Advances in Colloid and Interface Science 205 (2014) 187–206

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Advances in Colloid and Interface Science

journal homepage: www.elsevier.com/locate/cis

Niosomes from 80s to present: The state of the art

Carlotta Marianecci a,1, Luisa Di Marzio b,1, Federica Rinaldi a, Christian Celia b,c, Donatella Paolino d,
Franco Alhaique a, Sara Esposito b, Maria Carafa a,⁎
a
Department of Drug Chemistry and Technologies, University “Sapienza” of Rome, Rome, Italy
b
Department of Pharmacy, University “G. d'Annunzio”, Chieti, Italy
c
Department of Nanomedicine, The Methodist Hospital Research Institute, Houston, USA
d
Department of Health Sciences, University “Magna Graecia” of Catanzaro, University Campus “S. Venuta”, Catanzaro, Italy

a r t i c l e i n f o a b s t r a c t

Available online 11 December 2013 Efficient and safe drug delivery has always been a challenge in medicine. The use of nanotechnology, such as the
development of nanocarriers for drug delivery, has received great attention owing to the potential that
Keywords: nanocarriers can theoretically act as “magic bullets” and selectively target affected organs and cells while sparing
Surfactants normal tissues. During the last decades the formulation of surfactant vesicles, as a tool to improve drug delivery,
Vesicular carriers brought an ever increasing interest among the scientists working in the area of drug delivery systems. Niosomes
Niosomes
are self assembled vesicular nanocarriers obtained by hydration of synthetic surfactants and appropriate
Drug delivery
Drug targeting
amounts of cholesterol or other amphiphilic molecules. Just like liposomes, niosomes can be unilamellar or
multilamellar, are suitable as carriers of both hydrophilic and lipophilic drugs and are able to deliver drugs to
the target site. Furthermore, niosomal vesicles, that are usually non-toxic, require less production costs and are
stable over a longer period of time in different conditions, so overcoming some drawbacks of liposomes.
The niosome properties are specifically dictated by size, shape, and surface chemistry which are able to modify
the drug's intrinsic pharmacokinetics and eventual drug targeting to the areas of pathology.
This up-to-date review deals with composition, preparation, characterization/evaluation, advantages, disadvan-
tages and application of niosomes.
© 2013 Elsevier B.V. All rights reserved.

Contents

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 188
2. Factors leading to niosome formation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 189
3. Niosomal formulation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 190
3.1. Non-ionic surfactants . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 190
3.2. Cholesterol . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 190
3.3. Charged molecules . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 191
4. Issues related to niosomal formulation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 191
4.1. Vesicle preparation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 191
4.1.1. Proniosome . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 192
4.1.2. Heating method (HM) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 192
4.1.3. Supercritical carbon dioxide fluid (scCO2) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 192
4.1.4. Membrane contactor . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 192
4.1.5. Microfluidic hydrodynamic focusing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 192
4.2. Vesicle purification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 193
4.2.1. Dialysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 193
4.2.2. Gel filtration . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 193
4.2.3. Centrifugation/ultracentrifugation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 193

⁎ Corresponding author at: Department of Chemistry and Technologies, University of Rome “Sapienza”, P.le A. Moro 5, 00185 Roma, Italy. Tel.: +39 0649913603; fax: +39 0649913133.
E-mail address: maria.carafa@uniroma1.it (M. Carafa).
1
These authors equally contributed to the work.

0001-8686/$ – see front matter © 2013 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.cis.2013.11.018
188 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206

4.3.Vesicle characterization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 194

4.3.1. Vesicle size . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 194
4.3.2. ζ-potential and surface properties . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 194
4.3.3. Bilayer characterization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 194
4.3.4. Vesicle stability . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 194
4.3.5. Entrapment efficiency . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 194
4.3.6. pH-sensitivity assessment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 195
5. Niosomes in drug delivery and targeting . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 195
5.1. Dermal and transdermal delivery . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 195
5.2. Ocular delivery . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 196
5.3. Oral delivery . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 197
5.4. Pulmonary delivery . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 198
5.5. Parenteral delivery . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 199
5.6. Gene delivery . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 201
5.7. Diagnostics/theranostics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 201
6. Natural products delivery . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 201
7. Niosomes vs liposomes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 202
8. Niosome-based patents and pre-clinical and clinical trials . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 203
9. Concluding remarks and perspectives . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 203
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 204

1. Introduction
The use of both conventional and new drug delivery systems (DDS)
allows to face the major issues in drug release: (1) unfavorable pharma-
cokinetics and biodistribution which lead to unwanted side effects
(e.g. chemotherapy), (2) early drug degradation in the bloodstream by
reticulo-endothelial system (RES), and (3) inefficient uptake at target
sites that leads to low drug efficacy. In this sense, nanocarriers offer an
innovative approach to drug delivery, providing a range of features
including cargo protection and increased dose delivery to target sites.
Actually, the complexity of human diseases may lead to ineffective
in vivo drug treatments, consequently the development of biologically
functional nanocarriers that incorporate drugs to selectively bind and
target specific cells becomes crucial. There have been many attempts
to functionalize and design nanocarriers so as to increase their efficacy;
nevertheless, although the criteria for a successful nanocarrier are quite
numerous, only few nanocarriers have actually shown significant clinical
potential. Inspired by the unique properties of non-ionic surfactant ves-
icles (NSVs), our review expands on the versatility and flexibility of NSVs
and how such nanostructures can be used for therapeutic purposes.
Due to the rapid growth in nanotechnology, it can be asserted that
the study of drug delivery and disease treatment has been revolution-
ized. By molding nanomaterials into vesicles, numerous nanocarriers
(Fig. 1) have been developed to securely deliver drugs and various
other therapeutic agents specifically into target sites. Many of the
conventional nano DDS (e.g. liposomes, micelles, and polymer-based
nanodevices) have reached the late stages of development, and some
of them were approved. Over the last two decades, the development
of synthesis and characterization techniques including the ability to
manipulate molecules and supramolecular structures for beneficial
functions blossomed into innovative-engineered nanomaterials suit-
able for biopharmaceutical/therapeutic applications. Another important
development is that nanocarriers are becoming highly integrated and
multifunctional so as to include a range of applications such as on-
demand release, specific tissue/cell type targeting, in vivo imaging and
diagnosis, and photothermal treatment [1–4].
The ultimate goal of nano DDS is to generate therapeutically and
clinically useful formulations for disease treatments in patients. While
actively studied in academia and industry, the pursuit of optimized
nanocarriers is in continuous evolution and the topic requires innova-
tion and skill in order to translate them into human trials. A successful
nanocarrier should ideally meet the following requirements:
(1) formulation with biocompatible/biodegradable/bioexcretable
materials, (2) high drug and cargo loading capacity, (3) site-specific
delivery mechanism to avoid normal cells and tissues, (4) zero or negli-
gible premature drug release, and (5) controlled release mechanism
to provide an effective dose to the target site. To date, there is a very
limited number of nanocarriers which can achieve such prerequisites.

Liposome Micelle Dendrimer Polymeric Nanoparticle

Fe3O4

Carbon Nanotube Quantum Dots Iron Oxide Gold Nanoparticle Mesoporous Silica

Fig. 1. The leading nanocarriers for drug delivery: the top row shows the representative conventional nanocarriers while the bottom row shows novel inorganic nanocarriers.
 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206
At the present state of the art, numerous scientists throughout the
world are involved in the study of vesicles and other colloidal structures
as carriers for drug delivery.
Liposomes, described by Bangham in 1965, have been prepared with
a variety of phospholipids and have been extensively studied as drug
carriers [5–7].
Although phospholipids are biodegradable and non-toxic amphi-
philes, problems arise in practical applications of liposomes, because
of the low physical and chemical stability of aqueous suspensions of
this type of vesicles. For these reasons, other amphiphilic molecules,
alternative to phospholipids, have been studied to obtain vesicular
carriers: non-ionic surfactants [8,9].
Vesicles prepared by surfactants are known as niosomes or NSVs.
Niosomes are similar, in terms of structure and physical properties, to
liposomes [8]. They are also prepared, as unilamellar or multilamellar
vesicles, following the same procedures and under the same variety of
conditions.
The research interest in niosomal formulations is recently widening
because niosomes are able to overcome some disadvantages associated
with liposomes, surfactants are easily derivatized and give a higher
versatility to the vesicular structure and moreover they have lower
costs than phospholipids.
The self-assembly of non-ionic surfactant into vesicles was first
reported in the seventies by researchers in the cosmetic industry [10].
Since then an ever increasing interest has been exhibited on the applica-
tion of niosomes (Fig. 2) in the field of pharmaceutics, cosmetics and
food industry, leading to the publication of more than 1200 research ar-
ticles, about 200 patents and 6 clinical trials from 1980 (from SciFinder).
The aim of this paper is to present an overview on niosome prepara-
tion and characterization as well as a description of their applications,
with particular attention to more recent pharmaceutical formulations.
Considering that this is a fast-moving research field, all aspects of
niosomal applications cannot be included or discussed. This review
will provide an overview on the increasing interest on niosomes in the
field of disease diagnosis and therapy as well as in the cosmetic field.
2. Factors leading to niosome formation
The self-assembly of amphiphilic molecules into closed bilayers,
both in the case of liposomes and niosomes, is not spontaneous but it
involves some input of energy, for instance by means of physical
shaking (hand-shaking, ultrasound, heat, etc.) [11,12]. Moreover, ther-
modynamically stable vesicles are formed only in presence of appropri-
ate mixtures of surfactants and charge inducing agents. Although
previous studies showed that niosomes have a higher resistance to
micellar solubilization than liposomes [13,14], similar factors lead to
the formation of both types of vesicles.
Fig. 2. General stage of the development of niosomal carriers in pharmaceutical applications
from 1980 (from SciFinder).
189
The main factors that play a role in amphiphile self-assembling
are thermodynamic and physicochemical parameters such as the
hydrophilic-lipophilic balance (HLB) and the geometric feature of the
amphiphilic molecules; but several other factors are relevant for vesicle
formation and properties and must be taken into consideration: in
particular aqueous interlayer, lipid chain-length, chain-packing and
membrane asymmetry. The energy required to form vesicles with am-
phiphilic molecules has three contributors related to the surface energy,
the mechanical energy due to overpressure and the chemical potential
excess. The association of non-ionic surfactant monomers into vesicles
upon hydration is a result of the high interfacial tension between
water and hydrocarbon portion (or any other hydrophobic group) of
the amphiphile which causes the association of these groups. At the
same time the steric, hydrophilic, and/or ionic repulsion among the
head groups ensures that these groups are in contact with water. Of
course also monomer concentration and temperature play an important
role in vesicle formation.
Thermodynamically this self-assembly must contend against a
negative entropy component (ΔS) and the reduction in free energy
(ΔG) and is only achieved by the favorable enthalpy (ΔH) contribu-
tion arising from the van der Waals attractions, the hydrophobic
forces, the hydrogen bond formation and the screened electrostatic
interactions.
As above pointed out, vesicle formation may depend on the HLB
value; thus the guidance offered by the HLB number is useful in the
evaluation of new classes of compounds for their vesicles forming ability.
With the sorbitan monostearate surfactants (Sp), an HLB number
between 4 and 8 was found to be compatible with vesicle formation
[8]. On the other hand, it was actually shown that, owing to their high
aqueous solubility, hydrophilic surfactants cannot form free hydrated
units (vesicles) but these free units rather aggregate and coalesce to
form lamellar structures [15,16]. Consequently, niosomes are not gener-
ally formed with surfactants with a HLB value between 14 and 17 [17]
and the addition of cholesterol plays a fundamental role in vesicle
formation with surfactants with a HLB value around 10 [18].
The geometric feature of the single vesicle-forming unit may be
another important factor in determining the type of aggregate formed
in aqueous environments [19]. To explain the behavior of non-ionic
surfactants in water, a hydrophobic effect was proposed several years
ago, as the essential strength leading to an enhancement of global
system-free energy. Shapes of the spontaneously formed association
colloids can be predicted with considerable certainty using nominal
geometric parameters of the surfactant molecule.
The critical packing parameter (CPP also named as Ps) was discussed
by Israelachvili et al. in 1976 [20] and defined by relation:
V
CPP ¼ ð1Þ
a0 lc
where:
– V is the tail volume of the molecule,
– a0 is the surface area per molecule at the hydrocarbon–water inter-
face,
– lc is the critical, i.e. the greatest, span of a fluid molecular chain in an
aggregate.
According to the CPP value, the shape and size of the equilibrium
aggregate should evolve from spherical micelles (CPP ≤ 1/3) to cylin-
drical micelles (1/3 ≤ CPP ≤ 1/2), bilayers (1/2 ≤ CPP ≤ 1) or inverse
micelles (CPP N 1) (Fig. 3).
The influence of polar head surface, a0, on the ability to form vesicles
was confirmed by a comparison between Polysorbates (Tw) 21 and 20
bearing the same alkyl chain (dodecanoic acid) but different hydrophilic
head groups (Tw21:4 PEG units; Tw20:20 PEG units) [21], thus
influencing also the HLB value (Tw20 HLB: 16.7; Tw21 HLB: 13.3).
190 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206
Fig. 3. The relationship between the CPP and the morphology of self-assembled amphiphilic molecules.
Recently it was reported by Cevc [22] that scrutinizing amphiphile
aggregation in terms of the popular molecular descriptors (esp. the
CPP or HLB number) could be “too static” to foretell reliably and quanti-
tatively bilayer vesicle formation. Consequently, a different predictor
was proposed: the effective area per lipid chain (cross-section of a
“tail”, Ac), which seems to correlate, quasi-exponentially, with the
ease of bilayer vesicle formation and bilayer deformability.
According to this parameter, an Ac value above 0.43 nm2 will identify
a micellar structure; a value slightly below this limiting area should
imply bilayer vesicle formation. On the other side, amphiphiles with
an appreciably smaller area per chain, Ac ≪ 0.43 nm2, prefer to form
multilamellar structures.
It must be underlined that the assigning of an absolute value to
geometric or molecular parameters is at least misleading, if not wrong.
This is due to such molecular and geometric descriptors sensitivity to
the boundary conditions and to the different possibilities to combine
components and/or experimental conditions in surfactant vesicle
preparation.
It should be more correct to consider all reported parameters as
adjustable and it can consequently be useful to combine them in
predicting amphiphilic molecules capability to aggregate into bilayer
vesicles.
3. Niosomal formulation
The efficacy of a drug delivery system is strictly dependent on
its components, that should be characterized in terms of physical–
chemical and clinical properties needed for the formation of niosomal
systems with specific characteristics, together with biocompatibility
and relevance in clinics. The basic components of niosomes include
non-ionic surfactants, hydration medium and lipids such as cholesterol.
3.1. Non-ionic surfactants
Just as ionic surfactants, non-ionic surfactants are amphiphilic (or
amphipathic) molecules that have two distinct regions in their chemical
structure, one of which is water-liking or hydrophilic and the other is
water-hating or hydrophobic. The two portions of such molecules may
be linked by ether, amide or ester bonds.
Non-ionic surfactant vesicles can be prepared from different types of
molecules, such as: amino acids, fatty acids, amides, alkyl esters and
alkyl ether surfactants, the last one being mostly employed for such
purposes [23–28].
Alkyl ether surfactants can be broadly divided into two classes based
on the nature of the hydrophilic head group: alkyl ethers in which the
hydrophilic head group consists of repeating glycerol subunits, related
isomers or larger sugar molecules and those in which the hydrophilic
head group consists of repeating ethylene oxide subunits.
The first niosomes were formulated using cholesterol and single-
chain surfactants such as alkyl (usually from C12 to C18) oxyethylenes
[8]. Polyglycerol monoalkyl ethers and polyoxylate analogs are the
most widely used single-chain surfactants. Ether surfactants, suitable
for noisome formulations are composed, in their lipophilic moiety, of
monoalkyl or dialkyl chains. The latest ones, being similar to phospho-
lipids, and possess higher encapsulation efficiency. Esther type amphi-
philic surfactants are also used for niosome formulation. They are
degraded by estherases to triglycerides and fatty acids. Although these
types of surfactants are less stable than ether type ones, they are usually
less toxic. Finally, also glucosides of myristil, cethyl and stearyl alcohols
can be used to form niosomes.
While the number of hydrophobic permutations is at present quite
limited, there has been a wide variety of hydrophilic head groups in
vesicle forming surfactants, and the design of new molecules is in con-
tinuous evolution, according to the specific vesicle structure that are
needed. Some recent studies report the synthesis and the characteriza-
tion of new surfactants with peculiar physical–chemical properties,
showing promising applications in the field pharmaceutical colloid
science. As an example, a new non-ionic surfactant molecule, in which
the hydrophilic region consists of azacrown ether units, was synthesized
by Muzzalupo and co-workers. This compound, known as bola-
surfactant, is composed of two identical azacrown ether units, as polar
heads, linked to a long alkyl chain [29–31]. The authors demonstrated
that the bola-surfactant is able to assemble in colloidal structures if
associated with cholesterol or other amphiphilic molecules [30].
3.2. Cholesterol
It is well known that cholesterol (CHOL) influences the physical
properties and structure of niosomes because of its interaction with
non-ionic surfactants. Several surfactants form vesicles only after
CHOL addition (up to 30–50 mol%). The amount of CHOL to be added
depends on the HLB value of the surfactants. As the HLB value increases
above 10, it is necessary to increase the CHOL concentration in order to
compensate the effect of the larger head groups on the critical packing
parameter (CPP), previously discussed [32,33].
 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206
Tween® 20
x+y+z+w = 20
Fig. 4. Possible hydrogen bonding interaction between the 3-OH group of the cholesterol
and the carbonyl oxygen and also weaker interaction at oxygen of the ester bond.
The water soluble detergent Tw20 (HLB value = 16.7) forms stable
non-ionic surfactant vesicles in the presence of equimolar CHOL con-
centration because in absence of CHOL it would be well hydrated and
can be found as free monomers in solution (CMC = 60 mg/l, in water
at 20 °C) [34–37]. It was suggested that an interaction occurs between
the hydrophobic portion of the amphiphile (phospholipid/surfactant)
next to head group and the 3-OH group of CHOL in vesicular systems
at an equimolar ratio (CHOL:amphiphile) (Fig. 4) [38] and this interac-
tion could explain the effect of CHOL on the formation and hydration
behavior of Tw20 niosomal membranes that were found to coexist
with unsolubilized CHOL crystals [39].
The role of CHOL in the surfactant vesicle formation is also supported
by the work of Manconi et al. [40]. In this study it was reported that a
hydrophilic non-ionic surfactant, decylpolyglucoside, is able to form
stable and spherical vesicles only in the presence of high amounts of
CHOL. These results also suggest that CHOL increases the cohesion
among the non-polar portions of bilayer, thus improving the cohesion
of niosomal membrane [40].
The effect of CHOL in amphiphile bilayers is mostly the modulation
of their mechanical strength and their permeability to water [41,42],
properties that become particularly important under severe stress
conditions (e.g. in vivo) [42], nevertheless it should be pointed that pub-
lished results are not always in agreement. The inclusion of cholesterol
was successfully used by Manosroi et al. (2003) to increases the hydro-
dynamic diameter and the entrapment efficiency of niosomes prepared
by various non-ionic surfactants [43].
On the other side, it was recently observed that by decreasing
the molar ratio of cholesterol to Sp 60, the entrapment of a model
A B
Fig. 5. The average hydrodynamic diameter 2R and electrophoretic mobility course of polylysine/niosome structure as a function of the positive/negative charge ratio (A). Schematic
representation of niosomal/polyelectrolite «patchwork like» interaction (B).
191
hydrophilic within the obtained niosomes correspondingly increased,
regardless of the type of stabilizer (Dicetyl phosphate or Solutol HS
15) [44]. Furthermore, CHOL stabilizes niosomes towards destabilizing
effects of plasma and serum components and decreases the permeabil-
ity of vesicles to entrapped solute thus inhibiting leakage [11,45].
According to the above reported results, the still controversial role of
CHOL is evident. The opportunity to add CHOL, and its amounts, needs
to be evaluated case by case, depending on the physical–chemical char-
acteristic of surfactants and loaded drugs.
3.3. Charged molecules
One of the methods used to stabilize niosomes is based on the addi-
tion of a charged molecule to the bilayer. Dicetyl phosphate (DCP) and
phosphatidic acid are known as negatively charged molecules, while
stearylamine (SA) and cetylpyridinium chloride are positively charged
molecules which are both commonly used for preventing aggregation
of niosomes [9]. Normally, the charged molecule is added in niosomal
formulation in an amount of 2.5–5 mol% because a higher concentration
of charged molecules could inhibit the formation of niosomes [46,47]. It
is widely reported that zeta (ζ)-potentials over |30| mV are necessary
for full electrostatic stabilization; potentials between |5| and |15| mV
represent the region of limited flocculation; while between |5| and |3|
mV correspond to the maximum flocculation. Thus, particle aggregation
is less likely to occur for charged particles (high ζ-potential) due to their
electric repulsion. However, this rule cannot strictly be applied for
systems containing steric stabilizers, because the adsorption of such
steric stabilizers decreases the ζ-potential because of the shift in the
shear plane of the particle. Moreover, the strong dependence of the
ζ-potential on the ions present in the medium should be taken into
account [48]. Charged molecules can be also added to prepare charged
niosomes useful for an increase of drug encapsulation efficiency [49],
for skin permeation enhancement [50] and for hybrid niosomal com-
plex formation [51], (Fig. 5).
4. Issues related to niosomal formulation
4.1. Vesicle preparation
Numerous methods for non-ionic surfactant vesicle preparation are
reported, however only some of these are actually used to manufacture
liposomes and have been adapted/used as such. Unlike liposomes, the
niosome preparation is usually easier given to the surfactant resistance
192 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206

against air oxidation. The hydrating temperatures used to make
niosomes should usually be above the gel to liquid phase transition tem-
perature of the system. The formation of vesicular assemblies requires
the input of some form of energy [52] and all the experimental methods
surveyed consist of the hydration of a mixture of the surfactant: lipid at
elevated temperature followed by optional size reduction to obtain a
colloidal dispersion. Moreover, the elevated temperature used does
not represent an issue, because non-ionic surfactants are more stable
than phospholipids. Furthermore the hydrating temperature must be
chosen evaluating drug melting point. Many techniques such as the
thin film hydration, the organic solvent injection method, the reverse
phase evaporation method and the bubble method have been previous-
ly described for the preparation of niosomes [9,53] (Table 1).
Below the more recent and innovative methods for niosome prepa-
ration are reported:
4.1.1. Proniosome
Proniosome-produced niosomes involve coating a water-soluble
carrier (e.g. sucrose stearate, maltodextrin or mannitol) with non-
ionic surfactants. 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” [54,55].
Important advantages of proniosome-derived niosome are the
reduction of niosome physical instability such as aggregation, fusion
and leaking and the increase of drug entrapment efficiency [56].
In addition, proniosomes are obtained as a dry power, so further
processing is possible. To provide convenient unit dosing, the
proniosome powder may be processed to make beads, tablets, or cap-
sules. The proniosome powder can be implemented in a ‘point-of-use’
application.
4.1.2. Heating method (HM)
The heating method developed by Mozafari to produce vesicle
involves the hydration of an amphipathic molecule in an aqueous solu-
tion containing 3 vol.% polyol at high temperature. This is a non-toxic,
scalable and one-step method [57].
4.1.3. Supercritical carbon dioxide fluid (scCO2)
In the last years a new method, the supercritical reverse phase
evaporation (scRPE) method, for the preparation of niosomes using
supercritical fluids has been described by Manosroi [58]. The main
advantages of this method are: the use of no inflammable, volatile,
and non-toxic organic solvent, one step and easy scale-up production
of surfactant vesicles. However, the niosomes prepared with the scCO2
method have large unilamellar structure with size between 100 and
440 nm. A smaller size distribution of the vesicles could be obtained
by the combination of the scCO2 technique with other methods such
as sonication or polycarbonate extrusion.
4.1.4. Membrane contactor
The scaling-up ability of niosome production, from laboratory scale
using a syringe-pump device to a pilot scale using the membrane
contactor module (Fig. 6), was reported by Pham [59].
4.1.5. Microfluidic hydrodynamic focusing
Conventional bulk method of niosome preparation requires bulk
mixing of two liquid phases, which is time-consuming and not well-
controlled. Such mixing conditions often lead to large niosomes with
high size polydispersity, thus affecting the consistency of niosome
dosage or imaging quality. The new method of niosome by microfluidic
hydrodynamic (Fig. 7) leads to improve the size and size distributions of
niosomes, by taking advantage of the rapid and controlled mixing of

Table 1
Niosome preparation by “conventional” techniques.

Preparation Method description Reference

Thin layer evaporation (TLE)
Melted amphiphile injection
Solid amphiphiles/hot water
Micellar solution/enzymes
Ether injection
Trans membrane pH gradient method
Reversed phase evaporation
Bubbling of nitrogen
Microfluidization
The formation of a surfactant:lipid film by the evaporation of an organic solution of surfactants:lipids. This film is then [61]
hydrated with aqueous solutions to obtain multilamellar vesicles which are further treated for size/lamellae reduction.
Disadvantage: use of organic solvent.
Advantage: easy technique for bench researches.
The injection of melted lipids:surfactants into a highly stirred heated aqueous phase. [62]
Disadvantage: no applicability for thermosensitive drugs.
Advantage: no organic solvents, which are expensive, difficult to remove completely and hazardous.
The addition of the warmed aqueous phase to a mixture of the solid lipids:surfactants. [63]
Disadvantage: no applicability for thermosensitive drugs.
Advantage: no organic solvents, which are expensive, difficult to remove completely and hazardous.
Niosomes may also be formed from a mixed micellar solution by the use of enzymes. [64]
Disadvantage: enzymatic degradation of active substances.
Advantage: no organic solvents, which are expensive, difficult to remove completely and hazardous.
In this method, the surfactans and drug are dissolved in diethyl ether and injected slowly into an aqueous phase, which is [65]
heated above the boiling point of the organic solvent. This produces large unilamellar vesicles, which are further treated for
size/lamellae reduction
Disadvantage: no applicability for thermosensitive drugs.
Advantage: easy technique for bench researches.
After film formation (TLE) a solution of an acidic compound (generally citric acid) is added and mechanically stirred Freeze- [11]
thawing cycles are then carried out on the obtained product after which an aqueous solution of the drug is added and the
mixture is vortexed. pH is then finally adjusted to 7–7.2 by addition of a buffer solution.
Disadvantage: use of organic solvent.
Advantage: high drug entrapment efficiency.
The surfactants are dissolved in a mixture of ether and chloroform and an aqueous phase containing the drug is added. The [11]
resulting two-phase system is then homogenized and the organic phase evaporated under reduced pressure to form
niosomes dispersed in the aqueous phase.
Disadvantage: use of organic solvent
Advantage: high drug entrapment efficiency
Nitrogen is passed through a sample of homogenized surfactants leading to the formation of large unilamellar vesicles, which [66]
are further treated for size/lamellae reduction.
Advantage: no organic solvent.
A solution of surfactants and drug is pumped under pressure from a reservoir through an interaction chamber packed in ice at [67]
a rate of 100 ml/min. From the interaction chamber, the solution is passed through a cooling loop to remove the heat
produced during micro fluidization.
Disadvantage: no applicability for thermosensitive drugs.
Advantage: no organic solvents, which are expensive, difficult to remove completely and hazardous.
 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206 193

Permeation under
Pressurized vessel Organic phase
applied pressure
Dispersed phase

Tangential flow Membrane

N2 bottle

Niosome

Pump
Magnetic
stirrer

Aqueous phase

Fig. 6. Schematic drawing of the membrane experimental set-up for niosome preparation.

two miscible fluids (i.e., alcohol and water) in microchannels. Different
parameters might affect assembly of niosomes, such as (1) conditions
for the microfluidic mixing, (2) chemical structures of the surfactant
used (i.e., Sp 20, Sp 60, and Sp 80), and (3) device materials for the
microchannel fabrication [60].
removed. Although it may be argued that the use of systems in which
the drug is partially encapsulated in niosomes may eventually yield
systems with a beneficial biphasic biodistribution profile. This drug
delivery system could lead to an initial burst effect to initiate therapy
followed by a sustained maintenance dose. The methods used for the
removal of unentrapped material include different techniques:

4.2. Vesicle purification 4.2.1. Dialysis

The hydration of niosome mixtures seldom leads to complete drug
encapsulation, regardless of the drug loading optimization steps under-
taken. It is thus often a requirement that unencapsulated drug must be
Dialysis is based on diffusion and osmosis of solutes and fluid across
a semi-permeable membrane. Several authors used this technique
to purify their vesicle dispersion from unentrapped materials: the
prepared niosomes are filled into dialysis bags and the free drug is
dialyzed against buffer saline [30,68,69].

4.2.2. Gel filtration

In order to separate loaded NSVs from unentrapped substances,
Phosphate Phosphate purified vesicle dispersion can be obtained by means of gel-filtration
Buffer Buffer on Sephadex G75 or G50 or Sephadex G25 [70–76].
Gel-filtration chromatography is a popular and versatile technique
that allows the effective separation of free molecules in high yield.

Fig. 7. Schematics of niosome self-assembly by microfluidic hydrodynamic focusing. A
central stream containing the surfactant mixture in isopropyl alcohol is focused by
adjacent streams of phosphate buffer in a microfluidic format. FRR = flow rate; QB =
buffer volumetric flow rates; QS = alcohol volumetric flow rate; Wf = width of the
focusing stream; τmix = diffusive mixing time.
4.2.3. Centrifugation/ultracentrifugation
To purify niosomes, the centrifugation and ultracentrifugation sepa-
ration was reported [77].
In particular the centrifugation process was carried out to remove
the unentrapped genetic material from niosomes, following the gradi-
ent density centrifugation approach [78–81].
Some authors combined the previously mentioned centrifugation
processes with Sephadex chromatography mini-columns (Minicolumn
centrifugation method) [80,81]. This procedure compared to other
purification methods is applicable to a variety of solutes and 92 to
100% recovery can be achieved for niosomes with no dilution of the
niosomal preparation.
Since all these methods show, at the same time, advantages and
disadvantages, consequently the choice of the method must take into
account different factors. For instance, for industrial purposes it may
be more appropriate to concentrate efforts and resources on the
achievement of high levels of drug loading, thus avoiding separation
steps or to consider systems in which the unentrapped drug may be
useful as a specific priming dose.
194 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206
4.3. Vesicle characterization
A rational characterization of nanosystems is needed for the control
of the quality of the product, which is a prerequisite for their develop-
ment and clinical applications. Having direct impact on the stability
and critical importance on their in vivo performance, niosomes param-
eters such as morphology, size, polydispersity index, number of lamel-
lae, ζ-potential, bilayer fluidity, composition, encapsulation efficiency,
carrier-bioactive interaction and chemical stability must be evaluated
(Table 2).
4.3.1. Vesicle size
Determination of nanocarrier size distribution is a fundamental
parameter in the determination of physical properties and stability of
bioactive delivery formulations. Size distribution, along with composi-
tion, defines plasma pharmacokinetics, biodistribution, and stability of
nanocarriers and their associated/entrapped substances in plasma and
other organs [82].
Niosome diameters can be determined using microscopy or dynamic
light scattering (DLS or photon correlation microscopy, PCS).
DLS provides simultaneously cumulative information of the average
particle and of the particle size distribution. One problem with DLS is
that it does not provide information on the shape of the nanosystem
and it assumes any aggregation of more than one particle as one single
unit. The microscopic approach is commonly used to characterize the
structure/morphology/geometry of the nanocarriers.
Several electron microscopy techniques can be employed for
nanocarrier characterization. These include: scanning electron micros-
copy (SEM), preferentially for solid products; transmission electron
microscope (TEM), for liquid state sample; negative-staining transmis-
sion electron microscope (NS-TEM); freeze fracture transmission elec-
tron microscopy (FF-TEM); cryo-transmission electron microscopy
(cryo-TEM), atomic force microscopy (AFM) and scanning tunneling
microscope (STM), used in 1982 by Binnig and coworkers, for the
study of micro- and nano-scale structures. Furthermore, STM in partic-
ular is very useful in determining the bilayer thickness of liposomes,
nano-liposomes, and niosomes by its analytical ability in the vertical
axis [30,31,70,71,83–88].
It must be pointed out that microscopy techniques can give artifacts
and hence it can be useful to combine different techniques to obtain
more reliable results.
4.3.2. ζ-potential and surface properties
ζ-potential value has critical importance on vesicle stability and
in vivo fate. Several authors confirmed the importance of ζ-potential
measurements to assess vesicle formation, to study drug/vesicle inter-
action and formulation stability [70,71,89].
ζ-potential is also a critical parameter to be taken carefully into
account in the formation of aggregates, from charged niosomes and
oppositely charged polyions, when these vesicles are proposed as drug
carriers [90,91].
Table 2
Different approaches to niosome characterization.
Niosome parameter/property Applied technique
Size DLS/PCS; SEM (preferentially for solid products);
TEM (liquid state sample); NS-TEM; FF-TEM; cryo-TEM; AFM; STM
ζ-potential Electrophoretic mobility.
Bilayer characterization Fluorescence polarization.
Fluorescence anisotropy.
AFM; NMR; SAXS; EDXD.
Stability DLS; Turbiscan; microscopy techniques.
Release studies.
Entrapment efficiency UV/VIS; HPLC; fluorescence.
For genetic material: gel electrophoresis followed by UV densitometry.
pH-sensitivity assessment Fluorescence spectroscopy; NRET; SAXS
4.3.3. Bilayer characterization
Niosomes, as liposomes, can be morphologically classified according
to the number of membrane bilayers (lamellae) in uni- and multi-
lamellar vesicles. Uni-lamellar vesicles are characterized by the pres-
ence of one surfactant bilayer while multi-lamellar vesicles by several
concentric surfactant bilayers in an onion-like skin arrangement. The
number of lamellae can be determined by AFM, NMR and small angle
X-ray scattering (SAXS). The latter method, together with the in situ
energy-dispersive X-ray diffraction (EDXD), has been also used to char-
acterize the bilayer thickness [84,85,92–95].
Another feature of the niosomes is the fluidity of their membrane,
which allows membrane deformation without disrupting bilayer integ-
rity (vesicle stability) and drug-release ability.
These parameters can be measured by means of the mobility of fluo-
rescence probes (e.g. 1,6-diphenyl-1,3,5-hexatrine, DPH, and pyrene) as
a function of temperature and/or time [85,96–98].
The microviscosity of niosomal membrane can also be determined
by fluorescence polarization in order to study packing structure of the
vesicular membrane [58].
4.3.4. Vesicle stability
Vesicle stability is a complex issue and involves chemical stability,
physical stability and biological stability, which are all inter-related.
The evaluation of these parameters is fundamental to determine the
potential in vitro/in vivo applications in nanomedicine. Biological stabil-
ity, however, depends on the presence of agents that interact with the
vesicular structure after administration and it therefore depends also
on the administration route. Generally, stability is determined by
means of size and ζ-potential variations (DLS, Turbiscan Lab® Expert
or microscopy techniques) or by evaluation of the release rate of differ-
ent probes as a function of time and/or temperature, in absence or in
presence of biological fluids [99].
4.3.5. Entrapment efficiency
The entrapment efficiency (EE) of vesicular systems can be defined
as the amount of active substances loaded within the niosomal structure
and can be expressed as:
amount entrapped
EE ¼  100 ð2Þ
total amount
where “total amount” is the total amount of drug used in the prepa-
ration. The entrapment efficiency can be determined by spectropho-
tometry or, in the case of genetic materials (e.g. luciferase plasmid),
by gel electrophoresis followed by UV densitometry. In addition, the
entrapment efficiency can be fluorometrically evaluated using a hydro-
philic fluorescent marker (e.g. calcein) and then calculated according to
the following equations
Moleprobe entrapped
EE ¼  100 ð3Þ
Molesurfactant
Reference
[30,31,70,71,82–88]
[70,71,89–91]
[59,71,84,85,92–95]
[154,99]
[30,37,68]
[91,102–104]
 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206 195

lin −ltx  r  100
EE ¼  100
ltotal −ltx  r
where Itotal and Iin are the fluorescence intensity before (Itotal) and after
(Iin) addition of cobalt chloride (CoCl2) respectively, Itx is the fluores-
cence intensity after addition of CoCl2 and a detergent such as
n-propanol or Triton X-100, r is the volume correction factor (~1.04).
The second equation does not require vesicle purification [30,37,68,100].
and targeting vary by cell or tissue type, so the behavior of niosomes
ð4Þ
in vitro/in vivo may not be predictive of another.
An ever increasing number of publications on potential and practical
niosome applications in nanomedicine appeared in the scientific litera-
ture during the last years and most of them take into account the impor-
tance of nanovector characterization.
To highlight the state of the art of niosome pharmacological and
diagnostic applications, we attempt to summarize in this section cur-
rently available information on niosomal carriers that are endowed
with desiderable properties.

4.3.6. pH-sensitivity assessment 5.1. Dermal and transdermal delivery

Stimuli-sensitive NSVs, which release their cargo in response to
external stimuli, represent interesting alternatives for therapies direct-
ed towards solid tumors and other spatially well defined targets. The
gradual decrease in pH, experienced by niosomes that are internalized
via endocytosis, is actually a very useful intrinsic stimulus and several
pH-sensitive niosome formulations, based on this strategy, have been
developed and biologically evaluated [91,101,102].
The steady-state fluorescence spectroscopy was also used to charac-
terize the pH-sensitivity of the surfactant vesicles. The fluorescent
probes chosen for such experiments were hydroxypyrene-1,3,6-
trisulfonic acid (HPTS) and Nile Red. The maintenance of pH-sensitivity
in serum must be appropriately verified by monitoring fluorescent
probe release at different pH in the presence of serum. The pH effect
on vesicle bilayer can be also evaluated by SAXS. This technique is able
to evaluate bilayer thickness changes in different pH environments
[91,103].
An assay, performed at different pH, for vesicle-vesicle fusion evalu-
ation, is based on the non radioactive resonance energy transfer (NRET)
between two fluorescent lipid analogs N-(7-nitrobenz-2-oxa-1,3-
diazol-4-yl)-1,2-dihexadecanoyl-snglycero-3-phosphoethanolamine
(NBD-PE) (energy donor), and N-(lissamine rhodamine B sulfonyl)-
phosphatidylethanolamine (N-Rh-PE) (energy acceptor). This tech-
nique provides a means by which lipid mixing during vesicle fusion
can be followed since fusion of the vesicles will result in intermixing
of the membrane lipids and probe dilution [104].
5. Niosomes in drug delivery and targeting
Niosomal carriers are suitable for the delivery of numerous pharma-
cological and diagnostic agents. According to this statement it must be
pointed out that an emerging paradigm for the design of effective
nanocarriers is the modification of physico-chemical parameters to
influence the drug delivery and targeting. These factors are often
difficult to vary independently, so the contribution of each is difficult
to generalize. Furthermore, processes involved in efficient drug delivery
Dermal drug delivery is the topical application of drugs to the skin in
the treatment of skin diseases.
This approach has the advantage that high drug concentrations can
be localized at the site of action, minimizing systemic absorption, thus
reducing systemic side effects. On the other side, transdermal drug
delivery uses the skin as an alternative route for the drug towards the
systemic circulation. This drug delivery route shows several advantages
over the conventional oral and parental routes such as: avoidance of the
risk and inconvenience of intravenous therapy (non invasive), avoid-
ance of first pass hepatic metabolism (avoiding the deactivation by
digestive and liver enzymes), which leads to an increase of drug bio-
availability and efficacy, no gastrointestinal degradation (pH, enzymatic
activity, drug interaction with food, beverages and other orally adminis-
tered drugs), alternative to oral administration when such route is
unsuitable (e.g., vomiting and diarrhea). However, transdermal drug
delivery shows the major disadvantage of a low penetration rate
through the skin. Actually, only limited number of drugs can be formu-
lated as transdermal delivery systems due to the functions of the stra-
tum corneum (SC), which provides the main barrier to permeation.
Although a number of penetration enhancement techniques like
iontophorosis, ultrasound (phonophoresis and sonophoresis),
magnetophoresis, electroporation, laser radiation and photomechanical
waves have been used, the number of drugs that can be administered by
transdermal route is still limited. During the last decade, vesicle applica-
tion in drug delivery through the skin has undergone intense investiga-
tions and it seems to be promising [105–109] (Table 3). Both a
permeation enhancer effect and direct vesicle fusion with the SC may
contribute to the enhanced permeation of drugs loaded in niosomes.
NSVs are becoming popular in the field of topical drug delivery due to
their outstanding characteristics and properties induced by their pres-
ence in the formulation such as enhanced drug penetration, sustained
drug release, increased drug stability and ability to carry both hydrophil-
ic and lipophilic drugs [110]. Fang et al. showed that enhancing effect for
skin permeation of enoxacin for niosomes was greater than that of

Table 3
Dermal and transdermal application of niosomes.

Compositions Drug Experimental model Results Reference

Span40/Chol, Span60/Chol,
Span60/DCP/Chol
Span40/Chol/DCP, Span60/Chol/DCP,
Brij30/Chol/DCP,
TritonCG110/Chol/DCP
Oramix®CG110 or
Oramix®NS10/Chol/DCP or SA
Tw20/Chol, Tw20/Chol/Chems,
Sp60/Chol, Sp60/Chol/Chems
Span60/Chol in hydrogel
Span40/Chol niosomal gel
Enoxacin In vitro diffusion: Franz cells — nude mouse skin. Niosome fusion at the interface of the stratum [111]
corneum: high local drug concentration.
Tretinoin In vitro permeation: Franz cells — silicone membrane High local drug concentration. High trenitoin [74,75]
(TRA) (Perthese®). permeation Best formulation: TritonCG 110-
Niosomes
In vitro permeation: Franz cells — new-born pig skin. Drug delivery modulated by varying the structure
and/or bilayer composition: highly hydrophilic
surfactants (HLB = 16) improve TRA diffusion
through pig skin; Brij® 30 or Oramix NS 10 (HLB 9 and
11, respectively) enhance TRA cutaneous retention
Ibuprofen In vitro permeation: Gummer cells — rat skin. The more lipophilic the surfactant, the greater its [113]
ability to penetrate into the skin
Meloxicam Carrageenan induced rat paw edema method. Marked increase in the percentage inhibition of edema [124]
in animals treated with meloxicam vesicular gel.
Lopinavir Ex vivo skin permeation studies; histopathological Better safety, bioavailability and deposition in [123]
studies; in vivo bioavailability study in male Wistar comparison to ethosomal gel
rats.
196 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206

liposomes composed of dimyristoyl phosphatidylcholine. Manconi et al.
showed that higher tretinoin stability was obtained when it was incor-
porated in niosomes rather than in liposomes [111,112].
Recently pH-sensitive NSVs obtained with Tw20 or Sp 60 mixed
with CHOL and cholesteryl hemisuccinate (CHEMS), a derivative of
cholesterol as a pH-sensitive molecule, were proposed for topical
delivery of ibuprofen. Only niosomes with Sp 60 and CHEMS showed a
significant increase of in vitro skin permeation of the drug [113].
Other drugs loaded into niosome for skin administration were:
gallidermin; terbinafine; curcumin; vinpocetine; ellagic acid and am-
monium glycyrrhizinate [114–120].
An interesting study reports a comparison between lopinavir
niosomal gel and ethosomal gel formulations. The niosomal formulation
containing lopinavir, Sp 40, and CHOL possessed a high entrapment
efficiency for the drug with a minimum mean vesicular diameter.
Ex vivo skin permeation studies of lopinavir, as well as those of the
fluorescent probe coumarin, revealed a better deposition of ethosomal
carriers but a better release with niosomal carriers. Histopathological
studies indicated the better safety profile of niosomes over ethosomes.
In vivo bioavailability study in male wistar rats showed a significantly
higher extent of absorption of lopinavir via transdermally applied
niosomal gel when compared with its oral suspension. Taken together,
these findings suggested that niosomal gel holds a great potential of
being used as novel, nanosized drug delivery vehicle for transdermal
lopinavir delivery [121].
To enhance transdermal adsorption of drugs, elastic NSVs, niosome
loaded hydrogel systems [122–124] and proniosomal gel were also
proposed [125–130]. Topical immunization is novel and needle free
strategy involving vaccine delivery through topical application of anti-
gen and adjuvants directly or via a suitable carrier system on intact
skin. Anatomy and physiology of skin attracts scientists in developing
topical carrier systems for enhanced delivery of bioactive substances
and niosomes have been exploited for topical immunization [131,132].
The application of niosomes as dermal/transdermal delivery systems
is receiving an increasing attention and they are becoming popular in
the field of topical delivery due to their outstanding characteristics
like enhancing the penetration of drugs and providing a sustained pat-
tern of drug release [133].
5.2. Ocular delivery
There are many anatomical and physiological barriers to overcome
in topical ocular drug delivery and a strategy to cross them implies the
use of bioadhesive carriers (Table 4).
Mucoadhesive timolol maleate NSVs coated with chitosan or
carbopol have been used to extend optimal ocular pharmacodynamics
over a prolonged period, and they showed only limited systemic
absorption and side effects [134,135].
Niosomal formulations, prepared using various surfactants (Tw20,
Tw60, Tw80 or Brij 35) showed a slower in vitro release of gentamicin,
when compared to the drug solution or promote absorption of
cyclopentolate by preferentially modifying the permeability character-
istics of the conjunctival and scleral membranes [136,137].
Recently, Sp 60-based oval giant niosomes were reported to be
capable of controlling naltrexone hydrochloride permeation and
enhance its corneal permeability and were found practically non irritant
[138].
In the study NSVs and discomes for ocular delivery of naltrexone
hydrochloride (NTX) were evaluated for their spreading, rheological
properties and their ability to prevent NTX autoxidation in aqueous
solutions. The prepared niosome formulation was significantly more
viscous than the aqueous solution. The lipid content, size and composi-
tion of NSVs were the main factors affecting the viscosity of the
niosomal dispersions. Exposure of NTX solution to artificial daylight
illumination can produce extensive degradation of NTX due to oxida-
tion, while the prepared formulations were able to significantly protect
the encapsulated NTX from the photo-induced oxidation when
compared with free NTX solutions. The investigated niosomes showed
a potential ocular delivery for NTX.
Furthermore, NTX loaded-niosomes were proposed as a promising
treatment for corneal disorders linked to diabetes mellitus (diabetic
keratopathy) [139].
Niosomal formulations showed a fairly high retention of acetazol-
amide inside the vesicles (75%) at a refrigerated temperature for up to
three months. Each of the tested acetazolamide niosomes produced
significantly less intraocular pressure than the free drug and basic
niosome solution [140]. Studies of aqueous humor pharmacokinetics
and acetazolamide niosomes revealed that the Cmax of the drug from
the niosome formulation was double than that of the drug suspension,
showing a significant broadening of the peak from 80 to 120 min. The
concentration remained over 13 μg throughout this period [141].
It was demonstrated that deformable elastic liposomes can enter the
corneal structure that is similar to the SC [142] and that elastic niosomes
show advantages over liposomes and over conventional NSVs as well. In
that, the former can reach even the posterior eye segment due to its
elasticity and nanosize. This elasticity is attributed to the use of edge
activators (EAs). EAs are the surfactant molecules that provide flexibility
to the lipid bilayer membranes of these vesicles. These surfactant

Table 4
Niosomes in ocular delivery.

Compositions Drug Experimental model Results Reference

Chitosan or carbopol coated niosomes
(Span 60:Chol 1:1)
Tween 60 or Tween 80 or Brij 35/Chol/DCP
Tween 20/Chol
Niosomes: Span 60/DCP or Sodium Cholate
Discosomes: Span 60/Solulan C24
Span 40 or Span 60, or Span 80/edge
activators (EAs): sodium taurocholate or
sodium cholate or sodium deoxycholate,
or Tween 80
Timolol maleate In vitro release: dialysis bag; in vivo
pharmacodynamic studies (intra ocular
pressure, IOP) on adult male normotensive
rabbits
Gentamicin In vitro release: dialysis bag; in vivo evaluation
of potential ocular irritancy and/or damaging
effects on albino rabbits.
Cyclopentolate In vitro permeation — rabbit corneas; in vivo
evaluation of mydriatic activity in rabbits
Naltrexone hydrochloride Evaluations on surface tension, contact angle,
spreading coefficient, viscosity and effect of
niosomal encapsulation on NTX oxidation.
Fluconazole In vitro genotoxicity and cytotoxicity; ex vivo
corneal permeability studies (porcine cornea);
in vivo ocular irritation studies in rabbits.
Significant decrease in IOP [134,135]
Niosomes exhibit an alkyl chain length- [136]
dependent release, the higher the chain
length, the lower the release rate. DCP stabi-
lizes the niosomal membrane and delays the
drug release
In vitro enhanced transcorneal permeation; [135]
in vivo improved drug ocular bioavailability
Enhanced corneal uptake of the hydrophilic [137,138]
drug (NTX). Enhanced NTX protection from
photo-oxidation compared with conventional
NTX aqueous solutions.
Elastic niosomes safety in terms of [144]
genotoxicity, cytotoxicity, acute dermal/eye
irritation/corrosion, and chronic eye irritation/
corrosion test. Enhanced drug corneal
permeability
 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206 197

molecules destabilize the lipid bilayers and, in turn, increase the
deformability of the vesicles [143]. The surfactant present in these
vesicles induces pores in lipid structures, such as membranes, and also
leads to a solubilization (lysis) at the higher concentration range.
In the study of Kaur et al. [144] Sp-based elastic (spanlastic vesicles)
novel vesicular systems were developed and proposed as carrier
systems for fluconazole, which is a hydrophilic drug with a short
ocular elimination half-life and low log P value (log P = 0.25). A better
permeation was achieved by these systems and can be explained ac-
cording to two transport mechanisms. First, the elastic vesicles interact
with the epithelial cell membrane and act as penetration enhancers,
and subsequently modify the intercellular lipid lamellae (mechanism
1). Second, the elastic vesicles can act as drug-carrier systems, whereby
intact vesicles carrying the drug pass through the intercellular spaces
and reach the biological membrane (mechanism 2).
The developed spanlastic systems were found to be safe in terms of
genotoxicity, cytotoxicity, acute dermal/eye irritation/corrosion, and
chronic eye irritation/corrosion tests.
5.3. Oral delivery
Oral route is the preferred route of administration for most drugs;
however, the majority of the newly discovered and existing drugs
administered by oral route frequently must face bioavailability prob-
lems due to several reasons such as poor solubility and low dissolution
rate, unpredictable absorption, inter and intra subject variability
and lack of dose proportionality. Several strategies have been adopted
for the improvement of dissolution behavior of insoluble drugs;
e.g., complexation, drug derivatization, solid state manipulation, inclu-
sion of surfactants, increase of the surface area by micronization or
nanonization, spray drying and microencapsulation (Table 5).
NSVs are also capable to enhance drug stability in the gastrointesti-
nal environment; in this sense it was reported that the in vitro gastroin-
testinal stability of paclitaxel was well preserved with Sp 40 NSVs [145];
moreover vesicles from Sp 20, Sp 40, and Sp 60 significantly improved
the oral bioavailability of griseofulvin in albino rats after a single oral
dose [146].
Recently, NSVs from Sp 40, Sp 60, and CHOL were proposed as
potential oral delivery systems for the effective delivery of ganciclovir.
In vivo study on rats revealed a five-time increase in bioavailability of
the drug after oral administration with respect to tablets [147].
Metformin is an oral hypoglycemic which is widely used as a first
line treatment of type II diabetes, but at a very high dose 2–3 times a
day; moreover it suffers from a number of side effects such as lactic
acidosis, gastric discomfort, chest pain, allergic reactions. To decrease
metformin side effects, and dose frequency, niosomal formulations
obtained with Sp 60 and Sp 40 and CHOL in equimolar ratio were pro-
posed. The obtained vesicles showed a high entrapment efficiency and
a sustained release of the drug over extended period of time; thus signif-
icantly improving the performances of the drug [148].
NSVs were also used for oxcarbazepine loading with the aim of
improving its anticonvulsant activity. The in vivo study indicated how
the dispersion of drug-loaded vesicles, prepared by the thin film hydra-
tion method, showed an improved anticonvulsant activity in albino
wistar rats together with a significant increase in the mean residence
time, thus reflecting sustained release properties [149].
Lornoxicam loaded niosomes were prepared by thin film hydration
method with different Sp 60:CHOL ratios and the obtained formulations
were evaluated by and for their entrapment efficiency. In vitro release,
kinetic data analysis and stability studies suggested that multilamellar
niosomal formulation could provide consistent and prolonged release
of lornoxicam from different niosomal formulations. Such sustained
activity of the entrapped drug may lead to a reduction of the side effects
associated with frequent administration and correspondingly potenti-
ate its overall the therapeutic effects [150]. An interesting study focused
on the evaluation of the stability of NSVs composed by Tw20 and CHOL,
in the artificial gastrointestinal fluid was performed by Di Marzio et al.
Tw20 is a non-ionic surfactant, the stability and relative non-toxicity
of which, allows it to be used as excipient for oral administration.
From in vitro simulations, it was evidenced that Tw20 vesicles show

Table 5
Niosome in oral delivery.

Compositions Drug Experimental model Results Reference

Tweens or Spans or Brijs/Chol and DCP Paclitaxel In vitro PCT release; stability studies in GI en- Fast drug release from Tween niosomes (more flexible [145]
(PCT) zymes. structure). Best PCT protection against GI enzymes with
Span 40 niosomes
Span 40 or Span 60/Chol Ganciclovir In vitro drug release, release kinetic study, and In vivo study on rats reveals five-time increment in bio- [147]
in vivo performance availability of Ganciclovir after oral administration of op-
timized formulation (Span60:Chol 3:2) as compared with
tablet.
Spans or Tweens or Brijs/Chol/DCP Metformin In vitro release studies with USP dissolution Span 60 having longer chain length provide more stable [148]
apparatus. vesicles, higher entrapment and delayed release.
Spans/Chol Oxcarbazepine Pharmacokinetic studies in Albino Wistar rats Increased elimination half life and AUC when compared [149]
with free drug
Span 60/Chol Lornoxicam In vitro drug release studies. Consistent and prolonged release of lornoxicam. [150]
Span 60/Chol/coating agent (maltodextrin Glipizide In-vitro release — dialysis. Consistent and prolonged release with maltodextrin- [152]
or sorbitol or mannitol): proniosomes based proniosome
Span 60/Chol/DCP: proniosomes Celecoxib Dissolution studies; pharmacokinetic Slow drug release; Significantly higher values and [153]
evaluations capsules in comparison to significantly lower values for absorption rate constant
Celebrex®, Pfizer, on six healthy male. (Ka) of the proniosomal system compared with
Celebrex® capsule.
Span 60/Chol/Maltodextrin:proniosomes Valsartan In vitro dissolution studies in SGF and SIF; ex- Drug improved dissolution rate; enhanced drug [155]
vivo permeation studies on male Wistar rats' absorption across gastrointestinal membrane.
ileum.
Monopalmitoylglycerol/Chol/DCP/bile salts: Influenza In vivo biodistribution and vaccine efficacy Increase uptake within the Peyer's patches and reduction [157]
bilosomes vaccine studies in viral cell load in an influenza challenge study.
Tw20/Chol Tw20/Chol/Chems = Stability evaluation in SIF and SGF. Assess- Good stability in SIF and SGF; good mucoadhesive prop- [151]
ment of mucoadhesive properties. erties in SIF rather than in SGF
Span 60/Chol/stearylamine niosomes, Plasmid DNA In vitro ligand-specific activity by concanava- Measurable humoral and cellular immune response upon [158]
emulsificated with aqueous solution of encoding for lin A agglutination assay. Stability studies in oral administration.
plasmid DNA, coated with o-palmitoyl HbsAg. SGF and SIF. Biodistribution studies on BALB/c
mannan. mice
198 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206

good stability in simulated gastric and intestinal fluid, also in the pres-
ence of digestive enzymes and bile salts, as evidenced by the release
of a fluorescent probe which is most likely related to small membrane
damages rather than to a disintegration of the vesicles. Furthermore,
these vesicles were able to adhere to mucin in the intestinal medium
more than in gastric one [151].
Approaches to stabilize niosomal drug delivery systems without
affecting their properties led to the development of the promising
drug carriers called “Proniosomes”. Proniosomes are a dry formulation
obtained from a suitable carrier coated with non-ionic surfactants
which can be converted into NSVs immediately before use by hydration.
These proniosome-derived niosomes are as good as or even better than
conventional niosomes. Glipizide loaded sorbitol, maltodextrin and
mannitol based proniosomes were prepared by a slurry method with
different surfactant:CHOL ratios [152].
Another proniosome formulation (Sp 60:CHOL:DCP, 1:1:0.1) was
studied to improve the extent of celecoxib absorption, compared to
that of a conventional marketed celecoxib capsule. Experiments, carried
out on human volunteers, showed that the mean relative oral bioavail-
ability of the proniosomal formulation increased with respect to that of
the conventional capsule [153]. Similar results were reported by Xiao
et al. who studied the effect of oral proliposome on silymarin bioavail-
ability [154].
The oral administration of valsartan proniosomes was also studied to
overcome its low absolute bioavailability (25%) due to the poor aqueous
solubility, as well as the drug side effects in the gastrointestinal tract and
the fact that two third of the dose is excreted by renal and biliary trans-
port. Jukanti et al. reported the formulation and characterization of
valsartan loaded maltodextrin based proniosomal powders. Ex-vivo
studies, carried out on rat intestine to assess the permeation of valsartan
from proniosomes showed a significant permeation enhancement
across rat intestine thus suggesting the potential of proniosome carriers
for improved oral delivery of valsartan [155].
NSVs have also been tested as stabilizers for therapeutic proteins
and peptides in the gastrointestinal tract and for a possible improve-
ment of their permeation through the intestinal mucosa. In an in vitro
study by Yoshida et al. oral delivery of a vasopressin derivative
entrapped in niosomes showed that vesicle inclusion significantly
increased the stability of the peptide, in an intestinal loop model [156].
Oral vaccines offer significant benefits due to the ease of administra-
tion, better patient compliance and non-invasive, needle-free adminis-
tration. However, this route is marred by the harsh environment
which is detrimental to many vaccine formats. To overcome such prob-
lems, bilosomes have been considered; these are bilayer vesicles
constructed from non-ionic surfactants combined with the inclusion
of bile salts which can stabilize the vesicles in the gastrointestinal tract
by preventing membrane destabilization. Bilosomes were actually
constructed from monopalmitoylglycerol, CHOL, DCP and sodium
deoxycholate at different blend ratios. The optimized bilosome formula-
tion was identified and the potential of this formulation as an oral
vaccine delivery system was assessed by biodistribution and vaccine
efficacy studies. Obtained results showed that the larger bilosomes
vesicles increased uptake within the Peyer's patches and promoted a
reduction in viral cell load in an influenza challenge study [157].
Mannosylated niosomes were used as oral DNA vaccine carriers for
the induction of humoral, cellular and mucosal immunity. NSVs, com-
posed of Sp 60 with the addition of CHOL and SA as constitutive lipids,
were prepared by reverse phase evaporation and were coated with a
modified polysaccharide o-palmitoyl mannan (OPM) in order to protect
them from bile salt-caused dissolution and enzymatic degradation in
the gastrointestinal tract and to enhance their affinity towards the anti-
gen presenting cells of Peyer's patches. OPM coated niosomes produced
humoral (both systemic and mucosal) and cellular immune response
upon oral administration. This study signifies the potential of OPM
coated niosomes as DNA vaccine carriers and adjuvants for effective
oral immunization [158].
5.4. Pulmonary delivery
In inflammatory diseases, infections or cancer of the respiratory
tract, the therapeutic value of pulmonary administration may exceed
that of an oral or parenteral administration (Table 6) and in diseases
characterized by bronchial mucus hypersecretion, lipophilic substances,
such as corticosteroids, can be remarkably impeded in reaching their
receptors, which are localized within the cytoplasm of bronchial epithe-
lial cells. Surfactant vesicles were proposed to circumvent this prob-
lem [73]. Niosomal delivery of beclomethasone dipropionate (BDP) in
asthma and chronic obstructive pulmonary disease was evaluated by
different research groups.
Terzano et al. [73] and later Marianecci et al. [159] carried out formu-
lation, physical–chemical and in vitro cell characterization studies on
BDP loaded Tw20 vesicles in order to evaluate the possible advantages
of this new type of NSV system that can lead to an improved mucus per-
meation. Actually, such delivery system should enhance permeation
through mucosal barriers because of the presence of vesicles formed
with a remarkably hydrophilic non-ionic surfactant usually considered
as unsuitable for the formation of vesicular structures because of its
high HLB value (HLB = 16.7). From the obtained results, it was
highlighted that the vesicular dispersions show rheological characteris-
tics and nebulization efficiency (i.e. aerodynamic diameter and drug
mass output) comparable with that of a commercial product. Further-
more, the novel preparation, ensuring a better penetration through
the mucus layer, can provide an efficient system for the treatment of
obstructive pulmonary diseases. The aim of the second part of the
same study, carried out by Marianecci et al., was the evaluation of the
interaction between the innovative NSVs and human lung fibroblast

Table 6
Niosomes in lung delivery.

Compositions Drug Experimental model Results Reference

Tween 20/Chol Beclomethasone In vitro permeation experiments (mucin solution 0.1% Increased permeation rate through the model [73,159]
dipropionate w/v). Aerosolized droplet size studies. Cellular tolerability mucosal barrier. Good in vitro tolerability on HLFs.
and cytotoxic on primary human lung fibroblasts (HLFs). Improved drug anti- inflammatory activity in HLFs.
Evaluation of antiinflammatory effect on HLFs.
Span 60/Chol/sucrose: Beclomethasone Nebulization by Pari LC Sprint (Air-jet), Aeroneb Pro High drug output and fine particle fraction (FPF) by all [56]
proniosomes dipropionate (Actively vibrating-mesh) nebulizer and Omron MicroAir devices
NEU22: aerosol mass output gravimetrical determination
and aerosol droplet size analysis.
Tween80/Chol niosomes Amphotericin B In vitro antifungal and antileishmanial activity of AMB- Significant reduction in fungal lung burdens in a rat [169]
encapsulating niosomes; in vitro AMB delivery studies. In vivo real time model of invasive pulmonary aspergillosis and in a
hydroxypropyl-γ- delivery studies; in vivo antileishmanial and anti asper- significant suppression of Leishmania donovani liver
cyclodextrin/AMB gillosis activity of AMB-niosomes parasite burden
Span 60 and Tween 60/Chol Ciprofloxacin HCl MIC determination against some pulmonary pathogens. In vitro safety and efficacy of Ciprofloxacin loaded [170]
MTT assay in human lung carcinoma cell line. niosomes
 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206 199

(HLF) cells, the carrier tolerability, the vesicle localization within the
cells and the amount of BDP actually internalized by the cells. The
study confirmed that BDP-loaded NSVs were able to improve the drug
anti-inflammatory activity in HLFs due to the carrier ability to increase
the intracellular delivery of BDP and hence to determine a great amount
of drug interacting with its cytoplasm receptor. The overall conclusion
of this investigation was that these innovative NSVs could be considered
as effective and safe carriers for the BDP pulmonary delivery, thus suit-
able for the treatment of pulmonary inflammatory diseases.
Proniosomes were investigated also for pulmonary delivery of BDP.
Hence, proniosomes represent an approach to formulate stable pow-
dered formulations that can form NSVs by hydration prior to adminis-
tration [160]. The formation of niosomes using air-jet nebulizers has
been demonstrated [161,162], however, the actual feasibility, for drug
delivery, of niosomes obtained via vibrating-mesh nebulizers has not
yet been fully investigated. On the other side, proniosomes loaded
with the anti-asthma steroid BDP were developed to generate niosomes
that are suitable for aerosolization by air-jet or vibrating-mesh
nebulization.
The Sp 60 niosomes, obtained from the corresponding proniosomes,
were characterized and their aerosol delivery was investigated using
Omron MicroAir (passively vibrating-mesh) nebulizer, aeroneb pro
(actively vibrating-mesh) nebulizer and Pari LC print (air-jet) nebulizer.
The generated aerosols by all tested devices showed high drug output
and fine particle fraction (FPF). The drug output and FPF of the aeroneb
pro vibrating-mesh nebulizer were similar to those of the standard air-
jet nebulizer Pari LC sprint. In vivo investigations still need to explore
the suitability of niosomes as alternatives to liposomes for pulmonary
delivery [163].
Recently Sp and Tw niosomes were studied for the pulmonary deliv-
ery of anti-tuberculosis drugs [164–168] and of amphotericin B (AMB)
[169]. AMB is used to treat both fungal and leishmanial infections,
which are of major significance to human health. Clinical use of free
AMB is limited by its nephrotoxicity, whereas liposomal AMB is costly
and requires parenteral administration. On the other hand, treatment
with AMB-loaded niosomes showed a significant reduction in fungal
lung burdens in a rat model of invasive pulmonary aspergillosis and in
a significant suppression of Leishmania donovani liver parasite burdens.
The results of this study indicate that AMB aerosolised niosomal formu-
lations enhanced pulmonary and hepatic delivery while minimizing
systemic exposure and toxicity [169].
In order to develop a niosome-encapsulated ciprofloxacin HCl
(CPFX) formulation for pulmonary delivery, the feasibility of CPFX en-
capsulation in niosomes, their stability and nebulization process
effectiveness was evaluated by Moazeni et al. [170]. Various combina-
tions of non-ionic surfactants with cholesterol were used to prepare
the formulations. The in vitro deposition data of the niosomal formula-
tions were examined using an andersen cascade impactor. Formulations
composed of Sp 60 and Tw60 in combination with 40 mol% of CHOL
exhibited high encapsulation efficacy and stability and also had fine
particle fraction and high nebulization efficiency. Minimal inhibitory
concentration of the niosomal CPFX against some pulmonary pathogens
was lower than that of free CPFX. Using the cell viability assay in human
lung carcinoma cell line (A549), niosome-entrapped CPFX showed
significantly lower cytotoxicity in comparison to the free drug. These
results indicate that NSVs can be used as carriers for pulmonary delivery
of CPFX via nebulization [170].
5.5. Parenteral delivery
The parenteral route of administration is one of the most effective
routes for the delivery of the active pharmaceutical substances. To
maintain a therapeutic effective concentration of a drug, it requires
frequent injections which ultimately lead to low patient compliance.
In parenteral drug delivery, major progresses were obtained in the
field of vesicular formulation technologies which were in most cases
capable to provide a targeted and sustained drug release in predictable
manner to overcome the problems associated with conventional paren-
teral delivery systems, especially for drugs with narrow therapeutic
index and poor bioavailability (Table 7).
In 2007, Mukherjee et al. presented a study aimed to develop and
compare acyclovir containing nano-vesicular liposomes and niosomes
based on CHOL, soya L-α-lecithin and the non-ionic surfactant Sp 20.
Furthermore, in vitro investigations were performed in order to find
out whether acyclovir-loaded nanovesicles could sustain the release of
the drug by increasing residence time and thus, acyclovir could reduce
its dose-related systemic toxicity. Obtained results indicated that NSVs
showed better stability compared to liposomes [171].
Nystatin (NYS) is a polyene antifungal drug that has been used in the
treatment of cutaneous, vaginal and oral fungal infections since the
1950s. NYS encapsulation in niosomes was proposed to obtain a safe
and effective formulation which can be parenterally administered for

Table 7
Niosomes in parenteral delivery.

Compositions Drug Experimental model Results References

Span 60 or Span 40/Chol/SA or DCP.
Nanohybrid systems of non-ionic
surfactant (Solutol® HS 15, pluronic
F68 and cremophor EL) inserting
liposomes loading paclitaxel
Freeze dried Span 40/Chol niosomes
Tween 61⁄Chol⁄dodecyl dimethyl
ammonium
pH sensitive niosomes
Spans/, Chol/cetyl trimethyl
ammonium bromide
Nystatin (NYS) In vitro release studies, in vivo evaluation of Reduced NYS toxicity and enhanced antifun- [172]
nephrotoxicity and hepatotoxicity, analysis of gal activity, after parenteral administration.
the fungal burden in experimental animals
infected with Candida albicans
Paclitaxel (PTX) In vitro internalization studies (A549 and Empty nanohybrids interfere P-gp expression [178]
A549/T cells). Effect of blank nanohybrid on the membrane of drug resistant cells and
structures on A 549 cells and on P-gp expres- decrease ATP level. Higher cytotoxicity than
sion, on ATP levels. In vitro cytotoxicity ex- liposomes
periment. Apoptosis assay and cell cycle
analysis.
Cisplatin Anticancer efficacy in rats bearing VX2 Inhibition of tumor growth and of tumor [179]
sarcoma metastasis to inguinal lymph nodes and liver,
lower mortality when compared to free drug
Human tyrosinase plasmid Cytotoxicity on B16F10 cells Evaluation of No cytotoxic effect; high stability of the loaded [180]
(PE) and Tat ⁄human melanin concentration and tyrosinase enzyme plasmid when kept at various temperatures;
tyrosinase plasmid (TPE) activity. enhancement of human tyrosinase gene
expression and melanin production
5 Fluorouracil In vitro release studies. In vivo distribution to Remarkable high drug concentration at tumor [181]
tumor sites site
Autoclaved Leishmania major Induction of the immune response against Moderate effect in the prevention of [183,184]
(ALM) cutaneous leishmaniasis in BALB/c mice cutaneous leishmaniasis in BALB/c mice
200 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206
neutropenic patients. NYS niosomes were prepared using Sp 60 or Sp
40 and CHOL. SA and DCP were added as the positive and negative
charge-inducing agents, respectively. Neutral and positively charged
NSVs gave the highest encapsulation efficiencies. The release of neutral
and negatively charged NYS niosomes was estimated, and it showed
a slow sustained release profile. A 25-kGy γ-irradiation dose was
sufficient to sterilize the investigated vesicles. NYS niosomes exerted
less nephrotoxicity and hepatotoxicity in vivo, showed higher level
of drug in vital organs and revealed pronounced efficacy in elimination
of the fungal burden in experimental animals infected with Candida
albicans compared with those treated with free NYS. Niosomal
encapsulation thus provided means for parenteral administration of
NYS, reducing its toxicity and making it a more active antifungal agent
[172].
The parenteral route was also proposed for antineoplastic drug
administration. In this case the advantage of drug loading in a surfactant
carrier could be the selective delivery of the cytotoxic agent to
the tumor site, thus decreasing toxic effects. Drug loaded vesicles
could improve the therapeutic outcome and low drug resistance of
traditional chemotherapeutics by altering drug pharmacokinetics
and biodistribution. For example intravenous administration of
hydroxycamptothecin entrapped in niosomes to sarcoma 180 ascites
tumor bearing mice led to a total regression of the tumor and also higher
plasma level and slower elimination [173].
Niosomes have great potential in drug nanoformulations for
melanoma therapy [174–176]. New nanohybrid systems of non-ionic
surfactant and lipid loading paclitaxel (PTX) were prepared to over-
come multidrug resistance in PTX-resistant human lung cancer cell
line. Three non-ionic surfactants, solutol® HS 15, pluronic F68 and
cremophor EL, were inserted into liposomes. The apoptotic assay and
the cell cycle analysis showed that the nanohybrid systems were able
to induce more apoptotic cells in drug resistant cells when compared
with liposomes [177]. Cisplatin-loaded niosomes (CP-NMs) were pre-
pared using Sp 40 and CHOL as excipients, and then lyophilized and
characterized. Their anticancer efficacy was tested in rabbits bearing
VX2 sarcoma. The rabbits, locally injected with the CP-NMs, gave signif-
icantly superior results of inhibition of tumor growth, much lower mor-
tality and improved results of body weight variation and inhibition of
tumor metastasis to inguinal lymph nodes and liver compared to
those treated in the same way with the drug solution. The inspiring
anticancer results indicated that the CP-NMs might be developed as
an effective anticancer preparation with low toxicity [178].
Potent melanin production enhancement of human tyrosinase plas-
mid (pAH7⁄Tyr) in mouse melanoma cells (B16F10) by Tat peptide and
an entrapment in elastic cationic niosomes were described by Manosroi
et al. for vitiligo treatment. The NSVs, composed of Tw61, CHOL and
dodecyl dimethyl ammonium bromide, were prepared by freeze-dried
emptying liposomes method. The B16F10 cells transfected with
pAH7⁄Tyr –Tat peptide-elastic cationic niosomes (TPE) complexes in
a specific ratio showed the highest enhancement of the tyrosinase
enzyme activity and melanin production. For all TPE complexes a slight
or no cytotoxic effect on the cell line was obtained. The pAH7⁄Tyr-
loaded elastic cationic niosomes and TPE complexes exhibited high
stability of the loaded plasmid when kept at various temperatures for
8 weeks. This study indicated the enhancement of human tyrosinase
gene expression and melanin production with relatively low cytotoxic-
ity by Tat and entrapment in elastic cationic niosomes as well as the
enhancement of stability of the plasmid loaded in the TPE complexes
[179].
Wang et al. [180] investigated preparation, characterization and
tumor-targeted effect of pH-sensitive niosomes, composed of a non-
ionic surfactant mixed with CHEMS.
Both normal NSVs and pH-sensitive niosomes showed spherical
morphology under TEM with an appropriate average particle sizes.
The thermotropic behavior, structure changes and interaction of
5-fluorouracil were characterized by differential scanning calorimetry,
and the disappearance of the melting peak of the drug clearly indicated
that the drug was encapsulated within NSVs. Tumor-targeted effect
was proven by the remarkable high concentration of the pH-sensitive
niosomes in the tumor site of the mice transplanted with tumor cells.
The use of NSVs was also evaluated for injectable vaccines formula-
tions. It is known that a promising strategy for overcoming the poor
immunogenicity of antigen is the development of new vaccine adju-
vants, or carriers that enhance the effectiveness of vaccines. This occurs
because the adjuvants act as immunostimulatory agents. Given the ever
increasing interest in vesicle-based vaccines, it is important to under-
stand precisely how vesicular carriers interact with the immune system
and stimulate immunity [181]. The first report showing the adjuvant
effect of surfactant vesicles was provided by Brewer and Alexander.
These authors reported that niosomes are potent adjuvant in terms of
immunological selectivity, low toxicity and stability [182]. Sp niosomes
were also proposed as DNA vaccine carriers, after conjugation with OPM
[183].
More recently, NSVs were investigated for potential intranasal
immunization; the vesicles contained either the secretory recombinant
form of glycoprotein B (gBs) of herpes simplex virus type 1 or a related
polylysine-rich peptide (DTK) for the induction of protective immunity
against genital herpes infection in mice [184]. Furthermore, different
positively charged niosomal formulations containing Sps, CHOL and
cetyl trimethyl ammonium bromide were used for the induction of
the immune response against cutaneous leishmaniasis in BALB/c mice
[185].
Different charged niosomal formulations containing Sps, CHOL and
cetyl trimethyl ammonium bromide were prepared for the entrapment
of autoclaved Leishmania major (ALM). The selected formulation was
used for the induction of the immune response against cutaneous
leishmaniasis in BALB/c mice. In vivo studies showed that the niosomes
containing ALM have a moderate effect in the prevention of cutaneous
leishmaniasis in the tested animals [185].
Another interesting goal in niosomal formulations for parenteral
administration is the possibility to obtain successful targeting strategies,
avoiding effects on healthy tissues and organs and reaching only
interested area in the body. An emerging paradigm for the design of
effective drug targeting is the modification of vesicular carrier
parameters to encourage the preferential in vivo biodistribution. These
parameters include size, shape, charge, chemistry, and ligand modifica-
tion. In particular, after intravenous administration, the surfactant
vesicles, according to their physic-chemical properties, are surrounded
by blood components including many types of circulating serum
factors known as opsonins which mark them for clearance by RES
macrophages. For these reasons niosomes can be used for drug targeting
in the treatment of diseases in which the infecting organism resides
in the RES, exerting a sort of passive targeting. The study of antimony
distribution in mice, performed by Hunter et al. showed high liver
level after intravenous administration of the drug carried by NSVs, with-
out increasing the damage on heart and kidney [186].
More recently, Sp 60/Tw61 niosomes conjugated with a purified
monoclonal antibody to CD44 were proposed for immuno-targeting
[187] and “hydrid” Tw20 NSVs were reported to show intrinsic selectiv-
ity towards macrophages [91].
Attaching target molecules to niosome surface is one the approach to
overcome the limitation of passive targeting exploiting the EPR effect
(e.g. different degree of vascularization and porosity of tumor vessels).
Equipping the surfactant vesicles with a ligand it becomes able to be
guided to the intended target.
Ligands and techniques used to modify niosome surface are similar
to those used in liposomal formulations [188]: it was reported
that glucose and transferrin conjugation of niosomes enhance vesicle
distribution to the brain or to solid tumors [189,190] and that
niosome functionalization by N-palmitoylglucosamine (NPG), allowed
a reduction of doxorubicin accumulation in the heart, longer blood
circulation times and increased brain concentrations with respect to
 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206
the commercial formulation of the drug, after i.v. administration to rats
[191].
Although ligand-mediated targeting niosomes have not yet made a
considerable progress, it will be feasible and stimulating to expand
researches in this field, taking advantages of knowledge collected in
studying targeting liposomes to overtake their limitations.
5.6. Gene delivery
Gene therapies, using genetic material (DNA, oligonucleotides, small
interfering RNAs, ribozymes, DNAzymes), rather than traditional drugs
to patients, are being investigated for treatment of a number of different
inherited or acquired disorders. Unlike traditional pharmaceuticals,
gene therapy has the theoretical potential to treat almost any disease.
Although gene therapy is believed by several investigators as the
therapy of the 21st century, numerous barriers still exist that limit the
efficient delivery or expression of genetic material into our body cells
[192,193]. Different delivery systems are being developed to enhance
the stability, to increase the efficiency of cellular uptake and intracellu-
lar trafficking, or to alter the biodistribution of genetic materials. An
approach involves the use of cationic niosomes as novel non-viral
carriers for plasmid and oligonucleotide delivery [194,195]. Among
various nonionic surfactants, Sps and Tws have been commonly used
as additives for the formulation of lipid-based gene carriers. Compared
with Sps, it is worth noting that TWs not only are a critical class of
emulsifier and stabilizer in those formulations, but they also play an
active role in biological integration with DNA, thereby promoting gene
expression.
Recently, non-ionic surfactant vesicles, from cationic lipid and Sp 80
were synthesized and evaluated as promising vehicles for small inter-
fering RNA delivery to efficiently and specifically silence expression of
both green fluorescent protein (66% knockdown) and aromatase (77%
knockdown) genes in breast cancer cell lines [196].
Khositsuntiwong et al. investigated the enhancement of tyrosinase
gene expression and melanin production in tyrosinase gene knocked
out human melanoma cells and in B16F10 cells by loading the plasmid
in elastic cationic niosomes [197].
5.7. Diagnostics/theranostics
Nanocarrier-mediated delivery has emerged also as a successful
strategy to enhance delivery of imaging agents for tumors, thereby
increasing the potential for diagnosis at an earlier stage. It was showed
that combination of PEG and glucose conjugates on the surface of
niosomes significantly improved tumor targeting of an encapsulated
paramagnetic agent assessed with nuclear magnetic resonance (NMR)
imaging in a human carcinoma xenograft mode [198].
Niosomes composed of Tw61, CHOL and DCP were proposed as drug
delivery and imaging in combination with ultrasound. It was evidenced
that ultrasound at specific frequencies can reversibly permeabilize the
lipophilic membrane of niosomes to allow the controlled release of a
compound without destroying the vesicular structure [199].
Table 8
Recently proposed pharmaceutical application for natural product-loaded niosomes.
Natural product Niosome formulation
Ammonium glycyrrhizinate Sp 20:Tw85:CHOL Tw20:DCP: CHOL
Curcumin Sp 60 or Sp 80 or Tw 20:CHOL
Ellagic acid Sp 60 or Tw60:CHOL
Ginkgo biloba extract Tw80 or Sp 80:CHOL
Gymnema sylvestre extract Sp 40:CHOL
Papain Tw61:CHOL:sodium cholate
Rice bran extract Tw61:CHOL:cetyl trimethylammonium bromide
Silymarin Sp 40 or Sp 60:CHOL
Rutin Sp 60 or Sp 60/Gelucire®
201
Magneto niosomes, i.e. vesicles based on non-ionic surfactants with
their aqueous core loaded with both a magnetic material and a model
drug were proposed by Tavano et al. [200] as theranostic nanocarriers.
Magneto NSVs containing doxorubicin were prepared using Tw60 and
pluronic L64, a copolymer of ethylene oxide and propylene oxide blocks
arranged in a triblock structure, also known, more simply, as pluronic.
Both surfactants present in their structures ethylene oxide chains, that
are claimed to prevent particle opsonisation, render them ‘unrecogniz-
able’ by RES, and significantly reduce hepatic uptake. Moreover, these
moieties were used to sterically stabilize the vesicular carriers and,
hence, to prolong their half-life. The loading of magnetic ferrofluid did
not change the morphology and shape of niosomes (both empty and
drug loaded) and doxorubicin was efficiently encapsulated in both for-
mulations, due to favorable interactions between magnetic materials
and drug. Doxorubicin loading and release behavior of Tw60- and
pluronic-based magneto-niosomes could probably promote them as
effective functional materials for magnetically-controlled cancer therapy.
NSVs prepared using Tw20 or Sp 20 and CHOL loaded with lipophilic
and hydrophilic magnetic nanoparticles (MNPs) were prepared and
deeply characterized by Marianecci et al. as promising carriers for
the development of theranostic strategies. In this study the analyzed
formulations confirm the importance of surfactant chemical-physical
characteristics in the entrapment of MNPs of different polarity, thus
highlighting the high versatility of niosomal bilayers and structures
property that make such structures so appealing among drug delivery
nanocarriers [201].
6. Natural products delivery
In spite of the high costs related to new drug discovery, there is
today a great deal of interest in investigating new plant constituents
and/or new delivery systems for natural products.
As above pointed out, novel delivery systems, such as NSVs, that
modify the pharmacokinetics of phytochemicals can be used to enhance
or modulate the delivery to target sites; consequently the use of NSVs
was proposed also for several natural products and some of them are
reported (Table 8).
Ginkgo biloba extract (GbE) was investigated for human brain and
breast cancer prevention; GbE entrapped in niosomes (Tw80, Sp 80,
and CHOL) showed an increase of in vivo distribution, after oral admin-
istration, when compared to GbE tablet [202].
Niosome formulations loading Gymnema sylvestre extract showed
significant blood glucose level reduction and increased antihyper-
glycemic activity compared with the parent extract [203].
Based on the poor bioavailability of silymarin and on the advantages
of niosomes, a silymarin niosomal preparation was proposed, showing a
significant decrease in both transaminase levels as well as in serum
alkaline phosphatase level in comparison with administered silymarin
suspension in albino rats [204].
In a recent study the transdermal penetration of papain from gel (8%
carbopol®980) formulations containing elastic niosomes (Tw61:CHOL:
sodium cholate at 1:1:0.1) and from nanospheres (Poly(lactic-co-
Route of administration Reference
Dermal/Transdermal [119,120]
Transdermal [208]
Transdermal [118]
Oral [202]
Oral [203]
Transdermal [205]
Transfollicular [206,207]
Subcutaneous [204]
Rectal [215]
202 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206
glycolic acid), PLGA) loaded with papain was compared. The higher po-
tential of elastic niosomes for the enhancement of rat skin transdermal
absorption of papain and the improvement of scar reduction, when
compared to nanospheres, was demonstrated [205].
The physico-chemical properties of cationic niosomes (Tw61/
CHOLl/cetyl trimethylammonium bromide) loaded with rice bran ex-
tract were investigated [206]. These carriers appeared to be a suitable
system for topical anti-androgenic alopecia application because of the
convenient use and high transfollicular penetration in the skin, accom-
panied by a low systemic effect [207].
Different proniosomal gel bases were prepared using Sp 60, Sp 80,
Tw20 and CHOL for the encapsulation of curcumin. The investigated
formula was not irritant, not toxic, but had lower anti-inflammatory
and anti-arthritic effects than the marketed indomethacin products
[208].
Ellagic acid (EA) is a potent antioxidant phytochemical substance
which has limited applications due to its poor biopharmaceutical prop-
erties (i.e., low solubility and low permeability). EA loaded niosomes
were prepared from Sp 60 and Tw60 mixtures for transdermal delivery.
Skin distribution studies evidenced that EA-loaded niosomes were
more efficient in the delivery of EA through human epidermis and der-
mis than EA solutions [118].
NSVs made up of Tw85, Sp 20 and CHOL at various concentrations
were prepared to investigate the possible application of niosomes for
the delivery of ammonium glycyrrhizinate (AG), useful for the treat-
ment of various inflammatory-based diseases: the AG-loaded NSVs
showed no toxicity, good skin tolerability and were able to improve
the drug anti-inflammatory activity in mice. Furthermore, an improve-
ment of the anti-inflammatory activity of the niosome delivered drug
was observed on chemically induced skin erythema in humans
[119,120].
Niosomal formulations were proposed, in preliminary studies, for
the delivery of colchicine [209], punicalagin [210] and neem seed oil
[211].
Recently, resveratrol was loaded in Sp niosomes to modulate its
release in functional foods [212,213] and a black tea extract/multilamellar
niosome formulation (Brij 52:CHOL, 1:1) was investigated as sunscreen
agent [214].
Niosome
(A)
Drug
(B)
Stratum
corneum
Viable
Epidermis
Dermis
Fig. 8. Possible mechanisms of action of surfactant vesicles for dermal and transdermal applications: (A) drug molecules are released by niosomes, (B) niosome constituents act as
penetration enhancer, (C) niosome adsorption and/or fusion with Stratum Corneum; (D) intact niosome penetration through the intact skin; (E) niosome penetration through hair follicles
and/or pilosebaceous units.
The increasing number of studies on natural product (Table 8) can
lead to consider niosomes as promising carriers, by different routes of
administration [215], in pharmaceutics, in cosmetics as well as for
food applications.
7. Niosomes vs liposomes
Taking into account the enormous prospective of vesicular systems
in drug delivery and targeting, an increasing interest is arising in evalu-
ating the possible benefits that niosome entrapment can offer in com-
parison with liposomes in addition to general considerations as low
costs and higher stability towards oxidative degradation.
Most of published results concern the ability of vesicles to enhance
percutaneous permeation, thus increasing drug effectiveness after topi-
cal administration. Formulations with niosomes demonstrated a better
skin permeation potential, sustained release characteristic, and higher
stability as compared to liposomes.
Two mechanism were proposed for the enhancement of permeation
of drugs loaded in niosomes: a permeation enhancer effect and direct
vesicle fusion with the Stratum Corneum (SC) (Fig. 8). This mechanism
is supported by Fang et al. who investigated the feasibility of niosomes
for the transdermal administration of enoxacin and showed that
enhancing effect for skin permeation of niosomes was greater than that
of liposomes composed of dimyristoyl phosphatidylcholine (DMPC)
[111]. These results were confirmed by several authors for different sur-
factant vesicles and different drugs, regardless of the physico-chemical
properties of analyzed nanocarriers: tretinoin [112], resveratrol [216]
and bacitracin zinc [217] in niosomes showed a better behavior than
liposomal formulations in the cutaneous delivery.
On the other hand, comparative studies of liposomes and niosomes
for topical delivery of anticancer drug to cancer cells reported opposing
results. When ciclopirox olamine (CPO) was encapsulated in liposomes
and niosomes, niosomal CPO in comparison to liposomal and pure CPO
demonstrated cytotoxic effect at significantly lower concentration
selectively on cancer cells (KB-oral cancer, PC3-prostate cancer,
Siha-cervical cancer and Vero-kidney epithelial cell lines), improving
the anticancer potential of CPO [218]. On the contrary, in a comparative
study of liposomes and niosomes for topical delivery of 5-fluorouracil
(D)
(E)
(C)
 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206
(5-FU) to skin cancer cells (HaCaT), liposomes were found to be more
cytotoxic on the cell line in comparison with niosomes [219].
8. Niosome-based patents and pre-clinical and clinical trials
In spite of the large number of publications and the wide research
carried out over the last few decades of niosome formulations and
possible applications, as above reported, only few studies have stepped
into patents and pre-clinical and clinical trials, and these are exclusively
oriented towards topical delivery.
About 200 patents concern vesicular carriers, including niosomes,
but patents exclusively based on surfactant vesicles are very few and
have been developed in the last five years: most of them reported that
drug inclusion in niosomes has the advantages of improved curative
effect, reduced adverse side effect and convenient route of administra-
tion. The “niosomal approach” was proposed for different drugs: tacro-
limus for ophthalmic application after corneal transplantation [220];
beclomethasone for pulmonary delivery [221]; collagen for dermal
delivery [222]; metformin for oral delivery [223] and immunogenic
compounds, including an inactivated virus [224].
All proposed preparation processes are simple and suitable for com-
mercialization and it must be pointed out that the reported applications
for lyophilized niosomal formulation [221,224] could be useful in
further development of surfactant vesicular carriers.
Recent patented niosomal formulation concern surfactant vesicles
with targeting properties exerted by inducing a cholinesterase-
mediated disruption of niosome with time [225] or by surfactant deriv-
atization with folic acid [226].
Among niosome-based patents also peculiar applications have
been proposed such as the preparation of niosome probe for the detec-
tion of neomycin with advantages of high selectivity, high sensitivity
and wide range of application [227] or the possibility to use ultrasound
to mediate delivery non-invasively by altering the niosome membrane
structure [228].
Pre-clinical and clinical trials are exclusively oriented towards
dermal and transdermal delivery (Table 9).
Lakshmi et al. [129,229] reported the preparation of niosomal meth-
otrexate in chitosan gel to be tested for irritation and sensitization on
healthy human volunteers and for assessing the efficacy of the gel on
psoriasis patients. Obtained results suggested that the niosomal metho-
trexate gel is more effective than placebo (as expected) but also of the
marketed methotrexate gel.
NSVs loading urea, dispersed in chitosan gel, were tested on psoria-
sis affected patients with less than 25% severity of any category of pso-
riasis. The niosomal gel produced significant reduction in the lesion than
the plain urea gel. The niosomal urea gel produced greater reduction in
total score and desquamation score compared to erythema and infiltra-
tion score and proved that niosomal urea in chitosan gel can be used as
an adjuvant in the treatment of psoriasis [230].
A novel sustained release proniosomal system for transdermal
delivery of vinpocetine was designed using sugar esters as non-ionic
surfactants in which proniosomes were converted to NSVs upon skin
water hydration following topical application under occlusive condi-
tions. After in vitro characterization, an optimized formula was clinically
assessed for transdermal pharmacokinetic evaluations and a larger
Table 9
Niosomal formulations in pre-clinical and clinical trials (dermal and transdermal delivery).
Active compound Formulation
Finasteride Proniosomal gel
Griseofulvin Niosomal gel
Methotrexate Niosomal gel
Urea Niosomal gel
Vinpocetine Proniosomal gel
Rice bran bioactive compounds (RBBC) Niosomal gel, niosomal cream
Gallic acid Niosomal gel
203
absorption of the drug was reported when compared to oral tablet
administration [231].
Clinical trials by phototrichogram method were performed for
proniosomal formulation of finasteride on twenty healthy male volun-
teers and it was found to increase the anagen hair count of test group
by 42.85% when compared with the control group (6.6%), indicating
that the proniosomal gel formulation achieved the objective of enhanc-
ing the drug concentration in androgenic receptors of the hair shaft and
thus it can be used safely for treating androgenic alopecia [232].
The efficacy of a niosomal gel formulation of griseofulvin in the treat-
ment of tinea circinata was evaluated on sixteen patients, comparing it
to a griseofulvin gel form and to a placebo gel. The results evidenced an
effective and safe approach in treating tinea circinata, but underlined
that a further large-scale studies will be needed to establish the higher
efficacy of the niosomal gel formulation [233].
Recently, niosomal gel formulations were developed as anti-aging
agents: niosomes entrapping rice bran bioactive compounds or loaded
with a semi-purified fraction from terminalia chebula galls were tested
on human volunteers by measuring skin elasticity and roughness. In
both studies a superior clinical anti-aging activity of the niosomal gel
was reported [234,235].
9. Concluding remarks and perspectives
Niosomes have been extensively studied as an alternative to
liposomes. NSVs have some advantages over liposomes, such as their
relatively higher chemical stability, improved purity and relatively
lower cost in comparison with liposomes. The niosome bilayer is differ-
ent from the liposome bilayer in that NSVs are prepared from uncharged
single-chain surfactants and cholesterol whereas liposomes are
prepared from double-chain phospholipids (neutral or charged) and
cholesterol. As with liposomes, the in vitro/in vivo properties of
niosomes depend both on the composition of the bilayer and on method
of their production. Niosomal drug delivery has yet to make its clinical
debut even though these systems have been demonstrated to yield
promising therapeutics. NSVs alter the plasma clearance kinetics, tissue
distribution, metabolism and cellular interaction of the drug.
Although numerous papers, among those reported in this review,
show data related to cytotoxicity of niosomal vesicles as well as of
surfactant monomers; unfortunately, up to date there are no specific
wide studies aimed to investigate toxicity of NSVs after in vivo adminis-
tration, in particular in long term trials. From published information it is
possible to evaluate the safe concentration of unstructured and struc-
tured surfactants that can be used in cell interaction studies.
The lack of a specific toxicological study can represent an intriguing
opportunity for toxicologists to enrich the knowledge in drug delivery
system tolerability. For example there are no studies on the fate of
surfactant in cells, animals, humans, while the understanding of their
metabolism is a fundamental step in reaching the clinical trials.
Lipidic vesicles are the first nanomedicine delivery system to make
the transition from bench to clinical application, and they are now an
established technology platform with high clinical acceptance. We can
look forward to many more efforts in developing surfactant vesicles,
according to the evidence that, despite the many promising proof of
Application Control group Reference
Androgenetic alopecia Finasteride gel [232]
Treatment of tinea circinata Griseofulvin gel and placebo gel [233]
Psoriasis Marketed methotrexate gel [129,229]
Psoriasis Urea gel [230]
Cerebrovascular disorders Oral tablet administration [231]
Skin aging RBBC gel or cream [234]
Skin aging Gallic acid gel [235]
204 C. Marianecci et al. / Advances in Colloid and Interface Science 205 (2014) 187–206
concept studies there is still a long road ahead to becoming a clinical
reality.
In addition, the big chance to the field consists of its highly interdis-
ciplinary nature from basic chemistry and physic to clinical evaluations.
This means there are still many open challenges that need to be
overcome and there are still opportunities for academic and industrial
scientists to make a decisive impact.
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