Current Drug Delivery, 2007, 4, 297-305

297

Liposomal Drug Delivery Systems: An Update Review

Abdus Samad, Y. Sultana* and M. Aqil
Department of Pharmaceutics, Faculty of Pharmacy, Jamia Hamdard, New Delhi 110062, India
Abstract: The discovery of liposome or lipid vesicle emerged from self forming enclosed lipid bi-layer upon hydration; liposome drug
delivery systems have played a significant role in formulation of potent drug to improve therapeutics. Recently the liposome formulations
are targeted to reduce toxicity and increase accumulation at the target site. There are several new methods of liposome preparation based
on lipid drug interaction and liposome disposition mechanism including the inhibition of rapid clearance of liposome by controlling particle size, charge and surface hydration. Most clinical applications of liposomal drug delivery are targeting to tissue with or without expression of target recognition molecules on lipid membrane. The liposomes are characterized with respect to physical, chemical and biological parameters. The sizing of liposome is also critical parameter which helps characterize the liposome which is usually performed by
sequential extrusion at relatively low pressure through polycarbonate membrane (PCM). This mode of drug delivery lends more safety
and efficacy to administration of several classes of drugs like antiviral, antifungal, antimicrobial, vaccines, anti-tubercular drugs and gene
therapeutics. Present applications of the liposomes are in the immunology, dermatology, vaccine adjuvant, eye disorders, brain targeting,
infective disease and in tumour therapy. The new developments in this field are the specific binding properties of a drug-carrying
liposome to a target cell such as a tumor cell and specific molecules in the body (antibodies, proteins, peptides etc.); stealth liposomes
which are especially being used as carriers for hydrophilic (water soluble) anticancer drugs like doxorubicin, mitoxantrone; and bisphosphonate-liposome mediated depletion of macrophages. This review would be a help to the researchers working in the area of liposomal
drug delivery.

Keywords: Liposome, characterization, sizing of liposomes, surface hydration, targeted site, phospholipids.
INTRODUCTION
When phospholipids are dispersed in water, they spontaneously
form closed structure with internal aqueous environment bounded
by phospholipid bilayer membranes, this vesicular system is called
as liposome [1]. Liposomes are the small vesicle of spherical shape
that can be produced from cholesterols, non toxic surfactants,
sphingolipids, glycolipids, long chain fatty acids and even membrane proteins [2]. Liposomes are the drug carrier loaded with great
variety of molecules such as small drug molecules, proteins, nucleotides and even plasmids. Liposomes were discovered about 40
years ago by A.D. Bangham [3] which has become the versatile tool
in biology, biochemistry and medicine today. In 1960s, liposome
has been used as a carrier to deliver a wide variety of compounds in
its aqueous compartment. Liposome can be formulated and processed to differ in size, composition, charge and lamellarity. To date
liposomal formulations of anti-tumor drugs and antifungal agents
have been commercialized [4].
The clinical potential of liposomes as a vehicle for replacement
therapy in genetic deficiencies of lysosomal enzymes was first
demonstrated in 1970s [5, 6].
Considerable progress was made during 1970s and 1980s in the
field of liposome stability leading to long circulation times of
liposomes after intravenous administration resulting in the improvement in bio-distribution of liposome.
The important anti-tumour drug doxorubicin had been formulated as liposome in 1980s to improve the therapeutic index.
There are several mechanisms by which liposomes act within
and outside the body which are as follows [7]:
1- Liposome attaches to cellular membrane and appears to fuse
with them, releasing their content into the cell.
2- Some times they are taken up by the cell and their phospholipids are incorporated into the cell membrane by which the drug
trapped inside is released.

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Classification of Liposomes [8]

Liposomes are classified on the basis of:
1. Structure.
2. Method of preparation.
3. Composition and application.
4. Conventional liposome.
5. Specialty liposome.

1. Classification Based on Structure (Table 1)

Table-1.

Vesicle Types with their Size and Number of Lipid Layers

Vesicle Type

Abbreviation

Diameter Size

No of Lipid
Bilayer

Unilamellar vesicle

UV

All size range

One

Small Unilamellar
vesicle

SUV

20-100 nm

One

Medium Unilamellar
vesicle

MUV

More than 100nm

One

Large Unilamellar
vesicle

LUV

More than 100nm

One

Giant Unilamellar
vesicle

GUV

More than 1 micro

meter

One

Oligolamellar vesicle

OLV

0.1-1 micro meter

Approx. 5

Multilamellar vesicle

MLV

More than 0.5

5-25

Multi vesicular vesicle

MV

More than 1 micro

meter

Multi compartmental
structure

*Address correspondence to this author at the Department of Pharmaceutics,

Faculty of Pharmacy, Jamia Hamdard, New Delhi 110062, India;
Tel: +919899078661; +919811539489; E-mail: yas2312003@yahoo.com,
samadpharma@gmail.com
1567-2018/07 $50.00+.00

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3- In the case of phagocyte cell, the liposomes are taken up, the
phospholipid walls are acted upon by organelles called lysosomes
and the active pharmaceutical ingredients are released.

2007 Bentham Science Publishers Ltd.

298 Current Drug Delivery, 2007, Vol. 4, No. 4

Samad et al.

2. Based on Method of Preparation (Table 2)

Table-2.

Different Preparation Methods and the Vesicles Formed by

these Methods

Preparation Method

Vesicle Type

Single or oligo lamellar vesicle made by reverse phase

evaporation method

REV

Multi lamellar vesicle made by reverse phase evaporation

method

MLV-REV

Stable pluri lamellar vesicle

SPLV

Frozen and thawed multi lamellar vesicle

FATMLV

Vesicle prepared by extrusion technique

VET

Dehydration- Rehydration method

DR V

3. Based on Composition and Application (Table 3)

Table-3.

Different Liposome with their Compositions

Type of Liposome

Abbreviation

Composition

Conventional liposome

CL

Neutral or negatively charge phospholipids and cholesterol

Fusogenic liposome

RSVE

Reconstituted sendai virus envelops

PH sensitive liposomes

Phospholipids such as PER or DOPE

with either CHEMS or OA

Cationic lipid with DOPE

Long circulatory
liposome

LCL

Neutral high temp, cholesterol, and 510% PEG, DSP

Immuno liposome

IL

CL or LCL with attached monoclonal

antibody or recognition sequences

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Based Upon Speciality Liposome
1- Bipolar fatty acid
2- Antibody directed liposome.
3- Methyl/ Methylene x- linked liposome.
4- Lipoprotein coated liposome.
5- Carbohydrate coated liposome.
6- Multiple encapsulated liposome.

PREPARATION OF LIPOSOMES
An important parameter to be considered when preparing the
liposome is the rigidity of bi-layers. There are several groups of
phospholipids that can be used for the liposome preparation which
are as follows:
1. Phospholipids from natural source
2. Phospholipids modified from natural source
3. Semi synthetic phospholipids
4. Fully synthetic phospholipids and
5. Phospholipids with natural head groups
Dilauryl phosphotidyl choline (DLPC), Dimyristoyl phosphotidyl choline (DMPC), Dipalmitoyl phosphotidyl choline (DPPC),
Distearoyl phosphotidyl choline (DSPC), Dioleolyl phosphotidyl
choline (DOPC), Dilauryl phosphotidyl ethanolamine (DLPE),
Dimyristoyl phosphotidyl ethanolamine (DMPE), Distearoyl phos-

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Cationic liposome

Based Upon Conventional Liposome

1- Stabilize natural lecithin (PC) mixtures
2- Synthetic identical, chain phospholipids
3- Glycolipids containing liposome

photidyl ethanolamine (DSPE), Dioleoyl phosphotidyl ethanolamine (DOPE), Dilauryl phosphotidyl glycerol (DLPG), Distearoyl
phosphotidyl serine (DSPS) are the commonly used phospholipids
for liposome preparation (Table 4). Cholesterol can be added to the
bilayers mixture for the following purposes:
1. Act as a fluidity buffer
2. Act as intercalator with phospholipids molecules
Alters the freedom of formation of carbon molecule in the acyl
chain
3. Restrict the transformation of Trans to gauche conformation.
Rajendran et al. [9] prepared liposome from egg lecithin by
modified ether injection technique using stearylamine added dicetylphosphate as the charge inducing agent. The charged liposomes
were larger in the size and showed better drug entrapment efficiency and were found more entrapped in the organs of the reticuloendothelial system than neutral liposome.
It was also shown that cholesterol decreased the permeability
coefficients of negative, neutral as well as positively charged membranes to Na+, K+, Cl- and glucose. Cholesterol also stabilized the
membranes against temperature changes, leading to lower permeability at elevated temperatures [11].
Cholesterol is essential for lowering membrane permeability,
and imparting better stability. Cholesterol also modulates membrane-protein interactions. The polymorphic phase behavior of
lipids is modulated by a variety of factors including hydration, temperature and divalent cations, degree of unsaturation of the acyl
chains and the presence of other lipids such as sterols [12, 13].
Lasic et al. [14] tried active trapping of amphipathic weak bases
inside liposomes by using pH induced transmembrane potential
establishment. Doxorubicin was used as a model drug.
Triton X-100 has been observed to affect the physical properties of liposomes [15]. Solubilizing effect of Triton X-100 on the
membranes composed of different phospholipids and cholesterol
has been evaluated [16]. The measured changes indicated a drastic
structure transformation into entities of fairly high dipolar moment
leading to solubilization of phospholipids membranes.
The three different strategies for the preparation of liposomes
are as follows:
1. Mechanical methods
2. Methods based on replacement of organic solvent
3. Methods based on size transformation or fusion of prepared vesicle

1. MECHANICAL METHODS
A. Film Method
The original method of Bangham et al. [2] is still the simplest
procedure for the liposome formation but having some limitation
because of its low encapsulation efficiency. In this technique
liposome are prepared by hydrating the thin lipid film in an organic
solvent and organic solvent is then removed by film deposition
under vacuum. When all the solvent get removed, the solid lipid
mixture is hydrated using aqueous buffer. The lipids spontaneously
swell and hydrate to form liposome. This methods yields a heterogeneous sized population of MLVs over 1 micro meter in diameter.
Nagarsenkar et al. [17] prepared lidocaine encapsulated
liposomes employing the conventional lipid film hydration technique. The prepared liposomal dispersions were investigated, the
results showed that lidocaine incorporated into the liposomes got
selectively partitioned and localized in the skin to a greater extent.
A topical liposomal gel formulation containing 2% w/w lidocaine
was prepared using cabopol-934 as the gelling agent. The liposome
preparation of lidocaine gave a much longer duration of action
compared to the conventional topical formulation.

Current Drug Delivery, 2007, Vol. 4, No. 4

Liposomal Drug Delivery Systems

Table 4.

299

Phospholipids and Cholesterol which are Used for the Preparation of Liposome and their Transition Temperature (Tc) and Molecular
Weights [10]

Phospholipids

Abbreviation

Charge

Tc (0c)

Mol. Wt.

Brain Sphingomyellin

BSPM

32

814

Cholesterol

CH

--

387

Dimyristoyl phosphatidylcholine

DMPC

23

Dipalmitoyl phosphatidylcholine

DPPC

041

Distearoyl phosphatidylcholine

DSPC

58

790

Dioleyl phosphatidylcholine

DOPC

-22

786

Dilaruryl phosphatidylglycerol

DLPG

-1

633

Dimyristoyl phosphatidylglycerol

DMPG

-1

23

689

678
734

Dipalmitoyl phosphatidylglycerol

DPPG

-1

41

745

Distearoyl phosphatidylglycerol

DSPG

-1

55

796

Phosphatidyl ethanolamine

PE

48

Dimyristoyl phosphatidyl ethanolamine

DMPE

--

Dipalmitoyl phosphatidyl ethanolamine

DPPE

60

Distearoyl phosphatidyl ethanolamine

DSPE

Dimyristoyl PA

DMPA

-2

Dipalmitoyl PA

DPPA

-2

Dicetyl phosphate

DOPA

-2

DPPS

--

DCP

-1

Ridy et al. [18] prepared oxamniquine liposomes with different

compositions and surface charge were prepared by the chloroform
film method. The amount of oxamniquine entrapped was estimated.
Negatively charged liposomes exhibited the highest percentage
entrapment (23.09%). The maximum oxamniquine entrapment was
achieved in liposomes prepared from phospholipid molar ratio
(7:4:1). The chemoprophylactic effect of free and oxamiquine
liposome formulations was estimated and showed that drug encapsulated in liposomes provided more efficient prophylaxis.

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B. Ultrasonic Method
This method is used for the preparation of SUVs with diameter
in the range of 15-25
m. Ultrasonication of an aqueous dispersion
of phospholipids is done by two types of sonicators i.e. either probe
sonicators or bath sonicators. The probe sonicators are used for the
small volume which requires high energy while the bath sonicators
are employed for the large volume [19].
2. METHODS BASED ON REPLACEMENT OF ORGANIC
SOLVENTS
In this method lipids are co-solvated in organic solution, which
is then dispersed into aqueous phase containing material to be entrapped within the liposome. This method is of two types:
A. Reverse Phase Evaporation
The lipid mixture is added to a round bottom flask and the solvent is removed under reduced pressure by a rotary evaporator. The
system is purged with nitrogen and lipids are re-dissolved in the
organic phase which is the phase in which the reverse phase vesicle
will form. Diethyl ether and isopropyl ether are the usual solvents
of choice. After the lipids are redissolved the emulsion are obtained
and than the solvent is removed from an emulsion by evaporation to
a semisolid gel under reduced pressure. Non encapsulated material
is then removed. The resulting liposomes are called reverse phase
evaporation vesicles (REV). This method is used for the preparation

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Dioleoyl PA
Dipalmitoyl phosphatidyl serine

--

51

67
--

48
--

720

636

692

748

618

649

701

758

547

of large uni-lamellar and oligo-lamellar vesicles formulation and it

has the ability to encapsulate large macromolecules with high efficiency [20].
Pleumchitt et al. [21] showed the physicochemical properties of
phospatidylcholine cholesterol liposomes containing amphotericin
B prepared by reverse phase evaporation method. The liposomes
containing amphotericin B 2.0 mol % of total lipid demonstrated
the highest percentage of drug entrapment. The highest drug entrapment efficiency (approx. 95%) with particle size range of 13071451 nm was obtained with the formulation containing 1:1 molar
ratio of phosphatidycholine to cholesterol.
B. Ether Vaporization Method
There are two method according to the solvent used:
i. Ethanol injection method.
ii. Ether injection method.
In ethanol injection method, the lipid is injected rapidly through
a fine needle into an excess of saline or other aqueous medium. In
ether injection method the lipid is injected very slowly through a
fine needle into an excess of saline or other aqueous medium.
3. METHODS BASED ON SIZE TRANSFORMATION OR
FUSION OF PREFORMED VESICLE
A. Freeze Thaw Extrusion Method
The freeze thaw method is an extension of the classical DRV
method. Liposomes formed by the film method are vortexed with
the solute to be entrapped until the entire film is suspended and the
resulted MLVs are frozen in luke warm water and than vortexed
again. After two cycles of freeze thaw and vortexing the sample is
extruded three times. This is followed by six freeze thaw cycle and
addition eight extrusions. This process ruptures and defuses SUVs
during which the solute equilibrates between inside and outside and
liposome themselves fuse and increase in size to form large Uni
lamellar vesicle by extrusion technique (LUVET). For the encapsulation of protein this method is widely used [22].

300 Current Drug Delivery, 2007, Vol. 4, No. 4

Samad et al.

angular correlation (PAC) spectroscopy. In this 111In-label diethyene triamine penta acetic acid (DTPA) derivative dipalmitoyl
phosphatidyl ethanolamine (DPPE) lipid were incorporated in the
SUVs. This helped in the continuous non- invasive monitoring of
the microenvironment of the lipid bilayer.
Kolchens [29] used quasi elastic light scattering (QELS) or
photon correlate spectroscopic (PCS) technique for determining
distribution of vesicles prepared by freeze-thaw extrusion method.
The influence of filter pore size, extrusion press and lipid concentration on the size and size distribution extruded vesicles was studied.
Chemical characterization includes those studies which established the purity and potency of various liposomal constituents.
Biological characterization is helpful in establishing the safety
and suitability of formulation for the in vivo use for therapeutic
application [30]. The characteristics of the carrier through appropriate choice of membrane components, size and charge determines
the final behavior of liposomes both in vitro and in vivo as well [31,
32].
Jousma et al. [33] characterized the vesicles obtained by repetitive extrusion through polycarbonate membranes using freezefracture electron microscopy, small angle X-ray scattering (SAXS),
and encapsulated volume using 6-carboxy flurescein (6-CF) and
31
p-NMR as aqueous markers.

Canto et al. [23] prepared liposomes of soya phosphatidylcholine, cholesterol and stearylamine (molar ratio 6/3/1) and 0.1 %

to-copherol by the extrusion of multi-lamellar vesicles through 0.2

m polycarbonate membrane. They analyzed free piroxicam at 4
mg/kg, piroxicam encapsulated in liposomes and added to 1.5%
hydroxyethylcellulose (HEC) gel at 1.6 mg/kg, and piroxicam encapsulated in liposomes and added to HEC gel at 4 mg/kg. The
inhibition of inflammation obtained was 21.1%, 32.8% and 47.4%
respectively. These results showed that the encapsulation of piroxicam produced an increase of topical anti-inflammatory effect.
B. The Dehydration- Rehydration Method
In this method the empty buffer containing SUVs and rehydrating it with the aqueous fluid containing the material to be entrapped
after which they are dried. This leads to a dispersion of solid lipids
in finely subdivided form. Freeze drying is often the method of
choice. The vesicles are than rehydrated. Liposomes obtained by
this method are usually oligo lamellar vesicle [24].
Bhalerao et al. [25] prepared liposome by the conventional thin
film hydration technique. The result showed that the formation of
bi-layered liposomes in the particle size range of 0.2-0.8276
m
occurred with a maximum entrapment efficiency of 42.6%. The
liposomes stored at 4-5oC showed maximum stability as compared
to those stored at any other temperature.
SIZING OF LIPOSOMES:
Size characteristic of liposome have a major effect on their fate
i.e. for which application they can be used for. The therapeutic
applications of liposome are dependent on physical integrity and
stability of lipid bilayers structure. Therefore liposome production
procedure must be predictable and reproducible with particle size
distribution within a certain size range. Lipid based formulation can
be devised as site specific drug delivery vesicles that are:
1. Relatively cleared by kupffer cells of liver and the macrophages [26].
2. Evade detection of active substance by the reticulo endothelial system (RES) and efficiency deliver liposome incorporated
material to target tissue, organ or tumor [27]. Sizing of liposome are
usually performed by sequential extrusion at relatively low pressure
through polycarbonate membrane (PCM). The membrane extrusion
technique can be used to process LUVs as well as MLVs to form
LUVETs. The membrane (PCM) is of pore size 0.27 micrometer.
The other procedure of sizing of the liposomes are gel chromatography , mainly used to size liposome but more typically used to
remove encapsulated components by separation. Sonication is the
third method of sizing of liposomes, but this method is related with
following disadvantages:
1. Exclusion of oxygen is difficult which result in per oxidation
reaction
2. Titanium probes shed metal particle resulting in contamination
3. They can generate aerosols, which exclude them with from use
with certain agents
These above problems are mainly related with the probe sonication but these problems can be removed by using the bath sonication.

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Table-5.

CHARACTERIZATION OF LIPOSOMES (TABLE 5)

After preparation and before use in immunoassay the liposome
must be characterized. Evaluation could be classified into three
broad categories which are physical, chemical and biological methods. The physical methods include various parameters, which are
size, shape, surface features, lamellarity phase behaviors and drug
release profile.
Ma et al. [28] evaluated structural integrity of liposomal phospholipids membrane by a new technique of gamma-ray perturb

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Characterization of Liposomes with their Quality Control

Assays [34-52]

A. Biological characterization
Characterization parameters

Instrument for analysis

Sterility

Aerobic/anaerobic culture

Pyrogenicity

Rabbit fever response

Animal toxicity

Monitoring survival rats

B. Chemical characterization
Characterization parameter

Instrument for analysis

Phospholipids concentration

HPLC/Barrlet assay

Cholesterol concentration

HPLC / cholesterol oxide assay

Drug concentration

Assay method

Phospholipids per oxidation

UV observance

Phospholipids hydrolysis

HPLC/ TLC

Cholesterol auto-oxidation

HPLC/ TLC

Anti-oxidant degradation

HPLC/TLC

PH

PH meter

Osmolarity

Osmometer

C. Physical Characterization
Characterization parameter

Instrument for analysis

Vesicle shape, and surface morphology

TEM and SEM

Vesicle size and size distribution

Dynamic light scattering ,TEM

Surface charge

Free flow electrophoresis

Electrical surface potential and surface

PH

Zeta potential measurement and PH

sensitive probes

Lamellarity

P31 NMR

Phase behavior

DSC, freeze fracture electron microscopy

Percent capture

Mini column centrifugation, gel exclusion

Drug release

Diffuse cell/ dialysis

Current Drug Delivery, 2007, Vol. 4, No. 4

Liposomal Drug Delivery Systems

STABILIZATION OF LIPOSOME
The stability of liposome should meet the same standard as
conventional pharmaceutical formulation. The stability of any
pharmaceutical product is the capabilities of the delivery system in
the prescribed formulation to remain within defined or pre established limits for predetermined period of time. Chemical stability
involves prevention of both the hydrolysis of ester bonds in the
phospholipids bilayers and the oxidation of unsaturated sites in the
lipid chain. Chemical instability leads to physical instability or
leakage of encapsulated drug from the bilayers and fusion and finally aggregation of vesicles [53].
Chen et al. introduced the pro-liposome concept of liposome
preparation to avoid physicochemical instability encountered in
liposome suspension such as aggregation, fusion, hydrolysis, and/or
oxidation [54].
Approaches that can be taken to increase liposomal stability
involve efficient formulation and lyophillization. Formulation involves the selection of the appropriate lipid composition, concentration of bilayers, aqueous phase ingredients such as buffers, antioxidant, metal chelators and cryo protectants. Charge inducing lipid
such as phosphotidyl glycerol can be incorporated into liposome
bilayers to decrease fusion while cholesterol and sphingomyellin
can be included in the formulation to decrease permeability and
leakage of encapsulated drugs. Buffers at neutral pH can decrease
hydrolysis; addition of antioxidant such as sodium ascorbate can
decrease oxidation. Oxygen potential is kept to minimum during
processing by nitrogen purging solution [55].
In general successful formulation of stable liposomal drug
product requires the following precautions:
1. Processing with fresh, purified lipids and solvents.
2. Avoidance of high temperature and excessive shear forces
3. Maintenance of low oxygen potential (Nitrogen purging)
4. Use of antioxidant or metal chelators
5. Formulating at neutral pH.
6. Use of lyo-protectant when freeze drying

2. Protection of sensitive drug molecules (Cytosine arabinosa,

DNA, RNA, Anti-sense oligo-nucleotides, Ribozymes)
3. Enhance intracellular uptake (Anticancer, anti viral and antimicrobial drugs)
4. Altered pharmacokinetics and bio-distribution (prolonged or
sustained released drugs with short circulatory half life)
Several recent applications of liposomal drug delivery system
are as follows:
A. Liposome for Respiratory Drug Delivery System
Liposome is widely used in several types of respiratory disorders. Liposomal aerosol has several advantages over ordinary aerosol which are [57] as follows:
1. Sustained release
2. Prevention of local irritation
3. Reduced toxicity and
4. Improved stability in the large aqueous core.
Several injectable liposome based product are now in the market including ambisome, Fungisome and Myocet. To be effective,
liposomal drug delivery system for the lung is dependent on the
following parameters:
1. Lipid composition
2. Size
3. Charge
4. Drug and Lipid ratio and
5. Method of delivery
The recent use of liposome for the delivery of DNA to the lung
means that a greater understanding of their use in macromolecular
delivery via inhalational is now emerging. Much of this new
knowledge, including new lipids and analytical techniques, can be
used in the development of liposome based protein formulations.
For inhalation of liposome the liquid or dry form is taken and the
drug release occurs during nebulization. Drug powder liposome has
been produced by milling or by spry drying. Drugs which are formulated in the form of liposome are presented in Table 6.

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APPLICATION OF LIPOSOME
The field of liposome research has expanded considerably over
last 30 years. It is now possible to engineer a wide range of
liposome of varying size, phospholipids composition, cholesterol
composition, surface morphology suitable for wide range of application [56].
Liposomes interact with cells in many ways to cause liposomal
components to be associated with target cells [7].
The liposome carrier can be targeted to liver and spleen and
distinction can be made between normal and tumors tissue using
tomography. In case of transdermal drug delivery system, liposome
has a great application. Liposomal drug delivery system when used
to target the tumor cells leads to reduction in the toxic effect and
enhances the effectiveness of drugs. The targeting of the liposome
to the site of action takes place by the attachment of amino acid
fragment, such as antibody or protein or appropriate fragments that
target specific receptors cell. Liposomal DNA delivery vectors and
further enhancement in the form of LPDI -I and LPD-II are some of
the safest and potential most versatile transfer vectors which are
used to date.
DNA vaccination and improved efficiency of gene therapy are
just a few of the recent application of liposome. Several modes of
drug delivery application have been purposed for the liposomal
drug delivery system, few of them are as follows:
1. Enhance drug solublisation (Amphotericin-B, Minoxidil, Paclitaxels, and Cyclosporins)

301

Table-6.

Liposomal Formulation for the Respiratory Disorder [58]

Active constituent

Effect

Insulin

Facilitated pulmonary adsorption and enhanced hypoglycemic effect

Catalase

Conferred resistance to pulmonary oxygen toxicity

Super oxide dusmutase

Minimize toxicity to subsequent hyperoxia and improved survival

Cyclosporins

Preferentially adsorbed by lung and shows sustained

release

Ricin vaccine

Improved safety profile for intra pulmonary vaccination

Interleukin-2

The lungs Facilitated bioactivity and reduce toxicity

Isoniazid and Rifampicin

Improved the effect of drugs for the tuberculosis

B. Liposome in Nucleic Acid Therapy

Recombinant DNA technologies and studies of gene function
and gene therapy all depends on the successful delivery of nucleic
acid into cells in vitro and in vivo. Non viral vectors will be developed for the selective delivery of the gene to the malignant cells.
The vector will exploit the increase requirements of rapidly growing cells for more nutrients by attaching a nutrients ligand onto the
vector (liposome). The vector additionally will have a passively
charged lipid to enhance nucleic acid binding along with novel pH

302 Current Drug Delivery, 2007, Vol. 4, No. 4

Samad et al.

sensitive surfactants. The advantages and disadvantages of these

technologies are given in Table 7. The role of surfactants is to increase the amount of nucleic acid escaping the endosome and correspondingly increase the transfection efficiency [59, 60]. The vector
system will be evaluated first in an animal model of cancer. Lipid
based gene delivery is the focus of several specialized high technology companies, of which Vical ( San Diago, CA, USA), Genzyme ( Farmington, MA, USA) and Megabios (Burhingam, CA,
USA) have products in clinical trials.
Preliminary studies will be carried out using a marker gene
(BETA GALCTOSIDASE) with later experiments using a gene
encoding for cytosine deaminase. Cytosine deaminase can catalyze
the conversion of the innocuous agent 5- Flourocytosine to the anticancer agent 5- Fluorouracil. By selective delivery of this gene to
only cancer cells, the therapeutic index of 5-Flourouracil can then
be increased. Some of the engineered liposomal and non liposomal
versions like pH sensitive cationic and anionic liposome, pH sensitive immuno-liposome, fusagenic liposome, genosomes, lipofections and recently cochleats are being investigated as the major
gene vectors (Table 8). This agent will become activated at endosomal pH, has membrane disrupting effects and will be inactivated before reaching the lysosome [61]. For this agent to work, the
surfactant must enter the cell by endocytosis. The soft surfactant
will be incorporated into liposome that has shown to be entering
cell in this manner. The novelty of this delivery system stems from
soft pH sensitive surfactants (SPS). The pH sensitive liposomes
have been reported as plasmid expression vectors for the cytosolic
delivery of DNA. It is also effective carrier for intracellular trafficking of anti sense oligo nucleotides [62].

Viral vectors ( Adeno

virus, Retro vrus and
Adeno-associated virus)

Advantage

Disadvantage

Relative high transfection efficiency

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Non viral vectors
(Liposome, polymers
and peptides)

Table-8.

Favourable pharmaceutical issue

Plasmid ind4ependent
structure
Low immunogenicity
Opportunity for chemical / physical manipulation

Immunogenicity, presence of contaminants

and safety
Vector restricted size
limitation for recombinant gene
Unfavourable pharmaceutical issue

Relatively low transfection efficiency

Some of the Widely Cationic Liposome Formulations

Name

D. Liposome as Vaccine Adjuvant [64]

Liposome has been firmly established as immuno-adjuvant,
potentiating both cell mediated and non cell mediated (humoral)
immunity (Table 9).
Liposome acts as immuno-adjuvant by the following therapeutic points of view:
1. Liposomes as an immunological (vaccine) adjuvant
2. Liposomal vaccines
3. Liposomes as carrier of immuno modulation for
4. Liposomes as a tool in immuno diagnostics
Liposomal immuno-adjuvant act by slowly releasing encapsulated antigen on intramuscular injection and also by passively accumulating within regional lymph node [65].
The accumulation of liposome to lymphoid is done by the targeting of liposome with the help of phosphotidyl serine. Liposomal
vaccine can be prepared by inoculating microbes, soluble antigen,
cytokinesis of deoxyribonucleic acid with liposome. The latter
stimulating an immune response on expression of antigenic protein.
Antigens can be covalently coupled to liposomal membrane [66].

Composition

Lipofectin

DOTMA:DOPE(1:1)

Lipofetamine

DOSPA:DOPE(3:1)

LipofectASE

DDAB:DOPE(1:2.1)

TranfectASE

DDAB:DOPE(1:3)

Transfectam

DOGS

DC-chol

DC-Chol:DOPE(3:2)

DOTMA:N-[1-(2,3-Dioleoyloxy)propyl]N,N,N-trimethyl ammonium chloride

DOPE: Dioleoylphosphatidylethanolamine
DOSPA:2,3-Dioleoyloxy[2(sperminecarboxamido)ethyl]N,N- dimethyl-1-propanammonium triflouroacetate
DDAB: Dimethyl-dioctadecylethanolamine
DOGS: Dioctadecylamido glycyl spermine
DC-Chol: Chol-dimethyl amino ethane carbamoyl cholesterol.

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Table-7. Advantages and Disadvantages of Recombinant Technologies

Types of Vectors

C. Liposome in Eye Disorders

Liposome has been widely used to treat disorder of both anterior and posterior segment. The disease of eye includes dry eyes,
keratitis, corneal transplant rejection, uveitis, endopthelmitis and
proliferative vitro retinopathy. Retinal diseases are leading cause of
blindness in advanced countries. Liposome is used as vector for
genetic transfection and monoclonal antibody directed vehicle. The
recent techniques of the treatment like applying of focal laser to
heat induced release of liposomal drugs and dyes are used in the
treatment of selective tumour and neo-vascular vessels occlusion,
angiography, retinal and choroidal blood vessel stasis. Liposomal
drug formulations have been approved for the two of patent drugs
to date and several other products are under clinical trials. The
liposomal drugs currently approved are verteporfin for the use in
the eye , the benefit of the liposome will be applied in treatment,
diagnostic and research aspect of ophthalmology in future [63].

Table-9.

Some Antigens as Liposomal Preparation and their Applications

Antigens as Liposomal Preparation

Applications

Rabies glycoproteins

Interleukin -2 enhancement

Cholera toxin

Enhanced Ab* level

Diphtheria toxoid

Superior immunoadjuvant

Herpes simplex virus

Enhanced Ab level

Hepatitis B virus

Higher Ab response

Bacterial polysaccharides

Superior immunoadjuvants

Tetanus toxoids

Increased Ab titre

Influenza subunit antigen

Intranasal, protects animal from virus

Carbohydrate antigen

Increased Ab titre in salivary gland

*Ab = Antibody

Liposomes which are encapsulating antigen are second time

encapsulated within alginate lysine microcapsules, to control antigen release and improve the antibodies responses. Liposomal vaccines can be store at the refrigerated condition for about 12 months.
E. Liposomes for Brain Targeting
The biocompatible and biodegradable behaviour of liposomes
have recently led to their exploration as drug delivery system to
brain [67].

Current Drug Delivery, 2007, Vol. 4, No. 4

Liposomal Drug Delivery Systems

picin in lung targeted liposome modulates the toxicity and improve

the efficiency of these drugs [72, 73]. Various formulations of the
Amphotericin (Table 11) had been approved by several clinical
trials and are now marketed at different European countries [74].

Liposomes with a small diameter (100 nm) as well as large

diameter undergo free diffusion through the Blood Brain Barrier
(BBB). However it is possible that a small unilamellar vesicles
(SUVS) coupled to brain drug transport vectors may be transported
through the BBB by receptor mediated or absorptive mediated transcytosis. Similarly, cationic liposomes which were developed recently showed these structures to undergo absorptive mediated
endocytosis into cells. Whether cationic liposomes successfully
undergo absorptive mediated transcytosis through the BBB has not
yet been determined. The transport of substances through BBB by
liposomes was extensively studied. The important finding issues
from their studies are that the addition of the sulphatide (a sulphur
ester of galactocerebroside) to liposome composition increases their
several recent applications ability to cross BBB [68]. Wang et al.
reported that liposomes coated with the mannose reach brain tissue
and the mannose coat assist transport of loaded drug through BBB
[69]. The neutropeptides, leu-enkephaline and mefenkephalin kyoforphin normally do not cross BBB when given systemically. The
anti depressant amitriptylline normally penetrate the BBB, due to
versatility of this method. Nanoparticles (NP) were fabricated with
different stabilizers. It was found that amitriptyline level was significantly enhanced in brain when the substance was adsorbed onto
the NP and coated or particle stabilized with polysorbate 85 [70].
F. Liposome as Anti-Infective Agents
Intracellular pathogen like protozoal, bacterial, and fungal reside in the liver and spleen and thus to remove these pathogen the
therapeutic agent may be targeted to these organ using liposome as
vehicle system [71]. The disease like leishmaniasis, candidiasis,
aspergelosis, histoplasmosis, erythrococosis, gerardiasis, malaria
and tuberculosis are targeted by the respective therapeutic agent
using liposome as carrier shown in Table 10.
Table-10.

Active Constituents

F
t
o

Active targeting approach

Pentamidin

Leishmaniasis

Antisense oligo-nucleotides

Leishmaniasis

Anamycin

Table-11.

Application

Leishmaniasis

Asiaticoside

Tuberculosis and Leprosy

Rifampicin

Tuberculosis

Passive targeting approach

Amphotericin B

Meningitis, Leishmaniasis, Candidiasis

Praziquantal

Macrophage activation

Sparfloxacin

M. avium, M. Intracellularie complex

Gentamycin

Staphylococcal pneumonias

Use of Amphotericin B, a polyene antibiotic, in the treatment of

systemic fungal infection is associated with extensive renal toxicity.
Amphotericin B act by the mechanism, in which it binds to sterol in
the membrane of sensitive fungi, thus increasing the membrane
permeability. The toxicity of this compound is due to non specificity and binding to the mammalian cell cholesterol. Recently the first
preparation of Amphotericin B (ambisome) in the form of liposome
had passed all clinical trials and now it is used for the treatment of
fungal infections. Liposomal Amphotericin B, by passively targeting the liver and spleen, reduces the renal and general toxicity at
normal dose but renal toxicity appears when the drug is given at
elevated dose due to the saturation of liver and spleen macrophages.
Liposome can also be targeted to lungs by coating vesicle with ostearoyl amylopectin, polyoxylethylene or mono-sialogangliocyte.
The encapsulation of antitubercular agent like isoniazid and rifam-

Various Liposomal Preparation of Amphotericin B

Preparation

Drug

Targeted Site

Liposome (Am Bisome)

Amphotericin B

Systemic fungal infection,

Visceral lieshmaniaisis

Liposome (Amphocil)

Amphotericin B

Systemic fungal infection

Liposome (ABLC)

Amphotericin B

Systemic fungal infection

LIPOSOME IN TUMOUR THERAPY

The long term therapy of anticancer drug leads to several toxic
side effect. The liposomal therapy for the targeting to the tumour
cell have been revolutionized the world of cancer therapy with least
side effect. It has been said that the small and stable liposome are
passively targeted to different tumour because they can circulate for
longer time and they can extra vasate in tissue with enhanced vascular permeability [75, 76]. Liposome macrophage uptake by liver
and spleen hampered the development of liposome as drug delivery
for aver 20 years.
Several formulations of liposomal anticancer drug which are in
clinical use are given in Table 12.
Table-12.

Various Intravenous
Neoplastics

Preparation

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Some Liposomal Preparation for Infective Disease

303

Drug

Liposomal

Antibiotics

Anti-

Targeted Site

Liposome (Doxil)

Doxorubicin

Kaposi sarcoma

Liposome
(EVACTTM)

Doxorubicin

Refractory tumour

Daunosome

Advanced Kaposi sarcoma, breast ,

small cell lung cancer, leukaemia and
solid tumour

Liposome
(DaunoXome)

Metastatic breast cancer

Liposome

Nystatin

Liposome

Anamycin

Kaposi sarcoma, Refractory breast

cancer

Systemic fungal infection

Liposome (VincaXome)

Vincristine

Solid tumour

Liposome ( Mikasome)

Amikacin

Serious bacterial infection

Doxil is the liposomal formulation of doxorubicin, intravenous,

chemotherapeutic agent. Doxil is prepared by the new technology
called stealth technology, stealth liposome. These are the long circulatory liposome which is prepared by several means. Caelyx and
myocet are the liposomal formulations of doxorubicin. Caelyx is
used for treatment of metastatic ovarian cancer but now in now in
advanced breast cancer. Myocet s approved for metastatic breast
cancer [77-79].
CONCLUSION
Almost from the time of their discovery in 1960s and the demonstration of their entrapment potential, liposome vesicle have
drawn attention of researchers as potential carriers of various bioactive molecules that could be used for therapeutic applications in
human and animals. Many factors contribute to their success as
drug delivery vehicles. Liposomes solubilise lipophillic drug candidates that would otherwise be difficult to administer intravenously.
The encapsulated drug is inaccessible to metabolizing enzyme;
conversely, body component such as erythrocyte and tissue injection site are not directly exposed to full dose of the drug. Liposomes

304 Current Drug Delivery, 2007, Vol. 4, No. 4

Samad et al.

can cross the BBB because of the lipophillic nature of the phospholipids, so even the hydrophilic drugs (which otherwise can not easily
cross the BBB) might be formulated as liposomes. Liposome can
prolong the drug action by slowly releasing the drug in the body.
Targeting option change the distribution of the drug in the body.
They can also be used as adjuvant in vaccine formulation.
Liposome are prepared by various methods in which the most
common method applied for research purpose are film method and
dehydration rehydration method and many more. Stabilization of
liposomes has been an area of concern for optimum shelf life of the
liposomal formulation. Nowadays, stealth liposome (Pegylated
liposome) are under development, with prolonged circulation and
residence time in the body. The new developments in the liposome
are the specific binding properties of a drug-carrying liposome to a
target cell (tumor cell and specific molecules), stealth liposomes for
targeting hydrophilic (water soluble) anticancer drugs like doxorubicin, mitoxantrone which leads to decrease in side effects because
the drug is mostly concentrated at the site of action. Other development is bisphosphonate-liposome mediated depletion of macrophages. Several commercial liposomes have already been discovered, registered and introduced with great success in pharmaceutical
market. There is even greater promise in future for marketing of
more sophisticated and highly stabilized liposomal formulations. In
future the liposomal drug delivery system will revolutionize the
vesicular systems with wide application specially in the treatment
of cancer.

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Accepted: February 14, 2007

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