Summary of Study ST000595
This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR000434. The data can be accessed directly via it's Project DOI: 10.21228/M8ZC8D This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.
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| Study ID | ST000595 |
| Study Title | Emory University high-resolution metabolomic profiling CHEAR pooled reference materials: Urine |
| Study Type | Untargeted HRMS for cross-laboratory characterization of common reference material. |
| Study Summary | Proper QA/QC is an essential component of any analytical procedure. The variability in platforms and approaches used by the untargeted cores requires that each lab develop specific QA/QC protocols, making cross-laboratory assessments challenging. To address the difference in measures across the untargeted cores, the untargeted quality assurance working group (UQAWG) has identified the importance of using the same biological reference sample across the different laboratories. To reach this goal, Dr. Jones and Mr. Walker of Emory University volunteered to take the lead in identifying vendors, requesting quotes, purchasing the pooled material, delivering the specified volumes to each of the lab hubs and storing additional material on-site at Emory University. Based upon discussion with the UQAWG, it was decided that each participating laboratory (five total) will receive 500 mL of plasma and 2000 mL of urine, providing sufficient volume for daily analysis over the next five years. Initially, NIST standard reference materials were considered; however, limited supply and cost makes the use of NIST materials impractical. Vendors, including BioreclamationIVT, Lee Biosolutions Inc and Gemini Bio-Products were contacted to request quotes for preparing pooled plasma and urine meeting the following requirements: K2EDTA recovered whole blood (plasma), equal distribution male/female, equal distribution of Black/Hispanic/Caucasian donors and be collected from greater than 40 donors. In addition, we requested all blood samples be collected in FDA inspected facilities, provide collection details, provide general demographics for each donor and be delivered in 500 mL aliquots. BioreclamationIVT, who also provided the biological samples for the NIH metabolomics ring trial, was selected as the most appropriate vendor due to the ability to provide the requested materials, lead-time to deliver and price. To date, both pools have been purchased. The urine pool was received on July 21, 2016 and the plasma pool will be received July 26, 2016. Upon receipt, both pools will be stored at -80°C. Summary demographics were provided by BioreclamationIVT, the urine pool was created by combining urine samples from 60 males and 60 females with an average age of 38 (range 18-60) and equal distribution of races. The plasma pool was created by combining recovered plasma obtained from 50 males and 50 females with an average age of 38.4 (range 19-72) and equal distribution of races. Both urine and plasma pools will be delivered to the lab hubs frozen in units of 500 mL. Upon receipt, plasma and urine pools should be aliquoted into volumes that provide sufficient material so that freeze-thaw of the pooled material is minimized. For unambiguous identification, the urine pool has been named UrineRef_20160721 and the plasma pool has been named PlasmaRef_20160726. The purpose of providing each untargeted lab hub with the pooled material is two-fold. Through analysis of UrineRef_20160721 (if study samples are urine) or PlasmaRef_20160726 (if study samples are plasma) in each batch of samples analyzed for CHEAR, it will be possible to have a common reference across the different lab hubs. Thus, the ability to compare analyses on different platforms will be greatly improved. The use of a consistent reference material in each batch will also facilitate reference standardization, a concept developed for retrospective quantification of high-resolution metabolomics (Go, Walker et al. 2015). Using reference standardization, quantification post-data acquisition is possible by referencing the pooled material analyzed with each series of samples. The pooled material can be characterized and analytes quantified using traditional analytical chemistry techniques, such as quantification by methods of addition or standardization with a certified reference material, such as NIST SRM 1950. Known concentrations within the pooled material can be used to determine an analyte response factor and calculate sample concentrations based on single-point calibration. The benefit of this approach is that targeted quantification is only required in the pooled sample (i.e. UrineRef_20160721 or PlasmaRef_20160726), chemicals selected for quantification do not need to be selected a priori, and population wide estimates of plasma chemical concentrations can be determined without having to re-analyze samples using a targeted approach. Current Clinical Biomarker Laboratory protocol requires three analytical replicates of pooled reference material per day (three technical replicates per analytical replicate per column configuration, or 18 injections/instrument-day), resulting in analyses of the reference material every 10 samples. Results from untargeted LC-HRMS analyses of pooled CHEAR reference materials. PlasmaRef_20160726 was analyzed with additional NIST reference materials: SRM 1950, SRM 1957 and SRM 1958. UrineRef_20160721 was analyzed with NIST reference materials SRM 3672 and 3673. |
| Institute | Emory University |
| Department | School of Medicine, Division of Pulmonary, Allergy, Critical Care Medicine |
| Laboratory | Clincal Biomarkers Laboratory |
| Last Name | Walker |
| First Name | Douglas |
| Address | 615 Michael St. Ste 225, Atlanta, GA, 30322, USA |
| douglas.walker@emory.edu | |
| Phone | 4047275984 |
| Submit Date | 2017-02-21 |
| Num Groups | 2 |
| Study Comments | Only pooled material was analyzed. |
| Raw Data Available | Yes |
| Raw Data File Type(s) | raw(Thermo) |
| Chear Study | Yes |
| Analysis Type Detail | LC-MS |
| Release Date | 2024-12-31 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR000434 |
| Project DOI: | doi: 10.21228/M8ZC8D |
| Project Title: | Characterization of CHEAR reference material UrineRef_20160721 |
| Project Type: | Untargeted HRMS profiling |
| Project Summary: | Results from untargeted LC-HRMS analyses of pooled CHEAR reference materials. UrineRef_20160721 was analyzed with additional NIST reference materials: SRM 3672 and SRM3673. |
| Institute: | Emory University |
| Department: | Division of Pulmonary, Allergy, Critical Care, and Sleep Medicine |
| Laboratory: | Clinical Biomarkers Laboratory |
| Last Name: | Walker |
| First Name: | Douglas |
| Address: | 625 Michael St. Ste 225, Atlanta, GA, 30322, USA |
| Email: | douglas.walker@emory.edu |
| Phone: | 4047275984 |
| Funding Source: | NIEHS ES026560 |
Subject:
| Subject ID: | SU000618 |
| Subject Type: | Human Urine |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
| Species Group: | Mammals |
Factors:
Subject type: Human Urine; Subject species: Homo sapiens (Factor headings shown in green)
| mb_sample_id | local_sample_id | Source |
|---|---|---|
| SA032668 | Uri-q3June2014_1a | Plasma |
| SA032669 | Uri-q3June2014_1e | Plasma |
| SA032670 | Uri-q3June2014_1f | Plasma |
| SA032671 | Uri-q3June2014_1c | Plasma |
| SA032672 | Uri-q3June2014_1d | Plasma |
| SA032673 | Uri-q3June2014_1b | Plasma |
| SA032674 | ChearUrine_08 | Urine |
| SA032675 | NIST_SRM 3672_09 | Urine |
| SA032676 | NIST_SRM 3673_08 | Urine |
| SA032677 | ChearUrine_07 | Urine |
| SA032678 | NIST_SRM 3673_07 | Urine |
| SA032679 | NIST_SRM 3673_09 | Urine |
| SA032680 | NIST_SRM 3672_08 | Urine |
| SA032681 | ChearUrine_09 | Urine |
| SA032682 | NIST_SRM 3673_01 | Urine |
| SA032683 | NIST_SRM 3672_01 | Urine |
| SA032684 | ChearUrine_10 | Urine |
| SA032685 | NIST_SRM 3673_10 | Urine |
| SA032686 | NIST_SRM 3672_10 | Urine |
| SA032687 | NIST_SRM 3672_07 | Urine |
| SA032688 | ChearUrine_06 | Urine |
| SA032689 | NIST_SRM 3672_04 | Urine |
| SA032690 | NIST_SRM 3673_04 | Urine |
| SA032691 | ChearUrine_03 | Urine |
| SA032692 | NIST_SRM 3673_03 | Urine |
| SA032693 | ChearUrine_02 | Urine |
| SA032694 | NIST_SRM 3672_03 | Urine |
| SA032695 | ChearUrine_04 | Urine |
| SA032696 | NIST_SRM 3672_05 | Urine |
| SA032697 | NIST_SRM 3672_06 | Urine |
| SA032698 | NIST_SRM 3673_06 | Urine |
| SA032699 | ChearUrine_01 | Urine |
| SA032700 | NIST_SRM 3672_02 | Urine |
| SA032701 | NIST_SRM 3673_05 | Urine |
| SA032702 | ChearUrine_05 | Urine |
| SA032703 | NIST_SRM 3673_02 | Urine |
| Showing results 1 to 36 of 36 |
Collection:
| Collection ID: | CO000612 |
| Collection Summary: | The urine pool was received by Emory University on July 21, 2016 and stored at -80°C. Summary demographics were provided by BioreclamationIVT. The urine pool was created by combining urine samples from 60 males and 60 females with an average age of 38 (range 18-60) and equal distribution of races. Prior to analysis, the urine was aliquoted into 0.5 mL units. For unambiguous identification, the urine pool has been named UrineRef_20160721. |
| Sample Type: | Human urine |
| Blood Serum Or Plasma: | Urine |
Treatment:
| Treatment ID: | TR000632 |
| Treatment Summary: | Samples were delivered in individual units of 500 mL. Aliquots of 0.5 mL were prepared after receiving the material from BioreclamationIVT and stored at -80C |
Sample Preparation:
| Sampleprep ID: | SP000625 |
| Sampleprep Summary: | Samples were prepared for metabolomics analysis using established methods (Johnson et al. (2010). Analyst; Go et al. (2015). Tox Sci). Prior to analysis, urine aliquots were removed from storage at -80°C and thawed on ice. Each cryotube was then vortexed briefly to ensure homogeneity, and 50 μL was transferred to a clean microfuge tube. Immediately after, the urine was treated with 100 μL of ice-cold LC-MS grade acetonitrile (Sigma Aldrich) containing 2.5 μL of internal standard solution with eight stable isotopic chemicals selected to cover a range of chemical properties. Following addition of acetonitrile, urine was equilibrated for 30 min on ice, upon which precipitated proteins were removed by centrifuge (16.1 ×g at 4°C for 10 min). The resulting supernatant (100 μL) was removed, added to a low volume autosampler vial and maintained at 4°C until analysis (<22 h). |
| Sampleprep Protocol ID: | HRM_SP_082016_01 |
| Sampleprep Protocol Filename: | EmoryUniversity_HRM_SP_082016_01.pdf |
| Sampleprep Protocol Comments: | Date effective: 30 July 2016 |
| Extraction Method: | 2:1 acetonitrile: sample followed by vortexing and centrifugation |
| Sample Spiking: | 2.5 uL [13C6]-D-glucose, [15N,13C5]-L-methionine, [13C5]-L-glutamic acid, [15N]-L-tyrosine, [3,3-13C2]-cystine, [trimethyl-13C3]-caffeine, [U-13C5, U-15N2]-L-glutamine, [15N]-indole |
Combined analysis:
| Analysis ID | AN000911 | AN000912 |
|---|---|---|
| Analysis type | MS | MS |
| Chromatography type | HILIC | Reversed phase |
| Chromatography system | Thermo Dionex Ultimate 3000 RS | Thermo Dionex Ultimate 3000 RS |
| Column | Waters XBridge Amide (50 x 2.1mm,2.5um) | Thermo Higgins C18 (50 x 2.1mm,3um) |
| MS Type | ESI | ESI |
| MS instrument type | Orbitrap | Orbitrap |
| MS instrument name | Thermo Fusion Orbitrap | Thermo Fusion Orbitrap |
| Ion Mode | POSITIVE | NEGATIVE |
| Units | unspecified | unspecified |
Chromatography:
| Chromatography ID: | CH000648 |
| Chromatography Summary: | The HILIC column is operated parallel to reverse phase column for simultaneous analytical separation and column flushing through the use of a dual head HPLC pump equipped with 10-port and 6-port switching valves. During operation of HILIC separation method, the MS is operated in positive ion mode and 10 μL of sample is injected onto the HILIC column while the reverse phase column is flushing with wash solution. Flow rate is maintained at 0.35 mL/min until 1.5 min, increased to 0.4 mL/min at 4 min and held for 1 min. Solvent A is 100% LC-MS grade water, solvent B is 100% LC-MS grade acetonitrile and solvent C is 2% formic acid (v/v) in LC-MS grade water. Initial mobile phase conditions are 22.5% A, 75% B, 2.5% C hold for 1.5 min, with linear gradient to 77.5% A, 20% B, 2.5% C at 4 min, hold for 1 min, resulting in a total analytical run time of 5 min. During the flushing phase (reverse phase analytical separation), the HILIC column is equilibrated with a wash solution of 77.5% A, 20% B, 2.5% C. |
| Methods ID: | 2% formic acid in LC-MS grade water |
| Methods Filename: | 20160728_HILICpos_c18negwash_FullScan_5min |
| Chromatography Comments: | Triplicate injections for each chromatography mode |
| Instrument Name: | Thermo Dionex Ultimate 3000 RS |
| Column Name: | Waters XBridge Amide (50 x 2.1mm,2.5um) |
| Column Temperature: | 40C |
| Flow Gradient: | A= water, B= acetontrile, C= 2% formic acid in water; 22.5% A, 75% B, 2.5% C hold for 1.5 min, linear gradient to 77.5% A, 20% B, 2.5% C at 4 min, hold for 1 min |
| Flow Rate: | 0.35 mL/min for 1.5 min; linear increase to 0.4 mL/min at 4 min, hold for 1 min |
| Sample Injection: | 10 uL |
| Solvent A: | 100% water |
| Solvent B: | 100% acetonitrile |
| Analytical Time: | 5 min |
| Sample Loop Size: | 15 uL |
| Sample Syringe Size: | 25 uL |
| Chromatography Type: | HILIC |
| Chromatography ID: | CH000649 |
| Chromatography Summary: | The C18 column is operated parallel to the HILIC column for simultaneous analytical separation and column flushing through the use of a dual head HPLC pump equipped with 10-port and 6-port switching valves. During operation of the C18 method, the MS is operated in negative ion mode and 10 μL of sample is injected onto the C18 column while the HILIC column is flushing with wash solution. Flow rate is maintained at 0.4 mL/min until 1.5 min, increased to 0.5 mL/min at 2 min and held for 3 min. Solvent A is 100% LC-MS grade water, solvent B is 100% LC-MS grade acetonitrile and solvent C is 10mM ammonium acetate in LC-MS grade water. Initial mobile phase conditions are 60% A, 35% B, 5% C hold for 0.5 min, with linear gradient to 0% A, 95% B, 5% C at 1.5 min, hold for 3.5 min, resulting in a total analytical run time of 5 min. During the flushing phase (HILIC analytical separation), the C18 column is equilibrated with a wash solution of 0% A, 95% B, 5% C until 2.5 min, followed by an equilibration solution of 60% A, 35% B, 5% C for 2.5 min. |
| Methods ID: | 10mM ammonium acetate in LC-MS grade water |
| Methods Filename: | 20160728_c18neg_HILICposwash_FullScan_5min |
| Chromatography Comments: | Triplicate injections for each chromatography mode |
| Instrument Name: | Thermo Dionex Ultimate 3000 RS |
| Column Name: | Thermo Higgins C18 (50 x 2.1mm,3um) |
| Column Temperature: | 40C |
| Flow Gradient: | A= water, B= acetontrile, C= 10mM ammonium acetate in water; 60% A, 35% B, 5% C hold for 0.5 min, linear gradient to 0% A, 95% B, 5% C at 1.5 min, hold for 3 min |
| Flow Rate: | 0.4 mL/min for 1.5 min; linear increase to 0.5 mL/min at 2 min held for 3 min |
| Sample Injection: | 10 uL |
| Solvent A: | 100% water |
| Solvent B: | 100% acetonitrile |
| Analytical Time: | 5 min |
| Sample Loop Size: | 15 uL |
| Sample Syringe Size: | 25 uL |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS000810 |
| Analysis ID: | AN000911 |
| Instrument Name: | Thermo Fusion Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| Ion Mode: | POSITIVE |
| Capillary Temperature: | 300C |
| Collision Gas: | N2 |
| Dry Gas Flow: | 45 |
| Dry Gas Temp: | 250C |
| Mass Accuracy: | < 3ppm |
| Spray Voltage: | +3500 |
| Activation Parameter: | 5e5 |
| Activation Time: | 118ms |
| Interface Voltage: | S-Lens RF level= 69 |
| Resolution Setting: | 60,000 |
| Scanning Range: | 85-1275 |
| Analysis Protocol File: | EmoryUniversity_HRM_FusionMS_082016_01.pdf |
| MS ID: | MS000811 |
| Analysis ID: | AN000912 |
| Instrument Name: | Thermo Fusion Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| Ion Mode: | NEGATIVE |
| Capillary Temperature: | 300C |
| Collision Gas: | N2 |
| Dry Gas Flow: | 45 |
| Dry Gas Temp: | 250C |
| Mass Accuracy: | < 3ppm |
| Spray Voltage: | -4000 |
| Activation Parameter: | 5e5 |
| Activation Time: | 118ms |
| Interface Voltage: | S-Lens RF level= 69 |
| Resolution Setting: | 60,000 |
| Scanning Range: | 85-1275 |
| Analysis Protocol File: | EmoryUniversity_HRM_FusionMS_082016_01.pdf |
