LIPOSOMES

DEFINITION
The name liposome is derived from two
Greek words:
Lipo: meaning Fat
Soma: meaning body

Liposomes are simple microscopic
vesicles in which an aqueous volume
is entirely enclosed by a membrane
composed of a lipid molecule
History
Liposomes were first described
by British haematologist Dr
Alec D Bangham and
colleagues in 1961 (published
1964).
Structure
Structurally liposomes are concentric bilayered
vesicles in which an aqueous volume is
entirely enclosed by a membranous lipid
bilayer mainly composed of natural or
synthetic phospholipids
COMPONENTS
Phospholipids
major structural components
of biological membranes.
In the membrane cholesterol
increases separation between
choline head groups which
reduces the normal hydrogen
bonding and electrostatic
interaction
Drug is encapsulated in
i. Phospholipid bilayer
ii. In the entrapped aqueous
iii. At bilayer interface.
Activity
Head. Hydrophillic
Tail. Hydrophobic)
Hydrophobic portion is repelled by water
Hydrophilic portion is attracted by water
Activity of liposomes is enhancesd by modifying
the surface of liposomes with different
molecules such as glycolipids.

ACTIVITY
Hydrophilic drug is
entrapped in
hydrophilic portion
of liposomes.

Hydrophobic drug is
entrapped in
lipophillic portion.

Modes of Liposome/Cell Interaction
Adsorption
Endocytosis
Fusion
Lipid transfer
Mechanism of Drug Release
Liposome attaches to plasma membrane and
appears to fuse with them, releasing their
content into cell.


Membrane of phagolysosyme have proton pumps which
decrease pH of phagolysosyme & the enzyme
phospholipase destruct the liposomal membrane
Macrophages engulf liposomes (endocytosis)
Phagosome + lysosyme = phagolysosyme
Liposomes in blood stream
Taken by reticulo-endothelial system
Improved solubility of lipophilic and amphiphilic drugs .
Liposomes have both a lipophilic and aqueous
environment making it useful for delivering
hydrophobic, amphipathic, and hydrophilic medicines
It shows drug effect at its specific targeted site
Shows sustained release action.
Liposomes improved penetration of various drugs into
tissues
Liposomes increases efficacy and therapeutic index.
Liposomes protect the drug from environment and the
sensitive areas from drug.

Need For Liposomes
Disadvantages
High
production
cost
Liposome
phospholipid
may undergo
oxidation &
hydrolysis
Shorter Half
life
May have
leakage of
encapsulated
drug
Liposomes are
quickly taken
up by the
reticular
endothelial
cells
CLASSIFICATION
OF LIPOSOMES
According to Size

Name Diameter No. of lamella
Small unilamellar vesicles 20-100nm Single
Large unilamellar vesicles 100 nm - 400 nm Single
Intermediate-sized unilamellar
vesicles
100-200 nm

Single
Giant Unilamellar Vesicles 1 m & larger Single
Multivesicular Vesicles 200 nm - ~3 m Multiple

Names Diameter

Number of lamella
Unilamellar vesicles All sizes Single
Multilamellar vesicles 200 nm - ~3 m Multiple(5 & 20)
Multivesicular
liposomes
- -
ACCORDING TO MORPHOLOGY
According To Composition
Immunoliposomes
Liposomes using immunological
molecules particularly,
immunoglobulins for targeting purposes
may be attached to liposomes surface
by covalent linkage through membrane
possessing functional group




s

Proteosomes
A term applied to lipid
vesicles incorporating
proteins in or on the outer
membrane






According to Function
Stealth Tranfersomes
Long circulatory liposome
Any liposome that avoids uptake by the
RES as a result of coating the surface of
the liposomes with hydroxylated polymers
Particular composition that are capable
of transferring equeous content across
the skin
There membrane contains a certain
proportion of the bile salt distributed
among the phospholipid, which confers
the increased flexibility on membrane

Methods Of
Preparation
METHODS OF PREPARATION
All the methods of preparing the liposomes involve four
basic stages:








The difference between these methods is the steps by which
lipids are drying down from organic solvents and then
redispersed in aqueous media.


1. Drying down lipids
from organic solvent
2. Dispersing the lipid
in aqueous media
3. Purifying the
resultant liposome
4. Analyzing the final
product
SELECTION CRITERIA
Choice of method for liposomes production:

Physicochemical characteristics of drug to be loaded and
ingredients.
Nature of dispersion medium in which liposomes are
dispersed.
Effective concentration and toxicity of entrapped substance.
Size, dispersity and shelf life of vesicles for intended
application.
Batch to batch reproducibility.
Large-scale production of safe and efficient liposomal
products.

Mechanical Dispersion Method
1. Lipid Hydration Method
2. Lyophillization
3. Proliposomes
To Reduce Liposome
Size
Sonication
French Pressure Cell
Microemulsification
Membrane Extrusion
To Increase Liposome
Size
Freeze thawing
Direct reconstituted
vesicles

1. Mechanical Dispersion Method
Lipid dissolve in organic solvent/co-solvent

Remove organic solvent under vacuum

Film deposition

Solid lipid mixture is hydrated by using aqueous buffer

Lipid spontaneously swell and hydrate

Liposome

Post hydration vortexing, sonication, freeze thawing and high pressure extrusion
i. Lipid Hydration Method
Hand shaken vesicles
most widely used for the preparation of MLV



Drying of lipid solution forms a thin film at bottom
Hydrating this film by adding aqueous buffer
Vortexing it for some time
MLVs prepared
Lipids +Solvent
Non-shaking vesicles
Solution
of Lipids +
solvent
MLV are
floating on
surface
remaining
fluid
produces
LUV

Nitrogen
gas is
passed
Hydration
Until the opacitiy
of the dried lipid
film disappears.
Flask is slowly
returned to
upright position.
Take care not to
disturb the flask
in any way.
ii. Lyophilization
Freeze-drying involves the removal of water
from products in the frozen state at
extremely low pressures.
The process is generally used for dry
products.
iii. PROLIPOSOMES
Lipid is dried over the
finely divided
particulate support i.e.-
NaCl, Sorbitol, or other
polysaccharides.
These dried lipid coated
particulates are called
as proliposomes.
Drying lipid
on finely
divided
support
(NaCl)
Surface area
increases and
continuous
hydration
hydration swelling
MLVs

To Reduce Liposome Size
Sonication
French Pressure Cell
Micro emulsification
Membrane Extrusion

Sonication

most extensively used for the preparation of small
unilamellar vesicle (SUV). Sonication is the act of
applying sound energy to agitate particles in a
sample to reduce the size.


French pressure cell: extrusion
MLV dispersion are placed in the French Pressure
Cell and extruded at about 20,000psi at 45C

95% of MLVs get converted to SUVs which can be
determined by size extrusion chromatoraphy
Dispersion of MLVs can be
converted to SUVs by passage
through a small orifice under
high pressure.

MICRO EMULSIFICATION LIPOSOMES
The lipids can be introduced into fluidizers, either
as a dispersion of large mlvs or as a slurry of
unhydrated lipids in organic medium.
Microfluidizer pumps the fluid at very high
pressure through a 5um orifice.
Forced along defined micro channels,
Two streams of fluid to collide together at right
angles at a very high velocity.
The fluid collected of can be recycled through the
pump and interaction chamber until vesicles of the
spherical dimension are obtained.
Membrane extrusion

Size of liposomes is reduced by gently
passing them through polycarbonate
membrane filter of defined pore size
(100nm) at much lower pressure (<100psi)

To Increase Liposome Size
Freeze thawing
Direct reconstituted vesicles

Freeze-thawing
This method is based
upon freezing of SUV
then thawing by standing
at room temperature for
15mins.
Finally subjecting to a
brief sonication cycle.
It results in a high
proportion of large
unilamellar vesicles
formation.

Freeze SUVs
Thaw at
room temp.
for 15 min
Sonicate
SUVs
rupture
LUVs
Dried Reconstituted Vesicles
Empty SUVs
Hydrate with
entrapping material
Freeze drying
Rehydrate with
material to be
encapsulated
Uni or oligolamellar
vesicles
2. Solvent Dispersion Method:
Lipid dissolve in organic solvent

Excess addition of aqueous phase

Lipids align at interface of aqueous and organic layer

Formation of monolayer and bilayer of phospholipids

Liposome
Solvent
Dispersion
Method
solvent
injection (ether
and ethanol)
Double
emulsification
Reverse phase
evaporation
i. SOLVENT (Ether or Ethanol)
INJECTION TECHNIQUE
Ethanol injection
Lipid +ethanol

Rapid injection

Saline buffer + material to be
entrapped

SUVs
Ether injection
Lipid + ether

Slow injection

In aqueous phase (heat in
water bath 60C)

SUVs

Disadvantage:
Longer time
low efficiency



ii. Double Emulsification

iv. Reverse phase evaporation
method
Drawback: These conditions may possibly result in the
breakage of DNA strands or the denaturation of
some proteins.
3.Detergent Solubilization

The detergents at their critical micelles concentrations
have been used to solubilize lipids.


Phospholipid brought into intimate contact with aqueous phase
By addition optimized concentration of detergent
Formation of micelles (liposome)
Detergent is removed by following
methods:
i. dialysis
ii. column chromatography
ii. use of biobeads
ACTIVE LOADING TECHNIQUE
This technique employs the principle that certain type of
compounds with ionizable groups and those with both lipid and
water solubility can be introduced into the liposomes after the
formation of intact vesicle.

Loads drug molecules into preformed liposomes using pH
gradients and potential difference across cell membranes.

The transmembrane pH gradient can be developed using
various methods depending on nature of the drug to be
encapsulated.

2 step process generates the pH imbalance
and remote loading:
The vesicles are prepared in low pH
solution
It is followed by addition of base to the
extraliposomal medium
ADVANTAGES OVER THE
PASSIVE LOADING METHODS
A high encapsulation efficiency and
capacity.

A reduced leakage of the encapsulated
compounds.
REFERENCES
Drug Delivery Systems By Vasant V. Ranade, John B. Cannon. Third Edition.
Page no. 1 and 2 @ 2011 by Taylor and Francis Group, LLC.
(http://www.news-medical.net/health/What-is-a-Liposome.aspx) visited on
1st April, 2014 at 8:16pm. This article is licensed under the Creative
Commons Attribution-ShareAlike License.
(http://www.avantilipids.com/index.php?option=com_content&view=article
&id=1384&Itemid=372) visited on 1st April, 2014 at 8:50pm. Avanti Polar
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Comments.
By Jean R. Philippot, F. S. (1994). Liposomes as tools in basic research and
industry . The liposomes . Florida: CRC Press.
J.S. Dua1, P. A. (2012, april-june). International Journal of Pharmaceutical
Studies and Research. LIPOSOME: METHODS OF PREPARATION AND
APPLICATIONS . Jodhpur, Rajasthan,Nawanshahr, Panjab, India: IJPSR.
Liposome Drug Delivery: A Review By Chauhan Tikshdeep*, Arora Sonia,
Parashar Bharat and Chandel Abhishek. INTERNATIONAL JOURNAL OF
PHARMACEUTICAL AND CHEMICAL SCIENCES ISSN: 2277-5005
Liposome: classification, preparation, and applications. BY Abolfazl
Akbarzadeh, Rogaie Rezaei-Sadabady Soodabeh Davaran,Sang Woo Joo,
Nosratollah Zarghami,Younes Hanifehpour, Mohammad Samiei, Mohammad
Kouhi and Kazem Nejati-Koshki. Nanoscale Research Letters 2013
Akbarzadeh et al.; licensee Springer