bims-metlip 2023-07-30 papers

bims-metlip

Biomed News

on Methods and protocols in metabolomics and lipidomics

Issue of 2023–07–30
twenty-one papers selected by
Sofia Costa, Matterworks

Quantitative Analytical and Computational Workflow for Large-Scale Targeted Plasma Metabolomics.
LC-MS/MS-MS3 for the Determination and Quantification of Δ9-Tetrahydrocannabinol and Metabolites in Blood Samples.
Ion-to-Image, i2i, a Mass Spectrometry Imaging Data Analysis Platform for Continuous Ionization Techniques.
A Structure-Guided Molecular Network Strategy for Global Untargeted Metabolomics Data Annotation.
Determination of cortisone and cortisol in human scalp hair using an improved LC-MS/MS-based method.
A simple and sensitive LC-MS/MS method for the quantitation of sertraline and N-desmethylsertraline in human plasma.
ipaPy2: Integrated Probabilistic Annotation (IPA) 2.0-an improved Bayesian-based method for the annotation of LC-MS/MS untargeted metabolomics data.
An eco-friendly sample preparation procedure based on air-assisted liquid-liquid microextraction for the rapid determination of phthalate metabolites in urine samples by liquid chromatography-tandem mass spectrometry.
A Mass Spectrometry Database for Sea Cucumber Triterpene Glycosides.
Using HPLC for the determination of platinum drugs in biological matrixes after derivatization with diethyldithiocarbamate.
Simultaneous determination of water-soluble herbicides using hydrophilic interaction liquid chromatography-mass spectrometry.
Fast and Sensitive Bioanalytical Method for the Determination of Deucravacitinib in Human Plasma Using HPLC-MS/MS: Application and Greenness Evaluation.
Comparison of Compound Identification Tools Using Data Dependent and Data Independent High-Resolution Mass Spectrometry Spectra.
NanoLC-EI-MS: Perspectives in Biochemical Analysis.
Improved separation of fentanyl isomers using metal cation adducts and high-resolution ion mobility-mass spectrometry.
A Reliable Method Based on Liquid Chromatography-Tandem Mass Spectrometry for the Simultaneous Quantification of Neurotransmitters in Caenorhabditis elegans.
Flow-injection MS analysis as a simplified approach to pesticide screening in apples.
Advancements in Analyzing Tumor Metabolites through Chemical Derivatization-Based Chromatography.
Development and validation for bioanalysis of VK2809, its active metabolite VK2809A and glutathione-conjugated metabolite MB06588 in rat liver using LC-MS/MS.
Application to several suspected poisoning cases of a validated analytical method for the determination of muscarine in human biological matrices using liquid chromatography with high-resolution mass spectrometry detection.
HiSorb sorptive extraction for determining salivary short chain fatty acids and hydroxy acids in heart failure patients.

Metabolites. 2023 Jul 13. pii: 844. [Epub ahead of print]13(7):

Quantitative Analytical and Computational Workflow for Large-Scale Targeted Plasma Metabolomics.

Antonia Fecke, Nay Min Min Thaw Saw, Dipali Kale, Siva Swapna Kasarla, Albert Sickmann, Prasad Phapale.

  Quantifying metabolites from various biological samples is necessary for the clinical and biomedical translation of metabolomics research. One of the ongoing challenges in biomedical metabolomics studies is the large-scale quantification of targeted metabolites, mainly due to the complexity of biological sample matrices. Furthermore, in LC-MS analysis, the response of compounds is influenced by their physicochemical properties, chromatographic conditions, eluent composition, sample preparation, type of MS ionization source, and analyzer used. To facilitate large-scale metabolite quantification, we evaluated the relative response factor (RRF) approach combined with an integrated analytical and computational workflow. This approach considers a compound's individual response in LC-MS analysis relative to that of a non-endogenous reference compound to correct matrix effects. We created a quantitative LC-MS library using the Skyline/Panorama web platform for data processing and public sharing of data. In this study, we developed and validated a metabolomics method for over 280 standard metabolites and quantified over 90 metabolites. The RRF quantification was validated and compared with conventional external calibration approaches as well as literature reports. The Skyline software environment was adapted for processing such metabolomics data, and the results are shared as a "quantitative chromatogram library" with the Panorama web application. This new workflow was found to be suitable for large-scale quantification of metabolites in human plasma samples. In conclusion, we report a novel quantitative chromatogram library with a targeted data analysis workflow for biomedical metabolomic applications.

Keywords:  LC-MS; metabolite quantification; metabolomics; quantitative spectral library; relative response factor

DOI:  https://doi.org/10.3390/metabo13070844
J Anal Toxicol. 2023 Jul 26. pii: bkad046. [Epub ahead of print]

LC-MS/MS-MS3 for the Determination and Quantification of Δ9-Tetrahydrocannabinol and Metabolites in Blood Samples.

Paula Proença, Helena M Teixeira, Beatriz Martinho, Carla Monteiro, João Franco, Francisco Corte-Real.

  Due to the high prevalence of cannabinoids in forensic toxicology analysis, it is crucial to have an efficient method that allows the use of a small sample amount and that requires a minimal sample preparation, for the determination and quantification of low concentrations. A simple, highly selective and high throughput liquid chromatography tandem mass spectrometry methodology (LC-MS/MS-MS3) was developed for the determination and quantification of ∆9-tetrahydrocannabinol (THC), 11-hydroxy-∆9- tetrahydrocannabinol (THC-OH) and 11-nor-9-carboxy-∆9-tetrahydrocannabinol (THC-COOH), in blood samples. Chromatographic analysis of THC, THC-OH, and THC-COOH and their deuterated internal standards was preceded by protein precipitation (PPT) of 0.1 mL of blood samples with acetonitrile. Chromatographic separation was achieved by use of an Acquity UPLC® HHS T3 (100 mm x 2.1mm i.d., 1.8 μm) reversed-phase column, using a gradient elution of 2 mM aqueous ammonium formate, 0.1% formic acid, and methanol at a flow rate of 0.4 mL/min, with a run time of 10 minutes. For the MS/MS-MS3 analysis, a SCIEX QTRAP® 6500+ triple quadrupole linear ion trap mass spectrometer was used via electrospray ionization (ESI), operated in multiple reaction monitoring (MRM) and linear ion trap mode (MS3). The method was validated in accordance with internationally accepted criteria and guidelines, and proved to be selective and linear between 0.5-100 ng/mL (r2>0.995). The lower limits of quantification (LLOQ) corresponded to the lowest concentrations used for the calibration curves. The coefficients of variation obtained for accuracy and precision were <15%. The mean recoveries were between 88.0-117.2%, for the studied concentration levels (1 ng/mL, 5 ng/mL and 50 ng/mL). No significant interfering compounds, matrix effects or carryover were observed. The validated method provides a sensitive, efficient and robust procedure for the quantification of cannabinoids in blood, using LC-MS/MS-MS3 and a sample volume of 0.1 mL. This work is also a proof of concept for using LC-MS3 technique to determine drugs in biological samples.

Keywords:  LC-MS/MS-MS3; cannabinoids; protein precipitation; whole blood

DOI:  https://doi.org/10.1093/jat/bkad046
Anal Chem. 2023 Jul 28.

Ion-to-Image, i2i, a Mass Spectrometry Imaging Data Analysis Platform for Continuous Ionization Techniques.

Johan Lillja, Kyle D Duncan, Ingela Lanekoff.

Mass spectrometry imaging (MSI) techniques generate data that reveal spatial distributions of molecules on a surface with high sensitivity and selectivity. However, processing large volumes of mass spectrometry data into useful ion images is not trivial. Furthermore, data from MSI techniques using continuous ionization sources where data are acquired in line scans require different data handling strategies compared to data collected from pulsed ionization sources where data are acquired in grids. In addition, for continuous ionization sources, the pixel dimensions are influenced by the mass spectrometer duty cycle, which, in turn, can be controlled by the automatic gain control (AGC) for each spectrum (pixel). Currently, there is a lack of data-handling software for MSI data generated with continuous ionization sources and AGC. Here, we present ion-to-image (i2i), which is a MATLAB-based application for MSI data acquired with continuous ionization sources, AGC, high resolution, and one or several scan filters. The source code and a compiled installer are available at https://github.com/LanekoffLab/i2i. The application includes both quantitative, targeted, and nontargeted data processing strategies and enables complex data sets to be processed in minutes. The i2i application has high flexibility for generating, processing, and exporting MSI data both from simple full scans and more complex scan functions interlacing MSn and SIM scan data sets, and we anticipate that it will become a valuable addition to the existing MSI software toolbox.

DOI: https://doi.org/10.1021/acs.analchem.3c01615
Anal Chem. 2023 Jul 26.

A Structure-Guided Molecular Network Strategy for Global Untargeted Metabolomics Data Annotation.

Xinxin Wang, Chao Li, Zaifang Li, Yanpeng Qi, Xiuqiong Zhang, Xinjie Zhao, Chunxia Zhao, Xiaohui Lin, Xin Lu, Guowang Xu.

Large-scale metabolite annotation is a bottleneck in untargeted metabolomics. Here, we present a structure-guided molecular network strategy (SGMNS) for deep annotation of untargeted ultra-performance liquid chromatography-high resolution mass spectrometry (MS) metabolomics data. Different from the current network-based metabolite annotation method, SGMNS is based on a global connectivity molecular network (GCMN), which was constructed by molecular fingerprint similarity of chemical structures in metabolome databases. Neighbor metabolites with similar structures in GCMN are expected to produce similar spectra. Network annotation propagation of SGMNS is performed using known metabolites as seeds. The experimental MS/MS spectra of seeds are assigned to corresponding neighbor metabolites in GCMN as their "pseudo" spectra; the propagation is done by searching predicted retention times, MS1, and "pseudo" spectra against metabolite features in untargeted metabolomics data. Then, the annotated metabolite features were used as new seeds for annotation propagation again. Performance evaluation of SGMNS showed its unique advantages for metabolome annotation. The developed method was applied to annotate six typical biological samples; a total of 701, 1557, 1147, 1095, 1237, and 2041 metabolites were annotated from the cell, feces, plasma (NIST SRM 1950), tissue, urine, and their pooled sample, respectively, and the annotation accuracy was >83% with RSD <2%. The results show that SGMNS fully exploits the chemical space of the existing metabolomes for metabolite deep annotation and overcomes the shortcoming of insufficient reference MS/MS spectra.

DOI: https://doi.org/10.1021/acs.analchem.3c00849
Clin Chem Lab Med. 2023 Jul 25.

Determination of cortisone and cortisol in human scalp hair using an improved LC-MS/MS-based method.

Mina Mirzaian, Sofie K M van Zundert, Wim F Schilleman, Mostafa Mohseni, Susanne Kuckuck, Elisabeth F C van Rossum, Ron H N van Schaik, Sjoerd A A van den Berg.

   OBJECTIVES: Human scalp hair is an easily available but complex matrix for determination of cortisone and cortisol, and has been shown to reflect long-term glucocorticoid exposure. Hair glucocorticoid analysis has been used to detect hypo- and hypercortisolism. In this study, we describe the development and validation of a LC-MS/MS method for quantification of cortisone and cortisol in human scalp hair, and provide a novel approach for analysis and interpretation of the results.
METHODS: Improved sample preparation using pulverization and solid phase extraction allowed for low sample volumes (10 mg). Baseline chromatographic separation without matrix interference was achieved by reversed phase chromatography and MRM measurement in negative ion mode. Run-to-run time was 8 min. Mixed model analyses were performed to create individual patterns of cortisone and cortisol concentrations.
RESULTS: Matrix matched calibration curves showed excellent linearity up to 100 pg (analyte)/mg (hair) for both cortisone and cortisol (R2>0.995). LLOQ was 1.5 and 1.0 pg/mg for cortisone and cortisol, respectively. Matrix effect was negligible for hair color (recoveries 95-105 %). Cortisone and cortisol concentrations decreased from proximal to distal hair segments, following a predictable, but subject-specific pattern, with less individual variation for cortisone than for cortisol.
CONCLUSIONS: This improved LC-MS/MS method is able to accurately quantify cortisone and cortisol in human hair with minimum matrix interference. This new way of data analysis and interpretation including individual patterns of cortisone and cortisol will be of help with detection of pathological concentrations in both the high - and the low ranges of glucocorticoids.

Keywords:  cortisol; cortisone; human hair; liquid chromatography; mass spectrometry

DOI:  https://doi.org/10.1515/cclm-2023-0341
J Chromatogr B Analyt Technol Biomed Life Sci. 2023 Jul 19. pii: S1570-0232(23)00237-4. [Epub ahead of print]1228 123827

A simple and sensitive LC-MS/MS method for the quantitation of sertraline and N-desmethylsertraline in human plasma.

Cassius M Phogole, Roxanne Hastie, Tracy Kellermann.

  A simple and sensitive LC-MS/MS method for the simultaneous quantification of the second-generation antidepressant sertraline, and its active metabolite, N-desmethylsertraline, in human plasma was developed and validated. The analytes were extracted from 200 µL human plasma using a simple protein precipitation method. A gradient elution mode of water and 0.1% formic acid and acetonitrile was used for chromatographic separation on a Poroshell EC-C18 column (3 × 100 mm, 2.7 µm) at a flow rate of 0.450 mL/min. MS/MS analysis was performed in positive ionization mode using the transitions of m/z 306.1 → 159.1, 309.1 → 275.2, 292.1 → 159.1, and 296.2 → 279.0 for sertraline, sertraline-d3, N-desmethylsertraline, and N-desmethylsertraline-d4, respectively. The calibration curves for sertraline and N-desmethylsertraline in human plasma ranged from 2.50-320 ng/mL and 10.0-1280 ng/mL, respectively, with correlation coefficients (r) of ≥ 0.9992. The intra- and inter-assay precisions for both analytes at four concentrations (LLOQ, QCL, QCM, and QCH) ranged between 2.2% and 12.2%, respectively, while their accuracies ranged between 92.0 and 111.7%, with the exception of LLOQ, which ranged between 84.3 and 106.0%. The mean percentage recovery and process efficiency at three concentrations (QCL, QCM, and QCH) was 94.2 and 87.9% for sertraline, and 95.7 and 95.2% for N-desmethylsertraline, respectively. Both analytes were stable in plasma at room temperature for 2 h, at -80 °C for 28 days and through three freeze/thaw cycles. This method was successfully applied to a clinical study investigating the use of sertraline during pregnancy.

Keywords:  And pregnancy; Clinical study; Human plasma; LC-MS/MS; N-desmethylsertraline; Sertraline

DOI:  https://doi.org/10.1016/j.jchromb.2023.123827
Bioinformatics. 2023 Jul 01. pii: btad455. [Epub ahead of print]39(7):

ipaPy2: Integrated Probabilistic Annotation (IPA) 2.0-an improved Bayesian-based method for the annotation of LC-MS/MS untargeted metabolomics data.

Francesco Del Carratore, William Eagles, Juraj Borka, Rainer Breitling.

SUMMARY: The Integrated Probabilistic Annotation (IPA) is an automated annotation method for LC-MS-based untargeted metabolomics experiments that provides statistically rigorous estimates of the probabilities associated with each annotation. Here, we introduce ipaPy2, a substantially improved and completely refactored Python implementation of the IPA method. The revised method is now able to integrate tandem MS fragmentation data, which increases the accuracy of the identifications. Moreover, ipaPy2 provides a much more user-friendly interface, and isotope peaks are no longer treated as individual features but integrated into isotope fingerprints, greatly speeding up the calculations. The method has also been fully integrated with the mzMatch pipeline, so that the results of the annotation can be explored through the newly developed PeakMLViewerPy tool available at https://github.com/UoMMIB/PeakMLViewerPy.
AVAILABILITY AND IMPLEMENTATION: The source code, extensive documentation, and tutorials are freely available on GitHub at https://github.com/francescodc87/ipaPy2.

DOI: https://doi.org/10.1093/bioinformatics/btad455
Talanta. 2023 Jul 20. pii: S0039-9140(23)00725-7. [Epub ahead of print]266(Pt 1): 124974

An eco-friendly sample preparation procedure based on air-assisted liquid-liquid microextraction for the rapid determination of phthalate metabolites in urine samples by liquid chromatography-tandem mass spectrometry.

Bruno Alves Rocha, Matheus Gallimberti, João Paulo Bianchi Ximenez, Carla Giovana Basso, Anderson Joel Martino-Andrade, Holger Martin Koch, Leandro Augusto Calixto, Fernando Barbosa.

  Urinary phthalate metabolite (mPAEs) analysis is a reliable tool for assessing human exposure to phthalates. With growing interest in urinary biomonitoring of these metabolites, there is a need for fast and sensitive analytical methods. Therefore, a simple, rapid procedure for simultaneous determination of fifteen phthalate metabolites in human urine samples by liquid chromatography-tandem mass spectrometry was developed. The novelty of the present procedure is based on the use of diethyl carbonate as a green biobased extraction solvent and air-assisted liquid-liquid microextraction (AALLME) as a sample preparation step. A Plackett-Burman design was used for screening the factors that influence the AALLME extraction efficiency of mPAEs. The effective factors were then optimized by response surface methodology using a central composite rotatable design. Under the optimized conditions, good linearity can be achieved in a concentration range of 1.0-20.0 ng mL-1 with correlation coefficients higher than 0.99. The repeatability and reproducibility precision were in the range of 2-12% and 1-10% respectively. Recoveries ranging from 90% to 110%. This, and the low limits of detection, ranging from 0.01 to 0.05 ng mL-1, make the proposed procedure sensitive and suitable for human biomonitoring of phthalate exposures. For proof-of-principle, the new method was used to measure the urinary concentrations of mPAEs in 20 urine samples from Brazilian women. The high frequency of detections and in part high concentrations of mPAEs indicate to widespread exposure to several phthalates among Brazilian women.

Keywords:  Air-assisted liquid-liquid microextraction; Diethyl carbonate; Green solvent; Human biomonitoring; Liquid chromatography-tandem mass spectrometry; Phthalate metabolites

DOI:  https://doi.org/10.1016/j.talanta.2023.124974
Metabolites. 2023 Jun 23. pii: 783. [Epub ahead of print]13(7):

A Mass Spectrometry Database for Sea Cucumber Triterpene Glycosides.

Roman S Popov, Natalia V Ivanchina, Alexandra S Silchenko, Sergey A Avilov, Vladimir I Kalinin, Timofey V Malyarenko, Valentin A Stonik, Pavel S Dmitrenok.

  Sea cucumber triterpene glycosides are a class of secondary metabolites that possess distinctive chemical structures and exhibit a variety of biological and pharmacological activities. The application of MS-based approaches for the study of triterpene glycosides allows rapid evaluation of the structural diversity of metabolites in complex mixtures. However, the identification of the detected triterpene glycosides can be challenging. The objective of this study is to establish the first spectral library containing the mass spectra of sea cucumber triterpene glycosides using ultraperformance liquid chromatography-quadrupole time-of-flight mass spectrometry. The library contains the electrospray ionization tandem mass spectra and retention times of 191 triterpene glycosides previously isolated from 15 sea cucumber species and one starfish at the Laboratory of the Chemistry of Marine Natural Products of the G.B. Elyakov Pacific Institute of Bioorganic Chemistry. In addition, the chromatographic behavior and some structure-related neutral losses in tandem MS are discussed. The obtained data will accelerate the accurate dereplication of known triterpene glycosides and the annotation of novel compounds, as we demonstrated by the processing of LC-MS/MS data of Eupentacta fraudatrix extract.

Keywords:  Eupentacta fraudatrix; mass spectrometry; mass spectrometry database; metabolite annotation and identification; sea cucumber; triterpene glycosides; ultraperformance liquid chromatography

DOI:  https://doi.org/10.3390/metabo13070783
J Sep Sci. 2023 Jul 28. e2300392

Using HPLC for the determination of platinum drugs in biological matrixes after derivatization with diethyldithiocarbamate.

Dorota Turoňová, Lenka Kujovská Krčmová, Milan Vošmik, Bohuslav Melichar, František Švec.

  Challenges and pitfalls in the application of diethyldithiocarbamate derivatization for LC analysis of cisplatin and oxaliplatin, as well as the suitability of this method for different biological matrices with implications for use in routine practice have been identified. The LC of platinum drugs presents a significant challenge. They are polar compounds with poor retention on reverse phase packings. Cisplatin also exhibits poor absorption in UV and ionization in mass spectrometry. Therefore, we developed and optimized a derivatization approach for the LC analysis of total platinum in plasma, plasma ultrafiltrate, peritoneal fluid, and urine. Derivatization in urine proved to be difficult due to the complexity of the matrix, and extended testing was required. Our results highlight the important issues affecting the efficiency, reliability, and suitability of platinum drug derivatization. Although precolumn derivatization is less selective than its postcolumn counterpart, the application of precolumn derivatization is a simple, rapid, and universal approach for the determination of platinum drugs by HPLC. One of its major advantages is that it allows a more affordable analysis using UV detection without the need for additional high-end instrumentation such as a MS detector.

Keywords:  derivatization; diethyldithiocarbamate; peritoneal fluid; plasma; platinum drugs

DOI:  https://doi.org/10.1002/jssc.202300392
Forensic Toxicol. 2023 Jul 22.

Simultaneous determination of water-soluble herbicides using hydrophilic interaction liquid chromatography-mass spectrometry.

Daisuke Watanabe, Shuhei Sonoda, Hikoto Ohta.

   PURPOSE: The analysis of water-soluble herbicides, including glyphosate (Glyp), glufosinate (Gluf), paraquat (PQ), and diquat (DQ), is time-consuming and expensive because they cannot be analyzed using general toxicological screening methods. Thus, this study aimed to develop a simple and rapid method to simultaneously analyze these compounds without any derivatization nor ion-pairing reagents.
METHODS: The analytes were separated using hydrophilic interaction liquid chromatography and detected using tandem mass spectrometry. The developed method was applied to plant and biological samples assuming criminal damage and poisoning cases, respectively.
RESULTS: All analytes were separated well and detected with good peak shapes. For plant samples, the herbicides were specifically detected from withered leaves using a simple extraction method. For biological samples, quantitative analysis was successfully validated, and the limit of quantification values of Glyp and Gluf were 0.2 µg/mL, and those of PQ and DQ were 1 ng/mL.
CONCLUSION: The developed method had sufficient performance for practical forensic applications including poisoning cases and malicious uses to damage commercial crops.

Keywords:  Diquat; Glufosinate; Glyphosate; HILIC analysis; Paraquat

DOI:  https://doi.org/10.1007/s11419-023-00669-7
Molecules. 2023 Jul 17. pii: 5471. [Epub ahead of print]28(14):

Fast and Sensitive Bioanalytical Method for the Determination of Deucravacitinib in Human Plasma Using HPLC-MS/MS: Application and Greenness Evaluation.

Pottabattula Mahesh, M Akiful Haque, Baher I Salman, Tarek S Belal, Adel Ehab Ibrahim, Sami El Deeb.

  Plaque psoriasis is a common, long-lasting illness that affects the immune system and causes significant negative impacts on a patient's physical health, well-being, and ability to work effectively. Deucravacitinib (DEU) is the first oral medication used in the treatment of plaque psoriasis, a chronic skin condition that causes red, scaly patches on the skin. DEU is a type of medication called an oral Janus kinase (JAK) inhibitor, which works by blocking specific enzymes that play a role in the inflammation and immune response associated with psoriasis. Therefore, a quick, easy, novel, reliable, sensitive, and straightforward liquid chromatography-tandem mass spectrometry (LC-MS/MS) approach was used to analyze DEU in plasma samples. The LC-MS/MS method for the determination of DEU in human plasma was based on using trimethoprim as an internal standard (IS). The separation of DEU and IS was carried out via liquid-liquid extraction (LLE). The extract was then subjected to the chromatographic system separation using the ACE-C18 column (4.6 × 100 mm, 5 µm). The mobile phase employed consisted of methanol and a solution of 2 mM ammonium formate (80:20 v/v, respectively). The flow rate used was set at 0.9 mL min-1. The creative strategy was performed by running an ABSCIEX API 4000 mass spectrometer with an electron spray ionization source in multiple reaction monitoring (MRM) mode. The ion transitions m/z 426.3 → 358.2 were used for DEU quantitation, while the ion transitions m/z 291.1 → 261.1 were used for trimethoprim quantitation. The accuracy, precision, linearity, recovery, and selectivity of DEU were deemed acceptable when validated for a concentration range between 0.500 and 601.050 ng/mL, utilizing a weighting factor of 1/x2.

Keywords:  HPLC-MS/MS; deucravacitinib; human plasma; psoriasis

DOI:  https://doi.org/10.3390/molecules28145471
Metabolites. 2023 Jun 21. pii: 777. [Epub ahead of print]13(7):

Comparison of Compound Identification Tools Using Data Dependent and Data Independent High-Resolution Mass Spectrometry Spectra.

Rosalie Nijssen, Marco H Blokland, Robin S Wegh, Erik de Lange, Stefan P J van Leeuwen, Bjorn J A Berendsen, Milou G M van de Schans.

  Liquid chromatography combined with high-resolution mass spectrometry (LC-HRMS) is a frequently applied technique for suspect screening (SS) and non-target screening (NTS) in metabolomics and environmental toxicology. However, correctly identifying compounds based on SS or NTS approaches remains challenging, especially when using data-independent acquisition (DIA). This study assessed the performance of four HRMS-spectra identification tools to annotate in-house generated data-dependent acquisition (DDA) and DIA HRMS spectra of 32 pesticides, veterinary drugs, and their metabolites. The identification tools were challenged with a diversity of compounds, including isomeric compounds. The identification power was evaluated in solvent standards and spiked feed extract. In DDA spectra, the mass spectral library mzCloud provided the highest success rate, with 84% and 88% of the compounds correctly identified in the top three in solvent standard and spiked feed extract, respectively. The in silico tools MSfinder, CFM-ID, and Chemdistiller also performed well in DDA data, with identification success rates above 75% for both solvent standard and spiked feed extract. MSfinder provided the highest identification success rates using DIA spectra with 72% and 75% (solvent standard and spiked feed extract, respectively), and CFM-ID performed almost similarly in solvent standard and slightly less in spiked feed extract (72% and 63%). The identification success rates for Chemdistiller (66% and 38%) and mzCloud (66% and 31%) were lower, especially in spiked feed extract. The difference in success rates between DDA and DIA is most likely caused by the higher complexity of the DIA spectra, making direct spectral matching more complex. However, this study demonstrates that DIA spectra can be used for compound annotation in certain software tools, although the success rate is lower than for DDA spectra.

Keywords:  annotation; data dependent acquisition; data independent acquisition; high-resolution mass spectrometry; identification; metabolites; pesticides; veterinary drugs

DOI:  https://doi.org/10.3390/metabo13070777
Int J Mol Sci. 2023 Jul 21. pii: 11746. [Epub ahead of print]24(14):

NanoLC-EI-MS: Perspectives in Biochemical Analysis.

Natalia Gabrielly Pereira Dos Santos, Edvaldo Vasconcelos Soares Maciel, Deyber Arley Vargas Medina, Fernando Mauro Lanças.

  Although LC-MS with atmospheric pressure ionization (API) sources is the primary technique used in modern bioanalytical studies, electron ionization mass spectrometry (EI-MS) can provide some substantial advantages over it. EI-MS is a matrix effect-free technique that provides reproducible and comparable mass spectra, serving as a compound fingerprint for easy identification through automated comparison with spectral libraries. Leveraging EI-MS in biochemical studies can yield critical analytical benefits for targeted and untargeted analyses. However, to fully utilize EI-MS for heavy and non-volatile molecules, a new technology that enables the coupling of liquid chromatography with EI-MS is needed. Recent advancements in nanoLC have addressed the compatibility issues between LC and EI-MS, and innovative interfacing strategies such as Direct-EI, liquid electron ionization (LEI), and Cold-EI have extended the application of EI-MS beyond the determination of volatile organic molecules. This review provides an overview of the latest developments in nanoLC-EI-MS interfacing technologies, discussing their scope and limitations. Additionally, selected examples of nanoLC-EI-MS applications in the field of biochemical analysis are presented, highlighting the potential prospects and benefits that the establishment of this technique can bring to this field.

Keywords:  biochemical analysis; electron ionization; mass spectrometry; metabolomics; nano-liquid chromatography

DOI:  https://doi.org/10.3390/ijms241411746
Drug Test Anal. 2023 Jul 25.

Improved separation of fentanyl isomers using metal cation adducts and high-resolution ion mobility-mass spectrometry.

Ralph Aderorho, Christopher D Chouinard.

  Fentanyl is a potent synthetic opioid that has attracted significant attention due to its illegal production and distribution, resulting in misuse, overdose, and fatalities. Because numerous fentanyl analogs, including structural isomers, with different potency have been discovered in the field, there is a critical need to continue developing analytical methodologies capable of accurate identification in forensic and clinical laboratories. This study aimed to develop a rapid method for detecting and separating fentanyl isomers based on ion mobility-mass spectrometry (IM-MS), where IM separates gas-phase ions based on differences in their size, shape, and charge. Several strategies for improved differentiation were implemented, including using unconventional cation adducts (e.g., alkali and transition metals) and data post-processing by high-resolution demultiplexing. A collection of collision cross section (CCS) values for the various metal ion adducts was gathered, which can be used to improve confidence of identification in future samples. Notable examples, such as [M + Cu]+ and [M + Ag]+ adducts, contributed to significant improvement of resolution between isomers. Furthermore, the addition of high-resolution post-processing provided resolving power of >150, which constitutes a significant increase in comparison with the normal 50-60 obtained with low-resolution drift tube instruments. Collectively, these improved separation strategies allowed for confident detection and subsequent quantitative analysis. The optimized IM-MS method resulted in quantification of fentanyl in human urine with limits of detection and quantification of 13 pg/mL and 40 pg/mL, respectively.

Keywords:  fentanyl; ion mobility-mass spectrometry (IM-MS); isomers

DOI:  https://doi.org/10.1002/dta.3550
Molecules. 2023 Jul 13. pii: 5373. [Epub ahead of print]28(14):

A Reliable Method Based on Liquid Chromatography-Tandem Mass Spectrometry for the Simultaneous Quantification of Neurotransmitters in Caenorhabditis elegans.

Ann-Kathrin Weishaupt, Laura Kubens, Lysann Ruecker, Tanja Schwerdtle, Michael Aschner, Julia Bornhorst.

  Neurotransmitters like dopamine (DA), serotonin (SRT), γ-aminobutyric acid (GABA) and acetylcholine (ACh) are messenger molecules that play a pivotal role in transmitting excitation between neurons across chemical synapses, thus enabling complex processes in the central nervous system (CNS). Balance in neurotransmitter homeostasis is essential, and altered neurotransmitter levels are associated with various neurological disorders, e.g., loss of dopaminergic neurons (Parkinson's disease) or altered ACh synthesis (Alzheimer's disease). Therefore, it is crucial to possess adequate tools to assess precise neurotransmitter levels, and to apply targeted therapies. An established in vivo model to study neurotoxicity is the model organism Caenorhabditis elegans (C. elegans), as its neurons have been well characterized and functionally are analogous to mammals. We have developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method including a sample preparation assuring neurotransmitter stability, which allows a simultaneous neurotransmitter quantification of DA, SRT, GABA and ACh in C. elegans, but can easily be applied to other matrices. LC-MS/MS combined with isotope-labeled standards is the tool of choice, due to its otherwise unattainable sensitivity and specificity. Using C. elegans together with our analytically validated and verified method provides a powerful tool to evaluate mechanisms of neurotoxicity, and furthermore to identify possible therapeutic approaches.

Keywords:  C. elegans; liquid chromatography; mass spectrometry; neurodegenerative diseases; neurotransmitters

DOI:  https://doi.org/10.3390/molecules28145373
Food Addit Contam Part A Chem Anal Control Expo Risk Assess. 2023 Jul 28. 1-14

Flow-injection MS analysis as a simplified approach to pesticide screening in apples.

Darko Andjelkovic, Milica Brankovic.

  A fast and simplified method based on mass spectrometry analysis was developed for pesticide qualification in apple peel. The main feature of the method is flow-injection of the loop pre-defined sample volume directly to the MS source, with no chromatographic separation. Method performances were evaluated for five pesticides regularly used in apple orchard treatments. First, primary features of the method such as the loop injection dynamics, total analysis time, chronogramic peaks integrity and sensitivity were evaluated. Then the applicability of the method for qualitative and quantitative screening was assessed on citrate buffered QuEChERS cleaned-up apple peel samples. The developed method offers a possibility for simplified, more economic high throughput screening for pesticides in apples. The method is intended to be a tool for preliminary qualitative assessment of pesticides residues in fruit. Quantitative features of the method are analyte and sample preparation dependent, most certainly due to the lack of chromatography.

Keywords:  Loop injection; QuEChERS; full scan; mass spectrometry; qualitative analysis

DOI:  https://doi.org/10.1080/19440049.2023.2238842
J Chromatogr A. 2023 Jul 22. pii: S0021-9673(23)00461-2. [Epub ahead of print]1706 464236

Advancements in Analyzing Tumor Metabolites through Chemical Derivatization-Based Chromatography.

Lu Ye, Hua-Min Zhang, Bing-Jun Zhou, Weiyang Tang, Jian-Liang Zhou.

  Understanding the metabolic abnormalities of tumors is crucial for early diagnosis, prognosis, and treatment. Accurate identification and quantification of metabolites in biological samples are essential to investigate the relationship between metabolite variations and tumor development. Common techniques like LC-MS and GC-MS face challenges in measuring aberrant metabolites in tumors due to their strong polarity, isomerism, or low ionization efficiency during MS detection. Chemical derivatization of metabolites offers an effective solution to overcome these challenges. This review focuses on the difficulties encountered in analyzing aberrant metabolites in tumors, the principles behind chemical derivatization methods, and the advancements in analyzing tumor metabolites using derivatization-based chromatography. It serves as a comprehensive reference for understanding the analysis and detection of tumor metabolites, particularly those that are highly polar and exhibit low ionization efficiency.

Keywords:  Chemical derivatization; Chromatography; Mass spectrometry; Tumor metabolites

DOI:  https://doi.org/10.1016/j.chroma.2023.464236
J Pharm Biomed Anal. 2023 Jul 21. pii: S0731-7085(23)00364-3. [Epub ahead of print]234 115595

Development and validation for bioanalysis of VK2809, its active metabolite VK2809A and glutathione-conjugated metabolite MB06588 in rat liver using LC-MS/MS.

Si-Tao Xu, Hao-Wen Jin, Xin Jin, Bi-Xin Xu, Yu Zhang, Tao Xie, Guangji Wang, Jiankun Wang, Le Zhen.

  VK2809 is a promising drug candidate in Phase II clinical trials for the treatment of non-alcoholic steatohepatitis (NASH). It is a prodrug with a HepDirect strategy, which can achieve selective hepatic metabolic activation, generating an active metabolite VK2809A as a potent and selective agonist for thyroid hormone receptor beta (TRβ), a concomitant reactive metabolite VK2809B, and a glutathione (GSH) conjugate MB06588. Currently, there is no convenient and sensitive bioanalytical method for the simultaneous determination of the above three metabolites. Herein, we established an LC-MS/MS method to separate VK2809 and its metabolites on the XSelect HSS T3 column and quantified them in negative electrospray ionization mode. Subsequently, several factors were investigated such as the use of 60% acetonitrile for homogenization to stabilize the analytes, the addition of 20 mM glutathione for the derivation of VK2809B, and the protein precipitation with methanol containing Sobetirome as the internal standard (IS). The method exhibited good linearity for all compounds (19.4-388.4 nM for VK2809; 27.4-2744.4 nM for VK2809A and 10.6-211.0 nM for MB06588) with great correlation coefficients (r > 0.996). The method validation also demonstrated acceptable precision (RSD < 13.0% for VK2809, RSD < 7.9% for VK2809A, RSD < 14.4% for MB06588) and accuracy (92.7%-103% for VK2809, 91.2%-107.3% for VK2809A, 96%-106.7% for MB06588). The matrix effect, recovery, and stability were also suitable to determine all the analytes. This method is suitable for the bioanalysis of VK2809 and its metabolites and has been successfully applied to the study of intrahepatic exposure in rats. It is expected to be further practiced in drug design, optimization, and metabolism study in the following research.

Keywords:  CYP 3A; Drug metabolite; HepDirect prodrugs; Nonalcoholic steatohepatitis; Pharmacokinetics; Thyroid hormone receptor agonist

DOI:  https://doi.org/10.1016/j.jpba.2023.115595
Drug Test Anal. 2023 Jul 24.

Application to several suspected poisoning cases of a validated analytical method for the determination of muscarine in human biological matrices using liquid chromatography with high-resolution mass spectrometry detection.

Estelle Flament, Jean-Michel Gaulier, Nathalie Paret, Nathalie Jarrier, Chloé Bruneau, Gaelle Creusat, Jérôme Guitton, Yvan Gaillard.

  Despite prevention efforts, many cases of mushroom poisoning are reported around the world every year. Among the different toxins implicated in these poisonings, muscarine may induce parasympathetic neurological damage. Muscarine poisonings are poorly reported in the current literature, implying a lack of available data on muscarine concentrations in human matrices. A validated liquid chromatography with high-resolution mass spectrometry detection (Orbitrap technology) method was developed to determine muscarine concentrations in human urine, plasma, and whole blood samples. Muscarine was determined using 100 μL of biological fluids, and precipitation was used for sample preparation. Liquid chromatography-mass spectrometry was performed using an Accucore Phenyl-X analytical column with the electrospray source in positive ion mode. Muscarine was quantitated in parallel reaction monitoring (PRM) mode with D9-muscarine as the internal standard. The method was validated successfully over the concentration range 0.1-100 μg/L for plasma and whole blood and 1-100 μg/L for urine, with acceptable precision and accuracy (<13.5%), including the lower limit of quantification. Ten real cases of suspected muscarine poisoning were successfully confirmed with this validated method. Muscarine concentrations in these cases ranged from 0.12 to 14 μg/L in whole blood, <LOQ to 43 μg/L in plasma, and <LOQ to 1537 μg/L in urine. This work provides a tool of choice for the diagnosis of muscarine poisoning when mushrooms are not available for identification, as well as the possibility of determining toxins in the blood, which is an important step toward understanding the pharmacokinetics of the mycotoxin.

Keywords:  blood; high-resolution mass spectrometry; muscarine; poisoning; urine

DOI:  https://doi.org/10.1002/dta.3542
J Chromatogr B Analyt Technol Biomed Life Sci. 2023 Jul 17. pii: S1570-0232(23)00236-2. [Epub ahead of print]1228 123826

HiSorb sorptive extraction for determining salivary short chain fatty acids and hydroxy acids in heart failure patients.

Alessio Lenzi, Denise Biagini, Silvia Ghimenti, Federico M Vivaldi, Pietro Salvo, Fabio Di Francesco, Tommaso Lomonaco.

  Variations in salivary short-chain fatty acids and hydroxy acids (e.g., lactic acid, and 3-hydroxybutyric acid) levels have been suggested to reflect the dysbiosis of human gut microbiota, which represents an additional factor involved in the onset of heart failure (HF) disease. The physical-chemical properties of these metabolites combined with the complex composition of biological matrices mean that sample pre-treatment procedures are almost unavoidable. This work describes a reliable, simple, and organic solvent free protocol for determining short-chain fatty acids and hydroxy acids in stimulated saliva samples collected from heart failure, obese, and hypertensive patients. The procedure is based on in-situ pentafluorobenzyl bromide (PFB-Br) derivatization and HiSorb sorptive extraction coupled to thermal desorption and gas chromatography-tandem mass spectrometry. The HiSorb extraction device is completely compatible with aqueous matrices, thus saving on time and materials associated with organic solvent-extraction methods. A Central Composite Face-Centred experimental design was used for the optimization of the molar ratio between PFB-Br and target analytes, the derivatization temperature, and the reaction time which were 100, 60 °C, and 180 min, respectively. Detection limits in the range 0.1-100 µM were reached using a small amount of saliva (20 µL). The use of sodium acetate-1-13C as an internal standard improved the intra- and inter-day precision of the method which ranged from 10 to 23%. The optimized protocol was successfully applied for what we believe is the first time to evaluate the salivary levels of short chain fatty acids and hydroxy acids in saliva samples of four groups of patients: i) patients admitted to hospital with acute HF symptoms, ii) patients with chronic HF symptoms, iii) patients without HF symptoms but with obesity, and iv) patients without HF symptoms but with hypertension. The first group of patients showed significantly higher levels of salivary acetic acid and lactic acid at hospital admission as well as the lowest values of hexanoic acid and heptanoic acid. Moreover, the significant high levels of acetic acid, propionic acid, and butyric acid observed in HF respect to the other patients suggest the potential link between oral bacteria and gut dysbiosis.

Keywords:  Gas chromatography tandem mass spectrometry; Heart failure; HiSorb; Pentafluorobenzyl bromide; Saliva; Short-chain fatty acids and hydroxy acids

DOI:  https://doi.org/10.1016/j.jchromb.2023.123826