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MB Sample ID: SA151718

Local Sample ID:KP_JS_C08.WT5_017
Subject ID:SU001727
Subject Type:Mammal
Subject Species:Mus musculus
Taxonomy ID:10090
Genotype Strain:Swiss Webster

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Subject:

Subject ID:SU001727
Subject Type:Mammal
Subject Species:Mus musculus
Taxonomy ID:10090
Genotype Strain:Swiss Webster

Factors:

Local Sample IDMB Sample IDFactor Level IDLevel ValueFactor Name
KP_JS_C08.WT5_017SA151718FL017808WTColonization

Collection:

Collection ID:CO001720
Collection Summary:Cecal contents from gnotobiotic mice were collected under sterile conditions, flash-frozen in liquid nitrogen and stored at -80 C.
Sample Type:Feces
Storage Vials:-80

Treatment:

Treatment ID:TR001740
Treatment Summary:Swiss Webster germ-free mice were tri-colonized with Bacteroides thetaiotaomicron, Clostridium sporogenes and Escherichia coli for 10 days. Mice were subsequently infected with wild-type Clostridium difficile R20291 (n=5), the isogenic toxin-deficient mutant TcdA-B-CDT- (n=5), or gavaged with PBS as an uninfected control (n=4). Cecal contents were colleged 5 days after C. difficile infection.

Sample Preparation:

Sampleprep ID:SP001733
Sampleprep Summary:Metabolites were extracted according to the methodology described in Fiehn 2017 Curr Protoc Mol Biol. ; 114: 30.4.1–30.4.32. doi:10.1002/0471142727.mb3004s114. Briefly, 1 mL 3:3:2 acetonitrile:isopropanol:water was added to ~4 mg cecal contents/feces. Samples were homogenized and then shaken at 4 C for 6 min prior to centrifugation at 14,000 x g for 2 minutes. An aliquot of 475 µl supernatant was taken for subsequent analysis, evaporated, cleaned with acetonitrile and water (1:1), and fully evaporated to dryness. 10 µl MeOH and 91 µl MSTFA were added to all samples for derivatization.

Combined analysis:

Analysis ID AN002696
Analysis type MS
Chromatography type GC
Chromatography system Agilent 6890 GC
Column Restek Rtx-5Sil (30m x 0.25mm,0.25um)
MS Type EI
MS instrument type GC-TOF
MS instrument name Leco Pegasus IV TOF
Ion Mode UNSPECIFIED
Units preak area

Chromatography:

Chromatography ID:CH001989
Chromatography Summary:Agilent 6890 GC is equipped with a Gerstel automatic liner exchange system (ALEX) that includes a multipurpose sample (MPS2) dual rail, and a Gerstel CIS cold injection system (Gerstel, Muehlheim, Germany) with temperature program as follows: 50°C to 275°C final temperature at a rate of 12 °C/s and hold for 3 minutes. Injection volume is 0.5 μl with 10 μl/s injection speed on a splitless injector with purge time of 25 seconds. Liner (Gerstel #011711-010-00) is changed after every 10 samples, (using the Maestro1 Gerstel software vs. 1.1.4.18). Before and after each injection, the 10 μl injection syringe is washed three times with 10 μl ethyl acetate A 30 m long, 0.25 mm i.d. Rtx-5Sil MS column (0.25 μm 95% dimethyl 5% diphenyl polysiloxane film) with additional 10 m integrated guard column is used (Restek, Bellefonte PA). 99.9999% pure Helium with built-in purifier (Airgas, Radnor PA) is set at constant flow of 1 ml/min. The oven temperature is held constant at 50°C for 1 min and then ramped at 20°C/min to 330°C at which it is held constant for 5 min.
Instrument Name:Agilent 6890 GC
Column Name:Restek Rtx-5Sil (30m x 0.25mm,0.25um)
Flow Rate:1 ml/min
Chromatography Type:GC

MS:

MS ID:MS002494
Analysis ID:AN002696
Instrument Name:Leco Pegasus IV TOF
Instrument Type:GC-TOF
MS Type:EI
MS Comments:A Leco Pegasus IV time of flight mass spectrometer is controlled by the Leco ChromaTOF software vs. 2.32 (St. Joseph, MI). The transfer line temperature between gas chromatograph and mass spectrometer is set to 280°C. Electron impact ionization at 70V is employed with an ion source temperature of 250°C. Acquisition rate is 17 spectra/second, with a scan mass range of 85-500 Da.
Ion Mode:UNSPECIFIED
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