Synthesis of poly(ethylene glycol)/polypeptide/poly(D, L -lactide) copolymers and their nanoparticles

Article abstract of DOI:10.1002/pola.24720

Core-shell structured nanoparticles of poly(ethylene glycol) (PEG)/polypeptide/poly(D,L-lactide) (PLA) copolymers were prepared and their properties were investigated. The copolymers had a poly(L-serine) or poly(L-phenylalanine) block as a linker between a hydrophilic PEG and a hydrophobic PLA unit. They formed core-shell structured nanoparticles, where the polypeptide block resided at the interface between a hydrophilic PEG shell and a hydrophobic PLA core. In the synthesis, poly(ethylene glycol)-b-poly(L-serine) (PEG-PSER) was prepared by ring opening polymerization of N-carboxyanhydride of O-(tert-butyl)-L-serine and subsequent removal of tert-butyl groups. Poly(ethylene glycol)-b-poly(L-phenylalanine) (PEG-PPA) was obtained by ring opening polymerization of N-carboxyanhydride of L-phenylalanine. Methoxy-poly(ethylene glycol)-amine with a MW of 5000 was used as an initiator for both polymerizations. The polymerization of D,L-lactide by initiation with PEG-PSER and PEG-PPA produced a comb-like copolymer, poly(ethylene glycol)-b-[poly(L-serine)-g-poly(D,L-lactide)] (PEG-PSER-PLA) and a linear copolymer, poly(ethylene glycol)-b-poly(L-phenylalanine)-b-poly(D,L-lactide) (PEG-PPA-PLA), respectively. The nanoparticles obtained from PEG-PPA-PLA showed a negative zeta potential value of -16.6 mV, while those of PEG-PSER-PLA exhibited a positive value of about 19.3 mV. In pH 7.0 phosphate buffer solution at 36 °C, the nanoparticles of PEG/polypeptide/PLA copolymers showed much better stability than those of a linear PEG-PLA copolymer having a comparable molecular weight.

Full text of DOI:10.1002/pola.24720

Synthesis of Poly(ethylene glycol)/Polypeptide/Poly(D,L-lactide)
Copolymers and Their Nanoparticles
HYUNPYO LEE, JUN BEUM PARK, JI YOUNG CHANG
Department of Materials Science and Engineering, College of Engineering, Seoul National University, Seoul 151-744, Korea
Received 11 December 2010; accepted 9 April 2011
DOI: 10.1002/pola.24720
Published online 10 May 2011 in Wiley Online Library (wileyonlinelibrary.com).
ABSTRACT: Core-shell structured nanoparticles of poly(ethylene
glycol) (PEG)/polypeptide/poly(D,L-lactide) (PLA) copolymers
were prepared and their properties were investigated. The
copolymers had a poly(L-serine) or poly(L-phenylalanine) block
as a linker between a hydrophilic PEG and a hydrophobic PLA
unit. They formed core-shell structured nanoparticles, where
tiation with PEG-PSER and PEG-PPA produced a comb-like
copolymer, poly(ethylene glycol)-b-[poly(L-serine)-g-poly(D,L-
lactide)] (PEG-PSER-PLA) and a linear copolymer, poly(ethyl-
ene glycol)-b-poly(L-phenylalanine)-b-poly(D,L-lactide) (PEG-
PPA-PLA), respectively. The nanoparticles obtained from
PEG-PPA-PLA showed
a negative zeta potential value of
the polypeptide block resided at the interface between
a
ꢀ16.6 mV, while those of PEG-PSER-PLA exhibited a positive
value of about 19.3 mV. In pH 7.0 phosphate buffer solution at
36 ꢁC, the nanoparticles of PEG/polypeptide/PLA copolymers
showed much better stability than those of a linear PEG-PLA
hydrophilic PEG shell and a hydrophobic PLA core. In the syn-
thesis, poly(ethylene glycol)-b-poly(L-serine) (PEG-PSER) was
prepared by ring opening polymerization of N-carboxyanhy-
dride of O-(tert-butyl)-L-serine and subsequent removal of tert-
butyl groups. Poly(ethylene glycol)-b-poly(L-phenylalanine)
(PEG-PPA) was obtained by ring opening polymerization of N-
carboxyanhydride of L-phenylalanine. Methoxy-poly(ethylene
glycol)-amine with a MW of 5000 was used as an initiator for
both polymerizations. The polymerization of D,L-lactide by ini-
C
V
copolymer having a comparable molecular weight.
2011
Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 49:
2859–2865, 2011
KEYWORDS: block copolymers; nanoparticles; biodegradable;
drug delivery systems
INTRODUCTION Core-shell-type nanoparticles with modified
surface by flexible hydrophilic chains have been extensively
studied as drug carriers for targeted delivery with long-cir-
culation time in blood after intravenous administration.1–8
The hydrophilic surface properties of the nanoparticles
increase the circulation time by reducing the adsorption of
plasma proteins and the recognition of phagocytes, and
thereby influence the distribution pattern in the organs. Pol-
ylactide (PLA) is widely used to form a core of the core-
shell-type nanoparticle.8–11 As a flexible hydrophilic chain,
poly(ethylene glycol) (PEG) is outstanding.12,13 The polymer
shows essentially good water solubility to have high degree
of hydration. When PEG chains are tethered densely at the
surface of the hydrophobic nanoparticle, the chains stretch
out from the surface to form a brush conformation. The
hydrophilicity and brush conformation of PEG chains prevent
adsorption of hydrophobic proteins to the nanoparticles via
steric repulsion.
ior. There have been a number of efforts to modify physical
properties of PLA by controlling the structural architecture.
In particular, PLAs with branched structures such as star-
shaped, comb-shaped, and hyperbranched polymers attracted
considerable attention.16–18 The physical stability of the nano-
particles is primarily dependent on the molecular weight of a
PLA block. As the size of the PLA block increases, the hydro-
phobic core becomes more solid-like and stable. Several stud-
ies demonstrated that the stability of the nanoparticles could
be further improved by chemical or physical crosslinking.19,20
In this study, we report the preparation of PEG/polypeptide/
poly(D,L-lactide) (PLA) copolymers and their nanoparticles.
The copolymers had a comb-like or a linear structure, where a
hydrophilic PEG was linked to a hydrophobic PLA unit through
a poly(L-serine) or a poly(L-phenylalanine) block. In their
nanoparticles, the polypeptide blocks were expected to reside
at the interface between a hydrophilic PEG shell and a hydro-
phobic PLA core and to form intermolecular hydrogen bonds.
Hydrophobic drugs are loaded in the PEG-PLA nanoparticles
through noncovalent interactions and released by a diffusion
mechanism.11,14,15 Hence, it is evident that the physicochemi-
cal properties of the interfacial region and the core of the
nanoparticle has a great influence on the drug release behav-
EXPERIMENTAL
Materials and Instruments
Triphosgene (98%), poly(vinyl alcohol) (MW ¼ 31,000–
50,000, 87–89% hydrolyzed), O-(tert-butyl)-L-serine, L-
Correspondence to: J. Y. Chang (E-mail: jichang@snu.ac.kr)
C
V
Journal of Polymer Science Part A: Polymer Chemistry, Vol. 49, 2859–2865 (2011) 2011 Wiley Periodicals, Inc.
SYNTHESIS OF PEG/POLYPEPTIDE/PLA COPOLYMERS, LEE, PARK, AND CHANG
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JOURNAL OF POLYMER SCIENCE PART A: POLYMER CHEMISTRY DOI 10.1002/POLA
phenylalanine, and tin(II) 2-ethyl hexanoate were purchased
from Aldrich and used as received. Methoxy-poly(ethylene
glycol)-amine (MW ¼ 5000) and D,L-lactide (3,6-dimethyl-
1,4-dioxane-2,5-dione) were purchased from SunBio and TCI,
respectively, and used as received. Tetrahydrofuran and tolu-
ene were dried over sodium metal and distilled. Other rea-
gent-grade solvents were used as received. 1H NMR spectra
were recorded on a Bruker Avance-300 spectrometer. 13C
NMR spectra were recorded on a Bruker Avance-500 spec-
trometer. Elemental analysis was performed using EA1110
(CE Instrument, Italy). FTIR measurements were recorded
on a PERKIN ELMER Spectrum GX I using KBr pellets. The
particle sizes and zeta potentials of the nanoparticles were
measured using an ELS-Z (Otsuka Electronics, Japan) spec-
trometer. A typical result was obtained based on the average
from 5 runs. SEM images were taken by a JEOL JSM6330F
microscope. UV–vis spectra were obtained with a SCINCO S-
3100 spectrometer.
mL) and further purified by reprecipitation of the polymer
solution in dichloromethane into diethyl ether.
1
Yield, 0.855 g. H NMR (DMSO-d6, 300 MHz): d ¼ 7.99 (NH),
4.0–4.5 (CH and CH2 of polyserine), 3.50 (OCH2CH2).
Synthesis of Poly(ethylene glycol)-b-[poly
(L-serine)-g-poly(D,L-lactide)]
Poly(ethylene glycol)-poly(L-serine) block copolymer (0.20 g)
and D,L-lactide (1.22 g) were dissolved in dry toluene (75
mL). To remove moisture in the monomer and the macroini-
tiator, the solution was further dried by azeotropic distilla-
tion for 1 day. The solution (50 mL) and tin(II) 2-ethylhexa-
noate (0.5 wt %) were put into a flame-dried polymerization
tube (100 mL). After three freeze-thaw cycles, the tube was
sealed under vacuum condition. The polymerization was car-
ꢁ
ried out at 110 C for 24 h. After evaporation of the solvent,
the solid residue was dissolved in dichloromethane. The
polymer was isolated by precipitation into an excess amount
of diethyl ether and further purified by reprecipitation from
the polymer solution in dichloromethane into diethyl ether
twice.
Synthesis of O-(tert-Butyl)-L-serine N-Carboxyanhydride
A solution of triphosgene (3.31 g, 11.2 mmol) in THF (10
mL) was added to a suspension of O-(tert-butyl)-L-serine
Yield, 0.23 g. 1H NMR (DMSO-d6, 300 MHz): d ¼ 8.20 (NH),
5.19 (CH of polylactide), 4.0–4.8 (CH and CH2 of polyserine),
3.50 (OCH2CH2), 1.46 (CH3 of polylactide); FTIR (KBr, cmꢀ1):
3300 (NAH), 2930–2870 (CAH), 1760 (C¼¼O), 1630 (amide
I), 1550 (amide II), 1190 (CAO).
ꢁ
(1.50 g, 9.31 mmol) in THF (20 mL) at 50 C, and the result-
ing solution was stirred for 2 h at the same temperature.21
After evaporation of the solvent, the crude product was iso-
lated by precipitation in n-hexane. The product was purified
by recrystallization from THF/n-hexane [yield: 1.33
g
(76.0%)].
Synthesis of Poly(ethylene glycol)-b-Poly(D,L-lactide)
The linear poly(ethylene glycol)-b-Poly(D,L-lactide) (PEG-PLA)
diblock copolymer was synthesized by ring opening polymer-
ization of D,L-lactide using methoxy-poly(ethylene glycol)-OH
(MW ¼ 5000) as an initiator according to the literatures.4,22
Anal. Cald for C8H13NO4: C, 51.33; H, 7.00; N, 7.48; found: C,
51.36; H, 6.99; N, 7.44; 13C NMR (CDCl3, 125 MHz): d ¼
168.07, 152.68, 74.67, 60.98, 58.97, 27.44; 1H NMR (DMSO-
d6, 300 MHz): d ¼ 8.94 (s, NH, 1H), 4.56 (t, CH, 1H), 3.63–
3.46 (m, CH2, 2H), 1.09 (s, ACCH3, 9H); FTIR (KBr, cmꢀ1):
3311 (NAH), 2976 (CAH), 1851 (C¼¼O), 1787 (C¼¼O).
Synthesis of L-Phenylalanine N-Carboxyanhydride
This compound was prepared in the same manner as the
preparation of O-(tert-butyl)-L-serine N-carboxyanhydride.
From L-phenylalanine (2.0 g, 12.1 mmol) and triphosgene
(4.65 g, 15.7 mmol), the product (1.63 g, 8.52 mmol) was
obtained in 70.4% yield.
Synthesis of Poly(ethylene glycol)-b-poly
[O-(tert-butyl)-L-serine] Diblock Copolymer
To a solution of O-(tert-butyl)-L-serine N-carboxyanhydride
(1.33 g, 7.10 mmol) in DMF (30.0 mL), a solution of
methoxy-poly(ethylene glycol)-amine (1.41 g, 0.28 mmol,
MW ¼ 5000) was added in DMF (20.0 mL). The reaction
Anal. Cald for C10H9NO3: C, 62.82; H, 4.74; N, 7.33; found: C,
62.85; H, 4.74; N, 7.29; 13C NMR (CDCl3, 125 MHz): d ¼
168.92, 152.21, 134.05, 129.41, 128.19, 59.04, 37.96; 1H
NMR (DMSO-d6, 300 MHz): d ¼ 9.07 (s, NH, 1H), 7.17–7.35
(m, AC6H5, 5H), 4.78 (t, CH, 1H), 3.01 (d, CH2, 2H); FTIR
(KBr, cmꢀ1): 3301 (NAH), 3030 (CAH from phenyl), 2923
(CAH), 1836 (C¼¼O), 1777 (C¼¼O).
ꢁ
mixture was stirred for 2 days at 40 C. After evaporation of
the solvent, the solid was dissolved in chloroform (40.0 mL).
The polymer was isolated by precipitation into diethyl ether
(500 mL) and further purified by reprecipitation from the
polymer solution in chloroform into diethyl ether twice.
Yield, 0.87 g. 1H NMR (DMSO-d6, 300 MHz): d ¼ 7.82 (NH),
4.0–4.5 (CH and CH2 of polyserine), 3.50 (OCH2CH2), 1.10
(CCH3).
Synthesis of Poly(ethylene glycol)-b-poly(L-
phenylalanine)
To a solution of L-phenylalanine N-carboxyanhydride (0.4 g,
0.68 mmol) in DMF (10.0 mL), a solution of methoxy-poly(-
ethylene glycol)-amine (0.2 g, 0.04 mmol, MW ¼ 5000) was
added in DMF (10.0 mL). The reaction mixture was stirred
Deprotection of tert-Butyl Group
To a solution of poly(ethylene glycol)-poly[O-(tert-butyl)-L-
serine] (0.87 g, 0.15 mmol) in dichloromethane (50 mL), a
solution of hydrogen bromide in acetic acid (33 wt %, 0.26
g) was added slowly. The reaction mixture was refluxed for
24 h. After concentration by evaporation of the solvent, the
polymer was isolated by precipitation into diethyl ether (500
ꢁ
for 3 days at 50 C. After evaporation of the solvent, the resi-
due was dissolved in dichloromethane (10.0 mL). The poly-
mer was isolated by precipitation into diethyl ether (400
mL) and further purified by reprecipitation from the poly-
mer solution in dichloromethane into diethyl ether twice.
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ARTICLE
SCHEME 1 Synthesis of poly(ethylene glycol)-b-[poly(L-serine)-g-poly(D,L-lactide)] (PEG-PSER-PLA).
Yield, 0.16 g. 1H NMR (DMSO-d6, 300 MHz): d ¼ 8.14 (NH),
measured by dynamic light scattering (DLS) at particular
time intervals.
7.15 (C6H5), 4.51 (CH), 3.50 (OCH2CH2), 2.90 (CH2).
Synthesis of Poly(ethylene glycol)-b-poly(L-
phenylalanine)-b-poly(D,L-lactide)
1-Aminopyrene-Loaded Nanoparticles
From a solution of 1-amionopyrene (8 mg) and a block co-
polymer (40 mg) in dichloromethane (2 mL), 1-aminopyr-
ene-loaded nanoparticles were prepared as described above.
Loading efficiency of the nanoparticles was determined as
follows: 1-Aminopyrene loaded nanoparticles (5 mg) were
dissolved in DMF (2 mL). A concentration of 1-aminopyrene
in the DMF solution was calculated by measuring the absorb-
ance at 370 nm with a UV–vis spectrometer. The molar
extinction coefficient at 370 nm was 15,000 Mꢀ1 cmꢀ1. The
loading efficiency was calculated using the following equa-
tion.
This polymer was prepared in the same manner as the prep-
aration of poly(ethylene glycol)-b-[poly(L-serine)-g-poly(D,L-
lactide)] (PEG-PSER-PLA). From poly(ethylene glycol)-b-
poly(L-phenylalanine) (PEG-PPA) (0.20 g) and D,L-lactide (0.9
g), the copolymer (0.24 g) was obtained.
1H NMR (DMSO-d6, 300 MHz): d ¼ 8.22 (NH), 7.14 (C6H5),
5.19 (CH of polylactide), 4.50 (CH of polyphenylalanine),
3.50 (OCH2CH2), 2.91 (CH2), 1.47 (CH3); FTIR (KBr, cmꢀ1):
3289 (NAH), 2996–2887 (CAH), 1758 (C¼¼O), 1634 (amide
I), 1548 (amide II), 1114 (CAO).
Loading efficiency
Fabrication of Nanoparticles
Nanoparticles were prepared by an emulsion-solvent evapo-
ration method.23 Typically, a block copolymer (40 mg) was
dissolved in dichloromethane (2 mL). The solution was
dropped into deionized water (20 mL) containing 0.7% (w/
v) poly(vinyl alcohol) with stirring under 600 rpm. After
stirring for 9 h, the nanoparticles were isolated by centrifu-
gation at 5000 rpm for 15 min and washed twice with
deionized water. The nanoparticles were then resuspended
in deionized water (5 mL), filtered with a 0.80-lm pore size
filter, and freeze-dried.
Weight of 1 ꢀ aminopyrene in nanoparticles
¼
ꢂ 100
Weight of nanoparticles
Release of 1-Aminopyrene from the Nanoparticles
A suspension of 1-amonopyrene-loaded nanoparticles (4 mg)
in 1 mL of pH 3 or pH 7 phosphate buffer solution was put
in a dialysis tube (cutoff Mn ¼ 3500) and then placed in a
glass vial filled with 15 mL of theꢁphosphate buffer solution.
At particular time intervals at 36 C, the concentration of 1-
aminopyrene in the glass vial was calculated by measuring
the absorbance at 370 nm with a UV–vis spectrometer, and
the phosphate buffer solution was exchanged with a fresh
one.
Stability Test of Nanoparticles
The nanoparticles (1 mg) were suspended in pH 7.0 phos-
phate buffer solution (5 mL) at 36 ꢁC. The particle size was
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JOURNAL OF POLYMER SCIENCE PART A: POLYMER CHEMISTRY DOI 10.1002/POLA
SCHEME 2 Synthesis of poly(ethylene glycol)-b-poly(L-phenylalanine)-b-poly(D,L-lactide) (PEG-PPA-PLA).
RESULTS AND DISCISSION
Poly(ethylene glycol)-b-poly(L-phenylalanine)-b-poly(D,L-lac-
tide) (PEG-PPA-PLA) was synthesized according to Scheme 2.
N-carboxyanhydride of L-phenylalanine was polymerized
using CH3O-PEG-NH2 with a MW of 5000 as an initiator to
give poly(ethylene glycol)-b-poly(L-phenylalanine) (PEG-PPA).
Synthesis of Copolymers
Scheme 1 shows the synthetic route of PEG-PSER-PLA. Poly-
(ethylene glycol)-b-poly(L-serine) (PEG-PSER) was prepared
by ring-opening polymerization of N-carboxyanhydride of O-
PEG-PPA had a primary amino group at the end of the poly-
(tert-butyl)-L-serine using methoxy-poly(ethylene glycol)-
mer chain, which was used for the initiation of the polymer-
amine (CH3O-PEG-NH2) with a MW of 5000 as an initiator. In
1
ization of D,L-lactide.24 In the H NMR spectrum, the peak for
the 1H NMR spectrum, methylene protons of PEG and tert-
phenyl protons of poly(L-phenylalanine) appeared at 7.15
butyl protons of polyserine appeared at 3.50 and 1.10 ppm,
respectively. tert-Butyl groups were removed with hydrogen
bromide to generate hydroxyl groups on the polymer back-
ppm. The average degrees of polymerization for PPA and
PLA blocks were 22 and 119, respectively, which were esti-
mated by 1H NMR spectroscopy. The molecular weights and
bone, which was confirmed by 1H NMR spectroscopy. The
compositions of the copolymers are summarized in Table 1.
hydroxyl groups were used for the initiation of the ring-
opening polymerization of D,L-lactide in the presence of
Preparation of Nanoparticles
tin(II) 2-ethylhexanoate as a catalyst to give PEG-PSER-PLA
The nanoparticles of the copolymers were prepared by an
having a comb-like structure. In the 1H NMR spectrum, the
emulsion-solvent evaporation method.9,23 A solution of a
peaks corresponding to methine and methyl protons of the
block copolymer in dichloromethane was dropped into
PLA segment appeared around 5.19 and 1.46 ppm,
deionized water containing poly(vinyl alcohol) with stirring.
respectively.
The average diameters of the nanoparticles measured by
The number of PLA chains and its average degree of poly-
merization were estimated to be 7 and 10, respectively,
DLS, were 208 6 34 nm (PEG-PSER-PLA) and 210 6 96 nm
(PEG-PPA-PLA) (Table 2). The average diameter of the nano-
particles obtained from linear PEG-PLA was 128 6 91 nm. It
is noticeable that PEG-PSER-PLA produced larger particles
than PEG-PLA, although weight fractions of PLA chains of
these two copolymers were comparable. We presume that
the comb-like structure of PEG-PSER-PLA and the rigidity of
the polyserine block prevented the close packing of hydro-
phobic PLA chains during the formation of nanoparticles.
Figure 1 shows a SEM image of the nanoparticles obtained
from PEG-PSER-PLA. The freeze-dried nanoparticles had a
spherical shape and were readily dispersed in water.
1
based on the H NMR spectrum. The peak from PEG (MW ¼
5000) at 3.50 ppm was used as an internal standard. For the
comparison, PEG-PLA was also synthesized by ring-opening
polymerization of D,L-lactide using MeO-PEG-OH with a MW
1
of 5000 as an initiator. The Mn determined by H NMR spec-
troscopy was 8900.
TABLE 1 Molecular Weights and Compositions of the
Copolymers
TABLE 2 Sizes and Zeta Potentials of the Nanoparticles
Weight Fraction
Copolymer
Mn
PEG
Polypeptide
Polylactide
Polymers
Size (nm)
Zeta Potential (mV)
PEG-PSER-PLA
PEG-PPA-PLA
PEG-PLA
10,200
16,800
8,900
0.49
0.30
0.56
0.05
0.20
–
0.46
0.50
0.44
PEG-PSER-PLA
PEG-PPA-PLA
PEG-PLA
208 6 34
210 6 96
128 6 91
19.3 6 0.37
ꢀ16.6 6 0.45
ꢀ8.3 6 0.75
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ARTICLE
FIGURE 2 Schematic drawing of the nanoparticle structure.
tide block, we expected that the polypeptide block should re-
side at the interface between a hydrophilic PEG shell and a
hydrophobic PLA core. The core-shell structures of the nano-
particles were examined by zeta potential measurement. It
has been reported that PLA nanoparticles have a negative
zeta potential value of below ꢀ20 mV at pH 7 because of the
terminal carboxyl group.25 The absolute value decreases as
the surface is coated by hydrophilic PEG chains in the core-
shell structure of PEG-PLA. The nanoparticles obtained from
PEG-PLA and PEG-PPA-PLA showed negative zeta potential
values, exhibiting ꢀ8.3 and ꢀ16.6 mV, respectively (Table 2).
FIGURE 1 FE-SEM image of freeze-dried nanoparticles obtained
from PEG-PSER-PLA.
Figure 2 shows a schematic drawing of the nanoparticle.
Because the hydrophilic PEG chain of PEG-PSER-PLA or PEG-
PPA-PLA was linked to the hydrophobic PLA through a pep-
FIGURE 3 Size changes of the nanoparticles as a function of time in a pH 7.0 phosphate buffer solution at 36 ꢁC (a) and diameter
profiles of the nanoparticles measured after 15 days (b).
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JOURNAL OF POLYMER SCIENCE PART A: POLYMER CHEMISTRY DOI 10.1002/POLA
below 30% in pH 7 buffer solution. This result is attribut-
able to the fact that the polypeptide chains at the surface of
the hydrophobic core as well as 1-aminopyrene were proto-
nated under acidic conditions.
CONCLUSIONS
We demonstrated the preparation of core-shell structured
nanoparticles having polypeptide interlayers from the comb-
like copolymer, PEG-PSER-PLA and the linear copolymer,
PEG-PPA-PLA. Their nanoparticles showed better stability
than those of a linear PEG-PLA copolymer having a compara-
ble molecular weight. PEG-PSER-PLA had a positive zeta
potential value, differently from the nanoparticles obtained
from linear PEG-PLA, suggesting that the polypeptide chains
formed an interface layer between a hydrophilic PEG corona
and a hydrophobic PLA core. There have been numerous
reports about the overexpression of proteases in tumor
cells.28,29 If the polypetptide chains in the interface layer
contained a specific amino acid residue, the overexpressed
proteases could facilitate the degradation of nanoparticles on
accumulation in tumor cells. Our future studies will explore
this possibility.
FIGURE 4 Release profiles of 1-aminopyrene from PEG-PSER-
PLA nanoparticles in pH 3 (ꢃ) and pH 7 (~) phosphate buffer
solutions.
On the contrary, the nanoparticles of PEG-PSER-PLA had a
positive value of about 19.3 mV. As only a few neutral or-
ganic polymers such as polyethylenimine exhibit positive
zeta potentials,26 this result strongly indicates that the sur-
face of the nanoparticles was covered with polypeptide
chains.
This study was supported by a grant of the Korea Health 21
R&D Project, Ministry of Health and Welfare, Republic of Korea
(Project No. A062254).
REFERENCES AND NOTES
The stability of the nanoparticles was examined by meas-
uring changes of the nanoparticle diameters in pH 7 phos-
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27
ꢁ
phate buffer solution at 36 C by DLS. Figure 3 shows the
stability test results. Nanoparticles of PEG-PLA degraded in 1
day. The relatively fast degradation of these nanoparticles is
attributable to a low molecular weight of the PLA block. The
nanoparticles of PEG-PPA-PLA and PEG-PSER-PLA showed
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PEG-PPA-PLA and PEG-PSER-PLA were stable until 5 and 12
days, respectively. In the diameter profiles measured after 15
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showed the multimodal shapes, suggesting that they became
degraded and then aggregated into larger particles. Nanopar-
ticles of PEG-PSER-PLA exhibited a more uniform size distri-
bution than the other two polymers [Fig. 3(b)].
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The release profile of the nanoparticles obtained from PEG-
PSER-PLA was studied by using 1-aminopyrene as a model
drug. The nanoparticles were prepared by an emulsion-sol-
vent evaporation method9,23 in the presence of 1-aminopyr-
ene. The loading efficiency was 6.7%, which was estimated
by measuring the concentration of 1-aminopyrene with a
UV–vis spectrometer after dissolving the nanoparticles in
DMF. The release test of 1-aminopyrene from the nanopar-
ticles was performed at 36 ꢁC in pH 3 and pH 7 phosphate
buffer solutions (Fig. 4). 1-Aminopyrene was more rapidly
released at pH 3 than at pH 7. After 24 h, more than 70% of
loaded 1-aminopyrene was released in pH 3 phosphate
buffer solution, whereas a cumulative release amount was
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