Figure 3-1 shows the flow chart of the experimental design for the fabrication of IPC.
In this study, IPC was fabricated using a templating sol-gel method. Monodisperse PS latex was prepared by oil-in-water emulsion polymerization. PS opal structure was then fabricated by heat-assisted sedimentation. A molecularly imprinted sol solution was then infiltrated into the interstitial spaces between PS microspheres. The well-organized inverse opal structure was obtained after solvent extraction. The preparation conditions of imprinted sol solution including cross-linker, solvent and functional monomer were optimized to obtain the optimal detection ability. The linear range, detection limit, selectivity and sensitivity of IPC were examined in terms of shifts in the wavelengths of diffractive UV light.
3-1 Chemicals
All solvents and chemicals used in this study were of reagent grade and were used without further purification unless special explanation. Styrene (C8H8, Sigma-Aldrich, GC grade, 99%) was used as the monomer of PS latex. Potassium persulfate (KPS, K2S2O8, Sigma-Aldrich, 99%) and sodium dodecyl sulfate (SDS, C12H25SO4Na, Merck, 99%) were used as the initiator and the capping agent, respectively for the synthesis of monodisperse PS microspheres. Zirconium (IV) propoxide (ZPO, Zr(OCH2CH2CH3)4, Sigma-Aldrich, 70 wt
% solution in 1-propanol), bisphenol A (BPA, C15H16O2, Sigma-Aldrich, 99%), phenyltrimethoxysilane (PTMOS, C9H14O3Si, Sigma-Aldrich, 97%) and absolute ethanol (EtOH, C2H6O, Sigma-Aldrich, HPLC grade, 99.8%) were used as the cross-linker, template, functional monomer and solvent for the sol-gel derived MIP, respectively. Toluene (C6H5CH3, J.T. Baker, HPLC grade, 99.9%) and methyl alcohol (CH3OH, Mallinckrodt, HPLC grade, 99.9%) were employed as extraction solvents to remove the PS and BPA from
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IPC. Phenol (C6H6O, Riedel-de Haën), 1-naphthaol (C10H8O, Riedel-de Haën, 99%) and 4-tert-butylphenol (BP, (CH3)3CC6H4OH, Sigma-Aldrich, 99%) were used as the analogues of BPA. Deionized water (DI, Millipore, 18 MΩ cm) obtained from Milli-Q water purification system was used throughout the experiments. Table 3-1 lists the structures of the major reagents used in the photonic crystal and the imprinted polymers.
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Systhesis of polystyrene microspheres Opal structure Inverse opal structure Characterization
Optimization
SEM DLS TEM
Characterization Preparation
Imprinted polymer infiltration
Polystyrene opal structure Templates removal
Photonic crystal
Detection
UV-visible
Fabrication of BPA imprinted photonic crystals
Molecularly imprinted polymers
Fabrication of IPC
Sensitivity Selectivity Equilibrium Application
SEM BET FTIR Adsoronption
Cross-linker Solvent Functional monomer
Figure 3-1 Flow chart of experimental design in this study.
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Table 3-1 Structures of the major reagents used in photonic crystal and imprinted polymers.
Reagents Chemicals Structure
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Cross-linker
Zirconium propoxide (ZPO)
Zr O
O O O
H3C
CH3
CH3 H3C
Monomer
Phenyltrimethoxysilane (PTOMS)
Si OCH3
OCH3 OCH3
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3-2 Photonic crystal
3-2-1 Synthesis of monodisperse PS microspheres
Monodisperse PS microspheres were synthesized using an oil-in-water emulsion polymerization technique. The reaction took place in a 50 mL round-bottomed flask equipped with a water-cooled reflux condenser. Figure 3-2 shows the experimental apparatus for the polymerization. Styrene was used as the monomer, while KPS and SDS were used as the initiator and the capping agent, respectively. Initially, 49 mL DI was added to a flask and then purged with nitrogen for 30 min with vigorously stirring at 350 rpm to establish an anoxic condition. Then, 10 mL anoxic water was taken out to prepare a surfactant stock solution through dissolving 0.923 g SDS at 40 critical micellar concentrations (CMCs). 1 mL of SDS stock solution was added into the remaining 39 mL anoxic water to reach 1 CMC. Following, 6.5 mL styrene was added into the SDS solution which had been pre-heated 80oC for 30 min. Polymerization was initiated when 21.6 mg KPS was added into the mixture and the reaction was maintained at 80oC and at 350 rpm for 10 hr. The temperature (± 2oC) was controlled using a thermostatic silicon oil bath. The resulting latex was dialyzed using a dialysis membrane (Spectrum Laboratories, Inc., Molecular porous MWCO: 6-8000) for one week to separate PS from other reactants. The PS microspheres remained suspension in their mother liquor until needed. Figure 3-3 displays the mechanism of polymerization of PS.
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Figure 3-2 Experimental apparatus for the polymerization.
Figure 3-3 Mechanism of oil-in-water emulsion polymerization of PS.
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3-2-2 Fabrication of the opal structure
Three-dimensional colloidal crystals were prepared using a heat-assisted sedimentation on 25 × 25 mm2 glass slides. Figure 3-4 illustrates the formation of PS colloidal crystals.
All the glass substrates were cleaned through ultrasonication for 30 min in acetone, ethanol and DI sequentially. The PS latex solution (10 L) was dropped onto the glass slide.
Afterwards, the slide was placed on a hot plate at temperature 40-70oC to evaporate water from the PS droplet. The PS nanoparticles were driven into a long-range ordered, opaline lattice by the attractive capillary forces generated during water evaporation.
Figure 3-4 Formation of PS array through a heat-assisted sedimentation on a glass substrate.
3-2-3 Fabrication of the inverse opal structure
ZrO2 sol solution was added on the top of the colloidal crystals, the sol solution seeped into the interstitial voids between the PS nanoparticles through capillary and gravitational forces. The ZrO2-PS composites were dried at room temperature for 30 min for solidification. Finally, inverse opal ZrO2 structures were obtained via dissolving the PS with toluene. In order to optimize the volume ratio of PS and imprinted sol solution, the volumes of ZrO2 solution (ZPO/EtOH) were changed from 2.5 to 10 L and the molar ratio of ZrO2
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was controlled at 0.4. Figure 3-5 shows the infiltration of the ZrO2 sol solution between the PS crystals.
Figure 3-5 Scheme of the infiltration of the ZrO2 sol solution between the PS microspheres.
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3-3 Preparation of molecularly imprinted polymer
Molecularly imprinted sol solutions were prepared using a sol-gel method. Figure 3-6 shows the preparation procedure of the molecularly imprinted sol solution. ZPO was used as the precursors of ZrO2 and the cross-linker in the imprinting process. PTMOS and BPA were used as the functional monomer and the template, respectively. Firstly, 0.23 g BPA (1mM) was dissolved in 4.7 mL EtOH with stirring for 10 min at 400 rpm. Then, 0.19 mL PTMOS (1mM) and 13.5 mL ZPO (30mM) were sequentially added into the above solution under stirring at 400 rpm at room temperature for 20 min. The sol solution turned gradually from transparent to light yellow during the addition of ZPO. Non-imprinted sol solutions were prepared following the same procedure except for the absence of BPA.
Figure 3-6 Preparation procedure of the molecularly imprinted sol solution.
The BPA imprinted and non-imprinted ZrO2 gel was obtained after the evaporation of the solvents at 120oC for 24 hr. The final product was crushed and ground into fine powders.
The imprinted BPA molecules were extracted from the ZrO2 matrix by means of immersing the imprinted powders into methanol with stirring for 2 hr. The imprinted ZrO2 powders were then harvested via centrifugation at 15000 rpm for 5 min. The extraction was repeated
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for several times till the BPA adsorption peak ( = 276 nm) in the supernatant became undetectable. Figure 3-7 shows the mechanism of the formation of BPA imprinted ZrO2.
Figure 3-7 Mechanism of the formation of BPA imprinted ZrO2.
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3-4 Fabrication of imprinted photonic crystal (IPC)
Molecularly imprinted sol solution was added on the top of the colloidal crystals, and let it seeping into the interstitial voids between PS nanoparticles through capillary forces. The composites were dried at 40oC for 30 min for solidification. Then, the PS templates were removed by solvent extraction with 20 mL toluene in a 100 mL vial for 2 hr. Finally, the composites were statically immersed in 20 mL methanol for 1 hr to remove the BPA templates to get IPC. For control experiments, non-imprinted photonic crystal (NIPC) was also prepared using the same procedure but without the addition of templates. Figure 3-8 shows the time scale of the preparation process of IPC.
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Figure 3-8 Time scale of the preparation process of IPC.
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3-5 Characterization
3-5-1 Scanning Electron Microscopy (SEM)
The morphology of the samples was characterized using a scanning electron microscopy (SEM, Hitachi S-4700, Tokyo, Japan). The observation was proceeded under AN electron accelerating voltage of 25 kV and a pressure below 3 × 10-6 Pa. Powder samples were prepared suspending in acetone via ultrasonic vibration for 20 min. Then the suspension was directly dropped on the glass and dried at 100oC. The samples were coated with a layer of Au by Ion coater (Eiko IB-2) for 3 min to enhance enough conductivity.
3-5-2 Transmission Electron Microscopy (TEM)
Three-dimensional microstructures of the inverse structures were identified using a high resolution transmission electron microscopy (HR-TEM, Philips, TECNAI 20) at an accelerating voltage of 200 kV. The samples were well dispersed in acetone by ultrasonic for 30 min. Afterwards, the suspension was dropped onto Cu grids (Ted Pella, Inc., 200 Mesh) followed by drying at 100oC.
3-5-3 UV-visible Spectrometry (UV-visible)
The UV-vis diffused reflectance spectra were recorded on an UV-visible spectrometry (Hitachi, U-3010, Japan). All of the samples were reference to aluminum oxide (Al2O3) which was considered to exhibit total reflections. The spectra were recorded from 400 to 200 nm at a scan rate of 120 nm/min with an interval of 0.2 nm.
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3-5-4 Dynamic Light Scattering (DLS)
The average particle sizes of the PS microspheres were determined by dynamic light scattering (DLS, Zetasizer Nano ZS, Malbern, Britian). The instrument was equipped with a 4 MW He-Ne laser operating at wavelength of 633 nm and non-invasive backscatter optics (NIBS). The PS latex was diluted 10X with DI and ultra-sonicated for 30 min. The DLS measurement were carried out using polarized scattered light at a wavelength of 633 nm and 25 ± 0.5oC with scattering angles of 173o. The measurement position within the quartz cuvette was automatically determined by the software.
3-5-5 Fourier Transform Infrared Spectrometer (FTIR)
The surface functional groups of the imprinted particles and non-imprinted particles were characterized using a Fourier transform infrared spectrometer (FTIR, Thermo Scientific Nicolet iS10). All spectra were scanned between 400 and 4000 cm-1 with a resolution of 4 cm-1 for 100 runs.
3-5-6 Specific Surface Area (BET)
The specific surface areas and pore volume of the hybrid materials were analyzed by N2
adsorption technique and calculated using the Brunauer-Emmett-Teller (BET) mode. N2
physisorption and desorption was measured at 77K under variety of relative pressure (p/po) (Micromeritics, Tri Star 3000). 0.2 g of samples was degassed at 120oC for 12 hr before all analysis.
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3-5-7 X-ray photoelectron Spectroscopy (XPS)
The surface chemical compositions of the MIP and IPC were determined by X-ray photoelectron spectroscopy (XPS, ESCA PHI 1600) using an Al Kα X-ray source (1486.6 eV).
The photoelectrons were collected into the analyzer with pass energy (23.5 eV). The collection step size in wide range scan and high-resolution analysis were 1.0 and 0.1 eV, respectively. All analytical process was controlled under ultrahigh vacuum conditions at pressure less than 1.4 × 10-9 Torr. In order to quantify and qualify each element, curve fitting of the XPS spectra was performed. The parameters used for the curve fitting of the Zr 3d, C 1s and Si 2p, including the binding energies, doublet separation, full-width at half maximum and curve area. The integrated peak areas were normalized with atomic sensitive factors to calculate atomic ratios. The equation for atomic ratio calculation is shown below (Eq. 3-1):
Eq. 3-1
n: the atomic numbers
I: the intensity of species on XPS spectra A: the integrated peak area
ASF: the atomic sensitivity factor and Arabic number represents element types
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3-5-8 Adsorption
The rebinding capacities of the imprinted ZrO2 particles were examined via mixing the 25.0 mg samples with 5 mL BPA solutions at 50 mg/L. After stirring for 3 hrs to reach adsorption equilibrium, the powders were separated from the solutions via centrifugation at 15000 rpm for 5 min. The remaining BPA in the supernatants was determined from the intensity of its adsorption at 276 nm using UV-visible spectrometer. The adsorption capacity (Sb, mg/g) were calculated by the following equation (Eq. 3-2):
Eq. 3-2
C0: the template concentrations in the solutions measured initially time (mg/L) Ct: the template concentrations in the solutions measured interval time, t (mg/L) V: the volume of the bulk solution (L)
W: was the weight of the particles (g)
Imprinted factor (α) is defined as the amounts of rebinding capacity of the imprinted sample to that non-imprinted sample and is expressed as the following equation (Eq. 3-3):
Eq. 3-3
Sb (MIP): the BPA binding amount to the imprinted particles (mg/g)
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Sb (NIP): the binding amount to the non-imprinted particles (mg/g)
3-5-9 Selectivity
The recognition capability of the imprinted particles for BPA was examined in terms of adsorptions of analogues including phenol, 1-naphthol and BP. The physicochemical properties of BPA and its analogues were summarized in table 3-2. The adsorption test was carried out by mixing the imprinted particles (25.0 mg) with 50 mg/L of 1-naphthol, BP and phenol solutions (5 mL) separately under stirring for 3 hr. Then, the suspensions were centrifuged at 15000 rpm for 5 min to remove the particles. The adsorbed amounts of the 1-naphthol, BP and phenol were determined by the decrease in the intensities of their absorption peaks at 323, 274, and 270 nm, respectively. The selectivity factors were obtained using the following equation (Eq. 3-4):
Eq. 3-4
Sb (BPA): the BPA binding amount to the imprinted particles (mg/g) Sb (analogous): the binding amount to the analogous molecules (mg/g)
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Table 3-2 Physicochemical properties of compounds used for selective adsorption test.
Compound Structure Aqueous solubility Log Kow
Bisphenol A
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3-6 Sensing capability of imprinted photonic crystal (IPC)
The sensing capabilities of IPC, including detection limit, equilibrium time and selectivity for BPA were examined. The IPC was immersed in 20 mL BPA solution (50 mg/L) for 5 min to reach adsorption equilibrium. After drying with nitrogen gas, the reflection peak of the IPC was measured using the UV-visible spectrometer. According to the Bragg’s law,[51] 2d111(n2efsin2)1/2, the adsorption of BPA changes the mean refractive index, n, of the IPC and consequently changing the diffractive wavelength. The shift in the diffraction peak position with BPA binding can be used to quantitatively estimate the amount of bound BPA. Figure 3-9 shows the detection procedure of BPA using the IPC.
Figure 3-9 Detection process of BPA using the IPC.
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The linear range and detection limit were obtained at the BPA concentrations of 0.01-100 mg/L. Wavelength shifts for 30 mg/L BPA in different intervals of adsorption time were monitored to understand the response kinetics. 1-naphthol, BP and phenol were used as the analogues of BPA to test the selectivity of the IPC toward the target compound under the concentration of 50 mg/L. For completive systems, the phenol, BP and BPA were dissolved in DI to reach 50 mg/L for each one.
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