bims-metlip 2024-06-02 papers

bims-metlip

Biomed News

on Methods and protocols in metabolomics and lipidomics

Issue of 2024–06–02
twenty papers selected by
Sofia Costa, Matterworks

A Targeted, Bioinert LC-MS/MS Method for Sensitive, Comprehensive Analysis of Signaling Lipids.
Development and application of a sensitive liquid chromatography-tandem mass spectrometry method for the quantitative analysis of 11 free fatty acids in human serum using a derivatisation strategy.
Recent additions and access to a multidimensional lipidomic database containing liquid chromatography, ion mobility spectrometry, and tandem mass spectrometry information.
Determination of six volatile fatty acids in human serum, urine and faeces by low temperature derivatisation combined with HPLC-MS/MS.
Rapid and reliable quantification of urinary malondialdehyde by HILIC-MS/MS: A derivatization-free breakthrough approach.
Simultaneous quantification of terpenes and cannabinoids by reversed-phase LC-APCI-MS/MS in Cannabis sativa L. samples combined with a subsequent chemometric analysis.
Comparison of data processing strategies using commercial vs. open-source software in GC-Orbitrap-HRMS untargeted metabolomics analysis for food authentication: thyme geographical differentiation and marker identification as a case study.
Characterization of spironolactone and metabolites derivatized using Girard's reagent P using mass spectrometry and ion mobility spectrometry.
Visualizing 13C-Labeled Metabolites in Maize Root Tips with Mass Spectrometry Imaging.
Optimization and validation of metabolomics methods for feline urine and serum towards application in veterinary medicine.
Improved detection in untargeted lipidomics through silver-doped infrared matrix-assisted laser desorption electrospray ionization.
The screening method for 39 phytotoxins and mycotoxins in blood and urine with liquid chromatography-high resolution mass spectrometry.
Development and validation of an HILIC/MS/MS method for determination of nusinersen in rabbit plasma.
TransExION: a transformer based explainable similarity metric for comparing IONS in tandem mass spectrometry.
Impact of conditioning runs on hydrophilic interaction chromatography repeatability and its application as a second dimension in online comprehensive two-dimensional liquid chromatography.
An LC-MS/MS method for the simultaneous determination of 6-cyanodopamine, 6-nitrodopamine, 6-nitrodopa, 6-nitroadrenaline and 6-bromodopamine in human plasma and its clinical application in patients with chronic kidney disease.
Measurement of very low-molecular weight metabolites by traveling wave ion mobility and its use in human urine samples.
Pharmacokinetics of IMM-H012 in rats using ultra-performance liquid chromatography-tandem mass spectrometry.
A mass defect-based approach for the automatic construction of peak lists for databases of mass spectra with limited resolution: Application to time-of-flight secondary ion mass spectrometry data.
A new approach towards highly sensitive detection of endogenous N-acetylaspartic acid, N-acetylglutamic acid, and N-acetylaspartylglutamic acid in brain tissues based on strong anion exchange monolith microextraction coupled with UHPLC-MS/MS.

Anal Chem. 2024 May 25.

A Targeted, Bioinert LC-MS/MS Method for Sensitive, Comprehensive Analysis of Signaling Lipids.

Stefanie Rubenzucker, Mailin-Christin Manke, Rainer Lehmann, Alice Assinger, Oliver Borst, Robert Ahrends.

Signaling lipids are key players in cellular processes. Despite their importance, no method currently allows their comprehensive monitoring in one analytical run. Challenges include a wide dynamic range, isomeric and isobaric species, and unwanted interaction along the separation path. Herein, we present a sensitive and robust targeted liquid chromatography-mass spectrometry (LC-MS/MS) approach to overcome these challenges, covering a broad panel of 17 different signaling lipid classes. It involves a simple one-phase sample extraction and lipid analysis using bioinert reversed-phase liquid chromatography coupled to targeted mass spectrometry. The workflow shows excellent sensitivity and repeatability in different biological matrices, enabling the sensitive and robust monitoring of 388 lipids in a single run of only 20 min. To benchmark our workflow, we characterized the human plasma signaling lipidome, quantifying 307 endogenous molecular lipid species. Furthermore, we investigated the signaling lipidome during platelet activation, identifying numerous regulations along important lipid signaling pathways. This highlights the potential of the presented method to investigate signaling lipids in complex biological systems, enabling unprecedentedly comprehensive analysis and direct insight into signaling pathways.

DOI: https://doi.org/10.1021/acs.analchem.4c01388
J Chromatogr A. 2024 May 23. pii: S0021-9673(24)00393-5. [Epub ahead of print]1728 465019

Development and application of a sensitive liquid chromatography-tandem mass spectrometry method for the quantitative analysis of 11 free fatty acids in human serum using a derivatisation strategy.

Yongfu Zeng, Qian Li, Ruoyu Zhang, Maolin Wei, Xiaochan Zhao, Liping Hao, Haoran Zhang, Ziqing Wang, Xiujuan Guo, Lianfeng Ai.

  A stable isotope dilution-liquid chromatography-tandem mass spectrometry method based on a derivatisation strategy involving an N,N'-carbonylimidazole solution (CDI) with 4-(dimethylamino)-benzenemethanamine was developed for the determination of 11 free fatty acids (FFAs) in human blood samples. Serum samples were subjected to liquid‒liquid extraction and centrifuged, and the supernatant was collected for a two-step derivatisation reaction with a CDI and 4-(dimethylamino)-aniline acetonitrile solution. The derivatised solution was separated on a ACQUITY UPLC HSS T3 column (2.1 × 50 mm, 1.8 µm) column with a mobile phase consisting of water-acetonitrile in gradient elution and then detected by tandem mass spectrometry using electrospray ionisation (ESI) and multiple reaction monitoring (MRM) in positive ion mode and quantified using the isotope internal standard method. The effects of the derivatisation reaction time, temperature and concentration of derivatisation reagents on the response values of the analytes were investigated. The optimal conditions were as follows: 1.0 mg mL-1 CDI acetonitrile solution at 25 °C for 25 min, followed by a reaction with a 1.0 mg mL-1 4-(dimethylamino)-benzenemethanamine acetonitrile solution at 70 °C for 30 min. Under the optimal conditions, the limits of detection (LODs) of the 11 FFAs were in the range of 3.0-14.0 ng mL-1; the limits of quantification (LOQs) were in the range of 8.0-45.0 ng mL-1; and the mean recoveries ranged from 83.4 to 112.8%, with intraday and interday precisions ranging from 0.7 to 9.1% and 3.7-9.5%, respectively. The experimental method is simple in terms of the pretreatment operation, accurate and reliable, and can be applied to the sensitive determination of FFAs in human blood samples.

Keywords:  Derivatisation; Free fatty acids; LC‒MS/MS; Serum

DOI:  https://doi.org/10.1016/j.chroma.2024.465019
Anal Bioanal Chem. 2024 May 30.

Recent additions and access to a multidimensional lipidomic database containing liquid chromatography, ion mobility spectrometry, and tandem mass spectrometry information.

Amie M Solosky, Kaylie I Kirkwood-Donelson, Melanie T Odenkirk, Erin S Baker.

  The importance of lipids in biology continues to grow with their recent linkages to more diseases and conditions, microbiome fluctuations, and environmental exposures. These associations have motivated researchers to evaluate lipidomic changes in numerous matrices and studies. Lipidomic analyses, however, present numerous challenges as lipid species have broad chemistries that require different extraction methods and instrumental analyses to evaluate and separate their many isomers and isobars. Increasing knowledge about different lipid characteristics is therefore crucial for improving their separation and identification. Here, we present a multidimensional database for lipids analyzed on a platform combining reversed-phase liquid chromatography, drift tube ion mobility spectrometry, collision-induced dissociation, and mass spectrometry (RPLC-DTIMS-CID-MS). This platform and the different separation characteristics it provides enables more confident lipid annotations when compared to traditional tandem mass spectrometry platforms, especially when analyzing highly isomeric molecules such as lipids. This database expands on our previous publication containing only human plasma and bronchoalveolar lavage fluid lipids and provides experimental RPLC retention times, IMS collision cross section (CCS) values, and m/z information for 877 unique lipids from additional biofluids and tissues. Specifically, the database contains 1504 precursor [M + H]+, [M + NH4]+, [M + Na]+, [M-H]-, [M-2H]2-, [M + HCOO]-, and [M + CH3COO]- ion species and their associated CID fragments which are commonly targeted in clinical and environmental studies, in addition to being present in the chloroform layer of Folch extractions. Furthermore, this multidimensional RPLC-DTIMS-CID-MS database spans 5 lipid categories (fatty acids, sterols, sphingolipids, glycerolipids, and glycerophospholipids) and 24 lipid classes. We have also created a webpage (tarheels.live/bakerlab/databases/) to enhance the accessibility of this resource which will be populated regularly with new lipids as we identify additional species and integrate novel standards.

Keywords:  Collision cross section; Database; Ion mobility spectrometry; Lipidomics; Lipids; Mass spectrometry; Reverse phase liquid chromatography (RPLC)

DOI:  https://doi.org/10.1007/s00216-024-05351-4
J Chromatogr B Analyt Technol Biomed Life Sci. 2024 May 24. pii: S1570-0232(24)00181-8. [Epub ahead of print]1241 124172

Determination of six volatile fatty acids in human serum, urine and faeces by low temperature derivatisation combined with HPLC-MS/MS.

Qian Li, Yongfu Zeng, Lianfeng Ai, Maolin Wei, Xiaomeng Liu, Xiaochan Zhao, Haoran Zhang, Xiujuan Guo.

  A stable isotope dilution-liquid chromatography tandem mass spectrometry method based on a low-temperature derivatization strategy with 3-nitrophenylhydrazine (3-NPH) was developed for the determination of six volatile fatty acids (VFAs) in serum, urine, and feces. Ice acetonitrile was used to precipitate proteins and extract the target analytes. The extract was derivatized with 3-NPH methanol solution at 4 °C. BEH C8 (1.7 μm, 2.1 × 100 mm) column was used for chromatographic separation, and acetonitrile-water (both containing 0.01 % formic acid) were used as the mobile phase with a gradient elution of 10 min. Electrospray ionization source (ESI) in negative ion multiple reaction monitoring (MRM) mode were used for analyte detection. The regression coefficients R2 of the calibration curves for the six VFAs were in the range of 0.9963-0.9994, and the LOQs were in the range of 0.02-0.5 μg mL-1, with the recoveries in the range of 85.3-104.3 %, and the intra- and inter-day precision in the range of 1.8-9.1 %. The method is simple, accurate and reliable, and has been applied in the sensitive determination of VFAs in complex biological samples.

Keywords:  3-Nitrophenylhydrazine; HPLC–MS; Intestinal flora; Volatile fatty acids

DOI:  https://doi.org/10.1016/j.jchromb.2024.124172
Anal Chim Acta. 2024 Jul 04. pii: S0003-2670(24)00538-5. [Epub ahead of print]1311 342737

Rapid and reliable quantification of urinary malondialdehyde by HILIC-MS/MS: A derivatization-free breakthrough approach.

Gabriela Chango, Diego García-Gómez, Carmelo García Pinto, Encarnación Rodríguez-Gonzalo, José Luis Pérez Pavón.

   BACKGROUND: The development of fast analytical methods is crucial for the research, discovery, and confirmation of crucial biomarkers. Furthermore, the implementation of fast analytical strategies contributes to efficient and time-effective procedures. In this sense, analysis of malondialdehyde (MDA) has become an important tool for understanding the role of oxidative stress in various diseases and for evaluating the efficacy of therapeutic interventions.
RESULTS: A rapid and robust liquid chromatography tandem mass spectrometry method (HPLC-MS/MS) has been developed to determine endogenous amounts of malondialdehyde (MDA) in human urine without any associated derivatization reaction. MDA was separated in 4 min through a Urea-HILIC column and was analyzed using a triple quadrupole mass spectrometer in negative electrospray ionization mode. With a 50-fold dilution as the only sample pretreatment after alkaline hydrolysis, no matrix effect was present, which allowed for a fast and simple quantification by means of an external standard calibration with a limit of detection of 0.20 ng mL-1. The whole methodology was validated by analyzing unspiked and spiked urine samples from ten healthy individuals and comparing with the results obtained by the standard addition method. MDA was detected in all cases, with natural concentrations varying from 0.11 ± 0.03 to 0.31 ± 0.03 mg g-1 creatinine. Accuracies were found to be satisfactory, ranging from 95 % to 101 %. The proposed method also exhibited good repeatability and reproducibility (RSD<15 %) for four quality control levels.
SIGNIFICANCE: The main significance of this method is the avoidance of a derivatization reaction for the determination of urinary MDA, this constituting a step forward when compared with previous literature. This breakthrough not only streamlines time analysis to less than 5 min per sample but also results in a more robust procedure. Consequently, the method here developed could be applied to subsequent future research involving the determination of MDA as a lipid peroxidation biomarker, where simple, rapid, and reliable methods could represent a significant improvement.

Keywords:  Lipid peroxidation; Liquid chromatography; Malondialdehyde; Mass spectrometry; Oxidative stress

DOI:  https://doi.org/10.1016/j.aca.2024.342737
Anal Bioanal Chem. 2024 May 25.

Simultaneous quantification of terpenes and cannabinoids by reversed-phase LC-APCI-MS/MS in Cannabis sativa L. samples combined with a subsequent chemometric analysis.

Justine Raeber, Michael Poetzsch, Anina Schmidli, Sina Favrod, Christian Steuer.

  Cannabis sativa L. has been the most discussed medicinal plant in recent years. In particular, the dynamic shift from a formerly illicit and tightly controlled substance to a plant recognized for both medicinal and recreational purposes has brought C. sativa into the global spotlight. Due to the ongoing international legalization processes, fast and convenient analytical methods for the quality control of C. sativa flowers for medicinal and recreational purposes are of tremendous interest. In this study, we report the development and validation of a liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based method applying atmospheric pressure chemical ionization (APCI) to fully quantify 16 terpenes and 7 cannabinoids including their acidic forms by a single chromatographic method. The method presented here is unique and simple, as it eliminates the need for derivatization reactions and includes the unconventional analysis of volatile compounds by liquid chromatography. Samples were prepared by a simple and fast ethanolic extraction. Separation was accomplished within 25 min on a reversed-phase C18 column. Method validation was conducted according to international guidelines regarding selectivity, accuracy, precision, robustness, and linearity. Detection was done in multiple reaction monitoring, which allowed the simultaneous quantification of co-eluting analytes applying two selective mass transitions. In addition, due to reproducible in-source decarboxylation, the acidic forms of cannabinoids were reliably quantified using mass transitions of the neutral forms. The accuracy given as the bias was below 15% for all analytes. Matrix effects for cannabinoids were studied by spiking Humulus lupulus extracts with the analytes at varying concentrations. APCI did not show susceptibility toward ion suppression or enhancement. In addition, the recovery effect after spiking was between 80 and 120% for terpenes. Further, 55 authentic C. sativa extracts were fully quantified, and the obtained results for the terpene profiles were compared to state-of-the-art gas chromatography coupled to flame ionization detection. Comparable results were achieved, emphasizing the method's applicability for cannabinoids and terpenes. Further, acquired metabolite patterns for C. sativa samples were studied, identifying a relationship between cannabinoid and terpene patterns, as well as the abundance of myrcene in CBD-dominant C. sativa strains.

Keywords:  APCI; Cannabinoids; HPLC; Mass spectrometry; Terpenes

DOI:  https://doi.org/10.1007/s00216-024-05349-y
Anal Bioanal Chem. 2024 May 28.

Comparison of data processing strategies using commercial vs. open-source software in GC-Orbitrap-HRMS untargeted metabolomics analysis for food authentication: thyme geographical differentiation and marker identification as a case study.

Araceli Rivera-Pérez, Antonia Garrido Frenich.

  Untargeted analysis of gas chromatography-high-resolution mass spectrometry (GC-HRMS) data is a key and time-consuming challenge for identifying metabolite markers in food authentication applications. Few studies have been performed to evaluate the capability of untargeted data processing tools for feature extraction, metabolite annotation, and marker selection from untargeted GC-HRMS data since most of them are focused on liquid chromatography (LC) analysis. In this framework, this study provides a comprehensive evaluation of data analysis tools for GC-Orbitrap-HRMS plant metabolomics data, including the open-source MS-DIAL software and commercial Compound Discoverer™ software (designed for Orbitrap data processing), applied for the geographical discrimination and search for thyme markers (Spanish vs. Polish differentiation) as the case study. Both approaches showed that the feature detection process is highly affected by unknown metabolites (Levels 4-5 of identification confidence), background signals, and duplicate features that must be carefully assessed before further multivariate data analysis for reliable putative identification of markers. As a result, Compound Discoverer™ and MS-DIAL putatively annotated 52 and 115 compounds at Level 2, respectively. Further multivariate data analysis allowed the identification of differential compounds, showing that the putative identification of markers, especially in challenging untargeted analysis, heavily depends on the data processing parameters, including available databases used during compound annotation. Overall, this method comparison pointed out both approaches as good options for untargeted analysis of GC-Orbitrap-HRMS data, and it is presented as a useful guide for users to implement these data processing approaches in food authenticity applications depending on their availability.

Keywords:  Compound Discoverer; Data analysis; Gas chromatography; High-resolution mass spectrometry; MS-DIAL; Metabolomics

DOI:  https://doi.org/10.1007/s00216-024-05347-0
Rapid Commun Mass Spectrom. 2024 Aug 15. 38(15): e9775

Characterization of spironolactone and metabolites derivatized using Girard's reagent P using mass spectrometry and ion mobility spectrometry.

Stephanie M Jones, Kaylie I Kirkwood-Donelson, Georgia M Alexander, Lalith Perera, Serena M Dudek, Alan K Jarmusch.

RATIONALE: Spironolactone is a steroidal drug prescribed for a variety of medical conditions and is extensively metabolized quickly after administration. Measurement of spironolactone and its metabolites remains challenging using mass spectrometry (MS) due to in-source fragmentation and relatively poor ionization using electrospray ionization. Therefore, improved methods of measurements are needed, particularly in the case of small sample volumes.
METHODS: Girard's reagent P (GP) derivatization of spironolactone was employed to improve response and provide an MS-based solution to the measurement of spironolactone and its metabolites. We performed ultra-high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UHPLC-ESI-MS/MS) and ion mobility spectrometry (IMS)-high-resolution mass spectrometry (HRMS) to fully characterize the GP derivatization products. Analytes were studied in positive ionization mode, and MS/MS was performed using nonresonance and resonance excitation collision-induced dissociation.
RESULTS: We observed the successful GP derivatization of spironolactone and its metabolites using authentic chemical standards. A signal enhancement of 1-2 orders of magnitude was observed for GP-derivatized versions of spironolactone and its metabolites. Further, GP derivatization eliminated in-source fragmentation. Finally, we performed GP derivatization and ultra-high-performance liquid chromatography-high-resolution mass spectrometry (UHPLC-HRMS) in a small volume of murine serum (20 μL) from spironolactone-treated and control animals and observed multiple spironolactone metabolites only in the spironolactone-treated group.
CONCLUSIONS: GP derivatization was proven to have advantageous mass spectral performance (e.g., limiting in-source fragmentation, enhancing signals, and eliminating isobaric analytes) for spironolactone and its metabolites. This work and the detailed characterization using ultra-high-performance liquid chromatography-high-resolution tandem mass spectrometry (UHPLC-HRMS/MS) and IMS serve as the foundation for future developments in reaction optimization and/or quantitative assay development.

DOI: https://doi.org/10.1002/rcm.9775
J Am Soc Mass Spectrom. 2024 May 31.

Visualizing 13C-Labeled Metabolites in Maize Root Tips with Mass Spectrometry Imaging.

Young Jin Lee, Pubudu Nuwan Perera Hapuarachchige, Evan A Larson, Ngoc Le, Trevor T Forsman.

  Tracing in vivo isotope-labeled metabolites has been used to study metabolic pathways or flux analysis. However, metabolic differences between the cells have been often ignored in these studies due to the limitation of solvent-based extraction. Here we demonstrate that the mass spectrometry imaging of in vivo isotope-labeled metabolites, referred to as MSIi, can provide important insights into metabolic dynamics with cellular resolution that may supplement the traditional metabolomics and flux analysis. Developing maize root tips are adopted as a model system for MSIi by supplementing 200 mM [U-13C]glucose in 0.1x Hoagland medium. MSIi data sets were acquired for longitudinal sections of newly grown maize root tips after growing 5 days in the medium. A total of 56 metabolite features were determined to have been 13C-labeled based on accurate mass and the number of carbon matching with the metabolite databases. Simple sugars and their derivatives were fully labeled, but some small metabolites were partially labeled with a significant amount of fully unlabeled metabolites still present, suggesting the recycling of "old" metabolites in the newly grown tissues. Some distinct localizations were found, including the low abundance of hexose and its derivatives in the meristem, the high abundance of amino acids in the meristem, and the localization to epidermal and endodermal cells for lipids and their intermediates. Fatty acids and lipids were slow in metabolic turnover and showed various isotopologue distributions with intermediate building blocks, which may provide flux information for their biosynthesis.

Keywords:  13C-labeling; MALDI; in vivo isotope labeling; maize; mass spectrometry imaging; root tip

DOI:  https://doi.org/10.1021/jasms.4c00042
Anal Chim Acta. 2024 Jun 29. pii: S0003-2670(24)00495-1. [Epub ahead of print]1310 342694

Optimization and validation of metabolomics methods for feline urine and serum towards application in veterinary medicine.

Ellen Vanden Broecke, Laurens Van Mulders, Ellen De Paepe, Sylvie Daminet, Lynn Vanhaecke.

   BACKGROUND: Metabolomics is an emerging and powerful technology that offers a comprehensive view of an organism's physiological status. Although widely applied in human medicine, it is only recently making its introduction in veterinary medicine. As a result, validated metabolomics protocols in feline medicine are lacking at the moment. Since biological interpretation of metabolomics data can be misled by the extraction method used, species and matrix-specific optimized and validated metabolomic protocols are sorely needed.
RESULTS: Systematic optimization was performed using fractional factorial experiments for both serum (n = 57) and urine (n = 24), evaluating dilution for both matrices, and aliquot and solvent volume, protein precipitation time and temperature for serum. For the targeted (n = 76) and untargeted (n = 1949) validation of serum respectively, excellent instrumental, intra-assay and inter-day precision were observed (CV ≤ 15% or 30%, respectively). Linearity deemed sufficient both targeted and untargeted (R2 ≥ 0.99 or 0.90, respectively). An appropriate targeted recovery between 70 and 130% was achieved. For the targeted (n = 69) and untargeted (n = 2348) validation of the urinary protocol, excellent instrumental and intra-assay precision were obtained (CV ≤ 15% or 30%, respectively). Subsequently, the discriminative ability of our metabolomics methods was confirmed for feline chronic kidney disease (CKD) by univariate statistics (n = 41 significant metabolites for serum, and n = 55 for urine, p-value<0.05) and validated OPLS-DA models (R2(Y) > 0.95, Q2(Y) > 0.65, p-value<0.001 for both matrices).
SIGNIFICANCE: This study is the first to present an optimized and validated wholistic metabolomics methods for feline serum and urine using ultra-high performance liquid chromatography coupled to quadrupole-Orbitrap high-resolution mass spectrometry. This robust methodology opens avenues for biomarker panel selection and a deeper understanding of feline CKD pathophysiology and other feline applications.

Keywords:  Biofluids; Feline nephrology; Metabotyping; UHPLC-Orbitrap-HRMS

DOI:  https://doi.org/10.1016/j.aca.2024.342694
Rapid Commun Mass Spectrom. 2024 Aug 15. 38(15): e9832

Improved detection in untargeted lipidomics through silver-doped infrared matrix-assisted laser desorption electrospray ionization.

Seth M Eisenberg, David C Muddiman.

RATIONALE: Silver doping of electrospray is known to increase the abundance of olefinic compounds detected by mass spectrometry. While demonstrated in targeted experiments, this has yet to be investigated in an untargeted study. Utilizing infrared matrix-assisted laser desorption electrospray ionization mass spectrometry imaging (IR-MALDESI-MSI), an untargeted lipidomics experiment on mouse liver was performed to evaluate the advantages of silver-doped electrospray.
METHODS: 10 ppm silver nitrate was doped into the IR-MALDESI solvent consisting of 60% acetonitrile and 0.2% formic acid. Using an Orbitrap mass spectrometer in positive ionization mode, MSI was performed, analyzing from m/z 150 to m/z 2000 to capture all lipids with potential silver adducts. The lipids detected in the control and silver-doped electrosprays were compared by annotating using the LIPID MAPS Structural Database and eliminating false positives using the metabolite annotation confidence score.
RESULTS: Silver-doped electrospray allowed for the detection of such ions of lipid molecules as [M + H]+ or [M + NH4]+ and as [M + Ag]+. Among the ions seen as [M + H]+ or [M + NH4]+, the signal was comparable between the control and silver-doped electrosprays. The silver-doped electrospray led to a 10% increase in the number of detected lipids, all of which contained a bay region increasing the interaction between silver and alkenes. Silver preferentially interacted with lipids that did not contain hard bases such as phosphates.
CONCLUSIONS: Silver-doped electrospray enabled detection of 10% more olefinic lipids, all containing bay regions in their putative structures. This technique is valuable for detecting previously unobserved lipids that have the potential to form bay regions, namely fatty acyls, glycerolipids, prenol lipids, and polyketides.

DOI: https://doi.org/10.1002/rcm.9832
J Chromatogr B Analyt Technol Biomed Life Sci. 2024 May 26. pii: S1570-0232(24)00182-X. [Epub ahead of print]1241 124173

The screening method for 39 phytotoxins and mycotoxins in blood and urine with liquid chromatography-high resolution mass spectrometry.

Yuqi Yin, Weiyang Sun, Xuan Wang, Jiayue Chen, Hongyan Zeng, Sifan Hao, Lin Ren, Li Yong, Chunying Luo, Xiaoli Zou.

   BACKGROUND: Poisonings caused by plant toxins and mycotoxins occur frequently, which do great harm to human health and social public health safety. When a poisoning incident occurs, biological samples are commonly be used to conduct the detection of toxic substances and their metabolites for targeted clinical treatment and incident analysis.
OBJECTIVE: To establish an efficient and accurate analysis method of 39 phytotoxins and mycotoxins in blood and urine by high performance liquid chromatography quadrupole tandem orbitrap mass spectrometry (HPLC-Orbitrap MS).
METHOD: After 3 mL of methanol being added to 1 mL blood and urine respectively for extraction and protein precipitation, the supernatant was injected into HPLC-Orbitrap MS for analysis. The phytotoxins and mycotoxins were separated by Hypersil GOLD PFP column with gradient elution using methanol-5 mmol/L ammonium acetate as mobile phase. The data were collected in ESI positive ion mode using Full MS/dd-MS2 for mass spectrometry detection.
RESULT: The mass database of 39 phytotoxins and mycotoxins was developed, and accurate qualitative analysis can be obtained by matching with the database using the proposed identification criteria. Limit of detections (LODs) were 1.34 × 10-4 ∼ 1.92 ng/mL and 1.92 × 10-4 ∼ 9.80 ng/mL for blood and urine samples, respectively. Limits of quantification (LOQ) of toxins in blood and urine ranged from 4.47 × 10-4 ∼ 6.32 ng/mL and 6.39 × 10-4 ∼ 32.67 ng/mL, respectively. Intra-day relative standard deviations (RSDs) were 0.79 % ∼ 10.90 %, and inter-day RSDs were 1.08 % ∼ 18.93 %. The recoveries can reach 90 % ∼ 110 % with matrix matching calibration curves.
CONCLUSION: The established method is simple and rapid to operate, which can complete the sample analysis within 30 min, providing technical support for clinical poisoning treatment and public health poisoning analysis.

Keywords:  Blood; High performance liquid chromatography-high resolution mass spectrometry; Mycotoxins; Phytotoxins; Urine

DOI:  https://doi.org/10.1016/j.jchromb.2024.124173
Heliyon. 2024 May 30. 10(10): e31213

Development and validation of an HILIC/MS/MS method for determination of nusinersen in rabbit plasma.

Xiao Zhang, Chunjie Sha, Wei Zhang, Fengjuan Zhao, Mingli Zhu, Guangyi Leng, Wanhui Liu.

  A hydrophilic interaction liquid chromatography tandem mass spectrometry (HILIC/MS/MS) method was developed and validated for the quantitative analysis of the fully phosphorothioate modified oligonucleotide nusinersen. HILIC/MS/MS method is more robust and compatible with mass spectrometry than ion pair reversed-phase liquid chromatography-tandem mass spectrometry (IP-RP-LC/MS/MS). Various types and concentrations of additives and different pH of mobile phase affected the mass spectrometry response, chromatographic peak shape and retention of nusinersen. The optimized extraction method of nusinersen employs hydrophilic-lipophilic balance solid phase extraction, with a recovery of up to 80 %. Chromatographic quantification was performed using a gradient system on an amide column and the mobile phase consisted of ammonium acetate, acetonitrile and water in a certain proportion. The fully phosphorothioate modified nusinersen can obtain a high mass spectrometry response by providing greater peak symmetry and high ionization efficiency in a high-pH mobile phase. Moreover, the significant carry over interference was observed at the pH 6.3 of the mobile phase. Adjusting the pH value up to 10, and the carry over interference disappeared. The lower limit of quantitation of this developed HILIC/MS/MS assay was 30.0 ng/mL and the method was systematic methodology validated. This HILIC/MS/MS method provides an attractive and robust alternative for the quantitative analysis of nusinersen and was applied in the pharmacokinetic study of nusinersen in rabbits.

Keywords:  HILIC/MS/MS; Nusinersen; Pharmacokinetics; Phosphorothioate modification; Quantitation

DOI:  https://doi.org/10.1016/j.heliyon.2024.e31213
J Cheminform. 2024 May 28. 16(1): 61

TransExION: a transformer based explainable similarity metric for comparing IONS in tandem mass spectrometry.

Danh Bui-Thi, Youzhong Liu, Jennifer L Lippens, Kris Laukens, Thomas De Vijlder.

  Small molecule identification is a crucial task in analytical chemistry and life sciences. One of the most commonly used technologies to elucidate small molecule structures is mass spectrometry. Spectral library search of product ion spectra (MS/MS) is a popular strategy to identify or find structural analogues. This approach relies on the assumption that spectral similarity and structural similarity are correlated. However, popular spectral similarity measures, usually calculated based on identical fragment matches between the MS/MS spectra, do not always accurately reflect the structural similarity. In this study, we propose TransExION, a Transformer based Explainable similarity metric for IONS. TransExION detects related fragments between MS/MS spectra through their mass difference and uses these to estimate spectral similarity. These related fragments can be nearly identical, but can also share a substructure. TransExION also provides a post-hoc explanation of its estimation, which can be used to support scientists in evaluating the spectral library search results and thus in structure elucidation of unknown molecules. Our model has a Transformer based architecture and it is trained on the data derived from GNPS MS/MS libraries. The experimental results show that it improves existing spectral similarity measures in searching and interpreting structural analogues as well as in molecular networking. SCIENTIFIC CONTRIBUTION: We propose a transformer-based spectral similarity metrics that improves the comparison of small molecule tandem mass spectra. We provide a post hoc explanation that can serve as a good starting point for unknown spectra annotation based on database spectra.

Keywords:  Explainable deep neural network; Small molecule identification; Spectral similarity; Structural similarity; Tandem mass spectrometry

DOI:  https://doi.org/10.1186/s13321-024-00858-5
J Sep Sci. 2024 May;47(9-10): e2300935

Impact of conditioning runs on hydrophilic interaction chromatography repeatability and its application as a second dimension in online comprehensive two-dimensional liquid chromatography.

Clémence Gély, Yoan R Monneau, Maggy Hologne, Karine Faure.

  A common separation approach for polar compounds involves coupling reversed-phase liquid chromatography (RPLC) with hydrophilic interaction chromatography (HILIC) in two-dimensional chromatography. The higher proportion of acetonitrile used in the HILIC mobile phase, which enhances mass spectrometry detection, encourages its use in the second dimension. Previous studies demonstrated that the HILIC column can be partially equilibrated within very short timeframes without compromising retention time stability, rendering it suitable in online comprehensive two-dimensional liquid chromatography (LC×LC) setups. In addition, a specific number of conditioning cycles seems necessary to establish stable retention times. Here, the repeatability of HILIC when employed as second dimension in LC×LC was investigated, with a focus on determining the required number of conditioning cycles to achieve repeatable retention times. Various parameters influenced by the LC×LC online modulation system were studied, such as steep gradient slopes up to 8%, and very short equilibration times, less than or equal to dead time, as well as injection volume and solvent, which depend on the first dimension. Finally, the use of HILIC as a second dimension with tailored conditioning runs was applied to the analysis of hyaluronic acid hydrogel digests. The application of an RPLC×HILIC method using five conditioning runs yielded exceptional stability in second-dimension retention times.

Keywords:  RPLC×HILIC; conditioning runs; hyaluronic hydrogel; repeatability; two‐dimensional liquid chromatography

DOI:  https://doi.org/10.1002/jssc.202300935
Biomed Chromatogr. 2024 May 26. e5896

An LC-MS/MS method for the simultaneous determination of 6-cyanodopamine, 6-nitrodopamine, 6-nitrodopa, 6-nitroadrenaline and 6-bromodopamine in human plasma and its clinical application in patients with chronic kidney disease.

Luiz Fernando Ribeiro, Tainah Babadopulos, Mariana G de Oliveira, Flávio Nishimaru, Roberto Zatz, Rosilene Motta Elias, Odorico Moraes, Elisabete Moraes, Larryn W Peterson, Gilberto De Nucci.

  The aim of this study was to develop a high-performance liquid chromatography-tandem mass spectrometry method for the determination of 6-cyanodopamine, 6-nitrodopamine, 6-nitrodopa, 6-nitroadrenaline and 6-bromodopamine in human plasma samples. Strata-X 33 μm solid-phase extraction cartridges were used for the extraction of the catecholamines from human plasma samples. The catecholamines were separated in a 150 × 3 mm Shim-pack GIST C18-AQ column with 3 μm particle size, placed in an oven at 40°C and perfused with 82% mobile phase A (acetonitrile-H2O; 90:10, v/v) + 0.4% acetic acid and 18% mobile phase B (deionized H2O) + 0.2% formic acid at a flow rate of 340 μl/min in isocratic mode. The injected volume was 4 μl and the run lasted 4 min. The method was linear from 0.1 to 20 ng/ml and the lower limit of quantification was 0.1 ng/ml for all analytes. The method was applied to evaluate the plasma levels of catecholamines in plasma of patients with chronic kidney disease and allowed the detection for the first time of circulating levels of the novel catecholamines 6-bromodopamine and 6-cyanodopamine.

Keywords:  LC–MS/MS; human plasma; novel catecholamines; solid‐phase extraction

DOI:  https://doi.org/10.1002/bmc.5896
J Pharm Anal. 2024 May;14(5): 100921

Measurement of very low-molecular weight metabolites by traveling wave ion mobility and its use in human urine samples.

Alongkorn Kurilung, Suphitcha Limjiasahapong, Khwanta Kaewnarin, Pattipong Wisanpitayakorn, Narumol Jariyasopit, Kwanjeera Wanichthanarak, Sitanan Sartyoungkul, Stephen Choong Chee Wong, Nuankanya Sathirapongsasuti, Chagriya Kitiyakara, Yongyut Sirivatanauksorn, Sakda Khoomrung.

  The collision cross-sections (CCS) measurement using ion mobility spectrometry (IMS) in combination with mass spectrometry (MS) offers a great opportunity to increase confidence in metabolite identification. However, owing to the lack of sensitivity and resolution, IMS has an analytical challenge in studying the CCS values of very low-molecular-weight metabolites (VLMs ≤ 250 Da). Here, we describe an analytical method using ultrahigh-performance liquid chromatography (UPLC) coupled to a traveling wave ion mobility-quadrupole-time-of-flight mass spectrometer optimized for the measurement of VLMs in human urine samples. The experimental CCS values, along with mass spectral properties, were reported for the 174 metabolites. The experimental data included the mass-to-charge ratio (m/z), retention time (RT), tandem MS (MS/MS) spectra, and CCS values. Among the studied metabolites, 263 traveling wave ion mobility spectrometry (TWIMS)-derived CCS values (TWCCSN2) were reported for the first time, and more than 70% of these were CCS values of VLMs. The TWCCSN2 values were highly repeatable, with inter-day variations of <1% relative standard deviation (RSD). The developed method revealed excellent TWCCSN2 accuracy with a CCS difference (ΔCCS) within ±2% of the reported drift tube IMS (DTIMS) and TWIMS CCS values. The complexity of the urine matrix did not affect the precision of the method, as evidenced by ΔCCS within ±1.92%. According to the Metabolomics Standards Initiative, 55 urinary metabolites were identified with a confidence level of 1. Among these 55 metabolites, 53 (96%) were VLMs. The larger number of confirmed compounds found in this study was a result of the addition of TWCCSN2 values, which clearly increased metabolite identification confidence.

Keywords:  Collision cross-section; Human urine; Traveling wave ion mobility; Very low-molecular-weight metabolites

DOI:  https://doi.org/10.1016/j.jpha.2023.12.011
Biomed Chromatogr. 2024 May 28. e5905

Pharmacokinetics of IMM-H012 in rats using ultra-performance liquid chromatography-tandem mass spectrometry.

Shujuan Wu, Jialei Wu, Longquan Lin, Rongbin Jiang, Xianqin Wang, Congcong Wen, Xia-Yin Zhu.

  The present study examined the pharmacokinetics of IMM-H012 in rat plasma, utilizing ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Internal standard cilostazol was employed, and plasma samples were processed using acetonitrile precipitation. A mobile phase (acetonitrile-0.1% formic acid in water) with gradient elution was used to achieve chromatographic separation using a UPLC BEH C18 column. In multiple reaction monitoring mode, electrospray ionization MS/MS was utilized in positive ionization mode. Based on findings, the lower limit of quantification was 2 ng/mL, and the linearity of IMM-H012 in rat plasma was found to be acceptable within the range of 2-2000 ng/mL (R2 > 0.995). The intra-day and inter-day precision relative standard deviation was less than 14% of IMM-H012 in rat plasma. The matrix effect was within the range of 102%-107%, and the accuracy ranged from 92% to 113%. Pharmacokinetics of IMM-H012 in rats after oral administration were successfully studied using UPLC-MS/MS.

Keywords:  IMM‐H012; UPLC‐MS/MS; pharmacokinetics; plasma; rat

DOI:  https://doi.org/10.1002/bmc.5905
Rapid Commun Mass Spectrom. 2024 Aug 15. 38(15): e9777

A mass defect-based approach for the automatic construction of peak lists for databases of mass spectra with limited resolution: Application to time-of-flight secondary ion mass spectrometry data.

Mouad Daoudi, Nicolas Nuns, Philipp Schiffmann, Arnaud Frobert, Benjamin Hanoune, Pascale Desgroux, Alessandro Faccinetto.

RATIONALE: This study has developed a data processing protocol based on mass defect analysis for the automatic construction of unique peak lists addressing the need for the fast and efficient treatment of databases of mass spectra with limited mass resolution.
METHODS: The data processing protocol, implemented in MATLAB, is tested on a database of 126 mass spectra obtained from time-of-flight secondary ion mass spectrometry analysis of the exhaust of a laboratory diesel miniCAST burner deposited on Ti substrates.
RESULTS: The data processing protocol converts the mass spectra into a data matrix suitable for chemometrics (peak list) by combining mass defect analysis and multivariate analysis. In particular, the role of the mass defect analysis is expanded to improve mass calibration and automate the construction of the peak list.
CONCLUSIONS: In this context, mass defect analysis becomes an invaluable technique for the efficient processing of databases of mass spectra with limited mass resolution by allowing the fast and automated construction of a peak list common to all mass spectra, by improving the mass calibration, and finally by reducing the number of molecular formulae consistent with a given accurate mass, thus facilitating the identification of unknown ions.

DOI: https://doi.org/10.1002/rcm.9777
Mikrochim Acta. 2024 05 31. 191(6): 360

A new approach towards highly sensitive detection of endogenous N-acetylaspartic acid, N-acetylglutamic acid, and N-acetylaspartylglutamic acid in brain tissues based on strong anion exchange monolith microextraction coupled with UHPLC-MS/MS.

Wenxiu Zhou, Xiaoyuan Lv, Shengman Zhang, Zhenye Gao, Bingjie Li, Xin Wang.

  A novel in-tube solid-phase microextraction coupled with an ultra-high performance liquid chromatography-mass spectrometry method has been established for simultaneous quantification of three crucial brain biomarkers N-acetylaspartic acid (NAA), N-acetylglutamic acid (NAG), and N-acetylaspartylglutamic acid (NAAG). A polymer monolith with quaternary ammonium as the functional group was designed and exhibited efficient enrichment of target analytes through strong anion exchange interaction. Under the optimized conditions, the proposed method displayed wide linear ranges (0.1-80 nM for NAA and NAG, 0.2-160 nM for NAAG) with good precision (RSDs were lower than 15%) and low limits of detection (0.019-0.052 nM), which is by far the most sensitive approach for NAA, NAG, and NAAG determination. Furthermore, this approach has been applied to measure the target analytes in mouse brain samples, and endogenous NAA, NAG, and NAAG were successfully detected and quantified from only around 5 mg of cerebral cortex, cerebellum, and hippocampus. Compared with existing methods, the newly developed method in the current study provides highest sensitivity and lowest sample consumption for NAA, NAG, and NAAG measurements, which would potentially be utilized in determining and tracking these meaningful brain biomarkers in diseases or treatment processes, benefiting the investigations of pathophysiology and treatment of brain disorders.

Keywords:  Brain biomarkers; In-tube solid-phase microextraction; Liquid chromatography-mass spectrometry; Polymer monolithic column; Strong anion exchange

DOI:  https://doi.org/10.1007/s00604-024-06431-z