeds the static limit and increases the lipid ion signal by nearly 3 orders of magnitude. Furthermore, this method enabled static TOF-SIMS imaging of lipid-related ions and ion-induced electron imaging beyond the conventional static limit. These advancements significantly improve TOF-SIMS-based MSI applications, offering new opportunities for lipidomics and biological research.

DOI: https://doi.org/10.1021/acsomega.6c03014
Food Addit Contam Part A Chem Anal Control Expo Risk Assess. 2026 Oct 01. 1-11

One-step SPE-UHPLC-MS/MS for simultaneous detection of fenbendazole and metabolites in animal-derived foods.

Can Guo, Haifeng Yang, Shouying Wang, Yiqing Song, Bing Bai, Kaixuan Huo, Gang Chen, Wenshuai Si.

  Fenbendazole (FBZ) is a highly effective antiparasitic agent widely used in veterinary medicine; however, its improper application can lead to residues in edible animal-derived products, posing potential health risks to humans through accumulation via the food chain. This study aimed to establish an ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method for the determination of FBZ and metabolites in animal-derived foods. The proposed method involves a one-step solid-phase extraction followed by UHPLC-MS/MS analysis for the simultaneous detection of FBZ, oxfendazole, and oxfendazole sulphone. The method demonstrated excellent linearity, with correlation coefficients (R2) exceeding 0.99. The limit of detection and limit of quantification were 2.0 μg/kg and 5.0 μg/kg, respectively. Average recoveries ranged from 85.0% to 108.4%, with relative standard deviations below 10%. The established method is simple, highly specific, and sensitive, offering a reliable approach for production monitoring, routine batch analysis, and confirmatory testing of FBZ and metabolites in animal-derived foods.

Keywords:  Fenbendazole; UHPLC–MS/MS; animal-derived foods; metabolites; one-step SPE

DOI:  https://doi.org/10.1080/19440049.2026.2729044
Talanta. 2026 Sep 24. pii: S0039-9140(26)01295-6. [Epub ahead of print]313(Pt B): 130639

TC-TopoRT: Tautomer-consistent and topology-aware retention time prediction for LC-MS metabolite annotation.

Runquan Lai, Zhangqiang Liu, Bingyi Wang, Weihua Li.

  Accurate retention time (RT) prediction provides chromatographic evidence complementary to precursor mass and MS/MS spectra in LC-MS metabolomics. Such evidence is particularly valuable for metabolite annotation, where multiple structurally plausible candidates may exhibit similar spectral signatures. Here, we present TC-TopoRT, a tautomer-consistent and topology-aware framework for RT prediction in LC-MS metabolite annotation. TC-TopoRT constructs paired molecular views from the dataset-provided SMILES and a conservative tautomer-standardized representation generated under identity-preserving checks. Each view is lifted into a ring-aware cell complex and encoded by a cellular Weisfeiler-Lehman network (CWN) to learn topology-aware molecular representations. The resulting paired-view predictions are then integrated through a leakage-free out-of-fold fusion protocol without using independent-test labels for model selection, calibration, or stacker fitting. On the retained-compound SMRT benchmark, TC-TopoRT achieved a mean absolute error of 25.06 s. A five-seed ensemble further reduced the error to 24.92 s. Across six PredRet chromatographic datasets, TC-TopoRT reported lower MAEs than the cited reference values on four systems and remained within 1.92 s on the other two. With RT thresholds calibrated on fixed development queries held out from calibration-model fitting and disjoint from final evaluation, TC-TopoRT reduced candidate spaces by 41.05% on MetaboBase and 30.35% on RIKEN-PlaSMA while retaining 42/45 (93.33%) and 84/85 (98.82%) true candidates, respectively. It also maintained or improved Top-k prioritization relative to the original MS-FINDER ranking. These results support the use of tautomer-consistent and topology-aware RT prediction as complementary chromatographic evidence for LC-MS metabolite annotation.

Keywords:  Graph neural networks; LC–MS; Metabolite annotation; Retention time prediction; Tautomer consistency; Topological learning

DOI:  https://doi.org/10.1016/j.talanta.2026.130639
J Pharm Biomed Anal. 2026 Sep 17. pii: S0731-7085(26)00396-1. [Epub ahead of print]283 117728

Application of an intracellular LC-MS/MS assay for the determination of carbovir-triphosphate and remdesivir-triphosphate in peripheral blood mononuclear cells and dried blood spots.

Stefanie K Schwab, Laura E Roon, Lane R Bushman, Ryan P Coyle, Bethany M Johnson, Lucas A Ellison, Peter L Anderson, Kristina M Brooks.

  Intracellular concentrations of antiviral nucleos(t)ide analogs are associated with a variety of clinical outcomes and toxicities, underscoring the need for novel quantification methods for commonly used antivirals in cellular matrices. Here, we extend a previously validated intracellular LC-MS/MS method to carbovir-triphosphate (CBV-TP) and remdesivir-TP (RDV-TP), the active anabolites of abacavir (ABC) and remdesivir (RDV), respectively, in peripheral blood mononuclear cells (PBMC) and dried blood spots (DBS). CBV-TP and RDV-TP concentrations in DBS were quantified using two 7 mm punches of spotted, dried whole blood extracted in 50:50 methanol:water solution (lysed intracellular matrix). Lysed PBMC samples were prepared in a 70:30 methanol:water solution. Phosphorylated intracellular analyte fractions were isolated from the lysed matrix using strong anion‑exchange solid‑phase extraction (SPE), enzymatically dephosphorylated to the parent drug, then desalted/concentrated with reversed phase SPE and quantified utilizing stable isotope labeled internal standards by LC‑MS/MS methods adapted from prior validated assays. Accuracy was within ±15% difference from nominal and precision was ≤ 15% coefficient of variation for all validation assessments for both CBV-TP and RDV-TP. The validated quantifiable range for CBV-TP was 5-2000 fmol/sample and 0.1-100 pmol/sample for RDV-TP. The methods were successfully applied to clinical research PBMC and DBS samples with reproducible results. Both analytes demonstrated accuracy and precision across tested concentration ranges and matrices, meeting regulatory criteria outlined in current bioanalytical guidance. These methods support novel intracellular data for CBV-TP and RDV-TP in PBMC and DBS, which will facilitate wider use in antiviral pharmacology research.

Keywords:  Analytical methods; Antiviral treatment; Dried blood spots; Intracellular pharmacology; Nucleoside analog; Peripheral blood mononuclear cells

DOI:  https://doi.org/10.1016/j.jpba.2026.117728
Int J Mass Spectrom. 2026 Oct;pii: 117672. [Epub ahead of print]528

Fabrication of thermoplastic microfluidic probes for nanospray desorption electrospray ionization mass spectrometry.

Renkai Dong, Anthony H Choi, Xiangtang Li, Xindi Tang, Li-Xue Jiang, Julia Laskin.

  Nanospray desorption electrospray ionization mass spectrometry (nano-DESI MS) uses specially designed probes to achieve a stable liquid bridge on the sample surface, ensuring consistent analyte extraction. Here, we present a mold-based fabrication protocol for cyclic olefin copolymer (COC) microfluidic probes (MFPs). We describe mold design, probe fabrication, and probe setup for nano-DESI MS. Four mold fabrication methods, including computer numerical control (CNC) machining, metal 3D printing, photolithography, and selective laser-assisted etching (SLE), were evaluated, with SLE providing the highest structural precision and enabling reproducible channel fabrication. Performance evaluation using lysophosphatidylethanolamine (LPE) 17:1 standard demonstrated stable liquid bridge formation, clear spatial separation, reproducible signal response, and low background interference. This mold-based fabrication strategy produces reproducible, low-cost probes and offers an accessible approach for nano-DESI MS.

Keywords:  Ambient ionization; Cyclic olefin copolymer (COC); Liquid bridge; Mass spectrometry imaging; Microfluidic probe; Mold imprinting; Nano-DESI; Thermal bonding

DOI:  https://doi.org/10.1016/j.ijms.2026.117672
J Chromatogr A. 2026 Sep 28. pii: S0021-9673(26)00832-0. [Epub ahead of print]1788 467506

DES-impregnated electrospun nanofiber microextraction for simultaneous determination of carvedilol and verapamil in human plasma by HPLC-fluorescence detection.

Fereshteh Partovi Ebrahimpour, Hassan Heidari.

  The development of efficient and environmentally compatible sample preparation strategies for the determination of pharmaceuticals in complex biological matrices remains an important challenge in bioanalytical chemistry. Herein, a deep eutectic solvent (DES)-impregnated electrospun nanofiber microextraction (DES-ENF-ME) approach was developed for the simultaneous extraction of carvedilol and verapamil from human plasma prior to HPLC-fluorescence detection. A hydrophobic fatty-acid-based DES composed of nonanoic acid and dodecanoic acid (3:1, molar ratio) was selected and impregnated onto electrospun polyacrylonitrile nanofibers. In this configuration, the DES acts as the primary extraction phase, while the nanofibrous matrix provides a porous support for retaining and handling the liquid extraction phase. Sample pH, nanofiber amount, salt concentration, and extraction time were simultaneously optimized using central composite design and a desirability function. The method exhibited a linear response over the range of 1-1000 ng/mL for both analytes, with coefficients of determination of 0.9988 and 0.9942 for carvedilol and verapamil, respectively. The corresponding limits of detection were 0.396 and 0.342 ng/mL. The method showed satisfactory precision and relative recoveries for spiked plasma samples. Moreover, the DES-impregnated nanofibrous material was reusable for at least four consecutive extraction cycles. Overall, the proposed configuration combines the extraction capability of a hydrophobic DES with the practical handling advantages of an electrospun nanofibrous support, providing a simple, miniaturized, and solvent-efficient platform for plasma bioanalysis.

Keywords:  Deep eutectic solvent; Electrospun nanofibers; Miniaturized sample preparation; Pharmaceutical analysis; Plasma bioanalysis

DOI:  https://doi.org/10.1016/j.chroma.2026.467506
Chem Sci. 2026 Sep 22.

X-Align: annotation-independent cross-platform alignment of untargeted metabolomics features.

Han Bao, Zengqi Yan, Bin Wang, Xinxin Wang, Xinjie Zhao, Chunxia Zhao, Wangshu Qin, Wenzhao Wang, Chang Liu, Xin Lu, Guowang Xu.

Massive untargeted metabolomics datasets are rapidly accumulating, but cross-study and cross-platform reuse remain limited by poor comparability. This limitation is exacerbated by the low rate of confident structural annotation, which restricts structure-based alignment to only a small fraction of detected metabolic features. Here, X-Align is presented, which is an annotation-independent framework for cross-platform alignment of untargeted metabolomics features. Its core component, Align-ID, is a residual attention-based deep regression model that learns reference chromatographic behavior directly from MS/MS (tandem mass spectrometry) spectrum embeddings and maps heterogeneous spectra onto a common reference chromatographic scale without prior structural identification. Align-ID is trained on 804 150 high-resolution MS/MS spectra associated with reference retention values in the METLIN SMRT chromatographic system. X-Align achieved 99.6% accuracy at 52.3% coverage in the controlled benchmark and maintained high accuracy in independent cross-platform and external library validations. X-Align produces a cross-center feature repository containing 1402 aligned features and 60 688 MS/MS spectra from public plasma datasets and supports more consistent biological interpretation across independent experiments in a cross-center high-fat diet case study. This work establishes MS/MS-derived reference retention behavior as a practical basis for annotation-independent comparison and integration of cross-platform untargeted metabolomics data.

DOI: https://doi.org/10.1039/d6sc04644d
Food Chem. 2026 Sep 28. pii: S0308-8146(26)03511-9. [Epub ahead of print]531 151351

Ballpoint tip as an integrated tool for extraction and ionization: a dual-workflow for rapid screening and quantification of multiple mycotoxins in edible oils.

Xia Huang, Wenlin Wu, Yu Wang, Lu Chen, Fan Wu, Chaoting Shi, Bing Xia, Yan Zhou.

  This study developed a dual screening-quantification workflow based on ballpoint tip liquid-phase microextraction (BT-LPME) for the high-throughput analysis of multiple mycotoxins in edible oils. By integrating BT-LPME with ballpoint tip electrospray ionization (BP-ESI), the method enables the acquisition of high-resolution mass spectrometry data within 30 s per sample, achieving rapid broad-spectrum screening of 254 mycotoxins. For screen-positive samples, the extract from the tip can be retrieved for accurate quantification via LC-MS/MS. Systematic validation using 55 representative mycotoxins confirmed that the screening sensitivity meets regulatory requirements. The quantitative method demonstrated good linearity (r >0.9970) with limits of detection ranging from 0.01 to 10 μg/kg. In real edible oil samples, 78 mycotoxins were detected, most of which are not covered by current regulatory systems, highlighting potential food safety risks. This approach is green, rapid, and efficient, making it suitable for batch risk monitoring of edible oil safety.

Keywords:  Ambient mass spectrometry; Edible oil; Miniaturized sample preparation; Mycotoxins; Rapid screening

DOI:  https://doi.org/10.1016/j.foodchem.2026.151351
J Hazard Mater. 2026 Sep 25. pii: S0304-3894(26)02727-5. [Epub ahead of print]517 143746

Determination of perfluorobutane sulfonyl fluoride in environmental water and soil using a novel quaternary ammonium derivatization LC-MS/MS method.

Junchen Zhu, Ruochen Zhang, Xinyi Shao, Siwei Rao, Ning Han, Yuanjiang Pan, Cuirong Sun.

  Although current derivatization methods have enabled the detection of perfluorooctane sulfonyl fluoride (PFOSF) and perfluorohexane sulfonyl fluoride (PFHxSF), the analysis of their short-chain substitute, perfluorobutane sulfonyl fluoride (PFBSF), remains challenging despite its increasing production. This study developed a novel quaternary ammonium derivatization reagent and established a highly sensitive liquid chromatography tandem mass spectrometry (LC-MS/MS) method for the trace quantification of PFBSF in water and soil, while enabling the simultaneous determination of PFHxSF and PFOSF. Method validation demonstrated exceptional sensitivity, with method detection limits (MDLs) for PFBSF reaching 0.84 ng L-1 in water and 0.022 ng g-1 in soil. For PFHxSF and PFOSF, the MDLs ranged from 0.93 to 0.98 ng L-1 in water and 0.027-0.030 ng g-1 in soil. The method exhibited excellent linearity (R2 > 0.99 over 5-100 ng L-1), stable recoveries (71%-116%), and satisfactory specificity and stability. This method was successfully applied to environmental samples collected near an abandoned fluorochemical plant in Wuhan. PFBSF, PFHxSF, and PFOSF were widely detected at respective concentrations of 12.12-147.94 ng L-1, 7.75-231.88 ng L-1, and 8.33-26.81 ng L-1 in water, and 0.71-4.19 ng g-1, 0.37-12.81 ng g-1, and 0.29-2.40 ng g-1 in soil. The concurrent detection of high perfluoroalkane sulfonic acid (PFSA) concentrations confirmed the plant as a major point source for both compound classes. Furthermore, temporal monitoring from 2023 to 2025 revealed that regional perfluoroalkyl sulfonyl fluorides (PFASFs) contamination has decreased to a certain extent, whereas PFSA levels remain elevated, warranting continued regulatory attention.

Keywords:  Chemical derivatization; Environmental soil; Environmental water; Liquid chromatography tandem mass spectrometry; Perfluorobutane sulfonyl fluoride; Quaternary ammonium

DOI:  https://doi.org/10.1016/j.jhazmat.2026.143746
Nat Methods. 2026 Oct 01.

Times are changing but order matters: transferable prediction of small-molecule liquid chromatography retention times.

Fleming Kretschmer, Eva-Maria Harrieder, Michael Witting, Sebastian Böcker.

Liquid chromatography is a predominant technology for the separation of small molecules. Hundreds of retention time prediction models have been published over the past decades, yet truly transferable prediction (requiring no training data from the target chromatographic system) remains an open challenge. Unfortunately, retention times may change massively, even for nominally identical chromatographic conditions. Retention order is considerably more conserved; but even retention order may change if chromatographic conditions vary. Here we present 2-step, a two-step method for the prediction of retention times in reversed-phase chromatography. In the first step, a machine-learning model predicts a retention order index, taking into account chromatographic conditions. In the second step, we map predicted indices to absolute retention times. Disentangling these two tasks finally enables transferable retention time prediction across chromatographic conditions and compound classes, without requiring any target-system training data. Our 2-step method outperforms existing methods that were trained on the target dataset. Finally, we systematically study what chromatographic conditions result in notable changes of retention order.

DOI: https://doi.org/10.1038/s41592-026-03243-2
Talanta. 2026 Sep 30. pii: S0039-9140(26)01370-6. [Epub ahead of print]313(Pt A): 130714

Rapid quantification of environmental pollutant metals in dried blood spots via direct microextraction coupled to ICP-MS (μEx-ICP-MS).

Cameron J Stouffer, R Kenneth Marcus.

  Dried blood spot (DBS) sampling is a valuable method in clinical analysis due to its minimally invasive nature, reduced sample volume requirements, and simple storage conditions. DBS analysis is of specific interest in assessing exposure to medically relevant heavy metals, such as cadmium (Cd), mercury (Hg), and lead (Pb). However, the limited sample volume presents analytical challenges, particularly for low-concentration analytes, necessitating a highly sensitive and selective detection method. Inductively coupled plasma-mass spectrometry (ICP-MS) is the predominant technique for DBS heavy metal analysis, yet the complexity of the blood matrix often complicates direct analysis. After extraction of the sample from the substrate via acid digestion or ashing, chromatographic separations and additional pre-treatment steps are frequently required before MS detection, resulting in time-consuming workflows. To address these issues, we present an alternative DBS analysis approach utilizing the Advion Plate Express microextraction device (μEx-ICP-MS) for direct sampling with minimal sample preparation. This method enables rapid (<4 min), sensitive (sub to low μg L-1 detection limits), and multi-element profiling of DBS substrates. In this approach, a ∼2 × 4 mm region of the DBS sample is exposed to an extraction solvent, efficiently removing adsorbed species with direct delivery to the ICP nebulizer. The use of aqueous calibration standards for Cd, Hg, and Pb in real human blood samples provides matrix matched quantification. Additionally, studies examining sample extraction reproducibility and metal recovery from standard samples are presented. This method offers a streamlined, accessible approach for microextraction from DBS substrates and elemental detection in clinical analyses.

Keywords:  Dried blood spots; Heavy metals; ICP-MS; Microextraction

DOI:  https://doi.org/10.1016/j.talanta.2026.130714