Lipids can be extracted from tissues using organic solvents. The extracted lipids are then separated using techniques like thin layer chromatography, gas liquid chromatography, or high-performance liquid chromatography. Individual lipids can be identified by their behavior during chromatography or through mass spectrometry. Lipidomic analysis determines the full lipid profile of a cell, tissue, or organism and how it changes in response to stressors or differentiation.

1. By : Hammad Wali
Students of Biotechnology 4th- E
Quaid e Azam University
Lipids
(Methods to study lipids)

2. Table of
Contents
01 Introduction – Extraction
02 Seperation
03 Analysis

3. Introduction
• The word lipid is derived from a Greek word “lipos”
• They are soluble in non-polar solvents
• Lipids are hydrophobic
• Unlike the proteins, nucleic acids, and polysaccharides,
lipids are not polymer
• insoluble in water
• chief storage form of energy
• Fat and oils are the principle stored forms of energy
in many organism.

4. Functions Of Lipids
• Storage form of metabolic fuel.
• Transport form of metabolic fuel
• Provide the structural components of
membranes
• Serve as pigment
• Detergent
• Cofactors
• Hormone
• Protective functions

5. Compound Lipids (HeteroLipids)
Simple Lipids ( MonoLipids)
Classification
of Lipids
They are Esters of Fatty acids
with various Alcohols
They are Esters of Fatty acids
with various Alcohols and
possess additional Groups
also
Derived Lipids
These are the substances
derived from simple and
compound lipids by hyrolysis

6. Derived Lipids
(Fatty Acids)
Saturated Fatty Acids
● Saturated fatty acids have no double bonds
● They have higher melting points
● solid at room temperature
Unsaturated Fatty Acids
● Contain one or more double bonds
● Liquid at room temperature
● Low melting point

7. Working with
Lipids
When the lipids are
separated , ,We
analyze and identify
the lipids by various
methods
E.G Mass spectrometry
, Hydrolysis
Analysis
The lipids needs to be
separated from the
organic solvent when
separated
E.G HPLC , TLC , GLC
Separation
Extracting lipids
from a cell or tissue
by using organic
solvents
Preparing a solution
enriched in lipids
E.G Solvent
Extraction

8. Extraction of Lipids
(Solvent Extraction Method)
This method requires organic solvent thus called ‘Solvent Extraction Method ‘
Commonly used for Extraction from Biological Molecules or Food samples
Polar Organic Solvents Used
• Ethanol
• Methanol
Solvents Used :
• Ethyl ether
• Chloroform
• Benzene
Clustering of Lipids due to Hydrophobic
interactions is prevented by these
solvents

9. • Mixture of Chloroform, Methanol and
Water is the most used solvent for
extraction from Tissues
• Tissue is homogenized in the above
solvent mixture
• When adding water, we see 2 phases
Top phase consists of water
Lower phase consists of chloroform
• Sugar and Proteins, the polar molecules
partition into the top phase
• While Lipids stays at the lower phase
Extraction with Commonly used Organic Solvent
(Steps of Extraction)

10. GAS LIQUID
CHROMATOGRAPHY Chromatography is an analytical technique
where in a sample, mixture is separated into
different components based on difference in
their affinity for a stationary phase and mobile
phase.

11. GLC (SEPARATION TECHNIQUE)
STATIONARY PHASE MOBILE PHASE
Non polar. Non volatile liquid Inert carrier gas
Bonded on to a solid support He, Ar

12. GLC PRINCIPLE
PARTITION CHROMATOGRAPHY
Non polar components present in
the carrier gas have a greater
affinity towards stationary
phase
Polar components remain in
the carrier gas
Retains on the stationary phase Carried through the column

13. GAS CHROMATOGRAPH

14. GAS CHROMATOGRAPHY MASS
SPECTROSCOPY
• Instrument that allows to separate individual
components in an essential oil according to its
volatility and mass.
• There are many factors that go into analyzing an
essential oil for authentication.
• Gold standard for determining the purity in essential
oils

15.  Prepared sample is injected.
 Port is heated.
 Components pass through column.
 Column provides a surface for components to
interact.
 It allows the components “slow” so separation
can occur.
 Column is made of different affinities.
 Oil passes through column and temperature of
oven increased.
 Components hit the detector.
 Proportional peaks are recorded on a
chromatograph.
 Ions travel through an electromagnetic field that
filters them based on their mass.
 Detectors counts each number of ions.
 Information is sent to computer where a mass
spectrum is created.

16. ● Lipidomic is a large-scale study of pathways and networks of cellular
lipids in biological systems.
● Lipidomic is used to describe:
● The complete lipid profile within a cell, tissue or organism
LIPIDOMICS
● Lipidomic research involves;
● Analyzing which lipid are and in
what proportions.
● Analyzing interactions of lipids
with other molecules.
● Quantitatively describing the
alterations in lipid content and
their composition in response to a
perturbation of a cell.

17. SUMMARY
• The first step in determining lipid composition from
tissues is extraction of lipids with organic solvents.
• Extracted lipids are then separated by thin layer, gas
liquid or high-performance liquid chromatography
techniques.
• Individual lipids can be identified by either their
behavior on the chromatography columns or by mass
spectrometry.
• Lipidomic is an extremely strong technique used to
determine the lipid profile in a cell/tissue/organism in
response to different stress conditions or during
differentiation.