Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA072518

Local Sample ID:	SB12
Subject ID:	SU001119
Subject Type:	Mammal
Subject Species:	Rattus norvegicus
Taxonomy ID:	10116
Genotype Strain:	F344/Ntac-Apc-+/Pirc
Age Or Age Range:	30
Animal Animal Supplier:	Taconic and Envigo
Animal Housing:	Conventional,
Animal Feed:	Labdiet 5058
Animal Water:	Autoclaved, purified with sulfuric acid

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Sample Preparation:

Sampleprep ID:	SP001126
Sampleprep Summary:	Fecal samples were lyophilized at -20 ˚C using 0.1 millibar of vacuum pressure, following which dried samples (30 mg) were extracted sequentially for both UHPLC-MS and GC-MS. The dried samples were first treated with 1.0 mL of 80% MeOH containing 18 µg/mL umbelliferone, sonicated for 5 minutes and centrifuged for 40 minutes at 3000 g at 10 ºC. 0.5 mL of supernatant was used for UHPLC-MS analysis after a subsequent spin at 5000 g at 10 ºC for 20 minutes and transferring 250 µL of the sample into glass autosampler vials with inserts. For GC-MS (Gas Chromatography-Mass Spectrometry) analyses of primary polar metabolites, 0.5 mL water was added the remaining extract used above for the UHPLC preparation, sonicated for 5 min, extracted for 30min, and centrifuged at 3000 g. 0.5 mL of the polar extract was subsequently dried under nitrogen and derivatized using previously established protocols (84). Briefly, N-Methyl-N-(trimethylsilyl) trifluoroacetamide (MSTFA) with 1 % TMCS (2,2,2-Trifluoro-N-methyl-N-(trimethylsilyl)-acetamide, Chlorotrimethylsilane) was used to derivatize the polar metabolites, after treatment with methoxyamine-HCl-pyridine.

Sampleprep ID:

SP001126

Sampleprep Summary:

Fecal samples were lyophilized at -20 ˚C using 0.1 millibar of vacuum pressure, following which dried samples (30 mg) were extracted sequentially for both UHPLC-MS and GC-MS. The dried samples were first treated with 1.0 mL of 80% MeOH containing 18 µg/mL umbelliferone, sonicated for 5 minutes and centrifuged for 40 minutes at 3000 g at 10 ºC. 0.5 mL of supernatant was used for UHPLC-MS analysis after a subsequent spin at 5000 g at 10 ºC for 20 minutes and transferring 250 µL of the sample into glass autosampler vials with inserts. For GC-MS (Gas Chromatography-Mass Spectrometry) analyses of primary polar metabolites, 0.5 mL water was added the remaining extract used above for the UHPLC preparation, sonicated for 5 min, extracted for 30min, and centrifuged at 3000 g. 0.5 mL of the polar extract was subsequently dried under nitrogen and derivatized using previously established protocols (84). Briefly, N-Methyl-N-(trimethylsilyl) trifluoroacetamide (MSTFA) with 1 % TMCS (2,2,2-Trifluoro-N-methyl-N-(trimethylsilyl)-acetamide, Chlorotrimethylsilane) was used to derivatize the polar metabolites, after treatment with methoxyamine-HCl-pyridine.