Summary of Study ST001816

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 PR001149. The data can be accessed directly via it's Project DOI: 10.21228/M8611G 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 IDST001816
Study TitleQuantification of PIPs species in mouse tissues.
Study SummaryPhosphoinositides (PIPs) species, differing in phosphorylation at hydroxyls of the inositol head group, play roles in various cellular events. Here we developed a supercritical fluid chromatography-mass spectrometry (SFC-MS) method that allows us to quantify molecular species of all seven PIP regioisomers in culture cells and tissues. Using this methodology, we quantified PIPs species in 13 mouse tissues.
Institute
Grad Sch of Pharmaceut Sci, Univ of Tokyo
Last NameKono
First NameNozomu
AddressHongo 7-3-1,, Bunyo-ku, Tokyo, 113-0033, Japan
Emailnozomu@mol.f.u-tokyo.ac.jp
Phone+81-3-5841-4723
Submit Date2021-06-01
Raw Data AvailableYes
Raw Data File Type(s)mzML
Analysis Type DetailLC-MS
Release Date2022-05-04
Release Version1
Nozomu Kono Nozomu Kono
https://dx.doi.org/10.21228/M8611G
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR001149
Project DOI:doi: 10.21228/M8611G
Project Title:Quantification of PIPs species in biological samples.
Project Summary:Phosphoinositides (PIPs) species, differing in phosphorylation at hydroxyls of the inositol head group, play roles in various cellular events. Here we developed a supercritical fluid chromatography-mass spectrometry (SFC-MS) method that allows us to quantify molecular species of all seven PIP regioisomers in culture cells and tissues.
Institute:Grad Sch of Pharmaceut Sci, Univ of Tokyo
Last Name:Kono
First Name:Nozomu
Address:Hongo 7-3-1,, Bunyo-ku, Tokyo, 113-0033, Japan
Email:nozomu@mol.f.u-tokyo.ac.jp
Phone:+81-3-5841-4723

Subject:

Subject ID:SU001893
Subject Type:Mammal
Subject Species:Mus musculus
Taxonomy ID:10090
Age Or Age Range:8 weeks old
Gender:Male

Factors:

Subject type: Mammal; Subject species: Mus musculus (Factor headings shown in green)

mb_sample_id local_sample_id Tissue
SA168974Brain3Brain
SA168975Brain2Brain
SA168976Brain1Brain
SA168977Colon3Colon
SA168978Colon1Colon
SA168979Colon2Colon
SA168980Heart3Heart
SA168981Heart1Heart
SA168982Heart2Heart
SA168983Kidney3Kidney
SA168984Kidney2Kidney
SA168985Kidney1Kidney
SA168986Liver2Liver
SA168987Liver1Liver
SA168988Liver3Liver
SA168989Lung2Lung
SA168990Lung1Lung
SA168991Lung3Lung
SA168992Pancreas2Pancreas
SA168993Pancreas1Pancreas
SA168994Pancreas3Pancreas
SA168995Skeletal Muscle1Skeletal Muscle
SA168996Skeletal Muscle2Skeletal Muscle
SA168997Skeletal Muscle3Skeletal Muscle
SA168998Small Intestine3Small Intestine
SA168999Small Intestine2Small Intestine
SA169000Small Intestine1Small Intestine
SA169001Spleen2Spleen
SA169002Spleen3Spleen
SA169003Spleen1Spleen
SA169004Stomach2Stomach
SA169005Stomach3Stomach
SA169006Stomach1Stomach
SA169007Testis1Testis
SA169008Testis2Testis
SA169009Testis3Testis
SA169010White Adipose Tissue3White Adipose Tissue
SA169011White Adipose Tissue2White Adipose Tissue
SA169012White Adipose Tissue1White Adipose Tissue
Showing results 1 to 39 of 39

Collection:

Collection ID:CO001886
Collection Summary:Wild-type male mice (C57BL/6J) were anaesthetized, perfused and tissues were harvested. Isolated mouse tissues were rinsed with cold PBS and immediately frozen in liquid nitrogen.
Sample Type:Various Tissues

Treatment:

Treatment ID:TR001906
Treatment Summary:Thirteen tissues were harvested from 8-week-old wild-type male mice (C57BL/6J).

Sample Preparation:

Sampleprep ID:SP001899
Sampleprep Summary:The following solutions were prepared for lipid extractions; the quench mixture comprising 484 ml MeOH, 242 ml CHCl3, and 23.55 ml 1 M HCl; the pre-derivatization wash composed of 240 ml CHCl3, 120 ml MeOH and 90 ml 0.01 M HCl; and the post-derivatization wash made up of 240 ml CHCl3, 120 ml MeOH, and 90 ml H2O. Wash mixtures were shaken and allowed to separate into two phases. For pre/post-derivatization wash, the upper phase of each solution was used. Frozen tissues were ground into powder in liquid N2 using a pestle and mortar. Frozen tissues were collected into a safe-lock poly-propylene tube (2 ml), and resuspended with 750 μl of quench mix, 170 μl of H2O and internal standards [10 μl containing 2 ng 12:0/13:0 PI, 17:0/20:4 PI (4)P, 17:0/20:4 PI (4,5)P2 and 17:0/20:4 PI (3,4,5)P3], followed by vortex-mixing and lipid extraction. Lipid extraction was performed, based on procedures described by J Clark et al. The single-phase sample (a mixture of 170 μl of an aqueous sample, 2 ng of internal standards, and 750 μl of quench mix) were mixed with 725 μl of CHCl3 and 170 μl of 2M HCl, followed by vortex-mixing and centrifugation (15,000 g, 5 min at room temperature). The lower organic phase was collected into a fresh safe-lock poly-propylene tube (2 ml) and mixed with 708 μl of prederivatization wash, followed by vortex-mixing and centrifugation (15,000 g, 3 min at room temperature). The lower phase was collected into another fresh tube and subjected to derivatization. Derivatization of lipids was performed in a fume hood with adequate personal safety equipment as follows, based on procedures described by J Clark et al7. Fifty μl trimethylsilyl diazomethane in hexane (2 M solution; Sigma-Aldrich) was added to the lipid extracts (~1 ml), and after leaving to stand 10 min at room temperature, the reaction was quenched with 6 μl of acetic acid. Next, 700 μl post-derivatization wash solution was added to the mixture, followed by centrifugation (1,500 g, 3 min). The resultant lower phase was collected and rewashed with a 700 μl post-derivatization wash solution. Then 90 μl of MeOH and 10 μl of H2O were added to the final collected lower phase. The samples were dried up with a centrifugal evaporator. Finally, samples were dissolved in 80μl MeOH, sonicated briefly, and 20 μl H2O was added. To avoid degradation, the samples were stored at -80ºC until use.

Combined analysis:

Analysis ID AN002948
Analysis type MS
Chromatography type Other
Chromatography system Shimadzu Nexera UC
Column ULTRON AF-HILIC-CD (250 mm × 2.1 mm, 5.0 µm; Shinwa Chemical Industries)
MS Type ESI
MS instrument type Ion trap
MS instrument name ABI SCIEX 4500 QTrap
Ion Mode POSITIVE
Units pmol/mg tissue

Chromatography:

Chromatography ID:CH002183
Instrument Name:Shimadzu Nexera UC
Column Name:ULTRON AF-HILIC-CD (250 mm × 2.1 mm, 5.0 µm; Shinwa Chemical Industries)
Column Temperature:4℃
Flow Gradient:0–16 min: 5% B→20% B; 16.01–18 min: 40% B; 18.01–22 min: 5% B
Flow Rate:1.5 ml/min
Solvent A:supercritical carbon dioxide (SCCO2)
Solvent B:water/methanol (2.5/97.5, v/v) containing 0.1% (v/v) formic acid
Chromatography Type:Other

MS:

MS ID:MS002738
Analysis ID:AN002948
Instrument Name:ABI SCIEX 4500 QTrap
Instrument Type:Ion trap
MS Type:ESI
MS Comments:The instrument parameters of QTRAP4500 for positive ion mode were as follows: curtain gas, 20 psi; ionspray voltage, 4500 V; temperature, 300 ºC; ion source gas 1, 18 psi; ion source gas 2, 20 psi. and quantification was performed using MultiQuant™ software (SCIEX).
Ion Mode:POSITIVE
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