FORMULATION OF NANOSUSPENSION AS A CARRIER FREE DRUG
DELIVERY SYSTEM TO ENHANCE BIOAVAILABILITY OF POORLY
SOLUBLE DRUGS
Neha Vishal Gandhi1*, Dr. Uday Arvind Deokate2, Dr. Rajendra Panditrao Marathe2.
1
Department of Pharmaceutics, Government College of pharmacy, Aurangabad (M.S.), India.
2
Department of Chemistry, Government College of pharmacy, Aurangabad (M.S.), India.
*Corresponding Auther :Neha Vishal Gandhi
Phone No: 08446144354
E-mail ID: [email protected]
ABSTRACT: Solubility is an essential factor for drug effectiveness, independent of the route of
administration. Poorly soluble drugs are often a challenging task for formulators in the industry.
Conventional approaches for enhancement of solubility have limited applicability, especially
when the drugs are poorly soluble simultaneously in aqueous and in non‐aqueous media. Drug
nanosuspensions emerged as one promising solution for delivering such poorly soluble drugs. A
pharmaceutical nanosuspension is defined as very finely colloid ,biphasic, dispersed, solid drug
particles in an aqueous vehicle , bearing size below 1um ,without any matrix material , stabilized
by surfactants and polymers, prepared by suitable methods for drug delivery applications,
through various routes of administration like oral, parenteral ,ocular and pulmonary routes.
Scaling down to nanoparticles enhances drug aqueous solubility, bioavailability and thus
therapeutic efficacy by increasing drug surface area that comes into contact with biological
media. These are simple to prepare and bear several beneficial aspects intended to improve in
vivo performances i.e. pharmacokinetics, pharmacodynamics, safety and targeted delivery which
are discussed in this review. The present review also describes formulation criteria, different
preparation methods with their merits and demerits, characterization techniques and
achievements of nanosuspensions in drug delivery system. Additionally, transformation of
nanocrystals to final formulations and future trends of nanocrystals have also been described in
this review article.
KEYWORDS:
Nanosuspension,
bioavailability,
poorly soluble
drugs,
techniques, characterization, in vitro and in vivo performances, applications.
manufacturing
1. INTRODUCTION
The poor solubility of drug is a major problem which limits the development of highly potent
pharmaceutical drugs. The drugs with low solubility lead to low oral bioavailability and erratic
absorption which is particularly pertinent to drugs within class II of the Biopharmaceutical
Classification System (BCS) as mentioned in table no.1. Generally, the rate-limiting step for
absorption of the drugs in this class is the dissolution velocity arising from low solubility
(1)
.
Currently, poorly soluble drugs make up 1/3 of United States Pharmacopeia recognized drugs.
Among the new chemical entities (NCE) discovered by pharmaceutical chemist, 40% of NCE
were poorly soluble compounds
(2, 3)
. Also, 40% of the active pharmaceutical ingredients (API)
obtained by high-throughout screen have been found to be poorly soluble molecules. Lower
bioavailability resulting from poor solubility and incomplete dissolution in vivo, often holds
back continuous development and coming into the market of some promising NCE, or elicits
insufficient therapeutic effects of certain drugs. Therefore, formulating new poorly soluble
molecules to obtain an adequate bioavailability response has become a serious and challenging
scientific, industrial, and medical issue (4).
Poorly soluble compounds can be usually classified into two types: “grease ball” and “brick
dust” compounds
(3)
. Grease ball molecules are highly lipophilic compounds with a high log P
due to no interactions with water, whereas brick dust molecules usually are compounds with a
high melting point above 2000C and a low log P. These compounds have high lattice energy and
their solubility is limited by solid-state, in which the strong intermolecular bonds within the
crystal structure restrict their solubility in water and in oils. For example, 78% of compounds
with a log P of 6 and low melting point are grease ball, whereas 52% of compounds with melting
point of 2500C and a log P of 2 belong to brick dust
(5)
. Log P or partition coefficient is defined
as logarithm of the ratio of concentration of a compound in a mixture of two immiscible solvents
which are typically octanol and water. Log P determines the hydrophobic or lipophilic nature of
molecules (5, 6).
Grease ball molecules have easily passed through the drug development process pipeline to reach
the market by adopting appropriate formulation strategies, including incorporation of the proper
excipients like cosolvents such as PEG-400
(7)
, wetting agents such as sorbitan ester derivatives
(8)
, disintegrants such as croscarmellose sodium
, cyclodextrins such as β-cyclodextrins
(9)
(10)
,
micelles and lipid-based systems such as emulsions, microemulsions, liposomes, solid lipid
nanoparticles, and solid dispersions
(11)
. The use of such excipients in formulations of poorly
soluble drugs has been demonstrated to increase dissolution rate by increasing active drug
surface area in contact with the dissolution medium. However, there are some limitations with
the above formulations, such as low drug loading, high toxicity, poor stability, potential drug
expulsion during storage and complex manufacturing methods
(12)
. By contrast, brick dust
molecules are poorly soluble not only in water but also in oils. So the above formulation
strategies do not work effectively due to low encapsulation efficiency and low loading. Therefore
to overcome these limitations Muller et al. (13, 14) firstly developed nanosuspension for brick dust
molecules with improved dissolution and good absorption.
Nanosuspensions are defined as the submicron colloidal dispersions of pharmaceutical active
ingredient particles in a liquid phase (size below 1μm) without any matrix material which are
stabilized by surfactants and polymeric steric stabilizers. Nanosuspensions differ from
nanoparticles (polymeric colloidal carriers of drug) and solid lipid nanoparticles (lipid carrier of
drugs)
(15)
. A nanosuspension consists of drug nanocrystals, a stabilizing agent (typically
surfactants or polymeric stabilizers) and a liquid dispersion medium. Drug nanocrystals are pure
solid drug particles with a mean particle size below 1 µm, generally between 200 nm to 500 nm.
Although the term nanocrystal implies a crystalline structure, but particles can be crystalline,
partially crystalline or completely amorphous. Nanocrystals possess enhanced saturation
solubility, consequently an increased dissolution velocity as well as enhanced mucoadhesion (16).
These are composed of 100% drug without carriers and typically stabilized with surfactants or
polymeric steric stabilizers (1). The dispersion medium can be water, mixtures of water and other
non-aqueous media (e.g. water–ethanol mixtures) or non-aqueous media (e.g. polyethylene
glycol, oils) (4).
Over the last decade, drug nanosuspensions are considered as a novel approach to improve the
solubility of hydrophobic drugs since the technique is simple and effective which can quickly
launch product to the market. These were invented at the beginning of the 1990s and the first
products appeared very fast on the market from the year 2000 onwards. Additionally,
formulation of nanosuspension emerged to be a universal approach generally applied to all
poorly soluble drugs for the reason that all drugs can be disintegrated into nanometer-sized
particles (17). Nanosuspensions have been demonstrated to have a number of advantages as
compared to traditional forms of drug delivery. These advantages are summarized below:

an improved drug dissolution, absorption and thus improved bioavailability

simple production methods (18)

applicability to most drugs which are poorly soluble in both aqueous and organic media.

possibility to incorporate nanosuspensions in various dosage formats such as tablets,
pellets, and capsules following standard manufacturing techniques. For example,
ketoprofen nanosuspension has been successfully transformed into pellets (19)

improved dose proportionality

reduced fed/fasted state variability (20)

reduced inter-subject variability

better patient compliance (21, 22).
2. FORMULATION OF NANOSUSPENSION
Table 2: Formulation of nanosuspension [Modified from (23, 24)]
3. MANUFACTURING TECHNIQUES FOR NANOSUSPENSION
For the manufacturing of nanosuspensions, there are two converse methods: ‘bottom-up’ and the
‘top-down’ technologies (Fig. 1). The bottom-up technology is an assembling method from
molecules to nano-sized particles, including controlled precipitation, microemulsion, melt
emulsification method and so on. The top-down technology is a disintegration approach from
large particles, microparticles to nanoparticles, such as high-pressure homogenization and media
milling method. The principles and processes of these methods are described in detail in the text
ahead and their advantages and disadvantages have been mentioned in Table 3.
3.1 High pressure homogenization
It is most widely used method for preparing nanosuspensions of many poorly soluble drugs
(15)
.
The process can be summarized into three steps: firstly, drug powders are dispersed in a
stabilizer solution to form pre-suspension; then pre-suspension was homogenized by the high-
pressure homogenizer at a low pressure for several times as a kind of premilling, and finally was
homogenized at a high pressure for 10-25 cycles until the nanosuspensions with the desired size
were prepared. According to the liquids used to suspend drug powders, the method is classified
into homogenization in water (Dissocubes), homogenization in water-free media and water
mixtures (Nanopure) (25).
Homogenization in aqueous media (Disso cubes): This technology was developed by RH
Muller using a piston-gap type high pressure homogenizer in 1999 that can produce
nanosuspensions in water at room temperature. A drug powder is dispersed in an aqueous
surfactant solution and subsequently forced by a piston through the tiny homogenization gap
with pressure up to 4000 bar, typically 1500-2000 bar. The resulting high streaming velocity of
the suspension causes an increase in the dynamic pressure which is compensated by a reduction
in the static pressure below the vapor pressure of the aqueous phase (according to Bernoulli's
law). The simplified form of Bernoulli's law is shown below p + q = p0
where p0 is total pressure, p is static pressure, q is dynamic pressure.
Formation of gas bubbles occurs because the water starts boiling at room temperature. The gas
bubbles collapse immediately when the liquid leaves the homogenization gap being again under
normal air pressure of 1 bar. The phenomenon of formation and implosion of the gas bubbles is
called cavitation resulting in shockwaves. The drug particles are reduced in size due to high
shear forces, turbulent flow and the enormous power of these shockwaves. However, the use of
water leads to many disadvantages such as hydrolysis of water-sensitive drugs and problem
during subsequent drying steps (1).
Homogenization in nonaqueous media (Nanopure): Nanopure involves homogenization of
suspensions in water-free media having low vapour pressure like PEG 400, PEG 1000 etc. The
homogenization can be done at room temperature, 00C and below freezing point (-200C), hence it
is known as “deep freeze” homogenization (26). The cavitation in the homogenization gap is very
little or nonexistent. Even without cavitation, the size diminution to achieve nanoparticles is
sufficient by the remaining shear forces, particle collisions and turbulences. A low temperature
while homogenizing makes this process suitable for temperature labile drugs. Also, it is possible
to carry out the whole process in nonaqueous media to protect the drug from hydrolysis. The
obtained nanosuspensions from Nanopure® technology can directly be filled into soft gelatin
capsules or into hard gelatin or HPMC capsules which are then sealed. In addition, drug
nanocrystals in solid PEG can be used as powder for tablet production (1).
Nanojet: This is also a preparative technique for nanosuspension based on the principle of HighPressure Homogenization It is also called as opposite stream technology. It uses a chamber
where a stream of suspension is divided into two or more parts, which colloid with each other at
high pressure and due to the high shear forces produced during the process, particle size is
reduced (24).
To obtain an optimized formulation by the high pressure homogenization method, the following
process parameters that influence the properties of nanocrystals must be considered such as:
1. Applied pressure
2. Number of homogenization cycles
3. Temperature
The pressure is provided by the pump converting the kinetic energy of the fluid into the pressure
in the gap. The static pressure will drop to a larger extent leading to generation of more bubbles
and then higher energy to comminute the particles. This is consistent with the law of
conservation of energy. Therefore, it is anticipated that the higher homogenization pressure, the
smaller particle sizes are obtained. Usually for the production of the drug nanocrystals, a
maximum pressure (for most lab homogenizers this value is 1500 bar) is required. The fluid
passing through the gap is performed instantaneously, generally within several milliseconds. The
energy generated in such short time is not sufficient to comminute all particles into uniform drug
nanocrystals even at the highest applied pressure 1500 bar; thus more homogenization cycles are
needed to perform. The increased cycle numbers provide more energy to break down the
crystals. Therefore, homogenization is often performed in five, ten, or more cycles depending on
the hardness of drug and the desired particle size. Apart from reducing the particle size, more
cycles lead to more homogenous nanocrystal suspensions, i.e. a narrow size distribution. Because
the flow rate of fluid in the gap is not identical among different zones and the fluid in central
zone of the pipe has the higher velocity than the fluid near the wall, the energy dispersed among
the fluid is not uniform, leading to an inhomogeneous particle size distribution. By increasing
number of cycles, the probability that larger particles pass the zone of high-power density in the
middle of the gap increases; thus these particles are also diminished. Therefore, the particle size
is a function of pressure and number of cycles and thus the desired particle size can be achieved
by adjusting these procedure parameters. Temperature is also an important parameter which
should be strictly controlled when the drug is temperature sensitive. High pressure processing
increases the temperature of the sample (approximately 10 0C at 500 bar). An increasing
temperature in the homogenization process is not favorable to temperature-sensitive drugs. In
that case, the temperature can be promptly reduced by placing a heat exchanger ahead of the
homogenizer valve. In general, the sample temperature can be maintained at about 10 0C and
even below so that the process is applicable to the temperature-sensitive drugs. Obtainment of an
optimized procedure is achieved by adjustment of the production parameters and this leads to
generation of high quality nanosuspensions with little batch-to-batch variation (1).
3.2 Media milling
The media milling technique was developed by Liversidge et al.
(27)
. In this method high-shear
media mills or pearl mills are used to produce nanosuspension. The media mill consists of a
milling chamber, a milling shaft, and a recirculation chamber (Fig. 3). The milling media or balls
are framed in ceramic-sintered aluminium oxide or highly cross-linked polystyrene resin (11). The
method can be divided into three steps: firstly, drug and surfactants are mixed to form a
homogeneous mixture and inputted into the milling chamber; secondly, the milling ball or
milling pearls are introduced into and certain amount of water is or not added into the milling
chamber; thirdly, the miller is turned on for several hours to prepare the nanosuspensions. This
method is called as wet-milling method, if the water and other liquid are used, otherwise called
as dry-milling method. Alternatively, it can be further divided into jet-milling, pearl-milling and
ball-milling method according to the types of the milling media used. This procedure can be
carried out under controlled temperature. The friction and collision among drug particles and
pearls generate nanoparticles (28).
3.3 Controlled precipitation or microprecipitation
In this process the drug is firstly dissolved in a solvent. Then this solution is mixed with a
miscible antisolvent in the presence of surfactants. Rapid addition of a drug solution to the
antisolvent (usually water) leads to sudden supersaturation of drug in the mixed solution, and
generation of ultrafine crystalline or amorphous drug solids. This process involves two phases:
nuclei formation and crystal growth. When preparing a stable suspension with the minimum
particle size, a high nucleation rate but low growth rate is necessary. Both rates are dependent on
temperature: the optimum temperature for nucleation might lie below that for crystal growth,
which permits temperature optimization. High-supersaturation conditions are needed for rapid
nucleation by adding drug in minimum volume of a water-miscible organic solution into the
water under rapid mixing at low temperature. This rapid dilution with water results in highsupersaturation conditions, and causes spontaneous nucleation and a subsequent reduction of the
supersaturation condition in the vicinity of the nucleating crystals, and reduction of crystal
growth rates. In this process, the ratio of mixing time (τ
mix)
to precipitation time (τ
very important parameter influencing the resulting particle size. As τ
precip,
mix
precip)
is a
is reduced relative to τ
the greater supersaturation (more rapid nucleation) and longer time for precipitation
(including condensation and coagulation) may produce the minimum particle size (25).
3.4 Microemulsion
There are two methods to fabricate drug nanosuspensions by this technique. First include
particles precipitation by evaporating low–medium boiling point solvents with negligible water
solubility and second method involves particles precipitation by a quenching technique using
partially water-miscible solvents, such as benzyl alcohol and butyl lactate. Such solvents are
used as the dispersed phase of the emulsion to load the solute.
In the first method, an organic solvent or mixture of solvents loaded with drug is dispersed in the
aqueous phase containing suitable surfactants to form an emulsion or microemulsion. The
organic phase is then evaporated under reduced pressure so that the drug particles precipitate
instantaneously to form a nanosuspensions stabilized by surfactants. Since a particle is formed in
an emulsion droplet, it is flexible to control the nanoparticle size by controlling the droplet size.
Organic solvents usually used are acetone, methylene chloride, chloroform
safer ethyl acetate and ethyl formate etc (30).
(25)
and relatively
The second method uses partially water-miscible solvents such as butyl lactate, benzyl alcohol,
triacetin and ethyl acetate as the dispersed phase
(19, 20)
. The emulsion or microemulsion is
formed by the conventional dispersion method and the drug nanosuspensions are obtained by
diluting the emulsion or microemulsion with relatively large amount of water. The dilution
causes complete diffusion of the internal phase into the external phase, and leads to
instantaneous formation of the nanosuspensions
(11)
. The manner of diluting the emulsion
includes high-pressure homogenization or magnetic stirring. However the first method is much
more efficient than the latter one (20).
3.5 Melt emulsification method
The melt emulsification method traditionally was used to prepare solid lipid nanoparticles, but
Kocbek et al.
(31)
firstly used it to prepare ibuprofen nanosuspensions. The first step in this
method involves dispersing the drug in aqueous solution with stabilizer. Secondly, the
nanosuspension is heated above the melting point of the drug and homogenized with a highspeed homogenizer to produce an emulsion. During this procedure the temperature must be
controlled and maintained above the melting point of the drug. The final step of the melt
emulsification method is cooling off the emulsion to a suitable temperature, either at room
temperature or in an ice bath. Factors affecting particle size include process parameters like drug
and stabilizer concentrations, type of stabilizer, and cooling condition. Solvent free, prepared
nanosuspensions are particularly important from toxicity point of view (24).
3.6 Microprecipitation-High pressure homogenization (Nanoedge)
Nanoedge is a combination of microprecipitation and high-pressure homogenization techniques.
Method includes precipitation of friable materials followed by fragmentation under high shear
and/or thermal energy (26). The preparation process can be summarized into five steps. Firstly, the
drug is dissolved in a drug-soluble organic solvent to form a solution. Secondly, the stabilizers
are dissolved in the second solvent in which drug is insoluble. Thirdly, the drug solution is added
to the second solvent under high-speed agitation. This leads to the precipitation of drug as
microparticles during the diffusion from the miscible solvent to the second solvent. At the same
time, the surfactants in the solution get adsorbed into the surface of the freshly formed
microparticles rapidly to protect them from aggregation or growth and then pre-suspension gets
prepared. Finally, pre-suspension is homogenized under a suitable pressure for several times.
This is a process of adding energy, and can further reduce the size of particles and reinforce
interactions between drug-nanoparticles and stabilizers (32).
3.7 Supercritical fluid technology
Various methods attempted in supercritical fluid technology
include
rapid expansion of
supercritical solution (RESS) process, supercritical antisolvent process, and precipitation with
compressed antisolvent (PCA) process are used to produce nanoparticles. In RESS technique,
drug solution is expanded through a nozzle into supercritical fluid, resulting in precipitation of
the drug as fine particles by loss of solvent power of the supercritical fluid. By using RESS
method, Young et al. prepared cyclosporine nanoparticles having diameter of 400 to 700 nm. In
the PCA method, the drug solution is atomized into the CO2 compressed chamber. As the
removal of solvent occurs, the solution gets supersaturated and finally precipitation occurs. In
supercritical antisolvent process, drug solution is injected into the supercritical fluid and the
solvent gets extracted as well as the drug solution becomes supersaturated (33).
4. CHARACTERIZATION TECHNIQUES
4.1 Size: The most important characteristics of nanosuspensions are particle size and
polydispersity index (PI: particle size distribution). Particles size of nanosuspensions critically
determines the following characteristics of nanosuspensions (34):
(i) Drug saturation solubility
(ii) Dissolution rate
(iii) Bioavailability
(iv) Physical stability
According to Noyes-Whitney equation (1), which is based on Fick’s first law of diffusion,
decreasing particle size causes an increase in particle surface area that in turn increases drug
solubility in aqueous media contributing to an enhanced dissolution rate (35, 36):
dM/dt = DA/h (CBulk – CEq)
(1)
where dM/dt is the rate of dissolution, D is the average diffusion coefficient, A is the surface
area of the solid, CBulk is the concentration of drug in the bulk solution, CEq is the concentration
of drug in the diffusion layer surrounding the drug, and h is the diffusion layer thickness.
Increased solubility with reduction of particles size is also demonstrated by Ostwald-Freundlich
equation (2) (37):
C(r) = C (∞) exp (2γM / rρRT)
(2)
where C(r), C(∞) are the solubilities of a particle of radius r and infinite size, M is the molecular
weight, ρ is the density of the particle, γ is the interfacial tension, r is the particle radius, R is the
gas constant and T is the temperature.
Photon Correlation Spectroscopy (PCS) (also known as Dynamic Light Scattering) is a technique
often used to determine particle size and PI of drug nanosuspensions. PCS is capable of accurate
measurements of particle sizes in range of 3 nm to 3 µm. In this technique, the Brownian motion
(movement in random direction) of particles is measured as a function of time. Larger particles
move with lower velocity than smaller particles. In addition, larger particles may settle out of the
measurement zone. Hence, these factors limit capability for measuring particle sizes above 3 µm
(38)
. Laser Diffraction (LD) is typically used to measure particle size range of 0.05-80 µm up to
2000 µm. This technique can also be used to detect and quantify particle size ranges during the
production procedure
(34, 39)
. Other techniques routinely used for measuring particle size are
optical and electron microscopy. Scanning Electron Microscopy (SEM), Atomic Force
Microscopy (AFM)
(40, 41)
, and Transmission Electron Microscopy (TEM)
(41, 42)
are also
routinely used to characterize nanoparticles size and morphology. Furthermore, the Coulter
Counter analysis can be used to determine the absolute number of particles per unit volume for
different particle sizes (39).
4.2 Crystalline state and particle morphology: The high energy amorphous form of drugs is
thermodynamically unstable and changes to a crystalline form during storage. The amorphous
form is preferred due to superior dissolution characteristics and consequently higher
bioavailability of the drugs. Transformation from amorphous to crystalline forms over storage is
one of the issues that should be considered while formulating nanosuspensions. In order to
investigate amorphous and crystalline fractions X-ray powder diffraction (XRPD) is used. XRPD
is sometimes considered to be the most appropriate method for evaluating drug crystalline
structure, since each crystal has a specific diffraction pattern
(43)
. Differential Scanning
Calorimetry (DSC) is another commonly used technique for determining crystalline and
amorphous fractions. It measures the temperatures and heat flows associated with the transition
in drugs from crystalline to amorphous state as a function of time and temperature in a controlled
atmosphere. DSC can also be used in conjunction with XRPD (44, 45).
4.3 Particle charge (Zeta potential): Particle charge plays an important role in ensuring stable
nanosuspensions. The electric charge on a particle surface provides electrostatic repulsion
between the drug nanoparticles and in this way prevents particles from aggregation and
precipitation. The schematic presentation in figure (Fig. 6) provides an illustration of the electric
double layer around a charged particle. The double layer consists of a stern layer and a diffusion
layer of opposite ions. The electric potential at the shear plane is known as the zeta potential
(46)
.
It is considered that a minimum zeta potential of ± 30 mV is required to ensure pure electrostatic
stabilization. When electrostatic stabilization is combined with steric stabilization (by using
appropriate polymers), zeta potential of ± 20 mV could be sufficient to prevent drug particles
from aggregation and precipitation
(47)
. Steric stabilization is defined as stabilization caused by
the adsorbed and hydrated polymer layers on the dispersed particle (48). The zeta potential values
are commonly assessed by determining the particle electrophoretic mobility using the Zetasizer
(Malvern Instruments Ltd., UK) and converting the electrophoretic mobility to the zeta potential
via the Helmholtz-Smoluchowski equation
(72)
. In the field of materials science, an
electroacoustic technique (Acoustosizer, Matec Applied Sciences, USA) can also be used for the
determination of the zeta potential (49).
4.4 Stability: Reduction in particle size results in increased surface energy due to the greater
number of unstable surface atoms and molecules. This destabilizes the colloidal suspension.
Therefore, the use of stabilizers is often necessary to avoid particle agglomeration and reduce the
possibility for Ostwald ripening. Common stabilizers used to formulate nanosuspensions include
polysorbates, povidones, poloxamer, lecithin, polyoleate, and cellulose polymers. Mixture of
surfactants and polymers has been found to be beneficial for long-term stabilization of
nanosuspensions
(50, 51)
. Polymeric materials and surfactants act as an ionic barrier and/or
inhibitors of the close interaction between particles. Surfactants can increase the electrostatic
repulsion and improve particle stability by altering the zeta potential
(52)
. Precipitation of
particles is another phenomenon that should be taken into account when considering stability of
nanosuspensions. According to Stoke’s law (equation 3), decreasing particle size, reducing the
density difference of solid phase, and increasing the viscosity of the medium decrease the
precipitation velocity (53):
V = 2r2 (ρ1- ρ2) g / (9η)
(3)
where V is the precipitation velocity, r is the particle size, ρ1 is the mass density of particles, ρ2 is
the mass density of fluid, g is the gravitational acceleration, and η is the viscosity of the medium.
The stability of nanosuspension system can also be increased by increasing the uniformity of
particle sizes by using centrifugation or other techniques to remove larger particles (54).
5. IN VITRO PROPERTIES
Drug nanocrystals possess outstanding features enabling to overcome the solubility problems
including an increase in saturation solubility and dissolution rate. These features are resulted
from transferring of particle size from macroparticle to nanodimension that changes their
physicochemical properties on the basis of nanotechnology. A detailed description of the
physical background of these effects is reflected in the text ahead (25):
5.1 Saturation solubility: In general, saturation solubility is a compound-specific constant,
which is depending on physicochemical properties like crystalline structure (i.e. lattice energy)
and particle size of the compound, dissolution medium and temperature. However, these
specifications are only valid for drug particles with a minimum particle size in the micrometer
range. The polymorphic modification with highest energy and lowest melting point leads to the
best solubility. The saturation solubility is also a function of particle size with a critical size
below 1–2 μm and it increases with decreasing particle size below 1000 nm. This phenomenon
can be explained by the Kelvin and the Ostwald–Freundlich equations (32).
The Kelvin equation (4) is applicable to explain the relation between the dissolution pressure and
the curvature of the solid particles in liquid. The dissolution pressure is equivalent to the vapor
pressure. At saturation solubility state, the dissolving molecules and recrystallizing molecules are
in equilibrium. The dissolution pressure can be increased with increasing curvature (decreasing
particle size). Therefore, the equilibrium is shifted toward dissolution, and thus the saturation
solubility increases. The curvature is especially immense when the particle size is in the
nanometer range.
ln Pr / P∞ = 2γMr / rRTρ
(4)
where Pr is the dissolution pressure of a particle with the radius r, P∞ is the dissolution pressure
of an infinitely large particle, γ is the surface tension, R is the gas constant, T is the absolute
temperature, r is the radius of the particle, Mr is the molecular weight, ρ is the density of the
particle.
The Ostwald–Freundlich equation (5) directly describes the relation between the saturation
solubility of the drug and the particle size.
Log Cs / Cα = 2σV / 2.303RTρr
(5)
where Cs is the saturation solubility, Cα is the solubility of the solid consisting of large particles,
σ is the interfacial tension of substance, V is the molar volume of the particle material, R is the
gas constant, T is the absolute temperature, ρ is the density of the solid, r is the radius.
Thus Ostwald-Freundlich equation shows that the saturation solubility (Cs) of drug increases
with a decrease in the particle size (r). However, this effect is not substantial for larger particles
but will be pronounced for materials that have a mean particle size of less than 1-2 µm,
especially well under 200 nm (1).
5.2 Dissolution rate: Nanocrystals possess an increased dissolution velocity or rate that can be
explained by the Noyes - Whitney equation (6).
dX / dt = (DA / hD) x (Cs-Ct)
(6)
where dX/dt is the dissolution velocity, D is the diffusion coefficient, A is the surface area, hD is
the diffusional distance, Cs is the saturation solubility, Ct is the concentration around the
particles.
The dissolution velocity (dX/dt) of drug nanocrystals increases due to the greater surface area
(A) and the increase in saturation solubility (Cs) of the compound. The size reduction of
nanocrystals leads to an increased surface area and thus according to the Noyes-Whitney
equation the dissolution velocity is increased (55).
6. IN VIVO PHARMACOKINETIC PERFORMANCE
The establishment of relationship between in vitro release and in vivo absorption and the
monitoring of the in vivo performance of the nanosuspensions are essential to a successful
preparation, irrespective of the administration route and the delivery systems. For oral
nanosuspensions, the drug dissolution rate can influence in vivo biological performance of
formulations to a larger extent. For instance, albendazole nanosuspensions enhanced the Cmax
to1.5-2 times than that of microsuspension, and increased the AUC0-∞ and the relative
bioavailability by 1-2 times than those of microsuspension (Fig. 7) (56).
The formulation of drug nanocrystals can impressively improve the bioavailability of per orally
administered poorly soluble drugs as shown by changes in pharmacokinetic parameters of blood
profiles including, an increase in area under the blood concentration-time curve (AUC), an
increase in maximum plasma concentration (Cmax), a decrease in time to maximum plasma
concentration (Tmax). For example, Liversidge and Cundy reported that danazol, a gonadotropin
inhibitor, showed the absolute bioavailability of marketed danazol microsuspension (200 mg, 10
µm) only 5.1 ± 1.9%. Meanwhile, the absolute bioavailability of danazol nanosuspension (200
mg, 169 nm) was 82.3 ± 10.1% which was equal to 16-fold increase in bioavailability.
Additionally, the Tmax was reduced and the Cmax was 15-fold increased
(57)
. Additionally, the
formulation of drug nanocrystals can provide advantage whenever a quick onset of a poorly
soluble drug is required. For instance, an analgesic drug naproxen was formulated as
nanosuspension (270 nm) for oral administration. This nanosuspension gave approximately 3fold increase in AUC when compared to an unmilled suspension (20 µm) and also concurrent
reduced in Tmax was observed. The data showed that the time for nanosuspension to reach Cmax
was only about 8 min whereas the unmilled naproxen suspension achieved the Cmax at 33.5 min.
It was suggested that an increase in 4-fold faster absorption rate of nanosuspension on
comparison with unmilled suspension was contributed to the increased solubility and dissolution
rate of nanocrystals (58).
When poorly soluble drugs are formulated as a uniform nanosuspension, the variation in
bioavailability resulting from fasted/fed state can be minimized. The nanocrystals could
significantly increase dissolution rate because of the increase in solubility and enormous particle
surface. The dissolution rate of nanocrystals is fast enough even under the fasted state.
Therefore, the absorption in both fasted and fed state can be a permeability-limit, and the
absorption difference between the fasted and fed conditions due to the dissolution difference is
eliminated. For example the food effect on bioavailability of a new tablet formulation containing
fenofibrate nanoparticles was accessed in human. It was demonstrated that the peak and overall
exposures from the 145 mg nanoparticle fenofibrate tablet were not affected by food and the
result was concluded that the nanoparticle fenofibrate tablet can be taken regardless of the timing
of meals (59).
7. POST PRODUCTION PROCESSING
7.1. Solidification techniques: Nanoparticles are usually produced in the liquid media, that is,
nanosuspensions. The nanosuspensions usually have the stability issues involved in the physical
(e.g. Ostwald ripening and agglomeration) and chemical (e.g. hydrolysis) processes. In this case,
solid dosage forms are considered more attractive, due to their patient convenience (marketing
aspects) and good stability. Therefore, transformation of nanosuspensions into the solid dosage
form is desirable. Solidification methods of the nanosuspensions include some unit-operations
such as pelletization, granulation, spray drying or lyophilization. As the primary objective of the
nanoparticulate system is rapid dissolution, thus disintegration of the solid form and redispersion
of the individual nanoparticles should be rather rapid, so that it does not impose a barrier on the
integrated dissolution process
(25)
. On contrast drying of nanoparticles can create stress on the
particles that can cause aggregation. For example, drying may lead to crystallization of the
polymers such as poloxamers, thereby compromising their ability to prevent aggregation. Drying
can also create additional thermal stresses (due to heat for spray drying). In response to the above
considerations, adding matrix-formers to the suspension prior to solidification is necessary. At
present, except for traditional lyoprotectants (such as glucose, sucrose, trehalose, lactose,
mannitol, sorbitol, maltose and dextran),
(35, 36, 60)
some new matrix-formers have received
significant interest. Van Eerdenbrugh et al had successfully used microcrystalline cellulose to
displace sucrose as a matrix former during freeze-drying of itraconazole nanosuspensions(73), and
had again evaluated four alternative matrix formers [Avicel®PH101, Fujicalin® (CaHPO4),
Aerosil®200 (SiO2) and Inutec®SP1] for their capability in preserving rapid dissolution after
spray-drying of nanosuspensions (39).
7.2 Surface modification techniques: Rapid or burst release of nanosuspensions may cause
toxicity and severe side effects. Hence, surface modification is required in order to control drug
release and/or prolonged residence at the site of action. For instance, nanosuspensions used for
targeting the monocyte phagocytic system (MPS) in the treatment of lymphatic mediated
diseases
(61)
can cause toxicity due to accumulation of drug. Surface modification can be
understood if we compare drug release from coated and uncoated surfaces. Tan et al. showed that
layer-by-layer nanogels coating of procaine hydrochloride decreases the burst release of drug (62).
Another example is comparison between buparvaquone nanosuspensions with and without
mucoadhesive polymers. A significant reduction in the infective score of Cryptosporidium
parvum after oral administration of buparvaquone nanosuspensions with mucoadhesive polymers
was attributed to adhesion, higher concentration gradient development and prolonged residence
of drug particles at the absorption sites in the gastrointestinal tract (GIT)
(63, 64)
. The surface
engineering by surface coating is important for targeted drug delivery systems. PEG is
commonly used to modify nanoparticle surface. This leads to reduced protein adsorption and
opsonization of nanoparticles and leads to prolonged systematic circulation time. Longer
circulation time is required to allow nanoparticles sufficient time to leak out of vasculature in
infective and inflammatory areas including cancer tissues. Carefully engineered nanoparticle’s
surface can also effectively target the diseased tissue. For instance, Kreuter et al. have
demonstrated that polyisobutyl cyanoacrylate nanoparticles stabilised by classic surfactants
(Tween 20, 40, 60 and 80) can deliver peptide dalargin across blood-brain barrier (65).
8. APPLICATIONS
Nanosuspensions posses a wide application spectrum, as enables preparation of various dosage
forms by post-production processing. The smaller particle size and larger surface area lead to
increased dissolution rate and oral absorption. Additionally, the nanoparticulate nature facilitates
naturally targeting of the monocyte phagocytic system (MPS) and results in unusual
pharmacokinetic consequences. At present, there are many drugs in the form of nanosuspensions
to be reported and marketed (Table: 4) via the various administration routes, including oral,
parenteral, ophthalmic and pulmonary routes (25).
8.1 Oral drug delivery
In general, oral administration is first choice for various drugs due to painless-non invasive
administration, good patient compliance, readily transportation and simple manufacturing
process. The major problem associated with oral administration is low bioavailability and finally
its inadequate efficacy due to poor solubility and incomplete dissolution. Oral nanosuspensions
have been specifically used to increase the absorption rate and bioavailability of drugs due to
much larger surface to volume ratio. For example, when azithromycin was formulated as
nanosuspension, the dissolution rate was significantly enhanced in the nanometer-sized system,
that is more than 65% dissolved in 5 h, as opposed to only 20% of the micronized drugs. Apart
from improved oral absorption, nanosuspensions offer the following advantages: dose
proportionality, low inter-subject variability, flexibility to incorporate into various dosage forms
such as tablets, capsules and fast melts by means of standard manufacturing techniques.
Ketoprofen nanosuspensions have been successfully incorporated into pellets for the sustained
release the drug over a period of 24 h (66).
8.2 Parenteral drug delivery
Injections provide fast onset of action, accurate dose, reliable efficacy and avoidance of first-pass
metabolism. Previously, injections of poorly water-soluble drugs were usually approached by
formulating drugs with excessive amounts of co-solvents, solubilizers, surfactants and carrier
materials, which may induce serious toxicity. For instance, marketed paclitaxel injection
containing Cremophor EL often result in anaphylactoid reactions. To establish a more acceptable
formulation, an injectable nanosuspension formulation of poorly water-soluble drugs has
emerged. Successful formulations have been reported as applied to antineoplastic agents,
anaesthetic agents
hyperthermia
(67)
, antifungals and antibacterials
(68)
, as well as for agents for malignant
(69)
. Nanosuspensions can be administered via different parenteral routes, such as
intraarticular, intraperitoneal and intravenous injections. In some cases, their nanoparticulate
nature dictates MPS targeting. If intravenously administered nanoparticles do not dissolve
immediately, they will initially distribute to MPS organs, in particular the Kupffer cells in the
spleen and liver. Subsequent drug dissolution in MPS provides a depot effect of these organs.
The finding of initial sequestration by the MPS, followed by slow release is generally found for
intravenously administered itraconazole nanoparticulate dosage forms (Fig. 8)(69).
8.3 Pulmonary drug delivery
It aims at treating several respiratory conditions such as asthma and chronic obstructive
pulmonary diseases. Advantages of pulmonary drug delivery over oral and parenteral drug
administration include direct delivery to the site of action which leads to decreased dosage and
side effects. Conventional pulmonary delivery systems provide only rapid drug release, poor
residence time, and lack of selectivity. Nanosuspensions can solve problems of poor drug
solubility in pulmonary secretions and lack of selectivity through direct delivery to target
pulmonary cells. Adhesiveness of nanosuspensions to mucosal surfaces leads to improved
selectivity because of minimal drug loss and prolonged residence time at target site. Pulmonary
nanosuspensions improve drug diffusion and dissolution rate and consequently increase
bioavailability and also prevent undesirable drug deposition in the mouth and pharynx. Surface
engineered nanosuspensions may provide quick onset followed by controlled drug release which
is optimal drug delivery pattern for most pulmonary diseases. Moreover, nanosuspensions for
treating lung infections have demonstrated good proportion between actual and delivered drug
concentrations in each actuation. The internalization rate for nanoparticles of 0.5 µm diameter
into the pulmonary epithelial cell has been reported to be 10 times higher as compared to
particles of 1 µm and 100 times higher, when compared with particles of 2-3 µm (33).
8.4 Ophthalmic drug delivery
Major problems in ocular therapy include poor drug solubility in lachrymal fluids. Repeated
instillations of conventional eye drops due to drainage through the nasolacrimal duct often cause
side effects. Whereas nanosuspensions as ocular drug delivery systems offer several advantages
like (i) Nanocrystals with modified surface by appropriate bio-erodible polymer causes prolonged
residual time in cul-de-sac desired for effective treatment. Commonly reported polymers in
ocular nanosuspensions are poly (alkyl cyanoacrylates), polycaprolactone, and poly (lactic
acid)/poly(lactic-co-glycolic acid). Employing polymers in ocular drug delivery significantly
prolongs drug ocular residence time and improves bioavailability.
(ii) Positively charged nanoparticles have strong adhesion to negatively charged mucin which
extends the drug release. For example, polymer Eudragit RS 100 was used in ibuprofen
nanosuspensions to increase drug residence time by creating positively charged surface which
resulted in improved corneal adhesion (34). Flurbiprofen nanosuspensions formulated by Eudragit
polymers RS 100 and RL 100 exhibited prolonged drug release (70).
(iii) Reduced drug loss because of the natural adhesiveness of drug nanocrystals (34)
(iv) Enhanced rate and extent of drug absorption: for instance, in a study by Kassem et al.,
nanosuspensions of hydrocortisone, prednisolone, and dexamethasone were prepared by high
pressure homogenization. Measured intraocular pressure of normotensive Albino rabbits
demonstrated that glucocorticoid drugs in the form of nanosuspensions unlike conventional
dosage forms, showed significant increase in the absorption rate and therapeutic efficiency (71).
Employing polymers with the ability of in situ gelling (instilled in a liquid form and transformed
to a gel in the cul-de-sac) controls the drug release. Study by Gupta et al. suggested that
formulating forskolin nanoparticles in conjunction with in situ gel forming polymers noveon
AA-1 polycarbophil/poloxamer 407 controls drug release through increased corneal contact time
and slower drug diffusion within the viscous polymer medium (32).
8.5 Targeted drug delivery
Nanosuspensions can be used for targeted delivery as their surface properties and in- vivo
behavior can easily be altered by changing either the stabilizer or the milieu. The engineering of
stealth nanosuspensions (analogous to stealth liposomes) by using various surface coatings for
active or passive targeting at the desired site is the future of targeted drug delivery systems.
Kayser formulated a nanosuspension of Aphidicolin to improve drug targeting against
leishmania infected macrophages. He stated that the drug in the conventional form had an
effective concentration (EC 50) of 0.16 µg/ml whereas the nanosuspension formulation had an
enhanced activity with an EC (50) of 0.003 µg/ml
(28)
. Scholer et al showed an improved drug
targeting to the brain in the treatment of toxoplasmic encephalitis in a new murine model
infected with Toxoplasma gondii using a nanosuspension formulation of Atovaquone (49).
Thus nanosuspension proves to be a highly potent novel drug delivery system for variety of
routes of administration with flexibility of transformation into wide verity of dosage forms
favoring the desired response.
CONCLUSION AND PROSPECTIVES
Poor aqueous solubility is rapidly becoming the leading hurdle for formulation scientists
working on drug delivery of various drug compounds and leads to employment of novel
formulation technologies. The use of drug nanosuspension is a universal formulation approach to
increase the therapeutic performance of these drugs in any route of administration. Almost any
drug can be reduced in size to the nanometer range. Nanosuspensions are considered as the most
promising delivery system for poorly soluble drugs, due to high bioavailability and less interand intra-subjects variances. It is proved suitable to formulate various poorly soluble drugs, in
particular for the brick-dust molecules poorly soluble in both water and oils, and can reduce the
social investment and enhance the success rate in insoluble drug development. Altered
pharmacokinetic profiles of drugs caused by nanosuspensions have become appreciable so-far as
they improve safety and efficacy. So the study on in-vivo biological performance is extremely
important, and the establishment of an in-vitro/in-vivo relationship will become a hot research
field in the further study of nanosuspensions. The combination of nanosuspension solidification
technique with traditional dosage forms, e.g. transformation of drug nanoparticles into pellets,
tablets or gels, will readily widen application of nanosuspensions. Additionally their applications
in buccal, nasal and topical delivery will represent a more appealing prospective. Surface
modification of the drug nanosuspensions can further increase the benefits, e.g. stabilizing blood
level of drugs by controlling drug release and targeting specific organ by using special surface
ligands in the production process. Controlled drug release and functionalized surface coatings
capable of eliciting passive or active targeting, will be regarded as the future promising step in
the nanosuspensions research. This review presents the recent progress of therapeutic
nanosuspensions produced by various techniques such as high pressure homogenization, media
milling and microemulsion with the potential of these drug delivery vehicles in parenteral, oral,
ocular, and pulmonary administration routes. However, the research on drug nanosuspensions is
in its infancy. These systems carry flexibility and opportunity for further tailoring particle
surface properties to optimize in vivo responses and generation of new clinical approaches for
treating a number of diseases (cardiac, cancer, diabetes, Parkinson’s, Alzheimer’s, etc.).
ACKNOWLEDGEMENT
The authors express sincere thanks to all the dignitaries of Government College of Pharmacy,
Aurangabad, (M.S.) for their guidance, encouragement and availability of all the required
amenities.
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Table 1: Traits and examples of drugs in different BCS classes [Modified from (11)]
BCS class
Drug traits
Drug examples
I
High solubility,
high permeability
Low solubility, high
permeability
High solubility, low
permeability
Low solubility, low
permeability
Propranolol, metoprolol, and
theophylline
Piroxicam, naproxen, and
cyclosporine,
Ranitidine, cimetidine, and
metformin
Furosemide,
hydrochlorothiazide
II
III
IV
Table 2: Formulation of nanosuspension [Modified from (23, 24)]
Formulation ingredients
Function
Examples
Stabilizers
Cosurfactants
(preferred with microemulsion
technique)
Wet the drug particles
thoroughly, prevent
Ostwald’s ripening and
agglomeration of
nanosuspensions, providing
steric or ionic barrier
Lecithins, poloxamers,
polysorbate, cellulosics,
povidones
Influence phase behavior
when micro emulsions are
used to formulate
nanosuspensions
Bile salts, dipotassium
glycerrhizinate, Ethanol,
isopropanol,
Organic solvent
Pharmaceutically acceptable
less hazardous solvents are
used for preparation of
formulation
Methanol, ethanol,
chloroform, isopropanol,
(water miscible solvents)
ethyl acetate, ethyl Formate,
butyl lactate, triacetin,
benzyl alcohol
(partially water miscible
solvents)
Other additives
According to the
requirement of the route of
administration or the
properties of the drug
moiety
Buffers, salts, polyols,
osmogens, cryoprotectant
Table 3: The merits and demerits of different manufacturing processes for nanosuspensions
[Modified from (26)]
Technique
Merits
Demerits
High-Pressure
Simple technique
High no. of homogenization cycles
Homogenization
General applicability to most drugs
Pretreatment of micronized drug particles and
Useful for formation of very dilute as
presuspending materials before subjecting it to
well as highly concentrated
homogenization
nanosuspensions
Possible contamination of product could occur
Aseptic production possible
from metal ions coming through wall of the
Low risk of product contamination
homogenizer
Ease of scale-up
Media Milling
High flexibility in handling, even large
Possible erosion of material from the milling
quantities of drugs
pearls
Very few batch to batch variation in
Require milling process for hr. to days
particle size
Prolonged milling may induce the formation of
Ease of scale-up
amorphous leading to instability
Controlled
Simple process
Growing of drug crystals needs to be limit by
Precipitation
Yields stable products
surfactant addition
Low need of energy
Drug must be soluble in at least one solvent
Low cost of equipment
Narrowly applying space
Ease of scale-up
Wide size distribution
Potential toxicity of non aqueous solvents
Microemulsion
Low need of energy
Use of high concentration of surfactant and
Yields stable products
stabilizer
Simple process
Use of hazardous solvents
Small and uniform particle size
distribution
High drug solubilization
Ease of manufacture
Nanoedge
Small, uniform and stable products as
Complicated manufacturing process
compared to controlled -precipitation
Low need of mechanical force
and energy as compared to high
pressure homogenization
Melt Emulsification
Avoidance of organic solvents
Formation of large crystals
Table 4: Overview of drug nanocrystals for oral administration in current marketed form and
during pharmaceutical researches
Drug
Aprepitant
Finofibrate
Finofibrate
Griseofulvin
Danazol
Naproxen
Itraconazole
Ketoprofen
Tradename/
Company
Emend/ Merck
Indication
Applied
technology
Antiemetic
Top-down,
media milling
Tricor/ Abbott Hypercholesterolemia Top-down,
media milling
Triglide/First
Hypercholesterolemia Top-down, high
Horizon
pressure
Pharmaceutical
homogenization
Gris-PEG/
Antifungal
Bottom-up,
Novartis
precipitation
__
Estrogen antagonist
Top-down,
media milling
__
Anti-inflammatory
Top-down,
media milling
__
Antifungal
Bottom-up,
precipitation
__
Anti-inflammatory
Top-down,
media milling
Dosage form
Status
Capsule
Marketed
Tablet
Marketed
Tablet
Marketed
Tablet
Marketed
Nanosuspension In vivo
(dog)
Nanosuspension In vivo
(rat)
Nanosuspension In vivo
(rat)
Pellets
In vivo
containing dried (dog)
nanocrystal
powder
Bottom up process
(Assembling method)
Top down process
(Disintegration method)
Drug is blended with
surfactant and water
Crude drug powder
(molecules of drug)
Drug dissolution in aq. solution
containing stabilizer and then
heating to fascilitate its fusion
Drug dissolution in
suitable solvent
High pressure
homogenization
High pressure homogenization
at optimum temperature
Adding drug solution to
aq. surfactant solution
Or drug heating to fuse
Controlled
precipitation
Emulsification
Nanoedge
Supercritical
fluid
Nanosuspension
Figure 1: Various approaches for preparation of nanosuspension
Melt
emulsification
Milling
A
A describes the basic
principle of high
pressure
homogenization
technique.
And
B depicts the structural
set –up for the process.
Figure 2: Schematic representation of high-pressure homogenization
Figure 3: Schematic representation of media milling process. [Adopted from (29)]
Figure 4: Sequential procedure for controlled precipitation technique [Adopted from (26)].
Figure 5: Supercritical fluid technology process
Figure 6: Diagrammatic depiction of zeta potential
Figure 7: Mean plasma concentration of albendazole sulfoxide-time curves after oral
administration of albendazole formulations to rats at a dose of 50 mg/kg (n = 6).
ABZ-Sup:
microsuspension
including
polysorbate
80
(0.5%);
ABZ-T:
nanosuspension stabilized by polysorbate 80 (0.5%); ABZ-P: nanosuspension
stabilized by poloxamer 188 (0.5%); ABZ-HT: nanosuspension stabilized by HPMC
K4MCR P (0.5%) & Polysorbate 80 (0.5%) [Adopted from (56)].
Figure 8: Concentration of radioactivity in
tissues, post-dosing with IV itraconazole
nanosuspensions as a single dose of 10 mg/kg
[Adopted from (69)].