Summary of Study ST003500
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 PR002146. The data can be accessed directly via it's Project DOI: 10.21228/M8783Z 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.
Study ID | ST003500 |
Study Title | A UHPLC-MS/MS Method for Profiling of Urinary Mercapturic Acids using Positive Ion Mode |
Study Summary | We report the first application of a UHPLC-MS/MS method using positive ion mode detection for the unbiased characterization of mercapturic acids. The proposed method utilizes a neutral loss monitoring paradigm to monitor for two diagnostic fragmentation pathways for this class of compound. Using a cohort of 20 nonsmokers and 20 smokers, we detected 180 putative mercapturic acid signatures that exhibited a high degree of reproducibility from the complex urine metabolome background. Following a combination of multivariate and univariate statistics, we found 33 putative mercapturic acids associated with smoking status. |
Institute | University of Minnesota |
Last Name | Murray |
First Name | Kevin |
Address | 2-210 CCRB, 2231 6th St SE, Minneapolis, MN 55455 |
murra668@umn.edu | |
Phone | 612-626-2182 |
Submit Date | 2024-08-01 |
Num Groups | 2 |
Total Subjects | 20 |
Raw Data Available | Yes |
Raw Data File Type(s) | mzML |
Analysis Type Detail | LC-MS |
Release Date | 2025-02-03 |
Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
Project ID: | PR002146 |
Project DOI: | doi: 10.21228/M8783Z |
Project Title: | A UHPLC-MS/MS Method for Profiling of Urinary Mercapturic Acids using Positive Ion Mode |
Project Summary: | We describe an analytical global profiling approach with machine learning predicted structural annotations for the characterization of mercapturic acids, a detoxification product of chemical environmental exposure. |
Institute: | University of Minnesota |
Department: | School of Public Health, Division of Environmental Health Sciences |
Laboratory: | Balbo Research Group |
Last Name: | Murray |
First Name: | Kevin |
Address: | 2-210 CCRB, 2231 6th St SE, Minneapolis, MN 55455 |
Email: | murra668@umn.edu |
Phone: | 612-626-2182 |
Funding Source: | National Cancer Institute (5R01CA222005-05) |
Subject:
Subject ID: | SU003629 |
Subject Type: | Human |
Subject Species: | Homo sapiens |
Taxonomy ID: | 9606 |
Age Or Age Range: | 21-61 |
Gender: | Male and female |
Human Smoking Status: | Smoking and Nonsmoking |
Species Group: | Mammals |
Factors:
Subject type: Human; Subject species: Homo sapiens (Factor headings shown in green)
mb_sample_id | local_sample_id | Sample source | Smoking Status |
---|---|---|---|
SA385691 | N149 | Urine | Non-Smoker |
SA385692 | N082 | Urine | Non-Smoker |
SA385693 | N215 | Urine | Non-Smoker |
SA385694 | N207 | Urine | Non-Smoker |
SA385695 | N194 | Urine | Non-Smoker |
SA385696 | N189 | Urine | Non-Smoker |
SA385697 | N167 | Urine | Non-Smoker |
SA385698 | N162 | Urine | Non-Smoker |
SA385699 | N161 | Urine | Non-Smoker |
SA385700 | N079 | Urine | Non-Smoker |
SA385701 | N142 | Urine | Non-Smoker |
SA385702 | N087 | Urine | Non-Smoker |
SA385703 | N140 | Urine | Non-Smoker |
SA385704 | N134 | Urine | Non-Smoker |
SA385705 | N133 | Urine | Non-Smoker |
SA385706 | N128 | Urine | Non-Smoker |
SA385707 | N096 | Urine | Non-Smoker |
SA385708 | N094 | Urine | Non-Smoker |
SA385709 | N088 | Urine | Non-Smoker |
SA385710 | N141 | Urine | Non-Smoker |
SA385711 | pool_10 | Urine | QC |
SA385712 | pool_09 | Urine | QC |
SA385713 | pool_08 | Urine | QC |
SA385714 | pool_07 | Urine | QC |
SA385715 | pool_06 | Urine | QC |
SA385716 | pool_05 | Urine | QC |
SA385717 | pool_03 | Urine | QC |
SA385718 | pool_02 | Urine | QC |
SA385719 | pool_01 | Urine | QC |
SA385720 | pool_04 | Urine | QC |
SA385721 | S147 | Urine | Smoker |
SA385722 | S190 | Urine | Smoker |
SA385723 | S171 | Urine | Smoker |
SA385724 | S163 | Urine | Smoker |
SA385725 | S157 | Urine | Smoker |
SA385726 | S156 | Urine | Smoker |
SA385727 | S152 | Urine | Smoker |
SA385728 | S151 | Urine | Smoker |
SA385729 | S150 | Urine | Smoker |
SA385730 | S144 | Urine | Smoker |
SA385731 | S146 | Urine | Smoker |
SA385732 | S145 | Urine | Smoker |
SA385733 | S138 | Urine | Smoker |
SA385734 | S136 | Urine | Smoker |
SA385735 | S135 | Urine | Smoker |
SA385736 | S132 | Urine | Smoker |
SA385737 | S131 | Urine | Smoker |
SA385738 | S117 | Urine | Smoker |
SA385739 | S102 | Urine | Smoker |
SA385740 | S110 | Urine | Smoker |
Showing results 1 to 50 of 50 |
Collection:
Collection ID: | CO003622 |
Collection Summary: | Adult participants were recruited as part of an ongoing study conducted at the University of Minnesota. Demographic information including gender, race, age, and tobacco usage was obtained through a questionnaire. Urine samples were collected throughout a 24-hour period and stored at -80 °C until the LC-MS/MS analysis. Urine samples were normalized by total 24-hour collection volume prior to sample preparation. Urinary creatinine levels measured using the RayBio Creatinine Assay kit (RayBiotech, Norcross, GA). This study was approved by the University of Minnesota Institutional Review Board. |
Sample Type: | Urine |
Storage Conditions: | -80℃ |
Treatment:
Treatment ID: | TR003638 |
Treatment Summary: | Urine samples were collected from non smokers and smokers. Urine samples were collected from patients over a 24-hour period. Sample were snap frozen after collection. No additional sample treatment was applied. |
Sample Preparation:
Sampleprep ID: | SP003636 |
Sampleprep Summary: | Urine samples (200 μL) were acidified using 40 μL of a 30% aqueous HCl solution and vortexed gently. A mixture of 10 ng D5-PhMA, 20 ng D3-2CaEMA, 20 ng D3-2CyEMA, 10 ng D3-2HPMA, 50 ng D3-3HPMA, and 50 ng D3-3HMPMA was added to each sample. Oasis MAX mixed mode reverse phase anion exchange solid phase extraction cartridges (60 mg, 60 u, 2 mL reservoir size) were obtained from Waters Corp. (Milford, MA, USA). Before sample introduction, the plate was preconditioned with MeOH, water, and 2% aqueous NH4OH solutions. The samples were applied and washed with 0.7 mL 2% aqueous NH4OH and 0.7 mL MeOH, then dried with nitrogen gas for 20 min. The plate was washed with 0.7 mL 2% aqueous formic acid before sample collection. Finally, the unfractionated urine extracts were collected using 0.7 mL 90% MeOH in 2% formic acid wash. The samples were transferred from the 96-well collection plate to fresh 1.2 mL silanized vials and dried to completeness in a SpeedVac without heat. The dried samples were stored at -80 °C until ready for LC-MS/MS analysis. |
Processing Storage Conditions: | Room temperature |
Extract Storage: | -80℃ |
Combined analysis:
Analysis ID | AN005745 |
---|---|
Analysis type | MS |
Chromatography type | Reversed phase |
Chromatography system | Thermo Dionex Ultimate 3000 RS |
Column | Self-packed C18 column Dr. Maisch GmbH ReproSil-PUR (450 mm x 100 µm, 1.9 µm, 120 Å C18aq) |
MS Type | ESI |
MS instrument type | Orbitrap |
MS instrument name | Thermo Orbitrap Fusion Lumos Tribrid |
Ion Mode | POSITIVE |
Units | Peak heights |
Chromatography:
Chromatography ID: | CH004360 |
Methods Filename: | MA_18786_hcdOT_30K_CNL-MS3-50CE_77min_20230510 |
Instrument Name: | Thermo Dionex Ultimate 3000 RS |
Column Name: | Self-packed C18 column Dr. Maisch GmbH ReproSil-PUR (450 mm x 100 µm, 1.9 µm, 120 Å C18aq) |
Column Temperature: | 55 °C |
Flow Gradient: | Chromatographic separation was performed using a linear gradient starting at 0% B and increased to 50% B at 40 min, 90% B at 60 min, and held for 8 min followed by a return to starting conditions. |
Flow Rate: | 325 nL/min |
Sample Injection: | 1% sample equivalent load in 1 μL reconstituted using a load solvent mixture of 0.1% aqueous formic acid. |
Solvent A: | 99.9% water; 0.1% formic acid |
Solvent B: | 99.9% acetonitrile; 0.1% formic acid |
Analytical Time: | 68 min |
Sample Loop Size: | 10 µL injection loop |
Chromatography Type: | Reversed phase |
MS:
MS ID: | MS005468 |
Analysis ID: | AN005745 |
Instrument Name: | Thermo Orbitrap Fusion Lumos Tribrid |
Instrument Type: | Orbitrap |
MS Type: | ESI |
MS Comments: | Discovery DDA-CNL-MS3 analyses of experimental samples were performed using full-scan detection followed by data-dependent MS2 acquisition and a conditional neutral loss-triggered MS3 acquisition. All analyses were performed in positive ion mode. Full-scan detection was performed using Orbitrap detection at a resolution of 120,000, AGC targeted setting of 4 × 105, a maximum ion injection time of 50 ms, and an S-Lens RF setting of 40%. Scan ranges of 170 m/z – 600 m/z were used for full-scan detection. MS2 spectra were collected using a DDA design with a 2 sec cycle time in centroid mode with a 15 sec dynamic exclusion list. Fragment spectra were acquired with quadrupole isolation of 0.8 m/z, Orbitrap detection at a resolution of 30,000, an AGC setting of 1 × 105, and a 100 ms maximum injection time. The analysis utilized HCD fragmentation at a fixed collision energy of 30%. MS3 spectra were collected using a conditional neutral loss design that acquired an additional fragmentation scan on ions exhibiting a neutral loss of 131.0582 Da or 105.0426 Da from the selected precursor. MS3 spectra were acquired with a full-scan MS isolation of 1.5 m/z and MS2 isolation of 2.0 m/z, Orbitrap detection at a resolution of 30,000, AGC setting of 2 × 105, and a 200 ms maximum injection time. The MS3 analysis utilized HCD fragmentation at a fixed collision energy of 50%. Data processing: Fragmentation filtering was performed using the DFBuilder module incorporated in the open-source software MZmine to monitor diagnostic neutral loss and fragment ions characteristic of mercapturic acid conjugates. Putative mercapturic acid detection required the observation of a neutral loss of 131.0582 Da (C5H9NO3) or 105.0426 Da (C3H7NO3) in acquired MS/MS spectrum. A complete list of the processing parameters is summarized in Table S2. Fragmentation filtering using the DFBuilder module was performed for the specified neutral loss using a 5 ppm mass tolerance and minimum fragment ion intensity of 1 x 104 with a 1% base peak threshold. A targeted chromatogram was drawn around detected precursor masses using a 1.0 min retention time window. Masses of interest were detected using the centroid detector and noise threshold of 1 x 105. Chromatogram traces were computed using the ADAP chromatogram builder module with a mass tolerance of 5 ppm. Chromatographic peaks were detected using the Local Minimum search algorithm using a 30% chromatographic threshold, 0.1 min retention time minimum search range, a minimum peak height of 1 x 105, a minimum peak top/edge ratio of 10, and minimum peak width tolerance of 0.1 min. Duplicate peaks were removed using a 0.1 min retention time tolerance. Feature lists were deisotoped using the 13C isotope filter module. All samples and pooled replicate feature lists were aligned using the Join Aligner module with a mass tolerance of 5 ppm and retention time tolerance of 0.4 min. Commonly observed in-source fragments, adducts, and multimer complexes were detected and removed from consideration. Missing peak annotations were replaced using the Gap-Filling algorithm. Acquired MS/MS spectra were matched against the NIST mass spectral library (version 2020) to remove unrelated metabolic signatures. Finally, the remaining features were exported as CSV files for downstream analysis. |
Ion Mode: | POSITIVE |
Collision Energy: | NCE 30% |
Fragmentation Method: | HCD |
Ion Spray Voltage: | 2.1 kV |
Ionization: | ESI |
Mass Accuracy: | 5 ppm |
Precursor Type: | [M+H]+ |