Summary of Study ST003999

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 PR002504. The data can be accessed directly via it's Project DOI: 10.21228/M8ZV87 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	ST003999
Study Title	Measuring PEMT activity in HCMV infected fibroblasts using 13C-methionine.
Study Summary	HCMV promotes the levels of PC lipids in infection. The Kennedy pathway is the canonical host pathway for de novo PC lipid synthesis. Host cells encode alternative pathways for PC synthesis such as the Phosphatidylethanolamine N-methyltransferase (PEMT) pathway. Here, we tested if HCMV promotes metabolic activity in the PEMT pathway. We used 13C-methionine to track the metabolism of labeled tracer molecule into the head group of PC lipids thus, measuring PEMT pathway activity. We find that HCMV and uninfected cells exhibit similarly low PEMT metabolic activity and percentage of labeled PCs. Moreover, when cells were engineered to overexpress the host PEMT enzyme, PC labeling via PEMT increased when expression was induced with doxycycline. Overexpressing GFP and feeding labeled molecule resulted in labeling similar to mock and infected labeling percentages, <5%. Thus, PEMT is not an active pathway in the primary human fibroblasts used in our studies and HCMV infection does not increase metabolic activity of PEMT to synthesize PCs.
Institute	University of Arizona
Department	Immunobiology
Laboratory	John G. Purdy, PhD
Last Name	Kline
First Name	Ian
Address	1657 E Helen St, Tucson, AZ 85721
Email	ikline@arizona.edu
Phone	5209092596
Submit Date	2025-06-09
Raw Data Available	Yes
Raw Data File Type(s)	mzXML
Analysis Type Detail	LC-MS
Release Date	2025-06-25
Release Version	1

Select appropriate tab below to view additional metadata details:

Project:

Project ID:	PR002504
Project DOI:	doi: 10.21228/M8ZV87
Project Title:	Measuring PEMT activity in HCMV infected fibroblasts using 13C-methionine.
Project Summary:	Human cytomegalovirus (HCMV) is a common herpesvirus that establishes a lifelong and persistent infection in its human host. HCMV infection in most people does not cause overt disease. However, in immunocompromised individuals, severe CMV-associated disease can lead to permanent disabilities and even death. Additionally, congenital CMV is the leading infectious cause of birth defects. Viruses have evolved to hijack host metabolic pathways to facilitate their replication cycle. We previously reported HCMV infection increases phosphatidylcholine (PC) lipid levels, including PCs with VLCFAs. To expand upon the previously reported PC phenotype in HCMV infection, we determined the PC lipidome of several infected cell types grown under various growth conditions. Additionally, we determined which host pathways HCMV reprograms to induce PC lipid synthesis and describe when during infection PC lipids changes occur.
Institute:	University of Arizona
Department:	Immunobiology
Laboratory:	John G. Purdy, PhD
Last Name:	Kline
First Name:	Ian
Address:	1657 E Helen St, Tucson, AZ 85721
Email:	ikline@arizona.edu
Phone:	5209092596
Funding Source:	National Institute of Health (NIH) National Institute of Allergy and Infectious Disease (NIAID) R01AI162671, R01AI155539, F32AI178919, and National Institute of Aging (NIA) T32AG058503 award.

Subject:

Subject ID:	SU004136
Subject Type:	Cultured cells
Subject Species:	Homo sapiens
Taxonomy ID:	9606
Gender:	Male
Cell Strain Details:	HFF
Cell Passage Number:	<30

Subject ID:	SU004137
Subject Type:	Cultured cells
Subject Species:	Homo sapiens
Taxonomy ID:	9606
Gender:	Male
Cell Strain Details:	HFF
Cell Passage Number:	<30

Factors:

Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)

mb_sample_id	local_sample_id	Sample source	Time (hpi)	Infection status	Inducible gene	Doxycycline
SA461266	20230316_pos_24hpi_pLV-GFP_1ug_a	Primary human fibroblast cells (HFF)	24	Uninfected	GFP	1ug/mL
SA461267	20230316_pos_24hpi_pLV-GFP_1ug_b	Primary human fibroblast cells (HFF)	24	Uninfected	GFP	1ug/mL
SA461310	20230316_pos_24hpi_pLV-GFP_1ug_a	Primary human fibroblast cells (HFF)	24	Uninfected	GFP	1ug/mL
SA461311	20230316_pos_24hpi_pLV-GFP_1ug_b	Primary human fibroblast cells (HFF)	24	Uninfected	GFP	1ug/mL
SA461268	20230316_pos_24hpi_pLV-PEMT_1ug_a	Primary human fibroblast cells (HFF)	24	Uninfected	PEMT	1ug/mL
SA461269	20230316_pos_24hpi_pLV-PEMT_1ug_b	Primary human fibroblast cells (HFF)	24	Uninfected	PEMT	1ug/mL
SA461312	20230316_pos_24hpi_pLV-PEMT_1ug_a	Primary human fibroblast cells (HFF)	24	Uninfected	PEMT	1ug/mL
SA461313	20230316_pos_24hpi_pLV-PEMT_1ug_b	Primary human fibroblast cells (HFF)	24	Uninfected	PEMT	1ug/mL
SA461270	20230806_13C-methionine_pos_TB40E_choline_48_a	Primary human fibroblast cells (HFF)	48	HCMV	n/a	n/a
SA461271	20231007_13C-methionine_pos_TB40E_48_b	Primary human fibroblast cells (HFF)	48	HCMV	n/a	n/a
SA461272	20231007_13C-methionine_pos_TB40E_48_a	Primary human fibroblast cells (HFF)	48	HCMV	n/a	n/a
SA461273	20230905_13C_methionine_pos_choline_TB40E_48_b	Primary human fibroblast cells (HFF)	48	HCMV	n/a	n/a
SA461274	20230806_13C-methionine_pos_TB40E_choline_48_b	Primary human fibroblast cells (HFF)	48	HCMV	n/a	n/a
SA461275	20230905_13C_methionine_pos_choline_TB40E_48_a	Primary human fibroblast cells (HFF)	48	HCMV	n/a	n/a
SA461314	20230806_13C-methionine_pos_TB40E_choline_48_a	Primary human fibroblast cells (HFF)	48	HCMV	n/a	n/a
SA461315	20231007_13C-methionine_pos_TB40E_48_b	Primary human fibroblast cells (HFF)	48	HCMV	n/a	n/a
SA461316	20231007_13C-methionine_pos_TB40E_48_a	Primary human fibroblast cells (HFF)	48	HCMV	n/a	n/a
SA461317	20230905_13C_methionine_pos_choline_TB40E_48_b	Primary human fibroblast cells (HFF)	48	HCMV	n/a	n/a
SA461318	20230806_13C-methionine_pos_TB40E_choline_48_b	Primary human fibroblast cells (HFF)	48	HCMV	n/a	n/a
SA461319	20230905_13C_methionine_pos_choline_TB40E_48_a	Primary human fibroblast cells (HFF)	48	HCMV	n/a	n/a
SA461276	20230905_13C_methionine_pos_GFP_1ug_48_a	Primary human fibroblast cells (HFF)	48	Uninfected	GFP	1ug/mL
SA461277	20230905_13C_methionine_pos_GFP_1ug_48_b	Primary human fibroblast cells (HFF)	48	Uninfected	GFP	1ug/mL
SA461320	20230905_13C_methionine_pos_GFP_1ug_48_a	Primary human fibroblast cells (HFF)	48	Uninfected	GFP	1ug/mL
SA461321	20230905_13C_methionine_pos_GFP_1ug_48_b	Primary human fibroblast cells (HFF)	48	Uninfected	GFP	1ug/mL
SA461280	20230905_13C_methionine_pos_choline_mock_48_b	Primary human fibroblast cells (HFF)	48	Uninfected	n/a	n/a
SA461281	20230806_13C-methionine_pos_mock_choline_48_b	Primary human fibroblast cells (HFF)	48	Uninfected	n/a	n/a
SA461282	20231007_13C-methionine_pos_mock_48_b	Primary human fibroblast cells (HFF)	48	Uninfected	n/a	n/a
SA461283	20231007_13C-methionine_pos_mock_48_a	Primary human fibroblast cells (HFF)	48	Uninfected	n/a	n/a
SA461284	20230806_13C-methionine_pos_mock_choline_48_a	Primary human fibroblast cells (HFF)	48	Uninfected	n/a	n/a
SA461285	20230905_13C_methionine_pos_choline_mock_48_a	Primary human fibroblast cells (HFF)	48	Uninfected	n/a	n/a
SA461324	20230905_13C_methionine_pos_choline_mock_48_b	Primary human fibroblast cells (HFF)	48	Uninfected	n/a	n/a
SA461325	20230806_13C-methionine_pos_mock_choline_48_b	Primary human fibroblast cells (HFF)	48	Uninfected	n/a	n/a
SA461326	20231007_13C-methionine_pos_mock_48_b	Primary human fibroblast cells (HFF)	48	Uninfected	n/a	n/a
SA461327	20231007_13C-methionine_pos_mock_48_a	Primary human fibroblast cells (HFF)	48	Uninfected	n/a	n/a
SA461328	20230806_13C-methionine_pos_mock_choline_48_a	Primary human fibroblast cells (HFF)	48	Uninfected	n/a	n/a
SA461329	20230905_13C_methionine_pos_choline_mock_48_a	Primary human fibroblast cells (HFF)	48	Uninfected	n/a	n/a
SA461278	20230905_13C_methionine_pos_PEMT_1ug_48_b	Primary human fibroblast cells (HFF)	48	Uninfected	PEMT	1ug/mL
SA461279	20230905_13C_methionine_pos_PEMT_1ug_48_a	Primary human fibroblast cells (HFF)	48	Uninfected	PEMT	1ug/mL
SA461322	20230905_13C_methionine_pos_PEMT_1ug_48_b	Primary human fibroblast cells (HFF)	48	Uninfected	PEMT	1ug/mL
SA461323	20230905_13C_methionine_pos_PEMT_1ug_48_a	Primary human fibroblast cells (HFF)	48	Uninfected	PEMT	1ug/mL
SA461286	20230316_pos_6hpi_pLV-GFP_1ug_a	Primary human fibroblast cells (HFF)	6	Uninfected	GFP	1ug/mL
SA461287	20230316_pos_6hpi_pLV-GFP_1ug_b	Primary human fibroblast cells (HFF)	6	Uninfected	GFP	1ug/mL
SA461330	20230316_pos_6hpi_pLV-GFP_1ug_a	Primary human fibroblast cells (HFF)	6	Uninfected	GFP	1ug/mL
SA461331	20230316_pos_6hpi_pLV-GFP_1ug_b	Primary human fibroblast cells (HFF)	6	Uninfected	GFP	1ug/mL
SA461288	20230316_pos_6hpi_pLV-PEMT_1ug_a	Primary human fibroblast cells (HFF)	6	Uninfected	PEMT	1ug/mL
SA461289	20230316_pos_6hpi_pLV-PEMT_1ug_b	Primary human fibroblast cells (HFF)	6	Uninfected	PEMT	1ug/mL
SA461332	20230316_pos_6hpi_pLV-PEMT_1ug_a	Primary human fibroblast cells (HFF)	6	Uninfected	PEMT	1ug/mL
SA461333	20230316_pos_6hpi_pLV-PEMT_1ug_b	Primary human fibroblast cells (HFF)	6	Uninfected	PEMT	1ug/mL
SA461290	20231007_13C-methionine_pos_TB40E_72_b	Primary human fibroblast cells (HFF)	72	HCMV	n/a	n/a
SA461291	20231007_13C-methionine_pos_TB40E_72_a	Primary human fibroblast cells (HFF)	72	HCMV	n/a	n/a
SA461292	20230905_13C_methionine_pos_choline_TB40E_72_a	Primary human fibroblast cells (HFF)	72	HCMV	n/a	n/a
SA461293	20230905_13C_methionine_pos_choline_TB40E_72_b	Primary human fibroblast cells (HFF)	72	HCMV	n/a	n/a
SA461294	20230806_13C-methionine_pos_TB40E_choline_72_b	Primary human fibroblast cells (HFF)	72	HCMV	n/a	n/a
SA461295	20230806_13C-methionine_pos_TB40E_choline_72_a	Primary human fibroblast cells (HFF)	72	HCMV	n/a	n/a
SA461334	20231007_13C-methionine_pos_TB40E_72_b	Primary human fibroblast cells (HFF)	72	HCMV	n/a	n/a
SA461335	20231007_13C-methionine_pos_TB40E_72_a	Primary human fibroblast cells (HFF)	72	HCMV	n/a	n/a
SA461336	20230905_13C_methionine_pos_choline_TB40E_72_a	Primary human fibroblast cells (HFF)	72	HCMV	n/a	n/a
SA461337	20230905_13C_methionine_pos_choline_TB40E_72_b	Primary human fibroblast cells (HFF)	72	HCMV	n/a	n/a
SA461338	20230806_13C-methionine_pos_TB40E_choline_72_b	Primary human fibroblast cells (HFF)	72	HCMV	n/a	n/a
SA461339	20230806_13C-methionine_pos_TB40E_choline_72_a	Primary human fibroblast cells (HFF)	72	HCMV	n/a	n/a
SA461296	20231007_13C-methionine_pos_mock_72_a	Primary human fibroblast cells (HFF)	72	Uninfected	-	n/a
SA461340	20231007_13C-methionine_pos_mock_72_a	Primary human fibroblast cells (HFF)	72	Uninfected	-	n/a
SA461297	20230316_pos_72hpi_pLV-GFP_1ug_b	Primary human fibroblast cells (HFF)	72	Uninfected	GFP	1ug/mL
SA461298	20230905_13C_methionine_pos_GFP_1ug_72_b	Primary human fibroblast cells (HFF)	72	Uninfected	GFP	1ug/mL
SA461299	20230316_pos_72hpi_pLV-GFP_1ug_a	Primary human fibroblast cells (HFF)	72	Uninfected	GFP	1ug/mL
SA461300	20230905_13C_methionine_pos_GFP_1ug_72_a	Primary human fibroblast cells (HFF)	72	Uninfected	GFP	1ug/mL
SA461341	20230316_pos_72hpi_pLV-GFP_1ug_b	Primary human fibroblast cells (HFF)	72	Uninfected	GFP	1ug/mL
SA461342	20230905_13C_methionine_pos_GFP_1ug_72_b	Primary human fibroblast cells (HFF)	72	Uninfected	GFP	1ug/mL
SA461343	20230316_pos_72hpi_pLV-GFP_1ug_a	Primary human fibroblast cells (HFF)	72	Uninfected	GFP	1ug/mL
SA461344	20230905_13C_methionine_pos_GFP_1ug_72_a	Primary human fibroblast cells (HFF)	72	Uninfected	GFP	1ug/mL
SA461305	20230806_13C-methionine_pos_mock_choline_72_a	Primary human fibroblast cells (HFF)	72	Uninfected	n/a	n/a
SA461306	20230806_13C-methionine_pos_mock_choline_72_b	Primary human fibroblast cells (HFF)	72	Uninfected	n/a	n/a
SA461307	20231007_13C-methionine_pos_mock_72_b	Primary human fibroblast cells (HFF)	72	Uninfected	n/a	n/a
SA461308	20230905_13C_methionine_pos_choline_mock_72_b	Primary human fibroblast cells (HFF)	72	Uninfected	n/a	n/a
SA461309	20230905_13C_methionine_pos_choline_mock_72_a	Primary human fibroblast cells (HFF)	72	Uninfected	n/a	n/a
SA461349	20230806_13C-methionine_pos_mock_choline_72_a	Primary human fibroblast cells (HFF)	72	Uninfected	n/a	n/a
SA461350	20230806_13C-methionine_pos_mock_choline_72_b	Primary human fibroblast cells (HFF)	72	Uninfected	n/a	n/a
SA461351	20231007_13C-methionine_pos_mock_72_b	Primary human fibroblast cells (HFF)	72	Uninfected	n/a	n/a
SA461352	20230905_13C_methionine_pos_choline_mock_72_b	Primary human fibroblast cells (HFF)	72	Uninfected	n/a	n/a
SA461353	20230905_13C_methionine_pos_choline_mock_72_a	Primary human fibroblast cells (HFF)	72	Uninfected	n/a	n/a
SA461301	20230905_13C_methionine_pos_PEMT_1ug_72_a	Primary human fibroblast cells (HFF)	72	Uninfected	PEMT	1ug/mL
SA461302	20230905_13C_methionine_pos_PEMT_1ug_72_b	Primary human fibroblast cells (HFF)	72	Uninfected	PEMT	1ug/mL
SA461303	20230316_pos_72hpi_pLV-PEMT_1ug_a	Primary human fibroblast cells (HFF)	72	Uninfected	PEMT	1ug/mL
SA461304	20230316_pos_72hpi_pLV-PEMT_1ug_b	Primary human fibroblast cells (HFF)	72	Uninfected	PEMT	1ug/mL
SA461345	20230905_13C_methionine_pos_PEMT_1ug_72_a	Primary human fibroblast cells (HFF)	72	Uninfected	PEMT	1ug/mL
SA461346	20230905_13C_methionine_pos_PEMT_1ug_72_b	Primary human fibroblast cells (HFF)	72	Uninfected	PEMT	1ug/mL
SA461347	20230316_pos_72hpi_pLV-PEMT_1ug_a	Primary human fibroblast cells (HFF)	72	Uninfected	PEMT	1ug/mL
SA461348	20230316_pos_72hpi_pLV-PEMT_1ug_b	Primary human fibroblast cells (HFF)	72	Uninfected	PEMT	1ug/mL

Showing results 1 to 88 of 88

Showing results 1 to 88 of 88

Collection:

Collection ID:	CO004129
Collection Summary:	General Notes: -Work over ice when possible (during scraping and between vortex steps etc.). -Chloroform leeches plastics. Avoid contact with plastic caps, tubes and gloves around glass vial tops. -Work in manageable batch numbers. A batch of 8-16 samples at a time is common. -Clean syringes using chloroform before and after collection. Use a unique syringe for each sample (the same syringe can be used if A/B technical replicates are used. Recommend a quick flush of the syringe using 500uL chloroform before moving from A to B). Begin by counting cell numbers for each sample using a dedicated well meant for MS normalization based on cell count. In a 6-well plate, wash cells 2x with cold PBS. Add 1mL of cold 50% methanol to each well and scrape cells into glass vials. Add 500uL of chloroform. Vortex on low setting (careful to avoid splashing chloroform onto plastic caps and liners). Centrifuge @1000g for 5 mins. There should be a clear phase separation of methanol and cell debris on top, while the lower phase contains chloroform and lipids. Use a syringe to carefully extract the lower phase without transferring cell debris from the top layer. Transfer to clean vial and place on ice. Once all lipids have been extracted, add 500uL of chloroform and repeat the process again once more. In total each sample should contain ~1mL of chloroform and lipids from two extractions. Carefully, dry lipids under nitrogen gas. Avoid direct high pressure air flow onto chloroform:lipid solution as it can splash high up in vial walls. Store at -80C for up to 30 days. Samples may become unstable and degrade over time.
Sample Type:	Cultured cells
Collection Location:	Fume hood required when working with chloroform.
Collection Frequency:	Perform extraction twice on each sample.
Collection Duration:	1-2 hours dependent upon batch size
Volumeoramount Collected:	~1mL chloroform:lipid soluition
Storage Conditions:	-80℃
Collection Vials:	Glass vials, no plastic! (plastic cap is okay)
Storage Vials:	Glass vials, no plastic! (plastic cap is okay)
Collection Tube Temp:	25
Additives:	UHPLC 100% chloroform

Collection ID:	CO004130
Collection Summary:	General Notes: -Work over ice when possible (during scraping and between vortex steps etc.). -Chloroform leeches plastics. Avoid contact with plastic caps, tubes and gloves around glass vial tops. -Work in manageable batch numbers. A batch of 8-16 samples at a time is common. -Clean syringes using chloroform before and after collection. Use a unique syringe for each sample (the same syringe can be used if A/B technical replicates are used. Recommend a quick flush of the syringe using 500 μL chloroform before moving from A to B). Begin by counting cell numbers for each sample using a dedicated well meant for MS normalization based on cell count. In a 6-well plate, wash cells 2x with cold PBS. Add 1 mL of cold 50% methanol to each well and scrape cells into glass vials. Add 500 μL of chloroform. Vortex on low setting (careful to avoid splashing chloroform onto plastic caps and liners). Centrifuge @1000g for 5 mins. There should be a clear phase separation of methanol and cell debris on top, while the lower phase contains chloroform and lipids. Use a syringe to carefully extract the lower phase without transferring cell debris from the top layer. Transfer to clean vial and place on ice. Once all lipids have been extracted, add 500 μL of chloroform and repeat the process again once more. In total each sample should contain ~1 mL of chloroform and lipids from two extractions. Carefully, dry lipids under nitrogen gas. Avoid direct high pressure air flow onto chloroform:lipid solution as it can splash high up in vial walls. Store at -80℃ for up to 30 days. Samples may become unstable and degrade over time.
Sample Type:	Cultured cells
Collection Location:	Fume hood required when working with chloroform.
Collection Frequency:	Perform extraction twice on each sample.
Collection Duration:	1-2 hours dependent upon batch size
Volumeoramount Collected:	~1 mL chloroform:lipid soluition
Storage Conditions:	-80℃
Collection Vials:	Glass vials, no plastic! (plastic cap is okay)
Storage Vials:	Glass vials, no plastic! (plastic cap is okay)
Collection Tube Temp:	25
Additives:	UHPLC 100% chloroform

Treatment:

Treatment ID:	TR004145
Treatment Summary:	Cells were grown to full confluence and held for 3-days in Dulbecco's modified eagle medium (DMEM) with 10% fetal bovine serum (FBS). 24 hours prior to infection, cells were starved of serum in DMEM lacking FBS. Cells were HCMV-infected or mock-infected in serum-free DMEM for 1 hour. 1 hpi, 13C-methionine isotopic tracer was fed to cells in serum-free growth medium we previously prepared (based on the published DMEM Gibco recipe). Tracer media contains 13C-methionine in place 12C-methionine. Tracer molecule was added at the same concentration non-tracer molecule is normally present in DMEM (13C-methionine (30mg/L)). Medium was replaced at 48 hpi.

Treatment ID:	TR004146
Treatment Summary:	Cells were grown to full confluence and held for 3-days in Dulbecco's modified eagle medium (DMEM) with 10% fetal bovine serum (FBS). 24 hours prior to infection, cells were starved of serum in DMEM lacking FBS. Cells were HCMV-infected or mock-infected in serum-free DMEM for 1 hour. 1 hpi, 13C-methionine isotopic tracer was fed to cells in serum-free growth medium we previously prepared (based on the published DMEM Gibco recipe). Tracer media contains 13C-methionine in place 12C-methionine. Tracer molecule was added at the same concentration non-tracer molecule is normally present in DMEM (13C-methionine (30mg/L)). Medium was replaced at 48 hpi.

Sample Preparation:

Sampleprep ID:	SP004142
Sampleprep Summary:	Before starting, calculate the volume of 1:1:1 choloroform:methanol:isopropanol resuspension buffer needed for each sample. Volume of 1:1:1 is dependent on normalize cell count from experiment. Use 200uL of 1:1:1 for every 2E5 cells. When ready to start resuspension, remove dried lipids from -80C storage and resuspend in volume of 1:1:1 solution calculated for each sample. No cell conditions use 200uL. Use gentle vortex to allow for dried lipids from vial wall to get into solution. Prepare "blank" vials of 1:1:1 for buffer background analysis. Store lipids in autosampler between 4-7C.

Sampleprep ID:	SP004143
Sampleprep Summary:	Before starting, calculate the volume of 1:1:1 choloroform:methanol:isopropanol resuspension buffer needed for each sample. Volume of 1:1:1 is dependent on normalize cell count from experiment. Use 200uL of 1:1:1 for every 2E5 cells. When ready to start resuspension, remove dried lipids from -80C storage and resuspend in volume of 1:1:1 solution calculated for each sample. No cell conditions use 200uL. Use gentle vortex to allow for dried lipids from vial wall to get into solution. Prepare "blank" vials of 1:1:1 for buffer background analysis. Store lipids in autosampler between 4-7C.

Chromatography:

Chromatography ID:	CH005005
Instrument Name:	Thermo Vanquish
Column Name:	Phenomenex Kinetex C18 (100 x 2.1mm,2.6um)
Column Temperature:	60℃
Flow Gradient:	75% solvent A–25% solvent B for 2 min, 35% solvent A–65% solvent B for 2min at a curve value of 4, a hold at 35% solvent A–65% solvent B for 1min, 0% solvent A–100% solvent B for 11min at a curve value of 4, and a hold at 0% solvent A–100% solvent B for 4 min.
Flow Rate:	0.25 mL/min
Solvent A:	40% Water/60% Methanol; 10mM Ammonium formate; 0.1% Formic acid
Solvent B:	10% Methanol/90% Isopropanol; 10mM Ammonium formate; 0.1% Formic acid
Chromatography Type:	Reversed phase

Chromatography ID:	CH005006
Instrument Name:	Thermo Vanquish
Column Name:	Phenomenex Kinetex C18 (100 x 2.1mm,2.6um)
Column Temperature:	60℃
Flow Gradient:	75% solvent A–25% solvent B for 2 min, 35% solvent A–65% solvent B for 2min at a curve value of 4, a hold at 35% solvent A–65% solvent B for 1min, 0% solvent A–100% solvent B for 11min at a curve value of 4, and a hold at 0% solvent A–100% solvent B for 4 min.
Flow Rate:	0.25 mL/min
Solvent A:	40% Water/60% Methanol; 10mM Ammonium formate; 0.1% Formic acid
Solvent B:	10% Methanol/90% Isopropanol; 10mM Ammonium formate; 0.1% Formic acid
Chromatography Type:	Reversed phase

Analysis:

Analysis ID:	AN006593
Analysis Type:	MS
Analysis Protocol File:	Cambridge_isotope_13C-methionine_MSDS.pdf
Chromatography ID:	CH005005
Num Factors:	13
Num Metabolites:	216
Units:	Peak Area

Analysis ID:	AN006594
Analysis Type:	MS
Analysis Protocol File:	Cambridge_isotope_13C-methionine_MSDS.pdf
Chromatography ID:	CH005006
Num Factors:	13
Num Metabolites:	216
Units:	Peak Area