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MB Sample ID: SA060920
| Local Sample ID: | GLA_QEx_Lipids_SO059_SA060_1003bG2 |
| Subject ID: | SU001027 |
| Subject Type: | Human |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU001027 |
| Subject Type: | Human |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| GLA_QEx_Lipids_SO059_SA060_1003bG2 | SA060920 | FL010628 | GF | Phenotype |
| GLA_QEx_Lipids_SO059_SA060_1003bG2 | SA060920 | FL010628 | 2-h | Collection Time (hours) |
Collection:
| Collection ID: | CO001021 |
| Collection Summary: | We deliberately used samples from a recently published study in order to investigate if statistical and biological conclusions would differ from the published results, depending on the instrumentation used. In a blinded, placebo-controlled, crossover designed study, seven healthy subjects consumed a test meal containing high amounts of gamma-linolenic acid (GLA, 18:3n6) compared to a control meal. Each subject underwent the nutritional test on three separate test days for each test meal, and samples were taken each time over an 8 h period. |
| Collection Protocol Filename: | acs_analchem_7b03404.pdf |
| Sample Type: | Blood (plasma) |
| Collection Frequency: | 0, 2, and 4 hours |
Treatment:
| Treatment ID: | TR001041 |
| Treatment Summary: | For this study, we used a subset of samples from our recent study focused on nutritional phenotyping in response to a test meal containing gamma-linolenic acid. Briefly, in a single blind, placebo-controlled, crossover design, seven healthy subjects consumed a test meal that consisted of GLA fat (borage oil [denoted GF]) or a control fat (a mixture of corn, safflower, sunflower, and extra-virgin light olive oils [denoted CF]). Compared to the original study, where all subjects were fed on three separate test days for each test meal, a small modification was needed due to sample limitation. Thus, for this study, six subjects were fed on three separate test days for each test meal, while one subject was fed on two separate test days for a control fat meal and four test days for GLA fat (the fourth set was not used in the original study). Plasma samples collected at 0, 2, and 4h in response to the test meals were used for analysis. In total, 126 samples were analyzed out of which 42 were baseline samples (time 0 h), 40 were control fat samples (time 2 and 4 h), and 44 were GLA fat samples (time 2 and 4 h). For quality control, a pool sample consisted of a mixture of nonfasting blood plasma (both control and GLA fat) was used. Also, standard reference material SRM 1950 Metabolites in Frozen Human Plasma (NIST, Gaithersburg, MD) was used. |
| Treatment Protocol Filename: | acs_analchem_7b03404.pdf |
Sample Preparation:
| Sampleprep ID: | SP001034 |
| Sampleprep Summary: | Extraction of plasma lipids was carried out using a biphasic solvent system of cold methanol, methyl tertbutyl ether (MTBE), and water with some modifications. In more detail, 300 μL of cold methanol containing a mixture of odd chain and deuterated lipid internal standards [LPE(17:1), LPC(17:0), PC(12:0/13:0), PE(17:0/17:0), PG(17:0/17:0), d7-cholesterol, SM(d18:1/17:0), Cer(d18:1/17:0), sphingosine (d17:1), DG(12:0/12:0/0:0), DG(18:1/2:0/0:0), and d5-TG(17:0/17:1/17:0)] was added to a 40 μL blood plasma aliquot in a 2 mL Eppendorf tube and then vortexed (10 s). Then, 1000 μL of cold MTBE containing CE 22:1 (internal standard) was added, followed by vortexing (10 s) and shaking (6 min) at 4 °C. Phase separation was induced by adding 250 μL of LC−MS grade water followed by centrifugation at 14000 rpm for 2 min. |
| Sampleprep Protocol Filename: | acs_analchem_7b03404.pdf |
Combined analysis:
| Analysis ID | AN001614 |
|---|---|
| Analysis type | MS |
| Chromatography type | Reversed phase |
| Chromatography system | Thermo Q-Exactive Quadrupole Orbitrap |
| Column | Waters Acquity CSH C18 (100 x 2.1mm,1.7um) |
| MS Type | ESI |
| MS instrument type | QTOF |
| MS instrument name | Thermo Q Exactive HF hybrid Orbitrap |
| Ion Mode | POSITIVE |
| Units | nanograms (absolute) |
Chromatography:
| Chromatography ID: | CH001136 |
| Methods Filename: | Data_Dictionary_Fiehn_laboratory_CSH_QTOF_lipidomics_05-29-2014.pdf |
| Instrument Name: | Thermo Q-Exactive Quadrupole Orbitrap |
| Column Name: | Waters Acquity CSH C18 (100 x 2.1mm,1.7um) |
| Column Pressure: | 450-850 bar |
| Column Temperature: | 65 C |
| Flow Gradient: | 15% B to 99% B |
| Flow Rate: | 0.6 mL/min |
| Injection Temperature: | 4 C |
| Internal Standard: | See data dictionary |
| Retention Time: | See data dictionary |
| Sample Injection: | 1.67 uL |
| Solvent A: | 60% acetonitrile/40% water; 10mM formic acid; 10mM ammonium formate |
| Solvent B: | 90% isopropanol/10% acetonitrile; 10mM formic acid; 10mM ammonium formate |
| Analytical Time: | 13 min |
| Capillary Voltage: | 3500 eV |
| Oven Temperature: | 50°C for 1 min, then ramped at 20°C/min to 330°C, held constant for 5 min |
| Time Program: | 15 min |
| Washing Buffer: | Ethyl Acetate |
| Weak Wash Solvent Name: | Isopropanol |
| Strong Wash Solvent Name: | Isopropanol |
| Target Sample Temperature: | Autosampler temp 4 C |
| Sample Loop Size: | 30 m length x 0.25 mm internal diameter |
| Randomization Order: | Excel generated |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS001492 |
| Analysis ID: | AN001614 |
| Instrument Name: | Thermo Q Exactive HF hybrid Orbitrap |
| Instrument Type: | QTOF |
| MS Type: | ESI |
| Ion Mode: | POSITIVE |
| Capillary Voltage: | 3500 eV |
| Collision Energy: | 25 eV |
| Collision Gas: | Nitrogen |
| Dry Gas Flow: | 8L/min |
| Dry Gas Temp: | 325 C |
| Fragment Voltage: | 120 eV |
| Fragmentation Method: | Auto MS/MS |
| Ion Source Temperature: | 325 C |
| Ion Spray Voltage: | 1000 |
| Ionization: | Pos |
| Ionization Energy: | 70eV |
| Mass Accuracy: | Accurate |
| Reagent Gas: | Nitrogen |
| Source Temperature: | 325 C |
| Dataformat: | .d |
| Desolvation Gas Flow: | 11 L/min |
| Desolvation Temperature: | 350 C |
| Nebulizer: | 35 psig |
| Octpole Voltage: | 750 eV |
| Resolution Setting: | Extended Dyamic Range |
| Scan Range Moverz: | 60-1700 Da |
| Scanning Cycle: | 2 Hz |
| Scanning Range: | 60-1700 Da |
| Skimmer Voltage: | 65 |
