Summary of Study ST001710

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 PR001095. The data can be accessed directly via it's Project DOI: 10.21228/M85976 This work is supported by NIH grant, U2C- DK119886.

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This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

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Study IDST001710
Study TitleMetabolic signatures of NAFLD - Lipidomics data (part 1 of 3)
Study SummarySerum samples were randomized and extracted using a modified version of the previously-published Folch procedure, as applied recently [20]. The maternal samples were analysed as one batch and the cord blood samples as a second batch. In short, 10 µL of 0.9% NaCl and, 120 µL of CHCl3: MeOH (2:1, v/v) containing the internal standards (c = 2.5 µg/mL) was added to 10 µL of each serum sample. The standard solution contained the following compounds: 1,2-diheptadecanoyl-sn-glycero-3-phosphoethanolamine (PE(17:0/17:0)), N-heptadecanoyl-D-erythro-sphingosylphosphorylcholine (SM(d18:1/17:0)), N-heptadecanoyl-D-erythro-sphingosine (Cer(d18:1/17:0)), 1,2-diheptadecanoyl-sn-glycero-3-phosphocholine (PC(17:0/17:0)), 1-heptadecanoyl-2-hydroxy-sn-glycero-3-phosphocholine (LPC(17:0)) and 1-palmitoyl-d31-2-oleoyl-sn-glycero-3-phosphocholine (PC(16:0/d31/18:1)), were purchased from Avanti Polar Lipids, Inc. (Alabaster, AL, USA), and, triheptadecanoylglycerol (TG(17:0/17:0/17:0)) was purchased from Larodan AB (Solna, Sweden). The samples were vortex mixed and incubated on ice for 30 min after which they were centrifuged (9400 × g, 3 min). 60 µL from the lower layer of each sample was then transferred to a glass vial with an insert and 60 µL of CHCl3: MeOH (2:1, v/v) was added to each sample. The samples were stored at -80 °C until analysis. Calibration curves using 1-hexadecyl-2-(9Z-octadecenoyl)-sn-glycero-3-phosphocholine (PC(16:0e/18:1(9Z))), 1-(1Z-octadecenyl)-2-(9Z-octadecenoyl)-sn-glycero-3-phosphocholine (PC(18:0p/18:1(9Z))), 1-stearoyl-2-hydroxy-sn-glycero-3-phosphocholine (LPC(18:0)), 1-oleoyl-2-hydroxy-sn-glycero-3-phosphocholine (LPC(18:1)), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (PE(16:0/18:1)), 1-(1Z-octadecenyl)-2-docosahexaenoyl-sn-glycero-3-phosphocholine (PC(18:0p/22:6)) and 1-stearoyl-2-linoleoyl-sn-glycerol (DG(18:0/18:2)), 1-(9Z-octadecenoyl)-sn-glycero-3-phosphoethanolamine (LPE(18:1)), N-(9Z-octadecenoyl)-sphinganine (Cer(d18:0/18:1(9Z))), 1-hexadecyl-2-(9Z-octadecenoyl)-sn-glycero-3-phosphoethanolamine (PE(16:0/18:1)) from Avanti Polar Lipids, 1-Palmitoyl-2-Hydroxy-sn-Glycero-3-Phosphatidylcholine (LPC(16:0)), 1,2,3 trihexadecanoalglycerol (TG(16:0/16:0/16:0)), 1,2,3-trioctadecanoylglycerol (TG(18:0/18:0/18:)) and 3β-hydroxy-5-cholestene-3-stearate (ChoE(18:0)), 3β-Hydroxy-5-cholestene-3-linoleate (ChoE(18:2)) from Larodan, were prepared to the following concentration levels: 100, 500, 1000, 1500, 2000 and 2500 ng/mL (in CHCl3:MeOH, 2:1, v/v) including 1250 ng/mL of each internal standard. The samples were analyzed by ultra-high-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UHPLC-QTOFMS). Briefly, the UHPLC system used in this work was a 1290 Infinity II system from Agilent Technologies (Santa Clara, CA, USA). The system was equipped with a multi sampler (maintained at 10 °C), a quaternary solvent manager and a column thermostat (maintained at 50 °C). Injection volume was 1 µL and the separations were performed on an ACQUITY UPLC® BEH C18 column (2.1 mm × 100 mm, particle size 1.7 µm) by Waters (Milford, MA, USA). The mass spectrometer coupled to the UHPLC was a 6545 QTOF from Agilent Technologies interfaced with a dual jet stream electrospray (Ddual ESI) ion source. All analyses were performed in positive ion mode and MassHunter B.06.01 (Agilent Technologies) was used for all data acquisition. Quality control was performed throughout the dataset by including blanks, pure standard samples, extracted standard samples and control serum samples, including in-house serum and a pooled QC with an aliquot of each sample was collected and pooled and used as quality control sample. Relative standard deviations (% RSDs) for identified lipids in the control serum samples (n = 13) and in the pooled serum samples (n = 54) were on average 22.4% and 17.5%, respectively.
Institute
Örebro University
Last NameMcGlinchey
First NameAidan
AddressSchool of Medical Sciences, Örebro, Örebro, 70281, Sweden
Emailaidan.mcglinchey@oru.se
Phone+46736485638
Submit Date2021-02-10
Raw Data AvailableYes
Raw Data File Type(s)mzdata.xml
Analysis Type DetailLC-MS
Release Date2022-01-03
Release Version1
Aidan McGlinchey Aidan McGlinchey
https://dx.doi.org/10.21228/M85976
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Sample Preparation:

Sampleprep ID:SP001793
Sampleprep Summary:Serum samples were randomized and extracted using a modified version of the previously-published Folch procedure, as applied recently. The maternal samples were analysed as one batch and the cord blood samples as a second batch. In short, 10 µL of 0.9% NaCl and, 120 µL of CHCl3: MeOH (2:1, v/v) containing the internal standards (c = 2.5 µg/mL) was added to 10 µL of each serum sample. The standard solution contained the following compounds: 1,2-diheptadecanoyl-sn-glycero-3-phosphoethanolamine (PE(17:0/17:0)), N-heptadecanoyl-D-erythro-sphingosylphosphorylcholine (SM(d18:1/17:0)), N-heptadecanoyl-D-erythro-sphingosine (Cer(d18:1/17:0)), 1,2-diheptadecanoyl-sn-glycero-3-phosphocholine (PC(17:0/17:0)), 1-heptadecanoyl-2-hydroxy-sn-glycero-3-phosphocholine (LPC(17:0)) and 1-palmitoyl-d31-2-oleoyl-sn-glycero-3-phosphocholine (PC(16:0/d31/18:1)), were purchased from Avanti Polar Lipids, Inc. (Alabaster, AL, USA), and, triheptadecanoylglycerol (TG(17:0/17:0/17:0)) was purchased from Larodan AB (Solna, Sweden). The samples were vortex mixed and incubated on ice for 30 min after which they were centrifuged (9400 × g, 3 min). 60 µL from the lower layer of each sample was then transferred to a glass vial with an insert and 60 µL of CHCl3: MeOH (2:1, v/v) was added to each sample. The samples were stored at -80 °C until analysis. Calibration curves using 1-hexadecyl-2-(9Z-octadecenoyl)-sn-glycero-3-phosphocholine (PC(16:0e/18:1(9Z))), 1-(1Z-octadecenyl)-2-(9Z-octadecenoyl)-sn-glycero-3-phosphocholine (PC(18:0p/18:1(9Z))), 1-stearoyl-2-hydroxy-sn-glycero-3-phosphocholine (LPC(18:0)), 1-oleoyl-2-hydroxy-sn-glycero-3-phosphocholine (LPC(18:1)), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (PE(16:0/18:1)), 1-(1Z-octadecenyl)-2-docosahexaenoyl-sn-glycero-3-phosphocholine (PC(18:0p/22:6)) and 1-stearoyl-2-linoleoyl-sn-glycerol (DG(18:0/18:2)), 1-(9Z-octadecenoyl)-sn-glycero-3-phosphoethanolamine (LPE(18:1)), N-(9Z-octadecenoyl)-sphinganine (Cer(d18:0/18:1(9Z))), 1-hexadecyl-2-(9Z-octadecenoyl)-sn-glycero-3-phosphoethanolamine (PE(16:0/18:1)) from Avanti Polar Lipids, 1-Palmitoyl-2-Hydroxy-sn-Glycero-3-Phosphatidylcholine (LPC(16:0)), 1,2,3 trihexadecanoalglycerol (TG(16:0/16:0/16:0)), 1,2,3-trioctadecanoylglycerol (TG(18:0/18:0/18:)) and 3β-hydroxy-5-cholestene-3-stearate (ChoE(18:0)), 3β-Hydroxy-5-cholestene-3-linoleate (ChoE(18:2)) from Larodan, were prepared to the following concentration levels: 100, 500, 1000, 1500, 2000 and 2500 ng/mL (in CHCl3:MeOH, 2:1, v/v) including 1250 ng/mL of each internal standard. The samples were analyzed by ultra-high-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UHPLC-QTOFMS). Briefly, the UHPLC system used in this work was a 1290 Infinity II system from Agilent Technologies (Santa Clara, CA, USA). The system was equipped with a multi sampler (maintained at 10 °C), a quaternary solvent manager and a column thermostat (maintained at 50 °C). Injection volume was 1 µL and the separations were performed on an ACQUITY UPLC® BEH C18 column (2.1 mm × 100 mm, particle size 1.7 µm) by Waters (Milford, MA, USA). The mass spectrometer coupled to the UHPLC was a 6545 QTOF from Agilent Technologies interfaced with a dual jet stream electrospray (Ddual ESI) ion source. All analyses were performed in positive ion mode and MassHunter B.06.01 (Agilent Technologies) was used for all data acquisition. Quality control was performed throughout the dataset by including blanks, pure standard samples, extracted standard samples and control serum samples, including in-house serum and a pooled QC with an aliquot of each sample was collected and pooled and used as quality control sample. Relative standard deviations (% RSDs) for identified lipids in the control serum samples (n = 13) and in the pooled serum samples (n = 54) were on average 22.4% and 17.5%, respectively. Mass spectrometry data processing was performed using the open source software package MZmine 2.18. The following steps were applied in this processing: (i) Crop filtering with a m/z range of 350 – 1200 m/z and an RT range of 2.0 to 12 minutes, (ii) Mass detection with a noise level of 750, (iii) Chromatogram builder with a minimum time span of 0.08 min, minimum height of 1000 and a m/z tolerance of 0.006 m/z or 10.0 ppm, (iv) Chromatogram deconvolution using the local minimum search algorithm with a 70% chromatographic threshold, 0.05 min minimum RT range, 5% minimum relative height, 1200 minimum absolute height, a minimum ration of peak top/edge of 1.2 and a peak duration range of 0.08 - 5.0, (v), Isotopic peak grouper with a m/z tolerance of 5.0 ppm, RT tolerance of 0.05 min, maximum charge of 2 and with the most intense isotope set as the representative isotope, (vi) Peak filter with minimum 12 data points, a FWHM between 0.0 and 0.2, tailing factor between 0.45 and 2.22 and asymmetry factor between 0.40 and 2.50, (vii) Join aligner with a m/z tolerance of 0.009 or 10.0 ppm and a weight for of 2, a RT tolerance of 0.1 min and a weight of 1 and with no requirement of charge state or ID and no comparison of isotope pattern, (viii) Peak list row filter with a minimum of 10% of the samples (ix) Gap filling using the same RT and m/z range gap filler algorithm with an m/z tolerance of 0.009 m/z or 11.0 ppm, (x) Identification of lipids using a custom database search with an m/z tolerance of 0.009 m/z or 10.0 ppm and a RT tolerance of 0.1 min, and (xi) Normalization using internal standards PE(17:0/17:0), SM(d18:1/17:0), Cer(d18:1/17:0), LPC(17:0), TG(17:0/17:0/17:0) and PC(16:0/d30/18:1)) for identified lipids and closest ISTD for the unknown lipids followed by calculation of the concentrations based on lipid-class concentration curves.
Processing Storage Conditions:-80℃
Extract Storage:-80℃
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