Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA351643

Local Sample ID:	2789-1
Subject ID:	SU003330
Subject Type:	Mammal
Subject Species:	Mus musculus
Taxonomy ID:	10090

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Subject:

Subject ID:	SU003330
Subject Type:	Mammal
Subject Species:	Mus musculus
Taxonomy ID:	10090

Factors:

Local Sample ID	MB Sample ID	Factor Level ID	Level Value	Factor Name
2789-1	SA351643	FL041043	fetus blood	Sample source
2789-1	SA351643	FL041043	HET-het	Genotype

Collection:

Collection ID:	CO003323
Collection Summary:	Fetuses of mice embryo at E17.5 developmental stage were isolated from pregnant females and decapitated on ice. ~50ul blood was collected from each fetus using a pipette and transferred into tubes containing 2ul 0.5M EDTA. collected by centrifugation and frozen at -80C before metabolic analysis. Cardiac blood and serum samples were also collected from the mothers of each litter using the same approach.
Sample Type:	fetus blood or cardiac blood

Treatment:

Treatment ID:	TR003339
Treatment Summary:	N/A

Sample Preparation:

Sampleprep ID:	SP003337
Sampleprep Summary:	A monophasic extraction protocol was used to extract the metabolites from the serum. To 20 µL of serum, 140 µL of chilled 6:1 MeOH/Milli-Q water containing 0.8 nmol of 13C515N valine and 0.8 nmol of 13C6 sorbitol was added. Each sample was vortexed and then incubated at 4°C for 10 min with continuous agitation (950 rpm) using an Eppendorf Thermomixer C. The samples were centrifuged at 4°C for 10 min at 12700 rpm using an Eppendorf centrifuge 5430 R. The supernatant was transferred into a fresh 1.5 mL Eppendorf tube and the cell debris was discarded. A 16 µL aliquot of each sample was pooled to create the pooled biological quality control (PBQC). Sixteen µL of each study sample and the PBQC were transferred into HPLC inserts and evaporated at 30 °C to complete dryness, using a CHRIST RVC 2-33 CD plus speed vacuum. To limit the amount of moisture present in the insert, 20 µL 100% methanol (LCMS grade) was added to each insert and evaporated using a speed vacuum.

Sampleprep ID:

SP003337

Sampleprep Summary:

A monophasic extraction protocol was used to extract the metabolites from the serum. To 20 µL of serum, 140 µL of chilled 6:1 MeOH/Milli-Q water containing 0.8 nmol of 13C515N valine and 0.8 nmol of 13C6 sorbitol was added. Each sample was vortexed and then incubated at 4°C for 10 min with continuous agitation (950 rpm) using an Eppendorf Thermomixer C. The samples were centrifuged at 4°C for 10 min at 12700 rpm using an Eppendorf centrifuge 5430 R. The supernatant was transferred into a fresh 1.5 mL Eppendorf tube and the cell debris was discarded. A 16 µL aliquot of each sample was pooled to create the pooled biological quality control (PBQC). Sixteen µL of each study sample and the PBQC were transferred into HPLC inserts and evaporated at 30 °C to complete dryness, using a CHRIST RVC 2-33 CD plus speed vacuum. To limit the amount of moisture present in the insert, 20 µL 100% methanol (LCMS grade) was added to each insert and evaporated using a speed vacuum.

Combined analysis:

Analysis ID	AN005266
Analysis type	MS
Chromatography type	GC
Chromatography system	Shimadzu GC-2030
Column	Agilent DB5-MS (30m x 0.25mm, 0.25um)
MS Type	EI
MS instrument type	Triple quadrupole
MS instrument name	Shimadzu TQ8050NX
Ion Mode	POSITIVE
Units	Relative abundance

Chromatography:

Chromatography ID:	CH003985
Chromatography Summary:	The GC-MS system used comprised of an AOC6000 autosampler, a 2030 Shimadzu gas chromatograph and a TQ8050NX triple quadrupole mass spectrometer (Shimadzu, Japan)with an electron ionisation source(-70eV). The mass spectrometer was tuned according to the manufacturer’s recommendations using tris-(perfluorobutyl)-amine (CF43). GC-MS was performed on a 30m Agilent DB-5 column with 0.25mm internal diameter column and 1µm film thickness. The injection temperature (inlet) was set at 280°C, the MS transfer line at 280°C and the ion source adjusted to 200°C. Helium was used as the carrier gas at a flow rate of 1 mL/min and argon gas was used in the collision cell to generate the MRM product ion. The analysis of the derivatised samples was performed under the following oven temperature program; 100°C start temperature, hold for 4 minutes, followed by a 10°C/min oven temperature ramp to 320°C with a following final hold for 11 minutes.
Instrument Name:	Shimadzu GC-2030
Column Name:	Agilent DB5-MS (30m x 0.25mm, 0.25um)
Column Temperature:	100 - 320
Flow Gradient:	N/A
Flow Rate:	1 mL/min
Solvent A:	N/A
Solvent B:	N/A
Chromatography Type:	GC

MS:

MS ID:	MS004997
Analysis ID:	AN005266
Instrument Name:	Shimadzu TQ8050NX
Instrument Type:	Triple quadrupole
MS Type:	EI
MS Comments:	The GC-MS system used comprised of an AOC6000 autosampler, a 2030 Shimadzu gas chromatograph and a TQ8050NX triple quadrupole mass spectrometer (Shimadzu, Japan)with an electron ionisation source(-70eV). The mass spectrometer was tuned according to the manufacturer’s recommendations using tris-(perfluorobutyl)-amine (CF43). GC-MS was performed on a 30m Agilent DB-5 column with 0.25mm internal diameter column and 1µm film thickness. The injection temperature (inlet) was set at 280°C, the MS transfer line at 280°C and the ion source adjusted to 200°C. Helium was used as the carrier gas at a flow rate of 1 mL/min and argon gas was used in the collision cell to generate the MRM product ion. The analysis of the derivatised samples was performed under the following oven temperature program; 100°C start temperature, hold for 4 minutes, followed by a 10°C/min oven temperature ramp to 320°C with a following final hold for 11 minutes.
Ion Mode:	POSITIVE