Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA235671

Local Sample ID:	JM SQ 4
Subject ID:	SU002437
Subject Type:	Bacteria
Subject Species:	Escherichia coli
Taxonomy ID:	562
Genotype Strain:	BW25113

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Subject:

Subject ID:	SU002437
Subject Type:	Bacteria
Subject Species:	Escherichia coli
Taxonomy ID:	562
Genotype Strain:	BW25113

Factors:

Local Sample ID	MB Sample ID	Factor Level ID	Level Value	Factor Name
JM SQ 4	SA235671	FL029240	SQ	Factor

Collection:

Collection ID:	CO002430
Collection Summary:	Cells were metabolically arrested by infusing with ice-cold PBS and washed with PBS.
Sample Type:	Bacterial cells

Treatment:

Treatment ID:	TR002449
Treatment Summary:	E. coli adapted to growth on the relevant substrate was grown on M9 minimal medium containing either Glc or SQ as a sole carbon source until mid-log phase was achieved.

Sample Preparation:

Sampleprep ID:	SP002443
Sampleprep Summary:	Cell pellets were resuspended in chilled extraction solution (500 µL) comprised of 3:1 MeOH: H2O and (13C5, 15N)valine (1 mM, 0.5 µL) and (13C6)sorbitol (1 mM, 0.5 µL) as internal standards. The suspensions were subjected to freeze-thaw cycles to facilitate lysis of the cells (30 s in liquid N2, 30 s in a dry ice/EtOH bath for 10 cycles), and shaken (9000 rpm, 10 min, 2°C). The samples were then centrifuged (12700 rpm, 5 min, 1°C). The cell lysate supernatant was transferred into glass inserts and dried. All samples were washed with MeOH (50 µL). All samples were methoximated with methoxylamine hydrochloride solution (30 mg/mL in pyridine, 20 µL) for 2 h at 37°C, followed by trimethylsilyation in BSTFA + 1% TMCS (20 µL) for 1 h at 37°C with continuous mixing. Samples were incubated at rt for 1 h prior to GC-MS analysis.

Sampleprep ID:

SP002443

Sampleprep Summary:

Cell pellets were resuspended in chilled extraction solution (500 µL) comprised of 3:1 MeOH: H2O and (13C5, 15N)valine (1 mM, 0.5 µL) and (13C6)sorbitol (1 mM, 0.5 µL) as internal standards. The suspensions were subjected to freeze-thaw cycles to facilitate lysis of the cells (30 s in liquid N2, 30 s in a dry ice/EtOH bath for 10 cycles), and shaken (9000 rpm, 10 min, 2°C). The samples were then centrifuged (12700 rpm, 5 min, 1°C). The cell lysate supernatant was transferred into glass inserts and dried. All samples were washed with MeOH (50 µL). All samples were methoximated with methoxylamine hydrochloride solution (30 mg/mL in pyridine, 20 µL) for 2 h at 37°C, followed by trimethylsilyation in BSTFA + 1% TMCS (20 µL) for 1 h at 37°C with continuous mixing. Samples were incubated at rt for 1 h prior to GC-MS analysis.

Combined analysis:

Analysis ID	AN003833
Analysis type	MS
Chromatography type	GC
Chromatography system	Shimadzu GC-2010
Column	J&W DB-5 (30 m × 0.25mm,1.00um)
MS Type	EI
MS instrument type	Triple quadrupole
MS instrument name	Shimazu TQ8040
Ion Mode	UNSPECIFIED
Units	a.u.

Chromatography:

Chromatography ID:	CH002838
Instrument Name:	Shimadzu GC-2010
Column Name:	J&W DB-5 (30 m × 0.25mm,1.00um)
Chromatography Type:	GC

MS:

MS ID:	MS003575
Analysis ID:	AN003833
Instrument Name:	Shimazu TQ8040
Instrument Type:	Triple quadrupole
MS Type:	EI
MS Comments:	Please see attached file.
Ion Mode:	UNSPECIFIED