Advanced Drug Delivery Reviews 56 (2004) 1257 1272

www.elsevier.com/locate/addr

Solid lipid nanoparticles for parenteral drug delivery

S.A. Wissing a, O. Kayser b, R.H. Muller b,*
b

a
DDS, Drug Delivery Services, GmbH, Kronskamp 11, 24119 Kronshagen, Germany
Department of Pharmaceutics, Biopharmaceutics and Biotechnology, Free University of Berlin, Kelchstrasse 31, 12169 Berlin, Germany

Received 13 August 2003; accepted 20 December 2003

Abstract
This review describes the use of nanoparticles based on solid lipids for the parenteral application of drugs. Firstly, different
types of nanoparticles based on solid lipids such as solid lipid nanoparticles (SLN), nanostructured lipid carriers (NLC)
and lipid drug conjugate (LDC) nanoparticles are introduced and structural differences are pointed out. Different production
methods including the suitability for large scale production are described. Stability issues and drug incorporation mechanisms
into the particles are discussed. In the second part, the biological activity of parenterally applied SLN and biopharmaceutical
aspects such as pharmacokinetic profiles as well as toxicity aspects are reviewed.
D 2004 Elsevier B.V. All rights reserved.
Keywords: SLN; Solid lipid nanoparticles; NLC; LDC; Toxicity; Pharmacokinetics; Degradation; Parenterals

Contents
1.
2.

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
Nanoparticles based on solid lipids . . . . . . . . . . . . . . . . . . . . . .
2.1.
Definitions and structural features. . . . . . . . . . . . . . . . . . .
2.1.1. SLN. . . . . . . . . . . . . . . . . . . . . . . . . . . .
2.1.2. NLC . . . . . . . . . . . . . . . . . . . . . . . . . . .
2.1.3. LDC . . . . . . . . . . . . . . . . . . . . . . . . . . .
2.2.
Production of lipid nanoparticles . . . . . . . . . . . . . . . . . . .
2.2.1. High pressure homogenisation (HPH) . . . . . . . . . . . .
2.2.2. Production of SLN via microemulsions. . . . . . . . . . . .
2.2.3. Preparation by solvent emulsification-evaporation or -diffusion .
2.2.4. Preparation by w/o/w double emulsion method . . . . . . . .
2.2.5. Preparation by high speed stirring and/or ultra sonication . . .
2.2.6. Scale up feasibility . . . . . . . . . . . . . . . . . . . . .
2.3.
Stability of SLN dispersions . . . . . . . . . . . . . . . . . . . . .
2.4.
Incorporation of drugs . . . . . . . . . . . . . . . . . . . . . . . .
2.5.
Release of incorporated drugs . . . . . . . . . . . . . . . . . . . .

* Corresponding author. Tel.: +49-30-838-50678; fax: +49-30-838-50616.

E-mail address: mpharma@zedat.fu-berlin.de (R.H. Muller).
0169-409X/$ - see front matter D 2004 Elsevier B.V. All rights reserved.
doi:10.1016/j.addr.2003.12.002

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S.A. Wissing et al. / Advanced Drug Delivery Reviews 56 (2004) 12571272

2.5.1. Drug release models. . . . . . . . . . . . . . .

2.5.2. In vitro release of drugs from SLN . . . . . . . .
3. Biological activity and biopharmaceutical aspects . . . . . . . . .
3.1. General aspects . . . . . . . . . . . . . . . . . . . . .
3.2. Administration and drug liberation. . . . . . . . . . . . .
3.2.1. Pharmacokinetic profile . . . . . . . . . . . . .
3.3. Safety aspects . . . . . . . . . . . . . . . . . . . . . .
3.3.1. Toxicity and status of excipients . . . . . . . . .
3.3.2. Blood clotting and interaction with serum proteins .
3.3.3. Allergic reactions . . . . . . . . . . . . . . . .
3.4. Tissue distribution and drug targeting . . . . . . . . . . .
4. Outlook . . . . . . . . . . . . . . . . . . . . . . . . . . . .
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

1. Introduction
In the 1960s, the first safe parenteral fat emulsion
(Intralipid) was developed by Wretlind for parenteral
nutrition [1]. This was the beginning of a new delivery
system for lipophilic drugs, which can be incorporated
easily into the oil droplets. Successful market products
are, e.g. Diazemuls (1970s) and Diprivan (1980s). The
main advantage of this carrier system is the reduction
of side effects caused at the injection side [2]. A major
disadvantage however is the critical physical stability
of the drug containing emulsions due to a reduction of
the zeta potential (ZP) which can lead to agglomeration, drug expulsion and eventually breaking of the
emulsion [3].
Another interesting parenteral carrier systems are
the liposomes. They have been described for the first
time by Bangham et al. in the 1960s and were
introduced as drug delivery vehicles in the 1970s
[4,5]. Trade products are, e.g. Ambisome (1990,
Europe, 1997, USA), DaunoXome (1996, Europe
and USA) and Doxil (1995, USA, 1996, Europe).
These products have been developed in order to
reduce toxic side effects of the incorporated highly
potent drugs and increase the efficacy of the treatment
[6,7]. Major obstacles for the development of liposomal formulations wereand partly still arelimited
physical stability of the dispersions, drug leakage, low
activity due to no specific tumour targeting, non
specific clearance by the mononuclear phagocytic
system (MPS) and difficulties in upscaling [6,8].
Polymeric nanoparticles made from non-biodegradable and biodegradable polymers are yet another
innovative parenteral carrier system. Advantages of
these particles are site-specific targeting and con-

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trolled release of the incorporated drugs [9]. However,

the cytotoxicity of the polymers after internalisation
into cells is a crucial and often discussed aspect [10].
Also, large scale production of polymeric nanoparticles is problematic. Therefore, this carrier system has
so far not been relevant for the pharmaceutic market.
In the middle of the 1990s, the attention of
different research groups has focussed on alternative
nanoparticles made from solid lipids, the so-called
solid lipid nanoparticles (SLN or lipospheres or
nanospheres) [11 26]. The SLN combine the advantages of other innovative carrier systems (e.g. physical stability, protection of incorporated labile drugs
from degradation, controlled release, excellent tolerability) while at the same time minimising the
associated problems. SLN formulations for various
application routes (parenteral, oral, dermal, ocular,
pulmonar, rectal) have been developed and thoroughly characterised in vitro and in vivo [27 41]. A first
product has recently been introduced to the Polish
market (Nanobase, Yamanouchi) as a topically applied moisturiser.
At the turn of the millenium, modifications of
SLN, the so-called nanostructured lipid carriers
(NLC) and the lipid drug conjugate (LDC) nanoparticles have been introduced to the literature [42
45]. These carrier systems overcome observed limitations of conventional SLN.
This paper intends to describe briefly the different
lipid based carrier systems SLN, NLC and LDC,
structure and associated features, stability, applied
production methods, drug incorporation and drug
release mechanisms. The bioactivity of SLN after
parenteral application, i.e. tolerability, toxicology,
cellular uptake, albumin adsorption, pharmacokinet-

S.A. Wissing et al. / Advanced Drug Delivery Reviews 56 (2004) 12571272

ics, tissue distribution and drug targeting is reviewed

in detail.

2. Nanoparticles based on solid lipids

2.1. Definitions and structural features
SLN, NLC and LDC are particles with a solid lipid
matrix with an average diameter in the nanometer
range. In addition to lipid and drug, the particle
dispersions contain surfactants as stabilisers. All excipients are GRAS substances or have an accepted
GRAS status, therefore, a wide variety of substances
can be used for formulating purposes [46].
2.1.1. SLN
SLN are particles made from solid lipids (i.e. lipids
solid at room temperature and also at body temperature) and stabilised by surfactant(s). By definition, the
lipids can be highly purified triglycerides, complex
glyceride mixtures or even waxes [14]. Recently, SLN
based on para-acyl-calix[4]arenes have been prepared
and studied [47,48]. Through the work of various
research groups, the carrier system SLN has been
characterised intensively. For detailed information
on production, characterisation and application, the
reader is referred to the main review papers up to date
[17,49,50]. The first patents have been granted in
1993 and 1996 and contain claims on different production methods of SLN [12,14].
The main features of SLN with regard to parenteral
application are the excellent physical stability, protection of incorporated labile drugs from degradation,
controlled drug release (fast or sustained) depending
on the incorporation model, good tolerability and sitespecific targeting. Potential disadvantages such as
insufficient loading capacity, drug expulsion after
polymorphic transition during storage and releatively
high water content of the dispersions (70 99.9%)
have been observed.
The drug loading capacity of conventional SLN is
limited (generally up to approximately 25% with
regard to the lipid matrix, up to 50% for special
actives such as Ubidecarenone) by the solubility of
drug in the lipid melt, the structure of the lipid matrix
and the polymorphic state of the lipid matrix
[13,15,18,19,21,49,51 53]. If the lipid matrix con-

1259

sists of specially similar molecules (i.e. tristearin or

tripalmitin), a perfect crystal with few imperfections is
formed. Since incorporated drugs are located between
fatty acid chains, between the lipid layers and also in
crystal imperfections, a highly ordered crystal lattice
cannot accommodate large amounts of drug [19].
Therefore, the use of more complex lipids (mono-,
di-, triglycerides, different chain lenghths) is more
sensible for higher drug loading.
The transition to highly ordered lipid particles is
also the reason for drug expulsion. Directly after
production, lipids crystallisepartiallyin higher
energy modifications (a, hV) with more imperfections
in the crystal lattice [54 57]. The preservation of the
a-modification during storage and transformation after administration (e.g. by temperature changes) could
lead to a triggered and controlled release and has
recently been investigated for topical formulations
[58]. If however a polymorphic transition to h takes
place during storage, the drug will be expelled from
the lipid matrix and it can then neither be protected
from degradation nor released in a controlled way.
2.1.2. NLC
NLC have been introduced at the end of the 1990s
in order to overcome the potential difficulties of SLN
described above [43,44,57]. The goal was the development of a nanoparticulate lipid carrier with a certain
nanostructure in order to increase the payload and
prevent drug expulsion.
This could be realised in three ways. In the first
model, spatially different lipids, e.g. glycerides composed of different fatty acids are mixed. Using spatially different lipids leads to larger distances between
the fatty acid chains of the glycerides and general
imperfections in the crystal and thus to more room for
the accommodation of guest molecules. The highest
drug load could be achieved by mixing solid lipids
with small amounts of liquid lipids (oils). This model
is called imperfect type NLC.
If higher amounts of oil are mixed with the solid
lipid, a different type of nanostructure is present.
Here, the solubility of the oil molecules in the solid
lipid is exceeded; this leads to phase separation and
the formation of oily nanocompartments within the
solid lipid matrix [59 61]. Many drugs show a higher
solubility in oils than in solid lipids so that they can be
dissolved in the oil and still be protected from

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S.A. Wissing et al. / Advanced Drug Delivery Reviews 56 (2004) 12571272

degradation by the surrounding solid lipids. This type

of NLC is called multiple type NLC and can be
regarded as an analogue to w/o/w emulsions since it is
an oil-in-solid lipid-in-water dispersion.
Since drug expulsion is caused by ongoing crystallisation or transformation of the solid lipid, this can
be prevented by the formation of a third type, the
amorphous type NLC. Here, the particles are solid
but crystallisation upon cooling is avoided by mixing
special lipids (e.g. hydroxyoctacosanylhydroxystearate and isopropylmyristate) [60,61].
So far, NLC have only been exploited for the
topical delivery, however their advantages over conventional SLN are also of interest for parenteral
application routes.
2.1.3. LDC
SLN are useful for the incorporation of lipophilic
drugs. Due to partitioning effects during the production
process, only highly potent hydrophilic drugs which
are effective in low concentrations (e.g. LHRH or EPO)
can be firmly incorporated in the solid lipid matrix
[32,62]. In order to overcome this limitation, the socalled LDC nanoparticles with drug loading capacities
of up to 33% have been developed at the turn of the
millenium [45,63]. Here, an insoluble drug lipid conjugate bulk is prepared either by salt formation (e.g.
with a fatty acid) or by covalent linking (e.g. to esters or
ethers). In the salt formation process, the free drug base
and fatty acid are dissolved in a suitable solvent. The
solvent is then consequently evaporated under reduced
pressure. For the covalent linking, the drug (salt) and a
fatty alcohol react in presence of a catalyst and the LDC
bulk is then purified by recrystallisation. The obtained
LDC bulk is then processed with an aqueous surfactant
solution to a nanoparticle formulation using high
pressure homogenisation (HPH).
In the literature, studies on the cytotoxicity of
surface-modified diminazenestearate and -oleate
LDC, plasma protein absorption (2-DE) and activity
against trypanosomiasis via brain targeting have been
performed [45,64,65]. The results will be presented in
the context of this review.
2.2. Production of lipid nanoparticles
Different approaches exist for the production of
finely dispersed lipid nanoparticle dispersions. In this

section, the various methods are described briefly,

also with regard to scaling up possibility, a prerequisite for the introduction of a product to the
market.
2.2.1. High pressure homogenisation (HPH)
HPH is a suitable method for the preparation of
SLN, NLC and LDC and can be performed at elevated
temperature (hot HPH technique) or at or below room
temperature (cold HPH technique) [13,14,18,66 68].
The particle size is decreased by cavitation and
turbulences.
Briefly, for the hot HPH, the lipid and drug are
melted (approximately 5 jC above the melting point
of the lipid) and combined with an aqueous surfactant
solution having the same temperature. A hot preemulsion is formed by high speed stirring. The hot
pre-emulsion is then processed in a temperature controlled high pressure homogeniser, generally a maximum of three cycles at 500 bar are sufficient. The
obtained nanoemulsion recrystallises upon cooling
down to room temperature forming SLN, NLC or
LDC.
The cold HPH is a suitable technique for processing temperature labile drugs or hydrophilic drugs.
Here, lipid and drug are melted together and then
rapidly ground under liquid nitrogen forming solid
lipid microparticles. A pre-suspension is formed by
high speed stirring of the particles in a cold aqueous
surfactant solution. This pre-suspension is then homogenised at or below room temperature forming SLN,
NLC or LDC, the homogenising conditions are generally five cycles at 500 bar.
The influence of homogeniser type, applied pressure, homogenisation cycles and temperature on particle size distribution has been studied extensively
[13,18,69,70]. Both HPH techniques are suitable for
processing lipid concentrations of up to 40% and
generally yield very narrow particle size distributions
(polydispersity index < 0.2) [71,72].
2.2.2. Production of SLN via microemulsions
The group of Gasco has developed and optimised a
suitable method for the preparation of SLN via microemulsions which has been adapted and/or modified by
different labs [12,16,22,66,73,74]. Firstly, a warm
microemulsion is prepared by stirring, containing
typically c 10% molten solid lipid, 15% surfactant

S.A. Wissing et al. / Advanced Drug Delivery Reviews 56 (2004) 12571272

and up to 10% cosurfactant. This warm microemulsion is then dispersed under stirring in excess cold
water (typical ratio c 1:50) using an especially
developed thermostated syringe. The excess water is
removed either by ultra-filtration or by lyophilisation
in order to increase the particle concentration.
Experimental factors such as microemulsion composition, dispersing device, temperature and lyophilisation on size and structure of the obtained SLN have
been studied intensively. It has to be remarked critically, that the removal of excess water from the
prepared SLN dispersion is a difficult task with regard
to the particle size. Also, high concentrations of
surfactants and cosurfactants (e.g. butanol) are necessary for formulating purposes, however less desirable
with respect to regulatory purposes and application.
2.2.3. Preparation by solvent emulsification-evaporation or -diffusion
Different academic groups have attempted the
production of SLN via precipitation. In the solvent
emulsification-evaporation [25,47,48], the lipid is
dissolved in a water-immiscible organic solvent (e.g.
toluene, chloroform) which is then emulsified in an
aqueous phase before evaporation of the solvent under
reduced pressure. Upon evaporation of the solvent, the
lipid precipitates forming SLN. An important advantage of this method is the avoidance of heat during the
preparation, which makes it suitable for the incorporation of highly thermolabile drugs. Problems might
arise due to solvent residues in the final dispersion;
Sjostrom et al. have calculated the amount of toluene
residues as 20 100 ppm in final dispersions. Also,
these dispersions are generally quite dilute, because of
the limited solubility of lipid in the organic material.
Typically, lipid concentrations in the final SLN dispersion range around 0.1 g/l, therefore, the particle
concentration has to be increased by means of, e.g.
ultra-filtration or evaporation.
In the solvent-diffusion technique, partially watermiscible solvents (e.g. benzyl alcohol, ethyl formate)
are used [75,76]. Initially, they are mutually saturated
with water to ensure initial thermodynamic equilibrium of both liquids. Then, the lipid is dissolved in the
water-saturated solvent and subsequently emulsified
with solvent-saturated aqueous surfactant solution at
elevated temperatures. The SLN precipitate after the
addition of excess water (typical ratio: 1:5 1:10) due

1261

to diffusion of the organic solvent from the emulsion

droplets to the continuous phase. Similar to the
production of SLN via microemulsions, the dispersion
is fairly dilute and needs to be concentrated by means
of ultra-filtration or lyophilisation.
Average particle sizes around 100 nm and very
narrow particle size distributions can be achieved by
both solvent evaporation methods.
2.2.4. Preparation by w/o/w double emulsion method
Recently, a novel method based on solvent emulsification evaporation for the preparation of SLN
loaded with hydrophilic drugs has been introduced
to the scientific community [66]. Here, the hydrophilic drug is encapsulatedalong with a stabiliser to
prevent drug partitioning to the external water phase
during solvent evaporationin the internal water
phase of a w/o/w double emulsion. This technique
has been used for the preparation of sodium cromoglycate-containing SLN, however, the average size
was in the micrometer range so that the term lipospheres in the sense as a term for nanoparticles is not
used correctly for these particles.
2.2.5. Preparation by high speed stirring and/or ultra
sonication
The SLN were developed from lipid microparticles produced by spray congealing followed by
lipid nanopellets produced by high speed stirring or
sonication [77,78]. A great advantage of this method is the fact that the equipment is common in
every lab and the production can easily be done.
The problem of high speed stirring was a broader
particle size distribution ranging into the micrometer range. This lead to physical instabilities such as
particle growth upon storage. This could be improved by higher surfactant concentrations, which
in order might be correlated with toxicological
problems after parenteral administration. A further
disadvantage is potential metal contamination due
to ultra sonication.
Therefore, studies have been performed by various
research groups in order to improve the stability of the
obtained SLN dispersions. Generally, high speed
stirring and ultra sonication are combined and performed at elevated temperatures for some time
[79,80]. Quite narrow and physically stable distributions can be achieved, however the lipid concentration

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S.A. Wissing et al. / Advanced Drug Delivery Reviews 56 (2004) 12571272

is low ( < 1%) and the surfactant concentration is

comparably high.
2.2.6. Scale up feasibility
For the introduction of a product to the market,
scaling up feasibility is of uttermost importance. For
the two primarily used methods for the preparation
of lipid nanoparticles, HPH and production via
microemulsions, scaling up possibilities have been
investigated.
For HPH, a GMP unit for clinical batch production
was developed and validated which can achieve batch
sizes between 2 and 10 kg SLN dispersion. Further, a
large scale production line was designed having a
capacity of 50 150 kg SLN dispersion per h by
placing two homogenisers in series. The production
of SLN with these lines can be performed in discontinuous or continuous modes [18,81 84].
For the production via microemulsions, a system
has been developed allowing the production of 100 ml
microemulsion which is then poured in excess cold
water forming SLN. The dispersing water ration
ranged from 1:1 to 1:10, leading to batch sizes of
up to 1.1 l. Experimental factors which have been
investigated were the pressure of the pneumatic cylinder, the temperature and the needle gauge of the
syringe containing the microemulsion as well as the
volume of dispersing water [85].
2.3. Stability of SLN dispersions
The physical stability of SLN dispersions has been
investigated intensively, e.g. by measurements of
particle size (photon correlation spectroscopy, PCS;
laser diffraction, LD), charge (ZP) and thermal analysis (differential scanning calorimetry, DSC).
Physical stability of optimised aqueous SLN dispersions is generally more than 1 year ([51,53] and
Muller et al. could show stability for SLN made from
glyceryl palmitostearate or tribehenate for up to 3
years by PCS [20]. The average diameter of the main
population remained between 160 and 220 nm for the
investigated period.
Freitas and Muller investigated the effect of light
and temperature on the physical stability of SLN
dispersions composed of 10% tribehenate and 1.2%
poloxamer 188 [86]. They found that particle growth
could be induced by an input of kinetic energy (light,

temperature) to the system. Storage under artificial

light lead to gelation of the system within 7 days of
storage, under day light within 3 months and in
darkness particle growth started after 4 months storage. The gelation was accompanied by a decrease in
ZP from 24.7 to below 18 mV. The influence of
the storage temperature on particle size has also been
analysed. The authors found that the particle size
measured by LD increased rapidly at elevated temperatures and remained stable for more than 180 days
when refrigerated. Again, particle growth could be
correlated to a decrease in ZP from 24.7 to approximately 15 mV.
Freitas and Muller have also correlated the physical stability of the aforementioned SLN formulation
with the polymorphic state of the lipid [54]. After
hot HPH, the lipid is present in a mixture of hV, a
and sub a polymorphs. The input of kinetic energy
causes a transformation to hV accompanied by gel
formation. By inhibition of this transformation (refrigerated, dark storage), this transformation could be
avoided. These studies show that the development of
optimal storage conditions can improve the physical
stability of previously regarded unstable SLN formulations tremendously.
For the recently developed SLN based on calixarenes, stability data of more than 1 year have been
published. Shahgaldian et al. have further investigated
the influence of the ionic strength on stability, observing strongest destabilisation by sulphate ions [47].
Apart from optimised storage conditions of labile
SLN dispersions, they can also be spray-dried or
lyophilised. For spray-drying, a melting point of the
lipid matrix of >70 jC is a pre-requisite. Typically,
protectors such as trehalose are added to the dispersion
in concentrations of about 20 25%. For best reconstitution effects, SLN concentration in the spraying medium should be approximately 1%. The influence of
lipid type and concentration, carbohydrate type and
concentration, redispersion medium and spraying medium have been investigated by Freitas et al. [20,87].
Lyophilisation can be employed as an alternative
very sensitive drying method. The process has been
optimised with regard to operating conditions, lipid
concentration, type and concentration of cryoprotectant
and redispersing conditions [20,88]. Heiati et al. have
investigated the effect of cryoprotective sugars on the
size of neutral and negatively charged SLN after

S.A. Wissing et al. / Advanced Drug Delivery Reviews 56 (2004) 12571272

lyophilisation and reconstitution [89]. The azidothymidine palmitate (AZT-P) loaded SLN were composed of
trilaurin, stabilised with lecithin and they were prepared by solvent emulsification evaporation and subsequent HPH. They found that trehalose was the most
effective cryoprotectant in a sugar/lipid ratio of 3:9 for
neutral SLN and 2:6 for negatively charged SLN. Also,
trehalose was most effective for preventing drug expulsion upon reconstitution. Lim et al. showed for alltrans retinoic acid-loaded SLN excellent redispersion
characteristics [67]. Here, the PCS diameter increased
merely from 182 to 265 nm and the polydispersity
index from 0.173 to 0.200 upon redispering. No
changes in ZP and in drug loading were observed.
For SLN based on calixarenes, difficulties in redispersing after lyophilisation have been observed, i.e. these
particles require up to 1 h of ultrasonic treatment,
however no cryoprotectant was added to the formulations, so that these results have to be regarded as
preliminary.
Radomska et al. have developed an analytical method for the determination of the chemical stability of the
lipid matrix of SLN [90]. They extracted the lipid from
SLN dispersions and analysed it qualitatively and
quantitatively by gas chromatography. For SLN made
from cetyl palmitate and tristearate, they could show
chemical stability of the lipids of H90% after 2 years
storage at room temperature as well as physical stability
for up to 2 years when stored at 4 8 jC.
Regarding the parenteral application of SLN, sterility has to be ensured. Schwarz et al. have studied the
influence of autoclaving conditions on particle size and
ZP of different SLN formulations [13]. They observed
that steric stabilisation of trilaurin SLN with poloxamer
188 was not suitable for autoclaving due to its critical
flocculation temperature. For lecithin stabilised SLN
formulations, autoclaving was possible. No increase in
particle size was observed by PCS. Alternatively, SLN
could be sterilised by gamma irradiation or (if the size is
well below 200 nm) by filtration. Various other research groups have also published particle size and ZP
data of sterilised SLN dispersions revealing excellent
stability of the formulations [47,67,89,91].
2.4. Incorporation of drugs
An innovative and successful carrier system should
allow a high loading capacity for incorporated drugs

1263

as well as long-term incorporation. Many different

drugs have been incorporated in SLN, NLC or LDC.
Table 1 lists examples relevant for the parenteral
application including the corresponding references.
The drug can be incorporated between fatty acid
chains, between lipid layers or in imperfections.
Depending on the drug/lipid ratio and solubility, the
drug is located mainly in the core of the particles, in
the shell or molecularly dispersed throughout the
matrix. This influences the drug release and will be
explained in more detail in Section 2.5. The influence
of drug incorporation on particle size and physical
stability has been described before [19,51]. By optimising of the formulation, long term physical stability
can be achieved [20].
The solid lipid core of SLN should increase the
chemical stability of incorporated drugs and protect
them from degradation. Therefore, entrapment efficiency (E.E.) and long term retention of the drug in
the lipid matrix have to be ensured. Heiati et al. have
studied the incorporation and retention of AZT-P in

Table 1
Examples of drugs relevant for parenteral application incorporated
into SLN
Drug

References

AZT-P and derivatives

Camptothecin
Clobetasol propionate
Cortisone
Sodium cromoglycate
Cyclosporin A
Diazepam
Diminazenediaceturate
Doxorubicin
Etomidate
3V5V-Dioctanoyl-5-fluoro-2V-deoxuridine
Hydrocortisone
Idarubicin
[D-Trp-6]LHRH
Magnetite
Mifepristone
Paclitaxel
Pilocarpine
Piribedil
Prednisolone
Progesterone
Retinoic acid
Tetracaine
Thymopentin
Tobramycin

[89,99,100]
[40,98]
[76]
[51]
[66]
[97]
[41,51]
[45]
[31,101 103]
[93]
[79]
[34]
[31]
[32]
[74]
[80]
[91,103,104]
[33]
[29]
[51,93,94]
[34,66]
[67]
[93]
[105]
[35,37,106]

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S.A. Wissing et al. / Advanced Drug Delivery Reviews 56 (2004) 12571272

trilaurin SLN [89]. They found a dependence of the

E.E. on the phospholipid content of the formulation
and on the charge of the SLN. The formation of
phospholipid bilayers and the ability of amphiphilic
drugs such as AZT-P to integrate within these bilayers
is postulated as the reason for the increased E.E.
However, it should be kept in mind that phospholipids
can also form independent vesicles and integrate
drugs. These vesicles scatter only weakly and might
not be detected with PCS or LD. Ahlin et al. have
studied the location and E.E. of lipophilic spin probes
(C14- and C18-Tempo) by electron paramagnetic resonance (EPR) [92]. They could locate the spin probes
between the solid glyceride core and the soft phospholipid layer on the particle surface or at the lipid/
water interface and found E.E.s close to 100%. Hou et
al. have investigated mifepristone-loaded SLN based
on glycerol monostearate, stabilised with Tween 80
regarding E.E. and retention after 1 month [80]. For
formulations optimised regarding physical stability,
they found an E.E. of 87.89% on day 1 and 82.05%
after 1 month storage at 4 8 jC, the decrease correlated with lipid transitions. It has to be pointed out that
lipid transition during storage and subsequent drug
expulsion from the matrix can be avoided by formulation as NLC. Also, for hydrophilic drugs the formulation as LDC is more suitable for higher loading
capacities and retention within the particle matrix
upon storage.
2.5. Release of incorporated drugs
Next to the characterisation of the carrier system
SLN, the release characteristics have been studied by
various research groups. It could be shown that the
release profile can be influenced by modifications in
the lipid matrix, surfactant concentration and production parameters. Mehnert et al. have performed intensive in vitro release studies and developed structural
models for different release characteristics [93]. This
section of the paper presents in vitro and in vivo
release data published by different groups.
2.5.1. Drug release models
Mehnert et al. have observed variable release
profiles for SLN loaded with different drugs
[19,93 95]. They found the fastest release for tetracaine, followed by etomidate and extremely sustained

release for prednisolone. The proposed structural

models represented in Fig. 1 serve as an explanation.
Tetracaine SLN were prepared produced by hot
HPH at a drug concentration well below saturation
solubility in the lipid, i.e. during the production, the
drug partitioned to the water phase. Upon cooling, the
lipid precipitates first due to phase separation. Simultaneously, the drug re-partitions into the liquid lipid
phase and its concentration in the outer shell liquid
lipid increases continuously. Finally, the drugenriched shell crystallises. Atomic force microscopy
(AFM) studies on coenzyme Q10 loaded SLN have
shown a soft surface of the particles undermining this
thesis [96]. If the drug is located primarily in the shell
of the particles, a burst release is highly likely. Other
factors contributing to a fast release are large surface
area, high diffusion coefficient (small molecular size),
low matrix viscosity and short diffusion distance of
the drug.
Etomidate SLN represent the homogeneous solid
solution matrix model [93]. A homogeneous distribution of drugs in the particle matrix can be achieved by
cold HPH or by the avoidance of potentially drugsolubilising surfactants.
SLN loaded with prednisolone released the drug in
vitro (i.e. in absence of enzymes) over a period of
more than 5 weeks [93,94]. For them, the drugenriched core model is proposed. The prednisolone
concentration in the matrix material was close to its
saturation solubility. Cooling of the hot nanoemulsion
prepared by HPH leads to supersaturation of the drug
in the liquid lipid and subsequently to drug precipitation prior to lipid crystallisation. The drug-enriched
core is surrounded by a practically drug-free lipid
shell. Due to the increased diffusional distance and

Fig. 1. Proposed structural models for drug-containing SLN

(modified after [93]).

S.A. Wissing et al. / Advanced Drug Delivery Reviews 56 (2004) 12571272

hindering effects by the surrounding solid lipid shell,

the drug has a sustained release profile.
2.5.2. In vitro release of drugs from SLN
Apart from the in vitro release data presented in the
above section, various other studies have been published with regard to potential parenteral application.
Cavalli et al. have prepared stealth and non-stealth
tripalmitin SLN loaded with paclitaxel in order to
provide an alternative for the parenteral administration
[91]. The commercially available product TaxolR is a
toxicologically critical micellar solution of the drug in
Cremophor EL. Cavalli et al. report sustained in vitro
release: 0.1% of the paclitaxel is released into the
receptor medium (phosphate buffer, pH 7.4) after 120
min, this is correlated to first pseudo zero oder
kinetics. The same academic group has previously
shown similarly sustained in vitro release profiles for
doxorubicin and idarubicin (0.1% after 120 min) in
contrast to burst release from reference solutions [31].
For stearic acid SLN containing cyclosporin A, they
determined an in vitro release of < 4% after 2 h compared to >60% from solution [97].
Yang et al. determined the in vitro release of
camptothecin from stearic acid SLN in conjunction
with potential targeting to the brain using a dialysis
bag technique at 37 jC [98]. The data revealed a
sustained release and could be fitted to a Weibull
distribution (t1/2 = 23.1 h).
Heiati et al. have studied the in vitro release of 3Vazido-3V-deoxythymidine palmitate (AZT-P) from trilaurin SLN using a bulk-equilibrium reverse dialysis
sac technique at 37 jC [99]. The observed initial burst
is attributed to partial AZT-P localisation in phospholipid micelles. Further, a dependence of release profile
on the type of phospholipid could be shown, i.e.
phospholipids with phase transition temperatures
(PTT) below 37 jC lead to fast release, PTT >37
jC represented a stronger diffusional barrier causing
slower release.
Wang et al. have prepared an optimised SLN
formulation containing 3V,5V-dioctanoyl-5-fluoro-2Vdeoxyuridine (DO-FUdR) for enhanced brain targeting [79]. They studied the in vitro release by a bulkequilibrium reverse dialysis sac technique in at 37 jC
compared to DO-FUdR and also FUdR reference
solutions. Hundred parcent drug were released from
the FUdR solution after less than 2 h. After 4 h,

1265

approximately 80% drug were released from DOFUdR solution and only 60% from the SLN formulation. The SLN release profile was biphasic, the initial
burst release was followed by a prolonged release
(80% drug released after 48 h). This can be correlated
to the drug-enriched shell model. The drug present in
the shell is released fast, followed by gradually release
from the lipid core.
Hu et al. have studied in vitro release kinetics of
clobetasol propionate from SLN prepared by solvent
emulsification-diffusion [76]. The lyophilised product
was dispersed in aqueous dissolution medium (PEG
400 solution containing Tween 80) without dividing
membranes. The authors observed a biphasic release
profile following Higuchi (45% release after 3 h,
followed by 5.9% release per day for 4 days). However, the chosen release model is not quite suitable for
lipophilic drugs. Therefore, these data have to be
judged critically.
For detailed information of the in vitro release of
the mentioned drugs from SLN, the reader is kindly
referred to the corresponding references. Concluding,
it can be stated that depending on the formulation,
sustained in vitro release can be achieved for various
drugs that are of interest for the parenteral application.

3. Biological activity and biopharmaceutical

aspects
3.1. General aspects
Because of their small size, SLN may be injected
intravenously and used to target drugs to particular
organs. The particles, as with all intravenously
injected and colloidal particulates, are cleared from
the circulation by the liver and spleen. To facilitate
drug targeting, in tumour tissue for example, a reticuloendothelial system avoidance (stealth) facility may
be incorporated using polyoxyethylene as described
well for classical polymeric nanoparticels in the past.
This may be achieved using block polyoxyethylene
polypropylene copolymers like Pluronic F188 in
which the hydrophobic portion of the molecule forms
the nanoparticle matrix while the water soluble polyoxyethylene block forms a hydrophilic coating on the
particle Stealth SLN increase the tumour accumulation [104], antibacterial activity [106] of antiparasitic

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S.A. Wissing et al. / Advanced Drug Delivery Reviews 56 (2004) 12571272

and antifungal drugs and allow brain delivery of

anticancer drugs not capable of crossing the blood
brain barrier (BBB) [102]. Besides of new attractive
drug delivery strategies we have also addressed biopharmaceutical aspects regarding safety and toxicity
of the drug delivery system relevant for patients.
3.2. Administration and drug liberation
SLN are generally injected either intravenously,
intramuscularly or subcutaneously or to the target
organ. Because of their minimum size below 1 Am,
SLN formulations can be used for systemic body
distribution with a minimised risk of blood clotting
and aggregation leading to embolism. Also SLN
provide a sustained release depot of the drug when
administered subcutaneously or accumulated in the
MPS. Incorporated drug is gradually released on
erosion (e.g. degradation by enzymes) or by diffusion
from the particles. The rate of release may be controlled by the nature of the lipid material [108], particle
size [109], and choice of surfactant [107,109,110] and
also by inner structure of SLN as discussed above. The
particle size of intravenously administered drug must
be below 5 Am to avoid blocking of fine capillaries
leading to embolism. By production techniques described before, the size is mostly below this value.
Special problems regarding blood clotting and aggregation under high electrolyte conditions in vivo are
discussed in Section 3.3.2.
3.2.1. Pharmacokinetic profile
Administration of drugs incorporated in SLN versus free drugs leads to different pharmacokinetic
profiles as well described for doxorubicin [101,102]
and paclitaxel [103]. Using SLN as carriers mostly a
three to 5-fold enhancement of plasma peaks is
observed. Interestingly plasma formulations for doxorubicin SLN showed a bi-exponential curve [101,102]
with high AUC, a lower rate of clearance, and a
smaller volume of distribution in comparison to the
free drug. The biphasic behaviour is explained probably due to the slow distribution of doxorubicin in
SLN. SLN formulations reduced cardiotoxic side
effects of doxorubicin in Wistar rats and prolonged
the metabolisation of doxorubicin to doxorubicinol.
Uptake of the SLN in cells of the MPS differed
from the size and composition of the particles. Uptake

of SLN can be avoided by PEG-ylation leading to

long circulating particles, so-called Stealth SLN. Several authors have worked on Stealth SLN, but finally
only a few reports on the in vivo behaviour have been
published yet. Interestingly, stealth doxorubicin solid
lipid nanoparticels showed similar circulating time
and pharmacokinetic behaviour in comparison to
unmodified nanoparticels [102]. Main reasons might
be explained by similar low surface hydrophobicity of
both types of particles avoiding adsorption of blood
proteins mediating liver uptake. The modified tissue
distribution of doxorubicin in SLN was related to a
slower distribution and passage of particles through
biological barriers and slow drug release from the
lipid matrix into the blood [49]. This biopharmaceutical behaviour may explain lower drug blood concentrations and reduction of severe side effects.
3.3. Safety aspects
3.3.1. Toxicity and status of excipients
Since today no SLN product for parenteral use is
on the market, but intensive studies for SLN in
different bioassays have been published recently
[49,50]. Most studies have been conducted with
glycerides composed of fatty acids, which are mostly
accepted as safe. Good tolerability depends in the first
line of the used surfactant and secondarily on the
lipid. To formulate parental SLN, surfactant of GRAS
status must be employed, e.g. lecithin, Tween 80,
Poloxamer 188, Span 85, and sodium glycocholate.
When performing bolus injections into mice good
tolerability was found for most of these surfactants
coating SLN. As described, for cetyl palmitate SLN
with different surfactants no acute toxicity, and no
increase in liver and spleen weight was observed
[111]. After autopsy and histopathology no significant
evidence was documented that SLN were acute toxic
to tested animals.
3.3.2. Blood clotting and interaction with serum
proteins
Cellular binding of SLN formulations, with emphasis on erythrocytes, is important because it affects
not only blood clotting, embolic effects but also may
change pharmacokinetic behaviour. SLN have distinct
affinities to red blood cells depending on the surfactant used. By Flow Cytometry, Olbrich et al. showed

S.A. Wissing et al. / Advanced Drug Delivery Reviews 56 (2004) 12571272

that SLN formulation consisting of Compritol as

matrix material and Tween 80 and Poloxamer 188
showed no binding to erythrocytes (SLN binding
< 10.0%) [112]. In contrast, when Span 85 was used,
blood cell affinity of labelled SLN was increased
leading to significant aggregation of red blood cells
(75.3%).
The interaction of SLN with the major circulatory
protein, serum albumin, has been investigated recently. By PCS and AFM albumin adsorption on the
particle surface formed a capping layer of 17 nm
increasing the size of tested particle populations only
slightly (final particle size: 150, 183, 193 nm, respectively) [113]. AFM imaging revealed that the SLNs
are protected by this layer against flattening on
surfaces. At physiological albumin concentrations
(35 50 g/l) the increased size was not important
enough to explain blood cell aggregation [113].

1267

trations of 20 ppm accepted by the German Federal

Institute for Drugs (BfArM).
Since Cremophor EL was recognised as allergic
principle in paclitaxel emulsions for parenteral use
[117], the use surfactants for i.v. administration is
under critical debate. Today tested SLN were proved
to be not toxic in in vivo and in vitro systems. Muller
et al. showed in in vivo test that test animals did not
suffer from anaphylatic shock or related symptoms
[111]. Detailed investigations regarding the immunostimulation and release of interleukins from activated
macrophages showed no significant interleukin concentrations (IL 6, IL 12, TNF, IFN) causing proinflammatory reactions [110,115]. Even the absence of
IL 12, that is mostly associated with severe toxic
effects in vivo, indicates an acceptable non acute toxic
profile of SLN [110,115].
3.4. Tissue distribution and drug targeting

3.3.3. Allergic reactions

SLN has been used as drug delivery systems as
discussed here intensively, but also drug unloaded
as Vaccine adjuvants. Focussing on drug delivery
systems mainly, we want to describe SLN as
vaccine adjuvants briefly. For further detailed
reviews, we refer to contributions by Muller et al.
[114,115]. Shortly, antigens adsorbed on the surface
or entrapped in the matrix of SLN induce an
enhanced immunological response. SLN showed to
be equivalent to Freud incomplete adjuvants (FIA)
protecting against different infections in chicken
and sheep. Decreasing particle size and increasing
hydrophobicity improves the adjuvancy of these
particles [114]. The particles offer a prolonged
and controlled presentation of the antigen to the
immune system.
Immunologic and finally allergic reactions are not
desired for SLN as drug delivery systems. Immunologic reactions can be affected by the production
process or by the lipids and additives used in the
formulation. Applying HPH and producing with a
LAB 40 homogeniser made of V2A steel may lead
to a final product that contains traces of nickel with a
risk of allergic reaction. Krause et al. showed by
atomic absorption spectroscopy that no iron as the
dominant metal in steel was detectable in traces in
nanosuspension formulations, e.g. Fe < 1 ppm [116].
All detectable metals were distinctly below concen-

The accumulation of SLN within the Kupffer

cells of the liver is predominantly found after
intravenously injection in case non-stealth SLN
are injected. With the exception of liver diseases
like hepatic neoplasms, liver infections like hepatitis
and visceral leishmaniasis and systemic candidasis,
and physiologic disorders (e.g. hypercholesterinemia), passive liver targeting should be avoided.
The systemic use of colloidal carriers is limited
by the presence of MPS, and it is consequently
necessary to avoid such recognition. SLN carriers
are mostly recognised by macrophages due to the
physicochemical characteristics in particle size, surface charge and surface hydrophobicity as discussed
before [108,109]. Various attempts have been made
to achieve long circulation times by avoiding MPS
uptake as discussed for Stealth SLN before. One
outstanding example of targeting specific organs is
brain delivery. The uptake of SLN by the brain
might be explained by adsorption of blood proteins
like apolipoproteins on particle surfaces mediating
the adherence to endothelial cells of the BBB. This
effect was well described for the trypanocidal drug
diminazene formulated as LDC. Diminazen aceturate does not cross the BBB because of its hydrophilicity. Formulation as LDC increased crossing of
the BBB and reducing CNS infection of Trypanosoma brucei infected mice [45].

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S.A. Wissing et al. / Advanced Drug Delivery Reviews 56 (2004) 12571272

4. Outlook
Most of the discussed drugs belong to classical
low molecular weight drugs. In the future we can
expect an increasing number of therapeutic molecules like proteins [62], oligonucleotides [118] and
DNA as vaccine or as drug for gene therapy [119].
Because of their physical and chemical instability in
the GIT, these drugs must be administered parenterally. With the exception of gene therapy, up to today
only a few publications have reported about protein
or nucleotide formulation with SLN [62,105]. But, as
SLN are a new and innovative therapeutic delivery
system, we can expect in the future an increasing
number of contributions describing delivery of recombinant proteins.
In contrast to recombinant proteins, first studies
have been published about gene delivery we want to
discuss here briefly. This platform technologycalled
TransoplexRhas been developed as an alternative
gene delivery system for lipoplexes. Olbrich et al.
showed their efficacy in transfection of COS-1 cells
with the beta Galactosidase gene [119]. Transoplex
were produced in a mixture of a cationic lipids like
cetylpyridinium chloride, benzalkonium chloride or
cetrimide and a Compritol or cetyl palmitate as helper
lipid. Main advantages of the technology are easy
production, higher physical stability, lower cytotoxicity and increased transfection in comparison to
commonly used liposome formulations like [119].
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