NIOSOMES: A NOVEL DRUG

DELIVERY SYSTEM

Presented by:

Anirban Saha

M.Pharm (Pharmaceutics)

Amity Institute of Pharmacy (AIP)

AMITY INSTITUTE OF

PHARMACY

Introduction

Factors Affecting Niosomes Preparation

Methods of Preparation

Characterization of Niosomes

Stability of Niosomes

Applications of Niosomes

Toxicity of Niosomes

PRESENTATION FLOW

NOVEL DRUG DELIVERY SYSTEM (NDDS)

Refers to approaches, formulations, technologies, and

systems for transporting a pharmaceutical compound in the

body as needed to safely achieve its desired therapeutic

effect

May involve scientific site-targeting within the body, or

facilitating systemic pharmacokinetics

Technologies modify drug release profile, absorption,

distribution and elimination for the benefit of

Improving product efficacy and safety

Patient convenience and compliance

INTRODUCTION

EXAMPLES OF NDDS

• Niosomes

• Liposomes

• Nanoparticles

• Resealed erythrocytes

• Microspheres

• Monoclonal antibodies

• Micro emulsions

• Antibody-loaded drug delivery

• Magnetic microcapsules

• Implantable pumps

Figure 1: various drug delivery systems (Aitha S, 2013)

Novel drug delivery system, in

which the medication is

encapsulated in a vesicle which is

composed of a bilayer of non-ionic

surface active agents (Nasir A, 2012)

Are very small, and microscopic in

size.

Although structurally similar to

liposomes, they offer several

advantages over them.

NIOSOMES

Figure 2: Niosomes Vesicles (Aitha S, 2013)

The vesicles forming

amphiphile is a non-ionic

surfactant stabilized by

addition of cholesterol and

small amount of anionic

surfactant such as dicetyl

phosphate

NIOSOMES

Figure 3: Vesicle of niosome (Aitha S, 2013)

Figure 4: Structure of Niosomes

STRUCTURE

OF NIOSOMES similar to liposomes, in that they are also

made up of a bilayer.

However, the bilayer in the case of

Niosomes is made up of non-ionic

surface active agents rather than

phospholipids.

Made of a surfactant bilayer with its

hydrophilic ends exposed on the outside

and inside of the vesicle, while the

hydrophobic chains face each other

within the bilayer.

(Patel SM et al, 2012)

(Makeshwar KB, 2013)

STRUCTURE

OF NIOSOMES

vesicle holds hydrophilic

drugs within the space

enclosed in the vesicle,

while hydrophobic drugs

are embedded within the

bilayer itself.

Niosomes vesicle would

consist of a vesicle

forming amphiphile i.e. a

non-ionic surfactant such

as Span- 60, which is

usually stabilized by the

addition of cholesterol

(Makeshwar KB, 2013)

Figure 5: Structure of niosome (Makeshwar KB, 2013)

Entrap solutes in a manner analogous to liposomes.

Osmotically active and stable.

Accommodate the drug molecules with a wide range ofsolubility.

Exhibits flexibility in their structural characteristics(composition, fluidity and size)

Performance of the drug molecules is increased.

Better availability to the particular site by protecting thedrug from biological environment.

Surfactants used in preparation are biodegradable,biocompatible and non-immunogenic

SALIENT FEATURES OF

NIOSOMES (Makeshwar KB, 2013)

Improve the therapeutic performance of the drug molecules by

Delayed clearance from the circulation

Protecting the drug from biological environment

Restricting effects to target cells

Niosomal dispersion in an aqueous phase can be emulsified in a

nonaqueous phase to

Regulate the delivery rate of drug

Administer normal vesicle in external non-aqueous phase.

Handling and storage of surfactants requires no special conditions.

Bioavailability of poorly absorbed drugs is increased.

Targeted to the site of action by oral, parenteral as well as topical

routes.

ADVANTAGES OF NIOSOMES

DELIVERY SYSTEM (Makeshwar KB, 2013)

According to the nature of lamellarity

1. Multilamellar vesicles (MLV) 1-5 μm in size.

2. Large Unilamellar vesicles (LUV) 0.1 – 1μm in size

3. Small Unilamellar vesicles (SUV) 25 – 500 nm in size.

According to the size

1. Small Niosomes (100 nm – 200 nm)

2. Large Niosomes (800 nm – 900 nm)

3. Big Niosomes (2 μm – 4 μm)

TYPES OF NIOSOMES

FACTORS AFFECTING THE

FORMATION OF NIOSOMES

Type of surfactant influences encapsulation efficiency,

toxicity, and stability of Niosomes

NATURE OF SURFACTANT

The surfactant/lipid ratio is generally 10-30 mM (1-2.5%

w/w)

Increasing the surfactant/lipid level increases the total

amount of drug encapsulated

SURFACTANT AND LIPID LEVELS

NATURE OF THE DRUG

The Physio-chemical propertiesof encapsulated drug influencecharge and rigidity of theNiosome bilayer.

The drug interacts withsurfactant head groups anddevelops the charge that createsmutual repulsion betweensurfactant bilayers, and henceincreases vesicle size.

The aggregation of vesicles isprevented due to the chargedevelopment on bilayer.

Effect of the nature of drug on

formation vesicle

CHOLESTEROL(Tamizharas S et al, 2009)

Addition of cholesterol molecule to

Niosomal system

• Makes the membrane rigid

• Reduces leakage of drug from the Niosome

• Increases the chain order of bilayer

• Strengthen the non-polar tail of the non-ionic

surfactant

• Increase in the entrapment efficiency

• Leads to the transition from the gel state to

liquid phase in Niosomes systems

MEMBRANE ADDITIVES

Cholesterol

Charge inducers are one of the membrane

additives which are often included in Niosomes

because

Increase surface charge density

Prevent vesicles flocculation, Aggregation and

Fusion.

Examples: Dicetyl phosphate (DCP) and Stearyl

amine (SA)

MEMBRANE ADDITIVES(Nasir A, 2012)

Film Method

Ether Injection Method

Sonication

Reverse Phase Evaporation

Heating Method

Microfluidization

Multiple Membrane Extrusion Method

Transmembrane pH gradient (inside acidic) DrugUptake Process (remote Loading)

The “Bubble” Method

Formation of Niosomes from Proniosomes

METHODS OF PREPARATION (Madhav NVS, 2011)

•Mixture ofSurfactant andCholesterol

Dissolved in an organic solvent

in a round-bottomed flask.

(e.g. diethyl ether,

chloroform, etc.)

•organic solvent isremoved by lowpressure/vacuum atroom temperature

example using a rotary evaporator.

• The resultantdry surfactantfilm is hydratedby agitation at50–60°C

Multilamellar vesicles

(MLV) are formed

FILM METHOD • Also known as hand shaking method

FILM METHOD

Figure 6: Steps of Film method (Madhav NVS, 2011)

A solution of the surfactant ismade by dissolving it in diethylether.

This solution is then introduced using aninjection (14 gauge needle) into warm wateror aqueous media containing the drugmaintained at 60°C.

Vaporization of the etherleads to the formation ofsingle layered vesicles.

• The particle size of the Niosomes formed depend on theconditions used, and can range anywhere between 50-1000μm. (Madhav NVS, 2011)

ETHER INJECTION METHOD

Figure 7: Steps of Ether injection method (Madhav NVS, 2011)

The mixture is probe sonicated

at 60°C for 3 minutes using a sonicator with a titanium probe to yield Niosomes.

Added to the surfactant/ cholesterol mixture in a 10 ml glass

vial

Aliquot of drug solution in buffer

SONICATION

Figure 8: Sonication method (Madhav NVS, 2011)

Creation of a solutionof cholesterol andsurfactant (1:1 ratio)in a mixture of etherand chloroform

An aqueous phasecontaining the drugto be loaded isadded to this

Resulting twophases aresonicated at 4-5°C

A clear gel isformed which isfurther sonicatedafter the additionof phosphatebuffered saline(PBS)

Temperature israised to 40°C andpressure is reducedto remove theorganic phase

Viscous Niosomesuspension is formedwhich can be dilutedwith PBS and heatedon a water bath at60°C for 10 minutesto yield Niosomes

REVERSE PHASE EVAPORATION

Non-toxic, Scalable and one-step method.

HEATING METHOD

Mixtures of non-ionicsurfactant, cholesteroland/or charge inducingmolecules are added to anaqueous medium e.g.buffer, distilled H2O, etc

• In the presence of aPolyol such as glycerol.

The mixture is heated while stirring at low shear forces

• Until vesicles are formed

Recent technique used to prepare Unilamellar vesicles of

defined size distribution.

based on submerged jet principle

MICROFLUIDIZATION

Two fluidizedstreams interact atultra high velocities,in precisely definedmicro channelswithin the interactionchamber

The impingement of thinliquid sheet along acommon front is arrangedsuch that the energysupplied to the systemremains within the area ofNiosomes formation

The result is a greateruniformity, smallersize and betterreproducibility ofNiosome are formed

MICROFLUIDIZATION

Figure 9: Steps of microfludization method (Madhav NVS, 2011)

Good method for controlling Niosomes size.

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

Resultant suspension is extruded through polycarbonate membranes which are placed in series for upto 8 passages

Figure 10: Multiple membrane

extrusion method (Madhav NVS, 2011)

Solution of surfactantand cholesterol is madein chloroform

Solvent is then evaporatedunder reduced pressure to geta thin film on the wall of theround bottom flask, similar tothe hand shaking method

This film is thenhydrated using citric acidsolution by vortexmixing

Resulting Multilamellar vesicles are then treated

to three freeze thaw cycles and sonicated

To the Niosomal suspension, aqueous solution containing 10mg/ml of drug is added and vortexed

pH of the sample is then raised to 7.0-7.2 using 1M disodium

phosphate

Mixture is heated at 60°C for 10 minutes to

give Niosomes

TRANSMEMBRANE pH GRADIENT DRUG

UPTAKE PROCESS

A recently developed technique which allows the preparation of

Niosomes without the use of organic solvents.

BUBBLE METHOD

The bubbling unit consists of a round bottom flask with three necks, and this is positioned in a water bath to control the temperature.

Water-cooled reflux and thermometer is positioned in the first and second neck, while the third neck is used to supply nitrogen.

Cholesterol and surfactant are dispersed together in a buffer (pH 7.4) at 70°C.

This dispersion is mixed for a period of 15 seconds with high shear homogenizer and immediately afterwards, it is bubbled at 70°C using the nitrogen gas to yield Niosomes.

FORMATION OF NIOSOMES FROM

PRONIOSOMES (Makeshwar KB, 2013)

Water soluble carrier such as

sorbitol is coated 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”.

The Niosomesare recognized bythe addition ofaqueous phase atT > Tm and briefagitation.

T=Temperature.Tm = mean phase transition temperature

POST-PREPARATION PROCESSES

1) Dialysis:

The aqueous niosomal dispersion is dialyzed in a dialysis tubing

against phosphate buffer or normal saline or glucose solution.

2) Gel Filtration:

The unentrapped drug is removed by gel filtration of niosomal

dispersion through a Sephadex-G -50 column and elution with

phosphate buffered saline or normal saline.

3) Centrifugation:

The niosomal suspension is centrifuged and the supernatant is

separated. The pellet is washed and then resuspended to obtain a

niosomal suspension free from unentrapped drug.

POST-PREPARATION PROCESSES (Makeshwar KB, 2013)

a) Size, Shape and Morphology

b) Entrapment efficiency

c) Vesicle diameter

d) In vitro release

e) Vesicle charge

f) Bilayer rigidity and Homogeneity

g) Osmotic Shrinkage

h) Physical stability of vesicles at different temperature

i) Turbidity Measurement

CHARACTERIZATION OF NIOSOMES

Structure of surfactant based vesicles has been visualized

and established using freeze fracture microscopy

Photon correlation spectroscopy used to determine mean

diameter of the vesicles.

Electron microscopy used for morphological studies of

vesicles

Laser beam is generally used to determine size distribution,

mean surface diameter and mass distribution of Niosomes.

SIZE, SHAPE AND MORPHOLOGY

After preparing Niosomal dispersion, unentrapped drug is

separated by

Dialysis

Centrifugation

Gel filtration

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. (Bragagnia M, 2012)

ENTRAPMENT EFFICIENCY

To determine drug loading and encapsulation efficiency,

the niosomal aqueous suspension was ultracentrifuged,

supernatant was removed and sediment was washed

twice with distilled water in order to remove the

adsorbed drug.

The Niosomal recovery was calculated as:

NIOSOMAL DRUG LOADING

(Makeshwar KB, 2013)

Niosomes diameter can be determined using

Light microscopy

Photon correlation microscopy

Freeze fracture electron microscopy.

Freeze thawing

VESICLE DIAMETER (Shirsand SB, 2012)

Figure 11: Microphotograph of niosomes (Shrisand SB, 2012)

At various time intervals, the buffer is analysed for the drug content by an appropriate assay method.

The bag containing the vesicles is placed in 200 ml of buffer solution in a 250 ml beaker with constant shaking at 25°C or 37°C.

The vesicle suspension is pipetted into a bag made up of the tubing and sealed.

A dialysis sac is washed and soaked in distilled water.

A method of in-vitro release rate study includes the use of dialysis tubing.

IN VITRO RELEASE (Makeshwar KB, 2013)

The vesicle surface charge can play an important role in thebehaviour of Niosomes in vitro and in vivo.

Charged Niosomes are more stable against aggregation andfusion than uncharged vesicles.

In order to obtain an estimate of the surface potential, the zetapotential of individual Niosomes can be measured byMicroelectrophoresis, Fluorophores, and Dynamic lightscattering.

Zeta potential is calculated by using Henry equation (S P Vyas, 2011)

ζ =µ𝐸4πη

ΣWhere ζ is Zeta potential, µ𝐸 is electrophoretic mobility, η isviscosity of the medium and Σ is dielectric constant

VESICLE CHARGE

(Makeshwar KB, 2013)

The biodistribution and biodegradation of Niosomes are

influenced by rigidity of the bilayer.

Homogeneity can occur both within Niosomes structures

themselves and between Niosomes in dispersion and

could be identified via. NMR, Differential Scanning

Calorimetry (DSC) and Fourier transform-infra red

spectroscopy (FT-IR) techniques.

Membrane rigidity can be measured by means of

mobility of fluorescence probe as a function of

temperature. (Patel SM et al, 2012)

BILAYER RIGIDITY AND HOMOGENEITY

Osmotic shrinkage of vesicles can be determined by

monitoring reductions in vesicle diameter, initiated by

addition of hypertonic salt solution to suspension of

Niosomes.

Niosomes prepared from pure surfactant are osmotically

more sensitive in contrast to vesicles containing cholesterol.

OSMOTIC SHRINKAGE

Aggregation or fusion of vesicles as a function of

temperature was determined as the changes in vesicle

diameter by laser light scattering method.

The vesicles were stored in glass vials at room

temperature or kept in refrigerator (4oC) for 3 months.

The changes in morphology of Multilamellar vesicles

(MLVs) and also the constituent separation were assessed

by an optical microscope.

The retention of entrapped drug were measured 72 hours

after preparation and after 1, 2 or 3 months in same

formulations

PHYSICAL STABILITY OF VESICLES

AT DIFFERENT TEMPERATURE

Niosomes were diluted with bidistilled water to give a total

lipid concentration of 0.312 mM

After rapid mixing by sonication for 5 min

Turbidity was measured as the absorbance with an

ultraviolet-visible diode array spectrophotometer.

TURBIDITY MEASUREMENT

Vesicles are stabilized based upon formation of 4 different

forces:

1. Van der Waals forces among surfactant molecules

2. Repulsive forces emerging from the electrostatic

interactions among charged groups of surfactant

molecules

3. Entropic repulsive forces of the head groups of

surfactants

4. Short-acting repulsive forces.

STABILITY OF NIOSOMES

FACTORS

Nature of surfactant

Structure of surfactant

Temperature of hydration

Nature of encapsulate

d drug

Inclusion of a charged molecule

FACTORS AFFECTING STABILITY OF NIOSOMES

A surfactant used for preparation of Niosomes must have ahydrophilic head and hydrophobic tail.

The hydrophobic tail may consist of one or two alkyl orperfluoroalkyl groups or in some cases a single steroidalgroup.

The ether type surfactants with single chain alkyl ashydrophobic tail is more toxic than corresponding dialkyletherchain.

The ester type surfactants are chemically less stable than ethertype surfactants and the former is less toxic than the latter dueto ester-linked surfactant degraded by esterases totriglycerides and fatty acid in vivo.

The surfactants with alkyl chain length from C12-C18 aresuitable for preparation of Niosome.

NATURE OF SURFACTANT (Singh CH, 2011)

The geometry of vesicle to be formed from surfactants is affected by itsstructure, which is related to critical packing parameters

Critical packing parameters can be defined using following equation,

𝐶𝑝𝑝 =𝑣

lc∗ a0

Where

v = hydrophobic group volume,

lc = the critical hydrophobic group length

a0 = the area of hydrophilic head group

From the critical packing parameter value type of miceller structureformed can be ascertained as given below,

If CPP < ½ then formation of spherical micelles,

If ½ < CPP < 1 formation of bilayer micelles,

If CPP > 1 formation inverted micelles23.

surfactants with longer alkyl chains generally give larger vesicles

STRUCTURE OF SURFACTANT

(Madhav NVS, 2011)

The physico-chemical properties of encapsulated drug

influence charge and rigidity of the Niosome bilayer.

The drug interacts with surfactant head groups and

develops the charge that creates mutual repulsion between

surfactant bilayers and hence increases vesicle size.

NATURE OF ENCAPSULATED DRUG(Singh CH, 2011)

Hydration temperature influences the shape and size of

the Niosome.

For ideal condition it should be above the gel to liquid

phase transition temperature of system.

Temperature change of Niosomal system affects

assembly of surfactants into vesicles and also induces

vesicle shape transformation

TEMPERATURE OF HYDRATION (Madhav NVS, 2011)

Niosomes as Drug Carriers

Diagnostic imaging with Niosomes

Drug Targeting

Delivery to the brain

Anti cancer drugs

Anti infectives

Targeting of bioactive agents

To Reticulo-endothelial system (RES)

To organs other than RES

NIOSOME DELIVERY APPLICATIONS(Malhotra M et al, 1994)

Ophthalmic drug delivery

Delivery of peptide drugs

Immunological application of Niosomes

Transdermal delivery of drugs by Niosomes

Delivery system for the vasoactive intestinal peptide

(VIP)

Niosomes as carriers for Hemoglobin

Niosomal vaccines

NIOSOME DELIVERY APPLICATIONS

Sustained Release

Localized Drug Action

OTHER APPLICATIONS

Unfortunately, there is not enough research conducted to

investigate toxicity of Niosomes.

It was determined that the ester type surfactants are less

toxic than ether type surfactants.

In general, the physical form of Niosomes did not

influence their toxicity as evident in a study comparing

the formulations prepared in the form of liquid crystals

and gels.

Nasal applications of these formulations caused toxicity in

the case of liquid crystal type Niosomes.

TOXICITY OF NIOSOMES

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REFERENCE