Journal of Controlled Release 61 (1999) 107–112

A method for the preparation of polymeric nanocapsules

without stabilizer
Ze Lu, Jianzhong Bei, Shenguo Wang*
PCLCC, Institute of Chemistry, The Chinese Academy of Sciences, Beijing 100080, China

Received 22 July 1998; received in revised form 25 November 1998; accepted 17 April 1999

Abstract

In this communication poly( L-lactide) (PLLA) nanocapsules containing bovine serum albumin (BSA) were prepared by
means of a modified W/ O / W double emulsion technology. A mixture of glycerin and water was used instead of the
traditional stabilizer system in the preparation of polymeric nanocapsules. The preliminary results showed that the high
viscosity of the mixture and the hydroxyl group of the glycerin were helpful to the formation of the nanocapsules. The
prepared nanocapsules had a similar spherical form. By comparison of different polymers of poly( L-lactide) and
polycaprolactone-poly(ethylene oxide) block copolymer (PCE), it was found that the entrapment efficiency of the BSA was
strongly dependent on the hydrophilicity of the polymer. A lower entrapment efficiency of BSA and nanocapsules with
smaller size were obtained when the relative hydrophilic PCE polymer was used as the entrapping material.  1999
Elsevier Science B.V. All rights reserved.

Keywords: Polymeric nanocapsules; W/ O / W emulsion; Bovine serum albumin; Poly( L-lactide); Polycaprolactone-poly(ethylene oxide)
block copolymer (PCE)

1. Introduction would produce toxic residues in the polymerization

Recently, the special properties and wide applica-
tions of nanoparticles have gradually gained under-
standing and research on polymeric nanocapsules as
drug delivery systems has been of interest to many
groups. Polymeric nanocapsules show significant
advantages over microcapsules [1,2]. Some inves-
tigations show that the poly(isobutylcyanoacrylate)
nanocapsules containing insulin prepared by an
interfacial polymerization are able to cross the
intestinal mucosa into the blood system by oral
administration [3]. However this preparation method
*Corresponding author.
process such as monomers, oligomers or reagent. At
the same time, cross-linked reaction between the
drug and entrapping materials in the polymerization
process may exist and the biological activity of the
drug would be destroyed. Therefore the potential use
of the nanocapsules produced is limited [4]. Fessi
etc. proposed another preparation method of
nanocapsules involving the interfacial deposition of
poly( D,L-lactide) (PDLLA) [5]. This method would
avoid the risk of toxic residues and cross-linked
reaction between the drug and the entrapping materi-
als. However, the method is only suitable for the
entrapment of lipophilic drugs [6]. On the other
hand, as is well known, the stabilizer plays the role

0168-3659 / 99 / $ – see front matter  1999 Elsevier Science B.V. All rights reserved.
PII: S0168-3659( 99 )00112-1
108 Z. Lu et al. / Journal of Controlled Release 61 (1999) 107 – 112
of dispersing the emulsion drops and preventing the
emulsion drops from coagulating and precipitating in
the emulsion system. This is one of the important
components for the preparation of micro- and
nanocapsules. The first stage in the preparation of
micro- and nanocapsules using the solvent evapora-
tion method is the formation of an emulsion. The
encapsulating material is dissolved in organic solvent
to form the oil phase. In this stage, small-molecular-
weight emulsifier is of help to the formation of
emulsion. The second stage, the evaporation of
solvent, is more important. With the evaporation of
organic solvent, the oil phase is made thicker. It is
very easy to coacervate when the collisions occur.
The small-molecular-weight emulsifier such as Span
80 and Tween 80 cannot effectively prevent the
coacervation of emulsion drops because of the low
viscosity of the water phase and the weak interaction
with the emulsion drops. So macromolecular stabi-
lizers are often used due to the high viscosity of their
aqueous solution and a stronger adsorption around
the emulsion drops. The traditional stabilizers for the
preparation of micro- and nanocapsules are usually a
type of macromolecular substance such as gelatin,
poly(vinyl alcohol) and so on. They are not easily
washed from the micro- and nanocapsules. As a
result they may cause trouble when purifying the
product and lead to reduced yield and entrapment
efficiency of the drug.
In this study, polymeric nanocapsules containing
bovine serum albumin (BSA) were prepared by
means of a modified W/ O / W double emulsion
technology without stabilizer. In this method a
mixture of glycerin and water was used instead of
the traditional stabilizer, poly( L-lactide) (PLLA) and
polycaprolactone-poly(ethylene oxide) block copoly-
mer (PCE) were used as the entrapping material. The
effect of glycerin and the properties of the polymer
on the formation of nanocapsules is also discussed.
2. Experimental
2.1. Materials
Poly( L-lactide) (PLA) (m.w. 16 000 and 51 000)
was synthesized by ring-opening polymerization of
L-lactide in our laboratory. Polycaprolactone-poly-
(ethylene oxide) block copolymer (PCE(60 / 40))
(m.w. 79 000, with 6000 of PEO block and molar
ratio of [CL] to [EG] was 60 to 40) was synthesized
according to the method previously outlined [7].
Bovine serum albumin (BSA) (Shanghai Biochemi-
cal Drug Factory) was used without further purifica-
tion. All other reagents were of analytical grade.
2.2. Molecular weight measurement
Molecular weight was determined by room-tem-
perature Gel Permeation Chromatography (GPC)
(Waters 510 HPLC Pump, Waters R401 Differential
Refractometer, Waters 746 Date Module and Shodex
columns). Chloroform was used as eluant with a
flow-rate of 1.0 ml / min. The molecular weight of the
polymer was calibrated by standard polystyrene
samples.
2.3. Preparation of polymeric nanocapsules
The PLA and PCE nanocapsules containing BSA
were prepared as follows. A mass of 25 mg BSA
was dissolved in distilled water, then the solution
was emulsified with methylene chloride solution of
500 mg PLA or PCE containing Span 80 by ul-
trasonication. The prepared water-in-oil emulsion
was then added into a mixture of glycerin and
distilled water (50 / 50 by volume) containing another
emulsifier Tween 80 and stirred at 10 000 rpm for 1
mm to form W/ O / W emulsion. After that, 200 ml of
distilled water was added into it, the system was
stirred for an additional 30 min. The produced
nanocapsules were collected by centrifugation. The
collected nanocapsules were washed two times with
distilled water, and freeze-dried. The final product
was stored in a desiccator below 08C.
2.4. Characterization of nanocapsules
The freeze-dried nanocapsules were dispersed in
distilled water containing Tween 80 and then air-
dried onto a metal stub. After the sample was coated
with gold, the nanocapsules were observed with a
Hitachi S-530 scanning electron microscope. The
size distribution of the nanocapsules was analyzed by
computer.
 Z. Lu et al. / Journal of Controlled Release 61 (1999) 107 – 112 109

2.5. Measurement of contact angle

Polymer was dissolved in chloroform, and then the

polymer solution was cast on a glass plate. After the
solvent was evaporated at room temperature, the
polymer film was dried completely in vacuum. The
contact angle of the film to water and the mixture of
glycerin and water were determined on the air
surface of the film by a CAD type contact-angle
meter (Kyowa Kaimenkagaku).

2.6. Determination of the entrapment of BSA

A mass of 10 mg of PLA (or PCE) nanocapsules

containing BSA was dissolved with tetrahydrofuran
(THF) and then centrifuged. After the supernatant
layer was poured out, the residues were dissolved in
the distilled water. The entrapped BSA content was
determined by the Folin–phenol method.

3. Results and discussion

PLA and PCE nanocapsules containing BSA were

prepared according to the method described above.
The morphology of prepared polymeric nanocapsules
containing BSA was identified by SEM technique. It
could be seen that PLA and PCE nanocapsules had a
similar spherical form (Fig. 1). In the preparation of
nanocapsules, the traditional stabilizer was omitted
and a mixture of glycerin and water was used. It was
considered that this glycerin–water mixture had a
high viscosity, hence the opportunity for the coalesc-
ence of the emulsion drops to decrease and as a
result, this was helpful to the formation of the
nanocapsules. In the meantime, the multi-hydroxyl
structure of glycerin could accelerate the diffusion of
the organic solvent from the nanodrops to the extra
water. Thus the solvent can evaporate immediately to
form the nanocapsules after the water is added. On
the other hand, in comparison with other methods
that used the traditional stabilizer, since the glycerin
is a small molecular substance, it could be easily
washed. In addition, the glycerin is not harmful; it is
also not necessary for it to be washed from the
nanocapsules. Therefore, this method would make Fig. 1. The SEM photographs of PLA and PCE nanocapsules
the purifying of the nanocapsules produced easier. containing BSA. (a) PLLA (m.w. 16 000), (b) PLLA (m.w.
The size distribution of the nanocapsules, the 51 000), (c) PCE (m.w. 79 000).
110 Z. Lu et al. / Journal of Controlled Release 61 (1999) 107 – 112

Scheme 1. Polycaprolactone-poly(ethylene oxide) block copoly-

mer.

seen that under the same preparation conditions,

Fig. 2. The particle size distribution profile of polymeric nanocap-
sules. (a) PCE nanocapsules, (b) PLA nanocapsules (m.w.
16 000).
mean size and the entrapment efficiency of BSA in
nanocapsules are shown in Fig. 2 and Table 1
respectively. Comparing the results of the two kinds
of entrapping material of PCE and PLA, it could be
there were many differences in the particle size,
particle size distribution of nanocapsules and the
entrapment efficiency of BSA in nanocapsules. When
PCE was used a the entrapping material, the size of
the prepared nanocapsules was smaller and the
particle size distribution was narrower in comparison
with those of the nanocapsules prepared with PLA.
The entrapment efficiency of BSA of PCE
nanocapsules was also much lower than that of PLA
nanocapsules. It revealed that the morphology of the
nanocapsules and the entrapment efficiency of the
drug strongly depended on the property of the
entrapping materials. Considering the difference in
the chemical structure of the two polymers, it was
found that the special structure of PCE played an
important role in the forming of nanocapsules. There
is a hydrophilic segment of PEG with a molecular
weight of 6000 in the middle of the PCE molecules
as shown in Scheme 1. The contact angle data
showed that the surface of PCE film is more
hydrophilic than that of PLA. It suggested that the
surface property of polymer film is greatly affected
by the introduction of the hydrophilic PEG segment.
In the formation of nanocapsules, the PEG segment
showed a tendency to enter the aqueous phase. Since
the PEG segment was long and soft enough to adjust
the conformation of the PCE by the segment motion,
this resulted in nanocapsules with smaller size being
obtained by the arrangement of the PEG segment. A
similar phenomenon of PEG segment arrangement

Table 1
Effect of the encapsulating polymer on the particle size and entrapment efficiency of BSA in nanocapsules
Materials Molecular Contact angle Particle size Entrapment
weight of film (degree) (nm) efficiency (%)
PLLA 16 000 a 70 5636195 51
PLLA 51 000 a 72 8906214 60
PCE 79 000 b 60 317681 29
a
Calculated by measurement of inherent viscosity.
b
Determined by GPC measurement.
 Z. Lu et al. / Journal of Controlled Release 61 (1999) 107 – 112
had also been observed for PCE films [8]. Based on
the same reason, the leakage of BSA to the outer
aqueous solution was increased with the arrangement
of the PEG segment and the entrapment efficiency of
BSA was very low. It was suggested that the size of
the nanocapsules and the entrapment efficiency of
BSA were strongly dependent on the hydrophilicity
of the polymer used. A lower entrapment efficiency
of drug and nanocapsules with smaller size would be
obtained in the case of using a relatively hydrophilic
polymer.
The morphology of the prepared nanocapsules was
also affected by the molecular weight of the polymer
used. When the concentration of the polymer solution
was identical, the solution of polymer with higher
molecular weight had a higher viscosity. Therefore
the size of the nanocapsules increased with increas-
ing molecular weight of the polymer. On the other
hand, it was more difficult for the BSA to pass
through the oil phase consisting of polymer with
higher molecular weight because of the high viscosi-
ty. The entrapment efficiency of BSA increased with
Fig. 3. The release profile of BSA from nanocapsules prepared with different molecular weight of PLLA in vitro. – d –; m.w. 16 000:
– j –; m.w. 51 000.
111
increasing molecular weight of the PLLA. So the
particle size and the entrapment efficiency of BSA
also depended on the molecular weight of the
polymer used.
The release profile of BSA from nanocapsules is
shown in Fig. 3. It was considered that the release
rate of BSA from PLLA nanocapsules was depen-
dent on the molecular weight of PLLA. For the
nanocapsules prepared with higher molecular weight
of PLLA, the release rate of BSA was lower than
that from nanocapsules prepared with lower molecu-
lar weight PLLA. However because the entrapment
efficiency of BSA increased with increasing molecu-
lar weight of PLLA, and the driving force of drug
release increased with increasing the drug entrap-
ment efficiency, the release rate of the two kinds of
nanocapsules with different PLLA was only different
in the first few hours, but later was nearly the same.
Therefore, the drug release rate of nanocapsules can
be controlled by adjusting the polymer type and
molecular weight of the polymer used, as well as the
entrapment efficiency of the drug.
112 Z. Lu et al. / Journal of Controlled Release 61 (1999) 107 – 112
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