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MB Sample ID: SA307036

Local Sample ID:Neg_BK3
Subject ID:SU002943
Subject Type:Bacteria
Subject Species:Bacteroides thetaiotaomicron
Taxonomy ID:8188
Subject Comments:Fecal derived communities and isolates, supernatant was assayed

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Combined analysis:

Analysis ID AN004629 AN004630
Analysis type MS MS
Chromatography type Reversed phase Reversed phase
Chromatography system Thermo Vanquish Thermo Vanquish
Column Agilent Zorbax SB-C18 (100 x 3.0 mm, 1.8 um) Agilent Zorbax SB-C18 (100 x 3.0 mm, 1.8 um)
MS Type ESI ESI
MS instrument type Orbitrap Orbitrap
MS instrument name Thermo Q Exactive HF hybrid Orbitrap Thermo Q Exactive HF hybrid Orbitrap
Ion Mode POSITIVE NEGATIVE
Units counts, height counts, height

Chromatography:

Chromatography ID:CH003483
Chromatography Summary:Bacterial supernatant were analyzed via reversed phase (C18) coupled to a Thermo Q-Exactive HF high resolution mass spectrometer. Prepared samples were injected onto an Agilent Zorbax SB-C18 column (100 mm length × 3 mm id; 1.8 μm particle size) with an additional Phenomenex KrudKatcher pre-column (2 μm depth x 0.004in ID) maintained at 40°C coupled to an Thermo Vanquish UPLC. The mobile phases were prepared with 0.1% formic acid in either 100% LC-MS grade water for mobile phase (A), 100% water or mobile phase (B), 100% acetonitrile. Gradient elution was performed as follows 3% (B) maintained 0–0.43 min to 97% (B) at 9 min, maintained until 11 min, returning to initial conditions at 11.5 min and equilibrating until the end of the run at 14 min. Flow rate is maintained at 0.4 mL/min. Each sample was analyzed in both positive and negative ionization modes (ESI+, ESI-) via subsequent injections. Full MS-ddMS2 data was collected, an inclusion list was used to prioritize MS2 selection of metabolites from our in-house ‘local’ library, when additional scan bandwidth was available MS2 was collected in a data-dependent manner. Mass range was 60-900 mz, resolution was 60k (MS1) and 15k (MS2), centroid data was collected, loop count was 4, isolation window was 1.5 Da. Metabolomics data was processed using MS-DIAL v4.60 (https://www.nature.com/articles/s41587-020-0531-2) and queried against a combination of our in-house MS2 library and MassBank of North America, the largest freely available spectral repository (https://doi.org/10.1002/mas.21535). Features were excluded from analysis if peak height was not at least 5-fold greater in one or more samples compared to the procedural blank average.
Instrument Name:Thermo Vanquish
Column Name:Agilent Zorbax SB-C18 (100 x 3.0 mm, 1.8 um)
Column Temperature:40C
Flow Gradient:Gradient elution was performed from 100% (B) at 0–2 min to 70% (B) at 7.7 min, 40% (B) at 9.5 min, 30% (B) at 10.25 min, 100% (B) at 12.75 min, isocratic until 16.75 min with a column flow ofGradient elution was performed as follows 3% (B) maintained 0–0.43 min to 97% (B) at 9 min, maintained until 11 min, returning to initial conditions at 11.5 min and equilibrating until the end of the run at 14 min.
Flow Rate:0.4 mL/min.
Solvent A:Water + 0.1% formic acid
Solvent B:Acetonitrile + 0.1% formic acid
Chromatography Type:Reversed phase
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