Summary of Study ST001332

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 PR000904. The data can be accessed directly via it's Project DOI: 10.21228/M8TX11 This work is supported by NIH grant, U2C- DK119886.

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Study IDST001332
Study TitleLCMS lipid and acyl-carnitine analysis
Study SummaryLCMS Lipidomics and acyl-carnitine analysis.
Institute
University of Cambridge
LaboratoryCMaLL
Last NameJenkins
First NameBenjamin
AddressDepartment of Biochemistry, University of Cambridge, c/o Level 4, Pathology Building
Emailbjj25@medschl.cam.ac.uk
Phone07731103718
Submit Date2020-01-09
Raw Data AvailableYes
Raw Data File Type(s)raw(Thermo)
Analysis Type DetailLC-MS
Release Date2020-03-30
Release Version1
Benjamin Jenkins Benjamin Jenkins
https://dx.doi.org/10.21228/M8TX11
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR000904
Project DOI:doi: 10.21228/M8TX11
Project Title:LCMS lipid and acyl-carnitine analysis
Project Summary:Lipidomics and acyl-carnitine analysis.
Institute:University of Cambridge
Department:Department of Biochemistry
Last Name:Jenkins
First Name:Benjamin
Address:Department of Biochemistry, University of Cambridge, c/o Level 4, Pathology Building
Email:bjj25@medschl.cam.ac.uk
Phone:07731103718

Subject:

Subject ID:SU001406
Subject Type:Invertebrate
Subject Species:Caenorhabditis elegans
Taxonomy ID:6239

Factors:

Subject type: Invertebrate; Subject species: Caenorhabditis elegans (Factor headings shown in green)

mb_sample_id local_sample_id Treatment
SA09655020190326_BJJ_Acyl_carnitines_Sample_8_BIGBIG
SA09655120190326_BJJ_Lipids_Sample_8_BIGBIG
SA09655220190326_BJJ_Lipids_Sample_5_BIGBIG
SA09655320190326_BJJ_Lipids_Sample_7_BIGBIG
SA09655420190326_BJJ_Lipids_Sample_6_BIGBIG
SA09655520190326_BJJ_Acyl_carnitines_Sample_7_BIGBIG
SA09655620190326_BJJ_Acyl_carnitines_Sample_6_BIGBIG
SA09655720190326_BJJ_Acyl_carnitines_Sample_5_BIGBIG
SA09655820190326_BJJ_Acyl_carnitines_Sample_2_HBHB
SA09655920190326_BJJ_Acyl_carnitines_Sample_4_HBHB
SA09656020190326_BJJ_Lipids_Sample_4_HBHB
SA09656120190326_BJJ_Lipids_Sample_2_HBHB
SA09656220190510_BJJ_Lipids_Sample_1_HB101HB101
SA09656320190510_BJJ_Lipids_Sample_3_HB101HB101
SA09656420190510_BJJ_Acyl_carnitines_Sample_3_HB101HB101
SA09656520190510_BJJ_Lipids_Sample_2_HB101HB101
SA09656620190510_BJJ_Acyl_carnitines_Sample_2_HB101HB101
SA09656720190510_BJJ_Acyl_carnitines_Sample_1_HB101HB101
SA09656820190326_BJJ_Acyl_carnitines_Sample_1_HBSHBS
SA09656920190326_BJJ_Acyl_carnitines_Sample_3_HBSHBS
SA09657020190326_BJJ_Lipids_Sample_1_HBSHBS
SA09657120190326_BJJ_Lipids_Sample_3_HBSHBS
SA09657820190510_BJJ_Acyl_carnitines_Sample_4_PA14PA14
SA09657920190510_BJJ_Lipids_Sample_4_PA14PA14
SA09658020190510_BJJ_Acyl_carnitines_Sample_6_PA14PA14
SA09658120190510_BJJ_Acyl_carnitines_Sample_5_PA14PA14
SA09658220190510_BJJ_Lipids_Sample_5_PA14PA14
SA09658320190510_BJJ_Lipids_Sample_6_PA14PA14
SA09657220190510_BJJ_Lipids_Sample_9_P.luminP.Lumin
SA09657320190510_BJJ_Acyl_carnitines_Sample_8_P.LUMINP.Lumin
SA09657420190510_BJJ_Lipids_Sample_7_P.luminP.Lumin
SA09657520190510_BJJ_Acyl_carnitines_Sample_7_P.LUMINP.Lumin
SA09657620190510_BJJ_Acyl_carnitines_Sample_9_P.LUMINP.Lumin
SA09657720190510_BJJ_Lipids_Sample_8_P.luminP.Lumin
Showing results 1 to 34 of 34

Collection:

Collection ID:CO001401
Collection Summary:LCMS lipid and acyl-carnitine analysis.
Sample Type:C. elegans

Treatment:

Treatment ID:TR001421
Treatment Summary:Parental exposure to environmental stress can program adaptive changes in offspring in diverse organisms (1–4). The mechanisms by which parental exposure to environmental stresses can program predictive adaptive responses in offspring remain almost completely unknown. Here we report that the soil bacteria Pseudomonas vranovensis is a natural pathogen of the nematode Caenorhabditis elegans and that parental exposure of C. elegans to P. vranovensis promotes offspring resistance to infection. This adaptation can be transmitted transgenerationally such that infection of adults can enhance the immunity of their descendants four generations later. We find that parental infection by P. vranovensis results in increased expression of the cysteine synthases CYSL-1 and CYSL-2 and the regulator of hypoxia inducible factor RHY-1 in progeny, that the expression of these three genes in offspring is required for adaptation to P. vranovensis, and that the expression of these genes is regulated by the WD40 repeat protein WDR-23.

Sample Preparation:

Sampleprep ID:SP001414
Sampleprep Summary:C. elegans was prepared for LC-MS lipidomics and acyl-carnitine analysis as previously described (3) with minor modifications. Briefly, ~40 µL of concentrated embryos were re-suspended in 100 µL of water, then 0.4 mL of chloroform was added to each sample followed by 0.2 mL of methanol containing the stable isotope labelled acyl-carnitine internal standards (Butyryl-L-carnitine-d7 at 5 µM and Hexadecanoyl-L-carnitine-d3 at 5 µM). The samples were then homogenised by vortexing then transferred into a 2 mL Eppendorf screw-cap tube. The original container was washed out with 0.5 mL of chloroform: methanol (2: 1, respectively) and added to the appropriate 2 mL Eppendorf screw-cap tube. This was followed by the addition of 150 µL of the following stable isotope labelled internal standards (approximately 10 to 50 µM in methanol): Ceramide_C16d31, LPC_(C14:0d42), PC_(C16:0d31 / C18:1), PE_(C16:0d31 / C18:1), PG_(C16:0d31 / C18:1), PI_(C16:0d31 / C18:1), PS_(C16:0d62), SM_(C16:0d31), TG_(45:0d29) and TG_(48:0d31). Then, 400 µL of sterile water was added. The samples were vortexed for 1 min, and then centrifuged at ~20,000 rpm for 5 minutes. For the intact lipid sample preparation, 0.3 mL of the organic layer (the lower chloroform layer) was collected into a 2 mL amber glass vial (Agilent Technologies, Santa Clara California, USA) and dried down to dryness in an Eppendorf Concentrator Plus system (Eppendorf, Stevenage, UK) run for 60 minutes at 45 °C. The dried lipid samples were then reconstituted with 100 µL of 2:1:1 solution of propan-2-ol, acetonitrile and water, respectively, and then vortexed thoroughly. The lipid samples were then transferred into a 300 μL low-volume vial insert inside a 2 mL amber glass auto-sample vial ready for liquid chromatography separation with mass spectrometry detection (LC-MS) of intact lipid species. For the acyl-carnitine sample preparation, 0.2 mL of the organic layer (the lower chloroform layer) and 0.2 mL of the aqueous layer (the top water layer) were mixed into a 2 mL amber glass vial and dried down to dryness. The dried acyl-carnitine samples were then reconstituted with 100 µL of water and acetonitrile (4: 1, respectively) and thoroughly vortexed. The acyl-carnitine samples were then transferred into a 300 μL low-volume vial insert inside a 2 mL amber glass auto-sample vial ready for liquid chromatography separation with mass spectrometry detection (LC-MS) of the acyl-carnitine species.

Combined analysis:

Analysis ID AN002221
Analysis type MS
Chromatography type Reversed phase
Chromatography system Shimadzu 20AD
Column Waters Acquity UPLC CSH C18(50 x 2.1mm ,1.7um)
MS Type ESI
MS instrument type Orbitrap
MS instrument name Thermo Exactive
Ion Mode UNSPECIFIED
Units Area ratio

Chromatography:

Chromatography ID:CH001629
Chromatography Summary:Full chromatographic separation of intact lipids (4) was achieved using Shimadzu HPLC System (Shimadzu UK Limited, Milton Keynes, United Kingdom) with the injection of 10 µL onto a Waters Acquity UPLC® CSH C18 column; 1.7 µm, I.D. 2.1 mm X 50 mm, maintained at 55 °C. Mobile phase A was 6:4, acetonitrile and water with 10 mM ammonium formate. Mobile phase B was 9:1, propan-2-ol and acetonitrile with 10 mM ammonium formate. The flow was maintained at 500 µL per minute through the following gradient: 0.00 minutes_40% mobile phase B; 0.40 minutes_43% mobile phase B; 0.45 minutes_50% mobile phase B; 2.40 minutes_54% mobile phase B; 2.45 minutes_70% mobile phase B; 7.00 minutes_99% mobile phase B; 8.00 minutes_99% mobile phase B; 8.3 minutes_40% mobile phase B; 10 minutes_40% mobile phase B; 10.00 minutes_40% mobile phase B. The sample injection needle was washed using 9:1, 2-propan-2-ol and acetonitrile with 0.1 % formic acid. The mass spectrometer used was the Thermo Scientific Exactive Orbitrap with a heated electrospray ionisation source (Thermo Fisher Scientific, Hemel Hempstead, UK). The mass spectrometer was calibrated immediately before sample analysis using positive and negative ionisation calibration solution (recommended by Thermo Scientific). Additionally, the heated electrospray ionisation source was optimised at 50:50 mobile phase A to mobile phase B for spray stability (capillary temperature; 380 °C, source heater temperature; 420 °C, sheath gas flow; 60 (arbitrary), auxiliary gas flow; 20 (arbitrary), sweep gas; 5 (arbitrary), source voltage; 3.5 kV. The mass spectrometer resolution was set to 25,000 with a full-scan range of m/z 100 to 1,800 Da, with continuous switching between positive and negative mode. Lipid quantification was achieved using the area under the curve (AUC) of the corresponding high resolution extracted ion chromatogram (with a window of ± 8 ppm) at the indicative retention time. The lipid analyte AUC relative to the associated internal standard AUC for that lipid class was used to semi-quantify and correct for any extraction/instrument variation.
Instrument Name:Shimadzu 20AD
Column Name:Waters Acquity UPLC CSH C18(50 x 2.1mm ,1.7um)
Column Temperature:55
Flow Gradient:0.00 minutes_40% mobile phase B; 0.40 minutes_43% mobile phase B; 0.45 minutes_50% mobile phase B; 2.40 minutes_54% mobile phase B; 2.45 minutes_70% mobile phase B; 7.00 minutes_99% mobile phase B; 8.00 minutes_99% mobile phase B; 8.3 minutes_40% mobile phase B; 10 minutes_40% mobile phase B; 10.00 minutes_40% mobile phase B. The sample injection needle was washed using 9:1, 2-propan-2-ol and acetonitrile with 0.1 % formic acid.
Flow Rate:500 µL/min
Solvent A:60% acetonitrile/40% water; 10 mM ammonium formate
Solvent B:90% isopropanol/10% acetonitrile; 10 mM ammonium formate
Chromatography Type:Reversed phase

MS:

MS ID:MS002067
Analysis ID:AN002221
Instrument Name:Thermo Exactive
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:The mass spectrometer used was the Thermo Scientific Exactive Orbitrap with a heated electrospray ionisation source (Thermo Fisher Scientific, Hemel Hempstead, UK). The mass spectrometer was calibrated immediately before sample analysis using positive and negative ionisation calibration solution (recommended by Thermo Scientific). Additionally, the heated electrospray ionisation source was optimised at 50:50 mobile phase A to mobile phase B for spray stability (capillary temperature; 380 °C, source heater temperature; 420 °C, sheath gas flow; 60 (arbitrary), auxiliary gas flow; 20 (arbitrary), sweep gas; 5 (arbitrary), source voltage; 3.5 kV. The mass spectrometer resolution was set to 25,000 with a full-scan range of m/z 100 to 1,800 Da, with continuous switching between positive and negative mode. Lipid quantification was achieved using the area under the curve (AUC) of the corresponding high resolution extracted ion chromatogram (with a window of ± 8 ppm) at the indicative retention time. The lipid analyte AUC relative to the associated internal standard AUC for that lipid class was used to semi-quantify and correct for any extraction/instrument variation.
Ion Mode:UNSPECIFIED
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