Membrane separations using molecularly imprinted polymers

This review presents an overview on the promising field of molecularly imprinted membranes (MIM). The focus is onto the separation of molecules in liquid mixtures via membrane transport selectivity. First, the status of synthetic membranes and membrane separation technology is briefly summarized, emphasizing the need for novel membranes with higher selectivities. Innovative principles for the preparation of membranes with improved or novel functionality include self-assembly or supramolecular aggregation as well as the use of templates. Based on a detailed analysis of the literature, the main established preparation methods for MIM are outlined: simultaneous membrane formation and imprinting, or preparation of imprinted composite membranes. Then, the separation capability of MIM is discussed for two different types, as a function of their barrier structure. Microporous MIM can continuously separate mixtures based on facilitated diffusion of the template, or they can change their permeability in the presence of the template ("gate effect"). Macroporous MIM can be developed towards molecule-specific membrane adsorbers. Emerging further combinations of molecularly imprinted polymers (MIPs), especially MIP nanoparticles or microgels, with membranes and membrane processes are briefly outlined as well. Finally, the application potential for advanced MIM separation technologies is summarized.     

HPLC-based bioseparations using molecularly imprinted polymers

HPLC-based separations of amino acids and peptides, nucleotide bases, drugs, sugars and steroids using molecularly imprinted polymers (MIPs) have been reviewed in this article. The molecular recognition mechanisms of the template molecules on the MIPs in organic and aqueous eluents were discussed. Furthermore, new polymerization methods suitable for preparations of HPLC columns and packing materials using molecular imprinting techniques, and their applications to HPLC-based separations are also dealt with.     

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.     

Capacitive detection of glucose using molecularly imprinted polymers

A novel glucose biosensor based on capacitive detection has been developed using molecularly imprinted polymers. The sensitive layer was prepared by electropolymerization of o-phenylenediamine on a gold electrode in the presence of the template (glucose). Cyclic voltammetry and capacitive measurements monitored the process of electropolymerization. Surface uncovered areas were plugged with 1-dodecanethiol to make the layer dense, and the insulating properties of the layer were studied in the presence of redox couples. The template molecules and the nonbound thiol were removed from the modified electrode surface by washing with distilled water. A capacitance decrease could be obtained after injection of glucose. The electrode constructed similarly but with ascorbic acid or fructose only showed a small response compared with glucose. The stability and reproducibility of the biosensor were also investigated.     

The role of living/controlled radical polymerization in the formation of improved imprinted polymers

In this work, living/controlled radical polymerization (LRP) is compared with conventional free radical polymerization in the creation of highly and weakly cross-linked imprinted poly(methacrylic acid-co-ethylene glycol dimethacrylate) networks. It elucidates, for the first time, the effect of LRP on the chain level and begins to explain why the efficiency of the imprinting process is improved using LRP. Imprinted polymers produced via LRP exhibited significantly higher template affinity and capacity compared with polymers prepared using conventional methods. The use of LRP in the creation of highly cross-linked imprinted polymers resulted in a fourfold increase in binding capacity without a decrease in affinity; whereas weakly cross-linked gels demonstrated a nearly threefold increase in binding capacity at equivalent affinity when LRP was used. In addition, by adjusting the double bond conversion, we can choose to increase either the capacity or the affinity in highly cross-linked imprinted polymers, thus allowing the creation of imprinted polymers with tailorable binding parameters. Using free radical polymerization in the creation of polymer chains, as the template-monomer ratio increased, the average molecular weight of the polymer chains decreased despite a slight increase in the double bond conversion. Thus, the polymer chains formed were shorter but greater in number. Using LRP neutralized the effect of the template. The addition of chain transfer agent resulted in slow, uniform, simultaneous chain growth, resulting in the formation of longer more monodisperse chains. Reaction analysis revealed that propagation time was extended threefold in the formation of highly cross-linked polymers when LRP techniques were used. This delayed the transition to the diffusion-controlled stage of the reaction, which in turn led to the observed enhanced binding properties, decreased polydispersity in the chains, and a more homogeneous macromolecular architecture.     

Tools for fluorescent molecularly imprinted polymers

A linear co-polymer of hexyl acrylate and quinine acrylate was prepared anchored to cellulose filtration membranes. These were used to probe quenching of the tethered fluorophore by test compounds in solution for the validation of imprinted polymer fluorescence studies. The results are compared with simple solution phase quenching studies and also for two membrane-bound imprinted polymers containing the same fluorophore.     

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.     

Mobile phase effects on enantiomer resolution using molecularly imprinted polymers

The aim of this study was to rationalise retention behaviour of a chiral solute on molecularly imprinted polymer (MIP) HPLC stationary phases in terms of variation of the mobile phase. It is generally held that the most important interaction governing the separation of enantiomers on such materials is H-bonding, and that retention times increase with decreasing H-bonding potential of the mobile phase. Previous studies have largely concerned mobile phases containing chloroform with acetic acid as a polar modifier. Boc-L-Phenylalanine (Boc-L-Phe-OH) MIPs were prepared, processed, and packed into HPLC columns, which were then used to investigate the retention characteristics of Boc-L-Phe-OH and Boc-D-Phe-OH with a range of mobile phases. The main observations were as follows: (1) in chloroform-based mobile phases there was generally a linear relationship between the H-bond donator factor of the polar modifier and capacity (K′). Results also indicated a hydrogen bond donor parameter value for a polar modifier at which retention became concentration independent; (2) For given values of K′_L, K′_D varied depending on the polar modifier, indicating that enantiomer resolution was solvent dependent; (3) Using mobile phases based on solvents of lower polarity/H-bonding potential than chloroform, substantial increases in K′ were observed, although enantioselectivity was greatly reduced. Chirality 9:238–242, 1997. © 1997 Wiley-Liss, Inc.     

Investigation of polyacrylamide hydrogel-based molecularly imprinted polymers using protein gel electrophoresis

In conjunction with polyacrylamide gel electrophoresis (PAGE), molecular imprinting methods have been applied to produce a multilayer mini-slab in order to evaluate how selectively and specifically a hydrogel-based molecularly imprinted polymer (MIP) binds bovine haemoglobin (BHb, ~64.5 kDa). A three-layer mini-slab comprising an upper and lower layer and a MIP, or a non-imprinted control polymer dispersion middle layer has been investigated. The discriminating MIP layer, also based on polyacrylamide, was able to specifically bind BHb molecules in preference to a protein similar in molecular weight such as bovine serum albumin (BSA, ~66 kDa). Protein staining allowed us to visualise the protein retention strength of the MIP layer under the influence of an electric field. This method could be applied to other proteins with implications in effective protein capture, disease diagnostics, and protein analysis.     

Separation and sensing based on molecular recognition using molecularly imprinted polymers

Molecular recognition-based separation and sensing systems have received much attention in various fields because of their high selectivity for target molecules. Molecular imprinting has been recognized as a promising technique for the development of such systems, where the molecule to be recognized is added to a reaction mixture of a cross-linker(s), a solvent(s), and a functional monomer(s) that possesses a functional groups(s) capable of interacting with the target molecule. Binding sites in the resultant polymers involve functional groups originating from the added functional monomer(s), which can be constructed according to the shape and chemical properties of the target molecules. After removal of the target molecules, these molecularly imprinted complementary binding sites exhibit high selectivity and affinity for the template molecule. In this article, recent developments in molecularly imprinted polymers are described with their applications as separation media in liquid chromatography, capillary electrophoresis, solid-phase extraction, and membranes. Examples of binding assays and sensing systems using molecularly imprinted polymers are also presented.     

Selective solid-phase extraction of bio- and environmental samples using molecularly imprinted polymers

Of the many applications of molecular imprinting in analytical separation science, the one with highest potential of soon being used in routine analysis is that of solid-phase extraction. Already several examples of selective pre-concentration of biological and environmental samples have been reported. The interest in imprinted extraction sorbents originates from the high selectivities and affinities obtainable, properties which can be qualitatively and quantitatively pre-determined for a particular analyte and separation by the imprinting process. This review summarises work published on molecular imprinted solid-phase extraction and discusses some imprinted-sorbent specific method development issues.     

Separation and screening of compounds of biological origin using molecularly imprinted polymers

Molecularly imprinted polymers (MIPs) represent a new class of materials possessing high selectivity and affinity for the target molecule. Since their discovery in 1972, molecularly imprinted polymers have attracted considerable interest from bio- and chemical laboratories to pharmaceutical institutes. They have been utilized as sensors, catalysts, sorbents for solid-phase extraction, stationary phase for liquid chromatography, mimics of enzymes, receptors and antibodies. Among which, the application of molecularly imprinted polymers for molecular recognition-based separation and screening of compounds has undergone rapid extension and received much attention in recent years. This article mainly focuses on the separation and screening of certain pharmacophoric compounds of interests from biological origin using molecular imprinting technology. Examples of extraction and recognition of active components as anti-tumors or anti-Hepatitis C virus inhibitors from Chinese traditional herbs using molecularly imprinting technology are particularized in this article. Comparison between the screening effect based on MIPs and that based on antibodies is also represented. Consequently the merits and demerits of these two technologies are highlighted.     

Characterization of molecularly imprinted polymers using a new polar solvent titration method

A new method of characterizing molecularly imprinted polymers (MIPs) was developed and tested, which provides a more accurate means of identifying and measuring the molecular imprinting effect. In the new polar solvent titration method, a series of imprinted and non‐imprinted polymers were prepared in solutions containing increasing concentrations of a polar solvent. The polar solvent additives systematically disrupted the templation and monomer aggregation processes in the prepolymerization solutions, and the extent of disruption was captured by the polymerization process. The changes in binding capacity within each series of polymers were measured, providing a quantitative assessment of the templation and monomer aggregation processes in the imprinted and non‐imprinted polymers. The new method was tested using three different diphenyl phosphate imprinted polymers made using three different urea functional monomers. Each monomer had varying efficiencies of templation and monomer aggregation. The new MIP characterization method was found to have several advantages. To independently verify the new characterization method, the MIPs were also characterized using traditional binding isotherm analyses. The two methods appeared to give consistent conclusions. First, the polar solvent titration method is less susceptible to false positives in identifying the imprinting effect. Second, the method is able to differentiate and quantify changes in binding capacity, as measured at a fixed guest and polymer concentration, arising from templation or monomer aggregation processes in the prepolymerization solution. Third, the method was also easy to carry out, taking advantage of the ease of preparing MIPs. Copyright © 2014 John Wiley & Sons, Ltd.     

Fluorescence anisotropy studies of molecularly imprinted polymers.

A molecularly imprinted polymer (MIP) is a biomimetic material that can be used as a biochemical sensing element. We studied the steady-state and time-resolved fluorescence and fluorescence anisotropy of anthracene-imprinted polyurethane. We compared MIPs with imprinted analytes present, MIPs with the imprinted analytes extracted, MIPs with rebound analytes, non-imprinted control polymers (non-MIPs) and non-MIPs bound with analytes to understand MIP's binding behaviour. MIPs and non-MIPs had similar steady-state fluorescence anisotropy in the range 0.11-0.24. Anthracene rebound in MIPs and non-MIPs had a fluorescence lifetime of tau = 0.64 ns and a rotational correlation time of phi(F) = 1.2-1.5 ns, both of which were shorter than that of MIPs with imprinted analytes present (tau = 2.03 ns and phi(F) = 2.7 ns). The steady-state anisotropy of polymer solutions increased exponentially with polymerization time and might be used to characterize the polymerization extent in situ.     

Investigation of disaccharide recognition by molecularly imprinted polymers

The selectivity of carbohydrate-imprinted polymers for several disaccharides, namely cellobiose, maltose, lactose and gentiobiose, is investigated. An ternary ligand–Cu(II)–carbohydrate complex was formed in alkaline solution and captured afterwards in the polymer. The accessibility of the polymer matrix for disaccharides was investigated by HPLC analysis, refractometry and ¹H NMR spectroscopy applying excess of the original template during rebinding experiments under saturation conditions in unbuffered, aqueous solution at neutral pH and 20°C. The selective discrimination of the - and -glycosidic linkage of cellobiose and maltose is demonstrated. It is further shown, that the disaccharide-imprinted polymers slightly distinguish between the 1,4-- and the 1,6--glycosidic linkage of cellobiose and gentiobiose, while cellobiose and lactose are not selectively recognized. Due to the weak apparent binding constant of the functional Cu(II) monomers with the targeted disaccharides at physiological pH, the recognition process is dominated by the shape of the created imprinted cavity under the applied conditions.     

Towards the rational design of molecularly imprinted polymers

Efforts to elucidate the mechanisms underlying the formation of binding sites in molecularly imprinted polymers (MIPs) and of MIP-ligand binding events are presented in the context of a thermodynamic treatment of MIP recognition phenomena.     

Preparation of molecularly imprinted polymers for artemisinin based on the surfaces of silica gel

Artemisinin is an effective antimalarial drug isolated from the herbal medicine Artemisia annua L. Molecular imprinting is a technique of preparing molecularly imprinted polymers (MIPs) which can specifically recognize the imprinted template molecules. In this work, silica gel were used as supporting matrix, and vinyltriethoxysilane (VTES) was grafted onto its surface. The preparation of MIPs for artemisinin was performed on the surfaces of the modified silica gel using artemisinin as the template, acrylamide (AM) and methacrylic acid (MAA) as the functional monomers, ethylene glycol dimethacrylate (EGDMA) as the cross-linker and 2,2'-azo-bis-isobutyronitrile (AIBN) as the initiator. Scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FT-IR) and pore size analysis were used to characterize the prepared MIPs. The adsorption kinetic curve, adsorption isotherm and selective adsorption were measured by static method. The adsorption reached equilibrium at about 10 h, while fast adsorption took place during the first 2-3 h. The maximum adsorption capacity has been found to be 37.13 mg/g according to calculation with Langmuir-Freundlich isotherm. The electivity coefficients of MIPs for artemisinin with respect to artemether and arteether were 2.88 and 3.38, respectively. The results showed that the MIPs possessed good specific adsorption capacity and selectivity for artemisinin.     

Grafting of molecularly imprinted polymers on iniferter-modified carbon nanotube

Molecularly imprinted polymers (MIPs) were grafted on iniferter-modified carbon nanotube (CNT). Tween 20 was first immobilized on CNT by hydrophobic interactions. The hydroxyl-functionalized CNT was modified by silanisation with 3-chloropropyl trimethoxysilane. The iniferter groups were then introduced by reacting the CNT-bound chloropropyl groups with sodium N,N-diethyldithiocarbamate. UV light-initiated copolymerization of ethylene glycol dimethacrylate (crosslinking agent) and methacrylic acid (functional monomer) resulted in grafting of MIP on CNT for theophylline as a model template. MIPs grafted on CNT were characterized with elemental analysis, scanning electron microscopy, and thermogravimetric analysis. The theophylline-imprinted polymer on CNT showed higher binding capacity for theophylline than non-imprinted polymer on CNT and selectivity for theophylline over caffeine and theobromine (similar structure molecules). The data of theophylline and caffeine binding into the theophylline-imprinted polymer correlated well with the Scatchard plot. These MIPs on CNT can potentially be applied to probe materials in biosensor system based on CNT field effect transistor.     

Binding cross-reactivity of Boc-phenylalanine enantiomers on molecularly imprinted polymers

A potential problem associated with molecularly imprinted polymer (MIP) sorbents is that of cross-reactivity. In this study three MIPs (imprinted with Boc-L-phenylalanine, Boc-L-alanine, Boc-L-glutamic acid) plus a non-imprinted control were prepared and examined for their ability to bind differentially the enantiomers of boc-protected phenylalanine in an effort to quantify cross-reactivity and to develop a predictive model. Batch rebinding studies showed a degree of predictability for a number of MIP-ligand pairs, but other combinations showed unexpectedly high levels of cross-reactivity. Despite the general acceptance of heterogeneity of MIP binding sites, many previous studies report linear Scatchard plots, which is indicative of binding site homogeneity. The data from this study produced curves, clearly demonstrating heterogeneity. The theoretical and practical implications of this heterogeneity are discussed. Chirality 9:233–237, 1997. © 1997 Wiley-Liss, Inc.