bims-metlip 2019-12-08 papers

bims-metlip

Biomed News

on Methods and protocols in metabolomics and lipidomics

Issue of 2019–12–08
33 papers selected by
Sofia Costa, Cold Spring Harbor Laboratory

VOCCluster: Untargeted Metabolomics Feature Clustering Approach for Clinical Breath Gas Chromatography - Mass Spectrometry Data.
Ion exchange in supercritical fluid chromatography tandem mass spectrometry (SFC-MS/MS): Application for polar and ionic drugs and metabolites in forensic and anti-doping analysis.
Development and validation of LC-ESI-MS/MS methods for quantification of 27 free and conjugated estrogen-related metabolites.
A validated UHPLC-MS/MS method for simultaneous quantification of 9 exocyclic DNA adducts induced by 8 aldehydes.
Ion Mobility Spectrometry and the Omics: Distinguishing Isomers, Molecular Classes and Contaminant Ions in Complex Samples.
Multifunctional isotopic standards based steroidomics strategy: Exploration of cancer screening model.
Predicting Breast Cancer by Paper Spray Ion Mobility Spectrometry Mass Spectrometry and Machine Learning.
Simple and sensitive method for analysis of 14 anti-tuberculosis drugs using LC/MS/MS in human plasma.
Improvement of Tryptophan Analysis by Liquid Chromatography-Single Quadrupole Mass Spectrometry Through the Evaluation of Multiple Parameters.
Machine learning distilled metabolite biomarkers for early stage renal injury.
Enhancement of On-tissue Chemical Derivatization by Laser-Assisted Tissue Transfer for MALDI MS Imaging.
Using the chemical noise background in MALDI mass spectrometry imaging for mass alignment and calibration.
Clustering and variable selection evaluation of 13 unsupervised methods for multi-omics data integration.
Shotgun lipidomics-based characterization of the landscape of lipid metabolism in colorectal cancer.
Quantitative analysis of progesterone using isotope dilution-matrix-assisted laser desorption ionization-time of flight mass spectrometry as a reference procedure for radioimmunoassay.
A rapid and adaptable lipidomics method for quantitative UPLC-mass spectrometric analysis of phosphatidylethanolamine and phosphatidylcholine in vitro, and in cells.
Metabolomics analysis of the saliva in patients with chronic hepatitis B using nuclear magnetic resonance: a pilot study.
Differential adsorption of analyte and deuterated analogue onto the rotor seal of the injector in high-performance liquid chromatography/electrospray ionisation mass spectrometry.
Differences in the lipid patterns during maturation of 3T3-L1 adipocytes investigated by thin-layer chromatography, gas chromatography, and mass spectrometric approaches.
spectrum_utils: A Python package for mass spectrometry data processing and visualization.
Identification of Suvorexant in Blood Using LC-MS-MS: Important Considerations for Matrix Effects and Quantitative Interferences in Targeted Assays.
Development, validation, comparison, and implementation of a highly efficient and effective method using magnetic solid-phase extraction with hydrophilic-lipophilic-balanced materials for LC-MS/MS analysis of pesticides in seawater.
Simultaneous determination of thirteen substances related to NAFLD in mice brain tissue using 3-Aminobutyric acid as internal standard by HPLC-FLD.
Targeted metabolomics approach for identification of relapsing-remitting multiple sclerosis markers and evaluation of diagnostic models.
MINIATURIZATION IN MASS SPECTROMETRY.
Sustainable and Selective Extraction of Lipids and Bioactive Compounds from Microalgae.
Assessment of coverage for endogenous metabolites and exogenous chemical compounds using an untargeted metabolomics platform.
UPLC-MS based urine untargeted metabolomic analyses to differentiate bladder cancer from renal cell carcinoma.
Experimental design for parameter estimation in steady-state linear models of metabolic networks.
Ultra-high Performance Supercritical Fluid Chromatography Combined with Quadrupole Time-of-Flight Mass Spectrometry for the Characterization of Pentaerythritol Fatty Acid Esters.
Stir membrane liquid phase microextraction of tetracyclines using switchable hydrophilicity solvents followed by high-performance liquid chromatography.
Piloting hyperpolarized 13C-pyruvate MRI for imaging advanced prostate cancer.
Metabolite Profiling of Ortho-, Meta- and Para-Fluorofentanyl by Hepatocytes and High-Resolution Mass Spectrometry.

Anal Chem. 2019 Dec 02.

VOCCluster: Untargeted Metabolomics Feature Clustering Approach for Clinical Breath Gas Chromatography - Mass Spectrometry Data.

Yaser Alkhalifah, Iain Phillips, Andrea Soltoggio, Kareen Darnley, William H Nailon, Duncan McLaren, Michael Eddleston, C L Paul Thomas, Dahlia Salman.

Metabolic profiling of breath analysis involves processing, alignment, scaling and clustering of thousands of features ex-tracted from Gas Chromatography Mass spectrometry (GC-MS) data from hundreds of participants. The multi-step data processing is complicated, operator error-prone and time-consuming. Automated algorithmic clustering methods that are able to cluster features in a fast and reliable way are necessary. These accelerate metabolic profiling and discovery plat-forms for next generation medical diagnostic tools. Our unsupervised clustering technique, VOCCluster, prototyped in Py-thon, handles features of deconvolved GC-MS breath data. VOCCluster was created from a heuristic ontology based on the observation of experts undertaking data processing with a suite of software packages. VOCCluster identifies and clusters groups of volatile organic compounds (VOCs) from deconvolved GC-MS breath with similar mass spectra and retention index profiles. VOCCluster was used to cluster more than 15,000 features extracted from 74 GC-MS clinical breath samples obtained from participants with cancer before and after a radiation therapy. Results were evaluated against a panel of ground truth compounds and compared to other clustering methods (DBSCAN and OPTICS) that were used in previous metabolomics studies. VOCCluster was able to cluster those features into 1081 groups (including endogenous, exogenous compounds and instrumental artefacts) with an accuracy rate of 96% (± 0.04 at 95% confidence interval).

DOI: https://doi.org/10.1021/acs.analchem.9b03084
J Chromatogr A. 2019 Nov 19. pii: S0021-9673(19)31158-6. [Epub ahead of print] 460726

Ion exchange in supercritical fluid chromatography tandem mass spectrometry (SFC-MS/MS): Application for polar and ionic drugs and metabolites in forensic and anti-doping analysis.

Majlinda Xhaferaj, Edgar Naegele, Maria Kristina Parr.

  Supercritical fluid chromatography (SFC) is an orthogonal technique to UHPLC. In recent years, SFC has demonstrated potential for use in the analysis of a broad variety of analytes of different polarities, if modifiers and additives are utilized as additional mobile phase constituents. However, to date, little research has been carried out on ion-exchange separation of highly polar and ionic analytes using SFC. The objective of this work was to investigate the elution characteristics of polar compounds using SFC combined with tandem mass spectrometry. Highly polar and even ionic drugs and metabolites, with a diversity of functional groups such as gamma-hydroxybutyrate (GHB), gamma-butyrolactone, GHB-glucuronide, ethyl sulfate, ethyl glucuronide as well as meldonium and gamma-butyrobetaine, were selected for the study. To investigate the chromatographic behavior of the solutes using SFC, a systematic chromatographic method development workflow including a basic validation in human urine was implemented. To ensure the best selectivity, columns with different stationary phase chemistries (silica, NH2, CN, SCX, Diol, EP and amide) were screened. Furthermore, different modifier compositions were evaluated, including pure methanol and methanol with various additives (ammonium acetate or ammonium formate, water, and/or ammonia or formic acid in varying amounts). Trends in retention time shifts were investigated by the systematic variation of gradients, backpressure and column compartment temperature. The quality and reliability of the method has been tested in the course of a basic validation by evaluation of selectivity, linearity, limit of identification, limit of quantification, carry-over, precision, and matrix effect. The highest chromatographic selectivity (especially with respect to the separation of the critical pair of zwitterionic meldonium and γ-butyrobetaine) and suitable peak shapes for all target analytes were obtained using the strong cation exchange column. We observed that increasing buffer concentrations improved the peak shape. Ion-exchange stationary phase and polar additives (buffers, bases, acids and water) mixed with organic modifiers enabled separation of small ionic molecules using SFC. Hence, SFC further demonstrated its wide applicability in small molecule analysis, and may be considered an alternative separation technique to improve the separation of polar analytes.

Keywords:  Anti-doping analysis; Forensics; Ion-exchange chromatography; Method development; SFC-ESI-MS/MS

DOI:  https://doi.org/10.1016/j.chroma.2019.460726
Anal Biochem. 2019 Dec 02. pii: S0003-2697(19)30850-4. [Epub ahead of print] 113531

Development and validation of LC-ESI-MS/MS methods for quantification of 27 free and conjugated estrogen-related metabolites.

Carien van der Berg, Gerda Venter, Francois H van der Westhuizen, Elardus Erasmus.

  An imbalance in the estrogen metabolism has been associated with an increased risk of breast cancer development. Evaluation of the estrogen biotransformation capacity requires monitoring of various estrogen metabolites. Up to now, only some estrogen metabolites could be measured in urine. However, in order to offer tailor made nutritional support or therapies, a complete estrogen metabolite profile is required in order to identify specific deficiencies in this pathway for each patient individually. Here, we focused on this need to quantify as many as possible of the estrogen-related metabolites excreted in urine. The method was developed to quantify 27 estrogen-related metabolites in small urine quantities. This entailed sample clean-up with a multi-step solid phase extraction procedure, derivatisation of the metabolites in the less water-soluble fraction through dansylation, and analyses using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The metabolites accurately quantified by the method devised included parent estrogens, hydroxylated and methylated forms, metabolites of the 16α-hydroxyestrogen pathway, sulphate and glucuronide conjugated forms, precursors and a related steroid hormone. This method was validated and enabled quantification in the high picograms and low nanograms per millilitre range. Finally, analyses of urine samples confirmed detection and quantification of each of the metabolites.

Keywords:  Biotransformation; Dansylation; Estrogen metabolism; LC-MS/MS; Method development

DOI:  https://doi.org/10.1016/j.ab.2019.113531
J Pharm Biomed Anal. 2019 Nov 23. pii: S0731-7085(19)31580-8. [Epub ahead of print] 113007

A validated UHPLC-MS/MS method for simultaneous quantification of 9 exocyclic DNA adducts induced by 8 aldehydes.

Héléna Alamil, Mathilde Lechevrel, Stéphanie Lagadu, Laurence Galanti, Zeina Dagher, Raphaël Delépée.

  Human exposure to aldehydes is implicated in several diseases including cancer. These strong electrophilic compounds can react with nucleophilic sites in DNA to form reversible and irreversible modifications. These modifications, if not repaired, can contribute to pathogenesis. The aim of our study was to provide a mass spectrometry (MS)-based profiling method for identifying potential biomarkers of aldehydes exposure. We have developed and validated a highly sensitive method using ultra high performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UHPLC-ESI-MS/MS) for the simultaneous quantitation of 9 exocyclic DNA adducts derived from 8 main exogenous and endogenous aldehydes, namely formaldehyde, acetaldehyde, acrolein, crotonaldehyde, malondialdehyde, 4-hydroxy-2-nonenal, glyoxal and methylglyoxal. Finally, we applied the established method to quantify adducts in genomic DNA isolated from the blood of a smoker and a non-smoker blood samples in order to demonstrate its applicability.

Keywords:  Adductomic; Aldehydes; Analytical method validation; Cancer; Exposure biomarkers; Oxidative stress; Ultrahigh performance liquid chromatography -electrospray ionization- tandem mass spectrometry

DOI:  https://doi.org/10.1016/j.jpba.2019.113007
Trends Analyt Chem. 2019 Jul;116 292-299

Ion Mobility Spectrometry and the Omics: Distinguishing Isomers, Molecular Classes and Contaminant Ions in Complex Samples.

Kristin E Burnum-Johnson, Xueyun Zheng, James N Dodds, Jeremy Ash, Denis Fourches, Carrie D Nicora, Jason P Wendler, Thomas O Metz, Katrina M Waters, Janet K Jansson, Richard D Smith, Erin S Baker.

  Ion mobility spectrometry (IMS) is a widely used analytical technique providing rapid gas phase separations. IMS alone is useful, but its coupling with mass spectrometry (IMS-MS) and various front-end separation techniques has greatly increased the molecular information achievable from different omic analyses. IMS-MS analyses are specifically gaining attention for improving metabolomic, lipidomic, glycomic, proteomic and exposomic analyses by increasing measurement sensitivity (e.g. S/N ratio), reducing the detection limit, and amplifying peak capacity. Numerous studies including national security-related analyses, disease screenings and environmental evaluations are illustrating that IMS-MS is able to extract information not possible with MS alone. Furthermore, IMS-MS has shown great utility in salvaging molecular information for low abundance molecules of interest when high concentration contaminant ions are present in the sample by reducing detector suppression. This review highlights how IMS-MS is currently being used in omic analyses to distinguish structurally similar molecules, isomers, molecular classes and contaminant ions.

Keywords:  Exposomics; Glycomics; Ion Mobility Spectrometry; Lipidomics; Mass Spectrometry; Metabolomics; Omics; Proteomics

DOI:  https://doi.org/10.1016/j.trac.2019.04.022
J Chromatogr A. 2019 Nov 18. pii: S0021-9673(19)31155-0. [Epub ahead of print] 460723

Multifunctional isotopic standards based steroidomics strategy: Exploration of cancer screening model.

Qian Zhang, Ran Liu, Huarong Xu, Xiao Yang, Yiwen Zhang, Qian Wang, Peng Gao, Kaishun Bi, Tao Han, Qing Li.

  Quantitative determination of endogenous compounds in biological samples has still been challenged by the absence of authentic blank matrix. Alternative strategy of surrogate matrix for preparing reference samples are prevalent due to its low cost and high availability. However, the evaluation system of surrogate matrix is not perfect. Herein, a novel multifunctional isotopic standards based steroidomics strategy was developed. Isotope-labeled standards were used not only as internal standards but also for the evaluation the feasibility of surrogate matrix, which improved the accuracy of assessment and could provide a new prospect for the quantitative analysis of endogenous compounds. Based on this approach, a detailed optimization from LC separation, MS detection to extraction conditions for global steroids in the steroidogenesis was firstly carried out. Characteristics and regularities of steroids in LC-MS were summarized to make references for further targeted or untargeted steroidomics study. Then eighteen steroids were quantified with high accuracy and high sensitivity in plasma from four types of cancer patients and healthy subjects using 1% BSA in PBS as surrogate matrix. And multi-steroids indexes with the best discriminating ability for lung, colorectal, breast and gastric cancer in different genders were identified successfully with Student's t-test, PLS-DA and logistic regression- ROC curve analysis. Finally, efficient cancer screening workflow was established by integrating the amine submetabolomics and lipidomics data of our previous studies. Taken together, the integrated steroidomics strategy could shed a light on the guidance for further steroidome as well as other endogenous compounds analysis and may provide a powerful tool for cancer diagnosis.

Keywords:  Cancer biomarkers; Multifunctional isotopic standards; Steroidomics; Surrogate matrix

DOI:  https://doi.org/10.1016/j.chroma.2019.460723
Anal Chem. 2019 Dec 06.

Predicting Breast Cancer by Paper Spray Ion Mobility Spectrometry Mass Spectrometry and Machine Learning.

Ying-Chen Huang, Hsin-Hsiang Chung, Ewelina Dutkiewicz, Chih-Lin Chen, Hua-Yi Hsieh, Bo-Rong Chen, Ming-Yang Wang, Cheng-Chih Hsu.

Paper spray ionization has been used as a fast sampling/ionization method for the direct mass spectrometric analysis of biological samples at ambient conditions. Here, we demonstrated that by utilizing paper spray ionization mass spectrometry (PSI-MS) coupled with field asymmetric waveform ion mobility spectrometry (FAIMS), predictive metabolic and lipidomic profiles of routine breast core needle biopsies could be obtained effectively. By the combination of machine learning and pathological examination reports, we developed a classification model, which has an overall accuracy of 87.5% for an in-stantaneous differentiation between cancerous and non-cancerous breast tissues utilizing metabolic and lipidomic profiles. Our results suggested that PSI-FAIMS-MS is a powerful approach for rapid breast cancer diagnosis based on altered meta-bolic and lipidomic profiles.

DOI: https://doi.org/10.1021/acs.analchem.9b03966
Rapid Commun Mass Spectrom. 2019 Dec 04.

Simple and sensitive method for analysis of 14 anti-tuberculosis drugs using LC/MS/MS in human plasma.

Qian Lei, Yuan Zhao, Hao Wang, Jun Zhou, Xiaohui Lv, Liyun Dang, Changsheng Zhu.

RATIONALE: The monitoring of the plasma concentration and dose adjustment of anti-tuberculosis (TB) drugs are beneficial for improving responses to drug treatment and avoiding drug adverse reactions. A simple and sensitive liquid chromatography-tandem mass spectrometry (LC/MS/MS) method was developed to measure the plasma concentrations of 14 anti-TB drugs: ethambutol, isoniazid, pyrazinamide, levofloxacin, gatifloxacin, moxifloxacin, prothionamide, linezolid, rifampin, rifapentine, rifabutin, cycloserine, p-aminosalicylic acid (PAS) and clofazimine.
METHODS: Human plasma was precipitated by acetonitrile and was subsequently separated by an AQ-C18 column with a gradient elution. Drug concentrations were determined using multiple reaction monitoring (MRM) in positive ion ESI mode. According to pharmacokinetic data of patients, the peak concentration ranges and timing of blood collection were determined.
RESULTS: Intra- and inter-day precision was less than 14.8%. Linearity, accuracy, extraction recovery and matrix effect were acceptable for each drug. Stability of the method satisfied different storage conditions.
CONCLUSIONS: The method applied the sensitive and reproducible determination of 14 frequently-used anti-TB drugs, which already showed benefit for some TB patients.

DOI: https://doi.org/10.1002/rcm.8667
Front Chem. 2019 ;7 797

Improvement of Tryptophan Analysis by Liquid Chromatography-Single Quadrupole Mass Spectrometry Through the Evaluation of Multiple Parameters.

Rasmus la Cour, Henning Jørgensen, Jan K Schjoerring.

  Tryptophan is a key component in many biological processes and an essential amino acid in food and feed materials. Analysis of the tryptophan content in proteins or protein-containing matrices has always been a challenge. We show here that the preparation of samples prior to tryptophan analysis can be significantly simplified, and the time consumption reduced, by using ascorbic acid as antioxidant to eliminate the problem of tryptophan degradation during alkaline hydrolysis. Combined with separation by HPLC and detection by Single Quadrupole Mass Spectrometry, this allows the analytical run time to be reduced to 10 min. The alkaline hydrolysate obtained in the method presented here may be combined with the oxidized hydrolysate obtained when sulfur-containing amino acids are to be measured, thus essentially providing two analyses for the time of one.

Keywords:  HPLC; LC-MS; amino acid profile; high-throughput; protein

DOI:  https://doi.org/10.3389/fchem.2019.00797
Metabolomics. 2019 Dec 05. 16(1): 4

Machine learning distilled metabolite biomarkers for early stage renal injury.

Yan Guo, Hui Yu, Danqian Chen, Ying-Yong Zhao.

   INTRODUCTION: With chronic kidney disease (CKD), kidney becomes damaged overtime and fails to clean blood. Around 15% of US adults have CKD and nine in ten adults with CKD do not know they have it.
OBJECTIVE: Early prediction and accurate monitoring of CKD may improve care and decrease the frequent progression to end-stage renal disease. There is an urgent demand to discover specific biomarkers that allow for monitoring of early-stage CKD, and response to treatment.
METHOD: To discover such biomarkers, shotgun high throughput was applied to the detection of serum metabolites biomarker discovery for early stages of CKD from 703 participants. Ultra performance liquid chromatography coupled with high-definition mass spectrometry (UPLC-HDMS)-based metabolomics was used for the determination of 703 fasting serum samples from five stages of CKD patients and age-matched healthy controls.
RESULTS AND CONCLUSION: We discovered a set of metabolite biomarkers using a series of classic and neural network based machine learning techniques. This set of metabolites can separate early CKD stage patents from normal subjects with high accuracy. Our study illustrates the power of machine learning methods in metabolite biomarker study.

Keywords:  Chronic kidney disease; Deep learning; Glomerular filtration rate; Machine learning; Metabolite

DOI:  https://doi.org/10.1007/s11306-019-1624-0
Anal Chem. 2019 Dec 04.

Enhancement of On-tissue Chemical Derivatization by Laser-Assisted Tissue Transfer for MALDI MS Imaging.

Shuai Guo, Weiwei Tang, Yu Hu, Yanwen Chen, Andrew Gordon, Bin Li, Ping Li.

The development of on-tissue chemical derivatization methods for matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) of small endogenous metabolites in tissues has attracted great attention for their advantages in improving detection sensitivity and ionization efficiency of poorly ionized and low abundant metabolites. Herein, a laser-assisted tissue transfer (LATT) technique was developed to enhance on-tissue derivatization of small molecules. Using a focused blue laser, a thin-layer tissue film (~ 1 µm) was transferred to an acceptor slide from a 6 µm dry tissue section preliminarily coated with derivatization and matrix reagents. The acceptor slide with its ablated constituents was then imaged by MALDI MS. On-tissue chemical derivatization with amino-specific derivatization reagent 4-hydroxy-3-methoxycinnamaldehyde (CA) was carried out on LATT system. 20-235 folds increase in signal intensity for CA derivatized metabolites such as amino acids, neurotransmitters and dipeptides were observed from rat brain tissues in comparison with conventional incubation-based derivatization. This technique was further extended to derivatize steroids with Girard reagent T (GirT). The remarkable derivatization efficiency can mainly be attributed to the minimization of ion suppression effects due to the reduced thickness of tissue section and endogenous components. Additionally, shorter derivatization time with no obvious metabolite delocalization was achieved using LATT method. These results demonstrate the advantages of LATT in the enhancement of on-tissue derivatization for the more specific and sensitive imaging of small metabolites in tissues with MALDI MS.

DOI: https://doi.org/10.1021/acs.analchem.9b04618
Anal Chem. 2019 Dec 03.

Using the chemical noise background in MALDI mass spectrometry imaging for mass alignment and calibration.

Tobias Boskamp, Delf Lachmund, Rita Casadonte, Lena Hauberg-Lotte, Jan Hendrik Kobarg, Jörg Kriegsmann, Peter Maass.

Matrix-assisted laser desorption/ ionization mass spectrometry imaging (MALDI MSI) is an established tool for the investigation of formalin fixed paraffin embedded (FFPE) tissue samples and shows a high potential for applications in clinical research and histopathological diagnosis. The applicability and accuracy of this method, however, heavily depends on the quality of the acquired data, and in particular mass misalignment in axial time-of-flight (TOF) MSI continues to be a serious issue. We present a mass alignment and recalibration method that is specifically designed to operate on MALDI peptide imaging data. The proposed method exploits statistical properties of the characteristic chemical noise background observed in peptide imaging experiments. By comparing these properties to a theoretical peptide mass model, the effective mass shift of each spectrum is estimated and corrected. The method was evaluated on a cohort of 31 FFPE tissue samples, pursuing a statistical validation approach to estimate both the reduction of relative misalignment, as well as the increase in absolute mass accuracy. Our results suggest that a relative mass precision of approx. 5 ppm and an absolute accuracy of approx. 30 ppm are achievable using our method.

DOI: https://doi.org/10.1021/acs.analchem.9b04473
Brief Bioinform. 2019 Dec 03. pii: bbz138. [Epub ahead of print]

Clustering and variable selection evaluation of 13 unsupervised methods for multi-omics data integration.

Morgane Pierre-Jean, Jean-François Deleuze, Edith Le Floch, Florence Mauger.

  Recent advances in NGS sequencing, microarrays and mass spectrometry for omics data production have enabled the generation and collection of different modalities of high-dimensional molecular data. The integration of multiple omics datasets is a statistical challenge, due to the limited number of individuals, the high number of variables and the heterogeneity of the datasets to integrate. Recently, a lot of tools have been developed to solve the problem of integrating omics data including canonical correlation analysis, matrix factorization and SM. These commonly used techniques aim to analyze simultaneously two or more types of omics. In this article, we compare a panel of 13 unsupervised methods based on these different approaches to integrate various types of multi-omics datasets: iClusterPlus, regularized generalized canonical correlation analysis, sparse generalized canonical correlation analysis, multiple co-inertia analysis (MCIA), integrative-NMF (intNMF), SNF, MoCluster, mixKernel, CIMLR, LRAcluster, ConsensusClustering, PINSPlus and multi-omics factor analysis (MOFA). We evaluate the ability of the methods to recover the subgroups and the variables that drive the clustering on eight benchmarks of simulation. MOFA does not provide any results on these benchmarks. For clustering, SNF, MoCluster, CIMLR, LRAcluster, ConsensusClustering and intNMF provide the best results. For variable selection, MoCluster outperforms the others. However, the performance of the methods seems to depend on the heterogeneity of the datasets (especially for MCIA, intNMF and iClusterPlus). Finally, we apply the methods on three real studies with heterogeneous data and various phenotypes. We conclude that MoCluster is the best method to analyze these omics data. Availability: An R package named CrIMMix is available on GitHub at https://github.com/CNRGH/crimmix to reproduce all the results of this article.

Keywords:  benchmarks; multi-omics; performance evaluation; real data; unsupervised integrative methods

DOI:  https://doi.org/10.1093/bib/bbz138
Biochim Biophys Acta Mol Cell Biol Lipids. 2019 Nov 30. pii: S1388-1981(19)30230-6. [Epub ahead of print] 158579

Shotgun lipidomics-based characterization of the landscape of lipid metabolism in colorectal cancer.

Yuting Wang, Sebastian Hinz, Ortrud Uckermann, Pia Hönscheid, Witigo von Schönfels, Greta Burmeister, Alexander Hendricks, Jacobo Miranda Ackerman, Gustavo B Baretton, Jochen Hampe, Mario Brosch, Clemens Schafmayer, Andrej Shevchenko, Sebastian Zeissig.

  Solid tumors are characterized by global metabolic alterations which contribute to their growth and progression. Altered gene expression profiles and plasma lipid composition suggested a role for metabolic reprogramming in colorectal cancer (CRC) development. However, a conclusive picture of CRC-associated lipidome alterations in the tumor tissue has not emerged. Here, we determined molar abundances of 342 species from 20 lipid classes in matched biopsies of CRC and adjacent normal mucosa. We demonstrate that in contrast to previous reports, CRC shows a largely preserved lipidome composition that resembles that of normal colonic mucosa. Important exceptions include increased levels of lyso-phosphatidylinositols in CRC and reduced abundance of ether phospholipids in advanced stages of CRC. As such, our observations challenge the concept of widespread alterations in lipid metabolism in CRC and rather suggest changes in the cellular lipid profile that are limited to selected lipids involved in signaling and the scavenging of reactive oxygen species.

Keywords:  Cancer; Colon; Lipid metabolism; Mass spectrometry; Shotgun lipidomics

DOI:  https://doi.org/10.1016/j.bbalip.2019.158579
Clin Chim Acta. 2019 Nov 29. pii: S0009-8981(19)32142-4. [Epub ahead of print]

Quantitative analysis of progesterone using isotope dilution-matrix-assisted laser desorption ionization-time of flight mass spectrometry as a reference procedure for radioimmunoassay.

Ming-Hui Yang, Shih-Cheng Chen, Ko-Chin Chen, Han-Ping You, Hsin-Yi Wu, Yi-Ming Arthur Chen, Ying-Fong Huang, Ming-Yii Huang, Cheng-Hui Yuan, Po-Chiao Lin, Yu-Chang Tyan.

   BACKGROUND: Progesterone is one of the female steroid hormones and plays an important role in the menstrual cycle and during pregnancy. It is especially important in preparing the uterus for the implantation of the blastocyst and maintaining pregnancy. The concentration in human serum is measured to determine the ovarian function retroactively and the cause of abortion in early pregnancy.
METHODS: A quantification assay based on isotope dilution mass spectrometry to determine the concentration of progesterone in human serum is reported. Incorporated with 13C3-progesterone, serum samples were subjected to progesterone extraction and clean-up by C4 solid-phase-extraction columns and hexane-based liquid/liquid extraction, respectively. The cleaned-up serum samples were then subjected to MALDI-TOF mass spectrometry for the quantification of progesterone.
RESULTS: Progesterone and the internal standard, 13C3-progesterone, were measured in the selected reaction monitoring mode for the transitions m/z 315.4 to 108.9 and m/z 318.4 to 111.9, respectively. We calculated the peak area ratio of progesterone to 13C3-progesterone. The progesterone concentration in human serum was calculated by substituting the peak area ratio into an isotope dilution calibration curve, and then compared with the radioimmunoassay.
CONCLUSIONS: In the study, the concentrations of serum progesterone were measured, and the recovered progesterone concentration determined by the assay showed good robustness and consistency in comparison to the conventional radioimmunologic assay. We concluded that the 13C3-progesterone-based quantification assay is a robust method for the measurement of serum progesterone.

Keywords:  MALDI-TOF/MS; isotope dilution; progesterone; radioimmunologic assay

DOI:  https://doi.org/10.1016/j.cca.2019.11.020
Anal Methods. 2019 Apr 07. 11(13): 1765-1776

A rapid and adaptable lipidomics method for quantitative UPLC-mass spectrometric analysis of phosphatidylethanolamine and phosphatidylcholine in vitro, and in cells.

Daniel J Stephenson, H Patrick MacKnight, L Alexis Hoeferlin, Margaret Park, Jeremy Allegood, Christopher L Cardona, Charles E Chalfant.

  Phosphatidylethanolamine (PE) and phosphatidylcholine (PC) are highly prevalent phospholipids in mammalian membranes. There are currently no methods for detection of minute levels of these phospholipids or simultaneously with products of the utilization of these phospholipid substrates by phospholipase A2 (PLA2) enzymes. To examine the substrate utilization of PE and PC by PLA2, we developed a method to accurately detect and measure specific forms of PE and PC as low as 50 femtomoles. Validation of this method consisted of an enzymatic assay to monitor docosahexaenoic acid and arachidonic acid release from the hydrolysis of PE and PC by group IV phospholipase A2 (cPLA2α) coupled to the generation of lyso-PE (LPE) and lyso-PC (LPC). In addition, the PE and PC profiles of RAW 264.7 macrophages were monitored with zymosan/lipopolysaccharide-treatment. Finally, genetic validation for the specificity of the method consisted of the downregulation of two biosynthetic enzymes responsible for the production of PE and PC, choline kinase A (CHKA) and ethanolamine kinase 1 (ETNK1). This new UPLC ESI-MS/MS method provides accurate and highly sensitive detection of PE and PC species containing AA and DHA allowing for the specific examination of the substrate utilization of these phospholipids by PLA2 in vitro and in cells.

Keywords:  Group IVA cytosolic phospholipase A2; QTRAP 5500 LC-MS/MS; mass spectrometry; phosphatidylcholine; phosphatidylethanolamine

DOI:  https://doi.org/10.1039/C9AY00052F
Iran J Basic Med Sci. 2019 Sep;22(9): 1044-1049

Metabolomics analysis of the saliva in patients with chronic hepatitis B using nuclear magnetic resonance: a pilot study.

Kambiz Gilany, Ashraf Mohamadkhani, Saeed Chashmniam, Parisa Shahnazari, Mehdi Amini, Babak Arjmand, Reza Malekzadeh, Bibi Fatemeh Nobakht Motlagh Ghoochani.

   Objectives: Hepatitis B virus infection causes chronic disease such as cirrhosis and hepatocellular carcinoma. The metabolomics investigations have been demonstrated to be related to pathophysiologic mechanisms in many disorders such as hepatitis B infection. The aim of this study was to investigate the saliva metabolic profile of patients with chronic hepatitis B infection and to identify underlying mechanisms as well as potential biomarkers associated with the disease.
Materials and Methods: Saliva from 16 healthy subjects and 20 patients with chronic hepatitis B virus were analyzed by nuclear magnetic resonance (NMR). Then, multivariate statistical analysis was performed to identify discriminative metabolites between two groups.
Results: A set of metabolites were detected, including propionic acid, putrescine, acetic acid, succinic acid, tyrosine, lactic acid, butyric acid, pyruvic acid, 4-pyridoxic acid and 4-hydroxybenzoic acid, which in combination with one another could accurately distinguish patients from healthy controls. Our results clearly demonstrated altered metabolites are involved in nine metabolic pathways.
Conclusion: Metabolomics has the potential to be considered as a novel clinical tool for hepatitis B diagnosis while contributing to a comprehensive understanding of disease mechanisms.

Keywords:  Diagnostic biomarkers Hepatitis B virus; Metabolomics; NMR; Saliva

DOI:  https://doi.org/10.22038/ijbms.2019.36669.8733
Rapid Commun Mass Spectrom. 2019 Dec 04.

Differential adsorption of analyte and deuterated analogue onto the rotor seal of the injector in high-performance liquid chromatography/electrospray ionisation mass spectrometry.

Gwyn A Lord.

RATIONALE: Quantitation of analytes by LC/MS is now very widely used, usually with an appropriate internal standard, but this does not guarantee problem-free analyses. Investigation of a problem that arose during method development is described, where it was surmised that differential adsorption onto the HPLC injector of the analyte (Rac GTPase inhibitor NSC23766) and its deuterated analogue, as internal standard, was taking place.
METHODS: Samples were injected onto the HPLC via either a Valco C14W.1 internal sample injector or a Rheodyne 7125 injector, both with dissimilar Vespel rotor seals, and linked to a Q-Tof mass spectrometer. Alternatively, samples were infused directly into the mass spectrometer via a syringe-driver. Electrospray ionisation in positive mode was used in both cases.
RESULTS: Experiments demonstrated significant differential adsorption of the analyte (RAC Inhibitor, NSC23766) and its tri-deuterated internal standard onto the Vespel rotor seal, with the latter seeming to be preferentially adsorbed. The deuterated analogue also showed lower electrospray sensitivity, as demonstrated by syringe infusion of a mixture of the compounds, compared with their separate infusion at the same concentration.
CONCLUSIONS: This study has demonstrated the problem of differential adsorption onto HPLC Vespel rotor seals, and that the use of a stable isotope analogue as internal standard does not guarantee the constancy of the analyte/internal response ratio in quantitative methods. The partial solution was to work at much higher concentration where adsorption, whilst still apparent, was relatively insignificant.

DOI: https://doi.org/10.1002/rcm.8672
Anal Bioanal Chem. 2019 Dec 03.

Differences in the lipid patterns during maturation of 3T3-L1 adipocytes investigated by thin-layer chromatography, gas chromatography, and mass spectrometric approaches.

Yulia Popkova, Dirk Dannenberger, Jürgen Schiller, Kathrin M Engel.

  Populations of industrialized countries have registered a dramatically increasing prevalence in obesity for many years. Despite continuous research, mechanisms involved in the storage and utilization of chemical energy in adipocytes are still under investigation. Adipocytes have the task to store excessive energy in the form of triacylglycerols (TG) and it is already well-known that the fatty acyl composition of TG is largely determined by the composition of the diet. In contrast to TG, the composition of adipocyte phospholipids was less comprehensively investigated. In this study, the compositions of the most abundant phospholipid classes of 3T3-L1 undifferentiated (preadipocytes) and differentiated cells (adipocytes) were determined. The lipid fractions were isolated by normal phase high-performance thin-layer chromatography and subsequently analyzed by electrospray ionization mass spectrometry. Additionally, the fatty acyl (FA) compositions were determined by gas chromatography. The positions of the FA residues were additionally confirmed by phospholipase A2 digestion. The advantages and disadvantages of the different analytical approaches will be discussed. It will be shown that undifferentiated 3T3-L1 and mature adipocytes differ extremely regarding their compositions. This goes along with an increase in odd-chain fatty acids. Graphical abstract.

Keywords:  3T3-L1 adipocytes; ESI mass spectrometry; Gas chromatography; Odd-chain fatty acids; Phospholipase A2 digestion; Thin-layer chromatography

DOI:  https://doi.org/10.1007/s00216-019-02243-w
Anal Chem. 2019 Dec 06.

spectrum_utils: A Python package for mass spectrometry data processing and visualization.

Wout Bittremieux.

Given the wide diversity in applications of biological mass spectrometry, custom data analyses are often needed to fully interpret the results of an experiment. Such bioinformatics scripts necessarily include similar basic functionality to read mass spectral data from standard file formats, process it, and visualize it. Rather than having to reimplement this functionality, to facilitate this task, spectrum_utils is a Python package for mass spectrometry data processing and visualization. Its high-level functionality enables developers to quickly prototype ideas for computational mass spectrometry projects in only a few lines of code. Notably, the data processing functionality is highly optimized for computational efficiency to be able to deal with the large volumes of data that are generated during mass spectrometry experiments. The visualization functionality makes it possible to easily produce publication-quality figures as well as interactive spectrum plots for inclusion on web pages. spectrum_utils is available for Python 3.6+, includes extensive online documentation and examples, and can be easily installed using conda. It is freely available as open source under the Apache 2.0 license at https://github.com/bittremieux/spectrum_utils.

DOI: https://doi.org/10.1021/acs.analchem.9b04884
J Anal Toxicol. 2019 Nov 29. pii: bkz083. [Epub ahead of print]

Identification of Suvorexant in Blood Using LC-MS-MS: Important Considerations for Matrix Effects and Quantitative Interferences in Targeted Assays.

Britni Skillman, Sarah Kerrigan.

  Suvorexant (Belsomra®) is a novel dual orexin receptor antagonist used for the treatment of insomnia. The prevalence of suvorexant in forensic samples is relatively unknown, which demonstrates the need for robust analytical assays for the detection of this sedative hypnotic in forensic toxicology laboratories. In this study, suvorexant was isolated from whole blood using a simple acidic/neutral liquid-liquid extraction followed by analysis by liquid chromatography tandem mass spectrometry (LC-MS/MS). Matrix effects were evaluated qualitatively and quantitatively using various extraction solvents, proprietary lipid clean-up devices and source conditions. The method was validated in terms of limit of detection, limit of quantitation, precision, bias, calibration model, carryover, matrix effects and drug interferences. Electrospray is a competitive ionization process whereby compounds in the droplet compete for a limited number of charged sites at the surface. As such, it is capacity-limited, and LC-MS-based techniques must be carefully evaluated to ensure that matrix effects or coeluting drugs do not impact quantitative assay performance. In this report, we describe efforts to ameliorate such effects in the absence of an isotopically labeled internal standard. Matrix effects are highly variable and heavily dependent on the physico-chemical properties of the substance. Although there is no universal solution to their resolution, conditions at the electrospray interface can mitigate these issues. Using this approach, the LC-MS/MS assay was fully validated and limits of detection and quantitation of 0.1 and 0.5 ng/mL suvorexant were achieved in blood.

Keywords:  LC–MS/MS; LC–Q/TOF-MS; Suvorexant; ion suppression; matrix effects

DOI:  https://doi.org/10.1093/jat/bkz083
Sci Total Environ. 2019 Nov 22. pii: S0048-9697(19)35213-1. [Epub ahead of print] 135221

Development, validation, comparison, and implementation of a highly efficient and effective method using magnetic solid-phase extraction with hydrophilic-lipophilic-balanced materials for LC-MS/MS analysis of pesticides in seawater.

Zhenzhen Liu, Peipei Qi, Jiao Wang, Zhiwei Wang, Shanshan Di, Hao Xu, Huiyu Zhao, Qiang Wang, Xinquan Wang, Xinhong Wang.

  To achieve multi-pesticides residue analysis in seawater, hydrophilic-lipophilic-balanced magnetic particles were designed and fabricated by swelling polymerization of divinyl benzene (DVB) and N-vinyl pyrrolidone (NVP) on the surface of Fe3O4@SiO2 magnetic particles. The ratio of DVB to NVP was adjusted to achieve a proper balance in hydrophilicity and lipophilicity. The obtained magnetic particles were systematically characterized by TEM, SEM, FT-IR and vibrating sample magnetization. Based on the optimized magnetic nanoparticles, a sensitive magnetic solid-phase extraction method was developed for the simultaneous pre-concentration and determination of 96-pesticide residues from large-volume seawater samples prior to being detected by liquid chromatography-tandem mass spectrometry. Recoveries of pesticides in spiked seawater samples (0.001, 0.01, 0.1, 1.0 μg L-1) ranged from 62% to 112% with RSDs less than 21%. The method limits of detection of 96 pesticides ranged from 0.13 to 0.42 ng L-1, the method limits of quantification of 96 pesticides ranged from 1.0 to 10 ng L-1. The method was successfully applied to pesticide residue analysis in water samples from Jiulong River Estuary of China, demonstrating the prospects of this technique as a potential method for the rapid determination of trace levels of multi-pesticide residues in seawater.

Keywords:  Divinyl benzene; Liquid chromatography-tandem mass spectrometry; Magnetic particle adsorbents; N-vinyl pyrrolidone; Pesticides residue analysis; Seawater

DOI:  https://doi.org/10.1016/j.scitotenv.2019.135221
Biomed Chromatogr. 2019 Nov 30. e4767

Simultaneous determination of thirteen substances related to NAFLD in mice brain tissue using 3-Aminobutyric acid as internal standard by HPLC-FLD.

Yong-Hong Liu, Ting-Ting Du, Jing-Yuan Wan, Hua Zhao, Dan Huang, Jian-Sha Li, Xin-Hui Jiang.

  Disorders of certain branched-chain amino acids (BCAAs) may be associated with the occurrence and development of non-alcoholic fatty liver disease (NAFLD). Measurement of related BCAAs levels could provide a reference for the clinical and scientific research of the NAFLD. An established HPLC-FLD method was used to quantify Aspartic acid (Asp), Glutamate (Glu), Glutamine (Gln), Glycine (Gly), Taurine (Tau), Tyrosine (Tyr), 4-Amino butanoic acid (GABA), Tryptophan (Trp), Methionine (Met), Valine (Val), Phenylalanine (Phe), Isoleucine (Ile), and Leucine (Leu) in mice brain tissue. Brain tissue samples mixed with internal standard (DL-3-Aminobutyric acid) were processed, then derivatized with 2-O-phthaldialdehyde (OPA), and finally separated on an ODS2 column through gradient elution at a flow rate of 1.0 mL·min-1 . The excitation and emission wavelengths were set at 340 nm and 455 nm respectively. The mobile phase A was 100% methanol and the mobile phase B consisted of 30 mmol·L-1 sodium acetate (pH 6.8). The injection volume was 20 μL and the single run time was 45 min. Several parameters, accuracy, precision, and stability, were verified and the results showed the established method had good sensitivity and resolution for all of the 13 compounds and internal standard (IS) in mice brain.

Keywords:  3-Aminobutyric acid; Aromatic amino acids; Branched chain amino acids; HPLC-FLD; mice brain tissue; nonalcoholic fatty liver disease

DOI:  https://doi.org/10.1002/bmc.4767
Medchemcomm. 2019 Oct 01. 10(10): 1803-1809

Targeted metabolomics approach for identification of relapsing-remitting multiple sclerosis markers and evaluation of diagnostic models.

Marat F Kasakin, Artem D Rogachev, Elena V Predtechenskaya, Vladimir J Zaigraev, Vladimir V Koval, Andrey G Pokrovsky.

Multiple sclerosis (MS) is an inflammatory autoimmune disease that causes demyelination of nerve cell axons. This paper is devoted to the study of relapsing-remitting multiple sclerosis (RRMS) biomarkers using an LC-MS/MS-based targeted metabolomics approach and the assessment of changes in the profile of 13 amino acids and 29 acylcarnitines in plasma during the relapse of the disease. A significant increase (p < 0.05) in the concentration of glutamate in plasma in patients with RRMS was detected, while the sum of leucine and isoleucine was reduced. A decrease in the concentration of decenoylcarnitine (C10:1, p < 0.05) was observed among acylcarnitines, and this metabolite was detected as a biomarker for the disease for the first time. Several models based on a single marker or multiple pre-selected markers and multivariate analysis with a dimension reduction technique were compared in their effectiveness for the classification of RRMS and healthy controls. The best results for cross-validation showed models of general linear regression (GLM, AUC = 0.783) and random forest model (RF, AUC = 0.769) based on pre-selected biomarkers. Validation of the models on the test set showed that the RF model based on selected metabolites was the most effective (AUC = 0.72). The results obtained are promising for further development of the system of clinical decision support for the diagnosis of RRMS based on metabolic data.

DOI: https://doi.org/10.1039/c9md00253g
Mass Spectrom Rev. 2019 Dec 03.

MINIATURIZATION IN MASS SPECTROMETRY.

Przemyslaw Mielczarek, Jerzy Silberring, Marek Smoluch.

  Expectations for continuous miniaturization in mass spectrometry are not declining for years. Portable instruments are highly welcome by the industry, science, space agencies, forensic laboratories, and many other units. All are striving for the small, cheap, and as good as possible instruments. This review describes the recent developments of miniature mass spectrometers and also provides selected applications where these devices are used. Upcoming perspectives of further development are also discussed. @ 2019 Wiley Periodicals, Inc. Mass Spec Rev.

Keywords:  chromatography; hand-held; hyphenated techniques; mass spectrometer; miniature; miniaturization; portable

DOI:  https://doi.org/10.1002/mas.21614
Molecules. 2019 Nov 28. pii: E4347. [Epub ahead of print]24(23):

Sustainable and Selective Extraction of Lipids and Bioactive Compounds from Microalgae.

Ilaria Santoro, Monica Nardi, Cinzia Benincasa, Paola Costanzo, Girolamo Giordano, Antonio Procopio, Giovanni Sindona.

  The procedures for the extraction and separation of lipids and nutraceutics from microalgae using classic solvents have been frequently used over the years. However, these production methods usually require expensive and toxic solvents. Based on our studies involving the use of eco-sustainable methodologies and alternative solvents, we selected ethanol (EtOH) and cyclopentyl methyl ether (CPME) for extracting bio-oil and lipids from algae. Different percentages of EtOH in CPME favor the production of an oil rich in saturated fatty acids (SFA), useful to biofuel production or rich in bioactive compounds. The proposed method for obtaining an extract rich in saturated or unsaturated fatty acids from dry algal biomass is disclosed as eco-friendly and allows a good extraction yield. The method is compared both in extracted oil percentage yield and in extracted fatty acids selectivity to extraction by supercritical carbon dioxide (SC-CO2).

Keywords:  algal oil; bio compound; biofuel; extraction; green chemistry; green solvents

DOI:  https://doi.org/10.3390/molecules24234347
Pac Symp Biocomput. 2020 ;25 587-598

Assessment of coverage for endogenous metabolites and exogenous chemical compounds using an untargeted metabolomics platform.

Sek Won Kong, Carles Hernandez-Ferrer.

Physiological status and pathological changes in an individual can be captured by metabolic state that reflects the influence of both genetic variants and environmental factors such as diet, lifestyle and gut microbiome. The totality of environmental exposure throughout lifetime - i.e., exposome - is difficult to measure with current technologies. However, targeted measurement of exogenous chemicals and untargeted profiling of endogenous metabolites have been widely used to discover biomarkers of pathophysiologic changes and to understand functional impacts of genetic variants. To investigate the coverage of chemical space and interindividual variation related to demographic and pathological conditions, we profiled 169 plasma samples using an untargeted metabolomics platform. On average, 1,009 metabolites were quantified in each individual (range 906 - 1,038) out of 1,244 total chemical compounds detected in our cohort. Of note, age was positively correlated with the total number of detected metabolites in both males and females. Using the robust Qn estimator, we found metabolite outliers in each sample (mean 22, range from 7 to 86). A total of 50 metabolites were outliers in a patient with phenylketonuria including the ones known for phenylalanine pathway suggesting multiple metabolic pathways perturbed in this patient. The largest number of outliers (N=86) was found in a 5-year-old boy with alpha-1-antitrypsin deficiency who were waiting for liver transplantation due to cirrhosis. Xenobiotics including drugs, diets and environmental chemicals were significantly correlated with diverse endogenous metabolites and the use of antibiotics significantly changed gut microbial products detected in host circulation. Several challenges such as annotation of features, reference range and variance for each feature per age group and gender, and population scale reference datasets need to be addressed; however, untargeted metabolomics could be immediately deployed as a biomarker discovery platform and to evaluate the impact of genomic variants and exposures on metabolic pathways for some diseases.
BMC Cancer. 2019 Dec 05. 19(1): 1195

UPLC-MS based urine untargeted metabolomic analyses to differentiate bladder cancer from renal cell carcinoma.

Zhan Wang, Xiaoyan Liu, Xiang Liu, Haidan Sun, Zhengguang Guo, Guoyang Zheng, Yushi Zhang, Wei Sun.

   BACKGROUND: To discover biomarker panels that could distinguish cancers (BC and RCC) from healthy controls (HCs) and bladder cancers (BC) from renal cell carcinoma (RCC), regardless of whether the patients have haematuria. In addition, we also explored the altered metabolomic pathways of BC and RCC.
METHODS: In total, 403 participants were enrolled in our study, which included 146 BC patients (77 without haematuria and 69 with haematuria), 115 RCC patients (94 without haematuria and 21 with haematuria) and 142 sex- and age-matched HCs. Their midstream urine samples were collected and analysed by performing UPLC-MS. The statistical methods and pathway analyses were applied to discover potential biomarker panels and altered metabolic pathways.
RESULTS: The panel of α-CEHC, β-cortolone, deoxyinosine, flunisolide, 11b,17a,21-trihydroxypreg-nenolone and glycerol tripropanoate could distinguish the patients with cancer from the HCs (the AUC was 0.950) and the external validation also displayed a good predictive ability (the AUC was 0.867). The panel of 4-ethoxymethylphenol, prostaglandin F2b, thromboxane B3, hydroxybutyrylcarnitine, 3-hydroxyphloretin and N'-formylkynurenine could differentiate BC from RCC without haematuria. The AUC was 0.829 in the discovering group and 0.76 in the external validation. The metabolite panel comprising 1-hydroxy-2-oxopropyl tetrahydropterin, 1-acetoxy-2-hydroxy-16-heptadecyn-4-one, 1,2-dehydrosalsolinol and L-tyrosine could significantly discriminate BC from RCC with haematuria (AUC was 0.913). Pathway analyses revealed altered lipid and purine metabolisms between cancer patients and HCs, together with disordered amino acid and purine metabolisms between BC and RCC with haematuria.
CONCLUSIONS: UPLC-MS urine metabolomic analyses could not only differentiate cancers from HCs but also discriminate BC from RCC. In addition, pathway analyses demonstrated a deeper metabolic mechanism of BC and RCC.

Keywords:  Biomarkers; Bladder cancer; Metabolomics; Renal cell carcinoma; UPLC-MS

DOI:  https://doi.org/10.1186/s12885-019-6354-1
Math Biosci. 2019 Nov 28. pii: S0025-5564(19)30532-2. [Epub ahead of print] 108291

Experimental design for parameter estimation in steady-state linear models of metabolic networks.

Håvard G Frøysa, Hans J Skaug, Guttorm Alendal.

  Metabolic networks are typically large, containing many metabolites and reactions. Dynamical models that aim to simulate such networks will consist of a large number of ordinary differential equations, with many kinetic parameters that must be estimated from experimental data. We assume these data to be metabolomics measurements made under steady-state conditions for different input fluxes. Assuming linear kinetics, analytical criteria for parameter identifiability are provided. For normally distributed error terms, we also calculate the Fisher information matrix analytically to be used in the D-optimality criterion. A test network illustrates the developed tool chain for finding an optimal experimental design. The first stage is to verify global or pointwise parameter identifiability, the second stage to find optimal input fluxes, and finally remove redundant measurements.

Keywords:  D-optimality; Experimental design; Fisher information; Metabolic networks; Parameter identifiability; Systems biology

DOI:  https://doi.org/10.1016/j.mbs.2019.108291
Rapid Commun Mass Spectrom. 2019 Dec 04.

Ultra-high Performance Supercritical Fluid Chromatography Combined with Quadrupole Time-of-Flight Mass Spectrometry for the Characterization of Pentaerythritol Fatty Acid Esters.

Yan Tang, Jinheng Pan, Tangqiang Sun, Yajing Hu, Zhenxia Du.

RATIONALE: Pentaerythritol fatty acid esters, are excellent lubricant oil additive, for their good biodegradability, thermal ability, anti-wear, and friction. However, in order to meet the application requirements, fatty acids with different alkyl chain lengths are reacted with pentaerythritol, resulting in complex ester compositions. In order to reveal the relation between the functionalities and the composition of esters, it is important to develop a method for their analysis.
METHODS: We developed a method using ultra-high performance supercritical fluid chromatography combined with quadrupole time-of-flight mass spectrometry (UHPSFC-QTof-MS) to separate and characterize pentaerythritol fatty acid esters. This method has the advantages of short analysis time and high separation efficiency for such weakly polar compounds; high resolution MS provides exact mass information, enabling the structure of the pentaerythritol fatty acid esters to be identified.
RESULTS: Based on the exact masses and characteristic ions, the pentaerythritol fatty acid esters were identified. The main fragmentation pathways of pentaerythritol fatty acid esters were identified and the fatty acid composition were deduced from characteristic product ions. Through dihydrogen rearrangement reaction, the neutral fatty acid fragment was los; [M+Na-FA]+ product ions (a stable six-member ring structure) were produced due to the absence of aa γ hydrogen in pentaerythritol fatty acid esters.
CONCLUSIONS: A UHPSFC/QTof-MS method was successfully employed for the separation of pentaerythritol fatty acid esters. Exact masses and product ion information were given by high resolution mass spectrometry. The composition of the fatty acids was effectively deduced by characteristic ions and their relative abundances. This method is an effective means for the quality control and process optimization of this type of product, serving as a positive reference for further study on pentaerythritol fatty acid esters.

DOI: https://doi.org/10.1002/rcm.8664
J Chromatogr A. 2019 Nov 28. pii: S0021-9673(19)31175-6. [Epub ahead of print] 460743

Stir membrane liquid phase microextraction of tetracyclines using switchable hydrophilicity solvents followed by high-performance liquid chromatography.

Sofya Lebedinets, Christina Vakh, Ksenya Cherkashina, Aleksei Pochivalov, Leonid Moskvin, Andrey Bulatov.

  A novel approach for stir membrane liquid phase microextraction of tetracyclines from biological fluids was developed. The microextraction procedure assumed in situ formation of microdroplets of a hydrophobic medium-chain fatty acid (extraction solvent) from homogeneous sample solution containing water-soluble medium-chain fatty acid salt by acidification and simultaneous analytes extraction and organic phase separation into pores of stir membrane disk. Obtained large surface area between the extraction solvent and aqueous phase and high porosity of the membrane provided fast extraction and phase separation (extraction time - 5 min) and reducing extraction solvent volume to the order of several µL. The developed approach was applied for the HPLC-UV determination of tetracycline, oxytetracycline and chlortetracycline in biological fluids samples. The calibration graphs were linear over the concentration ranges of 0.1-100 mg L-1 for oxytetracycline, tetracycline and chlortetracycline. Regression coefficients were in the range from 0.994 to 0.998. The LOD values, calculated from the blank tests based on 3σ, were 30 µg L-1 for tetracycline, oxytetracycline and chlortetracycline. The RSD values expressing intra-day and inter-day repeatability were lower than 5% and 8%, respectively.

Keywords:  Antibiotics; Hexanoic acid; Liquid chromatography; Liquid phase microextraction; Stir membrane disk; Urine

DOI:  https://doi.org/10.1016/j.chroma.2019.460743
Nat Rev Urol. 2019 Dec 06.

Piloting hyperpolarized 13C-pyruvate MRI for imaging advanced prostate cancer.

Louise Stone.

DOI: https://doi.org/10.1038/s41585-019-0269-0
J Anal Toxicol. 2019 Nov 29. pii: bkz081. [Epub ahead of print]

Metabolite Profiling of Ortho-, Meta- and Para-Fluorofentanyl by Hepatocytes and High-Resolution Mass Spectrometry.

Per Ole M Gundersen, Anna Åstrand, Henrik Gréen, Martin Josefsson, Olav Spigset, Svante Vikingsson.

  New psychoactive substances are emerging on the illegal drug market. Synthetic opioids including fentanyl analogues are of special concern due to their high potency. This indicates the possibility of low drug concentrations in vivo and calls for sensitive analytical methods and identification of the most appropriate analytical targets. In this study the in vitro metabolism of ortho-, meta- and para-fluorofentanyl, three fluorinated derivatives of fentanyl, has been investigated using human hepatocytes and compared to the results from an authentic human urine sample. Based on knowledge on the metabolism of similar fentanyl analogues N-dealkylation and hydroxylation was hypothesized to be the most central pathways. The three fluorofentanyl isomers were incubated with pooled human hepatocytes at 1, 3 and 5 h. Liquid chromatography quadrupole time of flight mass spectrometry operating in data-dependent mode was used to analyse the hepatocyte samples, as well as the hydrolysed and non-hydrolysed authentic urine sample. Data were analysed by a targeted approach with a database of potential metabolites. The major metabolite formed in vitro was the N-dealkylation product norfluorofentanyl. In addition various hydroxylated metabolites, a N-oxide, dihydrodiol metabolites and a hydroxymethoxy metabolite were found. In total, 14 different metabolites were identified for each fluorofentanyl isomer. In the authentic urine sample, three metabolites were detected in addition to the ortho-fluorofentanyl parent compound, with hydroxymethoxy metabolite having the highest abundance followed by norfluorofentanyl and a metabolite hydroxylated on the ethylphenyl ring. This in vitro study showed that the metabolic pattern for ortho-, meta-, and para-fluorofentanyl was close to those previously reported for other fentanyl analogues. We suggest that the hydroxymethoxy metabolite and the metabolite hydroxylated on the ethylphenyl ring should be the metabolites primarily investigated in further studies to determine the most appropriate marker for intake of fluorofentanyl derivatives in urine drug screening for human subjects.

Keywords:  Fluorofentanyl; high-resolution mass spectrometry; human hepatocytes; metabolism

DOI:  https://doi.org/10.1093/jat/bkz081