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MB Sample ID: SA247719
Local Sample ID: | L_SM2 |
Subject ID: | SU002562 |
Subject Type: | Bacteria |
Subject Species: | Veillonella parvula |
Taxonomy ID: | 29466 |
Select appropriate tab below to view additional metadata details:
Subject:
Subject ID: | SU002562 |
Subject Type: | Bacteria |
Subject Species: | Veillonella parvula |
Taxonomy ID: | 29466 |
Factors:
Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
---|---|---|---|---|
L_SM2 | SA247719 | FL031140 | Spent Media | Matrix |
L_SM2 | SA247719 | FL031140 | Experiment | Sample Type |
L_SM2 | SA247719 | FL031140 | Lactate | Tretment |
Collection:
Collection ID: | CO002555 |
Collection Summary: | The strain Veillonella parvula SKV38 xdh::cat*. Briefly, V. parvula was grown on SK agar (composition: yeast extract 10 gL-1, casitone 10 gL-1, NaCl 2 gL-1, K2HPO4 0.4 gL-1) supplemented with either DL-lactate (50 mM lactate), potassium nitrate (40 mM KNO3), or both, incubated under anaerobic conditions at 37ºC overnight and then inoculated into the respective liquid media in biological triplicates. Cells and supernatants were harvested at mid-exponential phase (OD600=0.3), after centrifugation at 10,000g for 5 min. Both cell pellets and resulting supernatants were stored at -80C until processing for metabolite extraction and metabolomic analysis. A titration of cell pools and media pools was conducted (10,20,40,60,80,100%) and used to filter the data based on the correlation of the abundance of detected features with |
Sample Type: | Bacterial cells |
Treatment:
Treatment ID: | TR002574 |
Treatment Summary: | The strain was first streaked on an agar plate with SK media (composition: yeast extract 10 gL-1, casitone 10 gL-1, NaCl 2 gL-1, K2HPO4 0.4 gL-1) supplemented with 50 mM lactate and 40 mM KNO3 (SKLN medium) and antibiotics if required. From this agar plate, a single colony was selected and inoculated in 5mL SKLN media and grown for 24 hours. Next, overnight cells from this inoculum were grown using a 1/50 inoculum of the overnight culture, on either SK, SK + 50 mM lactate (SKL), SK + 40 mM nitrate (SKN), and SKLN. |
Sample Preparation:
Sampleprep ID: | SP002568 |
Sampleprep Summary: | Bacterial metabolites were profiled using the HILIC-pos and C8-pos methods for mapping Veillonella metabolites present in human stool. Bacterial samples were prepared as follows: mid-exponential Veillonella cultures (OD600 = 0.3-0.4) were harvested by centrifugation at 20,000g at 4°C for 1 minute, supernatants (spent media) were aliquoted, while cell pellets were resuspended in ice-cold PBS. Cells were then spun twice at 20,000g for 1 minute and all PBS supernatant removed and discarded. Cell pellet weights were estimated and all the samples harvested were stored at -80°C until metabolite profiling was conducted. For cells profiled in the C8-pos, cell pellets were resuspended in 20 µL of H2O and metabolites extracted using 380 µL of isopropanol containing 1,2-didodecanoyl-sn-glycero-3-phosphocholine (Avanti Polar Lipids; Alabaster, AL), 10 ul of media was extracted using 190 µL of isopropanol containing internal standards. Extracts were incubated at room temperature in the dark before centrifugation (10 min, 9,000 x g, Room Temperature). For cells profiled in the HILIC-pos mode, cell pellets were resuspended in 20 µL of H2O and metabolites extracted using180 µL HILIC extraction solution with internal standards (valine-d8, Sigma-Aldrich; St. Louis, MO; and phenylalanine-d8, Cambridge Isotope Laboratories; Andover, MA), while 10 µL of media was extracted with 90 µL of extraction solution. Extracts were cleared by centrifugation (10 min, 9,000 x g, 4C) prior to analysis. |
Combined analysis:
Analysis ID | AN004037 | AN004038 |
---|---|---|
Analysis type | MS | MS |
Chromatography type | HILIC | Reversed phase |
Chromatography system | Shimadzu Nexera X2 | Shimadzu Nexera X2 |
Column | Waters Atlantis HILIC (150 x 2 mm, 3 μm) | Waters Acquity BEH C8 (100 x 2.1mm, 1.7um) |
MS Type | ESI | ESI |
MS instrument type | Orbitrap | Orbitrap |
MS instrument name | Thermo Q Exactive Plus Orbitrap | Thermo Q Exactive Plus Orbitrap |
Ion Mode | POSITIVE | POSITIVE |
Units | Abundance | Abundance |
Chromatography:
Chromatography ID: | CH002985 |
Instrument Name: | Shimadzu Nexera X2 |
Column Name: | Waters Atlantis HILIC (150 x 2 mm, 3 μm) |
Column Temperature: | 30C |
Flow Gradient: | Isocratically with 5% mobile phase A for 1 minute followed by a linear gradient to 40% mobile phase B over 10 minutes |
Flow Rate: | 250 µL/min |
Solvent A: | 100% water; 10 mM ammonium formate; 0.1% formic acid |
Solvent B: | 100% acetonitrile; 0.1% formic acid |
Chromatography Type: | HILIC |
Chromatography ID: | CH002986 |
Instrument Name: | Shimadzu Nexera X2 |
Column Name: | Waters Acquity BEH C8 (100 x 2.1mm, 1.7um) |
Column Temperature: | 40C |
Flow Gradient: | The column was eluted at a flow rate of 450 µL/min isocratically for 1 minute at 80% mobile phase A, followed by a linear gradient to 80% mobile-phase B over 2 minutes, a linear gradient to 100% mobile phase B over 7 minutes, and then 3 minutes at 100% mobile-phase B. |
Flow Rate: | 450 µL/min |
Solvent A: | 95% water/5% methanol; 10 mM ammonium acetate; 0.1% acetic acid |
Solvent B: | 100% methanol; 0.1% acetic acid |
Chromatography Type: | Reversed phase |
MS:
MS ID: | MS003784 |
Analysis ID: | AN004037 |
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: | MS003785 |
Analysis ID: | AN004038 |
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 |