GNPS - American Gut Phase 1 - Metabolomics
Ontology highlight
ABSTRACT:
INSTRUMENT(S): maXis impact HD
ORGANISM(S): Homo Sapiens (ncbitaxon:9606)
SUBMITTER: 
Alan Jarmusch Pieter Dorrestein Rob Knight
PROVIDER: MSV000081981 | GNPS | Fri Jan 26 13:59:00 GMT 2018
REPOSITORIES: GNPS
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Project description:Sample Collection
Samples used for this study were obtained as part of the HoloFish project (Norwegian Seafood Research Fund, project no. 901436). This cohort has been described previously 19. Briefly, we sampled 460 ready-to-harvest Atlantic salmon from a commercial production site close to Bergen, Norway, owned by Leroy Seafood Group in April 2018. Samples were obtained from two groups reared in separate sea pens and fed two different standard commercial diets. These diets have been anonymised but were manufactured respectively by BioMar and EWOS in 2018.
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Extraction
Gut content samples were cryo-homogenised in 25% water, 25% methanol and 50% dichloromethane in a 1:15 sample: solvent ratio (w:v). Homogenisation was carried out using an OMNI Bead Ruptor 24, using liquid nitrogen to keep homogenised samples below 0 degrees Celsius to minimise degradation of metabolites during extraction. Homogenates were centrifuged at 20,000 g (0 degrees Celsius) and the polar phase from all samples was concentrated using SpeedVac (ThermoFisher Scientific) and resuspended in 200 microL 5% methanol. Four procedural blanks were included in homogenisation. A volume of 100 microL of all samples was collected into a Quality Control sample used for normalisation to enhance the detection of metabolites.
Chromatography
Samples were measured on a nano-flow ultra-high pressure liquid chromatography tandem high-resolution mass spectrometry analysis.
Mass spectrometry
Metabolites were detected using a Q Exactive HF Hybrid Quadrupole-Orbitrap Mass Spectrometer (ThermoFisher Scientific) operated in positive ion data-dependent acquisition mode.
Data Transformation
ThermoFisher Scientific UHPLC-Orbitrap-MS/MS RAW files were converted into mzML files using Proteo Wizard. For an increased deciphering of molecular spectres, MZmine2 was applied for mass detection of MS1 and MS2 spectres, followed by chromatogram detection and deconvolution. Subsequently, detected isotopes and features were grouped according to a tolerance of mass-charges (5 ppm for m/z) and retention time (6 sec.) and the features were further aligned according to retention time and m/z. Lastly, only features with an MS2 spectrum were kept for further substructural analysis and in silico analysis.
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