Summary of Study ST001613
This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001033. The data can be accessed directly via it's Project DOI: 10.21228/M85M45 This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
Study ID | ST001613 |
Study Title | Comparing gas chromatography with time-of-flight, quadrupole time-of-light and quadrupole mass spectrometry for stable isotope tracing (part-II) |
Study Summary | Stable isotope tracers are applied in vivo and in vitro studies to reveal the activity of enzymes and intracellular metabolic pathways. Most often, such tracers are used with gas chromatography coupled to mass spectrometry (GC-MS) due to its ease of operation and reproducible mass spectral databases. Differences in isotope tracer performance of classic GC-quadrupole MS instrument and newer time-of-flight instruments are not well-studied. Here, we used three commercially available instruments for the analysis of identical samples from a stable isotope labeling study that used [U-13C6] d-glucose to investigate the metabolism of Rothia mucilaginosa with respect to 29 amino acids and hydroxyl acids involved in primary metabolism. Overall, all three GC-MS instruments (low-resolution GC-SQ-MS, low-resolution GC-TOF-MS, and high-resolution GC-Q-TOF-MS) can be used to perform stable isotope tracing studies for glycolytic intermediates, TCA metabolites and amino acids, yielding similar biological results, with high-resolution GC-Q-TOF-MS offering additional capabilities to identify chemical structures of unknown compounds that might show significant isotope enrichments in biological studies. |
Institute | University of California, Davis |
Last Name | Zhang |
First Name | Ying |
Address | 451 East Health Science Drive, Davis, CA, 95616, USA |
ythzhang@ucdavis.edu | |
Phone | 1-530-752-8129 |
Submit Date | 2020-11-05 |
Raw Data Available | Yes |
Raw Data File Type(s) | d |
Analysis Type Detail | GC-MS |
Release Date | 2020-12-10 |
Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
Project ID: | PR001033 |
Project DOI: | doi: 10.21228/M85M45 |
Project Title: | Comparing gas chromatography with time-of-flight, quadrupole time-of-light and quadrupole mass spectrometry for stable isotope tracing |
Project Summary: | Stable isotope tracers are applied in vivo and in vitro studies to reveal the activity of enzymes and intracellular metabolic pathways. Most often, such tracers are used with gas chromatography coupled to mass spectrometry (GC-MS) due to its ease of operation and reproducible mass spectral databases. Differences in isotope tracer performance of classic GC-quadrupole MS instrument and newer time-of-flight instruments are not well-studied. Here, we used three commercially available instruments for the analysis of identical samples from a stable isotope labeling study that used [U-13C6] d-glucose to investigate the metabolism of Rothia mucilaginosa with respect to 29 amino acids and hydroxyl acids involved in primary metabolism. Overall, all three GC-MS instruments (low-resolution GC-SQ-MS, low-resolution GC-TOF-MS, and high-resolution GC-Q-TOF-MS) can be used to perform stable isotope tracing studies for glycolytic intermediates, TCA metabolites and amino acids, yielding similar biological results, with high-resolution GC-Q-TOF-MS offering additional capabilities to identify chemical structures of unknown compounds that might show significant isotope enrichments in biological studies. |
Institute: | University of California, Davis |
Laboratory: | Oliver Fiehn |
Last Name: | Ying |
First Name: | Zhang |
Address: | West Coast Metabolomics Center, University of California, Davis |
Email: | ythzhang@ucdavis.edu |
Phone: | +1-530-754-8258 |
Subject:
Subject ID: | SU001690 |
Subject Type: | Bacteria |
Subject Species: | Rothia mucilaginosa |
Taxonomy ID: | 43675 |
Genotype Strain: | RmFLR01 |
Factors:
Subject type: Bacteria; Subject species: Rothia mucilaginosa (Factor headings shown in green)
mb_sample_id | local_sample_id | Condition | Collection Time |
---|---|---|---|
SA136818 | 171015bBG_41_010_1 | Ambient | 12h |
SA136819 | 171015bBG_42_010_1 | Ambient | 12h |
SA136820 | 171015bBG_41_009_1 | Ambient | 12h |
SA136821 | 171015bBG_41_008_1 | Ambient | 12h |
SA136822 | 171015bBG_41_006_1 | Ambient | 12h |
SA136823 | 171015bBG_41_007_1 | Ambient | 12h |
SA136824 | 171015bBG_42_009_1 | Ambient | 12h |
SA136825 | 171015bBG_42_003_1 | Ambient | 12h |
SA136826 | 171015bBG_41_005_1 | Ambient | 12h |
SA136827 | 171015bBG_42_002_1 | Ambient | 12h |
SA136828 | 171015bBG_42_004_1 | Ambient | 12h |
SA136829 | 171015bBG_42_005_1 | Ambient | 12h |
SA136830 | 171015bBG_42_007_1 | Ambient | 12h |
SA136831 | 171015bBG_42_006_1 | Ambient | 12h |
SA136832 | 171015bBG_42_008_1 | Ambient | 12h |
SA136833 | 171015bBG_40_003_1 | Ambient | 12h |
SA136834 | 171015bBG_40_007_1 | Ambient | 12h |
SA136835 | 171015bBG_40_006_1 | Ambient | 12h |
SA136836 | 171015bBG_40_008_1 | Ambient | 12h |
SA136837 | 171015bBG_40_009_1 | Ambient | 12h |
SA136838 | 171015bBG_42_001_1 | Ambient | 12h |
SA136839 | 171015bBG_40_010_1 | Ambient | 12h |
SA136840 | 171015bBG_40_005_1 | Ambient | 12h |
SA136841 | 171015bBG_40_004_1 | Ambient | 12h |
SA136842 | 171015bBG_41_002_1 | Ambient | 12h |
SA136843 | 171015bBG_41_003_1 | Ambient | 12h |
SA136844 | 171015bBG_41_001_1 | Ambient | 12h |
SA136845 | 171015bBG_40_001_1 | Ambient | 12h |
SA136846 | 171015bBG_40_002_1 | Ambient | 12h |
SA136847 | 171015bBG_41_004_1 | Ambient | 12h |
SA136848 | 171015bBG_43_009_1 | Ambient | 24h |
SA136849 | 171015bBG_45_001_1 | Ambient | 24h |
SA136850 | 171015bBG_45_002_1 | Ambient | 24h |
SA136851 | 171015bBG_44_001_1 | Ambient | 24h |
SA136852 | 171015bBG_44_002_1 | Ambient | 24h |
SA136853 | 171015bBG_44_004_1 | Ambient | 24h |
SA136854 | 171015bBG_44_003_1 | Ambient | 24h |
SA136855 | 171015bBG_45_003_1 | Ambient | 24h |
SA136856 | 171015bBG_45_004_1 | Ambient | 24h |
SA136857 | 171015bBG_43_010_1 | Ambient | 24h |
SA136858 | 171015bBG_45_010_1 | Ambient | 24h |
SA136859 | 171015bBG_45_008_1 | Ambient | 24h |
SA136860 | 171015bBG_45_007_1 | Ambient | 24h |
SA136861 | 171015bBG_45_005_1 | Ambient | 24h |
SA136862 | 171015bBG_45_006_1 | Ambient | 24h |
SA136863 | 171015bBG_44_005_1 | Ambient | 24h |
SA136864 | 171015bBG_45_009_1 | Ambient | 24h |
SA136865 | 171015bBG_44_006_1 | Ambient | 24h |
SA136866 | 171015bBG_43_004_1 | Ambient | 24h |
SA136867 | 171015bBG_43_006_1 | Ambient | 24h |
SA136868 | 171015bBG_43_007_1 | Ambient | 24h |
SA136869 | 171015bBG_43_008_1 | Ambient | 24h |
SA136870 | 171015bBG_43_003_1 | Ambient | 24h |
SA136871 | 171015bBG_43_005_1 | Ambient | 24h |
SA136872 | 171015bBG_44_008_1 | Ambient | 24h |
SA136873 | 171015bBG_43_002_1 | Ambient | 24h |
SA136874 | 171015bBG_44_009_1 | Ambient | 24h |
SA136875 | 171015bBG_44_007_1 | Ambient | 24h |
SA136876 | 171015bBG_44_010_1 | Ambient | 24h |
SA136877 | 171015bBG_43_001_1 | Ambient | 24h |
SA136878 | 171012bBG_34_010_1 | Ambient | 4h |
SA136879 | 171012bBG_34_009_1 | Ambient | 4h |
SA136880 | 171012bBG_35_001_1 | Ambient | 4h |
SA136881 | 171012bBG_35_003_1 | Ambient | 4h |
SA136882 | 171012bBG_35_004_1 | Ambient | 4h |
SA136883 | 171012bBG_35_002_1 | Ambient | 4h |
SA136884 | 171012bBG_34_002_1 | Ambient | 4h |
SA136885 | 171012bBG_34_003_1 | Ambient | 4h |
SA136886 | 171012bBG_35_006_1 | Ambient | 4h |
SA136887 | 171012bBG_34_004_1 | Ambient | 4h |
SA136888 | 171012bBG_34_005_1 | Ambient | 4h |
SA136889 | 171012bBG_34_007_1 | Ambient | 4h |
SA136890 | 171012bBG_34_006_1 | Ambient | 4h |
SA136891 | 171012bBG_34_008_1 | Ambient | 4h |
SA136892 | 171012bBG_36_007_1 | Ambient | 4h |
SA136893 | 171012bBG_36_004_1 | Ambient | 4h |
SA136894 | 171012bBG_36_005_1 | Ambient | 4h |
SA136895 | 171012bBG_36_003_1 | Ambient | 4h |
SA136896 | 171012bBG_36_002_1 | Ambient | 4h |
SA136897 | 171012bBG_34_001_1 | Ambient | 4h |
SA136898 | 171012bBG_36_001_1 | Ambient | 4h |
SA136899 | 171012bBG_36_006_1 | Ambient | 4h |
SA136900 | 171012bBG_36_008_1 | Ambient | 4h |
SA136901 | 171012bBG_35_009_1 | Ambient | 4h |
SA136902 | 171012bBG_35_008_1 | Ambient | 4h |
SA136903 | 171012bBG_35_010_1 | Ambient | 4h |
SA136904 | 171012bBG_36_010_1 | Ambient | 4h |
SA136905 | 171012bBG_36_009_1 | Ambient | 4h |
SA136906 | 171012bBG_35_007_1 | Ambient | 4h |
SA136907 | 171012bBG_35_005_1 | Ambient | 4h |
SA136908 | 171012bBG_37_010_1 | Ambient | 8h |
SA136909 | 171012bBG_37_009_1 | Ambient | 8h |
SA136910 | 171012bBG_38_001_1 | Ambient | 8h |
SA136911 | 171012bBG_38_002_1 | Ambient | 8h |
SA136912 | 171012bBG_38_004_1 | Ambient | 8h |
SA136913 | 171012bBG_38_003_1 | Ambient | 8h |
SA136914 | 171012bBG_37_008_1 | Ambient | 8h |
SA136915 | 171012bBG_37_007_1 | Ambient | 8h |
SA136916 | 171012bBG_37_002_1 | Ambient | 8h |
SA136917 | 171012bBG_39_010_1 | Ambient | 8h |
Collection:
Collection ID: | CO001683 |
Collection Summary: | A quality control mixture of 29 unlabeled metabolite standards (Supporting information Table SI 1) was prepared to reach a final concentration of 1 mg/mL as stock solution. 50 µL aliquot from each stock solution was combined into a new tube and dried down. Then, the mixture was diluted to reach a final concentration of 50 µg/mL working solution. Rothia mucilaginosa strain RmFLR01 was isolated from a cystic fibrosis (CF) patient at the UC San Diego Adult CF Clinic [19, 32]. R. mucilaginosa cultures were grown in triplicates in artificial-sputum medium [33] spiked with 100 mM [U-13C6] d-glucose (Cambridge Isotope Laboratory, Tewksbury, MA, USA) under anaerobic and aerobic conditions (5% CO2) at 37°C and harvested at 4, 8, 12 and 24 h for isotope tracer analyses. |
Sample Type: | Bacterial cells |
Treatment:
Treatment ID: | TR001703 |
Treatment Summary: | under anaerobic and aerobic conditions (5% CO2) at 37°C and harvested at 4, 8, 12 and 24 h for isotope tracer analyses |
Sample Preparation:
Sampleprep ID: | SP001696 |
Sampleprep Summary: | For the 29 unlabeled standards mixture, 10 µL of the final solution was dried down for GC-MS measurement. Derivatization and data acquisition of mixture aliquots by GC-MS were reproduced on three days for inter-day precision. Dried mixtures were derivatized by adding 10 µL of 40 mg/mL methoxyamine hydrochloride (Sigma-Aldrich, St. Louis, MO, USA) in pyridine (Sigma-Aldrich, St. Louis, MO, USA) and shaking at 30°C for 1.5 h. Subsequently, 90 µL MTBSTFA (Sigma-Aldrich, St. Louis, MO, USA) was added with 13 fatty acid methyl esters (FAMEs) as retention index markers and shaken at 80°C for 30 min. Samples were immediately transferred to crimp top vials and injected onto each GC-MS instrument. Same samples of R. mucilaginosa cultures were extracted using published methods [19]. Samples were added 1 mL pre-chilled, degassed acetonitrile: isopropanol: water (v/v/v 3:3:2, Fisher Scientific) followed by vortexing 30 s and shaking at 4°C for 5 min. Samples were centrifuged for 2 min at 12,210 × g to precipitate debris from extracts. Supernatants were collected and split into two equal portions. One aliquot was dried to completeness in a Labconco cold trap centrifuge evaporator and then resuspended in 0.5 mL degassed acetonitrile: water (v/v 1:1, Fisher Scientific) to remove triacylglycerides. Resuspension solutions were vortexed for 30s and centrifuged for 2 min. Supernatants were transferred into clean Eppendorf tubes and dried down completely. Dried extracts were derivatized as given above. |
Combined analysis:
Analysis ID | AN002648 |
---|---|
Analysis type | MS |
Chromatography type | GC |
Chromatography system | Leco Pegasus IV GC |
Column | Restek Rtx-5Sil (30m x 0.25mm,0.25um) |
MS Type | EI |
MS instrument type | GC-TOF |
MS instrument name | Leco Pegasus IV TOF |
Ion Mode | POSITIVE |
Units | peak height |
Chromatography:
Chromatography ID: | CH001956 |
Chromatography Summary: | untargeted metabolomics metadata |
Instrument Name: | Leco Pegasus IV GC |
Column Name: | Restek Rtx-5Sil (30m x 0.25mm,0.25um) |
Chromatography Type: | GC |
MS:
MS ID: | MS002460 |
Analysis ID: | AN002648 |
Instrument Name: | Leco Pegasus IV TOF |
Instrument Type: | GC-TOF |
MS Type: | EI |
MS Comments: | LECO TOF-MS raw data files (.CDF format) were converted to MassHunter formats (.D format) using Agilent GCMS translator software. MassHunter Quantitative Analysis B.07.00 version was used to process the data |
Ion Mode: | POSITIVE |