LIPOSOMESLIPOSOMES-MICROPARTICULATE DRUG CARRIERS PART 1

DIGPATI ROY M Pharm. Assistant Professor KVSR SIDDHARTHA COLLEGE OF PHARMACEUTICAL SCIENCES,VIJAYAWADA 520010

CONTENTS INTRODUCTION PASSIVE LOADING TECHNIQUES

LIPOSOMES Concentric bilayered vesicles-an aqueous vesiclesvolume enclosed by a lipid bilayer membrane The membrane composed of natural or synthetic phospholipids Versatile drug delivery and targeting system Discovered by Dr Alec D Bangham in 1961(Published in 1964)

BENEFITS OF LIPOSOMES
Increased stability via encapsulation Reduction in toxicity of encapsulated agent Selective passive targeting to tumour tissues Active targeting by coupling with site specific ligands Site avoidance effect Improved pharmacokinetic effects-increased effectscirculation, decreased elimination Increased efficacy and therapeutic index

MECHANISM OF LIPOSOME FORMATION

Phospholipids are amphipathic in nature. The polar part consists of phosphoric acid, bound to a water soluble molecule. The non-polar part contains two fatty acid nonchains having 10-24 carbon atoms and 0-6 100double bonds in each chain. The polar lipids transform into stable colloidal nanoconstructs referred to as liposomes,upon dilution with an excess of water .

TYPICAL PHOSPHOLIPID STRUCTURE

LIPOSOMES FROM PHOSPHATIDYLCHOLINE

Phosphatidylcholine is a natural phospholipid. A glycerol bridge links to pair of hydrophobic acyl hydrocarbon chains with a hydrophilic polar head group,phosphocholine. In aqueous medium, the molecules align themselves closely in planner bilayer sheets. These sheets fold over themselves to form closed, sealed and concentric vesicles. This distinctive behavior derives in the lowest free energy state.

PARAMETERS AFFECTING BILAYER FORMATION

The large free energy difference between the aqueous and hydrophobic environment promotes the bilayer structures in order to achieve the lowest free energy level. The driving force for bilayer configuration of liposomes is the hydrophobic interaction coupled with the amphiphilic nature of the principal phospholipid molecules. Supramolecular self-assemblages mediated selfthrough specific molecular geometry.

PHASE TRANSITION OF LIPID BILAYERS INTO LIPOSOMES

Homogeneous,parallel lipid bilayers,closely packed,more ordered, lyotropic state(Gel phase)

Phase transition temperature

Water

Heterogeneous,multilamellar /unilamellar liposomes,loosely packed, less ordered,liquid crystal state(Fluid phase)

RIGIDIZATION OF FLUID PHASE VESICLES WITH CHOLESTEROL

CHOLESTEROL Inserts into the bilayer membrane formed by the phospholipid in very high concentrations. Orients its hydroxyl group towards the aqueous surface and the aliphatic chain aligned parallel to the acyl chains in the center of the bilayer membrane. Tends to condense and rigidize the membrane by suppressing the tilt of the acyl chains

ROLE OF CHOLESTEROL IN BILAYER FORMATION

Acts as a fluidity buffer. After intercalation with phospholipid molecules alters the freedom of motion of carbon molecules in the acyl chain. Restricts the transformations of trans- to transgauchegauche- conformations.

CHOLESTEROL ORIENTATION IN PHOSPHOLIPID BILAYERS

MOLECULAR GEOMETRY & LIPOSOME FORMATION (Israelachvili Hypothesis) MicelleMicelle-forming amphiphiles (e.g. surfactants) -CMC about 10-3 mol L-1 -High solubility in water MembraneMembrane-forming amphiphiles (e.g.phospholipids) -CMC about 10-8 mol L-1 -Less solubility in water

CRITICAL PACKING PARAMETER(CPP)

CPP=v /Ic Ap=Ahp/Ap /Ic Ap=Ahp/ Where v=hydrophobic group volume, Ic =the critical hydrophobic group length , Ap =the cross-sectional area of the hydrophilic head group crossand Ahp =the cross-sectional area of hydrophobic group crossA CPP below 0.5 (indicating a large contribution from the hydrophilic group area) is reported to give spherical micelles and above 1(indicating a large contribution from the hydrophobic group volume) should produce inverted micelles.

FORMATION OF LIPOSOMES

CLASSIFICATION OF LIPOSOMES

METHODS OF LIPOSOME PREPARATION

-Water soluble(hydrophilic) materials entrapped inside the liposomes by using aqueous solution of these materials as hydrating fluid or by the addition of drug/drug solution at some stage during the manufacturing of the liposomes. liposomes. -Lipid soluble (lipophilic) materials solubilized in the organic solution of the constitutive lipid(s) and then evaporated to a dry drug containing lipid film followed by its hydration. hydration. -These methods involve the loading of the entrapped agents before or during the manufacturing procedure (PASSIVE LOADING). -Certain types of compounds with ionizable groups and those which display both lipid and water solubility can be introduced into the liposomes after the formation of intact vesicles(ACTIVE LOADING).

METHODS OF LIPOSOMES PREPARATION

PASSIVE LOADING TECHNIQUES

MECHANICAL DISPERSION SOLVENT DISPERSION DETERGENT DEPLETION

MECHANICAL DISPERSION METHODS

- Co-dissolve the lipids in an organic solvent. -Remove the organic solvent by film deposition under vacuum. -Hydrate the solid lipid mixture using an aqueous buffer. -The lipids spontaneously swell and form liposomes.

THIN FILM HYDRATION USING HAND SHAKING (MLVs) -Lipids casted as stacks of film . -The casted film dispersed in an aqueous medium by hand shaking -Upon hydration, the lipids swell and peel off -MLVs are formed NON-SHAKING METHODS(ULVs) -Expose the film to a stream of water saturated nitrogen for 15 min -Swell in an aqueous medium without hand shaking -ULVs are formed

HAND SHAKING TECHNIQUE

PROPRO-LIPOSOMES -Lipids are dried over a finely divided particulate support (e.g. Sodium chloride or Sorbitol in powder form) -These lipid coated particulates are called pro- liposomes. pro- liposomes. -In water,the support is rapidly dissolved. -The lipid film is hydrated to form MLVs.

MECHANICAL TREATMENTS OF MLVs

MICROMICRO-EMULSIFICATION -Force out an MLV dispersion through a 5 m orifice in a
microfluidizer at very high pressure. -Then separate the fluid into two streams. -Allow the streams to collide together at right angles in an interaction chamber. -Collect SUVs.

MICROMICRO-EMULSIFICATION

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SONICATION -Place test tube containing an MLV dispersion in a bath

sonicator or place the tip of the sonicatorin the test tube in a probe sonicator. -Sonicate for 5-10 min above the transition temperature 5of the constituent lipid. -Centrifuge to remove residual large particles. -Collect a clear SUV dispersion.

BATH/PROBE SONICATION

FRENCH PRESSURE CELL

-Extrude MLVs in a French press under high pressure. -Get ULVs or OLVs.

MEMBRANE EXTRUSION
-Pass an MLV dispersion through a polycarbonate membrane filter . -Get SUV s / LUV s from MLVs.

Dried Reconstituted Vesicles (DRV)

-Freeze and lyophilize empty SUVs. -Re-hydrate them with the aqueous fluid containing Rethe material to be entrapped. -Get ULVs or OLVs.

Freeze Thawed Sonication (FTS) Method

-Freeze a SUV dispersion and thaw it at room temperature for 15 min. -Subject it to a brief sonication cycle -Obtain ULVs.

DRV vs FTS METHOD

SOLVENT DISPERSION METHODS FOR PASSIVE LOADING

-Dissolve lipids in an organic solution -Bring the solution into contact with an aqueous phase containing materials to be entrapped within the liposomes.

Ethanol Injection Injection -Inject an ethanolic lipid solution rapidly

,through a fine needle into an excess of saline. -Collect a SUV dispersion.

Ether Injection
-Inject an ethereal lipid solution very slowly ,through a narrow needle into an aqueous phase, at the temperature of vaporizing the organic solvent. -Collect a LUV dispersion.

SOLVENT DISPERSION METHODS

DOUBLE EMULSION
-Inject organic solution containing water droplets rapidly into hot aqueous solution of Tris -buffer by a 22-gauge hypodermic needle. 22-Evaporate the organic solvent using strong jet of nitrogen , thus forming double emulsion (W/O/W system). -Obtain ULVs.

DOUBLE EMULSION

REVERSE PHASE EVAPORATION VESICLES

-Sonicate a mixture of two phases in a bath sonicator. -Dry down the emulsion to a semisolid gel in a rotary evaporator under reduced pressure. -Collapse a certain proportion of the water droplets by vigorous mechanical shaking using a vortex mixture. -Get MLVs or LUVs.

REVERSE PHASE EVAPORATION

STABLE PLURIMELLAR VESICLES

-Prepare water-in-organic phase dispersion with an water-inexcess of lipid. -Dry it under continued batch sonication with an intermittent stream of nitrogen. -Obtain plurilamellar vesicles-entrapped aqueous vesiclesmedium being located in the compartment in between adjacent lamellae.

DETERGENT DEPLETION METHODS OF PASSIVE LOADING

-Bring the phospholipids intimate contact with the aqueous phase via detergents. -Get micelles containing other participating components known as mixed micelles. -Remove the detergent by dialysis or by column chromatography or by the use of biobeads. -Collect ULVs.

ACTIVE LOADING