‘Bite-and-Switch’ approach using computationally designed molecularly imprinted polymers for sensing of creatinine

A method for the selective detection of creatinine is reported, which is based on the reaction between polymerised hemithioacetal, formed by allyl mercaptan, o-phthalic aldehyde, and primary amine leading to the formation of fluorescent isoindole complex. This method has been demonstrated previously for the detection of creatine using creatine-imprinted molecularly imprinted polymers (MIPs) Since MIPs created using traditional methods were unable to differentiate between creatine and creatinine, a new approach to the rational design of a molecularly imprinted polymer (MIP) selective for creatinine was developed using computer simulation. A virtual library of functional monomers was assigned and screened against the target molecule, creatinine, using molecular modelling software. The monomers giving the highest binding score were further tested using simulated annealing in order to mimic the complexation of the functional monomers with template in the monomer mixture. The result of this simulation gave an optimised MIP composition. The computationally designed polymer demonstrated superior selectivity in comparison to the polymer prepared using traditional approach, a detection limit of 25 μM and good stability. The ‘Bite-and-Switch’ approach combined with molecular imprinting can be used for the design of assays and sensors, selective for amino containing substances.     

Combinatorial screening of polymer precursors for preparation of benzo[α] pyrene imprinted polymer: an ab initio computational approach

A combinatorial screening procedure was used for the selection of polymer precursors in the preparation of molecularly imprinted polymer (MIP), which is useful in the detection of the air pollution marker molecule benzo[a]pyrene (BAP). Molecular imprinting is a technique for the preparation of polymer materials with specific molecular recognition receptors. The preparation of imprinted polymers requires polymer precursors such as functional monomer, cross-linking monomer, solvent, an initiator of polymerization and thermal or UV radiation. A virtual library of functional monomers was prepared based on interaction binding scores computed using HyperChem Release 8.0 software. Initially, the possible minimum energy conformation of the monomers and BAP were optimized using the semi-empirical (PM3) quantum method. The binding energy between the functional monomer and the template (BAP) was computed using the Hartree-Fock (HF) method with 6-31 G basis set, which is an ab initio approach based on Moller-Plesset second order perturbation theory (MP2). From the computations, methacrylic acid (MAA) and ethylene glycol dimethacrylate (EGDMA) were selected for preparation of BAP imprinted polymer. The larger interaction energy (ΔE) represents possibility of more affinity binding sites formation in the polymer, which provides high binding capacity. The theoretical predictions were complimented through adsorption experiments. There is a good agreement between experimental binding results and theoretical computations, which provides further evidence of the validity of the usefulness of computational screening procedures in the selection of appropriate MIP precursors in an experiment-free way.     

Novel chiral recognition elements for molecularly imprinted polymer preparation

The use of a novel chiral functional monomer system in molecular imprinting protocols is described. The monomer, dibenzyl (2R,3R)-O-monoacryloyl tartrate, possesses a hydroxyl moiety which can be used to direct template-functional monomer interactions during molecular imprinting polymerization. This system simultaneously positions benzyl ester-protected carboxyl groups in close proximity to the template, which upon deprotection yield recognition sites with stronger ligand-binding capacities. Furthermore, the inherent chirality of the monomer engenders the polymer with an inbuilt preference for a given stereoisomer. Application of the system to the molecular imprinting of the cinchonidine alkaloids (+)-cinchonine and (-)-cinchonidine yielded stereoselective polymers. The effect of imprinting (+)-cinchonine produced a polymer which more than reversed the inherent chiral selectivity of the chiral monomer residues present in the matrix.     

A computational approach to study functional monomer‐protein molecular interactions to optimize protein molecular imprinting

Molecular imprinting has become a promising approach for synthesis of polymeric materials having binding sites with a predetermined selectivity for a given analyte, the so‐called molecularly imprinted polymers (MIPs), which can be used as artificial receptors in various application fields. Realization of binding sites in a MIP involves the formation of prepolymerization complexes between a template molecule and monomers, their subsequent polymerization, and the removal of the template. It is believed that the strength of the monomer‐template interactions in the prepolymerization mixture influences directly on the quality of the binding sites in a MIP and consequently on its performance. In this study, a computational approach allowing the rational selection of an appropriate monomer for building a MIP capable of selectively rebinding macromolecular analytes has been developed. Molecular docking combined with quantum chemical calculations was used for modeling and comparing molecular interactions among a model macromolecular template, immunoglobulin G (IgG), and 1 of 3 electropolymerizable functional monomers: m‐phenylenediamine (mPD), dopamine, and 3,4‐ethylenedioxythiophene, as well as to predict the probable arrangement of multiple monomers around the protein. It was revealed that mPD was arranged more uniformly around IgG participating in multiple H‐bond interactions with its polar residues and, therefore, could be considered as more advantageous for synthesis of a MIP for IgG recognition (IgG‐MIP). These theoretical predictions were verified by the experimental results and found to be in good agreement showing higher binding affinity of the mPD‐based IgG‐MIP toward IgG as compared with the IgG‐MIPs generated from the other 2 monomers.     

Effective removal of rhodamine B from contaminated water using non-covalent imprinted microspheres designed by computational approach

Molecular dynamics simulations and computational screening were used to identify functional monomers capable of interacting with rhodamine B (RhB). A library of 24 kinds of common functional monomers for preparing molecular imprinted polymer (MIP) was built and their interactions with RhB in acetonitrile were calculated using the molecular dynamics software (Gromacs 3.3). It was anticipated that the monomers giving the highest binding energy are suitable for preparing the affinity polymers. According to the theoretical calculation results, the MIP microspheres with RhB as template was prepared by reverse microemulsion polymerization method using acrylamide (AAm) as functional monomer and divinylbenzene as cross-linker in acetonitrile. Microspheres have been characterized by scanning electron microscopy (SEM). The proper adsorption and selective recognition ability of the MIP were studied by an equilibrium-adsorption method. The MIP showed outstanding affinity towards RhB in aqueous solution and the optimum pH value for binding has been found around neutral range. The molecular recognition of RhB was analyzed in detail by using molecular modeling software (Gaussian03). In addition, the MIP reusability without obviously deterioration in performance was demonstrated at least five repeated cycles.     

N-isopropylacrylamide as a functional monomer for noncovalent molecular imprinting

Although N-isopropylacrylamide (NIPAM) has previously been used in molecular imprinting, it has mostly been considered as an 'inert' monomer, or included for its temperature-responsive nature, rather than as a functional monomer responsible for the interactions with the template at the recognition site. A comparative study of NIPAM and other traditional, functional monomers for the imprinting of a hydrogen bond donor template, bisphenol A (BPA), is reported. Nuclear magnetic resonance titration data suggest that NIPAM forms a stronger complex with BPA than either acrylamide or methacrylic acid but a weaker complex than vinylpyridine. Molecular imprinted polymers (MIPs) were prepared using each functional monomer and compared as stationary phases for the separation of BPA from structural analogues. The NIPAM-containing MIP bound BPA with better selectivity than those prepared using acrylamide or methacrylic acid. Using NIPAM also reduces the nonspecific binding, which is found with MIPs using vinylpyridine as functional monomer.     

Rational synthesis of pindolol imprinted polymer by non-covalent protocol based on computational approach

Pindolol (PDL) is a potent and specific adrenoreceptor blocking agent. It is widely used in the treatment of hypertension, cardiac arrhythmia and angina pectoris. Molecularly imprinted polymers (MIPs) are synthetic receptors having potential applications in drug delivery systems and devices such as diagnostic sensors. In the present work, ab initio quantum mechanical simulations and computational screening were used to identify functional monomer having best interactions with PDL. A virtual library of 16 functional monomers was built and the possible minimum energy conformation of the monomers and PDL were calculated using Hartree-Fock (HF) method for the synthesis of PDL imprinted polymer. The interaction energy between functional monomer and the template were corrected by means of basis set superposition error (BSSE) in all pre-polymerization complexes. The hydrogen bonding between PDL and functional monomer was evaluated by changes in bond lengths before and after complex formation. The virtual template-monomer complex with highest interaction energy is more stable during the polymerization and leads to high selectivity and specificity toward the template. The interaction energy of PDL was found to be the highest with itaconic acid followed by 4-vinyl pyridine and least with acrylonitrile. Taking a spectroscopic viewpoint, results obtained from analysis of the harmonic infrared spectrum were examined. Red and blue shifts related to the stretching frequencies of either donors or acceptors of protons were identified and compared experimentally. Stoichiometric mole ratio of template to functional monomer was optimized and confirmed by UV visible spectra titrations. The theoretical results were correlated by evaluation of binding parameters of MIPs. The experimental binding results were in good agreement with theoretical computations.     

Molecular imprinting of nitrophenol and hydroxybenzoic acid isomers: effect of molecular structure and acidity on imprinting

Three nitrophenol isomer-imprinted polymers were prepared under the same conditions using 4-vinylpyridine as a functional monomer. Different recognition capacities for template molecules were observed for the three polymers. Another imprinting system with stronger acidity than nitrophenol isomers, 2-hydroxybenzoic acid (salicylic acid) and 4-hydroxybenzoic acid, was imprinted using 4-vinylpyridine or acrylamide as functional monomer respectively. Both 4-hydroxybenzoic acid-imprinted polymers using the two monomers showed recognition ability for the template molecule. However, when acrylamide was chosen as functional monomer, the salicylic acid-imprinted polymer showed very weak recognition for the template molecule, whereas strong recognition ability of the resultant polymer for salicylic acid was observed with 4-vinylpyridine as functional monomer. It seems that the structure and acidity of template molecules is responsible for the difference in recognition, by influencing the formation and strength of interaction between template molecule and functional monomer during the imprinting process. An understanding of the mechanism of molecular imprinting and molecular recognition of MIPs will help to predict the selectivity of MIPs on the basis of template molecule properties.     

Validation of computational approach to study monomer selectivity toward the template Gallic acid for rational molecularly imprinted polymer design

Gallic acid (GA) is important for pharmaceutical industries as an antioxidant. It also finds use in tanning, ink dyes and manufacturing of paper. Molecularly imprinted polymers (MIP), which are tailor made materials, can play an excellent role in separation of GA from complex matrices. Molecular recognition being the most important property of MIP, the present work proposes a methodology based on density functional theory (DFT) calculations for selection of suitable functional monomer for a rational design of MIP with a high binding capacity for GA. A virtual library of 18 functional monomers was created and screened for the template GA. The prepolymerization template-monomer complexes were optimized at B3LYP/6-31G(d) model chemistry and the changes in the Gibbs free energy (ΔG) due to complex formation were determined on the optimized structures. The monomer with the highest Gibbs free energy gain forms most stable complex with the template resulting in formation of more selective binding sites in the polymeric matrix in MIPs. This can lead to high binding capacity of MIP for GA. Amongst the 18 monomers, acrylic acid (AA) and acrylamide (AAm) gave the highest value of ΔG due to complex formation with GA. 4-vinyl pyridine (4-Vp) had intermediate value of ΔG while, methyl methacrylate (MMA) gave least value of ΔG due to complex formation with GA. Based on this study, the MIPs were synthesized and rebinding performance was evaluated using Langmuir-Freundlich model. The imprinting factor for AA and AAm based MIPs were 5.28 and 4.80 respectively, 4-Vp based MIP had imprinting factor of 2.59 while MMA based MIP exhibited an imprinting factor of 1.95. The experimental results were in good agreement with the computational predictions. The experimental data validated the DFT based computational approach.     

Insight into molecular imprinting in precipitation polymerization systems using solution NMR and dynamic light scattering

Molecular imprinting is a powerful synthetic technique for generating template-defined binding sites in cross-linked polymers. One scientific challenge in molecular imprinting research is to understand the intermolecular interactions leading to molecular complexation and the process of binding site formation during polymerization. In this work, we present a novel method for studying the molecular imprinting process in precipitation polymerization systems. This method employs solution (1) H NMR and dynamic light scattering (DLS) to investigate the association of template molecules with colloidal particles and the dynamic process of particle growth. Under precipitation polymerization conditions, the colloidal particles formed did not interfere with NMR signals from the soluble components, allowing unreacted monomers and free template to be easily quantified. To examine the process of particle nucleation and growth, DLS was used to measure the hydrodynamic particle size at different reaction times. To corroborate the interpretation of the NMR and DLS results, imprinted nanoparticles were collected at different reaction times and their binding characteristics were evaluated using radioligand-binding analysis. Our experimental results provide new insights into the molecular imprinting process that will be useful in the development of new imprinted nanoparticles.     

Chiral separation using molecularly imprinted heteroaromatic polymers

Novel molecularly imprinted polymer systems utilizing 4-vinylpyridine and 1-vinylimidazole as functional monomers have been developed for enantioselective recognition of carboxylic and N-protected amino acids. Non-covalent interactions between the functional monomers and the template molecules were the source of the subsequent recognition sites in the resultant polymers. The capacity of the polymers for molecular recognition was investigated by using them as stationary phases in the HPLC mode. Polymers prepared with 4-vinylpyridine were found to be more efficient in racemic resolution than those prepared with 1-vinylimidazole. When applying a racemic mixture of the template molecule, the polymers showed highest affinity for the enantiomer used as template. Imprints of a racemic template molecule, as expected, did not exhibit enantioselectivity. The optimal molar ratio of 4-vinylpyridine to the template Cbz-L -Asp-OH in the polymerization mixture was determined to be 12:1. In addition to enantioselectivity, the investigated polymers demonstrated ‘ligand selectivity’ e.g., a Cbz-L -Asp-OH-imprinted polymer was able to separate Cbz-D ,L -Asp-OH, but was unable to separate Cbz-D ,L -Glu-OH.     

Enzyme-based molecular imprinting with metals

Aromatic monomers with various functional groups were utilized in horseradish peroxidase-catalyzed polymerization reactions with metal ions Cu(II), Ni(II), and Fe(III) as imprinting templates. The approach described combines molecular imprinting with enzymatic free radical coupling. Selectivity in metal ion affinity between the various polymer products was assessed and found to depend on the metal used in the imprinting process using aniline, tyramine, and phenol as monomers. Selectivity in binding metals was found when polymers imprinted with copper, nickel, or iron were screened against the three metals, with preference for the metal used in the imprinting step. A model for the structural features of the putative imprinted polymers is proposed based on electron paramagnetic resonance, NMR, and IR analysis. Specific potential benefits to this imprinting method include reactivity with a wide range of aromatic monomers to provide more diverse options for molecular recognition with the target analyte and thus polymer products with higher selectivity, mild reaction conditions for the enzyme polymerization step to enable imprinting against labile substrates, imprinted polymeric products that contain conjugated backbones that could be suitable for electronics-based biosensor applications, and a potential for combinatorial selection to further enhance specificity.     

Fluorescent competitive flow-through assay for chloramphenicol using molecularly imprinted polymers

It is a fact that molecular imprinting techniques have reached tremendous importance in the research of new artificial recognition systems. These methods resemble the mechanism of natural recognition, generally based on non-covalent interactions, but improving their stability by means of a simple and inexpensive technique. Molecular imprinting polymers (MIPs) are easily obtained by copolymerisation of suitable functional monomers and crosslinkers in the presence of the print molecule. Removal of the template leaves a polymer that selectively recognises it. In this work, different imprinted polymers for chloramphenicol (CAP) obtained using different monomers and polymerisation conditions were tested in a HPLC system, in order to obtain a highly selective material for CAP. The optimised MIP was then used as recognition phase in a fluorescent competitive flow assay to determine chloramphenicol.     

Molecularly imprinted polymers—A closer look at the control polymer used in determining the imprinting effect: A mini review

Molecular recognition displayed by naturally occurring receptors has continued to inspire new innovations aimed at developing systems that can mimic this natural phenomenon. Since 1930s, a technology called molecular imprinting for producing biomimetic receptors has been in place. In this technology, tailor made binding sites that selectively bind a given target analyte (also called template) are incorporated in a polymer matrix by polymerizing functional monomers and cross‐linking monomers around a target analyte followed by removal of the analyte to leave behind cavities specific to the analyte. The success of the imprinting process is defined by two main figures of merit, that is, the imprinting factor, and selectivity, which are determined by comparing the amount of target analyte or structural analogue bound by the molecularly imprinted polymer (MIP) and the nonimprinted polymer (NIP). NIP is a control synthesized alongside the MIP but in the absence of the template. However, questions arise on whether these figures of merit are reliable measures of the imprinting effect because of the significant differences between the MIP and the NIP in terms of their physical and chemical characteristics. Therefore, this review critically looks into this subject, with a view of defining the best approaches for determining the imprinting effect.     

Design and synthesis of molecularly imprinted polypyrrole based on nanoreactor SBA-15 for recognition of ascorbic acid

Molecular imprinting is an attractive technique for preparing mimics of natural and biological receptors. Nevertheless, molecular imprinting for aqueous systems remains a challenge due to the hydrogen bonding between templates and functional monomers destroyed in the bulk water. The hydrogen bonding between templates and monomers are the most crucial factor governing recognition, particularly in non-covalent molecularly imprinted polymers. Using mesoporous materials for molecular imprinting is an effective approach to overcome this barrier and to remove the limitations of the traditional molecularly imprinted polymers which include incomplete template removal, small binding capacity, slow mass transfer, and irregular materials shape. Here, SBA-15 was used as a mesoporous silica material for synthesis of molecularly imprinted polypyrrole. The pyrrole monomers and template molecules were immobilized onto the SBA-15 hexagonal channels, and then polymerization occurred. The resulting nanocomposites were characterized by Fourier transform infrared (FT-IR) analysis, scanning electron microscopy (SEM) and transmission electron microscopy (TEM) methods. In batch rebinding tests, the imprinted nanocomposites reached saturated adsorption within 100min and exhibited significant specific recognition toward the ascorbic acid (AA) with high adsorption capacity (83.7mgg(-1)). To further illustrate the recognition property of the imprinted nanocomposites, binary competitive and non-competitive adsorption experiments were performed with ascorbic acid, dopamine, paracetamol and epinephrine. The imprinting factors for these compounds in non-competitive adsorption experiments were 3.2, 1.5, 1.4 and 1.3, respectively. The results showed that the imprinted nanocomposites exhibited significant adsorption selectivity for the ascorbic acid against the related compounds.     

Building Fluorescent Sensors by Template Polymerization: The Preparation of a Fluorescent Sensor for -Tryptophan

The development of fluorescent sensors for organic molecules is of great practical importance in chemical, biological, and pharmaceutical sciences. Using -tryptophan as an example, we have studied a new way of making polymeric fluorescent sensors using template polymerization or molecular imprinting techniques. The fluorescent polymers were prepared using functional monomers with a fluorescent probe attached to it. The fluorescence of this polymer could be quenched by 4-nitrobenzaldehyde. Addition of the template molecules, -tryptophan, increased the fluorescence intensity of the imprinted polymer/quencher mixture in a concentration-dependent fashion, presumably through the displacement of the quencher from the binding sites by -tryptophan. This fluorescence intensity change upon mixing with -tryptophan allows the binding event to be detected easily. The sensor also exhibited enantioselectivity for the template molecules. For example, the effect of -tryptophan on the fluorescence intensity of the polymer is about 70% that of its -enantiomer. Furthermore, the effect of -phenylalanine and -alanine on the fluorescence intensity change is much smaller than that of -tryptophan. Because the approach used does not require the de novo design of the complementary binding site and does not rely on any specific structural features of the template molecule or prior knowledge of its three-dimensional structure, the same principle could potentially be useful for the future construction of practical fluorescent sensors for many other compounds.     

Stoichiometric noncovalent interaction in molecular imprinting

In this review article the function of the binding site monomers in the molecular imprinting procedure is discussed. Especially, new developments towards stoichiometric noncovalent interactions are highlighted. In stoichiometric noncovalent interactions template and binding site monomer in an 1:1 molar ratio are nearly completely bound to each other. This is only possible if the association constants are considerably high (K _ass < 900 M^–1). Using this type of interaction in molecular imprinting no excess of binding sites is necessary and binding sites are only located inside the imprinted cavity. Since all cavities can be reloaded these polymers show high capacity (e.g., for preparative application) and are especially suited for the synthesis of catalytically active imprinted polymers. Discussed are binding site interactions based on amidines (and guanidines), multiple hydrogen bonding, charge-transfer interactions, and host–guest inclusion. The systematic investigation of the underlying binding reaction is described in detail. With low-molecular weight model substances the thermodynamics of the association can be conveniently investigated, e.g., by NMR spectroscopy.     

Towards the rational development of molecularly imprinted polymers: 1H NMR studies on hydrophobicity and ion-pair interactions as driving forces for selectivity

The preparation of molecularly imprinted polymers (MIP) based on non-covalent interactions has become a widely used technique for creating highly specific sorbent materials predominantly used in separation chemistry. A crucial factor in a successful imprinting protocol is the optimisation of the template/functional monomer interaction in the pre-polymerisation mixture, eventually leading to a maximum of high-affinity binding sites in the resulting polymer matrix. In order to develop more efficient preparation technologies for imprinted polymers, two separate pre-polymerisation complexes were investigated by NMR spectroscopic techniques in order to identify the types of interactions occurring in the pre-polymerisation mixture, and their implications for the subsequently formed imprinted polymer. In particular, hydrophobic effects have been followed by NMR spectroscopy and their contribution to the selectivity of the resulting MIP has been investigated. The 2,4-D imprint system is used as an example to fundamentally study whether observations at the pre-polymerisation stage correlate with properties of the finally prepared MIP, and which parameters govern success of an imprinting protocol.     

Aflatoxin-selective molecularly-imprinted polymer membranes based on acrylate-polyurethane semi-interpenetrating polymer networks

Synthetic analogs of biological receptors able to group-selective recognition of aflatoxins were obtained using the combination of the technique of molecular imprinting with the method of computer modeling. The synthetic receptors were obtained in a form of thin and porous membranes based on semi-interpenetrating polymer networks. The selection of functional monomers able to noncovalent interactions with aflatoxins B1, B2, and G2 was based on the data of computer modeling. Allylamine, diethylaminoethylmethacrylate, and N,N'-methylenebisacrylamide, providing high binding energies with aflatoxins B1, B2, and G2 were selected as functional monomers for the formation of aflatoxin B1-imprinted polymer membranes. It was shown that aflatoxin-B1-imprinted polymeric membranes synthesized using N,N'-methylenebisacrylamide as a functional monomer were characterized with good physico-mechanical properties as well as good adsorbtion capability towards aflatoxin B1. Neglidgible levels of aflatoxin B1 adsorbtion on the surface of blank membranes were observed. High adsorbtion capability of the MIP membranes towards mycotoxins affiliated to the group of aflatoxins was demonstrated, while negligible adsorbtion of ochratoxin A was observed. Therefore, synthetic analogs of biological receptors able to group-selective recognition of aflatoxins in the range 1-1000 ppb were developed.