bims-metlip 2024-10-20 papers

bims-metlip

Biomed News

on Methods and protocols in metabolomics and lipidomics

Issue of 2024–10–20
thirty papers selected by
Sofia Costa, Matterworks

EMBL-MCF 2.0: an LC-MS/MS method and corresponding library for high-confidence targeted and untargeted metabolomics using low-adsorption HILIC chromatography.
Met4DX: A Unified and Versatile Data Processing Tool for Multidimensional Untargeted Metabolomics Data.
A charge reversal approach for the sensitive quantification of dicarboxylic acids using Liquid chromatography-tandem mass spectrometry.
HPLC-MS/MS based method for the determination of 2-(3-hydroxy-5-phosphonooxymethyl-2-methyl-4-pyridyl)-1,3-thiazolidine-4-carboxylic acid in human plasma.
Highly reliable LC-MS lipidomics database for efficient human plasma profiling based on NIST SRM 1950.
Quantification of Letrozole, Palbociclib, Ribociclib, Abemaciclib, and Metabolites in Volumetric Dried Blood Spots: Development and Validation of an LC-MS/MS Method for Therapeutic Drug Monitoring.
Validation of a global method for the simultaneous analysis of polar and non-polar pesticides by online extraction and LC-MS/MS.
Molecular Imaging of Human Brain Organoids Using Mass Spectrometry.
Current approaches and outstanding challenges of functional annotation of metabolites: a comprehensive review.
Quantification of relevant metabolites in apoptotic bodies from HK-2 cells by targeted metabolomics based on liquid chromatography-tandem mass spectrometry.
CMSSP: A Contrastive Mass Spectra-Structure Pretraining Model for Metabolite Identification.
An integrated strategy for in-depth profiling of N-acylethanolamines in biological samples by UHPLC-HRMS.
MSBERT: Embedding Tandem Mass Spectra into Chemically Rational Space by Mask Learning and Contrastive Learning.
Facile fabrication of the immuno-MALDI-MS chip for the enrichment of abused drug in human urine integrated with MALDI-MS analysis.
Rapid quantification of polymyxin B in human pulmonary epithelial lining fluid by LC-MS/MS and its clinical application.
Advancing Bioanalytical Method Validation: A Comprehensive ICH M10 Approach for Validating LC-MS/MS to Quantify Fluoxetine in Human Plasma and Its Application in Pharmacokinetic Studies.
Application of UHPLC-QqQ-MS/MS Method for Quantification of Beta-Adrenergic Blocking Agents (β-Blockers) in Human Postmortem Specimens.
Targeted and Untargeted Amine Metabolite Quantitation in Single Cells with Isobaric Multiplexing.
Empirically establishing drug exposure records directly from untargeted metabolomics data.
SPME arrow-based extraction for enhanced targeted and untargeted urinary volatilomics.
LC-MS simultaneous profiling of acyl-CoA and acyl-carnitine in dynamic metabolic status.
Melt-blown polypropylene nonwoven as an efficient and eco-economic sorbent for pipette tip micro-solid phase extraction for the determination of tyrosine kinase inhibitors.
Development of LC-FAIMS-MS and its application to lipidomics study of Acinetobacter baumannii infection.
gcxgclab: An R Package for Two-Dimensional Gas Chromatography Preprocessing and Analysis.
Quality by design-based approach for the development of an analytical method for quantifying ponatinib in rat plasma.
Comprehensive two-dimensional gas chromatography as a tool for targeted and non-targeted analysis of contaminants of emerging concern in wastewater.
A Procedure for Solid-Phase Extractions Using Metal-Oxide-Coated Silica Column in Lipidomics.
Comprehensive characterization of active components in Salvia miltiorrhiza using polarity-partitioned two-dimensional liquid chromatography coupled with mass spectrometry.
Two-Dimensional Liquid Chromatography Tandem Mass Spectrometry Untangles the Deep Metabolome of Marine Dissolved Organic Matter.
Quantitative determination by UHPLC-MS/MS of 18 common drugs of abuse and metabolites, including THC and OH-THC, in volumetric dried blood spots: a sustainable method with minimally invasive sampling.

Metabolomics. 2024 Oct 13. 20(6): 114

EMBL-MCF 2.0: an LC-MS/MS method and corresponding library for high-confidence targeted and untargeted metabolomics using low-adsorption HILIC chromatography.

Svitlana Dekina, Theodore Alexandrov, Bernhard Drotleff.

   INTRODUCTION: Over the past two decades, liquid chromatography-mass spectrometry (LC-MS)-based metabolomics has experienced significant growth, playing a crucial role in various scientific disciplines. However, despite these advance-ments, metabolite identification (MetID) remains a significant challenge. To address this, stringent MetID requirements were established, emphasizing the necessity of aligning experimental data with authentic reference standards using multiple criteria. Establishing dependable methods and corresponding libraries is crucial for instilling confidence in MetID and driving further progress in metabolomics.
OBJECTIVE: The EMBL-MCF 2.0 LC-MS/MS method and public library was designed to facilitate both targeted and untargeted metabolomics with exclusive focus on endogenous, polar metabolites, which are known to be challenging to analyze due to their hydrophilic nature. By accompanying spectral data with robust retention times obtained from authentic standards and low-adsorption chromatography, high confidence MetID is achieved and accessible to the metabolomics community.
METHODS: The library is built on hydrophilic interaction liquid chromatography (HILIC) and state-of-the-art low adsorption LC hardware. Both high-resolution tandem mass spectra and manually optimized multiple reaction monitoring (MRM) transitions were acquired on an Orbitrap Exploris 240 and a QTRAP 6500+, respectively.
RESULTS: Implementation of biocompatible HILIC has facilitated the separation of isomeric metabolites with significant enhancements in both selectivity and sensitivity. The resulting library comprises a diverse collection of more than 250 biologically relevant metabolites. The methodology was successfully applied to investigate a variety of biological matrices, with exemplary findings showcased using murine plasma samples.
CONCLUSIONS: Our work has resulted in the development of the EMBL-MCF 2.0 library, a powerful resource for sensitive metabolomics analyses and high-confidence MetID. The library is freely accessible and available in the universal .msp file format under the CC-BY 4.0 license: mona.fiehnlab.ucdavis.edu https://mona.fiehnlab.ucdavis.edu/spectra/browse?query=exists(tags.text:%27EMBL-MCF_2.0_HRMS_Library%27) , EMBL-MCF 2.0 HRMS https://www.embl.org/groups/metabolomics/instrumentation-and-software/#MCF-library .

Keywords:  LC-MS/MS; Low-adsorption chromatography; Spectral library; Targeted metabolomics; Untargeted metabolomics

DOI:  https://doi.org/10.1007/s11306-024-02176-1
J Am Soc Mass Spectrom. 2024 Oct 14.

Met4DX: A Unified and Versatile Data Processing Tool for Multidimensional Untargeted Metabolomics Data.

Yandong Yin, Mingdu Luo, Zheng-Jiang Zhu.

  Liquid chromatography-mass spectrometry (LC-MS) is a powerful tool in untargeted metabolomics, enabling the high-sensitivity and high-specificity characterization of metabolites. The integration of ion mobility (IM) with LC-MS, known as LC-IM-MS, enhances the analytical depth, facilitating more comprehensive metabolite profiling. However, the complexity of data generated by these technologies presents significant challenges in data processing. Addressing these challenges, we developed Met4DX, a unified and versatile software tool for processing both 3D and 4D untargeted metabolomics data. Met4DX incorporates a new MS1-oriented peak detection approach coupled with our bottom-up assembly algorithm, enabling highly sensitive and comprehensive peak detection in untargeted metabolomics data. Additionally, Met4DX employs a uniform quantification strategy to enhance the precision of peak integration across different samples. The software provides a user-friendly interface that simplifies data processing with default parameter sets, consolidating peak detection, alignment, quantification, and other procedures into a single streamlined workflow. Together, Met4DX offers a comprehensive solution for multidimensional metabolomics data processing, transforming raw data from diverse MS instruments into a final feature table containing quantification and identification results. We postulate Met4DX facilitates metabolite discovery in biological samples by deciphering the complex untargeted metabolomics data. Met4DX is freely available on the Internet (https://met4dx.zhulab.cn/).

Keywords:  data processing; ion mobility-mass spectrometry; mass spectrometry; untargeted metabolomics

DOI:  https://doi.org/10.1021/jasms.4c00290
J Chromatogr A. 2024 Oct 09. pii: S0021-9673(24)00800-8. [Epub ahead of print]1737 465426

A charge reversal approach for the sensitive quantification of dicarboxylic acids using Liquid chromatography-tandem mass spectrometry.

Joby Jose, Alfred N Fonteh.

  Dicarboxylic acids (DCAs) are essential for intermediate metabolism and are implicated in multiple processes associated with various diseases. Several DCAs contribute to energy metabolism, impact mitochondrial function, and play a crucial role in body function. However, the low abundance of some DCAs in various body fluids makes their quantification particularly challenging. Therefore, an extremely sensitive method is required to determine DCA level fluctuations in biological samples in different diseases. We developed and optimized an LC-MS/MS method to quantify DCAs. We achieved charge reversal of the compounds from negative to positive ionization through chemical derivatization with dimethylaminophenacyl bromide (DmPABr) targeting the carboxyl group (R-COOH) under mild basic conditions. Derivatization enhanced sensitivity, mass fragmentation, and chromatographic separation for LC-tandem mass spectrometric quantification. The method was analytically optimized and demonstrated excellent linearity for individual DCAs (R2>0.99), as well as an exceptionally lower limit of detection (LLOD<266 fg) and lower limit of quantification (LLOQ<805 fg) for all DCAs. Furthermore, most derivatized DCAs were stable at room temperature and after ten repeated freeze-thaw cycles. After DCA extraction and quantification detection, we found differences in their distribution in plasma and urine. The rank order for DCAs in plasma is C4>C6>C7>C9>C5>C8>C22, whereas in the urine sample, the order is C4>C7>C6>C9>C5>C8>C10. For longer chains (C > 16), their proportions were >10x higher in plasma than in urine. Our optimized method using LC-MS/MS enables the quantification of DCAs with excellent sensitivity. The method will help in future studies investigating dicarboxylic acids' crucial role in health and biomarker discovery studies using targeted metabolomics.

Keywords:  Analytical validation; Charge reversal; Dicarboxylic acids; Liquid chromatography-tandem mass spectrometry (LC-MS/MS); Multiple reaction monitoring (MRM)

DOI:  https://doi.org/10.1016/j.chroma.2024.465426
Sci Rep. 2024 Oct 18. 14(1): 24425

HPLC-MS/MS based method for the determination of 2-(3-hydroxy-5-phosphonooxymethyl-2-methyl-4-pyridyl)-1,3-thiazolidine-4-carboxylic acid in human plasma.

Marta Gaweł, Rafał Głowacki, Paweł Kubalczyk, Justyna Piechocka.

  The report presents first high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) based method for the determination of plasma 2-(3-hydroxy-5-phosphonooxymethyl-2-methyl-4-pyridyl)-1,3-thiazolidine-4-carboxylic acid (HPPTCA), an adduct of cysteine and active form of vitamin B6 pyridoxal 5'-phosphate. The assay employs 4-deoxypyridoxine (4-DPD) as an internal standard. Sample preparation procedure primarily involves acetonitrile (ACN) extraction of HPPTCA from plasma proteins, sample deproteinization by ultrafiltration, and dilution of ultrafiltrate with mobile phase prior to chromatographic analysis. The chromatographic separations of HPPTCA and 4-DPD are achieved within 6 min at 20 °C on X-Bridge Glycan BEH Amide (100 × 2.1 mm, 2.5 μm) column using gradient elution. The eluent consists of 0.1% formic acid in a mixture of solvent A (water and ACN (95:5, v: v)) and solvent B (water and ACN (5:95, v: v)) delivered at a flow rate of 0.3 mL/min. The assay linearity was observed within 0.25-10 µmol/L in plasma. The limit of quantification was found to be 0.25 µmol/L. The method was successfully applied to plasma samples delivered by apparently healthy donors showing that the HPLC-MS/MS assay is suitable for human plasma screening. The presence of HPPTCA was confirmed in eleven of fifteen study samples. The HPPTCA concentration ranged from 0.55 to 8.39 µmol/L.

Keywords:  2-(3-hydroxy-5-phosphonooxymethyl-2-methyl-4-pyridyl)-1,3-thiazolidine-4-carboxylic acid; Cysteine; Hydrophilic interactions liquid chromatography; Plasma; Pyridoxal 5’-phosphate, tandem mass spectrometry

DOI:  https://doi.org/10.1038/s41598-024-75760-z
J Lipid Res. 2024 Oct 10. pii: S0022-2275(24)00176-7. [Epub ahead of print] 100671

Highly reliable LC-MS lipidomics database for efficient human plasma profiling based on NIST SRM 1950.

Sara Martínez, Miguel Fernández-García, Sara Londoño-Osorio, Coral Barbas, Ana Gradillas.

  Liquid chromatography coupled to high resolution mass spectrometry (LC-HRMS)-based methods have become the gold standard methodology for the comprehensive profiling of the human plasma lipidome. However, both the complexity of lipid chemistry and LC-HRMS-associated data pose challenges to the characterization of this biological matrix. In accordance with the current consensus of quality requirements for LC-HRMS lipidomics data, we aimed to characterize the NIST® Standard Reference Material for Human Plasma (SRM 1950) using an LC-ESI(+/-)-MS method compatible with high-throughput lipidome profiling. We generated a highly curated lipid database with increased coverage, quality, and consistency, including additional quality assurance procedures involving adduct formation, within-method m/z evaluation, retention behavior of species within lipid chain isomers, and expert-driven resolution of isomeric and isobaric interferences. As a proof-of-concept, we showed the utility of our in-house LC-MS lipidomic database -consisting of 592 lipid entries- for the fast, comprehensive, and reliable lipidomic profiling of the human plasma from healthy human volunteers. We are confident that the implementation of this robust resource and methodology will have a significant impact by reducing data redundancy and the current delays and bottlenecks in untargeted plasma lipidomic studies.

Keywords:  NIST SRM 1950; adduct profile; glycerolipids; glycerophospholipids; human plasma lipidome; in-house database; lipid annotation; lipidomic profiling; sphingolipids

DOI:  https://doi.org/10.1016/j.jlr.2024.100671
Int J Mol Sci. 2024 Sep 27. pii: 10453. [Epub ahead of print]25(19):

Quantification of Letrozole, Palbociclib, Ribociclib, Abemaciclib, and Metabolites in Volumetric Dried Blood Spots: Development and Validation of an LC-MS/MS Method for Therapeutic Drug Monitoring.

Eleonora Cecchin, Marco Orleni, Sara Gagno, Marcella Montico, Elena Peruzzi, Rossana Roncato, Lorenzo Gerratana, Serena Corsetti, Fabio Puglisi, Giuseppe Toffoli, Erika Cecchin, Bianca Posocco.

  Therapeutic drug monitoring (TDM) may be beneficial for cyclin-dependent kinase 4/6 inhibitors (CDK4/6is), such as palbociclib, ribociclib, and abemaciclib, due to established exposure-toxicity relationships and the potential for monitoring treatment adherence. Developing a method for quantifying CDK4/6is, abemaciclib metabolites (M2, M20), and letrozole in dried blood spots (DBS) could be useful to enhance the feasibility of TDM. Thus, an optimized LC-MS/MS method was developed using the HemaXis DB10 device for volumetric (10 µL) DBS collection. Chromatographic separation was achieved using a reversed-phase XBridge BEH C18 column. Detection was performed with a triple quadrupole mass spectrometer, utilizing ESI source switching between negative and positive ionization modes and multiple reaction monitoring acquisition. Analytical validation followed FDA, EMA, and IATDMCT guidelines, demonstrating high selectivity, adequate sensitivity (LLOQ S/N ≥ 30), and linearity (r ≥ 0.997). Accuracy and precision met acceptance criteria (between-run: accuracy 95-106%, CV ≤ 10.6%). Haematocrit independence was confirmed (22-55%),with high recovery rates (81-93%) and minimal matrix effects (ME 0.9-1.1%). The stability of analytes under home-sampling conditions was also verified. Clinical validation supports DBS-based TDM as feasible, with conversion models developed for estimating plasma concentrations (the reference for TDM target values) of letrozole, abemaciclib, and its metabolites. Preliminary data for palbociclib and ribociclib are also presented.

Keywords:  Ctrough; abemaciclib; dried blood spot; letrozole; mass spectrometry; palbociclib; plasma exposure; ribociclib; therapeutic drug monitoring

DOI:  https://doi.org/10.3390/ijms251910453
Anal Chim Acta. 2024 Nov 15. pii: S0003-2670(24)01032-8. [Epub ahead of print]1329 343231

Validation of a global method for the simultaneous analysis of polar and non-polar pesticides by online extraction and LC-MS/MS.

Susy Piovesana, Sara Elsa Aita, Chiara Cavaliere, Andrea Cerrato, Aldo Laganà, Carmela Maria Montone, Enrico Taglioni, Anna Laura Capriotti.

   BACKGROUND: Multi-residue methods for pesticide analysis in food are available for many compounds, but polar pesticides are not generally included due to their specific properties, which include high polarity and low molecular weight. Single residue methods are therefore needed for sample preparation, while chromatographic separation often requires derivatization, ion paring, or dedicated methods suitable for polar compounds, mostly ion chromatography and hydrophilic interaction liquid chromatography (HILIC). These challenges affect the important pesticide glyphosate and the related compounds aminomethylphosphonic acid (AMPA) and glufosinate. There are only a few methods including these compounds in large-scale analysis, mostly complex methods based on multidimensional chromatography.
RESULTS: A new method, for the global online extraction and analysis of pesticides in beer was developed and validated. The method exploited an online trapping device, with reversed-phase (RP) and anion exchange properties, that can trap small molecules from liquid samples. The ion exchange mechanism was used to retain the very polar pesticides glyphosate, AMPA, and glufosinate. The hydrophobic properties of the trapping column were also exploited to trap pesticides suitable for multi-residue investigations. The chromatographic separation was optimized by comparison of HILIC and RP C30, which could separate pesticides, including the polar ones, with modulation by the trapping column after proper selection of the mobile phase composition and basic modifier. The validation for beer provided recoveries in the range 71-112 %, with <15 % RSD, and LOD and LOQ values of 0.02-1 and 0.3-3 μg L-1, respectively. The result was competitive with previous methods on polar pesticide analysis in beer.
SIGNIFICANCE: The method was validated for 15 pesticides, over the log Kow range from -4.4 to 4.5, using a methodology with single and fast chromatographic separation under conditions compatible with multi-residue analysis by RP-LC-MS/MS. In the case of beer, for which the method was validated, the sample preparation was also performed online, after simple degassing, and sample dilution.

Keywords:  AMPA; Beer; Glyphosate; Online sample preparation; Pesticides; Reversed-phase chromatography

DOI:  https://doi.org/10.1016/j.aca.2024.343231
J Vis Exp. 2024 Sep 27.

Molecular Imaging of Human Brain Organoids Using Mass Spectrometry.

Saleh M Khalil, Gerarda Cappuccio, Feng Li, Mirjana Maletic-Savatic.

Brain organoid models serve as a powerful tool for studying human brain development and function. Mass spectrometry imaging (MSI), a cutting-edge technology, allows us to map the spatial distribution of diverse molecules such as lipids, sugars, amino acids, drugs, and their metabolites within these organoids, all without the need for specific molecular probes. High-quality MSI data hinge on meticulous sample preparation. Fixatives play a pivotal role, but conventional options such as glutaraldehyde, paraformaldehyde, and cryopreserving such as sucrose may inadvertently impact tissue metabolites. Optimal fixation entails flash freezing in liquid nitrogen. However, for small organoids, a more suitable approach involves transitioning the organoids directly from the incubator into a warmed embedding solution, followed by freezing in dry ice-cooled ethanol. Another critical step is the embedding prior to cryosectioning, which also requires materials compatible with MSI, as traditional options can interfere with matrix deposition and ionization. Here, an optimized protocol for high resolution-MALDI-MSI of human brain organoids is presented, encompassing sample preparation, sectioning, and imaging using mass spectrometry. This method showcases the molecular distribution of small metabolites, such as amino acids, with high mass accuracy and sensitivity. As such, coupled with complementary studies of brain organoids, it can assist in illuminating complex processes governing early brain development, metabolic cell fate trajectories, and distinctive metabolite signatures. Furthermore, it provides insights into the precise locations of molecules within the organoid, enriching our understanding of the spatial organization of 3D brain organoid models. As the field continues to advance, a growing number of studies leveraging MSI to delve into brain organoids and complex biological systems is anticipated, thereby deepening the understanding of the metabolic aspects of human brain function and development.

DOI: https://doi.org/10.3791/66997
Brief Bioinform. 2024 Sep 23. pii: bbae498. [Epub ahead of print]25(6):

Current approaches and outstanding challenges of functional annotation of metabolites: a comprehensive review.

Quang-Huy Nguyen, Ha Nguyen, Edwin C Oh, Tin Nguyen.

  Metabolite profiling is a powerful approach for the clinical diagnosis of complex diseases, ranging from cardiometabolic diseases, cancer, and cognitive disorders to respiratory pathologies and conditions that involve dysregulated metabolism. Because of the importance of systems-level interpretation, many methods have been developed to identify biologically significant pathways using metabolomics data. In this review, we first describe a complete metabolomics workflow (sample preparation, data acquisition, pre-processing, downstream analysis, etc.). We then comprehensively review 24 approaches capable of performing functional analysis, including those that combine metabolomics data with other types of data to investigate the disease-relevant changes at multiple omics layers. We discuss their availability, implementation, capability for pre-processing and quality control, supported omics types, embedded databases, pathway analysis methodologies, and integration techniques. We also provide a rating and evaluation of each software, focusing on their key technique, software accessibility, documentation, and user-friendliness. Following our guideline, life scientists can easily choose a suitable method depending on method rating, available data, input format, and method category. More importantly, we highlight outstanding challenges and potential solutions that need to be addressed by future research. To further assist users in executing the reviewed methods, we provide wrappers of the software packages at https://github.com/tinnlab/metabolite-pathway-review-docker.

Keywords:  functional analysis; liquid chromatography; mass spectrometry; metabolic pathways; metabolomics

DOI:  https://doi.org/10.1093/bib/bbae498
Anal Chim Acta. 2024 Nov 15. pii: S0003-2670(24)00991-7. [Epub ahead of print]1329 343190

Quantification of relevant metabolites in apoptotic bodies from HK-2 cells by targeted metabolomics based on liquid chromatography-tandem mass spectrometry.

Samuel Bernardo-Bermejo, Ana B Fernández-Martínez, Francisco Javier Lucio-Cazaña, María Castro-Puyana, María Luisa Marina.

   BACKGROUND: Apoptotic bodies play an important role in the cellular communication as a consequence of the great variety of biomolecules they harbor. There is evidence that 1st generation apoptotic bodies from HK-2 cells induced by cisplatin or UV light trigger apoptosis in naïve HK-2 cells whereas 2nd generation apoptotic bodies activate cell proliferation showing an opposite effect. Thus, the development of new analytical strategies to quantify the changes in the involved metabolites is imperative to shed light on the biological mechanisms which trigger apoptosis and cell proliferation.
RESULTS: A LC-(Q-Orbitrap)MS method has been developed to quantify the metabolites unequivocally identified in the apoptotic body fluid from HK-2 cells in our previous works based on untargeted metabolomics. Thus, two different columns and gradients were tested and the HILIC column was selected taking into account the retention times and chromatographic separation. Also, different normal collision energies were tested for each metabolite and the parallel reaction monitoring was chosen to carry out the quantitative analysis. Once the method was optimized, it was evaluated in terms of linearity, limits of detection and quantification, matrix effects, accuracy, and precision, for each metabolite. Limits of detection ranged from 0.02 to 1.4 ng mL-1. A total of 9 relevant metabolites proposed as potential biomarkers to reveal metabolic differences among apoptotic bodies from HK-2 cells were quantified and some insights about the biological relevance were discussed.
SIGNIFICANCE: The first targeted metabolomics methodology enabling the quantification of relevant metabolites in apoptotic bodies from HK-2 cells was developed using LC-(Q-Orbitrap)MS. Pyridoxine, kynurenine, and creatine concentrations were determined in apoptotic bodies from HK-2 cells treated with cisplatin and UV light. Phenylacetylglycine, hippuric acid, butyrylcarnitine, acetylcarnitine, carnitine, and phenylalanine were determined in 1st and 2nd generation apoptotic bodies from HK-2 cells treated with cisplatin. Concentrations determined were useful to establish their biological role in the metabolism.

Keywords:  Apoptotic bodies; HK-2 cells; Liquid chromatography; Q-orbitrap-mass spectrometry; Quantitative analysis; Targeted metabolomics

DOI:  https://doi.org/10.1016/j.aca.2024.343190
Anal Chem. 2024 Oct 14.

CMSSP: A Contrastive Mass Spectra-Structure Pretraining Model for Metabolite Identification.

Lu Chen, Bing Xia, Yu Wang, Xia Huang, Yucheng Gu, Wenlin Wu, Yan Zhou.

A pivotal challenge in metabolite research is the structural annotation of metabolites from tandem mass spectrometry (MS/MS) data. The integration of artificial intelligence (AI) has revolutionized the interpretation of MS data, facilitating the identification of elusive metabolites within the metabolomics landscape. Innovative methodologies are primarily focusing on transforming MS/MS spectra or molecular structures into a unified modality to enable similarity-based comparison and interpretation. In this work, we present CMSSP, a novel Contrastive Mass Spectra-Structure Pretraining framework designed for metabolite annotation. The primary objective of CMSSP is to establish a representation space that facilitates a direct comparison between MS/MS spectra and molecular structures, transcending the limitations of distinct modalities. The evaluation on two benchmark test sets demonstrates the efficacy of the approach. CMSSP achieved a remarkable enhancement in annotation accuracy, outperforming the state-of-the-art methods by a significant margin. Specifically, it improved the top-1 accuracy by 30% on the CASMI 2017 data set and realized a 16% increase in top-10 accuracy on an independent test set. Moreover, the model displayed superior identification accuracy across all seven chemical categories, showcasing its robustness and versatility. Finally, the MS/MS data of 30 metabolites from Glycyrrhiza glabra were analyzed, achieving top-1 and top-3 accuracies of 86.7 and 100%, respectively. The CMSSP model serves as a potent tool for the dissection and interpretation of intricate MS/MS data, propelling the field toward more accurate and efficient metabolite annotation. This not only augments the analytical capabilities of metabolomics but also paves the way for future discoveries in understanding of complex biological systems.

DOI: https://doi.org/10.1021/acs.analchem.4c03724
Anal Chim Acta. 2024 Nov 15. pii: S0003-2670(24)01063-8. [Epub ahead of print]1329 343262

An integrated strategy for in-depth profiling of N-acylethanolamines in biological samples by UHPLC-HRMS.

Jun-Yi Zhou, Yan-Qing Chen, Guang Hu, Haiyu Zhao, Jian-Bo Wan.

   BACKGROUND: N-acylethanolamines (NAEs) are a class of naturally occurring bioactive lipids that play crucial roles in various physiological processes, particularly exhibiting neuroprotective and anti-inflammatory properties. However, the comprehensive profiling of endogenous NAEs in complex biological matrices is challenging due to their low abundance, structural similarity and the limited availability of commercial standards. Here, we propose an integrated strategy for comprehensive profiling of NAEs that combines chemical derivatization and a three-dimensional (3D) prediction model based on quantitative structure-retention time relationship (QSRR) using liquid chromatography coupled with high-resolution tandem mass spectrometry (LC-HRMS).
RESULTS: After acetyl chloride (ACC) derivatization, the detection sensitivity of NAEs was significantly improved. We developed a QSRR prediction model to construct an in-house database for 141 NAEs, encompassing information on RT, MS1 (m/z), and MS/MS spectra. Propargylamine-labeled fatty acids were synthesized as RT calibrants across various analytical conditions to enhance the robustness of the RT prediction model. NAEs in biological samples were then in-depth profiled using parallel reaction monitoring (PRM) acquisition. This integrated strategy identified and annotated a total of 50 NAEs across serum, hippocampus and cortex tissues from a 5xFAD mouse model of Alzheimer's disease (AD). Notably, the levels of polyunsaturated NAEs, particularly NAE 20:5 and NAE 22:6, were significantly decreased in 5xFAD mice compared to WT mice, as confirmed by accurate quantitation using ACC-d0/d3 derivatization.
SIGNIFICANCE: Our integrated strategy exhibits great potential for the in-depth profiling of NAEs in complex biological samples, facilitating the elucidation of NAE functions in diverse physiological and pathological processes.

Keywords:  Alzheimer's disease; Chemical derivatization; N-acylethanolamides; Parallel reaction monitoring; UHPLC-HRMS

DOI:  https://doi.org/10.1016/j.aca.2024.343262
Anal Chem. 2024 Oct 14.

MSBERT: Embedding Tandem Mass Spectra into Chemically Rational Space by Mask Learning and Contrastive Learning.

Hailiang Zhang, Qiong Yang, Ting Xie, Yue Wang, Zhimin Zhang, Hongmei Lu.

Tandem mass spectrometry (MS/MS) is a powerful technique for chemical analysis in many areas of science. The vast MS/MS spectral data generated in liquid chromatography-mass spectrometry (LC-MS) experiments require efficient analysis and interpretation methods for the following compound identification. In this study, we propose MSBERT based on self-supervised learning strategies to embed MS/MS spectra into reasonable embeddings for efficient compound identification. It adopts the transformer encoder as the backbone for mask learning and uses the same spectra with different masks for contrastive learning. MSBERT is trained on the GNPS data set and tested on the GNPS data set, the MoNA data set, and the MTBLS1572 data set. It exhibits enhanced library matching and analogous compound searching capabilities compared to existing methods. The recalls at 1, 5, and 10 on a GNPS test subset with structures not in the training set are 0.7871, 0.8950, and 0.9080, respectively. The results are better than those of Spec2Vec with 0.6898, 0.8276, and 0.8620, and DreaMS with 0.7158, 0.8327, and 0.8635. The rationality of embeddings is demonstrated by t-SNE visualization, structural similarity, spectra clustering, compound identification, and analogous compound searching. A user-friendly web server is provided for efficient spectral analysis, and the source code for MSBERT is available at https://github.com/zhanghailiangcsu/MSBERT.

DOI: https://doi.org/10.1021/acs.analchem.4c02426
Anal Chim Acta. 2024 Nov 15. pii: S0003-2670(24)01025-0. [Epub ahead of print]1329 343224

Facile fabrication of the immuno-MALDI-MS chip for the enrichment of abused drug in human urine integrated with MALDI-MS analysis.

Hsin-Ping Chen, He-Hsuan Hsiao.

   BACKGROUND: Drug abuse can result in both physical and mental health issues for individuals, and can also contribute to broader societal problems. The number of drug abuse cases rose to 296 million in 2021. The sample pretreatment methods commonly employed typically require longer processing times and occasionally necessitate derivatization. Furthermore, with the increase in sample sizes, traditional chromatography-mass spectrometry methods for analyzing abused drugs were no longer sufficient to handle such numerous samples. In this study, immuno-MALDI-MS chip were fabricated for specific enrichment of illicit drugs, integrating with the rapid and accurate capabilities of MALDI-MS for high-throughput analysis of drug abuse.
RESULTS: The immuno-MALDI-MS chip was successfully prepared by coating an aluminum chip with antibody-conjugated boronic acid-modified gold nanoparticles. Ketamine, a frequently abused illicit drug, served as the proof of concept for this study. The immuno-MALDI-MS chip was employed to selectively enrich ketamine in human urine samples, facilitating direct MALDI-MS analysis with the addition of α-CHCA matrix solution. The challenge of detecting abused drugs, exacerbated by interfering peaks in the low m/z region from salts and small molecules in human urine samples, was successfully overcome. The developed method exhibited a wide linear range of 10-5000 ng/mL with a limit of detection of 3.3 ng/mL for ketamine. Notably, the proposed method enabled high-throughput screening and accurate confirmation of ketamine concentrations in suspects' urine samples within few minutes, requiring a minimal sample volume of 1 μL. The obtained data were in complete agreement with the previous GC/MS analysis.
SIGNIFICANCE: A straightforward, cost-effective and sensitive method for the selective enrichment and absolute quantification of abused drugs was developed using a homemade immuno-MALDI-MS chip integrated with MALDI-MS analysis. This method combines the advantages of immunoassay and mass spectrometry, offering both speed and accuracy. The reported method for the quantification of ketamine in human urine offers a practical approach and has the potential to analyze emerging new psychoactive substances in the future.

Keywords:  Abused drug; Immuno-MALDI-MS chip; Ketamine; Matrix-assisted laser desorption ionization mass spectrometer; Single drop analysis

DOI:  https://doi.org/10.1016/j.aca.2024.343224
J Chromatogr B Analyt Technol Biomed Life Sci. 2024 Oct 01. pii: S1570-0232(24)00341-6. [Epub ahead of print]1247 124332

Rapid quantification of polymyxin B in human pulmonary epithelial lining fluid by LC-MS/MS and its clinical application.

Yu Wang, Xiaofen Liu, Yan Chen, Beining Guo, Jiao Liu, Jing Zhang.

  Pulmonary epithelial lining fluid (ELF) was commonly used for the pharmacokinetic study in lower respiratory tract infections. To characterize the intrapulmonary pharmacokinetic properties of polymyxin B following aerosol delivery, we developed and fully validated a liquid chromatography/tandem mass spectrometry (LC/MS/MS) method for quantifying polymyxin B in human bronchoalveolar lavage fluid (BALF). The ELF concentrations were calculated by the BALF values of polymyxin B using urea as a volume normalizer. Chromatographic separation was achieved on a Phonomenex Kinetex XB-C18 column(100 mm × 2.1 mm I.D., 2.6 μm)in acetonitrile and water both containing 0.2 % formic acid. The flow rate was set as 0.4 mL/min for a 3.5 min running time. Protein precipitation was used in preparing BALF samples with polymyxin E1 as an internal standard. Polymyxin B was detected under multiple reaction monitoring conditions using the electrospray ionization interface running in the positive ionization mode. The assay showed a good linear relationship over the tested concentration ranges of 0.0300/0.00306---10.0/1.02 mg/L for polymyxin B1/B2 in bronchoalveolar lavage fluid (R2 > 0.99). The inter- and inter-day precisions (RSD, %) were < 12.2 %(15.2 % for LLOQ samples)and the accuracies (%) were within the range of 94.3 ∼ 110.4 %. This reliable LC-MS/MS method for detection of polymyxin B was successfully applied to conduct a pulmonary penetration study in patients following aerosol administration.

Keywords:  Human pulmonary epithelial lining fluid; LC-MS/MS; Pneumonia patients; polymyxin B

DOI:  https://doi.org/10.1016/j.jchromb.2024.124332
Molecules. 2024 Sep 27. pii: 4588. [Epub ahead of print]29(19):

Advancing Bioanalytical Method Validation: A Comprehensive ICH M10 Approach for Validating LC-MS/MS to Quantify Fluoxetine in Human Plasma and Its Application in Pharmacokinetic Studies.

Aimen El Orche, Amine Cheikh, Choukri El Khabbaz, Houda Bouchafra, My El Abbes Faouzi, Yahya Cherrah, Siddique Akber Ansari, Hamad M Alkahtani, Shoeb Anwar Ansari, Mustapha Bouatia.

  A fast and sample cleanup approach for fluoxetine in human plasma was developed using protein precipitation coupled with LC-MS-MS. Samples were treated with methanol prior to LC-MS-MS analysis. Chromatographic separation was performed on a reverse phase column with an isocratic mobile phase of methanol and 10 mM ammonium formate pH acidified with formic acid (80:20, v/v) at a flow rate of 0.2 mL/min. The run time was 4 min. Mass parameters were optimized to monitor transitions at m/z [M + H]+ 310 > > 148 for fluoxetine and m/z [M + H]+ 315.1 > > 153 for fluoxetine-d5 as an internal standard. The lower limit of quantification and the dynamic range were 0.25 and 0.25-50 ng/mL, respectively. Linearity was good for intra-day and inter-day validations (R2 = 0.999). The matrix effect was acceptable with CV% < 15 and accuracy% < 15. The hemolytic effect was negligible. Fluoxetine was stable in human plasma for 48 h at room temperature (25 °C), for 12 months frozen at -25 °C, for 48 h in an auto-sampler at 6 °C, and for three freeze/thaw cycles. The validated method was applied in a pharmacokinetic study to determine the concentration of fluoxetine in plasma samples. The study provides a fast and simple bioanalytical method for routine analysis and may be particularly useful for bioequivalence studies. The method was successfully applied to a pharmacokinetic study of fixed-dose fluoxetine in nine healthy volunteers.

Keywords:  LC-MS-MS; bioanalysis; fluoxetine; pharmacokinetics application; validation

DOI:  https://doi.org/10.3390/molecules29194588
Molecules. 2024 Sep 27. pii: 4585. [Epub ahead of print]29(19):

Application of UHPLC-QqQ-MS/MS Method for Quantification of Beta-Adrenergic Blocking Agents (β-Blockers) in Human Postmortem Specimens.

Paweł Szpot, Kaja Tusiewicz, Olga Wachełko, Marcin Zawadzki.

  Betablockers are one of the most frequently used medications in cardiology. They can lead to fatal drops in blood pressure and heart rhythm disturbances. Death is functional, and poisoning with this group of drugs can be difficult to detect. The liquid-liquid extraction (LLE) method developed using ethyl acetate at pH 9 successfully identified 18 β-blockers in human blood. The method's limit of quantification (LOQ) was in the range of 0.1 to 0.5 ng/mL. No carryover of substances between samples was detected, and no interfering ion current signals were observed in the biological samples at the retention times of the compounds or internal standards. All compounds had a coefficient of determination (R2) above 0.995. Intraday and interday precision (RSD%) and accuracy (RE%) for low and high QC levels were within 1.7-12.3% and -14.4 to 14.1%, respectively. Very good recovery (80.0-119.6%) and matrix effect (±20.0%) values were achieved for all compounds. In addition, fragmentation spectra were collected for all the examined substances, and high-resolution spectra were presented for landiolol and metipranolol, because they are not available in commercial HRMS spectra databases. The developed method was applied in authentic postmortem samples.

Keywords:  UHPLC-QqQ-MS/MS; beta-adrenergic receptor antagonists; betablockers; drugs intoxication; forensic toxicology

DOI:  https://doi.org/10.3390/molecules29194585
Chemistry. 2024 Oct 18. e202403278

Targeted and Untargeted Amine Metabolite Quantitation in Single Cells with Isobaric Multiplexing.

Juho Heininen, Parisa Movahedi, Tapio Kotiaho, Risto Kostiainen, Tapio Pahikkala, Jaakko Teppo.

  We developed a single cell amine analysis approach utilizing isobarically multiplexed samples of 6 individual cells along with analyte abundant carrier. This methodology was applied for absolute quantitation of amino acids and untargeted relative quantitation of amines in a total of 108 individual cells using nanoflow LC with high-resolution mass spectrometry. Together with individually determined cell sizes, this provides quantification of intracellular concentrations within individual cells. The targeted method was partially validated for 10 amino acids with limits of detection in low attomoles, linear calibration range covering analyte amounts typically from 30 amol to 120 fmol, and correlation coefficients (R) above 0.99. This was applied with cell sizes recorded during dispensing to determine millimolar intracellular amino acid concentrations. The untargeted approach yielded 249 features that were detected in at least 25% of the single cells, providing modest cell type separation on principal component analysis. Using Greedy forward selection with regularized least squares, a sub-selection of 100 features explaining most of the difference was determined. These features were annotated using MS2 from analyte standards and accurate mass with library search. The approach provides accessible, sensitive, and high-throughput method with the potential to be expanded also to other forms of ultrasensitive analysis.

Keywords:  Mass spectrometry; Metabolomics; Single Cell; absolute quantitation; liquid chromatography

DOI:  https://doi.org/10.1002/chem.202403278
bioRxiv. 2024 Oct 10. pii: 2024.10.07.617109. [Epub ahead of print]

Empirically establishing drug exposure records directly from untargeted metabolomics data.

Haoqi Nina Zhao, Kine Eide Kvitne, Corinna Brungs, Siddharth Mohan, Vincent Charron-Lamoureux, Wout Bittremieux, Runbang Tang, Robin Schmid, Santosh Lamichhane, Yasin El Abiead, Mohammadsobhan S Andalibi, Helena Mannochio-Russo, Madison Ambre, Nicole E Avalon, MacKenzie Bryant, Andrés Mauricio Caraballo-Rodríguez, Martin Casas Maya, Loryn Chin, Ronald J Ellis, Donald Franklin, Sagan Girod, Paulo Wender P Gomes, Lauren Hansen, Robert Heaton, Jennifer E Iudicello, Alan K Jarmusch, Lora Khatib, Scott Letendre, Sarolt Magyari, Daniel McDonald, Ipsita Mohanty, Andrés Cumsille, David J Moore, Prajit Rajkumar, Dylan H Ross, Harshada Sapre, Mohammad Reza Zare Shahneh, Sydney P Thomas, Caitlin Tribelhorn, Helena M Tubb, Corinn Walker, Crystal X Wang, Shipei Xing, Jasmine Zemlin, Simone Zuffa, David S Wishart, Rima Kaddurah-Daouk, Mingxun Wang, Manuela Raffatellu, Karsten Zengler, Tomáš Pluskal, Libin Xu, Rob Knight, Shirley M Tsunoda, Pieter C Dorrestein.

Despite extensive efforts, extracting information on medication exposure from clinical records remains challenging. To complement this approach, we developed the tandem mass spectrometry (MS/MS) based GNPS Drug Library. This resource integrates MS/MS data for drugs and their metabolites/analogs with controlled vocabularies on exposure sources, pharmacologic classes, therapeutic indications, and mechanisms of action. It enables direct analysis of drug exposure and metabolism from untargeted metabolomics data independent of clinical records. Our library facilitates stratification of individuals in clinical studies based on the empirically detected medications, exemplified by drug-dependent microbiota-derived N -acyl lipid changes in a human immunodeficiency virus cohort. The GNPS Drug Library holds potential for broader applications in drug discovery and precision medicine.

DOI: https://doi.org/10.1101/2024.10.07.617109
Anal Chim Acta. 2024 Nov 15. pii: S0003-2670(24)01062-6. [Epub ahead of print]1329 343261

SPME arrow-based extraction for enhanced targeted and untargeted urinary volatilomics.

Maria Llambrich, Noelia Ramírez, Raquel Cumeras, Jesús Brezmes.

   BACKGROUND: Volatile organic compounds (VOCs) present in human urine are promising biomarkers for various health conditions and environmental exposures. However, their reliable detection is challenging due to the complexity of urinary matrices and the low concentrations of VOCs. Moreover, untargeted approaches present considerable challenges in terms of data interpretation, increasing the complexity of method development. Here we address these challenges by developing a new method that combines solid-phase microextraction (SPME) Arrow with gas chromatography-high resolution mass spectrometry (GC-HRMS), using a design of experiments (DOE) approach for targeted and untargeted compounds. This methodology, specifically tailored for SPME Arrow, represents a significant advancement in untargeted urinary analysis.
RESULTS: The method was developed based on targeted and untargeted outcomes, were ranking results focus on the highest response area of 11 spiked target VOCs representative of urinary volatilomics, and on identifying the maximum untargeted number of VOCs. The method was developed focusing on the highest response area of 11 spiked target VOCs representative of urinary volatilomics and identifying the maximum number of VOCs. A univariate method determined the optimal coating type, urine volume, and salt addition. Subsequently, a central composite design (CCD) DOE was used to determine ideal temperature, extraction, and incubation times. The best method obtained has an extraction time of 60 min at a temperature of 53 °C, with an SPME Arrow CAR/PDMS using 2 mL of urine, with 0.25 % w/v of NaCl and a pH of 2. Compared to conventional SPME fibers, the SPME Arrow showed improved extraction efficiency, detecting more VOCs. Finally, the enhanced method was successfully applied to urine samples from children exposed and non-exposed to tobacco smoke, identifying specific VOCs, like p-cymene and p-isopropenyl toluene related to tobacco exposure.
SIGNIFICANCE: By integrating both targeted and untargeted approaches, the developed method comprehensively captures the complexity of urinary metabolomics. This dual strategy ensures the precise identification of known compounds and the discovery of novel biomarkers, thereby providing a more complete metabolic profile. Such an approach is crucial for advancing in non-invasive diagnostics and environmental health studies, as it offers deeper insights into the intricate relationships between metabolic processes and various health conditions.

Keywords:  Metabolomics; SPME arrow; Tobacco smoke exposure; Untargeted analysis; Urine; Volatilomics

DOI:  https://doi.org/10.1016/j.aca.2024.343261
Anal Chim Acta. 2024 Nov 15. pii: S0003-2670(24)01036-5. [Epub ahead of print]1329 343235

LC-MS simultaneous profiling of acyl-CoA and acyl-carnitine in dynamic metabolic status.

Jiangang Zhang, Juan Lei, Xudong Liu, Nan Zhang, Lei Wu, Yongsheng Li.

DOI: https://doi.org/10.1016/j.aca.2024.343235
Anal Chim Acta. 2024 Nov 15. pii: S0003-2670(24)01041-9. [Epub ahead of print]1329 343240

Melt-blown polypropylene nonwoven as an efficient and eco-economic sorbent for pipette tip micro-solid phase extraction for the determination of tyrosine kinase inhibitors.

Jing Ye, Yuanyuan Zheng, Xiangyu Li, Xingyi Jiang, Di Chen.

   BACKGROUND: The detection of tyrosine kinase inhibitors (TKIs) in biological fluids is essential due to their critical role in cancer therapy and the variability in individual drug metabolism, which necessitates precise dosing. Traditional methods for analyzing TKIs in biological fluids, such as blood plasma, typically involve complex sample preparation techniques that can be resource-intensive, environmentally burdensome, and not sufficiently sensitive for low-concentration analytes. There is a pressing need for more efficient, economical, and environmentally friendly methods that can enhance sensitivity and throughput without compromising accuracy.
RESULTS: This study explores the use of melt-blown polypropylene nonwoven (MBPP), commonly found in face masks, as a novel sorbent for pipette-tip micro-solid phase extraction (PT-μSPE). MBPP demonstrated excellent hydrophobicity and significant mesoporous adsorption capacity. An extraction device was fashioned by inserting a segment of MBPP (15 mg) into a 200 μL disposable plastic pipette tip, which was then attached to a 2.5 mL disposable plastic syringe. The MBPP's fabric form removes the need for a frit, allowing the extraction process to be completed in just 3 min through simple plunger manipulation. The method achieved extraction recoveries ranging from 60.5 % to nearly 100 %. Subsequent method validation using liquid chromatography-tandem mass spectrometry (LC-MS/MS) showed satisfactory linearity (coefficient of determination R2 > 0.993), accuracy (relative recoveries: 86.3%-114.8 %), and precision (relative standard deviation: 3.4%-11.3 %), with detection limits between 0.022 and 0.135 ng mL-1.
SIGNIFICANCE: The introduction of MBPP for PT-μSPE represents a significant advancement in the bioanalytical detection of TKIs, offering a highly efficient, cost-effective, and environmentally sustainable method. It compares favorably with existing techniques, offering advantages in terms of cost, environmental impact, and ease of use. This approach has the potential to be widely adopted for routine monitoring of TKIs in clinical settings.

Keywords:  Liquid chromatography-tandem mass spectrometry; Mask; Melt-blown polypropylene nonwoven; Pipette-tip micro-solid phase extraction; Plasma; Tyrosine kinase inhibitors

DOI:  https://doi.org/10.1016/j.aca.2024.343240
J Lipid Res. 2024 Oct 10. pii: S0022-2275(24)00173-1. [Epub ahead of print] 100668

Development of LC-FAIMS-MS and its application to lipidomics study of Acinetobacter baumannii infection.

Jianjun Li, Jacek Stupak, Arsalan S Haqqani, Greg Harris, Hongyan Zhou, Sam Williamson, Rui Chen, H Howard Xu, Wangxue Chen.

  The recent advances in mass spectrometry (MS) technologies have enabled comprehensive lipid profiling in biological samples. However, the robustness and efficiency of MS-based lipidomics is compromised by the complexity of biological samples. High-field asymmetric waveform ion mobility spectrometry (FAIMS) is a technology that can continuously transmit one type of ion, independent of mass-to-charge ratio. Here we present the development and application of LC-FAIMS-MS/MS based platform for untargeted lipidomics. We used 3 optimally balanced compensation voltages, i.e., 29 V, 34 V and 39 V, to analyse all subclasses of glycerophospholipids. The reproducibility of the method was evaluated using reference standards. The reproducibility of retention times ranged from 0.9 to 1.5 % RSD; whereas RSD values of 5-10 % were observed for peak areas. More importantly, the coupling of a FAIMS device can significantly improve the robustness and efficiency. We exploited this NPLC-FAIMS-HRMS to analyze the serum lipid profiles in mice infected intranasally with Acinetobacter baumannii. The temporal profiles of serum lipids after A. baumannii inoculation were obtained for 4 h, 8 h and 24 h. We found that nearly all ether PC and ether PE lipids were significantly decreased 8 h after inoculation. The resultant volcano plot illustrated the distribution of 28 increased and 28 decreased lipid species in mouse sera 24 h after inoculation. We also found that a single ether PE composition can comprise multiple isomeric structures, and the relative abundance of each isomer could be quantified using the newly developed NPLC-FAIMS-PRM method.

Keywords:  Acinetobacter baumannii; FAIMS; NPLC; bacterial infection; lipidomics; mass spectrometry; phospholipid

DOI:  https://doi.org/10.1016/j.jlr.2024.100668
Anal Chem. 2024 Oct 16.

gcxgclab: An R Package for Two-Dimensional Gas Chromatography Preprocessing and Analysis.

Stephanie N Gamble, Caroline O Granger, Joseph M Mannion.

Comprehensive two-dimensional gas chromatography (GC×GC) is an established technique capable of chromatographically separating thousands of analytes in complex matrices. When coupled with highly sensitive detectors such as a high-resolution mass spectrometer, these instruments produce large multidimensional data sets. A prevailing challenge for GC×GC users is efficient data handling and analysis. Although commercial software packages exist for GC×GC data analysis, these platforms are typically optimized for low-throughput qualitative data interpretation utilizing desktop computer systems. Additionally, commercial GC×GC data processing packages offer little flexibility to explore custom or novel data processing concepts. In this work, an open-source R package, called "gcxgclab", was developed as an alternative data processing package for GC×GC users and offers data preprocessing and analysis functions including baseline correction, smoothing, phase shift, peak detection, peak alignment, extracted ion chromatogram, mass spectra extraction, mass defect analysis, and targeted and nontargeted analysis. This package was designed to be customizable and allows for batch data processing on desktops or high-performance computing systems to increase the throughput. gcxgclab is available for free on the Comprehensive R Archive Network (CRAN) at https://cran.r-project.org/web/packages/gcxgclab/index.html.

DOI: https://doi.org/10.1021/acs.analchem.4c03126
Heliyon. 2024 Oct 15. 10(19): e38637

Quality by design-based approach for the development of an analytical method for quantifying ponatinib in rat plasma.

Nahyun Koo, Eun Ji Lee, Min Ju Kim, Minjung Park, Kyeong-Ryoon Lee, Yoon-Jee Chae.

  Ponatinib is a potent tyrosine kinase inhibitor that is approved for the treatment of chronic myeloid leukemia and Philadelphia chromosome-positive acute lymphoblastic leukemia. To further expand its clinical applications, accurate quantification of ponatinib in plasma is essential. In this study, we developed and validated a sensitive and selective high-performance liquid chromatography (HPLC) method coupled with a fluorescence detector (FLD) to measure ponatinib concentrations in rat plasma using the Analytical Quality by Design approach. Briefly, we screened and optimized the critical method parameters using the Taguchi and Box-Behnken designs. The developed method had excellent linearity in the range of 1-1000 ng/mL, sensitivity, and reproducibility, and required minimal sample volume and a short run time. Compared with previously reported HPLC-ultraviolet (UV) and liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods, this HPLC-FLD method offers superior sensitivity, simpler sample preparation, and greater efficiency. We successfully used this method in a pharmacokinetic study in rats to obtain reliable data on ponatinib plasma concentrations. Altogether, this analytical method will be applicable in several analytical conditions and will support further pharmacokinetic and clinical investigations of ponatinib for various cancer treatments.

Keywords:  Analytical quality by design; Box-Behnken design; Method development and validation; Ponatinib; Taguchi method

DOI:  https://doi.org/10.1016/j.heliyon.2024.e38637
Talanta. 2024 Oct 10. pii: S0039-9140(24)01411-5. [Epub ahead of print]282 127032

Comprehensive two-dimensional gas chromatography as a tool for targeted and non-targeted analysis of contaminants of emerging concern in wastewater.

Jason Devers, David I Pattison, Asger B Hansen, Jan H Christensen.

  Wastewater is a major reservoir for chemical contaminants, both anthropogenic and biogenic. Recent chemical and toxicological analysis reveals the abundance and impact of these compounds, often termed contaminants of emerging concern (CECs). Concurrently, incomplete removal of these compounds in wastewater treatment plants sets a precedent for detailed characterisation and monitoring of such substances. Although liquid chromatography (LC) is frequently used for analysis of CECs in wastewater, gas chromatography (GC) maintains its significance for non-polar to mid-polar analytes. GC offers advantages such as increased separation efficiency, fewer matrix effects, and greater availability and reliability of reference mass spectra compared to LC. Comprehensive two-dimensional gas chromatography (GC × GC) delivers unmatched peak capacity and separational capabilities, critical in the resolution of diverse compound groups present within wastewater. When coupled with high resolution mass spectrometry, it provides a powerful identification tool with spectral databases and both 1st and 2nd dimensional retention indices, and has allowed for the separation, reliable annotation and characterisation of diverse CECs within wastewater in recent years. Herein, on the basis of recent studies from the last fifteen years, we outline cutting-edge methodologies and strategies for wastewater analysis using GC × GC. This includes sample preparation, derivatization of polar analytes, instrumental setup, and data analysis, ultimately providing the reader a framework for future non-targeted analysis of wastewater and other complex environmental matrices.

Keywords:  Derivatization; GC×GC; Non-targeted analysis; Wastewater

DOI:  https://doi.org/10.1016/j.talanta.2024.127032
Anal Chem. 2024 Oct 15.

A Procedure for Solid-Phase Extractions Using Metal-Oxide-Coated Silica Column in Lipidomics.

Hiroaki Takeda, Manami Takeuchi, Mayu Hasegawa, Junki Miyamoto, Hiroshi Tsugawa.

Lipid enrichment is indispensable for enhancing the coverage of targeted molecules in mass spectrometry (MS)-based lipidomics studies. In this study, we developed a simple stepwise fractionation method using a titanium- and zirconium-dioxide-coated solid-phase extraction (SPE) silica column that separates neutral lipids, phospholipids, and other lipids, including fatty acids (FAs) and glycolipids. Chloroform was used to dissolve the lipids, and neutral lipids, including steryl esters, diacylglycerols, and triacylglycerols, were collected in the loading fraction. Second, methanol with formic acid (99:1, v/v) was used to retrieve FAs, ceramides, and glycolipids, including glycosylated ceramides and glycosylated diacylglycerols, by competing for affinity with the Lewis acid sites on the metal oxide surface. Finally, phospholipids strongly retained via chemoaffinity interactions were eluted using a solution containing 5% ammonia and high water content (45:50 v/v, 2-propanol:water), which canceled the electrostatic and chelating interactions with the SPE column. High average reproducibility of <10% and coverage of ∼100% compared to those of the non-SPE samples were demonstrated by untargeted lipidomics of human plasma and mouse brain, testis, and feces. The advantage of our procedure was showcased by characterizing minor lipid subclasses, including dihexosylceramides containing very long-chain polyunsaturated FA in the testis, monogalactosyl and digalactosyl monoacylglycerols in feces, and acetylated and glycolylated derivatives of gangliosides in the brain that were not detected using conventional solvent extraction methods. Likewise, the value of our method in biology is maximized during glycolipidome profiling in the absence of neutral lipids and phospholipids that cover more than 80% of the chromatographic peaks.

DOI: https://doi.org/10.1021/acs.analchem.4c03230
J Chromatogr A. 2024 Oct 06. pii: S0021-9673(24)00798-2. [Epub ahead of print]1736 465424

Comprehensive characterization of active components in Salvia miltiorrhiza using polarity-partitioned two-dimensional liquid chromatography coupled with mass spectrometry.

Liping Zhang, Yuye Cai, Qisheng Zhong, Shijing Zhang, Lingling Shen, Taohong Huang, Ting Zhou.

  Salvia miltiorrhiza, a widely used traditional Chinese medicine, contains a complex matrix of hydrophobic diterpenoids and hydrophilic phenolic acids, presenting significant challenges in comprehensive analysis. In this study, an online polarity-partitioned two-dimensional liquid chromatography coupled with mass spectrometry (2D-LC-MS) method was developed for comprehensive analysis of both lipophilic and hydrophilic active components in Salvia miltiorrhiza. The method integrated hydrophilic interaction liquid chromatography (HILIC) and reversed-phase liquid chromatography (RPLC), facilitating the efficient separation of compounds across a wide range of polarities. An online dilution strategy was implemented, minimizing sample loss and enhancing the method's utility for quality control and chemical characterization of complex herbal matrices. Compared with other LC methods, this approach significantly improved analyte coverage, resolution, and analysis efficiency. Under optimal conditions, 150 active components were successfully identified, including 33 compounds newly discovered in Salvia miltiorrhiza. Additionally, the validated online method was applied to the quantitative determination of 16 quality markers of Salvia miltiorrhiza from different sources. The results demonstrated the online method's potential as a superior alternative to existing techniques, offering broader applicability in traditional Chinese medicine research.

Keywords:  Active components; Danshen; Online two-dimensional liquid chromatography; Polarity-partition; Salvia miltiorrhiza

DOI:  https://doi.org/10.1016/j.chroma.2024.465424
Environ Sci Technol. 2024 Oct 16.

Two-Dimensional Liquid Chromatography Tandem Mass Spectrometry Untangles the Deep Metabolome of Marine Dissolved Organic Matter.

Stilianos Papadopoulos Lambidis, Tilman Schramm, Karoline Steuer-Lodd, Shane Farrell, Paolo Stincone, Robin Schmid, Irina Koester, Ralph Torres, Thorsten Dittmar, Lihini Aluwihare, Carsten Simon, Daniel Petras.

  Dissolved organic matter (DOM) is an ultracomplex mixture that plays a central role in global biogeochemical cycles. Despite its importance, DOM remains poorly understood at the molecular level. Over the last decades, significant efforts have been made to decipher the chemical composition of DOM by high-resolution mass spectrometry (HR-MS) and liquid chromatography (LC) coupled with tandem mass spectrometry (MS/MS). Yet, the complexity and high degree of nonresolved isomers still hamper the full structural analysis of DOM. To address this challenge, we developed an offline two-dimensional (2D) LC approach using two reversed-phase dimensions with orthogonal pH levels, followed by MS/MS data acquisition and molecular networking. 2D-LC-MS/MS reduced the complexity of DOM, enhancing the quality of MS/MS spectra and increasing spectral annotation rates. Applying our approach to analyze coastal-surface DOM from Southern California (USA) and open-ocean DOM from the central North Pacific (Hawaii), we annotated in total more than 600 structures via MS/MS spectrum matching, which was up to 90% more than that in iterative 1D LC-MS/MS analysis with the same total run time. Our data offer unprecedented insights into the molecular composition of marine DOM and highlight the potential of 2D-LC-MS/MS approaches to decipher the chemical composition of ultracomplex samples.

Keywords:  2D chromatography; 2D-LC-MS/MS; dissolved organic matter; environmental metabolomics; molecular networking; tandem mass spectrometry

DOI:  https://doi.org/10.1021/acs.est.4c07173
J Chromatogr B Analyt Technol Biomed Life Sci. 2024 Oct 09. pii: S1570-0232(24)00346-5. [Epub ahead of print]1247 124337

Quantitative determination by UHPLC-MS/MS of 18 common drugs of abuse and metabolites, including THC and OH-THC, in volumetric dried blood spots: a sustainable method with minimally invasive sampling.

Christina Ververi, Claudia Gentile, Marta Massano, Alberto Salomone, Marco Vincenti.

  The increased use of drugs of abuse urges forensic toxicologists to create quick, simple, minimally invasive sampling techniques for biological fluids combined with analytical methods assuring accurate results. To this purpose, a method was developed aimed at quantifying 18 drugs of abuse and metabolites in DBS. Validation of the method was conducted by spiking blank whole blood with the analytes on Capitainer® B cards. The extracts were analyzed by a targeted UHPLC-MS/MS method. Linear calibration was achieved in the range of 1-100 ng/mL for: amphetamine, MDA, MDMA, methamphetamine, cocaine, codeine, benzoylecgonine, cocaethylene, morphine, 6-MAM, buprenorphine, methadone, EDDP, ketamine, norbuprenorphine norketamine, THC, and OH-THC. Experimental LOD was found at 0.5 ng/mL for all analytes except for norbuprenorphine, THC and THC-OH which yielded a LOD of 1 ng/mL. Intra- and inter-day accuracy was satisfactory with bias% resulting within 5%. Evaluation of intra- and inter-day precision yielded CV% values within 20%, for all compounds except EDDP. Average extraction recovery calculated at low (2 ng/mL) and high (75 ng/mL) concentration levels was 63% while average matrix effect determined at the same levels was found to be within 85% - 115% for all analytes except from codeine (70%) and MDMA (131%). The method was applied to authentic blood samples spotted onto the DBS card and the minimum value detected was 1.3 ng/mL. HPLC-MS/MS proved capable to identify all the targeted analytes at low concentrations in the small blood volumes obtainable from DBS cards, which in turn confirmed to be effective and sustainable micro-sampling devices.

Keywords:  DBS; Drugs of abuse; HPLC–MS/MS; THC

DOI:  https://doi.org/10.1016/j.jchromb.2024.124337