Summary of Study ST002543

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench,, where it has been assigned Project ID PR001638. The data can be accessed directly via it's Project DOI: 10.21228/M8ZX4D This work is supported by NIH grant, U2C- DK119886.


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Study IDST002543
Study TitleGC/MS analysis of hypoxic volatile metabolic markers in the MDA-MB-231 breast cancer cell line
Study SummaryHypoxia in disease describes persistent low oxygen conditions, observed in a range of pathologies, including cancer. In the discovery of biomarkers in biological models, pathophysiological traits present a source of translatable metabolic products for the diagnosis of disease in humans. Part of the metabolome is represented by its volatile, gaseous fraction; the volatilome. Human volatile profiles, such as those found in breath, are able to diagnose disease, however accurate volatile biomarker discovery is required to target reliable biomarkers to develop new diagnostic tools. Using custom chambers to control oxygen levels and facilitate headspace sampling, the MDA-MB-231 breast cancer cell line was exposed to hypoxia (1% oxygen) for 24 hours. The maintenance of hypoxic conditions in the system was successfully validated over this time period. Targeted and untargeted gas chromatography mass spectrometry approaches revealed four significantly altered volatile organic compounds when compared to control cells. Three compounds were actively consumed by cells: methyl chloride, acetone and n-Hexane. Cells under hypoxia also produced significant amounts of styrene. This work presents a novel methodology for identification of volatile metabolisms under controlled gas conditions with novel observations of volatile metabolisms by breast cancer cells.
University of York
Last NameIssitt
First NameTheo
AddressBiology Dept. University of York, Personal
Submit Date2023-03-31
Num Groups4
PublicationsT. Issitt et al., Volatile compounds in human breath: critical review and meta-analysis Journal of Breath Research, Volume 16, Number 2 (2022)
Analysis Type DetailGC-MS
Release Date2023-04-21
Release Version1
Theo Issitt Theo Issitt application/zip

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Sample Preparation:

Sampleprep ID:SP002649
Sampleprep Summary:Collected canister samples were transferred to a liquid nitrogen trap through pressure differential. Pressure change between beginning and end of “injection” was measured, allowing calculation of the moles of canister collected air injected. Sample in the trap was then transferred, via heated helium flow, to an Aglient/HP 5972 MSD system (Santa Clara, CA, USA) equipped with a PoraBond Q column (25 m x 0.32 mm x 0.5 μm film thickness) (Restek©, Bellefonte, PN, USA). Targeted samples were analyzed in selected ion monitoring (SIM) mode, and untargeted samples in full scan (SCAN) mode with the mass range of 45-200 amu. The mass spectrometer was operated in electron impact ionization mode with 70 eV ionization energy, and transfer line, ion source, and quadrupole temperatures of 250, 280 and 280, respectively. For details on SIM and significantly altered, identified SCAN compounds, see Table 1. All samples were analysed within 6 days of collection. The oven program for both SIM and SCAN analyses were identical and are as follows: 35 ˚C for 2 min, 10 ˚C/min to 155 ˚C, 1 ˚C/min to 131 ˚C and 25 ˚C/min to 250 with a 5 min 30 second hold. Calibration was performed using standard gases (BOC Specialty Gases, Woking, UK). Linear regression of calibration curves confirmed strong, positive linear relationships between observed compound peak areas and moles of gas injected for each VOC (r2 > 0.9 in all cases). For compounds not purchased in gaseous state (BOC Specialty gases, as above), 1–2 mL of compound in liquid phase was injected neat into butyl sealed Wheaton-style glass vials (100 mL) and allowed to equilibrate for 1 h. 1 mL of headspace air was then removed from neat vial headspace using a gas tight syringe (Trajan, SGE) and injected into the headspace of a second 100 mL butyl sealed Wheaton-style glass vial. This was then repeated, and 1 mL of the 2nd serial dilution vial was injected into the GCMS system with 29 mL of lab air to give ppb concentrations. This was performed for methanethiol (MeSH (SPEXorganics, St Neots, UK)), isoprene (Alfa Aesar, Ward Hill, MA, USA), acetone (Sigma-Aldrich, Burlington, MA, USA), 2- & 3-methyl pentane and n-hexane (Thermo Scientific, Waltham, MA, USA). Reported compounds detected by the GC/-MS were confirmed by matching retention times and mass–charge (m/z) ratios with known standards.
Processing Storage Conditions:Room temperature