How Are Lipids Extracted in Lipid Metabolomics Research?

Lipid metabolomics involves the comprehensive study of lipid molecules within cells and their dynamic changes. This field investigates lipid biosynthesis, metabolism, and their functions in physiological and pathological processes. Lipid extraction is a critical step in metabolomics research, and the following procedure outlines the standard method:

 

1. Sample Preparation

Biological samples, such as blood, urine, cells, or tissues, are collected. To prevent degradation during collection and processing, samples may require storage at low temperatures or the addition of specific stabilizers.

 

2. Physical Disruption

For solid samples (e.g., tissues), physical disruption is necessary to improve lipid extraction efficiency. Common methods include homogenization, grinding, or ultrasonic treatment.

 

3. Internal Standard Addition

To enable accurate quantification and correct for extraction efficiency or instrument-related losses, internal standards with known concentrations (typically stable isotope-labeled lipids) are added to the sample.

 

4. Lipid Extraction

The most widely used lipid extraction methods include the Folch, Bligh and Dyer, or methanol-chloroform extraction techniques:

(1) Chloroform-methanol (typically in a 2:1 ratio) is added to the sample and thoroughly mixed to extract lipids, allowing the efficient recovery of diverse lipid classes.

(2) If required, water is added to induce phase separation, typically at a ratio of 1:1:0.9 (chloroform:methanol:water). After mixing, lipids predominantly partition into the chloroform (organic) layer.

 

5. Centrifugation and Phase Separation

After water addition, centrifugation is used to separate the organic phase (lower, lipid-rich) from the aqueous phase (upper, containing proteins and other polar compounds).

 

6. Collection of the Organic Phase

The lipid-containing organic phase (chloroform layer) is carefully collected without disturbing the aqueous layer to ensure the integrity of lipid extracts.

 

7. Solvent Removal

Organic solvents are evaporated using nitrogen gas purging or a vacuum rotary evaporator, resulting in purified lipids.

 

8. Lipid Reconstitution

The dried lipid extract is reconstituted in an appropriate solvent (such as methanol or a chloroform/methanol mixture) for further lipid analysis, including liquid chromatography-mass spectrometry (LC-MS).

 

These steps can be tailored and optimized based on specific research needs to ensure high-quality lipid extraction. As a fundamental step in lipid metabolomics, efficient lipid extraction is essential for generating high-quality samples for mass spectrometry, chromatography, or other analytical techniques.

 

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