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MB Sample ID: SA172831
Local Sample ID: | CE086-2 |
Subject ID: | SU001928 |
Subject Type: | Human |
Subject Species: | Homo sapiens |
Taxonomy ID: | 9606 |
Select appropriate tab below to view additional metadata details:
Subject:
Subject ID: | SU001928 |
Subject Type: | Human |
Subject Species: | Homo sapiens |
Taxonomy ID: | 9606 |
Factors:
Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
---|---|---|---|---|
CE086-2 | SA172831 | FL020831 | Centenarian | Category |
Collection:
Collection ID: | CO001921 |
Collection Summary: | Stool samples were collected by the individuals using sterile faeces containers and transported to the lab in cooling-box with ice and Anaero pack within 24 hours of sample collection. Immediately after arriving, samples were aliquoted and stored at -80°C until further analysis. |
Sample Type: | Feces |
Storage Conditions: | -80℃ |
Treatment:
Treatment ID: | TR001940 |
Treatment Summary: | Frozen feces were crushed with a metal bead using a Multi-Beads Shocker (Yasui Kikai). |
Sample Preparation:
Sampleprep ID: | SP001934 |
Sampleprep Summary: | Accurately weighed freeze-crushed faecal samples were resuspended in 20 times the volume of water (e.g. 12.5 mg of faece in 250 μL water). 250 μL of faecal suspension was homogenized in 747 μL of 0.27 N NaOH by ultrasonication for 1 hr in a screw-cap glass vial containing 3 μL of deuterium-labelled internal standards (d4-CA, d4-GCDCA, d4-TCDCA, d4-CDCA-3S, and d4-LCA, 50 μM each). After incubation for 1 hr at room temperature, pH was adjusted to 8.0 using 8 N HCl, mixed with 110 μL of 0.5 M EDTA/0.5 M TrisHCl. The solution was centrifuged at 15,000 rpm and the supernatant was transferred onto a solid-phase extraction cartridge (Agilent Bond Elut C18, 100 mg/3 mL, preconditioned with 1 mL of methanol and 3 mL of water, three times). The cartridge was washed with 1 mL of water and captured bile acids were eluted with 300 μL of 90% ethanol. |
Combined analysis:
Analysis ID | AN002999 |
---|---|
Analysis type | MS |
Chromatography type | Reversed phase |
Chromatography system | Exion LC |
Column | InertSustain C18 |
MS Type | ESI |
MS instrument type | Triple quadrupole |
MS instrument name | ABI Sciex Triple Quad 6500+ |
Ion Mode | UNSPECIFIED |
Units | nM |
Chromatography:
Chromatography ID: | CH002224 |
Chromatography Summary: | 2 μL was injected to LC/ESI-MS/MS (Triple Quad 6500+ tandem mass spectrometer, equipped with an ESI probe and Exion LC AD ultra-high pressure liquid chromatography system; SCIEX). A separation column, InertSustain C18 (150 mm × 2.1 mm ID, 3 μm particle size; GL Sciences Inc.), was utilized at 40 °C. A mixture of 10 mM ammonium acetate and acetonitrile was used as the eluent and the separation was carried out by linear gradient elution at a flow rate of 0.2 mL/min. The mobile phase composition was gradually changed as follows: ammonium acetate-acetonitrile (86:14, v/v) for 0.5 min, (78:22, v/v) for 0.5-5 min, (72:28, v/v) for 5-28 min, (46:54, v/v) for 28-55 min, (2:98, v/v) for 55-66 min, and (2:98, v/v) for 4 min. The total run time was 70 min. |
Instrument Name: | Exion LC |
Column Name: | InertSustain C18 |
Column Temperature: | 40 |
Flow Gradient: | ammonium acetate-acetonitrile (86:14, v/v) for 0.5 min, (78:22, v/v) for 0.5-5 min, (72:28, v/v) for 5-28 min, (46:54, v/v) for 28-55 min, (2:98, v/v) for 55-66 min, and (2:98, v/v) for 4 min. The total run time was 70 min. |
Flow Rate: | 0.2 mL/min |
Solvent A: | 100% water; 10mM ammonium acetate |
Solvent B: | 100% acetonitrile |
Chromatography Type: | Reversed phase |
MS:
MS ID: | MS002788 |
Analysis ID: | AN002999 |
Instrument Name: | ABI Sciex Triple Quad 6500+ |
Instrument Type: | Triple quadrupole |
MS Type: | ESI |
MS Comments: | The following MS parameters were used for the positive ion MRM mode: ion spray voltage; 5,500 V, interface temperature; 500 °C, curtain gas; 30 psi, collision gas (nitrogen); 9 psi, nebulizing gas; 80 psi, and heater gas; 80 psi. For the negative ion MRM mode: ion spray voltage; -4,500 V, interface temperature; 500 °C, curtain gas; 30 psi, collision gas (nitrogen); 9 psi, nebulizing gas; 80 psi, and heater gas; 80 psi. Samples were obtained using Analyst software ver1.71 and analysed using SCIEX OS-MQ software ver1.7.0.36606. |
Ion Mode: | UNSPECIFIED |