bims-metlip Biomed News
on Methods and protocols in metabolomics and lipidomics
Issue of 2019‒11‒10
twenty-four papers selected by
Sofia Costa
Cold Spring Harbor Laboratory


  1. Metabolites. 2019 Nov 01. pii: E261. [Epub ahead of print]9(11):
    Anesi A, Rubert J, Oluwagbemigun K, Orozco-Ruiz X, Nöthlings U, Breteler MMB, Mattivi F.
      Tryptophan and tyrosine metabolism has a major effect on human health, and disorders have been associated with the development of several pathologies. Recently, gut microbial metabolism was found to be important for maintaining correct physiology. Here, we describe the development and validation of a UHPLC-ESI-MS/MS method for targeted quantification of 39 metabolites related to tryptophan and tyrosine metabolism, branched chain amino acids and gut-derived metabolites in human plasma and urine. Extraction from plasma was optimised using 96-well plates, shown to be effective in removing phospholipids. Urine was filtered and diluted ten-fold. Metabolites were separated with reverse phase chromatography and detected using triple quadrupole MS. Linear ranges (from ppb to ppm) and correlation coefficients (r2 > 0.990) were established for both matrices independently and the method was shown to be linear for all tested metabolites. At medium spiked concentration, recovery was over 80% in both matrices, while analytical precision was excellent (CV < 15%). Matrix effects were minimal and retention time stability was excellent. The applicability of the methods was tested on biological samples, and metabolite concentrations were found to be in agreement with available data. The method allows the analysis of up to 96 samples per day and was demonstrated to be stable for up to three weeks from acquisition.
    Keywords:  LC-MS/MS; branched chain amino acids; clinical studies; gut microbiota metabolites; human plasma; targeted metabolomics; tryptophan metabolism; tyrosine metabolism; urine
    DOI:  https://doi.org/10.3390/metabo9110261
  2. Metabolites. 2019 Nov 07. pii: E270. [Epub ahead of print]9(11):
    Bearden DW, Sheen DA, Simón-Manso Y, Benner BA, Rocha WFC, Blonder N, Lippa KA, Beger RD, Schnackenberg LK, Sun J, Mehta KY, Cheema AK, Gu H, Marupaka R, Nagana Gowda GA, Raftery D.
      There is a lack of experimental reference materials and standards for metabolomics measurements, such as urine, plasma, and other human fluid samples. Reasons include difficulties with supply, distribution, and dissemination of information about the materials. Additionally, there is a long lead time because reference materials need their compositions to be fully characterized with uncertainty, a labor-intensive process for material containing thousands of relevant compounds. Furthermore, data analysis can be hampered by different methods using different software by different vendors. In this work, we propose an alternative implementation of reference materials. Instead of characterizing biological materials based on their composition, we propose using untargeted metabolomic data such as nuclear magnetic resonance (NMR) or gas and liquid chromatography-mass spectrometry (GC-MS and LC-MS) profiles. The profiles are then distributed with the material accompanying the certificate, so that researchers can compare their own metabolomic measurements with the reference profiles. To demonstrate this approach, we conducted an interlaboratory study (ILS) in which seven National Institute of Standards and Technology (NIST) urine Standard Reference Material®s (SRM®s) were distributed to participants, who then returned the metabolomic data to us. We then implemented chemometric methods to analyze the data together to estimate the uncertainties in the current measurement techniques. The participants identified similar patterns in the profiles that distinguished the seven samples. Even when the number of spectral features is substantially different between platforms, a collective analysis still shows significant overlap that allows reliable comparison between participants. Our results show that a urine suite such as that used in this ILS could be employed for testing and harmonization among different platforms. A limited quantity of test materials will be made available for researchers who are willing to repeat the protocols presented here and contribute their data.
    Keywords:  chromatography; interlaboratory study; nuclear magnetic resonance; principal components analysis; reproducibility
    DOI:  https://doi.org/10.3390/metabo9110270
  3. J Chromatogr B Analyt Technol Biomed Life Sci. 2019 Oct 22. pii: S1570-0232(19)31210-3. [Epub ahead of print]1132 121819
    Ferré S, González-Ruiz V, Guillarme D, Rudaz S.
      This review describes the analytical methods that have been developed over the years to tackle the high polarity and non-chromophoric nature of amino acids (AAs). First, the historical methods are briefly presented, with a strong focus on the use of derivatization reagents to make AAs detectable with spectroscopic techniques (ultraviolet and fluorescence) and/or sufficiently retained in reversed phase liquid chromatography. Then, an overview of the current analytical strategies for achiral separation of AAs is provided, in which mass spectrometry (MS) becomes the most widely used detection mode in combination with innovative liquid chromatography or capillary electrophoresis conditions to detect AAs at very low concentration in complex matrixes. Finally, some future trends of AA analysis are provided in the last section of the review, including the use of supercritical fluid chromatography (SFC), multidimensional liquid chromatography and electrophoretic separations, hyphenation of ion exchange chromatography to mass spectrometry, and use of ion mobility spectrometry mass spectrometry (IM-MS). Various application examples will also be presented throughout the review to highlight the benefits and limitations of these different analytical approaches for AAs determination.
    Keywords:  Amino acids; Capillary electrophoresis; Derivatization; Liquid chromatography; Mass spectrometry
    DOI:  https://doi.org/10.1016/j.jchromb.2019.121819
  4. Metabolites. 2019 Nov 01. pii: E259. [Epub ahead of print]9(11):
    Sun Y, Saito K, Saito Y.
      Extracellular vesicles (EVs) consist of lipid bilayers, occur in various biofluids, and are invaluable in biomarker screening. Liquid chromatography coupled with high-resolution mass spectrometry (LC-MS) was recently used to study comprehensive EV lipid profiles in vitro. The aim of this study was to establish a lipidomics platform for human plasma and serum EVs for comprehensive characterization of their lipid profiles, and to compare them with those of other lipid-containing particles, such as high-density lipoproteins (HDL), and low/very low-density lipoproteins (LDL/VLDL). Isolation was validated by specific protein markers; CD9 and MHC class for EVs, apoA-I for HDL, and apoB-100 for LDL/VLDL. Lipidomics identified 264 lipids from isolated plasma EVs, HDL, and LDL/VLDL. The absolute lipid levels per unit protein content in the EVs were more than eight times lower than those of the lipoproteins. Moreover, the EVs had higher lysoglycerophospholipid levels than HDL or LDL/VLDL. Similar profiles were also determined for human serum. The present study found that the lipid profiles of EVs are unique and distinctly different from those of lipoproteins. The lipidomics platform applied to human plasma and serum EVs could generate important information for the exploration and qualification of biomarkers in disease diagnosis.
    Keywords:  extracellular vesicles; lipidomics; lipoproteins; mass spectrometry
    DOI:  https://doi.org/10.3390/metabo9110259
  5. J Chromatogr A. 2019 Oct 31. pii: S0021-9673(19)31086-6. [Epub ahead of print] 460664
    Martín-Ortiz A, Carrero-Carralero C, Hernández-Hernández O, Lebrón-Aguilar R, Moreno FJ, Sanz ML, Ruiz-Matute AI.
      Oligosaccharides are gaining importance because of their beneficial properties in human health. They normally appear in natural and synthetic products as complex mixtures of different monomeric units, glycosidic linkages and degrees of polymerization, being disaccharides and trisaccharides usually the most abundant ones. Although liquid chromatography-mass spectrometry is a useful technique for oligosaccharides analysis, the similarity of their structures makes difficult their characterization. Moreover, there is still scarce information about the relationship between carbohydrate chemical structure, mass spectra and chromatographic data. Then, in this work, chromatographic parameters for 23 disaccharides with different linkages and monomeric units (glucose, galactose, mannose and fructose) were determined using porous graphitized and hydrophilic interaction liquid chromatography columns. Moreover, diagnostic ions of these disaccharides obtained by tandem mass spectra (MS2) were established by stepwise linear discriminant analysis. The relationship between carbohydrate chemical structure and their chromatographic retention data and characteristic ions obtained by multiple-stage mass spectrometry (MSn) was successful in establishing some specific criteria that allowed the characterization of trisaccharides with different structural features.
    Keywords:  Diagnostic fragment ions; Disaccharides; Glycosidic linkage; Liquid chromatography-multiple-stage mass spectrometry; Monomeric units; Trisaccharides
    DOI:  https://doi.org/10.1016/j.chroma.2019.460664
  6. J Steroid Biochem Mol Biol. 2019 Nov 04. pii: S0960-0760(19)30403-0. [Epub ahead of print] 105520
    Gomez-Gomez A, Miranda J, Feixas G, Arranz Betegon A, Crispi F, Gratacós E, Pozo OJ.
      The simultaneous determination of a broad panel of steroids provides more accurate information about the hormonal status than the detection of a single hormone. For that reason, the determination of the steroid profile, i.e. the endogenous steroid hormones and their main metabolites, has become the most powerful tool for the study of hormonal imbalances. The usefulness of the evaluation of the steroid profile in urine and plasma is widely accepted. However, despite its broad potential applicability, the evaluation of the whole steroid profile in alternative matrices such as amniotic fluid, saliva and breast milk remains almost unexplored. In this research we developed and validated a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the quantification of several steroids and their metabolites in amniotic fluid (28 analytes), saliva (15) and breast milk (12). Sample preparation, chromatographic conditions and mass spectrometric conditions (e.g. ionization species or ion source parameters) were optimized. The method was shown to be linear in the range of endogenous concentrations for all studied metabolites. Intra- and inter-assay accuracies were between 80% and 120% while intra- and inter-precisions were below 20% for all analytes in all matrices. The applicability of the method was evaluated by the comparison between the concentration ranges obtained in healthy volunteers (n = 30 per matrix) and the scarce data previously reported in literature. The concentration ranges for several analytes are reported for the first time. The present methodology represents a useful tool for the comprehensive evaluation of the steroid profile in alternative matrices and can be applicable for different clinical purposes.
    Keywords:  Alternative matrices; Amniotic fluid.; Breast milk; LC-MS/MS; Saliva; Steroid profile
    DOI:  https://doi.org/10.1016/j.jsbmb.2019.105520
  7. J Clin Exp Hepatol. 2019 Sep-Oct;9(5):9(5): 597-606
    Alsaleh M, Barbera TA, Andrews RH, Sithithaworn P, Khuntikeo N, Loilome W, Yongvanit P, Cox IJ, Syms RRA, Holmes E, Taylor-Robinson SD.
      Metabolic profiling, metabonomics and metabolomics are terms coined in the late 1990s as they emerged as the newest 'omics' technology at the time. This line of research enquiry uses spectroscopic analytical platforms, which are mainly nuclear magnetic resonance spectroscopy and mass spectrometry (MS), to acquire a snapshot of metabolites, the end products of a complex biological system. Metabolic profiling enables the detection, quantification and characterisation of metabolites in biofluids, cells and tissues. The source of these compounds can be of endogenous, microbial or exogenous origin, such as dietary or xenobiotic. This results in generating extensive, multivariate spectroscopic data that require specific statistical manipulation, typically performed using chemometric and pattern recognition techniques to reduce its dimensions, facilitate its biological interpretation and allow sample classification and biomarker discovery. Consequently, it is possible to study the dynamic metabolic changes in response to disease, intervention or environmental conditions. In this review, we describe the fundamentals of MS so that clinicians can be literate in the field and are able to interrogate the right scientific questions.
    Keywords:  CID, collision-induced dissociation; DC, direct current; ESI, electrospray ionisation; FC, fold change; GC, gas chromatography; HILIC, hydrophilic interaction liquid chromatography; LC, liquid chromatography; MS, mass spectrometry; MWA, metabolome-wide association; NMR, nuclear magnetic resonance; OPLS-DA, orthogonal partial least squared-discriminant analysis; PC, principal component; PCA, principal components analysis; Q-TOF, quadrupole coupled with time-of-flight; RF, radio frequency; RP, reversed-phase; UPLC, ultra-performance liquid chromatography; VIP, variable importance of projection; mass spectroscopy; mass-charge ratio; metabolic profiling; metabolomics; targeted profiling
    DOI:  https://doi.org/10.1016/j.jceh.2019.04.053
  8. J Chromatogr B Analyt Technol Biomed Life Sci. 2019 Sep 09. pii: S1570-0232(19)30460-X. [Epub ahead of print]1132 121792
    Li J, Wang QL, Liu Y, Ke Y, Fan QQ, Zhou P, An MC, Liu HM.
      Amino acids play key roles in cellular protein biosynthesis and energy metabolism pathways. In this study, a simple, rapid and sensitive method was developed for the simultaneous determination of 24 free amino acids in cell samples using hydrophilic interaction liquid chromatography coupled to tandem mass spectrometry (HILIC-MS/MS). Cell samples were deproteinized with methanol/H2O (80:20, v/v) without intricate derivatization process. The analytes were separated on a Waters BEH Amide column (2.1 mm × 100 mm, 1.7 μm), and accomplished within 5 min at a flow rate of 0.2 mL/min. The good linearity was obtained for all analytes (r2 > 0.99) with the limits of quantification from 0.1 to 25 ng/mL. The intra- and inter-day precision ranged from 0.35 to 10.36% and from 2.22 to 9.93%, respectively. The recoveries of most analytes were between 80% and 120% with RSD less than 10.0%. The developed method was then applied to the direct analysis of 24 underivatized amino acids in human gastric cancer cell line MGC803 treated with the antitumor candidate drug J3, and significant differences in the concentration levels of amino acids were also assessed.
    Keywords:  Free amino acids; Hydrophilic interaction liquid chromatography tandem mass spectrometry; MGC803 cells; Method validation
    DOI:  https://doi.org/10.1016/j.jchromb.2019.121792
  9. Metabolites. 2019 Oct 31. pii: E257. [Epub ahead of print]9(11):
    Izumi Y, Matsuda F, Hirayama A, Ikeda K, Kita Y, Horie K, Saigusa D, Saito K, Sawada Y, Nakanishi H, Okahashi N, Takahashi M, Nakao M, Hata K, Hoshi Y, Morihara M, Tanabe K, Bamba T, Oda Y.
      BACKGROUND: One of the current problems in the field of metabolomics is the difficulty in integrating data collected using different equipment at different facilities, because many metabolomic methods have been developed independently and are unique to each laboratory.METHODS: In this study, we examined whether different analytical methods among 12 different laboratories provided comparable relative quantification data for certain metabolites. Identical samples extracted from two cell lines (HT-29 and AsPc-1) were distributed to each facility, and hydrophilic and hydrophobic metabolite analyses were performed using the daily routine protocols of each laboratory.
    RESULTS: The results indicate that there was no difference in the relative quantitative data (HT-29/AsPc-1) for about half of the measured metabolites among the laboratories and assay methods. Data review also revealed that errors in relative quantification were derived from issues such as erroneous peak identification, insufficient peak separation, a difference in detection sensitivity, derivatization reactions, and extraction solvent interference.
    CONCLUSION: The results indicated that relative quantification data obtained at different facilities and at different times would be integrated and compared by using a reference materials shared for data normalization.
    Keywords:  data integration; inter-laboratory comparison; metabolomics; method validation; quality control sample; relative quantification
    DOI:  https://doi.org/10.3390/metabo9110257
  10. Rapid Commun Mass Spectrom. 2019 Nov 08.
    Li Y, Li L.
      RATIONALE: In chemical isotope labeling (CIL) LC/MS metabolome analysis, the peak pairs of the same metabolite detected from different samples are aligned according to their mass and retention time (RT). Any RT shift of a peak pair in one of the sample files that falls outside the tolerance window will result in misalignment of the pair as a different metabolite. Thus, determination and correction of any significant RT shift are important to ensure the generation of high-quality metabolome results.METHODS: In CIL LC/MS, the heavy-isotope-labeled pooled sample is spiked into all light-isotope-labeled individual samples. As a result, in the analysis of labeled samples of the same type, many common metabolites are detectable with high intensity in all LC/MS runs. We have developed a method to select a few of these metabolites as internal RT reference markers to check the occurrence of any RT shift in a LC/MS run. If significant shift is found, an expanded list of these markers with their RT values covering the entire LC retention time window is selected to serve as internal RT calibrants to re-calibrate the chromatogram to correct any RT shift.
    RESULTS: We wrote a software program in R to perform RT check (RTC) and re-calibration (RT-calib). This program can quickly determine the occurrence of any RT shift falling outside a user-defined threshold in a LC/MS run, thereby triggering a timely intervention to correct the problem (e.g., fixing a small leak or changing a column). In the analysis of 278 dansylation LC/MS runs of human urine samples, we show that the RT values can be corrected to be within a 30-s window.
    CONCLUSIONS: A RT-check method and program tailored to CIL LC/MS metabolome analysis have been developed for quick detection and correction of RT shifts during the course of running many metabolome samples.
    DOI:  https://doi.org/10.1002/rcm.8643
  11. Metabolites. 2019 Oct 31. pii: E256. [Epub ahead of print]9(11):
    Petrova I, Xu S, Joesten WC, Ni S, Kennedy MA.
      Metabolic profiling of cell line and tissue extracts involves sample processing that includes a drying step prior to re-dissolving the cell or tissue extracts in a buffer for analysis by GC/LC-MS or NMR. Two of the most commonly used drying techniques are centrifugal evaporation under vacuum (SpeedVac) and lyophilization. Here, NMR spectroscopy was used to determine how the metabolic profiles of hydrophilic extracts of three human pancreatic cancer cell lines, MiaPaCa-2, Panc-1 and AsPC-1, were influenced by the choice of drying technique. In each of the three cell lines, 40-50 metabolites were identified as having statistically significant differences in abundance in redissolved extract samples depending on the drying technique used during sample preparation. In addition to these differences, some metabolites were only present in the lyophilized samples, for example, n-methyl-α-aminoisobutyric acid, n-methylnicotimamide, sarcosine and 3-hydroxyisovaleric acid, whereas some metabolites were only present in SpeedVac dried samples, for example, trimethylamine. This research demonstrates that the choice of drying technique used during the preparation of samples of human cell lines or tissue extracts can significantly influence the observed metabolome, making it important to carefully consider the selection of a drying method prior to preparation of such samples for metabolic profiling.
    Keywords:  AsPC-1; MiaPaCa-2; NMR; Panc-1; cell line; human cell line; metabolic profiling; metabolomics; metabonomics; nuclear magnetic resonance spectroscopy
    DOI:  https://doi.org/10.3390/metabo9110256
  12. J Chromatogr B Analyt Technol Biomed Life Sci. 2019 Oct 21. pii: S1570-0232(19)30809-8. [Epub ahead of print]1132 121844
    Jolibois J, Schmitt A, Royer B.
      Targeted therapies such as cabozantinib (CABO), pazopanib (PAZO), sorafenib (SORA), sunitinib (SUNI) and its main active metabolite N-desethyl-sunitinib (DST-SUNI), olaparib (OLA) and palbociclib (PALBO) display large pharmacokinetics variability impacting their responses in terms of efficacy or toxicity. For the monitoring of these drugs, an analytical method allowing to routinely measure their concentrations in human plasma is needed. Such a method has been developed and validated and is presented here. The chromatographic separation is achieved on a Zorbax Bonus-RP analytical column using an isocratic elution of 92% V/V of acetonitrile and 8% of water in 0.1% formic acid at a flow rate of 500 µl/min for 0.5 min and then 300 µl/min for 2 min. After a liquid-liquid extraction of plasma samples, a step of filtration is performed. This method was validated based on the EMA and French committee of accreditation guidelines. The analysis time is 2.5 min per run, and all analytes eluted within 0.53-1.61 min. The standard curves are linear over the range from 1 to 380 ng/ml for SUNI; from 4.3 to 450 ng/ml for DST-SUNI; from 6 to 1000 ng/ml for PALBO; from 75 to 5000 ng/ml for CABO, from 0.17 to 20 µg/ml for OLA; from 0.35 to 40 µg/ml for SORA and from 1.7 to 200 µg/ml for PAZO. The method also showed satisfactory results in terms precision (below 9.5% for within-run and below 13% for between-run) and accuracy (below 13.5% for within-run and below 14% for between-run). After sampling, all the compounds are stable in whole blood at ambient temperature at least for 6 h and plasma are stable for 48 h at ambient temperature or 4 °C. The method presented here allows to measure the concentrations of 7 targeted therapies in a routine setting. We moreover present here a method that is, to our knowledge, one of the first detailed method aimed at the measurement of palbociclib in human plasma in a routine setting, together with data useful for the management of samples in routine hospital practice.
    Keywords:  Anticancer targeted therapies; LC–MS/MS; Routine analysis; Whole blood stability
    DOI:  https://doi.org/10.1016/j.jchromb.2019.121844
  13. J Pharm Biomed Anal. 2019 Oct 30. pii: S0731-7085(19)31970-3. [Epub ahead of print] 112956
    Liao HW, Kuo CH, Chao HC, Chen GY.
      Various lipidomics studies have revealed the potential of using phospholipids as disease biomarkers for conditions such as Alzheimer's disease, cancer, and sepsis. Establishing accurate quantification methods for targeted phospholipid analysis is important for making these potential markers more clinically relevant. Although a stable isotope labelled-internal standard method can provide good quantification accuracy for endogenous metabolite quantification, there are limited isotope labelled phosphatidylcholines (PCs) commercially available. For this reason, this study proposed a postcolumn infused-internal standard (PCI-IS) method for the accurate quantification of PCs. To demonstrate the quantification accuracy of the PCI-IS method combined with the matrix normalization factor (MNF), 2 LPCs and 6 PCs have been quantified in the human plasma specimens, and the results showed that the PCI-IS combined with MNF method can provide quantification results as accurate as those of the standard addition method (SAM) but without the need for the labor-intensive SAM procedure. We additionally applied the PCI-IS method for improving the PC profiling accuracy, and the results indicated that the biased estimation of the PC composition caused by the MEs can be resolved by PCI-IS correction. Finally, the method was applied to investigate drug resistance in lung cancer cells. Decreased levels of PCs in drug resistant cells disclose the potential role of PCs in drug resistance. We anticipate that the PCI-IS strategy could help quantitative lipidomics move forward and further contribute to various clinical and biomedical studies.
    Keywords:  LC-ESI-MS; Lipidomics; Matrix normalization factor (MNF); Phosphatidylcholine; Postcolumn infused-internal standard (PCI-IS)
    DOI:  https://doi.org/10.1016/j.jpba.2019.112956
  14. Mol Cell Proteomics. 2019 Nov 07. pii: mcp.RA119.001714. [Epub ahead of print]
    Barkovits K, Pacharra S, Pfeiffer K, Steinbach S, Eisenacher M, Marcus K, Uszkoreit J.
      Currently data-dependent acquisition (DDA) is the method of choice for mass spectrometry-based proteomics discovery experiments, but data-independent acquisition (DIA) is steadily becoming more important. One of the most important requirements to perform a DIA analysis is the availability of suitable spectral libraries for peptide identification and quantification. Several studies were performed addressing the evaluation of spectral library performance for protein identification in DIA measurements. But so far only few experiments estimate the effect of these libraries on the quantitative level.In this work we created a gold standard spike-in sample set with known contents and ratios of proteins in a complex protein matrix that allowed a detailed comparison of DIA quantification data obtained with different spectral library approaches. We utilized in-house generated sample-specific spectral libraries created using varying sample preparation approaches and repeated DDA measurement. In addition, two different search engines were tested for protein identification from DDA data and subsequent library generation. In total, eight different spectral libraries were generated and the quantification results compared to a library free method, as well as a default DDA analysis. Not only the number of identifications on peptide and protein level in the spectral libraries and the corresponding DIA analysis results was inspected, but also the number of expected and identified differentially abundant protein groups and their ratios.We found, that while libraries of prefractionated samples were generally larger, there was no significant increase in DIA identifications compared to repetitive non-fractionated measurements. Furthermore, we show that the accuracy of the quantification is strongly dependent on the applied spectral library and whether the quantification is based on peptide or protein level. Overall, the reproducibility and accuracy of DIA quantification is superior to DDA in all applied approaches.Data has been deposited to the ProteomeXchange repository with identifiers PXD012986, PXD012987, PXD012988 and PXD014956.
    Keywords:  Bioinformatics software; Label-free quantification; Mass Spectrometry; Quantification; Target identification; data-independent acquisition (DIA); peptide identification; proteomics; spectral library
    DOI:  https://doi.org/10.1074/mcp.RA119.001714
  15. BMC Bioinformatics. 2019 Nov 06. 20(1): 549
    Li T, Chen L, Gan M.
      BACKGROUND: Mass spectra are usually acquired from the Liquid Chromatography-Mass Spectrometry (LC-MS) analysis for isotope labeled proteomics experiments. In such experiments, the mass profiles of labeled (heavy) and unlabeled (light) peptide pairs are represented by isotope clusters (2D or 3D) that provide valuable information about the studied biological samples in different conditions. The core task of quality control in quantitative LC-MS experiment is to filter out low-quality peptides with questionable profiles. The commonly used methods for this problem are the classification approaches. However, the data imbalance problems in previous control methods are often ignored or mishandled. In this study, we introduced a quality control framework based on the extreme gradient boosting machine (XGBoost), and carefully addressed the imbalanced data problem in this framework.RESULTS: In the XGBoost based framework, we suggest the application of the Synthetic minority over-sampling technique (SMOTE) to re-balance data and use the balanced data to train the boosted trees as the classifier. Then the classifier is applied to other data for the peptide quality assessment. Experimental results show that our proposed framework increases the reliability of peptide heavy-light ratio estimation significantly.
    CONCLUSIONS: Our results indicate that this framework is a powerful method for the peptide quality assessment. For the feature extraction part, the extracted ion chromatogram (XIC) based features contribute to the peptide quality assessment. To solve the imbalanced data problem, SMOTE brings a much better classification performance. Finally, the XGBoost is capable for the peptide quality control. Overall, our proposed framework provides reliable results for the further proteomics studies.
    Keywords:  Gradient Boosting; Imbalanced Data; Mass Spectra; Proteomics; Quality Control
    DOI:  https://doi.org/10.1186/s12859-019-3170-1
  16. Angew Chem Int Ed Engl. 2019 Nov 07.
    Pawliszyn J, Vuckovic D, Bojko B, Napylov A, Reyes Garces N, Gomez-Rios G, Olkowicz M, Lendor S, Monnin C, Hamani C.
      Oxylipins are key lipid mediators of important brain processes, including pain, sleep, oxidative stress and inflammation. For the first-time, an in-depth profile of up to 52 oxylipins can be obtained from the brains of awake moving animals using in vivo solid-phase microextraction (SPME) chemical biopsy tool in combination with liquid chromatography - high resolution mass spectrometry. Among these, 23 oxylipins are detectable in the majority of healthy wildtype samples. This new approach successfully eliminates the changes in oxylipin concentrations routinely observed during the analysis of post-mortem samples, allows time-course monitoring of their concentrations with high spatial resolution in specific brain regions of interest and can be performed using the same experimental set-up as in vivo microdialysis (MD) thus providing a new and exciting tool in neuroscience and drug discovery.
    Keywords:  Oxylipins, in-vivo sampling, solid phase micro extraction, brain, chemical biopsy, unstable compounds
    DOI:  https://doi.org/10.1002/anie.201909430
  17. J Phys Chem A. 2019 Nov 04.
    Kusaka Y, Hasegawa T, Kaji H.
      A noise reduction method was developed for solid-state nuclear magnetic resonance spectroscopy using multivariate analysis. Principal component analysis was first applied for cross polarization/magic angle spinning and 13C spin-lattice relaxation measurements of solid-state nuclear magnetic resonance array spectra. The contact time of cross polarization/magic angle spinning and the delay time in spin-lattice relaxation measurements were continuously changed to obtain a series of spectra, which were used for noise reduction using principal component analysis. The noise reduction method successfully produced spectra with improved signal-to-noise ratios. This noise reduction method shortens the measurement time and allows for detection of components with minute signals.
    DOI:  https://doi.org/10.1021/acs.jpca.9b04437
  18. J Chromatogr A. 2019 Oct 24. pii: S0021-9673(19)31077-5. [Epub ahead of print] 460655
    McCalley DV.
      For a wide variety of hydrophilic interaction chromatography stationary phases, a repeatable partial equilibration was demonstrated in gradient elution after purging with as little as 12 column volumes of mobile phase. Relative standard deviations of retention time of on average ~0.15% could be obtained after 1 or 2 conditioning (blank) runs. The equilibration period must be kept strictly constant, otherwise selectivity changes occur, but this is not problematic on modern instruments. Partial equilibration was largely independent of stationary phase or gradient slope. Alternatively, full column equilibration is favoured for stationary phases that do not trap extensive water layers, and for materials with a wider pore size that have a lower surface area. Temperatures somewhat above ambient also shorten the equilibration time. Some stationary phases under optimum conditions can achieve full column equilibration using purging with ~12 column volumes, which is useful for rapid set-up of isocratic separations or for conventional gradient analysis.
    Keywords:  Equilibration; HILIC; Hydrophilic interaction chromatography
    DOI:  https://doi.org/10.1016/j.chroma.2019.460655
  19. Anal Chem. 2019 Nov 06.
    Foreman D, McLuckey SA.
      While the study of reactions between oppositely-charged ions has a long history, particularly within the context of plasmas and interstellar chemistry, the exploitation of such reactions for analytical applications was made possible with the advent of electrospray ionization (ESI). , ESI provides a means for generating abundant multiply-charged ions from a wide variety of molecular classes. It was then possible to use ESI to generate multiply-charged analyte or reagent ions for reactions with ions of opposite polarities without resulting in complete neutralization. The first description of ion/ion reaction studies using ESI was given by Loo et al. using a Y-tube reactor leading to the inlet of an atmosphere/vacuum interface coupled with a quadrupole mass filter. , This was an early example of ion/ion chemistry being effected prior to sampling ions into a mass spectrometer. The first implementation of ion/ion reactions within the context of a MSn experiment was demonstrated at Oak Ridge National Lab. , This was accomplished through the use of electrodynamic ion traps, which allow ions of opposite polarity to be stored in overlapping regions of space. Since those early reports, the use of ion/ion reactions, both in vacuo and ex vacuo, within the context of molecular analysis has expanded significantly. A number of characteristics make ion/ion reactions attractive for analytical mass spectrometry. For example, given the long-range Coulomb attraction associated with oppositely-charged ions, the cross-sections for ion/ion collisions are very large such that reactions can be driven on the millisecond to sub-millisecond time scales, depending upon ion densities and extents of ion overlap. Another important characteristic is that mutual neutralization is highly exothermic for virtually all combinations of oppositely-charged gas-phase ion/ion reactions. Hence, ion/ion reactions are highly efficient and always result in some kind of reaction. Given the diversity of ion types that can be generated by the suite of ionization methods now available, the range of ion/ion reactions that can be effected is extremely large, even larger than that of ion/molecule reactions, which are limited by volatility constraints. In the case of ion/ion reactions that take place within a tandem mass spectrometer, such as an ion trap or hybrid instrument, the fact that ions are readily manipulated on a time-dependent basis (i.e., they can be selected or ejected either selectively or non-selectively) enables a high degree of control over the identities of the reactants and the times over which they are exposed to one another. The attractive characteristics of ion/ion reactions for multiple analytical applications have been illustrated in reports going back to the original works described above. Many of these have been reviewed previously. - In this review, we emphasize developments that have taken place largely within the past decade with particular emphasis on the last five years. These have included developments in tools used to implement ion/ion reactions for one or more types of mass spectrometry experiments, an expansion in the range of ion/ion reaction types, and the growth in analytical applications of ion/ion chemistry. The review is organized with descriptions of instrument development, proton transfer chemistry, electron transfer chemistry, and reactions that can proceed only through long-lived complexes.
    DOI:  https://doi.org/10.1021/acs.analchem.9b05014
  20. BMC Bioinformatics. 2019 Nov 04. 20(1): 546
    Ma J, Shojaie A, Michailidis G.
      BACKGROUND: Pathway enrichment extensively used in the analysis of Omics data for gaining biological insights into the functional roles of pre-defined subsets of genes, proteins and metabolites. A large number of methods have been proposed in the literature for this task. The vast majority of these methods use as input expression levels of the biomolecules under study together with their membership in pathways of interest. The latest generation of pathway enrichment methods also leverages information on the topology of the underlying pathways, which as evidence from their evaluation reveals, lead to improved sensitivity and specificity. Nevertheless, a systematic empirical comparison of such methods is still lacking, making selection of the most suitable method for a specific experimental setting challenging. This comparative study of nine network-based methods for pathway enrichment analysis aims to provide a systematic evaluation of their performance based on three real data sets with different number of features (genes/metabolites) and number of samples.RESULTS: The findings highlight both methodological and empirical differences across the nine methods. In particular, certain methods assess pathway enrichment due to differences both across expression levels and in the strength of the interconnectedness of the members of the pathway, while others only leverage differential expression levels. In the more challenging setting involving a metabolomics data set, the results show that methods that utilize both pieces of information (with NetGSA being a prototypical one) exhibit superior statistical power in detecting pathway enrichment.
    CONCLUSION: The analysis reveals that a number of methods perform equally well when testing large size pathways, which is the case with genomic data. On the other hand, NetGSA that takes into consideration both differential expression of the biomolecules in the pathway, as well as changes in the topology exhibits a superior performance when testing small size pathways, which is usually the case for metabolomics data.
    Keywords:  Differential network biology; Pathway enrichment analysis; Pathway topology; Power; Type I error
    DOI:  https://doi.org/10.1186/s12859-019-3146-1
  21. Nat Methods. 2019 Nov 04.
    Morton JT, Aksenov AA, Nothias LF, Foulds JR, Quinn RA, Badri MH, Swenson TL, Van Goethem MW, Northen TR, Vazquez-Baeza Y, Wang M, Bokulich NA, Watters A, Song SJ, Bonneau R, Dorrestein PC, Knight R.
      Integrating multiomics datasets is critical for microbiome research; however, inferring interactions across omics datasets has multiple statistical challenges. We solve this problem by using neural networks ( https://github.com/biocore/mmvec ) to estimate the conditional probability that each molecule is present given the presence of a specific microorganism. We show with known environmental (desert soil biocrust wetting) and clinical (cystic fibrosis lung) examples, our ability to recover microbe-metabolite relationships, and demonstrate how the method can discover relationships between microbially produced metabolites and inflammatory bowel disease.
    DOI:  https://doi.org/10.1038/s41592-019-0616-3
  22. J Chromatogr Sci. 2019 Nov 04. pii: bmz080. [Epub ahead of print]
    Shi Z, Huai Q, Li X, Ma H, Zhou C, Chu X, Zhang H.
      In this paper, counter current salting-out homogenous liquid-liquid extraction was combined with dispersive liquid-liquid microextraction for the determination of environmental estrogens in water samples by high-performance liquid chromatography. In this method, initially, sodium chloride was filled into a syringe and a mixture of water sample and acetonitrile was driven to pass through the syringe. Due to salting-out effect, fine droplets of acetonitrile went up through the remaining mixture and aggregated as a separated layer on the top. Then, the collected organic phase (acetonitrile) was removed with a syringe and mixed with carbon tetrachloride (extraction solvent). In the second step, the mixed organic phase was rapidly injected into 5 mL of distilled water to further enrich the analytes. Good linearity was obtained in the concentration range of 2.0~200 ng/mL for diethylstilbestrol (DES) and 8.0~200 ng/mL for octylphenol (OP), respectively. Limits of detection were 0.09 ng/mL for DES and 0.20 ng/mL for OP, respectively. Relative standard deviations for intra- and inter-day precisions were less than 2.1 and 3.1%, respectively. Finally, the established method was successfully applied to determine DES and OP in river water, well water, bottled water and campus drinking water samples with recoveries in the range from 81.0 to 105.9%.
    DOI:  https://doi.org/10.1093/chromsci/bmz080
  23. Plant Methods. 2019 ;15 126
    Brunoni F, Collani S, Šimura J, Schmid M, Bellini C, Ljung K.
      Background: Plants rely on concentration gradients of the native auxin, indole-3-acetic acid (IAA), to modulate plant growth and development. Both metabolic and transport processes participate in the dynamic regulation of IAA homeostasis. Free IAA levels can be reduced by inactivation mechanisms, such as conjugation and degradation. IAA can be conjugated via ester linkage to glucose, or via amide linkage to amino acids, and degraded via oxidation. Members of the UDP glucosyl transferase (UGT) family catalyze the conversion of IAA to indole-3-acetyl-1-glucosyl ester (IAGlc); by contrast, IAA is irreversibly converted to indole-3-acetyl-l-aspartic acid (IAAsp) and indole-3-acetyl glutamic acid (IAGlu) by Group II of the GRETCHEN HAGEN3 (GH3) family of acyl amido synthetases. Dioxygenase for auxin oxidation (DAO) irreversibly oxidizes IAA to oxindole-3-acetic acid (oxIAA) and, in turn, oxIAA can be further glucosylated to oxindole-3-acetyl-1-glucosyl ester (oxIAGlc) by UGTs. These metabolic pathways have been identified based on mutant analyses, in vitro activity measurements, and in planta feeding assays. In vitro assays for studying protein activity are based on producing Arabidopsis enzymes in a recombinant form in bacteria or yeast followed by recombinant protein purification. However, the need to extract and purify the recombinant proteins represents a major obstacle when performing in vitro assays.Results: In this work we report a rapid, reproducible and cheap method to screen the enzymatic activity of recombinant proteins that are known to inactivate IAA. The enzymatic reactions are carried out directly in bacteria that produce the recombinant protein. The enzymatic products can be measured by direct injection of a small supernatant fraction from the bacterial culture on ultrahigh-performance liquid chromatography coupled to electrospray ionization tandem spectrometry (UHPLC-ESI-MS/MS). Experimental procedures were optimized for testing the activity of different classes of IAA-modifying enzymes without the need to purify recombinant protein.
    Conclusions: This new method represents an alternative to existing in vitro assays. It can be applied to the analysis of IAA metabolites that are produced upon supplementation of substrate to engineered bacterial cultures and can be used for a rapid screening of orthologous candidate genes from non-model species.
    Keywords:  Arabidopsis; Conjugation; Degradation; Enzyme assay; IAA metabolism; UHPLC–ESI-MS/MS
    DOI:  https://doi.org/10.1186/s13007-019-0509-6
  24. Crit Rev Anal Chem. 2019 Nov 07. 1-12
    Hamidi S, Taghvimi A, Mazouchi N.
      Metal organic frameworks (MOFs) are classified as metal clusters with high surface area, commutable structure and pore size, chemical and thermal stability with wide applications in different scientific subjects. Designing of novel adsorbents for sample preparation methods attends high attention towards MOFs and they are good candidates for this purpose. In the present review, recently applied MOF-based materials as micro solid phase extraction technique adsorbents for extraction of environmental, food, and biological samples are comprehensively overviewed.
    Keywords:  Dispersive micro solid phase extraction; metal-organic frameworks; micro solid phase extraction
    DOI:  https://doi.org/10.1080/10408347.2019.1684235