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MB Sample ID: SA217733
| Local Sample ID: | Plasma_QC_2 |
| Subject ID: | SU002355 |
| Subject Type: | Human |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
| Age Or Age Range: | 8.7-18.6 (year) |
| Gender: | Male and female |
| Human Ethnicity: | Korean |
| Human Inclusion Criteria: | We recruited children and adolescent participants who visited the pediatric departments of the Hallym University Sacred Heart Hospital and Seoul National University Children’s Hospital from January 2019 to May 2020, after obtaining informed consent from the children and their parents. |
| Human Exclusion Criteria: | We excluded participants who were taking alcohol or medications known to affect the results of liver function tests. Participants with viral hepatitis, such as hepatitis A, B, or C, or with Epstein–Barr virus, Wilson’s disease, autoimmune hepatitis, or muscular disease were also excluded. |
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Subject:
| Subject ID: | SU002355 |
| Subject Type: | Human |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
| Age Or Age Range: | 8.7-18.6 (year) |
| Gender: | Male and female |
| Human Ethnicity: | Korean |
| Human Inclusion Criteria: | We recruited children and adolescent participants who visited the pediatric departments of the Hallym University Sacred Heart Hospital and Seoul National University Children’s Hospital from January 2019 to May 2020, after obtaining informed consent from the children and their parents. |
| Human Exclusion Criteria: | We excluded participants who were taking alcohol or medications known to affect the results of liver function tests. Participants with viral hepatitis, such as hepatitis A, B, or C, or with Epstein–Barr virus, Wilson’s disease, autoimmune hepatitis, or muscular disease were also excluded. |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| Plasma_QC_2 | SA217733 | FL026427 | QC | Group |
Collection:
| Collection ID: | CO002348 |
| Collection Summary: | For metabolomic analysis, 4 mL of blood was collected from each participant after overnight fasting and centrifuged at 4℃. Separated plasma samples were collected and stored until use at -80℃. |
| Sample Type: | Blood (plasma) |
| Storage Conditions: | -80℃ |
Treatment:
| Treatment ID: | TR002367 |
| Treatment Summary: | We recruited children and adolescent participants who visited the pediatric departments of the Hallym University Sacred Heart Hospital and Seoul National University Children’s Hospital from January 2019 to May 2020, and evaluated the presence and grade of fatty liver by ultrasonography. The grade of steatosis was evaluated as follows by comparing hepatic echogenicity to kidney parenchyma: normal, 0; mild, 1; moderate, 2; and severe, 3 14,15. The participants were categorized into four groups according to the steatosis grade determined by abdominal ultrasonography and body mass index (BMI) z-score based on the 2017 Korean National Growth Chart for children and adolescents 16: healthy control (HC), steatosis grade = 0 and BMI z-score ≤ 1; lean NAFLD (LN), steatosis grade ≥ 1 and BMI z-score ≤ 1; overweight control (OC), steatosis grade = 0 and BMI z-score > 1; and overweight NAFLD (ON), steatosis grade ≥ 1 and BMI z-score > 1. |
| Treatment: | Observational |
Sample Preparation:
| Sampleprep ID: | SP002361 |
| Sampleprep Summary: | frozen plasma samples were thawed on ice and vortexed, followed by centrifugation at 2,750 × g, 4℃ for 5 min before the samples were loaded onto a 96-well plate with a filter. After the addition of 10 μL of analytical and pooled plasma samples and calibration standards to each well, the plates were dried with a nitrogen evaporator and derivatized with phenyl isothiocyanate. Then, dried samples were extracted with ammonium acetate solution in methanol and aliquoted into two deep-well plates for liquid chromatography mode and flow injection analysis (FIA) mode (described in the manual), followed by dilution with water and an FIA solvent, respectively. Both deep-well plates were placed in an autosampler of Ultimate 3000 ultra-performance liquid chromatography coupled with a Q Exactive Plus hybrid quadrupole-orbitrap mass spectrometer (Thermo Fisher Scientific, MA, USA) and analyzed using the validated method. |
| Sampleprep Protocol Filename: | UM-p400-HR-Thermo-3.pdf |
| Processing Storage Conditions: | On ice |
| Extract Storage: | 4℃ |
Combined analysis:
| Analysis ID | AN003706 | AN003707 | AN003708 | AN003709 |
|---|---|---|---|---|
| Analysis type | MS | MS | MS | MS |
| Chromatography type | Reversed phase | Reversed phase | None (Direct infusion) | None (Direct infusion) |
| Chromatography system | Thermo Dionex Ultimate 3000 | Thermo Dionex Ultimate 3000 | Thermo Dionex Ultimate 3000 | Thermo Dionex Ultimate 3000 |
| Column | Biocrates AbsoluteIDQ p400 HR Kit UHPLC (PN 9120052121032) | Biocrates AbsoluteIDQ p400 HR Kit UHPLC (PN 9120052121032) | NA | NA |
| MS Type | ESI | ESI | ESI | ESI |
| MS instrument type | Orbitrap | Orbitrap | Orbitrap | Orbitrap |
| MS instrument name | Thermo Q Exactive Plus Orbitrap | Thermo Q Exactive Plus Orbitrap | Thermo Q Exactive Plus Orbitrap | Thermo Q Exactive Plus Orbitrap |
| Ion Mode | POSITIVE | POSITIVE | POSITIVE | POSITIVE |
| Units | μM | μM | μM | μM |
Chromatography:
| Chromatography ID: | CH002745 |
| Chromatography Summary: | LC1 method for analysis of amino acids and biogenic amines For more details, please refer sections 4.6, 4.7, 5, 9, 10.1, and 10.2 of the method file attached. |
| Methods Filename: | UM-p400-HR-Thermo-3.pdf |
| Instrument Name: | Thermo Dionex Ultimate 3000 |
| Column Name: | Biocrates AbsoluteIDQ p400 HR Kit UHPLC (PN 9120052121032) |
| Column Temperature: | 50 |
| Solvent A: | 100% water; 0.2% formic acid |
| Solvent B: | 100% acetonitrile; 0.2% formic acid |
| Chromatography Type: | Reversed phase |
| Chromatography ID: | CH002746 |
| Chromatography Summary: | LC2 method for analysis of isoleucine For more details, please refer sections 4.6, 4.7, 5, 9, 10.1, and 10.2 of the method file attached. |
| Methods Filename: | UM-p400-HR-Thermo-3.pdf |
| Instrument Name: | Thermo Dionex Ultimate 3000 |
| Column Name: | Biocrates AbsoluteIDQ p400 HR Kit UHPLC (PN 9120052121032) |
| Column Temperature: | 50 |
| Solvent A: | 100% water; 0.2% formic acid |
| Solvent B: | 100% acetonitrile; 0.2% formic acid |
| Chromatography Type: | Reversed phase |
| Chromatography ID: | CH002747 |
| Chromatography Summary: | FIA1 method for analysis of acylcarnitines and lipids For more details, please refer sections 4.6, 4.7, 5, 9, 10.1, and 10.2 of the method file attached. |
| Methods Filename: | UM-p400-HR-Thermo-3.pdf |
| Instrument Name: | Thermo Dionex Ultimate 3000 |
| Column Name: | NA |
| Solvent A: | 290mL Methanol + 1 ampule FIA mobile phase additive |
| Chromatography Type: | None (Direct infusion) |
| Chromatography ID: | CH002748 |
| Chromatography Summary: | FIA2 method for analysis of acylcarnitines For more details, please refer sections 4.6, 4.7, 5, 9, 10.1, and 10.2 of the method file attached. |
| Methods Filename: | UM-p400-HR-Thermo-3.pdf |
| Instrument Name: | Thermo Dionex Ultimate 3000 |
| Column Name: | NA |
| Solvent A: | 290mL Methanol + 1 ampule FIA mobile phase additive |
| Chromatography Type: | None (Direct infusion) |
MS:
| MS ID: | MS003455 |
| Analysis ID: | AN003706 |
| Instrument Name: | Thermo Q Exactive Plus Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | MS acquisition method, data processing method and software used for feature assignments are described in analysis protocol file attached. Please refer sections 10.3 (MS acquisition parameters) and 9 (data processing) of the method file attached. The final quantitative results were exported to micromolar values with pooled quality control sample normalization by median. Subsequently, values under the lower limit of detection were imputed by one-fifth of the minimum positive values of their corresponding variables. These data were used for further analysis. |
| Ion Mode: | POSITIVE |
| Acquisition Parameters File: | UM-p400-HR-Thermo-3.pdf |
| Processing Parameters File: | UM-p400-HR-Thermo-3.pdf |
| Analysis Protocol File: | UM-p400-HR-Thermo-3.pdf |
| MS ID: | MS003456 |
| Analysis ID: | AN003707 |
| Instrument Name: | Thermo Q Exactive Plus Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | MS acquisition method, data processing method and software used for feature assignments are described in analysis protocol file attached. Please refer sections 10.3 (MS acquisition parameters) and 9 (data processing) of the method file attached. The final quantitative results were exported to micromolar values with pooled quality control sample normalization by median. Subsequently, values under the lower limit of detection were imputed by one-fifth of the minimum positive values of their corresponding variables. These data were used for further analysis. |
| Ion Mode: | POSITIVE |
| Acquisition Parameters File: | UM-p400-HR-Thermo-3.pdf |
| Processing Parameters File: | UM-p400-HR-Thermo-3.pdf |
| Analysis Protocol File: | UM-p400-HR-Thermo-3.pdf |
| MS ID: | MS003457 |
| Analysis ID: | AN003708 |
| Instrument Name: | Thermo Q Exactive Plus Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | MS acquisition method, data processing method and software used for feature assignments are described in analysis protocol file attached. Please refer sections 10.3 (MS acquisition parameters) and 5.3 (data processing) of the method file attached. The final quantitative results were exported to micromolar values with pooled quality control sample normalization by median. Subsequently, values under the lower limit of detection were imputed by one-fifth of the minimum positive values of their corresponding variables. These data were used for further analysis. |
| Ion Mode: | POSITIVE |
| Acquisition Parameters File: | UM-p400-HR-Thermo-3.pdf |
| Processing Parameters File: | UM-MetIDQ-Oxygen-9.pdf |
| Analysis Protocol File: | UM-p400-HR-Thermo-3.pdf |
| MS ID: | MS003458 |
| Analysis ID: | AN003709 |
| Instrument Name: | Thermo Q Exactive Plus Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | MS acquisition method, data processing method and software used for feature assignments are described in analysis protocol file attached. Please refer sections 10.3 (MS acquisition parameters) and 5.3 (data processing) of the method file attached. The final quantitative results were exported to micromolar values with pooled quality control sample normalization by median. Subsequently, values under the lower limit of detection were imputed by one-fifth of the minimum positive values of their corresponding variables. These data were used for further analysis. |
| Ion Mode: | POSITIVE |
| Acquisition Parameters File: | UM-p400-HR-Thermo-3.pdf |
| Processing Parameters File: | UM-MetIDQ-Oxygen-9.pdf |
| Analysis Protocol File: | UM-p400-HR-Thermo-3.pdf |
