seminar on liposomes in drug delivery

M.Pharm (Pharmaceutics) 2010-2011 Liposomes in Drug Delivery

Maliba Pharmacy College, Tarsadi

Seminar on

LIPOSOMES IN DRUG

DELIVERy

Presented By: - Dobariya Jayesh P.

M.Pharm Sem III (Pharmaceutics)

Roll no. 04,

Year : 2010-2011

Department of Pharmaceutics,

Maliba Pharmacy College, Tarsadi, Bardoli.



CONTENTS

1. Introduction

2. Structural components

3. Advantages and Disadvantages

4. Types of Liposomes

5. Preparation of Liposomes

- Handling of Liposomes

- Drying of liposomes

- Mechanism of liposomes preparation

- Methods of preparation

6. Characterization of Liposomes

7. Pharmacokinetics of Liposome encapsulated drugs

8. Pharmacodynamics of Liposome encapsulated drugs

9. Stability of Liposomes

10. Applications of Liposomes

11. Recent advances

12. Liposome products in Market as well as in clinical trials

13. Conclusion

14. References



Number of carriers was utilized to carry drug at the target organ/tissue which include

immunoglobulins, serum proteins, synthetic polymers, lipid vesicles (liposomes),

microspheres, erythrocytes, reversed micelles, niosomes, pharmacosomes, etc.

Among them, liposomes show strong potential for efficient drug delivery to the site of action.

Because they are - Biologically inert in nature,

- Devoid of any antigenic, pyrogenic or allergenic reactions

- Do not cause unfavorable side effects as well

Liposomes were first described by British haematologist Dr. Alec D Bangham in 1961

(published 1964), at the Babraham institute, Cambridge.

The name liposome is derived from two Greek words 'Lipid' meaning fat and 'Soma' meaning

body.

Definitions:

Bangham et al.,1965 : Simple microscopic vesicles in which an aqueous volume is

entirely enclosed by a membrane composed of lipid molecule.

Weiner N. et al.,1989 : As a microstructure consisting of one or more concentric

spheres of lipid bilayer separated by water or aqueous buffer compartments.

The drug molecules can encapsulated in aqueous space or intercalated into the lipid bilayer.

The particle size of liposomes ranges from 20 nm to 10 μm in diameter. Pharmaceutical

researchers use the tools of biophysics in evaluating liposomal dosage forms. Liposomes

have covered predominantly medical, albeit some non-medical areas like bioreactors,

catalysts, cosmetics and ecology.



Phospholipids and cholesterol are main components of liposomes.

PHOSPHOLIPIDS

Phospholipids are the major components of biological membranes, where two types of

phospholipids exist – phosphoglycerides and sphingolipids, together with their corresponding

hydrolysis products. The common phospholipid is phosphotidylcholine (PC) molecule. PC is

an amphipathic molecule in which a glycerol bridge links a pair of hydrophobic acyl

hydrocarbon chains, with a hydrophilic polar headgroup, phosphor choline.

Molecules of PC are not soluble in water and in aqueous media ,they align themselves closely

in planar bilayer sheets in order to minimize the unfavourable action between the bulk

aqueous phase and the long hydrocarbon fatty chain. Such unfavorable interactions are

completely eliminated when the sheets fold on themselves to form closed sealed vesicles. PC

molecule contrast markedly with other amphipathic molecule (detergents, lysolecithin) in that

they forms bilayer sheets but not micellar structures. This is thought to be because the

double fatty acid chain gives the molecule an overall tubular shape, more suitable for

aggregation in planar sheets compared with detergents with a polar head anf single chain

whose conical shape fits nicely into a spherical micellar structure



Lipid bilayer sheets Micellar

PC (or lecithin) can be derived from natural and synthetic sources. At various temperatures,

lecithin membranes can exist in different phases (phases are states such as solid gel state or

fluid liquid state). The transition from one phase to another can be detected by physical

techniques as the temperature is increased. At elevated temperature lipid membrane passes

from tightly ordered gel to a liquid crystal phase where freedom of movement of individual

molecule is higher. Most widely used method for determining the phase transition

temperature (Tc) is micro-calorimetry. In general, increasing chain length, or increasing the

saturation of the chains, increasing the transition temperature and also the stability of

molecule.

Some other commonly used phospholipids:-

Naturally occurring phospholipids:

- Phosphotidylcholine

- Phosphotidylethanolamine

- Phosphotidylserine

Synthetic phospholipids :

- DOPC : Dioleoyl phosphotidylcholine

- DSPC : Distearoyl phosphotidylcholine

- DOPE : Dioleoyl phosphotidylethanolamine

- DSPE : Distearoyl phosphotidylethanolamine

CHOLESTEROL

Cholesterol does not by itself form bilayer structure, but cholesterol can be incorporated in

very high concentration upto 1:1 or even 2:1 molar ratios of cholesterol to PC.

It acts as a fluidity buffer, i.e. below the phase transition temperature; it makes the membrane

less ordered and slightly more permeable; while above the phase transition temp. It makes the

membrane more ordered and stable.



Cholesterol inserts the membrane with its hydroxyl groups oriented towards the aqueous

surface and aliphatic chain aligned parallel to the acyl chians in the center of the bilayer.

Mechanism of cholesterol acting as a fluidity buffer:

It increases the separation between the choline head groups and eliminates the normal

electrostatic and hydrogen bonding interactions – thus pushing the phospholipids apart,

making layer less ordered at lower temperature.

However, in the higher concentrations that cholesterol is used, the membrane area occupied

by the combination of acyl chains and cholesterol is greater than that taken by

phosphocholine head group. This difference in area retards chain tilt.

Above the transition temperature, the reduction in the freedom of the acyl chains causes the

membrane to remain condensed and rigidized with a reduction in area through doser packing

and resultant decrease in fluidity.

Why use liposomes ?

Direction:

Liposomes can target a drug to the intended site of action in the body, thus enhancing its

therapeutic efficacy (drug targeting, site-specific delivery).

Liposomes may also direct a drug away from those body sites that are particularly sensitive to

the toxic action of it (site-avoidance delivery).

Duration:

Liposomes can act as a depot from which the entrapped compound is slowly released over

time. Such a sustained release process can be exploited to maintain therapeutic (but nontoxic)

drug levels in the bloodstream or at the local administration site for prolonged periods of

time.

Thus, an increased duration of action and a decreased frequency of administration are

beneficial consequences.



Protection:

Drugs incorporated in liposomes, in particular those entrapped in the aqueous interior, are

protected against the action of detrimental factors (e.g. degradative enzymes) present in the

host.

Conversely, the patient can be protected against detrimental toxic effects of drugs

Internalization:

Liposomes can interact with target cells in variousways and are therefore able to promote the

intracellular delivery of drug molecules that in their „free‟ form (i.e. non-encapsulated) would

not be able to enter the cellular interior due to unfavorable physicochemical characteristics

(e.g. DNA molecules).

Amplification:

If the drug is an antigen, liposomes can act as immunological adjuvant in vaccine

formulations.

Advantages of Liposomes

1. Liposomes are biocompatible, completely biodegradable, non-toxic, flexible and

nonimmunogenic for systemic and non-systemic administrations.

2. Liposomes supply both a lipophilic environment and aqueous “milieu interne” in one

system and are therefore suitable for delivery of hydrophobic, amphipathic and

hydrophilic drugs and agents.

3. Liposomes have the ability to protect their encapsulated drug from the external

environment and to act as sustained release depots (Propranolol, Cyclosporin).

4. Liposomes can be formulated as a suspension, as an aerosol, or in a semisolid form

such as gel, cream and lotion, as a dry vesicular powder (proliposome) for

reconstitution or they can be administered through most routes of administration

including ocular, pulmonary, nasal, oral, intramuscular, subcutaneous, topical and

intravenous.

5. Liposomes could encapsulate not only small molecules but also macromolecules like

superoxide dismutase, haemoglobin, erythropoietin, interleukin-2 and interferon-g.

6. Liposomes are reduced toxicity and increased stability of entrapped drug via

encapsulation. (Amphotericin B, Taxol).

7. Liposomes are increased efficacy and therapeutic index of drug (Actinomycin-D).

8. Liposomes help to reduce exposure of sensitive tissues to toxic drugs.

9. Alter the pharmacokinetic and pharmacodynamic property of drugs (reduced

elimination, increased circulation life time).

10. Flexibility to couple with site-specific ligands to achieve active targeting (Anticancer

and Antimicrobial drugs).

Disadvantages of liposomes

1. Production cost is high.

2. Leakage and fusion of encapsulated drug / molecules.

3. Sometimes phospholipid undergoes oxidation and hydrolysis like reaction.

4. Short half-life, Low solubility and Fewer stables.



1. Conventional liposomes

These can be defined as liposomes that are typically composed of only phospholipids (neutral

and/or negatively charged) and/or cholesterol. Most early work on liposomes as a drug-

carrier system employed this type of liposomes. Conventional liposomes are a family of

vesicular structures based on lipid bilayers surrounding aqueous compartments. Conventional

liposomes are characterized by a relatively short blood circulation time due to rapid uptake by

MPS system. They are useful for macrophage targeting, as local depot and for vaccination

purpose.

2. Long-circulating liposomes

The fast and efficient elimination of conventional liposomes from the circulation by liver and

spleen macrophages has seriously compromised their application for the treatment of the

wide range of diseases involving other tissues. The advent of new formulations of liposomes

that can persist for prolonged periods of time in the bloodstream led to a revival of interest in

liposomal delivery systems at the end of the 1980s. In fact, the long-circulating liposomes

opened a realm of new therapeutic opportunities that were up to then unrealistic because of

efficient MPS uptake of conventional liposomes. Perhaps the most important key feature of

long circulating liposomes is that they are able to extravasate at body sites where the

permeability of the vascular wall is increased. Fortunately, regions of increased capillary

permeability include pathological areas such as solid tumors and sites of infection and

inflammation.



It is illustrative for the importance of the long-circulation concept that the only two liposomal

anticancer products that are approved for human use are based on the use of long-circulating

liposomes for tumor-selective delivery of antitumor drugs (Doxil,DaunoXome).

At present the most popular way to produce long-circulating liposomes is to attach

hydrophilic polymer polyethylene glycol (PEG) covalently to the outer surface.

3. Immunoliposomes

Immunoliposomes have specific antibodies or antibody fragments (like Fab9 or single chain-

antibodies) on their surface to enhance target site binding. They are useful for site specific

targeting.

4. Cationic liposomes

These delivery systems are under development for improving the delivery of genetic material.

Their cationic lipid components interact with, and neutralize, the negatively-charged DNA,

thereby condensing the DNA into a more compact structure. The resulting lipid–DNA

complexes, rather than DNA encapsulated within liposomes, provide protection and promote

cellular internalization and expression of the condensed Plasmid.

5. Temp.-sensitive immunoliposomes

The heat induced drug release concept is based on the large increase in the permeability of

liposomal bilayers around their phase transition temperature. Local heating of tumor tissue

up to this phase transition temp. will enhance drug release from liposomes present in the

heated area. Both the degree of extravasation and the rate of drug release increases in this

case.

6. pH-sensitive immunoliposomes

pH sensitive IL targeted to internalizing receptors will end up in endosomes, where

acidification will trigger liposome destabilization and possible fusion with endosomal

membrane. They have been successfully applied in vitro for the delivery of antitumor drugs

into cytoplasm of tumor cells.

Lamella : Lamella is a thin flat plate like structure that appears during the formation of

liposomes. The phospholipid bilayer first exists as a lamella before getting converted into

spheres.

Several lamella of phospholipid bilayers are stacked one on top of the other during formation

of liposomes to form a multilamellar structure.



Handling of liposomes

In general, it is assumed that liposomes have a standard composition – egg lecithin :

cholesterol : phosphatidyl glycerol in molar ratio 0.9 : 1.0 : 0.1. This lipids can be stored

either as solids or in organic solution at -20ºC or at -70ºC in order to reduce the chances of

oxidation.

The solvent most widely used is a mixture of chloroform and methanol in a volume ratio of

2:1. Compounds which are sparingly soluble in either chloroform or methanol alone will

often dissolve readily in this 2:1 sovent mixture.

Solvent of the highest purity should be used, particularly since some contaminants may be

chemically reactive and may cause lipids to deteriorate. Ether degrades over time to form

peroxides, while chloroform gives rise to phosgene on standing. Formation of latter can be

prevented by addition of 1% ethanol to stabilize the chloroform and most commercial sourses

of chloroform are sold in this form.

All lipid solutions should be stored in dark, in glass vessels with a securely fastened ground

glass stopper. Polypropylene containers may also be used, although it is difficult to find caps

which fasten tightly enough to prevent evaporation of the solvents, which can take place

under refrigeration. Inert rubber (e.g. neoprene) can be used as a seat, but it does not tend to

swell in chloroform. In order to reduce the possibility of oxidation of lipids, nitrogen is most

commonly used. Since nitrogen is lighter than air, yet, a strong flow of gas is needed to

ensure complete exchange with air. The use of argon gas is preferable since this is heavier

than air and forms an effective blanket with just a very gentle stream of gas.

Drying of liposomes

Large volume of organic solution of lipids is most easily dried in a rotary evaporator fitted

with a cooling coil and a thermostatistically controlled water bath. Rapid evaporation of

solvent is carried out by gentle warming (20 ºC - 40 ºC) under reduced pressure (400 – 700

mmHg). Rapid rotation of the solvent containing flask increases the surface area for

evaporation.

To remove last traces of chloroform, attachment of flask to the manifold of lyophilizer, and

overnight exposure to high vacuum is a good method.

General methods of preparation of liposomes

All methods include three or four basic stages:-

1. Drying down lipids from organic solvent,

2. Dispersion of lipids in aqueous media,

3. Purification of resultant liposomes, and

4. Analysis of final product.



Mechanism of Liposome Preparation:

The budding theory

- Stress induced hydration of phospholipids.

- Organization in to lamellar arrays.

- Results in to budding of lipid bilayer leading to down sizing.

The bilayer phospholipids theory.

- Liposomes (lipid vesicles) are formed when thin lipid films or lipid cakes are

hydrated and stacks of liquid crystalline bilayers become fluid and swell.

- The hydrated lipid sheets detach during agitation and self-close to form large,

multilamellar vesicles (LMV).



- Once these particles have formed, reducing the size of the particle requires energy

input in the form of sonic energy (sonication) or mechanical energy (extrusion).

Figure: - Mechanism of vesicle formation

MECHANICAL DISPERSION METHODS FOR PASSIVE LOADING:

Lipid hydration method

In this method, lipid mixture are dissolved in solvent mixture of chloroform : methanol (2:1)

in rotary evaporator flask and dried thin film of lipid is made using rotary evaporator under

reduced pressure (60 rpm, 30ºC, and about 15 min).



Flask is flushed with nitrogen and Hydration of lipid is done by adding 5ml of saline

phosphate buffer containing drug/solute to be encapsulated and again use of rotary evaporator

for making homogeneous milky white suspension.

It is allowed to stand for 2 hr at RT/above Tc for complete swelling process. This will give

MLVs.

Sonication method

At high energy level, preformed MLVs are sonicated using either probe or bath ultrasonic

disintegrator.

Using Probe:

Used for suspensions which require high energy in a small volume. And contamination of

preparation with metal can lead to degradation of lipid.

Using Bath:

Used for large volume of dilute lipids where may not necessary to reach the vesicle size limit.

Finally, they are purified into the SUVs by ultracentrifugation and collected from supernant

of centrifuge tube.

Size of liposome is influenced by temperature, composition, and concentration, sonication

time & power, volume of product.



Microfluidization / Microemulsification method

In this method, Microfluidizer pumps the fluid at very high pressure through a 5µm screen.

Then, it is forced along defined micro channels which direct two streams of fluid to collide

together at right angles at a very high velocity, thereby effecting a very efficient transfer of

energy.

The lipid can be introduced into the fluidizer, either as a suspension of large MLVs, or as a

slurry of unhydrated lipid in a organic medium. The fluid collected can be recycled through

the pump and interaction chamber until vesicles of the spherical dimension are obtained.

Advantages: Excellent size reduction upto 0.2mm, High rate of production, for encapsulation

of water soluble materials due to high proportion of lipid.

French pressure cell method

In this method, liquid sample of preformed MLVs are introduced into the sample cavity, then

the position of piston and pressure is set up to fill sample upto the outlet hole. Then power is

switched on. At high pressure (2000 psi) and at 40ºC, MLVs are extruded through small

orifice, which is collected in suitable container.



This technique yields uni- or oligo lamellar liposome of intermediate size.

More stable than they obtained by sonication method and also leakage of the content from the

liposomes are lesser.

Drawback: High cost of the pressure cell.

Membrane extrusion method

Size of prepared liposomes is reduced by gentley passing them through membrane filter of

defined pore size and this can be achieved at much lower pressure.

In this process, the vesicles content are extruded with the dispersion medium during breaking

and resealing of phospholipids as they pass through the polycarbonate membrane in order to

achieve high entrapment.

The liposomes produced by this method have been termed as LUVETs and 30%

encapsulation can be obtained using high lipid concentration.

Dried reconstituted vesicles



It starts with freeze drying of a dispersion of empty SUVs and rehydrating it with the aqueous

fluid containing the materials to be entapped. This leads to dispersion of solid lipids in finely

subdivided form.

Freeze drying is used to freeze and lyophilize the preformed SUVs dispersion rather than to

dry the lipids from an organic solution. This leads to organized membrane structure which on

addition of water can rehydrate, fuse and reseal to form vesicle with high capture capacity.

It is used for manufacturing of uni - or oligo lamellar of the order of 1.0µm or less in

diameter.

Advantages: high entrapment of water soluble content and use of mild condition for

preparation & loading of bioactive.

Freeze thaw sonication method

This method is based on freezing of unilamellar dispersion and thawing (melting) by standing

at RT for 15 min. and finally subjected to a sonication cycle.

This process ruptures and refuses SUVs during which the solute equilibrates between inside

and outside, and liposomes themselves fuse and markedly increase in size.

The second step of the sonication considerably reduces the permeability of the liposome

membrane, by accelerating the rate at which the packing defects are eliminated.

For producing giant vesicles of diameter having 10 – 50 µm, the sonication step is replaced

by the dialysis against hypo-osmolar buffer. In this case, SUVs are mixed with salt solution

followed by freeze thawing. During this dialysis, the large vesicles formed by freeze thawing

swell and rupture as a result of the osmotic lysis, where the fuse and prepare as giant vesicles.

Disadvantage:

- Lesser encapsulation efficiency,

- Presence of charge particle for the formation of ice crystal to aid in the rupture or

fusion process, so neutral liposomes can not be resulted.

Advantage:

- simple, rapid, result in proportion of large unilamellar vesicles formation.



SOLVENT DISPERSION METHOD:

Ethanol injection method

Ethanol is used to dissolve the lipids and solution is rapidly injected through a fine needle

into an excess of buffer solution.

SUVs form spontaneously. Method is restricted to the production of relatively dilute SUVs

suspension.

Removal of residual ethanol is also present a problem. This can be done by ultrafilteration or

vacuum distillation

Ether infusion method

In this method, solution of lipids in diethyl ether or ether : methanol mixture is slowly

injected to aqueous solution of materials to be encapsulated at 55 - 65ºC. Subsequent removal

of ether under vacuum leads to the formation of liposomes.

Drawbacks: Heterogeneous size (70 - 190µm), exposure of compounds to organic solvents

or high temperature.

Double emulsion vesicles

When organic solution which already contain water droplet, is introduced into excess

aqueous phase followed by mechanical dispersion, multi compartment vesicles are obtained.

The ordered dispersion so obtained is desirable as a w/o/w system. The vesicles with aqueous

core are suspended in aqueous medium. So two aqueous compartments being separated from

each other by pair of phospholipids monolayer whose hydrophobic surface face each other

across a thin film of organic solvent. Removal of this solvent clearly results in intermediate

sized unilamellar vesicle. The theoretical entrapment may reach up to 90%.



Reversed phase evaporation vesicles

The essential feature of this method is the removal of solvent from emulsion by evaporation.

In this method, lipids dissolved in organic solvents are sonicated by bath sonication which

form emulsion (w/o) and then emulsion is dried down to a semi solid gel using rotary

evaporator under reduced pressure.

The next step is to bing about the collapse of a certain proportion of water droplets by

vigorous mechanical shaking with a vortex mixer. This will give LUVs.

Encapsulation percentage: upto 50%

Stable plurilamellar vesicles

In this method, w/o dispersion is prepared as described in REV method with excess lipid, but

drying process is accompanied by continued bath sonication with a stream of nitrogen. The

redistribution and equilibration of aqueous solvent and solute occur during this time in

between the various bilayer in each plurilamellar vesicle.

Entrapment percentage: 30%.

DETERGENT SOLUBILIZATION FOR PASSIVE LOADING:-

In this method, the phospholipids are brought into intimate contact with the aqueous phase

via the intermediary of detergents, which associate with phospholipid molecules and serve to

screen the hydrophobic portions of the molecule from water.

The detergent depletion method is used for preparation of a variety of liposomes and

proteoliposome formulations.

Detergents can be depleted from a mixed detergent-lipid micelles by various

techniques which leads to the formation of very homogeneous liposomes.

The most popular detergent is sodium cholate, alkyl(thio)glucoside, and

alkyloxypolyethylenes.

The use of different detergents results in different size distributions of the vesicles

formed.



A faster depletion rate produces smaller size liposomes.

The use of different detergents also results in different ratios of large unilamellar

vesicles/ oligolamellar vesicles/multilamellar vesicles.

Detergent depletion is achieved by four following approaches:

A. Dialysis: The dialysis can be preformed in dialysis bags immersed in large detergent

free buffers (equilibrium dialysis) or by using continuous flow cells, diafiltration and

cross filtration.

B. Gel filtration: In this method the detergent is depleted by size exclusive

chromatography. Sephadex G-50, Sephadex G-100, Sepharose 2B-6B and Sephacryl

S200-S1000 can be used for gel filtration. The liposomes do not penetrate into the

pores of the beads packed in a column.

C. Adsorption using biobeads: Detergent adsorption is achieved by shaking of mixed

micelle solution with beaded organic polystyrene adsorbers such as XAD-2 beads and

Bio-beads SM2. The great advantage of the using detergent adsorbers is that they can

remove detergents with a very low critical micelle concentration (CMC) which are not

completely depleted by dialysis or gel filtration methods.

D. Dilution: Upon dilution of aqueous mixed micellar solution of detergent and

phospholipids with buffer the micellar size and the polydispersity increases

dramatically, and, as the system is diluted beyond the mixed micellar phase

boundary, a spontaneous transition from polydisperse micelles to monodisperse

vesicles occurs.

ACTIVE (REMOTE) LOADING TECHNIQUE:

Certain types of drugs with ionisable groups and those with both lipid and water solubility

can be introduced into liposomes after the formation of the intact vesicles. Drug is loaded into

the preformed liposomes using pH gradient and potential difference across liposomal

membrane.

Approach for remote loading :

Vesicles are prepared in low pH solution, thus generating low pH within liposome interior

followed by addition of the base to external mediun of liposomes. Basic compounds with

amino group are relatively lipophilic at high pH and hydrophilic at low pH. Unprotonated

form of basic drug can diffuse through the bilayer. At the low pH side, the molecules are

predominantly protonated, which lower the concentration of the drug in the unprotonated

form. Dynamic equilibrium promotes the diffusion of more drug molecules at low pH side of

the bilayer. Exchange of external medium by gel-exclusion chromatography with a neutral

solution to remove remaining basic compound.

Advantages:

High encapsulation efficiency and capacity.

Reduced leakage of the encapsulated materials.



„Bed side‟ loading of the drugs, thus limiting loss of retention of drugs by diffusion or

chemical degradation during storage.

Flexibility for the use of constitutive lipids, as drug is loaded after the preparation of

carrier units.

Avoidance of biological reactive compounds during preparation steps in the

dispersion thus reducing safety hazards.

Weak base such as doxorubicin, adriamycin, vincristine and short modified peptides &

insulin have been successfully encapsulated by remote loading method.

The behaviour of liposomes in both physical and biological systems is governed by the

factors such as physical size, membrane permeability, percent entrapped solutes, chemical

composition as well as the quantity and purity of the starting materials. Therefore, the

liposomes are characterized for physical attributes i.e shape, size and its distribution,

percentage drug capture, entrapped volume, lamellarity, percentage drug release and

chemical composition (estimation of phospholipids, phospholipids oxidation and analysis of

cholesterol).

Physical Characterization

Characterization parameters Analytical method/Instrument

1 Vesicle shape and surface

morphology

Transmission electron microscopy,

Freeze-fracture electron microscopy

2 Mean vesicle size and size

distribution

(submicron and micron range)

Dynamic light scattering, zetasizer,

Photon correlation spectroscopy, laser light scattering,

gel permeation and gel exclusion

3 Surface charge Free-flow electrophoresis

4. Electrical surface potential and

surface pH

Zetapotential measurements & pH sensitive probes

5 Lamellarity Small angle X-ray scattering, 31

P-NMR, Freeze-

fracture electron microscopy

6 Phase behaviour Freeze-fracture electron microscopy, Differential

scanning colorimetery

7 Percent of free drug/ percent

capture

Minicolumn centrifugation, ion-exchange

chromatography, radiolabelling

8 Drug release Diffusion cell/ dialysis

Chemical Characterization

1 Phospholipid concentration Barlett assay, Stewart assay, HPLC



2 Cholesterol concentration Cholesterol oxidase assay and HPLC

3 Phopholipid peroxidation UV absorbance, Iodometric and GLC

4 Phospholipid hydrolysis,

Cholesterol auto-oxidation.

HPLC and TLC

5 Osmolarity Osmometer

Biological Characterization

1 Sterility Aerobic or anaerobic cultures

2 Pyrogenicity Limulus Amebocyte Lysate (LAL) test

3 Animal toxicity Monitoring survival rates, histology and pathology

These systems are designed to control following parameters:

1. The rate of input of drug into particular body compartment.

2. The distribution & localization of drug in to body.

3. The persistence or rate of metabolism of drug.

Liposomes generally perform some of same functions polymeric controlled release systems

i.e. they can act as a drug reservoir with an output controlled and limited by the permeability

characteristics of the liposome membrane. An important difference is that, at present, the

release rate of drug from liposomes is much faster than that from polymeric systems. Thus,

liposomes can only act as sustained release preparations for hours to days as compared with

days to months for polymeric devices.

Clearance and distribution of liposome in vivo:-

Two major determinants of liposome clearance: - Vesicle size & Surface size

Vesicle size:

SUVs persist in the circulation for longer periods than large MLVs of same

composition.

In addition, the clearance rate of liposome samples, homogeneous in size, can be

described by a simple exponential function, whereas the clearance of heterogeneous

samples can only be fitted by sum of exponentials, suggesting that liposomes are

cleared by a single type of process and that this process is size dependent.

Surface charge:

SUVs with ve+ and ve– charge retained in circulation for long periods, whereas small

negative vesicles are rapidly cleared.



After clearance from circulation, they are sequestered in various tissues and organs.

MLVs :- liver & spleen are primary sites of uptake (due to presence of phagocytic

reticuloendothelial cells and blood flow is through open sinusoids rather than through

capillaries in these organs).

Also preferentially retained in lung (due to physical entrapment of liposomes in the

capillary beds of this organ).

SUVs :- broader tissue distribution, however, in liver & spleen also.

Pharmacodynamic effects:-

Retardation of drug clearance from the circulation.

High drug accumulation in tissues rich in reticuloendothelial cells, especially liver and

spleen.

Retention of drug in tissue for large periods.

Protection of drug against metabolic degradation and elimination.

Localized drug delivery primarily for cancer therapy

Chemical degradation:-

It includes mainly liposomal phospholipid oxidation and hydrolysis.

Prevention:-

1. Start with freshly purified lipids & freshly distilled solvents.

2. Avoid procedure which involves high temperature.

3. Carry out manufacturing in absence of oxygen.

4. Deoxygenate aqueous solution with nitrogen.

5. Store all liposome suspensions in inert atmosphere.

6. Include anti-oxidants as a component of the lipid membrane.

Apart from these, saturated lipids reduce level of oxidizable lipid in membrane.



The hydrolysis may be avoided altogether by the use of lipids which contain either instead of

ester linkage such as found in membrane of halophilic bacteria. Hydrolysis in vivo as a result

of enzymatic attack can be prevented by the use of sphingomyelin, or phospholipid

derivatives with the 2-ester linkage replaced by a carbomyloxy function.

Physical degradation:-

It mainly includes leakage and fusion of vesicles.

Prevention:-

1. SUVs (prone to fusion) is stored at temperature away from the Tc.

2. Avoid high conc. of metal ions for liposome having negative charge in the membrane

and use of metal ion chelater in the suspending buffer.

3. High molar ratio of cholesterol is most stable with regard to leakage of solute for

large polar or ionic molecule and low MW lipophilic compound.

Freezing / lyophillization / cryopreservation – most suitable method to prevent degradation.

Liposomes in AIDS Therapy

Phillips and Tsoukas were the first to demonstrate the liposomes as carrier for the delivery of

anti-HIV drug AZT Azidothymidine and reported the decreased hemotopoitic toxicity and

enhanced activity against Murine Acquired Immunodeficiency Syndrome of Azidothymidine

by encapsulation in liposomes.

Desormeaux et al. demonstrated the accumulation of free and liposome-encapsulated ddI (2‟,

3‟ dideoxyinosine) in murine monocyte-macrophage RAW264.7 cells and human

premonocytoid U937 cells. The results of this study suggested that encapsulation of ddI in

liposomes modified the tissue distribution and plasma pharmacokinetic of the antiviral agent.

Smith et al. in their study established that the incorporation of neutralizing agents in anti-

HLA-DR immunoliposome could represent a novel therapeutic strategy to specifically target

cell free HIV particles and virally infected cells to treat HIV infection more efficiently.

Duzgunes et al. reported that intracellular delivery of novel macromolecular drugs against

human immunodeficiency virus type -1 (HIV-1), inducing antisense oligodeoxynucleotides,

ribozymes and therapeutic genes, may be achieved by encapsulation in or association with

certain types of liposomes. An HIV-1 protease inhibitor encapsulated in conventional

negatively charged multilamellar liposomes was about 10-fold more effective and had a



lower EC90 than the free drug in inhibiting HIV-1 production in human monocyte-derived

macrophages.

In a recent study Mareuli et al. compared the antiviral effects of free-SPC3 and liposome-

associated SPC3 in cultured human lymphocytes infected with HIV-1. SPC3 is a potent

antiviral drug, which blocks lymphocyte and macrophage infections with various HIV strains

in vitro. Liposomal entrapment was found to increase the antiviral efficacy of SPC3 by more

than 10-fold and 5-fold in C8166 and PBLs, respectively so data of present study suggest that

the liposome approach may be used successfully to improve SPC3 antiviral efficacy.

Pretzer et al. examined the effect of free and liposome encapsulated protease inhibitor L-

689502, on virus production by monocyte-derived macrophages infected with HIV-1BaL.

Continuous treatment with L-689502 drastically inhibited virus production in a dose

dependent manner in the range of 10-200 Nm, in some cases by more than 1000-fold,

compared to untreated cells. Since liposomes can be targeted to macrophages in vivo. The

inhibitor encapsulated in multilamellar liposomes was more effective than the free drug in

inhibiting virus production macrophages, throughout the concentration range studied. These

studies indicate that liposomes can be used to facilitate the intracellular delivery of certain

anti-HIV agents and to enhance their therapeutic effects.

Liposomes in Cancer Therapy

Colbern et al. reported the antitumor activity of the combination of Herceptin and

nonliposomal cisplatin or stealth liposomal cisplatin in two xenograft tumor models, initiated

from the cell lines, BT474 and MDA453, that overxpress the oncogene, HER2. Herceptin

alone had significant antitumor activity in all three experiments. Nonliposomal cisplatin and

stealth liposomal cisplatin were both effective antitumor agents but, at tolerable dose levels,

stealth lipsomal cisplatin was reported superior to nonliposomal cisplatin.

Hamilton et al. performed a phase I study doxorubicin liposomes (Caelyx, Doxil) using a

prolonged (6-week) dose interval to reduce the incidence of skin toxicity that was dose-

limiting at more conventional dose intervals and which appeared to be schedule dependent.

Metastatic breast cancer patients who had received a maximum of one prior therapy for

metastatic disease were administered defined dose levels of 60, 70, 80 and 90 mg/m2. Severe

skin toxicity was not observed at the 60 mg/m2 dose level, and occurred in only one patient

treated at 70 mg/m2.

Mukhopadhyay et al. developed conjugate of antineoplastic drug daunomycin (DNM) with

maleylated bovine serum albumin. It was taken up with high efficiency by multi drug

resistant variant JD100 of the murine - macrophage tumor cell line J774A.1 through the

scavenger receptors resulting in cessation of DNA synthesis.

A thermosensitive liposomal taxol formulation (heat mediated targeted drug delivery) in

murine melanoma was developed and studied by another group of workers. Cremophor

which is used as excipient due to the low aqueous solubility of taxol has toxic side effects.

Temperature sensitive liposomes encapsulating taxol were prepared using egg

phosphatidylcholine and cholesterol in combination with ethanol. A significant reduction in

tumor volume was noted in tumour bearing mice treated with a combination of hyperthermia

and theromosensitive liposome encapsulated taxol, compared to animals treated with free

http://www.pharmainfo.net/keywords/targeted-drug-delivery-systems



taxol with or without hyperthermia in B16F 10 murine melanoma transplanted into C57BI/6

mice.

Sharma et al. also investigated the use of polyvinylpyrrolidone liposomes containing taxol

prepared by reverse micro-emulsion method. The size of liposomes was found to be 50-60

nm. The antitumor effect of taxol was evaluated in B16F10 murine melanoma transplanted in

C57 B 1/6 mice. In vivo efficacy of taxol containing liposomes as measured by reduction in

tumor volume and increased survival time was significantly greater than that of an equivalent

concentration of free taxol.

Liposomes in Malaria Therapy

Pirson et al. reported use of liposomes for safe and effective delivery of primaquine. Liver

and spleen accumulation of labeled primaquine in negative charged liposomes was found

more as compared to its accumulation in lungs, kidneys, heart and brain, which lead to its

diminished toxicity as compared to free labeled primaquine. The uptake of liposomes

entrapped drug was gradual, reaching a plateau of 60 % of initial load after 20 minutes of

perfusion.

Peeter et al. examined the maximum permissible dose of chloroquine per intraperitonial

injection was 0.8 and 10 mg for chloroquine and liposomal chloroquine. An increase in

therapeutic and prophylactic efficacy of liposomal chloroquine in comparison with free

chloroquine at a 0.8 mg chloroquine dose level was found. It was possible to obtain 100 %

efficacy with one single intraperitonial injection of 6 mg liposomal chloroquine. Moreover,

the stability to increase the doses of chloroquine per injection after liposome encapsulation

allowed successful treatment of infections with chloroquine resistant P. berghei, which could

not be cured by a 7-day course with the maximum tolerable dose of free chloroquine of 0.8

mg/mouse/day.

Bayomi et al. demonstrated that arteether was successfully administered intravenously in

liposomal formulations. These had shown longer elimination half-life with respected to other

artemisinin derivatives. Also, an optimal oral liposomal formulation for arteether was found,

which was absorbed fastly and completely from GI tract. The liposomes were found suitable

for three months storage and entrapment efficiency of 100 %.

Moll et al. studied the effect of Trp-N-formylated gramicidin and gramicidin A incorporation

in liposomes on the growth of P. falciparum in an in vitro culture. Incorporation of Trp-N-

formylated gramicidin in the membranes of s0-called „stealth‟ vesicles strongly decrease the

concentration needed to induce 50 % inhibition of parasite growth. Trp-N-formylated

gramicidin incorporation in “stealth” vesicles ends up specifically in the infected cell, thereby

targeting and inhibiting the growth of the malaria parasite.

Liposome in lung therapy

Labana et al. reported co-administartion of isoniazid and rifampicin encapsulated in lung

specific stalth lipsomes at one tghird of their recommended doses of 12 and 10 mg/kg

respectively, exhibited a sustained release of these drugs in plasma (5 days) and lungs, liver

and spleen (7 days). Chemotherapeutic efficacy of once weekly administered liposomal drugs

for 6 weeks reduced the mycobacterial load significantly in lungs, liver and spllen of infected

mice compared with untreated animals.



Konduri et al. performed comparative study of budesonide encapsulated in liposomes with

free budesonide therapy in reducing allergic inflammation. Weekly therapy with budesonide

encapsulated in liposomes was found as effective as daily budesonide therapy in decreasing

lung inflammation and lowering eosinophil peroxidase activity, peripheral blood eosinophils

and total serum IgE levels. This novel strategy offers an effective alterative to standard daily

budesonide therapy in asthma and has the potential to reduce toxicity and improve

compliance.

In Chandigarh lung specific liposomes was developed and investigated in animal models of

tuberculosis. Liposomes tagged with O. stearlylamylopectin (O-SAP) resulted in increased

affinity towards lung tissue of mice. Liposomes containing egg phosphatidylcholine

cholesterol dicetylphosphate, O-SAP, monosialo-ganglioside (GMI)/DSPE PEG 2000 were

found to be more stable in serum. These liposomes accumulated more in lungs than in

reticulo endothelial system of normal and tuberculosis mice.

Liposomes in Infectious Diseases

Bacchawat and coworkers developed liposomal amphotericin and investigated it in animal

models of fungal infection and leishmaniasis. Kshirsagar and coworkers modified the

formulation, developed a “Patient Worthy” sterile pyrogen free liposomal amphotericin

preparation and investigated it in patients with systemic fungal infections and leishmaniasis.

It was found to be safe producing significantly less adverse effects compared to plain

amphotericin in patients with systemic fungal infection, did not produce nephrotoxicity and

could be given to patients with renal damage. It was effective in patients resistant to

fluconazole and plain amphotericin. Unlike Ambisome (USA) which needs to be used in dose

of 3 mg/kg/ day this is effective at 1 mg/kg/day dose. The same group studied different

dosage regimens of liposomal amphotericin using Aspergillus murine model. It was found

that liposomal amphotericin was more effective than equal dose of free amphotericin B given

after fungal spore challenge. A large single dose of liposomal amphotericin was more

effective, whether given before or after spore challenge, than given as two divided doses.

It was investigated in patients with visceral leishmaniasis and found to be effective in patients

who had not responded to antimony, pentamidine and amphotericin. Because of its safety, it

can be given at 3 mg/kg/day dose thus reducing total duration of treatment. It was

successfully used in a child suffering from visceral leishmaniasis. This is the first liposomal

preparation developed outside of USA, which has been used in patients.

In an attempt to improve efficacy and reduce toxicity further, liposomes with grafted ligand

have been developed. Pentamidine isethionate and its methoxy derivative were encapsulated

in sugar grafted liposomes and tested against experimental leishmaniasis in vivo. It was seen

that sugar grafted liposomes specially the mannose grafted ones were potent in comparison to

normal liposome encapsulated drug or free drug.

Liposomes in Dermatology and Cosmetology

The advantage of the liposomal form over the conventional dermatological form was

particularly striking when the activity of local anesthetic agents was evaluated in cream,

ointment or lotion form versus liposomal form. A liposomal product containing 0.5 %

tetracaine produced more intensive activity (6-8 folds) than 1.0 % tetracaine in cream form,

which was a commercial preparation, i.e. pentocaine cream. Similar results were obtained



with liposomal forms of other local anesthetic agents, e.g. lidocaine, dibucaine and

benzocaine.

Graysan et al. tested gentamycin encapsulating multivesicular liposomes (DepoForm) as a

prophylactic anti-infective treatment for surgical wounds. The liposomal formulation was

injected subcutaneously to provide local depots and was challenged 48 hr later by a bacterial

inoculum, also injected subcutaneously to the same location. Evaluation of bioburden

reduction 48 hr later showed that the liposomal formulation was significantly superior to

empty liposomes and saline.

Brown et al. have tested liposomes as a growth factor delivery system for the topical

treatment of incisional wounds in mice. An interesting dual carrier system was devised in

which insulin, serving as an intra liposomal carrier was first complexed with EGF (Epidermal

growth factor), then the insulin-EGF complex was encapsulated within liposomes. The

liposomal formulation provided a transient increase in tensile strength, compared with empty

loposomes or saline. Combined with exceptionally high intraliposomal retention of the EGF

reported in this study, it could be that the internal carrier induced a sustained release that was

too slow for effective therapy.

Margalit et al. examined the response of infected wounds to treatment with cefazolin-

encapsulating bioadhesive liposomes were studied in full thickness wounds in mice infected

with staphylococcus aureus. After 3 days treatment, the wound bacterial counts after 3 days

treatment with 1 mg of cefazolin in the bioadhesive liposomes (80 % encapsulated) were

down 100-fold from untreated controls, to the colonization – infection boundary. Although all

these studies can be taken as encouragement for exploring liposomes as drug delivery

systems for topical treatment of wounds and burns.

According the patent literature almost every kind of active ingredient might be suitable to be

encapsulated in topical liposomes (DermosomeTM

, InovitaTM

). However, among the great

variety of candidates, e.g. antibiotics, antifungal, disinfectant, immunosuppressive agents,

several hydrophilic and hydrophobic peptides are available in market as liposomal

preparation. Liposomes based anti-ageing topical formulations (creams, lotions, gels and

hydrogels) have been formulated launched in cosmetic market in 1986 by L‟Oreal in the form

of niosomes and then by Christian Dior in the form of liposomes (CaptureTM

). Currently,

various liposome based formulations for facial and body care, make-up, mascara and

foundations as well as hair care, self tanning and sun screen products and even perfumes are

being launched in cosmetic market.

Liposome as an immunological (Vaccine) Adjuvant

New generation vaccines that are based on recombinant protein subunits and synthetic-

peptide antigens are usually non-immunogenic hence the need of immuno-potentiation is well

realized. Although many structurally unrelated agents are capable of inducing immune

responses to vaccine antigens, most of them are toxic. Apart from alum, which is the only

immunological adjuvant used for last few decades, others are not clinically useful. After

being established the immunoadjuvant properties of liposomes, several liposome based

vaccines have been either approved or licensed for use in human. Vaccines based on

novasomes have been licensed for the immunization of fowl against Newcastle diseases virus

and avian retrovirus56

. However, the first liposome based vaccine (against hepatitis A) that

has been licensed for use in human is an IRIV vaccine produced by Swiss serum and vaccine



institute, Switzerland known as “Epaxal Berna Vaccine” is an IRIV vaccine. IRIVs are

spherical unilamellar vesicles with a mean diameter of -150 nm. IRIVs are prepared by

detergent removal of influenza surface glycoprotein and a mixture of natural and synthetic

phospholipids containing 70 % egg yolk phosphatidylcholine, 20 % synthetic PE and 10 %

envelope phospholipids originating from HINI influenza virus.

Fusogenic liposomes are specially engineered liposomes that fuse and merge with cell

membranes and directly introduce molecules into cytoplasm thus avoiding the route followed

by conventional liposomes. The fusogenic liposomes mimic the way by which several viruses

(HIV, Sendai virus) bind and merge with cell membrane at neutral pH and subsequently

release their genome into cytoplasm. Fusion spike glycoprotein of Sendai virus, rabies virus,

meales virus, influenza virus, herpes virus, HIV-1 and vesicular stomatitis virus have been

incorporated in liposome and these virosomes have been investigated for their

imunoadjuvant, gene and oligonucleotide delivery potentials.

Very recently, De Jonge et al. demonstrated that small-interfering RNA (siRNA),

encapsulated in virosomes, are able to downregulate the synthesis of newly induced and

consitutively expressed proteins, overcoming the lack of suitable delivery methods for these

molecules. Moreover the authors show that intraperitoneal injection of siRNA loaded

virosomes resulted in delivery of the nucleotides to cell in the peritoneal cavity.

Immunoassays such as ELISA, RIA or LILA are based on the selective interaction of an

analyte antigen with corresponding antibodies. Liposomes can be useful analytical reagents

as they can encapsulate upto a million mrkers and can therefore, serve as signal amplifiers.

Liposome is well recognized as a model membrane and its lytic ability inculcated through

various lytic agents such as complement led to the development of an efficient assay system

known as “liposome innume lysis assay (LILA)”. The marker could be a fluorescent dye such

as carboxyfluorescein, calcein or some markers that could be monitored enzymatically. LILA

assay has been implicated in the detection of serum components such as α-fetoprotein,

carcinoembryonic antigen, C-reactive protein and other serum proteins, which serve as

diagnostic tools especially for cancer.

Liposome as Radiodiagnostic Carriers

Liposomes are used in different imaging modalities to locate the sites specifically. Their

radiodioagnostic applications include liver, brain and spleen imaging, lymphatic imaging,

tumor imaging, blood pool imaging, visualization of inflammation and infection sites,

visualization of bone marrow and eye vasculature and imaging cardiovascular pathologies.

Liposome based imaging agents have already been successfully used for γ-scintigraphy,

magnetic resonance (MR), computer tomography (CT) and ultrasonography (US) of tumours. 111

In-labelled liposomes for tumour imaging (VenCan®, Vester) are already in phase II-III

clinical trials. Liposomal uptake by reticuloendothelial system (RES), which is useful

strategy in localization of contrast agents in RES-rich organs like liver, spleen and bone

marrow but it is not useful for localization to non-RES organs.



Miscellenous Application

Nabar et al. studied the effect of size and charge of liposome in the bio-distribution of 99m

TC-DTPA encapsulated in liposome after intravenous injection in rats. They observed that

multilamellar vesicles (MLV) were taken up to a greater extent as compared to SUVs in liver

spleen and lungs. Positively charged MLVs than negative or neutral ones, were taken up

more in liver, positively charged SUVs were taken up more in kidneys and neutral MLVs

were taken up more in lungs than charged ones. An attempt was made to improve stability of

liposome by coupling the drug with the lipid bilayer using a cross linking agent. Soya

phosphatidylcholine (SPC) containing liposomes were prepared by calcium induced fusion

method. Positively charged stearylamine was introduced in the bilayer. The liposomes were

coupled to entrapped ibuprofen by EDAC (1-ethyl 3-(3-dimethyl aminopropyl) carbodiimide

HCI) and the coupling was confirmed by UV spectrum. It was observed that EDAC in SPC

containing stearylamine liposomes retarded the release of ibuprofen significantly.

In albino rats, the various factors affecting systemic absorption of nasally applied gentamycin

sulphate using in situ nasal perfusion technique was studied by Martin et al. Tween 80 which

is a surfactant increases permeation by altering membrane structure and permeability. In this

study Tween 80 upto 1% W/V concentrations, increased permeability. Betacyclodextrin at

0.25% W/V concentration, another permeability enhancer was found to significantly increase

permeability initially but was found to plateau off later on. However both these permeability

enhancer were found to decrease stability and potency of gentamycin.

Jain et al. developed dopamine hydrochloride bearing positively charged small liposomes by

sonicating multilamellar vesicles and studied their physical attributes and drug leakage and

release pattern. In vivo performance was assessed by periodic measurement of

chlorpromazine induced catatonia in Sprague Dawley rats and was compared with plain

dopamine hydrochloride, dopamine levodopa and carbidopa. The studies showed that

dopamine can be effectively delivered into the brain and its degradation in circulation can be

prevented by incorporating it into liposomes.

Tsuchida et al. reported that the aggregation and fusion of hemoglobin vesicles (Hb-vesicles)

and the leakage on long-term storage can be prevented by using either polymerized

phospholipids or polyphospholipids or by introduction of oligosaccharides type of glycolipid

in the bilayer membrane. Recent studies suggest that the sterically stabilized liposome

bearing haemoglobin (PEG-PE-LEH) are even better than LEH as artificial blood substitutes

as they manifest less toxicity, less platelet and aggregation and less haemoststic generation.

1. Provesicles in drug delivery systems

To overcome the limitations (especially chemical and physical stability) of vesicular drug

delivery systems like liposomes, niosomes, transferosomes, and pharmacosomes,

provesicular approach was introduced.



This includes-

a. Proliposomes

Proliposomes are the products which are mixed with water phase containing drug

before use, liposomes formed automatically and load the drug.

Three different types of proliposomes are formulated.

b. Dry granular liposomes

Dry, free flowing granular product, which can be hydrated immediately before use.

Composed of water soluble porous powder coated with drug and lipids.

Dry granular type of liposomes has been studied for effective delivery of various

drugs like 5-fluorourasil, ibuprofen, indomethacin, adriamycin, doxorubicin,

glyburide, and hydrocortisone.

c. Mixed micellar proliposomes

Mixed micelles contain bile salts, cholesterol, and phospholipids, which upon

dilution, undergo micelles to vesicle transition to form liposomes.

Liquid crystalline proliposomes.

Involves organization of lipid/ethanol/water mixture into lamellar structure.

d. Protransferosomes

Protransferosomes are proultraflexible vesicles, which can be converted into

ultraflexible vesicles.

Characterization of provesicular system

Morphology.

Angle of repose.

Size and size distribution.

Rate of hydration.

Entrapment efficiency.

Degree of deformability and permeability measurement.

In vitro release rate.

In vivo fate and pharmacokinetic.

2. Lipopolyplexes

A combination of DNA, polymers and liposomes has been prepared with a view to

enhance transfection ability by utilization of their individual properties.

It has been reported that this method has resulted in better gene transfer and lower

toxicity as compare to cationic liposomes alone.

3. Transferosomes Modified liposomes developed to increase the transdermal permeation of drug.

Deformability is achieved by using surface active agent in proper ratio.

Concentration of surfactant is very crucial because at sublytic conc. This agent

provides flexibility of transferosomal membrane and at higher conc. and cause

destruction of vesicles.



4. Ethosomes Ethosomal system is a vesicular system composed mainly of phospholipids & alcohol

(ethanol or IPA, sometimes polyols; glycol) in relatively high concentration & water.

Better membrane permeability.

5. Discosomes

Small et al first observed discoidal mixed micelle in phase behavior of PC in cholate-

water system.

6. Virosomes

Reconstituted lipid vesicles equipped with viral glycoprotein is used for DNA

transfer.

7. Emulsomes

New generation colloidal drug carrier unit.

The emulsomes can be explicitly distinguished from fat emulsion or lipid microsphere

as they are distinctively sphere vesicular graft like system due to utilization of higher

quantities of PC both as emulsifying agent as well as surface modifier.

8. Cochleates

Cochleates are cigar-like microstructures that consist of a series of lipid bilayers

which are formed as a result of the condensation of small unilamellar negatively

charged liposomes.

In the presence of calcium, the small phosphatidylserine (PS) liposomes fuse and

form large sheets.

These sheets have hydrophobic surfaces and, in order to minimize their interactions

with water, tend to roll-up into the cigar-like cochleate.

Discovered in 1975 by Dr. D. Papahadjoupoulos.

9. Depofoam technology

Depofoam particles include hundred of bilayer enclosed aqueous compound.

Formed by first emulsifying a mixture of an aqueous phase containing the compound

to be encapsulated & an organic phase containing lipid.

The first emulsion is then dispersed & emulsified in a 2nd aqueous phase.



After the organic solvent is evaporated, numerous submicron to micrometer sized

water compartment are separated by lipid layer & take on a closed packed polyhedral

structure From which the comp slowly permeate.

Marketed

product

Drug used Target diseases Company

DoxilTM

or

CaelyxTM

Doxorubicin Kaposi‟s sarcoma SEQUUS, USA

DaunoXomeTM

Daunorubicin Kaposi‟s sarcoma, breast

& lung cancer

NeXstar, USA

AmphotecTM

Amphotericin-B fungal infections,

Leishmaniasis

SEQUUS, USA

Fungizone® Amphotericin-B fungal infections,

Leishmaniasis

Bristol-squibb, Netherland

VENTUSTM

Prostaglandin-E1 Systemic inflammatory

diseases

The liposome company,

USA

ALECTM

Dry protein free

powder of

DPPC-PG

Expanding lung diseases

in babies

Britannia Pharm, UK



Topex-Br Terbutaline

sulphate

Asthma Ozone, USA

Depocyt Cytarabine Cancer therapy Skye Pharm, USA

Novasome® Smallpox

vaccine

Smallpox Novavax, USA

Avian retrovirus

vaccine

Killed avian

retrovirus

Chicken pox Vineland lab, USA

Epaxal –Berna

Vaccine

Inactivated

hepatitis-A

Virions

Hepatitis A Swiss serum & vaccine

institute, Switzerland

Doxil®

Doxorubicin Hcl Refractory ovarian cancer ALZA, USA

EvacetTM

Doxorubicin Metastatic breast cancer The liposome company,

USA

VincaXome Vincristine Solid Tumours NeXstar, USA

Mikasome® Amikacin Bacterial infection NeXstar, USA

AutragenTM

Tretinoin Kaposi‟s sarcoma Aronex

Pharm, USA

Shigella Flexneri

2A Vaccine

Shigella flexneri

2A

Shigella Flexneri 2A

infections

Novavax, USA

NyotranTM

Nystatin Systemic fungal

infections

Aronex Pharm, USA

CONCLUSION

Liposomes are one of the unique drug delivery system, which can be of potential use in

controlling and targeting drug delivery. Liposomes are administrated orally, parenterally and

topically as well as used in cosmetic and hair technologies, sustained release formulations,

diagnostic purpose and as good carriers in gene delivery. One major problem associated in

the formulation of liposome due to physicochemical and biological instability. These stability

problems can be alleviated by using various methods like lyophilization, proliposome, pH

sensitive liposome, microencapsulation and steric stabilization. Now a days liposomes are

used as versatile carriers for targeted delivery of drug.



1. Sanjay K. Jain and N. K. Jain, Controlled and Novel Drug Delivery Systems “Chapter

15: Liposomes as drug carriers” published by CBS publishers & distributors, reprint:

2002, pg. 304-352.

2. www.pharmainfo.com (Sanjay S. Patel, Liposomes: A versatile platform for targeted

delivery of drugs, volume 4. Issue 5, 2006)

3. http.//en.wikipedia.org/wiki/liposomes

4. Gert Storm, et al., Liposomes: Quo vadis?, Pharm. Sci. Tech. Today 1 (1998) 19-31.

5. D.J.A. Crommelin, et al., Liposomes: vesicles for the targeted and controlled delivery

of peptides and proteins, J. Control. Release 46 (1997) 165-175.

6. Andreas Wagner, et al., Liposomes produced in a pilot scale: Producton, Purification

and efficiency aspects, Eur. J. Pharm. Biopharm. 54 (2002) 213-219.

7. Sugi S. Chrai, et al., Liposomes, part II: drug delivery systems, Pharm. Technol.

Europe (February 2003) 53-56.

8. Jessy Shaji, et al., Immunoliposomes: targeted delivery for cancer, Pharma Times 39

(2007) 17-20.

9. Gregoriaadis G, Liposomes in drug delivery: clinical, diagnosticand ophthalmic

potential, Drugs 45 (1993) 15-28.

http://www.pharmainfo.com/

seminar on liposomes in drug delivery

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