Naked-eye cadmium sensor: Using chromoionophore arrays of Langmuir-Blodgett molecular assemblies

Article abstract of DOI:10.1021/ac0623540

This study demonstrates the possibility of a reversible naked-eye detection method for submicromolar levels of cadmium(II) using the Langmuir-Blodgett (L-B) technique. Molecular assemblies of 4-n-dodecyl-6-(2-thiazolylazo) resorcinol are transferred on precleaned microscopic glass slides, to act as a sensing probe. Isotherm (π-A) measurements were performed to ensure the films' structural rigidity and homogeneity during sensor fabrication. The sensor surface morphology was characterized using atomic force microscopy and scanning electron microscopy. The probe membrane exhibits visual color transition, forming a series of reddish-orange to pinkish-purple complexes with cadmium, over a wide concentration range (0.04-44.5 μM). Cadmium response kinetics and the changes in the sensors' intrinsic optical properties were monitored using absorption spectroscopy and further confirmed using X-ray photoelectron spectroscopy. A hybrid L-B film composite of poly(vinyl stearate) and pory(vinyl-N-octadecylcarbamate) were investigated for enhancing sensor performance. The sensor was tested for its practical approach to prove its cadmium selectivity and sensitivity amid common matrix constituents using synthetic mixtures and real water samples. Using the sensor strips, the respective lower limits of cadmium detection and quantification are 0.039 and 0.050 μM, as estimated from a normalized linear calibration plot.

Full text of DOI:10.1021/ac0623540

Anal. Chem. 2007, 79, 4056-4065
Naked-Eye Cadmium Sensor: Using
Chromoionophore Arrays of Langmuir-Blodgett
Molecular Assemblies
Deivasigamani Prabhakaran, Ma Yuehong, Hiroshi Nanjo, and Hideyuki Matsunaga*
Research Center for Compact Chemical Process, National Institute of Advanced Industrial Science and Technology (AIST
Tohoku), Sendai 983-8551, Japan
applications in the production of nickel-cadmium rechargeable
batteries, smelting, phosphate fertilizers, ceramic enamels, and
paint pigments, thereby rendering it a species that is certain to
be dispersed into the environment.7,8 It is also estimated that
cadmium ions are released into the atmosphere through natural
phenomena such as erosion, abrasion, and volcanic eruption.
Consequently, cadmium is naturally found in water, soil, and food,
with its average content placed generally between 0.1 and 0.5 ppm
in the earth’s crust.9 Numerous quantitative testing methods are
available for cadmium(II), but because of the high setup and
running costs of sophisticated instrumental methods such as
atomic absorption spectrometry (AAS),10 inductively coupled
plasma atomic emission spectrometry (ICP-AES),11 inductively
coupled plasma mass spectrometry,12 and electrochemical de-
vices,13 small local governments and developing countries cannot
realistically make practical use of them. Although good analytical
This study demonstrates the possibility of a reversible
naked-eye detection method for submicromolar levels of
cadmium(II) using the Langmuir-Blodgett (L-B) tech-
nique. Molecular assemblies of 4-n-dodecyl-6-(2-thiaz-
olylazo)resorcinol are transferred on precleaned micro-
scopic glass slides, to act as a sensing probe. Isotherm
(π-A) measurements were performed to ensure the films’
structural rigidity and homogeneity during sensor fabrica-
tion. The sensor surface morphology was characterized
using atomic force microscopy and scanning electron
microscopy. The probe membrane exhibits visual color
transition, forming a series of reddish-orange to pinkish-
purple complexes with cadmium, over a wide concentra-
tion range (0.04-44.5 µM). Cadmium response kinetics
and the changes in the sensors’ intrinsic optical properties
were monitored using absorption spectroscopy and fur-
ther confirmed using X-ray photoelectron spectroscopy.
A hybrid L-B film composite of poly(vinyl stearate) and
poly(vinyl-N-octadecylcarbamate) were investigated for
enhancing sensor performance. The sensor was tested
for its practical approach to prove its cadmium selec-
tivity and sensitivity amid common matrix constituents
using synthetic mixtures and real water samples.
Using the sensor strips, the respective lower limits of
cadmium detection and quantification are 0.039 and
0.050 µM, as estimated from a normalized linear calibra-
tion plot.
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Fax: (+81) 22-237-7027. E-mail: hide.matsunaga@aist.go.jp.
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4056 Analytical Chemistry, Vol. 79, No. 11, June 1, 2007
10.1021/ac0623540 CCC: $37.00 © 2007 American Chemical Society
Published on Web 04/21/2007

performance is obtainable with pretreatment techniques such
as solvent extraction14,15 and solid-phase extraction,16,17 they are
normally associated with complicated treatment procedures, large
volumes of reagent/solvent consumption, and unsatisfactory
enrichment factors.18 For those reasons, development of smart,
simple, and compact analytical methods for toxicity validation
has been an area of prime research concern. In recent years,
sensor technology is considered as one of the few potential
candidates for environmental monitoring.19 Considerable
attention has been devoted to the design of supramolecules and
chromoionophores that can recognize target analytes
selectively through visual detection and optical responses.20-23
However, developing such sensing materials has not been
an easy task because of many factors, which include tedious
synthesis, supporting electronic devices, insufficient ion
selectivity, and sensitivity. Analytical researchers are intent
upon developing new minimized and green methods using
nanotechnology and nanoscale materials, which constitute a new
and exciting field of research to overcome these looming prob-
lems.
Langmuir-Blodgett (L-B) thin-film technique has been a key
manufacturing tool in nanotechnology, with applications ranging
from biotechnology (membranes and biosensors), gas sensors,
and electronic devices to lithography.24-26 Interest in L-B technique
is fueled by its merits, which include (a) precise control of the
monolayer thickness, (b) homogeneous deposition of the mono-
layers over large areas, and (c) the possibility of generating
multilayer structures with varying layer composition on almost
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reveal meaningful applications in sensing techniques and other
fundamental studies. However, such implications of L-B film
monolayers in toxicity validation have remained only vague
conjecture, especially in the emerging areas of naked-eye sensing.
In that context, we intend to fabricate a smart visual ion sensor
based on L-B methodology that can fulfill major analytical needs
in environmental monitoring.
In this study, we report for the first time the use of chemically
synthesized amphiphilic 4-n-dodecyl-6-(2-thiazolylazo)resorcinol
(DTAR) monolayers as a potential solid-state chromoionophore
sensor for detecting trace levels of cadmium(II). The sensor kit
provides two means of cadmium detection: one by mere naked-
eye detection of the visual color transition, and the other by precise
measurement of the relative changes of the sensor optical
properties. The sensor strips were selective and sensitive for
cadmium ions without major interference from coexisting transi-
tion ions and other cationic and anionic species. The sensor strips
were reversible and reusable without marked loss in their sensing
efficiency up to four repeated cycles. It was inferred that the
combined use of polymer composite with the chromoionophore
molecular assemblies provided a better way to produce mechani-
cally stable optical sensors. The fabricated LB film sensor was
found to be both promising and reliable for rapid and selective
detection of submicromolar cadmium ions from highly saline
samples. To our knowledge, this sensor marks the first step in
developing a naked-eye cadmium sensor using L-B nanoassem-
blies.
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Analytical Chemistry, Vol. 79, No. 11, June 1, 2007 4057

was used for measuring the pressure (π)-area (A) isotherm and
the transfer ratio (τ). Structural features of the synthesized probe
molecules were characterized using an FT-IR spectrophotometer
(IRPrestige-21; Shimadzu Corp.); its purity was ascertained using
CHN elemental analysis (JM10 Microcorder; J-Science Lab.). A
UV-vis spectrophotometer (UV-3150; Shimadzu Corp.) equipped
with a detachable solid sample compartment was used to measure
the absorption spectra of the sensor’s optical changes. The surface
morphology of the probe membrane was studied using tapping-
mode atomic force microscopy (AFM; NanoScope IIIa; Digital
Instruments) and by scanning electron microscopy (SEM) (Mini-
scope TM-1000; Hitachi Ltd.) imaging. The confirmation of
cadmium chelation with an L-B film probe was studied using XPS
(PHI 5601ci; Ulvac) analysis, using Mg KR X-ray source (14 kV,
400 W). The residual metal ions in the aqueous phase after
equilibration were determined using ICP-AES (SPS-1500; Seiko
Instruments Inc.) analysis. A water bath incubator built into a
mechanical shaker (BT-100; Yamato Co. Ltd.) was used for batch
equilibration studies. A portable pH/Ion meter (D-53; Horiba Ltd.)
was used for measuring the subphase (water) pH during L-B
isotherm and film transfer.
Reagents and Procedures. All reagents, solvents, and
chemicals procured for probe synthesis and subsequently for
sensor fabrication were of high-purity grade (Wako Pure Chemical
Industries Ltd., Osaka, Japan) and were used without further
purification. For sensor protection, four amphiphilic polymers
(poly(vinyl-N-octadecylcarbamate) (PVOC), poly(vinyl stearate)
(PVS), poly(maleic anhydride-alt-1-octadecene) (PMO), and poly-
(octadecyl methacrylate) (POMA)) were studied; they were
procured from Aldrich Chemical Co. Inc. (Milwaukee, WI).
Individual metal ion stock solutions were prepared from AAS-grade
standard solutions (1000 ppm (µg/mL) in 0.01 mol/L HNO3),
purchased from Wako Pure Chemical Industries Ltd. For pH
adjustment, Good’s buffers (0.2 mol/L, Dojindo Laboratories,
Kumamoto, Japan), consisting of 3-morpholinopropanesulfonic
acid (MOPS)-NaOH (pH 6-9), 2-(cyclohexylamino)ethanesulfon-
ic acid (CHES)-NaOH (pH 9-10), and N-cyclohexyl-3-aminopro-
panesulfonic acid (CAPS)-NaOH (pH 10-12), were used. Pre-
cleaned microslide glass plates (0.8-1.0 mm thick and 38 × 13
mm, S-111 type; Matsunami Glass Ind. Ltd.) were used as solid
substrates for probe-anchoring.
Chart 1. Structure of Sensing Probe and Its Four
Protecting Polymers
molecular structure of the chromoionophore and the four poly-
mers studied for sensor stability are depicted in Chart 1.
Fabrication of DTAR Multilayer Film Assembly. For sensor
fabrication, the probe and the four polymers were dissolved
respectively in high-purity grade benzene and chloroform. The
pressure (π)-area (A) isotherm plots were measured to elucidate
the phase transition of the structural orientation and film stability
of the probe and polymer layers. On the basis of their individual
limiting collapse pressure (from the isotherm plot), the target
pressures were set to achieve a film transfer of greater homogene-
ity and uniformity. The fabrication of L-B films and the layer-by-
layer transfer of molecular assemblies involve the following
sequence: (a) microdroplets of probe and polymer solutions were
spread evenly on the water subphase of a moving-wall L-B film
fabrication system; (b) after solvent evaporation, the surface
pressure was increased to its target pressure by film compression,
using a Teflon barrier; and (c) upon attaining equilibration, the
compressed film layers were eventually transferred as self-
organized L-B monolayers at the air-water interface by vertical
dipping of the planar glass substrate. During film transfer, the
surface pressure was maintained using a computer-controlled
feedback system.
Isotherm Plots and Transfer Ratios. The pressure (π)-area
(A) isotherm for the DTAR, PVOC, PVS, PMO, and POMA
monolayers generated at the air-water interface is shown in
Figure 1A. The isotherm plot for DTAR film exhibits a first phase
transition at 36.3 Å2 that should correspond to the nonplanar to
planar conformation of the thiazole and phenyl rings, with the
dodecyl chain oriented outward from the water surface. With
increasing compression, the isotherm observed after 25 mN/m
with a molecular area of ∼32.9 Å2 corresponds to the transition
from liquid-expanded to a more condensed state, reaching to its
limiting pressure at 37.5 mN/m. In the case of polymer films, their
isotherm exhibits a two-phase transition of molecular orientation,
first from the liquid-expanded gaseous phase to a liquid-expanded
phase, with respective mean molecular areas of 27.0, 35.1, 39.2,
and 50.7 Å,2 for POMA, PMO, PVS, and PVOC. This phase
transition possibly explains the gauche conformation acquired
using the long alkyl units. The second transition to an ordered,
liquid-expanded condensed state was observed at 22.1, 27.9, 33.0,
and 43.3 Å2 for POMA, PMO, PVS, and PVOC, respectively,
corresponding to surface pressure of 15.2, 47.1, 51.7, and 45.5 mN/
The amphiphilic chromophore (DTAR) was synthesized under
freezing conditions using equimolar coupling of prediazotized
2-aminothiazole with 4-dodecylresorcinol. The product was ob-
tained as a dark red precipitate and was purified through repeated
recrystallization from hot ethanol.31 The details of probe synthesis
and its characterization are provided in Supporting Information.
Based on spectral characteristics of 2-thiazolylazoresorcinol, at pH
<6.9, the probe (DTAR) exists as a neutral species (H2L) with a
λmax at 455 nm (ꢀ - 1.99 × 104 L mol-1 cm-1). The visible
absorption spectra of DTAR (25 µM) were measured after
extraction into 4-methyl-2-pentanone. Solvent extraction studies
confirmed that the DTAR molecules can be extracted as a neutral
H2L species, under these conditions. The monoanionic species
(HL-) and dianionic species (L2-) were extracted respectively as
ion pairs with sodium ion, with a λmax at 495 and 539 nm. The
(31) Ueno, K.; Imamura, T.; Cheng, K. L. Handbook of Organic Analytical Reagents,
2nd ed.; CRC Press: Boca Raton, FL, 1992; pp 205-206.
4058 Analytical Chemistry, Vol. 79, No. 11, June 1, 2007

Figure 1. (A) Surface pressure (π) vs area (A) isotherm of DTAR, PVOC, POMA, PMO, and PVS films at the air-water interface at 20 °C
obtained by compression at a rate of 10 mm/min. (B) (%) Transfer ratio (τ) values observed for DTAR 10 layers, and for PVOC, PMO, and PVS
(3 layers each) on DTAR10.
that PMO monolayers are unsuitable for sensor protection. Table
1 shows optimized instrumental parameters for generating an
efficient sensing system; precedent studies of the mixing ratios
of hybrid probe-polymer composite will be described in subse-
quent sections.
Table 1. Optimized Instrumental and Experimental
Parameters
mixed
parameters
DTAR PVOC
PVS
ratio
limiting pressure (mN/m)
target pressure (mN/m)
barrier speed (mm/min)
dipper upstroke (mm/min)
dipper downstroke (mm/min)
concentration (mM)
38
28
45
40
10
0.10
-
51.5
43
37
32
10
Analytical Method for Ion Sensing. Aqueous-phase equili-
bration of sensor strips with cadmium ions was performed using
a batch equilibration method. For ion sensing, sensor strips were
equilibrated for specific time durations at 200 rpm in 50-mL-
capacity containers holding 30 mL of overall sample volume,
including 3-4 mL of MOPS buffer (pH 7.5). Optimization of sensor
working parameters was normally performed at 30-40 °C, unless
otherwise specified. Buffer solutions were adjusted to ambient pH
using a microcomputerized pH/Ion meter (F-24 model; Horiba
Ltd.). After phase equilibration, the residual aqueous-phase metal
ion concentrations were estimated using ICP-AES. Metal ion con-
centrations are expressed either in terms of parts per million (ppm,
mg/L or µg/mL), in parts per billion (ppb, µg/L), or in molarity
(M, mol/L). For sensor regeneration, a 20-mL volume of 0.1 M
HCl or EDTA was used as a cadmium decomplexing agent.
10
10
0.25
0.25
1
0.10
0.10
1
0.15
0.10
1
1
volume (µL) range
85-100 75-80 75-85 120-150
600
60
dipper up stay time (s)
dipper down stay time (s)
600
60
600
60
600
60
m, prior to film collapse. The isotherm plot for POMA indicates
that the polymer does not form stable layers on the water
subphase; hence, PVOC, PVS and PMO films were investigated
to ensure sufficient cohesion of the polymer monolayers on the
transferred probe monolayers.
The mechanism of monolayer transfer onto hydrophilic SiO2
(glass) substrate was investigated, where the probe monolayers
exhibits a homogeneous Y-type film deposition pattern. The extent
of deposition was measured in terms of transfer ratio (τ), defined
as the ratio of monolayer area occupied on the water surface (AL)
and the solid substrate (AS): τ ) AL/AS.32 For a homogeneous
film coating, a τ value of 1 ( 0.05 was achieved during each
(DTAR) monolayer transfer (Figure 1B). However, with polymer
films, τ values of e0.97 and g0.25 for PVOC and e0.89 and g0.15
for PVS were achieved, respectively, for the upstroke and
downstroke (Figure 1B). This helps in predicting that both
polymers exhibit a mixed Z-type (head-to-tail) pattern, with a
prominent monolayer transfer during the upstroke. However, for
PMO, a τ value of e0.5 and g0.15 was observed for upstrokes
and downstrokes, despite greater film rigidity. For the efficient
protection of the probe molecular assemblies, the polymer
monolayers are expected to have higher τ values to ensure greater
surface coverage. Based on preliminary studies, it was predicted
RESULTS AND DISCUSSION
Characterization of Film Sensor Surface Morphology. XPS
analyses of the DTAR 10-layered film sensor assembly confirms
the elemental presence of NdN and OH groups along with the
thiazole unit. The analysis was performed in a sample chamber
pressure maintained under 10-7 MPa and the binding energy
scales were adjusted to the highest C(1s) peak position equal to
285 eV. Figure 2A depicts the XPS plot for a Cd2+ chelate probe
assembly, with a characteristic Cd(3d) peak corresponding to the
binding energy of 405 eV. The surface topographic image of a
cadmium-complexed, five-layered DTAR assembly, observed using
tapping-mode AFM (Figure 2B), reflects a distorted peak-valley
domain structure with few holes, associated with the distorted
molecular orientation of probe monolayers, attributable to the
steric influence of cadmium inclusion. In addition, AFM imaging
of 10-layer probe strips at different cadmium(II) concentrations
(1.25, 0.75, and 0.25 µM) clearly illustrates the diminishing size
of the membrane cavity with increasing cadmium chelation (right
(32) Ullman, A. An Introduction to Ultra Thin Organic Films from Langmuir-
Blodgett to Self-Assembly; Academic Press Inc.: New York, 1991.
Analytical Chemistry, Vol. 79, No. 11, June 1, 2007 4059

Figure 2. (A) XPS spectra of Cd2+ complexed 10-layer DTAR film using Mg KR X-ray source (14 kV, 400 W). (B) AFM images in tapping
mode, (i) 10-layer DTAR film with Cd2+ions, (area, 1 µm XY, 10 nm Z), (ii) its surface morphology (area, 3 µm XY, 30 nm Z) at various cadmium
concentrations (1.25, 0.75, and 0.25 µM), and (iii) aggregate structure of a polymer-probe composite observed using AFM. (C) SEM imaging,
(i) at the membrane-glass interface (700 µm), (ii) DTAR film surface topography with Cd2+ on glass substrate (20 µm), with 4000 × order of
magnification, and (iii) Cd2+ complexed DTAR monolayer pattern, a closer look.
to left). An AFM image of a polymer-probe L-B composite reveals
a sheetlike aggregate structure, which might be caused by the
certain degree of phase separation observed between the probe-
polymer monolayers. SEM images recorded at the probe-
substrate interface (marked as a thick line) reveal a smooth
surface morphology, which showed some roughness with cad-
mium inclusion (Figure 2C).
Polymer Influence on Sensor Stability. For studies of the
sensor stability, membrane strips of 22 probe layers were
uniformly prepared under default conditions and their relative film
stability was checked in terms of its absorbance intensity at λmax
539 nm, after 45 min of equilibration at different pH conditions.
Figure 3A depicts an unaltered film stability profile between pH
6.0 and 7.0. However, under extreme solution pH, desorption
(leaching) of film monolayers was observed because of the fragile
nature of the outer DTAR multilayers, held by a weak van der
Waals force. The easy desorption of hydrophilic H3L+ and L2-
species, under acidic and alkaline conditions, was predicted as
the major reason for such behavior. Consequently, for the intact
position of the DTAR layers, a systematic study of the use of
polymeric film coating for sensor protection was performed.
First, the nonspectral interference property of the PVOC
polymer was checked by transferring PVOC3 film on a D22 (D
denotes DTAR, and 22 indicates no. of monolayers) membrane
strips. Changes in the absorption spectra were measured with and
without PVOC. The peak intensity of the probe and its spectral
properties were apparently unaltered in the presence of polymer
monolayers, which showed similarity with PVS coatings. Study
4060 Analytical Chemistry, Vol. 79, No. 11, June 1, 2007

Figure 3. (A) Stability profile of D22 film sensors as a function of solution pH; (B) relative stability profile of D22 film sensors protected with
polymer films, under extreme working conditions.
Figure 4. (A) Variation in absorption spectral profile for a D16-PVOC1 sensing system, with increasing cadmium concentration (mg/L) at pH
7.5 (10 min, 40 °C). (B) concentration-proportionate signal response for cadmium ions on D31-PVOC1 strips, batch equilibrated at pH 7.5 (20
min, 40 °C).
of the durability of a D22-PVOC3 film molecular assembly at
extreme pH conditions of 2.0 and 9.5 revealed no great change in
the signal intensity, thereby confirming the absence of probe
leaching. Additional studies also confirmed that a single polymer
layer (PVOC1) was sufficiently efficient for sensor protection.
Because L-B films are prone to deteriorate in the presence of
detergents and organics, a comprehensive study was performed
on the sensor’s stability in the presence of various surfactants (see
Supporting Information). Interestingly, the polymer (PVOC)
coating ensures a stable sensor assembly, even in the presence
of major surfactants. However, similar studies performed with
D22-PVS3 strips reveal a measurable probe loss attributable to
the insufficient molecular interaction of the PVS monolayers (τ
>0.9) with the DTAR membrane. Figure 3B represents the
stability profile of a D22 L-B film sensor coated with a PVOC1
and PVS3 system, under rigorous experimental conditions. The
surfactant presented in Figure 3B is 0.14 mM sodium lauryl
sulfate, an anionic surfactant that is commonly used as an
emulsifier in soaps and detergents.
cadmium chelation was studied before and after equilibration in
terms of the relative changes in the sensor absorption spectra.
Absorption spectra of the sensor assembly showed a bathochromic
shift from 539 to 607 nm upon addition of Cd2+, as a result of the
formation of the charge-transfer complex. The relative absorbance
at λ607 nm was at its maximum at pH 7.5 (MOPS buffer), with
prominent color transition. However, an optimum pH range of
7.0-7.7 was selected for both quantitative and visual detection of
cadmium (see Supporting Information). Because the ion-sensing
property of the probe layers was prominent at near-neutral and
slightly alkaline conditions, we recommend that the sample pH
be adjusted within the optimum working range to achieve
distinctive color transition and sensitive analysis for cadmium ions.
Panels A and B in Figure 4 respectively show that the relative
signal response for various cadmium concentrations as a function
of probe monolayers was examined using D16-PVOC1 and D31-
PVOC1 sensor strips.
The visual color transition profile observed on a D16-PVOC1
sensor system for various cadmium(II) concentrations (µg/mL)
is shown in Figure 5A. The sensor strips exhibit a prominent color
transition from its original yellowish orange to form a series of
reddish-pink, pinkish-violet, and deep-purple complexes, with
increasing cadmium content. The observed color change was
Efficiency of Cadmium Sensing. The ion-sensing property
of the sensor for cadmium ions as a function of solution pH (1-
12) was tested with a series of D16-PVOC1 strips, using 5 µM
cadmium solutions batch equilibrated for 45 min. The extent of
Analytical Chemistry, Vol. 79, No. 11, June 1, 2007 4061

Figure 5. (A) Visible color transition pattern observed on a D16-PVOC1 multilayer sensor assembly mounted on the glass substrates, with
increasing concentrations of cadmium ions (pH 7.5, 10 min, 40 °C). (B) A photographic image depicting the sensor instant regeneration from its
cadmium complexes, with 0.1 M HCl.
stable even after 3 days’ storage, with reproducible spectral data
obtained on subsequent trials. Regeneration of the L-B film sensors
was studied using both protolysis and a liquid-exchange reaction.
In protolysis, a solution pH of <1 engendered spontaneous
cadmium desorption from the sensor phase. However, to prolong
the sensor life span, ligand exchange using 0.01 M EDTA was
used for effective removal of cadmium ions after continuous
equilibration for 40-45 min. The restored yellowish-orange strips
were washed with water, and their absorption spectra were
measured to elucidate the unaltered signal intensity at λmax, 539
nm. The sensor was reused for ion sensing up to 3-4 repeated
cycles without loss of efficiency. The reversibility of the L-B film
sensor strips, upon equilibration with a 20-mL volume of 0.1 mol/L
hydrochloric acid, is depicted in Figure 5B.
Cadmium Response with Increasing Monolayers. With the
Figure 6. Temperature-dependent cadmium response kinetics for
intention of fabricating the most sensitive film sensor, six L-B film
D16-PVOC1 sensor. The inset graph denotes the signal response
for cadmium with D31-PVOC1 film sensor at pH 7.5.
sensor assemblies, i.e., D62-PVOC1, D31-PVOC1, D24-PVOC1,
D16-PVOC1, D10-PVOC1, and D6-PVOC1, were fabricated and
their relative signal responses for cadmium ions were tested. With
the D62-PVOC1 and D31-PVOC1 film sensors, the initial color
contrast of the probe film was too intense to show the visual color
transition for micromolar concentrations of cadmium ions. In
addition, with increasing monolayers (>18), the sensitivity of
cadmium detection <0.05 µM was prolonged to 40 min of
equilibration at 30 °C. However, sensor assemblies ranging
between D16-PVOC1 and D10-PVOC1 offer quantitative sensing
of cadmium down to 0.05 µM, within 10 min of equilibration at
40 °C. The sensitivity and speed of reaction between cadmium
and L-B film sensors increases considerably with decreasing
DTAR monolayers. Using fewer monolayers enables faster and
greater metal ion penetration through the probe molecular
channels, which is not easily feasible with a dense membrane
system, in spite of the sensor system exposure to high surface
area. However, with the D6-PVOC1 system, because of the low
chromophore content, it was difficult to perform naked-eye
detection and an instrumental analysis was necessary for detection.
Consequently, L-B film sensor strips of D10-PVOC1 and D16-
PVOC1 assemblies were considered preferable for reliable visual
and quantitative detection of cadmium ions at a rapid time scale.
The maximum response time required for cadmium signal
saturation was monitored using a series of D16-PVOC1 and
D31-PVOC1 glass strips, using 1 µM cadmium solution. Figure
6 shows that, to achieve prominent color transition and signal
saturation at 20 °C, a time of 25-30 min was required for both
D16-PVOC1 and D31-PVOC1 sensors. Experiments were per-
formed at different temperatures (30, 40, and 50 °C) to elucidate
the influence of the solution temperature on the rate of cadmium
sensing. The time-proportionate signal response for cadmium ions
at various solution temperatures was plotted against its relative
absorbance at λ607 nm, with respect to the blank sensor (D31-
PVOC1 and D16-PVOC1). It was interesting to observe that the
response time was reduced to <10 and <20 min at 40 °C, for
D16-PVOC1 and D31-PVOC1 systems, which could not be
reduced further at even higher temperatures (>50 °C). It was
inferred that the raised solution temperature enhances the kinetic
energy of the aqueous-phase metal ions, thereby facilitating the
faster diffusion-controlled exchange process through the dense
probe membrane channels. The diffusion coefficient values
(20 °C, 10 min) for cadmium ions were calculated to be 7.51 ×
10-9 and 1.98 × 10-9 cm2/s, with D16-PVOC1 and D31-PVOC1
strips, respectively, using the following expression, proposed by
Zhang and Davison.33
D ) M∆g/CbtA
(1)
Therein, D is the diffusion coefficient of Cd2+ through the
membrane of thickness ∆g, at any time t and A is the exposed
(33) Zhang, H.; Davison, W. Anal. Chem. 1995, 67, 3391-3400.
4062 Analytical Chemistry, Vol. 79, No. 11, June 1, 2007

For this, appropriate ratios of DTAR and PVOC were prepared
(90:10, 80:20, 60:40, 40:60, 20:80, 10:90) and transferred as hybrid
multilayer composites in the form of 10-16 monolayers. The (π-
A) isotherm plots for different ratios of DTAR/PVOC and DTAR/
PVS were measured (see Supporting Information); their relative
film stability and cadmium signal response were investigated. The
stability of DTAR-PVOC mixed layers was studied in 2 M HCl
solution, which revealed a mere e4% loss in the chromophore
signal intensity, compared to a >60% loss for 100% DTAR (see
Supporting Information). However, exposing the DTAR-PVS
hybrid sensor assemblies to extreme acidities revealed probe
leaching over time, underscoring the poor efficiency of the PVS
layers.
Fabrication of hybrid L-B film composites was quick, but when
subjected to time-dependent phase exchange, it exhibits a pro-
longed response time for cadmium sensing. Kinetic studies
performed using a series of hybrid-sensor composites (DTAR/
PVOC ratios of 85:15, 75:25, 70:30, 60:40, 50:50), to estimate the
relative rate of cadmium (5 µM) signal response, confirmed this
behavior. With lower DTAR ratios of 60:40 and 50:50, the elapsed
time for signal saturation was longer (>20 min), which is
apparently related to the net decline in the probe monolayers,
aside from increasing PVOC content that might impair the free
ion mobility through the probe molecular channels. In addition,
the low color contrast associated with these hybrid assemblies
hinders visual detection. However, at higher DTAR ratios (70:30,
75:25, 85:15), the cadmium visual sensing was good, with the
sensing time limit slashed to <15 min for detection of 0.10 µM
cadmium concentration at 40 °C. However, hybrid L-B composites
of 90:10 or 95:5 (DTAR/PVOC) assemblies were unstable to
sensing studies because of the inadequate polymer composition.
It was also predicted that hybrid L-B composites (75:25-85:15)
comprising more than 16 monolayers detected higher concentra-
tions (gµM) of cadmium, but were inappropriate for submicro-
molar cadmium concentrations.
Table 2. Tolerance Limitsa for Various Inorganic and
Complexing Species during Cadmium Sensing
tolerance
limit
tolerance
limit
cations
(mg/L)
anions
Cl-
(mg/L)
Na+/ K+
Ca2+/Mg2+
Ba2+/Sr2+
Fe3+/Fe2+
Co2+/Ni2+
Cu2+/Zn2+
Mn2+
8500/7525
1750/1520
150/14.8
∼12.5b
4.55b/4.80b
4.75b/5.05b
2.35
8960
7850
250
220
350
350
250
250
70
250
220
255
250
340
-
NO3
-
HCO3
3-
PO4
F-
Br-
-
NO2
-
Bi3+/Al3+
Cr6+
10.6/11.5
12.0
SO3
SCN-
As5+
11.7
CO3
2-
Sn4+/2+
Sb2+
∼11.2
citrate
9.95
tartrate
Mo6+
10.4
oxalate
Ti4+
9.5
potassium
biphthalate
phthalate
La3+
13.3
340
tolerance
surfactants
limit (mg/L)
cetyltrimethyl-
78.0
89.1
82.2
79.9
ammonium chloride
tetraethylammonium
-chloride
tetraamylammonium
chloride
dilauryldimethyl-
ammonium bromide
sodium dodecyl sulfate
Triton X-100
44.5
98.5
100.2
0.0011%
Triton N-101
humic acid
a Data denote quantities up to which no spectral interference
was observed in cadmium (0.5 µM) sensing. b Maximum tolerance
limit attained in the presence of masking agents during cadmium
sensing.
Ion Selectivity and Matrix Tolerance Limits. The ion
selectivity of the film sensor (D16-PVOC1) in detecting trace
cadmium ions from multicomponent system was studied in terms
of its matrix tolerance limits. For this study, 0.5 M each of K+,
membrane surface area. In addition, M is the mass of cadmium-
(II), which is defined as M ) CmVm, where Cm is the cadmium
ion concentration in the probe membrane of volume, Vm. Be-
cause the viscosity of the medium changes with temperature,
the cadmium diffusion coefficients values at 30 and 40 °C were
calculated using eq 2, where DT is the diffusion coefficient at any
-
Na+, Cl-, and NO3 and 0.01 M each of Ca2+, Mg2+, Ba2+, F-,
2-
Br-, and SO4 were used because they form a class of matrix
components that are commonly found in major environmental
samples. In addition, frequently encountered metal ions such as
(10 µM) Mn2+, Fe2+, Fe3+, Co2+, Ni2+, Cu2+, Zn2+, Bi3+, and Al3+
1.37023 (T - 20) + 8.36 × 10-4 (T - 20)2
log DT )
+
109 + T
-
and anions such as (2.5 mM) HCO3 and PO43-, which might
log D20 (273 + T)
(2)
coexist with cadmium ions in wastewater samples, were also
investigated. However, Table 2 shows that the tolerance limits
for the studied foreign ions were established individually by batch
equilibration with 0.5 µM cadmium solutions, to the maximum
limit to which cadmium ions were found to be spectrally free from
interference. The analytical data reveal that most of the matrix
species showed no significant interference, except Ba2+, Fe3+, and
293
temperature T and D20 is the diffusion coefficient of Cd2+ at
20 °C, obtained from eq 1. The diffusion coefficient values were
found to increase with temperature for both D16-PVOC1 and
D31-PVOC1 sensors. For instance, the D30 and D40 values for
cadmium(II) at 10 min with D16-PVOC1 sensor were calculated
respectively as 3.69 × 10-8 and 1.04 × 10-7 cm2/s.
2-
SO4 ions, which slightly prolonged the reaction time. Interfer-
ence from ferric ions was eliminated using 0.2 mM pyrophosphate
solution prior to analysis. Apart from this, Cu2+, Co2+, Ni2+, and
Zn2+ ions were found to exhibit positive interference when greater
than 0.61 ppm. However, these ions were eliminated quantitatively
up to e5.0 ppm by adding a mixture 0.25 mM thiosulfate, citrate,
Hybrid Probe-Polymer L-B Sensor. To enhance the
performance of the L-B film sensor and to reduce the possible
stages of time consumption without compromising the film quality,
the combined use of polymer and chromophore was investigated.
Analytical Chemistry, Vol. 79, No. 11, June 1, 2007 4063

Figure 7. (i) Ion-selectivity photographic profile depicting the degree
of tolerance achieved for various matrix constituents on a D16-
PVOC1 L-B film sensor, (ii) relative color transition observed
for interfering metal ions present in 100-fold excess over cadmium
ions, and (iii) cadmium selectivity with respect to other toxic ions
(20-90-fold excess), which were suppressed beyond their tolerance
limit.
Figure 8. Normalized linear calibration plot for a cadmium-
complexed D16-PVOC1 film assembly. Data points denote the
absorbance difference at λ 607 nm, with respect to a blank D16-
PVOC1 sensor.
µM cadmium using D16-PVOC1 sensor strips at pH 7.5 (40 °C,
10 min). The relative standard deviation for the analysis was
calculated as 0.259%.
and tartrate solution along with MOPS buffer. Tolerance studies
were also extended to some toxic heavy metal ions, Sn2+/4+, Sb2+
,
LD ) k1Sb/m
LQ ) k2 Sb/m
(3)
(4)
As5+, As3+, Mo6+, Cr6+, Bi3+, and Al3+, which were found non-
competent for the chelating sites and hence did not cause any
serious interference. However, beyond their tolerance limits (listed
in Table 2) they showed negative interference resulting from the
increasing ionic strength of the medium. However, Pb2+ and Hg2+
ions were found to be partly competitive under these conditions;
they were suppressed using 0.15 mM oxalate and citrate, up to
1.5 ppm. The sensor’s visual transition for cadmium (0.05 ppm)
is highlighted in Figure 7 in comparison with various foreign ions.
Because surfactants (cationic, anionic, and nonionic) are known
to play an influential role in both L-B film durability and the ion-
sensing process, prominent surfactants that are commonly used
were examined. Interestingly, aside from sodium dodecyl sulfate,
which, when greater than 45 mg/L concentration, was found to
enhance the probe initial optical intensity, the surfactants were
ineffectual, even beyond 75 mg/L, which is an added advantage
of using these sensor strips.
Therein, Sb represents the standard deviation of the signal
response for the blank, m is the slope of the linear calibration
range, k1 ) 3, and k2 ) 10.
APPLICATION
Sensing Cadmium from a Synthetic Mixture. The practical
utility of the sensor strip in detecting ultratrace cadmium ions
from a wastewater sample was tested using a simulated synthetic
solution containing 5500 mg/L Na+ and K+, 750 mg/L Ca2+ and
Mg2+, 2.0 mg/L Mn2+, Cu2+, Ni2+, and Co2+, and 10 mg/L Fe2+
and Zn2+ ions, along with 6000 mg/L Cl-and NO3- and 400 mg/L
SO42-, F-, and Br- ions. To this composite, 0.010 mg/L cadmium
ions was spiked and equilibrated under optimum conditions. The
corresponding spectral changes of D16-PVOC1 strip were
monitored, and the normalized absorbance at 607 nm was fitted
into the calibration plot, which showed a value of 9.88 ( 0.5 µg/L
(95% CL), for triplicate measurement (RSD 3.1%).
Targeting Cadmium from Spiked Real Samples. Quantita-
tive estimation of submicromolar levels of cadmium was attempted
from various real sample effluents collected from different sources
including a food processing factory, a semiconductor fabrication
facility, quartz manufacturing facility, and a local hospital. Because
the real samples subjected to the current analysis were pretreated,
cadmium-sensing amid the multicomponent system was viewed
using an internal standard addition method. The samples were
filtered and first subjected to ICP-AES analysis to measure the
possibility of detecting any toxic metal ions, but only 15.7-265
Calibration Graph and Statistical Analysis. The concentra-
tion-proportionate absorption spectra for cadmium were measured,
and their relative absorbance was normalized with respect to blank
sensors at λ607 nm. A linear calibration graph in the cadmium
concentration range of 0.025-4.46 µM was achieved, with a
coefficient of determination (r2) of 0.996 (Figure 8). The detection
(LD) and quantification (LQ) limits were determined from the plot
using eqs 3 and 4,34 which were estimated respectively as 0.039
and 0.050 µM. To validate the precision and accuracy of the
method, seven successive measurements were performed for 0.31
(34) Christian, G. D. Analytical Chemistry, 6th ed.; John Wiley & Sons Inc.: New
York, 2003; pp 112-113. Skoog, D. A.; Holler, F. J.; Nieman, T. A. Principles
of Instrumental Analysis, 5th ed.; Thomas Asia Pty. Ltd.: Singapore, 2005;
pp 13-14.
4064 Analytical Chemistry, Vol. 79, No. 11, June 1, 2007

Table 3. Determination of Spiked Cadmium Ions from Industrial and Domestic Effluents
Cd2+ (µg/L)
amount
amount
found
effluent source
spiked amount
added
food
10 mg/L Ca2+, Mg2+
;
10
10.90b (3.8)
20.29b (2.7)
processing
factory
1.0 mg/L Co2+, Cu2+, Ni2+, Zn2+, Fe2+
;
20
0.05 mg/L Mn2+
;
0.2 mg/L Sn2+, Sb2+
250 mg/L Na+, K+
;
quartz
0.5 mg/L Si4+
;
10
20
11.24 (3.6)
19.90 (2.8)
manufacturing
facility
0.15 mg/L Bi3+, Tl+, Ga3+, Ir3+
;
0.10 mg/L La3+, Ce4+, Nd3+, Sm3+
0.25 mg/L Bi3+, Al3+, Tl+, Ga3+, Ir3+
semiconductor
fab
;
15
20
35
13.38b (4.3)
18.73b (3.8)
36.32b (3.8)
0.10 mg/L La3+, Ce4+, Nd3+, Sm3+
;
0.5 mg/mL Ni2+, Co2+, Mn2+
;
10 mg/L Ca2+, Mg2+
;
0.5 mg/L - Si4+
local
1.75 mg/L Cu2+, Ni2+, Zn2+, Mn2+, Fe2+
;
6.5
15
5.93b (4.1)
hospital
effluent
15 mg/L Ca2+, Mg2+
275 mg/L Na+, K+
;
14.16b (4.0)
a Values in parentheses denote relative standard deviations for triplicate measurements. b Data obtained after eliminating foreign ions was beyond
their noninterference limit.
mg/L alkali and alkaline earth metal ions were detected, aside
from traces (0.02-0.083 mg/L) of Zn2+, Mn2+, and Fe2+ ions. The
studied samples were used as a precursor to test the sensor’s
practical implementation in real samples. Consequently, Table 3
shows that a known amount of foreign ions that might coexist in
the effluent solution was added to the effluent samples, along with
a trace amount of cadmium ions. The spiked effluent solutions
were batch-equilibrated with D16-PVOC1 membrane strips and
fabricated under default conditions. Then, the sensing was
performed at 30 °C, under optimum experimental conditions.
The analytical data indicate the promising aspect of the
fabricated L-B film sensor as a potential candidate for monitoring
environmental and synthetic samples. It is also noteworthy
that the levels of foreign ions spiked to these samples were many
times greater than any industrial or domestic effluent samples
that one might reasonably encounter in the present day, based
on the regulatory laws of waste disposal and environmental
pollution.
present film sensor offers good ion selectivity for cadmium that
is useful for sensitive detection of low microgram per liter levels
of cadmium ions from synthetic and environmental samples.
Interestingly, the film sensor exhibits greater durability and
complete reversibility, which markedly reduces costs for these
types of sensor. The salient features, such as naked-eye sensing,
faster exchange kinetics, portability, and storability, render the
present solid sensor a perfect testing kit for on-site field analyses.
ACKNOWLEDGMENT
This work was financially supported by the Japan Society for
the Promotion of Science (JSPS).
SUPPORTING INFORMATION AVAILABLE
Synthesis (Scheme S1) and spectral characterization of probe,
4-n-dodecyl-6-(2-thiazolylazo)-resorcinol). Characterization of probe
molecular assemblies (D2) by ATR-FT-IR spectra (Figure S1).
Influence of sample pH on cadmium signal response (Figure S2).
Isotherm plot for probe-polymer hybrid sensor (Figure S3) and
its relative stability (Figure S4). Influence of surfactants on sensor
stability (Figure S5). This material is available free of charge via
the Internet at http://pubs.acs.org.
SUMMARY
In this work, visual and quantitative sensing of trace cadmium
ions was undertaken using nanoassemblies of amphiphilic chro-
moionophore molecules anchored on microscopic glass slides and
covered with polymer monolayers. This demonstration of chro-
moionophore monolayers as a naked-eye sensing tool for cadmium
ions presents the first example of a new class of solid-state
colorimetric ion sensors using L-B thin-film methodology. The
Received for review December 13, 2006. Accepted March
23, 2007.
AC0623540
Analytical Chemistry, Vol. 79, No. 11, June 1, 2007 4065