Transcript Copy of Nanoparticle..

Nanoparticles
1
Nanoparticles are the most extensively investigated
drug delivery systems.
This includes:
Polymeric nanoparticles
&
Liposomes
2
Polymeric nanoparticles
Nanoparticles are solid colloidal particles ranging in
size from 10 to 1,000 nm.
They are made of a macromolecular material which can be of
synthetic or natural origin.
Depending on the process used for their preparation, two
different types of nanoparticles can be obtained,
nanospheres and nanocapsules.
Nanospheres have a matrix-type structure in which a drug is
dispersed,
whereas nanocapsules exhibit a membrane-wall structure
with a core containing the drug.
Because these systems have very high surface areas, drugs
may also be adsorbed on their surface.
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MANUFACTURE OF NANOPARTICLES
Methods of manufacturing nanoparticles
The choice of the manufacturing method depends on the
raw material intended to be used and on the solubility
characteristics of the active compound to be associated to
the particles.
The raw material, biocompatibility, the degradation
behavior, choice of the administration route, desired
release profile of the drug, and the type of biomedical
application determine its selection.
Thus nanoparticle formulation requires an initial and very
precise definition of the needs and objectives to be
achieved.
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1- In situ polymerization of a monomer
Nanospheres
Two
different
considered
approaches
for
the
have
been
preparation
of
nanospheres by in situ polymerization,
depending on whether the monomer to be
Polymerized:
is
emulsified
in
a
nonsolvent
phase
(emulsification polymerization),
or
dissolved
nonsolvent
in a
for
the
solvent
that
resulting
(dispersion polymerization)
is
a
polymer
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Emulsification Polymerization.
Depending on the nature of the continuous phase in the
emulsion, whether,
the continuous phase is aqueous
(o/w emulsion), or organic (w/o emulsion).
In
both
cases
the
monomer
is
emulsified
in
the
nonsolvent phase in presence of surfactant molecules,
leading to the formation of monomer-swollen micelles
and stabilized monomer droplets.
6
The polymerization reaction takes place in the presence
of a chemical or physical initiator.
The energy provided by the initiator creates free reactive
monomers in the continuous phase which then collide
with the surrounding unreactive monomers and initiate
the polymerization chain reaction.
The reaction generally stops once full consumption of
monomer or initiator is achieved.
7
The mechanism by which the polymeric particles are formed
during
emulsification
polymerization
is
by
micellar
polymerization, where the swollen-monomer micelles act as
the site of nucleation and polymerization.
Swollen micelles exhibit sizes in the nanometer range and
thus have a much larger surface area in comparison with
that of the monomer droplets.
Once generated in the continuous phase, free reactive
monomers would more probably initiate the reaction within
the micelles.
8
As the monomer molecules are
slightly soluble in the
surrounding phase, they reach
the micelles by diffusion from
the monomer droplets through
the continuous phase, thus
allowing the polymerization to be followed within the
micelles.
So, in this case, monomer droplets would essentially act as
monomer reservoirs.
9
The drug to be associated to the nanospheres may be
present during polymerization or can be subsequently added
to the preformed nanospheres, so that the drug can be
either incorporated into the matrix or simply adsorbed at the
surface of the nanospheres.
10
Micellar polymerization mechanism.11
Dispersion Polymerization
The monomer is no more emulsified but dissolved in an
aqueous medium which acts as a precipitant for the polymer
to be formed.
The nucleation is directly induced in the aqueous monomer
solution.
For Production of Polymethacrylic Nanospheres, water
soluble methyl methacrylate monomers are dissolved in an
aqueous medium and polymerized by
chemical
initiation
(ammonium
y-irradiation
or
or by
potassium
peroxodisulfate) combined with heating to temperatures
above 650C.
12
In the case of chemical initiation, the aqueous medium must
be previously flushed with nitrogen for 1 h in order to
remove
its
oxygen
content,
which
could
inhibit
the
polymerization by interfering with the initiated radicals.
Oligomers (primary polymer) are formed and above a certain
molecular weight precipitate in the form of primary particles.
Finally, nanospheres are obtained by the growth or the
fusion of primary particles in the aqueous phase
13
The removal of detergents is very important that produce
very slowly biodegradable and biocompatible nanoparticles
The technique can be used for vaccination purposes.
where initiation by y-irradiation can be useful for the
production of nanospheres by polymerization in the
presence of antigenic material at room temperature,
thus preventing its destruction.
Examples of antigenic materials used to produce
nanoparticulate were different influenza antigens.
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Nanocapsules
Nanocapsules is a colloidal carrier with a capsular
structure consisting of a polymeric envelope surrounding
an oily central cavity containing lipophilic drugs.
Interfacial Polymerization Mechanism
The monomer (isobutyl cyanoacrylate) and a lipophilic
drug (progesterone) are dissolved in an ethanolic phase
containing an oil (Myglyol®, Lipiodol®) or a non-miscible
organic solvent (benzylic alcohol).
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This mixture is slowly injected through a needle into a
magnetically stirred aqueous phase (pH 4—10) containing
an nonionic surfactant (poloxamer 188).
Upon mixing with the aqueous phase, ethanol rapidly
diffuses
out
spontaneous
of
the
organic
emulsification
of
phase
the
giving
rise
to
oil/monomer/drug
mixture.
Immediately the monomer molecules polymerize at the
water-oil interface, leading to the formation of solid wallstructured particles.
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The mixture immediately becomes milky and nanocapsules
with a mean diameter of 200-300 nm are formed.
The colloidal suspension can then be concentrated by
evaporation under reduced pressure and filtered.
17
For encapsulate hydrophilic compounds such as doxorubicin
and fluorescein, inverse emulsification polymerization
Technique can be used.
In this procedure, the drug was dissolved in a small volume
of water and emulsified in an organic external phase
( hexane) containing a surfactant.
An organic solution of cyanoacrylate monomers is added to
the w/o emulsion.
Nanocapsules are formed, resulting from an interfacial
polymerization process around the nanodroplets.
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Disadvantages of preparation by
in situ polymerization of a monomer
1. Most of the carriers produced by polymerization have
inadequate biodegradability properties preventing their
use for regular therapeutic administration.
Only for vaccination purposes is being suitable when
achievement of a very prolonged immune response is
desired.
2.
The
possible
interactions
inhibition
with
of
activated
polymerization processes.
drug
activity
monomers
due
to
present
in
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3. It is very difficult to calculate the molecular weight of
the
resulting
polymerized
material
due
to
the
multicomponent nature of the polymerization media.
However, the determination of molecular weight is very
important as it influences the biodistribution and release
of the polymeric carrier.
4. The presence of toxic residues due to the unreacted
monomer, initiator, and surfactant molecules whose
elimination requires time-consuming and not always
efficient procedures.
20
In
order
to
avoid
biodegradable,
those
limitations
well-characterized,
and
and
produce
nontoxic
nanoparticles., already polymerized materials have been
used.
These
materials
include
natural
macromolecules
(biopolymers) and synthetic polymers.
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2- Dispersion of a Preformed Polymer
Nanospheres Prepared From Natural Macromolecules
Due to their biodegradability and biocompatibility, example
of natural macromolecules used for the manufacture of
nanospheres are:
Proteins as albumin, gelatin,
(the most widely used)
Polysaccharides as alginate or agarose
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Two manufacturing techniques are used to produce
nanospheres from natural macromolecules.
1. The first technique is based on the formation
of a w/o emulsion followed by heat denaturation or
chemical cross-linking of the macromolecule.
2.The second technique is a phase separation process in
an aqueous medium followed by chemical crosslinking.
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Emulsification-Based Methods
An aqueous solution of albumin is emulsified at room
temperature in a vegetal oil (cottonseed oil) and
homogenized
either
by
a
homogenizer
or
an
ultrasonication.
 Once
a high degree of dispersion is achieved, the
emulsion is added drop wise to a large volume of
preheated oil (>120°C) under stirring.
This leads to the immediate vaporization of the water
contained in the droplets and to the irreversible
denaturation of the albumin which coagulates in the
form of solid nanospheres.
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The
suspension is then cooled at
room temperature or in an ice
bath.
For
complete removal of the oil,
wash
the
particles
using
large
amounts of organic solvent (e.g.,
ether, ethanol, acetone).
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Disadvantages of this technique:
The purification step may cause particle wastes.
The hardening step by heat denaturation may
be
harmful to heat-sensitive drugs. This can be avoided by
the use of a crosslinking agent.

Large amounts of organic solvents are required to
obtain nanospheres free of any oil or residues.

It is very difficult to produce small nanospheres
(<500 nm) with narrow-size distributions, due to the
instability of the emulsion prior to hardening by heat
or crosslinking.
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Preparation of nanospheres by thermal denaturation of albumin
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Phase Separation-Based Methods
in an Aqueous Medium
The particles are formed in an aqueous medium by a phase
separation process and are stabilized by cross linking with
glutaraldehye.

Gelatin and albumin nanospheres can be produced by the
slow addition of a desolvating agent (neutral salt or
alcohol) to the protein solution to the form protein
aggregates

The nanospheres are obtained by crosslinking of these
aggregates with glutaraldehyde.
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Preparation of nanospheres by desolvation of albumin.
29
The
major
disadvantage
of
this
technique is the necessity for using
hardening agents (glutaraldehyde) that
may react with the drug and may cause
toxicity
to
the
nanoparticle
formulations.
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Nanospheres Prepared From Synthetic Polymers
Examples of synthetic polymers used for the preparation
of nanospheres:
Polylactic acid (PLA), poly(glycolic acid) (PGA),
polylactic-co-glycolic acid) (PLGA), poly(e-caprolactone)
(PCL), and poly(Polyhydroxybutyrate) (PHB).
These polyesters polymers exhibit biodegradability and
biocompatibility.
Under physiological conditions, they are degraded into
safe products as glycolic acid and lactic acid.
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Polyesters nanoparticles can be produced using two
different methods.
Emulsification-Based Methods.
The method is based on the emulsification of an organic
solution of the polymer (chloroform, methylene chloride,
ethyl acetate), in an aqueous phase (o/w emulsion)
containing
surfactants
(e.g.,
polysorbate,
poloxamer,
sodium dodecyl sulfate).
The extraction of the solvent from the nanodroplets is
achieved by evaporation of the organic solvent at room
temperature under stirring.
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Emulsification-solvent evaporation method
33
A second method is based on the direct precipitation of
the solubilized polymer by salting out process
Two different salting-out agents, magnesium chloride and
magnesium acetate, were used, providing an acidic or a
basic aqueous phase, respectively.
Although the salting-out process has proved suitable for
the production of large quantities of highly drug-loaded
nanospheres, the use of large amounts of salts
may raise a problem concerning compatibility with active
compounds.
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Salting-out process
35
Direct Precipitation-Based Method
This method allows nanospheres to be obtained without
prior emulsification.
This technique involves the use of an organic solvent that is
completely miscible with the aqueous phase, typically
(acetone, ethanol or methanol).
In this case, the polymer precipitation is directly induced in
an aqueous medium (non solvent), by progressive
addition under stirring of the polymer solution.
This method is limited to drugs that are highly soluble
in polar solvents, but only slightly soluble in water (e.g.,
indomethacin).
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Direct precipitation method
37
There are some requirements for nanoparticles intended to
be used as pharmaceutical dosage forms in humans:
(1)to be free of any potentially toxic impurities
(2)to be easy to store and to administer
(3)to be sterile if for parenteral administration.
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Purification.
Depending on the preparation method, various toxic
impurities
can
be
found
in
the
nanoparticulate
suspensions including:
organic solvents, residual monomers, polymerization
initiators, electrolytes, surfactants, stabilizers, and large
polymer aggregates.
The
necessity
for
and
degree
of
purification
are
dependent on the final purpose of the formulation
developed.
39
For
example,
the
stabilizer
PVA,
frequently used to prepare polyester
nanoparticles, is not acceptable for
parenteral administration, whereas it is
not so critical for oral and ocular
administration.
Polymer aggregates can be easily removed by simple
filtration.
The
removal
of
other
impurities
requires
more
complicated procedures as gel filtration, dialysis, and
ultracentrifugation.
40
However, these methods are incapable of eliminating
molecules with high molecular weight.
Using cross-flow filtration technique, the nanoparticle
suspension is filtered through membranes, with the
direction of the fluid being tangential to the surface of
the membranes to avoid the clogging of the filters.
It was shown that by using a microfiltration membrane
(porosity of 100 nm), nanoparticles produced by the
salting out process could be purified of the salts.
41
Main Methods for the Purification of Nanoparticles on the Laboratory Scale
42
Freeze-drying.
Freeze-drying (lyophilization) represents one of the most
useful methodologies to ensure the long-term conservation
of polymeric nanoparticles
This technique involves the freezing of the suspension and
the elimination of its water content by sublimation under
reduced pressure, where nanoparticles are obtained in the
form of a dry powder that is easy to handle and to store.
Freeze-dried nanoparticles are usually readily redispersible
in water without modification of their physicochemical
properties
43
Nanocapsules composed of an oily core surrounded by a
tiny polymeric wall tend to aggregate during the freezedrying process.
This problem can be solved by desiccating these systems in
the
presence
of
an
appropriate
lyoprotective
and
cryoprotection agent such as mono- or disaccharides
(e.g., lactose, sucrose, glucose).
The mechanisms by which sugars protect nanoparticles
during freeze-drying is that during freezedrying sugars may
interact with the solute of interest (e.g., liposome, protein)
through hydrogen-bonding.
44
As a result, the solute might
be maintained in a "pseudohydrated“ state during the
dehydrating step of freezedrying, and would therefore
be protected from damage
during
dehydration
and
subsequent rehydration.
It has to be kept in mind that the addition of sugar may
affect
the
isotonicity
of
the
final
nanoparticulate
suspension, and that a subsequent step of tonicity
adjustment may be required prior to any parenteral or
ocular administration.
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Sterilization
 Nanoparticles intended to be used parenterally are
required to be sterile and apyrogenic.
 Filtration on 0.22 μm filters is not adequate for
nanoparticle suspensions because microorganisms and
nanoparticles are generally similar in size (0.25-1 μm).
 Sterilization may be achieved, either by using aseptic
conditions throughout formulation, or by sterilizing
treatments such as autoclaving or γ-irradiation.
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The choice of the sterilizing treatment depends on the
physical susceptibility of the system.
Autoclaving (moist heat sterilization) and γ - irradiation
May alter the physicochemical properties of the particles
in several systems.
These modifications occur as a consequence of the
cleavage or cross-linking of the polymeric chains.
The final formulation would therefore result from a
rational balance between conditions maintaining the
formulation integrity upon sterilization and the final
purpose of the formulation.
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