Summary of Study ST002136

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 PR001353. The data can be accessed directly via it's Project DOI: 10.21228/M8TQ43 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 IDST002136
Study TitleTargeted Microchip Capillary Electrophoresis-Orbitrap Mass Spectrometry Metabolomics to Monitor Ovarian Cancer Progression (calibration standards)
Study SummaryThe lack of effective screening strategies for high-grade serous carcinoma (HGSC), a subtype of ovarian cancer (OC) responsible for 80% of OC related deaths, emphasizes the need for new diagnostic markers and a better understanding of disease pathogenesis. Capillary electrophoresis (CE) coupled with high-resolution mass spectrometry (HRMS) offers high selectivity and sensitivity, thereby increasing metabolite coverage and consequently enhancing biomarker discovery. Recent advances in CE-MS include small, chip-based CE systems coupled with nanoelectrospray ionization (nanoESI) to provide rapid, high-resolution analysis of biological specimens. Here, we describe the development of a targeted microchip (µ) CE-HRMS method to analyze 40 target metabolites in serum samples from a triple-mutant (TKO) mouse model of HGSC, with an acquisition time of only 3 min. Extracted ion electropherograms showed sharp, highly resolved peak shapes, even for structural isomers such as leucine and isoleucine. All analytes maintained good linearity with an average R2 of 0.994, while detection limits were in the nM range. Thirty metabolites were detected in mice serum, with recoveries ranging from 78 to 120 %, indicating minimal ionization suppression and good accuracy. We applied the µCE-HRMS method to sequentially-collected serum samples from TKO and TKO-control mice. Time-resolved analysis revealed characteristic temporal trends for amino acids, nucleosides, and amino acids derivatives associated with HGSC progression. Comparison of the µCE-HRMS dataset with non-targeted ultra-high performance liquid chromatography (UHPLC) – MS results revealed identical temporal trends for the 5 metabolites detected on both platforms, indicating the µCE-HRMS method performed satisfactorily in terms of capturing metabolic reprogramming due to HGSC progression, while reducing the total analysis time 3-fold.
Institute
Georgia Institute of Technology
Last NameSah
First NameSamyukta
Address901 Atlantic Dr NW, Atlanta, GA, 30332, USA
Emailssah9@gatech.edu
Phone5746780124
Submit Date2022-04-11
Raw Data AvailableYes
Raw Data File Type(s)raw(Thermo)
Analysis Type DetailLC-MS
Release Date2022-05-02
Release Version1
Samyukta Sah Samyukta Sah
https://dx.doi.org/10.21228/M8TQ43
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:

Project:

Project ID:PR001353
Project DOI:doi: 10.21228/M8TQ43
Project Title:Targeted Microchip Capillary Electrophoresis-Orbitrap Mass Spectrometry Metabolomics to Monitor Ovarian Cancer Progression.
Project Summary:The lack of effective screening strategies for high-grade serous carcinoma (HGSC), a subtype of ovarian cancer (OC) responsible for 80% of OC related deaths, emphasizes the need for new diagnostic markers and a better understanding of disease pathogenesis. Capillary electrophoresis (CE) coupled with high-resolution mass spectrometry (HRMS) offers high selectivity and sensitivity, thereby increasing metabolite coverage and consequently enhancing biomarker discovery. Recent advances in CE-MS include small, chip-based CE systems coupled with nanoelectrospray ionization (nanoESI) to provide rapid, high-resolution analysis of biological specimens. Here, we describe the development of a targeted microchip (µ) CE-HRMS method to analyze 40 target metabolites in serum samples from a triple-mutant (TKO) mouse model of HGSC, with an acquisition time of only 3 min. Extracted ion electropherograms showed sharp, highly resolved peak shapes, even for structural isomers such as leucine and isoleucine. All analytes maintained good linearity with an average R2 of 0.994, while detection limits were in the nM range. Thirty metabolites were detected in mice serum, with recoveries ranging from 78 to 120 %, indicating minimal ionization suppression and good accuracy. We applied the µCE-HRMS method to sequentially-collected serum samples from TKO and TKO-control mice. Time-resolved analysis revealed characteristic temporal trends for amino acids, nucleosides, and amino acids derivatives associated with HGSC progression. Comparison of the µCE-HRMS dataset with non-targeted ultra-high performance liquid chromatography (UHPLC) – MS results revealed identical temporal trends for the 5 metabolites detected on both platforms, indicating the µCE-HRMS method performed satisfactorily in terms of capturing metabolic reprogramming due to HGSC progression, while reducing the total analysis time 3-fold.
Institute:Georgia Institute of Technology
Last Name:Sah
First Name:Samyukta
Address:901 Atlantic Dr NW, Atlanta, GA, 30332, USA
Email:ssah9@gatech.edu
Phone:5746780124

Subject:

Subject ID:SU002221
Subject Type:Synthetic sample

Factors:

Subject type: Synthetic sample; Subject species: - (Factor headings shown in green)

mb_sample_id local_sample_id factor
SA2052100.1uMstdmix_run1standard mix
SA2052110.1uMstdmix_run2standard mix
SA2052120.05uM_stdmix_run2standard mix
SA20521325uM_run3standard mix
SA2052140.05uM_stdmix_run1standard mix
SA20521525uM_run2standard mix
SA2052165uM_run2standard mix
SA20521710uM_run1standard mix
SA20521810uM_run2standard mix
SA2052191uM_stdmix_run1standard mix
SA2052201uM_stdmix_run2standard mix
SA20522110uM_stdmix_run2standard mix
SA205222sample2-2-run2standard mix
SA205223sample2-2-run3standard mix
SA20522410uM_stdmix_run1standard mix
SA20522525uM_stdmix_run2standard mix
SA2052262.5uM_stdmix_run1standard mix
SA2052272.5uM_stdmix_run2standard mix
SA20522825uM_stdmix_run1standard mix
SA2052295uM_run1standard mix
SA2052302.5uM_run2standard mix
SA205231sample4-1standard mix
SA205232sample4-2standard mix
SA205233sample5-1standard mix
SA205234sample5-2standard mix
SA205235sample3-2standard mix
SA205236sample3-1standard mix
SA205237sample1-3standard mix
SA205238sample2-1standard mix
SA205239sample2-2standard mix
SA205240sample6-1standard mix
SA205241sample6-2standard mix
SA2052420.025uM_run2standard mix
SA2052431uM_run1standard mix
SA2052441uM_run2standard mix
SA2052452.5uM_run1standard mix
SA2052460.025uM_run1standard mix
SA2052470.05uM_run2standard mix
SA2052480.1uM_run1standard mix
SA2052490.1uM_run2standard mix
SA2052500.05uM_run1standard mix
SA205251sample1-2standard mix
Showing results 1 to 42 of 42

Collection:

Collection ID:CO002214
Collection Summary:To improve CE peak shape and support electrophoretic focusing, a sample diluent consisting of 133 mM ammonium acetate, and 0.1% formic acid was prepared.13 This sample diluent was spiked with 1 µM 13C phenylalanine, 3 µM 13C6 arginine and 0.8 µM 13C methionine D3 as internal standards. Calibration mixtures were prepared from serial dilution of stock standard solutions using the spiked sample diluent in a 1:4 ratio. Each calibration standard was analyzed twice to yield calibration curves, calculate figures of merit, and perform metabolite quantification.
Sample Type:Synthetic Mixture

Treatment:

Treatment ID:TR002233
Treatment Summary:To improve CE peak shape and support electrophoretic focusing, a sample diluent consisting of 133 mM ammonium acetate, and 0.1% formic acid was prepared.13 This sample diluent was spiked with 1 µM 13C phenylalanine, 3 µM 13C6 arginine and 0.8 µM 13C methionine D3 as internal standards. Calibration mixtures were prepared from serial dilution of stock standard solutions using the spiked sample diluent in a 1:4 ratio. Each calibration standard was analyzed twice to yield calibration curves, calculate figures of merit, and perform metabolite quantification.

Sample Preparation:

Sampleprep ID:SP002227
Sampleprep Summary:To improve CE peak shape and support electrophoretic focusing, a sample diluent consisting of 133 mM ammonium acetate, and 0.1% formic acid was prepared.13 This sample diluent was spiked with 1 µM 13C phenylalanine, 3 µM 13C6 arginine and 0.8 µM 13C methionine D3 as internal standards. Calibration mixtures were prepared from serial dilution of stock standard solutions using the spiked sample diluent in a 1:4 ratio. Each calibration standard was analyzed twice to yield calibration curves, calculate figures of merit, and perform metabolite quantification.

Combined analysis:

Analysis ID AN003497
Analysis type MS
Chromatography type CE
Chromatography system ZipChip (908 Devices)
Column HS chip
MS Type ESI
MS instrument type Orbitrap
MS instrument name Thermo Exactive Plus Orbitrap
Ion Mode POSITIVE
Units Micromolars

Chromatography:

Chromatography ID:CH002583
Instrument Name:ZipChip (908 Devices)
Column Name:HS chip
Chromatography Type:CE

MS:

MS ID:MS003257
Analysis ID:AN003497
Instrument Name:Thermo Exactive Plus Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:All experiments were performed in positive ionization mode in the 50-500 m/z range at a mass resolution setting of 17,500. The capillary temperature was set to 200 °C and the sheath gas flow rate was 2 psi. The automatic gain control (AGC) target value was set to 3E6 and the maximum injection time was 20 ms. Data were acquired using Xcalibur 3.0 (Thermo Scientific) and were imported to Skyline software14 for peak picking and integration. The peak picking procedure used the analyte accurate m/z and migration time. Peak areas obtained from Skyline were exported as spreadsheets for further analysis. Quantitation was performed with the analyte peak areas relative to the peak area of one of the three isotopically labeled internal standards (13C6 arginine, 13C methionine D3 and 13C phenylalanine) chosen based on migration time similarities.
Ion Mode:POSITIVE
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