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Presented by;
AITHA SWETHA
M.PHARMACY 1ST YEAR 2ND SEM
• Introduction
• Structure of liposomes
• Advantages& disadvantages
• Components of liposome
• Mechanism of liposome
• Preparation methods of liposomes
• Characterization of liposomes
• Applications of liposomes
• Summary
• Niosomes Introduction
• Advantages& disadvantages
• Preparation methods of niosomes
• Characterisation of niosomes
• summary
• References
LIPOSOMES

liposomes are concentric bilayered vesicles in which an aqueous
volume is entirely enclosed by a membraneous lipid bilayer
mainly composed of natural or synthetic phospholipids.


   Liposomes were first produced in England in 1961 by
   Alec D. Bangham. The size of a liposome ranges from some
   20 nm up to several micrometers

                                                              1
Liposome =Phospholipid+
                                            cholesterol




                                             Hydrophillic head

                                             Hydrophobic tail

The lipid moecules are usually phospholipids-amphipathic

moieties with a hydrophilic head group and two hydrophobic tails.
                                                                 2
Advantages of
     liposomes:
Provides selective passive targeting to tumor tissues.
(liposomal doxorubicin) .
Increased efficacy and therapeutic index.
Reduction in toxicity of the encapsulated agent.
Site avoidance effect (avoids non-target tissues).
Improved pharmacokinetic effects .
Flexibility to couple with site-specific ligands to achieve
active targeting.
                                                              3
Disadvantages of liposomes:
  Production cost is high.
  Leakage and fusion of encapsulated drug /
  molecules.
  Sometimes phospholipid undergoes oxidation
  and hydrolysis like reaction.
  Short half-life.
  Low solubility.
                                               4
Cross-section of liposomes:

                                     Polar Lipids
                                   (Phospholipid)




                                    Lipid Soluble
                                    ingredients
                                    (Drugs,Nutrients
                                    & vitamins)
         H2O Layer   Water Soluble
                     ingredients
                     (Drugs, Nutrients              5
                            & vitamins)
components of liposomes:

 The structural components of liposomes
 include:
            A. Phospholipids
            B. cholesterol


                                          6
A. General representation of
   phospholipids:




                               7
Phospholipids

 Phosphatidylcholine- natural
 Amphipathic molecule
Hydrophilic polar head-
 Phosphoric acid bound to water
    soluble molecule.
Glyceryl bridge
Hydrophobic tail-
 2 fatty acid chain containing 10-24 carbon
 atoms and 0-6 double bond in each chain.

 The amphipathic molecule self organise
 in ordered supramolecular structure when
   confronted (meet face to face)
   with solvent.
                                               8
The most common natural phospholipid is the
phospatidylcholine (PC ).
                                               Polar Head Groups
Naturally occurring phospholipids used are :
   PC: Phosphatidylcholine.
   PE: Phosphatidylethanolamine.
                                               Three carbon glycerol
   PS: Phosphatidylserine
Synthetic phospholipids used are:
   DOPC: Dioleoyl phosphatidylcholine
   DSPC: Disteroyl phosphatidylcholine
   DOPE: Dioleoyl phosphatidylethanolamine
   DSPE: Distearoyl phosphatidylethanolamine
                                                                9
Molecular geometry on structure of amphiphillic aggregates:




                                                              10
Molecules of PC are not soluble in water.
In aqueous media they align themselves closely in planar bilayer sheets
in order to minimize the unfavorable 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




                                                                      11
PHASE TRANSITION
TEMPERATURE phospholipid membranes can exist
 At various temperatures,
in different phases.
The transition from one phase to another can be detected by
technique like micro calorimetry .
What exactly happens during phase transition?

 Tightly ordered At elevated temperature liquid crystal phase
     gel state
 ( lipid membrane)                         (movement is higher)
This is due to the fatty acid chain adopting a new
conformation other than all trans straight chain configuration,
         such as gauche configuration state( phenomenon- chain
          tilt )
                                                              12
B. Cholesterol:
 Cholesterol stabilizes the Membrane
  Steroid lipid
  Interdigitates between phospholipids.
 i.e. below Tc , it makes membrane less ordered & above Tc more ordered.




      Being an amphipathic molecule, cholesterol inserts into the
      membrane with its hydroxyl group of cholesterol oriented
      towards the aqueous surface and aliphatic chain aligned parallel to
      the acyl chains in the center of the bilayer .                   13
Role of cholesterol in bilayer
         formation:
Cholesterol act as fluidity buffer
After intercalation with phospholipid molecules alter the
freedom of motion of carbon molecules in the acyl
Chain
Restricts the transformations of trans to gauche
Conformations.
Incorporated into phospholipid membrane upto 1:1 or
2:1 of cholesterol to PC.


                                                       14
Mechanism of liposome formation:




                                   15
Classification of liposome :


                    Classification
                     of liposome




   Structural         Method of      Composition
   parameters         preparation    and application



                                                   16
Lamella :




  Types of vesicles based on lamella
                                       17
A. Structural
         parameters:
                   Based on structural
                       parameters

     MLV              OLV                    UV             MVV
 Multilamellar   oligolamellar                         Multivesicular
                                         Unilamellar
Large vesicles      vesicles                             vesicles
  (>0.5 um)       (>0.1-1.0 um)           Vesicles          (> 1.0 UM)


                                                   SUV
                                  MUV            20-100nm
                                  GUV                LUV
                                  >1um             >100nm                18
B. Based on         REV, SUV made
method of            by reverse
                        phase
preparation:         evaporation
                       method



          VET
                                        SPLV
        Vesicles     Based on
                     method of         Stable
      prepared by
                    preparation     plurilamenar
       extrusion
                                      vesicles
         tech.



                      FATMLV
                      Frozen &
                    thawed MLV                     19
Based on
composition
and                     convential
application:

           immuno                       fusogenic
                          Based on
                        composition
                        & application
             Long                          pH
          circulatory                   sensitive

                          cationic
                                                    20
MethodS of Liposome Preparation


        Passive
                       Loading of the entrapped agents
        loading        before/ during the manufacture
       technique       procedure.




      Active/remo      Certain types of compounds with
                       ionizable groups & those with both
       te loading      lipid & water solubility can be
       technique       Introduced into liposomes after the
                       formation of intact vesicles.



                                                             21
Methods of liposome preparation

     Passive loading techniques             Active loading techniques

 Mechanical dispersion            Solvent dispersion        Detergent removal
        methods                     methods                       technique
LIPID FILM HYDRATION        ETHANOL INJECTION            DETERGENT REMOVAL
 BY HAND SHAKING,FREEZE
 DRYING OR NON HAND           ETHER INJECTION               FORM MIXED MICELLES
 SHAKING
                              DOUBLE EMULSION               BY DIALYSIS
MICRO EMULSIFICATION
                             REVERSE PHASE                CHROMATIGRALPY
SONICATION
                                   VAPOURATION VESICLES   DIFFUSION
FRENCH PRESSURE CELL
                             STABLE PLURI LAMELLER         VESICLES LIKE….
MEMBRANE EXTRUSON                                                                 22
                              VESICLES                     RECONSTITUTED &
DRIED RECONSTITUTED
 VESICLES                                                  SANDAI VIRUS ENVELOPE
General Method Of Liposome
Preparation:




                             23
1. Mechanical dispersion method:
          Lipid dissolve in organic solvent/co-solvent

            Remove organic solvent under vacuum

                        Film deposition

    Solid lipid mixture is hydrated by using aqueous buffer

             Lipid spontaneously swell & Hydrate

                         Liposome
   Post Hydration vortexing, sonication, freeze thawing &
                  high pressure extrusion                     24
There are four basic methods of physical/mechanical
dispersion :
    Hand shaken method.
    Non shaking method.
    Pro – liposomes .
    Freeze drying .

                                                25
Lipidsform stacks of film
from organic solution
        (FE/HS)
Then film is treated with
aqueous medium

Upon hydration lipids
swell and peel out from
RB flask

vesiculate to form Multi
lamellar vesicles(MLVs)


                       26
Pro-liposomes:
    To increase the surface area of dried lipid film & to
      facilitate instantaneous hydration.

                  lipid Dried
                  over
                                Finely divided
     lipid                   particulate support    Pro - liposomes
                            like powdered NACL/
                                   sorbital

         Pro-                                Dispersion of MLV’S
                                water
      liposomes

      This Method overcome the stability problem.                 27
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 .
         Membrane extrusion Liposomes
          Dried reconstituted vesicles(DRVs)
          Freeze thaw sonification (FTS)
         pH induced vesiculation
         Cochleate method.                                   28
Sonicated unilamellar vesicles:
The exposure of MLVs to ultrasonic
irradation for producing small vesicles.



  Probe sonicator        Bath sonicator
Used for dispersions      large volume
 require high              of dilute lipids
energy in
small volumes
                       Sonication
             MLVs               hazy transparent
                        5-10 min solution

                             centrifugation 30 min

                                        clear SUV    29
                                       Dispersion.
Micro emulsification liposomes:




                                                    30
                                  Micro fluidizer
French pressure cell liposomes:

Extrusion of preformed large liposomes in french press under very
high pressure .

 uni or oligo lamellar liposomes of intermediate size (30-80nm ) .

Advantages
Less leakage and more stable liposomes are formed compared to
sonicated forms




                                                                     31
Vesicles prepared by extrusion technique :
 The size of liposomes is
 reduced by gently passing them
 through           polycarbonate
 membrane filter of defined
 pore size at lower pressure

 Used for preparation of LUVs
 and MLVs




                                             32
Dried reconstituted vesicles& freeze thaw sonication method




                                                     33
pH induced vesiculation:
The transient change in pH brings about
an increase in surface charge of the lipid
bilayer which induces spontaneous                             LUVs
vesiculation .

                                                 Reduced the pH
                                                 to 7.5
                              Exposed to high pH * Addition of
                              ~ (addition of 1M    0.1M Hcl
                  Preformed      NaoH)
                  MLV’S
                              ~Period of
                  (2.5-3.0)
                               exposure < 2min
     MLVs                                                         34
Cochleate method:


                                                           Cochleates

                                                  Removal
                                                  of Ca++ by
                                Cylindrical       EDTA
                                rolls(cochleate
                    Addition of cylinders)
                    Ca++ ions
        SUVs made
        from
        phosphatidylse
        rine(PS)
                                                                35
Solvent dispersion methods:
                Lipid dissolve in organic solvent


               Excess addition of aqueous phase


     Lipids allign at interface of aqueous and organic layer

     Formation of monolayer and bilayer of phospholipids


                        Liposome
    Note:- Organic solvent miscible with aqueous phase
                                                               36
Solvent dispersion methods:
 ETHANOL INJECTION/ETHER INJECTION:




                                      37
De-Emulsification method:

 Generally the liposome is made up in 2 steps:           Aqueous medium
                                                        containing material
 1 st the inner leaflet of the bilayer .                 to be entrapped
 Then the outer half.
                                                        Add to immiscible
                                                        organic solution of
                                                               lipid

                                                    Mechanical agitation

                                                        Microscopic water
                                                             droplets
            Methods to prepare the droplets:
                  ~Double emulsion vesicles
                  ~Reverse phase evaporation vesicles                   38
                  ~Sonication methods
Reverse phase evapouration method:




                                     39
DETERGENT SOLUBILISATIOIN METHODS

      Phospholipid brought into intimate contact with
                      aqueous phase

      By addition optimized concentration of detergent


               Formation of micelles (Liposome)

Below CMC, detergent molecules exist in free soln. As the
concentration is increased, micelles are formed.
    Note:- Liposome size and
                                         Methods to remove detergents:
    shape depend on chemical
                                              Dialysis
    nature                      of
                                              Column chromatography.
    detergent, concentration and                                     40
    other lipid involved
Active/remote loading technique:
  The lipid bilayer membrane is impermeable to ions & hydrophilic
  molecules. But, Permeation of hydrophobic molecules can be controlled
  by concentration gradients.
  Some weak acids or bases can be transported due to various
  transmembrane gradients
            Electrical gradients.
            Ionic(pH) gradients.
            Chemical potential gradients.
        Weak amphipathic bases accumulate in aq phase of lipid vesicles
        in response to difference in pH b/w Inside & outside of
        liposomes                                                 41
Solute bearing no               Liposomes with low
                                internal pH
charge at neutral pH                             pH gradient is created by preparing liposomes
                                                 with low internal pH.

                                                 Addtn of base to extraliposomal medium.

                                                 [Basic compds ( lipophilic (non ionic) at high
                                                 pH & hydrophilic(ionic) at low pH)]

           Neutral solute passes                 Lipophilic (UNPROTONATED) drug diffuse
           easily through bilayer                through the bilayer
           membrane by
           diffusion
                                                 At low pH side, the molecules are
                                                 predominantly protonated .

                                                 Exchange of external medium by gel extrusion
                                                 chromatorapghy with neutral solution.
                          Charge aquired by
                          solute inside               Weak bases like doxorubicine,
                          liposomes makes             adriamycin and vincristine are
                          them unable to exit         encapsulated.               42
Locus of drugs in liposomes:
                               Hydrophilic (DOXORUBICIN)
                               Low entrapment
                               Leakage
                               Hydrolytic degradation

                               Lipophilic (CYCLOSPORINE)
                               High entrapment
                               Low leakage
                               Chemical stability

                               Ampiphilic (VINBLASTIN)
                               High entrapment
                                Rapid leakage
                               Biphasic insoluble
                               (ALLOPURINOL, 6-
                               MERCAPTOPURINE)
                                Poor loading & entrapment 43
Characterization of liposomes:
PHYSICAL CHARACTERISATION            CHEMICA L CHARACTERISATION

→ Vesicles size/shape/morphology     → Phospholipids /lipid concentration

→ Surface -charge/electrical potential → Drug concentration
→ Phase behaviour/ lamellarity       → PH / Osmomolality

→ Drug release                       →Antioxidant degradation

→ % capture /free drug               → Phospholipids / cholesterols –
                                        peroxidation/oxidation/hydrolysis
            BIOLOGICAL CHARACTERISATION
            → Sterility
            → Pyrogenisity
            → Animal toxicity
                                                                  44
            →Plasma Stability:
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        Photon correlation spectroscopy, laser light
(submicron and micron range)                     scattering, gel permeation and gel exclusion

3. Surface charge                                Free-flow electrophoresis

4. Electrical surface potential and surface pH   Zetapotential measurements

5. Lamellarity                                   Small angle X-ray scattering, 31 P-NMR, Freeze-
                                                 fracture electron microscopy
6. Phase behavior Freeze-fracture                electron microscopy, Differential scanning
                                                 calorimetery
7. Percent of free drug/ percent capture         Minicolumn centrifugation, ion-exchange
                                                 chromatography, radio labelling
8. Drug release                                  Diffusion cell/ dialysis
                                                                                           45
Characterization parameters      Analytical method/Instrument

1. Phospholipid concentration    Barlett assay, stewart assay, HPLC

2. Cholesterol concentration     Cholesterol oxidase assay and HPLC

3. Phopholipid peroxidation      UV absorbance

4. Phospholipid hydrolysis,      HPLC and TLC
   Cholesterol auto-oxidation.

5. Osmolarity                    Osmomete


                                                                46
Characterization parameters           Analytical method/Instrument

1. Sterility                            Aerobic or anaerobic cultures
2. Pyrogenicity                         Limulus Amebocyte Lysate (LAL) test

3. Animal toxicity                      Monitoring survival rates, histology and
                                        pathology

           STABILITY OF LIPOSOMES:
           Stability invitro .
              ~ Lipid oxidation
              ~ Lipid peroxidation
              ~ Long term & accelerated stability
           Stability after systemic administration.
                                                                            47
1. Endocytosis     2. Adsorption




 3. fusion       4. Lipid transfer




                                     48
Encapsulation of drugs in liposomes:
• Encapsulation volume/Trapped volume
  Volume of aqueous solution entrapped in liposomes per mole of PL (µL/µmol PL)
• Encapsulation Efficiency
  Assessed by mini column centrifugation method & protamine aggregation method.
  protamine aggregation method used for neutral and negetively charged liposomes.
  Liposome dispersion can be precipitated with protamine solution and subsequent
  centrifugation at 2000RPM.
  By analysing the material in super natent & in liposome pellet ( after disrupting
  liposomal pellet with 0.6 ml of 10% triton x-100 ). The encapsulation efficiency of
  entrapped material can be estimated.
• % Encapsulation

   Drug entrapped in liposomes
                                   x 100
         Total drug added

                                                                             49
In gene delivery.
As drug delivery carriers.
Enzyme replacement therapy.
Chelation therapy for treatment of heavy metal poisoning.
Liposomes in antiviral/anti microbial therapy.
In multi drug resistance.
In tumour therapy.
In immunology.
In cosmetology

                                                      50
DNA delivery of Genes by Liposomes


   Cheaper than viruses


   No immune response



          Especially good
for in-lung delivery (cystic fibrosis)


          100-1000 times more plasmid DNA needed
       for the same transfer efficiency as for viral vector   51
Lipofection




              52
Liposomes could serve as tumor specific vehicles
       (even without special targeting)




    Liposomes better penetrate into tissues
       with disrupted endothelial lining           53
DRUG             ROUTE OF                      APPLICATION                        TARGETED DISEASES
                 ADMINISTRATION
Amphotericin B   Oral delivery                 Ergosterol membrane                Mycotic infection
Insulin          Oral,ocular,pulmonary         Decrease glucose level             Diabetic mellitus
                 And transdermal
Ketoprofen       Ocular delivary               Cyclooxygenase enzyme inhibitor    Pain muscle condition

Pentoxyfyllin    Pulmonary delivery            phosphodiesterase                  Asthama
Tobramycin       Pulmonary delivery            Protein synthesis inhibitor        Pseudomonas
                                                                                  infection,aeroginosa
Salbutamol       Pulmonary delivery            ß2-adrenoceptor antagonist         Asthama
Cytarabin        Pulmonary delivery            DNA-polymerase inhibition          Acute leukameias

Benzocaine       Transdermal                   Inhibition of nerve impulse from   Ulcer on mucous surface
                                               sensory nerves                     with pain

Ketaconazole     Transdermal                   Inhibit ergosterol membrane        Candida albicans

Levanogesterol   Transdermal                   Rhamnose receptor                  skin disorder
hydroxyzine      Transdermal                   H1-receptor antagonist             Urtecaria,allergic skin
                                                                                  disease
Ibuprofen        Oral delivery                 Chaemoceptor,free ending           Rheumatoid arthritis

triamcilonone    Ocular delivery,Transdermal   Inhibition of prostaglandin        Anti-inflammatory

                                                                                                  54
NAME                     TRADE NAME     COMPANY            INDICATION
Liposomal                Abelcet        Enzon              Fungal infections
amphotericin B
Liposomal                Ambisome       Gilead Sciences    Fungal and protozoal infections
amphotericin B
Liposomal cytarabine     Depocyt        Pacira (formerly   Malignant lymphomatous meningitis
                                        SkyePharma)
Liposomal                DaunoXome      Gilead Sciences    HIV-related Kaposi’s sarcoma
daunorubicin
Liposomal doxorubicin    Myocet         Zeneus             Combination therapy with cyclophosphamide in
                                                           metastatic breast cancer
Liposomal IRIV vaccine   Epaxal         Berna Biotech      Hepatitis A

Liposomal IRIV vaccine   Inflexal V     Berna Biotech      Influenza

Liposomal morphine       DepoDur        SkyePharma, Endo   Postsurgical analgesia
Liposomal verteporfin    Visudyne       QLT, Novartis      Age-related macular degeneration, pathologic
                                                           myopia, ocular
                                                           histoplasmosis
Liposome-PEG             Doxil/Caelyx   Ortho Biotech,     HIV-related Kaposi’s sarcoma, metastatic breast
doxorubicin                             Schering-Plough    cancer, metastatic
                                                           ovarian cancer
                                                                                                  55
Micellular estradiol     Estrasorb      Novavax            Menopausal therapy
summary:
 o liposomes are concentric bilayered vesicles in which an aqueous
   volume is entirely enclosed by a membraneous lipid bilayer
 o Liposomes are one of the unique drug delivery system, in controlling
   and targeting drug delivery.
 o Components of liposomes include phospholipid and cholesterol.
 o Method of preparation of liposomes include active loading technique
   and passive loading technique.
 o Passive loading techniques include solvent mechanical dispersion,
   solvent dispersion & detergent solubilisation
 o Characterization of liposomes include physical,chemical and
                                                                       56
   biological.
NIOSOMES
Niosomes are non-ionic surfactant based unilamellar or multilamellar
bilayer vesicles up on hydration of non ionic surfactants with or
without incorporation cholesterol .
The niosomes are very small, and microscopic in size. Their size lies
in the nanometric scale.
Niosomes are a novel drug delivery system, in which the medication is
encapsulated in a vesicle. Both hydrophilic
& lipophilic drugs ,entrap either in the
aqueous layer or in vesicular membrane
made of lipid materials.
                                                                 57
Hydrophilic drugs      Polar heads facing
Structure of niosomes:              located in             hydrophilic region
                                    aqueous regions
                   Head part        encapsulated
                   (hydrophillic)
                    Tail part
                    (hydrophobic)
                  Drug molecules
                                                        Hydrophobic drugs
                                                        localized in the
                             Phospholipids              hydrophobic
                                                        lamellae
These vesicular systems are similar to liposomes that can be
 used as carriers of amphiphilic and lipophilic drugs.

It is less toxic and improves the therapeutic index of drug by
 restricting its action to target cells.                      58
Advantages of niosomes:
  They are osmotically active and stable.
  They increase the stability of the entrapped drug.
  The vesicle suspension being water based offers greater patient
  compliance over oil based systems
  Since the structure of the niosome offers place to accommodate
  hydrophilic, lipophilic as well as ampiphilic drug moieties, they can be
  used for a variety of drugs.
  The vesicles can act as a depot to release the drug slowly and of
  controlled release.
  Biodegradable, non-immunogenic and biocompatible.                   59
Aggregation
Fusion
Leaking of entrapped drug
Hydrolysis of encapsulated drugs which limiting the shelf
life of the dispersion.



                                                        60
Classification of niosomes




  Small                   Large
  Unilamellar             Unilamellar              Multilamellar
  Vesicle                 Vesicle                  Vesicle
  (SUV)                   (LUV)                    (MLV)

   Typical Size Ranges: SLV: 20-50 nm – MLV:100-1000 nm


                                                                   61
Components of niosomes:

 Cholesterol and Non ionic surfactants are the two major components
 used for the preparation of niosomes.
 Cholesterol provides rigidity and proper shape. The surfactants play a
 major role in the formation of niosomes.
 non-ionic surfactants like spans(span 20,40,60,85,80), tweens (tween
 20,40,60,80) are generally used for the preparation of
 Niosomes.
  Few other surfactants that are reported to form niosomes are as follows :
                          Ether linked surfactant
                          Di-alkyl chain surfactant
                          Ester linked
                          Sorbitan Esters
                          Poly-sorbates
                                                                   62
alkyl group chain
                                                                       length : C12-C18
Shud be above                                                          Span surfactants
the gel to liquid                                                      with HLB values
phase transition                                                       4 and 8
temperature of               Hydration                   Non-ionic
the system                  Temperature                  surfactant
                                                           nature


                                              Factors
                                             affecting
                   Surfactants              niosomes
                    and lipid                                     Membrane
                                            formation             additives
                     levels

surfactant/lipid
ratio: 10-30 mM                             Nature of                 Cholesterol: Prevent
                                          encapsulated                vesicle aggregation.
                                              drug                    Dicetyl phosphate: -ve
                                                                      charge
                                                                                 63
Concept of Critical Packing Parameter

  Prediction of vesicle forming ability is not a simply a matter of HLB
          CPP = v/lca0
  where
          v - hydrophobic group volume,
          lc - critical hydrophobic group length and
          a0 - area of the hydrophilic head group

  CPP between 0.5 and 1 likely to form vesicles.

  < 0.5 (indicating a large contribution from the hydrophilic head group
  area) is said to give spherical micelles.

  >1 (indicating a large contribution from the hydrophobic group volume)
  should produce inverted micelles.
                                                                   64
Comparisition between liposomes &
niosomes:
Sl.              Liposomes                           Niosomes
No.
1.    Vesicles made up of concentric    Vesicles made up of surfactants
      bilayer of phospholipids          with or without incorporation of
                                        cholesterol.
2.    Size ranges from 10-3000nm        Size ranges from 10-100nm

3.    Comparatively expensive           Inexpensive

4.    Special storage condition are     No such special requirement
      required
5.    Phospholipids used are unstable   Non-ionic surfactants are stable

6.    Comparatively more toxic          Less toxic


                                                                       65
Methods of Niosome preparation:
  Hand Shaking method

   Reverse phase evaporation technique

   Ether Injection method

  Multiple membrane extrusion method

  Bubble method

   Sonication

   From Proniosomes
                                          66
Hand shaking method:




                       Rotary evaporator




                                   67
Reverse phase evaporation technique :
 Surfactant is dissolved in chloroform ond 0.25 volume of PBS buffer is
 emulsified to get a W/O emulsion.
                             sonicated
  chloroform is evaporated under reduced pressure.

 The lipid or surfactant forms a gel first and hydrates to form vesicles.

  Free drug (unentrapped) is generally removed by dialysis.

  sonication:
        Surfactant +cholesterol                  Mixture is sonicated for 3
      mixture is dispersed in 2 ml              min at 60 C using titanium
         aqueous phase in vial                        probe sonicator


                                                   Unilamellar niosomes       68
14 guage needle
Ether injection Method:




                                       69
Multiple membrane extrusion Method:
                 •Mixture of surfactant, cholesterol and
                 dicetyl phosphate in chloroform is made
                 into thin film by evaporation




                •The film is hydrated with aqueous drug
                solution and the resultant suspension
                extruded through polycarbonate membranes



                                                           70
Bubble method:
                                 RBF as bubbling unit with three necks in water
It is novel technique for the                         bath.

one step preparation of
liposomes and niosomes           Reflux , thermometer and nitrogen supply by
                                                 three necks
without the use of organic
solvents.                         Cholesterol+ Surfactant dispersed in buffer
                                                pH 7.4 at 70°C


                                Above dispersion is homogenized for 15 sec and
                                 then bubbled with nitrogen gas at 70°C to get
                                                   niosomes
                                                                                  71
proniosomes:

 • Bubble Method
 • Formation of niosomes from proniosomes:
 It is prepared by coating water-soluble carrier such as sorbitol with
 surfactant. The result of the coating process is a dry formulation. In
 which each water-soluble particle is covered with a thin film of dry
 surfactant. This preparation is termed “Proniosomes”.




                                                                    72
Separation of unentrapped drug:



      Gel filtration                 Separation of
                                     unentrapped                    Centrifugation
                                         drug                   The niosomal suspension
The unentrapped drug is
                                                                is centrifuged and the
removed by gel filtration of
                                                                supernatant is separated.
niosomal dispersion through a
                                                                The pellet is washed and
Sephadex-G-50 column and
                                                                then resuspended to obtain
elution     with   phosphate
                                                                a niosomal suspension free
buffered saline                           Dialysis              from unentrapped drug.

                                Dialyzed in a dialysis tubing
                                against phosphate buffer or
                                normal saline




        Gel Filtration                                           Centrifuser         73
a) Size, Shape and Morphology
Freeze Fracture Electron Microscopy:- Visualize the vesicular structure of
surfactant based vesicles.
Photon Correlation spectroscopy :- Determine mean diameter of the
vesicles.
Electron Microscopy :- Morphological studies of vesicles.
b) Entrapment efficiency
After preparing niosomal dispersion, unentrapped drug is separated by
dialysis and the drug remained entrapped in niosomes is determined by
complete vesicle disruption using 50% n-propanol or 0.1% Triton X-100
and analysing the resultant solution by appropriate assay method for the
drug.
c) Vesicle Suface Charge
Determined by measurement of electrophoretic mobility and expressed in
expressed in terms of zeta potential
d) In vitro studies                                                    74
Applications


                                            Oral
                           immunological                             Diagnostic
Leishmaniasis   Oncology                    drug       Transdermal
                             adjuvants                                imaging
                                            delivery




                                                                     75
Lancôme has come out with a variety of anti-ageing
products which are based on noisome formulations.
L‟Oreal is also conducting research on anti-ageing
cosmetic products.




                                                     76
Summary :
    Niosomes provide incorporating the drug into for a
  better targeting of the drug at appropriate tissue
  destination .
  They presents a structure similar to liposome and hence
  they can represent alternative vesicular systems with
  respect to liposomes
  Niosomes are thoughts to be better candidates drug
  delivery as compared to liposomes due to various factors
  like cost, stability etc. Various type of drug deliveries can
  be possible using niosomes like targeting, ophthalmic,
  topical, parenteral etc.
                                                             77
1. S.P. Vyas And R.K. Khar,targeted & Controlled Drug
     Delivery,liposomes,173-279.
2. Mohammad Riaz, Liposomes :Preparation Methods,
   Pakistan Journal Of Pharmaceutical Sciences, January
   1996,Vol.19(1),65-77.
3. Sharma Vijay K1*, Liposomes: Present Prospective and
   Future Challenges,International Journal Of Current
   Pharmaceutical Review And Research, oct 2010,vol1,
   issue 2,6-16
4. Himanshu Anwekar*, Liposome- as drug carriers,
   International Journal Of Pharmacy & Life Sciences,
   Vol.2, Issue 7: July: 2011, 945-951


                                                          78
5. Madhav Nvs* And Saini A, Niosomes: A Novel
   Drug Delivery System, International Journal Of
  Research In Pharmacy And Chemistry, 2011,
  1(3),498-511.
6. Lohumi Ashutosh, Rawat Suman, A Novel Drug
  Delivery System: Niosomes Review, Journal Of Drug
  Delivery & Therapeutics; 2012, 2(5), 129-135.
7. Pawar Sd *, Pawar Rg, Niosome: An Unique Drug
  Delivery System, International journal Of Pharmacy,
  Biology and Allied Sciences, April, 2012, 1(3): 406-416.
8. Rajesh Z. Mujoriya, Niosomal Drug Delivery System –
   A Review, International Journal Of Applied
  Pharmaceutics, Vol 3, Issue 3, 2011,7-10.


                                                             79
Success in life mostly depends on the power of
„CONCENTRATION‟
                            --- Swami Vivekananda

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liposomes and niosomes

  • 2. • Introduction • Structure of liposomes • Advantages& disadvantages • Components of liposome • Mechanism of liposome • Preparation methods of liposomes • Characterization of liposomes • Applications of liposomes • Summary • Niosomes Introduction • Advantages& disadvantages • Preparation methods of niosomes • Characterisation of niosomes • summary • References
  • 3. LIPOSOMES liposomes are concentric bilayered vesicles in which an aqueous volume is entirely enclosed by a membraneous lipid bilayer mainly composed of natural or synthetic phospholipids. Liposomes were first produced in England in 1961 by Alec D. Bangham. The size of a liposome ranges from some 20 nm up to several micrometers 1
  • 4. Liposome =Phospholipid+ cholesterol Hydrophillic head Hydrophobic tail The lipid moecules are usually phospholipids-amphipathic moieties with a hydrophilic head group and two hydrophobic tails. 2
  • 5. Advantages of liposomes: Provides selective passive targeting to tumor tissues. (liposomal doxorubicin) . Increased efficacy and therapeutic index. Reduction in toxicity of the encapsulated agent. Site avoidance effect (avoids non-target tissues). Improved pharmacokinetic effects . Flexibility to couple with site-specific ligands to achieve active targeting. 3
  • 6. Disadvantages of liposomes: Production cost is high. Leakage and fusion of encapsulated drug / molecules. Sometimes phospholipid undergoes oxidation and hydrolysis like reaction. Short half-life. Low solubility. 4
  • 7. Cross-section of liposomes: Polar Lipids (Phospholipid) Lipid Soluble ingredients (Drugs,Nutrients & vitamins) H2O Layer Water Soluble ingredients (Drugs, Nutrients 5 & vitamins)
  • 8. components of liposomes: The structural components of liposomes include: A. Phospholipids B. cholesterol 6
  • 9. A. General representation of phospholipids: 7
  • 10. Phospholipids Phosphatidylcholine- natural Amphipathic molecule Hydrophilic polar head- Phosphoric acid bound to water soluble molecule. Glyceryl bridge Hydrophobic tail- 2 fatty acid chain containing 10-24 carbon atoms and 0-6 double bond in each chain. The amphipathic molecule self organise in ordered supramolecular structure when confronted (meet face to face) with solvent. 8
  • 11. The most common natural phospholipid is the phospatidylcholine (PC ). Polar Head Groups Naturally occurring phospholipids used are : PC: Phosphatidylcholine. PE: Phosphatidylethanolamine. Three carbon glycerol PS: Phosphatidylserine Synthetic phospholipids used are: DOPC: Dioleoyl phosphatidylcholine DSPC: Disteroyl phosphatidylcholine DOPE: Dioleoyl phosphatidylethanolamine DSPE: Distearoyl phosphatidylethanolamine 9
  • 12. Molecular geometry on structure of amphiphillic aggregates: 10
  • 13. Molecules of PC are not soluble in water. In aqueous media they align themselves closely in planar bilayer sheets in order to minimize the unfavorable 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 11
  • 14. PHASE TRANSITION TEMPERATURE phospholipid membranes can exist At various temperatures, in different phases. The transition from one phase to another can be detected by technique like micro calorimetry . What exactly happens during phase transition? Tightly ordered At elevated temperature liquid crystal phase gel state ( lipid membrane) (movement is higher) This is due to the fatty acid chain adopting a new conformation other than all trans straight chain configuration, such as gauche configuration state( phenomenon- chain tilt ) 12
  • 15. B. Cholesterol: Cholesterol stabilizes the Membrane Steroid lipid Interdigitates between phospholipids. i.e. below Tc , it makes membrane less ordered & above Tc more ordered. Being an amphipathic molecule, cholesterol inserts into the membrane with its hydroxyl group of cholesterol oriented towards the aqueous surface and aliphatic chain aligned parallel to the acyl chains in the center of the bilayer . 13
  • 16. Role of cholesterol in bilayer formation: Cholesterol act as fluidity buffer After intercalation with phospholipid molecules alter the freedom of motion of carbon molecules in the acyl Chain Restricts the transformations of trans to gauche Conformations. Incorporated into phospholipid membrane upto 1:1 or 2:1 of cholesterol to PC. 14
  • 17. Mechanism of liposome formation: 15
  • 18. Classification of liposome : Classification of liposome Structural Method of Composition parameters preparation and application 16
  • 19. Lamella : Types of vesicles based on lamella 17
  • 20. A. Structural parameters: Based on structural parameters MLV OLV UV MVV Multilamellar oligolamellar Multivesicular Unilamellar Large vesicles vesicles vesicles (>0.5 um) (>0.1-1.0 um) Vesicles (> 1.0 UM) SUV MUV 20-100nm GUV LUV >1um >100nm 18
  • 21. B. Based on REV, SUV made method of by reverse phase preparation: evaporation method VET SPLV Vesicles Based on method of Stable prepared by preparation plurilamenar extrusion vesicles tech. FATMLV Frozen & thawed MLV 19
  • 22. Based on composition and convential application: immuno fusogenic Based on composition & application Long pH circulatory sensitive cationic 20
  • 23. MethodS of Liposome Preparation Passive Loading of the entrapped agents loading before/ during the manufacture technique procedure. Active/remo Certain types of compounds with ionizable groups & those with both te loading lipid & water solubility can be technique Introduced into liposomes after the formation of intact vesicles. 21
  • 24. Methods of liposome preparation Passive loading techniques Active loading techniques Mechanical dispersion Solvent dispersion Detergent removal methods methods technique LIPID FILM HYDRATION ETHANOL INJECTION DETERGENT REMOVAL BY HAND SHAKING,FREEZE DRYING OR NON HAND  ETHER INJECTION FORM MIXED MICELLES SHAKING  DOUBLE EMULSION BY DIALYSIS MICRO EMULSIFICATION REVERSE PHASE CHROMATIGRALPY SONICATION  VAPOURATION VESICLES DIFFUSION FRENCH PRESSURE CELL STABLE PLURI LAMELLER  VESICLES LIKE…. MEMBRANE EXTRUSON 22  VESICLES  RECONSTITUTED & DRIED RECONSTITUTED  VESICLES  SANDAI VIRUS ENVELOPE
  • 25. General Method Of Liposome Preparation: 23
  • 26. 1. Mechanical dispersion method: Lipid dissolve in organic solvent/co-solvent Remove organic solvent under vacuum Film deposition Solid lipid mixture is hydrated by using aqueous buffer Lipid spontaneously swell & Hydrate Liposome Post Hydration vortexing, sonication, freeze thawing & high pressure extrusion 24
  • 27. There are four basic methods of physical/mechanical dispersion : Hand shaken method. Non shaking method. Pro – liposomes . Freeze drying . 25
  • 28. Lipidsform stacks of film from organic solution (FE/HS) Then film is treated with aqueous medium Upon hydration lipids swell and peel out from RB flask vesiculate to form Multi lamellar vesicles(MLVs) 26
  • 29. Pro-liposomes:  To increase the surface area of dried lipid film & to facilitate instantaneous hydration. lipid Dried over Finely divided lipid particulate support Pro - liposomes like powdered NACL/ sorbital Pro- Dispersion of MLV’S water liposomes This Method overcome the stability problem. 27
  • 30. 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 . Membrane extrusion Liposomes Dried reconstituted vesicles(DRVs) Freeze thaw sonification (FTS) pH induced vesiculation Cochleate method. 28
  • 31. Sonicated unilamellar vesicles: The exposure of MLVs to ultrasonic irradation for producing small vesicles. Probe sonicator Bath sonicator Used for dispersions large volume require high of dilute lipids energy in small volumes Sonication MLVs hazy transparent 5-10 min solution centrifugation 30 min clear SUV 29 Dispersion.
  • 32. Micro emulsification liposomes: 30 Micro fluidizer
  • 33. French pressure cell liposomes: Extrusion of preformed large liposomes in french press under very high pressure . uni or oligo lamellar liposomes of intermediate size (30-80nm ) . Advantages Less leakage and more stable liposomes are formed compared to sonicated forms 31
  • 34. Vesicles prepared by extrusion technique : The size of liposomes is reduced by gently passing them through polycarbonate membrane filter of defined pore size at lower pressure Used for preparation of LUVs and MLVs 32
  • 35. Dried reconstituted vesicles& freeze thaw sonication method 33
  • 36. pH induced vesiculation: The transient change in pH brings about an increase in surface charge of the lipid bilayer which induces spontaneous LUVs vesiculation . Reduced the pH to 7.5 Exposed to high pH * Addition of ~ (addition of 1M 0.1M Hcl Preformed NaoH) MLV’S ~Period of (2.5-3.0) exposure < 2min MLVs 34
  • 37. Cochleate method: Cochleates Removal of Ca++ by Cylindrical EDTA rolls(cochleate Addition of cylinders) Ca++ ions SUVs made from phosphatidylse rine(PS) 35
  • 38. Solvent dispersion methods: Lipid dissolve in organic solvent Excess addition of aqueous phase Lipids allign at interface of aqueous and organic layer Formation of monolayer and bilayer of phospholipids Liposome Note:- Organic solvent miscible with aqueous phase 36
  • 39. Solvent dispersion methods: ETHANOL INJECTION/ETHER INJECTION: 37
  • 40. De-Emulsification method: Generally the liposome is made up in 2 steps: Aqueous medium containing material 1 st the inner leaflet of the bilayer . to be entrapped Then the outer half. Add to immiscible organic solution of lipid Mechanical agitation Microscopic water droplets Methods to prepare the droplets: ~Double emulsion vesicles ~Reverse phase evaporation vesicles 38 ~Sonication methods
  • 42. DETERGENT SOLUBILISATIOIN METHODS Phospholipid brought into intimate contact with aqueous phase By addition optimized concentration of detergent Formation of micelles (Liposome) Below CMC, detergent molecules exist in free soln. As the concentration is increased, micelles are formed. Note:- Liposome size and Methods to remove detergents: shape depend on chemical Dialysis nature of Column chromatography. detergent, concentration and 40 other lipid involved
  • 43. Active/remote loading technique: The lipid bilayer membrane is impermeable to ions & hydrophilic molecules. But, Permeation of hydrophobic molecules can be controlled by concentration gradients. Some weak acids or bases can be transported due to various transmembrane gradients Electrical gradients. Ionic(pH) gradients. Chemical potential gradients. Weak amphipathic bases accumulate in aq phase of lipid vesicles in response to difference in pH b/w Inside & outside of liposomes 41
  • 44. Solute bearing no Liposomes with low internal pH charge at neutral pH pH gradient is created by preparing liposomes with low internal pH. Addtn of base to extraliposomal medium. [Basic compds ( lipophilic (non ionic) at high pH & hydrophilic(ionic) at low pH)] Neutral solute passes Lipophilic (UNPROTONATED) drug diffuse easily through bilayer through the bilayer membrane by diffusion At low pH side, the molecules are predominantly protonated . Exchange of external medium by gel extrusion chromatorapghy with neutral solution. Charge aquired by solute inside Weak bases like doxorubicine, liposomes makes adriamycin and vincristine are them unable to exit encapsulated. 42
  • 45. Locus of drugs in liposomes: Hydrophilic (DOXORUBICIN) Low entrapment Leakage Hydrolytic degradation Lipophilic (CYCLOSPORINE) High entrapment Low leakage Chemical stability Ampiphilic (VINBLASTIN) High entrapment Rapid leakage Biphasic insoluble (ALLOPURINOL, 6- MERCAPTOPURINE) Poor loading & entrapment 43
  • 46. Characterization of liposomes: PHYSICAL CHARACTERISATION CHEMICA L CHARACTERISATION → Vesicles size/shape/morphology → Phospholipids /lipid concentration → Surface -charge/electrical potential → Drug concentration → Phase behaviour/ lamellarity → PH / Osmomolality → Drug release →Antioxidant degradation → % capture /free drug → Phospholipids / cholesterols – peroxidation/oxidation/hydrolysis BIOLOGICAL CHARACTERISATION → Sterility → Pyrogenisity → Animal toxicity 44 →Plasma Stability:
  • 47. 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 Photon correlation spectroscopy, laser light (submicron and micron range) scattering, gel permeation and gel exclusion 3. Surface charge Free-flow electrophoresis 4. Electrical surface potential and surface pH Zetapotential measurements 5. Lamellarity Small angle X-ray scattering, 31 P-NMR, Freeze- fracture electron microscopy 6. Phase behavior Freeze-fracture electron microscopy, Differential scanning calorimetery 7. Percent of free drug/ percent capture Minicolumn centrifugation, ion-exchange chromatography, radio labelling 8. Drug release Diffusion cell/ dialysis 45
  • 48. Characterization parameters Analytical method/Instrument 1. Phospholipid concentration Barlett assay, stewart assay, HPLC 2. Cholesterol concentration Cholesterol oxidase assay and HPLC 3. Phopholipid peroxidation UV absorbance 4. Phospholipid hydrolysis, HPLC and TLC Cholesterol auto-oxidation. 5. Osmolarity Osmomete 46
  • 49. Characterization parameters Analytical method/Instrument 1. Sterility Aerobic or anaerobic cultures 2. Pyrogenicity Limulus Amebocyte Lysate (LAL) test 3. Animal toxicity Monitoring survival rates, histology and pathology STABILITY OF LIPOSOMES: Stability invitro . ~ Lipid oxidation ~ Lipid peroxidation ~ Long term & accelerated stability Stability after systemic administration. 47
  • 50. 1. Endocytosis 2. Adsorption 3. fusion 4. Lipid transfer 48
  • 51. Encapsulation of drugs in liposomes: • Encapsulation volume/Trapped volume Volume of aqueous solution entrapped in liposomes per mole of PL (µL/µmol PL) • Encapsulation Efficiency Assessed by mini column centrifugation method & protamine aggregation method. protamine aggregation method used for neutral and negetively charged liposomes. Liposome dispersion can be precipitated with protamine solution and subsequent centrifugation at 2000RPM. By analysing the material in super natent & in liposome pellet ( after disrupting liposomal pellet with 0.6 ml of 10% triton x-100 ). The encapsulation efficiency of entrapped material can be estimated. • % Encapsulation Drug entrapped in liposomes x 100 Total drug added 49
  • 52. In gene delivery. As drug delivery carriers. Enzyme replacement therapy. Chelation therapy for treatment of heavy metal poisoning. Liposomes in antiviral/anti microbial therapy. In multi drug resistance. In tumour therapy. In immunology. In cosmetology 50
  • 53. DNA delivery of Genes by Liposomes Cheaper than viruses No immune response Especially good for in-lung delivery (cystic fibrosis) 100-1000 times more plasmid DNA needed for the same transfer efficiency as for viral vector 51
  • 55. Liposomes could serve as tumor specific vehicles (even without special targeting) Liposomes better penetrate into tissues with disrupted endothelial lining 53
  • 56. DRUG ROUTE OF APPLICATION TARGETED DISEASES ADMINISTRATION Amphotericin B Oral delivery Ergosterol membrane Mycotic infection Insulin Oral,ocular,pulmonary Decrease glucose level Diabetic mellitus And transdermal Ketoprofen Ocular delivary Cyclooxygenase enzyme inhibitor Pain muscle condition Pentoxyfyllin Pulmonary delivery phosphodiesterase Asthama Tobramycin Pulmonary delivery Protein synthesis inhibitor Pseudomonas infection,aeroginosa Salbutamol Pulmonary delivery ß2-adrenoceptor antagonist Asthama Cytarabin Pulmonary delivery DNA-polymerase inhibition Acute leukameias Benzocaine Transdermal Inhibition of nerve impulse from Ulcer on mucous surface sensory nerves with pain Ketaconazole Transdermal Inhibit ergosterol membrane Candida albicans Levanogesterol Transdermal Rhamnose receptor skin disorder hydroxyzine Transdermal H1-receptor antagonist Urtecaria,allergic skin disease Ibuprofen Oral delivery Chaemoceptor,free ending Rheumatoid arthritis triamcilonone Ocular delivery,Transdermal Inhibition of prostaglandin Anti-inflammatory 54
  • 57. NAME TRADE NAME COMPANY INDICATION Liposomal Abelcet Enzon Fungal infections amphotericin B Liposomal Ambisome Gilead Sciences Fungal and protozoal infections amphotericin B Liposomal cytarabine Depocyt Pacira (formerly Malignant lymphomatous meningitis SkyePharma) Liposomal DaunoXome Gilead Sciences HIV-related Kaposi’s sarcoma daunorubicin Liposomal doxorubicin Myocet Zeneus Combination therapy with cyclophosphamide in metastatic breast cancer Liposomal IRIV vaccine Epaxal Berna Biotech Hepatitis A Liposomal IRIV vaccine Inflexal V Berna Biotech Influenza Liposomal morphine DepoDur SkyePharma, Endo Postsurgical analgesia Liposomal verteporfin Visudyne QLT, Novartis Age-related macular degeneration, pathologic myopia, ocular histoplasmosis Liposome-PEG Doxil/Caelyx Ortho Biotech, HIV-related Kaposi’s sarcoma, metastatic breast doxorubicin Schering-Plough cancer, metastatic ovarian cancer 55 Micellular estradiol Estrasorb Novavax Menopausal therapy
  • 58. summary: o liposomes are concentric bilayered vesicles in which an aqueous volume is entirely enclosed by a membraneous lipid bilayer o Liposomes are one of the unique drug delivery system, in controlling and targeting drug delivery. o Components of liposomes include phospholipid and cholesterol. o Method of preparation of liposomes include active loading technique and passive loading technique. o Passive loading techniques include solvent mechanical dispersion, solvent dispersion & detergent solubilisation o Characterization of liposomes include physical,chemical and 56 biological.
  • 60. Niosomes are non-ionic surfactant based unilamellar or multilamellar bilayer vesicles up on hydration of non ionic surfactants with or without incorporation cholesterol . The niosomes are very small, and microscopic in size. Their size lies in the nanometric scale. Niosomes are a novel drug delivery system, in which the medication is encapsulated in a vesicle. Both hydrophilic & lipophilic drugs ,entrap either in the aqueous layer or in vesicular membrane made of lipid materials. 57
  • 61. Hydrophilic drugs Polar heads facing Structure of niosomes: located in hydrophilic region aqueous regions Head part encapsulated (hydrophillic) Tail part (hydrophobic) Drug molecules Hydrophobic drugs localized in the Phospholipids hydrophobic lamellae These vesicular systems are similar to liposomes that can be used as carriers of amphiphilic and lipophilic drugs. It is less toxic and improves the therapeutic index of drug by restricting its action to target cells. 58
  • 62. Advantages of niosomes: They are osmotically active and stable. They increase the stability of the entrapped drug. The vesicle suspension being water based offers greater patient compliance over oil based systems Since the structure of the niosome offers place to accommodate hydrophilic, lipophilic as well as ampiphilic drug moieties, they can be used for a variety of drugs. The vesicles can act as a depot to release the drug slowly and of controlled release. Biodegradable, non-immunogenic and biocompatible. 59
  • 63. Aggregation Fusion Leaking of entrapped drug Hydrolysis of encapsulated drugs which limiting the shelf life of the dispersion. 60
  • 64. Classification of niosomes Small Large Unilamellar Unilamellar Multilamellar Vesicle Vesicle Vesicle (SUV) (LUV) (MLV) Typical Size Ranges: SLV: 20-50 nm – MLV:100-1000 nm 61
  • 65. Components of niosomes: Cholesterol and Non ionic surfactants are the two major components used for the preparation of niosomes. Cholesterol provides rigidity and proper shape. The surfactants play a major role in the formation of niosomes. non-ionic surfactants like spans(span 20,40,60,85,80), tweens (tween 20,40,60,80) are generally used for the preparation of Niosomes. Few other surfactants that are reported to form niosomes are as follows : Ether linked surfactant Di-alkyl chain surfactant Ester linked Sorbitan Esters Poly-sorbates 62
  • 66. alkyl group chain length : C12-C18 Shud be above Span surfactants the gel to liquid with HLB values phase transition 4 and 8 temperature of Hydration Non-ionic the system Temperature surfactant nature Factors affecting Surfactants niosomes and lipid Membrane formation additives levels surfactant/lipid ratio: 10-30 mM Nature of Cholesterol: Prevent encapsulated vesicle aggregation. drug Dicetyl phosphate: -ve charge 63
  • 67. Concept of Critical Packing Parameter Prediction of vesicle forming ability is not a simply a matter of HLB CPP = v/lca0 where v - hydrophobic group volume, lc - critical hydrophobic group length and a0 - area of the hydrophilic head group CPP between 0.5 and 1 likely to form vesicles. < 0.5 (indicating a large contribution from the hydrophilic head group area) is said to give spherical micelles. >1 (indicating a large contribution from the hydrophobic group volume) should produce inverted micelles. 64
  • 68. Comparisition between liposomes & niosomes: Sl. Liposomes Niosomes No. 1. Vesicles made up of concentric Vesicles made up of surfactants bilayer of phospholipids with or without incorporation of cholesterol. 2. Size ranges from 10-3000nm Size ranges from 10-100nm 3. Comparatively expensive Inexpensive 4. Special storage condition are No such special requirement required 5. Phospholipids used are unstable Non-ionic surfactants are stable 6. Comparatively more toxic Less toxic 65
  • 69. Methods of Niosome preparation: Hand Shaking method  Reverse phase evaporation technique  Ether Injection method Multiple membrane extrusion method Bubble method  Sonication  From Proniosomes 66
  • 70. Hand shaking method: Rotary evaporator 67
  • 71. Reverse phase evaporation technique : Surfactant is dissolved in chloroform ond 0.25 volume of PBS buffer is emulsified to get a W/O emulsion. sonicated chloroform is evaporated under reduced pressure. The lipid or surfactant forms a gel first and hydrates to form vesicles. Free drug (unentrapped) is generally removed by dialysis. sonication: Surfactant +cholesterol Mixture is sonicated for 3 mixture is dispersed in 2 ml min at 60 C using titanium aqueous phase in vial probe sonicator Unilamellar niosomes 68
  • 72. 14 guage needle Ether injection Method: 69
  • 73. Multiple membrane extrusion Method: •Mixture of surfactant, cholesterol and dicetyl phosphate in chloroform is made into thin film by evaporation •The film is hydrated with aqueous drug solution and the resultant suspension extruded through polycarbonate membranes 70
  • 74. Bubble method: RBF as bubbling unit with three necks in water It is novel technique for the bath. one step preparation of liposomes and niosomes Reflux , thermometer and nitrogen supply by three necks without the use of organic solvents. Cholesterol+ Surfactant dispersed in buffer pH 7.4 at 70°C Above dispersion is homogenized for 15 sec and then bubbled with nitrogen gas at 70°C to get niosomes 71
  • 75. proniosomes: • Bubble Method • Formation of niosomes from proniosomes: It is prepared by coating water-soluble carrier such as sorbitol with surfactant. The result of the coating process is a dry formulation. In which each water-soluble particle is covered with a thin film of dry surfactant. This preparation is termed “Proniosomes”. 72
  • 76. Separation of unentrapped drug: Gel filtration Separation of unentrapped Centrifugation drug The niosomal suspension The unentrapped drug is is centrifuged and the removed by gel filtration of supernatant is separated. niosomal dispersion through a The pellet is washed and Sephadex-G-50 column and then resuspended to obtain elution with phosphate a niosomal suspension free buffered saline Dialysis from unentrapped drug. Dialyzed in a dialysis tubing against phosphate buffer or normal saline Gel Filtration Centrifuser 73
  • 77. a) Size, Shape and Morphology Freeze Fracture Electron Microscopy:- Visualize the vesicular structure of surfactant based vesicles. Photon Correlation spectroscopy :- Determine mean diameter of the vesicles. Electron Microscopy :- Morphological studies of vesicles. b) Entrapment efficiency After preparing niosomal dispersion, unentrapped drug is separated by dialysis and the drug remained entrapped in niosomes is determined by complete vesicle disruption using 50% n-propanol or 0.1% Triton X-100 and analysing the resultant solution by appropriate assay method for the drug. c) Vesicle Suface Charge Determined by measurement of electrophoretic mobility and expressed in expressed in terms of zeta potential d) In vitro studies 74
  • 78. Applications Oral immunological Diagnostic Leishmaniasis Oncology drug Transdermal adjuvants imaging delivery 75
  • 79. Lancôme has come out with a variety of anti-ageing products which are based on noisome formulations. L‟Oreal is also conducting research on anti-ageing cosmetic products. 76
  • 80. Summary :  Niosomes provide incorporating the drug into for a better targeting of the drug at appropriate tissue destination .  They presents a structure similar to liposome and hence they can represent alternative vesicular systems with respect to liposomes  Niosomes are thoughts to be better candidates drug delivery as compared to liposomes due to various factors like cost, stability etc. Various type of drug deliveries can be possible using niosomes like targeting, ophthalmic, topical, parenteral etc. 77
  • 81. 1. S.P. Vyas And R.K. Khar,targeted & Controlled Drug Delivery,liposomes,173-279. 2. Mohammad Riaz, Liposomes :Preparation Methods, Pakistan Journal Of Pharmaceutical Sciences, January 1996,Vol.19(1),65-77. 3. Sharma Vijay K1*, Liposomes: Present Prospective and Future Challenges,International Journal Of Current Pharmaceutical Review And Research, oct 2010,vol1, issue 2,6-16 4. Himanshu Anwekar*, Liposome- as drug carriers, International Journal Of Pharmacy & Life Sciences, Vol.2, Issue 7: July: 2011, 945-951 78
  • 82. 5. Madhav Nvs* And Saini A, Niosomes: A Novel Drug Delivery System, International Journal Of Research In Pharmacy And Chemistry, 2011, 1(3),498-511. 6. Lohumi Ashutosh, Rawat Suman, A Novel Drug Delivery System: Niosomes Review, Journal Of Drug Delivery & Therapeutics; 2012, 2(5), 129-135. 7. Pawar Sd *, Pawar Rg, Niosome: An Unique Drug Delivery System, International journal Of Pharmacy, Biology and Allied Sciences, April, 2012, 1(3): 406-416. 8. Rajesh Z. Mujoriya, Niosomal Drug Delivery System – A Review, International Journal Of Applied Pharmaceutics, Vol 3, Issue 3, 2011,7-10. 79
  • 83. Success in life mostly depends on the power of „CONCENTRATION‟ --- Swami Vivekananda

Editor's Notes

  1. Vesicle contents are exchanged with the dispersion medium during breaking and resealing of phospholipid bi layers as they pass into membrane
  2. polysorbate 20 ,should be above the gel to liquid phasetransition temperature of system.leads to gel to liq transition in niosomes,
  3. Niosomes for the treatment of Leishmaniasis-Niosomes are being used for the delivery of stilbogluconate an antileishmaniasis agent for its delivery to visceral organs. It may be related to passive delivery of the vesicles and the contents through RES recognition and uptake by the Kupffer cells. Niosomes in Oncology:- Various anticancer drugs like MTX, DOX, can be encapsulated inside the niosomes and bac easily be delivered to the tumor cells due to small size. Niosomes as immunological adjuvants:- The ability of niosomes to enhance antibody production in response to Bovine Seum Albumin was compared with Freud’s adjuvant in the Balb/c mice which revealed niosomes as potent stimulator of cellular immunity. 4. Niosomes and Oral drug delivery:-Niosomes can be used for oral delivery of drug thus protecting it from the hostile environment of the GIT and targeting to RE.5. Niosomes for Transdermal drug delivery:- They are being used in topical and transdermal products both contaning hydrophobic and hydrophillic drugs. The intracellular route is the main route of vesicle penetration across the skin.6. Niosomes in Diagnostic imaging:-Niosomes can act as carriers for radiopharmaceuticals and site specific vehicle for spleen and liver imaging.
  4. Niosomes for the treatment of Leishmaniasis-Niosomes are being used for the delivery of stilbogluconate an antileishmaniasis agent for its delivery to visceral organs. It may be related to passive delivery of the vesicles and the contents through RES recognition and uptake by the Kupffer cells. Niosomes in Oncology:- Various anticancer drugs like MTX, DOX, can be encapsulated inside the niosomes and bac easily be delivered to the tumor cells due to small size. Niosomes as immunological adjuvants:- The ability of niosomes to enhance antibody production in response to Bovine Seum Albumin was compared with Freud’s adjuvant in the Balb/c mice which revealed niosomes as potent stimulator of cellular immunity. 4. Niosomes and Oral drug delivery:-Niosomes can be used for oral delivery of drug thus protecting it from the hostile environment of the GIT and targeting to RE.5. Niosomes for Transdermal drug delivery:- They are being used in topical and transdermal products both contaning hydrophobic and hydrophillic drugs. The intracellular route is the main route of vesicle penetration across the skin.6. Niosomes in Diagnostic imaging:-Niosomes can act as carriers for radiopharmaceuticals and site specific vehicle for spleen and liver imaging.
  5. Niosomes for the treatment of Leishmaniasis-Niosomes are being used for the delivery of stilbogluconate an antileishmaniasis agent for its delivery to visceral organs. It may be related to passive delivery of the vesicles and the contents through RES recognition and uptake by the Kupffer cells. Niosomes in Oncology:- Various anticancer drugs like MTX, DOX, can be encapsulated inside the niosomes and bac easily be delivered to the tumor cells due to small size. Niosomes as immunological adjuvants:- The ability of niosomes to enhance antibody production in response to Bovine Seum Albumin was compared with Freud’s adjuvant in the Balb/c mice which revealed niosomes as potent stimulator of cellular immunity. 4. Niosomes and Oral drug delivery:-Niosomes can be used for oral delivery of drug thus protecting it from the hostile environment of the GIT and targeting to RE.5. Niosomes for Transdermal drug delivery:- They are being used in topical and transdermal products both contaning hydrophobic and hydrophillic drugs. The intracellular route is the main route of vesicle penetration across the skin.6. Niosomes in Diagnostic imaging:-Niosomes can act as carriers for radiopharmaceuticals and site specific vehicle for spleen and liver imaging.