Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA216113

Local Sample ID:	9510
Subject ID:	SU002333
Subject Type:	Human
Subject Species:	Homo sapiens
Taxonomy ID:	9606

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Subject:

Subject ID:	SU002333
Subject Type:	Human
Subject Species:	Homo sapiens
Taxonomy ID:	9606

Factors:

Local Sample ID	MB Sample ID	Factor Level ID	Level Value	Factor Name
9510	SA216113	FL026078	6 months	host_age

Collection:

Collection ID:	CO002326
Collection Summary:	Fecal samples were collected at birth (between 0 and 10 days post-delivery, median = 1 day, time point labeled as “birth”), during subsequent scheduled visits, and when possible during illness episodes. Stool samples were initially stored at -20°C at the International Centre for Diarrhoeal Disease Research (Bangladesh) and subsequently transferred to -80°C for shipment on dry ice to Nestlé Research where they were stored at -80°C until analysis.
Sample Type:	Feces

Treatment:

Treatment ID:	TR002345
Treatment Summary:	NA

Sample Preparation:

Sampleprep ID:	SP002339
Sampleprep Summary:	Stool samples were homogenized in 10 μL of water per milligram stool sample weight using a bead mill (TissueLyser II; Qiagen) and the aqueous homogenates were aliquoted for metabolite profiling analyses. LC-MS samples were prepared for four profiling methods as follows: HILIC-pos: Metabolites were extracted by adding 90 μL of 74.9:24.9:0.2 v/v/v acetonitrile/methanol/formic acid containing stable isotope-labeled internal standards (valine-d8, Isotec; and phenylalanine-d8, Cambridge Isotope Laboratories; Andover, MA) to a 10 μL aliquot of stool homogenate or plasma. Samples were vortexed and then centrifuged (10 min, 9,000 x g, 4°C) to pellet protein precipitates. Supernatants were transferred to glass autosampler vials containing inserts for LC-MS analysis. C8-pos: Lipids were extracted by adding 190 μL of isopropanol containing 1-dodecanoyl-2-tridecanoyl-sn-glycero-3-phosphocholine as an internal standard (Avanti Polar Lipids; Alabaster, AL) to a 10 μL aliquot of stool homogenate or plasma. Samples were vortexed and then centrifuged (10 min, 9,000 x g, ambient temperature) to pellet protein precipitates. Supernatants were transferred to glass autosampler vials containing inserts for LC-MS analysis. HILIC-neg: Metabolites were extracted by adding 120 μL of 80% methanol containing inosine-15N4, thymine-d4 and glycocholate-d4 internal standards (Cambridge Isotope Laboratories; Andover, MA) to a 30 μL aliquot of stool homogenate or plasma. Samples were vortexed and then centrifuged (10 min, 9,000 x g, 4°C) to pellet protein precipitates. Supernatants were transferred to glass autosampler vials containing inserts for LC-MS analysis. C18-neg: Metabolites were extracted by adding 90 μL of methanol containing PGE2-d4 as an internal standard (Cayman Chemical Co.; Ann Arbor, MI) to a 30 μL aliquot of stool homogenate or plasma. Samples were vortexed and then centrifuged (10 min, 9,000 x g, 4°C) to pellet protein precipitates. Supernatants were transferred to glass autosampler vials containing inserts for LC-MS analysis.

Sampleprep ID:

SP002339

Sampleprep Summary:

Stool samples were homogenized in 10 μL of water per milligram stool sample weight using a bead mill (TissueLyser II; Qiagen) and the aqueous homogenates were aliquoted for metabolite profiling analyses. LC-MS samples were prepared for four profiling methods as follows: HILIC-pos: Metabolites were extracted by adding 90 μL of 74.9:24.9:0.2 v/v/v acetonitrile/methanol/formic acid containing stable isotope-labeled internal standards (valine-d8, Isotec; and phenylalanine-d8, Cambridge Isotope Laboratories; Andover, MA) to a 10 μL aliquot of stool homogenate or plasma. Samples were vortexed and then centrifuged (10 min, 9,000 x g, 4°C) to pellet protein precipitates. Supernatants were transferred to glass autosampler vials containing inserts for LC-MS analysis. C8-pos: Lipids were extracted by adding 190 μL of isopropanol containing 1-dodecanoyl-2-tridecanoyl-sn-glycero-3-phosphocholine as an internal standard (Avanti Polar Lipids; Alabaster, AL) to a 10 μL aliquot of stool homogenate or plasma. Samples were vortexed and then centrifuged (10 min, 9,000 x g, ambient temperature) to pellet protein precipitates. Supernatants were transferred to glass autosampler vials containing inserts for LC-MS analysis. HILIC-neg: Metabolites were extracted by adding 120 μL of 80% methanol containing inosine-15N4, thymine-d4 and glycocholate-d4 internal standards (Cambridge Isotope Laboratories; Andover, MA) to a 30 μL aliquot of stool homogenate or plasma. Samples were vortexed and then centrifuged (10 min, 9,000 x g, 4°C) to pellet protein precipitates. Supernatants were transferred to glass autosampler vials containing inserts for LC-MS analysis. C18-neg: Metabolites were extracted by adding 90 μL of methanol containing PGE2-d4 as an internal standard (Cayman Chemical Co.; Ann Arbor, MI) to a 30 μL aliquot of stool homogenate or plasma. Samples were vortexed and then centrifuged (10 min, 9,000 x g, 4°C) to pellet protein precipitates. Supernatants were transferred to glass autosampler vials containing inserts for LC-MS analysis.

Combined analysis:

Analysis ID	AN003670	AN003671	AN003672	AN003673
Analysis type	MS	MS	MS	MS
Chromatography type	HILIC	Reversed phase	HILIC	Reversed phase
Chromatography system	Shimadzu Nexera X2	Shimadzu Nexera X2	Shimadzu Nexera X2	Shimadzu Nexera X2
Column	Waters Atlantis HILIC (150 x 2mm,3um)	Waters Acquity BEH C8 (100 x 2.1mm,1.7um)	Phenomenex Luna NH2 (150 x 2.1mm,3um)	Waters Acquity T3 (150 x 2.1 mm,1.7um)
MS Type	ESI	ESI	ESI	ESI
MS instrument type	Orbitrap	Orbitrap	Orbitrap	Orbitrap
MS instrument name	Thermo Q Exactive Plus Orbitrap	Thermo Q Exactive Plus Orbitrap	Thermo Q Exactive Plus Orbitrap	Thermo Q Exactive Orbitrap
Ion Mode	POSITIVE	POSITIVE	NEGATIVE	NEGATIVE
Units	Abundances	Abundances	Unitless Abundances	Abundances

Chromatography:

Chromatography ID:	CH002720
Instrument Name:	Shimadzu Nexera X2
Column Name:	Waters Atlantis HILIC (150 x 2mm,3um)
Chromatography Type:	HILIC

Chromatography ID:	CH002721
Instrument Name:	Shimadzu Nexera X2
Column Name:	Waters Acquity BEH C8 (100 x 2.1mm,1.7um)
Chromatography Type:	Reversed phase

Chromatography ID:	CH002722
Instrument Name:	Shimadzu Nexera X2
Column Name:	Phenomenex Luna NH2 (150 x 2.1mm,3um)
Chromatography Type:	HILIC

Chromatography ID:	CH002723
Instrument Name:	Shimadzu Nexera X2
Column Name:	Waters Acquity T3 (150 x 2.1 mm,1.7um)
Chromatography Type:	Reversed phase

MS:

MS ID:	MS003421
Analysis ID:	AN003670
Instrument Name:	Thermo Q Exactive Plus Orbitrap
Instrument Type:	Orbitrap
MS Type:	ESI
MS Comments:	Raw data were processed using TraceFinder 3.3 software (Thermo Fisher Scientific; Waltham, MA) and Progenesis QI (Nonlinear Dynamics; Newcastle upon Tyne, UK). Metabolite identities were confirmed using authentic reference standards or reference samples.
Ion Mode:	POSITIVE

MS ID:	MS003422
Analysis ID:	AN003671
Instrument Name:	Thermo Q Exactive Plus Orbitrap
Instrument Type:	Orbitrap
MS Type:	ESI
MS Comments:	Raw data were processed using TraceFinder 3.3 software (Thermo Fisher Scientific; Waltham, MA) and Progenesis QI (Nonlinear Dynamics; Newcastle upon Tyne, UK). Metabolite identities were confirmed using authentic reference standards or reference samples.
Ion Mode:	POSITIVE

MS ID:	MS003423
Analysis ID:	AN003672
Instrument Name:	Thermo Q Exactive Plus Orbitrap
Instrument Type:	Orbitrap
MS Type:	ESI
MS Comments:	Raw data were processed using TraceFinder 3.3 software (Thermo Fisher Scientific; Waltham, MA) and Progenesis QI (Nonlinear Dynamics; Newcastle upon Tyne, UK). Metabolite identities were confirmed using authentic reference standards or reference samples.
Ion Mode:	NEGATIVE

MS ID:	MS003424
Analysis ID:	AN003673
Instrument Name:	Thermo Q Exactive Orbitrap
Instrument Type:	Orbitrap
MS Type:	ESI
MS Comments:	Raw data were processed using TraceFinder 3.3 software (Thermo Fisher Scientific; Waltham, MA) and Progenesis QI (Nonlinear Dynamics; Newcastle upon Tyne, UK). Metabolite identities were confirmed using authentic reference standards or reference samples.
Ion Mode:	NEGATIVE