Liposomes converted

liposomes

1. Liposomes
Presented by,
Pratiksha C
M pharma 2
nd
SEM
Dept. of pharmaceutics
HSK COP Bagalkot
Facilitated to,
Dr. Anita Desai
HOD and professor
Dept. of pharmaceutics
HSK COP Bagalkot
1

2. •Introduction
•Structure ofliposomes
•Advantages&disadvantages
•Components ofliposome
•Preparation methods ofliposomes
•Characterization ofliposomes
•Applications ofliposomes
•References
2

3. Introduction:
Liposomesareconcentricbilayeredvesiclesinwhichanaqueous
volumeisentirelyenclosedbyamembraneouslipidbilayer
mainlycomposedofnaturalorsyntheticphospholipids.
LiposomeswerefirstproducedinEnglandin1961byAlecD.
Bangham.Thesizeofaliposomerangesfromsome20nmupto
severalmicrometers
3

4. Hydrophillichead
Hydrophobictail
The lipid moecules are usually phospholipids-amphipathic
moieties with a hydrophilic head group and two
hydrophobictails.
Liposome=Phospholipid+
cholesterol
4

5. Advantages of liposomes
1.Provides selective passive targeting to tumor tissues.
(liposomal doxorubicin).
2.Increased efficacy and therapeutic index.
3.Reduction in toxicity of the encapsulated agent.
4.Site avoidance effect (avoids non-target tissues).
5.Improved pharmacokinetic effects.
6.Flexibility to couple with site-specific ligandsto achieve
activetargeting.
5

6. Disadvantages ofliposomes:
Production cost ishigh.
Leakageandfusionofencapsulated drug/
molecules.
Sometimesphospholipid undergoesoxidation
and hydrolysis likereaction.
Short half-life.
Lowsolubility.
6

7. H
2OLayer
PolarLipids
(Phospholipid)
Water Soluble
ingredients
(Drugs,Nutrients
Lipid Soluble
ingredients
(Drugs,Nutrients
&vitamins)
Cross-sectionofliposomes:
&vitamins)
7

8. components ofliposomes:
The structural components ofliposomes
include:
A.Phospholipids
B.cholesterol
8

9. Classification of liposome:
Classification
ofliposome
Structural
parameters
Based on lamelle Composition and
application
9

10. Types of vesicles based onlamella
Lamella:
10

11. Based onstructural parameters
MLV
Multilamellar Large
vesicles (>0.5um)
OLV
oligolamellar vesicles
(>0.1-1.0um)
UV
Unilamellar
Vesicles
MVV
Multivesicular vesicles
(> 1.0UM)
MUV
GUV
>1um
SUV 20-
100nm
LUV
>100nm
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12. Based on
composition &
application
convential
fusogenic
pH
sensitive
cationic
Long
circulatory
immuno
Based on
composition
and
application:
12

13. Passive
loading
technique
Active/remo
te loading
technique
Loadingoftheentrappedagents
before/duringthemanufacture
procedure.
Certaintypesofcompoundswith
ionizable groups & those withboth
lipid&watersolubilitycanbe
Introduced into liposomes after the
formation of intactvesicles.
MethodS of LiposomePreparation
13

14. Methods of liposomepreparation
Passiveloadingtechniques Active loadingtechniques
Mechanicaldispersion
methods
❑LIPID FILMHYDRATION
BY HAND SHAKING,FREEZE
DRYINGORNON HAND
SHAKING
❑MICROEMULSIFICATION
❑SONICATION
❑FRENCH PRESSURECELL
❑MEMBRANEEXTRUSON
❑DRIEDRECONSTITUTEDVESICLES
Solvent dispersion
methods
❑ETHANOLINJECTION
❑ETHERINJECTION
❑DOUBLEEMULSION
❑REVERSEPHASE
VAPOURATIONVESICLES
❑STABLE PLURILAMELLERVESICLES
Detergentremoval
technique
14

15. General Method OfLiposome
Preparation:
2315

16. Liposome
Post Hydration vortexing, sonication, freeze thawing&
high pressure extrusion
Lipid dissolve in organicsolvent/co-solvent
Remove organic solvent undervacuum
Filmdeposition
Solidlipidmixture is hydrated by using aqueous buffer
Lipid spontaneously swell &Hydrate
1. Mechanical dispersionmethod:
2416

17. There are four basic methods of physical/mechanical
dispersion:
Handshakenmethod.
Nonshakingmethod.
Pro–liposomes.
Freezedrying.
17

18. Lipidsformstacksoffilm
fromorganicsolution
(FE/HS)
Then film is treatedwith
aqueousmedium
Uponhydrationlipids
swellandpeeloutfrom
RBflask
vesiculatetoformMulti
lamellarvesicles(MLVs)
18

19. Pro-liposomes:
✓Toincrease the surface area of dried lipid film &
to facilitate instantaneoushydration.
lipidDried
over
lipid
Finelydivided
particulate support
like powderedNACL/
sorbital
Pro -liposomes
Pro-
liposomes
water
Dispersion ofMLV’S
✓This Method overcome the stabilityproblem.
2719

20. Membrane extrusion Liposomes
Dried reconstituted vesicles(DRVs)
Freeze thaw sonification(FTS)
pH induced vesiculation
Cochleatemethod.
Processing of the lipids hydrated by physical means or the
mechanical treatments of MLVs:
Micro Emulsification liposomes (MEL)
Sonicated unilamellar vesicles (SUVs)
French Pressure Cell Liposomes.
2820

21. Sonicated unilamellarvesicles:
TheexposureofMLVstoultrasonic
irradation for producing smallvesicles.
Bathsonicator
largevolume
of dilutelipids
Probe sonicator
Used fordispersions
requirehigh
energyin
smallvolumes
Sonication
MLVs hazytransparent
5-10minsolution
centrifugation 30min
clear SUV
Dispersion.
2921

22. Micro emulsificationliposomes:
Microfluidizer
3022

23. French pressure cellliposomes:
Extrusion of preformed large liposomes in french press undervery
high pressure.
uni or oligo lamellar liposomes of intermediate size (30-80nm ).
Advantages
Lessleakageand more stable liposomes
are formed comparedto
sonicatedforms
3123

24. Vesicles prepared by extrusion technique:
Thesizeofliposomesis
reducedbygentlypassingthem
through polycarbonate
membranefilterofdefined
poresizeatlowerpressure
UsedforpreparationofLUVs
andMLVs
24

25. Dried reconstituted vesicles& freeze thaw sonicationmethod
25

26. pH inducedvesiculation:
Preformed
MLV’S
(2.5-3.0)
~ (addition of1M
NaoH)
~Period of
exposure <2min
Reduced the pH
to7.5
Exposed tohighpH* Additionof
0.1MHcl
ThetransientchangeinpHbringsabout
anincreaseinsurfacechargeofthelipid
bilayerwhichinducesspontaneous
vesiculation.
MLVs
LUVs
26

27. SUVs made
from
phosphatidylse
rine(PS)
Additionof
Ca++ions
Cylindrical
rolls(cochleate
cylinders)
Removal
of Ca++by
EDTA
Cochleatemethod:
Cochleates
27

28. Lipid dissolve in organicsolvent
Excess addition of aqueousphase
Lipids allign at interface of aqueous and organic layer
Formation of monolayer and bilayer ofphospholipids
Liposome
Note:-Organic solvent miscible with aqueousphase
Solvent dispersionmethods:
28

29. Solvent dispersion methods:
ETHANOL INJECTION/ETHERINJECTION:
29

30. De-Emulsificationmethod:
Generally the liposome is made up in 2steps:
1 st the inner leaflet of the bilayer.
Then the outerhalf.
~Reverse phase evaporationvesicles
~Sonicationmethods
Aqueous medium
containingmaterial
to beentrapped
Add toimmiscible
organic solutionof
lipid
Mechanical agitation
Microscopicwater
droplets
Methods to prepare thedroplets:
~Double emulsionvesicles
30

31. Reverse phase evapourationmethod:
31

32. Phospholipid brought into intimate contact with
aqueousphase
By addition optimized concentration ofdetergent
Formation of micelles(Liposome)
shapedependonchemical
nature of
detergent, concentrationand
other lipidinvolved
Below CMC, detergent molecules exist in free soln. Asthe
concentration is increased, micelles areformed.
Note:-Liposomesizeand
DETERGENT SOLUBILISATIOINMETHODS
Methods to removedetergents:
Dialysis
Columnchromatography.
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33. Active/remote loadingtechnique:
Thelipidbilayermembraneisimpermeabletoions&hydrophilic
molecules.But,Permeationofhydrophobicmoleculescanbecontrolled
byconcentrationgradients.
Someweakacidsorbasescanbetransportedduetovarious
transmembranegradients
Electrical gradients.
Ionic(pH)gradients.
Chemical potentialgradients.
Weak amphipathic bases accumulate in aq phase of lipid vesicles
in response to difference in pH b/w Inside & outsideof
liposomes
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34. pH gradient is created by preparingliposomes
with low internalpH.
Addtn of base to extraliposomalmedium.
[Basic compds ( lipophilic (non ionic) athigh
pH & hydrophilic(ionic) at lowpH)]
Lipophilic (UNPROTONATED) drug diffuse through
thebilayer
At low pH side, the moleculesare
predominantly protonated.
Exchange of external medium by gelextrusion
chromatorapghy with neutralsolution.
Weak bases like doxorubicine,
adriamycin and vincristineare
encapsulated.
Solute bearingno
charge at neutralpH
Liposomes withlow
internalpH
Neutral solute passes
easily through bilayer
membrane by diffusion
Charge aquired by solute
inside liposomes makes
them unable toexit
34

35. Characterization ofliposomes:
PHYSICALCHARACTERISATION
→ Vesiclessize/shape/morphology
→ Surface -charge/electricalpotential
→ Phase behaviour/lamellarity
→ Drugrelease
→ % capture /freedrug
CHEMICA LCHARACTERISATION
→ Phospholipids /lipidconcentration
→ Drugconcentration
→ PH /Osmomolality
→Antioxidantdegradation
→ Phospholipids / cholesterols–
peroxidation/oxidation/hydrolysis
BIOLOGICALCHARACTERISATION
→Sterility
→Pyrogenisity
→ Animal toxicity
→PlasmaStability:
35

36. CharacterizationparametersAnalyticalmethod/Instrument
1. Vesicle shape and surfacemorphology Transmission electron microscopy,Freeze-
fracture electronmicroscopy
2.Mean vesicle size and size distribution
(submicron and micronrange)
Photon correlation spectroscopy, laserlight
scattering, gel permeation and gelexclusion
3. Surfacecharge Free-flowelectrophoresis
4. Electrical surface potential and surfacepHZetapotentialmeasurements
5.Lamellarity Small angle X-ray scattering, 31 P-NMR,Freeze-
fracture electronmicroscopy
6. Phase behaviorFreeze-fracture electron microscopy, Differentialscanning
calorimetery
7. Percent of free drug/ percentcapture Minicolumn centrifugation, ion-exchange
chromatography, radiolabelling
8. Drugrelease
Diffusion cell/dialysis
36

37. CharacterizationparametersAnalyticalmethod/Instrument
1. PhospholipidconcentrationBarlett assay, stewart assay,HPLC
2. CholesterolconcentrationCholesterol oxidase assay andHPLC
3. PhopholipidperoxidationUVabsorbance
4. Phospholipid hydrolysis,
Cholesterolauto-oxidation.
HPLC andTLC
5.Osmolarity Osmometer
37

38. CharacterizationparametersAnalyticalmethod/Instrument
1.Sterility Aerobic or anaerobiccultures
2.Pyrogenicity Limulus Amebocyte Lysate (LAL)test
3. Animal toxicity Monitoring survival rates, histologyand
pathology
STABILITY OFLIPOSOMES:
❖Stability invitro.
~ Lipidoxidation
~ Lipidperoxidation
~ Long term & acceleratedstability
❖Stability after systemicadministration.
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39. In genedelivery.
As drug delivery carriers.
Enzyme replacementtherapy.
Chelation therapy for treatment of heavy metal poisoning.
Liposomes in antiviral/anti microbialtherapy.
In multi drugresistance.
In tumourtherapy.
Inimmunology.
Incosmetology
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40. DRUG ROUTEOF
ADMINISTRATION
APPLICATION TARGETEDDISEASES
AmphotericinBOraldelivery Ergosterolmembrane Mycoticinfection
Insulin Oral,ocular,pulmonary
Andtransdermal
Decrease glucoselevel Diabeticmellitus
Ketoprofen Oculardelivary Cyclooxygenase enzymeinhibitorPain musclecondition
PentoxyfyllinPulmonarydelivery phosphodiesterase Asthama
Tobramycin Pulmonarydelivery Protein synthesisinhibitor Pseudomonas
infection,aeroginosa
Salbutamol Pulmonarydelivery ß
2-adrenoceptorantagonist Asthama
Cytarabin Pulmonarydelivery DNA-polymeraseinhibition Acuteleukameias
Benzocaine Transdermal Inhibition of nerve impulsefrom
sensorynerves
Ulcer on mucoussurface
withpain
Ketaconazole Transdermal Inhibit ergosterolmembrane Candidaalbicans
LevanogesterolTransdermal Rhamnosereceptor skindisorder
hydroxyzine Transdermal H1-receptorantagonist Urtecaria,allergicskin
disease
Ibuprofen Oraldelivery Chaemoceptor,freeending Rheumatoidarthritis
triamcilononeOculardelivery,TransdermalInhibition ofprostaglandin Anti-inflammatory
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41. NAME TRADENAME COMPANY INDICATION
Liposomal
amphotericinB
Abelcet Enzon Fungalinfections
Liposomal
amphotericinB
Ambisome GileadSciences Fungal and protozoalinfections
LiposomalcytarabineDepocyt Pacira(formerly
SkyePharma)
Malignant lymphomatousmeningitis
Liposomal
daunorubicin
DaunoXome GileadSciences HIV-related Kaposi’ssarcoma
LiposomaldoxorubicinMyocet Zeneus Combination therapy with cyclophosphamide in
metastatic breastcancer
Liposomal IRIVvaccineEpaxal BernaBiotech HepatitisA
Liposomal IRIVvaccineInflexalV BernaBiotech Influenza
Liposomalmorphine DepoDur SkyePharma,Endo Postsurgicalanalgesia
LiposomalverteporfinVisudyne QLT,Novartis Age-related macular degeneration,pathologic
myopia,ocular
histoplasmosis
Liposome-PEG
doxorubicin
Doxil/CaelyxOrtho Biotech,
Schering-Plough
HIV-related Kaposi’s sarcoma, metastaticbreast
cancer,metastatic
ovariancancer
Micellularestradiol Estrasorb Novavax Menopausaltherapy
41

42. 1.S.P.VyasAndR.K.Khar,targeted&ControlledDrug
Delivery,liposomes,173-279.
2.MohammadRiaz,Liposomes:PreparationMethods,Pakistan
JournalOfPharmaceuticalSciences,January1996,Vol.19(1),65-
77.
3.SharmaVijayK1*,Liposomes:PresentProspectiveandFuture
Challenges,InternationalJournalOfCurrentPharmaceutical
ReviewAndResearch,oct2010,vol1,issue2,6-16
4.Himanshu Anwekar*, Liposome-as drug carriers,
International Journal Of Pharmacy & LifeSciences,Vol.2,
Issue 7: July: 2011,945-951.
5.Jain NK. Introduction to novel drug delivery systems.1
st
edition. VallabhPrakashanpublishers;2010.
6.Dr. BaviskarDT, Dr. Jain DK. Novel drug delivery systems.
2
nd
edition. NiraliPrakashanpublishers;2015.
7.Images are from google.
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