Signal-Responsive Gating
by Polyelectrolyte Brush
on Nanoporous Membrane
by
Yoshihiro Ito
Graduate School of Material Science, NAIST
8916-5 Takayama-cho, Ikoma, 630-01, JAPAN
telephone: +81-743-72-5903, fax: +81-743-72-5903
[email protected]
This is a draft paper
for a talk at the
Fifth
Foresight Conference on Molecular Nanotechnology.
The final version has been submitted
for publication in the special Conference issue of Nanotechnology.
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Abstract
Poly(L-glutamic acid) carrying a terminal disulfide group at
one end of the chain was self-assembled on a porous membrane. The
rate of water permeation through the surface-modified porous
membrane depended on pH. In the region of low pH, poly(L-glutamic
acid) chain is protonated and folded to form  -helical
structure; in the region of high pH, it is de-protonated to have
extended random structure. The ionic strength also regulate the
rate of water permeation. Increasing ionic strength reduced the
pH dependence of permeation due to shielding effect. The
permeation through the porous membrane, the surface of which was
covered with the self-assembled polypeptide brush, was sensibly
regulated by changing pH because of direct contact of brush
chains with environment.
Introduction
A variety of nano- or micro-structured materials have been
prepared to apply for catalysis, separation, and host compound
for template synthesis of other nanoscopic materials. These
materials have found many potential applications in the areas of
device technology and drug delivery systems. We have synthesized
various signal-responsive polymers-grafted porous membranes to
regulate substance permeations (Ito et al, 1992; Ito et al.,
1997a; Ito et al., 1997b).
In the present investigation, we synthesized a polypeptide
brush on a nanoporous polymeric membrane to have
stimulus-sensitive gating of channel. It is known that the
conformation of polyelectrolyte chain is dependent upon the
environmental conditions such as pH and ionic strength. In the
region of low pH, poly(L-glutamic acid) chain is protonated and
folded to form -helical structure; in the region of high pH, it is
de-protonated to have extended random structure. The permeation
through the porous membrane, the surface of which was covered
with the self-assembled polypeptide brush, was sensibly regulated
by changing pH because of direct contact of brush chains with
environment.
Materials and Methods
Synthesis of poly(L-glutamic acid) carrying a terminal
disulfide group.
Poly(L-glutamic acid) carrying a disulfide group at the amino
terminal was synthesized. The oxidation of 11-mercaptoundecanoic
acid (Aldrich) was attained by 24 h treatment in DMSO/1N HCl. The
product (216 mg) was dissolved in 15 mL of dimethylformamide
(DMF). 380 mg of
O-(7-azabenzotriazol-1-yl)-1,1,3,3-tetramethyluronium
hexafluorophosphate (HATU, PerSpective Biosystems, Hamburg,
Germany) and 210 microlitter of N,N-diisopropylethylamine (DIEA,
Aldrich) were added to the solution to make disulfide:HATU:DIEA
molar ratio 1:2:2.4. After addition of poly(g-benzyl-L-glutamate)
(PBLG, 1.0 g), the solution was stirred for 1 h in ice bath. PBLG
was purchased from Sigma Chemical Co. which provided the
following MW data: (viscosity) 22,000; (light scattering) 17,400.
The mixture solution was stirred for 24 h at room temperature.
The product, poly(benzyl-L-glutamate) carrying a terminal
disulfide group (PBLG-SS), was isolated by twice precipitation
with methanol, and then dried overnight in vacuo at room
temperature.
In order to hydrolyze PBLG, 500 mg of PBLG-SS were treated in
dioxane (6 mL)/ methanol (2 mL)/ 4N NaOH (2 mL) mixture for 2 h
at room temperature. The precipitate was again dissolved in 500
mL of distilled water, and then concentrated to 50 mL by
ultrafiltration using AMICON model 2000 (mw cut-off = 10,000).
The concentrated solution was lyophilized to obtain
poly(L-glutamic acid) carrying a terminal disulfide group
(PLGA-SS).
Preparation of PLGA immobilizd membrane.
Track-etched porous polycarbonate membrane (DuPont Nuclepore
membrane: average pore diameter, 200 nm) was coated with gold.
The Au-coated membrane was immersed in aqueous solution of
PLGA-SS (2.5 mM, pH = 3.0) for 24 h. The surface-modified
membrane was washed with deionized water until the pH of the
washing liquid became neutral.
FT-ATR-IR measurement.
FT-ATR-IR spectra were measured on Perkin-Elmer infrared
spectrometer using KRS-5 prism.
Permeation experiment.
Water permeation through the membrane was investigated using
an apparatus previously reported (Ito et al, 1997a). The prepared
membrane was mounted on a ultrafiltration cell (Toyo Roshi
UHP-25), and placed 200 cm below a water reservoir. The reservoir
was filled with an aqueous solution and adjusted to different pHs
using NaOH and HCl. The aqueous solution was allowed to flow
under a constant hydraulic pressure. The permeation rate was
calculated by measuring the mass of water permeating through the
membrane every minute.
Results and Discussion
The presence of immobilizated PLGA on the surface of Au-coated
membrane was confirmed by infrared spectroscopy. The FT-ATR-IR
spectrum of PLGA-immobilized membrane shows the amide I (1650
cm-1) and amide II (1550 cm-1) absorptions, and the ester
carbonyl absorption (1730 cm-1), which are characteristic of
PLGA.
The rate of water permeation through an nonimmobilized
membrane was independent of pH. The rate of water permeation
through the grafted membrane was dependent upon pH, which was
high at low pH, but low toward neutral pH. It is expected that in
the region of low pH, the poly(L-glutamic acid) chain is
protonated and folded to form a-helical structure, while in the
region of high pH, it is de-protonated to have extended random
structure. This type of conformational change of polypeptide
chain may have affected the porosity of the membrane, leading to
the pH-dependent permeability of water.
The permeation rate also depended on the ionic strength. With
increasing ionic strength, the pH dependence decreased. In the
high pH region, permeability was strongly dependent on the ionic
strength. The high concentration of ions should moderate
charge-to-charge interactions of polypeptide brush leading to
conformational change.
With the PLGA-grafted membrane, the change in permeability in
response to pH change occurred within a few minutes. The PLGA
brushes were put in direct contact with the media of different
pH; this situation allowed the brushes to quickly respond to the
environmental change. In the case of hydrogel-based system, the
fastest response so for reported is 20 min, and it usually takes
several hours to a day.
A reversible change of permeation rate with pH change between
2 and 7 was observed. The conformational change of self-assembled
PLGA brush reversibly regulates the pore size of the membrane.
Previously we devised some porous membranes having
polyelectrolyte brush on the surface, which was prepared by
plasma polymerization. It was difficult to control the length and
density of polymer brush. The present method enabled us to
fabricate intelligent membrane by combination of porous membrane
and polymer brush. This technique should be useful to prepare
micro-device and nano-devices in the near future.
References
Ito, Y.; Inaba, M.; Chung, D. J.; Imanishi, Y. (1992)
Macromolecules 25, pages 7313-7316. Control of water permeability
by pH and ionic strength through a porous membrane having
poly(carboxylic acid) surface-grafted.
Ito, Y.; Ochiai, Y.; Park, Y.S.; Imanishi, Y. (1997a) J. Am.
Chem. Soc. 119, pages 1619-1623. pH-sensitive gating by
conformational change of polypeptide brush grafted on porous
polymer membrane.
Ito, Y.; Park, Y. S.; Imanishi, Y. (1997b) J. Am. Chem. Soc.
119 , pages 2739-2740. Visualization of critical pH-controlled
gating of porous membrane grafted with polyelectrolyte brushes.
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