Mar 19, 2025

Lipid Extraction for Mass Spectrometry Lipidomics

DOI
https://dx.doi.org/10.17504/protocols.io.36wgqd2zxvk5/v1
  • Patricia Yuste-Checa1,
  • Barbara Steigenberger2,
  • F Ulrich Hartl1
  • 1Department of Cellular Biochemistry, Max Planck Institute of Biochemistry, Martinsried, Germany;
  • 2Mass Spectrometry Core Facility, Max Planck Institute of Biochemistry, Martinsried, Germany
Patricia Yuste-Checa
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DOI: https://dx.doi.org/10.17504/protocols.io.36wgqd2zxvk5/v1
Protocol Citation: Patricia Yuste-Checa, Barbara Steigenberger, F Ulrich Hartl 2025. Lipid Extraction for Mass Spectrometry Lipidomics. protocols.io https://dx.doi.org/10.17504/protocols.io.36wgqd2zxvk5/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: March 10, 2025
Last Modified: March 19, 2025
Protocol Integer ID: 124318
Keywords: ASAPCRN, lipid extraction for mass spectrometry lipidomic, mass spectrometry lipidomic, extraction method compatible with mass spectrometry, lipid extraction, mass spectrometry, extraction, butyl ether
Funders Acknowledgements:
Aligning Science Across Parkinson's
Grant ID: ASAP-000282
Abstract
This protocol details how to perform lipid extraction using a methyl tert-butyl ether (MTBE)-based extraction method compatible with mass spectrometry.
Lipid Extraction
Add 200 µL of cold methanol and 800 µL of cold methyl tert-butyl ether (MTBE) to the samples.

Vortex.

Add 200 µL of water. Phase separation into aqueous and organic phases should occur.

Centrifuge the extraction mixture at 10000 x g, 4°C, 00:10:00 to separate the organic and the aqueous phases.

10m
Collect the upper organic phase.

Dry it in a SpeedVac and store at -80 °C until analysis.

For mass spectrometry analysis, reconstitute the samples with 20 µL -40 µL of acetonitrile/isopropanol/water in a 65:30:5 ratio (v/v/v).

Note
For LC-MS measurement, a C8 column (Luna 3u, 2 x 100 mm, 3.0 µm, Phenomenex) is recommended. DMPC gets stuck in the C18 column.