Summary of Study ST003354
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 PR002085. The data can be accessed directly via it's Project DOI: 10.21228/M83R6P This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
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.
| Study ID | ST003354 |
| Study Title | Incorporation of co-treated oleate-d9 and elaidate-d17 in long-chain base of sphingolipids in Huh7 cells. |
| Study Summary | We analyzed hydrolyzed long-chain bases (LCBs) in Huh7 cells treated with a combination of BSA-oleate-d9 and BSA-elaidate-d17. We aimed to confirm the incorporation of oleate and elaidate in the LCB of sphingolipids using their deuterated versions and put them both in direct competition for SPT. |
| Institute | Salk Institute for Biological Studies |
| Last Name | Gengatharan |
| First Name | Jivani |
| Address | 10010 N Torrey Pines Rd, La Jolla, CA, 92037, USA |
| jivani14@gmail.com | |
| Phone | (858) 453-4100 |
| Submit Date | 2024-07-26 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | d |
| Analysis Type Detail | LC-MS |
| Release Date | 2024-08-09 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR002085 |
| Project DOI: | doi: 10.21228/M83R6P |
| Project Title: | Altered sphingolipid biosynthetic flux and lipoprotein trafficking contribute to trans fat-induced atherosclerosis |
| Project Summary: | The goal of the project is to determine the role of sphingolipid metabolism in atherosclerosis induced by dietary trans fat. We analyzed lipid metabolites in Huh7 cells following various fatty acid treatments, with specific focus on cis and trans unsaturated fatty acids. Additionally, we analyzed lipid metabolites in plasma and liver of Ldlr-/- mice fed high-fat diets enriched in cis or trans fatty acids in the presence or absence of myriocin, a pharmacological inhibitor of Serine palmitoyltransferase (SPT), the initial rate-limiting enzyme of sphingolipid biosynthesis. |
| Institute: | Salk Institute for Biological Studies |
| Last Name: | Gengatharan |
| First Name: | Jivani |
| Address: | 10010 N Torrey Pines Rd, La Jolla, CA, 92037, USA |
| Email: | jivani14@gmail.com |
| Phone: | (858) 453-4100 |
Subject:
| Subject ID: | SU003475 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
| Cell Strain Details: | Huh7 |
Factors:
Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)
| mb_sample_id | local_sample_id | Sample source | Treatment |
|---|---|---|---|
| SA365732 | Huh7_oleated9elaidated17_1 | Huh7 cells | BSA-oleate-d9 + BSA-elaidate-d17 |
| SA365733 | Huh7_oleated9elaidated17_2 | Huh7 cells | BSA-oleate-d9 + BSA-elaidate-d17 |
| SA365734 | Huh7_oleated9elaidated17_3 | Huh7 cells | BSA-oleate-d9 + BSA-elaidate-d17 |
| Showing results 1 to 3 of 3 |
Collection:
| Collection ID: | CO003468 |
| Collection Summary: | Cells (~400,000) were spiked with internal standards sphinganine-d7 (Avanti Polar Lipids, Cat# 860658) and sphingosine-d7 (Avanti Polar Lipids, Cat# 860657) and were scraped with 0.5 mL methanol. |
| Sample Type: | Cultured cells |
Treatment:
| Treatment ID: | TR003484 |
| Treatment Summary: | Huh7 cells were treated with a combination of 50 µM BSA-oleate-d9 and 50 µM BSA-elaidate-d17 for 48 hours in delipidated media. |
Sample Preparation:
| Sampleprep ID: | SP003482 |
| Sampleprep Summary: | Cells (~400,000) were spiked with internal standards sphinganine-d7 (Avanti Polar Lipids, Cat# 860658) and sphingosine-d7 (Avanti Polar Lipids, Cat# 860657) and were scraped with 0.5 mL methanol. Homogenate aliquot of 50 µL was taken to determine protein content using the BCA protein assay (Thermo Fisher Scientific). Samples were placed on a mixer for 1 hr at 37°C and then centrifuged for 5 min at 16,000g. The MeOH was supernatant transferred to a new Eppendorf tube and hydrolyzed for 16 hr at 65°C. 100 µL 10M KOH, 625 µL chloroform, 100 µL 2N NH4OH, and 500 µL alkaline water were added to samples followed by vortexing for 5 min and centrifugation for 5 min at 16,000g. The lower organic phase was washed 3 times with alkaline water and dried under air. After dried extracts were resuspended in 60 µl Buffer B, 5 µL of sample was injected. |
Combined analysis:
| Analysis ID | AN005495 |
|---|---|
| Analysis type | MS |
| Chromatography type | Reversed phase |
| Chromatography system | Agilent 1260 Infinity II |
| Column | Thermo Hypersil GOLD C18 Selectivity (100 x 2.1 mm, 1.9 µm) |
| MS Type | ESI |
| MS instrument type | Triple quadrupole |
| MS instrument name | Agilent 6460 QQQ |
| Ion Mode | POSITIVE |
| Units | Peak area |
Chromatography:
| Chromatography ID: | CH004178 |
| Instrument Name: | Agilent 1260 Infinity II |
| Column Name: | Thermo Hypersil GOLD C18 Selectivity (100 x 2.1 mm, 1.9 µm) |
| Column Temperature: | 40°C |
| Flow Gradient: | 0 min, 40%B; 0.5 min, 40%B; 16 min, 100%B; 25.5 min, 100%B; 26 min, 40%B |
| Flow Rate: | 0.2 mL/min |
| Solvent A: | 60% methanol/40% water; 0.1% formic acid; 5 mM ammonium formate |
| Solvent B: | 100% methanol; 0.1% formic acid; 5 mM ammonium formate |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS005221 |
| Analysis ID: | AN005495 |
| Instrument Name: | Agilent 6460 QQQ |
| Instrument Type: | Triple quadrupole |
| MS Type: | ESI |
| MS Comments: | Long-chain base (LCB) species were analyzed by multiple reaction monitoring of the transition from precursor to product ions at associated optimized collision energies, and fragmentor voltages using Agilent Masshunter. The m/z values of the precursor and product ions are provided in the metabolite metadata section. |
| Ion Mode: | POSITIVE |
