Enhancement of photoelectric response of bacteriorhodopsin by multilayered WO3 x H2O nanocrystals/PVA membrane.
ABSTRACT For the first time, a multilayered WO(3) x H(2)O/PVA membrane on bacteriorhodopsin (bR) is constructed to significantly enhance the photoelectric response of bR by the spillover effect of WO(3) x H(2)O nanocrystals, providing great potential in its important applications in bioelectronics and proton exchange membrane fuel cells.
Enhancement of photoelectric response of bacteriorhodopsin by
multilayered WO3?H2O nanocrystals/PVA membrane
Rui Li,abFengping Hu,abQiaoliang Bao,abShujuan Bao,abYan Qiao,abShucong Yu,a
Jun Guocand Chang Ming Li*ab
Received (in Cambridge, UK) 9th November 2009, Accepted 15th December 2009
First published as an Advance Article on the web 23rd December 2009
For the first time, a multilayered WO3?H2O/PVA membrane on
bacteriorhodopsin (bR) is constructed to significantly enhance
the photoelectric response of bR by the spillover effect of
WO3?H2O nanocrystals, providing great potential in its
important applications in bioelectronics and proton exchange
membrane fuel cells.
Bacteriorhodopsin (bR) is the only protein component of the
purple membrane (PM), a two-dimensional crystal lattice in
Halobacterium salinarum (H. salinarum) and one of the
simplest biological energy converters. Due to its extreme
stability to various environmental conditions,1extraordinary
photoelectric and photochromic properties and superior
ability to maintain biological activity on a solid support,2
bR has become one of the most promising biomaterials
for various applications in the bioelectronics field.3Light
absorption by bR leads to conformational change of the
protein, thus driving a proton translocation across the
membrane from the cytoplasmic side to the outer medium.4
The resulting proton gradient generates the electrochemical
photocurrent.5At this juncture, the enhancement of the
photoelectric response of a bR-based system is essential and
is the main challenge in its practical applications. Various
methods such as chemically induced enhancement6and
layer-by-layer enhancement7have been reported. Nano-
materials possess unique properties arising from the size
reduction and thus are attracting major attention in bio-
electronics applications.8Some nanomaterials have been
incorporated with bR to either increase the amplitude9or
change the pattern10of its photoelectric responses. Applica-
tion of WO3?H2O nanocrystals in a biomolecule-based system,
however, has never been studied to date. Based on their
spillover effect, WO3?H2O nanoparticles have been recently
employed to improve the performance of the proton exchange
membrane in fuel cells.11Since the photoelectric response of
bR originates from the protein’s unique proton translocation,
it can be expected to be enhanced by the effective proton
spillover of WO3?H2O nanocrystals. It remains a great
challenge to construct a unique bR/WO3?H2O nanostructure
for the enhancement.
In our work, we incorporate WO3?H2O nanocrystals into a
PVA matrix to engineer a novel bio-organic–inorganic
interface with bR. bR-embedded PM was isolated from
Halobacterial halobium S9 strain according to the standard
protocol,12of which a 5 mg ml?1suspension in DI water was
prepared before use. An electrophoretic sedimentation (EPS)
method5was used to uniformly immobilize the bR-embedded
PM with a favorable orientation on a gold electrode. 1 g
tungsten powder was dissolved in a mixture of 10 ml 30%
H2O2 solution, 10 ml DI water and 5 ml isopropanol to
prepare a precursor of WO3?nH2O nanocrystals. 2 ml of
the as-prepared precursor was then mixed with 6 ml of the
66.67 mg ml?1PVA (MW = 164000) solution (B18% (w/w)
WO3) followed by spin-coating on a freshly prepared
bR-modified electrode (the ratio was optimized by preliminary
experiments). The thickness of the WO3?nH2O precursor/PVA
layer could be tailored by repeating a number of spin-coating
operations, followed by one-hour heating at 60 1C to allow
formation of the integrated bR/WO3?nH2O/PVA membrane.
As controls, WO3?nH2O/PVA membrane and bR/PVA
membrane, respectively were also prepared.
The surface morphology of the as-prepared membrane and
the distribution of tungsten element across the membrane
were examined by field-effect scanning electron microscopy
(FESEM, JEOL JSM-6700F FESEM) and energy dispersive
respectively. The cross-section image of the WO3?nH2O/PVA
membrane (prepared by 12 times of spin-coating) (Fig. 1a)
illustrates that the thickness of WO3?nH2O/PVA membrane is
quite consistent. Inset I shows that the surface morphology of
the WO3?nH2O/PVA membrane is smooth with no obvious
protuberance. The EDX mapping of the membrane surface
(Inset II) and the EDX spectra (Fig. 1b) corresponding to the
measured points shown in the membrane cross-section
(Fig. 1a) demonstrate that the tungsten element is uniformly
distributed across both directions of the WO3?nH2O/PVA
membrane. High resolution transmission electron microscopy
(HRTEM, JEM 2100F, 200 kV, JEOL, Japan) was carried out
to study the microstructure of WO3?nH2O embedded in PVA
membrane, as shown in Fig. 1c–e. Fig. 1c clearly revealed that
WO3?nH2O nanocrystals with diameters of 3–5 nm are
spatially stacked in PVA membrane. The crystal lattice of
WO3?nH2O was further resolved in Fig. 1d, in which the lattice
spacing along (040) is determined to be d040= 0.27 ? 0.01 nm,
in excellent agreement with the lattice constant of the standard
aSchool of Chemical and Biomedical Engineering, Nanyang
Technological University, 70 Nanyang Drive, Singapore 637457,
Singapore. E-mail: ECMLi@ntu.edu.sg; Fax: +65 6791 1761;
Tel: +65 6790 4485
bCenter for Advanced Bionanosystems, Nanyang Technological
University, 70 Nanyang Drive, Singapore 637457, Singapore
cSchool of Materials Science and Engineering, Nanyang
Technological University, Nanyang Avenue, Singapore 639798,
This journal is ? c The Royal Society of Chemistry 2010Chem. Commun., 2010, 46, 689–691 | 689
COMMUNICATION www.rsc.org/chemcomm | ChemComm
WO3hydrate (JCPDS, card No. 20-1806). The selective area
electron diffraction (SAED) pattern in Fig. 1e displays
discontinuous rings composed of diffraction spots, which
further identifies the polycrystalline nature of the nano-
The crystal structure of the WO3?nH2O/PVA membrane was
also characterized by X-ray powder diffraction (XRD, Bruker
AXS X-ray diffractometer). In Fig. 2a, three relatively sharp
peaks are shown in curve II, two of which (2y = 25.631 and
2y = 33.441) can be indexed to the (111) and (040) reflection,
respectively, of the tungsten oxide hydrate (I, JCPDS, card
No. 20-1806), suggesting that the hydrate of WO3 forms
during the preparation of the membrane. Another prominent
peak (2y = 19.421) can be ascribed to the PVA component,
which shifts a little from the main peak (2y = 20.221) in the
XRD pattern of PVA (curve III) due to the concentration
change. No characteristic peaks of impurities are observed.
Hence, the prepared membrane is confirmed to be a
PVA/WO3hydrate hybrid. According to the Scherrer Formula,13
the crystallite size of the WO3hydrate was also calculated to
be B3.0 nm, which is highly consonant with the previous
The thermal properties of the WO3?nH2O/PVA membrane
were determined by thermogravimetric analysis (TGA, Perkin
Elmer Pyris Diamond thermogravimetric analyzer). The TGA
profile for the WO3?nH2O/PVA membrane in Fig. 2b presents
a first weight loss of B7% at around 200 1C, which is
attributed to the evaporation of the occluded water, followed
by a second sharp weight loss of B5% at around 250 1C,
which should be caused by the loss of chemically combined
water in the WO3hydrate. The TGA curve section of PVA
(inset) is rather flat between 200 1C and 300 1C, indicating
there is no decomposition of the polymer in the temperature
range. Between 250 1C and 440 1C, the third sharp weight loss
of B6% occurs in the WO3?nH2O/PVA membrane, which is
consistent with the obvious weight loss of B83% between 320
and 550 1C in the TGA curve of PVA (inset), demonstrating
the decomposition of the PVA component in the WO3?nH2O/
PVA membrane. From the TGA result, the number of
chemically combined water molecules could be calculated to
be 1, i.e., the WO3hydrate should be denoted as WO3?H2O.
The abundant water resource in the hydrated WO3has great
significance in maintaining the moisture of the integrated
membrane for higher stability11and a better proton transport
environment14in the integrated membrane.
The photoelectric responses of the integrated bR/WO3?
H2O/PVA membrane were measured by Autolab PGSTAT
30 with a Polychrome V system (Till photonics) as the light
source (power density = 6.5 mW cm?2).10Amperometric and
potentiometric chrono measurements at open circuit were
conducted for the photocurrent and photopotential response,
respectively, showing a capacitive nature identical to that
generated by pure bR due to the proton pumping effect10
(Fig. 3). Fig. 3a illustrates the effect of multilayer WO3?H2O/
PVA on the photoelectric responses of the integrated
photovoltage response of the membrane proportionally
increase with number increase of the coated WO3?H2O/PVA
layers on top of the bR layer. However, the photoelectric
response hits a maximum at B20 mm thickness of the
multilayered WO3?H2O/PVA membrane (corresponds to 12
spin-coated single layers), and then drops with further increase
of the modified layers. Fig. 3b and c depict the measured
typical responding photocurrent and photovoltage profiles for
membranes consisting of different components, showing that
both photocurrent and photovoltage responses of the inte-
grated bR/WO3?H2O/PVA membrane (signal II in both figures)
are enhanced sharply to around six times more than those of
pure bR (signal I in both figures). The photoelectric responses
for the integrated membrane exhibit significantly longer decay
time (Fig. 3d) than that of the pure bR film, suggesting a
longer neutralization time of the polarization caused by the
larger proton gradient formed by the proton transport in the
integrated system.5However, the PVA/bR membrane without
a WO3?H2O component (Fig. 3b and c, signal III) does not
generate obvious enhancement of the bR photoelectric
response (signal I) in comparison to the integrated bR/WO3?
H2O/PVA membrane with the same thickness (signal II). In
addition, the WO3?H2O/PVA membrane without bR cannot
produce any photoelectric response at all under the same
experimental conditions (signal IV in both figures). All these
results clearly indicate that WO3?H2O nanocrystal in the
integrated bR/WO3?H2O/PVA membrane is the only factor
to dramatically boost the photoelectric response of bR.
SEM micrograph and (II) EDX mapping of the surface, (b) EDX
spectra of the cross-section and HRTEM image at (c) low magnification
and (d) high magnification, (e) SAED pattern of the WO3?nH2O/PVA
(a) SEM micrograph of the cross-section, insets show the (I)
oxide hydrate (JCPDS 20-1806), (II) WO3?nH2O/PVA membrane, and
(III) plain PVA membrane; (b) TGA curves of the WO3?nH2O/PVA
membrane. Inset shows the TGA curves of the PVA membrane.
(a) XRD patterns of (I) the standard spectrum of tungsten
690 | Chem. Commun., 2010, 46, 689–691 This journal is ? c The Royal Society of Chemistry 2010
It is very important to understand the fundamental insight
of the photoelectric enhancement. It is known that WO3?H2O
can perform hydrogen spillover to produce protons,15,16
during which the color of WO3?H2O changes from light yellow
to dark yellow showing the hydrogen tungsten bronze forma-
tion, which is the direct index of the hydrogen spillover.15In
our work, the color change of the WO3?H2O/PVA membrane
in a photoelectric experiment with the bR presence (B5 h)
was much faster than that without bR (B48 h), possibly
evidencing that bR-introduced electric field could strengthen
the hydrogen spillover process. A mechanism based on light-
driven proton pumping of bR associated with hydrogen
spillover of WO3?H2O nanocrystals is therefore proposed, as
shown in Fig. 4. When green light irradiates the integrated
membrane, protons in the interface between the bR film and
the gold electrode are pumped out by bR molecules to produce
capacitive current while building a local electric field vertical to
the membrane to trick the hydrogen spillover16of WO3?H2O
nanocrystals and produce protons, which should move away
from the electrode in the same way as the protons generated
from bR. This larger amount of moving protons can definitely
build higher potential gradient in the double layer structure of
the electrode for a higher photoelectric current. That is why
the photoelectric response is proportional to the number of the
WO3?H2O/PVA layer in the integrated membrane within a
certain range. However, when the WO3?H2O/PVA layer number
increases to above 12, the photoelectric response decreases,
possibly caused by increased resistance inthe thick film(Fig. 3a).
In brief, construction of a multilayered WO3?H2O/PVA
membrane on bR boosts bR’s photoelectric response by
around six times. The mechanism for the photoelectric
enhancement is possibly due to the hydrogen spillover process
of the WO3?H2O. This work could provide a method to
significantly enhance thebR
bioelectronic devices and also render a promising proton
exchange membrane in bio fuel cell applications.
Notes and references
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(a) Effect of different layer numbers of WO3?H2O/PVA on the photo-
electric responses of integrated bR/WO3?H2O/PVA membrane (n = 4);
(b) photocurrent and (c) photovoltage of pure bR (I), bR integrated
with 12 layers of WO3?H2O/PVA component (II), bR integrated with
12 layers of PVA component (III) and 12 layers of WO3?H2O/PVA
membrane alone (IV); (d) decay time comparison of photocurrents
generated by pure bR and integrated bR/WO3?H2O/PVA membrane.
Photoelectric responses of the bR/WO3?H2O/PVA membrane.
integrated bR/WO3?H2O/PVA membrane.
Model of the photoelectric response generation of the
This journal is ? c The Royal Society of Chemistry 2010Chem. Commun., 2010, 46, 689–691 | 691