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MB Sample ID: SA274729
| Local Sample ID: | 1 |
| Subject ID: | SU002835 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU002835 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| 1 | SA274729 | FL035576 | db/+ | Animal |
| 1 | SA274729 | FL035576 | vehicle | Treatment |
Collection:
| Collection ID: | CO002828 |
| Collection Summary: | All mice were housed in a pathogen-free facility and maintained on a 12-h light and 12-h dark cycle with free access to normal chow diet and water. CANA was mixed at 0.03% (w/w) with CRF-1 and administered to 8-week-old db/+ and db/db mice for 4 weeks. Food intake was calculated for 3 consecutive days each week using the mouse feeder MF-4S. At the end of the experiments, 6 h-fasted mice were anesthetized with isoflurane and blood samples were obtained from the inferior vena cava. Skeletal muscles including soleus and extensor digitorum longus were immediately frozen and stored at -80°C for metabolome and lipidome analysis. |
| Sample Type: | Skeletal muscles |
Treatment:
| Treatment ID: | TR002844 |
| Treatment Summary: | All mice were free access to normal chow diet and water. CANA was mixed at 0.03% (w/w) with CRF-1 and administered to 8-week-old db/+ and db/db mice for 4 weeks. |
Sample Preparation:
| Sampleprep ID: | SP002841 |
| Sampleprep Summary: | Metabolites were extracted from frozen and crushed skeletal muscle (~10 mg) or serum (20 uL) with 1 mL of solvent mixture (methanol:chloroform:water = 10:4:4, v/v/v) containing piperazine-1,4-bis (2-ethanesulfonic acid) (PIPES) (0.63 umol/L), 2-bromohypoxanthine (0.13 umol/L), free fatty acid (FA) 16:0 (13C16) (0.13 umol/L), diacylglycerol (DG) 15:0-18:1 (d7) (0.15 umol/L), and triacylglycerol (TG) 15:0-18:1 (d7) -15:0 (0.35 umol/L) as internal standards. The samples were vigorously mixed for 1 min and sonicated for 5 min. The extracts were then centrifuged at 16,000 × g for 5 min at 4°C, and the resultant supernatant was collected. Protein concentrations in the pellets were determined using a Pierce BCA Protein Assay Kit (Thermo Fisher Scientific, Massachusetts, USA). The collected supernatant (700 uL) was mixed with 235 uL of chloroform, and 155 uL of water, and then centrifuged at 16,000 × g for 5 min at 4°C. The aqueous (upper) layer (400 µL) was evaporated under vacuum, dried extracts were stored at −80°C until targeted polar metablome analysis. Prior to analysis, the dried aqueous layer was reconstituted in 50 uL of water. The organic (lower) layer (200 µL) was dried under a nitrogen stream, dried extracts were stored at −80°C until targeted lipidome analysis. Prior to analysis, the organic aqueous layer was reconstituted in 80 µL of methanol/chloroform (1/1, v/v) |
Combined analysis:
| Analysis ID | AN004423 | AN004424 | AN004425 | AN004426 |
|---|---|---|---|---|
| Analysis type | MS | MS | MS | MS |
| Chromatography type | Ion exchange | Reversed phase | Reversed phase | Normal phase |
| Chromatography system | Thermo Dionex ICS-5000+ | Shimadzu Nexera X2 | Shimadzu Nexera X2 | Shimadzu Nexera UC |
| Column | Dionex IonPac AS11-HC (250 x 2mm,4um) | Merck Discovery HS F5 (PFP) (150 x 2.1mm,3um) | GL Sciences Inert Sustain C18 (150 × 2.1mm, 3um) | Waters ACQUITY UPC2 Torus DEA (100 x 3.0mm, 1.7um) |
| MS Type | ESI | ESI | ESI | ESI |
| MS instrument type | Orbitrap | Orbitrap | Orbitrap | Triple quadrupole |
| MS instrument name | Thermo Q Exactive Orbitrap | Thermo Q Exactive Orbitrap | Thermo Q Exactive Orbitrap | Shimadzu LCMS-8060 |
| Ion Mode | UNSPECIFIED | UNSPECIFIED | POSITIVE | UNSPECIFIED |
| Units | relative peak area | relative peak area | concentration (pmol/mg-protein) | relative peak area |
Chromatography:
| Chromatography ID: | CH003322 |
| Chromatography Summary: | Anionic polar metabolites (i.e., organic acids, sugar phosphates, nucleotides, etc.) were analyzed via IC/HRMS/MS. |
| Instrument Name: | Thermo Dionex ICS-5000+ |
| Column Name: | Dionex IonPac AS11-HC (250 x 2mm,4um) |
| Column Temperature: | 30℃ |
| Flow Gradient: | 10-100%A, 0-24.0 min, 100%A, 24.0-27.0 min, 100-10%A, 27.0-27.1 min, 10%A, 27.1-35.0 min |
| Flow Rate: | 0.3 mL/min |
| Solvent A: | 100 mM KOH in H2O |
| Solvent B: | 1 mM ammonium acetate in MeOH |
| Chromatography Type: | Ion exchange |
| Chromatography ID: | CH003323 |
| Chromatography Summary: | Cationic polar metabolites (i.e., amino acids, bases, nucleosides, etc) were analyzed via PFPP-LC/HRMS/MS. |
| Instrument Name: | Shimadzu Nexera X2 |
| Column Name: | Merck Discovery HS F5 (PFP) (150 x 2.1mm,3um) |
| Column Temperature: | 40℃ |
| Flow Gradient: | 0%B, 0-5.0 min, 0-40%B, 5.0-15.0 min, 40-100%B, 15.0-15.1 min, 100%B, 15.1-18.0 min, 100-0%B, 18.0-18.1 min, 0%B, 18.1-25.0 min |
| Flow Rate: | 0.25 mL/min |
| Solvent A: | 0.1% formic acid in H2O |
| Solvent B: | ACN |
| Chromatography Type: | Reversed phase |
| Chromatography ID: | CH003324 |
| Chromatography Summary: | Acyl-CoA and acyl-carnitine were analyzed via C18-LC/HRMS/MS. |
| Instrument Name: | Shimadzu Nexera X2 |
| Column Name: | GL Sciences Inert Sustain C18 (150 × 2.1mm, 3um) |
| Column Temperature: | 40℃ |
| Flow Gradient: | 0-13.0 min, 2-95%B, 13.0-20.0 min, 95%B, 20.0-20.1 min, 95-2%B, 20.1-25.0 min, 2%B |
| Flow Rate: | 0.3 mL/min |
| Solvent A: | 5 mM ammonium acetate in H2O |
| Solvent B: | ACN |
| Chromatography Type: | Reversed phase |
| Chromatography ID: | CH003325 |
| Chromatography Summary: | Fatty acids (FA), diacylglycerols (DG), and triacylglycerols (TG) were analyzed via Normal phase SFC/MS/MS. |
| Instrument Name: | Shimadzu Nexera UC |
| Column Name: | Waters ACQUITY UPC2 Torus DEA (100 x 3.0mm, 1.7um) |
| Column Temperature: | 50℃ |
| Flow Gradient: | 0-1.0 min, 1%B, 1.0-24.0 min, 1-75%B, 24.0-26.0 min, 75%B, 26.0-26.1 min, 75-1%B, 26.1-30.0 min, 1%B |
| Flow Rate: | 1.0 mL/min |
| Solvent A: | CO2 |
| Solvent B: | 0.1% (w/v) ammonium acetate in MeOH/H2O (95/5) |
| Chromatography Type: | Normal phase |
| Solvent C: | 0.1% (w/v) ammonium acetate in MeOH/H2O (95/5) |
MS:
| MS ID: | MS004170 |
| Analysis ID: | AN004423 |
| Instrument Name: | Thermo Q Exactive Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | MS acquisition: FullMS (posi/nega) Data processing/software: Reifycs Cascade ver. 1.0 |
| Ion Mode: | UNSPECIFIED |
| MS ID: | MS004171 |
| Analysis ID: | AN004424 |
| Instrument Name: | Thermo Q Exactive Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | MS acquisition: FullMS (posi/nega) Data processing/software: Reifycs Cascade ver. 1.0 |
| Ion Mode: | UNSPECIFIED |
| MS ID: | MS004172 |
| Analysis ID: | AN004425 |
| Instrument Name: | Thermo Q Exactive Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | MS acquisition: FullMS (posi) Data processing/software: Reifycs Cascade ver. 1.0 |
| Ion Mode: | POSITIVE |
| MS ID: | MS004173 |
| Analysis ID: | AN004426 |
| Instrument Name: | Shimadzu LCMS-8060 |
| Instrument Type: | Triple quadrupole |
| MS Type: | ESI |
| MS Comments: | MS acquisition: MRM mode (posi/nega) Data processing/software: Multi-ChromatoAnalyst (mCAT) ver.23 |
| Ion Mode: | UNSPECIFIED |
