Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA235200

Local Sample ID:	QC_pos_08
Subject ID:	SU002430
Subject Type:	Cultured cells
Subject Species:	Homo sapiens
Taxonomy ID:	9606

Select appropriate tab below to view additional metadata details:

Subject:

Subject ID:	SU002430
Subject Type:	Cultured cells
Subject Species:	Homo sapiens
Taxonomy ID:	9606

Factors:

Local Sample ID	MB Sample ID	Factor Level ID	Level Value	Factor Name
QC_pos_08	SA235200	FL029178	QC	Class

Collection:

Collection ID:	CO002423
Collection Summary:	Two hours before metabolite extraction, the cell culture medium was replaced with fresh medium. After discarding the medium in each culture dish, the cells were quickly rinsed twice with cold isotonic saline (0.9% NaCl [w/v], 4 °C). Water (1.5 mL) was added to each dish, and then the dishes were stored in a freezer (−80 °C) for 20 min before extraction.
Sample Type:	Cultured cells

Treatment:

Treatment ID:	TR002442
Treatment Summary:	SW480 cells and SW620 cells were cultured in DMEM medium supplemented with 10% FBS at 37 °C under a 5% CO2 atmosphere, HT-29 and COLO 205 cells were cultured in RPMI 1640 medium supplemented with 10% FBS at 37 °C under a 5% CO2 atmosphere.
Cell Media:	DMEM medium supplemented with 10% FBS is for SW480 cells and SW620 cells, while RPMI 1640 medium supplemented with 10% FBS is for HT-29 and COLO 205 cells.
Cell Envir Cond:	at 37 °C under a 5% CO2 atmosphere
Cell Harvesting:	After discarding the medium in each culture dish, the cells were quickly rinsed twice with cold isotonic saline (0.9% NaCl [w/v], 4 °C). Water (1.5 mL) was added to each dish, and then the dishes were stored in a freezer (−80 °C) for 20 min before extraction. Then, cells were collected by scraping with a cell scraper.
Cell Pct Confluence:	70%-80%

Sample Preparation:

Sampleprep ID:	SP002436
Sampleprep Summary:	Cell lysates (20 μL) were removed for a protein assay, and the remaining cell lysates were extracted by the addition of 4.5 mL of methanol containing 3 μg of internal standard (acetaminophen). The cells were completely collected with a cell scraper. Finally, the suspension of cell debris in each dish was transferred to an Eppendorf tube, vigorously vortexed for 5 min, and then centrifuged at 10,000×g for 10 min at 4 °C. The supernatant was transferred to another tube and evaporated to dryness in a Refrigerated CentriVap Benchtop Vacuum Concentrator (Labconco Corporation, Kansas City, MO). The pellets were reconstituted with methanol-water (75:25) and centrifuged twice at 12,000×g for 15 min at 4 °C.

Sampleprep ID:

SP002436

Sampleprep Summary:

Cell lysates (20 μL) were removed for a protein assay, and the remaining cell lysates were extracted by the addition of 4.5 mL of methanol containing 3 μg of internal standard (acetaminophen). The cells were completely collected with a cell scraper. Finally, the suspension of cell debris in each dish was transferred to an Eppendorf tube, vigorously vortexed for 5 min, and then centrifuged at 10,000×g for 10 min at 4 °C. The supernatant was transferred to another tube and evaporated to dryness in a Refrigerated CentriVap Benchtop Vacuum Concentrator (Labconco Corporation, Kansas City, MO). The pellets were reconstituted with methanol-water (75:25) and centrifuged twice at 12,000×g for 15 min at 4 °C.

Combined analysis:

Analysis ID	AN003824	AN003825
Analysis type	MS	MS
Chromatography type	Reversed phase	Reversed phase
Chromatography system	UPLC Ultimate 3000 system	UPLC Ultimate 3000 system
Column	Phenomenex Kinetex C18 (100 x 2.1mm,2.6um)	Phenomenex Kinetex C18 (100 x 2.1mm,2.6um)
MS Type	ESI	ESI
MS instrument type	Orbitrap	Orbitrap
MS instrument name	Thermo Q Exactive HF hybrid Orbitrap	Thermo Q Exactive HF hybrid Orbitrap
Ion Mode	POSITIVE	NEGATIVE
Units	Peak area	Peak area

Chromatography:

Chromatography ID:	CH002830
Instrument Name:	UPLC Ultimate 3000 system
Column Name:	Phenomenex Kinetex C18 (100 x 2.1mm,2.6um)
Column Temperature:	40 °C
Flow Gradient:	10% B (0 min) → 30% B (1 min) → 95% B (19 min) → 95% B (20 min)
Flow Rate:	0.4 mL/min
Injection Temperature:	4 °C
Solvent A:	100% water; 0.1% formic acid
Solvent B:	100% acetonitrile; 0.1% formic acid
Chromatography Type:	Reversed phase

MS:

MS ID:	MS003566
Analysis ID:	AN003824
Instrument Name:	Thermo Q Exactive HF hybrid Orbitrap
Instrument Type:	Orbitrap
MS Type:	ESI
MS Comments:	All operations and acquisitions were controlled by Xcalibur 2.0.7 (Thermo Fisher Scientific, Bremen, Germany).UPLC-HRMS data were analyzed using MS-DIAL software after conversion to mzML file format using MSConvert (Version 3.0, ProteiWizard).
Ion Mode:	POSITIVE
Capillary Temperature:	300 °C
Capillary Voltage:	+3.5 kV

MS ID:	MS003567
Analysis ID:	AN003825
Instrument Name:	Thermo Q Exactive HF hybrid Orbitrap
Instrument Type:	Orbitrap
MS Type:	ESI
MS Comments:	All operations and acquisitions were controlled by Xcalibur 2.0.7 (Thermo Fisher Scientific, Bremen, Germany).UPLC-HRMS data were analyzed using MS-DIAL software after conversion to mzML file format using MSConvert (Version 3.0, ProteiWizard).
Ion Mode:	NEGATIVE
Capillary Temperature:	300 °C
Capillary Voltage:	-2.5 kV