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ELSEVI ER Advanced Drug Delivery Reviews 16 (1995) 61-73

advanced
drug delivery
reviews
Microspheres and nanoparticles used in ocular delivery
systems
Andreas Zimmer, J&g Kreuter*
I nstitute of Pharmaceutical Technology, Biocenter, J ohann Wolfgang Goethe University.
Marie-Curie-Strasse 9, D-60439 Frankfurt am Main. Germany
(Received 2 March 1995: accepted 16 March 1995)
Abstract
Microparticles and nanoparticles are colloidal drug carriers in the micro- and submicron range. These systems
were developed to overcome solubility problems of poorly soluble drugs as well as for long acting injectable depot
formulations and specific drug targeting options. These carriers were also evaluated for ophthalmic drug delivery
purposes over the past 15 years. The main objective for these systems was to improve the classical aqueous eye drop
formulations which have major disadvantages like a rapid elimination of the drugs from the precorneal area.
Consequently. colloidal carrier suspensions were designed to combine ophthalmic prolonged action with the ease of
the application of liquid eye drops. The review summarises the present manufacturing methods and the materials
used for these delivery systems with respect to ophthalmic purposes. The distribution and penetration pathways of
the particulate delivery systems are also described. The applications of drug-loaded particles are presented with
focus on ophthalmic diseases like glaucoma, inflammations or infections of the eye. Accordingly, particulate carriers
outlined in the review include systems loaded with pilocarpine, beta-blockers, hydrocortisone, amikacin, and
miscellaneous drugs. Finally, the biodegradation and toxicity of the carrier materials under evaluation are
reviewed.
Keywords: Eye: Drug delivery; Nanoparticle; Nanosphere; Microparticles; Microsphere; Glaucoma; Pilocarpine;
Beta-blocker; Hydrocortisone
Contents
Abstract ..................................................
1. Introduction ............................................
2. Manufacturing methods .................................
3. Distribution and penetration of particles in the eye ........
4. Applications of drug-loaded particles .....................
5. Biodegradation and toxicity .............................
6. Conclusions ............................................
References ................................................
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* Corresponding author.
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0169-409X/95/$29.00 @ 1995 Elsevier Science B.V. All rights reserved
SSDI 0169-409X(95)00017-8
62 A. Zinmer. J. Kreurer / Advanced Drug Ddiverv Reviews 16 (iYY5,51 61- 7.?
1. Introduction
One of the main problems in ophthalmic drug
delivery, the rapid elimination of conventional
liquid eye drops from the eye, still remains
unsolved. A number of factors, namely rapid tear
turnover and the resulting precorneal loss, induc-
tion of tear flow due to irritation caused by the
drug preparation, as well as the relatively large
volume of the administered eye drop (-50 ~1
versus 7 ~1 of cornea1 tear film), lead to a high
rate of lacrimal drainage. Due to the resulting
elimination rate. the precorneal half life of drugs
applied by these pharmaceutical formulations is
considered to be between about 1-3 min. As a
consequence, only the very small amount of
about l-3% of the dosage actually penetrates
through the cornea and is able to reach intraocu-
lar tissues [1,2,3]. The poor productive absorp-
tion, on the other hand, results in a high amount
of drug that is drained into the nose or into the
gut. Especially the nose but also the gut are very
efficient absorption organs of the body. This in
turn leads to an extensive systemic absorption
and may result in unwanted side effects and
toxicity of the drug [3]. Although these problems
have been recognised for a long time, surprising-
ly little effort has been made by drug companies
to improve the situation, and only very few
alternative ocular drug delivery systems are on
the market.
One possibility for such systems is the employ-
ment of small particles. These colloidal particles
have the advantage that they may be applied in
liquid form just like eye drop solutions. Thus
they avoid the discomfort that is combined with
the application of viscous or sticky preparations
such as ointments. The latter preparations lead
to a total blurring of vision if they are properly
utilised. Large inserts, on the other hand. are
difficult to administer or if they are designed as
non-dissolving inserts they are even more dif-
ficult to remove, especially by elderly patients.
Another alternative to small particle systems are
liposomes. Liposomes. however. may have the
disadvantage of lower stability.
Suitable small particle systems include micro-
spheres and microcapsules as well as nanoparti-
cles and nanocapsules. Microparticles and mi-
crocapsules have a particle size above 1 ,um.
Microspheres are monolithic particles possessing
a porous or solid polymer matrix, whereas mi-
crocapsules consist of a polymeric membrane
surrounding a solid or liquid drug reservoir.
Nanoparticles have a particle size in the nanome-
ter size range below 1 pm [4]. The definitions
[4,S] for these particles include nanospheres as
well as nanocapsules.
In micro- or in nanocapsules the drug can be
present in a liquid form, it can be dissolved or
suspended in a liquid or solid core or it may be
present in the core itself. Monolithic micro- or
nanoparticles contain the drug either dissolved in
the polymer matrix in form of a solid solution or
suspended in form of a solid dispersion. Alter-
natively the drug may be adsorbed to the particle
surface.
It is important to note that the particle size for
ophthalmic applications should not exceed
10 pm because with larger sizes a scratching
feeling might occur. Therefore, a reduction in
particle size improves the patient comfort during
administration.
2. Manufacturing methods
A large number of production methods exist
for microspheres, microcapsules and nanoparti-
cles. The optimal method that may be used for
the preparation depends mainly on the drug to
be used. Since the description and the discussion
of the various production methods would by far
exceed the space allocated to the present review,
the reader who is interested in further details is
referred to various books and reviews that exten-
sively describe these topics [3-91.
A high incorporation ratio, in combination
with a considerable amount of free drug in order
to obtain an adequate initial drug level, as well as
a sustained drug release during the residence
time of the carrier systems in the precorneal area
are required for ophthalmic use. So far, various
synthetic and natural biocompatible polymers
have been used in order to manufacture micro-
spheres for ocular drug delivery (Table 1). Prep-
aration techniques using natural biopolymers like
albumin are summarised in Table 2.
A. Zimmer, .I. Kreuter I Advanced Drug Delivery Reviews 16 (1995) 61-73
Table 1
Nano- and microparticulate carriers in ophthalmology
63
Drug Polymer Reference
Amikacin
Betaxolol
Carte0101
Chloramphenicol
Hydrocortisone
Indomethacin
Pilocarpine
Progesterone
Timolol
Poly(butyl)cyanoacrylate
Poly(epsilon)caprolacton
Poly( isobutyl)cyanoacrylate
Polylactic-co-glycolic acid
Poly(epsilon)caprolacton
Polylactic acid
Albumin
Poly(epsilon)caprolacton
Albumin
Cellulose-acetate-hydrogen-
phthalate
Gelatine
Poly(butyl)cyanoacrylate
Poly( hexyl)cyanoacrylate
Polylactic acid
Poly(methyl)methacrylate-
acrylic-acid copolymer
Poly(methyl)methacrylate
Polyamide
Polyphthalamide
Poly(hexyl)cyanoacrylate
Poly( butyl)cyanoacrylate
Poly(alkyl)cyanoacrylate
Alonso et al.
Maincent et al.
Shell et al.
Zimmer et al.
Marchal-Heussler et al.
Masson et al.
Marchal-Heussler et al.
Leucuta et al.
Diepold et al.
Marchal-Heussler et al.
Zimmer et al.
Boye et al.
Gurny et al.
Leucuta et al.
Harmia et al.
Diepold et al.
Kreuter
Zimmer et al.
Harmia et al.
Kreuter et al.
Vidmar et al.
Andermann et al.
i14j
]I01
[I51
[I61
]Il,l2]
1171
[I81
[l2,13]
[I91
[20,21,22]
1121
[231
[24,25,26]
[W
[27,28,29.30]
[311
1321
[331
[2&30]
[331
[341
[35,36]
Blumenthal et al.
[371
Duzman et al.
[381
Klein et al.
[391
Mazor et al. [40,41]
Robinson et al.
[421
Stodtmeister et al.
[431
Ticho et al. L44.451
Harmia et al. (28.301
Evans et al.
[461
Beal et al.
I471
Kreuter et al.
[481
Li et al.
[491
Harmia et al.
[501
Table 2
Manufacturing techniques used for preparing micro- and nanospheres
Manufacturing process
Denaturation or cross-linking of macromolecules
in emulsion form
Interfacial polymerisation
Formation in an aerosol phase
Reference
Burgess et al.
Kim et al.
Senyei et al.
Due-Mauger et al.
Fickat et al.
Przyborowski et al.
[511
[521
I531
]541
]551
[561
Desolvation
Aggregation by pH adjustment and heat treatment
Formation of nanoparticles via microemulsions
Marty and Oppenheim [57]
Oppenheim et al.
1581
Zimmer et al.
]2ll
Taplin et al.
]591
Dechow
I601
64 A. Zimmer, J. Kreuter I Advanced Drug Delivery Reviews 16 (199.5) 61-73
Nanoparticles for ophthalmic drug delivery
prepared from synthetic polymers (i.e. poly-
(alkyl)cyanoacrylates) have been mainly pro-
duced by emulsion polymerisation. In this pro-
cess, a poorly soluble monomer is dissolved in
the continuous phase. This continuous phase can
be aqueous or organic. Additional monomer may
be emulsified in emulsion droplets that are stabil-
ised by emulsifiers. The polymerisation is started
by chemical initiation, pH shift, or by irradiation
with gamma-rays, ultraviolet (UV) or visible
light. The location of the polymerisation is in the
continuous phase where dissolved monomer
molecules react with each other and grow until
particle formation by phase separation occurs.
Additional monomer molecules diffuse to the
resulting growing polymer particles and maintain
the polymerisation. The emulsion droplets main-
ly act as monomer reservoirs. In later stages, the
emulsifiers stabilise the resulting polymer par-
ticles. It has to be noted, that the nanoparticles
are not formed by one single polymer molecule
but consist of a large number of macromolecules
[S]. Nanocapsules with an oily core were pre-
pared mainly by the addition of benzylbenzoate
or mineral oils. In this case, the polymerisation of
the poly(alkyl)cyanoacrylates (i.e. poly( iso-
butyl)cyanoacrylate) is located at the interface
between the oily and the aqueous phase resulting
in a small capsule covered by a polymer mem-
brane [61].
As shown in Table 1, a number of studies deal
with the preparation and application of ophthal-
mic drugs loaded to micro- and nanospheres.
Pilocarpine is still among one of the most im-
portant drugs for ocular delivery and glaucoma
therapy. Also, beta-blockers like timolol and
more recently betaxolol were investigated as
potent candidates for particulate ophthalmic
drug delivery systems. In most cases, the drug is
present during the polymerisation- or particle
manufacturing process leading to an incorpora-
tion of the drug in form of a solid solution or
solid dispersion. Alternatively, a drug can be
adsorbed onto the particle surface after the
manufacturing process. In the case of hydro-
cortisone it was shown that a very high loading
capacity can be reached. Similar results were
observed for other steroids like progesterone.
3. Distribution and penetration of particles in
the eye
First attempts to address the problem of pre-
cornea1 elimination of particles were made by
Sieg and Tripplett [62]. In this study, ‘41Ce-la-
belled polystyrene microspheres of 3 and 25 pm
diameter were used. With the smaller particles,
the elimination rate in rabbits was dependent on
the instilled volume into the eye, i.e., 25 and
50 ~1, the larger volume being eliminated about
twice as fast as the smaller volume. For both
dose volumes a very rapid elimination phase was
observed for the first 3 min with an elimination
of 25% or 55%, respectively, of the dose. This
rapid elimination phase was followed by a much
slower phase. Consequently, the total elimination
after 10 min was measured to amount to 35% or
65%, respectively. No significant elimination was
observable for the larger 25 pm particles over 20
min at both dosages (25 or 50 ,ul).
In another study with 30 pm sized poly-
phthalamide microcapsules containing encapsu-
lated yyTechnetium-labelled albumin, Beal et al.
[47] observed that 60% of the instilled dose was
still present after 90 min, whereas in an aqueous
control solution of albumin, 90% of the dose
cleared within 10 min.
The elimination and ophthalmic distribution of
[ “C]poly( hexyl)cyanoacrylate nanoparticles in
rabbits was extensively studied by Wood et al.
[63]. The observed elimination kinetic from the
tear film was not first order but was linear after
plotting the log of the concentration over the
square root of time. From these data an approxi-
mate elimination half-life of 20 min was esti-
mated [64]. A shorter ocular residence time of
about 10 min was observed with ” ‘Indium-ox-
ine-labelled poly( butyl)cyanoacrylate nanoparti-
cles [65]. This shorter half-like, however, may be
to some degree attributable to the release of the
label into the tear-fluid [64].
A more detailed study was the comparative
investigation of the distribution of [‘“Cl poly-
(hexyl)cyanoacrylate nanoparticles between
healthy and inflamed eyes by Diepold et al. [66].
The highest portion of the instilled nanoparticle
dose was observed with 7% in conjunctival tissue
after 10 min. A rapid decrease followed, and
A. Zimmer, J. Kreuter I Advanced Drug Delivery Reviews 16 (1995) 61-73 65
about 1% of the applied dosage remained in
ocular tissues for about 4 h. Interestingly, in
inflamed eyes the tissue concentrations were
about three to five times higher than in normal
eyes with the exception of the conjunctiva at
early time points (Fig. 1). The observed con-
siderably higher association of the targeted drug
carriers to inflamed tissues renders these par-
ticles very promising as drug carriers for steroids
to inflamed regions of the eye.
Additionally it was found [16] that in all ocular
tissues the level of [3H] hydrocortisone was
higher in inflamed than in healthy eyes due to an
increased cell permeability as a result of the
inflammatory process. The application of
[“Hlhydrocortisone-loaded albumin nanoparti-
cles led to lower hydrocortisone tissue concen-
trations than with a reference solution due to
i&
Time Iminl
Humor
kk-
Fig. 1. Concentration of [‘4C]poly(hexyl)cyanoacrylate nano-
particles in healthy (open symbols) or inflamed (closed
symbols) rabbit eyes. Reprinted from Ref. [3] with permis-
sion of the copyright holder, M. Dekker, Inc., New York.
strong binding of hydrocortisone to a particle
system intended for slow release. An exception
occurred with the reference hydrocortisone solu-
tion in the conjunctiva, as less drug was found in
the inflamed than in the normal ‘tissue, since
enhanced lacrimation led to increased drug
drainage. In contrast, the corresponding
nanoparticle preparation was more efficiently
retained at the inflamed than at the normal
conjunctiva. Consequently, in the inflamed eye,
nanoparticles enabled hydrocortisone targeting
to the precorneal area away from the inner
segments of the eye.
In another study the penetration of fluorescent
labelled nanoparticles in rabbit eyes was investi-
gated [67,68]. Correspondingly to the data ob-
served with radioactively labelled particles, it was
found that the particles mainly remained in the
precorneal area. Conjunctiva as well as cornea
tissue were examined by laser-confocal micro-
scopy and showed an intracellular uptake of
particles into the first epithelium cell layers. An
uptake via endocytosis, lysis of cell walls by
particle degradation products or self-permeation
enhancing effects of the particles were discussed
in these references. However, a complete per-
meation of particles through the cornea into the
aqueous humor as well as a penetration through
intercellular junctions was not observable. There-
fore, minor radioactivity found in the aqueous
humor by Diepold et al. [66] can be ascribed to
the degradation products of the nanoparticles.
Corresponding results were observed by Calvo
et al. [69]. In this study the previously mentioned
results, demonstrating the cornea1 uptake of
nanoparticles were confirmed with poly(ep-
silon)caprolacton nanocapsules, leading to an
assumption that a general uptake mechanism for
small particles, probably based on phagocytosis,
exists in percorneal tissues. These results are also
in correlation with earlier observations made by
Kaye et al. [70]. This research group evaluated
the transport of colloidal TiO, particles. It was
found that particles were associated only with the
outermost layer of the epithelium and also were
visualised in some large vacuoles. No particles
were detected in the intercellular space, and no
transport of colloidal tracer into deeper layers of
the epithelium was noticed.
66 A. Zimmer. J. Kreuter I Advanced Drug Delivery Reviews 16 (1995) 61-73
4. Applications of drug-loaded particles
The most applications of drug-loaded ophthal-
mic delivery systems are for glaucoma therapy,
especially cholinergic agonists like pilocarpine.
Among the first particulate systems with pilocar-
pine that were developed was a cellulose acetate
hydrogen phthalate (CAP) pseudolatex formula-
tion introduced by Gurny [24]. In this formula-
tion, the pH of the nanoparticulate pseudolatex
was maintained at 4.5 during the storage. After
instillation into the eye, the tear fluids rapidly
shifted the pH of the applied liquid to 7.2,
causing a dissolution of the polymeric particles.
The resulting polymer solution had a very high
viscosity that prevented a rapid wash-out of the
formulation and the action of pilocarpine was
considerably prolonged. As a result, both the
miosis fmax and the area under the curve (AUC)
of the miosis versus time curve were increased by
50%, compared to an aqueous reference solu-
tion. This pharmacological enhancement was
mainly due to the strong increase of the viscosity
and was not caused by a slow release of the
nanoparticles.
The pseudolatex preparation was considerably
more effective in rabbits than ophthalmic rods or
a thermosetting gel [71]. Compared to the pseu-
dolatex system, a further enhancement was ob-
served with a bioadhesive formulation based on a
solution of 0.125% hyaluronic acid. This prepara-
tion showed a more pronounced decrease of drug
elimination of pilocarpine which was also con-
firmed by elimination kinetics analysed with
“Tc-1abelled sulphur colloids.
In another study by Vidmar et al. [34], poly-
(lactic acid) microcapsules containing micronised
pilocarpine hydrochloride were prepared by a
modified solvent deposition evaporation process.
Microcapsules of a size between 63 and 100 pm
were obtained after sieving. Freshly prepared
microcapsules containing pilocarpine prolonged
the miosis .time in rabbits from 2 h with a
pilocarpine solution up to 4 h. However, in-
stability of the preparation and leakage of the
encapsulated drug was responsible for the lack of
response after 3 days of storage.
Binding of pilocarpine to poly( butyl)-
cyanoacrylate (PBCA) nanoparticles by adsorp-
tion [29] or by partial incorporation (-15%) [31]
enhanced the miotic response by about 22% [29]
or 33% [31]. The intraocular pressure (IOP)-
lowering effect was investigated in three rabbit
models and its prolongation was even more
pronounced than the miotic response. These
models, water loading model, alpha chymotryp-
sin model, and betamethasone model, created an
artificial increase of the IOP but were used
because pilocarpine has little effect on the IOP in
normostatic eyes. The results of this study con-
cerning the duration of the IOP lowering effect
and the relative AUCs are shown in Table 3.
Diepold et al. [31] concluded that the betametha-
sone model most closely resembles the course of
the IOP curve in humans.
Both studies, Harmia et al. [29] and Diepold et
al. [31], are in agreement with the results of
Zimmer et al. [33]. In this study with pilocarpine-
loaded PBCA nanoparticles the aqueous humor
pilocarpine pharmacokinetics as well as the phar-
macodynamic response were determined as IOP-
lowering effects and as miosis (Fig. 2). Formula-
tions with different drug concentrations (2-6%)
as well as different particle concentrations were
investigated. Pilocarpine binding to the particles
was achieved at a level of lo-18% of the total
drug content. All preparations were applied to
rabbit eyes with enhanced IOP by pretreatment
with betamethasone prior to pilocarpine usage.
The results of the pilocarpine pharmacokinetics
showed a 23% increase of the aqueous humor
pilocarpine concentrations (AUC) for nanoparti-
cle suspensions compared to the aqueous refer-
ence solutions. Additionally, t, ,2 was prolonged
Table 3
Duration of activity and relative AUC (area under the curve
activity vs. time curve) of pilocarpine (2%) in three different
rabbit models for the determination of the intraocular pres-
sure
Model Duration Relative AUC
S N S N
Betamethasone 5 >9 1 3.35
Alpha-chymotrypsine >lO >lO I 1.28
Water loading >9 >9 1 1.42
Pilocarpine nitrate was instilled into the eyes in the form of
an eye drop solution (S) or loaded to nanoparticles (N). Data
from Ref. 1311.
A. Zimmer, J. Kreuter I Advanced Drug Delivery Reviews 16 (199-f) 61-73 67
8
7
z 6
i
5
a 4
8
Q 3
2
1
0
n 246NP/c Suspension
Y ’ I ’ I ’ I ’ I ’ I ’
0 30 60 90 120 150 180 [min]
I’I’I’l’l’I’I’I’I’l’l~1
0 3oT 60 90 120 150 180 210 240 270 300 330 [mill]
T ’ I
1
I ’ I ’
0 2 4 6
8 [hl
Time
Fig. 2. Pilocarpine aqueous humor levels, miotic response,
expressed as A pupillary diameter, and A intraocular pressure
(TOP) of 2% pilocarpine nitrate applied as aqueous reference
solution, poly(butyl)cyanoacrylate nanoparticle suspension
containing 1% particles (NP), respectively 8% particles (NPI
c). (Values statistically significantly different from the refer-
ence solution: *P < 0.05, **P < 0.01.)
and the elimination coefficient was significantly
decreased. The t,,, values of aqueous humor
concentration were observed to be in a similar
time range as the miosis t,,, readings. It was
found that at lower drug levels a more pro-
nounced prolongation of miosis was achieved
with nanoparticles in comparison to the standard
solution. Additionally, the IOP-reduction was
highly significantly prolonged with nanoparticle
preparations. The maximum reduction was ob-
tained with the reference solution after 1-2
hours, whereas with nanoparticles it was reached
after about 2-3 hours. Differences between
nanoparticle suspensions and aqueous solutions
again especially occurred at lower drug concen-
trations.
In a recent study (Zimmer et al. [21]) the
loading of pilocarpine was optimised employing
albumin nano- and microspheres. Optimal load-
ing conditions were obtained with 2% pilocar-
pine and a particle concentration of 20 to 40 mg
particles/ml. Due to their high surface area
nanoparticles exhibited the highest loading. The
in vitro release of the albumin particle carrier
systems differing in size was measured by
simulating the rapid drug dilution as occurring in
the eye. All particle fractions released pilocar-
pine completely within a few minutes. The sys-
tems were further tested in vivo by determi-
nation of their miotic and IOP-lowering prop-
erties. Pilocarpine-loaded albumin nano- and
microparticles increased the pharmacodynamic
response of pilocarpine by about 50-90%
(miosis), and .50-70% (IOP) respectively, com-
pared to a pilocarpine reference solution. The
best results were obtained with pilocarpine-
loaded albumin nanoparticles (2% drug content)
at a particle concentration of about 40 mg/ml.
A possibility to increase the efficacy of pilocar-
pine-loaded particle systems is the approach to
coat them with bioadhesive or viscous polymers
[22]. In this study different polymers, solutions,
and coated albumin nanoparticles possessing pre-
defined comparable viscosities were investigated.
The polymers methylcellulose, polyvinylalcohol
and hydroxypropylmethylcellulose possessed
mainly viscosity enhancing properties, whereas
hyaluronic acid, mucin, sodiumcarboxymethylcel-
lulose and Carbopol@941 were chosen because
of their bioadhesive properties. After coating
pilocarpine-loaded albumin nanoparticles with
various viscous but not bioadhesive polymers at
different concentrations, the particles and the
pure polymer solution yielded superior but not
additive effects in comparison to the pilocarpine
68 A. Zimmer. J. Kreuter I Advanced Drug Delivery Reviews 16 (199-f) 61-73
reference solution. However, with the bioadhe-
sive polymers an additional improvement in
miotic response and IOP reduction compared to
both, the pure nanoparticle suspension and the
pure polymeric vehicles was observed after coat-
ing of the particles. The bioadhesive polymer
mucin displayed the best effects concerning im-
provements in miotic response and in IOP reduc-
tion (see Fig. 3). Obviously the particle-coating
with the latter polymers led to an improved
bioadhesion. The coated particles seem to adhere
to conjunctival mucin thus prolonging the resi-
dence time of the drug-loaded particles in the
precorneal area.
The shown pharmacological differences con-
cerning miosis and IOP reduction between the
albumin nanoparticles [21,22] or the
cyanoacrylate nanoparticles [33] and the corre-
sponding reference solutions are especially sig-
nificant since it has to be taken into considera-
tion that only about 15-18% of pilocarpine is
0 60 120 180 240 Km 360 420
Time @in)
Fig 3. Miotic response, expressed as A pupillary diameter.
and A intraocular pressure (IOP) of 2% pilocarpine nitrate
applied as mucin reference solution containing 2.5% mucin
and mucin coated albumin nanoparticles (20 mgiml). *,
values statistically significantly different from the baseline; + ~
statistically different from the reference solution.
bound to the particles, whereas about 82-85%
free drug is present in solution. It seems to be
likely that this unbound drug portion can favor-
ably act as an initial drug dose. Additionally, the
release time frame should correspond to the
residence time of the particles in the eye. Conse-
quently, a study by Li et al. [49] demonstrated
that a too efficient loading may reduce the
bioavailability of the drug. Li et al. [49] sorbed
progesterone to polycyanoacrylate nanoparticles,
yielding a solid solution with a polymer/water
distribution coefficient of more than 80000. As a
result, the drug was released too slowly and the
nanoparticles were eliminated from the eye be-
fore effective progesterone delivery could take
place.
Microspheres loaded with pilocarpine which
were used for glaucoma therapy were prepared
with gelatin and with albumin by Leucuta [18].
These carriers were manufactured by crosslink-
ing either with formaldehyde in the case of
gelatine or by heat denuaturation of albumin in
an oily phase. The particles obtained exhibited a
mean size of about 30 pm. Pilocarpine nitrate
incorporated in gelatin particles caused a 2.3-fold
increase of the miotic response (AUC) and a
3.3-fold increase in the case of albumin micro-
spheres.
As mentioned before, anti-inflammatory
glucocorticoids delivered to the eye with particu-
late systems included [3H]hydrocortisone [16]
and [3H]labelled hydrocortisone-17-butyrate-21-
propionate ([3H]HBP). The [3H]HBP was in-
corporated into lipid microspheres [72]. The
concentrations in the cornea, anterior sclera, and
aqueous humor 1 h after instillation and in the
cornea 3 h after instillation were significantly
higher in the rabbits eyes treated with liquid
solutions than with a microsphere suspension.
Other drugs which were applied with particu-
late carrier systems were mainly beta-blockers
including betaxolol [11,12], carteolol [14], and
metipranolol [73]. Betaxolol chlorhydrate was
adsorbed to poly( isobutyl)cyanoacrylate nano-
particles of a size of about 240 nm by Marchal-
Heussler et al. [11,12]. An S-shaped sorption
isotherm was obtained from the drug loading
data. Additionally, drug release by dialysis and
the reduction of the IOP in alpha-chymotrypsin
A. Zitnmer, J. Kreuter I Advanced Drug Delivery Reviews 16 (1995) 61-73 69
pretreated rabbits were determined. Betaxolol
loading was achieved between 25 and 45%
(drug/polymer). An optimal prolonged release
compared to a reference solution of about 45%
as well as a most significantly decreased IOP
were observed by a preparation containing 25%
drug bound to the particles using 1% dextran
70 000 as a polymerisation stabiliser. However,
the difference to a commercial eyedrop solution
(Betoptic@) was not statistically significant.
A significant improvement of the betaxolol
response compared to the Betoptic eyedrops was
achieved after adsorption of the drug to poly-
(epsilon)caprolactone nanocapsules [12] manu-
factured by the method of Al Khouri et al. [61].
While the 0.5% commercial betaxolol solution
(Betoptic) achieved a maximal reduction of the
IOP in alpha-chymotrypsin pretreated rabbits of
about 20% and approached baseline after about
8 h, betaxolol at the same concentration encapsu-
lated into the poly(epsilon)caprolactone
nanocapsules yielded about a 30% IOP reduction
and a much longer pharmacodynamic response.
After 8 h, the IOP reduction was still about 25%
below baseline values. Even a 0.1% betaxolol
nanocapsule preparation with this polymer still
was significantly better than the commercial
0.5% betaxolol solution. Moreover, the systemic
side effects of betaxolol were reduced by en-
capsulation into the nanocapsules. Betoptic in-
hibited the isoprenalin induced increase in heart
frequence by about 50%, whereas no significant
difference from the control was observed with
the same amount of encapsulated drug [12].
Similar results were observed with Carteolol
[14]. Again poly(epsilon)caprolactone nanocap-
sules loaded with this beta-blocker increased the
therapeutic effect demonstrated by a prolonged
IOP decrease over 8 h. Systemic side effects on
cardiac rate and arterial pressure were signifi-
cantly reduced in comparison to commercial eye
drop preparations.
Encapsulation of another beta-blocker, meti-
pranolol, into a number of nanocapsules consist-
ing of different polymers and different nanocap-
sule oil core materials did not lead to a signifi-
cant change in the reduction of the IOP in
rabbits in comparison to commercial eyedrops
containing the same amount of metipranolol
(Betamann@) [73]. However, again cardiovascu-
lar side effects determined by heart rate mea-
surements were drastically reduced by loading
metipranolol to nanocapsules.
Binding of amikacin sulphate by adsorption to
previously polymerised and resuspended freeze-
dried empty poly( butyl)cyanoacrylate nanopar-
titles significantly improved the delivery of this
antiinfective drug to the cornea and into the
aqueous humor of rabbits [lo]. The efficacy of
delivery to these ocular compartments depended
on the polymerisation stabiliser used. 1% dex-
tran 70 000 in the polymerisation mixture yielded
nanoparticles that increased the concentrations
of amikacin in the cornea about two-fold and in
the aqueous humor about three-fold. On the
other hand, the combination of dextran with
sodium lauryl sulphate or the use of poloxamer
188 as stabiliser did not significantly increase the
cornea1 uptake or aqueous humor amikacin
levels compared to the aqueous reference solu-
tion of the drug [lo].
5. Biodegradation and toxicity
The materials most extensively studied as
particulate carrier materials for the ophthalmic
drug delivery are poly(alkyl)cyanoacrylate and
albumin. Poly( butyl)cyanoacrylate has been used
as an artificial tissue and bone glue for over 20
years [74]. It is well tolerated and significantly
less histotoxic than other comparable tissue glues
[75]. This material can be degraded following
two different metabolisation pathways. One
possibility is the erosion of the polymer back-
bone under formation of formaldehyde [76,77].
The second pathway is the cleavage of the ester
resulting in a water soluble polymer backbone
and a corresponding alcohol [78]. Due to the
presence of enzymes in vivo this pathway is much
faster than the first one [79] and poly-
(butyl)cyanoacrylate nanoparticles are biode-
gradable within a few hours.
The acute and subacute toxicity of poly-
(butyl)cyanoacrylate nanoparticles were tested
by Couvreur et al. [80]. A low toxicity was
demonstrated and the LD,, was determined with
500 mg/kg after an i.v. application. In the sub-
70 A. Zimmer. J. Kreuter I Advanced Drug Delivery Reviews 16 (1YY.5) 61-73
acute toxicity tests, no significant effects on
either the histological pattern of the tissues
studied, the blood parameters. or on the body
weight were observable after i.v. injection of
poly(isobutyl)- and of poly(hexyl)cyanoacrylate
nanoparticles [64,80]. No mutagenicity was ob-
served for these nanoparticles and their degra-
dation products when analysed with the Ames
test [80].
The ocular application of poly(isobutyl)- and
of poly( hexyl)cyanoacrylate nanoparticles was
well tolerated in rabbits. No reddening of the
eyes or other adverse effects were observed even
after multiple dosing. The treatment of human
volunteers with empty poly( butyl)cyanoacrylate
nanoparticles in saline did not cause any adverse
reaction [64].
Albumin nanoparticles as well as albumin
microspheres also are biodegradable and well
tolerated particle systems. Particles of this ma-
terial were applied i.v. in various forms in nu-
clear medicine and for ocular purposes without
causing any adverse effects [21]. The biodegrada-
tion of albumin in vivo is dependent on the
enzymatic activity. In vitro human serum al-
bumin nanoparticles and microparticles of a
mean size of 1.5 pm and a maximal size of about
5 pm degrade from their centre
after
phagocytosis by human macrophages [Sl] within
a few days. So far it is unknown if this degra-
dation pathway also is possible after ocular
application. Possible allergic effects are expected
to be reduced due to the cross-linking process
during the preparation and by using human
serum albumin instead of bovine serum albumin.
Other types of nanoparticles may degrade
somewhat differently. Polylactic acid micro-
spheres, for instance, degrade from their centre
by hydrolysis [82]. In this case, of course, hy-
drolysis results in the erosion of the polyester
backbone. It is unknown whether nanoparticles
made from polylactic acid also degrade from the
center. It is important to note that this material
seems to be one of the most biocompatible.
biodegradable, and non toxic materials used for
preparing nano- and microparticles. However,
when considering toxicity it has to be kept in
mind that ocular application of particles is some-
what different from systemic application and that
most of the particles are eliminated from the eye
via the tear flow followed by a gut elimination
pathway with a possible gut absorption.
6. Conclusions
Nanoparticles and microspheres represent
promising drug carriers for ophthalmic applica-
tions. The binding of drugs depends on the
physicochemical properties of the drugs as well
as of the nano- and microparticle material and
also on the manufacturing process for these
particles. After optimal drug binding to these
particles, the ocular bioavailability of a number
of drugs is significantly enhanced in comparison
to normal aqueous eye drop solutions.
Generally, smaller particles are better toler-
ated by the patients than larger particles. For this
reason especially nanoparticles may be preferred
for long-acting ocular drug delivery systems,
although larger microparticles showed slower
elimination kinetics from the precorneal com-
partment.
In order to be of commercial use drug-loaded
particle systems have to be further developed by
optimising the drug payload and by controlling
the release rate. In addition, nanoparticles were
shown to adhere preferentially to inflamed pre-
cornea1 tissues of the eye, which seems to be that
drug targeting to inflamed parts of the eye might
be a promising application for commercial
formulations.
Furthermore, particles retention and, there-
fore, ocular bioavailability of drugs can be en-
hanced by a coating of nanoparticles with non-
irritating polymers, tissue antibodies, or by creat-
ing systems which coagulate in the conjunctival
sac due to electrostatic interactions or pH
changes.
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