Thin Molecularly Imprinted Polymer Research Articles

Plasmonic Optical Fiber Sensors and Molecularly Imprinted Polymers for Glyphosate Detection at an Ultra-Wide Range

In this study, a surface plasmon resonance (SPR) sensor based on modified plastic optical fibers (POFs) was combined with a specific molecularly imprinted polymer (MIP), used as a synthetic receptor, for glyphosate (GLY) determination in aqueous solutions. Since GLY is a non-selective herbicide associated with severe environmental and health problems, detecting glyphosate in environmental and biological samples remains challenging. The selective interaction between the MIP layer and GLY is monitored by exploiting the SPR phenomenon at the POF’s gold surface. Experimental results show that in about ten minutes and by dropping microliter volume samples, the presented optical–chemical sensor can quantify up to three orders of magnitude of GLY concentrations, from nanomolar to micromolar, due to a thin MIP layer over the SPR surface. The developed optical–chemical sensor presents a detection limit of about 1 nM and can be used for onsite GLY measurements. Moreover, the experimental analysis demonstrated the high selectivity of the proposed POF-based chemical sensor.

On-Demand, Reversible, Ultrasensitive Polymer Membrane Based on Molecular Imprinting Polymer.

The development of in vivo, longitudinal, real-time monitoring devices is an essential step toward continuous, precision health monitoring. Molecularly imprinted polymers (MIPs) are popular sensor capture agents that are more robust than antibodies and have been used for sensors, drug delivery, affinity separations, assays, and solid-phase extraction. However, MIP sensors are typically limited to one-time use due to their high binding affinity (>107 M-1) and slow-release kinetics (<10-4 μM/sec). To overcome this challenge, current research has focused on stimuli-responsive MIPs (SR-MIPs), which undergo a conformational change induced by external stimuli to reverse molecular binding, requiring additional chemicals or outside stimuli. Here, we demonstrate fully reversible MIP sensors based on electrostatic repulsion. Once the target analyte is bound within a thin film MIP on an electrode, a small electrical potential successfully releases the bound molecules, enabling repeated, accurate measurements. We demonstrate an electrostatically refreshed dopamine sensor with a 760 pM limit of detection, linear response profile, and accuracy even after 30 sensing-release cycles. These sensors could repeatedly detect <1 nM dopamine released from PC-12 cells in vitro, demonstrating they can longitudinally measure low concentrations in complex biological environments without clogging. Our work provides a simple and effective strategy for enhancing the use of MIPs-based biosensors for all charged molecules in continuous, real-time health monitoring and other sensing applications.

Design of molecularly imprinted polymer materials relying on hydrophobic interactions

The design of molecularly imprinted polymers (MIPs) for the specific binding of hydrophobic molecules (mitotane) was investigated by considering monomers and cross-linking agents with variable hydrophobic character, methyl methacrylate, butyl methacrylate and lauryl methacrylate as functional monomers, and ethylene glycol dimethacrylate and 1,6-hexanediol dimethacrylate as cross-linking agents. MIPs were bound to a porous silica support by means of a radical transfer reaction with grafted 3-mercaptopropyl groups. Equilibrium adsorption isotherms to molecularly imprinted and non-imprinted materials were modeled with the Volmer and Langmuir–Volmer isotherms, giving the thermodynamic parameters of adsorption. The most hydrophobic monomer provided the highest selective adsorption although strong non-selective adsorption and the intrinsic softness of poly(lauryl methacrylate) is detrimental to selectivity. Adsorption was exothermic with a predominant enthalpic contribution. Adsorption kinetics were fast due to the high accessibility of molecular imprints on the surface of the thin MIP layer coating the porous silica support.

Molecularly Imprinted Polymer-Coated Optical Waveguide for Attogram Sensing.

Ultrahigh sensitivity and selectivity are the ultimate goals of sensor development. For such purposes, we propose a sensing platform in which an optical fiber-waveguide-fiber (OFWF) structure is integrated with a molecularly imprinted polymer (MIP). The OFWF works as a highly efficient probe light launcher and signal light collector, and the MIP layer acts as a highly selective and sensitive sensing interface. In the MIP design, a high-molecular refractive index monomer (2-phenylphenoxyethyl acrylate) was copolymerized with a MIP functional monomer (acrylic acid). The resulting high-refractive index MIP layers could effectively extract the probe light from the waveguide and send it to the MIP sensing interface. Moreover, a highly elastic cross-linker (poly(ethylene glycol) 600 diacrylate) was employed to increase the MIP mesh size, which could effectively increase the penetrability of the analyte. Rhodamine B (Rh B) is widely used in the textile industry, and its contamination may lead to serious public health problems. As a proof of concept, the Rh B chromophore was used as a molecular template, and the thin MIP layer was cured on the waveguide surface by utilizing the evanescent wave of the 405 nm propagating light in the waveguide. The MIP-OFWF sensing platform afforded highly selective monitoring of the absorption spectra of the components in a mixture solution of Rh B and methyl blue. It also afforded an extremely low detection limit of approximately 6.5 × 10-17 g/mL, with an absolute mass of 20-30 ag.

Micro-gel thin film molecularly imprinted polymer coating for extraction of organophosphorus pesticides from water and beverage samples

Molecularly imprinted polymers (MIPs) have become an important class of materials for selective and efficient adsorption of target analytes. Despite versatility of MIPs for fabrication in numerous formats, these materials have been primarily reported as solid phase extraction packing materials. An effective thin film MIP prepared on stainless steel substrate is reported here for high throughput enrichment of organophosphorus pesticides (OPPs) from water and beverage samples followed by liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis. The key factors controlling performance as well as best practices for optimized fabrication of thin film MIPs are presented. A pseudo-phase diagram is introduced to evaluate and predict the effect of the ratio of porogen (solvent, 1-octanol) volume to relative crosslinker mass on the desired polymer features (i.e., porosity, surface area, capacity, and selectivity). At low porogen ratios, a macroporous polymer with insignificant selectivity is formed, whereas at high porogen ratios a micro-gel polymer with superior selectivity towards targets is obtained. The porosity and morphology determined with nitrogen adsorption and scanning electron microscopy were attributed to specific regions in the pseudo-phase diagram. Other factors influencing selectivity and stability of the polymer, such as type of the template and its ratios with monomer (methacrylic acid) and crosslinker (ethylene glycol dimethacrylate) were optimized. The prepared thin film MIPs were characterized using adsorption isotherms and adsorption kinetics, and evaluated for matrix effects (high humic acid content) and cross-reactivity in presence of other pesticides and pharmaceuticals. The optimized method provided limits of quantitation (LOQs) ranged from 0.002 to 0.02 ng mL−1 in water and from 0.095 to 0.48 ng g−1 in apple juice. Regarding inter-device variability (CV∼10% without normalization), excellent linearity (R2 > 0.99), satisfactory accuracies (90–110%) and precisions (<15%) were obtained.

Thin Film Plastic Antibody-Based Microplate Assay for Human Serum Albumin Determination.

Herein we demonstrate molecularly imprinted polymers (MIP) as plastic antibodies for a microplate-based assay. As the most abundant plasma protein, human serum albumin (HSA) was selected as the target analyte model. Thin film MIP was synthesized by the surface molecular imprinting approach using HSA as the template. The optimized polymer consisted of acrylic acid (AA) and N-vinylpyrrolidone (VP) in a 2:3 (w/w) ratio, crosslinked with N,N′-(1,2-dihydroxyethylene) bisacrylamide (DHEBA) and then coated on the microplate well. The binding of MIP toward the bound HSA was achieved via the Bradford reaction. The assay revealed a dynamic detection range toward HSA standards in the clinically relevant 1–10 g/dL range, with a 0.01 g/dL detection limit. HSA-MIP showed minimal interference from other serum protein components: γ-globulin had 11% of the HSA response, α-globulin of high-density lipoprotein had 9%, and β-globulin of low-density lipoprotein had 7%. The analytical accuracy of the assay was 89–106% at the 95% confidence interval, with precision at 4–9%. The MIP-coated microplate was stored for 2 months at room temperature without losing its binding ability. The results suggest that the thin film plastic antibody system can be successfully applied to analytical/pseudoimmunological HSA determinations in clinical applications.

How Reliable Is the Electrochemical Readout of MIP Sensors?

Electrochemical methods offer the simple characterization of the synthesis of molecularly imprinted polymers (MIPs) and the readouts of target binding. The binding of electroinactive analytes can be detected indirectly by their modulating effect on the diffusional permeability of a redox marker through thin MIP films. However, this process generates an overall signal, which may include nonspecific interactions with the nonimprinted surface and adsorption at the electrode surface in addition to (specific) binding to the cavities. Redox-active low-molecular-weight targets and metalloproteins enable a more specific direct quantification of their binding to MIPs by measuring the faradaic current. The in situ characterization of enzymes, MIP-based mimics of redox enzymes or enzyme-labeled targets, is based on the indication of an electroactive product. This approach allows the determination of both the activity of the bio(mimetic) catalyst and of the substrate concentration.

Selective PQQPFPQQ Gluten Epitope Chemical Sensor with a Molecularly Imprinted Polymer Recognition Unit and an Extended-Gate Field-Effect Transistor Transduction Unit.

A molecularly imprinted polymer (MIP) recognition system was devised for selective determination of an immunogenic gluten octamer epitope, PQQPFPQQ. For that, a thin MIP film was devised, guided by density functional theory calculations, and then synthesized to become the chemosensor recognition unit. Bis(bithiophene)-based cross-linking and functional monomers were used for this synthesis. An extended-gate field-effect transistor (EG-FET) was used as the transduction unit. The EG-FET gate surface was coated with the PQQPFPQQ-templated MIP film, by electropolymerization, to result in a complete chemosensor. X-ray photoelectron spectroscopy analysis confirmed the presence of the PQQPFPQQ epitope, and its removal from the MIP film. The chemosensor selectively discriminated between the octamer analyte and another peptide of the same number of amino acids but with two of them mismatched (PQQQFPPQ). The chemosensor was validated with respect to both the PQQPFPQQ analyte and a real gluten extract from semolina flour. It was capable to determine PQQPFPQQ in the concentration range of 0.5-45 ppm with the limit of detection (LOD) = 0.11 ppm. Moreover, it was capable of determining gluten in real samples in the concentration range of 4-25 ppm with LOD = 4 ppm, which is a value sufficient for discriminating between gluten-free and non-gluten-free food products. The gluten content in semolina flour determined with the chemosensor well correlated with that determined with a commercial ELISA gluten kit. The Langmuir, Freundlich, and Langmuir-Freundlich isotherms were fitted to the epitope sorption data. The sorption parameters determined from these isotherms indicated that the imprinted cavities were quite homogeneous and that the epitope analyte was chemisorbed in them.

Molecularly imprinted polymer-based electrochemical sensors for biopolymers

Abstract Electrochemical synthesis and signal generation dominate among the almost 1200 articles published annually on protein-imprinted polymers. Such polymers can be easily prepared directly on the electrode surface, and the polymer thickness can be precisely adjusted to the size of the target to enable its free exchange. In this architecture, the molecularly imprinted polymer (MIP) layer represents only one ‘separation plate’; thus, the selectivity does not reach the values of ‘bulk’ measurements. The binding of target proteins can be detected straightforwardly by their modulating effect on the diffusional permeability of a redox marker through the thin MIP films. However, this generates an ‘overall apparent’ signal, which may include nonspecific interactions in the polymer layer and at the electrode surface. Certain targets, such as enzymes or redox active proteins, enables a more specific direct quantification of their binding to MIPs by in situ determination of the enzyme activity or direct electron transfer, respectively.

Magnetic Field-Induced Polymerization of Molecularly Imprinted Polymers

In this work, we developed a novel approach for the preparation of molecularly imprinted polymer (MIP) coatings directly onto magnetic multicore nanoparticles (MMCs) using alternating magnetic fields to trigger the polymerization reaction. MIPs were synthesized with rhodamine 123 (R123) as model template molecule, methacrylic acid as functional monomer, and trimethylolpropane trimethacrylate as cross-linker. The amount of iron oxide nanoparticles and the composition of the polymerization mixture were optimized to enable the thermal polymerization of a thin MIP shell on each MMC using electromagnetic heating without altering the properties of the recognition layer. The thickness of the polymerized MIP layer grafted onto the MMCs was fine-tuned by adjusting the dose of electromagnetic field (101.4 kHz, total power dissipation = 105 W). The resulting magnetic multicore MIP nanoparticles (MMC-MIPs) were characterized by Fourier transform infrared and X-ray diffraction spectroscopy, transmission electron micro...

Integrating fluorescent molecularly imprinted polymer (MIP) sensor particles with a modular microfluidic platform for nanomolar small-molecule detection directly in aqueous samples

Fluorescent sensory MIP (molecularly imprinted polymer) particles were combined with a droplet-based 3D microfluidic system for the selective determination of a prototype small-molecule analyte of environmental concern, 2,4-dichlorophenoxyacetic acid or 2,4-D, at nanomolar concentration directly in water samples. A tailor-made fluorescent indicator cross-linker was thus designed that translates the binding event directly into an enhanced fluorescence signal. The phenoxazinone-type cross-linker was co-polymerized into a thin MIP layer grafted from the surface of silica microparticles following a RAFT (reversible addition-fragmentation chain transfer) polymerization protocol. While the indicator cross-linker outperformed its corresponding monomer twin, establishment of a phase-transfer protocol was essential to guarantee that the hydrogen bond-mediated signalling mechanism between the urea binding site on the indicator cross-linker and the carboxylate group of the analyte was still operative upon real sample analysis. The latter was achieved by integration of the fluorescent core-shell MIP sensor particles into a modular microfluidic platform that allows for an in-line phase-transfer assay, extracting the analyte from aqueous sample droplets into the organic phase that contains the sensor particles. Real-time fluorescence determination of 2,4-D down to 20nM was realized with the system and applied for the analysis of various surface water samples collected from different parts of the world.

A NIR-responsive up-conversion nanoparticle probe of the NaYF4:Er,Yb type and coated with a molecularly imprinted polymer for fluorometric determination of enrofloxacin

The paper describes the preparation of a fluorescent probe based on the use of up-conversion particles (UCPs) of the type NaYF4: Er, Yb and shelled with a molecularly imprinted polymer (MIP) that is specific for the antibiotic enrofloxacin. The upconverted green luminescence emitted by the UCPs upon photoexcitation at 980 nm was used to locally generate a thin MIP shell around the UCPs via photopolymerization. Enrofloxacin is used as the template, and ethylene glycol dimethacrylate (EGDMA) as the cross-linking agent. It is found that enrofloxacin is well captured by the MIP and then quenches the fluorescence of the UCPs. Fluorescence decreases linearly in the 63 ng·L−1 to 60 μg·L−1 enrofloxacin concentration range, with a 8 ng·L−1 limit of detection. The method was applied to the determination of enrofloxacin in spiked fish samples. Recoveries ranged from 83.7 to 87.7%, with relative standard deviations between 2.0 and 5.5% (for n = 3).

Conducting Molecularly Imprinted Polymer (MIP) Chemical Sensors for Toxic N-Nitrosamines Selective Determination in Heat Processed Food of Animal Origin

High-temperature processing of meat food generates in it toxic heteroaromatic amines and nitrosamines (NAs)1,2 causing chronic diseases including hormonal dysfunctions and cancer.3 Therefore, we devised and fabricated chemical sensors for selective determination of the N-nitroso-L-proline (Pro-NO)4 and N-nitroso-thiazolidine-4-carboxylic acid (NT4A) toxins in this food. We applied films of conducting polymers molecularly imprinted (MIPs) with either the Pro-NO or NT4A template as recognition units of these chemosensors. For selection of most appropriate functional monomers, we used DFT at the B3LYP/3-21G(*) level modeling to find phenolic derivative of bis(bithiophene) and a thiosalenCo(III) complex (CS)5,6 suitable to form pre-polymerization complexes with Pro-NO and NT4A, respectively, at the 1:2 template/monomer stoichiometry. The calculated Gibbs free energy gain due to formation of these complexes with a series of NAs was proportional to the UV-vis spectroscopy determined stability constants of these complexes. These constants were relatively high equalling 3.14×109 M-2 and 2.72×1010 M-2for Pro-NO and NT4A, respectively. Then, we electrochemically polymerized these complexes under potentiodynamic conditions. This electropolymerization resulted in depositing thin MIP films on different electrode substrates. After subsequent extraction of the Pro-NO and NT4A templates from MIPs, we used the resulting MIPs with empty imprinted cavities for determination of Pro-NO and NT4A analytes in grilled pork neck samples. We confirmed the presence and then absence of NA templates in the MIP films before and after extraction, respectively, with FTIR spectroscopy and XPS, and then characterized morphology of the films with AFM. For analytical signal transduction, we used piezoelectric microgravimetry (PM) at the Au-QCR/MIP electrodes of EQCM as well as DPV and EIS at the Pt/MIP electrodes. Limits of detection of the chemosensors were 36.9 nM Pro-NO (with EIS) and 46 nM NT4A (with DPV). The chemosensors selectively responded to the target toxins in the presence of interfering compounds of similar composition and structure. Successful selective determination of Pro-NO and NT4A in the food extract samples indicates that the chemosensors are promising as tools for determination of these toxins in food of animal origin.

A novel quartz crystal microbalance sensor array based on molecular imprinted polymers for simultaneous detection of clenbuterol and its metabolites

For the rapid and robust detection of both parent clenbuterol (CLB) and its metabolites in swine urine samples, a novel quartz crystal microbalance (QCM) sensor array for CLB detection based on molecularly imprinted polymers (MIPs) was developed in this investigation. At first, clenbuterol and the structural analogs of its metabolites, 4-Aminohippuric acid (AHA) and 4-hydroxymandelic acid (HMA), were chosen as molecular templates. Through computational molecular modeling, the optimum ratio between the functional monomer and molecular template was selected. The surface imprinting method was applied to modify QCM electrode surface to graft a thin MIP film. The grafting polymer was characterized by Fourier-transformed infrared spectrometry (FTIR) and atomic force microscopy (AFM), respectively. After then, an array system composed of three sensors was employed to test the responses with different solutions and the principal component analysis (PCA) was adopted to analyze the corresponding data. As a result, for the designed sensor to clenbuterol, a linear equation y=100.07x-722.96 (R2=0.9928) was found between the sensor frequency shift ΔF and negative logarithm of clenbuterol concentration (-lgC). The limitation of detection (LOD) was 3.0ng/mL, which is lower than the Codex Alimentarius Commission regulations residue limit 10μg/L. The corresponding data of the three template solutions were analyzed by PCA, obtaining 100% recognition. The result demonstrated the feasibility that the developed method could be applied to detect whether the livestock was feed with CLB nutrient redistribution agent by checking the urine samples.

Molecularly Imprinted Mesoporous SBA-15 Synthesized by Surface-Initiated Atom Transfer Radical Polymerization for Bisphenol A Recognition

Ordered mesoporous material has good application potential in solid-phase extractions due to its high surface area and rapid binding kinetics. Molecularly imprinted mesoporous SBA-15 microspheres (MIP-SBA-15) with both advantages of molecularly imprinted polymer (MIP) and mesoporous material have been synthesized in the study. The surface-initiated atom transfer radical polymerization was employed in the imprinting process in order to create more homogeneous polymer layer on the surface of SBA-15. The bisphenol A (BPA) was used as imprinting template to provide selective extraction material with potential application in the BPA determinations. The results demonstrated the spherical MIP-grafted-SBA-15 with ordered mesoporous structure has been synthesized which is good for chromatographic applications. The material has good selectivity, high binding capacity and kinetics. The BPA and its analogs have less peak tailings on the MIP column in HPLC, which is attributed to the thin MIP film and less mass transfer resistance of the material. An online solid-phase extraction/HPLC experiment was established using the MIP-SBA-15 as the extraction material. In the BPA concentration range of 0.04–40 ng mL−1, a linear regression curve with R 2 > 0.9999 has been obtained. The LOD and LOQ are 30.84 and 102.79 pg mL−1, respectively. The recoveries were higher than 97 % determined using spiked tap water samples with different concentrations. It demonstrated the MIP-SBA-15 can be used in the BPA extraction from water samples with good efficiency.

Changes in the Porosity and Permeability of a Molecularly Imprinted Membrane Induced by the Adsorption of a Trace Quantity of Template

It is known that the diffusive permeability of solutes within a thin layer of molecularly imprinted polymer (MIP) may be affected by specific binding of the MIP with its template molecule. This phenomenon, termed the gate effect, shows promise for the development of novel biomimetic sensors. However, the mechanism underlying this effect is still unclear; although the relationship between the specific adsorption of a template and the corresponding porosity and permeability of the polymeric film or membrane is very important, this association has not yet been examined in detail. We therefore studied this relationship using a molecularly imprinted self-supporting membrane (MISSM) possessing chiral specificity, specially developed as a tool for investigating the gate effect. Both the diffusive permeability and volume porosity of the MISSM were sensitive to the presence of the template compounds (D and L-phenylalanine) at concentrations as low as 5 µM, while, at the same time, insensitive to the enantiomer of the template. The relationship between the amount of adsorbed template and the equilibrium template concentration followed the expected Langmuir isotherm pattern, which indicates the thermodynamic homogeneity of binding sites in the MISSM. This study also demonstrated that the relative concentration of the adsorbed template in the membrane was only 3 ppm and relative site occupation was only 1% following exposure to a 5 µM concentration of the template. These results show that the gate effect may be advantageously exploited during application of MIPs in amplifiers or sensors offering high sensitivity. Molecularly imprinted polymers (MIPs) are synthetic polymers that contain specific binding sites formed by imprinting of a target molecule (or template) during the polymerization process. An MIP layer can be prepared by a simple and economical procedure (1-3). Before such MIPs may be employed as molecular recognition elements in chemical-sensing devices, however, it is necessary to develop a means of translating specific binding events at the MIP into an electric signal. The so-called gate effect, which refers to changes in the diffusive permeability of solutes within the MIP layer resulting from specific binding at the template, can be used as a mechanism for signal transference based on conductometry, utilizing changes in the ionic permeability of the MIP membrane associated with a specific interaction with the template (4-6). In this gate analogy, the template corresponds to the key while the MIP site which allows specific rebinding with the template corresponds to the keyhole. An amperometric method is also applicable, using a thin MIP layer grafted onto an electrode, in which the template can be detected by following changes in the faradic current resulting from the change in permeability of a redox marker across the MIP layer (7-11). The gate effect is a selective process capable of discriminating between

The complete replacement of antibodies by protein-imprinted poly(ethylene-co-vinyl alcohol) in sandwich fluoroimmunoassays

The replacement of antibodies by molecularly imprinted polymers (MIPs) has been investigated for many decades. However, indirect protocols (including natural primary and secondary antibodies) are still utilized to evaluate the ability of MIP thin films to recognize target molecules. MIPs can be prepared as either a thin film or as particles, and cavities that are complementary to the template can be generated on their surfaces. We have prepared thin film MIPs and particle MIPs prepared by solvent evaporation and phase inversion, respectively, from solutions of poly(ethylene-co-vinyl alcohol) (pEVAL) in the presence of the target analytes amylase, lysozyme, and lipase. These were first adsorbed on MIP thin films and by MIP particles that contain fluorescent quantum dots. Sandwich fluoroimmunoassays were then conducted to quantify them in MIP-coated 96-well microplates. The method was applied to determine amylase in saliva, and results were compared with a commercial analytical system.

Colorimetric test-systems for creatinine detection based on composite molecularly imprinted polymer membranes

An easy-to-use colorimetric test-system for the efficient detection of creatinine in aqueous samples was developed. The test-system is based on composite molecularly imprinted polymer (MIP) membranes with artificial receptor sites capable of creatinine recognition. A thin MIP layer was created on the surface of microfiltration polyvinylidene fluoride (PVDF) membranes using method of photo-initiated grafting polymerization. The MIP layer was obtained by co-polymerization of a functional monomer (e.g. 2-acrylamido-2-methyl-1-propanesulfonic acid, itaconic acid or methacrylic acid) with N, N′-methylenebisacrylamide as a cross-linker. The choice of the functional monomer was based on the results of computational modeling. The creatinine-selective composite MIP membranes were used for measuring creatinine in aqueous samples. Creatinine molecules were selectively adsorbed by the MIP membranes and quantified using color reaction with picrates. The intensity of MIP membranes staining was proportional to creatinine concentration in an analyzed sample. The colorimetric test-system based on the composite MIP membranes was characterized with 0.25mM detection limit and 0.25–2.5mM linear dynamic range. Storage stability of the MIP membranes was estimated as at least 1 year at room temperature. As compared to the traditional methods of creatinine detection the developed test-system is characterized by simplicity of operation, small size and low cost.

Protein Crystallization and Biosensor Applications of Hydrogel-Based Molecularly Imprinted Polymers

We have characterized the imprinting capability of a family of acrylamide polymer-based molecularly imprinted polymers (MIPs) for bovine hemoglobin (BHb) and trypsin (Tryp) using spectrophotometric and quartz crystal microbalance (QCM) sensor techniques. Bulk gel characterization on acrylamide (AA), N-hydroxymethylacrylamide (NHMA), and N-isopropylacrylamide (NiPAM) gave varied selectivities when compared with nonimprinted polymers. We have also harnessed the ability of the MIPs to facilitate protein crystallization as a means of evaluating their selectivity for cognate and noncognate proteins. Crystallization trials indicated improved crystal formation in the order NiPAM<AA<NHMA. QCM studies of thin film MIPs confirm this trend with N-hydroxymethyl acrylamide MIPs exhibiting best discrimination between MIP and NIP and also cognate/noncognate protein loading. Equivalent results for acrylamide MIPs suggested that the cavities were equally selective for both proteins, while N-isopropylacrylamide MIPs were not selective for either cognate BHb or noncognate BSA. All BHb MIP-QCM sensors based on AA, NHMA, or NiPAM were essentially nonresponsive to smaller, noncognate proteins. Protein crystallization studies validated the hydrophilic efficacy of MIPS indicated in the QCM studies.

Thin Walled Imprinted Polymer Beads Featuring Both Uniform and Accessible Binding Sites

A novel approach addressing the classical deficiencies of molecularly imprinted polymers (MIPs), that is, low binding capacity and nonuniform binding sites, is reported. The thin walled beads were produced in two steps by first grafting thin MIP films, under controlled (RAFT) or noncontrolled conditions, from porous silica beads following previously reported procedures. The resulting composites were compared in terms of film thickness, the grafted layer homogeneity, the effect of different support morphologies, and for their ability to recognize the template in chromatographic or static binding tests. Thus, using l-Phenylalanine anilide (L-PA) as template to imprint poly(MAA-co-EDMA) in such a way led to nanometer thick films where the resulting composite were able to selectively retain the template in relation to the thickness of the grafted film. In the second step, removing the silica supports from the above composites by etching, led to nanometer thin walled beads with structure, morphology and recognition properties strongly depending on grafting chemistry (RAFT or non-RAFT), monomer dilution and on the film thickness of the original composite. Thus whereas the thicker walled materials retained their mesoporous morphology and displayed enhanced enantioselectivity, load capacity, and higher surface areas compared to their composite precursors, the thin walled beads showed lower surface areas indicating network collapse. The thin walled beads prepared under dilute conditions in absence of RAFT displayed a perfectly uniform binding site distribution and a saturation capacity exceeding that of a conventional monolithic MIP. The beads prepared by RAFT control showed a further enhanced saturation capacity significantly exceeding that of the reference material. Finally, the reduced hydrophobic character of the thin walled materials indicated the existence of two separate pore systems with different pore wettabilities.